81,9(56,7<2),168%5,$
'RFWRUDO6FKRRORI%LRORJLFDODQG0HGLFDO6FLHQFHV
3K'3URJUDPLQ1HXURELRORJ\
)XQFWLRQDODQGPROHFXODUGHIHFWVRIKXPDQLQGXFHG
SOXULSRWHQWVWHPFHOOGHULYHGQHXURQVIURPSDWLHQWV
ZLWKWKHQHXURGHJHQHUDWLYHGLVRUGHU
DWD[LDWHODQJLHFWDVLD
7XWRU'U'RPHQLFR'HOLD
6XSHUYLVRUDQG&RRUGLQDWRU3URI'DQLHOD3DURODUR
3K'WKHVLVRI
'U(OHQD)XVDU3ROL
;;9,,&\FOH²
7KHUHVHDUFKZRUNSUHVHQWHGLQWKLVWKHVLVZDVFDUULHGRXWDWWKH
)RQGD]LRQH,5&&6,VWLWXWR1D]LRQDOHGHL7XPRUL
'HSDUWPHQWRI([SHULPHQWDO2QFRORJ\
9LD$PDGHR
0LODQR
$FNQRZOHGJHPHQWV
$FNQRZOHGJHPHQWV
,ZRXOGOLNHWRH[SUHVVP\YHU\JUHDWDSSUHFLDWLRQWR'U'RPHQLFR'HOLDIRUKLVYDOXDEOH
DQG FRQVWUXFWLYH VXJJHVWLRQV GXULQJ WKH SODQQLQJ DQG GHYHORSPHQW RI WKLV UHVHDUFK ZRUN WR
'U/XLJL &DUOHVVL IRU SUHFLRXV FROODERUDWLRQ WKURXJKRXW DOO DVSHFWV RI WKH SURMHFW DQG WR
3URI'DQLHOD3DURODURIRUKHUSURIHVVLRQDOJXLGDQFHDQGYDOXDEOHVXSSRUW
, ZLVK WR DFNQRZOHGJH WKH KHOS SURYLGHG E\ 'U *LXOLD %HFKL 'HSDUWPHQW RI
1HXURSK\VLRSDWKRORJ\ )RQGD]LRQH ,5&&6 ,VWLWXWR 1HXURORJLFR &DUOR %HVWD 0LODQ ,WDO\ IRU
HOHFWURSK\VLRORJLFDO UHFRUGLQJV DQG DQDO\VLV DQG E\ 'U%DUEDUD 3DVFXFFL &15 ,QVWLWXWH RI
&U\VWDOORJUDSK\5RPH,WDO\IRUEDVHH[FLVLRQUHSDLUDQDO\VLV,DOVRZLVKWRWKDQN'U&KLDUD
9HUSHOOL &15 ,QVWLWXWH RI 1HXURVFLHQFH DQG 'HSDUWPHQW RI %LRWHFKQRORJ\ DQG 7UDQVODWLRQDO
0HGLFLQH 0LODQ ,WDO\ IRU SURYLGLQJ XV ZLWK FRQWURO SULPDU\ GHUPDO ILEUREODVWV DQG
'U&KULVWLDQ8QJHU&HQWUHIRU6WHP&HOO%LRORJ\8QLYHUVLW\RI6KHIILHOG8.IRUSURYLGLQJXV
ZLWKDQWL66($DQGDQWL75$DQWLERGLHV
7KLV ZRUN ZDV ILQDQFLDOO\ VXSSRUWHG E\ &RPLWDWR 7HOHWKRQ )RQGD]LRQH 2QOXV JUDQW
QR**3DQGE\WKH,WDOLDQ$VVRFLDWLRQIRU&DQFHU5HVHDUFK$,5&JUDQWQR,*
7DEOHRIFRQWHQWV
7DEOHRIFRQWHQWV
$EVWUDFW««««««««««««««««««««««««««««««««««««««««««««««
/LVWRIDEEUHYLDWLRQV«««««««««««««««««««««««««««««««««««««
,QWURGXFWLRQ««««««««««««««««««««««««««««««««««««««««««
$WD[LDWHODQJLHFWDVLD««««««««««««««««««««««««««««««««««««««««««
1HXURORJLFDOPDQLIHVWDWLRQVDQGQHXURSDWKRORJLFDOILQGLQJV«««««««««««««««««
1RQQHXURORJLFDOFOLQLFDOIHDWXUHV««««««««««««««««««««««««««««««««
'LDJQRVLVDQGWUHDWPHQW««««««««««««««««««««««««««««««««««««
$70JHQHDQGSURWHLQ«««««««««««««««««««««««««««««««««««««««««
)XQFWLRQVRI$70DQGSRVVLEOHFDXVHVRIQHXURGHJHQHUDWLRQLQDWD[LDWHODQJLHFWDVLD«««
5ROHRI$70LQWKH'1$GDPDJHUHVSRQVHDQGLQFHOOF\FOHFRQWURO«««««««««««««
$GGLWLRQDOUROHVRI$70LQWKHF\WRSODVPDQGLQWKHQXFOHXV««««««««««««««««
6SHFLILFUROHVRI$70LQQHXURQV««««««««««««««««««««««««««««««««
0RGHOVRIQHXURGHJHQHUDWLRQLQDWD[LDWHODQJLHFWDVLD««««««««««««««««««««««
$70GHILFLHQWDQLPDOPRGHOV«««««««««««««««««««««««««««««««««
$70GHILFLHQWKXPDQFHOOXODUPRGHOV««««««««««««««««««««««««««««
+XPDQLQGXFHGSOXULSRWHQWVWHPFHOOGHULYHGQHXUDOFHOOVIURPSDWLHQWVZLWKDWD[LD
WHODQJLHFWDVLD«««««««««««««««««««««««««««««««««««««««««««««
$LPRIWKHSURMHFW«««««««««««««««««««««««««««««««««««««
0DWHULDOVDQGPHWKRGV«««««««««««««««««««««««««««««««««
3ULPDU\FHOOFXOWXUHV«««««««««««««««««««««««««««««««««««««««««
*HQHUDWLRQDQGFXOWXUHRIKXPDQLQGXFHGSOXULSRWHQWVWHPFHOOV«««««««««««««««
/HQWLYLUDOLQIHFWLRQ«««««««««««««««««««««««««««««««««««««««
7DEOHRIFRQWHQWV
7UDQVLHQWWUDQVIHFWLRQ«««««««««««««««««««««««««««««««««««««
0DLQWHQDQFHRIKXPDQLQGXFHGSOXULSRWHQWVWHPFHOOFXOWXUHV««««««««««««««««
,QYLWURSOXULSRWHQF\DVVD\««««««««««««««««««««««««««««««««««««««
*HQHUDWLRQDQGGLIIHUHQWLDWLRQRIKXPDQQHXUDOSUHFXUVRUFHOOV««««««««««««««««
:HVWHUQEORWDQDO\VLV«««««««««««««««««««««««««««««««««««««««««
:KROHFHOOH[WUDFWVDQGSURWHLQTXDQWLILFDWLRQ««««««««««««««««««««««««
6'6SDJHDQG:HVWHUQEORW«««««««««««««««««««««««««««««««««««
,PPXQRIOXRUHVFHQFH«««««««««««««««««««««««««««««««««««««««««
(OHFWURSK\VLRORJLFDOUHFRUGLQJVDQGDQDO\VLV«««««««««««««««««««««««««««
&RPHWDVVD\««««««««««««««««««««««««««««««««««««««««««««««
1HXWUDOFRPHWDVVD\««««««««««««««««««««««««««««««««««««««
$ONDOLQHFRPHWDVVD\««««««««««««««««««««««««««««««««««««««
0RGLILHGDONDOLQHFRPHWDVVD\«««««««««««««««««««««««««««««««««
'HWHFWLRQRIDŽ+$;DQG%3QXFOHDUIRFL««««««««««««««««««««««««««««
,QYLWUREDVHH[FLVLRQUHSDLUDVVD\«««««««««««««««««««««««««««««««««
5DSLGDSSURDFKWR'1$DGGXFWUHFRYHU\DVVD\««««««««««««««««««««««««
1HXURQDOYLDELOLW\DVVD\««««««««««««««««««««««««««««««««««««««
&HOOF\FOHSKDVHDQDO\VLV««««««««««««««««««««««««««««««««««««««
5HVXOWV««««««««««««««««««««««««««««««««««««««««««««
*HQHUDWLRQDQGFKDUDFWHUL]DWLRQRIKXPDQLQGXFHGSOXULSRWHQWVWHPFHOOVGHULYHGIURP
DWD[LDWHODQJLHFWDVLDDQGFRQWUROILEUREODVWV««««««««««««««««««««««««««
*HQHUDWLRQRIQHXUDOSUHFXUVRUFHOOVDQGQHXURQVIURPDWD[LDWHODQJLHFWDVLDDQGFRQWURO
KXPDQLQGXFHGSOXULSRWHQWVWHPFHOOV«««««««««««««««««««««««««««««««
(OHFWURSK\VLRORJLFDOFKDUDFWHUL]DWLRQ«««««««««««««««««««««««««««««««
7DEOHRIFRQWHQWV
'1$GDPDJHUHVSRQVHDQGUHSDLULQSRVWPLWRWLFDWD[LDWHODQJLHFWDVLDQHXURQV««««««
%DVHH[FLVLRQUHSDLUFDSDFLW\LQXQGLIIHUHQWLDWHGDQGGLIIHUHQWLDWHGQHXUDOSUHFXUVRUFHOOV
DQGUHSDLURIVLQJOHVWUDQGEUHDNVLQSRVWPLWRWLFQHXURQV«««««««««««««««««««
7UHDWPHQWRIDWD[LDWHODQJLHFWDVLDDQGFRQWUROQHXURQDOFHOOVZLWKJHQRWR[LFDQGR[LGDWLYH
DJHQWV««««««««««««««««««««««««««««««««««««««««««««««««««
$FFXPXODWLRQRIWRSRLVRPHUDVH'1$FOHDYDJHFRPSOH[HVLQDWD[LDWHODQJLHFWDVLD
SRVWPLWRWLFQHXURQV««««««««««««««««««««««««««««««««««««««««««
'1$GDPDJHLQGXFHG*6FKHFNSRLQWLQUHODWLRQWRGHYHORSPHQWVWDJH««««««««««
'LVFXVVLRQ«««««««««««««««««««««««««««««««««««««««««««
)XWXUHSHUVSHFWLYHV«««««««««««««««««««««««««««««««««««
5HIHUHQFHV«««««««««««««««««««««««««««««««««««««««««««
3XEOLFDWLRQVLQLQWHUQDWLRQDO,6,MRXUQDOV«««««««««««««««««««
$EVWUDFW
$EVWUDFW
$WD[LDWHODQJLHFWDVLD$7LVDUDUHKHUHGLWDU\HDUO\RQVHWQHXURGHJHQHUDWLYHGLVRUGHU
FDXVHG E\ LQDFWLYDWLRQ RI WKH $70 VHULQHWKUHRQLQH SURWHLQ NLQDVH ZKLFK LV WKH PDMRU
UHJXODWRU RI WKH '1$ GDPDJH UHVSRQVH WR GRXEOHVWUDQG EUHDNV '6%VDQGZRUNV LQ VHQVLQJ
DQG VLJQDOLQJ R[LGDWLYH VWUHVV 'LVHDVH PRGHOV DUH HVVHQWLDO IRU XQUDYHOLQJ WKH PHFKDQLVPV
XQGHUO\LQJ WKH QHXURSDWKRORJ\ EXW $70 NQRFNRXW PLFH GR QRW UHFDSLWXODWH WKH FHQWUDO
QHUYRXV V\VWHP SKHQRW\SH DQG QHXUDO VWHP FHOOV DUH YHU\ KHWHURJHQHRXV ,Q WKLV ZRUN ZH
DSSOLHG D UHSURJUDPPLQJ DSSURDFK WR JHQHUDWH DQ LQ YLWUR KXPDQ $7 PRGHO LQ RUGHU WR
LQYHVWLJDWHWKHRXWFRPHRI$70DEODWLRQLQQHXURQV
:HGHULYHGLQGXFHGSOXULSRWHQWVWHPFHOOVIURP$7DQGQRUPDOFRQWUROILEUREODVWVE\
LQWURGXFLQJ WKH UHSURJUDPPLQJ IDFWRUV 2&7 ./) 62; DQG O0<& DQG IURP WKHVH ZH
REWDLQHG H[SDQGDEOH QHVWLQSRVLWLYH QHXUDO SUHFXUVRU FHOOV 13&V YLD HPEU\RLG ERG\ DQG
QHXUDO URVHWWH IRUPDWLRQ )ROORZLQJ GLIIHUHQWLDWLRQ 13&V JDYH ULVH WR D KLJK SHUFHQWDJH RI
PDWXUH QHXURQV H[SUHVVLQJ ǃWXEXOLQ ,,, DQG PLFURWXEXOHDVVRFLDWHG SURWHLQ 2XU UHVXOWV
VKRZ WKDW HOHFWURSK\VLRORJLFDO SURSHUWLHV ZHUH VLPLODU LQ FRQWURO DQG $7 FHOOV $OVR EDVH
H[FLVLRQ UHSDLU ZKLFK LV LPSRUWDQW IRU WKH UHPRYDO RI '1$ VLQJOH VWUDQG EUHDNV FDXVHG E\
R[LGDWLYHVWUHVVDQGVLQJOHVWUDQGEUHDNUHSDLUZHUHQRUPDOLQ$7FHOOV2QWKHRWKHUKDQG$
7 QHXURQV GLVSOD\HG DWWHQXDWHG '1$ GDPDJH UHVSRQVH DQG UHSDLU IROORZLQJ LUUDGLDWLRQ DQG
ZHUHUHVLVWDQWWRDSRSWRVLVLQGXFHGE\JHQRWR[LFDJHQWVEXWQRWE\R[LGDWLYHDJHQWV0RUHRYHU
PXWDQWQHXURQVGLVSOD\HGGHILFLWVLQWKHH[SUHVVLRQRIWKHV\QDSWLFPDUNHUVV\QDSWRSK\VLQDQG
SRVWV\QDSWLFGHQVLW\SURWHLQDVZHOODVRIWKHQHXURQDOJURZWKDVVRFLDWHGSURWHLQ6&*DQG
.FKDQQHOLQWHUDFWLQJSURWHLQV.&K,31RWDEO\$7QHXURQVH[KLELWHGDEQRUPDODFFXPXODWLRQ
RI WRSRLVRPHUDVH '1$ FRYDOHQW FRPSOH[HV ZKLFK PD\ FDXVH LPSDLUPHQW RI WUDQVFULSWLRQ
HORQJDWLRQ7KHVHILQGLQJVUHYHDOWKDWLQQHXUDOFHOOV$70GHILFLHQF\VXSSUHVVHVWKHUHVSRQVH
DQG UHSDLU RI '1$ '6%V LPSDLUV QHXURQDO PDWXUDWLRQ DQG FRXOG SURPRWH WUDQVFULSWLRQDO
GHFOLQHSRVVLEO\FRQWULEXWLQJWRQHXURGHJHQHUDWLRQLQ$7
$EVWUDFW
7KLV SUHYLRXVO\ XQGRFXPHQWHG IXQFWLRQDO DQG PROHFXODU FKDUDFWHUL]DWLRQ RI SDWLHQW
GHULYHG QHXURQDO FHOOV UHSUHVHQWV WKH ILUVW VWHS LQ WKH HPSOR\PHQW RI WKHVH FHOOV DV WKH PRVW
LQIRUPDWLYH LQ YLWUR PRGHO WR LQYHVWLJDWH QHXURGHJHQHUDWLRQ LQ $7 0RUHRYHU WKLV PRGHO
UHSUHVHQW D SRZHUIXO WRRO IRU IXUWKHU H[SORULQJ WKH PHFKDQLVPV WKDW FDXVH GHJHQHUDWLRQ RI
KXPDQ$70GHILFLHQWQHXURQVDQGLWPD\EHH[SORLWHGIRULGHQWLI\LQJWKHUDSHXWLFWDUJHWVDQG
IRUVFUHHQLQJPROHFXOHVZLWKSRWHQWLDOQHXURSURWHFWLYHSURSHUWLHV
/LVWRIDEEUHYLDWLRQV
/LVWRIDEEUHYLDWLRQVDQGV\PEROV
$7DWD[LDWHODQJLHFWDVLD
$70DWD[LDWHODQJLHFWDVLDPXWDWHG
%(5EDVHH[FLVLRQUHSDLU
E)*)EDVLFILEUREODVWJURZWKIDFWRU
&+.FKHFNSRLQWNLQDVH
&37FDPSWRWKHFLQ
'$3,·GLDPLGLQRSKHQ\OLQGROH
'6%GRXEOHVWUDQGEUHDN
(%HPEU\RLGERG\
(*)HSLGHUPDOJURZWKIDFWRU
(6&HPEU\RQLFVWHPFHOO
+5KRPRORJRXVUHFRPELQDWLRQ
L36&LQGXFHGSOXULSRWHQWVWHPFHOO
,5LRQL]LQJUDGLDWLRQ
05105(5$'1%6
1+(-QRQKRPRORJRXVHQGMRLQLQJ
13&QHXUDOSUHFXUVRUFHOO
16&QHXUDOVWHPFHOO
30()&)/SULPDU\PRXVHHPEU\RQLFILEUREODVWVVWUDLQ&)
5$'$5UDSLGDSSURDFKWR'1$DGGXFWUHFRYHU\
526UHDFWLYHR[\JHQVSHFLHV
66%VLQJOHVWUDQGEUHDN
7RSWRSRLVRPHUDVH,
7RSFF7RSFOHDYDJHFRPSOH[
,QWURGXFWLRQ
,QWURGXFWLRQ
$WD[LDWHODQJLHFWDVLD
$WD[LDWHODQJLHFWDVLD $7 DOVR NQRZQ DV /RXLV%DU V\QGURPH LV D UDUH DXWRVRPDO
UHFHVVLYH GLVRUGHU ZKLFK OHDGV WR D FRPSOH[ DQG YDULDEO\ VHYHUH FRPELQDWLRQ RI GLVHDVH
SKHQRW\SHV
LQFOXGLQJ
QHXURGHJHQHUDWLRQ
WHODQJLHFWDVLD
FDQFHU
SUHGLVSRVLWLRQ
UDGLRVHQVLWLYLW\FKURPRVRPDOLQVWDELOLW\DQGLPPXQRGHILFLHQF\$7ZDVILUVWUHSRUWHGLQ
WKUHHSDWLHQWVE\6\OODEDDQG+HQQHULQIXUWKHUGHVFULEHGLQDQDGGLWLRQDOSDWLHQWE\
/RXLV%DULQDQGILQDOO\UHFRJQL]HGDVDVHSDUDWHGLVHDVHHQWLW\LQE\%RGHUDQG
6HGJZLFN 7KH GLVHDVH LV EURDGO\ GLVWULEXWHG WKURXJKRXW WKH ZRUOG DQG RFFXUV HTXDOO\
DPRQJPDOHVDQGIHPDOHVRIDOOUDFHVZLWKDQLQFLGHQFHRIDERXWLQOLYHELUWKV
DQG D FDUULHU IUHTXHQF\ HVWLPDWHG WR EH DSSUR[LPDWHO\ 7KH PRQRJHQLF QDWXUH RI WKH
GLVRUGHU ZDV RQO\ HVWDEOLVKHG LQ ZKHQ 6DYLWVN\ HW DO GLVFRYHUHG WKDW $7 LV FDXVHG E\
YDU\LQJ GHJUHHV RI PXWDWLRQDO LQDFWLYDWLRQ RI WKH DWD[LDWHODQJLHFWDVLD PXWDWHG $70
VHULQHWKUHRQLQHSURWHLQNLQDVH
1HXURORJLFDOPDQLIHVWDWLRQVDQGQHXURSDWKRORJLFDOILQGLQJV
7KHFOLQLFDOKDOOPDUNDQGPRVWGHYDVWDWLQJIHDWXUHRI$7LVHDUO\RQVHWFHUHEHOODUDWD[LD
ZKLFK FKDUDFWHUL]HV DOO FDVHV DQG LV XVXDOO\ WKH ILUVW SUHVHQWLQJ V\PSWRP EHFRPLQJ PDQLIHVW
EHWZHHQ DQG PRQWKV RI DJH DQG DIIHFWLQJ VLWWLQJ EDODQFH DQG ZDONLQJ &HUHEHOODU
G\VIXQFWLRQ VXEVHTXHQWO\ VSUHDGV WR LQYROYH WKH H[WUHPLWLHV DQG VSHHFK ZLWK FRQVHTXHQW
G\VDUWKULDRFXORPRWRUDSUD[LDQ\VWDJPXVLPSDVVLYHIDFLHVG\VSKDJLDDQGLPSDLUPHQWRIILQH
PRWRU IXQFWLRQV &KRUHRDWKHWRVLV DQG G\VWRQLD DUH DOVR W\SLFDO PDQLIHVWDWLRQV GXULQJ WKH
SURJUHVVLRQ RI WKH GLVHDVH $OWKRXJK WKH JURVV PLOHVWRQHV RI HDUO\ PRWRU GHYHORSPHQW DUH
XVXDOO\ DFTXLUHG RQ WLPH WKDQNV WR GHYHORSPHQWDO SURJUHVV WKH VORZO\ DGYDQFLQJ
QHXURGHJHQHUDWLRQ FDXVHV DIIHFWHG FKLOGUHQ WR UHTXLUH D ZKHHOFKDLU E\ WKHLU HDUO\ WHHQV
,QWURGXFWLRQ
6WULNLQJO\ WKH UHOHQWOHVV QHXURORJLFDO LPSDLUPHQW
LPSDLUPHQW VHHPV WR VSDUH FRJQLWLYH IXQFWLRQ DQG PRVW
$7LQGLYLGXDOV DSSHDUWR KDYH D QRUPDO ,4
,4 DOWKRXJKWKHUH DUHLQGLFDWLRQV RI PHQWDO GHFOLQH
ZLWKRQJRLQJGLVHDVHFRXUVH
)URPDKLVWRDQDWRPLFDOSRLQWRIYLHZWKHGHYDVWDWLQJQHXURGHJH
DQDWRPLFDOSRLQWRIYLHZWKHGHYDVWDWLQJQHXURGHJH
DQDWRPLFDOSRLQWRIYLHZWKHGHYDVWDWLQJQHXURGHJHQHUDWLRQZKLFKOHDGV
WR WKH FRPSOH[ QHXURORJLFDO SKHQRW\SH LQGXFHV VHYHUH
VHYHUH DEQRUPDOLWLHV LQ PDQ\ GLVWULFWV RI WKH
FHQWUDO DQG SHULSKHUDO QHUYRXV V\VWHP RI DIIHFWHG LQGLYLGXDOV $FFRUGLQJ ZLWK WKH IDFW
WKDW$7LVSULPDULO\DV\QGURPHRISURJUHVVLYHFHUHEHOODU
7LVSULPDULO\DV\QGURPHRISURJUHVVLYHFHUHEHOODUDWD[LDKLVWRORJLFDOVWXGLHVRIDXWRSV\
KLVWRORJLFDOVWXGLHVRIDXWRSV\
VDPSOHV IURP SDWLHQWV VKRZ DWURSK\ RI DOO FHUHEHOODU
FHUHEHOODU FRUWLFDO OD\HUV ZLWK D WKLQQLQJ RI
FHUHEHOODU KHPLVSKHULF IROLDH DQG FHUHEHOODU YHUPLV
YHUPLV DQG IUHTXHQW YDVFXODU FKDQJHV 7KHVH
GHJHQHUDWLYH PRGLILFDWLRQV DUH DFFRPSDQLHG E\ H[WHQVLYH DQG SURJUHVVLYH ORVV RI 3XUNLQMH
QHXURQV DQG WR D OHVVHU H[WHQW RI JUDQXOH QHXURQV
QHXURQV ,QWHUHVWLQJO\ ZKLOH
ZK
VRPHWLPHV
GLVSODFHGLQWKHPROHFXODUOD\HURIWKHFHUHEHOOXP
GLVSODFHGLQWKHPROHFXODUOD\HURIWKHFHUHEHOOXPWKHPDMRULW\RIWKHUHPDLQLQJ
WKHUHPDLQLQJDWURSKLF
3XUNLQMHFHOOVDUHIRXQGLQWKHVLQJOHFHOOOD\HURI3XUNLQMH
WKHVLQJOHFHOOOD\HURI3XUNLQMHFHOOV)LJXUHLQGLFDWLQJWKDWWKH\
DUHDEOHWRPLJUDWHFRUUHFWO\LQ
FRUUHFWO\LQWKHGHYHORSLQJEUDLQDQGLQGHHGWKHEDVNHWFHOOVW
WKHGHYHORSLQJEUDLQDQGLQGHHGWKHEDVNHWFHOOVWKDWRQO\IRUP
DURXQGWKHPDUHLQWKHLUSURSHUSODFH
DURXQGWKHPDUHLQWKHLUSURSHUSODFH7KLVVXJJHVWVWKDW3XUNLQMHFHOOV
3XUNLQMHFHOOVDUHDIIHFWHGDWDQ
HDUO\VWDJHEXWRQO\DIWHUWKH\DUHIXOO\GHYHORSHG
O\DIWHUWKH\DUHIXOO\GHYHORSHG7KHVHREVHUYDWLRQVDUHLQDFFRUGDQFH
DUHLQDFFRUGDQFHZLWK
PDJQHWLF UHVRQDQFH LPDJLQJ VWXGLHV RI WKH FHUHEHOOXP
FHUHEHOOXP ZKLFK DUH JHQHUDOO\ ZLWKRXW
SDWKRORJLFDO ILQGLQJV GXULQJ HDUO\ FKLOGKRRG EXW VKRZ VLJQLILFDQW WKLQQLQJ RI WKH PROHFXODU
OD\HUDQGFHUHEHOODUDWURSK\E\WKHHQGRIWKHILUV
OD\HUDQGFHUHEHOODUDWURSK\E\WKHHQGRIWKHILUVWGHFDGHRIOLIH
)LJ&RPSDULVRQRI3XUNLQMHFHOOVIURPDQ$
3XUNLQMHFHOOVIURPDQ$7SDWLHQW$DQGDQDJHPDWFKHGFRQWUROVXEMHFW%GHWHFWHGE\
PDWFKHGFRQWUROVXEMHFW%GHWHFWHGE\
DQWLFDOELQGLQ
FDOELQGLQLPPXQRVWDLQLQJRIFHUHEHOODUVHFWLRQV
,QWURGXFWLRQ
$V FKLOGUHQ JURZ LQWR DGXOWKRRG GHJHQHUDWLYH FKDQJHV EHFRPH PRUH H[WHQVLYH
$XWRSV\VWXGLHVUHSRUWQHXURQDOORVVLQWKHVWULDWXPDQGVXEVWDQWLDQLJUDQHXURGHJHQHUDWLRQRI
FUDQLDO QHUYHV DWURSK\ RI WKH SRVWHULRU FROXPQV DQG RI WKH DQWHULRU KRUQ ORVV RI P\HOLQDWHG
ILEHUV LQ FRUWLFRVSLQDO DQG VSLQRFHUHEHOODU WUDFWV GHFUHDVHG QXPEHU RI P\HOLQDWHG ILEHUV DQG
VHYHUH DWURSKLF FKDQJHV LQ SHULSKHUDO QHUYHV OHDGLQJ WR VHYHUH VHQVRULPRWRU D[RQDO
GHJHQHUDWLRQSHULSKHUDOQHXURSDWK\DQGFRQVHFXWLYHVSLQDOPXVFXODUDWURSK\
1RQQHXURORJLFDOFOLQLFDOIHDWXUHV
2FXORFXWDQHRXVWHODQJLHFWDVLDZKLFKFRQVLVWVLQFKURQLFGLODWLRQRIFDSLOODULHVOHDGLQJWR
WKH GHYHORSPHQW RI GDUN UHG VSRWV HVSHFLDOO\ LQ WKH EXOEDU FRQMXQFWLYDH DQG LQ WKH EXWWHUIO\
DUHDRIWKHIDFHLVWKHVHFRQGFDUGLQDOIHDWXUHRI$7DQGGHYHORSVVHYHUDO\HDUVDIWHUWKHRQVHW
RIWKHQHXURORJLFDOV\PSWRPV)RUWXQDWHO\LWLVQRWDVVRFLDWHGWRIXUWKHUFRPSOLFDWLRQVDQG
LVWKXVPHGLFDOO\LUUHOHYDQW$QRWKHUWUDJLFPDQLIHVWDWLRQRI$7LVWKHJUHDWO\LQFUHDVHGULVNRI
GHYHORSLQJ FDQFHU SULPDULO\ OHXNHPLDV DQG O\PSKRPDV ZLWK D OLIHWLPH IUHTXHQF\ WKDW
DSSURDFKHV,WLVQRWHZRUWK\WKDWDOVRFDUULHUVRIGLVHDVHFDXVLQJPXWDWLRQVKDYHDQ
LQFUHDVHG ULVN RI FRQWUDFWLQJ EUHDVW WXPRUV VXJJHVWLQJ GRPLQDQFH RI WKH GHIHFWLYH JHQH ,PSRUWDQWO\PLVVLQJWKHGLDJQRVLVRI$7PD\OHDGWRDGPLQLVWUDWLRQRIFKHPRWKHUDS\RUHYHQ
UDGLRWKHUDS\ZLWKVHYHUHDQGIDWDOFOLQLFDOFRQVHTXHQFHVLQGHHGSDWLHQWVDUHK\SHUVHQVLWLYHWR
JHQRWR[LF DJHQWV DQG LRQL]LQJ UDGLDWLRQ ,5 ZKLFK UHQGHUV WKHVH WKHUDSHXWLF DSSURDFKHV
SDUWLFXODUO\KD]DUGRXVDQGFRPSOLFDWHVWKHPDQDJHPHQWRIDQ\PDOLJQDQF\3DWLHQWVDOVR
GLVSOD\YDULDEOHFHOOXODUDQGKXPRUDOLPPXQRGHILFLHQF\ZLWKDEVHQWRUORZOHYHOVRI,J$DQG
,J( DEQRUPDOLWLHV LQ ,J* DQG ,J0 DQG DQ DWURSKLF RU DEVHQW WK\PXV &RQVHTXHQWO\
DIIHFWHG LQGLYLGXDOV DUH SDUWLFXODUO\ YXOQHUDEOH WR LQIHFWLRQV DQG VXIIHU IURP UHFXUUHQW
EURQFKRSXOPRQDU\GLVHDVHV,QDGGLWLRQWKHGLVRUGHULVFKDUDFWHUL]HGE\ULVLQJOHYHOVRIDOSKD
IHWRSURWHLQFKURPRVRPDOLQVWDELOLW\LQYROYLQJSULPDULO\WKHORFLRILPPXQRJOREXOLQDQG7
,QWURGXFWLRQ
FHOOUHFHSWRUJHQHVRQFKURPRVRPHVDQGJURZWKUHWDUGDWLRQLQVXOLQUHVLVWDQFH
ZKLFKFDXVHVVXVFHSWLELOLW\WRW\SH,,GLDEHWHVDQGJRQDGDODWURSK\([SHFWHGOLIHVSDQ
LVEHWZHHQDQG\HDUVRIDJHDQGGHDWKLVFDXVHGSUHGRPLQDQWO\E\FDQFHUFKURQLFOXQJ
GLVHDVH UHVXOWLQJ IURP LPPXQRGHILFLHQF\ DQG GHIHFWV LQ FKHZLQJ DQG VZDOORZLQJ GXH WR
SURJUHVVLYHQHXURORJLFDOLPSDLUPHQW
'LDJQRVLVDQGWUHDWPHQW
,W LV LPSRUWDQW WR DFFXUDWHO\ GLDJQRVH $7 DV UDSLGO\ DV SRVVLEOH LQ RUGHU WR DOORZ
DSSURSULDWHFDUHDQGJHQHWLFFRXQVHOLQJHVSHFLDOO\JLYHQWKHLQFUHDVHGULVNRIPDOLJQDQFLHVDQG
WKH KLJKHU ULVN RI VLGH HIIHFWV RI VXEVHTXHQW FDQFHU WUHDWPHQW QRW RQO\ LQ SDWLHQWV EXW DOVR LQ
KHWHUR]\JRXV UHODWLYHV 7KH SUHVHQFH RI $7 LV XVXDOO\ UHFRJQL]HG E\ WKH FRPELQDWLRQ RI
QHXURORJLFDO IHDWXUHV ZLWK WHODQJLHFWDVLD DQG LQIHFWLRQV EXW XQIRUWXQDWHO\ LQ YHU\ \RXQJ
FKLOGUHQWKHSKHQRW\SHLVRIWHQLQFRPSOHWHDQGWHODQJLHFWDVLDRQO\DSSHDUVDURXQGVL[\HDUVRI
DJHPDNLQJHDUO\GLIIHUHQWLDOGLDJQRVLVGLIILFXOW0RUHRYHULWPXVWEHQRWHGWKDWWKH$7
SKHQRW\SH LQFOXGHV D ZLGH DQG FRQWLQXRXV VSHFWUXP RI PDQLIHVWDWLRQV WKDW VSDQ IURP WKH
SUHYLRXVO\ GHVFULEHG VHYHUH FODVVLFDOIRUP WR D OHVV VHYHUH DGXOWRQVHW GLVRUGHU UHIHUUHG WR DV
YDULDQW $7 ZKLFK LV FKDUDFWHUL]HG E\ PLOGHU V\PSWRPV H[WUDS\UDPLGDO VLJQV DQG H[WHQGHG
OLIHVSDQZLWKFRQVHTXHQWIXUWKHUGLIILFXOW\LQUHFRJQL]LQJWKHGLVHDVH
/DERUDWRU\ LQYHVWLJDWLRQV LQ SDWLHQWV ZKR DUH VXVSHFWHG WR VXIIHU IURP $7 FDQ
DFFHOHUDWH GLDJQRVLV 7KHVH LQFOXGH PHDVXUHPHQW RI $)3 OHYHOV F\WRJHQHWLF DQDO\VLV RI
FKURPRVRPDOLQVWDELOLWLHVZLWKVSHFLDODWWHQWLRQWRWUDQVORFDWLRQVLQYROYLQJFKURPRVRPHVDQG
HYDOXDWLRQRIVHUXPLPPXQRJOREXOLQOHYHOVDVVHVVPHQWRIWKHOHYHOVRI$70SURWHLQRURI
$70 SURWHLQ NLQDVH DFWLYLW\ FRORQ\ VXUYLYDO DVVD\ DIWHU H[SRVXUH WR DŽUDGLDWLRQ DQG PRVW
UHFHQWO\PXWDWLRQGHWHFWLRQ
,QWURGXFWLRQ
6DGO\ WKHUH LV FXUUHQWO\ QR WUHDWPHQW DYDLODEOH WR FXUH SUHYHQW RU SRVWSRQH WKH
SURJUHVVLRQ RI $7 3DWLHQWV DUH RQO\ RIIHUHG V\PSWRPDWLF DQG VXSSRUWLYH WKHUDSLHV VXFK DV
SK\VLRWKHUDS\ ORJRSHGLFV DQWLELRWLF WUHDWPHQW JDVWURVWRP\ DQG UHVSLUDWRU\ VXSSRUW ZKHQ
QHHGHG
$70JHQHDQGSURWHLQ
7KHJHQHUHVSRQVLEOHIRU$7ZDVLQLWLDOO\ORFDOL]HGWRFKURPRVRPDOUHJLRQT
LQE\*DWWLHWDODQGLQZDVGHILQLWHO\LGHQWLILHGE\6DYLWVN\HWDODVWKH$70
JHQH ZKLFK H[WHQGV RYHU NE RI JHQRPLF '1$ DQG FRQWDLQV H[RQV HQFRGLQJ D NE
WUDQVFULSW6LQFHWKHQPRUHWKDQXQLTXHJHQHPXWDWLRQVUHSRUWHGWKURXJKRXWWKHZKROH
VHTXHQFH RI $70 KDYH EHHQ DVVRFLDWHG WR $7 7KHVH PDLQO\ FRPSULVH WUXQFDWLQJ
PXWDWLRQV ZKLFK KDYH GHWULPHQWDO HIIHFWV RQ WKH SURWHLQ DQG DUH DVVRFLDWHG ZLWK WKH PRVW
VHYHUH HDUO\RQVHW IRUP RI WKH GLVRUGHU ZKLOH D ORZHU SHUFHQWDJH RI PXWDWLRQV DOORZLQJ IRU
UHVLGXDO $70 DFWLYLW\ VXFK DV PLVVHQVH PXWDWLRQV RU LQIUDPH GHOHWLRQV RU LQVHUWLRQV FDXVH
PLOGHUIRUPVRIWKHGLVHDVH
7KH $70 JHQH HQFRGHV D ODUJH VHULQHWKUHRQLQH SURWHLQ NLQDVH PDGH XS RI UHVLGXHVDQGZLWKDPROHFXODUZHLJKWRIN'DZKLFKLVFRQVWLWXWLYHO\H[SUHVVHGLQDOOWLVVXHV
DQG LV SUHGRPLQDQWO\ ORFDWHG LQ WKH QXFOHXV DOWKRXJK D VPDOO DPRXQW LV DOVR SUHVHQW LQ WKH
F\WRSODVP RI PDQ\ FHOO W\SHV ,Q DFFRXQW RI LWV FDUER[\WHUPLQXV ZKLFK FRQWDLQV D
FDWDO\WLFDOO\ DFWLYH SKRVSKDWLG\OLQRVLWRONLQDVH 3,. GRPDLQ WKH $70 SURWHLQ KDV EHHQ
FODVVLILHG DV PHPEHU RI WKH 3,.UHODWHG SURWHLQ NLQDVH 3,.. IDPLO\ ZKLFK DOVR LQFOXGHV
$70DQG5$' UHODWHG$75'1$GHSHQGHQWSURWHLQNLQDVH'1$3.PDPPDOLDQWDUJHW
RI UDSDP\FLQ P725 VXSSUHVVRU RI PXWDJHQHVLV LQ JHQLWDOLD 60* DQG
WUDQVIRUPDWLRQWUDQVFULSWLRQGRPDLQDVVRFLDWHGSURWHLQ755$3$70SKRVSKRU\ODWHVLWV
VXEVWUDWHVRQDVHULQHRUWKUHRQLQHWKDWLVIROORZHGE\JOXWDPLQHQDPHO\DQ64RU74>67@4
,QWURGXFWLRQ
PRWLI,QDGGLWLRQWRWKHNLQDVHGRPDLQWKUHHUHJXODWRU\GRPDLQVDUHDOVRFRQVHUYHGLQWKH
3,..IDPLO\DQGDUHWKXVIRXQGLQ$70D)5$3$7075$3)$7GRPDLQD)$7&WHUPLQDO
)$7&GRPDLQDQGD3,..UHJXODWRU\GRPDLQ35'ZKLFKLVWKH1WHUPLQDOSRUWLRQRIWKH
)$7& GRPDLQ 2WKHU SURWHLQ UHJLRQV ZKLFK KDYH EHHQ FKDUDFWHUL]HG LQFOXGH VLWHV RI
LQWHUDFWLRQZLWKRWKHUSURWHLQVVXFKDVWKHVXEVWUDWHELQGLQJVLWH6%6QHDUWKH1WHUPLQXVRI
WKHSURWHLQDQGSRVWWUDQVODWLRQDOPRGLILFDWLRQVLWHVVXFKDV6HU6HU6HU6HU
DQG /\V )LJXUH EXW XQIRUWXQDWHO\ WKH VWUXFWXUH RI WKH SURWHLQ KDV QRW \HW EHHQ
HQWLUHO\GHILQHGFRPSOLFDWLQJWKHIXOOFRPSUHKHQVLRQRI$70IXQFWLRQ
)LJ6FKHPDWLFUHSUHVHQWDWLRQRI$70
)XQFWLRQV RI $70 DQG SRVVLEOH FDXVHV RI QHXURGHJHQHUDWLRQ
LQDWD[LDWHODQJLHFWDVLD
,WKDVORQJEHHQNQRZQWKDW$70SOD\VDFHQWUDOUROHLQWKHFHOOXODUUHVSRQVHWR'1$
GDPDJHDQGLQFHOOF\FOHFKHFNSRLQWFRQWURODQGLWLVQRZHYLGHQWWKDWLWVDFWLYLW\LVHVVHQWLDOLQ
DQ LQFUHDVLQJ QXPEHU RI RWKHU FHOOXODU SDWKZD\V LQYROYHG LQ WKH PDLQWHQDQFH RI FHOOXODU
KRPHRVWDVLV $FFRUGLQJO\ WKH PXOWLSOH IXQFWLRQV RI $70 DFFRXQW IRU WKH FRPSOH[
SKHQRW\SLFPDQLIHVWDWLRQVGHULYLQJIURPLQDFWLYDWLQJPXWDWLRQVLQWKHJHQH1RQHWKHOHVVRXU
FXUUHQW NQRZOHGJH RQ $70 DFWLYLW\ FDQQRW \HW IXOO\ H[SODLQ WKH WUDJLF QHXURORJLFDO GHFD\
ZKLFK DIIHFWV $7 SDWLHQWV 7KH UHOHQWOHVV QHXURGHJHQHUDWLRQ FDXVHG E\ $70 GHILFLHQF\
SUHVXPDEO\ UHVXOWV IURP WKH FRQFXUUHQFH RI GLIIHUHQW SDWKRJHQHWLF PHFKDQLVPV ZKLFK DULVH
,QWURGXFWLRQ
IROORZLQJ WKH GLVUXSWLRQ RI WKH PXOWLSOH SDWKZD\V LQ ZKLFK WKH VHULQHWKUHRQLQH NLQDVH LV
LQYROYHG 7KHUHIRUH LW LV H[WUHPHO\ LPSRUWDQW WKDW $70 IXQFWLRQV EH GHILQHGLQ GHWDLOLQ
RUGHUWREHDEOHWRWKRURXJKO\FKDUDFWHUL]HWKHSDWKRJHQHVLVRIWKHGLVRUGHUDQGFRQVHTXHQWO\
GHOLQHDWHSRWHQWLDOWKHUDSHXWLFVWUDWHJLHV
5ROHRI$70LQWKH'1$GDPDJHUHVSRQVHDQGLQFHOOF\FOHFRQWURO
$V FHOOV DUH FRQVWDQWO\ H[SRVHG WR D PXOWLWXGH RI VWUHVVHV LQFOXGLQJ PDQ\ RI ZKLFK
WKUHDWHQJHQRPHVWDELOLW\VHYHUDOGLIIHUHQWVLJQDOWUDQVGXFWLRQSDWKZD\VDUHUHDGLO\DFWLYDWHG
ZKHQQHFHVVDU\LQRUGHUWRSURWHFWFHOOXODULQWHJULW\,QSDUWLFXODU'1$GRXEOHVWUDQGEUHDNV
'6%V UHSUHVHQW WKH PRVW GDQJHURXV IRUP RI '1$ GDPDJH DQG FDQ OHDG WR FHOO GHDWK
GHOHWHULRXV FKURPRVRPDO LQVWDELOLW\ DQG PDOLJQDQW WUDQVIRUPDWLRQ 7KHVH OHVLRQV DUH
JHQHUDWHGIROORZLQJH[SRVXUHWRH[RJHQRXVDJHQWVVXFKDV,5DQGUDGLRPLPHWLFFKHPLFDOVEXW
FDQ DOVR EH FDXVHG E\ HQGRJHQRXV SURGXFWV OLNH R[\JHQ UDGLFDOV RU RFFXU GXULQJ QRUPDO
SK\VLRORJLFDO FHOO SURFHVVHV VXFK DV '1$ UHSOLFDWLRQ PHLRWLF UHFRPELQDWLRQ DQG 9'-
UHFRPELQDWLRQ7KHUHSDLURI'6%VFDQRFFXUHLWKHUE\QRQKRPRORJRXVHQGMRLQLQJ1+(-RU
E\ KRPRORJRXV UHFRPELQDWLRQ +5 7KH ILUVW LV WKH PDMRU '1$ UHSDLU SURFHVV LQ
PDPPDOLDQFHOOVEHLQJDFWLYHWKURXJKRXWWKHFHOOF\FOHDQGFRQVLVWVLQWKHGLUHFWOLJDWLRQRIWKH
WZR'1$HQGVWKHODWWHULQWHUYHQHVLQWKHODWH6SKDVHRU*SKDVHRIWKHFHOOF\FOHDQGUHSDLUV
WKHGDPDJHG'1$VWUDQGZLWKWKHDLGRIWKHLQWDFWVLVWHUFKURPDWLG1RWDEO\QRWDOO'6%VDUH
UHSDLUHGDWWKHVDPHVSHHGDSSUR[LPDWHO\DUHUHSDLUHGZLWKIDVWNLQHWLFVZKLOH²DUH
UHSDLUHGPRUHVORZO\
:KHQ '1$ FDQQRW EH UHDGLO\ UHSDLUHG DV ZKHQ WKH OHVLRQ LV VLWXDWHG LQ D WLJKWO\
FRPSDFWHGKHWHURFKURPDWLFDUHDRUZKHQKLJKOHYHOVRIGDPDJHRFFXUDQ$70GHSHQGHQW'6%
UHVSRQVHEHFRPHVQHFHVVDU\,QGHHGWKH$70SURWHLQNLQDVHSDUWLFLSDWHVLQWKHVORZHU
UHSDLURI'1$OHVLRQVDQGPRVWLPSRUWDQWO\SOD\VDFUXFLDOUROHLQWKHLQLWLDWLRQRIWKHFHOOXODU
,QWURGXFWLRQ
UHVSRQVH WR '6%V EHLQJ DEOH WR FUHDWH D FRPSOH[ \HW FRRUGLQDWHG VLJQDOLQJ QHWZRUN WKDW
WULJJHUV DUUHVW RI FHOO F\FOH SURJUHVVLRQ WKURXJK FKHFNSRLQW DFWLYDWLRQ 7KLV LV FUXFLDO IRU
DOORZLQJ WKH FHOO WR SURFHVV WKH '1$ GDPDJH DQG ZKHQ WKLV LV QRW SRVVLEOH IRU LQGXFLQJ
VHQHVFHQFHRUDSRSWRWLFGHDWK
,Q XQGDPDJHG FHOOV $70 LV IRXQGLQ WKHQXFOHXV LQ WKHIRUP RI LQDFWLYH KRPRGLPHUV
)ROORZLQJ WKH LQGXFWLRQ RI '6%V $70 LV DFWLYDWHG WKURXJK LQWHUPROHFXODU
DXWRSKRVSKRU\ODWLRQ DW 6HU ZKLFK FDXVHV FRQIRUPDWLRQDO FKDQJHV OHDGLQJ WR
PRQRPHUL]DWLRQ DQG LV UHFUXLWHG WR WKH OHVLRQV E\ WKH SULPDU\ VHQVRU FRPSOH[ 05(
5$'1%6051ZKLFKDVVHPEOHVDWWKHGDPDJHGVLWHVXQZLQGVWKH'1$HQGVDQGELQGV
DQG UHWDLQV$70 WKURXJK WKH &WHUPLQDO GRPDLQ RI 1%6$OWKRXJK LW LV FOHDU WKDW WKHVH
HYHQWV DUH HVVHQWLDO IRU WULJJHULQJ $70 DFWLYDWLRQ WKHLU SUHFLVH VHTXHQFH LV VWLOO GHEDWHG
0RUHRYHU RWKHU SURFHVVHV LQFOXGLQJ FKURPDWLQ UHPRGHOLQJ DQG SRVWWUDQVODWLRQDO
PRGLILFDWLRQV VXFK DV DXWRSKRVSKRU\ODWLRQ RQ WKH WKUHH DGGLWLRQDO VLWHV 6HU 6HU DQG
6HU
7,3 PHGLDWHG
DFHW\ODWLRQ
DW
/\V
DQG
33GHSHQGHQW
GHSKRVSKRU\ODWLRQ RI $70 DOO FRRSHUDWH LQ WKH LQLWLDWLRQ RI LWV NLQDVH DFWLYLW\ 2QFH
DFWLYDWHG$70IXQFWLRQVDVWKHDSLFDONLQDVHLQWKHFRPSOH[LQWUDFHOOXODUVLJQDOLQJFDVFDGHWKDW
FKDUDFWHUL]HVWKH'1$GDPDJHUHVSRQVHWR'6%V
$WWKHGDPDJHGVLWHV$70SKRVSKRU\ODWHVWKH051FRPSOH[DQGKLVWRQH+$;DW
6HU SURGXFLQJ DŽ+$; LQGXFLQJ FKURPDWLQ PRGLILFDWLRQV WKDW DOORZ WKH UHFUXLWPHQW
DQGSKRVSKRU\ODWLRQRIWKHPHGLDWRUSURWHLQVPHGLDWRURI'1$GDPDJHFKHFNSRLQWSURWHLQ
0'&SELQGLQJSURWHLQ%3DQGEUHDVWFDQFHUW\SHVXVFHSWLELOLW\SURWHLQ
%5&$ZKLFKDVVHPEOHRQWKH'1$FUHDWLQJDXWRDOLPHQWHGIRFLWKDWIRUPDVFDIIROGIRU
WKH UHFUXLWPHQW RI UHSDLU SURWHLQV VXVWDLQ DQG DPSOLI\ GDPDJH VLJQDOLQJ DQG PDLQWDLQ WKH
DFWLYDWLRQRI$70LQDSRVLWLYHIHHGEDFNORRS,QWULJXLQJO\WKHDVVHPEO\RIIRFLDWWKH
VLWH RI '1$ '6%V FDQ DOVR RFFXU LQ WKH DEVHQFH RI $70 DFWLYLW\ WKDQNV WR WKH UHGXQGDQW
,QWURGXFWLRQ
SKRVSKRU\ODWLRQ RI +$; RSHUDWHG E\ '1$3. DQRWKHU 3,.. LQYROYHG LQ '6% SURFHVVLQJ
