Discussion
The aim of this preliminary phase 1 study was to evaluate the potential
efficacy of interventions that might be used to mitigate ISRs following
the SC administration of 60 mg elamipretide in healthy subjects as well
as any potential impact on PK and safety. We focused on treatments that
target mast cells or factors upstream or downstream to their activation,
due to our preclinical data that strongly suggest that
elamipretide-associated ISRs are caused by direct activation of mast
cells through MRGPRX2. Overall, mometasone appeared to favorably
mitigate the presumed pseudo-allergic reactions observed following SC
elamipretide, specifically reducing pruritus and induration/swelling
with no effect on PK. Ice application ameliorated early injection site
pain and itching, but reduced the absorption of elamipretide, while
diphenhydramine demonstrated potential in reducing induration/swelling
but caused sedation in some cases. Tacrolimus and doxepin, as utilized
in this study, demonstrated little impact on reported ISRs compared to
elamipretide alone and are not likely to be recommended for ISR
mitigation, though additional investigation could be considered. There
were no significant changes in elamipretide or M1 and M2 metabolite
plasma exposures with any of the treatments except for ice application,
which reduced exposure to elamipretide and its metabolites.
Although ice application and diphenhydramine showed some reduction in
ISR signs and symptoms, mometasone revealed the most promise. The
reduction in pruritus and induration/swelling demonstrated by mometasone
enabled patients to be more comfortable with the elamipretide treatment.
In addition, mometasone may further improve elamipretide tolerability by
reducing scratching and subsequent scratching-related skin damage. In
some subjects, the hydrocolloid occlusive dressing applied over the
mometasone ointment resulted in redness of the covered area (i.e. the
injection site area), confounding the ability to identify erythema due
to ISRs. The capture of ISR photographs proved useful in deciphering
erythema grading in subjects where the pattern of redness appeared to be
related to the use of the occlusive dressing. However, it is possible
that erythema as an ISR was overreported in the mometasone treatment arm
of this study.
Despite ice application and diphenhydramine reducing some ISR signs and
symptoms, both interventions presented undesirable effects.
Specifically, the simple intervention of ice application reduced early
injection site pain and itching, but reduced the Cmaxand AUC0-6h of elamipretide and its metabolites (M1 and
M2), possibly due to vasoconstriction. Neither M1 nor M2 are
biologically active and the potential reduction in plasma concentrations
of these metabolites is not therefore anticipated to have a clinical
impact [data on file, Stealth BioTherapeutics Inc.]. However, while
the potential ramification on efficacy of the disruption in absorption
of the active parent drug is unclear, lowering plasma exposure of
elamipretide is not desirable, making ice application a less appropriate
mitigation option. Similarly, diphenhydramine showed some potential in
reducing induration/swelling, but a significant incidence (50%) of mild
somnolence was reported in this treatment arm. While second generation
antihistamines were not included in this study, the observations with
diphenhydramine suggest that other antihistamines that are less sedating
could provide utility in mitigating ISRs.
Based on the outcomes of mometasone and antihistamine use, further
investigation is warranted to include separate and combined
interventions with oral second-generation antihistamines (such as
fexofenadine) and topical mometasone without the use of an occlusive
dressing. Fexofenadine is a selective peripheral H1 receptor antagonist
that does not readily cross the blood–brain barrier, causing less
drowsiness in comparison to the first-generation antihistamines, such as
diphenhydramine. Several international dermatology and allergy
organizations issued a common guideline on chronic urticaria management,
recommending the regular use of second-generation antihistamines as
first-line treatment. Fexofenadine appears safe and well tolerated and
daily doses can be titrated upwards in case of no improvement
(Zuberbier, 2018). Chronic urticaria could be considered as the closest
model to chronic ISRs in terms of available data on therapeutic
management. With regard to mometasone, given that elamipretide is
injected daily at alternating sites, applications with mometasone twice
a day (one prior and one post-injection) appear to be a more practical
alternative to the use of mometasone with occlusive dressing.
Our focus on mast cells arose from preclinical experiments which
established that elamipretide, in addition to its intended function,
also acts as an agonist of the human G protein-coupled receptor MRGPRX2.
MRGPRX2 is primarily expressed by mast cells, which are constitutive
residents of the skin and other tissues, trigger rapid tissue
inflammation, and mediate many of the symptoms of allergic diseases. We
reasoned that MRGPRX2 might be involved because it can be activated by
many cationic peptides and small molecules with properties similar to
elamipretide and which also cause ISRs and other pseudo-allergic
reactions (McNeil 2021a). We have several pieces of evidence to support
the hypothesis that elamipretide-associated ISRs are due to MRGPRX2
activation. First, elamipretide triggers intracellular signaling
pathways, as measured by calcium mobilization, in cell lines forced to
express MRGPRX2 or its mouse ortholog Mrgprb2, but not in unmodified
cell lines that do not natively express the receptors (Figures 2 A, B).
Notably, this could be blocked by a molecule reported to inhibit MRGPRX2
signaling (Figure 2C). MRGRPX2 activation by elamipretide is
physiologically relevant, as the calculated EC50 is over
1,000-fold lower than the injection concentration in our clinical trial.
Second, elamipretide causes intracellular signaling, again as measured
by calcium flux, in primary mouse mast cells from wild type but not
Mrgprb2 knockout mice (Figure 3). Third, SC elamipretide injection into
mouse hindpaws triggered rapid swelling via fluid extravasation from the
bloodstream, similar to human ISRs, in wild type mice, while
extravasation was nearly absent in mice lacking Mrgprb2 (Figure 4).
These effects almost certainly are due to the parent drug, as
elamipretide’s metabolites have no effect on MRGPRX2 signaling (Figures
2 D, E).
While itch produced in humans by injection of MRGPRX2 agonists can be
blocked by antihistamines (Hasbak et al., 2006), development of
inhibitors is still in its infancy with no candidates in clinical trial
yet (McNeil, 2021a). Indeed, a combination of H1 and H2 histamine
receptor antagonists have demonstrated efficacy in blocking
MRGPRX2-driven systemic and local reactions (McNeil 2021b) and could be
considered in future elamipretide-driven ISR studies. Although the
topical steroid mometasone reduced pruritus and induration/swelling
following SC elamipretide in this study, its anti-inflammatory mechanism
is unclear but is thought to act by inhibition of the arachidonic acid
pathway (Spada, et al., 2018; ELOCON® Prescribing
Information, 2018). In addition to anti-inflammatory effects, topical
steroids, such as mometasone, possess anti-mitotic, immunosuppressive,
and vasoconstrictive effects (Gabros et al., 2021), which may have
played a role in mitigating ISR signs in this study.
Overall, the data collected in this study support prior findings that SC
elamipretide is generally safe and well-tolerated (Karaa et al., 2020;
Thompson et al., 2021; Mettu et al., 2022). With the exception of data
relating to ISRs, very few TEAEs were identified in this study. The only
TEAE seen in more than one subject was that of somnolence in the
diphenhydramine treatment arm. Given that somnolence is a well-known
side effect of diphenhydramine (Sicari & Zabbo 2021), this adverse
event was considered likely related to diphenhydramine and not to
elamipretide treatment.
A limitation of the study was the relatively low number of subjects
evaluated (N=10), which impacted the potential to demonstrate
statistically significant results with respect to efficacy of
mitigation. This study was not powered to show a statistically
significant difference in the ISR profile between treatment arms but was
meant to identify signals that could warrant further investigation in
other clinical settings.