Tenofovir-associated nephrotoxicity in patients with chronic hepatitis B: two cases

Tenofovir disoproxil fumarate (TDF) is effective against chronic hepatitis B (CHB) infection and its use is increasing rapidly worldwide. However, it has been established that TDF is associated with renal toxicity in human immunodeficiency virus-infected patients, while severe or symptomatic TDF-associated nephrotoxicity has rarely been reported in patients with CHB. Here we present two patients with TDF-associated nephrotoxicity who were being treated for CHB infection. The first patient was found to have clinical manifestations of proximal renal tubular dysfunction and histopathologic evidence of acute tubular necrosis at 5 months after starting TDF treatment. The second patient developed acute kidney injury at 17 days after commencing TDF, and he was found to have membranoproliferative glomerulonephritis with acute tubular injury. The renal function improved in both patients after discontinuing TDF. We discuss the risk factors for TDF-associated renal toxicity and present recommendations for monitoring renal function during TDF therapy

3-bromopyruvate and buthionine sulfoximine effectively kill anoikis-resistant hepatocellular carcinoma cells.

Acquisition of anoikis resistance is a prerequisite for metastasis in hepatocellular carcinoma (HCC). However, little is known about how energy metabolism and antioxidant systems are altered in anoikis-resistant (AR) HCC cells. We evaluated anti-tumor effects of a combination treatment of 3-bromopyruvate (3-BP) and buthionine sulfoximine (BSO) in AR HCC cells.We compared glycolysis, reactive oxygen species (ROS) production, and chemoresistance among Huh-BAT, HepG2 HCC cells, and the corresponding AR cells. Expression of hexokinase II, gamma-glutamylcysteine synthetase (rGCS), and epithelial-mesenchymal transition (EMT) markers in AR cells was assessed. Anti-tumor effects of a combination treatment of 3-BP and BSO were evaluated in AR cells and an HCC xenograft mouse model.AR HCC cells showed significantly higher chemoresistance, glycolysis and lower ROS production than attached cells. Expression of hexokinase II, rGCS, and EMT markers was higher in AR HCC cells than attached cells. A combination treatment of 3-BP/BSO effectively suppressed proliferation of AR HCC cells through apoptosis by blocking glycolysis and enhancing ROS levels. In xenograft mouse models, tumor growth derived from AR HCC cells was significantly suppressed in the group treated with 3-BP/BSO compared to the group treated with 3-BP or sorafenib.These results demonstrated that a combination treatment of 3-BP/BSO had a synergistic anti-tumor effect in an AR HCC model. This strategy may be an effective adjuvant therapy for patients with sorafenib-resistant HCC

PI3Kδ Is a Therapeutic Target in Hepatocellular Carcinoma

Class I phosphoinositide 3-kinase (PI3K) signaling is a major pathway in human cancer development and progression. Among the four PI3K isoforms, PI3Kα and PI3Kβ are ubiquitously expressed, whereas PI3Kγ and PI3Kδ are found primarily in leukocytes. Until now, PI3K targeting in solid tumors has focused on inhibiting PI3Kα-mediated and PI3Kβ-mediated cancer cell-intrinsic PI3K activity. The role of PI3Kδ in solid tumors is unknown. Here, we evaluated the effects of PI3Kδ using established hepatocellular carcinoma (HCC) cells, malignant hepatocytes derived from patients with advanced HCC, murine models, and HCC tissues using RNA sequencing, quantitative PCR, immunoblotting, immunofluorescence, microarray, liquid chromatography-tandem mass spectrometry, and kinase assay. We established a chemical carcinogenesis model of liver malignancy that reflects the malignant phenotype and the in vivo environment of advanced HCC. In this in vivo advanced HCC-mimic system using HCC cells treated with hydrogen peroxide (H2 O2 ), we showed that H2 O2 selectively increases PI3Kδ activity while decreasing that of other class I PI3Ks. Blocking PI3Kδ activity with a PI3Kδ inhibitor or small interfering RNA-mediated PI3Kδ gene silencing inhibited HCC-cell proliferation and dampened key features of malignant HCC, including the up-regulation of telomerase reverse transcriptase (TERT). Mechanistically, H2 O2 induced oxidative modification of the serpin peptidase inhibitor, serpin peptidase inhibitor (SERPINA3), blocking its ubiquitin-dependent degradation and enhancing its activity as a transcriptional activator of PI3Kδ and TERT. High PI3Kδ levels in HCC were found to correlate with poor survival rates, with human advanced HCC showing positive correlations between the protein levels of oxidized SERPINA3, PI3Kδ, and TERT. Thus, PI3Kδ plays significant roles in malignant liver tumors. Conclusion: Our data identify PI3Kδ inhibition, recently approved for the treatment of human B-cell malignancies, as a potential treatment for HCC. © 2018 by the American Association for the Study of Liver Diseases

Suppression of HCC invasion by 3-BP, BSO, and a combination treatment.

