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EIF3D promotes sunitinib resistance of renal cell carcinoma by interacting with GRP78 and inhibiting its degradation.

ABSTRACT: BACKGROUND:Sunitinib is one of the multi-targeted tyrosine kinase inhibitors for the treatment of renal cell carcinoma (RCC) at present. However, its clinical efficacy is limited by chemoresistance of RCC. Our previous study found that eukaryotic translation initiation factor 3 subunit D (EIF3D) was an oncogene in the development and progression of RCC but little is known about whether EIF3D participated in sunitinib resistance of RCC. METHODS:The expression of EIF3D in the tumor tissue specimen was detected by immunohistochemistry. The effect of EIF3D on sunitinib-resistance of RCC cells was evaluated by colony formation, IC50 proliferation and in vivo tumor growth assays. The interaction between EIF3D and glucose regulated protein 78 (GRP78) was assessed by Co-IP and Western blot assays. FINDING:EIF3D expression was found higher in 786-OR and ACHN-R cells with acquired sunitinib resistance than that in 786-O and ACHN cells sunitinib to sensitive. The EIF3D level was also up-regulated in sunitinib-chemoresistant tumor tissues compared with chemosensitive tumor tissues. Functional study showed that EIF3D knockdown decreased cell viability with sunitinib treatment. Mechanistical study demonstrated that EIF3D interacted with GRP78 and enhanced protein stability through blocking the ubiquitin-mediated-proteasome degradation of GRP78. GRP78 overexpression induced sunitinib resistance of RCC cells by triggering the unfolded protein response, whereas GRP78 silencing inhibited cell viability. Forced expression of GRP78 eliminated the inhibitory effect of EIF3D silencing on cell growth in vitro and in vivo. INTERPRETATION:our results indicate that EIF3D played an important role in sunitinib resistance of RCC cells, suggesting that it may prove to be a potential therapeutic target for RCC.

SUBMITTER: Huang H

PROVIDER: S-EPMC6945244 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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EIF3D promotes sunitinib resistance of renal cell carcinoma by interacting with GRP78 and inhibiting its degradation.

Huang Hai H Gao Yi Y Liu Ao A Yang Xiaoqun X Huang Fang F Xu Le L Danfeng Xu X Chen Lu L

EBioMedicine 20191026

<h4>Background</h4>Sunitinib is one of the multi-targeted tyrosine kinase inhibitors for the treatment of renal cell carcinoma (RCC) at present. However, its clinical efficacy is limited by chemoresistance of RCC. Our previous study found that eukaryotic translation initiation factor 3 subunit D (EIF3D) was an oncogene in the development and progression of RCC but little is known about whether EIF3D participated in sunitinib resistance of RCC.<h4>Methods</h4>The expression of EIF3D in the tumor ...[more]

PMID: 31669222

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Project description: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-?B (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-?B (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.

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EIF3D promotes sunitinib resistance of renal cell carcinoma by interacting with GRP78 and inhibiting its degradation.

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EIF3D promotes sunitinib resistance of renal cell carcinoma by interacting with GRP78 and inhibiting its degradation.

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