EZH1/2 inhibition augments the anti-tumor effects of sorafenib in hepatocellular carcinoma.

Both EZH2 and its homolog EZH1 function as histone H3 Lysine 27 (H3K27) methyltransferases and repress the transcription of target genes. Dysregulation of H3K27 trimethylation (H3K27me3) plays an important role in the development and progression of cancers such as hepatocellular carcinoma (HCC). This study investigated the relationship between the expression of EZH1/2 and the level of H3K27me3 in HCC. Additionally, the role of EZH1/2 in cell growth, tumorigenicity, and resistance to sorafenib were also analyzed. Both the lentiviral knockdown and the pharmacological inhibition of EZH1/2 (UNC1999) diminished the level of H3K27me3 and suppressed cell growth in liver cancer cells, compared with EZH1 or EZH2 single knockdown. Although a significant association was observed between EZH2 expression and H3K27me3 levels in HCC samples, overexpression of EZH1 appeared to contribute to enhanced H3K27me3 levels in some EZH2lowH3K27me3high cases. Akt suppression following sorafenib treatment resulted in an increase of the H3K27me3 levels through a decrease in EZH2 phosphorylation at serine 21. The combined use of sorafenib and UNC1999 exhibited synergistic antitumor effects in vitro and in vivo. Combination treatment canceled the sorafenib-induced enhancement in H3K27me3 levels, indicating that activation of EZH2 function is one of the mechanisms of sorafenib-resistance in HCC. In conclusion, sorafenib plus EZH1/2 inhibitors may comprise a novel therapeutic approach in HCC.

Switching to systemic therapy after locoregional treatment failure: Definition and best timing.

In patients with unresectable hepatocellular carcinoma (HCC) without both macrovascular invasion and extrahepatic metastasis, the initial treatment choice recommended is transarterial chemoembolization (TACE). Before sorafenib came into wide use, TACE had been pointlessly carried out repeatedly. It was in the early 2010s that the concept of TACE refractory was advocated. Two retrospective studies from Japan indicated that conversion from TACE to sorafenib the day after patients were deemed as TACE refractory improved overall survival compared with continued TACE, according to the definition by the Japan Society of Hepatology. Nowadays, phase 3 trials have shown clinical benefits of several novel molecular target agents. Compared with the era of sorafenib, sequential treatments with these molecular target agents have gradually prolonged patients' survival and have become major strategies in patients with HCC. Taking these together, conversion from TACE to systemic therapies at the time of TACE refractory, compared with before, may have a greater impact on survival and may be considered deeper in the decisions-making process in patients with unresectable HCC who are candidate for TACE. Up-to-date information on the concept of TACE refractory is summarized in this review. We believe that the survival of patients with unresectable HCC without both macrovascular invasion and extrahepatic metastasis may be dramatically improved by optimal timing of TACE refractory and switching to systemic therapies.

Sequential therapy with sorafenib and regorafenib for advanced hepatocellular carcinoma: a multicenter retrospective study in Japan.

Background Conversion from sorafenib to regorafenib is primarily an evidence-based treatment strategy in patients with advanced hepatocellular carcinoma (HCC). This study aimed to assess the safety and efficacy of sequential therapy with sorafenib and regorafenib in patients with advanced HCC by analysis of outcomes in clinical practice with the aim to complement phase III findings. Methods The medical records of patients with advanced HCC receiving regorafenib were retrieved to collect data on sorafenib administration at seven Japanese institutions. Radiological responses and adverse events were evaluated using the Response Evaluation Criteria in Solid Tumors version 1.1 and the Common Terminology Criteria for Adverse Events version 4.0, respectively. Results Before March 2018, 44 patients were administered regorafenib for advanced HCC. The median sorafenib treatment duration was 8.4 months. The most common adverse events were similar to those reported by the RESORCE trial. The median overall survival (OS) was 17.3 months (95% confidence interval [CI] 11.4-22.9), and 17 of 37 patients (45.9%) discontinued regorafenib and received sequential systemic therapy after regorafenib. These patients had significantly longer OS than those who were treated by the best supportive care or sub-optimal therapy (not reached versus 8.7 months [95% CI 5.8-11.7]; P < 0.001). Conclusion The results based on Japanese clinical practices verified the tolerability of regorafenib in advanced HCC. Major regorafenib-associated adverse events were similar to those related to sorafenib. OS was significantly longer than expected, which might be associated with the sequential systemic therapies after regorafenib, mainly lenvatinib.

