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The intraneuronal mechanism of action of BTX-A involves the proteolytic cleavage by its light chain of the protein SNAP 25. With syntaxin, synaptobrevin and synaptostagmin, these neuronal proteins form the core complex postulated to coordinate regulated vesicular fusion for neurotransmitter release. By preventing the assembly of this synaptic core complex, botulinum toxin inhibits neurotransmitter release, notably for cholinergic and catecholaminergic cells, but this can also occur for several classes of neutransmitters at central and peripheral neurones, once the toxin is injected intraneuronally in vitro.
However, after intramuscular injection in vivo, BTX-A binds by its heavy chain to high affinity ectoacceptors located on the presynaptic membrane of the neuromuscular junction. The toxin is then internalized across the membrane and undergoes a pH-induced translocation allowing its entry into the cytosol where the light chain cleaves SNAP 25. This high affinity of the heavy chain for the neuromuscular junction explains the selectivity of the clinical effect of BTX-A and the relative lack of effects other than neuromuscular junction blockade after intramuscular injection."...
Botulinum toxin is an injectable drug used to treat muscle spasm.
After injection into a muscle, the toxin is absorbed by the local nerve fibers, innervating the muscle (i.e., supplying the muscle with nerves) and blocking the transmission of impulses between nerve and muscle.
Normal muscles contract after a signal is transmitted from the brain, through the neuronal network, by nerve structures called axons. These axons connect with muscles at the neuromuscular junction. Every time a signal to contract is sent, axon terminals at the neuromuscular junction release packets of acetylcholine, a neurotransmitter. These packets float toward the muscle and cause contraction. Botulinum toxin prevents the release of acetylcholine from the axonal terminal, and as a result, blocks muscular contraction.
The nerve reacts by sprouting new branches and creating new junctions, called synapses, with the muscle. This sprouting and creation of new synapses continues for several weeks to a few months.
When it is completed, the nerve can again communicate normally with the muscle; this marks the end of the clinical effect of the injection. During the whole process, no detectable amount of toxin has entered the general circulation of the body.
PRESENTATION AND PRINCIPLES OF USE
Botox R (Allergan): lyophilised form of purified botulinum toxin type A. It is supplied sterile in glass vials, each containing 100 units of botulinum toxin type A, 0.5 mg of human albumin and 0.9 mg of sodium chloride. It contains no preservative. It must be diluted in sterile, non-preserved saline prior to use for injection.
The english toxin, Dysportr (Porton Down) is presented in vials of 50ng BTX-A haemagglutinin complex.
Storage: Careful handling of purified toxin is important for maintenance of stability: BTX-A is readily denatured by heat at temperatures above 40øC. This denaturation also happens in an atmosphere of nitrogen or carbon dioxide. The most satisfactory method of storage is to leave the crystalline toxin at 4 degrees in the mother liquor of the last crystallization, in which toxicity has been retained for 10 or more years (Schantz & Johnson, 1992).
Reconstitution and administration, according to the drug company recommendations: After drawing up the diluent, care must be taken in injecting it into the vial to avoid violent bubbling or agitation that could denature the toxin. Reconstituted toxin must be clear, colourless and free of particulate matter. It should then be stored in a refrigerator and, used within 4 hours of reconstitution. For small facial muscles a concentration of at least 10 to 25 units per mL of diluent is recommended (the higher the concentration, the less likely the diffusion to other muscles). According to experienced clinicians, concentrations can be lower for injections into bigger muscles of the limb such as the 20 U/ml used by J. Dunne et al. (1995 a,b). This table gives examples of different concentrations :
|Diluent added (mL)||Resulting concentration (U/mL)|
|50 (Brin et al., 1995)|
The english toxin can be diluted to a concentration of 5 ng/mL (Dengler et al., 1992).
When ready for use, the solution is drawn up into a standard 1 mL sterile disposable syringe with a 27-30 gauge needle attached for the injection. If alcohol is used to cleanse the skin, it must be allowed to dry before injection, as it could deactivate the toxin.
Dosage: From 50-100 units of the toxin should be injected into spastic limb muscles, the dosage depending on the size of the muscle and on the severity of its spasticity.