1HYHUWKHOHVV EHLQJ D ODUJH DPRXQW RI WKH SURWHLQV LQ HDFK IRFXV DQ $70 VXEVWUDWH WKH
IXQFWLRQDO VLJQLILFDQFH RI WKHLU $70GHSHQGHQW SKRVSKRU\ODWLRQ LV ORVW ZKHQ WKH NLQDVH LV
PLVVLQJRULQDFWLYH
)XUWKHUPRUH$70GLUHFWO\FRRSHUDWHVLQWKHUHSDLURI'6%VLQKHWHURFURPDWLF'1$E\
SKRVSKRU\ODWLQJ WKH WUDQVFULSWLRQDO FRUHSUHVVRU .USSHODVVRFLDWHG ER[DVVRFLDWHG SURWHLQ
.$3DWWKHGDPDJHGVLWHFRQVHTXHQWO\HDVLQJWKHHQWU\RIWKH'1$UHSDLUPDFKLQHU\
DQG E\ K\SHUSKRVSKRU\ODWLQJ WKH HQGRQXFOHDVH $UWHPLV ZKLFK LV UHVSRQVLEOH IRU WKH
SURFHVVLQJ RI WKH GDPDJHG HQGV ,PSRUWDQWO\ WKH$70GHSHQGHQW'6%UHSDLU DOVR UHOLHV
RQWKH051FRPSOH[DŽ+$;0'&DQG%3
$ZD\ IURP WKH GDPDJHG VLWH $70 SKRVSKRU\ODWHV WKH PDLQ WUDQVGXFHU RI WKH $70
VLJQDOLQJSDWKZD\WKHFKHFNSRLQWNLQDVH&+.LQGXFLQJDVLJQDOLQJFDVFDGHWKDWOHDGVWRFHOO
F\FOHDUUHVWWKURXJK'1$GDPDJHFKHFNSRLQWDFWLYDWLRQ,PSRUWDQWO\&+.LVRQO\RQHRI
WKHQXPHURXVGLIIXVLEOHVXEVWUDWHVRIWKHDSLFDONLQDVH,QGHHG$70LVDEOHWRUHDFKWKHVDPH
HQGSRLQWV E\ WULJJHULQJ GLIIHUHQW SDWKZD\V WKHUHIRUH JXDUDQWHHLQJ D VWULFW UHJXODWLRQ RI HDFK
SURFHVV +HQFH WKH *6 FKHFNSRLQW LV DOVR DFWLYDWHG WKURXJK GLUHFW SKRVSKRU\ODWLRQ
VWDELOL]DWLRQDQGDFFXPXODWLRQRIWKHSWUDQVFULSWLRQIDFWRUWKHLQWUD6SKDVHFKHFNSRLQWFDQ
DOVREHHOLFLWHGWKURXJKSKRVSKRU\ODWLRQRI60&RU%5&$DQGWKH*0FKHFNSRLQWFDQEH
WULJJHUHGWKURXJKSKRVSKRU\ODWLRQRI%5&$
7KLV$70GHSHQGHQW'1$GDPDJHVLJQDOLQJSDWKZD\JLYHVULVHWRDFHOOXODUUHVSRQVH
WR'6%VWKDWJRHVEH\RQGUHSDLUDFWLYDWLQJERWKFHOOVXUYLYDODQGFHOOGHDWKSDWKZD\VWKURXJK
WKHPRGXODWLRQRIJHQHH[SUHVVLRQDQGRISURWHLQV\QWKHVLVDQGGHJUDGDWLRQ7KHRXWFRPHLVD
YHU\PHWLFXORXV'1$GDPDJHUHVSRQVHUHVXOWLQJHLWKHULQGDPDJHUHPRYDOFHOOVHQHVFHQFHRU
DSRSWRVLV)LJXUH
,QWURGXFWLRQ
)LJ6FKHPDWLFUHSUHVHQWDWLRQRIWKH$70GHSHQGHQW'1$GDPDJHUHVSRQVH
7KH DEVHQFH RI $70 DFWLYLW\ GXULQJ QHXURQDO GHYHORSPHQW DW YHU\ HDUO\ VWDJHV PD\
FDXVHWKHHPHUJHQFHRIJHQRPLFGDPDJHWKDWEHFRPHVIL[HGLQWKHQHUYRXVV\VWHPDQGDWODWHU
VWDJHV LW PD\ DOORZ WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQV WR IXUWKHU DFFXPXODWH GDPDJHG '1$
ZLWKRXW DQ HIILFLHQW UHSDLU PHFKDQLVP $70GHILFLHQW GDPDJHG QHXURQV PD\ HQWHU
DSRSWRVLV RU PD\ EH UHWDLQHG LQ WKH QHUYRXV V\VWHP DQG DV D FRQVHTXHQFH GHJHQHUDWH GXH WR
DFFXPXODWLRQ RI WR[LF SURWHLQV RU G\VIXQFWLRQDOJHQHV 0RUHRYHU ORVV RI $70GHSHQGHQW FHOO
F\FOHFRQWUROFRXOGDOVRFDXVHXQVFKHGXOHGUHHQWU\RIGLIIHUHQWLDWHGSRVWPLWRWLFQHXURQVLQWR
WKHFHOOF\FOHZKLFKUHVXOWVLQFHOOGHDWK
,QWURGXFWLRQ
$GGLWLRQDOUROHVRI$70LQWKHF\WRSODVPDQGLQWKHQXFOHXV
$OWKRXJKWKHPRVWVWULNLQJIXQFWLRQRI$70LVFHUWDLQO\LWVFHQWUDOUROHLQWKHUHVSRQVH
WR'6%VZKLFKLVFDUULHGRXWLQWKHQXFOHXVWKHVHULQHWKUHRQLQHSURWHLQNLQDVHLVDOVRSUHVHQW
LQWKHF\WRSODVPDQGSOD\VDQLPSRUWDQWSDUWLQRWKHUFHOOXODUSURFHVVHVLQWKHXQGDPDJHGFHOO
SDUWLFLSDWLQJ LQ VLJQDOLQJ SDWKZD\V WKDW PDLQWDLQ FHOOXODU KRPHRVWDVLV DQG UHVSRQG WR
SK\VLRORJLFDO VWLPXOL ,QGHHG $70 FDQ EH DFWLYDWHG LQ D '6% DQG 051LQGHSHQGHQW
PDQQHUDQGKDVEHHQIRXQGWRSKRVSKRU\ODWHRYHUDKXQGUHGVXEVWUDWHVPDQ\RIZKLFKGLIIHU
IURPWKRVHLQYROYHGLQWKH'1$GDPDJHUHVSRQVH
2QH RI WKH PRVW LPSRUWDQW F\WRSODVPLF IXQFWLRQV RI $70 UHJDUGV WKH VHQVLQJ DQG
VLJQDOLQJ RI UHDFWLYH R[\JHQ VSHFLHV 526 526 DUH FKHPLFDOO\ UHDFWLYH PROHFXOHV ZKLFK
DUHIRUPHGDVE\SURGXFWRIWKHSK\VLRORJLFDODFWLYLW\RIPDQ\HQGRJHQRXVHQ]\PHVDQGRIWKH
UHVSLUDWRU\ FKDLQ LQ WKH PLWRFKRQGULD DQG KDYH LPSRUWDQW UROHV LQ FHOO VLJQDOLQJ DQG
KRPHRVWDVLV :KHQ WKHUH LV DQ RYHUSURGXFWLRQ RI 526 IRU H[DPSOH IROORZLQJ
PLWRFKRQGULDO G\VIXQFWLRQ RU WKH H[SRVXUH WR H[RJHQRXV IDFWRUV VXFK DV 89 OLJKW DQG ,5 RU
ZKHQ WKH HIIHFWLYHQHVV RI FHOOXODU DQWLR[LGDQW GHIHQVHV GHFUHDVHV LQWUDFHOOXODU 526 OHYHOV FDQ
LQFUHDVHGUDPDWLFDOO\OHDGLQJWRR[LGDWLYHVWUHVVZKLFKSURGXFHVVLJQLILFDQWOHVLRQVWRFHOOXODU
VWUXFWXUHVDQGFDXVHV'1$GDPDJHLQWKHIRUPRIR[LGL]HG'1$EDVHVDSXULQLFDS\ULPLGLQLF
VLWHVVLQJOHVWUDQGEUHDNV66%VDQG'6%V,WKDVORQJEHHQNQRZQWKDW$70GHILFLHQWFHOOV
DUHPRUHVXVFHSWLEOHWRR[LGDWLYHGDPDJHSURGXFHGE\ERWKH[RJHQRXVDQGHQGRJHQRXVIDFWRUV
DQGWKDWWKH\GLVSOD\HOHYDWHGOHYHOVRIR[LGDWLYHGDPDJHUHVSRQVLYHSDWKZD\V
DQGUHGXFHGOHYHOVRIDQWLR[LGDQWVVXJJHVWLQJWKDW$70LVDVHQVRUIRUUHGR[KRPHRVWDVLV
DQG WKDW $7 LV D GLVRUGHU UHODWHG WR R[LGDWLYH VWUHVV ,QGHHG LWKDV UHFHQWO\ EHHQ VKRZQ WKDW
$70FDQEHGLUHFWO\DFWLYDWHGWKURXJKDXWRSKRVSKRU\ODWLRQLQUHVSRQVHWRWKHR[LGL]LQJDJHQW
K\GURJHQSHUR[LGH+2LQWKHDEVHQFHRI'1$GDPDJHDQGLQDQ051LQGHSHQGHQWIDVKLRQ
7KLVLVSRVVLEOHWKDQNVWRWKHIRUPDWLRQRIDQLQWHUPROHFXODUGLVXOILGHERQGDW&\VLQ
,QWURGXFWLRQ
WKH &WHUPLQXV RI WKH SURWHLQ ZKLFK JLYHV ULVH WR DQ DFWLYH FRYDOHQW GLPHU $70 DSSHDUV WR
UHJXODWH WKH DQWLR[LGDQW FDSDFLW\ RI FHOOV DOVR E\ LQGLUHFWO\ SDUWLFLSDWLQJ LQ WKH FRUUHFW
UHJHQHUDWLRQ RI WKH DQWLR[LGDQW JOXWDWKLRQH ,QGHHG $70 LV DEOH WR HQKDQFH WKH SHQWRVH
SKRVSKDWH SDWKZD\ DQG WKHUHIRUH WKH V\QWKHVLV RI WKH DQWLR[LGDQW FRIDFWRU 1$'3+ ZKLFK LV
UHTXLUHG E\ WKH JOXWDWKLRQH UHGXFWDVH HQ]\PH IRU WKH UHGXFWLRQ RI WKH R[LGL]HG IRUP RI
JOXWDWKLRQHEDFNWRLWVUHGXFHGDFWLYHYHUVLRQ$VQHXURQVKDYHDQLQWHQVLYHPHWDEROLFDFWLYLW\
WKH\ SK\VLRORJLFDOO\ DFFXPXODWH HOHYDWHG OHYHOV RI 526 DQG VLQFH WKH DEVHQFH RI $70
FDXVHVWKHORVVRIUHGR[EDODQFH$7QHXURQVPD\EHK\SHUVHQVLWLYHWRHQGRJHQRXVR[LGDWLYH
VWUHVV DQG H[RJHQRXV R[LGDWLYH VWUHVVLQGXFLQJ DJHQWV 7KLV PD\ FRQWULEXWH WR WKH
DFFXPXODWLRQRI'1$GDPDJHLQWKHVHFHOOVDQGQHXURGHJHQHUDWLRQLQ$7FRXOGWKHUHIRUHDOVR
UHVXOWIURPGHILFLHQW526KRPHRVWDVLV
0RUHRYHU $70 SOD\V DQ LPSRUWDQW UROH LQ PLWRFKRQGULDO KRPHRVWDVLV 0LWRFKRQGULDDUHRUJDQHOOHVZKLFKKDYHWKHLURZQ'1$DQGDUHWKHVLWHVRIFHOOXODUUHVSLUDWLRQ
ZKLFKLVHVVHQWLDOIRUWKHJHQHUDWLRQRI$73DQGWKHUHIRUHIRUVXSSO\LQJHQHUJ\WRWKHFHOO
,PSRUWDQWO\ WKLV SURFHVV FDQ OHDG WR WKH HQGRJHQRXV SURGXFWLRQ RI GDPDJLQJ 526 VSHFLHV
ZKLFK LV H[DFHUEDWHG E\ PLWRFKRQGULDO G\VIXQFWLRQ (DUO\ UHSRUWV KDYH GHVFULEHG LPSDLUHG
PLWRFKRQGULDO IXQFWLRQV LQ $70GHILFLHQW FHOOV DQG D PRUH UHFHQW LQYHVWLJDWLRQ KDV
VKRZQWKDWLQGHHGDIUDFWLRQRI$70ORFDOL]HVWRPLWRFKRQGULDLQWK\PRF\WHV,QSDUWLFXODU
DFFRUGLQJ WR WKLV VWXG\ $70 ORVV UHVXOWV LQ GHILFLHQFLHV LQ PLWRFKRQGULDO HOHFWURQ WUDQVSRUW
FKDLQDFWLYLW\LPSDLUHG$73OHYHOV HOHYDWHGPLWRFKRQGULDO 526 DQGLQFUHDVHG PLWRFKRQGULDO
QXPEHU ZKLFK LV GXH WR GHIHFWV LQ WKH VHOHFWLYH FOHDUDQFH RI DEQRUPDO PLWRFKRQGULD E\
DXWRSKDJ\ D SURFHVV DOVR NQRZQ DV PLWRSKDJ\ ,W KDV DOVR EHHQ VKRZQ WKDW $70 SOD\V DQ
LPSRUWDQWUROHLQPDLQWDLQLQJWKHLQWHJULW\RIPLWRFKRQGULDO'1$,QGHHGWKHODFNRI$70
FDXVHV GHFUHDVHG SURWHLQ OHYHOV RI '1$ OLJDVH ,,, ZKLFK LV WKH RQO\ NQRZQ '1$ OLJDVH
RSHUDWLQJ LQ PLWRFKRQGULD 7KLV LPSDLUV WKH EDVH H[FLVLRQ UHSDLU %(5 SDWKZD\ DQG WKXV WKH
,QWURGXFWLRQ
UHPRYDO RI R[LGDWLYH OHVLRQV IURP PLWRFKRQGULDO '1$ ZLWK FRQVHTXHQW SHUVLVWHQW
PLWRFKRQGULDO'1$GDPDJHDQGPLWRFKRQGULDOG\VIXQFWLRQ+HQFHWKHVHILQGLQJVLQGLFDWHWKDW
$70LVLQYROYHGLQWKHUHJXODWLRQRIPLWRFKRQGULDODFWLYLW\DQGPLWRFKRQGULDOG\VIXQFWLRQPD\
EHUHVSRQVLEOHIRUHOHYDWHG526SURGXFWLRQDQGR[LGDWLYHVWUHVVLQ$7QHXURQV
&\WRSODVPLF $70 DOVR SDUWLFLSDWHV LQ LQVXOLQ VLJQDOLQJ SDWKZD\V ,QVXOLQ LV D
FUXFLDO KRUPRQH LQ WKH UHJXODWLRQ RI PLWRJHQLF DQG PHWDEROLF DFWLRQV DV LW VWLPXODWHV FHOO
SUROLIHUDWLRQ ERRVWV SURWHLQ V\QWKHVLV LQKLELWV EUHDNGRZQ RI IDW DQG UHJXODWHV JOXFRVH
KRPHRVWDVLV E\ LQKLELWLQJ LWV KHSDWLF RXWSXW DQG SURPRWLQJ LWV XSWDNH HVSHFLDOO\ LQWR VWULDWHG
PXVFOH FHOOV DQG DGLSRF\WHV ,W KDV EHHQ VKRZQ WKDW $70 SDUWLFLSDWHV LQ WKH FHOOXODU
UHVSRQVH WR LQVXOLQ E\ SKRVSKRU\ODWLQJ WKH WUDQVODWLRQ UHSUHVVRU H,)( ELQGLQJ SURWHLQ (%3ZKLFKELQGVWRDQGLQKLELWVWKHHXNDU\RWLFWUDQVODWLRQLQLWLDWLRQIDFWRU(H,)(
)ROORZLQJ SKRVSKRU\ODWLRQ (%3 GLVVRFLDWHV IURP H,)( ZLWK FRQVHTXHQW HQKDQFHPHQW RI
SURWHLQ V\QWKHVLV $70 DOVR UHVSRQGV WR LQVXOLQ E\ SKRVSKRU\ODWLQJ DQG IXOO\ DFWLYDWLQJ WKH
SURWHLQNLQDVH$.7ZKLFKSDUWLFLSDWHVLQPXOWLSOHSK\VLRORJLFDOSURFHVVHVDQGLVLQYROYHG
LQ WKH HQKDQFHPHQW RI JOXFRVH XSWDNH E\ LQGXFLQJ WUDQVORFDWLRQ RI WKH JOXFRVH WUDQVSRUWHU
*/87WRWKHSODVPDPHPEUDQH7KHPHWDEROLFHIIHFWVRILQVXOLQDSSHDUWREHUHJXODWHGE\
$70DOVRLQDQR[LGDWLYHVWUHVVGHSHQGHQWPDQQHUWKURXJKWKHSKRVSKRU\ODWLRQRIS6HU
ZKLFKLQGXFHVFKDQJHVLQJHQHH[SUHVVLRQLQRUGHUWRPDLQWDLQLQVXOLQVHQVLWLYLW\DQGJOXFRVH
KRPHRVWDVLV,QGHHGORVVRIWKH$70SKRVSKRU\ODWLRQVLWHLQSLQGXFHVLQVXOLQUHVLVWDQFH
DQG JOXFRVH LQWROHUDQFH ZKLFK FDQ EH DPHOLRUDWHG E\ WUHDWPHQW ZLWK WKH DQWLR[LGDQW 1
DFHW\OF\VWHLQH 1$& 7KHUHIRUH WKH GLVUXSWLRQ RI DQ LQVXOLQGHSHQGHQW UHJXODWLRQ RI JHQH
H[SUHVVLRQRISURWHLQV\QWKHVLVDQGRIJOXFRVHXSWDNHLQWKHDEVHQFHRI$70FDQH[SODLQVRPH
RIWKHV\PSWRPVDVVRFLDWHGZLWK$7VXFKDVPHWDEROLFDEQRUPDOLWLHVJURZWKUHWDUGDWLRQDQG
LQVXOLQUHVLVWDQW GLDEHWHV ,Q DGGLWLRQ EHFDXVH LQVXOLQ VHUYHV DV D VXUYLYDO DQG GLIIHUHQWLDWLRQ
,QWURGXFWLRQ
IDFWRUIRUWKHGHYHORSLQJQHXURQDOWLVVXHWKHLPSDLUPHQWRILQVXOLQVLJQDOLQJLQ$70GHILFLHQW
QHXUDOFHOOVPD\FRQWULEXWHWRQHXURGHJHQHUDWLRQLQ$7
)XUWKHUPRUH$70SOD\VDQLPSRUWDQWUROHLQWKHPLWRWLFVSLQGOHFKHFNSRLQW7KLV
FKHFNSRLQWUHJXODWHVWKHWUDQVLWLRQIURPPHWDSKDVHWRDQDSKDVHDQGLVHVVHQWLDOIRUWKHFRUUHFW
SDUWLWLRQRIVLVWHUFKURPRVRPHVGXULQJPLWRVLVLQRUGHUWRDYRLGDQHXSORLG\LQGDXJKWHUFHOOV
5HFHQW ILQGLQJV KDYH GHPRQVWUDWHG WKDW PDQ\ SURWHLQV LQYROYHG LQ WKH PDLQWHQDQFH RI
JHQRPLF VWDELOLW\ WKURXJK WKH '1$ GDPDJH UHVSRQVH DOVR SDUWLFLSDWH LQ WKH UHJXODWLRQ RI WKH
PLWRWLF VSLQGOH FKHFNSRLQW LQFOXGLQJ $70 ,QGHHG GXULQJ PLWRVLV $70 LV
SKRVSKRU\ODWHG DQG DFWLYDWHG E\ WKH DXURUD % NLQDVH DQG LQ WXUQ SKRVSKRU\ODWHV FULWLFDO
FRPSRQHQWV RI WKH PLWRWLF VSLQGOH FKHFNSRLQW VXFK DV WKH EXGGLQJ XQLQKLELWHG E\
EHQ]LPLGD]ROHVKRPRORJ%8%SURWHLQNLQDVHDQGWKHPLWRWLFDUUHVWGHILFLHQWSURWHLQ
0$' LQGXFLQJ DFWLYDWLRQ RI WKHILUVW DQG GLPHUL]DWLRQ RI ODWWHU DOWRJHWKHU KDOWLQJ WKH
FHOO F\FOH XQWLO DOO FKURPRVRPHV DUH FRUUHFWO\ DWWDFKHG WR WKH PLWRWLF VSLQGOH ,Q FDVH RI $70
GHILFLHQF\ GHIHFWV LQ WKH UHJXODWLRQ RI WKH WUDQVLWLRQ IURP PHWDSKDVH WR DQDSKDVH FDXVH
DQHXSORLG\WKXVIXUWKHUVXSSRUWLQJWKHUROHRI$70LQWKHPDLQWHQDQFHRIJHQRPLFDQG
FKURPRVRPDOVWDELOLW\
)LQDOO\ $70 KDV EHHQ OLQNHG WR WKH UHVSRQVH WR DQG SURFHVVLQJ RI WRSRLVRPHUDVH 7RS'1$ FOHDYDJH FRPSOH[HV 7RSFFV LQ QHXUDO FHOOV 7RS LV DQ HQ]\PH WKDW
UHOD[HV '1$ VXSHUFRLOLQJ GXULQJ UHSOLFDWLRQ DQG WUDQVFULSWLRQ E\ JHQHUDWLQJ WUDQVLHQW '1$
66%V DQG UHOLJDWLQJ WKH VWUDQGV 7RSFFV DUH WKXV WHPSRUDU\ LQWHUPHGLDWHV FRQVLVWLQJ RI
7RSFRYDOHQWO\OLQNHGWRDQLFNHG'1$PROHFXOHDQGPXVWEHUHDGLO\UHPRYHG7KHSUHVHQFH
RIDOWHUHG'1$EDVHVIRUH[DPSOHGXHWRR[LGDWLRQRUWUHDWPHQWZLWK7RSLQKLELWRUVVXFKDV
FDPSWRWKHFLQ &37 VWDELOL]H WKH FOHDYDJH FRPSOH[HV DQG FRQVHTXHQWO\ EORFN '1$
WUDQVFULSWLRQ ,W KDV EHHQ VKRZQ WKDW WKH WUDSSLQJ RI WUDQVFULSWLRQGHSHQGHQW 7RSFFV
SUHYHQW '1$ UHOLJDWLRQ DQG LQGXFH '1$ '6%V ZKLFK OHDG WR WKH DFWLYDWLRQ RI WKH $70
,QWURGXFWLRQ
GHSHQGHQW'1$GDPDJHUHVSRQVH0RUHRYHU$70KDVDOVREHHQUHSRUWHGWRSUHYHQWWKH
DFFXPXODWLRQ RI 7RSFFV E\ UHJXODWLQJ WKH WXUQRYHU RI 7RS LQ D NLQDVHLQGHSHQGHQW
PDQQHU 7KHUHIRUH LQ WKH DEVHQFH RI $70 GHIHFWLYH 7RSFF SURFHVVLQJ FDQ
LQFUHDVHOHYHOVRI'1$GDPDJHDQGLPSDLUWUDQVFULSWLRQWKXVSDUWLFLSDWLQJLQFDXVLQJ
QHXURQDOGHDWKLQ$7
6SHFLILFUROHVRI$70LQQHXURQV
&RQFRUGDQW ZLWK WKH IDFW WKDW $7 LV FKDUDFWHUL]HG E\ VHOHFWLYH GHDWK RI QHXURQV DQ
LQFUHDVLQJQXPEHURIILQGLQJVVXSSRUWWKHQRWLRQWKDWLQQHXURQDOFHOOV$70FDUULHVRXWVSHFLILF
F\WRSODVPLFDQGQXFOHDUIXQFWLRQVZKLFKDUHHVVHQWLDOIRUWKHVXUYLYDODQGZHOOEHLQJRIWKHVH
FHOOV &RQVHTXHQWO\ WKH GLVUXSWLRQ RIWKHVHSURFHVVHV GXH WR $70 GHILFLHQF\ DGGV WR WKH
ORVVRIXELTXLWRXV$70DFWLYLWLHVDQGSDUWLFLSDWHVLQWKHRQVHWRIQHXURGHJHQHUDWLRQ
7KHILUVWQHXURQDOVSHFLILFIXQFWLRQRI$70WREHGHVFULEHGZDVLWVDVVRFLDWLRQZLWKDQG
UHJXODWLRQRIV\QDSWLFYHVLFOHF\FOLQJ6\QDSWLFYHVLFOHVDQGWKHLUUHF\FOLQJDUHHVVHQWLDOLQ
QHXURQV DV WKH\ VWRUH QHXURWUDQVPLWWHUV ZKLFK IROORZLQJ VWLPXODWLRQ DUH UHOHDVHG DW WKH
V\QDSVH IRU WKH SURSDJDWLRQ RI WKH QHUYH LPSXOVH ,Q WKH D[RQ WHUPLQDOV $70 WRJHWKHU
ZLWK $75 DVVRFLDWHV ZLWK WKH WZR V\QDSWLF YHVLFOH SURWHLQV V\QDSVLQ DQG YHVLFOHDVVRFLDWHG
PHPEUDQHSURWHLQ9$03DOVRNQRZQDVV\QDSWREUHYLQ7KHVHIRXUSURWHLQVWKXVIRUP
DFRPSOH[LQZKLFK$70DQG$75UHJXODWHWKHUHOHDVHRIYHVLFOHVE\SKRVSKRU\ODWLQJV\QDSVLQ
DQG9$03UHVSHFWLYHO\WKHUHIRUHPRGXODWLQJV\QDSWLFIXQFWLRQ0RUHUHFHQWO\LWKDVEHHQ
UHSRUWHG WKDW $70 FRRSHUDWHV ZLWK $75 DQG SRVVLEO\ RWKHU NLQDVHV LQ WKH JHQHUDWLRQ RI D
VSHFLILF>67@4SKRVSKRSURWHRPHWKDWORFDOL]HVWRGHQGULWHVV\QDSVHVDQGVRPD,QGHHG
$70 KDV EHHQ IRXQG WR EH DFWLYDWHG LQ UHVSRQVH WR GHSRODUL]LQJ SRVWV\QDSWLF VWLPXOL LQ WKH
VRPDDQGLQWKHSUR[LPDOSRUWLRQRIGHQGULWHVFRQVHTXHQWO\GULYLQJWKHSKRVSKRU\ODWLRQRID
,QWURGXFWLRQ
YDVWQXPEHURIVXEVWUDWHVZKLFKSDUWLFLSDWHLQWKHUHJXODWLRQRIQHXURQDOJHQHH[SUHVVLRQFHOO
YLDELOLW\ DQG V\QDSWLF SODVWLFLW\ 7KH GLVUXSWLRQ RI WKHVH F\WRSODVPLF IXQFWLRQV RI WKH
VHULQHWKUHRQLQH NLQDVH FRXOG FRQWULEXWH WR WKH QHXURORJLFDO V\PSWRPV RI $7 E\ LQGXFLQJ
V\QDSWLF GHJHQHUDWLRQ DV LW KDV EHHQ VKRZQ WKDW LQ WKH DEVHQFH RI $70 QHXURQV GLVSOD\
GHIHFWLYHYHVLFOHWUDIILFNLQJDQGUHGXFHGORQJWHUPSRWHQWLDWLRQZKLFKLVLQYROYHGLQV\QDSWLF
SODVWLFLW\
$QRWKHU UROH RI $70 LQ WKH QHXURQ UHJDUGV LWV LQYROYHPHQW LQ WKH SURWHFWLRQ DJDLQVW
JOXWDPDWHUJLFH[FLWRWR[LFLW\DSDWKRORJLFDOSURFHVVWKDWOHDGVWRQHXURQDOGHDWKIROORZLQJ
H[FHVVLYHVWLPXODWLRQE\H[FLWDWRU\QHXURWUDQVPLWWHUVDQGVXEVWDQFHV,WKDVEHHQVKRZQ
WKDW GXULQJ QHXURQDO GLIIHUHQWLDWLRQ LQGXFHG E\ UHWLQRLF DFLG $70 SKRVSKRU\ODWHV DQG
DFWLYDWHV WKH F$03UHVSRQVLYH HOHPHQWELQGLQJ SURWHLQ &5(% ZKLFK ELQGV WR WKH SURPRWHU
UHJLRQ RI WKH ]LQF ILQJHU KRPHRER[ =)+; WUDQVFULSWLRQ IDFWRU JHQH DQG LQGXFHV LWV
H[SUHVVLRQ,QWXUQ=)+;SURPRWHVWKHWUDQVFULSWLRQRIWKHSODWHOHWGHULYHGJURZWKIDFWRU
W\URVLQH NLQDVH UHFHSWRU ǃ 3'*)5% ZKLFK SDUWLFLSDWHV LQ WKH FHOOXODU GHIHQVH DJDLQVW
H[FLWRWR[LFLW\DQGWKXVSURPRWHVQHXURQDOVXUYLYDO$FFRUGLQJO\LQUHVSRQVHWRR[LGDWLYH
VWUHVVJHQHUDWHGE\H[FHVVLYHH[FLWDWLRQRIQHXURQV3'*)5%DFWLYDWHV$70LQD'1$GDPDJH
LQGHSHQGHQW PDQQHU WR LQKLELW P725& WKXV SURPRWLQJ DXWRSKDJ\ IRU WKH PDLQWHQDQFH RI
FHOOXODUKRPHRVWDVLV$OOWRJHWKHUWKHVHILQGLQJVIXUWKHULQGLFDWHWKDWDGHIHFWLYHSURWHFWLRQ
RIQHXURQVDJDLQVWR[LGDWLYHVWUHVVLQWKHDEVHQFHRI$70FDQSOD\DUROHLQLQGXFLQJFKURQLF
QHXURQDOGHJHQHUDWLRQ
$70 LV DOVR HQJDJHG LQ SUHVHUYLQJ WKH QHXURQDO HSLJHQHWLF VLJQDWXUH WKURXJK WKH
UHJXODWLRQRIKLVWRQHPRGLILFDWLRQZKLFKLVRISLYRWDOLPSRUWDQFHIRUPDLQWDLQLQJSURSHUJHQH
H[SUHVVLRQ LQ QHXURQV 6SHFLILFDOO\ RQH RI WKH F\WRSODVPLF UROHV RI $70 LQ QHXURQV
FRQVLVWVLQSKRVSKRU\ODWLQJDQGGRZQUHJXODWLQJWKHSURWHLQSKRVSKDWDVH33$WKXVPHGLDWLQJ
WKHUHWHQWLRQRIWKHSKRVSKRU\ODWHGIRUPRIKLVWRQHGHDFHW\ODVH+'$&LQWKHF\WRSODVPDQG
,QWURGXFWLRQ
FRQVHTXHQWO\ SUHYHQWLQJ KLVWRQH GHDFHW\ODWLRQ GXH WR LWV DFFXPXODWLRQ LQ WKH QXFOHXV )XUWKHUPRUH $70 UHGXFHV SRO\FRPE UHSUHVVLYH FRPSOH[ 35&GHSHQGHQW WULPHWK\ODWLRQ
RIKLVWRQH+RQ/\V+.PHE\SKRVSKRU\ODWLQJDQGGRZQUHJXODWLQJWKHKLVWRQHO\VLQH
1PHWK\OWUDQVIHUDVH HQKDQFHU RI ]HVWH KRPRORJ (=+ ZKLFK LV D FDWDO\WLF FRPSRQHQW RI
35& 7KHVH KLVWRQH PRGLILFDWLRQV IDYRU D PRUH RSHQ FKURPDWLQ FRQIRUPDWLRQ ZKLFK
IDFLOLWDWHV '1$ UHSDLU DQG SURPRWHV WKH WUDQVFULSWLRQ RI JHQHV LQYROYHG LQ QHXURQDO VXUYLYDO
DQGIXQFWLRQ$70GHILFLHQF\FRXOGLPSHGH'1$UHSDLUDQGFDXVHWUDQVFULSWLRQDOUHSUHVVLRQRI
PXOWLSOH QHXURQDO JHQHV WKURXJK GHFUHDVH RI KLVWRQH DFHW\ODWLRQ DQG LQFUHDVH RI KLVWRQH
PHWK\ODWLRQWKXVSURPRWLQJWKHQHXURGHJHQHUDWLYHSKHQRW\SHRI$7
0RGHOVRIQHXURGHJHQHUDWLRQLQDWD[LDWHODQJLHFWDVLD
)RUWKHSXUSRVHRIJDLQLQJLQVLJKWVLQWRWKHPHFKDQLVPVXQGHUO\LQJWKHQHXURSDWKRORJ\
RI $7 WKH GHVLJQ DQG XVH RI H[SHULPHQWDO PRGHOV LV HVVHQWLDO ,QGHHG VWXGLHV FRQGXFWHG LQ
YLYRLQZKROH DQLPDOVDQG LQ YLWUR LQ FHOO FXOWXUHV KDYH SURYLGHG LQYDOXDEOH LQIRUPDWLRQRQ
WKH FHOOXODU DQG PROHFXODU GHIHFWV WKDW FKDUDFWHUL]H WKH QHXURORJLFDO SKHQRW\SH 1RQHWKHOHVV
WKHODFNRIDSSURSULDWHDQLPDODQGFHOOXODUPRGHOVLVRQHRIWKHPDMRUGUDZEDFNVRIUHVHDUFKLQJ
WKHFDXVHVRIQHXURGHJHQHUDWLRQLQ$7
$70GHILFLHQWDQLPDOPRGHOV
7KHXVHRIJHQHWLFDOO\HQJLQHHUHGDQLPDOPRGHOVLVH[WUHPHO\LPSRUWDQWIRULQFUHDVLQJ
WKH NQRZOHGJH RQ WKH SDWKRSK\VLRORJLFDO PHFKDQLVPV RI QHXURGHJHQHUDWLYH GLVHDVHV 7KHVH PRGHOV DLP WR UHFDSLWXODWH WKH FDXVHV WKH SDWKRORJLFDO OHVLRQV DQG WKH V\PSWRPV RI D
VSHFLILF GLVHDVH DQG DUH JHQHUDOO\ XVHIXO IRU DVVHVVLQJ WKH HIILFDF\ RI SRWHQWLDO WKHUDSHXWLF
VWUDWHJLHVEHIRUHFRQGXFWLQJFOLQLFDOWULDOVLQKXPDQV
,QWURGXFWLRQ
'URVRSKLOD PHODQRJDVWHU LV DQ LQYHUWHEUDWH ZKLFK LV RIWHQ XVHG DV D PRGHO RUJDQLVP IRU
QHXURGHJHQHUDWLYH GLVRUGHUV ,QGHHG LWV VKRUW OLIH F\FOHDQG VLPSOH JHQRPH DOORZ UDSLG
FRQVWUXFWLRQRIWUDQVJHQLFDQLPDOVZKLFKVXFFHHGLQSDUWLDOO\UHSURGXFLQJWKHOHVLRQVIRXQGLQ
GLIIHUHQW GLVHDVHV ' PHODQRJDVWHU KDV SURYHQ LQVWUXPHQWDO LQ GLVVHFWLQJ EDVLF GLVHDVH
PHFKDQLVPV LQ $7 EXW LWV XVH LQ PHGLFDO UHVHDUFK LV OLPLWHG E\ WKH JUHDW
SK\VLRORJLFDOGLIIHUHQFHVEHWZHHQIOLHVDQGKXPDQV
6LQFH PLFH GLVSOD\ VWULNLQJ DQDWRPLFDO DQG SK\VLRORJLFDO VLPLODULWLHV WR KXPDQV ZLWK
ZKRP WKH\ VKDUH RI WKHLU JHQHV WKH\ DUH ZLGHO\ FRQVLGHUHG WR EH WKH SULPH PRGHO RI
LQKHULWHG KXPDQ GLVHDVH ,QGHHG WKH DELOLW\ WR HQJLQHHU WKH PRXVH JHQRPH HQDEOHV WKH
JHQHUDWLRQRIWUDQVJHQLFPLFHZKLFKLGHDOO\VKRXOGIDLWKIXOO\UHSUHVHQWWKHKXPDQSKHQRW\SH
7KH LGHQWLILFDWLRQ RI WKH $70 JHQH LQ DOORZHG WKH GHYHORSPHQW RI $70GHILFLHQW
NQRFNRXW PLFH LQ ZKLFK WKH $70 JHQH LV GLVUXSWHG DQG QR $70 SURWHLQ LV SURGXFHG DQG
NQRFNLQ PRXVH PRGHOV LQ ZKLFK D GHOHWLRQ LV LQWURGXFHG LQWR WKH $70 JHQH UHVXOWLQJ LQ WKH
SURGXFWLRQ RI DQ $70 SURWHLQ WKDW ODFNV NLQDVH DFWLYLW\ 7KH JRDO ZDV WR DFKLHYH D
JUHDWHUXQGHUVWDQGLQJRIWKHGLVRUGHUDQGSURYLGHDPHDQVRIVFUHHQLQJIRUGUXJVWRSUHYHQWRU
DOOHYLDWH WKH GHIHFWV LQ $7 SDWLHQWV SDUWLFXODUO\ WKH QHXURORJLFDO GHFD\ 8QIRUWXQDWHO\ ZKLOH
WKHVH PLFH H[KLELW WKH PDMRU FKDUDFWHULVWLFV RI KXPDQ $7 LQFOXGLQJ LPPXQRORJLFDO
DEQRUPDOLWLHVUDGLRVHQVLWLYLW\FDQFHUSUHGLVSRVLWLRQDQGLQIHUWLOLW\WKH\EDUHO\VKRZWKHPRVW
FDUGLQDO IHDWXUHV RI $7 QHXURGHJHQHUDWLRQ DQG DVVRFLDWHG DWD[LD ,QGHHG DOWKRXJK
PLFHODFNLQJ$70GLVSOD\VRPHQHXUREHKDYLRUDOGHIHFWVDQGVXEWOHQHXURORJLFDODEQRUPDOLWLHV
FRQVLVWHQWZLWKDFHQWUDOQHUYRXVV\VWHPWKDWLVQRWWRWDOO\QRUPDOWKH\IDLOWR
GHYHORSVHYHUHQHXURQDOGHJHQHUDWLRQDQGQHXURPRWRUG\VIXQFWLRQ7KHGLIIHUHQFHEHWZHHQWKH
QHXURQDO SKHQRW\SHV LQ $7 SDWLHQWV DQG LQ $70GHILFLHQW PLFH LV SUREDEO\ FDXVHG E\
GLIIHUHQFHV LQ WKH DPRXQWV RI '1$ GDPDJH WKDW FDXVH QHXURQDO GHPLVH DQG WKH VKRUW OLIH
H[SHFWDQF\RIWKHPRXVHZKLFKGRQRWDOORZWKHGHJHQHUDWLYHSURFHVVWRGHYHORSDVLWGRHVLQ
,QWURGXFWLRQ
KXPDQV ,W LV WUXH WKDW WKHVH PRXVH PRGHOV DQG WKH FHOO OLQHV GHULYHG IURP WKHP KDYH
SURYLGHGJUHDWHULQVLJKWLQWRWKHSDWKRSK\VLRORJ\RIWKHGLVRUGHULQFOXGLQJWKHUROHRI$70LQ
SUROLIHUDWLQJ DQG SRVWPLWRWLF FHOOV DQG GHIHFWV LQ VLJQDOOLQJ SDWKZD\V 1RQHWKHOHVV WKH
GLVDSSRLQWLQJ SKHQRW\SH KDV FDVW D VKDGRZ RYHU WKH XVH RI WKHVH DQLPDOV IRU VWXG\LQJ WKH
QHXURORJLFDODVSHFWVRI$7DQGFOHDUO\WKHUHLVDQHHGIRUDPRGHOWKDWPRUHFORVHO\UHIOHFWVWKH
KXPDQGLVRUGHU
$70GHILFLHQWKXPDQFHOOXODUPRGHOV
$VDNH\OLPLWDWLRQWRWKHVWXG\RIWKH$7QHUYRXVV\VWHPLVWKHLQDSSURSULDWHQHVVRI
WKHPRXVHPRGHORIWKHGLVHDVHYDULRXVLQYLWURKXPDQFHOOXODUPRGHOVKDYHEHHQHPSOR\HGIRU
LQYHVWLJDWLQJWKHFHOOXODUDQGPROHFXODUPHFKDQLVPVWKDWFKDUDFWHUL]H$70GHILFLHQWQHXURQDO
FHOOV LQ RUGHU WR RYHUFRPH WKLV REVWDFOH ,QGHHG LQ YLWUR PRGHOV RI QHXURGHJHQHUDWLYH
GLVHDVHVDUHXVHIXOIRUH[SORULQJWKHSDWKRORJLFDOSURFHVVHVOHDGLQJWRWKHGHIHFWLYHSKHQRW\SH
DQG FDQ DOVR EH HPSOR\HG IRU HYDOXDWLQJ SRVVLEOH GHOHWHULRXV RU SURWHFWLYH UROHV RI VSHFLILF
PROHFXOHVDQGFRPSRXQGV8QIRUWXQDWHO\OLYHDGXOWKXPDQQHXURQVDUHQRWUHDGLO\DYDLODEOH
IRU IXQFWLRQDO DQG LQWHUYHQWLRQDO VWXGLHV WKXV FHOOXODU PRGHOV RI $7 DUH EDVHG RQ LQGXFHG
QHXURQDOGLIIHUHQWLDWLRQRISOXULSRWHQWRUPXOWLSRWHQWFHOOVLQFXOWXUH
&HOO OLQHVWKDW DUH ZLGHO\ XVHG WR VWXG\ PHFKDQLVPV RIQHXURGHJHQHUDWLRQ DQG ZKLFK
DUH DOVR XVHG LQ WKH ILHOG RI $7 UHVHDUFK DUH WKRVH GHULYHG IURP KXPDQ
QHXUREODVWRPDFHOOV7KHVHFHOOOLQHVDUHRIWHQXVHGDVLQYLWURPRGHOVRIQHXURQDOIXQFWLRQDQG
GLIIHUHQWLDWLRQ EHFDXVH EHLQJ WUDQVIRUPHG FHOO OLQHV WKH\ FDQ EH SURSDJDWHG LQ YLWUR DQG
GLIIHUHQWLDWHGWRDPRUHPDWXUHQHXURQOLNHSKHQRW\SH+RZHYHUWRLQYHVWLJDWHWKHUROHRI
$70LQQHXURGHJHQHUDWLRQLWLVGHVLUDEOHWRKDYHPRGHOV\VWHPVPRUHUHSUHVHQWDWLYHRIKXPDQ
QHXURQV
,QWURGXFWLRQ
$ PRUH SHUWLQHQW LQ YLWUR PRGHO V\VWHP WR VWXG\ QHXURGHJHQHUDWLRQ LQ $7 LV
UHSUHVHQWHG E\ KXPDQ QHXUDO SUHFXUVRU FHOOV DQG PDWXUH QHXURQV GHULYHG IURP WKH
GLIIHUHQWLDWLRQ RI SOXULSRWHQW HPEU\RQLF VWHP FHOOV (6&V ,QGHHG WKHVH FHOOV DUH
GHULYHG IURP WKH LQQHU FHOO PDVV RI D EODVWRF\VW DQG DUH SOXULSRWHQW KDYLQJ WKH SRWHQWLDO WR
JHQHUDWHDOOWKHFHOOW\SHVRIWKHKXPDQRUJDQLVP8QIRUWXQDWHO\LVRODWLQJWKHLQQHUFHOOPDVV
UHVXOWVLQGHVWUXFWLRQRIWKHEODVWRF\VWZKLFKUDLVHVHWKLFDOLVVXHVWKDWKDPSHUWKHXVHRIKXPDQ
(6&VLQELRPHGLFDOUHVHDUFK
$QRWKHU SRWHQW H[SHULPHQWDO WRRO WR VWXG\ QHXURGHJHQHUDWLRQ LV RIIHUHG E\ JHQHWLFDOO\
VWDEOHKXPDQQHXUDO VWHP FHOOV16&V ZKLFK UHVLGH LQ VSHFLILF UHJLRQV RI WKH IHWDO DQG DGXOW
FHQWUDOQHUYRXVV\VWHPDQGSDUWLFLSDWHLQLWVKRPHRVWDVLV7KHVHFHOOVUHWDLQORQJWHUPLQ
YLWURVHOIUHQHZDOFDSDFLW\DQGFDQGLIIHUHQWLDWHLQWRQHXURQVDVWURF\WHVDQGROLJRGHQGURF\WHV
LQ SURSRUWLRQV WKDW PLUURU WKHLU SK\VLRORJLFDO GLVWULEXWLRQ EXW KDYH EHHQ DUJXHG WR EH
KHWHURJHQHRXV
7KHLPSDFWRI$70GHILFLHQF\RQWKHELRORJLFDOSURSHUWLHVRIWKHVHFHOOVLVVWXGLHGHLWKHU
E\FKHPLFDOLQKLELWLRQRI$70NLQDVHDFWLYLW\ZLWKWKHVSHFLILFLQKLELWRU.8RUE\
VWDEOH NQRFNGRZQ RI $70 PHGLDWHG E\ OHQWLYLUDO VKRUW KDLUSLQ 51$ YHFWRUV +RZHYHULWDSSHDUVWKDWLQKLELWLRQRI$70DFWLYLW\DQGNQRFNGRZQRIWKHSURWHLQGRQRWIXOO\
PLUURUWKHEURDGVSHFWUXPRI$7FDXVLQJPXWDWLRQV
,QWHUHVWLQJO\ QHXURQDO PRGHOV RI $7 EDVHG RQ WKH XVH RI SDWLHQWGHULYHG WLVVXHV DUH
FXUUHQWO\ EHLQJ GHYHORSHG )RU H[DPSOH ROIDFWRU\ PXFRVDGHULYHGQHXURVSKHUHV REWDLQHGE\
ELRSV\WKURXJKWKHQRVWULOKDYHUHFHQWO\EHHQHVWDEOLVKHGIURPVHYHUDO$7SDWLHQWVDQGKDYH
EHHQ VKRZQ WR GLIIHUHQWLDWH LQ YLWUR LQWR QHXURQV DVWURF\WHV DQG ROLJRGHQGURF\WHV +RZHUYHU RQH RI WKH PRVW SURPLVLQJ KXPDQ LQ YLWUR PRGHOV RI QHXURGHJHQHUDWLYH GLVHDVHV
LQFOXGLQJ $7 LV UHSUHVHQWHG E\ SDWLHQWGHULYHG QHXUDOVSHFLILHG LQGXFHG SOXULSRWHQW VWHP
FHOOV
,QWURGXFWLRQ
+XPDQ LQGXFHG SOXULSRWHQW VWHP FHOOGHULYHG QHXUDO FHOOV
IURPSDWLHQWVZLWKDWD[LDWHODQJLHFWDVLD
,QYHVWLJDWLRQ RI WKH FDXVHV RI PDQ\ KXPDQ QHXURORJLFDO GLVRUGHUV LQFOXGLQJ $7 KDV
EHHQ KDPSHUHG E\ WKH LQDGHTXDF\ RI DQLPDO DQG FHOOXODU PRGHOV DQG E\ WKH GLIILFXOW\ LQ
REWDLQLQJSDWLHQWGHULYHGQHXUDOFHOOVRUWLVVXHVEHFDXVHRIWKHOLPLWHGDFFHVVLELOLW\WRWKHEUDLQ
$ PDMRU EUHDNWKURXJK LQ UHVHDUFK FDPH LQ UHFHQW \HDUV ZKHQ 3URIHVVRU 6KLQ\D <DPDQDND RI
.\RWR8QLYHUVLW\HVWDEOLVKHGDUHSURJUDPPLQJPHWKRGIRUWKHJHQHUDWLRQRISOXULSRWHQWVWHP
FHOOVIURPPRXVHDQGKXPDQDGXOWFHOOVWKURXJKUHWURYLUDOGHOLYHU\RIIRXUJHQHVWKDWHQFRGH
WKHWUDQVFULSWLRQIDFWRUV2&762;./)DQGF0<&7KHVHFHOOVZHUHGHVLJQDWHG
DVLQGXFHGSOXULSRWHQWVWHPFHOOVL36&VDQGUHVHPEOH(6&VLQWKHLUVHOIUHQHZDOFDSDFLW\DQG
SOXULSRWHQF\EHLQJDEOHWRSURSDJDWHLQGHILQLWHO\DVZHOODVJLYHULVHWRHYHU\RWKHUFHOOW\SHLQ
WKHERG\7KLVWHFKQRORJ\ZKLFKLVQRZDGD\VEDVHGRQQRQYLUDOLQWHJUDWLRQIUHHGHOLYHU\RI
UHSURJUDPPLQJIDFWRUVKDVPDGHLWSRVVLEOHWRHVWDEOLVKL36&VIURPWKHVRPDWLFFHOOVRI
DQ\LQGLYLGXDOLUUHVSHFWLYHRIUDFHJHQHWLFEDFNJURXQGDQGVWDWHRIKHDOWKWKXVFUHDWLQJJUHDW
KRSH QRW RQO\ LQ WHUPV RI DSSOLFDWLRQV LQ UHJHQHUDWLYH PHGLFLQH VXFK DV FHOO WUDQVSODQWDWLRQ
WKHUDS\ EXW DOVR IRU PRGHOLQJ KXPDQ GLVHDVHVDQG IRUQHZ GUXJ GHYHORSPHQW ,QGHHG
SURWRFROV HVWDEOLVKHG IRU WKH LQ YLWUR GLIIHUHQWLDWLRQ RI (6&V WRZDUG DOO WKUHH HPEU\RQLF JHUP
OD\HUV FDQ EH DSSOLHG WR L36&V JUDQWLQJ WKH SURGXFWLRQ RI D YDULHW\ RI L36&GHULYHG FHOOV
LQFOXGLQJKHSDWRF\WHVFDUGLRP\RF\WHVDQGQHXURQV)LJXUH
,QWURGXFWLRQ
)LJ6FKHPDWLFUHSUHVHQWDWLRQRIKXPDQL36&JHQHUDWLRQGLIIHUHQWLDWLRQDQGSRWHQWLDODSSOLFDWLRQV
$VDUHVXOWWKHL36&WHFKQRORJ\KDVUDSLGO\H[SDQGHGZRUOGZLGHDQGWKHSRVVLELOLW\WR
UHSURJUDPSDWLHQWGHULYHGFHOOVWREHFRPHSOXULSRWHQWDQGWKHLUVXEVHTXHQWGLIIHUHQWLDWLRQLQWR
SXUH SRSXODWLRQV RI GLVHDVHUHOHYDQW VRPDWLF FHOOV KDV DOORZHG WKH FRQVWUXFWLRQ RI LQ YLWUR