A multicenter Phase II study of sorafenib in Japanese patients with advanced hepatocellular carcinoma and Child Pugh A and B class.

OBJECTIVE: To evaluate prospectively the efficacy and safety of sorafenib, which has been the first-line treatment for advanced hepatocellular carcinoma (HCC), in Japanese HCC patients (pts) with not only Child-Pugh (C-P) A class but also C-P B class. METHODS: Sorafenib was administered orally at the dose of 400 mg twice daily for pts with HCC and liver function of C-P score of 5-8. Administration was continued until the detection of disease progression or appearance of unacceptable toxicity. The primary endpoint was time to progression (TTP), and toxicity and the secondary endpoints included objective response, overall survival (OS). RESULTS: Forty C-P A pts and 12 C-P B pts were enrolled. The median TTP in the C-P A pts and C-P B pts was 3.3 months and 3.2 months, respectively. Among the pts with C-P A, complete response, partial response, and stable disease were achieved for 2.5%, 7.5% and 47.5%. Among the pts with C-P B, there were no treatment responses, 66.7% of pts had stable disease. The median OS in the C-P A pts and C-P B pts was 13.4 months and 7.4 months, respectively. With regard to toxicities, fewer C-P A pts experienced Grade 3/4 toxicities than C-P B pts (77.5% vs. 91.6%). There were no treatment-related deaths in either group of patients. CONCLUSIONS: This study shows sorafenib has similar effectiveness in the recent post-approval studies and is well-tolerated in Japanese pts with HCC and Child Pugh A class. Sorafenib should be used with great care for Child Pugh class B pts.

Characteristics of patients with sorafenib-treated advanced hepatocellular carcinoma eligible for second-line treatment.

Background Regorafenib has been investigated for its efficacy and safety as a second-line treatment in patients with advanced hepatocellular carcinoma (HCC). We assessed the characteristics of patients with HCC treated with sorafenib who might be eligible for second-line treatment in general and regorafenib in particular. Methods Patients with HCC treated with sorafenib were retrospectively analyzed. We defined second-line candidate patients as maintaining Child-Pugh A and ECOG-PS ≤1 at the time of sorafenib failure. We also defined regorafenib candidate patients as follows: 1) continuing sorafenib at the time of radiological progression, 2) maintaining Child-Pugh A and ECOG-PS ≤ 1 at the time of sorafenib failure, and 3) continuing sorafenib 400 mg or more without intolerable adverse events at least 20 days of the last 28 days of treatment. Results Of 185 patients, 130 (70%) and 69 (37%) were candidates for second-line treatment and regorafenib. Child-Pugh score 6 and ECOG-PS 1 at the time of starting sorafenib were significantly lower in both second-line treatment and regorafenib candidate patients. Moreover, hand-foot skin reaction and liver failure during sorafenib treatment were associated with significantly low and high probabilities, respectively, of both Child-Pugh score > 6 and ECOG-PS > 1 at the time of sorafenib failure. Conclusion Regorafenib candidate patients after sorafenib failure are limited, and generally fewer than those who are candidates for second-line treatment. A lower Child-Pugh score and a better ECOG-PS were predictors of eligibility for second-line therapy and regorafenib treatment in sorafenib-treated patients with advanced HCC patients.

Analysis of Sorafenib Outcome: Focusing on the Clinical Course in Patients with Hepatocellular Carcinoma.

