Incontro Congiunto SINC, SIRN, SIMFER
“Update su Tossina Botulinica, Spasticità e Dolore”
Quale ruolo della
Tossina Botulinica nel
Trattamento del
Dolore?
Valeria Tugnoli
U.O.S. di Neurofisiologia
D.A.I. Neuroscienze-Riabilitazione – Ferrara
[email protected]
Agenda
BoNT and Pain  Serendipitous Target
Clinical application on Pain
How do BoNT reduces pain
Agenda
BoNT and Pain  Serendipitous Target
Clinical application on Pain
How do BoNT reduces pain
Serendipity is the effect by which one
accidentally discovers something fortunate,
while looking for something else
BoNT and Pain
1) 2/3 pts affected by Cervical Dystonia (CD)  PAIN
 76-93% of Pain in CD has been reduced by
BoNT, also if dystonic pattern didn’t change,
 Different timing (Tsui ‘85, Jancovic ‘91, Freund ‘03)
2) …..Patients treated for
facial wrinkles
 ….. improved
migraine headache (Binder ‘00)
3) Chronic anal fissure
improvement before healing,
long-lasting resolution of pain
(Runfola et al 2006)
Pain severity
masseter
spasm
7
pain
3
Before
1 month after BoNT
follow up
Jancovic ‘91
Freund ‘03
Agenda
BoNT and Pain  Serendipitous Target
Clinical application on Pain
How do BoNT reduces pain
Jeynes LC et al 2008
Disease
BoNT
Level
Evidence
BoNT/a BoNT/B
1A
BONT/A
1B
BoNT/A BoNT/B
1B
Neuropathic pain, Phantom limb
BoNT/A
1C
Myofascial pain, Chronic LBP
BoNT/A
2A
CTS, Joint Pian, CRPS,
Trigeminal Neuralgia, PHN, Neuroma,
Brachial Plexopathy,
Peripheral Neuropathy, MS
BoNT/A
1C
2C
Cervical Dystonia
Pelvic Pain, Plantar Fasciitis,
Temporomandibular pain
Piriformis syndrome,
Whiplash
Notes
2B against MFP
Muscle-related pain and BoNT
 Dystonia (Jankovic J ‘91, Costa J ‘06)
 Spasticity (Group SS M&N ‘97)
 Myofascial pain and trigger points, Low Back Pain
(Porta 2001, Casale 2007, Forster 2007, Jabbari 2007, 2008)
 Chronic cervical pain (Chesire PIN ’94, Porta Pain’00, Wheeler Pain’01)
 Anal fissures, Chronic Pelvic pain, Hemorrhoidectomy pain
(Gui ’94, Zermann ’00, Thomson ’05,
Abbott ‘06, Patti ’06, Lai ’07, Sinha ’09, Apostolidis A’09)
 “Whiplash” neck pain
(Freund Headacke ’00)
 “Painless arms/moving fingers” (Singer ’07)
 Postoperative pain (muscle spasms, adductor muscle contracture)
(Adler ‘96, Racette ’98, Kern ‘03, Schofferman ‘05, Cannard ‘05, Hamdy ’07, Filipovic’09, Hamdy’09, Santamato ‘10)
 Temporomandibular Dysfunction
(Berardelli ’94, Brin ’95,Nixdorf 2002)
Not muscle-related pain and BoNT
 Primary headache prophylactic treatment:
Migraine, Chronic Daily and Tensive Headache (Silberstein ’01,’05,’06, Evers ’04,’06, Dodick ’05, SchulteMattler ’07, Relja ‘07, Straube ’08, Burstein R’09)
 Chronic Facial Pain
(Borodic ‘02)
Yan’09)
, Acute angle glucoma (Chien ‘10), Aphthous ulcer (
 Painful Neuropathies, CRPS
