A e s t h e t i c U s e s of t h e

B otulinum Toxin
Andrew Dorizas, MDa,*, Nils Krueger, PhDa,
Neil S. Sadick, MDa,b

KEYWORDS
 Botulinum neurotoxin  Fillers  Perioral rhytides  Facial rhytides  Glabellar lines  BoNTA

KEY POINTS
 The cosmetic use of botulinum toxin (BoNT) is the most common cosmetic procedure performed in
the world today.
 Thus far, the immunogenicity rates of onabotulinumtoxinA (BoNTA-ona) and incobotulinumtoxinA
(BoNTA-inco) have been approximately 1%, whereas the immunogenicity rate for abobotulinum-
toxinA (BoNTA-abo) is slightly higher, at 3%.
 Conversion ratios of 2.5:1.0 for BoNTA-abo: BoNTA-ona and 1:1 for BoNTA-inco: BoNTA-ona are
widely accepted in the field.
 Common adverse events seen in the aesthetic use of the BoNT include swelling, localized bruising,
headaches, injection site discomfort, excessive muscle weakness, and unintended paresis of adja-
cent muscles.
 BoNT has a wide array of cosmetic uses, including treatment of glabellar lines, chemical browlift,
forehead wrinkles, periorbital, and perioral lines.
 The future formulations and applications of BoNTA will be plentiful, and are exciting to consider.

BACKGROUND agent cannot be equivalent and, that differences

and similarities exist between them. All products
Aesthetic injection botulinum toxin (BoNT) is the are synthesized by the same strain of anaerobic
most common cosmetic procedure performed in Clostridium botulinum, a gram-positive, rod-
the world today.1 BoNT has proven to be one of shaped anaerobic bacterium. The therapeutic
the most versatile agents used in the aesthetic preparations of botulinum neurotoxin however,
field, and has also been used for numerous medi- vary in molecular weight, complexing proteins
cal applications, including arm and leg spasticity, and excipients.
blepharospasm, cervical dystonia, strabismus, se-
vere hyperhidrosis, chronic migraine, and overac- HISTORY OF BONT
tive bladder, to name a few.2
Although the only U.S. Food and Drug Adminis- C botulinum was discovered by professor of bacte-
tration (FDA)–approved cosmetic indication for riology, Emile Pierre van Ermengem, in 1895, after
BoNT is the treatment of dynamic rhytides in the a botulism outbreak in a Belgian village.2 The toxin
glabellar area, successful off-label uses of the itself, however, was not isolated until the 1920s.1
products have been vast. Currently, the aesthetic Purification of the toxin was attempted by Dr Her-
market has multiple formulations available, and man Sommer and colleagues at the University of
with the expected availability of more in the near California, San Francisco, but these attempts
future, it is important to understand that each were not successful until Dr Edward Schantz
derm.theclinics.com

a
Sadick Research Group, 911 Park Avenue, New York, NY 10075, USA; b Department of Dermatology, Weill
Medical College of Cornell University, 1300 York Avenue, New York, NY 10065, USA
* Corresponding author.
E-mail address: [email protected]

Dermatol Clin 32 (2014) 23–36

http://dx.doi.org/10.1016/j.det.2013.09.009
0733-8635/14/$ – see front matter Ó 2014 Elsevier Inc. All rights reserved.
24 Dorizas et al