BACKGROUND: Treatment outcomes of sorafenib therapy may greatly vary depending not only on tumor spread but also on past clinical processes prior to sorafenib therapy and timing of sorafenib administration in the past clinical course of hepatocellular carcinoma (HCC). We evaluated the efficacy of sorafenib in patients with HCC, taking into account of their past clinical courses. METHODS: Patients with HCC treated with sorafenib as a first-line systemic therapy, whose courses documented from the time of the initial diagnosis, were retrospectively analyzed. RESULTS: Of the 123 patients receiving sorafenib therapy at an advanced-stage, baseline characteristics differed including the rate of hepatitis C virus, Child-Pugh class, and status of intrahepatic lesions according to stage progression processes. Overall survival (OS) in patients progressed directly from the early-stage (15.3 months) was significantly longer than that in patients diagnosed at the advanced-stage (5.3 months, P = 0.022) and progressed from the intermediate-stages (6.0 months, P = 0.041). Of 105 patients diagnosed at the intermediate-stage on past clinical courses, OS of starting sorafenib therapy before progression to the advanced-stage (67 patients) was significantly longer than for patients starting sorafenib therapy only after progression to the advanced-stage (38 patients) (P = 0.015). CONCLUSION: Characteristic differences between past stage progression processes might affect prognosis in advanced-stage HCC patients receiving sorafenib. Switching to sorafenib therapy before progression to the advanced-stage appears more effective than that after progression to the advanced-stage in patients diagnosed in the intermediate-stage on past clinical courses prior to sorafenib administration.

Post-progression survival in patients with advanced hepatocellular carcinoma resistant to sorafenib.

BACKGROUND: Since the approval of sorafenib, no other agent has been proven to show survival benefits in clinical trials involving patients with advanced hepatocellular carcinoma (HCC) resistant to sorafenib. Prognostic factors for survival after tumor progression in sorafenib-treated patients are critical for designing second-line trials. METHODS: To determine the factors affecting the post-progression survival (PPS) after sorafenib treatment, additional analyses were conducted using fixed data obtained from our previous prospective study. Data on patients with advanced HCC treated with sorafenib were analyzed in view of patient characteristics at the time of tumor progression and the progression pattern (intra-/extrahepatic growth or emergence of new intra-/extrahepatic lesions). RESULTS: Of the 89 enrolled patients, 70 were diagnosed with disease progression according to the Response Evaluation Criteria in Solid Tumors version 1.1. Multivariate Cox's regression analysis revealed that Child-Pugh scores of ≥7, macrovascular invasion (MVI), and alpha-fetoprotein of >400 ng/mL were independent predictors of poor PPS. Although both extrahepatic metastasis (EHM) and MVI were characteristics of advanced HCC, EHM was not determined as a prognostic factor. Additionally, the emergence of new extrahepatic lesions also served as an independent indicator of a poor prognosis. The PPS of the patients was well stratified according to the index based on the sum of these prognostic factors, ranging from 0 to 4. CONCLUSIONS: Child-Pugh score of ≥7, AFP of >400 ng/mL, MVI, and new extrahepatic lesions at the time of progression may be utilized to assess the prognosis and taken into consideration when designing second-line trials.

Liver function assessment according to the Albumin-Bilirubin (ALBI) grade in sorafenib-treated patients with advanced hepatocellular carcinoma.

BACKGROUND: The Albumin-Bilirubin (ALBI) grade has been proposed as a new, simple, and objective method of assessing liver function. However, there is lack of data in sorafenib-treated patients with advanced hepatocellular carcinoma (HCC). METHODS: We evaluated the correlations between the ALBI grade and Child-Pugh score, adverse events, and survival in 89 patients with advanced HCC who were prospectively treated with sorafenib. RESULTS: Majority of patients with ALBI grade 1 (14/15 patients, 93%) had a Child-Pugh score of 5. Patients with ALBI grade 2 had a wide range of liver function according to the Child-Pugh scores, with scores of 5, 6, 7, and ≥ 8. We divided ALBI grade 2 patients into ALBI grade 2A and 2B groups according to the median ALBI score among patients with ALBI grade 2. Although no significant difference was observed, the incidence of liver dysfunction in sorafenib-treated patients with ALBI grades 1, 2A, and 2B was 7%, 19%, and 35%, respectively. Overall survival in the ALBI grade 2B group was significantly shorter than that in the ALBI grade 1 and 2A groups. Thus, ALBI grade 2B was an independent predictor of poor prognosis in addition to elevated serum aspartate aminotransferase levels, increased serum alpha-fetoprotein level, and macrovascular invasion. CONCLUSION: Sorafenib may be indicated for all patients with advanced HCC and ALBI grade 1 and for some with ALBI grade 2. The subdivision of patients with ALBI grade 2 increases the utility of ALBI in identifying patients indicated for sorafenib therapy.