GLVHDVH PRGHOV WKDW FRQWDLQ SDWLHQWVSHFLILF JHQHWLF LQIRUPDWLRQ WKXV SURYLGLQJ DQ
XQOLPLWHGVRXUFHRILQIRUPDWLYHPDWHULDOWRLQYHVWLJDWHWKHGLVHDVHRILQWHUHVW,QSDUWLFXODUWKH
UHSURJUDPPLQJRIDGXOWKXPDQVRPDWLFFHOOVWRDOWHUQDWLYHIDWHVKDVHQDEOHGWKHJHQHUDWLRQRI
QRYHOFHOOXODUPRGHOVRIFHQWUDOQHUYRXVV\VWHPGLVRUGHUVVXFKDV$O]KHLPHU·VGLVHDVH
3DUNLQVRQ·V GLVHDVH DP\RWURSKLF ODWHUDO VFOHURVLV +XQWLQJWRQ·V GLVHDVH
VSLQDOPXVFXODUDWURSK\DQG5HWWV\QGURPH
,Q WKH FDVH RI $7 WKH HVWDEOLVKPHQW RI SDWLHQWGHULYHG L36&V DQG WKHLU FRQYHUVLRQ WR
IXQFWLRQDOQHXURQVKDVEHHQUHFHQWO\DFKLHYHG$FFHVVWRWKHVHFHOOW\SHVFDQSURYLGHD
PHDQV RI HQKDQFLQJ RXU NQRZOHGJH UHJDUGLQJ WKH UROH RI $70 LQ WKH QHXURQDO FHOOXODU
,QWURGXFWLRQ
HQYLURQPHQW DQG UHSUHVHQWV D SRZHUIXO WRRO IRU H[SORULQJ WKH PHFKDQLVPV WKDW FDXVH
GHJHQHUDWLRQ RI KXPDQ $70GHILFLHQW QHXURQV 7KLV LQ WXUQ PD\ EH H[SORLWHG IRU GHWHFWLQJ
WKHUDSHXWLF WDUJHWV DQG IRU IXWXUH LQ YLWUR VFUHHQLQJV IRU WKH LGHQWLILFDWLRQ RI QHXURSURWHFWLYH
PROHFXOHVRISKDUPDFRORJLFDOUHOHYDQFH1HYHUWKHOHVVDWKRURXJKFKDUDFWHUL]DWLRQRIWKHQHXUDO
SURJHQ\ RI $7 SDWLHQWGHULYHG LQGXFHG SOXULSRWHQW VWHP FHOOV KDV QRW \HW EHHQ UHSRUWHG
7KHUHIRUH LQ WKLV VWXG\ ZH SHUIRUPHG D SUHYLRXVO\ XQGRFXPHQWHG IXQFWLRQDO DQG PROHFXODU
DQDO\VLV RI KXPDQ L36&GHULYHG $7 QHXURQDO FHOOV KDUERULQJ SDWLHQWVSHFLILF PXWDWLRQV LQ
RUGHUWRVKHGOLJKWRQWKHFDXVHVRIWKHQHXURSDWKRORJ\LQ$7
$LPRIWKH3URMHFW
$LPRIWKHSURMHFW
7KLV VWXG\ DLPV WR FODULI\ WKH PHFKDQLVWLF OLQN EHWZHHQ $70 DQG QHXURGHJHQHUDWLRQ
,QGHHGDVLGHIURPGHIHFWLYH'1$GDPDJHUHVSRQVHDQGFRQVHTXHQWDFFXPXODWLRQRIJHQRWR[LF
OHVLRQV LQ SUH DQG SRVWPLWRWLF QHXURQV RWKHU IDFWRUV VHHPLQJO\ FRQWULEXWH WR QHXURQDO FHOO
GHDWK 8QIRUWXQDWHO\ PRXVH PRGHOV RI $7 DQG LQ YLWUR $70GHILFLHQW FHOO OLQHV GR QRW IXOO\
UHFDSLWXODWH WKH KXPDQ FHQWUDO QHUYRXV V\VWHP GHIHFWV KDPSHULQJ VWXGLHV RQ QHXURORJLFDO
GHFD\ LQ $7 7KHUHIRUH WKH SXUSRVH RI WKLV SURMHFW LV WR LQYHVWLJDWH WKH UHODWLRQVKLS EHWZHHQ
$70 GHILFLHQF\ DQG QHXURGHJHQHUDWLRQ XVLQJ D QRYHO LQ YLWUR PRGHO RI $7 SDWLHQWGHULYHG
L36&V,QSDUWLFXODUWKHVSHFLILFDLPVRIWKLVWKHVLVDUHWKHIROORZLQJ
*HQHUDWLRQ RI KXPDQ L36&V WKURXJK WKH UHSURJUDPPLQJ RI KHDOWK\ FRQWURO DQG $7
SDWLHQWGHULYHGVNLQILEUREODVWV
'LIIHUHQWLDWLRQ RI KXPDQ L36&V LQWR QHXUDO SUHFXUVRU FHOOV DQG WHUPLQDOO\ GLIIHUHQWLDWHG
QHXURQV
'HWHUPLQDWLRQRIWKHRXWFRPHRI$70GHILFLHQF\RQQHXURQDOGLIIHUHQWLDWLRQVXUYLYDODQG
HOHFWURSK\VLRORJLFDODFWLYLW\LQRUGHUWRHYDOXDWHSRVVLEOHPDWXUDWLRQRUIXQFWLRQDOGHIHFWV
$QDO\VLV RI '1$ UHSDLU DFWLYLWLHV DQG UHVSRQVHV WR JHQRWR[LF RU R[LGDQW DJHQWV LQ
SRVWUHSOLFDWLYHGLIIHUHQWLDWHGQHXURQVLQUHODWLRQWR$70GHILFLHQF\
$VVHVVPHQW RI WKH YLDELOLW\ RI $70GHILFLHQW SRVWUHSOLFDWLYH GLIIHUHQWLDWHG QHXURQDO FHOOV
IROORZLQJH[SRVXUHWRYDULRXVGRVHVRIJHQRWR[LFRUR[LGDQWDJHQWV
0DWHULDOVDQG0HWKRGV
0DWHULDOVDQG0HWKRGV
3ULPDU\FHOOFXOWXUHV
*0DQG*0$7SDWLHQWGHULYHGSULPDU\GHUPDOILEUREODVWVZHUHSXUFKDVHG
IURP WKH &RULHOO ,QVWLWXWH IRU 0HGLFDO 5HVHDUFK &DPGHQ 1- 86$ %RWK FHOO OLQHV FDUULHG
FRPSRXQG KHWHUR]\JRXV PXWDWLRQV LQ WKH $70 JHQH LQ SDUWLFXODU *0 F$!7 DQG
FBLQV *0 FBGHO LQ WUDQV ZLWK F*!$ DQG FBGHO
3ULPDU\ GHUPDO ILEUREODVWV IURP WKH VNLQ ELRSV\ RI D KHDOWK\ GRQRU ZHUH NLQGO\ SURYLGHGE\
'U &KLDUD 9HUSHOOL &15 ,QVWLWXWH RI 1HXURVFLHQFH DQG 'HSDUWPHQW RI %LRWHFKQRORJ\ DQG
7UDQVODWLRQDO 0HGLFLQH0LODQ ,WDO\ 3ULPDU\ PRXVH HPEU\RQLF ILEUREODVWV VWUDLQ &) ZHUH
SXUFKDVHGIURP0LOOLSRUH30()&)/
$OO SULPDU\ ILEUREODVWV ZHUH FXOWXUHG DW ƒ& LQ D KXPLGLILHG &2 LQFXEDWRU LQ
FRQYHQWLRQDODWPRVSKHULF27KH$7DQGFRQWUROSULPDU\GHUPDOILEUREODVWVZHUHPDLQWDLQHG
DQG H[SDQGHG LQ (0(0 /RQ]D VXSSOHPHQWHG ZLWK KHDWLQDFWLYDWHG IHWDO FDOI VHUXP
)&6 /LIH 7HFKQRORJLHV ZKLOH 30()&)/ ZHUH PDLQWDLQHG DQG H[SDQGHG LQ '0(0)
/LIH 7HFKQRORJLHV VXSSOHPHQWHG ZLWK KHDWLQDFWLYDWHG )&6 %RWK PHGLD DOVR FRQWDLQHG
XQLWVPO SHQLFLOOLQ —JPO VWUHSWRP\FLQ /RQ]D Ǎ0 QRQHVVHQWLDO DPLQR DFLGV
/RQ]D DQG P0 JOXWDPLQH /RQ]D $OO ILEUREODVWV ZHUH SDVVDJHG LQ XQFRDWHG 7 IODVNV
ZKHQ WKH\ UHDFKHG FRQIOXHQFH %ULHIO\ DOO FXOWXUH PHGLXP ZDV UHPRYHG IURP WKH IODVN
FHOOV ZHUH ULQVHG ZLWK 3%6 /RQ]D DQG PO RI WU\SVLQ('7$ VROXWLRQ 8O WU\SVLQ
PJO ('7$ /RQ]D ZHUH DGGHG VR WKDW WKH HQWLUH VXUIDFH ZDV FRYHUHG 7KH IODVN ZDV
LQFXEDWHGDWƒ&IRUPLQDQGZKHQWKHFHOOVEHFDPHSDUWLDOO\GHWDFKHGDQGURXQGHGWKH\
ZKHUH GLVORGJHG IURP WKH VXUIDFH RI WKH IODVN E\ JHQWO\ WDSSLQJ WKH VLGH RI LW 7U\SVLQ ZDV
QHXWUDOL]HGE\DGGLQJPORIPHGLXPWRWKHIODVNDQGWKHGHWDFKHGFHOOVZHUHWUDQVIHUUHGWRD
VWHULOH PO FRQLFDO WXEH 7KH FHOOV ZHUH WKHQ FHQWULIXJHG DW USP IRU PLQ 7KH
VXSHUQDWDQW ZDV GLVFDUGHG IURP WKH WXEH ZLWKRXW GLVORGJLQJ WKH FHOO SHOOHW ZKLFK ZDV
0DWHULDOVDQG0HWKRGV
UHVXVSHQGHG LQ IUHVK JURZWK PHGLXP 7KH FHOOV ZHUH SLSHWWHG XS DQG GRZQ WR HQVXUH D
KRPRJHQHRXVFHOOVXVSHQVLRQDQGSODWHGLQQHZFXOWXUHIODVNVDWDVSOLWUDWLRRI
,Q RUGHU WR SUHSDUH D IHHGHU OD\HU IRU WKH JHQHUDWLRQ DQGRU PDLQWHQDQFH RI KXPDQ
L36&V30()&)/LQDFRQIOXHQWIODVNZHUHPLWRWLFDOO\LQDFWLYDWHGZLWK—JPOPLWRP\FLQ&
.\RZD +DNNR .LULQ 0LODQ ,WDO\ LQ '0(0 KLJK JOXFRVH (XURFORQH 3HUR ,WDO\ IRU K DW
ƒ&0HDQZKLOHWKHDSSURSULDWHQXPEHURIZHOOVRIDZHOOSODWHRUFPGLVKHVZHUHFRDWHG
ZLWKJHODWLQVROXWLRQIRUPLQDWƒ&DGVHWDVLGHXQWLOUHDG\WRUHFHLYHWKHLQDFWLYDWHG
30()&)/$IWHUWUHDWPHQWWKHIODVNFRQWDLQLQJWKH30()&)/ZDVZDVKHGZLWKSUHZDUPHG
'0(0) WKUHH WLPHV DVSLUDWLQJ DIWHU HDFK ZDVK &HOOV ZHUH FROOHFWHG DV SUHYLRXVO\
GHVFULEHG ZLWK WULSVLQ('7$ VROXWLRQ FHQWULIXJHG DQG UHVXVSHQGHG LQ D VPDOO YROXPH RI
PHGLXPDQGWKHLUFRQFHQWUDWLRQZDVGHWHUPLQHG7KHJHODWLQVROXWLRQZDVDVSLUDWHGIURPWKH
ZHOOVRUGLVKHVZKLFKZHUHWKHQVHHGHGZLWK[FHOOVZHOORU[FHOOVFPGLVKLQ
IUHVK 30()&)/ PHGLXP ,QDFWLYDWHG 30()&)/ ZHUH WKHQ LQFXEDWHG IRU RQH WR WZR GD\V
XQWLOWKH\ZHUHXVHG
*HQHUDWLRQ DQG FXOWXUH RI KXPDQ LQGXFHG SOXULSRWHQW VWHP
FHOOV
7KHSURGXFWLRQRIKXPDQL36&VZDVDFKLHYHGWKURXJKWZRWHFKQLTXHVEDVHGRQQXFOHLF
DFLG GHOLYHU\ RI WKH IRXU <DPDQDND WUDQVFULSWLRQ IDFWRUV 2&7 ./) 62; DQG F0<&
OHQWLYLUDO LQIHFWLRQ XVLQJ WKH KXPDQ 67(0&&$ K67(0&&$ &UH([FLVDEOH &RQVWLWXWLYH
3RO\FLVWURQLF /HQWLYLUXV NLW 0LOOLSRUH DQG WUDQVLHQW WUDQVIHFWLRQ WKURXJK HOHFWURSRUDWLRQ RI
HSLVRPDOSODVPLGYHFWRUV
0DWHULDOVDQG0HWKRGV
/HQWLYLUDOLQIHFWLRQ
,QLWLDOO\ KXPDQ SULPDU\ ILEUREODVWV ZHUH UHSURJUDPPHG XVLQJ WKH K67(0&&$ &UH
([FLVDEOH &RQVWLWXWLYH 3RO\FLVWURQLF /HQWLYLUXV NLW $ &UHH[FLVDEOH SRO\FLVWURQLF OHQWLYLUDO
YHFWRU DOORZV IRU H[SUHVVLRQ RI D VWHP FHOO FDVVHWWH RU 67(0&&$ FRPSULVHG RI DOO IRXU
WUDQVFULSWLRQIDFWRUV2&7./)62;DQGF0<&7KLVHQDEOHVDUHGXFHGQXPEHURIYLUDO
LQWHJUDWLRQVZLWKUHVSHFWWRFRLQIHFWLRQRIWKHVHIDFWRUVLQIRXUVHSDUDWHH[SUHVVLRQYHFWRUV
2QGD\WKHILEUREODVWVWREHUHSURJUDPPHGZHUHVHHGHGLQWZRZHOOVRIDZHOOSODWH
DWDFRQFHQWUDWLRQRI[FHOOVZHOOLQKXPDQILEUREODVWJURZWKPHGLXP&HOOVZHUHLQFXEDWHG
RYHUQLJKWDQGWKHIROORZLQJGD\EHIRUHWUDQVGXFWLRQWKHQXPEHURIFHOOVLQRQHZHOORIWKH
ZHOOSODWHZHUHFRXQWHGLQRUGHUWRFDOFXODWHWKHYROXPHRIYLUXVQHHGHGWRDFKLHYHD02,RI
XVLQJWKHIROORZLQJHTXDWLRQ
9LUXVYROXPH—OUHTXLUHG 1XPEHURIFHOOVð'HVLUHG02,ð—O
9LUXV7LWHU,)8POPO
7KH YLDO RI K67(0&&$/HQWLYLUXV ZDV WKDZHGDQG NHSW RQ LFH 7KH PHGLXP IURP HDFK ZHOO
ZDVUHSODFHGZLWKPOIUHVKKXPDQILEUREODVWPHGLXPDQGWKHUHTXLUHGYROXPHRIYLUXVZDV
DGGHGWRWKHZHOOVFRQWDLQLQJWKHILEUREODVWV7KHSODWHZDVLQFXEDWHGRYHUQLJKWDQGKODWHUD
VHFRQG YLUXV LQIHFWLRQ ZDV SHUIRUPHG 7KH IROORZLQJ GD\ FHOOV ZHUH ZDVKHG WKUHH WLPHV ZLWK
3%6DVSLUDWLQJDIWHUHDFKZDVKWKHQPOIUHVKKXPDQILEUREODVWPHGLXPZHUHDGGHGWRHDFK
ZHOO DQG FHOOV ZHUH WUDQVIHUUHG WR D 2 LQFXEDWRU 2Q GD\V DQG WKH PHGLXP ZDV
UHSODFHG2QGD\WKHDSSURSULDWHQXPEHURIZHOOVRILQDFWLYDWHG30()&)/IHHGHUOD\HUVZDV
SUHSDUHG DV GHVFULEHG LQ VHFWLRQ WR VXSSRUW WKH FHOOV EHLQJ UHSURJUDPPHG 2Q GD\ WKH
PHGLXPZDVUHPRYHGIURPWKHZHOOSODWHFRQWDLQLQJWKHIHHGHUOD\HUVDQGDIWHUULQVLQJZLWK
3%6 UHSODFHG ZLWK KXPDQ L36& PHGLXP FRQVLVWLQJ RI '0(0) VXSSOHPHQWHG ZLWK .QRFN2XW VHUXP UHSODFHPHQW /LIH 7HFKQRORJLHV XQLWVPO SHQLFLOOLQ —JPO
0DWHULDOVDQG0HWKRGV
VWUHSWRP\FLQ Ǎ0 QRQHVVHQWLDO DPLQR DFLGV P0 JOXWDPLQH P0 VRGLXP S\UXYDWH
Ǎ0 PHUFDSWRHWKDQRO DQG QJPO E)*) 3HSURWHFK 5RFN\ +LOO 1- 86$ 7KH YLUXV
LQIHFWHG FHOOV ZHUH GHWDFKHG IURP WKH ZHOOV ZLWK WU\SVLQ('7$ VROXWLRQ DQG DIWHU DGGLQJ
ILEUREODVWPHGLXPIRUWU\SVLQQHXWUDOL]DWLRQZHUHFROOHFWHGLQDPOWXEHDQGFHQWULIXJHGDW
USP IRU PLQ &HOOV ZHUH UHVXVSHQGHG LQ PO RI KXPDQ L36& PHGLXP FRXQWHG DQG
VHHGHGDWDFRQFHQWUDWLRQRI[FHOOZHOORQWRWKHZHOOSODWHFRQWDLQLQJLQDFWLYDWHG30()
&)/ 7KHQH[W GD\ WKHPHGLXP ZDVQRW FKDQJHG EXW LW ZDV H[FKDQJHG HYHU\ RWKHU GD\ ZLWK
IUHVKKXPDQL36&PHGLXPVWDUWLQJRQGD\)UHVKLQDFWLYDWHG30()&)/ZHUHDGGHGHYHU\
GD\V WR UHSOHQLVK ROGHU IHHGHUV GXULQJ WKH UHSURJUDPPLQJ WLPHFRXUVH &HOO JURZWK DQG
PRUSKRORJ\ZDVPRQLWRUHGGDLO\$IWHUGD\VKXPDQL36&FRORQLHVVWDUWHGWRDSSHDUDQG
ZKHQWKH\UHDFKHGDSSUR[LPDWHO\FHOOVRURYHULQVL]HWKH\ZHUHUHDG\WREHSLFNHG
)RU H[SDQVLRQ RQ D IHHGHU OD\HU WZR GD\V SULRU WR SLFNLQJ WKH KXPDQ L36& FRORQLHV
IUHVK ZHOOV IURP D ZHOO SODWH ZHUH SUHSDUHG ZLWK LQDFWLYDWHG 30()&)/ )RU H[SDQVLRQ LQ
IHHGHUIUHH FRQGLWLRQVIUHVKZHOOV IURP D ZHOOSODWH ZHUHFRDWHGZLWKKXPDQ (6&TXDOLILHG
0DWULJHO &RUQLQJ IRU PLQ DW ƒ& 7KH FRORQLHV ZHUH PDQXDOO\ SLFNHG XQGHU D WLVVXH
FXOWXUH KRRG FRQWDLQLQJ D GLVVHFWLQJ PLFURVFRSH XVLQJ D 3 3LSHWPDQ DQG HDFK ZDV
WUDQVIHUUHGLQWRDQHZZHOORIWKHSUHYLRXVO\SUHSDUHGZHOOSODWHLQKXPDQL36&PHGLXPRU
LQ 1XWULVWHP FXOWXUH PHGLXP 6WHPJHQW &DPEULGJH 0$ 86$ VXSSOHPHQWHG ZLWK XQLWVPOSHQLFLOOLQDQG—JPOVWUHSWRP\FLQIRUIHHGHUIUHHFXOWXUH
7UDQVLHQWWUDQVIHFWLRQ
,QRUGHUWRREWDLQLQWHJUDWLRQIUHHKXPDQL36&VZHSHUIRUPHGDWUDQVLHQWWUDQVIHFWLRQ
E\HOHFWURSRUDWLQJWKUHHHSLVRPDOSODVPLGYHFWRUVFDUU\LQJJHQHVIRUWKHIRXUUHSURJUDPPLQJ
IDFWRUV2&7./)62;DQGO0<&DQGDSVKRUWKDLUSLQ51$VK51$LQWRWKH$7DQG
FRQWURO SULPDU\ GHUPDO ILEUREODVWV DGDSWLQJ WKH SURWRFRO GHVFULEHG LQ 2NLWD HW DO 0DWHULDOVDQG0HWKRGV
)LEUREODVWV ZHUH HOHFWURSRUDWHG XVLQJ WKH $PD[D 1XFOHRIHFWRU 7HFKQRORJ\ /RQ]D ZKLFK
DOORZVWKHWUDQVIHFWLRQRIQXFOHLFDFLGVLQWRERWKWKHF\WRSODVPDQGQXFOHXVRIFHOOVDQGLVEDVHG
RQDQRSWLPL]HGFRPELQDWLRQRIHOHFWULFDOSDUDPHWHUVDQGFHOOW\SHVSHFLILFVROXWLRQVWRJHWKHU
ZLWK WKH $PD[D +XPDQ 'HUPDO )LEUREODVW 1XFOHRIHFWRU .LW /RQ]D 7KH SODVPLGV ZH
HPSOR\HG ZHUH WKH IROORZLQJ S&;/(K2&7VKS) $GGJHQH SODVPLG FDUU\LQJ
2&7DQGVKSS&;/(K6.$GGJHQHSODVPLGFDUU\LQJ62;DQG./)DQGS&;/(
K8/$GGJHQHSODVPLGFDUU\LQJO0<&
)LEUREODVWV ZHUH WUDQVIHFWHG RQ GD\ ZKHQ WKH\ UHDFKHG FRQIOXHQF\ LQ D 7
IODVN)RUHDFKFHOOVDPSOHWREHHOHFWURSRUDWHG—O1XFOHRIHFWRU6XSSOHPHQWIURPWKH$PD[D
+XPDQ 'HUPDO )LEUREODVW 1XFOHRIHFWRU .LW ZDV DGGHG WR —O 1XFOHRIHFWRU 6ROXWLRQ
0RUHRYHUDFPGLVKZDVILOOHGZLWKPORIKXPDQILEUREODVWPHGLXPZLWKRXWSHQLFLOOLQRU
VWUHSWRP\FLQ DQG HTXLOLEUDWHG LQ WKH LQFXEDWRU )LEUREODVWV ZHUH KDUYHVWHG E\ WU\SVLQL]DWLRQ
FHQWULIXJHG DQG FRXQWHG )RU HDFK VDPSOH [ FHOOV ZHUH WUDQVIHUUHG WR D PO WXEH DQG
FHQWULIXJHGDWUSPIRUPLQ7KHFHOOSHOOHWZDVUHVXVSHQGHGLQWKHSUHYLRXVO\SUHSDUHG
—O6XSSOHPHQW1XFOHRIHFWRU6ROXWLRQPL[DQG—J'1$—JSODVPLGZDVDGGHGWRWKH
FHOO VXVSHQVLRQ 7KH VXVSHQVLRQ ZDV WUDQVIHUUHG WR D FXYHWWH ZKLFK ZDV LQVHUWHG LQWR WKH
1XFOHRIHFWRU GHYLFH 7KH SURJUDP 8 ZDV VHOHFWHG DQG DSSOLHG 2QFH WKH SURJUDP ZDV
ILQLVKHG—ORISUHHTXLOLEUDWHGPHGLXPZDVDGGHGWRWKHFXYHWWH7KHVDPSOHZDVJHQWO\
WUDQVIHUUHG LQWR WKH FP GLVK DQG LQFXEDWHG LQ D 2 LQFXEDWRU DV PLOG K\SR[LD LV WKH
RSWLPDO FRQGLWLRQ IRU ERWK WKH JHQHUDWLRQ DQG SUROLIHUDWLRQ RI L36&V 7KH PHGLXP ZDV
UHSODFHGWKHIROORZLQJGD\ZLWKIUHVKILEUREODVWPHGLXPDQGHYHU\RWKHUGD\WKHUHDIWHUIRU
GD\V
1H[W WKH WUDQVIHFWHG FHOOV ZHUH VKLIWHG RQWR D 30()&)/ IHHGHU OD\HU LQ D PHGLXP
DSSURSULDWH IRUKXPDQ L36&V 7R GR WKLVWKUHH FP GLVKHV RILQDFWLYDWHG 30()&)/IHHGHU
OD\HUV ZHUH SUHSDUHG IRU HDFK ILEUREODVW OLQH DV GHVFULEHG LQ VHFWLRQ WR VXSSRUW WKH FHOOV
0DWHULDOVDQG0HWKRGV
EHLQJUHSURJUDPPHG7KHPHGLXPZDVUHPRYHGIURPWKHGLVKHVFRQWDLQLQJWKHIHHGHUOD\HUV
DQGDIWHUULQVLQJZLWK3%6LWZDVUHSODFHGZLWKKXPDQILEUREODVWPHGLXP7KHHOHFWURSRUDWHG
ILEUREODVWVZHUHKDUYHVWHGE\WU\SVLQL]DWLRQFHQWULIXJHGFRXQWHGDQG[FHOOVZHUHSODWHG
LQ HDFK FP GLVK LQILEUREODVW PHGLXP 7KHQH[W GD\ WKH PHGLXP ZDV UHSODFHG ZLWK PO
KXPDQL36&PHGLXPZKLFKZDVFKDQJHGHYHU\RWKHUGD\
)LQDOO\ DIWHU DERXW WZR ZHHNV KXPDQ L36& FRORQLHV EHJDQ WR DSSHDU DQG ZHUH
PDQXDOO\SLFNHGXQGHUDWLVVXHFXOWXUHKRRGFRQWDLQLQJDGLVVHFWLQJPLFURVFRSHXVLQJD3
3LSHWPDQ (DFK ZDV WUDQVIHUUHG LQWR D QHZZHOO RIDKXPDQ (6&TXDOLILHG 0DWULJHOFRDWHG
ZHOOSODWHLQ1XWULVWHPFXOWXUHPHGLXPVXSSOHPHQWHGZLWKXQLWVPOSHQLFLOOLQ—JPO
VWUHSWRP\FLQQJPOOHXNDHPLDLQKLELWRU\IDFWRU/,)0LOOLSRUHZKLFKKHOSVPDLQWDLQFHOOV
LQ D SOXULSRWHQW VWDWH DQG —0 UKRDVVRFLDWHG SURWHLQ NLQDVH 52&. LQKLELWRU <
&DUOR(UED0LODQ,WDO\ZKLFKLQKLELWVDSRSWRVLVRIQRQFOXVWHUHGL36FHOOV
0DLQWHQDQFHRIKXPDQLQGXFHGSOXULSRWHQWVWHPFHOOFXOWXUHV
$IHHGHU OD\HU RI LQDFWLYDWHG 30()&)/ ZDV HPSOR\HG IRU WKH PDLQWHQDQFH RIKXPDQ
L36&VXQWLODIHHGHUIUHHFXOWXUHFRQGLWLRQZDVRSWLPL]HG+XPDQL36&VZHUHSDVVDJHGZKHQ
WKH\ UHDFKHG FRQIOXHQFH DW D VSOLW UDWLR RI LQ ZHOOV RI D ZHOO SODWH 7R GR WKLV WKH
PHGLXP ZDV UHPRYHG IURP WKH ZHOO DQG FHOOVZHUH LQFXEDWHG ZLWK '0(0) FRQWDLQLQJ
PJPOFROODJHQDVH,96LJPDDWƒ&IRUPLQ:KHQWKHHGJHVRIWKHFRORQLHVEHJDQWRFXUO
WKHZHOOVZHUHVFUDSHGZLWKDPOSLSHWWHDQGFOXPSVRIFHOOVZHUHFROOHFWHGDQGWUDQVIHUUHGWR
D PO WXEH 7KH\ ZHUH FHQWULIXJHG DW USP IRU PLQ DQG VHHGHG LQ D QHZ SODWH RQ
LQDFWLYDWHG 30()&)/ LQ KXPDQ L36& PHGLXP )RU IHHGHUIUHH FXOWXUH KXPDQ L36&V ZHUH
VHHGHG RQ KXPDQ (6&TXDOLILHG 0DWULJHOFRDWHG SODWHV LQ 1XWULVWHP FXOWXUH PHGLXP
VXSSOHPHQWHGZLWKXQLWVPOSHQLFLOOLQDQG—JPOVWUHSWRP\FLQ
0DWHULDOVDQG0HWKRGV
$OWHUQDWLYHO\ KXPDQ L36&V ZHUH KDUYHVWHG XVLQJ $FFXWDVH FHOO GHWDFKPHQW VROXWLRQ
&DUOR (UED ZKLFK ZDV DGGHG WR WKH ZHOO DIWHU UHPRYDO RI WKH PHGLXP 7KH SODWH ZDV
LQFXEDWHG IRU PLQ XQWLO FHOOV KDG GHWDFKHG 7KH\ ZHUH FROOHFWHG LQ D PO WXEH DQG
FHQWULIXJHG DW USP IRU PLQ 7KH\ ZHUH WKHQ VHHGHG RQ KXPDQ (6&TXDOLILHG 0DWULJHO
FRDWHG SODWHV LQ 1XWULVWHP FXOWXUH PHGLXP VXSSOHPHQWHG ZLWK XQLWVPO SHQLFLOOLQ —JPOVWUHSWRP\FLQDQG—052&.LQKLELWRU%HWZHHQSDVVDJHVWKHPHGLXPZDVFKDQJHG
HYHU\RWKHUGD\
,QYLWURSOXULSRWHQF\DVVD\
)RUGLIIHUHQWLDWLRQLQWRGHULYDWLYHVRIDOOWKUHHSULPDU\JHUPOD\HUVKXPDQL36&VZHUH
FROOHFWHGXVLQJPJPOFROODJHQDVH,9DVSUHYLRXVO\GHVFULEHGFHQWULIXJHGDQGWUDQVIHUUHGWR
DJHODWLQFRDWHGSODWHLQ30()&)/PHGLXP0HGLXPZDVFKDQJHGHYHU\RWKHUGD\IRU
GD\VDQGFHOOVZHUHWKHQDQDO\]HG
*HQHUDWLRQ DQG GLIIHUHQWLDWLRQ RI KXPDQ QHXUDO SUHFXUVRU
FHOOV
7KH ILUVW VWHS IRU REWDLQLQJQHXUDOFHOOV ZDV WKHJHQHUDWLRQ RIKXPDQ HPEU\RLG ERGLHV
(%V (%V DUH WKUHHGLPHQVLRQDO DJJUHJDWHV ZKLFK UHFDSLWXODWH PDQ\ DVSHFWV RI FHOO
GLIIHUHQWLDWLRQGXULQJHDUO\PDPPDOLDQHPEU\RJHQHVLVDQGJLYHULVHWRPDWXUHFHOOVRIDOOWKUHH
JHUPOD\HUVDQGWKHUHIRUHUHSUHVHQWWKHLQLWLDOVWHSLQPDQ\QHXUDOLQGXFWLRQSURWRFROV
:KHQKXPDQL36&VUHDFKHGFRQIOXHQFHWKHPHGLXPZDVUHPRYHGIURPWKHZHOODQGFHOOV
ZHUHLQFXEDWHGLQ'0(0)FRQWDLQLQJPJPOFROODJHQDVH,96LJPD$OGULFKDWƒ&IRU
PLQ:KHQWKHHGJHVRIWKHFRORQLHVEHJDQWRFXUOWKHZHOOVZHUHVFUDSHGZLWKDPOSLSHWWH
DQGFOXPSVRIFHOOVZHUHFROOHFWHGDQGWUDQVIHUUHGWRDPOWXEH7KH\ZHUHFHQWULIXJHGDW
0DWHULDOVDQG0HWKRGV
USP IRU PLQ WKH VXSHUQDWDQW ZDV FDUHIXOO\ UHPRYHG DQG WKH FHOOV ZHUH UHVXVSHQGHG LQ
KXPDQ (% PHGLXP ZKLFK FRQVLVWHG RI KXPDQ L36& PHGLXP ZLWKRXW E)*) 7KH\ ZHUH WKHQ
SODWHGLQORZDWWDFKPHQWGLVKHV1XQF6LJPD$OGULFKIRUKXPDQ(%IRUPDWLRQLQVXVSHQVLRQ
FXOWXUH7KHH[DFWDPRXQWRIKXPDQ(%PHGLXPXVHGYDULHGGHSHQGLQJRQWKHQXPEHURIFHOOV
WKHFHOOVIURPRQHZHOORIDZHOOSODWHZHUHUHVXVSHQGHGLQPODQGSODWHGLQDPPGLVK
+DOI RI WKH PHGLXP ZDV FKDQJHG HYHU\ RWKHU GD\ WKH GLVK ZDV ODLG DW D ƒ DQJOH WR OHW WKH
KXPDQ (%V VLQN WKH PHGLXP ZDV DVSLUDWHG ZLWKRXW GLVWXUELQJ WKH DJJUHJDWHV DQG IUHVK
PHGLXPZDVDGGHG
7KHIROORZLQJVWHSFRQVLVWHGLQWKHJHQHUDWLRQRIQHXUDOURVHWWHV1HXUDOURVHWWHVFRQVLVW
RI QHXURSURJHQLWRUV DQG DUH UDGLDO DUUDQJHPHQWV RI FROXPQDU FHOOV WKDW H[SUHVV PDQ\ RI WKH
SURWHLQV H[SUHVVHG LQ QHXURHSLWKHOLDO FHOOV LQ WKH QHXUDO WXEH DQG GLIIHUHQWLDWH LQWR QHXURQV
ROLJRGHQGURF\WHV DQG DVWURF\WHV $IWHU GD\V RI VXVSHQVLRQ FXOWXUH KXPDQ (%V ZHUH
FROOHFWHG LQ D PO WXEH DQG OHW VLQN IRU PLQ 7KH VXSHUQDWDQW ZDV UHPRYHG DQG WKH
DJJUHJDWHV ZHUH UHVXVSHQGHG LQ KXPDQ L36& PHGLXP VXSSOHPHQWHG ZLWK 1
VXSSOHPHQW*LEFR/LIH7HFKQRORJLHVDQGSODWHGLQWRJURZWKIDFWRUUHGXFHG0DWULJHOFRDWHG
ZHOOVRIDZHOOSODWH7KHPHGLXPZDVFKDQJHGHYHU\RWKHUGD\IRUGD\VDVQHXUDOURVHWWHV
DSSHDUHG
)LQDOO\ KXPDQ QHXUDO SUHFXUVRU FHOOV 13&V ZHUH H[SDQGHG 1HXUDO URVHWWHV ZHUH
PDQXDOO\SLFNHGDQGFROOHFWHGLQDPOWXEHLQPO13&PHGLXPFRQVLVWLQJRI'0(0)
VXSSOHPHQWHGZLWKXQLWVPOSHQLFLOOLQ—JPOVWUHSWRP\FLQP0JOXWDPD[%
VXSSOHPHQW *LEFR /LIH 7HFKQRORJLHV 1 VXSSOHPHQW QJPO (*) DQG QJPO
E)*)6RFLHWj,WDOLDQD&KLPLFL5RPH,WDO\7KH\ZHUHPHFKDQLFDOO\GLVVRFLDWHGXVLQJDPO
SLSHWWHDQGSODWHGLQDJURZWKIDFWRUUHGXFHG0DWULJHOFRDWHG7IODVNLQPO13&PHGLXP
+XPDQ13&VZHUHSDVVDJHGZKHQWKH\UHDFKHGFRQIOXHQFHDWDVSOLWUDWLRRILQ7RU
7 IODVNV 7R GR WKLV WKH PHGLXP ZDV UHPRYHG DQGFHOOV ZHUH LQFXEDWHG ZLWK$FFXWDVH FHOO
0DWHULDOVDQG0HWKRGV
GHWDFKPHQWVROXWLRQDWƒ&IRUPLQ7KH\ZHUHWKHQFROOHFWHGLQDPOWXEHFHQWULIXJHG
DWUSPIRUPLQDQGVHHGHGLQDQHZJURZWKIDFWRUUHGXFHG0DWULJHOFRDWHGIODVNLQIUHVK
13&PHGLXP
,Q RUGHU WR REWDLQ WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQV SUROLIHUDWLQJ KXPDQ 13&V ZHUH
VHHGHGDWDFRQFHQWUDWLRQRI[FHOOVFPLQ13&PHGLXPZKLFKZDVUHSODFHGKODWHU
ZLWK 13& PHGLXP ZLWKRXW E)*) DQG (*) &HOOV ZHUH IHG HYHU\ GD\V WKHUHDIWHU /LNH
KXPDQ L36&V KXPDQ 13&V DQG QHXURQV ZHUH REWDLQHG DQG PDLQWDLQHG LQ 2 DV PLOG
K\SR[LDHQKDQFHVWKHSUROLIHUDWLRQDQGGLIIHUHQWLDWLRQRIKXPDQQHXUDOVWHPFHOOV
:HVWHUQEORWDQDO\VLV
:KROHFHOOH[WUDFWVDQGSURWHLQTXDQWLILFDWLRQ
)RUSURWHLQH[SUHVVLRQDQDO\VLVFHOOVZHUHKDUYHVWHGFHQWULIXJHGWUDQVIHUUHGWRDPO
WXEHZDVKHGRQFHLQ3%6DQGILQDOO\FHQWULIXJHGDWUSPIRUPLQDWƒ&7KHSHOOHWZDV
UHVXVSHQGHG LQ WZR YROXPHV RI O\VLV EXIIHU FRPSRVHG RI 0 7ULV²+&O S+ 6'6
DQGERLOHGIRUPLQ$QHTXDOYROXPHRIO\VLVEXIIHUFRQWDLQLQJ;SKRVSKDWDVHDQGSURWHDVH
LQKLELWRUVP0SKHQ\OPHWK\OVXOIRQ\OIOXRULGH—JPOSHSVWDWLQNDOOLNUHLQLQDFWLYDWLQJ
XQLWVPODSURWLQLQ—JPOOHXSHSWLQZDVWKHQDGGHGDQGO\VDWHVZHUHNHSWRQLFHIURPWKLV
PRPHQWRQ/\VDWHVZHUHVRQLFDWHGDWIRUVIRU'1$IUDJPHQWDWLRQDQGFHQWULIXJHGDW
USPIRUPLQ
7KH ILQDO VXSHUQDWDQW FRUUHVSRQGLQJ WR WKH WRWDO SURWHLQ H[WUDFW ZDV WUDQVIHUUHG WR D
QHZ PO WXEHDQGTXDQWLILHG XVLQJ WKH FRORULPHWULF 0LFUR %&$3URWHLQ$VVD\ .LW 7KHUPR
6FLHQWLILF3LHUFHDVIROORZVLQDZHOOSODWHD%6$VWDQGDUGRIPJPO7KHUPR6FLHQWLILF
3LHUFH ZDV XVHG WR FUHDWH D VWDQGDUG FRORU UHVSRQVH FXUYH E\ GLOXWLQJ JURZLQJ TXDQWLWLHV RI
%6$LQGLVWLOOHGZDWHUIURPWR—JZHOOWRDILQDOYROXPHRI—OZHOODQG—ORIHDFK
0DWHULDOVDQG0HWKRGV
VDPSOH WR EH TXDQWLILHG ZDV GLOXWHG LQ —O RI GLVWLOOHG ZDWHU 7KH DSSURSULDWH YROXPH RI
0LFUR %&$ZRUNLQJ UHDJHQW ZDV SUHSDUHG E\PL[LQJ UHDJHQW 0$ ZLWK UHDJHQW 0%
DQGUHDJHQW0&DQG—OZHUHDGGHGWRHDFKZHOORIWKHVWDQGDUGVDQGWKHVDPSOHV7KH
SODWHZDVLQFXEDWHGLQWKHGDUNIRUPLQDWURRPWHPSHUDWXUH$EVRUEDQFHZDVPHDVXUHGDW
QP RQ D *(1LRV SODWH UHDGHU 7HFDQ 7KHDEVRUEDQFH UHDGLQJ RI WKH EODQN VWDQGDUGZDV
VXEWUDFWHGIURPWKHUHDGLQJVRIDOORWKHULQGLYLGXDOVWDQGDUGVDQGVDPSOHV$VWDQGDUGFXUYH
ZDVSUHSDUHGE\SORWWLQJWKHEODQNFRUUHFWHGUHDGLQJIRUHDFK%6$VWDQGDUGYVLWVFRQWHQWLQ
—JDQGZDVXVHGWRGHWHUPLQHWKHSURWHLQFRQFHQWUDWLRQRIHDFKVDPSOH
6'6SDJHDQG:HVWHUQEORW
$ WRWDO RI —J RI SURWHLQV IRU HDFK VDPSOH ZHUH EURXJKW WR D WRWDO YROXPH RI —O LQ O\VLV
EXIIHUDQGǃPHUFDSWRHWKDQROZDVDGGHG7KHVDPSOHVZHUHWKHQERLOHGIRUPLQDQGVL]H
IUDFWLRQDWHG E\ 6'6²3$*( RQ D 1X3$*( SUHFDVW SRO\DFU\ODPLGH JHO RU DFFRUGLQJWRWKHPROHFXODUZHLJKWRIWKHSURWHLQVRILQWHUHVWLQ;7ULVDFHWDWH6'6RU;0(6
6'6 UXQQLQJ EXIIHU /LIH 7HFKQRORJLHV DFFRUGLQJ WR WKH JHO XVHG 5HVROYHG SURWHLQV ZHUH
HOHFWUREORWWHG DW 9 IRU K RQWR 39') PHPEUDQHV 0LOOLSRUH ZKLFK ZHUH SUHYLRXVO\
DFWLYDWHG E\ LQFXEDWLRQ LQ PHWKDQRO IRU PLQ DQG VXEVHTXHQWO\ LQ WUDQVIHU EXIIHU /LIH
7HFKQRORJLHVIRUPLQ
7KHPHPEUDQHVZHUHWKHQLQFXEDWHGRYHUQLJKWZLWKSULPDU\DQWLERGLHVGLOXWHGLQ
QRQIDW GU\ PLON GLVVROYHG LQ 3%6 FRQWDLQLQJ 7ZHHQ 3%67 LQ VHDOHG EDJV XVLQJ WKH
;%/273K\EULGL]DWLRQLQVWUXPHQW,VHQHW0LODQ,WDO\7KHIROORZLQJSULPDU\DQWLERGLHV
ZHUHXVHG
0DWHULDOVDQG0HWKRGV
$QWLERG\
+RVW
6XSSOLHU
DQWL2&7&ORQH&
0RXVH
6DQWD&UX]%LRWHFQRORJ\
DQWLS
0RXVH
6DQWD&UX]%LRWHFQRORJ\
DQWL&GFS
0RXVH
6DQWD&UX]%LRWHFQRORJ\
DQWLSZDI
0RXVH
6DQWD&UX]%LRWHFQRORJ\
DQWLǃWXEXOLQ,,,&ORQH*
0RXVH
6LJPD
DQWLV\QDSWRSK\VLQ
0RXVH
6LJPD
DQWLYLQFXOLQ
0RXVH
6LJPD
DQWLǃDFWLQ
0RXVH
6LJPD
DQWL75$
0RXVH
*LIWRI'U&KULVWLDQ8QJHU
DQWL62;
0RXVH
$EFDP
DQWL&'&$
0RXVH
$EFDP
DQWLF\FOLQ%&ORQH*16
0RXVH
%'3KDUPLQJHQ
DQWLDŽ+$;+$;S6HUFORQH-%:
0RXVH
0LOOLSRUH
DQWL36'
0RXVH
1HXUR0DE
DQWL6&*
0RXVH
1HXUR0DE
DQWLSDQ.&K,3
0RXVH
1HXUR0DE
DQWLV\QDSWRSK\VLQ
0RXVH
1HXUR0DE
DQWLQHVWLQ
0RXVH
&KHPLFRQ
DQWL0$3&ORQH$3
0RXVH
&KHPLFRQ
DQWL.$37,)ǃ&ORQH(
0RXVH
&HOO6LJQDOLQJ7HFKQRORJ\
DQWL&+.&ORQH'&6
0RXVH
0%/,QWO&RUS
DQWL7RS
0RXVH
%'%LRVFLHQFHV
DQWL$70&ORQH<
5DEELW
(SLWRPLFV
DQWL$70S6
5DEELW
5RFNODQG
0DWHULDOVDQG0HWKRGV
DQWLFOHDYHG3$53
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWLFDVSDVH
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWLFDVSDVH
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWL;5&&
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWL.$3S6
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWL&+.S7KU
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWLSS6HU
5DEELW
&HOO6LJQDOLQJ7HFKQRORJ\
DQWL*)$3
5DEELW
0LOOLSRUH
DQWL9*$7
5DEELW
6\QDSWLF6\VWHP
DQWL9*/87
5DEELW
6\QDSWLF6\VWHP
DQWLSROǃ
5DEELW
$EFDP
DQWL)(1
5DEELW
$EFDP
DQWL$3(
5DEELW
6DQWD&UX]%LRWHFKQRORJ\
DQWL60&
5DEELW
%HWK\O/DERUDWRULHV
DQWL60&S6HU
5DEELW
%HWK\O/DERUDWRULHV
DQWL.$3S6
5DEELW
%LRODEV
7KHPHPEUDQHVZHUHZDVKHGWKUHHWLPHVLQ3%67DQGLQFXEDWHGIRUPLQZLWKSHUR[LGDVH
FRQMXJDWHGJRDWDQWLPRXVHRUDQWLUDEELW,J*VHFRQGDU\DQWLERGLHV*(+HDOWKFDUHGLOXWHGLQ
QRQIDWGU\PLONGLVVROYHGLQ3%67)ROORZLQJWKUHHZDVKHVLQ3%67WKHSURWHLQVRILQWHUHVW
ZHUHGHWHFWHGXVLQJDQGDQ(&/ZHVWHUQEORWWLQJGHWHFWLRQV\VWHP7KHUPR6FLHQWLILF3LHUFHRQ
DXWRUDGLRJUDSKLFILOPV%DQGVZHUHDFTXLUHGZLWKDGLJLWDOVFDQQHU9LQFXOLQDQGǃDFWLQZHUH
XVHGDVORDGLQJFRQWUROV
0DWHULDOVDQG0HWKRGV
,PPXQRIOXRUHVFHQFH
)RU LPPXQRIOXRUHVFHQFH DQDO\VLV FHOOV ZHUH JURZQ RQ JODVV FRYHUVOLSV IL[HG LQ EXIIHUHGSDUDIRUPDOGHK\GHIRUPLQDWURRPWHPSHUDWXUHZDVKHGLQ3%6DQGSHUPHDELOL]HG
ZLWK 7ULWRQ ; LQ 3%6 IRU PLQ 7KH\ ZHUH WKHQ EORFNHG LQ 3%6 FRQWDLQLQJ QRUPDOJRDWVHUXP/LIH7HFKQRORJLHVIRUPLQDQGLQFXEDWHGRYHUQLJKWDWƒ&ZLWKSULPDU\
DQWLERGLHVGLOXWHGLQEORFNLQJVROXWLRQ7KHIROORZLQJSULPDU\DQWLERGLHVZHUHXVHG
$QWLERG\
+RVW
6XSSOLHU
DQWL2&7&ORQH&
0RXVH
6DQWD&UX]%LRWHFQRORJ\
DQWL66($
0RXVH
*LIWRI'U&KULVWLDQ8QJHU
DQWL75$
0RXVH
*LIWRI'U&KULVWLDQ8QJHU
DQWLǂ60$&ORQH$
0RXVH
6LJPD
DQWL62;&ORQH
0RXVH
5'6\VWHP
DQWLǃWXEXOLQ,,,&ORQH78-
0RXVH
&RYDQFH
DQWL1HVWLQ
0RXVH
&KHPLFRQ
DQWL0$3&ORQH$3
0RXVH
&KHPLFRQ
DQWL9LPHQWLQ
0RXVH
'$.2
DQWL*)$3
5DEELW
0LOOLSRUH
DQWL*$%$
5DEELW
6LJPD
DQWL.L
5DEELW
7KHUPR6FLHQWLILF
)ROORZLQJWKUHHZDVKHVZLWK3%6FHOOVZHUHLQFXEDWHGZLWK$OH[DIOXRUJUHHQHPLWWLQJRU
UHGHPLWWLQJ FRQMXJDWHG JRDW DQWLPRXVH ,J* RU DQWLUDEELW ,J* /LIH 7HFKQRORJLHV IRU
PLQ DW URRP WHPSHUDWXUH &HOOV ZHUH ZDVKHG WKUHH WLPHV LQ 3%6 FRXQWHUVWDLQHG ZLWK
—JPO'$3,IRUPLQDQGPRXQWHGRQJODVVVOLGHVZLWK3URORQJ*ROG$QWLIDGHPRXQWLQJ
0DWHULDOVDQG0HWKRGV
VROXWLRQ/LIH7HFKQRORJLHV)OXRUHVFHQWVLJQDOVZHUHGHWHFWHGXVLQJDIOXRUHVFHQFHPLFURVFRSH
HTXLSSHGZLWKD&&'FDPHUD
(OHFWURSK\VLRORJLFDOUHFRUGLQJVDQGDQDO\VLV
&HOOVZHUHXVHGIRUHOHFWURSK\VLRORJLFDOUHFRUGLQJVZLWKWKHZKROHFHOOFRQILJXUDWLRQRI
WKH SDWFKFODPS WHFKQLTXH GD\V DIWHU SODWLQJ 5HFRUGLQJV ZHUH SHUIRUPHG DW URRP
WHPSHUDWXUH XVLQJ D 0XOWLFODPS $ SDWFKFODPS DPSOLILHU DQG S&ODPS VRIWZDUH
0ROHFXODU'HYLFHV5HFRUGLQJVZHUHXVXDOO\VWDUWHGPLQDIWHUWKHUXSWXUHRIWKHPHPEUDQH
SDWFKWRDOORZLQWUDFHOOXODUGLDO\VLVZLWKWKHSLSHWWHVROXWLRQ([WHUQDOEDWKVROXWLRQFRQWDLQHG
P01D&OP01D+32P0JOXFRVHP00J62P0&D&OP0.&O
DQG P0 +(3(6 S+ ZLWK 1D2+ LQWHUQDO SLSHWWH VROXWLRQ FRQWDLQHG P0 .
