GNPNAT1 Regulation: A Key Role in Radioimmune Function and NK Cell Resistance in NSCLC

Background: Glucosamine-6-phosphate N-acetyltransferase 1 (GNPNAT1) is an enzyme involved in the hexosamine biosynthetic pathway, which is critical for glycosylation processes. In the context of non-small cell lung cancer (NSCLC), GNPNAT1 plays a significant role in modulating immune responses. The purpose of this study is to investigate the role of GNPNAT1 in regulating the efficacy of radiotherapy and resistance to natural killer (NK) cell-mediated cytotoxicity in patients with NSCLC.

Methods: To assess GNPNAT1's impact on radiotherapy efficacy, 122 lung cancer patients were categorized into radiosensitive and radioresistant groups. GNPNAT1 expression levels in cancerous tissues from both groups were measured using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blotting. This analysis was extended to various lung cancer cell lines (BEAS-2B, A549, LTEP-2, SPCA1, and H157) using the same molecular techniques. To investigate GNPNAT1's functional role in radioresistance, radioresistant A549 cells (A549R26-1) were established, and GNPNAT1 expression was genetically manipulated. Experimental groups included control, si-NC, si-GNPNAT1, Oe-NC, and Oe-GNPNAT1. Post-treatment, GNPNAT1 levels were measured via qRT-PCR and Western blotting. Cells were exposed to varying doses of radiation, and subsequent assessments included cell proliferation (Cell Counting Kit-8 (CCK-8) assay), radiosensitivity (plate cloning assays), and apoptosis rates (flow cytometry). Isolated and purified primary NK cells were co-cultured with lung cancer cells from each experimental group. The cytotoxicity of NK cells against lung cancer cells was assessed through lactate dehydrogenase (LDH) release and colony formation assays.

Results: Compared to the radiosensitive group, the radioresistant group exhibited significantly elevated GNPNAT1 expression levels ( < 0.05). The radioresistant cell line A549R26-1 demonstrated higher proliferation ability and lower apoptosis levels compared to its parental cell line, A549P. Subsequently, down-regulation of GNPNAT1 expression in A549R26-1 cells resulted in reduced proliferation, increased apoptosis, and weakened resistance to NK cell cytotoxicity. Conversely, up-regulation of GNPNAT1 expression in A549R26-1 cells following co-culture with NK cells led to increased proliferation and survival rates, and enhanced resistance to NK cell cytotoxicity. Notably, GNPNAT1 knockdown effectively attenuated the radioresistance of A549R26-1 cells.

Conclusion: Down-regulation of GNPNAT1 expression reduces the immune resistance of non-small cell lung cancer to radiotherapy and enhances susceptibility to NK cell cytotoxicity.