 Trigeminal
(Allam ‘05, Piovesan ‘05, Liu ’06) ,
 Post-herpetic Neuralgia
(Argoff ‘02, Bach.-Rojecky ‘07, Knoderer ‘07)
Occipital Neuralgia (Kapural ’07,Taylor’08)
(Dykstra ‘04, Schwartz ‘04, Liu ‘06, Ko ‘07)
 Notalgia paresthetica
(Weinfled ‘07),
 Painful Keloid
SCI Allodynia (Jabbari ‘03), Phantom Limb (Jin’09)
(Uyesugi ‘10),
 Interstizial Cystitis
(Heigelshoven ‘06),
(Smith et al ’04, Namazi ‘08)
 Refractory joint pain
Mahowald’09)
(Hou ‘07)
Thelalgia
(Monnier et al ’06, Mahowald ML et al’06, Singh ’06-’09, Monnier et al’07, Oskarsson et al’08, Singh’09,
,Plantar Fasciitis (Babcock et al ’05), Tennis Elbow (Lin ‘10), Patellofemoral Pain S.
 Stellate ganglion block for facial pain
(Wilkinson ‘06),
and CRPS (Carroll’09)
o 1 paziente con SM relapsing-remitting
o 1 paziente nevralgia post-herpetica
o 1 paziente con radiculopatia C8 da ED
o 1 paziente con neuropatia periferica
o Risultato:miglioramento del dolore
o Trattamento sicuro
o Ipotizzato meccanismo diverso dal blocco neuromuscolare
(blocco SP e/o del simpatico e/o altri sistemi…)
13 pazienti
To the Editor 
Studio non controllato
Effetto ipostenia….
 BTX-A (100 U of Clostridium botulinum type A neurotoxin complex, 5 mg gelatin, 25 mg
dextran, and 25 mg saccharose) Lanzhou
Biological Products Institute, China.
 Dilution = 2 mL saline solution
 22 pts 75 U (1.5 mL) of BTX-A, 15 points vs 20 pts placebo
 Epidermis or submucosally in the oral mucosa if the pain involved the oral
mucosa
 AE=5 short-term facial asymmetry, 3 oedema
Attacks frequency
Pain intensity
NEVRALGIA TRIGEMINALE E BONT
AUTORI
Sede
Anno
N° paz.
Micheli F et al
Italia
2002
1
Turk U et al
2005
8
Allam N et al
b
2005
Zuniga C et al
2008
12
Bohluli B et al
Iran
2011
15
Ngeow WC
Nair R
Malaysia
2010
1
Yoon SH et al
Korea
2010
1
WU C et al
China
2012
22 BoNT
20 Placeb
Dose
BoNT/A
Risultati
Dolore
Effettii
Collaterali
7 scomp/8 riduz
(5FANS-3Altri)
3 paz
Ipostenia mm.faciali
ridotto
(200 mg CBZ)
?
ridotto
?
100U
50-100U
(Trigger zone)
100U
trigger zone
(Cute)
75UI
(cute o mucosa)
5 paz
Ipostenia
Pain Medicine 2006
 Paziente di 80 aa
 Da 1 mese bruciore, gnawing tightness pain,Allodinia T2-4
 Terapia farmacologica VAS 104 ma effetti collaterali
 Catetere epidurale con anestetico inefficace
Arch Phys Med Rehabil 2004
 BoNT/A 100U  VAS 10 1 dopo 2 gg per 52 gg poi alla
ripresa dolore gestibille con terapia farmacologica classica
(amitriptilina e gabapentin)
 Effetto placebo??? Atipico l’andamento temporale
Arch Phys Med
Rehabil 2004
Arch Phys Med
Rehabil 2004
Pain Medicine 2006
ejpain 2007
 Studio randomizzato, controllato con placebo e lidocaina
 60 pazienti
 VAS, ore di sonno (basali,1-7-90 giorni), consumo di Farmaci
 RISULTATI


