Transitional cell carcinoma (TCC) is the one of the most commonly observed types of cancer globally. The identification of novel disease-associated genes in TCC has had a significant effect on the diagnosis and treatment of bladder cancer; however, there may be a large number of novel genes that have not been identified. In the present study, the exomes of two individuals who were diagnosed with muscle-invasive TCC (MI-TCC) were sequenced to investigate potential variants. Subsequently, following algorithm and filter analysis, Sanger sequencing was used to validate the results of deep sequencing. Immunohistochemistry (IHC) was employed to observe the differences in HECT, C2 and WW domain-containing E3 ubiquitin protein ligase 1 (HECW1) protein expression between tumor tissues and para-carcinoma tissues. A total of 6 nonsynonymous mutation genes were identified in MI-TCC, identified as copine VII, RNA binding motif protein, X-linked-like 3, acyl-CoA synthetase medium-chain family member 2A, HECW1, zinc finger protein 273 and trichohyalin. Furthermore, 5 cases were identified to possess a HECW1 gene mutation in 61 MI-TCC specimens, and all of these were point mutations located at exon 11 on chromosome 7. The mutation categories of HECW1 had 4 missense mutations and 1 nonsense mutation. IHC revealed that HECW1 protein was expressed at significantly increased levels in MI-TCC compared with normal bladder urothelium (P<0.001). The present study provided a novel approach for investigating genetic changes in the MI-TCC exome, and identified the novel mutant gene HECW1, which may possess a significant role in the pathogenesis of TCC.