DNA methylation-regulated QPCT promotes sunitinib resistance by increasing HRAS stability in renal cell carcinoma

Abstract

Rationale: Although sunitinib has been shown to improve the survival rate of advanced renal cell carcinoma (RCC) patients, poor drug response is a major challenge that reduces patient benefit. It is important to elucidate the underlying mechanism so that the therapeutic response to sunitinib can be restored.Methods: We used an Illumina HumanMethylation 850K microarray to find methylation-differentiated CpG sites between sunitinib-nonresponsive and -responsive RCC tissues and Sequenom MassARRAY methylation analysis to verify the methylation chip results. We verified glutaminyl peptide cyclotransferase (QPCT) expression in sunitinib-nonresponsive and -responsive RCC tissues via qRT-PCR, western blot and immunohistochemical assays. Then, cell counting kit 8 (CCK-8), plate colony formation and flow cytometric assays were used to verify the function of QPCT in RCC sunitinib resistance after QPCT intervention or overexpression. Chromatin immunoprecipitation (ChIP) was performed to clarify the upstream regulatory mechanism of QPCT. A human proteome microarray assay was used to identify downstream proteins that interact with QPCT, and co-immunoprecipitation (co-IP) and confocal laser microscopy were used to verify the protein chip results.Results: We found that the degree of methylation in the QPCT promoter region was significantly different between sunitinib-nonresponsive and -responsive RCC tissues. In the sunitinib-nonresponsive tissues, the degree of methylation in the QPCT promoter region was significantly reduced, and the expression of QPCT was upregulated, which correlated with a clinically poor response to sunitinib. A knockdown of QPCT conferred sunitinib sensitivity traits to RCC cells, whereas an overexpression of QPCT restored sunitinib resistance in RCC cells. Mechanistically, reducing the methylation degree of the QPCT promoter region by 5-aza-2'-deoxycytidine (decitabine) in RCC cells could increase the expression of QPCT and NF-κB (p65) bound to the QPCT promoter region, positively regulating its expression, while the hypermethylation in the QPCT promoter region could inhibit the binding of NF-κB (p65). QPCT could bind to HRAS and attenuate the ubiquitination of HRAS, thus increasing its stability and leading to the activation of the ERK pathway in RCC cells.Conclusion: QPCT may be a novel predictor of the response to sunitinib therapy in RCC patients and a potential therapeutic target.

Hypermethylation of GNA14 and its tumor-suppressive role in hepatitis B virus-related hepatocellular carcinoma

“…DNA methylation, an important epigenetic modification, affects the development of tumors in a variety of ways 17 - 19 . Upregulated DNA methylation of tumor suppressive genes leads to their decreased expression, which is a crucial inducement factor for HCC 20 .…”

Restoring the epigenetically silenced PCK2 suppresses renal cell carcinoma progression and increases sensitivity to sunitinib by promoting endoplasmic reticulum stress

“…However, with the use of sunitinib, its drawbacks have gradually emerged. In the clinical treatment process, the drug treatment efficiency often decreases, which results in insensitivity to the drug and even resistance in patients 55 , 56 . These are the significant challenges and difficulties faced in the clinical treatment of RCC.…”

Screening of Autoantibodies in Serum of Renal Cancer Patients Based on Human Proteome Microarray

“…HRAS is a member of the RAS family, and normal or mutated forms of HRAS are overexpressed in multiple tumors [28][29][30][31]. QPCT (Glutaminyl peptide cyclotransferase) is bound to HRAS and increases the stability of HRAS by reducing its ubiquitination degradation, thus activating the ERK signaling pathway and leading to sunitinib resistance in renal cell cancer [32]. The above results indicate that proteins abnormalities in the MAPK signaling pathway may exist in all types of kidney cancer.…”