Pregnancy: "category B2": As animal reproduction studies have not been conducted with Botox injection, safety for use in pregnancy and lactation has not been established. Therapy will therefore be withheld from pregnant and nursing women or women of childbearing age unless use of a suitable contraceptive is confirmed.
Should accidental injection or oral ingestion occur, the person should be medically supervised for several days on an office or outpatient basis for signs and symptoms of systemic weakness or muscle paralysis. The entire content of a vial is below the estimated dose for toxicity in humans weighing 6 kg or greater.
In the event of overdosage or injection into the wrong muscle, additional information may be obtained by contacting Allergan Australia on (008) 252224.
As with all biological products, epinephrine (adrenaline) and other precautions as necessary should be available should an anaphylactic reaction occur. Location of treatment in outpatient setting is appropriate with the proviso that appropriate emergency care is readily available if needed.
(other than spasticity)
Botulinum toxin acting on the final common pathway, in theory muscular hyperactivity of any cause could be temporarily relieved by this treatment.
Following about 10 years of experimental treatment on human volunteers, crystalline BTX-A (Oculinumr or Botoxr) was first licensed in the United States in 1989 by the Food and Drug Administration for treatment of three human spasmodic muscle disorders:
1- Strabismus: after the pioneering action of Scott (1973, 1980), the toxin became generally accepted for the treatment of strabismus as an alternative to conventional incisional surgery (American Academy of Ophthalmology, 1989). The indications have so far been concomitant strabismus, vertical strabismus (Dissociated Vertical Deviation, injection into the superior rectus muscle of the non-dominant eye, Mac Neer, 1989), lateral rectus palsy (injection in medial rectus, Metz, 1988), nystagmus and dysthyroid myopathy (for references see Jankovic & Brin, 1991).
The injections are usually performed under electromyographic guidance with a Teflon-coated needle to ensure accurate placement. Most patients are treated in the office with topical anaesthesia. Follow-up studies up to five years after the injection show that 85 percent of the patients available for reassessment had satisfactory improvement (American Academy of Ophthalmology, 1989). Smaller deviations are more effectively corrected than larger deviations (Magoon, 1995). Side effects, including partial ptosis and secondary vertical deviations, are transient and do not result in amblyopia. Scleral perforation rate was 0.11% in Scott's experience (Magoon, 1995).
The technique is considered to be especially effective for conditions when surgery is inappropriate such as active thyroid ophthalmopathy or acute nerve palsy, for strabismic angles under 40 prism diopters and for postoperative residual strabismus following surgery. Some authors remain sceptical (Biglan et al., 1989) and prospective randomized clinical trials are needed to answer the question of whether botulinum toxin can be a substitute for or an adjunct to surgery.
2- Essential blepharospasm: Intermittent or sustained closure of the eyes due to involuntary contractions of the orbicularis oculi. The impairment may range from increased blinking to functional blindness. The injection is made in orbicularis oculi, of ca. 20 U of average dose. Its beneficial effects have been demonstrated in one controlled (Jankovic & Orman, 1987) and several open trials: 70% to 90% of significant benefits in more than 8000 treatments. Side-effects were temporary and transient: ptosis, blurred vision, diplopias by diffusion to neighbouring muscles; no adverse long term effects were observed after 7 years or more of repeated injections in some patients. There was no demonstrable decline in efficacy after repeated injections (Jankovic & Brin, 1991).
Because of its safety and efficacy, botulinum toxin is now considered a primary form of therapy for blepharospasm (Report of the Therapeutics and Technology Assessment Subcommittee of the American Academy of Neurology, 1990).
3- Post-paretic hemifacial "spasm" in patients 12 years of age and older: The post paretic hemifacial contracture is a tonic contraction of some muscles of one side of the face, causing constant facial twitching, often socially or psychologically embarrassing. The hemifacial spasm is characterized by involuntary, recurrent, paroxysmal clonic twitches of the eyelids and perinasal, perioral, zygomatic or other facial muscles. It is often attributed to compression or irritation of the facial nerve by an abnormal arterial vasculature in the posterior fossa (occasionally by a tumor). In that condition, microvascular decompression of the facial nerve has more often provided relief than systemic pharmacological therapies (Janetta et al., 1977), but this surgery has risks (permanent ipsilateral facial palsy, deafness, stroke, death). Direct injection of the toxin into the hyperactive muscle (5 to 30 U generally), has brought a relief in more than 90% of the patients treated (see Schantz & Johnson, 1992). According to some experienced investigators, it is often safer to confine the injection to hyperactive muscles of the upper part of the hemiface, because regional diffusion of the toxin is more likely to occur towards lower facial muscles, which are more sensitive to transient weakness (Geller et al., 1989).
