Understanding Toxin Resistance

Botulinum toxin type A (BoNT-A) has been used since the 1970s,¹
and these anti-wrinkle injections are one of the most popular
cosmetic treatments in the world.²
With estimates of nearly seven
million injections per year globally,³
it’s no wonder that the discussion
surrounding BoNT-A resistance and concerns of its effectiveness,
have become more prevalent. The issue of antibody-induced
treatment failure is well-known in the therapeutic setting, but with more
use cosmetically we are seeing more reports of resistance.^4,5 At the
time of writing, I have just had my first experience with a patient who
seemed to be resistant to all FDA approved toxin brands, which were
tried with no effect.  

Understanding toxin

Firstly, to understand how/why resistance happens we must
understand how botulinum toxin works. BoNT is composed of a core
neurotoxic protein and nontoxic accessory proteins (NAPS) that have
a light chain and a heavy chain. A normal vial of the neurotoxin will
consist of 150kD that contains 100kD heavy chain (which helps bind
the neurons) and a 50kD light chain that helps entry into the cell.
These heavy and light chains are linked together by a disulfide bond.⁶

The NAPS contain hemagglutinin (50Kd) and non-hemagglutinin
proteins (130kD) to help prevent the toxin from breaking down.^7,8 NAPS
vary in different brands, but most include albumin, sucrose, lactose,
sodium chloride and disodium succinate.⁶ BoNT prevents the release
of acetylcholine at the axon endings at the neuromuscular junction.
It does this by affecting the SNARE (SNAP receptor) proteins that
transport the acetylcholine and their docking with the presynaptic
membrane, before it is released into synaptic cleft. BoNT-A and
BoNT-B work differently, as BoNT-A works on the SNAP-25 protein
and BoNT-B works on synaptobrevin, and impairing this process
prevents acetylcholine from being released, therefore making it hard
for the muscles to contract.^6,8 

Causes and categorisations
of resistance 

Resistance is defined as the process wherein
the body forms an antibody response to a
drug medication, or something that the body
regards as foreign. An antibody response can
happen because botulinum toxin is regarded
as foreign by the host and therefore has the
potential to induce an immune response, this
then results in the production of antibodies.⁸
These antibodies block the pharmacological
effects of the botulinum neurotoxin.^9,10 There
are two types of resistance to BoNT-A type A:
primary and secondary. 

Primary is where a patient has had a
first-time treatment and has no response
instantly, however this seems to be very rare,
and there are a limited number of cases
that have been reported.¹¹ Whilst there is no
evidence suggesting the main causes of
primary failure, it is thought it may be caused
by a cross-reaction of other antibodies such
as tetanus toxin A, pre-existing antibodies
to botulinum toxin, chronic exposure to
botulinum in childhood, and abnormalities of
BoNT-A acceptors.¹ The main cause of this
BoNT-A resistance currently identified in the
literature is that the presence of a foreign
protein in the body causes antigens to induce a biological immune
response.^9,12-14 Antibodies can block the pharmacologic effects of
the botulinum toxin, which are known as neutralising antibodies,
and some patients can lose their response to botulinum injections
over a period of time.^12,14 The antibodies that can cause resistance to
botulinum toxin have not yet been defined, and immune responses
can differ between patients.^11,14 

Secondary resistance is described as initial results/benefits from
treatments followed by a decrease in effect or no effectiveness at a
later treatment date. This is defined as being after two subsequent
treatments of botulinum toxin treatment.¹¹ The literature suggests
that secondary resistance is likely to develop within the first year of
treatment, and if it’s not seen after the first four years it is highly unlikely
that it will develop.¹⁵ Currently, it is thought from the evidence that
secondary resistance occurs due to NAPS, however the research is
limited.¹⁶ It is thought that NAPS can act like an adjuvant in a vaccine
which stimulate your immune system causing an antibody response,
which makes resistance more likely. There is very little evidence at
present for us to understand why some patients get resistance and
some don’t, and much more research is needed to valid the current
research which suggests that NAPS can cause antibodies to be
produced.¹⁷ The existing evidence shows that the factors that may
contribute to antibody formation include longer large doses, a shorter
time period between injections and decreased purity of botulinum
toxin preparation.^18,19,20  

Studies in current literature 

Preliminary research in animals injected with IncobotulinumtoxinA
suggests that BoNT-A, which has the absence of complexing
proteins (proteins that have no therapeutic function and don’t
influence the diffusion of neurotoxin),^21,22 is indeed associated with reduced immunogenicity. A study involving cynomolgus monkeys
that received repeated four-weekly injections with 4, 8, or 16 U/kg
of IncobotulinumtoxinA or control group were not associated with
the development of neutralising antibodies in any animal, despite
clear evidence of biologic activity of the neurotoxin, which was more
prominent in the highest dose group.²³ However, this study lacked a
positive control group, therefore more research and data is required to
show that complexing proteins do indeed result in antibodies forming.
A recent cross-sectional study in humans contained 59 patients that
had exclusively been treated with IncobotulinumtoxinA (mono group)
and ³² patients having been treated with other BoNT-A preparations
less than nine times, who were then switched to at least 14 sessions
of IncobotulinumtoxinA treatment (switch group). The study checked
for the prevalence of neutralising antibodies, and was tested by
means of the mouse hemi-diaphragm assay (MHDA). The study
found that none of the patients in the mono and only two in the
switch group had a positive MHDA-test, suggesting that there’s
an association between neurotoxin (complex) protein load and
neutralising antibody formation.²⁴ 