isolated a purified BoNT type A (BoNTA) in a crys- BoNT consists of 2 protein chains: a heavy chain
talline form in the 1940s. (100 kDa) and a light chain (50 kDa), connected by
Approximately 30 years later, Dr Alan Scott, a a single disulfide bridge and noncovalent bonds.
surgeon at the Kettlewell Eye Research Institute The heavy chain is made up of the binding and
in San Francisco, began testing BoNTA in mon- translocation domains. The light chain is the actual
keys as a potential therapy for strabismus. After toxin itself. The binding domain attaches to the re-
successful treatments on monkeys, Dr Scott per- ceptor on the neuronal end plate. The transloca-
formed the first large-scale clinical trial using ona- tion domain causes endocytosis of the BoNT into
botulinumtoxinA (BoNTA-Ona) (Oculinum) to treat the cytoplasm of the nerve. Simply, BoNT exerts
strabismus.2–5 It was during this time that Dr Jean its effects by inhibiting acetylcholine (Ach) release
Carruthers noticed what was to become the at the neuromuscular junction and autonomic
future of BoNTA for aesthetic use: diminishing of nerve terminals. The effects of BoNT are nonper-
wrinkles in the glabellar area. After Dr Scott’s suc- manent. All formulations of BoNT inhibit exocy-
cessful use of Oculinum, Allergan licensed his tosis and Ach release without affecting Ach
technology, named their product Botox, and production or conduction of signaling along the
received FDA approvals for both therapeutic nerve fiber.7
and cosmetic indications, including strabismus The process through which BoNT exerts its
and blepharospasm in 1989, cervical dystonia in effect is dependent on a series of steps that be-
2000, glabellar lines in 2002, axillary hyperhidro- gins with uptake of the toxin into the nerve termi-
sis in 2004, upper limb spasticity and chronic nal. BoNTA-ona, BoNTA-inco, and BoNTA-abo
migraines in 2010, and urinary incontinence in bind to the same cellular receptor synaptic vesicle
2011.2,3 BoNT was the first microbial protein protein 2 (SV-2), whereas BoNTB-rima binds to
injection used to treat human disease.6 In 1991, synaptotagmin II (Syt II) with great selectivity for
Speywood (now called Ispen) launched another these glycoproteins at the neuromuscular junc-
BoNTA formulation based on abobotulinumtoxinA tion. Once internalized in the cell, the light chain
(BoNTA-abo), Dysport, for the treatment of dissociates from the heavy chain and cleaves
blepharospasm, hemifacial spasm, and focal one or more of the soluble N-ethylmaleimide–sen-
spasticity. Dysport was then approved for the sitive factor attachment receptors (SNAREs),
treatment of glabellar lines in 2009. In 2000, which are part of the Ach transportation cascade
Elan (now Solstice Neurosciences) brought its ri- and are required for docking of vesicles with the
mabotulinumtoxinB (BoNTB-rima)-based BoNT neural cell membrane in the release of Ach from
type B product Myobloc to market for the treat- the nerve terminal.7 The light chain of each sero-
ment of cervical dystonia, currently its only type of BoNT cleaves a different peptide bond,
FDA-approved use. More recently, Merz Pharma- and no 2 serotypes do so at the same site. The
ceuticals received regulatory approval for Xeo- substrate for BoNTA is a 25-kDa synaptosomal-
min, a product based on incobotulinumtoxinA associated protein (SNAP-25). BoNTB targets
(BoNTA-inco) for the treatment of focal dystonias, and cleaves vesicle-associated membrane pro-
spasticity, and glabellar lines. teins (synaptobrevins).9
Once the neuromuscular junction has been
STRUCTURE AND MECHANISM OF ACTION blocked by BoNT, binding sites for the toxin are
decreased. As a result, repeated injection into an
Seven serologically distinct BoNTs (BoNT types A, already denervated muscle is not as effective,
B, C, D, E, F, and G) are produced by various because uptake into muscle is reduced. The
strains of C botulinum, C butyricum, and C baratii neurotoxic effect of the neuromuscular junction is
in nature. Only serotypes A and B are used thera- temporary. After injection, the effect is evident
peutically, with type A being the most neurotoxic.7 within 1 to 3 days. Maximal effect is seen approx-
The nontoxic accessory proteins are composed of imately 2 weeks after the toxin is introduced, and
hemagglutinin and nontoxin, nonhemagglutinin then gradually starts to decline after approximately
proteins, and spontaneously associate with the 3 months. Restoration of the neuromuscular junc-
core neurotoxin after their cosynthesis by the tion is seen at 2 levels.10 Axonal sprouting is a
bacteria.8 temporary recovery process, wherein new neuro-
The size of the neurotoxin complexes is deter- muscular junctions develop around the blocked
mined by serotype and accessory protein content. terminal. Once the original neuromuscular junction
For example, type A clostridial strains produce is restored, the sprouts are removed.
complex sizes of 300, 500, and 900 kDa, whereas This inhibitory mechanism of action is also used
type B strains produce complexes of only 300 and in the treatment of hyperhidrosis. In addition to
500 kDa.9 their effect on the neuromuscular junction, BoNTs
 Aesthetic Uses of Botulinum Toxin 25

have the ability to act on the innervated eccrine formulation of 300- and 500-kDa complexes,
secretory cells. Through inhibiting the sympathetic whereas BoNTB-rima manufacturing produces a
fibers to sweat glands, where Ach is used as the homogenous mixture of complexes. The produc-
neurotransmitter, BoNTs have been shown to tion of BoNTA-inco is distinctive, because the
effectively reduce sweat production.11 complexing proteins are separated and discarded
Other indications for the use of BoNT are based from the neurotoxin during the manufacturing pro-
on the fact that sensory nerves release mediators cess in several purification steps. The resulting
of pain and inflammation, including substance P pure neurotoxin has a molecular weight of only
and cGRP (calcitonin gene-related peptide). Vesi- 150 kDa. The excipient components of the vials,
cles located at sensory nerve terminals also intentionally added substances that do not exert
contain SNARE proteins that facilitate the release a therapeutic effect but may affect product deliv-
of these substances. In studies conducted using ery, vary among the formulations.13
animals, BoNTA-ona has been shown to reduce All the products contain human serum albumin.
cGRP. It has been suggested that blocking Human serum albumin acts as a stabilizer of
cGRP, glutamine, and substance P inhibits neuro- BoNT. As discussed by Pickett and Perrew,13 the
genic inflammation and can ameliorate pain syn- toxin is protected by the accessory proteins,
dromes, such as migraine headache or psoriatic which must dissociate for the core toxin to act.
skin lesions.9,12 This dissociation occurs under physiologic condi-
tions, including when the toxins are diluted with
NONINTERCHANGEABILITY OF DIFFERENT saline. Thus, the core toxin may be dissociated
FORMULATIONS from the accessory proteins before injection. The
amount of human serum albumin differs between
All BoNTA products cannot be generics, and each the formulations.
has a unique manufacturing process. BoNTA-ona, The pH of the formulations depends on the pH of
BoNTA-inco and BoNTA-abo are derived from a the preserved saline used and the ability of the ex-
strain of C botulinum type A strain called Hall cipients to buffer the solution. The number of units
strain. BoNTB-rima is derived from a type B strain and the amount of core neurotoxin differs between
called Bean strain. BoNTA-ona is produced using the formulations. BoNTA-ona and BoNTA-inco are
a crystallization process that yields a homogenous distributed in 100-U vials, BoNTA-abo in 300- and
formulation of 900-kDa complexes, whereas 500-U vials and BoNTB-rima in 2500-, 5000-, and
BoNTA-abo and BoNTB-rima are both manufac- 10,000-U vials. Therefore, the amount of neuro-
tured with column chromatography. The process toxin injected depends on the dose of the product
of BoNTA-abo production yields a heterogeneous used (Table 1).