JOXFRQDWHP0.&OP00J&OP0(*7$P0+(3(6P03FUHDWLQHP0
$731DP0*731DDQGP0/HXSHSWLQHS+ZLWK.2+&HOOFDSDFLWDQFHDQG
VHULHV UHVLVWDQFH HUURUV ZHUH FDUHIXOO\ FRPSHQVDWHG a WKURXJKRXW WKH H[SHULPHQW 7KH
UHPDLQLQJOLQHDUFDSDFLW\DQGOHDNDJHFXUUHQWVZHUHHOLPLQDWHGRQOLQHXVLQJD3VXEWUDFWLRQ
SDUDGLJP3LSHWWHUHVLVWDQFHZDVEHWZHHQDQG0ƺ)RUWKHUHFRUGLQJVRIWRWDOYROWDJH
JDWHGLRQFXUUHQWVVLJQDOVZHUHILOWHUHGDWN+]DQGVDPSOHGDWN+])RUWKHUHFRUGLQJV
RI SRVWV\QDSWLF FXUUHQWV VLJQDOV ZHUH ILOWHUHG DW N+] DQG VDPSOHG DW N+] :KHQ ZH
VZLWFKHG WKH DPSOLILHU WR FXUUHQWFODPS PRGH ZH DSSOLHG WKH EULGJH EDODQFH FRPSHQVDWLRQ
DQGKHOGWKHUHVWLQJSRWHQWLDODWP9E\LQMHFWLQJWKHDSSURSULDWHKROGLQJFXUUHQW1HXURQDO
ILULQJZDVUHFRUGHGLQMHFWLQJGHSRODUL]LQJFXUUHQWSXOVHVRILQFUHDVLQJDPSOLWXGH7KHQHXURQV
ZLWKXQVWDEOHUHVWLQJSRWHQWLDODQGRUXQVWDEOHILULQJZHUHGLVFDUGHG,QFXUUHQWFODPSPRGH
VLJQDOVZHUHILOWHUHGDWN+]DQGVDPSOHGDWN+]
0DWHULDOVDQG0HWKRGV
&RPHWDVVD\
7KH FRPHW DVVD\ RU VLQJOH FHOO JHO HOHFWURSKRUHVLV LV D PHWKRG IRU PHDVXULQJ '1$
GDPDJH LQ HXNDU\RWLF FHOOV 7KH DVVD\ GHSHQGV RQ WKH UHOD[DWLRQ RI VXSHUFRLOHG '1$ LQ
DJDURVHHPEHGGHGFHOOVDIWHUO\VLV'1$FRQWDLQLQJDEUHDNORVHVLWVVXSHUFRLOLQJDQGEHFRPHV
IUHH WR H[WHQG WRZDUG WKH DQRGH XQGHU HOHFWURSKRUHVLV IRUPLQJ FRPHWOLNH LPDJHV GHWHFWDEOH
XQGHU IOXRUHVFHQFH PLFURVFRS\ IROORZLQJ VWDLQLQJ RI '1$ 7KH LQWHQVLW\ RI WKH FRPHW WDLO
UHODWLYHWRWKHKHDGUHIOHFWVWKHQXPEHURI'1$EUHDNVDQGWKHH[WHQWRI'1$GDPDJHFDQEH
FDOFXODWHGDXWRPDWLFDOO\E\LPDJLQJVRIWZDUHV
7R SUHSDUH FRPHW VOLGHV WKH GD\ EHIRUH DQDO\VLV PLFURVFRSH JODVV VOLGHV ZHUH ZDVKHG
ZLWK PHWKDQRO DLU GULHG DQG LPPHUVHG LQ PROWHQ QRUPDOJHOOLQJWHPSHUDWXUH DJDURVH WR
REWDLQ WKH ILUVW DJDURVH OD\HU EHIRUH EHLQJ VWRUHG RYHUQLJKW DW ƒ& 7R HYDOXDWH '1$ VWUDQG
EUHDNV WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQDO FHOOV ZHUH LQFXEDWHG ZLWK —0 +2 6LJPD IRU
PLQRULUUDGLDWHGZLWK*\$IWHUWUHDWPHQWFHOOVZHUHFROOHFWHGZLWK$FFXWDVHUHGXFHGWRD
VLQJOHFHOOVXVSHQVLRQDQGUHVXVSHQGHGLQLFHFROG&DDQG0JIUHH3%6WRDFRQFHQWUDWLRQRI
[FHOOVPO$QDOLTXRWRIǍORIFHOOV[FHOOVZDVDGGHGWRǍORIPROWHQORZ
JHOOLQJWHPSHUDWXUHDJDURVHNHSWDWƒ&DQGZDVLPPHGLDWHO\VSUHDGRQWRWKHFRPHWVOLGHV
FRYHUHGZLWKDFRYHUVOLSDQGVWRUHGDWƒ&LQWKHGDUNWRDFFHOHUDWHJHOOLQJRIWKHDJDURVH$IWHU
PLQWKHFRYHUVOLSZDVJHQWO\UHPRYHGDQGDVHFRQGORZJHOOLQJWHPSHUDWXUHDJDURVHOD\HU
ZDV VSUHDG RQWR WKH VOLGHV 7KH VOLGHV ZHUH WKHQ WUDQVIHUUHG WR D SUHFKLOOHG O\VLV VROXWLRQ
FRPSRVHG RI 0 1D&O P0 ('7$ P0 7ULVEDVH P0 1D2+ S+ DQG 7ULWRQ;DGGHGMXVWEHIRUHXVLQJIRUKDWƒ&LQWKHGDUNWKHQSURFHVVHGDVIROORZV
0DWHULDOVDQG0HWKRGV
1HXWUDOFRPHWDVVD\
7KHQHXWUDOFRPHWDVVD\ZDVHPSO\HGIRUWKHGHWHFWLRQRI'6%VDVGHVFULEHGLQ&DOLQLHW
DO $IWHU WKH O\VLV VWHS WKH VOLGHV ZHUH ZDVKHG WZLFH ZLWK ; 7ULVERUDWH ('7$ EXIIHU
VROXWLRQ 7%( S+ IRU PLQ 7KH VOLGHV ZHUH SODFHG LQ D KRUL]RQWDO HOHFWURSKRUHVLV
FKDPEHUDQGFRYHUHGZLWK7%(EXIIHU(OHFWURSKRUHVLVZDVFDUULHGRXWDWWKHUDWHRI9FP
IRU PLQ 7KH VOLGHV ZHUH WKHQ UHPRYHG IURP WKH HOHFWURSKRUHVLV FKDPEHU DQG ZDVKHG LQ
GHLRQL]HGZDWHUIRUPLQ$IWHUZDVKLQJWKHVOLGHVZHUHLPPHUVHGLQSUHFKLOOHGPHWKDQROIRU
PLQ IRU FHOO IL[DWLRQ DQG DLU GULHG RYHUQLJKW '1$ ZDV VWDLQHG ZLWK ǍO RI PJPO
HWKLGLXP EURPLGH IRU PLQ WKHQ FRPHW VOLGHV ZHUH REVHUYHG XQGHU WKH PLFURVFRSH
)OXRUHVFHQW VLJQDOV ZHUH GHWHFWHG XVLQJ D IOXRUHVFHQFH PLFURVFRSH HTXLSSHG ZLWK D &&'
FDPHUD DQG FRPHW LPDJHV ZHUH DQDO\]HG ZLWK WKH &RPHW 6FRUH VRIWZDUH Y 7UL7HN
&RUSRUDWLRQ
$ONDOLQHFRPHWDVVD\
7KHQHXWUDOFRPHWDVVD\ZDVHPSO\HGIRUWKHGHWHFWLRQRIVLQJOHVWUDQGEUHDNV66%VDV
GHVFULEHGLQ&DOLQLHWDO7KHVOLGHVZHUHSODFHGLQDKRUL]RQWDOHOHFWURSKRUHVLVFKDPEHU
DQGDGHQDWXUDWLRQVWHSZDVSHUIRUPHGLQSUHFKLOOHGDONDOLQHHOHFWURSKRUHVLVVROXWLRQP0
1D2+ P0 ('7$ S+ ! DW ƒ& LQ WKH GDUN $IWHU PLQWKH VOLGHV ZHUH VXEMHFWHG WR
HOHFWURSKRUHVLVDW9FPP$IRUPLQ)ROORZLQJHOHFWURSKRUHVLVVOLGHVZHUHZDVKHG
IRU PLQ ZLWK FROG QHXWUDOL]DWLRQ EXIIHU 0 7ULV+&O S+ $IWHU ZDVKLQJ WKH VOLGHV
ZHUHLPPHUVHGLQSUHFKLOOHGPHWKDQROIRUPLQIRUFHOOIL[DWLRQDQGDLUGULHGRYHUQLJKW'1$
ZDV VWDLQHG ZLWK ǍO RI PJPO HWKLGLXP EURPLGH IRU PLQ WKHQ FRPHW VOLGHV ZHUH
REVHUYHG XQGHU WKH PLFURVFRSH )OXRUHVFHQW VLJQDOV ZHUH GHWHFWHG XVLQJ D IOXRUHVFHQFH
PLFURVFRSH HTXLSSHG ZLWK D &&' FDPHUD DQG FRPHW LPDJHV ZHUH DQDO\]HG ZLWK WKH &RPHW
6FRUHVRIWZDUHY7UL7HN&RUSRUDWLRQ
0DWHULDOVDQG0HWKRGV
0RGLILHGDONDOLQHFRPHWDVVD\
$PRGLILHGDONDOLQHFRPHWDVVD\ZDVHPSOR\HGIRUWKHLQGLUHFWGHWHFWLRQRI7RSFFVDV
GHVFULEHGLQ$ODJR]HWDO7HUPLQDOO\GLIIHUHQWLDWHGKXPDQ13&VZHUHLQFXEDWHGZLWK
—0 &37 6LJPD IRU PLQ &HOOV ZHUH ZDVKHG WKUHH WLPHV ZLWK 3%6 VXSSOHPHQWHG ZLWK P0 &37 DW URRP WHPSHUDWXUH &HOOV ZHUH ILQDOO\ VXVSHQGHG LQ 3%6 FRQWDLQLQJ P0 &37
VXSSOHPHQWHG ZLWK PJPO SURWHLQDVH . 6LJPD DQG NHSW DW URRP WHPSHUDWXUH &HOO
VXVSHQVLRQZDVPL[HGZLWKDQHTXDOYROXPHRIORZJHOOLQJWHPSHUDWXUHDJDURVH6LJPD
PDLQWDLQHG DW ƒ& WR EULQJ WKH ILQDO &37 FRQFHQWUDWLRQ WR P0 &HOO VXVSHQVLRQ ZDV
LPPHGLDWHO\ OD\HUHG RQWR IURVWHG JODVV VOLGHV SUHFRDWHG ZLWK DJDURVH FRYHUHG ZLWK D
FRYHUVOLSDQGPDLQWDLQHGLQWKHGDUNDWURRPWHPSHUDWXUHIRUPLQ7KHFRYHUVOLSZDVWKHQ
JHQWO\UHPRYHGDQGVOLGHVZHUHLPPHUVHGLQO\VLVVROXWLRQFRPSRVHGRIPJPOSURWHLQDVH
.P0&37P07ULV+&OP0('7$S+7ULWRQ;'062S+IRU
KDWƒ&/\VLVVROXWLRQZDVWKHQVXSSOHPHQWHGZLWK01D&OIRUDGGLWLRQDOKDWƒ&
&HOOV ZHUH LPPHGLDWHO\ LPPHUVHG LQ DONDOLQH HOHFWURSKRUHVLV EXIIHU FRPSRVHG RI P0
1D2+ P0 ('7$ '062 DQG HOHFWURSKRUHVLV ZDV FRQGXFWHG DW 9FP IRU PLQ
IROORZHGE\QHXWUDOL]DWLRQLQP07ULV+&OS+IRUK7KHVOLGHVZHUHWKHQLPPHUVHG
LQ SUHFKLOOHG PHWKDQRO IRU PLQ IRU FHOO IL[DWLRQ DQG DLU GULHG RYHUQLJKW '1$ ZDV VWDLQHG
ZLWKǍORIPJPOHWKLGLXPEURPLGHIRUPLQWKHQFRPHWVOLGHVZHUHREVHUYHGXQGHU
WKH PLFURVFRSH )OXRUHVFHQW VLJQDOV ZHUH GHWHFWHG XVLQJ D IOXRUHVFHQFH PLFURVFRSH HTXLSSHG
ZLWK D &&' FDPHUD DQG FRPHW LPDJHV ZHUH DQDO\]HG ZLWK WKH &RPHW 6FRUH VRIWZDUH Y
7UL7HN&RUSRUDWLRQ
0DWHULDOVDQG0HWKRGV
'HWHFWLRQRIDŽ+$;DQG%3QXFOHDUIRFL
)RU WKH DVVHVVPHQW RI QXFOHDU IRFL GD\ WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQV RQ
FRYHUVOLSVZHUHLUUDGLDWHGZLWK*\$IWHUPLQKDQGKFHOOVZHUHZDVKHGZLWKD
1D&O VROXWLRQ WR UHPRYH WKH PHGLXP DLU GULHG RYHUQLJKW IL[HG LQ EXIIHUHG
SDUDIRUPDOGHK\GHDQGSHUPHDELOL]HGIRUPLQDWƒ&LQDVROXWLRQFRPSRVHGRIP0+HSHV
S+P01D&OP00J&OP0VXFURVHDQG7ULWRQ&HOOVZHUHWKHQEORFNHGLQ
3%6FRQWDLQLQJ%6$DQG7ZHHQIRUPLQDQGODEHOHGZLWKDPRXVHDQWLDŽ+$;
DQWLERG\ FORQH -%: 8SVWDWH %LRWHFKQRORJ\ 1< 86$ RU D UDEELW DQWL%3 DQWLERG\
1% 1RYXV %LRORJLFDOV &DPEULGJH 8. RYHUQLJKW DW ƒ& )ROORZLQJ WKUHH ZDVKHV LQ
3%6 FHOOVZHUH LQFXEDWHG ZLWK$OH[D )OXRU JUHHQHPLWWLQJ FRQMXJDWHG JRDW DQWLPRXVH RU
DQWLUDEELW,J*VHFRQGDU\DQWLERGLHV/LIH7HFKQRORJLHVIRUPLQDWURRPWHPSHUDWXUH&HOOV
ZHUHZDVKHGWKUHHWLPHVLQ3%6FRXQWHUVWDLQHGZLWKXJPO'$3,IRUPLQDQGPRXQWHG
RQJODVVVOLGHVZLWK3URORQJ*ROG$QWLIDGHPRXQWLQJ)OXRUHVFHQWVLJQDOVZHUHGHWHFWHGXVLQJ
DIOXRUHVFHQFHPLFURVFRSHHTXLSSHGZLWKD&&'FDPHUD
,QYLWUREDVHH[FLVLRQUHSDLUDVVD\
7KHLQYLWUR%(5DVVD\ZDVHPSOR\HGIRUPHDVXULQJ%(5HIILFLHQF\DQGPRGHLQKXPDQ
13&V DQG WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQV )LUVW ZKROHFHOO H[WUDFWV ZHUH SUHSDUHG
$SSUR[LPDWHO\ [ H[SRQHQWLDOO\ JURZLQJFHOOV ZHUHKDUYHVWHG DQGJHQWO\ UHVXVSHQGHGLQ
EXIIHU , P0 7ULV S+ P0 .&O DW D FRQFHQWUDWLRQ RI [ FHOOVPO $Q HTXDO
YROXPHRIEXIIHU,,P07ULVS+P0.&OP0('7$JO\FHURO1RQLGHW
3P0'77DQGSURWHDVHLQKLELWRUVZDVDGGHG7KHFHOOVXVSHQVLRQZDVVWLUUHGE\URFNLQJ
IRU K DW ƒ& )ROORZLQJ FHQWULIXJDWLRQ DW USP IRU PLQ DW ƒ& WKH VXSHUQDWDQW ZDV
GLVSHQVHGLQWRDOLTXRWVDQGVWRUHGDWƒ&
0DWHULDOVDQG0HWKRGV
'1$VXEVWUDWHVZHUHWKHQSUHSDUHG&ORVHGFLUFXODU'1$PROHFXOHVFRQWDLQLQJDVLQJOH
OHVLRQ ZHUH FRQVWUXFWHG E\ SULPLQJ VLQJOHVWUDQGHG S*HP=I '1$ 3URPHJD ZLWK
WKH ROLJRQXFOHRWLGHV FRQWDLQLQJ WKH PRGLILHG EDVH RI LQWHUHVW XUDFLOH DQG WHWUDK\GURIXUDQ
7+) 7KHVH ROLJRQXFOHRWLGHV ZHUH >DŽ3@$73 · HQG ODEHOOHG 7KH LQ YLWUR '1$
V\QWKHVLV ZDV SHUIRUPHG E\ XVLQJ 7 '1$ SRO\PHUDVH KRORHQ]\PH VLQJOHVWUDQGHG '1$
ELQGLQJ SURWHLQ G173V DQG 7 '1$ OLJDVH 7KH SODVPLG '1$ FRQWDLQLQJ D VLQJOH XUDFLO
UHVLGXHZDVGLJHVWHGZLWK8'*WRSURGXFHDVLQJOHDEDVLFVLWH
)LQDOO\ UHSDLU RI WKH SODVPLG '1$ FRQWDLQLQJ WKH OHVLRQV S*(0$37+) ZDV
FRQGXFWHG DV GHVFULEHG LQ )URVLQD HW DO 5HDFWLRQ PL[WXUHV ǍO FRQWDLQHG P0
+(3(6.2+S+P0.&OP00J&OP0GLWKLRWKUHLWROX0RIHDFKG173
P0 $73 P0 SKRVSKRFUHDWLQH ǍJ RI FUHDWLQH SKRVSKRNLQDVH W\SH , 6LJPD JO\FHURO ǍJ RI ERYLQH VHUXP DOEXPLQ DQG —J FHOO H[WUDFWV $IWHU LQFXEDWLRQ DW ƒ& IRU
LQFUHDVLQJ SHULRGV RI WLPH WKH SODVPLG '1$ ZDV UHFRYHUHG DQG GLJHVWHG ZLWK UHVWULFWLRQ
HQ]\PHV 7KH GLJHVWLRQ SURGXFWV ZHUH HOHFWURSKRUHVHG RQ D GHQDWXULQJ SRO\DFU\ODPLGH
JHO 7KH UHSDLU SURGXFWV ZHUH YLVXDOL]HG DQG TXDQWLILHG E\ DXWRUDGLRJUDSK\ ,VWDQW ,PDJHU
3DFNDUGDQGWKHUHODWLYH\LHOGRIWKHGLIIHUHQWSODVPLGIRUPVZDVPHDVXUHG
5DSLGDSSURDFKWR'1$DGGXFWUHFRYHU\DVVD\
7KH UDSLG DSSURDFK WR '1$ DGGXFW UHFRYHU\ 5$'$5 DVVD\ DOORZV WKH
LPPXQRGHWHFWLRQ RI SURWHLQ'1$ DGGXFWV )LUVW RI DOO FHOOV ZHUH O\VHG DQG '1$SURWHLQ
FRYDOHQWFRPSOH[HV'3&&VZHUHLVRODWHGDVIROORZV1HXURQVWHUPLQDOO\GLIIHUHQWLDWHGIRU
GD\VLQZHOOSODWHVZHUHLQFXEDWHGZLWKǍ0&37IRUK)ROORZLQJWUHDWPHQWSDUWRIWKH
FHOOVZDVLPPHGLDWHO\O\VHGZKLOHSDUWZDVZDVKHGZLWK3%6DQGLQFXEDWHGLQIUHVKPHGLXP
IRUKEHIRUHO\VLV)RUWKHLVRODWLRQRI'3&&VFHOOVZHUHO\VHGLQWKHSODWHE\DGGLWLRQRIPO
O\VLV UHDJHQW FRPSRVHG RI 0 *7& P0 7ULV+&O S+ P0 ('7$ 7ULWRQ ;
0DWHULDOVDQG0HWKRGV
6DUNRV\ODQGGLWKLRWKUHLWRODQGSURFHVVHGDVIROORZV(DFKO\VDWHZDVFROOHFWHGLQ
WZR PO WXEHV FRQWDLQLQJ PO HDFK DQG QXFOHLF DFLGV DQG '3&&V ZHUH SUHFLSLWDWHG E\
DGGLWLRQRI—ORIHWKDQRODQGLQFXEDWLRQDWƒ&IRUPLQ7KHVROXWLRQVZHUHWKHQ
VRQLFDWHG DW IRU V DQG FHQWULIXJHG DW USP IRU PLQ 7KH VXSHUQDWDQW ZDV
GLVFDUGHGDQGWKHSUHFLSLWDWHZDVZDVKHGWZLFHLQHWKDQRODQGLPPHGLDWHO\UHVXVSHQGHG
LQ —O IUHVKO\ SUHSDUHG P0 1D2+ )RU FRPSOHWH VROXELOL]DWLRQ LW ZDV HVVHQWLDO QRW WR
DOORZWKHSHOOHWWRGU\5HFRYHUHG'1$ZDVTXDQWLILHGE\PHDVXULQJWKHDEVRUEDQFHDWQP
XVLQJD1DQR'URSFVSHFWURSKRWRPHWHU7KHUPR6FLHQWLILF6DPSOHVZHUHVWRUHGDWƒ&
1H[W '3&&V ZHUH LPPXQRGHWHFWHG 7KH GHVLUHG TXDQWLW\ RI SXULILHG '1$ IURP HDFK
VDPSOHZDVGLOXWHGLQ3%6WRDILQDOYROXPHRI—ODQGDSSOLHGWRDQLWURFHOOXORVHPHPEUDQH
%LR5DGXVLQJDYDFXXPVORWEORWPDQLIROG%LR5DG7KHPHPEUDQHZDVWKHQEORFNHGIRUK
LQ QRQIDW GU\ PLON GLVVROYHG 3%67 DQG LQFXEDWHG ZLWK D PRXVH PRQRFORQDO DQWLKXPDQ
7RS SULPDU\ DQWLERG\ %' %LRVFLHQFHV GLOXWHG LQ QRQIDW GU\ PLON RYHUQLJKW DW ƒ& LQ D
VHDOHGEDJXVLQJWKH;%/273K\EULGL]DWLRQLQVWUXPHQW7KHPHPEUDQHZDVZDVKHGWKUHH
WLPHV LQ 3%67 DQG LQFXEDWHG IRU PLQ ZLWK D JRDW DQWLPRXVH SHUR[LGDVHFRQMXJDWHG ,J*
VHFRQGDU\DQWLERG\*(+HDOWKFDUHGLOXWHGLQQRQIDWGU\PLON)ROORZLQJWKUHHZDVKHVLQ
3%67 7RS ZDV GHWHFWHG XVLQJ DQG DQ (&/ GHWHFWLRQ V\VWHP 7KHUPR 6FLHQWLILF 3LHUFH RQ
DXWRUDGLRJUDSKLF ILOPV6LJQDOVZHUH DFTXLUHG ZLWK D GLJLWDO VFDQQHU DQG VLJQDO LQWHQVLW\ZDV
TXDQWLILHG XVLQJ WKH ,PDJH4XDQW VRIWZDUH 6DPSOH ORDGLQJ ZDV QRUPDOL]HG EDVHG RQ '1$
FRQWHQW DV GHWHUPLQHG E\ '$3, IOXRUHVFHQFH 7KH SUHVHQFH RI FRQWDPLQDQW QRQFRYDOHQWO\
ERXQGSURWHLQVZDVH[FOXGHGE\LQFXEDWLRQZLWKDQDQWLǃDFWLQDQWLERG\
1HXURQDOYLDELOLW\DVVD\
1HXURQDO YLDELOLW\ ZDV HYDOXDWHG XVLQJ WKH &HOO 7LWHU*OR OXPLQHVFHQW FHOO YLDELOLW\
DVVD\ 3URPHJD ZKLFK DOORZV WR GHWHUPLQH WKH QXPEHU RI YLDEOH FHOOV LQ FXOWXUH EDVHG RQ
0DWHULDOVDQG0HWKRGV
TXDQWLWDWLRQRIWKH$73SUHVHQWDQLQGLFDWRURIPHWDEROLFDOO\DFWLYHFHOOVGLUHFWO\SURSRUWLRQDO
WR WKH QXPEHU RIFHOOV SUHVHQWLQFXOWXUH 1HXURQV WHUPLQDOO\ GLIIHUHQWLDWHG IRU GD\VLQ ZHOOSODWHVZHUHWUHDWHGZLWKLQFUHDVLQJGRVHVRI&37WUDEHFWHGLQ+2RU3DUDTXDWDOOIURP
6LJPD IRU K +2 ZDV UHPRYHG DIWHU PLQ 7KUHH ZHOOV ZHUH HPSOR\HG IRU HDFK
FRQFHQWUDWLRQ $IWHU WUHDWPHQW D YROXPH RI &HOO7LWHU*OR 5HDJHQW HTXDO WR WKH YROXPH RI FHOO
FXOWXUHPHGLXPSUHVHQWZDVDGGHGWRHDFKZHOO7KHFRQWHQWVZHUHWKHQPL[HGLQWKHGDUNDW
URRP WHPSHUDWXUH IRU PLQ RQ D VKDNHU WR LQGXFH FHOO O\VLV &HOOXODU OXPLQHVFHQFH ZDV
UHFRUGHGXVLQJD*(1LRVSODWHUHDGHU7HFDQ
&HOOF\FOHSKDVHDQDO\VLV
3ULPDU\ ILEUREODVWV KXPDQ L36&V DQG SUROLIHUDWLQJ DQG GLIIHUHQWLDWHG KXPDQ 13&V
JURZQLQDZHOOSODWHZHUHLUUDGLDWHGDW*\ZKHQWKH\UHDFKHGFRQIOXHQFH$IWHUK
FHOOVZHUHKDUYHVWHGDQGJHQWO\WULWXUDWHGWRREWDLQVLQJOHFHOOVXVSHQVLRQVZDVKHGZLWKFROG
3%6 FHQWULIXJHG DQG ILQDOO\ IL[HG ZLWK PO FROG HWKDQRO DGGHG GURSZLVH &HOOV ZHUH
VWRUHGDWƒ&DQGEHIRUHDQDO\VLVWKH\ZHUHFHQWULIXJHGDWUSPIRUPLQUHVXVSHQGHGLQ
PORI—JPO3%651$VH$LQFXEDWHGDWƒ&IRUPLQDQGWKHQVWDLQHGZLWK—JPO
SURSLGLXP LRGLGH 5DGLDWLRQLQGXFHG FHOO F\FOH SKDVH PRGLILFDWLRQV ZHUH H[DPLQHG E\ IORZ
F\WRIOXRULPHWU\ XVLQJ D )$&6&DOLEXU LQVWUXPHQW ILWWHG ZLWK D &HOO 4XHVW VRIWZDUH SDFNDJH
%HFWRQ'LFNLQVRQ
5HVXOWV
5HVXOWV
*HQHUDWLRQ DQG FKDUDFWHUL]DWLRQ RI KXPDQ LQGXFHG
SOXULSRWHQW VWHP FHOOV GHULYHG IURP DWD[LDWHODQJLHFWDVLD DQG
FRQWUROILEUREODVWV
+XPDQL36&OLQHVZHUHHVWDEOLVKHGIURPSULPDU\GHUPDOILEUREODVWVRIWZRXQUHODWHG$
7 SDWLHQWV DQG D KHDOWK\ FRQWURO &RORQLHV ZLWK HPEU\RQLF VWHP FHOOOLNH PRUSKRORJ\ ZHUH
H[SDQGHG RQ D 30()&)/ IHHGHU OD\HU )LJXUH $ OHIW :HVWHUQ EORW DQDO\VLV UHYHDOHG DV
H[SHFWHGDSRVLWLYHVLJQDOIRU$70LQILEUREODVWVDQGKXPDQL36&VIURPFRQWUROEXWQRWIURP
$7 FDVHV )LJXUH $ ULJKW 1R $70 SURWHLQ ZDV GHWHFWDEOH LQ 30()&)/ DV WKH DQWLERG\
XVHGZDVKXPDQVSHFLILF)XUWKHUPRUHRQO\FRQWURODQG$7KXPDQL36&OLQHVH[SUHVVHGWKH
SOXULSRWHQF\ PDUNHU 2&7 EXW QRW ILEUREODVWV RU 30()&)/ )LJXUH $ ULJKW
,PPXQRIOXRUHVFHQFHDQDO\VLVDWWHVWHGWKHSOXULSRWHQF\RIWKHKXPDQL36&VEHLQJSRVLWLYHIRU
2&7VWDJHVSHFLILFHPEU\RQLFDQWLJHQ66($DQG75$)LJXUH%
0RUHRYHUWKHKXPDQL36&VJDYHULVHWRWKHHPEU\RQLFGHULYDWLYHVHFWRGHUPHQGRGHUP
DQG PHVRGHUP ODEHOHG SRVLWLYH IRU ȕWXEXOLQ ,,, 62; DQG DOSKDVPRRWK PXVFOH DFWLQ ǂ
60$ UHVSHFWLYHO\ )LJXUH & $OVR WKURXJK (% JHQHUDWLRQ QHXUDO URVHWWH IRUPDWLRQ DQG
VXEVHTXHQW FXOWXUH LQ D PLWRJHQIUHH PHGLXP FRQWDLQLQJ 1 VXSSOHPHQW KXPDQ L36&V
GLIIHUHQWLDWHGLQWRPLFURWXEXOHDVVRFLDWHGSURWHLQ0$3QHXURQV)LJXUH'
5HVXOWV
)LJXUH
5HVXOWV
)LJ*HQHUDWLRQFKDUDFWHUL]DWLRQSOXULSRWHQF\YDOLGDWLRQDQGQHXURQDOGLIIHUHQWLDWLRQRIKXPDQL36&FRORQLHV
IURPFRQWURODQG$7SDWLHQWV,Q$UHSUHVHQWDWLYHLPDJHVRIQHZO\IRUPHGKXPDQL36&FRORQLHVEHIRUHSLFNLQJ
XSSHU SDQHO DQG DIWHU SLFNLQJ ERWWRP SDQHO 7KH KXPDQ L36& FRORQLHV ZHUH FKDUDFWHUL]HG E\ :HVWHUQ EORW
ULJKWWRHYDOXDWHWKHH[SUHVVLRQRIWKH$70SURWHLQDQGRIWKHSOXULSRWHQF\PDUNHU2&7DQGZHUHFRPSDUHG
ZLWK SULPDU\ ILEUREODVWV DQG 30()&)/ IHHGHUV 9LQFXOLQ ZDV XVHG DV D ORDGLQJ FRQWURO ,Q % KXPDQ L36&
FRORQLHV ZHUH ODEHOHG WR YLVXDOL]H WKH H[SUHVVLRQ RI WKH SOXULSRWHQF\ PDUNHUV 2&7 66($ DQG 75$
1XFOHL ZHUH FRXQWHUVWDLQHG ZLWK '$3, EOXH ,Q & FRQWURO DQG $7 KXPDQ L36&V ZHUH GLIIHUHQWLDWHG LQ YLWUR
LQWRWKHWKUHHJHUPOD\HUVIRUGD\V&HOOVZHUHODEHOHGZLWKDQWLERGLHVVSHFLILFIRUǂ60$PHVRGHUPPDUNHU
62; HQGRGHUP PDUNHU DQG ǃWXEXOLQ ,,, HFWRGHUP PDUNHU 1XFOHL ZHUH FRXQWHUVWDLQHG ZLWK '$3, EOXH
7KHDELOLW\RIKXPDQL36&VWRJHQHUDWHQHXURQDOFHOOVZDVFRQILUPHGWKURXJKWKHIRUPDWLRQRIIORDWLQJ(%VDQG
URVHWWHVDQGUHSUHVHQWDWLYHLPDJHVIRUHDFKGLIIHUHQWLDWLRQVWHSDUHVKRZQLQ'
*HQHUDWLRQRIQHXUDOSUHFXUVRUFHOOVDQGQHXURQVIURPDWD[LD
WHODQJLHFWDVLDDQGFRQWUROKXPDQLQGXFHGSOXULSRWHQWVWHPFHOOV
7R JHQHUDWH SUROLIHUDWLQJ 13&V KXPDQ L36&GHULYHG URVHWWHV ZHUH FXOWXUHG LQ D
PHGLXP FRQWDLQLQJ % 1 (*) DQG E)*) DV GHSLFWHG LQ )LJXUH $ 3KHQRW\SLFDOO\ ERWK
FRQWURO DQG $7 KXPDQ 13&V ZHUH QHJDWLYH IRU WKH SOXULSRWHQF\ PDUNHUV 75$ DQG
2&7 )LJXUH % OHIW DQG SRVLWLYH IRU WKH QHXUDO SUHFXUVRU PDUNHUV 62; QHVWLQ DQG
YLPHQWLQ DQG IRU WKH SUROLIHUDWLRQ PDUNHU .L )LJXUH % ULJKW 2Q :HVWHUQ EORWV KXPDQ
13&VVKRZHGORZHUH[SUHVVLRQRI75$DQG2&7WKDQKXPDQL36&VDQGKLJKHUOHYHOV
5HVXOWV
RI62;DQGQHVWLQZKLFKWKHQGHFUHDVHGGXULQJWHUPLQDOQHXURQDOGLIIHUHQWLDWLRQREWDLQHGE\
ZLWKGUDZDORIWKHJURZWKIDFWRUVIRUDWOHDVWGD\V)LJXUH%ERWWRPOHIW
7R GHWHUPLQH WKH FDSDFLW\ RI KXPDQ 13&V WR DFWLYDWH WKH '1$ GDPDJH UHVSRQVH ZH
LQYHVWLJDWHGWKHWLPHGHSHQGHQWUHVSRQVHRIFRQWURODQG$713&VWR,5ZKLFKLQGXFHV'6%V
DQG $70 VLJQDOOLQJ 8VLQJ SKRVSKRVSHFLILF DQWLERGLHV IRU WKH $70 VXEVWUDWHV 60&
S6 .$3S6 &+.S7 SS6 DQG DŽ+$; DQG IRU WKH &+. VXEVWUDWH .$3
S6ZHIRXQGWKDWDIWHUPLQRI,5WKHVHWDUJHWPROHFXOHVZHUHVWURQJO\SKRVSKRU\ODWHG
LQFRQWUROEXWQRWLQ$713&V)LJXUH&,QWHUHVWLQJO\DIWHU,5DQDFFXPXODWLRQRIFOHDYHG
SRO\$'3ULERVHSRO\PHUDVH3$53DPDUNHURIDSRSWRVLVZDVGHWHFWHGLQFRQWUROEXWQRWLQ
$7 KXPDQ 13&V VXJJHVWLQJ WKDW $70 GHILFLHQF\ FRQIHUV UDGLRUHVLVWDQFH LQ SUROLIHUDWLYH
13&V)LJXUH&
:KHQ KXPDQ 13&V ZHUH LQGXFHG WR GLIIHUHQWLDWH WKH\ DFTXLUHG D QHXURQDOOLNH
PRUSKRORJ\ DQG DW GD\ RI GLIIHUHQWLDWLRQ ' ZHUH SRVLWLYH IRU ȕWXEXOLQ ,,, DQG 0$3
ZLWK D IUDFWLRQ RI FHOOV DOVR H[SUHVVLQJ WKH PDUNHU RI LQKLELWRU\ QHXURQV DŽDPLQREXW\ULF DFLG
*$%$)LJXUH'ZKLFKZHZHUHQRORQJHUDEOHWRGHWHFWDWGD\RIGLIIHUHQWLDWLRQ'LQ
PDWXUHQHXURQDOFHOOVH[SUHVVLQJȕWXEXOLQ,,,RU0$3)LJXUH(
:HVWHUQEORWDQDO\VLVFRQILUPHGWKHGRZQUHJXODWLRQRIWKHVWHPFHOOPDUNHUQHVWLQDQG
WKHXSUHJXODWLRQRIWKHQHXURQDOPDUNHUȕWXEXOLQ,,,DWGD\RIGLIIHUHQWLDWLRQ')LJXUH
)OHIWDQGDW'DQG')LJXUH)ULJKWDQGWKHH[SUHVVLRQRIWKHSRVWV\QDSWLFPDUNHU
SRVWV\QDSWLF GHQVLW\ SURWHLQ 36' RI WKH LQKLELWRU\ V\QDSVH PDUNHU YHVLFXODU *$%$
WUDQVSRUWHU 9*$7 RI WKH QHXURQDO JURZWKDVVRFLDWHG SURWHLQ 6&* DQG RI SRWDVVLXP
FKDQQHOLQWHUDFWLQJ SURWHLQV .&K,3V $W ' DQG ' EXW QRW ' FHOOV DOVR H[SUHVVHG WKH
SUHV\QDSWLFPDUNHUV\QDSWRSK\VLQ6<3:HGHWHFWHGQRREYLRXVGLIIHUHQFHVEHWZHHQFRQWURO
DQG$7KXPDQ13&VFRQFHUQLQJWKHJHQHUDWLRQRIQHXURQDOFHOOVEXW$7QHXURQVGLVSOD\HG
5HVXOWV
GHILFLWV LQ WKH H[SUHVVLRQ RI 6<3 DQG 36' DV ZHOO DV RI 6&* DQG .&K,3 DQG KLJKHU
DPRXQWVRIJOLDOILEULOODU\DFLGLFSURWHLQ*)$3)LJXUH)
7KHSRVWPLWRWLFVWDWXVRIWKHGLIIHUHQWLDWHGFHOOVZDVFRQILUPHGE\WKHGRZQUHJXODWLRQ
LQ ' ' )LJXUH * DQG ' VDPSOHV )LJXUH + RI &'& DQG F\FOLQ % ZKLFK DUH
SURWHLQV LQYROYHG LQ WKH * FHOO F\FOH SKDVH 7R QRWH WKH H[SUHVVLRQ RI WKH DSRSWRVLV PDUNHU
FOHDYHG 3$53 ZDV FRQVLVWHQWO\ KLJKHU LQ FRQWURO WKDQ LQ $7 QHXURQDO FHOOV DQG DŽ+$; ZDV
DOVRKLJKO\H[SUHVVHGDW'DQGLWVOHYHOVZHUHORZHULQ$7WKDQLQFRQWUROFHOOV)LJXUH*
)LJXUH
5HVXOWV
5HVXOWV
5HVXOWV
)LJ*HQHUDWLRQRIKXPDQ13&VIURPFRQWURODQG$7KXPDQL36&V,Q$WKHSURWRFROZHIROORZHGWRREWDLQ
VWDEOHDQGSUROLIHUDWLQJKXPDQ13&VLVGHSLFWHGDQGDUHSUHVHQWDWLYHLPDJHRIWKHFHOOFXOWXUHREWDLQHGLVVKRZQ
,Q%WKHFKDUDFWHUL]DWLRQRIKXPDQ13&VE\LPPXQRIOXRUHVFHQFHGHPRQVWUDWHVORVVRIWKHSOXULSRWHQF\PDUNHUV
75$ DQG 2&7 H[SUHVVLRQ RI WKH QHXUDO PDUNHUV 62; QHVWLQ DQG YLPHQWLQ DQG SUROLIHUDWLRQ DELOLW\
.L$FRPSDUDWLYHDQDO\VLVEHWZHHQKXPDQL36&VSUROLIHUDWLQJKXPDQ13&VDQGGLIIHUHQWLDWHG'KXPDQ
13&VZDVSHUIRUPHGE\:HVWHUQEORW,Q&SUROLIHUDWLQJKXPDQ13&VZHUHH[SRVHGWRLRQL]LQJUDGLDWLRQ*\
DQGWKHWLPHGHSHQGHQWDFWLYDWLRQRIWKH'1$GDPDJHUHVSRQVHZDVHYDOXDWHGE\DQDO\]LQJWKHSKRVSKRU\ODWLRQ
RI WKH LQGLFDWHG $70 VXEVWUDWHV 7KH H[SUHVVLRQ RI FOHDYHG 3$53 ZDV DOVR DQDO\]HG ȕDFWLQ ZDV XVHG DV D
ORDGLQJ FRQWURO ,Q ' WKH UHDO FDSDFLW\ RI KXPDQ 13&V WR GLIIHUHQWLDWH LQWR QHXURQV LV VKRZQ ZKHUH D KLJK
QXPEHURI0$3ȕWXEXOLQ,,,DQG*$%$FHOOVDUHGHWHFWHGDWDQGLQ(DWGD\VRIGLIIHUHQWLDWLRQ,Q)
KXPDQ13&VDQGQHXURQVDWRUGD\VRIGLIIHUHQWLDWLRQZHUHFROOHFWHGDQGDQDO\]HGE\:HVWHUQEORWIRU
WKHH[SUHVVLRQRIGLIIHUHQWLDWLRQDQGPDWXUDWLRQPDUNHUVDQGLQ*DQG+IRUWKHLQGLFDWHGSURWHLQV
(OHFWURSK\VLRORJLFDOFKDUDFWHUL]DWLRQ
&RQWURODQG$7QHXURQVDW'“ZHUHDQDO\]HGE\ZKROHFHOOSDWFKFODPSUHFRUGLQJV
LQ RUGHU WR YHULI\ WKHLU IXQFWLRQDO PDWXUDWLRQ 9ROWDJH FODPS H[SHULPHQWV VKRZHG VLPLODU
YROWDJHJDWHG SRWDVVLXP FXUUHQWV )LJXUH $ DQG PRVW LPSRUWDQWO\ YROWDJHJDWHG VRGLXP
FXUUHQWV VHOHFWLYHO\ EORFNHG E\ WHWURGRWR[LQ 77; ZKLFK LV D VSHFLILF IHDWXUH RI PDWXUH
5HVXOWV
QHXURQV)LJXUH$ULJKWSDQHO7UDQVLHQWSHDNVRGLXPLQZDUGFXUUHQWVKDGVLPLODUDPSOLWXGH
LQ FRQWURO DQG $7 QHXURQV PD[LPDO SHDN FXUUHQW GHQVLW\ ZDV “ S$S) Qõ LQ
FRQWURO QHXURQV DQG “ S$S) Qõ LQ $7 QHXURQV )LJXUH % 0RUHRYHU FXUUHQW
FODPS H[SHULPHQWV VKRZHG WKDW ERWK FRQWURO DQG $7 QHXURQV FRXOG JHQHUDWH GLVFKDUJHV RI
DFWLRQSRWHQWLDOVLQUHVSRQVHWRLQMHFWLRQRIGHSRODUL]LQJFXUUHQWVWHSV)LJXUH&
)LQDOO\ WR GHWHUPLQH ZKHWKHU QHXURQV GHYHORSHG IXQFWLRQDO V\QDSVHV ZH SHUIRUPHG
YROWDJH FODPS H[SHULPHQWV IRU UHFRUGLQJ VSRQWDQHRXV H[FLWDWRU\ DQG LQKLELWRU\ SRVWV\QDSWLF
FXUUHQWVV(36&VDQGV,36&V%RWKFRQWURODQG$7QHXURQVH[KLELWHGV(36&V)LJXUH'ZLWK
VLPLODU IUHTXHQFLHV “ +] Qõ FHOOV DQG “ +] Qõ FHOOV UHVSHFWLYHO\ ZKLFK
ZHUHEORFNHGE\DSSOLFDWLRQRIN\QXUHQLFDFLGP0GDWDQRWVKRZQEXWQRV,36&V)LJXUH
(
)LJXUH
5HVXOWV
)LJ )XQFWLRQDO FKDUDFWHUL]DWLRQ RI FRQWURO DQG $7 QHXURQV DIWHU GD\V RI GLIIHUHQWLDWLRQ ,Q $ OHIW SDWFK
FODPS ZKROHFHOO UHFRUGLQJV RI UHSUHVHQWDWLYH WRWDO LRQLF FXUUHQWV HOLFLWHG ZLWK GHSRODUL]LQJ YROWDJH VWHSV
EHWZHHQ²DQGP9P9LQFUHPHQWVIURPDKROGLQJSRWHQWLDORI²P9LQFRQWURODQG$7KXPDQ13&
GHULYHGQHXURQVVFDOHEDUS$PV,Q$ULJKWUHSUHVHQWDWLYHFXUUHQWVHOLFLWHGZLWKDGHSRODUL]LQJYROWDJH
VWHS WR P9 LQ FRQWURO EODFN GXULQJ SHUIXVLRQ ZLWK 77; P0 UHG DQG DIWHU ZDVKRXW EOXH LQ FRQWURO
XSSHUSDQHOVDQG$7ORZHUSDQHOVKXPDQ13&GHULYHGQHXURQVVFDOHEDUS$PV,Q%EDUJUDSKRI