The study concluded that to reduce the risk of antibodies forming, a
toxin with the lowest amount of complex proteins should be used.²⁵
Research shows that in addition to selecting a low-risk antigen to
reduce the risk of antibodies developing, we should wait at least three
months in-between treatments and use the lowest most effective
dose.^8,26 There have been multiple studies that have shown higher
rates of resistance linked to treatments where patients have had
higher therapeutic doses, such as in ²² patients with cervical dystonia
or oromandibular dystonia who developed resistance and were found
to have NAPS. This showed that the mean dose per visit and the dose
of BoNT-A were significantly larger than those without NAPS, showing
a link between developing NAPS and high doses of BoNT.¹⁹ Another
study involving 616 patients receiving BoNT-A for cervical dystonia
of which nine patients had NAPS, showed a link between those
receiving higher doses at shorter treatment intervals. It also showed
that patients who were reinjected within six weeks were more likely to
develop NAPS.²⁰ In cosmetic use this could validate why the evidence
shows that resistance to botulinum toxin is less common in patients
who receive BoNT-A at lower doses. 

A recent systematic review was carried out using 43 studies, including
8,833 patients being treated with all available toxins on the UK market.
The research showed that the number of patients with NAPS
was 1.8% and there was a slight increase related to the duration the
patient had been receiving BoNT-A. Those being treated for medical
conditions using high doses was shown with the highest incidence of
NAPS. Patients being treated with Abobotulinumtoxin-A were shown
to have the most incidence of NAPs (7.4%) whilst Incobotulinumtoxin-A
and Onabotulinumtoxin-A were shown to have a rate of 0.3%. The
study concluded that whilst NAPS following BoNT-A is relatively low,
there is limited evidence on what can cause a secondary response.²⁷

Solutions to toxin resistance 

With the risk of BoNT-A resistance, albeit small, it’s important to know
what alternative treatments we can offer to our patients should the
problem arise. Currently there is limited evidence for determining if
antibodies disappear over time and whether as medical practitioners
we should reinject or not. According to the literature, it is however
important that the quality of the botulinum toxin used is neurotoxin free
of complexing proteins with low antigenicity, to prevent a resistance
happening again.²⁸ 

Although there are no current guidelines, it is recommended in the
literature that patients who develop immunoresistance to BoNT-A
can be treated with higher doses or by trying different brands,
although there is debate around whether complex proteins are a
help or a hindrance for the BoNT-A molecule.⁹ If there are no changes
after trying an alternative brand, another alternative could be to try
botulinum toxin type B (BoNT-B). The only commercially available
BoNT-B drug available in the UK is RimabotulinumtoxinB (Neurobloc),
but is indicated only for the treatment of cervical dystonia (torticollis) in
adults.¹¹ In terms of secondary resistance, a recent study containing 36
patients who had cervical dystonia (involuntary muscle contractions in
the neck) and a secondary non-response to BoNT-A showed that 36%
of the trial patients had a good clinical response to Neurobloc. The
other 23 patients either had no response, a poor response, or side
effects and stopped treatment. Another study containing 20 patients
that were resistant to BoNT-A showed that seven of these patients
showed some response to type B.²⁹ These findings show that BoNT-B
may have a place as an alternative treatment for patients who have
have become resistant to type A, but overall the research shows that
BoNT-B has no guarantee for an effective treatment.²⁹ 

There is some guidance from the British Neurotoxin Network30
in 2016, which recommends distinguishing no response to the
toxin and then considering a dose revision. Where resistance
is identified it recommends a switch to BoNT-B or a treatment
break. This guidance is not specific to cosmetic treatments or
primary causes.30 Another paper discusses the association between
secondary botulinum toxin treatment failure in cosmetic indication and
anti-complexing protein antibody production, and proposes a protocol
for the treatment of patients who are suspected to have resistance to
BoNT-A.³¹ The protocol recommends that instead of switching brands,
all practitioners should stop injecting BoNT-A for at least six months to
prevent further antibody production or immune responses.³¹
Practitioners may also use other aesthetic treatments such as
cosmeceuticals to help prevent wrinkles and skin tightening
treatments using energy-based devices.  

Conclusion 

From the current literature it is easy to see that there is limited
evidence to support how common resistance to BoNT-A is and why it
happens. However, based on the research available there is a small
amount of evidence that shows resistance may be linked to complex
proteins. Until there is more clinical evidence it is wise to follow best
treatment guidelines by using the lowest most effective dose and to
have intervals of at least ¹² weeks. In future, we need more long-term
clinical studies to follow up patients and to understand the causes of
botulinum toxin resistance and how we can treat them.