Exploration of the regulatory mechanism of PD-L1 expression in pMMR type gastric cancer cell by cytotoxic drugs and anti angiogenic drugs

Background and Objectives: Advanced gastric cancer with proficient mismatch repair (pMMR) status responds poorly to immune checkpoint inhibitors (ICIs) and constitutes a significant proportion of cases, posing a major clinical challenge. Studies suggest that cytotoxic and anti-angiogenic drugs may influence ICI efficacy by modulating programmed death-ligand 1 (PD-L1) expression; however, their specific role and mechanisms in pMMR gastric cancer remain unclear. This study aimed to investigate the regulatory effects of cytotoxic drugs (5-fluorouracil, 5-FU; cisplatin) and an anti-angiogenic drug (apatinib) on PD-L1 expression in human pMMR-type AGS gastric cancer cells and explore the underlying signaling pathway mechanisms. Methods The impact of drug treatments at various concentrations on AGS cell proliferation was first assessed using the CCK-8 assay to select appropriate concentrations for subsequent experiments. Real-time quantitative PCR (qPCR) and Western blotting were employed to evaluate the effects of 72-hour drug treatment on PD-L1 mRNA and protein expression levels, respectively. Furthermore, Western blot analysis was used to examine drug-induced changes in the expression of key proteins (PI3K, Akt, MEK1, ERK) within the PI3K/Akt/mTOR and RAS/RAF/MEK/ERK signaling pathways, which are potentially involved in PD-L1 regulation. Results 1. Regulation of PD-L1 Expression: qPCR results showed that 5-FU (64 µM), cisplatin (16 µM), and apatinib (320 nM, 640 nM) significantly upregulated PD-L1 mRNA expression in AGS cells ( P  < 0.05). Western blot analysis further confirmed that treatment with 5-FU (64 µM), cisplatin (16 µM), and apatinib (320 nM) significantly increased PD-L1 protein levels ( P  < 0.05). 2. Alterations in Signaling Pathway Proteins: Western blot analysis indicated that treatments with 5-FU, apatinib, and cisplatin all upregulated PI3K protein expression ( P  < 0.05). Regarding Akt protein, apatinib treatment increased its expression, whereas 5-FU and cisplatin decreased it ( P  < 0.05). Within the MAPK pathway, 5-FU treatment upregulated the expression of MEK1 and ERK proteins, while apatinib and cisplatin treatments downregulated the expression of both proteins ( P  < 0.05). Conclusion At specific concentrations, cytotoxic drugs (5-FU, cisplatin) and the anti-angiogenic drug (apatinib) can upregulate both mRNA and protein expression of PD-L1 in pMMR-type AGS gastric cancer cells and differentially modulate the expression of key proteins in the PI3K/Akt and MAPK signaling pathways. These findings provide preliminary in vitro experimental evidence and mechanistic insights supporting the potential of combining these drugs with ICIs to enhance therapeutic efficacy in pMMR gastric cancer.