Integrative computational and experimental characterisation of endometrial cancer alternations

Abstract

Endometrial cancer, the fourth most common female cancer in Hong Kong, warrants further research efforts to understand its genetic aetiology. The revolutionary reclassification of endometrial cancer proposed by The Cancer Genome Atlas completely changed the way to see endometrial cancer. The huge number of genetic alternations identified by omics approach urges for more precise computational analysis to identify true key players contributing to the development of endometrial cancer and assist experimental verification. About 500 endometrial cancer transcriptome and exome sequence data were acquired from The Cancer Genome Atlas. 244 potential oncogenes and 73 potential tumour suppressor genes were identified from the omics data. The many known oncogenes appearing on the potential oncogenes list and tumour suppressor genes on its respective list demonstrated the robustness of such computational method. ZNF555 and KIAA1462 from the potential tumour suppressor list were picked up for further studies. These genes were significantly downregulated in tumours and displayed low expression level in the aggressive copy number high group of endometrial cancer. Knowing how these genes can contribute to cancer development may give direction to curing the aggressive subtype. Gene set variation analysis revealed positive correlation of KIAA1462 expression to a number of oncogenic functions. ZNF555 was knocked down by shRNA in MFE-296 cells and such knockdown granted higher tumourigenicity to the cells in both anchorage-dependent and -independent colony formation assay. Metastasis capability was found to have increased upon ZNF555 knockdown in in-vitro migration assay. Xenograft model was employed to verify the loss of tumour suppressive effect upon ZNF555 and the results agreed with in-vitro evidence. shRNA knockdown of KIAA1462 in HEC-1-B cells leaded to a decrease in expression of Hippo signalling pathway product CTGF and BIRC5. 8XGTIIC luciferase reporter was used to provide extra evidence towards showing the role of KIAA1462 in Hippo signalling pathway. Luciferase reporter assay in MFE-296 cells upon knockdown of KIAA1462 showed a decline in Hippo signalling pathway transcription activity. Overexpression of KIAA1462 in the high expression HEC-1-B cells failed to produce higher transcriptional activity. Yet, overexpression of KIAA1462 in the low expression MFE-296 cells rendered an uprising of Hippo signalling pathway transcriptional activity. The protein product of KIAA1462, JCAD, is known to have physical interaction with Hippo signalling pathway core kinase LATS2. Experimental results in this study suggest JCAD is a negative regulator to LATS2 function and implying oncogenic role of KIAA1462 in endometrial cancer. This study identified ZNF555 as a potential tumour suppressor gene in endometrial cancer by computation and experimental methods. There is computational evidence suggesting KIAA1462 could be tumour suppressor gene or oncogene while experimental evidence showed KIAA1462 acts as oncogene in endometrial cancer.