Before 1989 and continuing until now, many clinical studies have been performed, trying to assess the efficacy and relative innocuity of BTX-A in various other conditions, all involving a focal increase in muscle tone or activity. Most of them are dystonias, i.e. syndromes dominated by involuntary sustained (tonic) or spasmodic (rapid, repetitive) patterned muscle contractions, frequently causing disabling movements or abnormal postures.
Before the introduction of botulinum toxin, some general medications could ameliorate the condition of some patients (benzodiazepines, some anticholinergic drugs, neuroleptics, carbamazepine, baclofen, reserpine, propanolol, primidone, etc...), but the majority failed to produce satisfactory relief, and adverse reactions were virtually universal, including even sometimes the induction of other movement disorders! Moreover, there was no scientific rationale for many of these pharmacological approaches. The alternative treatments were surgical (myectomies, tenotomies, rhizotomies or other functional neurosurgery, when possible).
These conditions are, approximately in decreasing order of number of patients treated (Jankovic & Brin, 1991):
Meige's syndrome (cranial-cervical dystonia), consisting of blepharospasm accompanied by spasms of facial, oromandibular, pharyngeal, laryngeal, or neck muscles. The results of botulinum toxin injection have been as good as in essential blepharospasm (Jankovic et al., 1990).
Spasmodic torticollis (cervical dystonia): This focal dystonia can result in twisting (torticollis, the most common form), tilting towards one shoulder (laterocollis), flexing (anterocollis) or extending (retrocollis) of the neck, but often a combination of these postures, making local treatment difficult. Physical therapy, relaxation techniques or properly fitted cervical braces can help, but usually not enough to improve patient's function and control of neck movement. After the first positive result from a controlled study of the efficacy of botulinum toxin in this indication (Tsui et al., 1986), others have been made, all clearly confirming the potency of the new treatment. Over 1000 cases had been treated by 1992 (Schantz & Johnson, 1992), with improvement of 50 to 90% of the patients, including most of the time relief of the frequently associated neck pain (Borodic et al., 1990). Transient side effects, usually resolving within two weeks, included swallowing difficulties (more frequently reported in studies with the english preparation of botulinum toxin), neck weakness and nausea (Jankovic & Brin, 1991).
Some authors favour an toxin action partly through a change in proprioceptive input (Kaji et al, 1995). A lateral effect of the treatment seems to be an improvement of brainstem auditory and trigeminal evoked potentials which were proven to be abnormal in patients with cranio-cervical dystonia (Giladi et al., 1995). Central effect of the toxin or artefact?
Prior to the availability of BTX-A, trihexyphenidyl was regarded as the first choice treatment. More recently, a prospective randomized, double-blind trial was performed in which BTX-A (EMG guided injections) was compared with trihexyphenidyl for the treatment of these patients (Brans et al., 1995). It showed a significantly better efficacy of BTX-A in disability and quality of life scores and a clinically meaningful difference in pain reduction even if not reaching statistical significance.
In addition, incidence of adverse effects was significantly lower in the BTX-A treatment group. The authors concluded that botulinum toxin should be considered as the treatment of choice in patients with cervical dystonia.
It is the unanimous opinion of all the physicians concerned that the most important determinant of a favorable response to treatment with botulinum toxin is proper selection of the involved muscles and choice of the dose (usually up to 150 units as a total dose per session). Careful examination of the patients by experienced investigators may become more reliable than electromyography in selecting the muscles to inject.
Oromandibular and lingual dystonia: It involves the masticatory, lower facial, and tongue muscles, resulting in trismus, bruxism, involuntary tongue movement and opening, closure or diduction of the jaw. The usual inefficacy of general pharmacotherapies led to the use of injections of botulinum toxin into the masseter, temporalis and internal pterigoid muscles (usually between 30 and 140 units per session). Improvement was obtained in approximately 70 percent of patients with jaw closure, the simplest pathological paradigm. A transient side effect could be weakness of the jaw, impeding closure (Jankovic et al., 1990).