Table 1
Summary of commercially available botulinum toxin formulations

Botox Dysport Xeomin Myobloc

First approval 1989 1991 2011 2000
Serotype A A A B
Strain Hall Hall Hall Bean
Receptor/target SV2/SNAP-25 SV2/SNAP-25 SV2/SNAP-25 Syt II/VAMP
Process Crystallization Chromatography Chromatography Chromatography
Complex size (kDa) 900 300 150 700
Uniformity Homogeneous Heterogeneous Free neurotoxin Homogeneous
Excipients HSA HSA HSA HSA
Sodium chloride Lactose Sucrose Sodium chloride
Sodium succinate
Stabilization Vacuum drying Lyophilization Vacuum drying Solution
Solubilization Normal saline Normal saline Normal saline N/A
Units per vial 100 300, 500 100 2500, 5000, 10,000

Abbreviations: HSA, human serum albumin; N/A, not applicable; SNAP-25, 25-kDa synaptosomal-associated protein; SV2,
synaptic vesicle protein 2; Syt II, synaptotagmin II; VAMP, vesicle-associated membrane protein.
Data from Pickett A, Perrow K. Formulation composition of botulinum toxins in clinical use. J Drugs Dermatol
2010;9(9):1081–4; and Xeomin [package insert]. Greensboro (NC): Merz Pharmaceuticals, LLC; 2013.
26 Dorizas et al

As a result of the differences discussed, among with BoNTA-inco or BoNTA-abo. Overall, BoNTA
others, it has been mandated that the product la- products exhibit low clinically detectable levels of
beling include specific language that states that antibodies when compared with other approved
the unit and dose of each product are not inter- biologic products, especially when used in low
changeable and cannot be simply converted be- doses for aesthetic indications.16
tween products.2,14,15
CONTRAINDICATIONS AND ADVERSE EVENTS
IMMUNOGENICITY
Although generally treatments with BoNTA are
Because BoNT is a protein-based drug that is considered safe, practitioners must be aware of
administered repeatedly for long-term effect, it is the contraindications and adverse events and
capable of triggering immunologic reactions and take the appropriate measures to minimize the
antibody formation. Various factors have an risk. BoNT is contraindicated in any patient with
impact on its immunogenicity, such as prior expo- known hypersensitivity to BoNT or any compo-
sure, manufacturing process, antigenic protein nent of the formulation and in patients who have
load, overall toxin dose administered, and pres- an active infection at the site to be in-
ence of accessory proteins.16,17 jected.14,15,24 Relative contraindications are in
Immunity in patients treated for indications patients who are pregnant or nursing and anyone
requiring high doses, such as cervical dystonia, with a preexisting neuromuscular disease, such
was as high as 20%.18 In 1998 the manufacturer as myasthenia gravis and Eaton-Lambert syn-
improved the purity of the BoNTA-ona formulation drome. Although BoNT is classified as pregnancy
to decrease the amount of protein accompanying category C in the United States, no teratogenicity
the toxin. Through decreasing the antigenic expo- has been reported as a result of women who
sure, the immunity rates with repeated injections inadvertently received up to 300 U of BoNTA dur-
for cervical dystonia decreased and the purifica- ing pregnancy.21
tion did not alter dosing requirements.10 Common adverse events seen in the aesthetic
Because the protein content of BoNTA-inco is use of the toxin include swelling, localized
less than that of BoNTA-ona when comparing a bruising, headaches, injection site discomfort,
1-to-1 U dose equivalence, BoNTA-inco is ex- excessive muscle weakness, and unintended
pected to have a lower risk of immunogenicity. paresis of adjacent muscles.14,15,24 Most of these
Although BoNTA-abo has a 2.5:1.0 equivalence adverse events have been comparable to those
ratio versus BoNTA-ona, the protein load is seen with placebo, as shown in several
considerably smaller. Therefore, BoNTA-abo studies.25–27 Avoiding anticoagulants or other
would also have a lower immunogenicity.19,20 blood-thinning supplements 2 weeks before the
However, currently no studies have directly treatment can reduce the risk of bruising. Injection
compared antigenicity rates between currently site discomfort can be avoided by using small-
available products. Factors that are thought to in- gauge needles and careful technique. If neces-
crease the incidence of antigenicity include sary, a topical anesthetic can be used or ice can
increased treatment number and cumulative be applied before the treatment. To avoid unin-
dose, frequent exposure, and genetic predisposi- tended muscle weakness, therapy should be initi-
tion.21–23 With these risk factors in mind, and given ated with lower doses and titrated upwards to
the increased doses used with BoNTA-abo and achieve the desired effect. Lastly, using diluents
the shorter dosing interval, one might expect containing preservatives has been shown to
higher rates of antigenicity because of greater reduce patient discomfort during the injection.28
exposure to the toxin than when using BoNTA- BoNT is among the most potent toxins in exis-
ona. However, thus far, the immunogenicity rates tence. Safety measures should be known and
of BoNTA-ona and BoNTA-inco have been around adhered to by all physicians, regardless of their
1%, whereas the immunogenicity rate for BoNTA- experience. Proper patient selection and evalua-
abo is slightly higher at 3%.16 BoNTA-inco has tion are crucial, along with precise injection tech-
also been shown to induce an immunogenic nique. In most cases, adhering to the safety
response, which was established during the clin- measures will avoid systemic exposure of the
ical trials performed in the United States. Twelve toxin.
subjects developed neutralizing antibodies during
the course of their trials.19 The purification process HANDLING AND STORAGE
for each BoNTA is different, and therefore experi-
ence with BoNTA-ona and immunogenicity cannot Studies have shown that BoNT formulations are not
be used to estimate the risk of immunogenicity as fragile as suggested in the package instructions.
 Aesthetic Uses of Botulinum Toxin 27

