Abstract 6810: The mechanism of PRMT5 in causing smoking-induced lung cancer and development of EGFR Tyrosine kinase inhibitor resistance

Abstract In 2024, lung cancer accounts for 20% of US cancer deaths and 80-90% of NSCLC patients are smokers. PRMT5 (protein arginine methyl transferase 5) is involved in various normal cellular pathways and activates the PI3k-Akt signaling pathways that increase lung tumorigenicity. In NSCLC, EGFR-TKI treatments with Osimertinib initially reduce tumor growth but often lead to resistance in about 19 months. Currently, PRMT5 inhibitors are in clinical trials for several malignancies; further studying PRMT5's role in EGFR-TKI resistance could reveal if these inhibitors enhance TKI efficacy in NSCLC. We hypothesize that modulating PRMT5 could reduce tumorigenicity and overcome OR resistance. We studied the expression of PRMT5 in both drug-resistant (OR) and drug-sensitive (parental) H3255 and PC9 NSCLC cell lines after treatment with CSE for 24 and 48 hours, followed by western blotting and qPCR. Also, using qPCR the effects of PRMT5i after CSE treatment were studied on inflammatory cytokines.The efficacy of PRMT5i (GSK591) on PC9OR cells were studied using bioprinted with GelMA with a R-Gen 200 bioprinter. After 24 and 48 hours of CSE treatment PRMT5 gene expression was increased as observed using qPCR in TKI-resistant H3255P/OR (1.5-11 fold) and PC9/OR (1.5-17 fold) (p<0.05) cells respectively. Immunoblotting results also indicated a 1.3-2.1-fold (p<0.01) upregulation of PRMT5 in these cell lines. PC9P/OR cells were treated with GSK591, and the MTT assay revealed a significant decrease in cell viability to 49% and 46% (p<0.01) respectively, in the GSK591 plus OR group as compared to OR treatment group. The cell viability was significantly lowered by 49% in the siRNA plus OR group compared to the siRNA group alone. The treatment groups in both siRNA and PRMT5i treated cells had additive inhibitory effect (p<0.01). Studies on inflammatory cytokines in PC9OR cells first treated with CSE and then with PRMT5i showed that CSE alone caused an upregulation of cytokines: TNFα (4-5 fold), IL-1β (1-2.3 fold), and IL-8 (7-9 fold) (p<0.05). However, treatment with PRMT5i resulted in a downregulation of these cytokines: TNFα (0.4-0.5 fold), IL-8 (0.5 fold), and IL-1β (0.4-0.6 fold) (p<0.05). These findings indicate that inhibiting or downregulating PRMT5 may be effective in reducing CSE-induced oncogenicity. To replicate patient lung tumor microenvironment and enhance the clinical relevance of our findings, we employed 3D bioprinting by accurately layering cells onto biological scaffolds. Preliminary results with PC9OR along with HLF (Human lung fibroblasts) bioprinted spheroids showed 40% inhibition of spheroid count/microscopic field by a combination of PRMT5i and OR compared to OR alone. In conclusion, we infer that PRMT5 is overexpressed in TKI-resistant cells, leading to increased oncogenic potential. Modulating PRMT5 levels could enhance the effectiveness of EGFR TKIs in NSCLC patients. Citation Format: Ayesha Khan, Usama Altayeh, Subaranjana Saravanaguru Vasanthi, Mohammad Fazle Alam, Neelu Puri. The mechanism of PRMT5 in causing smoking-induced lung cancer and development of EGFR Tyrosine kinase inhibitor resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6810.