PD[LPXP VRGLXP FXUUHQW GHQVLWLHV &' LQ FRQWURO DQG $7 KXPDQ 13&GHULYHG QHXURQV QR VWDWLVWLFDOO\
VLJQLILFDQWGLIIHUHQFH6WXGHQW·V7WHVW,Q&UHSUHVHQWDWLYHDFWLRQSRWHQWLDOGLVFKDUJHVUHFRUGHGLQFXUUHQWFODPS
GXULQJ LQMHFWLRQV RI V ORQJ GHSRODUL]LQJ FXUUHQW VWHSV IURP D KROGLQJ SRWHQWLDO RI P9 VFDOH EDU P9
PV,Q'WUDFHVVKRZLQJVSRQWDQHRXVH[FLWDWRU\SRVWV\QDSWLFFXUUHQWVUHFRUGHGDWWKHKROGLQJSRWHQWLDORI
P9 ZKLOH LQ ( WUDFHV VKRZLQJ WKH DEVHQFH RI VSRQWDQHRXV LQKLELWRU\ SRVWV\QDSWLF FXUUHQWV UHFRUGHG DW WKH
KROGLQJSRWHQWLDORIP9VFDOHEDUP9V
5HVXOWV
'1$ GDPDJH UHVSRQVH DQG UHSDLU LQ SRVWPLWRWLF DWD[LD
WHODQJLHFWDVLDQHXURQV
:H LQYHVWLJDWHG WKH $70GHSHQGHQW '1$ GDPDJH UHVSRQVH RI KXPDQ 13&GHULYHG
WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQV WKURXJK :HVWHUQ EORWWLQJ XVLQJ SKRVSKRVSHFLILF DQWLERGLHV
IRU$70VXEVWUDWHV,5WLPHFRXUVHDQDO\VLVRIFRQWUROQHXURQVDW'VKRZHGDYLJRURXV$70
GHSHQGHQWSKRVSKRU\ODWLRQRI60&6.$36&+.7S6DQGDŽ+$;ZKLOH
WKLVUHVSRQVHZDVUHSUHVVHGLQ$7QHXURQV)LJXUH$6LPLODUO\WKHSKRVSKRU\ODWLRQRIWKH
&+. VXEVWUDWH .$36 ZDV DEURJDWHG LQ $7 QHXURQV ,QWHUHVWLQJO\ DV SUHYLRXVO\
GHVFULEHGIRUSUROLIHUDWLQJKXPDQ13&V)LJXUH&KDIWHU,5DQDFFXPXODWLRQRIFOHDYHG
3$53ZDVGHWHFWHGLQFRQWUROEXWQRWLQ$7QHXURQVVXJJHVWLQJWKDW$70GHILFLHQF\FRQIHUV
VKRUWWHUP UDGLRUHVLVWDQFH DOVR LQ WHUPLQDOO\ GLIIHUHQWLDWHG FHOOV )LJXUH$ XSSHU SDQHO
$OWRJHWKHU WKHVH UHVXOWV LQGLFDWH WKDW QRUPDO SRVWPLWRWLF QHXURQV DFWLYDWH WKH '1$ GDPDJH
UHVSRQVH DV HIILFLHQWO\ DV WKHLU SUROLIHUDWLQJ SUHFXUVRUV DQG WKDW $70 GHILFLHQF\ DEODWHV WKLV
UHVSRQVH
7KHDELOLW\RISRVWPLWRWLFQHXURQVWRUHSDLU'6%VLQGXFHGE\,5ZDVLQYHVWLJDWHGZLWK
WKH QHXWUDO FRPHW DVVD\ DQG DSSHDUHG GHIHFWLYH LQ $7 QHXURQV ZKLFK GLVSOD\HG PRUH XQUHSDLUHG OHVLRQV WKDQ FRQWURO FHOOV )LJXUH % 7KLV ILQGLQJ LV FRQVLVWHQW ZLWK D
GHIHFWLYH$70GHSHQGHQW'1$GDPDJHUHVSRQVHSDWKZD\DFWLYDWLRQLQUHVSRQVHWR'6%VDV
VKRZQLQ)LJXUH$
7RLQYHVWLJDWHWKHUROHRI$70LQWKHUHVROXWLRQRI,5LQGXFHG'6%VLQ'SRVWPLWRWLF
QHXURQVZHVFRUHGWKHWLPHGHSHQGHQWIRUPDWLRQDQGFOHDUDQFHRIDŽ+$;DQG%3QXFOHDU
IRFLE\LPPXQRIOXRUHVFHQFH8QLUUDGLDWHGFRQWURO'QHXURQVVKRZHGDEDVDOQXPEHURI
DŽ+$; DQG %3 IRFL SHU FHOO “ DQG “ ZKLFK LQFUHDVHG WR “ DQG “
UHVSHFWLYHO\ DIWHU ,5 WR GHFOLQH DIWHU K DQG HYHQ IXUWKHU DIWHU K )LJXUHV & DQG '
5HVXOWV
8QWUHDWHG $7 FHOOV VKRZHG D QXPEHU RI IRFL SHU FHOO FRPSDUDEOH WR FRQWURO FHOOV ZKLFK
LQFUHDVHGPRUHPRGHVWO\“IRUDŽ+$;DQG“IRU%3DIWHUPLQDQGSHUVLVWHG
DIWHUK)LJXUHV&DQG'7KHVHILQGLQJVLQGLFDWHWKDW$7SRVWPLWRWLFQHXURQVVKRZVORZHU
NLQHWLFVRI'6%UHSDLUWKDQFRQWUROQHXURQV
7KH DSRSWRWLF HIIHFW RI UDGLDWLRQ RQ FRQWURO DQG $7 QHXURQV K DIWHU *\ ,5 ZDV
DVVHVVHG E\ IORZ F\WRPHWU\ DQDO\VLV RI WKH VXEGLSORLG '1$ SHDN RQ '1$ KLVWRJUDPV
)LJXUH()HZHUDSRSWRWLFFHOOVZHUHGHWHFWHGLQ,5WUHDWHG$7FHOOVWKDQLQFRQWURO
FHOOVVXJJHVWLQJWKDW$70GHILFLHQF\DWWHQXDWHVVKRUWWHUPDSRSWRVLVFRQFRUGDQWZLWK
WKHILQGLQJVRQFOHDYHG3$53)LJXUH$XSSHUSDQHO
)LJXUH
5HVXOWV
5HVXOWV
)LJ $7 SRVWPLWRWLF QHXURQV DUH GHIHFWLYH LQ '1$ GDPDJH UHVSRQVH ,Q $ SRVWPLWRWLF QHXURQV ' ZHUH
WHVWHG E\ :HVWHUQ EORW ZLWK DQWLERGLHV VSHFLILF IRU WKH YDULRXV $70 SKRVSKRVXEVWUDWHV DQG FOHDYHG 3$53 DW
YDULRXVWLPHVDIWHUWUHDWPHQWZLWK*\LRQL]LQJUDGLDWLRQȕDFWLQZDVXVHGDVDORDGLQJFRQWURO,Q%'1$'6%V
ZHUH DQDO\]HG LQ ' QHXURQV E\ QHXWUDO FRPHW DVVD\ DW WZR GLIIHUHQW WLPHV DIWHU WUHDWPHQW 5HSUHVHQWDWLYH
SKRWRVRIFRPHWVIURPXQWUHDWHGFHOOVDQGDWGLIIHUHQWWLPHSRLQWVIROORZLQJWUHDWPHQWDUHVKRZQ7KHWLPHSRLQW
ZLWK WKH PD[LPXP '1$ GDPDJH ZDV FRQVLGHUHG PLQ DIWHU LRQL]LQJ UDGLDWLRQ WUHDWPHQW 7KH UDWLR EHWZHHQ
WUHDWHGDQGXQWUHDWHGWDLOPRPHQWVRIDWOHDVW²FHOOVSHUH[SHULPHQWDOSRLQWLVVKRZQLQWKHJUDSKVYDOXHV
LQ 2QH UHSUHVHQWDWLYH H[SHULPHQW RXW RI WKUHH LV VKRZQ ,Q & DQG ' IRUPDWLRQ DQG UHVROXWLRQ RI LRQL]LQJ
UDGLDWLRQLQGXFHG QXFOHDU IRFL ' QHXURQV ZHUH LUUDGLDWHG ZLWK *\ FROOHFWHG DW WKH LQGLFDWHG WLPHV DQG
ODEHOHG IRU DŽ+$; &DQG %3 ' )RUHDFK PDUNHU WKH QXPEHURI IRFL ZDV VFRUHG IURP FHOOQXFOHLSHU
GXSOLFDWHSUHSDUDWLRQVDQG IURP WKUHH LQGHSHQGHQW H[SHULPHQWV PHDQ “ 6' :KHUHLQGLFDWHG WKH GLIIHUHQFH
EHWZHHQFRQWURODQG$7ZDVVWDWLVWLFDOO\VLJQLILFDQW3 6WXGHQW·V7WHVW,Q('SRVWPLWRWLFFRQWURODQG
$7FHOOVZHUHWUHDWHGZLWK*\LRQL]LQJUDGLDWLRQFROOHFWHGDIWHUKDQGDQDO\]HGIRU'1$FRQWHQWE\IORZ
F\WRPHWU\7KHSHUFHQWDJHRIVXEGLSORLGDSRSWRWLFFHOOVLVLQGLFDWHG
%DVH H[FLVLRQ UHSDLU FDSDFLW\ LQ XQGLIIHUHQWLDWHG DQG
GLIIHUHQWLDWHG QHXUDO SUHFXUVRU FHOOV DQG UHSDLU RI VLQJOH VWUDQG
EUHDNVLQSRVWPLWRWLFQHXURQV
,Q QHXURQV WKH %(5 SDWKZD\ LV HVVHQWLDO WR UHPRYH R[LGDWLYH '1$ GDPDJH DQG 66%V
JHQHUDWHGE\KLJKOHYHOVRI526DQGGHIHFWVLQ%(5FRQWULEXWHWRQHXURGHJHQHUDWLRQ
5HVXOWV
7RGHWHUPLQHKRZ%(5LVUHJXODWHGLQSUHDQGSRVWPLWRWLFQHXURQVZHDQDO\]HGWKHHIILFLHQF\
RIWKHVKRUWSDWFK63DQGORQJSDWFK/3%(5VXESDWKZD\V$QDO\VLVRI63%(5UHYHDOHGQR
VLJQLILFDQW GLIIHUHQFHV EHWZHHQ FRQWURO DQG $7 KXPDQ 13&V ERWK LQ WKH SUROLIHUDWLRQ DQG
SRVWPLWRWLF'VWDJH)LJXUH$7REHWWHUXQGHUVWDQGWKLVUHVXOWZHDQDO\]HGWKHOHYHOVRI
WKH 63%(5 SURWHLQV DSXULQLFDS\ULGLQLF HQGRQXFOHDVH $3( '1$ SRO\PHUDVH ȕ SRO ȕ
DQG ;5&& $IWHU GD\V RI GLIIHUHQWLDWLRQ ' RQO\ WKH OHYHOV RI ;5&& ZHUH UHGXFHG
)LJXUH&6LPLODUILQGLQJVZHUHVHHQLQ'QHXURQV
1H[WZHDQDO\]HG/3%(5DFWLYLW\ZKLFKDSSHDUHGPDUNHGO\ORZHULQERWKFRQWURODQG
$7 SRVWPLWRWLF QHXURQV WKDQ LQ WKHLU UHVSHFWLYH SUROLIHUDWLQJ KXPDQ 13&V )LJXUH % 7KLV
ILQGLQJ LV FRQFRUGDQW ZLWK WKH UHGXFHG H[SUHVVLRQ RI IODS HQGRQXFOHDVH )(1 D SURWHLQ
LQYROYHG LQ /3%(5 LQ QRQUHSOLFDWLQJ FHOOV )LJXUH & 2YHUDOO WKHVH UHVXOWV SRLQW RXW
GLIIHUHQFHV WKDW GHSHQG RQ GLIIHUHQW OHYHOV RI H[SUHVVLRQ RU GLIIHUHQWLDO DFWLYDWLRQ RI %(5
HQ]\PHVLQXQGLIIHUHQWLDWHGDQGGLIIHUHQWLDWHGFHOOVEXWDSSHDULQGHSHQGHQWRI$70
0RUHRYHUZHLQYHVWLJDWHGWKHDELOLW\RISRVWPLWRWLFQHXURQVWRUHSDLU66%VLQGXFHGE\
+2 ZLWK WKH DONDOLQH FRPHW DVVD\ &RQVLVWHQW ZLWK WKH UHVXOWV RQ %(5 HIILFLHQF\ ZH
IRXQGWKDWFRQWURODQG$7QHXURQVGLVSOD\HGQRVLJQLILFDQWGLIIHUHQFHVLQWKHUHSDLUNLQHWLFVRI
WKLVW\SHRIGDPDJH)LJXUH'
5HVXOWV
)LJXUH
5HVXOWV
)LJ%DVHH[FLVLRQUHSDLU%(5DFWLYLWLHVLQSUROLIHUDWLQJDQGSRVWPLWRWLFFRQWURODQG$7QHXURQDOFHOOV,Q$
DQG%LQYLWURUHSDLUUHDFWLRQVZHUHSHUIRUPHGE\XVLQJZKROHFHOOH[WUDFWVDQGDVVXEVWUDWHD3ODEHOHGFLUFXODU
SODVPLGFRQWDLQLQJDVLQJOH$3VLWHS*(0$3WRPHDVXUHVKRUWSDWFK%(5$RUD7+)UHVLGXHS*(07+)
IRUORQJSDWFK%(5%7KHFRUUHFWLQVHUWLRQRIDVLQJOHOHVLRQLQWKHSODVPLGPROHFXOHVZDVFKHFNHGE\GLJHVWLRQ
ZLWK XUDFLO '1$ JO\FRV\ODVH 8'* IROORZHG E\ HQGRQXFOHDVH ,,, 17+ IRU S*(0$3 $ DQG E\ $3
HQGRQXFOHDVH+$3IRUS*(07+)%5HSDLUNLQHWLFVRIWKH$3VLWH$RU7+)UHVLGXH%E\SUROLIHUDWLQJ
5HVXOWV
KXPDQ13&VDQGSRVWPLWRWLFQHXURQV'ZHUHPHDVXUHGIROORZLQJLQFXEDWLRQRIWKHVLQJOHOHVLRQFRQWDLQLQJ
SODVPLGV ZLWK H[WUDFWV IURP FRQWURO RU $7 GHIHFWLYH QHXURQDO FHOOV IRU LQFUHDVLQJ SHULRGV RI WLPH 5HSDLU
SURGXFWVZHUHDQDO\]HGE\DJDURVHJHOHOHFWURSKRUHVLVDQGWKHUDGLRDFWLYLW\RIWKHEDQGVFRUUHVSRQGLQJWRQLFNHG
IRUP,,DQGVXSHUFRLOHGIRUP,SODVPLGVZDVTXDQWLILHG5HSDLUHIILFLHQF\LVH[SUHVVHGDVUHODWLYHDPRXQWRI
)RUP , RYHU WRWDO UDGLRDFWLYLW\ LQ HDFK ODQH ,Q & WKH H[SUHVVLRQ OHYHO RI VKRUW SDWFK%(5 DQG ORQJ SDWFK%(5
SURWHLQV ZDV HYDOXDWHG E\ :HVWHUQ EORW LQ SUROLIHUDWLQJ DQG SRVWPLWRWLF 13&V ȕDFWLQ ZDV XVHG DV D ORDGLQJ
FRQWURO ,Q ' '1$ 66%V ZHUH DQDO\]HG LQ ' QHXURQV E\ DONDOLQH FRPHW DVVD\ DW WZR GLIIHUHQW WLPHV DIWHU
WUHDWPHQW5HSUHVHQWDWLYHSKRWRVRIFRPHWVIURPXQWUHDWHGFHOOVDQGDWGLIIHUHQWWLPHSRLQWVIROORZLQJWUHDWPHQW
DUH VKRZQ 7KH WLPH SRLQW ZLWK WKH PD[LPXP '1$ GDPDJH ZDV FRQVLGHUHG PLQ DIWHU +2 WUHDWPHQW 7KH
UDWLR EHWZHHQ WUHDWHG DQG XQWUHDWHG WDLO PRPHQWV RIDW OHDVW ² FHOOV SHUH[SHULPHQWDO SRLQW LV VKRZQ LQ WKH
JUDSKVYDOXHVLQ2QHUHSUHVHQWDWLYHH[SHULPHQWRXWRIWKUHHLVVKRZQ
7UHDWPHQWRIDWD[LDWHODQJLHFWDVLDDQGFRQWUROQHXURQDOFHOOV
ZLWKJHQRWR[LFDQGR[LGDWLYHDJHQWV
$V JHQRWR[LF DJHQWV DQG R[LGDWLYH VWUHVV PD\ VLPLODUO\ DIIHFW $70GHILFLHQW QHXURQV
ZHLQYHVWLJDWHGWKHYLDELOLW\RIFRQWURODQG$7'SRVWPLWRWLFQHXURQVXSRQWUHDWPHQWZLWK
FRPSRXQGV WKDW ZRUN WKURXJK GLIIHUHQW PHFKDQLVPV &37 ZKLFK LQKLELWV 7RS DQG WUDSV
7RSFFV WUDEHFWHGLQ ZKLFKEORFNV WUDQVFULSWLRQ IDFWRU DFWLYLW\ 3DUDTXDWZKLFK LQGXFHV
526WKURXJKGHSRODUL]DWLRQRIWKHLQQHUPLWRFKRQGULDOPHPEUDQHDQG+2&RQWURODQG
$7 SRVWPLWRWLF QHXURQV ZHUH HTXDOO\ VHQVLWLYH WR WKH F\WRWR[LF HIIHFWV RI +2 DQG 3DUDTXDW
)LJXUHV$DQG%DILQGLQJFRQFRUGDQWZLWKWKHUHVXOWVLQ)LJXUH'VKRZLQJWKDWWKHVHFHOOV
KDYHWKHVDPHDELOLW\WRUHVROYH66%V%\FRQWUDVW$7QHXURQVZHUHVLJQLILFDQWO\PRUHUHVLVWDQW
WKDQFRQWUROQHXURQVWR&37DQGWUDEHFWHGLQE\DERXWDQGUHVSHFWLYHO\)LJXUHV&
DQG'
5HVXOWV
)LJXUH
)LJ$7SRVWPLWRWLFFHOOVDUHUHVLVWDQWWR&37DQGWUDEHFWHGLQWUHDWPHQW'QHXURQVJURZQLQZHOOSODWHV
ZHUH DVVHVVHG IRU FHOO YLDELOLW\ E\ &HOO7LWHU*OR K DIWHU H[SRVXUH WR +2 IRU PLQ $ RU FRQWLQXRXV
WUHDWPHQWZLWK3DUDTXDW%&37&RUWUDEHFWHGLQ'DWWKHLQGLFDWHGGRVHV$OOWUHDWPHQWVZHUHSHUIRUPHGLQ
WULSOLFDWHZHOOV7KHJUDSKVVKRZPHDQ“6'DQGZKHUHLQGLFDWHGWKHGLIIHUHQFHEHWZHHQFRQWURODQG$7ZDV
VWDWLVWLFDOO\VLJQLILFDQW3 3 6WXGHQW·V7WHVW
$FFXPXODWLRQRIWRSRLVRPHUDVH'1$FOHDYDJHFRPSOH[HVLQ
DWD[LDWHODQJLHFWDVLDSRVWPLWRWLFQHXURQV
$VLQPXULQHQHXUDOFHOOV$70GHILFLHQF\UHVXOWVLQDFFXPXODWLRQRI7RSFFVDQGIDLOXUH
WRUHFRYHUJOREDOWUDQVFULSWLRQDIWHU7RSFFWUDSSLQJZHXVHGDPRGLILHGDONDOLQHFRPHW
DVVD\ ZKLFK LQGLUHFWO\ TXDQWLILHV 7RSFFV WR DQDO\]H KXPDQ 13&GHULYHG ' QHXURQV
XQWUHDWHG RU WUHDWHG IRU PLQ ZLWK P0 &37 )LJXUH $ OHIW 7KH DFFXPXODWLRQ RI
5HVXOWV
7RSFFVZDVPXFKJUHDWHULQ$7WKDQLQFRQWUROQHXURQVERWKEHIRUHDQGDIWHU&37WUHDWPHQW
2I QRWH WKH RYHUDOO OHYHOV RI 7RS LQ SUROLIHUDWLQJ DQG SRVWPLWRWLF QHXURQV ZHUH VLPLODU
)LJXUH$ULJKW
7R IXUWKHU FRQILUP WKHVH ILQGLQJV ZH DSSOLHG WKH 5$'$5 DVVD\ ZKLFK DOORZV WR
GLUHFWO\ GHWHFW 7RSFFVERXQG WR JHQRPLF '1$ ' QHXURQV LQFXEDWHG IRU K ZLWK P0
&37ZHUHKDUYHVWHGLPPHGLDWHO\RUKDIWHUZDVKRXWRIWKHGUXJWRDQDO\]HWKHUHFRYHU\RI
WKHGDPDJH7KHOHYHOVRI7RSFFV)LJXUH%OHIWSDQHOZHUHWKHQQRUPDOL]HGZLWKWKHWRWDO
DPRXQWRI'1$UHYHDOHGE\'$3,)LJXUH%PLGGOHSDQHOȕDFWLQZDVXVHGWRYHULI\WKDW
WKH'1$VDPSOHVZHUHIUHHRIQRQFRYDOHQWO\ERXQGFRQWDPLQDWLQJSURWHLQV)LJXUH%ULJKW
SDQHO $V H[SHFWHG XQWUHDWHG $7 QHXURQV GLVSOD\HG KLJKHU DFFXPXODWLRQ RI 7RSFFV WKDQ
FRQWURO FHOOV DQG DOVR DSSHDUHG GHIHFWLYH LQ 7RSFF UHFRYHU\ DIWHU &37 UHPRYDO )LJXUH %
JUDSK
)LJXUH
5HVXOWV
)LJ $7 SRVWPLWRWLF QHXURQV DFFXPXODWH DEQRUPDO OHYHOV RI WRSRLVRPHUDVH '1$ FOHDYDJH FRPSOH[HV
7RSFFV,Q$WKHOHYHOVRIHQGRJHQRXV7RSFFVZHUHTXDQWLILHGLQFRQWURODQG$7'SRVWPLWRWLFQHXURQV
WUHDWHG ZLWK P0 &37 IRU PLQ E\ PRGLILHG DONDOLQH FRPHW DVVD\ $ OHIW )LIW\ FHOOV SHU VDPSOH SHU
H[SHULPHQWZHUHDQDO\]HG'DWDDUHWKHDYHUDJHRIWKUHHLQGHSHQGHQWH[SHULPHQWVPHDQ“6'7KHGLIIHUHQFH
EHWZHHQ FRQWURO DQG $7 ZDV VWDWLVWLFDOO\ VLJQLILFDQW 3 7RWDO LQWUDFHOOXODU OHYHOV RI 7RS SURWHLQ ZHUH
DQDO\]HGE\:HVWHUQEORWLQSUROLIHUDWLQJKXPDQ13&VDQGSRVWPLWRWLFQHXURQV$ULJKW,Q%FRQWURODQG$7
'SRVWPLWRWLFQHXURQDOFHOOVZHUHXQWUHDWHGRUWUHDWHGZLWKP0&37IRUPLQRULQFXEDWHGDQGKDUYHVWHG
KDIWHUZDVKLQJ'1$ZDVSXULILHGIURPWKHFHOOVDIWHUO\VLVZLWKDGHQDWXULQJEXIIHU7KHLQGLFDWHGDPRXQWRI
'1$ZDVYDFXXPEORWWHGRQDQLWURFHOOXORVHPHPEUDQHZKLFKZDVWKHQWHVWHGZLWKDQDQWL7RSDQWLERG\DQG
(&/OHIWDQGZLWK'$3,PLGGOHȕDFWLQULJKWUHSUHVHQWVWKHFRQWUROIRUQRQFRYDOHQWO\ERXQGFRQWDPLQDQW
SURWHLQV 7KH GHQVLWRPHWULF DQDO\VLV RI 7RSFF VLJQDOV DIWHU QRUPDOL]DWLRQ IRU EORWWHG '1$ XVLQJ '$3, IURP
WKUHHELRORJLFDOUHSOLFDWHVPHDQ“6'LVUHSRUWHGLQWKHJUDSK<D[LVUHSUHVHQWVWKHUHODWLYHVLJQDOLQWHQVLW\
:KHUHLQGLFDWHGWKHGLIIHUHQFHEHWZHHQFRQWURODQG$7ZDVVWDWLVWLFDOO\VLJQLILFDQW3 6WXGHQW·V7WHVW
5HVXOWV
'1$ GDPDJHLQGXFHG *6 FKHFNSRLQW LQ UHODWLRQ WR
GHYHORSPHQWVWDJH
,Q VRPDWLF FHOOV $70 PHGLDWHV WKH *6 FHOO F\FOH FKHFNSRLQW DUUHVW WR SUHYHQWV FHOOV
ZLWK GDPDJHG '1$ IURP HQWHULQJ WKH 6 SKDVH WKHUHE\ SURWHFWLQJ JHQRPLF LQWHJULW\ +RZHYHUWKH*6FKHFNSRLQWVHHPVWREHGHYHORSPHQWDOO\UHJXODWHGDQGLVLQDFWLYHLQ(6&V
DQGL36&V*LYHQWKHUROHRIWKH*6FKHFNSRLQWLQJHQRPLFVWDELOLW\LWVDEVHQFHLQ
SOXULSRWHQWVWHPFHOOVXQGHUOLHVIXQGDPHQWDOGLIIHUHQFHVLQWKHZD\WKHVHFHOOVSUHVHUYHJHQRPH
LQWHJULW\UHODWLYHWRVRPDWLFFHOOV7RIXUWKHUGHOLQHDWHWKLVDFWLYLW\LQUHODWLRQWRGHYHORSPHQWDO
VWDJHZHDQDO\]HGWKHFHOOF\FOHSURILOHRISULPDU\ILEUREODVWKXPDQL36&VKXPDQ13&VDQG
WHUPLQDOO\GLIIHUHQWLDWHGQHXURQVE\IORZF\WRPHWU\EHIRUHDQGKDIWHU,5WUHDWPHQW3ULPDU\
FRQWURO ILEUREODVWV GLVSOD\HG D QRUPDO *6 FKHFNSRLQW DUUHVW DV HYLGHQW IURP WKH VPDOO
UHGLVWULEXWLRQRIWKHFHOOF\FOHSKDVHVDIWHU,5LQPDUNHGFRQWUDVWWR$7ILEUREODVWVLQZKLFK
WKLV FKHFNSRLQW GHIHFW UHVXOWHG LQ D VLJQLILFDQW GURS LQ WKH SHUFHQWDJH RI * DQG FRQFRPLWDQW
LQFUHDVH LQ * SKDVHFHOOV )LJXUH $ 1RWDEO\ QRW RQO\$7KXPDQ L36&V EXW DOVR FRQWURO
KXPDQL36&VODFNHGWKH*6FKHFNSRLQW,QWHUHVWLQJO\FRQWURO13&VUHJDLQHGWKHFDSDFLW\WR
DFWLYDWH WKH FKHFNSRLQW DIWHU ,5 ZKHUHDV $7 13&V PDLQWDLQHG WKLV GHIHFW 7KH DQDO\VLV RI
QHXURQV DW ' UHYHDOHG WKDW DOO FHOOV ZHUH LQ WKH ** SKDVH DQG WKLV FRQGLWLRQ ZDV QRW
SHUWXUEHGKDIWHU,57KHGLVWULEXWLRQRIFHOOVLQWKHGLIIHUHQWFHOOF\FOHSKDVHVLVVXPPDUL]HG
LQ)LJXUH%
7R EHWWHU XQGHUVWDQG WKH LPSOLFDWLRQ RI '1$ GDPDJH UHVSRQVH SURWHLQV LQ WKH
UHJXODWLRQRIFHOOF\FOHDUUHVWLQILEUREODVWVL36&VDQG13&VFRQWUROFHOOVZHUHKDUYHVWHGDWWKH
LQGLFDWHG WLPH SRLQWV DIWHU WUHDWPHQW ZLWK *\ ,5 DQG WKH H[SUHVVLRQ RI $70S6 S
S6 &'&$ DQG SZDI ZDV DQDO\]HG )LJXUH & ,QWHUHVWLQJO\ DOWKRXJK DOO FHOO W\SHV
SURILFLHQWO\DFWLYDWHG$70DQGLQGXFHGWKHDFFXPXODWLRQRISKXPDQL36&VZHUHXQDEOHWR
5HVXOWV
LQGXFH SZDI DQG GLG QRW H[SUHVV &'&$ 7KLV FRXOG H[SODLQ WKH LQFDSDFLW\ RI L36&V WR
UHJXODWH WKH FHOO F\FOH DIWHU '1$ GDPDJH :H DOVR LQYHVWLJDWHG WKH H[SUHVVLRQ RI RWKHU '1$
GDPDJH UHVSRQVH SURWHLQV VXFK DV 5$' WKH 051 FRPSOH[ DQG ;UD\ UHSDLU FURVV
FRPSOHPHQWLQJ ;5&& DQG IRXQG FKDQJHV LQ SURWHLQ H[SUHVVLRQ GXULQJ WKH GLIIHUHQW
GLIIHUHQWLDWLRQ VWHSV VXJJHVWLQJD GLIIHUHQW DELOLW\ WR UHSDLU '1$ GDPDJH LQ WKH GLIIHUHQW FHOO
W\SHV)LJXUH'
)LJXUH
5HVXOWV
5HVXOWV
)LJ &HOO F\FOH SKDVH GLVWULEXWLRQ FKDQJHV DIWHU LRQL]LQJ UDGLDWLRQ WUHDWPHQW ,Q $ FRQWURO DQG $7 FHOOV DW
YDULRXVGHYHORSPHQWDOVWDJHVSULPDU\ILEUREODVWVKXPDQL36&VKXPDQ13&V'SRVWPLWRWLFQHXURQVZHUH
WUHDWHGZLWK*\LRQL]LQJUDGLDWLRQFROOHFWHGDIWHUKDQGDQDO\]HGIRU'1$FRQWHQWE\IORZF\WRPHWU\,Q%
WKH**0UDWLREHIRUHDQGKDIWHULRQL]LQJUDGLDWLRQWUHDWPHQWIRUHDFKFHOOW\SHZDVGHWHUPLQHGIURPWKH
'1$ KLVWRJUDPV GHSLFWHG LQ $ ,Q & FRQWURO ILEUREODVWV L36&V DQG 13&V ZHUH WUHDWHG ZLWK *\ LRQL]LQJ
UDGLDWLRQ FROOHFWHG DW WKH LQGLFDWHG WLPHV DQG DQDO\]HG E\ :HVWHUQ EORW IRU WKH H[SUHVVLRQ RI WKH LQGLFDWHG
SURWHLQV ȕDFWLQ ZDV XVHG DV D ORDGLQJ FRQWURO ,Q ' SULPDU\ ILEUREODVWV KXPDQ L36&V SUROLIHUDWLQJ KXPDQ
13&V DQG SRVWPLWRWLF QHXURQV DW GD\V RI GLIIHUHQWLDWLRQ ' ZHUH FRPSDUHG E\ :HVWHUQ EORW IRU WKH
H[SUHVVLRQRIWKHLQGLFDWHG'1$GDPDJHUHVSRQVHSURWHLQVǃDFWLQZDVXVHGDVDORDGLQJFRQWURO
'LVFXVVLRQ
'LVFXVVLRQ
7KH PHFKDQLVPV XQGHUO\LQJ QHXURGHJHQHUDWLRQ LQ $7 VWLOO UHPDLQ HOXVLYH 8S WR WKH
SUHVHQW $70 NQRFNRXW DQG NQRFNLQ PLFH DQG LQ YLWUR $70GHILFLHQW KXPDQ FHOOXODU PRGHOV
KDYH EHHQ XVHIXO IRU HOXFLGDWLQJ PDQ\ DVSHFWV RI WKH QHXURSDWKRORJ\ EXW WKH ILUVW GR QRW
UHFDSLWXODWH WKH FHQWUDO QHUYRXV V\VWHP GLVHDVH ZKLOH WKH ODWWHU FDQQRW IXOO\ PLUURU WKH
HIIHFWVRI$7FDXVLQJPXWDWLRQV
,QRUGHUWRREWDLQKXPDQ$7QHXURQVLQYLWURLQWKLVVWXG\ZHSURGXFHGSUROLIHUDWLQJ
VHOIUHQHZLQJKXPDQ13&VWKURXJKWKHUHSURJUDPPLQJRI$7ILEUREODVWVLQWRKXPDQL36&V
7KHVH 13&V FDQ EH GLIIHUHQWLDWHG LQWR QHXURQV EXW DOVR H[SDQGHG IUR]HQ DQG WKDZHG
&RQVHTXHQWO\LQYLWURDQDO\VLVRIWKHPHFKDQLVPVOHDGLQJWRQHXURGHJHQHUDWLRQLQ$7FDQEH
LQYHVWLJDWHG LQ ERWK VHOIUHQHZLQJ QHXUDO SUHFXUVRUV DQG PDWXUH QHXURQV ZLWKRXW KDYLQJ WR
FRQVWDQWO\ XQGHUWDNH WKH ODERULRXV DQG WLPHFRQVXPLQJ QHXURQDO GLIIHUHQWLDWLRQ RI KXPDQ
L36&V7KLVWHFKQLTXHKDVSURYHQWREHDSRZHUIXOWRROIRUPRGHOLQJQHXURORJLFDOGLVHDVHVDQG
LQGHHG LW KDV EHHQ XVHIXO IRU VWXG\LQJ WKH SDWKRORJLFDO PHFKDQLVPV XQGHUO\LQJ PDQ\
QHXURGHJHQHUDWLYH GLVRUGHUV 7KH JHQHUDWLRQ RI KXPDQ L36&GHULYHG QHXUDO FHOOV KDV
SUHYLRXVO\EHHQGHVFULEHGIRU$7SDWLHQWVEXWDIXQFWLRQDODQDO\VLVRIWKHQHXURQDO
FHOOV KDUERULQJ SDWLHQWVSHFLILF PXWDWLRQV LQ $70 KDV QRW \HW EHHQ UHSRUWHG 7KXV ZH
SHUIRUPHG D WKRURXJK H[DPLQDWLRQ RI RXU KXPDQ L36&GHULYHG $7 13&V DQG WHUPLQDOO\
GLIIHUHQWLDWHGQHXURQVLQFXOWXUH
:HIRXQGWKDW$7KXPDQSUROLIHUDWLQJ13&VGLVSOD\HGDVWURQJO\DWWHQXDWHGUHVSRQVH
WR '6%V ZLWK UHVSHFW WR FRQWURO KXPDQ 13&V ZKLFK LV FRQFRUGDQW ZLWK D SUHYLRXV VWXG\ RQ
KXPDQ16&V0RUHRYHUDOWKRXJKHOHFWURSK\VLRORJLFDOSURSHUWLHVZHUHVLPLODULQFRQWURO
DQG$7FHOOVWHUPLQDOO\GLIIHUHQWLDWHG$7QHXURQVH[KLELWHGGHFUHDVHGH[SUHVVLRQRI6<3DQG
36'7KLVLVFRQFRUGDQWZLWKSUHYLRXVZRUNVVKRZLQJSUHDQGSRVWV\QDSWLFGHJHQHUDWLRQLQ
$70 NQRFNRXW PLFH WKH UHTXLUHPHQW RI F\WRSODVPLF $70 IRU SKRVSKRU\ODWLRQ RI WKH
V\QDSWLFYHVLFOHSURWHLQV9$03DQGV\QDSVLQDQGWKDW$70GHILFLHQF\DIIHFWVVSRQWDQHRXV
'LVFXVVLRQ
YHVLFOH UHOHDVH DQG HVWDEOLVKPHQW RI ORQJWHUP SRWHQWLDWLRQ :H DOVR VKRZHG WKDW $7
QHXURQVDUHGHIHFWLYHLQWKHH[SUHVVLRQRI6&*DQG.&K,3DOWRJHWKHUXQGHUSLQQLQJGHIHFWV
LQQHXURQDOPDWXUDWLRQ
,W KDV EHHQ SUHYLRXVO\ UHSRUWHG WKDW IHWDO EUDLQGHULYHG KXPDQ 16& H[KLELW D QRUPDO
UHVSRQVHWR,5ZKLFKLVDWWHQXDWHGE\DEODWLRQRULQKLELWLRQRI$706XEVHTXHQWO\1D\OHU
GHPRQVWUDWHGWKDW$7L36&VGLVSOD\GHIHFWLYH,5LQGXFHGVLJQDOLQJUDGLRVHQVLWLYLW\DQG
FHOO F\FOH FKHFNSRLQW GHIHFWV ,Q DFFRUGDQFH ZLWK WKHVH UHVXOWV KHUH ZH IRXQG WKDW WKH '1$
GDPDJHUHVSRQVHZDVYLJRURXVO\DFWLYDWHGLQSRVWPLWRWLFFRQWUROQHXURQVDVHYLGHQWIURPWKH
VWURQJ SKRVSKRU\ODWLRQRI $70 VXEVWUDWHVZKHUHDV LQ$7 SRVWPLWRWLFQHXURQV WKLV UHVSRQVH
ZDV DOPRVW DEODWHG GHPRQVWUDWLQJ WKDW QRUPDO KXPDQ L36&VGHULYHG QHXURQV PDLQWDLQ WKH
DELOLW\ WR DFWLYDWH WKH $70GHSHQGHQW '1$ GDPDJH UHVSRQVH 7KURXJK WKH QHXWUDO FRPHW
DVVD\ DQG WKH DVVHVVPHQW RI DŽ+$; DQG %3 IRFL ZH DOVR KLJKOLJKWHG UHOHYDQW '6% UHSDLU
GHIHFWV LQ $7 QHXURQV ZKLFK LQGLFDWHV WKDW $70 GHILFLHQF\ GHOD\V WKH UHSDLU RI '6%V LQ
DFFRUGDQFHZLWKSUHYLRXVUHSRUWVDQGZLWKWKHREVHUYDWLRQWKDW$70LVUHTXLUHGIRU
WKH UHSDLU RI KHWHURFKURPDWLF '6%V )XUWKHUPRUH ZH GHPRQVWUDWHG WKDW $7 QHXURQV
VKRZ OHVV DSRSWRVLVUHODWHG VXEGLSORLG '1$ FRQWHQW DQG D ORZHU FOHDYHG 3$53 LQGXFWLRQ
DIWHU ,5WUHDWPHQW WKDQ FRQWURO QHXURQV ,QGHHG $70 GHILFLHQF\ KDV EHHQ IRXQG WR DWWHQXDWH
WKH DSRSWRWLF UHVSRQVH WR ,5 LQ $7 O\PSKREODVWRLG FHOOV DQG LQ KXPDQ 16&V DQG
VWXGLHV XVLQJ $70GHILFLHQW PLFH KDYH VKRZQ WKDW $70 LV UHTXLUHG IRU WKH SPHGLDWHG
DSRSWRVLVRIGHYHORSLQJSRVWPLWRWLFQHXURQVH[SRVHGWR,5
2[LGDWLYHVWUHVVDQGHQVXLQJR[LGDWLYH'1$GDPDJHFDQFRQWULEXWHWRQHXUDOFHOOGHDWK
%(5 LV WKH PDMRU SDWKZD\ IRU WKH UHSDLU RI R[LGDWLYHO\ GDPDJHG '1$ DQG YDULRXV 66%
LQWHUPHGLDWHV DQG SUHYLRXV ILQGLQJV GHPRQVWUDWH WKDW $70 DQG &+. IDFLOLWDWH WKH
UHFUXLWPHQW RI GRZQVWUHDP %(5 SURWHLQV GXULQJ WKH LQLWLDO GDPDJH UHFRJQLWLRQ DQG H[FLVLRQ
VWHS WR SURPRWH %(5 +HUH ZH IRXQG QR VLJQLILFDQW GLIIHUHQFHV LQ 63%(5 DFWLYLW\
'LVFXVVLRQ
EHWZHHQFRQWURODQG$7SUROLIHUDWLYHKXPDQ13&VDQGSRVWPLWRWLFQHXURQVZKLOHFRQYHUVHO\
/3%(5DFWLYLW\ZDVUHGXFHGLQSRVWPLWRWLFQHXURQV:KHQZHDQDO\]HGWKHH[SUHVVLRQRI63
%(5SURWHLQVDIWHUGLIIHUHQWLDWLRQZHIRXQGWKDWWKHOHYHOVRI;5&&ZHUHGRZQUHJXODWHGDQG
WKHVDPHZDVWUXHIRUWKH/3%(5DVVRFLDWHG)(1SURWHLQ,WVKRXOGEHQRWHGWKDWERWK63%(5
DQG /3%(5 HQ]\PDWLF HIILFLHQFLHV KDYH EHHQ IRXQG WR GHFUHDVH DORQJ ZLWK WKH UHVSHFWLYH
SURWHLQFRPSRQHQWVLQDQHXUREODVWRPDFHOOOLQHXSRQGLIIHUHQWLDWLRQDQGZKLOHRXUGDWD
RQ /3%(5 FRQFRUG ZLWK WKHVH UHVXOWV WKRVH RQ 63%(5 DUH FOHDUO\ DW YDULDQFH 7KLV FRXOG EH
EHFDXVH WKH H[SUHVVLRQ RI WKH 63%(5 FRPSRQHQW ;5&& LV WRWDOO\ DEODWHG LQ GLIIHUHQWLDWHG
QHXUREODVWRPDFHOOVZKLOHEHLQJRQO\SDUWLDOO\GRZQUHJXODWHGLQRXUQHXURQV5HJDUGOHVV
RI WKLV GLVFUHSDQF\ WKH YDULDWLRQV LQ /3%(5 DFWLYLW\ EHWZHHQ SRVWPLWRWLF QHXURQV DQG
SUROLIHUDWLYH KXPDQ 13&V ZHUH WRWDOO\ LQGHSHQGHQW RI $70 H[SUHVVLRQ 0RUHRYHU DOWKRXJK
$70 DSSHDUV LQYROYHG LQ VHQVLQJ DQG PRGXODWLQJ WKH FHOOXODU UHVSRQVH WR 526 DONDOLQH
FRPHWDVVD\VVKRZHGWKDW$7DQGFRQWUROQHXURQVHTXDOO\UHSDLUHG66%V
&RQVLVWHQWZLWKWKHVHUHVXOWVLQRXUVWXG\$7SRVWPLWRWLFQHXURQVZHUHDVVHQVLWLYHWR
R[LGDWLYH DJHQWV DV FRQWURO SRVWPLWRWLF QHXURQV LQ DJUHHPHQW ZLWK UHVXOWV REWDLQHG ZLWK $7
O\PSKREODVWRLG FHOOV %\ FRQWUDVW $7 SRVWPLWRWLF QHXURQV ZHUH VLJQLILFDQWO\ PRUH
UHVLVWDQWWKDQFRQWUROQHXURQVWRWKHJHQRWR[LFDJHQWV&37DQGWUDEHFWHGLQSUREDEO\EHFDXVH
LQ WKH DEVHQFH RI $70 GUXJLQGXFHG '6%V LQ WUDQVFULEHG JHQHV IDLO WR DFWLYDWH D S
GHSHQGHQW DSRSWRWLF UHVSRQVH 7KLV ILQGLQJ LV FRQFRUGDQW ZLWK UHVXOWV VKRZLQJ WKDW $70
LQKLELWLRQVXSSUHVVHVWKHJHQRWR[LFUHVSRQVHLQUDWFHUHEHOODUJUDQXOHQHXURQV
,W KDV EHHQ UHFHQWO\ VKRZQ WKDW SRVWPLWRWLF QHXUDO FHOOV IURP $70GHILFLHQW PLFH
DFFXPXODWH7RSFFVDQGIDLOWRUHFRYHUJOREDOWUDQVFULSWLRQDIWHU7RSFFWUDSSLQJE\&37
*LYHQWKDW7RSFFVFDQSURPRWHWUDQVFULSWLRQHORQJDWLRQDUUHVWDQGGHFD\DQGFRQVHTXHQW
WUDQVFULSWLRQDO GHFOLQH D SKHQRPHQRQ XQGHUO\LQJ QHXURGHJHQHUDWLRQ ZH DQDO\]HG
7RSFFVE\WZRGLIIHUHQWDVVD\VDQGIRXQGJUHDWHUOHYHOVRIWKHVHLQWHUPHGLDWHVLQ$7WKDQLQ
'LVFXVVLRQ
FRQWUROQHXURQV7KLVILQGLQJZKLFKWRRXUNQRZOHGJHKDVQHYHUEHHQUHSRUWHGLQKXPDQ$7