Sorafenib treatment in Child-Pugh A and B patients with advanced hepatocellular carcinoma: safety, efficacy and prognostic factors.

BACKGROUND: We aimed to evaluate the safety, efficacy and prognostic impact of baseline and early clinical markers in both Child-Pugh A and B patients with advanced hepatocellular carcinoma (HCC). METHODS: We prospectively studied 89 Japanese patients with HCC (Child-Pugh A, n = 59; Child-Pugh B, n = 30) who were started with sorafenib between May 2010 and July 2013. RESULTS: Frequency of sorafenib-related adverse events was almost similar between Child-Pugh score 5, 6, and 7 patients. The rate of liver dysfunction, including any grade encephalopathy, ≥ grade 3 ascites, or ≥ grade 3 bilirubin increased, in Child-Pugh score ≥8 group was significantly higher than that in the other groups. The median overall survival of Child-Pugh score 5, 6, 7 and ≥8 patients were 14.5, 11.1, 8.7 and 4.6 months, respectively. Patients in Child-Pugh score 6 had significantly longer OS than those in Child-Pugh score 7 (P = 0.049). Multivariate analysis identified macrovascular invasion (MVI), alpha-fetoprotein (AFP), Child-Pugh score and aspartate aminotransferase (AST) as baseline predictors of survival. However, extrahepatic metastasis (EHM) was not a significant prognostic factor. In addition, decrease in AFP level and development of hand-foot skin reaction within 4 weeks after sorafenib initiation were closely associated with favorable survival. CONCLUSION: It is possible that not only Child-Pugh score 5 and 6 but also 7 patients are eligible for future clinical trials with sorafenib or similar drugs. Various survival predictors identified in this study might be considered as stratification factor. Although both MVI and EHM is a phenotype of advanced HCC, MVI should be discriminated from EHM because of the prognostic impact on survival in sorafenib-treated advanced HCC patients.

Efficacy of sorafenib in intermediate-stage hepatocellular carcinoma patients refractory to transarterial chemoembolization.

OBJECTIVE: We compared the benefits of sorafenib therapy with continued transarterial chemoembolization (TACE) in TACE-refractory patients with intermediate-stage hepatocellular carcinoma (HCC). METHODS: This retrospective study reviewed intermediate-stage HCC patients who underwent the first TACE. Patients were defined as TACE-refractory and divided into two cohorts: (1) patients who switched from TACE to sorafenib and (2) those who continued TACE. We evaluated the patient overall survival (OS) and time to disease progression (TTDP; the time patients reached Child-Pugh C or developed advanced-stage HCC). RESULTS: A total of 509 patients with HCC underwent TACE. Of 249 intermediate-stage HCC patients undergoing the first TACE, 122 were deemed refractory. At the time they were identified as refractory, 20 patients converted to sorafenib, whereas 36 patients continued TACE. We excluded patients with Child-Pugh scores of ≥ 8, those with advanced-stage HCC, those who had undergone hepatic arterial infusion chemotherapy or other systemic therapy, and those treated with best supportive care alone. The median TTDP and OS were 22.3 and 25.4 months, respectively, in the conversion group, and 7.7 and 11.5 months, respectively, in the continued group (p = 0.001 and p = 0.003, respectively). CONCLUSIONS: It is possible that sorafenib conversion might prolong OS and TTDP in TACE-refractory patients with intermediate-stage HCC.

A phase I/II trial of capecitabine combined with peginterferon α-2a in Patients with sorafenib-refractory advanced hepatocellular carcinoma.

