Abstract 4663: Establishing the role of PRMT5 in lung cancer tumorigenicity and modulation of expression in smokers
Authors: Ali Syed N. , Patel Meet , Dieng Bamby , Lund Cody , Puri Neelu 
Published: March 2024

Abstract
Studies show that PRMT5 (protein arginine methyl transferase 5) is responsible for increased tumorigenicity by activation of the PI3K/Akt signaling pathway promoting increased cancer cell proliferation. However, the role of PRMT5 in modulating EGFR TKI resistance is minimally studied. We hypothesize that over-expression of PRMT5 could promote tumorigenesis and TKI resistance by activating PI3K-Akt-mTOR signaling pathways. Hence, modulating this key biomarker may help reduce tumorigenicity and overcome TKI resistance. We performed qPCR to study the expression of PRMT5 in normal lung cells and NSCLC cell lines. We also compared the expression of these biomarkers in erlotinib resistant (ER) and Osimertinib resistant (OR) cells using Western blotting. PRMT5 was inhibited using GSK591 (PRMT5i) to evaluate its sensitivity to EGFR TKIs (ER and OR). PRMT5 expression in normal lung cells and NSCLC cells treated with Cigarette smoke extract (CSE) were also studied and correlated with immunostaining results from smoker and non-smoker lung tumor tissue samples. Changes in PRMT5 expression were analyzed in WT-EGFR lung cancer cell lines (H358, H2170), mutant EGFR lung cancer cell lines (H3255, H1975, PC9) and normal lung fibroblast cells (HLF). PRMT5 expression was upregulated by 1.6-2.1 fold in HLF, H2170 and H358 cells after 1 week of treatment with CSE. CSE treatment also upregulated expression of inflammatory cytokines (IL-1β, IL-8 and TNF-α) in H358 cells by 1.6-7 fold as analyzed by qPCR. PRMT5 expression was upregulated in EGFR-TKI resistant cell lines, by 1.5-1.9 fold in PC9ER, PC9OR and H2170ER cells, compared to parental cells. Similar results were found using qPCR, with 1.6-2.3 fold increase in PRMT5 mRNA expression in H2170OR, PC9ER/OR and H3255OR cells compared to parental cells. An MTT assay was done to assess percentage viability of cells after treatment of PRMT5i. While 2μM PRMT5i treatment of PC9OR cells decreased cell viability by 42%, an inhibitory effect that was more than additive was seen with a combination of 2μM of PRMT5i and 21nM of OR. Similar results were also observed in H2170ER cells. Migratory properties of OR resistant cells, were studied by wound healing assays. Percentage wound area in PRMT5i treated PC9OR cells was 29.3% as compared to 6.1% and 10.5% in diluent and OR treated cells respectively after 48 hrs. Combination treatment of PRMT5i and OR further inhibited cell migration with higher wound area of 41.7%. Immunostaining of lung tumor sections from smokers and non-smokers showed high expression of PRMT5 localized in the cytoplasm and nucleus. Microscopic analysis of 20 tumor sections from smokers showed higher expression of PRMT5 (p&lt;0.05) compared to non-smokers. We can conclude that PRMT5 is upregulated in TKI resistant cells causing tumorigenesis. Modulation of PRMT5 can prove to be groundbreaking and revolutionary, as it may increase the efficacy of EGFR TKIs.
Citation Format: Syed N. Ali, Meet Patel, Bamby Dieng, Cody Lund, Neelu Puri. Establishing the role of PRMT5 in lung cancer tumorigenicity and modulation of expression in smokers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4663.