Identification and validation of prognostic and tumor microenvironment characteristics of necroptosis index and BIRC3 in clear cell renal cell carcinoma.

Background: Necroptosis is a form of programmed cell death; it has an important role in tumorigenesis and metastasis. However, details of the regulation and function of necroptosis in clear cell renal cell carcinoma (ccRCC) remain unclear. It is necessary to explore the significance of necroptosis in ccRCC.

Methods: Necroptosis-related clusters were discerned through the application of Consensus Clustering. Based on the TCGA and GEO databases, we identified prognostic necroptosis-related genes (NRGs) with univariate COX regression analysis. The necroptosis-related model was constructed through the utilization of LASSO regression analysis, and the immune properties, tumor mutation burden, and immunotherapy characteristics of the model were assessed using multiple algorithms and datasets. Furthermore, we conducted comprehensive GO, KEGG, and GSVA analyses to probe into the functional aspects of biological pathways. To explore the expression and of hub gene (BIRC3) in different ccRCC cell types and cell lines, single-cell sequencing data was analysed and we performed Quantitative Real-time PCR to detect the expression of BIRC3 in ccRCC cell lines. Function of BIRC3 in ccRCC was assessed through Cell Counting Kit-8 (CCK8) assay (for proliferation), transwell and wound healing assays (for migration and invasion).

Results: Distinct necroptosis-related clusters exhibiting varying prognostic implications, and enrichment pathways were identified in ccRCC. A robust necroptosis-related model formulated based on the expression of six prognostic NRGs, presented substantial predictive capabilities of overall survival and was shown to be related with patients' immune profiles, tumor mutation burden, and response to immunotherapy. Notably, the hub gene BIRC3 was markedly upregulated in both ccRCC tissues and cell lines, and showed significant correlations with immunosuppressive cells, immune checkpoints, and oncogenic pathways. Downregulation of BIRC3 demonstrated a negative regulatory effect on ccRCC cell proliferation migration and invasion.

Conclusion: The necroptosis-related model assumed a pivotal role in determining the prognosis, tumor mutation burden, immunotherapy response, and immune cell infiltration characteristics among ccRCC patients. BIRC3 exhibited significant correlations with the immunosuppressive microenvironment, which highlighted its potential for informing the design of innovative immunotherapies for ccRCC patients.