Abstract 5270: Silibinin modulates migration and survival pathways in bone marrow mast cells via RAC2: Implications in its anti-cancer activity in basal cell carcinoma growth and progression

Abstract

Abstract Most of the non-melanoma skin cancers are basal cell carcinoma (BCC); thus, extensive efforts to discover chemopreventive agents against BCC are ongoing. Recently, we showed that silibinin (SB) exerts strong efficacy against ultraviolet B radiation (UVB)-induced BCC growth/progression, and mast cells (MCs) are one of the targets of its efficacy. Notably, 30 days of UVB exposure in patched (Ptch)+/- mouse model of UVB-induced BCC formation increased MC numbers by ~50% that was completely inhibited by SB. Hence, understanding pathways associated with MC regulation by SB seems valuable for its efficacy against BCC and other cancers. We generated bone marrow MCs (BMMCs) in presence of interleukin (IL)-3 and stem cell factor from C57BL/6 mice (confirmed using flow cytometry dual staining: cKit/Fc€RI). They were treated with two concentrations of SB (25 µM; SB 25 and 100 µM; SB 100) or untreated (control), and proteomics was performed (liquid chromatography mass spectrometry on Fusion Lumos mass spectrometer). Total 3575 proteins were identified and 166 were found to be statistically significant (ANOVA, Fisher’s posthoc analysis, p<0.05, FDR<0.01) amongst the three groups (n=4/group). Proteins with highest differential expression among the groups (greatest fold change expression; n=50), greatest contribution to the selection (variable importance; n=15), and common expression profiles in both SB 25 and SB 100 compared to control (n=21) were selected for further pathway analysis. Important nodes found for SB regulation of MCs were tumor necrosis factor, interferon-gamma, and IL-13; IL-13 also emerged as a key regulatory node in normal BMMCs. Furthermore, to determine the effect of SB in BCC associated MCs, a subset of proteins implicated with skin-associated MC activity (determined using IPA) was generated. Most important molecules implicated included IL-13 and Rac Family Small GTPase2 (RAC2; regulates all myeloid lineages). RAC2 was found to modulate the expression of mast cell protease4 (MCPT4), mast cell transcriptase1 (TPSAB1), and macrophage migration inhibition factor. RAC2 also activates MC proteases via JNK signaling cascade. Together, via the regulation of MCPT4, TPSAB1, and JNK signaling, RAC2 regulates inflammatory responses, and controls MC migration via actin cytoskeleton reorganization and lamellipodia formation. Phagosome maturation was also an important pathway implicated in our analysis; MCs lyse engulfed pathogens/debris via phagosome maturation and fusion with lysosomes. In conclusion, our findings demonstrate that SB increases RAC2 expression in BMMCs, which can regulate MC migration into the tumors and effectively increase the MC protease gene expression. Thus, SB treatment shifts BMMCs towards anti-tumorigenic pathways and RAC2 can be a novel therapeutic target in BCC and other cancers as well. Citation Format: Neha Mishra, Sandeep Paudel, Chapla Agarwal, Rajesh Agarwal. Silibinin modulates migration and survival pathways in bone marrow mast cells via RAC2: Implications in its anti-cancer activity in basal cell carcinoma growth and progression. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5270.