VAS diminuisce nei 3 gruppi, BoNT>lidocaina e placebo (7-90 gg)
Uso di farmaci: BoNT (21.1%), lidocaina (52.6%), placebo (66.7%)
 Randomized, controlled study vs placebo
 29 pts affected by post-herpetic neuralgia,




painfull neuropathy (trauma or surgery) since 6
months
VAS 3-10, area < 60cm2
Endpoints  50% decrease of daily pain (BPI),
previous 24 h pain, Allodinia area and intensity,
pressure and thermic pains
40% high improvement;7% mild improvement,
53% unchanged vs 7% placebo
Significant improvement of burning and
parossistic pain and allodynia (unchanged
deep pain, paresthesias)
Ann Neurol 2008

Conclusions: This pilot study found that botulinum
toxin type A significantly reduced diabetic
neuropathic pain (3 cm VAS  44% BoNT vs 0%
placebo) and transiently improved sleep quality.
Further large-scaled study is warranted.
………We are skeptical of the benefits of BoNT/A in
treating neuropathic pain. We also believe that Apfel’s
accompanying editorial was too positive regarding this
study involving only 18 patients…. BoNT/A has proven
indications, but the drive to expand them is industrydriven and is reminiscent of the marketing strategy of
Neurontin. Given its cost and mode of use, larger
studies funded by unrelated sources should be
pursued. (Torgovnick J et al, Neurology 2010)
Francisco GE 2012
 Rephractory joint pain  improved in 55%
knees-ankles, 71% shoulders (25-100U i.a.,
long duration 3-12 months)
 Active range of motion  increased 67% in
flexion and 42% in abduction (Mahowald 2006)
 Epicondylitis  contradictory results (Keizer 2002,
Placzek 2004, Hayton 2005, Oskarsson 2008)
 Safe treatment, no adverse events
 Controlled studies vs placebo are needed
Agenda
BoNT and Pain  Serendipitous Target
Clinical application on Neuropathic Pain
How do BoNT reduces pain
Blocco
esocitosi
Copyright
may apply.2001;285:1059-1070.
Arnon,
S. S.restrictions
et al. JAMA
BoNT and Pain:
effects on Muscular-related events
Casale R, Tugnoli V 2008
1.
2.
3.
4.
Inhibition of the Vicious Circle
muscle contraction-pain-muscle
contraction
Decompression of nociceptive
intramuscular fibres
Reduction of intrafusal afferents
from the muscle spindles to the
spinal cord
Central effect  reduction of
cord excitability and functional
central resetting  modulation
of sensorimotor descending
control system
BoNT and Pain:
effects on Non Muscular-related events
 Block of the PKC-induced surface expression of  Vanilloid receptors
TRPV1 (Morenilla-Palao ‘04)
 Inhibition of  neuropeptide release from primary sensory neurons (Glu
(Cui’00,’02),CGRP(Morris’01, Durham’04), Subst.P(Purkiss ‘’00), Norepinephrine (Shone’02), ATP(Smith
Hannover ‘02)), Trigeminal Ganglia (infraorbital nerve constriction IOC, Kitamura ‘09) in vitro and in vivo
(animals) models
 Reduction of peripheral sensitization (Formalin-pain models) and of the
related central sensitization (c-Fos,Wells ‘01)
 Different effects for BoNT/A and BoNT/B if administrated sc or ivc
(Luvisetto ‘06)
 Inhibition of Autonomic System and
Inflammatory cells (mastzellen)?
 Pre o post-injury treatment?
 Central effects?
 Acceleration of reparative proccesses?
of
Botulinum Toxin A inhibits the inflammatory pain
in rat formalin model. Cui et al. 2000
 BoNT A iniettato pianta via sottocutanea, poi formalina  licking
 Fase 1 (Dolore Acuto)  stimolazione diretta dei nocicettori e
infiammazione
 Interfase quiescienza modulazione centrale inibitoria del dolore
 Fase 2 (Dolore Cronico)  processi di sensibilizzazione centrale
 BoNT A causava una riduzione “licking”, solo con dosaggi alti fase 1,
per tutti i dosaggi Fase 2 (dosaggi alti= effetti sistemici)
Review of a proposed mechanism for the antinociceptive
action of botulinum toxin type A (Aoki KR ‘05)




Possibile ingresso per “pinocitosi”
release di sostanze eccitatorie dalle fibre nocicettive (Glu, CGRP,
Sost.P), e di sostanze attive pro-infiammatorie (BK, PGs,HA,5HT)
espressione dei recettori di superficie TRPV1
Conclusione:
infiammazione e sensibilizzazione nocicettori
periferici e
secondaria del processo di sensibilizzazione centrale
 Formalin-induced orofacial pain model (FT)
in male Rattus norvegicus
 The results support a temporary
antinociceptive effect of BoNT/A when used
as a preemptive treatment
Arq Neuropsiquiatr. 2011 Feb;69(1):56-6
***
10
VAS
***
***
8
2
1
6
4
2
***
*
c
protocol B
10
8
VAS
p2
p1
c
d1
d2
protocol A
6
4
2
0
0
5
10
15
20
25
30
35
40
45
5
Time (min)
10
15
20
25
30
35
40
45
Time (min)
protocol A
10
protocol
A
VAS
8
6
4
protocol B
protocol
B
2
0
25
30
35
2
Area (cm )
40
6
8
10
12
2
Area (cm )
14
(Neuroscience 2011)

The effects of low doses of BTX-A injected
into the rat whisker pad and sensory trigeminal
ganglion on formalin-induced facial pain.