QHXURQVZDUUDQWVIXUWKHUVWXGLHVWRGHWHUPLQHWKHLPSDFWRI7RSFFVRQJHQRPHZLGHJOREDO
WUDQVFULSWLRQ
)LQDOO\ LQ RUGHU WR GHOLQHDWH WKH DFWLYLW\ RI WKH *6 FKHFNSRLQW LQ UHODWLRQ WR
GHYHORSPHQWDO VWDJH ZH DQDO\]HG WKH FHOO F\FOH SURILOH RI SULPDU\ ILEUREODVW KXPDQ L36&V
KXPDQ 13&V DQG WHUPLQDOO\ GLIIHUHQWLDWHG QHXURQV ,Q DJUHHPHQW ZLWK WKH UROH RI $70 LQ
HVWDEOLVKLQJWKH*6FKHFNSRLQWDUUHVWWRSUHYHQWFHOOVZLWKGDPDJHG'1$IURPHQWHULQJWKH6
SKDVH SULPDU\ FRQWURO ILEUREODVWV HIIHFWLYHO\ DUUHVWHG DW WKH *6 ERXQGDU\ DIWHU ,5 EXW
RQFHUHSURJUDPPHGLQWRKXPDQL36&VWKH\ORVWWKLV*6FKHFNSRLQWDUUHVWWRUHJDLQLWDWWKH
VWDJH RI KXPDQ 13&V ,QVWHDG $7 FHOOV VKRZHG D GLVUXSWHG *6 FKHFNSRLQW ZKDWHYHU WKH
GHYHORSPHQWDO VWDJH 1RWDEO\ WKH DEVHQFH RI WKH *6 FKHFNSRLQW DUUHVW LQ FRQWURO KXPDQ
L36&VLVDFKDUDFWHULVWLFRI(6&VZKHUHFHOOVVXEMHFWHGWR'1$GDPDJHLQ*HQWHUWKH6SKDVH
WR EH HOLPLQDWHG E\ DSRSWRVLV WKXV DYRLGLQJ WKH SURSDJDWLRQ RI PXWDWLRQV ZKLFK ZRXOG EH
GHWULPHQWDO IRU WKH ZKROH RUJDQLVP $FFRUGLQJO\ KXPDQ L36&V VKRZHG D JUHDW
DFFXPXODWLRQ RI SS6 DQG IDLQW LQGXFWLRQ RI SZDI FRPSDWLEOH ZLWK D SGHSHQGHQW
DSRSWRWLF UHVSRQVH UDWKHU WKDQ ZLWK D *6 FKHFNSRLQW DFWLYDWLRQ 0RUHRYHU ZH IRXQG WKDW
FRQWUROKXPDQL36&VXSUHJXODWH+5FRPSRQHQWVVXFKDV5$'FRQFRUGDQWZLWKWKHIDFWWKDW
(6&VSUHGRPLQDQWO\XVHWKLVKLJKILGHOLW\SDWKZD\WRDYRLGWKHDFFXPXODWLRQRIPXWDWLRQVLQ
FRQWUDVWWRVRPDWLFFHOOVZKLFKUHO\RQWKHHUURUSURQH1+(-WRUHSDLU'1$GDPDJHRFFXUULQJ
LQ*
,PSRUWDQWO\ DOWKRXJK KXPDQ $7 QHXURQDO FHOOV UHSUHVHQW DQ DWWUDFWLYH LQ YLWUR PRGHO
IRUWKHVWXG\RIWKHQHXURGHJHQHUDWLYHSURFHVVHVLQ$7WKHFHOOW\SHZKLFKLVVSHFLILFDOO\ORVW
LQ SDWLHQWV LH KXPDQ 3XUNLQMH QHXURQV FDQQRW \HW EH JHQHUDWHG IURP ILEUREODVWGHULYHG
SOXULSRWHQWVWHPFHOOVGXHWRWKHODFNRIRSWLPL]HGSURWRFROV,QGHHGWKHHIILFLHQWJHQHUDWLRQRI
KXPDQ L36&GHULYHG QHXUDO VXEW\SHV ZLWK FRUUHFW SKHQRW\SHV UHPDLQV D FKDOOHQJH 'LVFXVVLRQ
0RUHRYHU RXU LQ YLWURPRGHO GRHV QRW WDNH LQWR DFFRXQW SRVVLEOHQRQFHOO DXWRQRPRXV HIIHFWV
WKDWWKHXELTXLWRXVODFNRI$70FRXOGH[HUWRQQHXURQVLQWKHFRPSOH[FRQWH[WRIWKHFHQWUDO
QHUYRXV V\VWHP 7KHUHIRUH DOWKRXJK WKLV PRGHO PD\ EH XVHIXO IRU XQUDYHOLQJ WKH
PHFKDQLVPVRI$70IXQFWLRQVLQQHXURQVWKHVHOLPLWDWLRQVPXVWEHNHSWLQWRDFFRXQW
7R VXPPDUL]H ZH KDYH VKRZQ WKDW KXPDQ $7 L36&GHULYHG SRVWPLWRWLF QHXURQV
GLVSOD\ QRUPDO HOHFWURSK\VLRORJLFDO DFWLYLW\ GHIHFWLYH H[SUHVVLRQ RI PDWXUDWLRQ PDUNHUV
DWWHQXDWHGUHVSRQVHWRDQGUHSDLURI'6%VEXWQRUPDOFDSDFLW\WRUHSDLU66%VDQGQRUPDO%(5
DFWLYLWLHV 6WULNLQJO\ $7 QHXURQV H[KLELW HOHYDWHG OHYHOV RI 7RSFFV ZKLFK PD\ SRWHQWLDOO\
LPSDLUWUDQVFULSWLRQDOILGHOLW\DSRVVLELOLW\WKDWUHTXLUHVIXUWKHULQYHVWLJDWLRQ)LQDOO\ZHKDYH
VKRZQWKDWKXPDQL36&VOLNH(6&VSURYLGHDXQLTXHLQYLWURPRGHOWRVWXG\WKHGHWDLOVRIWKH
*6FHOOF\FOHFKHFNSRLQWLQDGHYHORSPHQWDOFRQWH[W
,Q FRQFOXVLRQ WKH JHQHUDWLRQ RI $7 SDWLHQWGHULYHG QHXURQDO FHOOV DOORZV RYHUFRPLQJ
WKH REVWDFOH SRVHG E\ WKH LQDGHTXDF\ RI $70GHILFLHQW DQLPDOV DQG FHOO OLQHV 7KLV PRGHO
UHSUHVHQWV D SRZHUIXO WRRO IRU H[SORULQJ WKH PHFKDQLVPV WKDW FDXVH GHJHQHUDWLRQ RI SDWLHQW
$70GHILFLHQW QHXURQV DQG LW PD\ EH H[SORLWHG IRU LGHQWLI\LQJ WKHUDSHXWLF WDUJHWV DQG IRU
VFUHHQLQJPROHFXOHVZLWKSRWHQWLDOQHXURSURWHFWLYHSURSHUWLHV2XUSUHYLRXVO\XQGRFXPHQWHG
IXQFWLRQDODQGPROHFXODUFKDUDFWHUL]DWLRQRISDWLHQWGHULYHGQHXURQDOFHOOVUHSUHVHQWVWKHILUVW
VWHS LQ WKH HPSOR\PHQW RI WKHVH FHOOV DV WKH PRVW LQIRUPDWLYH LQ YLWUR PRGHO WR LQYHVWLJDWH
QHXURGHJHQHUDWLRQLQ$7
)XWXUHSHUVSHFWLYHV
)XWXUHSHUVSHFWLYHV
6HYHUHFHUHEHOODUDWURSK\DQGOHVVSURQRXQFHGGHJHQHUDWLRQLQWKHEUDLQVWHPDQGVSLQDO
FRUGWUDFWVFKDUDFWHUL]H$7EXWWKHVHOHFWLYLW\RIWKHVHFHOOXODUWDUJHWVRIGHJHQHUDWLRQLVVWLOO
XQGHWHUPLQHG,QWKHEUDLQKLJKR[\JHQFRQVXPSWLRQDQGPHWDEROLFDFWLYLW\H[SRVHQHXURQVWR
HOHYDWHG 526 OHYHOV WKDW FDQ LQGXFH DEXQGDQW 66%V 7KHVH OHVLRQV FDQ DOVR DULVH LQGLUHFWO\
GXULQJ %(5 RU DV WKH UHVXOW RI DERUWHG 7RS DFWLYLW\ DQG LI XQUHVROYHG XQGHUPLQH JOREDO
WUDQVFULSWLRQDQGYLDELOLW\2QHGLVWLQFWLYHIHDWXUHRI3XUNLQMHFHOOVUHODWLYHWRRWKHUQHXURQVLV
WKHLULQWULQVLFDOO\KHLJKWHQHGWUDQVFULSWLRQDOUDWHDVLQIHUUHGIURPWKHODUJHDQGDOPRVWHQWLUHO\
HXFKURPDWLFQXFOHLFKDUDFWHUL]HGE\HSLJHQHWLFPDUNVIRUDFWLYHFKURPDWLQ%DVHGRQ
WKHVH REVHUYDWLRQV ZH K\SRWKHVL]H WKDW LQ SRVWPLWRWLF $7 QHXURQV WZR FRPELQHG HIIHFWV RI
$70ORVVLQFUHDVHGOHYHOVRIHQGRJHQRXV526DQGRIWUDQVFULSWLRQEORFNLQJ7RSFFVGULYHWKH
DFFXPXODWLRQ RI 66%V SURPRWLQJ JHQH WUDQVFULSWLRQHORQJDWLRQ GHFOLQH DQG XOWLPDWHO\ FHOO
GHDWK 7KLV SKHQRPHQRQ LQ $7 3XUNLQMH QHXURQV FRXOG EH IXUWKHU H[DFHUEDWHG E\ WKHLU
H[FHVVLYH WUDQVFULSWLRQDO GHPDQG OLNHO\ WR LQFUHDVH WKH UDWH RI FKHPLFDO DQG JHQRWR[LF
PRGLILFDWLRQVRQWKHH[SRVHG'1$WHPSODWHDQGWRRYHUZKHOPWKHFDSDELOLWLHVRIWKH
WUDQVFULSWLRQDO PDFKLQHU\ 7R DGGUHVV WKLV SRVVLELOLW\ ZH ZLOO DVVHVV JHQRPHZLGH
WUDQVFULSWLRQDQG51$VSOLFLQJLQKXPDQL36&GHULYHGQHXURQVZLWKQRUPDORULQDFWLYH$70
DFWLYLW\ XVLQJ 51$VHT DQG DQDO\]H E\ 7RSFF,3VHT WKH RFFXSDQF\ RI 7RSFF DFURVV WKH
JHQRPH DQG JHQH ERGLHV DQG PDWFK WKLV LQIRUPDWLRQ ZLWK JHQH H[SUHVVLRQ GDWD REWDLQHG E\
51$VHT
7KHDFWLYDWLRQRILRQRWURSLFJOXWDPDWHUHFHSWRUVLQGXFHVPHPEUDQHGHSRODUL]DWLRQ&D
LQÁX[ DQG D VLJQDOLQJ FDVFDGH FXOPLQDWLQJ LQ WUDQVFULSWLRQDO LQGXFWLRQ RI LPPHGLDWHHDUO\
JHQHV <HW SURORQJHG H[SRVXUH WR JOXWDPDWH OHDGV WR H[FLWRWR[LFLW\ FKDUDFWHUL]HG E\ GHOD\HG
GHUHJXODWLRQ RI FHOOXODU &D KRPHRVWDVLV GUDPDWLF LQFUHDVH LQ 526 DQG FROODSVH RI
PLWRFKRQGULDO PHPEUDQH SRWHQWLDO 6LQFH $7 FHOOV SUHVHQW DQ LPSDLUHG 526 UHJXODWLRQ
DQG GHIHFWLYH PRELOL]DWLRQ RI LQWUDFHOOXODU &D FRQFHLYDEO\ WKHVH G\VIXQFWLRQV
)XWXUHSHUVSHFWLYHV
PLJKWIDYRUH[FLWRWR[LFLW\HVSHFLDOO\LQWKHFDVHRI3XUNLQMHFHOOVZKLFKXQOLNHRWKHUQHXURQV
DUHQRUPDOO\KLJKO\VXVFHSWLEOHWR$03$PHGLDWHGH[FLWRWR[LFLW\RZLQJDOVRWRSRRUKDQGOLQJ
RI&DORDGOHDGLQJWRFDOSDLQDFWLYDWLRQ:HZLOOWKHUHIRUHDVVHVVWKHH[FLWRWR[LFUHVSRQVH
RIQRUPDODQG$7KXPDQL36&GHULYHGQHXURQVWROLJDQGVIRULRQRWURSLFJOXWDPDWHUHFHSWRUV
+XPDQL36&VDOORZWRPRGHOJHQHWLFGLVHDVHVLQYLWURDQGWRHOXFLGDWHWKHLUXQGHUO\LQJ
PHFKDQLVPV<HWLQWKHFDVHRI$7DFXUUHQWGUDZEDFNLVWKHODFNRIDSURWRFROWRGLIIHUHQWLDWH
KXPDQL36&VLQWR3XUNLQMHQHXURQV$OEHLWWKHVHQHXURQVFDQEHJHQHUDWHGLQYLWURIURPPRXVH
(6&V XVLQJ DQ HVWDEOLVKHG SURWRFRO LW LV XQNQRZQ ZKHWKHU LW ZRXOG ZRUN ZLWK
KXPDQL36&VEXWLIVXFFHVVIXOWKLVZRXOGIDUDGYDQFHWKHNQRZOHGJHRI3XUNLQMHFHOOELRORJ\
LQ $7 +HUH WZR H[SHULPHQWDO DSSURDFKHV ZLOO EH XQGHUWDNHQ 2QH LQ YLWUR DLPHG DW
JHQHUDWLQJ3XUNLQMHQHXURQVIURPQRUPDODQG$7KXPDQL36&VDGDSWLQJSURWRFROVGHYHORSHG
IRU PRXVH (6&V 7KH RWKHU LQ YLYR LQYROYLQJ WKH RUWKRWRSLF WUDQVSODQWLRQ LQ XWHUR LQWR WKH
FHUHEHOODU SODWH RI UDW HPEU\RV RI KXPDQ L36&GHULYHG QHXUDO SUHFXUVRUV IDWHG ZLWK
FDXGDOL]LQJDQGGRUVDOL]LQJIDFWRUVWRILQGRXWLIWKHV\VWHPLVSHUPLVVLYHIRUWKHGHYHORSPHQW
RIKXPDQ3XUNLQMHFHOOVDQGLIVRWRLQYHVWLJDWHWKHLUSURSHUWLHVLQRUJDQRW\SLFFXOWXUHV
5HIHUHQFHV
5HIHUHQFHV
$PEURVH 0 *DWWL 5$ 3DWKRJHQHVLV RI DWD[LDWHODQJLHFWDVLD WKH QH[W JHQHUDWLRQ RI $70
IXQFWLRQV%ORRG
6\OODED / +HQQHU . &RQWULEXWLRQ D O·LQGHSHQGHQFH GH O·DWKHWRVH GRXEOH LGLRSDWKLTXH HW
FRQJHQLWDOH5HY1HXURO
/RXLV %DUU ' 6XU XQ V\QGURPH SURJUHVVLI FRPSUHQHQW GHV WHODQJLHFWDVLHV FDSLOODLUHV
FXWDQHHVHWFRQMRQFWLYDOHVV\PHWULTXHVDGLVSRVLWLRQQDHYRLGHHWGHVWURXEOHVFHUHEHOOHX[
&RQILQ1HXURO
%RGHU(6HGJZLFN53$WD[LDWHODQJLHFWDVLD$IDPLOLDOV\QGURPHRISURJUHVVLYHFHUHEHOODU
DWD[LD RFXORFXWDQHRXV WHODQJLHFWDVLD DQG IUHTXHQW SXOPRQDU\ LQIHFWLRQ $ SUHOLPLQDU\
UHSRUWRQFKLOGUHQDQDXWRSV\DQGDFDVHKLVWRU\8QLY6&DOLI0HG%XOO²
6ZLIW00RUUHOO'&URPDUWLH(&KDPEHUOLQ$56NROQLFN0+%LVKRS'77KHLQFLGHQFH
DQG JHQH IUHTXHQF\ RI DWD[LDWHODQJLHFWDVLD LQ WKH 8QLWHG 6WDWHV $P - +XP *HQHW 6DYLWVN\ . %DU6KLUD $ *LODG 6 5RWPDQ * =LY < 9DQDJDLWH / HW DO $ VLQJOH DWD[LD
WHODQJLHFWDVLDJHQHZLWKDSURGXFWVLPLODUWR3,NLQDVH6FLHQFH
%RGHU(6HGJZLFN53$WD[LDWHODQJLHFWDVLDDIDPLOLDOV\QGURPHRISURJUHVVLYHFHUHEHOODU
DWD[LD RFXORFXWDQHRXV WHODQJLHFWDVLD DQG IUHTXHQW SXOPRQDU\ LQIHFWLRQ 3HGLDWULFV +RFKH ) 6HLGHO . 7KHLV 0 9ODKR 6 6FKXEHUW 5 =LHOHQ 6 HW DO 1HXURGHJHQHUDWLRQ LQ
DWD[LDWHODQJLHFWDVLDZKDWLVQHZ":KDWLVHYLGHQW"1HXURSHGLDWULFV
9HUKDJHQ 00 0DUWLQ -- YDQ 'HXUHQ 0 &HXWHULFNGH *URRWH & :HHPDHV &0 .UHPHU
%+ HW DO 1HXURSDWKRORJ\ LQ FODVVLFDO DQG YDULDQW DWD[LDWHODQJLHFWDVLD 1HXURSDWKRORJ\
-XQ
5HIHUHQFHV
%RWWLQL $5 *DWWL 5$ :LUHQIHOGW 0 9LQWHUV +9 +HWHURWRSLF 3XUNLQMH FHOOV LQ DWD[LD
WHODQJLHFWDVLD1HXURSDWKRORJ\
9LQWHUV+9*DWWL5$5DNLF36HTXHQFHRIFHOOXODUHYHQWVLQFHUHEHOODURQWRJHQ\UHOHYDQW
WR H[SUHVVLRQ RI QHXURQDO DEQRUPDOLWLHV LQ DWD[LDWHODQJLHFWDVLD .URF )RXQG 6HU &KXQ++*DWWL5$$WD[LDWHODQJLHFWDVLDDQHYROYLQJSKHQRW\SH'1$5HSDLU
7DYDQL)=LPPHUPDQ5$%HUU\*76XOOLYDQ.*DWWL5%LQJKDP3$WD[LDWHODQJLHFWDVLD
WKHSDWWHUQRIFHUHEHOODUDWURSK\RQ05,1HXURUDGLRORJ\
$JXLODU0-.DPRVKLWD6/DQGLQJ%+%RGHU(6HGJZLFN533DWKRORJLFDOREVHUYDWLRQVLQ
DWD[LDWHODQJLHFWDVLD$UHSRUWRIILYHFDVHV-1HXURSDWKRO([S1HXURO²
*RRGPDQ:1&RRSHU:&.HVVOHU*%)LVFKHU06*DUGQHU0%$WD[LDWHODQJLHFWDVLD$
UHSRUWRIWZRFDVHVLQVLEOLQJVSUHVHQWLQJDSLFWXUHRISURJUHVVLYHVSLQDOPXVFXODUDWURSK\
%XOO/RV$QJHOHV1HXURO6RF²
'H /HRQ *$ *URYHU :' +XII '6 1HXURSDWKRORJLF FKDQJHV LQ DWD[LDWHODQJLHFWDVLD
1HXURORJ\²
.RHSS 0 6FKHORVN\ / &RUGHV , &RUGHV 0 3RHZH : '\VWRQLD LQ DWD[LD WHODQJLHFWDVLD
UHSRUW RI D FDVH ZLWK SXWDPLQDO OHVLRQV DQG GHFUHDVHG VWULDWDO >,@LRGREHQ]DPLGH
ELQGLQJ0RY'LVRUG²
2SHVNLQ.:DWHUVWRQ-1LUHQEHUJ$+DUH:6$WD[LDWHODQJLHFWDVLDZLWKORQJVXUYLYDO-
&OLQ1HXURVFL²
3HWHUVRQ 5' )XQNKRXVHU -' 7XFN0XOOHU &0 *DWWL 5$ &DQFHU VXVFHSWLELOLW\ LQ DWD[LD
WHODQJLHFWDVLD/HXNHPLD6XSSO²
$KPHG05DKPDQ1$70DQGEUHDVWFDQFHUVXVFHSWLELOLW\2QFRJHQH
5HIHUHQFHV
/DYLQ 0) *XHYHQ 1 %RWWOH 6 *DWWL 5$ &XUUHQW DQG SRWHQWLDO WKHUDSHXWLF VWUDWHJLHV IRU
WKHWUHDWPHQWRIDWD[LDWHODQJLHFWDVLD%U0HG%XOO
YDQ'HXUHQ09HUKDJHQ0:HHPHDV&+HUHGLWDU\SHUVLVWHQFHRIDOSKDIHWRSURWHLQ1HWK
-0HG
$XULDV $ 'XWULOODX[ % %XULRW ' /HMHXQH - +LJK IUHTXHQFLHV RI LQYHUVLRQV DQG
WUDQVORFDWLRQVRIFKURPRVRPHVDQGLQDWD[LDWHODQJLHFWDVLD0XWDW5HV
%DU56/HYLV:55HFKOHU00+DUULVRQ/&6LHEHUW&3RGVNDOQ\-HWDO([WUHPHLQVXOLQ
UHVLVWDQFHLQDWD[LDWHODQJLHFWDVLDGHIHFWLQDIILQLW\RILQVXOLQUHFHSWRUV1(QJO-0HG
&UDZIRUG726NRODVN\5/)HUQDQGH]55RVTXLVW.-/HGHUPDQ+06XUYLYDOSUREDELOLW\
LQDWD[LDWHODQJLHFWDVLD$UFK'LV&KLOG
&DEDQD0'&UDZIRUG72:LQNHOVWHLQ-$&KULVWHQVHQ-5/HGHUPDQ+0&RQVHTXHQFHV
RIWKHGHOD\HGGLDJQRVLVRIDWD[LDWHODQJLHFWDVLD3HGLDWULFV3W
3HUOPDQ 6 %HFNHU&DWDQLD 6 *DWWL 5$ $WD[LDWHODQJLHFWDVLD GLDJQRVLV DQG WUHDWPHQW
6HPLQ3HGLDWU1HXURO
*DWWL5$%HUNHO,%RGHU(%UDHGW*&KDUPOH\3&RQFDQQRQ3HWDO/RFDOL]DWLRQRIDQ
DWD[LDWHODQJLHFWDVLDJHQHWRFKURPRVRPHT1DWXUH
6DYLWVN\.6IH]67DJOH'$=LY<6DUWLHO$&ROOLQV)6HWDO7KHFRPSOHWHVHTXHQFHRI
WKH FRGLQJ UHJLRQ RI WKH $70 JHQH UHYHDOV VLPLODULW\ WR FHOO F\FOH UHJXODWRUV LQ GLIIHUHQW
VSHFLHV+XP0RO*HQHW
0F&RQYLOOH&06WDQNRYLF7%\UG3-0F*XLUH*0<DR4</HQQR[**HWDO0XWDWLRQV
DVVRFLDWHG ZLWK YDULDQW SKHQRW\SHV LQ DWD[LDWHODQJLHFWDVLD $P - +XP *HQHW 5HIHUHQFHV
&RQFDQQRQ3*DWWL5$'LYHUVLW\RI$70JHQHPXWDWLRQVGHWHFWHGLQSDWLHQWVZLWKDWD[LD
WHODQJLHFWDVLD+XP0XWDW
6KLORK < =LY < 7KH $70 SURWHLQ NLQDVH UHJXODWLQJ WKH FHOOXODU UHVSRQVH WR JHQRWR[LF
VWUHVVDQGPRUH1DW5HY0RO&HOO%LRO
/HPSLlLQHQ + +DOD]RQHWLV 7' (PHUJLQJ FRPPRQ WKHPHV LQ UHJXODWLRQ RI 3,..V DQG
3,.V(0%2-
.LP 67 /LP '6 &DQPDQ &( .DVWDQ 0% 6XEVWUDWH VSHFLILFLWLHV DQG LGHQWLILFDWLRQ RI
SXWDWLYHVXEVWUDWHVRI$70NLQDVHIDPLO\PHPEHUV-%LRO&KHP
/DYLQ0)$WD[LDWHODQJLHFWDVLDIURPDUDUHGLVRUGHUWRDSDUDGLJPIRUFHOOVLJQDOOLQJDQG
FDQFHU1DW5HY0RO&HOO%LRO
3ROR6(-DFNVRQ63'\QDPLFVRI'1$GDPDJHUHVSRQVHSURWHLQVDW'1$EUHDNVDIRFXV
RQSURWHLQPRGLILFDWLRQV*HQHV'HY
,\DPD 7 :LOVRQ '0 UG '1$ UHSDLU PHFKDQLVPV LQ GLYLGLQJ DQG QRQGLYLGLQJ FHOOV
'1$5HSDLU
0HW]JHU/,OLDNLV*.LQHWLFVRI'1$GRXEOHVWUDQGEUHDNUHSDLUWKURXJKRXWWKHFHOOF\FOH
DVDVVD\HGE\SXOVHGILHOGJHOHOHFWURSKRUHVLVLQ&+2FHOOV,QW-5DGLDW%LRO
*RRGDU]L$$-HJJR3/REULFK07KHLQIOXHQFHRIKHWHURFKURPDWLQRQ'1$GRXEOHVWUDQG
EUHDNUHSDLU*HWWLQJWKHVWURQJVLOHQWW\SHWRUHOD['1$5HSDLU
%DNNHQLVW &- .DVWDQ 0% '1$ GDPDJH DFWLYDWHV $70 WKURXJK LQWHUPROHFXODU
DXWRSKRVSKRU\ODWLRQDQGGLPHUGLVVRFLDWLRQ1DWXUH
/HH-+3DXOO77$70DFWLYDWLRQE\'1$GRXEOHVWUDQGEUHDNVWKURXJKWKH0UH5DG
1EVFRPSOH[6FLHQFH
5HIHUHQFHV
.R]ORY69*UDKDP0(-DNRE%7RELDV).LMDV$:7DQXML0HWDO$XWRSKRVSKRU\ODWLRQ
DQG $70 DFWLYDWLRQ DGGLWLRQDO VLWHV DGG WR WKH FRPSOH[LW\ - %LRO &KHP 0DU
6XQ < ;X < 5R\ . 3ULFH %' '1$ GDPDJHLQGXFHG DFHW\ODWLRQ RI O\VLQH RI $70
DFWLYDWHV$70NLQDVHDFWLYLW\0RO&HOO%LRO
$OL $ =KDQJ - %DR 6 /LX , 2WWHUQHVV ' 'HDQ 10 HW DO 5HTXLUHPHQW RI SURWHLQ
SKRVSKDWDVHLQ'1$GDPDJHLQGXFHG$70DFWLYDWLRQ*HQHV'HY
/DPDUFKH%-2UD]LR1,:HLW]PDQ0'7KH051FRPSOH[LQGRXEOHVWUDQGEUHDNUHSDLU
DQGWHORPHUHPDLQWHQDQFH)(%6/HWW
%XUPD6&KHQ%30XUSK\0.XULPDVD$&KHQ'-$70SKRVSKRU\ODWHVKLVWRQH+$;
LQUHVSRQVHWR'1$GRXEOHVWUDQGEUHDNV-%LRO&KHP
/RX = 0LQWHU'\NKRXVH . )UDQFR 6 *RVWLVVD 0 5LYHUD 0$ &HOHVWH $ HW DO 0'&
PDLQWDLQVJHQRPLFVWDELOLW\E\SDUWLFLSDWLQJLQWKHDPSOLILFDWLRQRI$70GHSHQGHQW'1$
GDPDJHVLJQDOV0RO&HOO
:DUG,00LQQ.-RUGD.*&KHQ-$FFXPXODWLRQRIFKHFNSRLQWSURWHLQ%3DW'1$
EUHDNV LQYROYHV LWV ELQGLQJ WR SKRVSKRU\ODWHG KLVWRQH +$; - %LRO &KHP 0D\
3DXOO 77 5RJDNRX (3 <DPD]DNL 9 .LUFKJHVVQHU &8 *HOOHUW 0 %RQQHU :0 $ FULWLFDO
UROH IRU KLVWRQH+$; LQ UHFUXLWPHQW RI UHSDLUIDFWRUV WR QXFOHDU IRFL DIWHU '1$ GDPDJH
&XUU%LRO
6KLORK < 7KH $70PHGLDWHG '1$GDPDJH UHVSRQVH WDNLQJ VKDSH 7UHQGV %LRFKHP 6FL
6WLII 7 2·'ULVFROO 0 5LHI 1 ,ZDEXFKL . /|EULFK 0 -HJJR 3$ $70 DQG '1$3.
IXQFWLRQUHGXQGDQWO\WRSKRVSKRU\ODWH+$;DIWHUH[SRVXUHWRLRQL]LQJUDGLDWLRQ&DQFHU
5HV
5HIHUHQFHV
1RRQ $7 6KLEDWD $ 5LHI 1 /|EULFK 0 6WHZDUW *6 -HJJR 3$ HW DO %3GHSHQGHQW
UREXVW ORFDOL]HG .$3 SKRVSKRU\ODWLRQ LV HVVHQWLDO IRU KHWHURFKURPDWLF '1$ GRXEOH
VWUDQGEUHDNUHSDLU1DW&HOO%LRO
5LEDOOR(.KQH05LHI1'RKHUW\$6PLWK*&5HFLR0-HWDO$SDWKZD\RIGRXEOH
VWUDQG EUHDN UHMRLQLQJ GHSHQGHQW XSRQ $70 $UWHPLV DQG SURWHLQV ORFDWLQJ WR JDPPD
+$;IRFL0RO&HOO
6PLWK - 7KR /0 ;X 1 *LOOHVSLH '$ 7KH$70&KN DQG$75&KNSDWKZD\V LQ'1$
GDPDJHVLJQDOLQJDQGFDQFHU$GY&DQFHU5HV
)LQQ . /RZQGHV 1) *UHQRQ 0 (XNDU\RWLF '1$ GDPDJH FKHFNSRLQW DFWLYDWLRQ LQ
UHVSRQVHWRGRXEOHVWUDQGEUHDNV&HOO0RO/LIH6FL
0F.LQQRQ 3- $70 DQG WKH PROHFXODU SDWKRJHQHVLV RI DWD[LD WHODQJLHFWDVLD $QQX 5HY
3DWKRO
+HUUXS.3RVWPLWRWLFUROHRIWKHFHOOF\FOHPDFKLQHU\&XUU2SLQ&HOO%LRO
9DONR 0 /HLEIULW] ' 0RQFRO - &URQLQ 07 0D]XU 0 7HOVHU - )UHH UDGLFDOV DQG
DQWLR[LGDQWVLQQRUPDOSK\VLRORJLFDOIXQFWLRQVDQGKXPDQGLVHDVH,QW-%LRFKHP&HOO%LRO
<L 0 5RVLQ 03 $QGHUVRQ &. 5HVSRQVH RI ILEUREODVW FXOWXUHV IURP DWD[LDWHODQJLHFWDVLD
SDWLHQWVWRR[LGDWLYHVWUHVV&DQFHU/HWW
:DUG $- 2OLYH 3/ %XUU $+ 5RVLQ 03 5HVSRQVH RI ILEUREODVW FXOWXUHV IURP DWD[LD
WHODQJLHFWDVLDSDWLHQWVWRUHDFWLYHR[\JHQVSHFLHVJHQHUDWHGGXULQJLQIODPPDWRU\UHDFWLRQV
(QYLURQ0RO0XWDJHQ
5HIHUHQFHV
6KDFNHOIRUG5(0DQXV]DN53-RKQVRQ&'+HOOUXQJ'-/LQN&-:DQJ6,URQFKHODWRUV
LQFUHDVHWKHUHVLVWDQFHRI$WD[LDWHODQJLHFWDVLDFHOOVWRR[LGDWLYHVWUHVV'1$5HSDLU
7D\ORU$6KDQJ)1RZHOO7*DODQW\<6KLORK<8ELTXLWLQDWLRQFDSDELOLWLHVLQUHVSRQVHWR
QHRFDU]LQRVWDWLQ DQG +2 VWUHVV LQ FHOO OLQHV IURP SDWLHQWV ZLWK DWD[LDWHODQJLHFWDVLD
2QFRJHQH
&DQPDQ &( :ROII $& &KHQ &< )RUQDFH $- -U .DVWDQ 0% 7KH SGHSHQGHQW * FHOO
F\FOHFKHFNSRLQWSDWKZD\DQGDWD[LDWHODQJLHFWDVLD&DQFHU5HV
-XQJ 0 =KDQJ < /HH 6 'ULWVFKLOR $ &RUUHFWLRQ RI UDGLDWLRQ VHQVLWLYLW\ LQ DWD[LD
WHODQJLHFWDVLDFHOOVE\DWUXQFDWHG,NDSSD%DOSKD6FLHQFH
%HDPLVK + :LOOLDPV 5 &KHQ 3 /DYLQ 0) 'HIHFW LQ PXOWLSOH FHOO F\FOH FKHFNSRLQWV LQ
DWD[LDWHODQJLHFWDVLDSRVWLUUDGLDWLRQ-%LRO&KHP
%DUORZ&'HQQHU\3$6KLJHQDJD0.6PLWK0$0RUURZ-'5REHUWV/-QGHWDO/RVV
RI WKH DWD[LDWHODQJLHFWDVLD JHQH SURGXFW FDXVHV R[LGDWLYH GDPDJH LQ WDUJHW RUJDQV 3URF
1DWO$FDG6FL86$
0HUHGLWK 0- 'RGVRQ 0/ ,PSDLUHG JOXWDWKLRQH ELRV\QWKHVLV LQ FXOWXUHG KXPDQ DWD[LD
WHODQJLHFWDVLDFHOOV&DQFHU5HV
*XR = .R]ORY 6 /DYLQ 0) 3HUVRQ 0' 3DXOO 77 $70 DFWLYDWLRQ E\ R[LGDWLYH VWUHVV
6FLHQFH
&RVHQWLQR & *ULHFR ' &RVWDQ]R 9 $70 DFWLYDWHV WKH SHQWRVH SKRVSKDWH SDWKZD\
SURPRWLQJDQWLR[LGDQWGHIHQFHDQG'1$UHSDLU(0%2-
/L-2:/L:-LDQJ=**KDQEDUL+$2[LGDWLYHVWUHVVDQGQHXURGHJHQHUDWLYHGLVRUGHUV
,QW-0RO6FL
5HIHUHQFHV
'HYLQ $ 5LJRXOHW 0 0HFKDQLVPV RI PLWRFKRQGULDO UHVSRQVH WR YDULDWLRQV LQ HQHUJ\
GHPDQGLQHXNDU\RWLFFHOOV$P-3K\VLRO&HOO3K\VLRO&
.DPVOHU$ 'DLO\ ' +RFKPDQ $ 6WHUQ 1 6KLORK < 5RWPDQ * HW DO ,QFUHDVHG R[LGDWLYH
VWUHVV LQ DWD[LD WHODQJLHFWDVLD HYLGHQFHG E\ DOWHUDWLRQV LQ UHGR[ VWDWH RI EUDLQV IURP
$WPGHILFLHQWPLFH&DQFHU5HV
$PEURVH0*ROGVWLQH-9*DWWL5$,QWULQVLFPLWRFKRQGULDOG\VIXQFWLRQLQ$70GHILFLHQW
O\PSKREODVWRLGFHOOV+XP0RO*HQHW
9DOHQWLQ9HJD <$ 0DFOHDQ .+ 7DLW0XOGHU - 0LODVWD 6 6WHHYHV 0 'RUVH\ )& HW DO
0LWRFKRQGULDOG\VIXQFWLRQLQDWD[LDWHODQJLHFWDVLD%ORRG
6KDUPD1./HEHGHYD07KRPDV7.RYDOHQNR2$6WXPSI-'6KDGHO*6HWDO,QWULQVLF
PLWRFKRQGULDO'1$UHSDLUGHIHFWVLQ$WD[LD7HODQJLHFWDVLD'1$5HSDLU
3HVVLQ -( 6DOWLHO $5 6LJQDOLQJ SDWKZD\V LQ LQVXOLQ DFWLRQ PROHFXODU WDUJHWV RI LQVXOLQ
UHVLVWDQFH-&OLQ,QYHVW
<DQJ'4.DVWDQ0%3DUWLFLSDWLRQRI$70LQLQVXOLQVLJQDOOLQJWKURXJKSKRVSKRU\ODWLRQ
RIH,)(ELQGLQJSURWHLQ1DW&HOO%LRO
9LQLHJUD -* 0DUWtQH] 1 0RGLUDVVDUL 3 +HUQiQGH] /RVD - 3DUDGD &RER & 6iQFKH]
$UpYDOR /RER 9- HW DO )XOO DFWLYDWLRQ RI 3.%$NW LQ UHVSRQVH WR LQVXOLQ RU LRQL]LQJ
UDGLDWLRQLVPHGLDWHGWKURXJK$70-%LRO&KHP
+DODE\0-+LEPD-&+H-<DQJ'4$70SURWHLQNLQDVHPHGLDWHVIXOODFWLYDWLRQRI$NW
DQG UHJXODWHV JOXFRVH WUDQVSRUWHU WUDQVORFDWLRQ E\ LQVXOLQ LQ PXVFOH FHOOV &HOO 6LJQDO
$UPDWD +/ *ROHELRZVNL ' -XQJ '< .R +- .LP -. 6OXVV +. 5HTXLUHPHQW RI WKH
$70S WXPRU VXSSUHVVRU SDWKZD\ IRU JOXFRVH KRPHRVWDVLV 0RO &HOO %LRO 5HIHUHQFHV
3HUHW] 6 -HQVHQ 5 %DVHUJD 5 *OD]HU 30 $70GHSHQGHQW H[SUHVVLRQ RI WKH LQVXOLQOLNH
JURZWK IDFWRU, UHFHSWRU LQ D SDWKZD\ UHJXODWLQJ UDGLDWLRQ UHVSRQVH 3URF 1DWO $FDG 6FL
86$
)ROH\($.DSRRU700LFURWXEXOHDWWDFKPHQWDQGVSLQGOHDVVHPEO\FKHFNSRLQWVLJQDOOLQJ
DWWKHNLQHWRFKRUH1DW5HY0RO&HOO%LRO 6WRO]$(UW\FK1.LHQLW]$9RJHO&6FKQHLGHU9)ULW]%HWDO7KH&+.%5&$WXPRXU
VXSSUHVVRUSDWKZD\HQVXUHVFKURPRVRPDOVWDELOLW\LQKXPDQVRPDWLFFHOOV1DW&HOO%LRO
;LRQJ%/L6$L-6<LQ62X\DQJ<&6XQ6&HWDO%5&$LVUHTXLUHGIRUPHLRWLFVSLQGOH
DVVHPEO\DQGVSLQGOHDVVHPEO\FKHFNSRLQWDFWLYDWLRQLQPRXVHRRF\WHV%LRO5HSURG
=DFKRV * %ODFN (- :DONHU 0 6FRWW 07 9DJQDUHOOL 3 (DUQVKDZ :& HW DO &KN LV
UHTXLUHGIRUVSLQGOHFKHFNSRLQWIXQFWLRQ'HY&HOO
2ULFFKLR ( 6DODGLQR & ,DFRYHOOL 6 6RGGX 6 &XQGDUL ( $70 LV DFWLYDWHG E\ GHIDXOW LQ
PLWRVLV ORFDOL]HV DW FHQWURVRPHV DQG PRQLWRUV PLWRWLF VSLQGOH LQWHJULW\ &HOO &\FOH <DQJ & 7DQJ ; *XR ; 1LLNXUD < .LWDJDZD . &XL . HW DO $XURUD% PHGLDWHG $70
VHULQH SKRVSKRU\ODWLRQ LV UHTXLUHG IRU PLWRWLF $70 DFWLYDWLRQ DQG WKH VSLQGOH
FKHFNSRLQW0RO&HOO
<DQJ&+DR-.RQJ'&XL;=KDQJ::DQJ+HWDO$70PHGLDWHG0DG6HULQH
SKRVSKRU\ODWLRQ UHJXODWHV 0DG GLPHUL]DWLRQ DQG WKH VSLQGOH DVVHPEO\ FKHFNSRLQW
&DUFLQRJHQHVLV
5HIHUHQFHV
6KLJHWD 7 7DNDJL 0 'HOLD ' &KHVVD / ,ZDWD 6 .DQNH < HW DO 'HIHFWLYH FRQWURO RI
DSRSWRVLV DQG PLWRWLF VSLQGOH FKHFNSRLQW LQ KHWHUR]\JRXV FDUULHUV RI $70 PXWDWLRQV
&DQFHU5HV 7DNDJL 0 'HOLD ' &KHVVD / ,ZDWD 6 6KLJHWD 7 .DQNH < HW DO 'HIHFWLYH FRQWURO RI
DSRSWRVLV UDGLRVHQVLWLYLW\ DQG VSLQGOH FKHFNSRLQW LQ DWD[LD WHODQJLHFWDVLD &DQFHU 5HV
6KHQ .& +HQJ + :DQJ < /X 6 /LX * 'HQJ &; HW DO $70 DQG S FRRSHUDWH WR
VXSSUHVVDQHXSORLG\DQGVXEVHTXHQWWXPRUGHYHORSPHQW&DQFHU5HV .DW\DO6/HH<1LWLVV.&'RZQLQJ60/L<6KLPDGD0HWDO$EHUUDQWWRSRLVRPHUDVH
'1$ OHVLRQV DUH SDWKRJHQLF LQ QHXURGHJHQHUDWLYH JHQRPH LQVWDELOLW\ V\QGURPHV 1DW
1HXURVFL
6RUGHW 2 5HGRQ &( *XLURXLOK%DUEDW - 6PLWK 6 6ROLHU 6 'RXDUUH & HW DO $WD[LD
WHODQJLHFWDVLD PXWDWHG DFWLYDWLRQ E\ WUDQVFULSWLRQ DQG WRSRLVRPHUDVH ,LQGXFHG '1$
GRXEOHVWUDQGEUHDNV(0%25HS
3RPPLHU<7RSRLVRPHUDVH,LQKLELWRUVFDPSWRWKHFLQVDQGEH\RQG1DW5HY&DQFHU
+HUUXS . /L - &KHQ - 7KH UROH RI $70 DQG '1$ GDPDJH LQ QHXURQV XSVWUHDP DQG
GRZQVWUHDPFRQQHFWLRQV'1$5HSDLU
/L-+DQ<53OXPPHU05+HUUXS.&\WRSODVPLF$70LQQHXURQVPRGXODWHVV\QDSWLF
IXQFWLRQ&XUU%LRO
6FKZHL]HU )( 5\DQ 7$ 7KH V\QDSWLF YHVLFOH F\FOH RI H[RF\WRVLV DQG HQGRF\WRVLV &XUU
2SLQ1HXURELRO
5HIHUHQFHV
6LGGRZD\%+RX+<DQJ+3HWUDOLD5;LD+6\QDSWLFDFWLYLW\ELGLUHFWLRQDOO\UHJXODWHVD
QRYHOVHTXHQFHVSHFLILF64SKRVSKRSURWHRPHLQQHXURQV-1HXURFKHP
.LP 76 .DZDJXFKL 0 6X]XNL 0 -XQJ &* $VDL . 6KLEDPRWR < HW DO 7KH =)+;
$7%) WUDQVFULSWLRQ IDFWRU LQGXFHV 3'*)5% ZKLFK DFWLYDWHV $70 LQ WKH F\WRSODVP WR
SURWHFWFHUHEHOODUQHXURQVIURPR[LGDWLYHVWUHVV'LV0RGHO0HFK
/DX$7\PLDQVNL0*OXWDPDWHUHFHSWRUVQHXURWR[LFLW\DQGQHXURGHJHQHUDWLRQ3IOXJHUV
$UFK
,VKLL < 2\D 7 =KHQJ / *DR = .DZDJXFKL 0 6DELW + HW DO 0RXVH EUDLQV GHILFLHQW LQ
QHXURQDO3'*)UHFHSWRUEHWDGHYHORSQRUPDOO\EXWDUHYXOQHUDEOHWRLQMXU\-1HXURFKHP
6ZHDWW-'7KHHPHUJLQJILHOGRIQHXURHSLJHQHWLFV1HXURQ
/L-&KHQ-5LFXSHUR&/+DUW536FKZDUW]06.XVQHFRY$HWDO1XFOHDUDFFXPXODWLRQ
RI +'$& LQ $70 GHILFLHQF\ SURPRWHV QHXURGHJHQHUDWLRQ LQ DWD[LD WHODQJLHFWDVLD 1DW
0HG
/L-+DUW530DOOLPR(06ZHUGHO05.XVQHFRY$:+HUUXS.(=+PHGLDWHG+.
