Defining the relationship between farsenoid X receptor, hepatitis B virus X protein and hepatocellular carcinoma: It's complicated
Authors: Lindsey Kennedy, Heather Francis
Journal: Hepatology (2016)

Potential conflict of interest: Nothing to report. Portions of this work were supported by (1) a VA Merit Award (1I01BX003031) from the United States Department of Veteran's Affairs, Biomedical Laboratory Research and Development Service and an RO1 from NIH NIDDK (DK108959) and (2) the Dr. Nicholas C. Hightower Centennial Chair of Gastroenterology from Baylor Scott & White Health. This material is the result of work supported with resources and the use of facilities at the Central Texas Veterans Health Care System, Temple, Texas. The content is the responsibility of the author(s) alone and does not necessarily reflect the views or policies of the Department of Veterans Affairs or the United States government. See Article on Page 893 Hepatocelluar carcinoma (HCC) is the second most common cause of cancer‐related death worldwide and is associated (∼50% of cases) with hepatitis B virus (HBV) infection.1 The high rate of metastasis or recurrence, coupled with low chemotherapy response, contributes to the dismal prognosis in HCC patients.3 Hepatocyte transformation is induced by inflammation, DNA damage, senescence, chromosomal instability, and epigenetic alterations.2 Patients with HBV infection have a greater chance of developing HCC, and reports estimate that the risk is increased 25‐fold over patients that are not carrying HBV.2 Hepatitis B virus X protein (HBV‐X or HBx) has been implicated in the progression of HCC and regulates transcriptional gene expression modulating oncogenesis, proliferation, inflammation, and immune responses.4 Patients with C‐terminal truncated HBx, in contrast to full length, have enhanced tumor metastasis and cell invasion.3 Full‐length HBx has also been implicated in HCC, but with contrasting findings. In one study, autotaxin (ATX) transgenic mice, where human α1‐antitrypsin gene promoter controls HBx expression, do not develop spontaneous HCC.5 However, HBx transgenic mice, where HBx is under control of its own promoter elements, have been shown to develop spontaneous HCC, indicating HBx as a promoter of HCC.6 The contrasting role of HBx during HCC progression has been controversial among researchers, and understanding the underlying mechanism whereby HBx either promotes or protects against HCC development is key to unlocking its therapeutic potential. Farsenoid X Receptor Farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily and is an essential transcription factor that regulates liver and intestinal functions. Bile acids are the primary endogenous ligands of FXR and include chenodeoxycholic acid, lithocholic acid, deoxycholic acid, and cholic acid.7 The role of FXR has been mostly defined as hepatoprotective given that knockout of FXR in mice results in cholestasis, nonalcoholic steatohepatitis, and liver tumors as well as colon tumors.4 Studies on the relationship between FXR and HBx are limited, and most discuss their separate roles during HCC progression. In the setting of HCC, 80%‐90% of patients have HBV integration and HBx genes are often partially lost during integration, resulting in C‐terminal truncation of HBx.8 A recent study of in vitro HCC cell lines raised from patients with HBV found that the C‐terminal truncated mutants of HBx have increased capacity to confer the expression of "stemness" genes through deregulation of FXR/retinoid X receptor (RXR) signaling.8 Furthermore, overexpression of HBx truncated mutants increased hepatosphere formation and migration, which was rescued through use of Z‐guggulsterone treatment.8 However, treatment with RXR inhibitor did not rescue these phenotypes, indicating an FXR‐specific event. These unique studies show that truncated HBx deregulates FXR signaling during cancer development and increases stemness, hepatosphere formation, and migration. Summary of Current Findings The finding that HBx is a modulator of HBV‐associated HCC has been demonstrated in numerous studies; however, the concept that HBx‐induced FXR activation decreases tumor incidence is innovative. In this article, Niu et al. demonstrate that full‐length HBx behaves as a transactivator of FXR that, when activated, decreases HCC development.9 The investigators elegantly demonstrate that full‐length HBx enhances FXR binding to its response elements, thereby increasing FXR transcriptional activity and target gene expression. Considering that FXR has been indicated as a regulator of anti‐inflammation, these findings support the argument that full‐length HBx