Axonal transport was prevented by colchicine
injection into the trigeminal ganglion
Both peripheral and intraganglionic BTX-A
reduce phase II of formalin-induced pain.
Antinociceptive effect of BTX-A was
prevented completely by colchicine



The axonal transport of BTX-A was
obligatory for its antinociceptive effects,
via sensory neurons, directed to sensory
nociceptive nuclei in the CNS
Immunofluorescently labeled truncated
SNAP-25 (light red) in coronal sections of
rat caudal medulla 5 d after BTX-A
injection into the whisker pad.
Contra—TNC contralateral and Ipsi—
TNC ipsilateral to the site of injection.
single injection

An ineffective dose of BoNT/A (2 pg/paw) combined with an ineffective dose of morphine
(1 mg/kg) exerted a significant analgesic action both during the early and the late phases
of formalin test.
•
Tolerance to morphine = complex physiological response reflecting adaptive changes at
multiple levels  initiated by the activation of the μ-opioid receptors; role for glutamatergic
system, phosphorylation- dependent desensitisation and endocytosis of G protein coupled
μ-opioid receptors, reduction in the number of functional binding sites and in the
decreased responsiveness to the μ-opioid agonist
preadministration of BoNT/A
on morphine-induced tolerance

A single intraplantar injection of BoNT/A (15 pg/paw),
administered the day before the beginning of chronic
morphine treatment (7 days of s.c. injections of 20
mg/kg), was able to counteract the occurrence
ofTolerance to morphine

Immunofluorescence (IF) staining of
inflammatory markers at the spinal cord level,
such as p38 MAPK (p38), whose
phosphorylation (p-p38)(colocalization with
GFAP) is increased in inflammatory
conditions, particularly in glial cells, and
modulated by opioids
A)
The formalin-induced inflammatory pain
produced a strong enhancement of the
astrocytes’ expression. NaiveBoNT/A is
sufficient per se to induce a reduction of the
astrocytes’ expression
BoNT/A reduces the enhancement of the
expression of astrocytes induced by
inflammatory formalin pain.
repeated M administration tolerance (s-M-M
=s-s-s), significantly counteracted by the
pretreatment with a single administration of
BoNT/A (A-M-M)
B)
C)




BoNT/A significantly reduced the amplitude of neuropathic pain symptoms,
such as Chronic Constriction Injury (sciatic nerve) -induced mechanical
allodynia and thermal hyperalgesia, both in mice and rats
The antiallodynic and antihyperalgesic effects were long-lasting and
observable after a single BoNT/A injection
Functional recovery appeared in mice as demonstrated by the equal loading of
the hind limbs and by the walking pattern analysis
Acceleration of the regenerative processes (proteins’ expression associated
with nerve injury and repair, Cdc2, GAP-43, and with SC proliferation,
S100, GFAP)
EVIDENCEBASEDMEDICINE
Section Editor: Jan Van Zundert, MD, PhD, FIPP
Evidence for the Use of Botulinum Toxin in the Chronic Pain Setting
A Review of the Literature
Jeynes LC; Gauci CA Pain Practice, Volume 8, Issue 4, 2008 269–276
BTX has both direct and indirect actions:
 Direct:


1. inhibition of alpha and gamma-motor neurons;
2. inhibition of release of local nociceptive neuropeptides/agents via vesicle-dependent exocytosis.
 Indirect:



1. reduction in neurogenic inflammation;
2. alterations within the autonomic nervous system resulting in changes to regional perfusion, as well
as central changes affecting behavior and stress;
3. alterations in sensory patterns within the central nervous system (CNS).
The effects of these actions are:
1.prevention of spasm and the subsequent sensitization and activation of nociceptors, and the
recruitment of mechanoreceptors as nociceptors;
2. reduction in central sensitization and wind-up;
3. possible neuroplastic reorganization within CNS.
Is time to
challenge this
“ancient” chapter
Molecular Medicine Today 1996
?
Pavone and Luvisetto 2010
 Lectina da Erythrina cristagalli (ECL) binds
Gal in nociceptive peripheral and central
terminals
 LHN/A-ECL  selectively blocks SNAP25
exocitosys in vitro DRG (SP, CGRP)
 In vivo: C fibers drive decrease, without
interferring with A-beta function
 Long duration of the effect
 Possible new drug for the selective use
LHN/A-ECL
in neuropatic pain
ECL


