Inhibition of NSUN3 suppresses immune escape in non-small cell lung cancer through stabilizing PD-L1 in a 5-methyladenosine dependent way

Abstract

Background: Alterations in 5-methyladenosine (m5 C) RNA methyltransferase have been implicated in the pathogenesis of lung cancer. However, the role of NSUN3, a key m5 C regulator, in modulating PD-L1 expression and immune evasion in Non-Small Cell Lung Cancer (NSCLC) remains poorly understood. Methods: NSUN3 expression levels in NSCLC tissues and cell lines were quantified using quantitative Reverse Transcription PCR (RT-qPCR). The impact of NSUN3 expression modulation on lung cancer cell viability and proliferation was assessed through CCK-8 assays and colony formation experiments. To evaluate immune cell interactions, CD8+ T-cells were co-cultured with NSCLC cells, and cytotoxic activity was measured by an LDH cytotoxicity detection kit. The in vivo tumorigenic role of NSUN3 was investigated using subcutaneous tumor xenograft models. Mechanistic insights into NSUN3-PD-L1 interactions were obtained through m5 C-RIP, RIP, and dual-luciferase reporter assays. Results: Elevated NSUN3 mRNA levels were observed in NSCLC tissues compared to normal controls. Functional studies demonstrated that NSUN3 knockdown significantly inhibited cell viability and proliferation in both A549 and PC9 cell lines. Furthermore, NSUN3 depletion enhanced CD8+ T-cell-mediated cytotoxicity against NSCLC cells and suppressed tumor growth in vivo. Mechanistically, NSUN3 inhibition was found to stabilize PD-L1 mRNA, suggesting a regulatory role in immune checkpoint modulation. Conclusions: The present findings establish NSUN3 as an oncogenic driver in NSCLC pathogenesis, functioning through PD-L1 mRNA stabilization. The identification of this NSUN3-PD-L1 regulatory axis provides new insights into the molecular mechanisms underlying immune escape in NSCLC and offers promising directions for developing targeted immunotherapies.