BACKGROUND: Several pilot studies have demonstrated the effectiveness of combination therapy with pyrimidine fluoride and interferon for advanced hepatocellular carcinoma.This study aimed to determine the recommended dose of capecitabine combined with peginterferon α-2a (Phase I) and evaluate its safety and efficacy for sorafenib-refractory advanced hepatocellular carcinoma (Phase II). METHODS: Capecitabine was administered daily on days 1-14, while peginterferon α-2a was administered on days 1, 8, and 15. The cycle was repeated every 21 days. The patients were scheduled to receive capecitabine [mg/(m(2)∙day)] and peginterferon α-2a (µg/week) at 3 dose levels in phase I: 1200 and 90 (level 1), 1600 and 90 (level 2), and 2000 and 90 (level 3), respectively. RESULTS: A total of 30 patients were enrolled. The recommended dose was level 3. Among the 24 patients receiving the drug at the recommended dosage, 2 (8 %) exhibited a partial response, 9 (38 %) exhibited stable disease, 10 (42 %) exhibited progressive disease, and 3 (13 %) were not evaluated. The median time to progression and overall survival were 3.0 months and 7.2 months, respectively. The most common toxicities were decreased white blood cell (88 %), neutrophil (88 %), and platelet counts (58 %); fatigue (50 %); and palmar-plantar erythrodysesthesia syndrome (42 %). Four patients (17 %) discontinued treatment because of severe adverse events. CONCLUSION: Capecitabine at 2000 mg/(m(2)∙day) combined with peginterferon α-2a (90 µg/week) exhibited moderate, albeit manageable, toxicity and was declared as the recommended phase II dose. Further research is required to refine the efficacy of this combination.

A phase I/II study of S-1 with sorafenib in patients with advanced hepatocellular carcinoma.

BACKGROUND: Sorafenib is the sole molecular-targeted agent showing a survival benefit in patients with advanced hepatocellular carcinoma (HCC). We evaluated the tolerability and effectiveness of a combination of S-1 with sorafenib in patients with advanced HCC. METHODS: S-1 was administered during days 1-14 and sorafenib was administered every day. This treatment was repeated every 21 days. In phase I, we determined the maximum tolerated dose (MTD) and dose-limiting toxicities (DLTs). The dose of each drug was planned as follows: cohort 1: S-1 48 mg/m(2)/day and sorafenib 400 mg/day, cohort 2a: S-1 48 mg/m(2)/day and sorafenib 800 mg/day, cohort 2b: S-1 64 mg/m(2)/day and sorafenib 400 mg/day, cohort 3: S-1 64 mg/m(2)/day and sorafenib 800 mg/day, and cohort 4: S-1 80 mg/m(2)/day and sorafenib 800 mg/day. In phase II, the patients were treated at the MTD to evaluate safety and efficacy. RESULTS: Nineteen patients were enrolled in phase I. One of the six patients in cohort 1 and one of the six patients in cohort 3 experienced DLT. None of the three patients in cohort 2a experienced DLT and three of the four patients in cohort 4 experienced DLT. Therefore, cohort 3 was considered the MTD. Subsequently, 26 patients were enrolled in phase II. The most common grade 3/4 toxicities were an increase of aspartate aminotransferase (38.5 %), thrombocytopenia (23.1 %), neutropenia (19.2 %), hyperbilirubinemia (15.4 %), an increase of alanine aminotransferase (15.4 %), hyponatremia (11.5 %), rash (11.5 %), and hypophosphatemia (11.5 %). Sudden death occurred in one patient (3.8 %). A patient (3.8 %) had a partial response, 15 (57.7 %) had stable disease, and 10 (38.5 %) had progressive disease. The median times to progression and overall survival were 2.4 and 10.5 months, respectively. CONCLUSION: The MTD of S-1 and sorafenib in patients with advanced HCC was 64 mg/m(2)/day and 800 mg/day, respectively. This dose/regimen demonstrated substantial clinical activity among patients with advanced HCC.

Phase I study of combination chemotherapy using sorafenib and transcatheter arterial infusion with cisplatin for advanced hepatocellular carcinoma.