Prescribing information for BoNTA-ona, BoNTA- 109 days.26 One study showed a difference in
abo, BoNTA-inco, and BoNTB-rima state that response rate at 16 weeks between 2 different for-
each vial is for single use. BoNTB-rima is available mulations; the relapse rate was 23% for BoNTA-
in a ready-to-use form with no reconstitution neces- ona and 40% for BoNTA-abo.35 BoNTA-inco has
sary. Handling instructions state that vials should been shown to have similar efficacy duration as
be refrigerated and used within 4 hours of reconsti- BoNTA-ona, although more clinical studies are
tution in the case of BoNTA-abo and BoNTB-rima, warranted for further comparison.36,37
and within 24 hours of reconstitution in the case A recent study of 45 patients using both patient
of BoNTA-ona and BoNTA-inco. BoNTA-inco, how- and physician assessments of response after
ever, does not require refrigeration. Exposure to treatment of glabellar rhytides examined the
direct sunlight is thought to inactivate the toxin onset of BoNTA-ona. Results showed that the
within 1 to 3 hours, and temperatures of 80 C to mean time to response after treatment with a
100 C denature the toxin in approximately 30 and 20-U dose of BoNTA-ona was less than 2 days.
10 minutes, respectively.14,15,24 In this study 48% of patients had onset on
Hexsel and colleagues29 published a study that 1 day after injection and 87% of patients experi-
showed that the efficacy of BoNT was not enced a response by day 2.38 Studies analyzing
compromised for up to 6 weeks after reconstitu- the onset of BoNTA-abo indicated that the mean
tion. Similarly, a different study showed no onset time was 3 days, with effects seen in as
contrast in the efficacy of freshly reconstituted early as 24 hours.39
BoNTA and BoNTA-ona that was reconstituted BoNTB-rima has been associated with a faster
2 weeks before being injected and then stored in onset but a shorter duration of action, with relapse
either a freezer or a refrigerator. This study, and times between 2 and 3 months.40,41
also one conducted by Sloop and colleagues,30
showed that proper storage could be in either a CONVERSION RATIOS
freezer or a refrigerator.31
A recent animal study examined the effect of Although different formulations cannot be directly
sustained agitation of BoNTA-ona for 6 weeks af- compared, knowing the approximate dose con-
ter reconstitution. The agitated BoNTA-ona was versions between BoNT products is essential to
subsequently injected into mice to determine if it incorporate them into practice, and for studies
was still lethal (LD50). The study found no loss of that seek to accurately compare these products.
efficacy in the vigorously agitated BoNTA-ona.32 The conversion ratio of 2.5:1.0 (BoNTA-abo:-
Additional studies have compared the efficacy of BoNTA-ona) is widely accepted in the field. In
gently and vigorously reconstituted BoNTA-ona an editorial commentary on the use of the 2 for-
used for the treatment of rhytides in human sub- mulations for cervical dystonia, Poewe42 stated
jects. These studies concluded that no loss of ef- that, based on clinical studies, the ratio of
fect occurred with vigorous reconstitution.33 BoNTA-abo to BoNTA-ona should be no greater
than 3.0:1.0. A study for the cosmetic use of
ONSET AND DURATION BoNT used electromyographic activity to deter-
mine more precisely the dose conversion. This
Awareness with respect to expected time to study enrolled 26 patients, each of whom was
relapse of a treated area after BoNT injections is randomly assigned to receive BoNTA-abo and
vital in scheduling the subsequent appointment. BoNTA-ona at a 3.0:1.0 ratio to each half of the
It is generally accepted that the average duration frontalis muscle at 3 points of injection. Electro-
of effect is between 3 and 4 months. A study con- myographic activity was then measured at inter-
ducted by Carruthers and colleagues34 evaluated vals as long as 20 weeks after the first
the efficacy of BoNTA-ona for the treatment of injections. Results showed that at a 3.0:1.0 con-
glabellar lines. The investigators determined that version ratio, the BoNTA-abo had a longer dura-
the rate of response at maximal frown lasted tion of effect. These results were interpreted as
3 months for most patients, and as long as an indication that the true conversion ratio must
4 months in 25% of the patients, showing that be less than 3.0:1.0, because an accurate conver-
3- to 4-month intervals between injections is sion ratio would have both formulations with
appropriate. Likewise, a study using BoNTA-abo equal duration of effect.43 Lowe and colleagues35
with a 50-U dose versus placebo determined compared the duration of effect between BoNTA-
that the optimal timing for a second injection was abo and BoNTA-ona in the glabella using a dose
between 3 and 4 months. Another study with ratio of 2.5:1.0. The study showed a greater dura-
more than 700 subjects treated with BoNTA-abo tion of effect in patients treated with the 20-U
found a median duration of effect of 85 to dose of BoNTA-ona.
28 Dorizas et al