acts as a protective agent during HCC progression through FXR transactivation. To further define HBx‐FXR interaction, the investigators thoroughly describe the shared binding sites between full‐length HBx and FXR. Whereas full‐length HBx seems to be protective against hepatocarcinogenesis through FXR transactivation, the investigators found that C‐terminal truncated HBx did not provide any protective benefits and was unable to bind to or transactivate FXR. In support of these data, Niu et al. found that there was a significant loss of full‐length HBx, but increased positive staining for C‐terminal truncated HBx, in tumors from patients with HBV‐positive HCC, suggesting that the truncated mutant of HBx may be tumorigenic, whereas the full length may play a protective role. This is further supported by previous work demonstrating that truncated HBx is critical to hepatocarcinogenesis.8 When clarifying the protective benefits associated with full‐length HBx‐FXR interaction, the investigators used HBx‐sensitized ATX mice, FXR–/– mice, and ATX‐FXR–/– mice. The investigators found that ATX mice did not develop spontaneous HCC tumors; however, almost half of the ATX‐FXR–/– mice developed tumors by 15 months. Unlike previous studies that have demonstrated that FXR–/– mice develop tumors, the current study found the opposite and explained that this discrepancy may be attributed to different backgrounds of mice when compared to previous reports.7 In the absence of FXR, ATX mice had increased inflammatory cytokine expression, indicating that activation of HBx/FXR signaling may behave as a protective mechanism inhibiting inflammation and tumorigenesis. The investigators further demonstrate that HBx may regulate other players involved in FXR signaling, including small heterodimer partner (SHP). The expression of SHP was absent in ATX‐FXR–/– mice, confirming that loss of SHP may be another mechanism promoting HCC formation in ATX‐FXR–/– mice given that SHP–/– mice develop spontaneous HCC whereas SHP overexpression partially protects FXR–/– mice from HCC formation.10 For every 1 female with HBV‐associated HCC, there are 3 afflicted males, suggesting underlying sex effects that reduce female HCC susceptibility.2 Surprisingly, the investigators herein found that 100% of female ATX‐FXR–/– mice had increased tumor burden compared to 44% of the male ATX‐FXR–/– mice. Interestingly, it has also been noted previously that both male and female FXR–/– mice develop equal numbers of tumors, suggesting that total knockout of FXR interrupts estrogen‐protected pathways.7 In addition, the investigators found that females had significantly higher levels of inflammatory cytokines compared to male ATX‐FXR–/– mice. Given that the authors found that 100% of their ATX‐FXR–/– females developed tumors and it has been demonstrated that total FXR knockout induces tumors in both male and female mice,7 the current study leaves one wondering what might happen if the investigators used ATX mice combined with FXR liver‐specific knockout mice, thus eliminating the detrimental effects of FXR deficiency on estrogen production. Conclusions The study from Niu et al. nicely demonstrates the relationship between either full‐length or C‐terminal truncated HBx and FXR during HBV‐induced HCC. This study provides compelling evidence for the hepatoprotective nature of full‐length HBx through FXR transactivation and further describes the direct binding sites between these two proteins, providing potential therapeutic targets for HCC. The complex relationship between full‐length versus truncated HBx and FXR is a novel and interesting finding of the article, which may help clear the confusion on the role of HBx during hepatocarcinogenesis. Therapies that combine chemotherapy and drugs that target molecules like HBx or FXR may offer promise for advanced treatment strategies, specifically during HBV‐induced HCC. It is obvious that the intricacies of the coactivation of FXR by HBx (truncated or full‐length) should be further examined to shed more light on the potential therapeutic benefits of this viral protein‐nuclear receptor interaction. In addition, further questions should be answered regarding the role of HBx‐activated FXR, including evaluation of other components that might alter tumor incidence. For example, what might happen with HBx‐FXR coactivation where liver functions are altered, such as cholestatic liver injury, bile acid feeding or fatty liver disease? While this paper begins to define the differential properties of full‐length versus truncated HBx during HCC, the exact relationship between these proteins and FXR‐mediated events remains complicated.