WULPHWK\ODWLRQ PHGLDWHV QHXURGHJHQHUDWLRQ LQ DWD[LDWHODQJLHFWDVLD 1DW 1HXURVFL 5LEHLUR)0&DPDUJRV(5GH6RX]D/&7HL[HLUD$/$QLPDOPRGHOVRIQHXURGHJHQHUDWLYH
GLVHDVHV5HY%UDV3VLTXLDWU6XSSO6
/HQ] 6 .DUVWHQ 3 6FKXO] -% 9RLJW $ 'URVRSKLOD DV D VFUHHQLQJ WRRO WR VWXG\ KXPDQ
QHXURGHJHQHUDWLYHGLVHDVHV-1HXURFKHP
5HIHUHQFHV
5LPNXV 6$ .DW]HQEHUJHU 5- 7ULQK $7 'RGVRQ *( 7LEEHWWV 56 :DVVDUPDQ '$
0XWDWLRQV LQ 6WULQJ&'& LQKLELW FHOO F\FOH UHHQWU\ DQG QHXURGHJHQHUDWLRQ LQ D
'URVRSKLODPRGHORI$WD[LDWHODQJLHFWDVLD*HQHV'HY
3HWHUVHQ$-:DVVDUPDQ'$'URVRSKLODLQQDWHLPPXQHUHVSRQVHSDWKZD\VPRRQOLJKWLQ
QHXURGHJHQHUDWLRQ)O\$XVWLQ
3HWHUVHQ$-.DW]HQEHUJHU5-:DVVDUPDQ'$7KHLQQDWHLPPXQHUHVSRQVHWUDQVFULSWLRQ
IDFWRU UHOLVK LV QHFHVVDU\ IRU QHXURGHJHQHUDWLRQ LQ D 'URVRSKLOD PRGHO RI DWD[LD
WHODQJLHFWDVLD*HQHWLFV
1JX\HQ';X77KHH[SDQGLQJUROHRIPRXVHJHQHWLFVIRUXQGHUVWDQGLQJKXPDQELRORJ\
DQGGLVHDVH'LV0RGHO0HFK
%DUORZ & +LURWVXQH 6 3D\ORU 5 /L\DQDJH 0 (FNKDXV 0 &ROOLQV ) HW DO $WPGHILFLHQW
PLFHDSDUDGLJPRIDWD[LDWHODQJLHFWDVLD&HOO
(OVRQ $ :DQJ < 'DXJKHUW\ &- 0RUWRQ && =KRX ) &DPSRV7RUUHV - HW DO 3OHLRWURSLF
GHIHFWV LQ DWD[LDWHODQJLHFWDVLD SURWHLQGHILFLHQW PLFH 3URF 1DWO $FDG 6FL 8 6 $ ;X < %DOWLPRUH ' 'XDO UROHV RI $70 LQ WKH FHOOXODU UHVSRQVH WR UDGLDWLRQ DQG LQ FHOO
JURZWKFRQWURO*HQHV'HY
+HU]RJ .+ &KRQJ 0- .DSVHWDNL 0 0RUJDQ -, 0F.LQQRQ 3- 5HTXLUHPHQW IRU $WP LQ
LRQL]LQJ UDGLDWLRQLQGXFHG FHOO GHDWK LQ WKH GHYHORSLQJ FHQWUDO QHUYRXV V\VWHP 6FLHQFH
6SULQJ . &URVV 6 /L & :DWWHUV ' %HQ6HQLRU / :DULQJ 3 HW DO $WP NQRFNLQ PLFH
KDUERULQJ DQ LQIUDPH GHOHWLRQ FRUUHVSRQGLQJ WR WKH KXPDQ $70 GHO FRPPRQ
PXWDWLRQH[KLELWDYDULDQWSKHQRW\SH&DQFHU5HV
5HIHUHQFHV
%RUJKHVDQL 35 $OW ): %RWWDUR $ 'DYLGVRQ / $NVR\ 6 5DWKEXQ *$ HW DO $EQRUPDO
GHYHORSPHQWRI3XUNLQMHFHOOVDQGO\PSKRF\WHVLQ$WPPXWDQWPLFH3URF1DWO$FDG6FL8
6$
.XOMLV52;X<$JXLOD0&%DOWLPRUH''HJHQHUDWLRQRIQHXURQVV\QDSVHVDQGQHXURSLO
DQG JOLDO DFWLYDWLRQ LQ D PXULQH $WP NQRFNRXW PRGHO RI DWD[LDWHODQJLHFWDVLD 3URF 1DWO
$FDG6FL86$
/DYLQ0)7KHDSSURSULDWHQHVVRIWKHPRXVHPRGHOIRUDWD[LDWHODQJLHFWDVLDQHXURORJLFDO
GHIHFWVEXWQRQHXURGHJHQHUDWLRQ'1$5HSDLU
%LWRQ6%DU]LODL$6KLORK<7KHQHXURORJLFDOSKHQRW\SHRIDWD[LDWHODQJLHFWDVLDVROYLQJD
SHUVLVWHQWSX]]OH'1$5HSDLU
&KLHVD 1 %DUORZ & :\QVKDZ%RULV $ 6WUDWD 3 7HPSLD ) $WPGHILFLHQW PLFH 3XUNLQMH
FHOOV VKRZ DJHGHSHQGHQW GHIHFWV LQ FDOFLXP VSLNH EXUVWV DQG FDOFLXP FXUUHQWV
1HXURVFLHQFH
(LODP53HWHU<*URQHU<6HJDO0/DWHGHJHQHUDWLRQRIQLJURVWULDWDOQHXURQVLQ$70
PLFH1HXURVFLHQFH
&DUOHVVL/)XVDU3ROL('H)LOLSSLV/'HOLD'$70GHILFLHQWKXPDQQHXUDOVWHPFHOOVDV
DQLQYLWURPRGHOV\VWHPWRVWXG\QHXURGHJHQHUDWLRQ'1$5HSDLU
%RHKUV -. +H - +DODE\ 0- <DQJ '4 &RQVWLWXWLYH H[SUHVVLRQ DQG F\WRSODVPLF
FRPSDUWPHQWDOL]DWLRQRI$70SURWHLQLQGLIIHUHQWLDWHGKXPDQQHXURQOLNH6+6<<FHOOV-
1HXURFKHP
%LWRQ 6 'DU , 0LWWHOPDQ / 3HUHJ < %DU]LODL $ 6KLORK < 1XFOHDU DWD[LDWHODQJLHFWDVLD
PXWDWHG $70 PHGLDWHV WKH FHOOXODU UHVSRQVH WR '1$ GRXEOH VWUDQG EUHDNV LQ KXPDQ
QHXURQOLNHFHOOV-%LRO&KHP
5HIHUHQFHV
/L < ;LRQJ + <DQJ '4 )XQFWLRQDO VZLWFKLQJ RI $70 VHQVRU RI '1$ GDPDJH LQ
SUROLIHUDWLQJ FHOOV DQG PHGLDWRU RI $NW VXUYLYDO VLJQDO LQ SRVWPLWRWLF KXPDQ QHXURQOLNH
FHOOV&KLQ-&DQFHU
$JKROPH / /LQGVWU|P 7 .nJHGDO . 0DUFXVVRQ - +DOOEHFN 0 $Q LQ YLWUR PRGHO IRU
QHXURVFLHQFH GLIIHUHQWLDWLRQ RI 6+6<< FHOOV LQWR FHOOV ZLWK PRUSKRORJLFDO DQG
ELRFKHPLFDOFKDUDFWHULVWLFVRIPDWXUHQHXURQV-$O]KHLPHUV'LV
%LWRQ6*URSS0,WV\NVRQ33HUHJ<0LWWHOPDQ/-RKH.HWDO$70PHGLDWHGUHVSRQVH
WR '1$ GRXEOH VWUDQG EUHDNV LQ KXPDQ QHXURQV GHULYHG IURP VWHP FHOOV '1$ 5HSDLU
$GDPV %5 *ROGLQJ 6( 5DR 55 9DOHULH . '\QDPLF GHSHQGHQFH RQ $75 DQG $70 IRU
GRXEOHVWUDQGEUHDNUHSDLULQKXPDQHPEU\RQLFVWHPFHOOVDQGQHXUDOGHVFHQGDQWV3/R6
2QHH
%URFFROL 9 *LDQQHOOL 6* 0D]]DUD 3* 0RGHOLQJ SK\VLRORJLFDO DQG SDWKRORJLFDO KXPDQ
QHXURJHQHVLVLQWKHGLVK)URQW1HXURVFL
%DOGZLQ 7 0RUDOLW\ DQG KXPDQ HPEU\R UHVHDUFK ,QWURGXFWLRQ WR WKH 7DONLQJ 3RLQW RQ
PRUDOLW\DQGKXPDQHPEU\RUHVHDUFK(0%25HS
'H)LOLSSLV/%LQGD(&RQFLVHUHYLHZVHOIUHQHZDOLQWKHFHQWUDOQHUYRXVV\VWHPQHXUDO
VWHPFHOOVIURPHPEU\RWRDGXOW6WHP&HOOV7UDQVO0HG
-DNHO5-6FKQHLGHU%/6YHQGVHQ&18VLQJKXPDQQHXUDOVWHPFHOOVWRPRGHOQHXURORJLFDO
GLVHDVH1DW5HY*HQHW
+LFNVRQ , =KDR < 5LFKDUGVRQ &- *UHHQ 6- 0DUWLQ 10 2UU $, 5HDSHU 30 HW DO
,GHQWLILFDWLRQ DQG FKDUDFWHUL]DWLRQ RI D QRYHO DQG VSHFLILF LQKLELWRU RI WKH DWD[LD
WHODQJLHFWDVLDPXWDWHGNLQDVH$70&DQFHU5HV
5HIHUHQFHV
&DUOHVVL/'H)LOLSSLV//HFLV'9HVFRYL$'HOLD''1$GDPDJHUHVSRQVHVXUYLYDODQG
GLIIHUHQWLDWLRQLQYLWURRIDKXPDQQHXUDOVWHPFHOOOLQHLQUHODWLRQWR$70H[SUHVVLRQ&HOO
'HDWK'LIIHU
*LODG 6 &KHVVD / .KRVUDYL 5 5XVVHOO 3 *DODQW\ < 3LDQH 0 HW DO *HQRW\SHSKHQRW\SH
UHODWLRQVKLSVLQDWD[LDWHODQJLHFWDVLDDQGYDULDQWV$P-+XP*HQHW
'DQLHO -$ 3HOOHJULQL 0 /HH %6 *XR = )LOVXI ' %HONLQD 19 HW DO /RVV RI $70 NLQDVH
DFWLYLW\OHDGVWRHPEU\RQLFOHWKDOLW\LQPLFH-&HOO%LRO
6WHZDUW 5 .R]ORY 6 0DWLJLDQ 1 :DOL * *DWHL 0 6XWKDUVDQ 5 HW DO $ SDWLHQWGHULYHG
ROIDFWRU\ VWHP FHOO GLVHDVH PRGHO IRU DWD[LDWHODQJLHFWDVLD +XP 0RO *HQHW 7DNDKDVKL.<DPDQDND6,QGXFWLRQRISOXULSRWHQWVWHPFHOOVIURPPRXVHHPEU\RQLFDQG
DGXOWILEUREODVWFXOWXUHVE\GHILQHGIDFWRUV&HOO
7DNDKDVKL . 7DQDEH . 2KQXNL 0 1DULWD 0 ,FKLVDND 7 7RPRGD . HW DO ,QGXFWLRQ RI
SOXULSRWHQW VWHP FHOOV IURP DGXOW KXPDQ ILEUREODVWV E\ GHILQHG IDFWRUV &HOO =KRX<<=HQJ),QWHJUDWLRQIUHHPHWKRGVIRUJHQHUDWLQJLQGXFHGSOXULSRWHQWVWHPFHOOV
*HQRPLFV3URWHRPLFV%LRLQIRUPDWLFV
2NDQR + <DPDQDND 6 L36 FHOO WHFKQRORJLHV VLJQLILFDQFH DQG DSSOLFDWLRQV WR &16
UHJHQHUDWLRQDQGGLVHDVH0RO%UDLQ
,VUDHO 0$ <XDQ 6+ %DUG\ & 5H\QD 60 0X < +HUUHUD & HW DO 3URELQJ VSRUDGLF DQG
IDPLOLDO $O]KHLPHU
V GLVHDVH XVLQJ LQGXFHG SOXULSRWHQW VWHP FHOOV 1DWXUH 5HIHUHQFHV
.RQGR7$VDL07VXNLWD..XWRNX<2KVDZD<6XQDGD<HWDO0RGHOLQJ$O]KHLPHU
V
GLVHDVH ZLWK L36&V UHYHDOV VWUHVV SKHQRW\SHV DVVRFLDWHG ZLWK LQWUDFHOOXODU $ǃ DQG
GLIIHUHQWLDOGUXJUHVSRQVLYHQHVV&HOO6WHP&HOO
,PDL]XPL < 2NDGD < $NDPDWVX : .RLNH 0 .X]XPDNL 1 +D\DNDZD + HW DO
0LWRFKRQGULDOG\VIXQFWLRQDVVRFLDWHGZLWKLQFUHDVHGR[LGDWLYHVWUHVVDQGDOSKDV\QXFOHLQ
DFFXPXODWLRQ LQ 3$5. L36&GHULYHG QHXURQV DQG SRVWPRUWHP EUDLQ WLVVXH 0RO %UDLQ
5HLQKDUGW 3 6FKPLG % %XUEXOOD /) 6FK|QGRUI '& :DJQHU / *ODW]D 0 HW DO *HQHWLF
FRUUHFWLRQRID/55.PXWDWLRQLQKXPDQL36&VOLQNVSDUNLQVRQLDQQHXURGHJHQHUDWLRQWR
(5.GHSHQGHQWFKDQJHVLQJHQHH[SUHVVLRQ&HOO6WHP&HOO
'LPRV-75RGROID.71LDNDQ..:HLVHQWKDO/00LWVXPRWR+&KXQJ:HWDO,QGXFHG
SOXULSRWHQW VWHP FHOOV JHQHUDWHG IURP SDWLHQWV ZLWK $/6 FDQ EH GLIIHUHQWLDWHG LQWR PRWRU
QHXURQV6FLHQFH²
(JDZD1.LWDRND67VXNLWD.1DLWRK07DNDKDVKL.<DPDPRWR7HWDO'UXJVFUHHQLQJ
IRU $/6 XVLQJ SDWLHQWVSHFLILF LQGXFHG SOXULSRWHQW VWHP FHOOV 6FL 7UDQVO 0HG UD
-HRQ,/HH1/L-<3DUN,+3DUN.60RRQ-HWDO1HXURQDOSURSHUWLHVLQYLYRHIIHFWV
DQG SDWKRORJ\ RI D +XQWLQJWRQ
V GLVHDVH SDWLHQWGHULYHG LQGXFHG SOXULSRWHQW VWHP FHOOV
6WHP&HOOV
-XRSSHUL 7$ .LP :5 &KLDQJ &+ <X + 0DUJROLV 5/ 5RVV &$ HW DO $VWURF\WHV
JHQHUDWHGIURPSDWLHQWLQGXFHGSOXULSRWHQWVWHPFHOOVUHFDSLWXODWHIHDWXUHVRI+XQWLQJWRQ
V
GLVHDVHSDWLHQWFHOOV0RO%UDLQ
5HIHUHQFHV
&KDQJ 7 =KHQJ : 7VDUN : %DWHV 6 +XDQJ + /LQ 5- HW DO %ULHI UHSRUW SKHQRW\SLF
UHVFXHRILQGXFHGSOXULSRWHQWVWHPFHOOGHULYHGPRWRQHXURQVRIDVSLQDOPXVFXODUDWURSK\
SDWLHQW6WHP&HOOV
5LFFLDUGL68QJDUR)+DPEURFN05DGHPDFKHU16WHIDQHOOL*%UDPELOOD'HWDO&'./
HQVXUHV H[FLWDWRU\ V\QDSVH VWDELOLW\ E\ UHLQIRUFLQJ 1*/36' LQWHUDFWLRQ LQ WKH
SRVWV\QDSWLFFRPSDUWPHQWDQGLVLPSDLUHGLQSDWLHQWL36&GHULYHGQHXURQV1DW&HOO%LRO
$QDQLHY*:LOOLDPV(&/L+&KDQJ4,VRJHQLFSDLUVRIZLOGW\SHDQGPXWDQWLQGXFHG
SOXULSRWHQW VWHP FHOO L36& OLQHV IURP 5HWW V\QGURPH SDWLHQWV DV LQ YLWUR GLVHDVH PRGHO
3/R62QHH
1D\OHU6*DWHL0.R]ORY6*DWWL5$0DU-&:HOOV&$HWDO,QGXFHGSOXULSRWHQWVWHP
FHOOVIURPDWD[LDWHODQJLHFWDVLDUHFDSLWXODWHWKHFHOOXODUSKHQRW\SH6WHP&HOOV7UDQVO0HG
/HH 3 0DUWLQ 17 1DNDPXUD . $]JKDGL 6 $PLUL 0 %HQ'DYLG 8 HW DO 6057
FRPSRXQGV DEURJDWH FHOOXODU SKHQRW\SHV RI DWD[LD WHODQJLHFWDVLD LQ QHXUDO GHULYDWLYHV RI
SDWLHQWVSHFLILFKL36&V1DW&RPPXQ
2NLWD.0DWVXPXUD<6DWR<2NDGD$0RUL]DQH$2NDPRWR6HWDO$PRUHHIILFLHQW
PHWKRGWRJHQHUDWHLQWHJUDWLRQIUHHKXPDQL36FHOOV1DW0HWKRGV
<RVKLGD < 7DNDKDVKL . 2NLWD . ,FKLVDND 7 <DPDQDND 6 +\SR[LD HQKDQFHV WKH
JHQHUDWLRQRILQGXFHGSOXULSRWHQWVWHPFHOOV&HOO6WHP&HOO
,WVNRYLW](OGRU - 6FKXOGLQHU 0 .DUVHQWL ' (GHQ $ <DQXND 2 $PLW 0 HW DO
'LIIHUHQWLDWLRQ RI KXPDQ HPEU\RQLF VWHP FHOOV LQWR HPEU\RLG ERGLHV FRPSURPLVLQJ WKH
WKUHHHPEU\RQLFJHUPOD\HUV0RO0HG
5HIHUHQFHV
:LOVRQ 3* 6WLFH 66 'HYHORSPHQW DQG GLIIHUHQWLDWLRQ RI QHXUDO URVHWWHV GHULYHG IURP
KXPDQHPEU\RQLFVWHPFHOOV6WHP&HOO5HY
6DQWLOOL * /DPRUWH * &DUOHVVL / )HUUDUL ' 5RWD 1RGDUL / %LQGD ( HW DO 0LOG K\SR[LD
HQKDQFHV SUROLIHUDWLRQ DQG PXOWLSRWHQF\ RI KXPDQ QHXUDO VWHP FHOOV 3/R6 2QH H
&DOLQL98UDQL&&DPDWLQL0&RPHWDVVD\HYDOXDWLRQRI'1$VLQJOHDQGGRXEOHVWUDQG
EUHDNVLQGXFWLRQDQGUHSDLULQ&+7FHOOV&HOO%LRO7R[LFRO
$ODJR]0&KLDQJ6&6KDUPD$(O.KDPLV\6)$70GHILFLHQF\UHVXOWVLQDFFXPXODWLRQ
RI '1$WRSRLVRPHUDVH , FRYDOHQW LQWHUPHGLDWHV LQ QHXUDO FHOOV 3/R6 2QH H
)URVLQD * &DSSHOOL ( 5RSROR 0 )RUWLQL3 3DVFXFFL % 'RJOLRWWL ( ,Q YLWUR EDVH H[FLVLRQ
UHSDLUDVVD\XVLQJPDPPDOLDQFHOOH[WUDFWV0HWKRGV0RO%LRO
0DWVXPRWR<.LP.%RJHQKDJHQ')3UROLIHUDWLQJFHOOQXFOHDUDQWLJHQGHSHQGHQWDEDVLF
VLWHUHSDLULQ;HQRSXVODHYLVRRF\WHVDQDOWHUQDWLYHSDWKZD\RIEDVHH[FLVLRQ'1$UHSDLU
0RO&HOO%LRO
.LLDQLWVD.0DL]HOV1$UDSLGDQGVHQVLWLYHDVVD\IRU'1$SURWHLQFRYDOHQWFRPSOH[HVLQ
OLYLQJFHOOV1XFOHLF$FLGV5HVH
9HUSHOOL & &DUOHVVL / %HFKL * )XVDU 3ROL ( 2UHOODQD ' +HLVH & HW DO &RPSDUDWLYH
QHXURQDOGLIIHUHQWLDWLRQRIVHOIUHQHZLQJQHXUDOSURJHQLWRUFHOOOLQHVREWDLQHGIURPKXPDQ
LQGXFHGSOXULSRWHQWVWHPFHOOV)URQW&HOO1HXURVFL
%HQLWH]%ULELHVFDO / 6DQFKH]6XDUH] 3 2[LGDWLYH GDPDJH EOHRP\FLQ DQG JDPPD
UDGLDWLRQ LQGXFH GLIIHUHQW W\SHV RI '1$ VWUDQG EUHDNV LQ QRUPDO O\PSKRF\WHV DQG
WK\PRF\WHVDFRPHWDVVD\VWXG\$QQ1<$FDG6FL²
5HIHUHQFHV
/|EULFK 0 6KLEDWD $ %HXFKHU $ )LVKHU $ (QVPLQJHU 0 *RRGDU]L $$ HW DO
*DPPD+$; IRFL DQDO\VLV IRU PRQLWRULQJ '1$ GRXEOHVWUDQG EUHDN UHSDLU VWUHQJWKV
OLPLWDWLRQVDQGRSWLPL]DWLRQ&HOO&\FOH
;X*+HU]LJ05RWUHNO9:DOWHU&$%DVHH[FLVLRQUHSDLUDJLQJDQGKHDOWKVSDQ0HFK
$JHLQJ'HY²
+HJGH 0/ +HJGH 30 5DR .6 0LWUD 6 2[LGDWLYH JHQRPH GDPDJH DQG LWV UHSDLU LQ
QHXURGHJHQHUDWLYH GLVHDVHV IXQFWLRQ RI WUDQVLWLRQ PHWDOV DV D GRXEOHHGJHG VZRUG -
$O]KHLPHUV'LV6XSSO²
6\NRUD3<DQJ-/)HUUDUHOOL/.7LDQ-7DGRNRUR7.XONDUQL$HWDO0RGXODWLRQRI'1$
EDVHH[FLVLRQUHSDLUGXULQJQHXURQDOGLIIHUHQWLDWLRQ1HXURELRO$JLQJ²
0LQX]]R00DUFKLQL6%URJJLQL0)DLUFORWK*'·,QFDOFL00DQWRYDQL5,QWHUIHUHQFHRI
WUDQVFULSWLRQDODFWLYDWLRQE\WKHDQWLQHRSODVWLFGUXJHFWHLQDVFLGLQ3URF1DWO$FDG6FL
86$²
0F&DUWK\66RPD\DMXOX06LNRUVND0%RURZ\%RURZVNL+3DQGH\63DUDTXDWLQGXFHV
R[LGDWLYHVWUHVVDQGQHXURQDOFHOOGHDWKQHXURSURWHFWLRQE\ZDWHUVROXEOHFRHQ]\PH4
7R[LFRO$SSO3KDUPDFRO²
6KLORK < $70 DQG UHODWHG SURWHLQ NLQDVHV VDIHJXDUGLQJ JHQRPH LQWHJULW\ 1DW 5HY
&DQFHU
0RPFLORYLþ 2 &KRL 6 9DUXP 6 %DNNHQLVW & 6FKDWWHQ * 1DYDUD & ,RQL]LQJ UDGLDWLRQ
LQGXFHV DWD[LD WHODQJLHFWDVLD PXWDWHGGHSHQGHQW FKHFNSRLQW VLJQDOLQJ DQG * EXW QRW
* FHOO F\FOH DUUHVW LQ SOXULSRWHQW KXPDQ HPEU\RQLF VWHP FHOOV 6WHP &HOOV 5HIHUHQFHV
0RPFLORYLþ 2 .QREORFK / )RUQVDJOLR - 9DUXP 6 (DVOH\ & 6FKDWWHQ * '1$ GDPDJH
UHVSRQVHV LQ KXPDQ LQGXFHG SOXULSRWHQW VWHP FHOOV DQG HPEU\RQLF VWHP FHOOV 3/R6 2QH
H
4LDQJ / )XMLWD 5 $EHOLRYLFK $ 5HPRGHOLQJ QHXURGHJHQHUDWLRQ VRPDWLF FHOO
UHSURJUDPPLQJEDVHGPRGHOVRIDGXOWQHXURORJLFDOGLVRUGHUV1HXURQ
:RRGELQH/%UXQWRQ+*RRGDU]L$$6KLEDWD$-HJJR3$(QGRJHQRXVO\LQGXFHG'1$
GRXEOH VWUDQG EUHDNV DULVH LQ KHWHURFKURPDWLF '1$ UHJLRQV DQG UHTXLUH DWD[LD
WHODQJLHFWDVLDPXWDWHGDQG$UWHPLVIRUWKHLUUHSDLU1XFOHLF$FLGV5HV²
/HH < &KRQJ 0- 0F.LQQRQ 3- $WD[LD WHODQJLHFWDVLD PXWDWHGGHSHQGHQW DSRSWRVLV DIWHU
JHQRWR[LFVWUHVVLQWKHGHYHORSLQJQHUYRXVV\VWHPLVGHWHUPLQHGE\FHOOXODUGLIIHUHQWLDWLRQ
VWDWXV-1HXURVFL²
&KRX :& :DQJ +& :RQJ )+ 'LQJ 6/ :X 3( 6KLHK 6< HW DO &KNGHSHQGHQW
SKRVSKRU\ODWLRQ RI ;5&& LQ WKH '1$ GDPDJH UHVSRQVH SURPRWHV EDVH H[FLVLRQ UHSDLU
(0%2-²
6XUDZHHUD $ %HFKHUHO 2- &KHQ 3 5XQGOH 1 :RRGV 5 1DNDPXUD - HW DO 6HQDWD[LQ
GHIHFWLYHLQDWD[LDRFXORPRWRUDSUD[LDW\SHLVLQYROYHGLQWKHGHIHQVHDJDLQVWR[LGDWLYH
'1$GDPDJH-&HOO%LRO²
7LDQ % <DQJ 4 0DR = 3KRVSKRU\ODWLRQ RI $70 E\ &GN PHGLDWHV '1$ GDPDJH
VLJQDOOLQJDQGUHJXODWHVQHXURQDOGHDWK1DW&HOO%LRO²
6ROLHU65\DQ0&0DUWLQ6(9DUPD6.RKQ.:/LX+HWDO7UDQVFULSWLRQSRLVRQLQJE\
7RSRLVRPHUDVH , LV FRQWUROOHG E\ JHQH OHQJWK VSOLFH VLWHV DQG PL5S &DQFHU 5HV
²
5HIHUHQFHV
*ODVV'9LQXHOD$'DYLHV015DPDVDP\$3DUWV/.QRZOHV'HWDO*HQHH[SUHVVLRQ
FKDQJHVZLWKDJHLQVNLQDGLSRVHWLVVXHEORRGDQGEUDLQ*HQRPH%LRO5
%HUFKWROG1&&ROHPDQ3'&ULEEV'+5RJHUV-*LOOHQ'/&RWPDQ&:6\QDSWLFJHQHV
DUH H[WHQVLYHO\ GRZQUHJXODWHG DFURVV PXOWLSOH EUDLQ UHJLRQV LQ QRUPDOKXPDQ DJLQJDQG
$O]KHLPHU·VGLVHDVH1HXURELRO$JLQJ²
/X73DQ<.DR6</L&.RKDQH,&KDQ-HWDO*HQHUHJXODWLRQDQG'1$GDPDJHLQWKH
DJHLQJKXPDQEUDLQ1DWXUH²
$EGHODOLP (0 0ROHFXODU PHFKDQLVPV FRQWUROOLQJ WKH FHOO F\FOH LQ HPEU\RQLF VWHP FHOOV
6WHP&HOO5HY²
0RPFLORYLþ21DYDUD&6FKDWWHQ*&HOOF\FOHDGDSWDWLRQVDQGPDLQWHQDQFHRIJHQRPLF
LQWHJULW\ LQ HPEU\RQLF VWHP FHOOV DQG LQGXFHG SOXULSRWHQW VWHP FHOOV 5HVXOWV 3UREO &HOO
'LIIHU²
7LFK\('3LOODL5'HQJ//LDQJ/7LVFKILHOG-6FKZHPEHUJHU6-HWDO0RXVHHPEU\RQLF
VWHP FHOOV EXW QRW VRPDWLF FHOOV SUHGRPLQDQWO\ XVH KRPRORJRXV UHFRPELQDWLRQ WR UHSDLU
GRXEOHVWUDQG'1$EUHDNV6WHP&HOOV'HY²
-RQJNDPRQZLZDW11RLVD3%LRPHGLFDODQGFOLQLFDOSURPLVHVRIKXPDQSOXULSRWHQWVWHP
FHOOVIRUQHXURORJLFDOGLVRUGHUV%LRPHG5HV,QW
%DU]LODL$7KHQHXURJOLDOYDVFXODULQWHUUHODWLRQVLQJHQRPLFLQVWDELOLW\V\PSWRPV0HFK
$JHLQJ'HY
%DOWDQiV)&&DVDIRQW,/DIDUJD9:HUXDJD($ORQVR-5%HUFLDQR07HWDO3XUNLQMHFHOO
GHJHQHUDWLRQLQSFGPLFHUHYHDOVODUJHVFDOHFKURPDWLQUHRUJDQL]DWLRQDQGJHQHVLOHQFLQJ
OLQNHGWRGHIHFWLYH'1$UHSDLU-%LRO&KHP
5HIHUHQFHV
7]XU*LODW $ =LY < 0LWWHOPDQ / %DU]LODL $ 6KLORK < 6WXG\LQJ WKH FHUHEHOODU '1$
GDPDJHUHVSRQVHLQWKHWLVVXHFXOWXUHGLVK0HFK$JHLQJ'HY
7DNDKDVKL7%XUJXLHUH6OH]DN*9DQGHU.HPS3$%RLWHX[67RSRLVRPHUDVHSURYRNHV
WKH IRUPDWLRQ RI VKRUW GHOHWLRQV LQ UHSHDWHG VHTXHQFHV XSRQ KLJK WUDQVFULSWLRQ LQ
6DFFKDURP\FHVFHUHYLVLDH3URF1DWO$FDG6FL86$
)RQJ<:&DWWRJOLR&7MLDQ57KHLQWHUWZLQHGUROHVRIWUDQVFULSWLRQDQGUHSDLUSURWHLQV
0RO&HOO
'HSNHQ 0 3DUURQGR -0 *ULOO 6: ,QWHUPLWWHQW WUDQVFULSWLRQ G\QDPLFV IRU WKH UDSLG
SURGXFWLRQRIORQJWUDQVFULSWVRIKLJKILGHOLW\&HOO5HS
'XFKHQ 05 0LWRFKRQGULD FDOFLXPGHSHQGHQW QHXURQDO GHDWK DQG QHXURGHJHQHUDWLYH
GLVHDVH3IOXJHUV$UFK
&KHQ33HQJ&/XII-6SULQJ.:DWWHUV'%RWWOH6HWDO2[LGDWLYHVWUHVVLVUHVSRQVLEOH
IRU GHILFLHQW VXUYLYDO DQG GHQGULWRJHQHVLV LQ 3XUNLQMH QHXURQV IURP DWD[LDWHODQJLHFWDVLD
PXWDWHGPXWDQWPLFH-1HXURVFL
<DQ - .KDQQD .. /DYLQ 0) 'HIHFWLYH UDGLDWLRQ VLJQDO WUDQVGXFWLRQ LQ DWD[LD
WHODQJLHFWDVLDFHOOV,QW-5DGLDW%LRO
*XJJHU 26 .DSIKDPPHU -3 5HGXFHG VL]H RI WKH GHQGULWLF WUHH GRHVQRW SURWHFW 3XUNLQMH
FHOOVIURPH[FLWRWR[LFGHDWK-1HXURVFL5HV
0XJXUXPD . 1LVKL\DPD $ 2QR < 0L\DZDNL + 0L]XKDUD ( +RUL 6 HW DO 2QWRJHQ\
UHFDSLWXODWLQJ JHQHUDWLRQ DQG WLVVXH LQWHJUDWLRQ RI (6 FHOOGHULYHG 3XUNLQMH FHOOV 1DW
1HXURVFL
/HWR . 5RODQGR & 5RVVL ) 7KH JHQHVLV RI FHUHEHOODU *$%$HUJLF QHXURQV IDWH SRWHQWLDO
DQGVSHFLILFDWLRQPHFKDQLVPV)URQW1HXURDQDW
5HIHUHQFHV
0XJXUXPD.6DVDL<,QYLWURUHFDSLWXODWLRQRIQHXUDOGHYHORSPHQWXVLQJHPEU\RQLFVWHP
FHOOVIURPQHXURJHQHVLVWRKLVWRJHQHVLV'HY*URZWK'LIIHU
0DJUDVVL / 9LVLRQJXLGHG WHFKQLTXH IRU FHOO WUDQVSODQWDWLRQ DQG LQMHFWLRQ RI DFWLYH
PROHFXOHVLQWRUDWDQGPRXVHHPEU\RV0HWKRGV0RO%LRO
3XEOLFDWLRQVLQ,QWHUQDWLRQDO,6,
-RXUQDOV
3XEOLFDWLRQVLQ,QWHUQDWLRQDO,6,-RXUQDOV
5HODWHGWRWKHSURMHFW
&DUOHVVL/)XVDU3ROL(%HFKL*0DQWHJD]]D03DVFXFFL%1DUFLVR/'RJOLRWWL(6DOD&
9HUSHOOL & /HFLV ' 'HOLD ' )XQFWLRQDO DQG PROHFXODU GHIHFWV RI KL36&GHULYHG QHXURQV
IURPSDWLHQWVZLWK$70GHILFLHQF\&HOO'HDWK'LVH
&DUOHVVL/)XVDU3ROL('HOLD'%UDLQDQGLQGXFHGSOXULSRWHQWVWHPFHOOGHULYHGQHXUDOVWHP
FHOOV DV DQ LQ YLWUR PRGHO RI QHXURGHJHQHUDWLRQ LQ DWD[LDWHODQJLHFWDVLD ([S %LRO 0HG
0D\ZRRG
&DUOHVVL/)XVDU3ROL('H)LOLSSLV/'HOLD'$70GHILFLHQWKXPDQQHXUDOVWHPFHOOVDVDQLQ
YLWURPRGHOV\VWHPWRVWXG\QHXURGHJHQHUDWLRQ'1$5HSDLU
9HUSHOOL&&DUOHVVL/%HFKL*)XVDU3ROL(2UHOODQD'+HLVH&)UDQFHVFKHWWL60DQWHJD]]D
5 0DQWHJD]]D 0 'HOLD ' 6DOD & &RPSDUDWLYH QHXURQDO GLIIHUHQWLDWLRQ RI VHOIUHQHZLQJ
QHXUDO SURJHQLWRU FHOO OLQHV REWDLQHG IURP KXPDQ LQGXFHG SOXULSRWHQW VWHP FHOOV )URQW &HOO
1HXURVFL
1RQUHODWHGWRWKHSURMHFW
)XVDU 3ROL ( =DOID & '·$YDQ]R ) 7RPDQLQ 5 &DUOHVVL / %RVVL 0 1RGDUL /5 %LQGD (
0DUPLUROL 3 6FDUSD 0 'HOLD ' 9HVFRYL $/ 'H )LOLSSLV / 0XULQH QHXUDO VWHP FHOOV PRGHO
+XQWHU GLVHDVH LQ YLWUR JOLDO FHOOPHGLDWHG QHXURGHJHQHUDWLRQ DV D SRVVLEOH PHFKDQLVP
LQYROYHG&HOO'HDWK'LVH
OPEN
Citation: Cell Death and Disease (2014) 5, e1342; doi:10.1038/cddis.2014.310
& 2014 Macmillan Publishers Limited All rights reserved 2041-4889/14
www.nature.com/cddis
Functional and molecular defects of hiPSC-derived
neurons from patients with ATM deficiency
L Carlessi1, E Fusar Poli1, G Bechi2, M Mantegazza2,3, B Pascucci4, L Narciso5, E Dogliotti6, C Sala7, C Verpelli7, D Lecis1 and
D Delia*,1
Loss of ataxia telangiectasia mutated (ATM) kinase, a key factor of the DNA damage response (DDR) pathway, causes the
cancer predisposing and neurodegenerative syndrome ataxia-telangiectasia (A-T). To investigate the mechanisms of
neurodegeneration, we have reprogrammed fibroblasts from ATM-null A-T patients and normal controls to pluripotency
(human-induced pluripotent stem cells), and derived from these neural precursor cells able to terminally differentiate into
post-mitotic neurons positive to 490% for b-tubulin III þ /microtubule-associated protein 2 þ . We show that A-T neurons
display similar voltage-gated potassium and sodium currents and discharges of action potentials as control neurons, but
defective expression of the maturation and synaptic markers SCG10, SYP and PSD95 (postsynaptic density protein 95).
A-T neurons exhibited defective repair of DNA double-strand breaks (DSBs) and repressed phosphorylation of ATM substrates
(e.g., cH2AX, Smc1-S966, Kap1-S824, Chk2-T68, p53-S15), but normal repair of single-strand breaks, and normal
short- and long-patch base excision repair activities. Moreover, A-T neurons were resistant to apoptosis induced by the
genotoxic agents camptothecin and trabectedin, but as sensitive as controls to the oxidative agents. Most notably, A-T neurons
exhibited abnormal accumulation of topoisomerase 1-DNA covalent complexes (Top1-ccs). These findings reveal that ATM
deficiency impairs neuronal maturation, suppresses the response and repair of DNA DSBs, and enhances Top1-cc accumulation.
Top1-cc could be a risk factor for neurodegeneration as they may interfere with transcription elongation and promote
transcriptional decline.
Cell Death and Disease (2014) 5, e1342; doi:10.1038/cddis.2014.310; published online 17 July 2014
Ataxia-telangiectasia (A-T) is a pleiotropic disorder with
predisposition to cancer and early onset neurodegeneration
as key features.1–3 The neuropathological abnormalities in
A-T include progressive loss of cerebellar Purkinje and
granule neurons, less pronounced degeneration of the bulbar
olivae in the brainstem and mild loss of myelinated fibers in
corticospinal and spinocerebellar tracts.4 A-T is caused by
germline mutations inactivating the ataxia telangiectasia
mutated (ATM) protein kinase, which has an apical role in
the DNA damage response (DDR) to double-strand breaks
(DSBs). Upon activation by DSBs, ATM phosphorylates a
plethora of substrates implicated in cell cycle arrest, DNA
processing and repair, chromatin remodeling, transcription
and apoptosis.1 Concordant with the role of ATM in the DDR,
the neurodegenerative phenotype in A-T has been attributed
to a defective DDR in pre- and post-mitotic neurons and
ensuing accumulation of genotoxic lesions. However, the fact
that cells from A-T patients and specimens from ATM knock-
out mice exhibit increased signatures of oxidative stress,5,6
together with the discovery that ATM senses and responds
to reactive oxygen species (ROS) through formation of
active, disulfide-cross-linked ATM dimers7 suggest that the
loss of redox balance may contribute to neurodegeneration
in A-T.
In neurons, a fraction of ATM is localized in the cytoplasm
where it phosphorylates the synaptic proteins VAMP2 and
synapsin-1.8 Recently, ATM has been detected in dendrites
and dendritic spines, and its activation by neuronal firing and
synaptic activity induces the phosphorylation of proteins
essential for neuronal function.9
Neurodegeneration in A-T has additionally been linked to
epigenetic modifications associated with deregulation of class
II histone deacetylase HDAC410 and hyperexpression of the
histone-lysine N-methyltransferase EZH2,11 altogether inducing transcriptional repression of multiple neuronal genes and
cell death.
1
Department of Experimental Oncology, Fondazione IRCCS Istituto Nazionale dei Tumori, Via Amadeo 42, 20133 Milano, Italy; 2Department of Neurophysiopathology,
Fondazione IRCCS Istituto Neurologico Carlo Besta, Via Amadeo 42, 20133 Milano, Italy; 3Institute of Molecular and Cellular Pharmacology (IPMC) CNRS UMR7275
and University of Nice-Sophia Antipolis, 660 Route des Lucioles, 06560 Valbonne, France; 4CNR Institute of Crystallography, Via Salaria, Km. 29.300, 00016
Monterotondo Scalo, Roma, Italy; 5Department of Food Safety and Veterinary Public Health, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy;
6
Department of Environment and Primary Prevention, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy and 7CNR Institute of Neuroscience and
Department of Biotechnology and Translational Medicine, Via Vanvitelli 32, 20129 Milano, Italy
*Corresponding author: D Delia, Department of Experimental Oncology, Fondazione IRCCS Istituto Nazionale dei Tumori, Via Amadeo 42, Milano 20133, Italy.
Tel: +39 02 23902641; Fax: +39 02 23902764; E-mail: [email protected]
Abbreviations: ATM, ataxia telangiectasia mutated; Chk2, checkpoint kinase 2; SSEA4, stage-specific embryonic antigen 4; a-SMA, a-smooth muscle actin; MAP2,
microtubule-associated protein 2; PARP, poly (ADP ribose) polymerase; GABA, g-aminobutyric acid; PSD95, discs large homolog 4; VGAT, vesicular GABA transporter;
KChIP, Kv channel-interacting protein 1; GFAP, glial fibrillary acidic protein; XRCC1, X-ray repair cross-complementing 1; CPT, camptothecin; DAPI, 40 ,6-diamidin-2fenilindolo; APE1, apurinic/apyridinic endonuclease 1; Pol-b, DNA polymerase b; bFGF, basic fibroblast growth factor; EGF, epidermal growth factor; NPCs, neural
precursor cells; hiPSCs, human-induced pluripotent stem cells; DDR, DNA damage response; DSBs, double-strand breaks; Top1-cc, topoisomerase 1-DNA covalent
complexes; IR, ionizing radiation
Received 24.3.14; revised 30.5.14; accepted 16.6.14; Edited by A Verkhratsky
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
2
It should be noted that germline mutations in ATM severely
impair glial cell functionality and vascular integrity, suggesting
that Purkinje cell death and cerebellar degeneration in A-T
may result from a dysfunctional neuro-astro-vascular unit.12
In summary, several factors are implicated in the neurodegenerative phenotype of A-T, but which of them has the most
crucial role is still unresolved, primarily because of the
unavailability of model systems able to recapitulate the
neurological disease. In this regard, ATM knockout mice do
not manifest the progressive ataxia seen in A-T nor the loss of
cerebellar Purkinje cells.13
The reprogramming of human somatic cells into induced
pluripotent stem cells (human-induced pluripotent stem cell
(hiPSCs)) by the introduction of pluripotency factors (Oct4/
Klf4/Sox2/cMyc) represents a prominent advance in stem cell
biology, owing to the capacity of these cells to differentiate to
virtually any cell type of the human body, and the possibility to
model patient- and allele-specific genetic diseases.14 In the
case of A-T, the establishment of hiPSCs and conversion to
functional neurons has been recently reported.15–17
In this study, we performed a previously undocumented
functional and molecular analysis of hiPSC-derived A-T
neuronal cells harboring patient-specific mutations, in order
to shed light on the causes of the neuropathology in A-T.
Results
Generation and characterization of hiPSCs derived from
A-T and Ctrl fibroblasts. The hiPSC lines were established
from primary fibroblasts of two unrelated A-T patients and a
healthy control (Ctrl), as detailed in Materials and methods
section. Colonies with embryonic stem cell-like morphology
were expanded on a mouse embryonic fibroblast (MEF)
feeder layer (Figure 1a, left). Western blot analysis revealed,
as expected, a positive signal for ATM in fibroblasts and
hiPSCs from Ctrl but not from A-T cases (Figure 1a, right).
No ATM protein was detectable in MEFs as the antibody
used was human specific. Furthermore, only Ctrl and A-T
hiPSC lines expressed the pluripotency marker Oct3/4, but
not fibroblasts or MEFs (Figure 1a, right). Immunofluorescence analysis attested the pluripotency of the hiPSCs, being
positive for Oct3/4, stage-specific embryonic antigen 4
(SSEA4) and Tra-1-81 (Figure 1b). Moreover, the hiPSCs
gave rise to the embryonic derivatives ectoderm, endoderm
and mesoderm, labeled positive for b-tubulin III, Sox17
and alpha-smooth muscle actin (a-SMA), respectively
(Figure 1c). Also, through embryoid body (EB) and neural
rosette formation and subsequent culture in a mitogen-free
medium, hiPSCs differentiated into b-tubulin III þ and
Figure 1 Generation, characterization, pluripotency validation and neuronal differentiation of hiPSC colonies from Ctrl and A-T patients. Representative images of newly
formed hiPSC colonies before picking (a, upper panel) and after picking (a, bottom panel). The hiPSC colonies were characterized by western blot (a, right) to evaluate the
expression of ATM protein and the pluripotency marker Oct 3/4 and were compared with primary fibroblasts and MEF feeders. Vinculin was used as a loading control. In b,
hiPSC colonies were labeled to visualize the expression of the pluripotency markers Oct 3/4, SSEA4 and TRA-1-81. Nuclei were counterstained with DAPI (blue). In c, Ctrl and
A-T hiPSCs were differentiated in vitro into the three germ layers. After 20 days of differentiation, cells were labeled with antibodies specific for a-SMA (mesoderm marker),
Sox17 (endoderm marker) and b-tubulin III (ectoderm marker). Nuclei were counterstained with DAPI (blue). The ability of hiPSCs to generate neuronal cells was confirmed
through the formation of floating EBs and rosette formation. Representative images for each differentiation step are shown in d
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
3
microtubule-associated protein 2 þ (MAP2 þ ) neurons
(Figure 1d and Supplementary Figure 1).
Generation of hNPCs and neurons from A-T and Ctrl
hiPSCs. Proliferating human neural precursor cells (hNPCs)
were generated as depicted in Figure 2a. Both Ctrl and A-T
hNPCs were negative for the pluripotency markers Tra-1-81
and Oct3/4 (Figure 2b, left) but positive for the neural
precursor markers Sox2, Nestin and Vimentin, and for the
proliferation marker Ki67 (Figure 2b, right). On western blots,
hNPCs showed lower expression of Tra-1-81 and Oct3/4
than hiPSCs, and higher levels of Sox2 and Nestin, which
then decreased during neuronal differentiation (Figure 2b,
bottom left).
The capacity of hNPCs to activate the DDR was investigated in response to ionizing radiation (IR), which induces
DSBs and ATM signalling.1 Using phospho-specific antibodies for the indicated substrates of ATM and checkpoint
kinase 2 (Chk2; KAP1-pS473), we found that after 15–30 min
of IR these molecules were strongly phosphorylated in Ctrl but
not in A-T hNPCs (Figure 2c). Interestingly, after IR an
accumulation of cleaved poly (ADP ribose) polymerase
Figure 2 Generation of hNPCs from Ctrl and A-T hiPSCs. To obtain stable and proliferating hNPCs, we followed the protocol depicted in a and a representative image of
the cell culture obtained is shown. In b, the characterization of the hNPCs by immunofluorescence demonstrates the loss of the pluripotency markers Tra-1-81 and Oct3/4, the
expression of neural markers (Sox2, Nestin and Vimentin) and the proliferation ability (Ki67). A comparative analysis between hiPSCs and proliferating (hNPCs) and
differentiated (D15) hNPCs was performed by western blot. In c, proliferating hNPCs were exposed to IR (5 Gy), and the time-dependent DDR activation was evaluated by
analyzing the phosphorylation of the indicated ATM substrates, and of cleaved PARP. b-Actin was used as a loading control. The real capacity of hNPCs to differentiate into
neurons is shown in d, where a high number of MAP2 þ , b-tubulin III þ and GABA þ cells are detected at D15 and D50 (e). hNPCs and neurons at 15–30 or 50 days of
differentiation were collected and analyzed by western blot for the expression of differentiation and maturation markers (f) or for the indicated proteins (g and h)
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
4
(PARP), a marker of apoptosis, was detected in Ctrl but not in
A-T hNPCs, suggesting that ATM deficiency confers radioresistance in proliferative cells (Figure 2c). When hNPCs
were induced to differentiate, they acquired a neuronal-like
morphology and at day 15 of differentiation (D15) were
positive for b-tubulin III or MAP2, with a fraction of cells also
expressing the marker of inhibitory neurons g-aminobutyric
acid (GABA; Figure 2d), which we were no longer able to
detect at D50 in mature neuronal cells18 expressing b-tubulin
III or MAP2 (Figure 2e). Western blot analysis confirmed the
downregulation of the stem cell marker Nestin, the upregulation of the neuronal marker b-tubulin III at D30 (Figure 2f, left)
and at D15 and D50 (Figure 2f, right), and the expression of
the postsynaptic marker PSD95 postsynaptic density
protein 95, the inhibitory synapse marker vesicular GABA
transporter (VGAT), the neuronal growth-associated protein
SCG10 and the potassium channel-interacting proteins
(KChIPs). At D30 and D50, but not D15, cells also expressed
the pre-synaptic marker Synaptophysin (SYP). We detected
no obvious differences between Ctrl and A-T hNPCs
concerning the generation of neuronal cells, but A-T neurons
displayed deficits in the expression of SYP and PSD95 as well
as of SCG10 and KChIP, and higher amounts of glial fibrillary
acidic protein (GFAP; Figure 2f).
The post-mitotic status of the differentiated cells was
confirmed by the downregulation in D15–D50 (Figure 2h)
and D30 samples (Figure 2g) of Cdc2 and cyclin B1.
Concordant with our previous results,19 cleaved poly (ADP
ribose) polymerase (PARP) was higher in Ctrl than in A-T
neuronal cells. To note, gH2AX (pS139) was also highly
expressed at D50, and its levels were lower in A-T than in Ctrl
cells (Figure 2g).
Electrophysiological characterization. Ctrl and A-T neurons at D53±6 were analyzed by whole-cell patch clamp
recordings in order to verify their functional maturation.
Voltage clamp experiments showed similar voltage-gated
potassium currents (Figure 3a) and, most importantly,
voltage-gated sodium currents, selectively blocked by
tetrodotoxin (TTX), which is a specific feature of mature
neurons (Figure 3a, left panel). Transient peak sodium
inward currents had similar amplitude in Ctrl and A-T
neurons: maximal peak current density was 84.9±19.6 pA/pF,
n ¼ 10 in Ctrl neurons, 82.7±13.5 pA/pF, n ¼ 13 in A-T
neurons (Figure 3b). Moreover, current clamp experiments
showed that both Ctrl and A-T neurons could generate
discharges of action potentials in response to injection of
depolarizing current steps (Figure 3c). Finally, to determine
whether neurons developed functional synapses, we performed voltage clamp experiments for recording spontaneous excitatory and inhibitory postsynaptic currents
(sEPSCs and sIPSCs). Both Ctrl and A-T neurons exhibited
sEPSCs (Figure 3d) with similar frequencies (0.02±0.01 Hz,
n ¼ 4 cells and 0.03±0.02 Hz, n ¼ 4 cells, respectively),
which were blocked by application of kynurenic acid (3 mM;
data not shown), but no sIPSCs (Figure 3e).
DNA damage-induced G1/S checkpoint in relation to
development stage. In somatic cells, ATM mediates the
G1/S checkpoint but is inactive in ESCs and iPSCs.20,21
Cell Death and Disease
To further delineate this activity in relation to the developmental stage, we analyzed the cell cycle profile of primary
fibroblast, hiPSCs, hNPCs and terminally differentiated
neurons by flow cytometry before and 24 h after IR treatment
(Supplementary Figure 2). Primary Ctrl fibroblasts displayed a
normal G1/S checkpoint arrest, in contrast to A-T fibroblasts in
which this checkpoint defect resulted in a significant increase
in the percentage of G2 phase cells (Figure 4a). Notably, not
only A-T hiPSCs, but also Ctrl hiPSCs lacked the G1/S
checkpoint although, interestingly, Ctrl hNPCs regained it,
whereas A-T hNPCs maintained this defect. The analysis of
neurons at D30 revealed that all cells were in the G0/G1
phase and this condition was not perturbed 24 h after IR.
To better understand the implication of DDR proteins in the
regulation of cell cycle arrest in fibroblasts, hiPSCs and
hNPCs, Ctrl cells were harvested at the indicated time points
after treatment with 5 Gy IR (Figure 4b) and the expression
of ATM-pS1981, p53-pS15, CDC25A and p21waf1 was
analyzed. Interestingly, although all cell types proficiently
activated ATM and induced the accumulation of p53, hiPSCs
were unable to induce p21waf1, and did not express CDC25A.
This could explain the incapacity of hiPSCs to regulate the cell
cycle after DNA damage. We also investigated other DDR
proteins such as RAD51, the MRN complex (RAD50-MRE11NBS1) and X-ray repair cross-complementing 1 (XRCC1),
and found changes in expression during the developmental
stages, suggesting a different ability to repair DNA damage in
the different cell types (Supplementary Figure 3).