<p>(A) Invasion capability of AR Huh-BAT cells significantly suppressed by BSO (200 μM), 3-BP (20 μM) and a combination by invasion assay using Boyden chambers (quantified at right panels). (B) Invasion capability of AR HepG2 cells significantly suppressed by BSO (200 μM), 3-BP (20 μM) and a combination by invasion assay using Boyden chambers (quantified at right panels). The concentrations of 3-BP (20 μM) and BSO (200 μM) which did not kill cancer cells were used in this assay. 3-BP and BSO was treated for 72 hours. In a combination treatment in this assay, 3-BP (20 μM) was treated for 72 hours with 24-hour pre-treatment of BSO (200 μM). *<i>P</i><0.05; **<i>P</i><0.01; ***<i>P</i><0.001. Abbreviation: AR, anoikis-resistant; BSO, buthionine sulfoximine; 3-BP, 3-bromopyruvate.</p

Oral Medications Enhance Adherence to Surveillance for Hepatocellular Carcinoma and Survival in Chronic Hepatitis B Patients

<div><p>Background/Aims</p><p>Regular surveillance for hepatocellular carcinoma (HCC) in chronic hepatitis B (CHB) patients is essential to detect HCC earlier and to improve prognosis. This study investigated whether prescription of oral medication contributes to adherence to surveillance, early tumor detection, and overall survival (OS).</p><p>Methods</p><p>A total of 401 CHB patients who were newly diagnosed with HCC were included: 134 patients received no medication (group 1), 151 received hepatoprotective agents such as ursodeoxycholic acid and silymarin (group 2), and 116 received antiviral agents (group 3) at two years before HCC diagnosis. The primary endpoint was OS, and secondary endpoints were compliance to regular surveillance and HCC status at diagnosis.</p><p>Results</p><p>Compared to group 1, both group 2 and 3 had higher rates of good compliance to regular surveillance (defined as participation in >80% of imaging intervals being ≤6 months) (58.2%, 90.1%, and 97.4%, respectively; <i>P</i><0.001), more HCC diagnosed at a very early stage (20.9%, 32.5%, and 36.2%; <i>P</i> = 0.019) and smaller tumor size (2.8±2.4cm, 1.9±1.1cm, and 1.8±0.9cm; <i>P</i><0.001). Finally, compared to group 1, both group 2 (hazard ratio, 0.63; 95% confidence interval, 0.41–0.97; <i>P</i> = 0.035) and group 3 (hazard ratio, 0.40; 95% confidence interval, 0.22–0.71; <i>P</i> = 0.002) had significantly longer OS. In mediation analysis, prolonged OS is resulted considerably from indirect effect mediated by shorter imaging interval (>100% in group 2 and 14.5% in group 3) rather than direct effect of medication itself.</p><p>Conclusions</p><p>Prescription of oral medication improves compliance to surveillance and enables early detection of HCC, which is associated with enhanced survival.</p></div

<i>In vivo</i> anti-tumor effects of 3-BP, sorafenib, and a combination treatment in xenograft nude mice bearing AR Huh-BAT cells.

<p>(A) Tumor growth rates in the combination treatment group were significantly lower than those in the control, sorafenib, or 3-BP treatment group (upper panel). Gross images of tumors before treatment, tumors from the control group, and tumors from the combination treatment group are shown (lower panel). (B) <i>In vivo</i> demonstration of the apoptosis-inducing efficacy in the control, 3-BP, sorafenib, and combination treatment group was shown: H & E and TUNEL staining of tumor tissues in the control, sorafenib, 3-BP, and combination-treated mice (×40 magnification). (C) TUNEL-positive cell percentages (apoptotic index) were determined in six different high power (×400 magnification) fields. (D) There was a significant difference in body weight between the control group and the combination treatment group (<i>P</i><0.05). (E) After matrix detachment, anoikis-resistant cancer cells decrease intracellular ROS levels through inducing enzymes involved in the glycolysis and antioxidant systems for their survival. The Warburg effect can be modulated by increased intracellular ROS levels. Increased ROS levels induce HK II expression and make cancer cells sensitive to 3-BP treatment, and thereby promote cell death via ROS-mediated apoptosis (the black box indicates monocarboxylate transporter-1, and the white box indicates monocarboxylate transporter-4). *<i>P</i><0.05; **<i>P</i><0.01; ***<i>P</i><0.001. Abbreviation: AR, anoikis-resistant; BSO, buthionine sulfoximine; 3-BP, 3-bromopyruvate.</p

Changes of lactic acid levels following 3-BP, BSO, or a combination treatment.

<p>(A) Lactic acid production in Huh-BAT, HepG2, and the corresponding AR cells was significantly increased after BSO treatment compared to the control. (B) Lactic acid production in Huh-BAT, HepG2, and the corresponding AR cells was significantly suppressed after 3-BP treatment compared to the control. (C) Lactic acid production was significantly suppressed after a combination treatment of 3-BP and BSO in AR Huh-BAT and AR HepG2 cells; there was a significant difference among baseline, 2, and 12 hours exposure to the combination treatment. Each assay and western blotting was performed at condition of 3-BP treatment 40 μM, 12 hour; BSO 200 μM, 24 hours; a combination treatment at 3-BP 40 μM, 12 hours after 24 hours pre-exposure of BSO 200 μM. *<i>P</i><0.05; **<i>P</i><0.01; ***<i>P</i><0.001. Abbreviation: AR, anoikis-resistant; BSO, buthionine sulfoximine; CTL, control; HK II, hexokinase II; rGCS, gamma-glutamylcysteine synthetase; ROS, reactive oxygen species; 3-BP, 3-bromopyruvate.</p