D1.324 - Evaluation of the Potential for Sebetralstat to Cause Pseudo-allergic Reactions

Injection-site reactions occurred in >90% of icatibant clinical trial participants driven primarily by pseudo-allergic reactions secondary to cutaneous mast cell degranulation and histamine release initiated through Mas-Related G-Protein Coupled Receptor X2 (MRGPRX2) activation (McNeil et al. Nature 2015). Sebetralstat, an investigational on-demand treatment for HAE-C1INH attacks, showed no evidence of causing allergic reactions in a recently completed phase 3 trial. Overall, sebetralstat was well tolerated in clinical trials and had a safety profile no different from placebo. To complement clinical safety, we conducted general safety profiling with sebetralstat utilizing in vitro assays.

The general safety profiling of sebetralstat (10 μM) was examined against commercial panels (AdverseReactionEnzymes, SafetyScreen) comprising 124 receptor, enzyme, transporter, and ion-channel binding targets (Eurofins Panlabs). MRGPRX2 activation was examined in the PathHunter^® beta-arrestin human CHO-K1 cell line stably expressing MRGPRX2. Cells were incubated with Cortistatin 14 (positive control), icatibant, or sebetralstat, and calcium mobilization following receptor activation was monitored in duplicate using a calcium-sensitive fluorescent dye in a live cell, non-imaging assay (Eurofins DiscoverX Corp). Half-maximal effective concentration (EC₅₀) and percentage activity at concentrations of 3 to 100 µM were calculated.

In the in vitro safety profiling panel assays, sebetralstat did not significantly inhibit the activity or binding at any of the 124 molecular targets (ie, <50% inhibition at 10 µM), representing a window of >1,656-fold selectivity of sebetralstat. The EC₅₀ of MRGPRX2 activation was 9.5 µM for icatibant and >100 µM for sebetralstat (Figure).

Corroborating the clinical safety observed in sebetralstat clinical trials, in vitro assays confirm the high selectivity of sebetralstat across a wide panel of molecular targets associated with drug adverse reactions and safety concerns. Sebetralstat also did not activate MRGPRX2 at clinically relevant concentrations, reducing the likelihood of pseudo-allergic reactions with broader use in a potential post-approval setting.