DIFFUSION
The toxin’s ability to remain relatively localized at
the site of injection is essential for the safety of
BoNT treatments. Uncontrolled spread, diffusion,
or migration of the toxin not only increases the
risk of distal and systemic effects but also makes
the aesthetic outcome unpredictable for the treat-
ing physician.
An often-cited study by Trinidade de Almeida
and colleagues44 compared the area of anhidrosis
of each side of the forehead after injection of
BoNTA-ona and BoNTA-abo to contrast their
diffusion properties. Either side of each subject’s
forehead was randomized to receive either
BoNTA-ona or BoNTA-abo at 2 injection points
(medial and lateral). The side of the forehead ran-
domized to BoNTA-ona received 3 U at each injec-
tion point. The alternate side was then randomized Fig. 1. BoNT injection points.
to receive an equal volume of injection of BoNTA-
abo but with a dose ratio of 1.0:2.5, 1.0:3.0, or
1.0:4.0. An iodine and starch test across the fore- Carruthers50 tested the safety, efficacy, and dura-
head was used to highlight the areas of anhidrosis. tion of response of 4 doses of BoNTA-ona on
The halos of anhidrosis could then be measured glabellar rhytides in men. The subjects were ran-
and compared. The important factor in this study domized to receive a total dose of 20, 40, 60, or
is that the injections were kept at constant volume 80 U of BoNTA-ona in the glabellar area. Seven in-
regardless of the dose ratio. The anhidrotic halo for jections points were used and the units were
BoNTA-abo was found to be larger at all dose ra- distributed evenly among the points. Results
tios from week 1 to month 6 compared with the ha- showed that increasing doses had more efficacy
los for BoNTA-ona. in reducing glabellar lines. They concluded that
Studies comparing BoNTA-ona and BoNTA- men with glabellar rhytides benefit from a starting
inco showed that the complex proteins in dose of at least 40 U.49
BoNTA-ona do not influence the diffusion pro- In a similar study, Carruthers28 examined the ef-
file.45,46 BoNTB consistently showed the highest fect of 4 doses of BoNTA-ona in female subjects.
local and systemic diffusion properties. The B In addition, an open-label arm of the trial evaluated
serotype not only produced a greater radius of the use of a 30-U dose in all subjects. Subjects
toxin diffusion but also had a significantly higher were randomly administered 10, 20, 30, or 40 U.
number of associated side effects.47,48 Brodsky The investigators concluded that 20 U was an
and colleagues49 summarize that neither molecu- optimal dose for treating glabellar lines in woman,
lar weight nor the presence of complexing proteins but that higher doses were not associated with an
seem to affect diffusion, but properties intrinsic to increase in adverse events and therefore can be
the drug, accurate muscle selection, and dilution, administered if necessary.28
volume, and dose injected are factors that influ- Frampton and Easthope51 compared BoNTA-
ence diffusion.50 ona with placebo in 2 large clinical trials in 537
subjects with moderate to severe glabellar lines.
Investigators and subjects both saw a clear
COSMETIC USES
improvement in the treatment group versus the
Glabellar Lines
control group throughout all the study visits. The
Treatment of the glabellar remains the only FDA- efficacy of BoNTA-ona has been concluded in
approved cosmetic use for BoNT. This treatment many publications.36,52,53
is by far the most common cosmetic use of In a study54 comparing the effects of BoNTA-
BoNTA, and its efficacy has been shown in ona versus BoNTA-inco in the treatment of
numerous clinical trials. The target site for the glabellar frown lines, response rates were similar
treatment is the procerus muscle and the corruga- for both formulations at the same dose (1 U).
tor supercilii muscle (Fig. 1). These muscles are Investigator assessment and high patient satis-
responsible for depressing the medial brow. In faction rates confirmed high treatment success
an early dose-ranging study, Carruthers and for both formulations at the same dose. Both
 Aesthetic Uses of Botulinum Toxin 29

