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SIRT6, a member of the class III histone deacetylase, has been shown to inhibit glycolysis and promote DNA double strand break repairs. Despite of its proposed tumor suppressor role, no significant differences in SIRT6 mRNA levels among normal bladder urothelium, non-muscle invasive, and muscle invasive urothelial carcinoma were noted in the two largest bladder cancer gene expression datasets available in OncomineTM. We therefore studied the expression and function of SIRT6 in muscle invasive urothelial carcinoma of the bladder. Immunohistochemistry studies of SIRT6 on radical cystectomy samples showed a dramatic decline of SIRT6 expression when bladder cancer progressed from T2 to T4. Functional study with bladder cancer cell lines confirmed its role in inhibiting glycolysis and cell proliferation. Reducing SIRT6 with siRNA, however, did not sensitize bladder cancer cells to drug induced DNA damage. The differential expression patterns of SIRT6 amongst different T stages of muscle invasive bladder cancers indicate less reliance on glycolysis when urothelial carcinoma invades deeper through the bladder and into the adjacent tissues.

Two discrete molecular pathways of bladder carcinogenesis are recognized, one leading to superficial papillary carcinoma and the other to more aggressive either flat (carcinoma in situ) or invasive carcinoma. The majority of aggressive urothelial carcinomas show alterations in the p53 (TP53)-Mdm2-p14 and the retinoblastoma (Rb)-p16 genes and pathways. Both p53 and Rb maintain cellular homeostasis and control normal cell cycle, cellular growth and proliferation. In response to cellular stress, Rb is the main regulator of cell cycle progression, while p53’s main function is to trigger apoptosis or growth arrest in the G1 phase. Tumor angiogenesis also plays a role in urothelial carcinoma progression by providing oxygen, nutrients and growth factors to the neoplastic cells. Distinct genetic events portray the interaction between the molecules involved in these pathways, lending to their use as prognostic indicators. Patients with unaltered wild type p53 and Rb bladder cancers show significant decreased risk of recurrence and mortality when compared to those who have mutational alteration in both p53 and Rb. Alterations of p53 and Rb may help identify patients with high risk superficial cancers more likely to progress to invasive carcinoma or to identify patients who may fail conventional treatment. These patients may benefit from therapies targeting specific altered pathway molecules. Within the past several years, components of these pathways have been shown to be important prognostic and therapeutic response indicators and probable therapeutic targets.

Unlike most other solid tumors, there is limited success of targeted agents in treating metastatic bladder cancer (BC), a lethal disease with a median overall survival of merely 13-15 months. No prevalent “drugable” driver mutations have been identified in muscle invasive bladder cancer (MIBC) so far. Several high profile publications in the past few years have put Sirt6, a member of the class III histone deacetylase in the center stage of cancer metabolism and DNA repair. We therefore assessed the expression and function of Sirt6 in MIBC. Inconsistent with its proposed function as a tumor suppressor in colon cancer, the gene expression datasets available in Oncomine did not reveal significant differences in Sirt6 mRNA levels among normal urothelium, superficial BC and MIBC. Compared to normal urothelium, carcinoma in situ, superficial BC, the median Sirt6 mRNA was the lowest in MIBC in the Dyrkjot bladder 3 dataset, but this was not statistically significant. We then studied Sirt6 protein expression with immunohistochemistry (IHC) in commercial tissue microarray as well as radical cystoprostatectomy samples in our total cancer care tumor bank. Sirt6 IHC were reviewed by pathologist and considered positive for Sirt6 if more than 5% of the tumor cells had at least 1+ Sirt6 nuclear staining. Positive Sirt6 IHC was detected in 33/56 (59%) of T2 (grade 1-3), 2/23 T3 (9%) and 0/3 T4 urothelial carcinoma. All the grade 3/high grade T3 urothelial carcinomas were negative for Sirt6 IHC. Negative Sirt6 IHC was also noted in 2 adenocarcinomas, 1 sarcomatoid carcinoma and 1 small cell MIBC. 1/18 (5%) lymph node metastasis had positive Sirt6 IHC. Consistent with its role of inhibiting aerobic glycolysis, bladder cancer cell lines with higher Srit6 have lower glucose uptake and lower lactate production. Knocking down Sirt6 expression in bladder cancer cell lines, however, did not sensitize bladder cancer lines to drug induced DNA damage. Among MIBC, much higher frequency of Sirt6 IHC positivity was noted in T2 urothelial carcinoma than T3, T4 and non-urothelial carcinomas. Data with bladder cancer cell lines suggest the function of Sirt6 in bladder cancer is primarily inhibiting aerobic glycolysis. Citation Format: Minghui Wu, Shohreh Dickinson, Jingsong Zhang. Expression and function of Sirt6 in muscle invasive bladder cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1590. doi:10.1158/1538-7445.AM2014-1590