primary CD34+ derived mast cells provide an excellent platform to assess single agent and combination therapies for mastocytosis Free

Mast cells are multi-functional tissue-dwelling cells involved in various pathological conditions such as allergy, asthma, cardiovascular diseases, rheumatoid arthritis, rosacea and mastocytosis. Multiple compounds targeting different receptors have been used to treat mastocytosis including Omalizumab (FcεR1A), Avapritinib (CD117), Budesonide (glucocorticoid receptor) and Salmetrol (β2 adrenergic receptors). Since mast cells are tissue-resident cells that are present only in small numbers and are thus poorly accessible for in vitro analyses, studies on human mast cell biology have typically utilized cell lines and primary mast cells. We evaluated mobilized peripheral blood (MPB) CD34+ derived mast cells from multiple donors, cultured in medium with IL-3, IL-6 and SCF for two weeks followed by a medium containing IL-6 and SCF. Following 7 weeks of culture mast cell phenotype (n=6 unique donors) was confirmed with expression of CD117 (85 ± 11%), MRGX2 (40 ± 23%), FcεR1A (61 ± 17%) and CD203c (80 ±17). Degranulation was induced by IgE, Substance P, Cortistatin 14 and Compound 48/80 using a flow cytometry-based CD63 readout as well as histamine and tryptase release via ELISA. Omalizumab, Avapritinib, Budesonide and Salmetrol were tested over a six-point curve on primary mast cell degranulation. The cells were plated at 40,000 cells per well and with an overnight incubation with an IgE/ anti-IgE stimulation. The mean IC₅₀ value for Omalizumab and Avapritinib were 5.7 nM and 7.3 µM respectively. Budesonide, and Salmetrol induced limited degranulation even at concentrations of 10 µM. Combinations of Avapritinib (dose response) with Omalizumb at 1 nM significantly increased the degranulation with a mean IC₅₀ value of 85 nM for Avapritinib. This data confirm primary MPB CD34+ cell derived mast cells provide an excellent in vitro platform with an unlimited supply of cells to assess single agent and combinations for mastocytosis and other mast cell diseases.