formulations of the toxin (BoNTA-ona and BoNTA- In clinical practice the chemical browlift is more
inco) are generally administered at comparable commonly performed in combination with other
doses for the same indications.54 BoNT treatments for other target areas. When tar-
International consensus recommendation for geting the orbicularis oculi together with the
the treatment of glabellar lines with BoNTA-abo glabella complex, a natural looking and aestheti-
is 5 injection points, with 10 U injected per site cally pleasing result can be achieved.
(Fig. 2).55 In addition to dynamic lines, lines pre-
sent at rest may soften or be eliminated with
Forehead Wrinkles
BoNT to the glabellar region.56
Horizontal forehead lines occur when the frontalis
Chemical Browlift muscle contracts to raise the eyebrows upward.
The frontalis muscle has a rear belly and an ante-
BoNT injections can lead to a desirable elevation in rior belly. The rear belly is continuous with and
the resting position of the eyebrow. This technique transitions to become the aponeurosis of the scalp
will give the eye a more open appearance and a via the tendinous galea aponeurotica. This transi-
youthful look. Through paralyzing the lateral brow tion point varies from person to person. Contrac-
depressor (orbicularis oculi), the authors allow tion of the rear belly does not cause a significant
the frontalis muscle to elevate the brow without effect on the anterior belly of the frontalis muscle,
opposition. Treating the glabellar complex leads and thus injections in the aponeurotic area have no
to a medial brow lift, and a lateral brow lift has clinical significance.62
also been shown.57 Carruthers and colleagues63 determined that
Frankel and Kamer58 first noted the medial BoNTA-ona can be used safely and effectively to
elevation and increase in interbrow distance with treat forehead wrinkles. They found that the effi-
the use of BoNTA-ona for the treatment of cacy improves with higher doses (24 U) injected
glabellar lines. In a study of BoNTA-ona injected into the frontalis compared with lower doses
in the procerus and corrugator muscles in 30 sub- (16 and 8 U). The consensus recommendations
jects, they found that 32% of patients had a raise regarding use of BoNTA-ona in the upper face
in medial eyebrow position, 48% had a higher advise a range of 6 to 15 U in women. In men,
eyebrow position in the midpupillary line, and the recommended dose is 6 to 15 U. The usual
59% had an increase in the interbrow distance. number of injections points is between 4 and 8.
Chen and Frankel59 discussed the various International consensus recommends doses
manipulations of the eyebrow position that can of 5 to 10 U of BoNTA-abo at 4 to 6 injection
be achieved with BoNT. They note that slowly in- points for forehead lines, up to a total of 20 to
jecting BoNTA-ona at doses up to 10 U above 60 U.55
the orbital rim may help avoid affecting the levator In a case series reporting on a man and a
palpebrae muscle and causing an undesirable lid woman with forehead, glabellar, and periorbital
ptosis. lines injected with BoNT at 4-month intervals
Maas and Kim60 examined the effect of 5 to 10 U over a 7-year period64 showed continued improve-
of BoNTA-ona or BoNTA-abo in 22 subjects in- ment in skin smoothening and effacement of
jected into the lateral portion of the orbicularis nonreducible horizontal forehead lines. The inves-
oculi. Their study, and others, show statistical sig- tigators concluded that this showed evidence of
nificant increases in the lateral brow height.57,61 ongoing dermal and epidermal remodeling, and
that forehead lines can show long-term reduction
when treated with BoNTA-ona injections. An
assessment of wrinkle depth using silicone masks
and multiphoton microscopy 3 weeks after injec-
tion with BoNTA-ona showed an objective
improvement compared with baseline.65
When treating horizontal forehead lines, injec-
tions must be placed 1 to 2 cm above the orbital
border to avoid eyebrow ptosis. The average
target sites for forehead lines are between 5 and
10, depending on the patient. The points are
generally in a V-shape or 2 horizontal lines approx-
imately 1 cm apart. The lateral forehead should
also be treated to avoid excessively arched eye-
Fig. 2. Injection points for glabellar lines. brows (“Spock” eyebrow) (Fig. 3).66
30 Dorizas et al

and colleagues69 showed that wrinkles in the peri-

orbital area were significantly improved for up to
6 months after injection of the crow’s feet with
12 U of BoNTA-ona on each side.
Adverse events that may occur with treatment of
the crow’s feet include periorbital hematoma, dry
eyes, eyelid ptosis, and mouth droop. The zygo-
maticus major muscle is located nearby and, as
a precaution, the injector should never inject
beneath the zygoma to avoid paralyzing it. Using
the minimal injection dose and correcting injection
target sites are essential to avoid and adverse
Fig. 3. Injection points for forehead wrinkles.
events.70

Bunny Lines
Periorbital Lines
Bunny lines are multiple dynamic rhytides located
Periorbital lines or crow’s feet are among the most on the upper nasal dorsum that result from the
popular treatment sites for BoNT. These dynamic contraction of the transverse nasalis muscle with
lateral canthal lines appear when smiling, and are smiling. These lines can become more prominent
not to be confused with the static lateral canthal after BoNT treatment in the glabella and periorbital
lines that are caused by photoaging. Static lines region. A single injection of 2 to 4 U of BoNTA-ona
are more resistant to BoNT treatments and require or 5 to 10 U of BoNTA-abo should be injected on
additional procedure to be corrected, including each side about 1 cm superior to the alar grove.71
laser resurfacing. Injecting into the levator labii superioris or alaeque
Consensus data, clinical studies, and experi- nasi can lead to upper lip ptosis (Fig. 5).
ence support that the usual number of injection
points for crow’s feet is 2 to 5 (average of 3) per Gummy Smile
side, and that dosing of BoNTA-ona and BoNTA-
inco ranges from 10 to 30 U for women and 20 An excessive gingival display or gummy smile is
to 30 U for men (Fig. 4). For BoNTA-abo, the rec- defined as gingival exposure greater than 2.0 mm
ommended dosing is 20 to 60 U divided among above the dental line with smiling. This condition
3 injection points per side.63 The injections should may be considered aesthetically unappealing and
be at least 1.0 to 1.5 cm from the external orbital is more common in women. In a study involving
rim to avoid any paralysis of the palpebral portion 52 subjects, Sucupira and Abramovitz72 injected
of the orbicularis oculi muscle.67
A study of 162 patients evaluated the treatment
of crow’s feet with doses of 3, 6, 12, and 18 U. The
investigators found that the optimal dose was 12 U
because it was more effective than the smaller
doses, but they noted no significant difference be-
tween the 12- and 18-U doses.68 A study by Levy