Base excision repair capacity in undifferentiated and
differentiated hNPCs. In neurons, the base excision repair
(BER) pathway is essential to remove oxidative DNA
damage and single-strand breaks (SSBs) generated by high
levels of ROS, and defects in BER contribute to neurodegeneration.22–25 To determine how BER is regulated in preand post-mitotic neurons, we analyzed the short-patch (SP)
and long-patch (LP) BER subpathways.23 Analysis of SPBER revealed no significant differences between Ctrl and
A-T both in the proliferation and post-mitotic (D15) stage
(Figure 5a). To better understand this result, we analyzed the
levels of the SP-BER proteins apurinic/apyridinic endonuclease 1 (APE1), DNA polymerase b (Pol-b) and XRCC1,
and after differentiation (D15) only the levels of XRCC1
appear reduced (Figure 5c). Similar findings were seen in
D30 neurons (data not shown).
LP-BER activity appeared markedly lower in both Ctrl and
A-T post-mitotic neurons than in their respective proliferating
hNPCs (Figure 5b). This finding is concordant with the
reduced expression of Flap endonuclease 1, a protein
involved in LP-BER in non-replicating cells (Figure 5c).
Overall, these results point out differences that depend on
different levels of expression or differential activation of BER
enzymes in undifferentiated and differentiated cells, but
appear independent of ATM.
DNA damage response and repair in post-mitotic
neurons. IR-time course analysis of Ctrl neurons at D30
showed a vigorous ATM-dependent phosphorylation of
SMC1-S966, KAP1-S824, Chk2-T68, p53-S15 and
g-H2AX, while this response was repressed in A-T neurons
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
5
Figure 3 Functional characterization of Ctrl and A-T neurons after 50 days of differentiation. In a, left, patch clamp whole-cell recordings of representative total ionic
currents elicited with depolarizing voltage steps between –70 and þ 10 mV (10 mV increments from a holding potential of –70 mV) in Ctrl and A-T hNP-derived neurons; scale
bar: 500 pA, 10 ms. In a, right, representative currents elicited with a depolarizing voltage step to ÿ 10 mV in control (black), during perfusion with TTX 3 mM (red) and after
washout (blue) in Ctrl (upper panels) and A-T (lower panels) hNP-derived neurons; scale bar: 500 pA, 10 ms. In b, bar graph of maximum sodium current densities (CD) in Ctrl
and A-T hNPC-derived neurons (no statistical significant difference: unpaired T-test). In c, representative action potential discharges recorded in current clamp during
injections of 1-s long depolarizing current steps from a holding potential of ÿ 70 mV; scale bar: 10 mV, 100 ms. In d, traces showing sEPSCs, recorded at the holding potential
of ÿ 70 mV, while in e traces showing the absence of sIPSCs, recorded at the holding potential of þ 30 mV; scale bar: 10 mV, 1 s
(Figure 6a). Similarly, the phosphorylation of the Chk2
substrate KAP1-S473 was abrogated in A-T neurons.
Interestingly, as previously described for proliferating
hNPCs (Figure 2c), 24 h after IR an accumulation of
cleaved PARP was detected in Ctrl but not in A-T neurons,
indicating that ATM deficiency confers short-term radioresistance also in terminally differentiated cells (Figure 6a,
upper panel). Altogether, these results indicate that normal
post-mitotic neurons activate the DDR as efficiently as their
proliferating precursors, and that ATM deficiency ablates
this response.
The ability of post-mitotic neurons to repair SSBs and DSBs
was investigated with the alkaline and neutral comet assay,
respectively. Although the repair of SSBs, induced by
hydrogen peroxide (H2O2),26 showed no major differences
between Ctrl and A-T post-mitotic neurons (Figure 6b), the
repair of DSBs, induced by IR,26 appeared defective in A-T
neurons, which displayed 30% more unrepaired lesions than
Ctrl cells (Figure 6c). The latter finding is consistent with a
defective DDR, as shown in Figure 6a.
To investigate the role of ATM in the resolution of IR-induced
DSBs in D30 neurons, we scored the time-dependent
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
6
Figure 4 Cell cycle phase distribution changes after IR treatment. Ctrl and A-T cells at various developmental stages (primary fibroblasts, hiPSCs, hNPCs, D30 postmitotic neurons) were treated with 5 Gy IR, collected after 24 h, and analyzed for DNA content by flow cytometry (a). In b, Ctrl fibroblasts, hiPSCs and hNPCs were treated with
5 Gy IR, collected at the indicated times and analyzed by western blot. b-Actin was used as a loading control
formation and clearance of g-H2AX and 53BP1 nuclear foci by
immunofluorescence, as described.27,28 Unirradiated Ctrl
D30 neurons showed a basal number of g-H2AX and 53BP1
foci per cell (1.4±1.2 and 0.9±1), which increased to
13.3±3.3 and 10.1±2.4, respectively, after IR, to decline
after 6 h and even further after 24 h (Figures 6d and e).
Untreated A-T cells showed a number of foci per cell
comparable to Ctrl cells, which increased more modestly,
5.6±2.6 for g-H2AX and 5.6±2.1 for 53BP1 after 15 min, and
persisted after 24 h (Figures 6d and e graphs). These findings
Cell Death and Disease
indicate that A-T post-mitotic neurons show slower kinetics of
DSBs repair than Ctrl, concordant with previous findings in
other ATM-deficient cells.19
The apoptotic effect of radiation on Ctrl and A-T neurons
24 h after 5 Gy IR was assessed by flow cytometry analysis of
the subdiploid DNA peak (Supplementary Figure 4). Fewer
apoptotic cells were detected in IR-treated A-T cells (22.2%)
than in Ctrl cells (32.3%), suggesting that ATM deficiency
attenuates short-term apoptosis, concordant with the findings
on cleaved PARP (Figure 6a) and our previous findings.19,29
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
7
Figure 5 BER activities in proliferating and post-mitotic Ctrl and A-T neuronal cells. In vitro repair reactions were performed by using whole-cell extracts and as substrate a
32P-labeled circular plasmid containing a single AP site (pGEM-AP) to measure SP-BER (a) or a THF residue (pGEM-THF) for LP-BER (b). The correct insertion of a single
lesion in the plasmid molecules was checked by digestion with uracil DNA glycosylase (UDG) followed by endonuclease III (NTH) for pGEM-AP (a), and by AP endonuclease
(HAP1) for pGEM-THF (b). Repair kinetics of the AP site (a) or THF residue (b) by proliferating hNPCs and post-mitotic neurons (D15) were measured following incubation of
the single lesion-containing plasmids with extracts from Ctrl or A-T defective neuronal cells for increasing periods of time. Repair products were analyzed by agarose gel
electrophoresis and the radioactivity of the bands corresponding to nicked (form II) and supercoiled (form I) plasmids was quantified. Repair efficiency is expressed as relative
amount of Form I over total radioactivity in each lane. Blue bar, Ctrl cell extracts; red bar, A-T cell extracts. The expression level of SP-BER and LP-BER proteins was
evaluated by western blot in proliferating and post-mitotic neuronal cells (c). b-Actin was used as a loading control
Treatment of A-T and Ctrl neuronal cells with genotoxic
and oxidative agents. As genotoxic agents and oxidative
stress may similarly affect ATM-deficient neurons, we
investigated the viability of Ctrl and A-T post-mitotic neurons
(D30) upon treatment with compounds that work through
different mechanisms: camptothecin (CPT), which inhibits
Topoisomerase I (Top1) and traps Top1-DNA covalent
complexes (Top1-ccs);30 trabectedin, which blocks transcription factor activity;31 Paraquat, which induces ROS through
depolarization of the inner mitochondrial membrane,32 and
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
8
Figure 6 A-T post-mitotic neurons are defective in DDR. In a, D30 post-mitotic neurons (D30) were tested by western blot with antibodies specific for the various ATM
phosphosubstrates and cleaved PARP at various times after treatment with 5 Gy IR. b-Actin was used as a loading control. DNA SSBs (b) and DSBs (c) were analyzed in
neurons (D30) by alkaline and neutral comet assay, respectively, at two different times after treatment. 20 min after H2O2 or IR treatment was considered the time point with the
maximum DNA damage. In b and c, representative photos of comets from untreated cells and at different time points following treatment. The ratio between treated and
untreated tail moments of at least 50–70 cells per experimental point is shown in the graphs (values in %). One representative experiment out of three is shown. In d and e,
formation and resolution of IR-induced nuclear foci. Neurons (D30) were irradiated with 0.5 Gy, collected at the indicated times and labeled for g-H2AX (d) and 53BP1 (e). For
each treatment, the number of foci was scored from 100 cell nuclei per duplicate preparations and from three independent experiments (mean±S.D.) (d and e, bottom) For
each time point, the difference between Ctrl and A-T was statistically significant (#Po0.01) (analysis performed by the Student’s t-test)
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
9
H2O2. Ctrl and A-T post-mitotic neurons were equally
sensitive to H2O2 and Paraquat (Figures 7a and b), a finding
concordant with previous data33 and with the results in
Figure 6b showing that these cells have the same ability to
resolve SSBs. By contrast, A-T neurons were significantly
more resistant than Ctrl neurons to CPT and trabectedin by
about 40% and 30%, respectively (Figures 7c and d).
Accumulation of Top1-ccs in A-T post-mitotic neurons.
As in murine neural cells ATM deficiency results in
accumulation of Top1-ccs and failure to recover global
transcription after Top1-cc trapping,34 we used a modified
alkaline comet assay (MACA),34 which indirectly quantifies
Top1-ccs, to analyze hNPC-derived neurons (D30) untreated
or treated for 45 min with 30 mM CPT (Figure 8a, left).
As shown in the graph, the accumulation of Top1-ccs was
much greater in A-T than in Ctrl neurons, both before and
after CPT treatment. Of note, the overall levels of Top1 in
proliferating and post-mitotic neurons (D30) were similar
(Figure 8a, right).
To further confirm these findings, we applied the rapid
approach to DNA adduct recovery (RADAR) assay that allows
to directly detect Top1-ccs bound to genomic DNA.35 D30
neurons, incubated for 1 h with 30 mM CPT, were harvested
immediately or 3 h after washout of the drug to analyze the
recovery of the damage. DNA was isolated, dot blotted and
Top1-ccs were revealed with an anti-Top1 antibody and ECL.
The levels of Top1-ccs (Figure 8b, left) were then normalized
with the total amount of DNA, revealed by 40 ,6-diamidin2-fenilindolo (DAPI; Figure 8b, middle). b-Actin was used to
verify that the DNA samples were free of non-covalently
bound contaminating proteins (Figure 8b, right). The graph
shows Top1 protein levels after normalization with DNA, and,
as expected, untreated A-T neurons displayed higher
accumulation of Top1-ccs than Ctrl cells, and also appeared
defective in Top1-cc recovery after CPT removal.
Discussion
The mechanisms underlying neurodegeneration in A-T still
remain elusive. Up to the present, ATM knockout and knockin
mice36,37 and in vitro ATM-deficient human neural stem cell
(hNSC) models19,38 have been useful for elucidating many
aspects of the neuropathology, but animal models do not
recapitulate the CNS disease,39 while hNSCs have been
argued to be very heterogeneous.40 To obtain a more reliable
in vitro model of neurodegeneration in A-T, two novel
approaches have recently been described: the establishment
of patient olfactory mucosa-derived neurospheres, which give
rise to neurons,41 and the reprogramming of patient fibroblasts to a pluripotent stage.15,17
As a functional analysis of the neuronal cells harboring
patient-specific mutations in ATM has not yet been reported,
in this study we examined hiPSC-derived A-T hNPCs and
terminally differentiated neurons in culture. We found that A-T
hNPCs displayed a strongly attenuated response to DSBs
with respect to Ctrl hNPCs, which is concordant with our
previous study on hNSCs.19
Moreover, terminally differentiated A-T neurons exhibited
decreased expression of SYP and PSD95, which is
Figure 7 A-T post-mitotic cells are resistant to CPT and trabectedin treatment. Neurons at D30 grown in 96-well plates were assessed for cell viability by CellTiter-Glo 72 h
after exposure to H2O2 for 20 min (a), or continuously with Paraquat (b), CPT (c) and trabectedin (d) at the indicated doses. All treatments were performed in triplicate wells.
The graphs show mean±S.D. and, where indicated, the difference between Ctrl and A-T was statistically significant (*Po0.05; #Po0.01) (analysis performed by the
Student’s t-test)
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
10
Figure 8 A-T post-mitotic neurons accumulate abnormal levels of Top1-cc. Total intracellular levels of Top1 protein were analyzed by western blot in proliferating hNPCs
and post-mitotic neurons (a left). The levels of endogenous Top1-ccs were quantified in Ctrl and A-T post-mitotic neurons (D30) subjected to 30 mM CPT for 40 min by MACA
from 50 cells per sample per experiment (a right). Data are the average of three independent experiments (mean±S.D.). For each time point, the difference between Ctrl and
A-T was statistically significant (#Po0.01) except where indicated. In b, Ctrl or A-T post-mitotic neuronal cells were untreated or treated with 30 mM CPT for 60 min or
incubated and harvested 3 h after washing. The DNA was purified from cells after lyses with a denaturing buffer (MB) as reported in Materials and methods section. The
indicated amount of DNA was vacuum blotted on a nitrocellulose membrane, which was then tested with anti-Top1 antibody and ECL (left) and with DAPI (middle). b-Actin
(left) represents the control for non-covalently bound contaminant proteins. The densitometry analysis of Top1-cc signals after normalization for blotted DNA (DAPI) from three
biological replicates (mean±S.D.) is reported in the graph. Y axis represents the relative signal intensity. Where indicated the difference between Ctrl and A-T was statistically
significant (#Po0.01; analysis performed by the Student’s t-test)
concordant with previous works showing pre- and postsynaptic degeneration in ATM knockout mice,42 the requirement of cytoplasmic ATM for phosphorylation of the synaptic
vesicle proteins VAMP2 and Synapsin-1, and that ATM
deficiency affects spontaneous vesicle release and establishment of long-term potentiation.8 We also showed that
A-T neurons are defective in the expression of SCG10
and KChip, altogether underpinning defects in neuronal
maturation.
Cell Death and Disease
In agreement with the role of ATM in establishing the G1/S
checkpoint arrest to prevent cells with damaged DNA from
entering the S-phase,1 primary Ctrl fibroblasts effectively
arrested at the G1/S boundary after IR, but once reprogrammed into hiPSCs lost this G1/S checkpoint arrest to
regain it at the stage of hNPCs. Instead, A-T cells showed a
disrupted G1/S checkpoint, whatever the developmental
stage. Notably, the absence of the G1/S checkpoint arrest in
Ctrl hiPSCs is a characteristic of ESCs, where the cells
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
11
subjected to DNA damage in G1 enter S-phase to be eliminated
by apoptosis, thus avoiding the propagation of mutations
detrimental for the whole organism.43,44 Accordingly, hiPSCs
showed a great accumulation of p53-pS15 and faint induction
of p21waf1, compatible with a p53-dependent apoptotic
response rather than with a G1/S checkpoint activation.
Moreover, we found that Ctrl hiPSCs upregulate homologous
recombination components such as Rad51, concordant with
the fact that ESCs predominantly use this high fidelity pathway
to avoid the accumulation of mutations, in contrast to somatic
cells, which rely on the error-prone non-homologous end
joining to repair DNA damage occurring in G1.45
BER is the major pathway for repair of oxidatively damaged
DNA and various SSB intermediates,22 and previous findings
demonstrate that ATM and Chk2 facilitate the recruitment of
downstream BER proteins to the initial damage recognition/
excision step to promote BER.46 Here, we found no significant
differences in SP-BER activity between Ctrl and A-T
proliferative hNPCs and post-mitotic neurons, while conversely, LP-BER activity was reduced in post-mitotic neurons. It
should be noted that both SP-BER and LP-BER enzymatic
efficiencies have been found to decrease, along with the
respective protein components, in a neuroblastoma cell line
upon differentiation,24 and while our data on LP-BER concord
with these results, those on SP-BER are clearly at variance.
This could be because the expression of the SP-BER
component XRCC1 is totally ablated in differentiated neuroblastoma cells,24 while being only partially downregulated in
our neurons. Regardless of this discrepancy, the variations in
LP-BER activity between post-mitotic neurons and proliferative hNPCs were totally independent of ATM expression.
In accordance with our previous findings using hNSCs,19
here we found that the DDR was vigorously activated in postmitotic Ctrl neurons, according to the strong phosphorylation
of ATM substrates, whereas in A-T post-mitotic neurons this
response was almost ablated. We also highlighted relevant
DSB repair defects in A-T neurons, which indicated that ATM
deficiency delays the repair of DSBs, in accordance with
previous reports19,47 and with the observation that ATM is
required for the repair of heterochromatic DSBs.48 Furthermore, we demonstrated that A-T neurons show less apoptosis-related sub-diploid DNA content and a lower cleaved
PARP induction after IR-treatment than Ctrl neurons. Indeed,
ATM deficiency has been found to attenuate the apoptotic
response to IR in A-T lymphoblastoid cells and in hNSCs19,29
and ATM is required for the p53-mediated apoptosis of
developing post-mitotic neurons exposed to IR.49,50
Although ATM appears involved in sensing and modulating
the cellular response to ROS,7 in our study A-T post-mitotic
neurons were as sensitive to oxidative agents as Ctrl postmitotic neurons, in agreement with the alkaline comet assays
showing that these neurons equally repair SSBs, and in the
case of H2O2, with results obtained with A-T lymphoblastoid
cells.33 By contrast, A-T post-mitotic neurons were significantly
more resistant than Ctrl neurons to the genotoxic agents CPT
and Trabectedin, probably because, in the absence of ATM,
drug-induced DSBs in transcribed genes fail to activate a p53dependent apoptotic response. This finding is concordant with
results showing that ATM inhibition suppresses the genotoxic
response in rat cerebellar granule neurons.51
Finally, as it has been recently shown that post-mitotic
neural cells from ATM-deficient mice accumulate Top1-ccs
and fail to recover global transcription after Top1-cc trapping
by CPT,34 and given that Top1-ccs can promote transcription
elongation arrest and decay,52 a phenomenon underlying
neurodegeneration,53–55 we analyzed Top1-ccs by two
different assays and found greater levels of these intermediates in A-T than in Ctrl neurons. This finding, which to our
knowledge has never been reported in human A-T neurons,
warrants further studies to determine the impact of Top1-cc on
genome-wide global transcription.
In conclusion, we have shown that A-T post-mitotic neurons
display normal electrophysiological activity, defective expression of maturation markers, attenuated response to and repair
of DSBs, but normal capacity to repair SSBs and normal BER
activities. Strikingly, A-T neurons exhibit elevated levels of
Top1-ccs, which may potentially impair transcriptional fidelity,
a possibility that requires further investigation. Finally, we
have shown that hiPSCs, like ESCs, provide a unique in vitro
model to study the G1/S cell cycle checkpoint in a developmental context.
Materials and Methods
Primary cell culture. A-T patient-derived primary dermal fibroblasts
(GM03487 and GM02530) were purchased from the Coriell Institute for Medical
Research (Camden, NJ, USA). Both carry compound heterozygous mutations in
ATM gene: GM03487 (c.8266A4T and c.1141_1142ins4), GM02530
(c.5675_5762del in trans with c.2251-1G4A and c.6573_6653del). Dermal
fibroblasts of a healthy, ethnically matched donor and their iPSCs derivatives were
previously described by us.18 Fibroblasts were cultured at 37 1C in a 5% CO2 in
E-MEM supplemented with 15% heat-inactivated FCS, 100 units/ml penicillin,
100 mg/ml streptomycin, 100 mM non-essential amino acids and 2 mM glutamine.
Primary MEFs (PMEF-CFL, Millipore, Bedford, MA, USA) were cultured in
DMEM/F12 supplemented with 20% heat-inactivated FCS, 100 units/ml penicillin,
100 mg/ml streptomycin, 100 mM non-essential amino acids and 2 mM glutamine.
PMEF-CFLs were mitotically inactivated (Mito-MEFs) with 1 mg/ml Mytomycin C
(Kiowa Hakko Kirin, Milan, Italy) in DMEM high glucose (Euroclone, Pero, Italy) for
3 h and were then collected and plated into 0.1% gelatin-coated six-well plates at a
concentration of 1.8 105 cells/cm2.
hiPSC generation and culture. Human primary fibroblasts were infected
with the STEMCCA Cre-Excisable Constitutive Polycistronic Lentivirus (Millipore)
following the manufacturer’s instructions. After 20–25 days, hiPSC clones were
picked and transferred onto a Mito-MEF feeder in human embryonic stem cell
(hESC) medium composed of DMEM/F12 supplemented with 20% KnockOut
serum replacement (Life Technologies, Monza, Italy), 100 units/ml penicillin,
100 mg/ml streptomycin, 100 mM non-essential amino acids, 2 mM glutamine,
1 mM sodium pyruvate, 100 mM 2-mercaptoethanol and 20 ng/ml basic fibroblast
growth factor (bFGF; Peprotech, Rocky Hill, NJ, USA). For passaging, hiPSCs
were incubated with DMEM/F12 containing 1 mg/ml collagenase IV (Sigma, Milan,
Italy) at 37 1C for 10 min. Cells were then scraped, collected and transferred at a
split ratio of 1 : 3 to a new plate on Mito-MEFs in hESC medium. For feeder-free
culture, hiPSCs were seeded on Matrigel (hESC-qualified matrix, BD Biosciences,
San Jose, CA, USA)-coated plates in Nutristem culture medium (Stemgent,
Cambridge, MA, USA) supplemented with 100 units/ml penicillin and 100 mg/ml
streptomycin. For differentiation into derivatives of all three primary germ layers,
hiPSCs were collected as for passaging and transferred to a gelatin-coated plate
in DMEM/F12 supplemented with 20% heat-inactivated FCS, 100 units/ml
penicillin, 100 mg/ml streptomycin, 100 mM non-essential amino acids and
2 mmol/l glutamine.
Generation of hNPCs and terminal differentiation. hiPSC colonies
were harvested as for passaging, resuspended in hESC medium without bFGF
and plated into a 6 cm low binding dish (HydroCell, Nunc, Cornaredo, Italy) for
floating cultivation. After 5 days in suspension culture, EBs were collected and
plated into a Matrigel (growth factor reduced, BD Biosciences)-coated dish for
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
12
additional 7 days in hESC media supplemented with 1X N2 (Gibco, Monza, Italy).
After 7 days, rosettes were manually picked, mechanically dissociated into single
cells and resuspended in Neural Precursor Medium (NPMedium) composed of
DMEM/F12 supplemented with 2 mM glutamax, 100 units/ml penicillin, 100 mg/ml
streptomycin, 1 : 500 B27 (Gibco), 1X N2, 20 ng/ml epidermal growth factor (EGF)
and 20 ng/ml bFGF (Società Italiana Chimici, Roma, Italy) and plated into Matrigelcoated flasks. NPMedium was changed every 2 days. For passaging, 90%
confluent cells were detached using Accutase (PAA; Piscataway, NJ, USA) and
split at a ratio of 1 : 4. hNPCs displayed a homogenous population starting at
passage 3. To obtain terminally differentiated neurons, proliferating hNPCs were
plated in NPMedium at a concentration of 5 103 cells/cm2; 24 h later medium
was changed to NPMedium without bFGF and EGF and replaced every 2–3 days
thereafter.
0.2 mM EGTA, 10 mM HEPES, 20 mM P-creatine, 2 mM ATP-Na2, 0.2 mM GTPNa2 and 0.1 mM Leupeptine, pH 7.2 with KOH. Cell capacitance and series
resistance errors were carefully compensated (B85%) throughout the experiment.
The remaining linear capacity and leakage currents were eliminated online using a
P/4 subtraction paradigm. Pipette resistance was between 2.6 and 3.0 MO. For
the recordings of total voltage-gated ion currents, signals were filtered at 10 kHz
and sampled at 100 kHz. For the recordings of postsynaptic currents, signals were
filtered at 3 kHz and sampled at 10 kHz. When we switched the amplifier to current
clamp mode, we applied the bridge balance compensation and held the resting
potential at ÿ 70 mV by injecting the appropriate holding current. Neuronal firing
was recorded injecting depolarizing current pulses of increasing amplitude. The
neurons with unstable resting potential and/or unstable firing were discarded. In
current clamp mode signals were filtered at 10 kHz and sampled at 20 kHz.
Western blotting. Western blot analysis was performed as previously
described.56 Briefly, cells were lysed in Laemmli buffer (0.125 M Tris-HCl pH 6.8,
5% SDS) and lysates were sonicated, size-fractionated by SDS-PAGE and
electroblotted onto PVDF membranes (Millipore), which were then incubated with
primary antibodies and binding detected by ECL (Pierce, Rockford, IL, USA) on
autoradiographic films. Bands were acquired with a digital scanner. The primary
mouse monoclonal antibodies used were against the following molecules: OCT3/
4(clone C-10), p53, Cdc2 (p34), p21waf1 (Santa Cruz Biotechnology, Inc., Dallas,
TX, USA), b-tubulin III (clone 2G10), SYP, Vinculin, b-actin (Sigma), Tra-1-81
(eBioscience, San Diego, CA, USA), anti-Sox2, CDC25A (Abcam, Cambridge,
UK), cyclin B1 (clone GNS-1, BD Pharmigen, Franklin Lakes, NJ, USA), gH2AX
(H2AX-pSer139, clone JBW301, Millipore), PSD95, SCG10, Pan-KChIP, SYP
(NeuroMab, Davis, CA, USA), Nestin, MAP2 (clone AP20, Chemicon International,
Billerica, MA, USA), KAP1 (TIF1b, clone 4E1, Cell Signaling Technology,
Danvers, MA, USA), Chk2 (clone DCS270-273, MBL Intl Corp., Des Plaines, IL,
USA) and Top1 (BD Biosciences); rabbit antibodies specific for ATM (clone Y170,
Epitomics, Burlingame, CA, USA), ATM pS1981 (Rockland Inc., Gilbertsville, PA,
USA), cleaved PARP, XRCC1, KAP1-pS824, Chk2-pThr68, p53-pSer15 (Cell
Signaling Technology), GFAP (Millipore), VGAT (Synaptic System, Goettingen,
Germany), Polb, Fen1 (Abcam), APE1 (Santa Cruz Biotechnology, Inc.),
SMC1, SMC1-pSer966 (Bethyl Laboratories, Inc., Montgomery, TX, USA) and
KAP1-pS473 (Biolabs, San Diego, CA, USA).
Cell cycle phase analysis. Cells at different developmental stages were
irradiated with 5 Gy IR, incubated for 24 h and then fixed with 70% ethanol,
washed, treated PBS-RNAse A, at 37 1C for 30 min and finally stained with
propidium iodide. Cells were analyzed with a FACSCalibur flow cytofluorimeter
instrument fitted with a Cell Quest software package (Becton Dickinson,
Sunnyvale, CA, USA).
Immunofluorescence. Cells grown on coverslips were fixed in 4% buffered
paraformaldehyde and permeabilized with 0.1% Triton X-100 in PBS for 10 min.19
After blocking with PBS plus 10% normal goat serum for 40 min, cells were
incubated overnight with primary antibodies at 4 1C. The following primary
antibodies were used: monoclonal anti-Oct 3/4 (Clone C-10, Santa Cruz
Biotechnology, Inc.), anti-SSEA4 (eBioscience), anti-Tra-1-81 (eBioscience),
anti-a-SMA (clone 1A4, Sigma), anti-Sox17 (clone 245013, R&D System,
Minneapolis, MN, USA), anti-b-tubulin III (clone TUJ1, Covance, Princeton, NJ,
USA), anti-Nestin, anti-MAP2 (clone AP20, Chemicon International), anti-Vimentin
(DAKO, Cernusco sul Naviglio, Italy), and polyclonal anti-GFAP (Millipore), antiGABA, and anti-Ki67 (Thermo Scientific, Milan, Italy). After three washes with
PBS, cells were incubated with Alexa fluor-labeled goat anti-mouse or anti-rabbit
antibodies (Life Technologies) for 45 min. Cells were washed, counterstained with
DAPI, mounted on glass-slides with Prolong Gold Antifade (Life Technologies) and
analyzed using an Eclipse E1000 Nikon fluorescence microscope equipped with a
DMX1200F CCD camera (Torino, Italy). For the assessment of nuclear foci, cells
on coverslips were washed with 0.9% NaCl, air dried, fixed in 3% buffered
paraformaldehyde and permeabilized for 5 min at 4 1C in 0.5% Triton. Cells were
then blocked in PBS containing 5% BSA and 0.2% Tween20, labeled with
antibodies specific for H2AX-pS139 antibody (clone JBW301, Upstate Biotechnology,
New York, NY, USA) and 53BP1 (NB100-304; Novus Biologicals, Cambridge, UK)
and foci from three independent experiments and duplicate slides were
enumerated.
Electrophysiology analysis. Electrophysiological recordings were done as
reported18,57 at room temperature (22–25 1C) using a Multiclamp 700 A patch
clamp amplifier and pClamp 10.2 software (Molecular Devices, Sunnyvale, CA,
USA). Recordings usually started 5 min after the rupture of the membrane patch,
to allow intracellular dialysis with the pipette solution. External bath solution
consisted of 129 mM NaCl, 1.25 mM NaH2PO4, 35 mM glucose, 1.8 mM MgSO2,
1.6 mM CaCl2, 3 mM KCl and 10 mM HEPES, pH 7.4 with NaOH. The internal
pipette solution consisted of 120 mM K gluconate, 15 mM KCl, 2 mM MgCl2,
Cell Death and Disease
In vitro BER assay. Whole-cell extracts for BER assays were prepared from
15 106 cells that were harvested, resuspended in 400 ml buffer I (10 mM Tris pH
7.8, 200 mM KCl) and then lysed by adding an equal volume of Buffer II (10 mM
Tris pH 7.8, 200 mM KCl, 2 mM EDTA, 40% glycerol, 0.2% Nonidet-P40, 2 mM
DTT and protease inhibitors). After stirring for 1 h at 4 1C and centrifugation, the
supernatant was aliquoted and stored at ÿ 80 1C. Closed circular DNA molecules
containing a single lesion were constructed as described58 by priming singlestranded pGem-3Zf( þ ) DNA with the oligonucleotides containing the modified
base of interest (uracil and tetrahydrofuran (THF). These oligonucleotides were [g32P]ATP 50 end-labeled. The in vitro DNA synthesis was performed by using DNA
T4 DNA polymerase holoenzyme, single-stranded DNA-binding protein, dNTPs
and T4 DNA ligase. The plasmid DNA containing a single uracil residue was
digested with UDG to produce a single abasic site. Repair of the plasmid DNA
containing the lesions (pGEM-AP/THF) was conducted as described.58 Briefly,
reaction mixtures (50 ml) contained 40 mM HEPES/KOH (pH 7.9), 75 mM KCl,
5 mM MgCl2, 0.5 mM dithiothreitol, 50 mM of each dNTP, 2 mM ATP, 40 mM
phosphocreatine, 2.5 mg of creatine phosphokinase (type I), 3.4% glycerol, 18 mg
of bovine serum albumin and 50 mg of cell extracts. After incubation at 30 1C, the
plasmid DNA was recovered and loaded onto agarose gels. The relative yield of
the different plasmid forms was measured. All experiments were repeated at least
three times, and representative experiments are shown.
Comet assay and MACA. To prepare the Comet slides, the day before
analysis microscope glass slides were washed with methanol, air dried and
immersed in molten Normal Melting-Agarose (1%) to obtain the first agarose layer,
before being stored overnight at 4 1C. To evaluate DNA strand breaks, post-mitotic
neurons were incubated with 20 mM H2O2 (Sigma) for 20 min or irradiated with
5 Gy. After treatment with Accutase, the cell suspension was added to 180 ml of
molten Low Melting-Agarose (0.7% LM-agarose) at 37 1C, and immediately
layered onto Comet slides, covered with a coverslip and stored at 4 1C in the dark.
After 25 min, the coverslip was removed and a second LM-Agarose layer was
layered onto the slides. The slides were then transferred to a pre-chilled lysis
solution (2.5 M NaCl, 100 mM EDTA, 10 mM Tris-base, 300 mM NaOH, pH 10,
and 1% freshly added Triton X-100) for 2 h at 4 1C in the dark. In the alkaline
condition, the slides were placed in a horizontal electrophoresis chamber and the
denaturation step was performed in pre-chilled electrophoresis solution (300 mM
NaOH, 1 mM EDTA, pH413) at 4 1C in the dark. After 20 min, the slides were
electrophoresed (1 V/cm, 300 mA) for 20 min, and washed with neutralization
buffer (0.4 M Tris-HCl, pH 7.4). In the neutral condition, after the lysis the slides
were washed twice with 1 Tris-borate EDTA buffer solution pH 8.3 (TBE) for
10 min. Electrophoresis was carried out at the rate of 1.0 V/cm for 20 min. The
slides were then washed in deionized water for 5 min. MACA on post-mitotic
neurons was performed as described.34 After electrophoresis, Comet slides were
fixed with pre-chilled methanol, air dried overnight, stained with ethidium bromide
(0.1 mg/ml) for 10 min, and images acquired with an Eclipse E1000 Nikon
fluorescence microscope equipped with a DMX1200F CCD camera. Comet
images were processed with Comet Score software (v1.5; TriTek Corporation,
Sumerduck, VA, USA).
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
13
Rapid approach to DNA adduct recovery. Post-mitotic neurons (D30)
were incubated with or without 30 mM CPT for 1 h. Following treatment, part of the
cells was immediately lysed, while the remaining washed with PBS and incubated
in fresh medium for 3 h before lysis. For DNA–protein covalent complexes
isolation, cells were lysed in the culture plates by addition of 2 ml of MB lysis
reagent (6 M GTC, 10 mM Tris-HCl, pH 6.5, 20 mM EDTA, 4% Triton X100, 1%
Sarkosyl and 1% dithiothreitol) and processed as described.35 Purified DNA was
vacuum blotted on nitrocellulose membranes and Top1 detected using a mouse
monoclonal anti-Top1 antibody (BD Biosciences) and ECL. Sample loadings were
normalized for DNA content using DAPI.
Neuronal viability assay. Neuronal viability was evaluated using the Cell
Titer-Glo luminescent assay (Promega, Madison, WI, USA). Briefly, hNPCs were
seeded into Matrigel-coated 96-well plates (10 000 cells per well) and let to
differentiate for 30 days, after which they were treated in triplicate with CPT, H2O2,
Paraquat (all from Sigma) or trabectedin (Yondelis; ET-743) for 72 h. H2O2 was
removed after 20 min. Afterward, a volume of CellTiter-Glo Reagent was added to
each well and luminescence measured using a Tecan Genios instrument
(Männedorf, Switzerland).
Conflict of Interest
The authors declare no conflict of interest.
Acknowledgements. This work was financially supported by Comitato
Telethon Fondazione Onlus (grant # GGP10066 to DD) and by the Italian Association
for Cancer Research (AIRC) (grant # IG-10248 to DD). Dr. Carlessi Luigi is recipient of
a post-doctoral fellowships award from Fondazione Umberto Veronesi. We wish to
thank Dr. Sherif El-Kamisy, Department of Molecular Biology and Biotechnology,
Sheffield University, UK, for kindly providing the detailed MACA protocol, and
Dr. Christian Unger, Centre for Stem Cell Biology, Sheffield University, UK, for
providing training (to DL) and expertise in the field of iPSCs, as well as reagents.
1. Shiloh Y, Ziv Y. The ATM protein kinase: regulating the cellular response to genotoxic
stress, and more. Nat Rev Mol Cell Biol 2013; 14: 197–210.
2. Ambrose M, Gatti RA. Pathogenesis of ataxia-telangiectasia: the next generation of ATM
functions. Blood 2013; 121: 4036–4045.
3. McKinnon PJ. DNA repair deficiency and neurological disease. Nat Rev Neurosci 2009; 10:
100–112.
4. Verhagen MM, Martin JJ, van Deuren M, Ceuterick-de Groote C, Weemaes CM, Kremer BH
et al. Neuropathology in classical and variant ataxia-telangiectasia. Neuropathology 2012;
32: 234–244.
5. Barzilai A, Rotman G, Shiloh Y. ATM deficiency and oxidative stress: a new dimension
of defective response to DNA damage. DNA Repair 2002; 1: 3–25.
6. Liu N, Stoica G, Yan M, Scofield VL, Qiang W, Lynn WS et al. ATM deficiency induces
oxidative stress and endoplasmic reticulum stress in astrocytes. Lab Invest 2005; 85:
1471–1480.
7. Guo Z, Kozlov S, Lavin MF, Person MD, Paull TT. ATM activation by oxidative stress.
Science 2010; 330: 517–521.
8. Li J, Han YR, Plummer MR, Herrup K. Cytoplasmic ATM in neurons modulates synaptic
function. Curr Biol 2009; 19: 2091–2096.
9. Siddoway B, Hou H, Yang H, Petralia R, Xia H. Synaptic activity bidirectionally regulates
a novel sequence-specific S-Q phosphoproteome in neurons. J Neurochem 2014; 128:
841–851.
10. Li J, Chen J, Ricupero CL, Hart RP, Schwartz MS, Kusnecov A et al. Nuclear accumulation
of HDAC4 in ATM deficiency promotes neurodegeneration in ataxia telangiectasia.
Nat Med 2012; 18: 783–790.
11. Li J, Hart RP, Mallimo EM, Swerdel MR, Kusnecov AW, Herrup K. EZH2-mediated H3K27
trimethylation mediates neurodegeneration in ataxia-telangiectasia. Nat Neurosci 2013;
16: 1745–1753.
12. Barzilai A. The neuro-glial-vascular interrelations in genomic instability symptoms. Mech
Ageing Dev 2011; 132: 395–404.
13. Borghesani PR, Alt FW, Bottaro A, Davidson L, Aksoy S, Rathbun GA et al. Abnormal
development of Purkinje cells and lymphocytes in Atm mutant mice. Proc Natl Acad Sci
USA 2000; 97: 3336–3341.
14. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K et al. Induction
of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 2007; 131:
861–872.
15. Nayler S, Gatei M, Kozlov S, Gatti R, Mar JC, Wells CA et al. Induced pluripotent stem cells
from ataxia-telangiectasia recapitulate the cellular phenotype. Stem Cells Transl Med
2012; 1: 523–535.
16. Carlessi L, Fusar Poli E, De Filippis L, Delia D. ATM-deficient human neural stem cells
as an in vitro model system to study neurodegeneration. DNA Repair (Amst) 2013; 12:
605–611.
17. Carlessi L, Fusar Poli E, Delia D. Brain and induced pluripotent stem cell-derived neural
stem cells as an in vitro model of neurodegeneration in ataxia-telangiectasia. Exp Biol Med
(Maywood) 2013; 238: 301–307.
18. Verpelli C, Carlessi L, Bechi G, Fusar Poli E, Orellana D, Heise C et al. Comparative
neuronal differentiation of self-renewing neural progenitor cell lines obtained from human
induced pluripotent stem cells. Front Cell Neurosci 2013; 7: 175.
19. Carlessi L, De Filippis L, Lecis D, Vescovi A, Delia D. DNA-damage response, survival
and differentiation in vitro of a human neural stem cell line in relation to ATM expression.
Cell Death Differ. 2009; 16: 795–806.
20. Momilovic O, Choi S, Varum S, Bakkenist C, Schatten G, Navara C. Ionizing radiation
induces ataxia telangiectasia mutated-dependent checkpoint signaling and G2 but not
G1 cell cycle arrest in pluripotent human embryonic stem cells. Stem Cells 2009; 27:
1822–1835.
21. Momcilovic O, Knobloch L, Fornsaglio J, Varum S, Easley C, Schatten G. DNA damage
responses in human induced pluripotent stem cells and embryonic stem cells. PLoS One
2010; 5: e13410.
22. Xu G, Herzig M, Rotrekl V, Walter CA. Base excision repair, aging and health span. Mech
Ageing Dev 2008; 129: 366–382.
23. Fortini P, Ferretti C, Pascucci B, Narciso L, Pajalunga D, Puggioni EMR et al. DNA damage
response by single-strand breaks in terminally differentiated muscle cells and the control of
muscle integrity. Cell Death Differ 2012; 19: 1741–1749.
24. Sykora P, Yang JL, Ferrarelli LK, Tian J, Tadokoro T, Kulkarni A et al. Modulation of DNA
base excision repair during neuronal differentiation. Neurobiol Aging 2013; 34: 1717–1727.
25. Hegde ML, Hegde PM, Rao KS, Mitra S. Oxidative genome damage and its repair in
neurodegenerative diseases: function of transition metals as a double-edged sword.
J Alzheimers Dis 2011; 24(Suppl 2): 183–198.
26. Benitez-Bribiescal L, Sanchez-Suarez P. Oxidative damage, bleomycin, and gamma
radiation induce different types of DNA strand breaks in normal lymphocytes and
thymocytes: a comet assay study. Ann. N Y Acad Sci 1999; 887: 133–149.
27. Riballo E, Kuhne M, Rief N, Doherty A, Smith GC, Recio MJ et al. A pathway of doublestrand break rejoining dependent upon ATM, Artemis, and proteins locating to gammaH2AX foci. Mol Cell 2004; 16: 715–724.
28. Schultz LB, Chehab NH, Malikzay A, Halazonetis TD. p53 binding protein 1 (53BP1) is an
early participant in the cellular response to DNA double-strand breaks. J Cell Biol 2000;
151: 1381–1390.
29. Takagi M, Delia D, Chessa L, Iwata S, Shigeta T, Kanke Y et al. Defective control of
apoptosis, radiosensitivity, and spindle checkpoint in ataxia telangiectasia. Cancer Res
1998; 58: 4923–4929.
30. Liu LF, Desai SD, Li T, Mao Y, Sun M, Sim S. Mechanism of action of camptothecin. Ann N
Y Acad Sci 2000; 922: 1–10.
31. Minuzzo M, Marchini S, Broggini M, Faircloth G, D’Incalci M, Mantovani R. Interference of
transcriptional activation by the antineoplastic drug ecteinascidin-743. Proc Natl Acad Sci
USA 2000; 97: 6780–6784.
32. McCarthy S, Somayajulu M, Sikorska M, Borowy-Borowski H, Pandey S. Paraquat induces
oxidative stress and neuronal cell death; neuroprotection by water-soluble coenzyme Q10.
Toxicol Appl Pharmacol 2004; 201: 21–31.
33. Suraweera A, Becherel OJ, Chen P, Rundle N, Woods R, Nakamura J et al. Senataxin,
defective in ataxia oculomotor apraxia type 2, is involved in the defense against oxidative
DNA damage. J Cell Biol 2007; 177: 969–979.
34. Alagoz M, Chiang SC, Sharma A, El-Khamisy SF. ATM deficiency results in accumulation
of DNA-topoisomerase I covalent intermediates in neural cells. PLoS One 2013; 8: e58239.
35. Kiianitsa K, Maizels N. A rapid and sensitive assay for DNA–protein covalent complexes in
living cells. Nucleic Acids Res 2013; 41: e104.
36. Spring K, Cross S, Li C, Watters D, Ben-Senior L, Waring P et al. Atm knock-in mice
harboring an in-frame deletion corresponding to the human ATM 7636del9 common
mutation exhibit a variant phenotype. Cancer Res 2001; 61: 4561–4568.
37. Elson A, Wang Y, Daugherty CJ, Morton CC, Zhou F, Campos-Torres J et al. Pleiotropic
defects in ataxia-telangiectasia protein-deficient mice. Proc Natl Acad Sci USA 1996; 93:
13084–13089.
38. Biton S, Gropp M, Itsykson P, Pereg Y, Mittelman L, Johe K et al. ATM-mediated response
to DNA double strand breaks in human neurons derived from stem cells. DNA Repair 2007;
6: 128–134.
39. Lavin MF. The appropriateness of the mouse model for ataxia-telangiectasia: neurological
defects but no neurodegeneration. DNA Repair (Amst) 2013; 12: 612–619.
40. Jakel RJ, Schneider BL, Svendsen CN. Using human neural stem cells to model
neurological disease. Nat Rev Genet 2004; 5: 136–144.
41. Stewart R, Kozlov S, Matigian N, Wali G, Gatei M, Sutharsan R et al. A patient-derived
olfactory stem cell disease model for ataxia-telangiectasia. Hum Mol Genet 2013; 22:
2495–2509.
42. Kuljis RO, Xu Y, Aguila MC, Baltimore D. Degeneration of neurons, synapses, and neuropil
and glial activation in a murine Atm knockout model of ataxia-telangiectasia. Proc Natl
Acad Sci USA 1997; 94: 12688–12693.
43. Abdelalim EM. Molecular mechanisms controlling the cell cycle in embryonic stem cells.
Stem Cell Rev 2013; 9: 764–773.
Cell Death and Disease
Defects of ATM-deficient neural-specified hiPSCs
L Carlessi et al
14
44. Momcilovic O, Navara C, Schatten G. Cell cycle adaptations and maintenance of genomic
integrity in embryonic stem cells and induced pluripotent stem cells. Results Probl Cell
Differ 2011; 53: 415–458.
45. Tichy ED, Pillai R, Deng L, Liang L, Tischfield J, Schwemberger SJ et al. Mouse embryonic
stem cells, but not somatic cells, predominantly use homologous recombination to repair
double-strand DNA breaks. Stem Cells Dev 2010; 19: 1699–1711.
46. Chou WC, Wang HC, Wong FH, Ding SL, Wu PE, Shieh SY et al. Chk2-dependent
phosphorylation of XRCC1 in the DNA damage response promotes base excision repair.
EMBO J 2008; 27: 3140–3150.
47. Adams BR, Golding SE, Rao RR, Valerie K. Dynamic dependence on ATR and ATM for
double-strand break repair in human embryonic stem cells and neural descendants. PLoS
One 2010; 5: e10001.
48. Woodbine L, Brunton H, Goodarzi AA, Shibata A, Jeggo PA. Endogenously induced DNA
double strand breaks arise in heterochromatic DNA regions and require ataxia telangiectasia
mutated and Artemis for their repair. Nucleic Acids Res 2011; 39: 6986–6997.
49. Herzog K, Chong MJ, Kapsetaki M, Morgan JI, McKinnon PJ. Requirement for Atm in
ionizing radiation-induced cell death in the developing central nervous system. Science
1998; 280: 1089–1091.
50. Lee Y, Chong MJ, McKinnon PJ. Ataxia telangiectasia mutated-dependent apoptosis after
genotoxic stress in the developing nervous system is determined by cellular differentiation
status. J Neurosci 2001; 21: 6687–6693.
51. Tian B, Yang Q, Mao Z. Phosphorylation of ATM by Cdk5 mediates DNA damage signalling
and regulates neuronal death. Nat Cell Biol 2009; 11: 211–218.
52. Solier S, Ryan MC, Martin SE, Varma S, Kohn KW, Liu H et al. Transcription poisoning by
Topoisomerase I is controlled by gene length, splice sites, and miR-142-3p. Cancer Res
2013; 73: 4830–4839.
53. Glass D, Vinuela A, Davies MN, Ramasamy A, Parts L, Knowles D et al. Gene expression
changes with age in skin, adipose tissue, blood and brain. Genome Biol 2013; 14: R75.
54. Berchtold NC, Coleman PD, Cribbs DH, Rogers J, Gillen DL, Cotman CW. Synaptic genes
are extensively downregulated across multiple brain regions in normal human aging and
Alzheimer’s disease. Neurobiol Aging 2013; 34: 1653–1661.
55. Lu T, Pan Y, Kao SY, Li C, Kohane I, Chan J et al. Gene regulation and DNA damage in the
ageing human brain. Nature 2004; 429: 883–891.
56. Carlessi L, Buscemi G, Fontanella E, Delia D. A protein phosphatase feedback mechanism
regulates the basal phosphorylation of Chk2 kinase in the absence of DNA damage.
Biochim Biophys Acta 2010; 1803: 1213–1223.
57. Cestele S, Schiavon E, Rusconi R, Franceschetti S, Mantegazza M. Nonfunctional
NaV1.1 familial hemiplegic migraine mutant transformed into gain of function
by partial rescue of folding defects. Proc Natl Acad Sci USA 2013; 110:
17546–17551.
58. Frosina G, Cappelli E, Ropolo M, Fortini P, Pascucci B, Dogliotti E. In vitro base excision
repair assay using mammalian cell extracts. Methods Mol Biol 2006; 314: 377–396.
Cell Death and Disease is an open-access journal
published by Nature Publishing Group. This work is
licensed under a Creative Commons Attribution 3.0 Unported License.
The images or other third party material in this article are included in
the article’s Creative Commons license, unless indicated otherwise in
the credit line; if the material is not included under the Creative
Commons license, users will need to obtain permission from the
license holder to reproduce the material. To view a copy of this license,
visit http://creativecommons.org/licenses/by/3.0/
Supplementary Information accompanies this paper on Cell Death and Disease website (http://www.nature.com/cddis)
Cell Death and Disease