In this study, we demonstrate a positive correlation between immunohistochemical expression of FAM83H and ZNF16 in gallbladder cancers. There was also a close association between the expression of FAM83H and ZNF16 and advanced clinicopathological factors. Moreover, the positivity for the expression of nuclear FAM83H, cytoplasmic FAM83H, nuclear ZNF16, and cytoplasmic ZNF16 were significantly associated with shorter survival of gallbladder carcinoma patients. Especially, nuclear FAM83H expression was an independent marker of poor prognosis of gallbladder carcinoma patients. Concerning the subcellular localization of FAM83H, FAM83H was present in the cytoplasmic membrane and the cytosol [3, 4, 23]. Subsequently, it was shown that the nuclear expression of FAM83H is present in some cases of colon carcinomas, and its nuclear localization has been suggested to involved in tumorigenesis . Furthermore, in hepatocellular carcinoma and clear cell renal cell carcinoma, nuclear FAM83H expression was an independent prognostic indicator of cancer patients [5, 8]. In addition, FAM83H had cooperative roles with MYC and Wnt/β-catenin pathways in cancer progression [5, 6]. MYC transcriptionally controlled the expression of FAM83H and FAM83H is involved in the stabilization of β-catenin and consequent transcriptional activation of the canonical Wnt pathway. Therefore, when considering the roles of the nuclear localization of MYC and β-catenin, nuclear FAM83H also has a role in the nuclei of cells in association with MYC/β-catenin. In addition, FAM83H localized to nuclear speckles and interacted with SON, a protein in nuclear speckle, and FAM83H is involved in nuclear recruitment of casein kinase 1α in colorectal cancer cells . Therefore, it has been suggested that FAM83H might be involved in cancer progression via extensive interaction with nuclear proteins. However, in contrast, cytoplasmic FAM83H expression was more predictive than nuclear FAM83H expression for the survival of osteosarcoma patients . Therefore, although the prognostic significance of FAM83H expression varied according to its subcellular localization with different cancer type, it has been suggested that the overall expression of FAM83H in the cell could be vital in the progression of cancers because knock-down of FAM83H suppressed cancer progression and overexpression of FAM83H stimulated cancer progression [5, 6]. However, further study is needed to clarify the mechanism and role of nuclear localization of FAM83H.
With regards to the subcellular localization of ZNF16, our results showed ZNF16 expression in both the cytoplasm and nuclei of tumor cells. Furthermore, the nuclear expression of ZNF16 was indicated to be a potential prognostic factor of gallbladder carcinoma patients. Supportively, it has been reported that ZNF16 is expressed in both the cytoplasm and nucleus and nuclear ZNF16 has an important role in the differentiation of cells [11, 25]. In addition, ZNF16 inhibited apoptosis and stimulated cell cycle progression by inhibiting INCA1 in K562 leukemia cells . Therefore, when considering our finding that there is a significant association between nuclear ZNF16-positivity and higher tumor stage and histologic grade, nuclear expression of ZNF16 might be significantly involved in the progression of gallbladder carcinomas. However, the reports concerning the role of ZNF16 in human cancers, especially with regards to the prognostic significance of ZNF16 expression in human cancers, is limited. Therefore, we have searched the public database. In a search of the GEPIA database (Accessed 2 March 2020) , the mRNA expression of ZNF16 was elevated in breast cancer, cholangiocarcinoma, esophageal carcinoma, colon adenocarcinoma, head and neck squamous cell carcinoma, prostatic cancer and lung carcinoma compared with their normal counterpart tissues. In addition, although there is no data for gallbladder cancers, The Human Protein Atlas database (https://www.proteinatlas.org. Accessed 2 March 2020)  indicates that higher expression of ZNF16 mRNA is an indicator of poor prognosis of liver cancer (Log-rank, p = 0.008) and breast cancer (Log-rank, p = 0.041). However, higher expression of ZNF16 mRNA was associated with favorable prognosis of head and neck cancer (Log-rank, p < 0.001) and gastric cancer (Log-rank, p = 0.017) . Therefore, further study is needed to clarify the role of ZNF16 in the progression of human cancers.
Another interesting finding of our study is that the expression of FAM83H and ZNF16 were closely associated with each other in gallbladder cancers. Furthermore, the co-expression pattern of nFAM83H/nZNF16 was a strong prognostic indicator of gallbladder carcinoma patients. Although no reports have investigated the relationship between FAM83H and ZNF16 in human cancers, we found a significant association between FAM83H and ZNF16 expression in human cancers in our search of the public database. Although there is no data for gallbladder cancers, a significant correlation between the expression of FAM83H and ZNF16 was seen in hepatobiliary cancers. The cBioPortal database (Accessed March 2, 2020) [12, 13] indicates that there is a significant correlation between the expression of mRNA of FAM83H and ZNF16 in cholangiocarcinoma (Spearman’s correlation, R = 0.70, p < 0.001). The GEPIA database (Accessed March 2, 2020)  showed a significant correlation between the expression of mRNA of FAM83H and ZNF16 in cholangiocarcinoma (Spearman’s correlation, R = 0.82, p < 0.001), hepatocellular carcinoma (Spearman’s correlation, R = 0.60, p < 0.001), and pancreatic adenocarcinoma (Spearman’s correlation, R = 0.40, p < 0.001). Moreover, the co-expression patterns of nuclear FAM83H and nuclear ZNF16 were independent indicators of poor prognosis of gallbladder carcinoma patients. Therefore, our results suggest that FAM83H and ZNF16 are cooperatively involved in the progression of gallbladder cancers. However, the exact mechanism of the relationship between FAM83H and ZNF16 is unclear. Therefore, further study of the mechanism(s) by which FAM83H/ZNF16 are involved in gallbladder cancer progression is needed.