Fig. 4. Injection points for crow’s feet. Fig. 5. Injection points for bunny lines.
 Aesthetic Uses of Botulinum Toxin 31

BoNTA-ona to the levator labii superioris alaeque drooling), administering the minimum dose is rec-
nasi muscle. The average dose was 1.95 U per ommended (0.5 U of BoNTA-ona and BoNTA-inco
side. The average gingival display was 3.62 mm and 1.0 U of BoNTA-abo) and then increased on a
before treatment, and 0.58 mm after treatment. case-by-case basis.
The results showed that injecting just the levator Additionally, other treatment modalities, in-
labii superioris can be effective in correcting exces- cluding laser resurfacing and injectable fillers,
sive gingival display. may be required for full correction. This location
Polo73 documented the treatment of 12 women should not be injected in patients who use their
with gummy smile. Each elevator muscle was in- mouth to play musical instruments, because these
jected with variable dosing in 3 phases. He used patients rely on the full function of the orbicularis
3 injection points on each side, injecting 2.5 U of oris.2 Consensus guidelines on the topic of perioral
BoNTA-ona into the levator labii superioris, the rhytides include similar recommendations, in-
junction of the levator labii superioris, and the cluding using low doses and avoiding central and
zygomaticus minor, and 1.5 U into the orbicularis lateral injections points that may flatten the cupid’s
oris at a point 2 to 3 mm below the nares and bow, and lead to drooling or drooping, respec-
2 to 3 mm away from midline. tively.77 These guidelines state that it is safer to
In a study of 16 patients, Mazzuco and Hexsel74 keep injections within 5 mm of the vermillion
categorized the type of gummy visibility present, border. Ice may be used as an anesthetic before
and then based the treatment on the muscles injections in this more sensitive area.
that were responsible for each type. Subjects
were categorized into 4 groups: anterior gummy Dimpled Chin
show defined as greater than 3 mm of gum expo-
The dimpling of the chin, or peau d’orange (orange
sure in the area between the canine teeth; poste-
peel) appearance, is caused by a combination of
rior gummy show defined as normal anterior
loss of volume and contraction of the mentalis
exposure but posterior to the canines; greater
muscle.78 A combination of BoNT and dermal
than 3 mm of gingival display mixed with gum
fillers is recommended to achieve optimal results.
exposure both anteriorly and posteriorly; and
Consensus recommendations state that generally
lastly, asymmetric gummy show. Subjects with
1 injection point per side is needed just below
anterior gummy show were treated with 5.0 U of
the tip of the chin close to the mandibular
BoNTA-abo into the levator labii superioris alae-
bone.68 A typical dose is 5 to 10 U of BoNTA-
que nasi bilaterally, whereas subjects with poste-
ona and BoNTA-inco and 10 to 20 U of BoNTA-
rior gummy smile were given injections of 2.5 U
abo; however, men may require higher doses.
into the zygomaticus major and 2.5 U into the zy-
Lowe and colleagues68 recommended careful
gomaticus minor. Subjects in the mixed group
avoidance of an injection point that is too close
were treated in all 3 areas, and subjects in the
to the lips, which could lead to loss of function of
asymmetric group were treated according to the
the lip depressors. Furthermore, when treating
asymmetry. These authors supported the principle
the dimpled chin along with the oral commissures,
that lower dilution volumes should be used for this
the dose must be decreased accordingly to avoid
indication given the proximity to important func-
overtreatment.63
tional muscles. Suggested doses for BoNTA-ona,
BoNTA-inco are 2.0 to 5.0 U and 5.0 to 15.0 U
Marionette Lines
for BoNTA-abo.
The marionette lines, or melomental folds, are also
best treated with a combination approach of BoNT
Perioral Lines
and dermal fillers.
Perioral lines, or lipstick lines, are vertical rhytides The depressor anguli oris contributes to
projecting in a perpendicular fashion from the depressing the mouth and draw it laterally, causing
vermillion border of the lips. Movements of the or- a sad face appearance. A recent retrospective
bicularis oris muscle, such as puckering, and analysis of 60 patients treated with BoNT found
smoking, photoaging, and hereditary factors that 10% to 20% received injections into the
contribute to the formation of these rhytides.75,76 depressor anguli oris.78 The injection should be at
In the upper lip, 4 injection points are recommen- the level of the mandible to avoid affecting the
ded, and 2 in the lower lip. The injection points depressor labii inferioris muscle. A dose of 3 to
should be at the vermillion border at least 1.5 cm 5 U of BoNTA-ona and BoNTA-inco or 5 to 10 U
away from the oral commissures. Because this is BoNTA-abo is recommended at the posterior
a very delicate area that can lead to functional portion of the depressor anguli oris near the ante-
impairment of the lips (eg, eating, drinking, rior portion of the masseter.79
32 Dorizas et al

