The tumor promoter role and molecular mechanism of C8orf76/CALB2 axis in clear cell renal cell carcinoma

Background: Renal cell carcinoma (RCC) is one of the most common malignant tumors of the urinary system, with clear cell renal cell carcinoma (ccRCC) accounting for 90% of all RCC subtypes. Chromosome 8 open reading frame 76 (C8orf76) is significantly upregulated in various tumor types and has been involved in tumor cell proliferation, migration, invasion, and is associated with poor prognosis. However, the expression profile and molecular mechanisms of C8orf76 in ccRCC have not been fully elucidated, and further investigations are required to clarify these aspects. Methods: We systematically investigated the mechanism of action of C8orf76 in ccRCC through in vitro biological function experiments. Cellular function assessments were performed, including CCK-8 assay, colony formation assay, flow cytometry, SA-β-gal staining, Transwell chamber assay, and wound healing assay. Additionally, combined with a subcutaneous xenograft mouse model and an in vivo imaging system, we studied the phenotypic changes following C8orf76 knockdown. Potential downstream targets of C8orf76 were screened via RNA-sequencing and bioinformatics analysis. Additionally, we utilized The Cancer Genome Atlas (TCGA) database to analyze the expression patterns of C8orf76 and CALB2 in ccRCC, as well as their correlations with clinical prognosis. Results: Both C8orf76 and CALB2 are highly expressed in ccRCC and correlate with poor prognosis. Knockdown of C8orf76 significantly inhibits the proliferation and migration of ccRCC cells both in vivo and in vitro. Specifically, Knockdown of C8orf76 downregulates the transcriptional level of CALB2, leading to G1-phase cell cycle arrest, enhanced cellular senescence, and subsequent suppression of ccRCC proliferation and migration. Furthermore, ectopic overexpression of CALB2 can partially reverse these effects. Dual-luciferase reporter assay confirms that C8orf76 directly binds to the promoter region of CALB2. Similarly, CALB2 knockdown also induces tumor cell cycle arrest and cellular senescence, accompanied by inhibited proliferation and migration of ccRCC. Notably, the aforementioned phenomena are partially rescued following further knockdown of CDKN2A. Conclusions: C8orf76 is highly expressed in clear cell renal cell carcinoma and correlates with poor prognosis. C8orf76 directly binds to the CALB2 promoter, thereby promoting CALB2 transcription and downstream biological behaviors. Inhibition of the C8orf76/CALB2 axis induces G1-phase cell cycle arrest and activates cellular senescence signaling pathways, which in turn suppresses the proliferation and migration of ccRCC.