Short synthetic peptide (TD-1) can facilitate effective transdermal
delivery of BoNT-A through intact skin useful to reduce inflammation
produced by activating nociceptors in the skin.
Peptide-mediated delivery of BoNT-A is an easy and noninvasive way
of administering the toxin that may prove to be useful in clinical
practice.
BoNT-A reduces saphenous
nerve-induced plasma extravasation
Take Home
message
 The “weight” of evidences for BoNT
efficacy in painful conditions is growing
but still debated
 It does appear that BoNT should be
useful in selected patients but we don’t
yet know what type of patients
 BoNT seems a promising and safe
treatment for different kind of pain, but
it must be further evaluated in wider
populations and in controlled studies
 Different mechanisms other than
cholinergic block do occur to justify
BoNT analgesia
Thanks
MODELLI ANIMALI
Dolore centrale: lesioni midollari
Dolore periferico: legatura/lesione
n.sciatico, radici spinali; malattie
(diabete, nevralgia post-herpetica,…)
Modello Formalina, Capsaicina….
Valutati allodinia termica e meccanica
Mechanisms of the antinociceptive effect of
subcutaneous BOTOX: inhibition of
peripheral and central nociceptive processing
(Cui et al Hannover 2002)
 Modello ratto trattato con formalina zampa non
varia soglia termica
 Diminuisce, con effetto dose-dipendente:
1)Release glutamato locale, 2)attività elettrica
neuroni corna dorsali MS, 3) espressione
spinale di C-fos (indicatore attività neuronale)
lamine I-II (circuiti monosinaptici), lamina V-VI
(circuiti polisinaptici)
 BoNTinvariato “early acute nociceptive (AN),
diminuisce late tonic nociceptive (TN)” (FIG.1)
 Effetto persistente 12 gg (FIG.2)
BONT/A
ivc
BONT/B
1.
Analgesic effect is different for BoNT/A and BoNT/B in relation to sc or ivc
administration
2.
3.
BoNT/A is able to inhibit Phase 2 both by sc and ivc administration
BoNT/B is able to inhibit Phase 1 only by sc administration and causes hyperalgesia if
administrated by ivc.
4.
BoNT/A blocks neuropeptide release (Glu, Subst P, CGRP) in peripheral and central
sensitization and inhibits inflammatory aspects (vasodilatation)
BoNT/B blocks central inhibitory GABA patways that express VAMP and not SNAP25 
hyperalgesia
5.
BoNT/A 
the reduction of pain and flare
induced by capsaicin
VAS
8 vs 5
(Tugnoli et al, 2007)
Flare Area
Pain Intensity
Pain Duration
Flowmetry
It is then suggested that BoNTA can exert anti-inflammatory actions through inhibition of
neurogenic vasodilation that may contribute to its analgesic effects in inflammatory pain models.
 BoNTA rapidly (within 2.5 h) exters the desensitizing effect on muscle nociceptors that doesn’t
result from a decrease in muscle blood flow
The results suggest that injection of BoNTA into craniofacial muscles acts to decrease migraine
headaches by rapidly decreasing the mechanical sensitivity of temporalis muscle nociceptors
through inhibition of glutamate release and by attenuating the provoked release of CGRP from
muscle nociceptors.
Spontaneous spikes GLU
Mechanical Thr
Temperature
MT after GLU
BoNT didn’t impair the withdrawal nociceptive reflex
Induced dose-dependent inhibition of carrageenan hyperalgesia (without
modifying oedema), only when applied in the treated carrageenan site
Induced dose-dependent inhibition of paclitaxel hyperalgesia in both sites
(no systemic diffusion). It remains to be clarified how BoNT may affect
proinflammatory cytokines cascade in DRG or spinal cord (synaptic plasticity or
retrograde axonal transport?)
Pharmacol Biochem Behav. 2009 Dec;94(2):234
Central origin of the antinociceptive action of botulinum toxin type A.
Bach-Rojecky L, Lacković Z
IASP Definition
of Neuropathic Pain
neuropathic pain
pain caused by primary
LESION of the nervous system 2003
peripheral neuropathic pain
pain caused by lesion of the
peripheral nervous system
pain neuropathic central
pain caused by lesion of the central
nervous system
Applicazione BoNT
Nevralgia post-herpetica, Nevralgia Trigeminale
Neuropatia diabetica, Neuropatia post-traumatica,
Merskey H et al. (Eds) In: Classification of Chronic Pain:
Descriptions of Chronic Pain Syndromes and Definitions of Pain Terms. 1994: 209-212.
Radiculopatia, SCI, (CRPS)
(Neuroscience 2011)


Axonal transport of BTX-A from periphery to CNS was previously identified only
in motoneurons, with unknown significance.
Occurrence of truncated SNAP-25 in TNC suggests that peripherally applied,
axonally transported BTX-A can affect second order central sensory neurons,


presynaptically by SNAP-25 cleavage in central terminals of primary afferent neurons
following transcytosis to second-order synapses (Antonucci et al., 2008, Kitamura et al. (2009)

BTX-A can alter central nociceptive
transmission of unknown neurotransmitters

The axonal transport of BTX-A was
obligatory for its antinociceptive effects,
via sensory neurons, directed to sensory
nuclei in the CNS
nociceptive