Genomic Study Of Renal Cell Carcinoma With Venous Tumor Thrombus And Role Of SETD2 In Drug Resistance In Renal Cell Carcinoma

Background and Objectives:Tumor thrombus of RCC(renal cell carcinoma)with venous tumor thrombus may have unique genomic and biological characteristics compared with primary tumor.We compared efficacy of sorafenib or sunitinib as neoadjuvant targeted therapy between primary tumor and tumor thrombus and performed WES(whole-exome sequencing)and m RNA sequencing on cc RCC(clear cell renal cell carcinoma)with venous tumor thrombus combined with tissue microarray immunochemistry in order to gain insight into characteristics of tumor thrombus.Further we explored function and role of SETD2 in targeted therapy resistance in cc RCC.Materials and Methods:The first part included RCC patients with tumor thrombus in our center from January2007 to December 2014.Wilcoxon signed rank test was used to compare reduction rate between primary tumor and tumor thrombus.Log-rank test was used to compare the survival outcomes of patients with or without neoadjuvant therapy.In the second part,we collected primary tumors,tumor thrombus,paired normal tissue and paired blood cells from 28 cc RCC patients with venous tumor thrombus and tissue microarrays of65 cc RCC patients with venous tumor in our center from January 2007 to December2018 and carried out WES,m RNA sequencing and immunohistochemistry test.In the third part,experiments in vitro were used to explore the role of SETD2 in drug resistance of targeted treatment of cc RCC and the prognostic role of its expression in targeted treatment of metastatic cc RCC was further investigated.Results:1.Reduction rate was significantly lower in tumor thrombus than primary tumor(6.9%vs.10.8%,P = 0.020).RCC with tumor thrombus patients with neoadjuvant therapy had no significantly longer median RFS(30 vs 28 months,P = 0.543)and OS(45 vs42 months,P = 0.939)than those without neoadjuvant therapy.2.A barely detectable statistically significant difference of higher mutation rate and significant lower expression of SETD2 were detected in tumor thrombus than primary tumor,TCGA-KIRC and COSMIC database.Moreover,signature 3(Homologous recombination deficiency)and signature 21(Microsatellite unstable tumors)were specially identified in the tumor thrombus.Platelet activation pathway(P = 0.009)was identified frequently altered in cc RCC tumor thrombus specially.Blood coagulations related pathways and inflammation and immune response related pathways were also significantly enriched in tumor thrombus.3.SETD2 was significantly differently expressed in RCC sunitinib-resistant and paracarcinoma tissue at m RNA and protein level.In vitro,knockdown of SETD2 could lead to low expression of H3K36me3 and could promote sunitinib resistance of RCC cells by activity of PI3K/AKT/mTOR pathway.Multivariate regression analysis confirmed that low SETD2 expression was an independent risk factor for poor prognosis of m RCC patients receiving TKI(Tyrosine Kinase Inhibitor)treatment.Conclusions:1.The response of tumor thrombus to TKI is worse than primary tumor.Neoadjuvant therapy of sorafenib or sunitinib might not improve survival outcomes for high risk RCC patients with tumor thrombus.2.Some heterogeneities have been observed between primary tumor and tumor thrombus in cc RCC.Novel mutated genes,copy number variations,mutational signatures,differential expression genes and enrichment pathways in tumor thrombus compared with primary tumor were identified.Notably,mutation rate of SETD2 was almost significantly higher and expression of SETD2 was significantly lower in tumor thrombus than primary tumor and databases.Further,homologous recombination deficiency and deficient mismatch repair were specially found in tumor thrombus.The plate activity pathways and blood coagulation pathways enrichments also played important roles in the formation of tumor thrombus.These findings provide further insight into the genetic underpinnings of RCC with thrombus development and potential therapeutic implications.3.SETD2 depletion potentiates drug resistance of sunitinib by activating PI3K/AKT/mTOR pathway in cc RCC which might be the reason of resistance to TKI therapy of tumor thrombus.Low expression of SETD2 is an independent risk factor for m RCC patients receiving TKI therapy.