Focal dystonias of the hand, like writer's cramp and other task-specific limb dystonias (typists, musicians etc..). In open trials, injections of botulinum toxin into selected muscles of the hand and forearm provided effective relief (Jankovic et al., 1990). In some studies, fine wire electrodes were used to localize bursts of muscle activation during the task and the toxin was injected into the belly of the most active muscle; in others the absence of electromyographic guidance did not prevent the investigators from obtaining as good results: between 80 and 90 percent improvement. In those cases, the selection of the muscles to be injected was usually based on clinical observation whilst the patient was writing or performing other involved tasks. Marion and MP Sheehy, relating their experience (Marion & Sheehy, 1995) insist on injecting only rarely more than two muscles, chosen as being likely the "trigger muscles", the other muscles involved in the spasm being often "compensatory features". They place an almost equal degree of importance upon reeducation, the retraining in this fine motor skill aiming at giving back the patient his schema for writing. Some transient hand weakness was possible but never considered as disabling as the cramps by the patients (Jankovic & Brin, 1991). Poewe and Wissel (1995) remark that none of their patients with various types of "musician's cramps" or those published in the literature has reached the premorbid performance level. A very documented study of the improvement of motor performance in 31 patients with writer's cramp showed that near normal movements parameters (speed or number of acceleration changes) were only reached in a subgroup whose base-line data were the least abnormal. The authors suggest that restoration of normal motor function in focal task specific dystonia could be possible when treatment is initiated as early as possible and when motor performance deterioration is still mild.
Adductor Laryngeal Dystonia (spastic dysphonia): It is an action-induced dystonia characterized by a strained, choked, strangled voice, possibly interrupted by voiceless pauses. There is another rarer type of spasmodic dysphonia due to abductor dystonia, clinically characterized by a more breathy, whispery voice. Again, treatments of these conditions (speech therapy, systemic pharmacotherapy, recurrent laryngeal nerve sections) were unrewarding before the advent of local injections of botulinum toxin.
The injection, through a monopolar, hollow, Teflon-coated needle, is electromyog-raphically guided into the thyroarytenoid vocalis complex (Brin et al., 1989). Injection is either made bilaterally, between 0.6 and 4 U for each vocal cord, or only on one side with a higher dose: 15 to 30 U (for references see Cohen et al., 1989). The studies report 80 to 100 percent improvement in voice symptoms, regardless of the technique. Side effects include transient breathy hypophonia, hoarseness and usually clinically unimportant aspiration of fluids. For abductor laryngeal dystonia, injection is electromyographically guided into posterior cricoarytenoid muscle (Jankovic et al., 1990).
Under these conditions, botulinum toxin is currently recommended as the primary therapy for laryngeal dystonia (American Academy of Otolaryngology, 1990)
Tremors (Essential tremor): Tremor is an oscillary movement produced by alternating or synchronous contractions of antagonistic muscles. Essential tremor is the most common movement disorder, becoming disabling when of high amplitude: patients often experience difficulty in daily tasks such as eating, writing and pouring of liquids. It is a more recent indication for BTX-A and pilot studies have first been completed: patients with hand tremor received approximately 50 U of BTX-A in forearm flexor and extensor muscles and those with head and neck tremor received about 240 U in cervical muscles. A functional improvement was reported in two thirds of the patients (Jankovic & Brin, 1991). Then Jankovic recently presented a double-blind placebo-controlled study (Jankovic et al., 1995) in which patients with essential hand tremor were treated with botulinum neurotoxin type A. Based on the physician-blinded tremor severity rating scale, patients receiving botulinum toxin experienced a significant improvement in postural and kinetic tremors at four weeks post injection, observation which was consistent with that from patients' self-assessment.
Seventh nerve palsy:
for a protective ptosis if corneal exposure, as an alternative to suturing: injection in levator palpebrae superioris, for references see Adams et al. 1987; - for correction of facial asymmetry (Clark, 1989). - Eyelid entropion, using the same technique of injection in levator palpebrae superioris (Clarke & Spalton, 1988).