Masseter Platysmal Bands

The appearance of a wide jaw, which can be unde- Contraction of the platysma muscle can result in
sirable in many cases, is caused by the hypertro- prominent vertical bands in patients who are thin.
phy of the masseter muscle. BoNT injections to Horizontal neck lines can also be caused by mus-
treat a hypertrophic masseter are not the most cle activity or skin laxity. BoNTA can be a safe and
common indication, but are seen more often in effective treatment to reduce both the platysmal
Asians. The paralysis of the muscle leads to a bands and the horizontal lines caused by muscle
smoother and slimmer appearance. Ahn and col- activity. Careful patient selection is important
leagues80 treated 19 Asian patients with injections when evaluating platysmal band treatment with
of BoNTA-ona at the inferior border of the BoNTA. The ideal patient should have good skin
masseter. The initial dose was 25 U per side, and elasticity in the neck and little or no fat.63,68
then at 1-week follow-up an additional 25-U The injection points should be superficially into
dose was administered if needed; 3 of the patients each platysmal band, beginning below the jaw
required a third dose. They found that maximum line and spacing each injection approximately
correction occurred at 1 to 2 months, and reinjec- 2 cm apart. A total of 3 to 5 injections per side is
tion was required at 6 to 8 months. A dosing study recommended with 2 to 4 U of BoNTA-ona and
evaluated treatment doses of 10, 20, and 30 U of BoNTA-inco and 5 to 10 U of BoNTA-abo.2,78
BoNTA-ona in 22 patients.81 The investigators The greatest risk associated with injection into
used ultrasonographic imaging to compare the the platysmal bands is dysphagia or weakening
thickness of the masseter muscles before and af- of the muscles of the neck responsible for holding
ter injection, and concluded that the optimal the head up. The maximum recommended dose
dose was 20 U. Results were maintained for should not be exceeded to avoid causing any
6 months after treatment. side effects (40–60 U of BoNTA-ona and BoNTA-
In a study evaluating the treatment of masseter inco and 100 U of BoNTA-abo).68
hypertrophy in non-Asian, Western populations, Another interesting cosmetic use of BoNT in-
Liew and Dar82 used doses of 25 to 30 U of jected into the platysma is for the technique
BoNTA-ona on each side and compared the called the Nefertiti lift. Levy85 published an article
results with those of previously treated Asian describing a method of contouring the jawline
patients. Results showed that BoNTA-ona injec- with BoNT. His method is to treat platysma fibers
tions were successful in treating masseter along the jawline laterally from the nasolabial fold
hypertrophy in Western patients. Similar to the ex- up to the mandibular angle and into the posterior
periences of other investigators, they cautioned platysmal band. A total of 130 patients were in-
about careful placement of the injections. They jected with a total dose of 20 U of BoNTA-ona,
recommended that the injection point should be distributed in 2 to 3 U per injection site. The
below the sigmoid notch, approximately 1 cm study concluded that this technique was suc-
above the mandibular angle border.80 They noted cessful in creating a mini-lift of the jawline in pa-
few cases of difficulty with eating that lasted tients who wanted an alternative to invasive
4 weeks. Improvement in the lower face resulted surgical correction.
in prominence of the cheek bones in the Western
population, an effect that is typically very desir- INDICATIONS IN MEDICAL DERMATOLOGY
able. Similarly, Choi and Park found that among
Similar to the facial aesthetic use of BoNT, its use
45 patients who were injected with 25 to 30 U
in medical dermatology has also grown impres-
of BoNTA-ona, most were satisfied. However,
sively. BoNTA-ona was approved for the use of
most patients also experienced difficulty with
axillary hyperhidrosis, and its use has expanded
mastication, commonly followed by muscle ach-
into other dermatologic uses of BoNT. Some
ing. To avoid any side effects, it is advisable to
off-label uses include treatment of palmar and
begin treatment with smaller doses and re-treat
plantar hyperhidrosis, Raynaud disease, dyshi-
as needed.83,84
drotic eczema, lichen simplex, Hailey-Hailey
Several studies have noted the longer duration
disease, piloleiomyomas and leiomyomas, anal
of effect when treating masseter hypertrophy.
fissures, and inverse psoriasis.86–94
This finding is probably because the muscle hy-
pertrophy is caused by the excessive muscle use ON THE HORIZON
in these subjects, which is mitigated after the
BoNT injection. Regaining the baseline muscle Other BoNT products that have been developed or
mass after attenuated contraction takes longer are currently in development are aiming to improve
than simply regaining function.71,72 on the toxin formulations currently available.
 Aesthetic Uses of Botulinum Toxin 33

A topical gel containing BoNTA, RT001 (Revance therapies with concurrent use of BoNTA, fillers,
Therapeutics, Newark, CA, USA), is currently under and energy-based devices are becoming more
development for the treatment of crow’s feet. The common in clinical practice, further studies are
topical get contains both the toxin and a peptide needed to evaluate the combinability of these
that helps facilitate transport through the skin. treatments for facial rejuvenation.
Two controlled studies evaluating the efficacy of
RT001, involving 126 patients overall, showed clin- REFERENCES
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