Nystagmus, with injection site in extraocular muscles (Magoon, 1995).
Hyperkinesias following hypoglossal-facial anastomosis (Dressler, 1990);
Other voice disorders: vocal tremor, stuttering, using, as for laryngeal dystonias, injections into the vocalis complex (Ludlow,1990).
Neurogenic bladder: Injections of botulinum toxin into sphincter muscles have been found to improve bladder function in patients with detrusor-shincter dyssynergia due to spinal cord injury (Dykstra et al., 1988; Dykstra & Sidi, 1990).
Anismus (spasm of the rectal shincter) associated with intractable constipation has been reported to respond favorably to local injections of botulinum toxin (Hallan et al., 1988).
Anal fissure: As an alternative to surgical procedures Jost et al. (1995) have treated patients with BTX-A. The injection produced paresis of the sphincters for approximately 3 months, allowing the fissure to heal completely in 78% of patients where 91% reported complete pain relief. No patients experienced incontinence.
Tardive dyskinetic syndrome (Truong et al., 1990).
Achalasia (Pasricha et al., 1995): It is a disorder of osophageal motil
Many of these dystonias are progressively adknowledged to be indications of BTX-A as primary treatment (National Institutes of Health consensus development statement, 1991). In all of these conditions, toxin injection generally relieves undesired muscle contraction for 3 to 4 months on average, after which a repeated injection is required, and so on. The latency period from injection to onset of improvement is 2 to 6 days for small head and neck muscles.
With the American form of the toxin, the LD50 in monkeys after intravenous (see Jankovic & Brin, 1991) or intramuscular administration has been estimated to be 40 U per kg of body weight (Herrero et al., 1967; Scott & Suzuki, 1988). The dosages used in human therapeutic applications are roughly proportional to the mass of the muscle being injected and are much lower than the estimated LD50 which would be, for an adult male (70 kg), 2000 ng or 5000 U (35.7 vials of Botoxr) on the basis of the data obtained in monkeys (Scott, 1981). Intramuscular doses as high as 6 U/kg have been administered to human adults without systemic symptoms (Koman et al., 1994).
No adverse clinical effects at a distance from the injection point have been reported in patients who received low doses of botulinum toxin, e.g. under 20 U. This has been interpreted physiologically by the rapid and high affinity binding of the toxin to the neuromuscular junction acceptors, which prevents significant systemic absorption (see Physiology; Drachman & Houk, 1969). The primary side effect was then weakening of nearby muscles, considerably dependent on the clinician's technical experience.
With use of relatively large quantities of botulinum toxin (140 to 165 U), single-fiber electromyography analyses have shown subclinical blockings of neuromuscular transmission (Sanders et al., 1986; Lange et al., 1991). Elsewhere, some case reports of weakening of distant muscles have been published (Borg-Stein et al., 1993), possibly associated with pain evoking then brachial plexopathy (2 cases in Glanzmann et al., 1990 and Sampaio et al., 1993) resembling neuralgic amyotrophy syndrome (2 cases after upper limb injections with the british toxin Dysportr partly resolving within a few weeks, Sheean et al., 1995) .
Local or minor
Whatever the dose, two minor complications have to be mentioned:
-the first one is hematoma formation at the injection site and can be due to puncture of small blood vessels during injection. Bleeding can be stopped by external pressure. Patients receiving anticoagulant therapy may be excluded of or at least carefully monitored in any study on BTX-A.
-the second is the possibility of a diffuse skin rash, especially after repeated injections, resembling neurodermatitis.
They have been partly evaluated:
- Toxin spread from the injection site: Borodic and colleagues (1990) have made attempts to quantify this phenomenon, in animals, establishing a denervation gradient using acetylcholinesterase staining, ATPase staining and muscle fiber size analyses. A significant gradient effect was found up to 3 cm from the site of injection of 2 to 3 U/kg (dose which would correspond to ca 120 to 180U in a standard human subject). The denervation was transient, following a similar 3 months time course as the clinical effect.
- Immunity to botulinum toxin: Although more work is needed to evaluate the incidence of antibody formation and its clinical impact on long-term treated patients, this does not seem to be an issue of concern so far. The dose of toxin required to trigger significant antibody formation in humans is not known, but recurrent episodes of botulism in patients have been reported (Beller & Middaugh, 1990), suggesting that repeated exposure to botulinum toxin may not impart long-term immunity. Two tests are available to detect and quantify toxin antibodies: an in vivo mouse neutralization assay (protected mice remain healthy after the injection of both serum and botulinum toxin), mostly used, and an enzyme-linked immuno-sorbent assay (Desfulian & Bartlett, 1984). From the model of laboratory workers who repeatedly receive toxoid injections, it is known that anti-A antibodies are slow to develop and do not rise steeply before the fourth year of immunization (Siegel, 1988). In patients injected with BTX-A, antibody formation has been observed in a very small number of cases (about 12 of more than 7000 patients treated in 1991, according to Jankovic & Brin, 1991; see also Hambleton et al., 1992). According to the test used, bioassay or ELISA, there are so far diverging preliminary answers to the question whether this phenomenon can reduce or not the beneficial effect of the treatment (Tsui et al., 1988; Schwartz & Jankovic, 1990; Kessler et al., 1995). However, it is recommended that the doses be kept as low as possible for repeated injections, in order to minimize the risk of seroconversion and reinjection on an as needed basis. The frequency of injections and booster injections are also factors that were incriminated in increasing this risk.
Two other theoretical problems have so far never been reported: hypersensitivity
to the toxin, and teratogenicity even though several women have been injected
during their pregnancy.
Side effects could be increased in patients who are being treated with drugs other than botulinum toxin that affect neuromuscular transmission (Argov & Mastaglia, 1979).
Antitoxin: There are interest and current studies among physicians and scientists in developing methods to prevent the effect of the toxin in neighboring muscles. Scott demonstrated that injection of antitoxin at the correct time following toxin injection partially prevented toxin migration (Scott, 1988). Since the currently available equine antitoxin could lead to undesirable antiserum reactions in some patients, it could be planned to use human immunoglobin G pooled from immunized human volunteers (potential treatment for infant botulism, Frankovich & Arnon, 1991) to try to alleviate these side effects.
Others? Physiological ways of reducing the action of BTX-A were found
- increase in the intracellular calcium concentration (by ionophore treatment in synaptosome preparations, Ashton & Dolly, 1991)
- aminopyridine (Gansel et al., 1987).
- possibly Enalapril (Captopril) that would reduce the activity of the toxin by an anti-zinc effect.
- the very desirable approach of confining the paralyzing action to the presynaptic nerve by using presynaptic intraneuronal injection of L chain mRNA, has been done successfully in Aplysia californica (depressing transmitter release in the bath, the synthesis of L chain being demonstrated to happen in the presynaptic neuron, Mochida & coll., 1990), but this is still far from possible in clinical practice.
Other serotypes of botulinum toxin
Evidence is accumulating to show that different types bind to different acceptors
and may have subtle variations in their modes of action. However, they all
cause chemical denervation at the myoneural junction by inhibiting acetylcholine
release. The other types could therefore :
- complement type A in some clinical applications;
- be combined to type A toxin to achieve better efficacy;
- also be used in some of the patients who develop immunity to type A
Indeed, preliminary studies (for references, see Schantz & Johnson, 1992) indicate that types B (Borodic et al., ) and F (Siegel, 1989; Scott, ) could control certain spasmodic muscle disorders.
Genetic synthesis of botulinum toxin ?
This seems difficult to achieve because the protein synthesized by Escherichia coli when transformed with plasmids containing genomic libraries of type A is only the neurotoxin protein, without other non toxic proteins of the natural toxin complex which guarantee its effective clinical toxicity and its stability through dilution (Binz et al., 1990).
The genes of these other non-toxic proteins have been identified, they have a similar genetic arrangement, maybe a common ancestral origin, but it has not proved possible yet to synthesize the total toxin complex, as occurs under natural conditions (for references see Schantz & Johnson, 1992). No clinical trial on primates have been performed with purified neurotoxins.
It also points out the need for research on methods for culturing, purification, genetic expression, and preservation of this toxin more easily applicable to its use for human treatment.
Jean-Michel Gracies MAJ 10/1997
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