The aims of this study were to evaluate the frequency of dose-limiting toxicities and to find the recommended dose of combination chemotherapy with sorafenib and transcatheter arterial infusion (TAI) using cisplatin for patients with advanced hepatocellular carcinoma (HCC), for whom surgical resection, local ablation therapy, or transcatheter arterial chemoembolization were not indicated. Patients received 800 mg sorafenib daily. Cisplatin was given at one of three dosages (level 1, 35 mg/m(2) /cycle; level 2, 50 mg/m(2) /cycle; and level 3, 65 mg/m(2) /cycle) from feeding arteries to the HCC. The treatment was repeated every 4-6 weeks up to a maximum of six cycles, until there were signs of tumor progression or unacceptable toxicity. The dose-limiting toxicities experienced by the 20 enrolled patients were grade 4 increased aspartate aminotransferase at level 1, grade 3 gastrointestinal hemorrhaging at level 1, and grade 3 hypertension at level 3. The common drug-related adverse events that were of severity grade 3 or 4 included the elevation of aspartate aminotransferase (30%), alanine aminotransferase (20%), amylase (30%), and lipase (30%). Partial response was seen in four patients (20%), and 13 patients (65%) had stable disease. The median overall survival and progression-free survival were 9.1 and 3.3 months, respectively. The combination of sorafenib at 800 mg/day with TAI of cisplatin at 65 mg/m(2) /cycle was determined to be the recommended regimen. A randomized phase II trial of sorafenib alone versus sorafenib plus TAI of cisplatin is currently underway. This study was registered at UMIN as trial number UMIN000001496.

Efficacy of transcatheter arterial chemoembolization with miriplatin-lipiodol water-soluble contrast agent emulsion in patients with hepatocellular carcinoma.

AIM: To evaluate the therapeutic efficacy of transcatheter arterial chemoembolization (TACE) using miriplatin emulsion in unresectable hepatocellular carcinoma (HCC). PATIENTS AND METHODS: The efficacy of TACE was evaluated by dynamic computed tomography or magnetic resonance imaging three months after TACE, according to the Response Evaluation Criteria in Cancer Study Group of Japan (RECICL). Adverse events were assessed using Common Terminology Criteria, version 4.0. RESULTS: Eighteen patients with 48 lesions received TACE with miriplatin-lipiodol (LPD) suspension (miriplatin suspension) and 53 patients with 114 HCC tumors received TACE with miriplatin-LPD water-soluble contrast agent emulsion (miriplatin emulsion). TACE with miriplatin emulsion enabled for administration of a higher dose of miriplatin compared to TACE with miriplatin suspension (p=0.016), although there were no significant differences in the frequency of adverse events between the two groups. The treatment effect per tumor was significantly higher in the emulsion group than in the suspension group (p=0.001). The time-to-progression per tumor was significantly shorter in the suspension group than in the emulsion group (p=0.001). CONCLUSION: TACE with miriplatin emulsion is more effective than that with miriplatin suspension.

Rapid identification of ligand-binding sites by using an assignment-free NMR approach.

In this study, we developed an assignment-free approach for rapid identification of ligand-binding sites in target proteins by using NMR. With a sophisticated cell-free stable isotope-labeling procedure that introduces (15)N- or (13)C-labels to specific atoms of target proteins, this approach requires only a single series of ligand titrations with labeled targets. Using titration data, ligand-binding sites in the target protein can be identified without time-consuming assignment procedures. We demonstrated the feasibility of this approach by using structurally well-characterized interactions between mitogen-activated protein (MAP) kinase p38α and its inhibitor 2-amino-3-benzyloxypyridine. Furthermore, we confirmed the recently proposed fatty acid binding to p38α and confirmed the fatty acid-binding site in the MAP kinase insert region.

31P NMR studies of energy metabolism in xanthosine-5'-monophosphate overproducing Corynebacterium ammoniagenes.

Corynebacterium ammoniagenes is an overproducer of xanthosine-5'-monophosphate (XMP) by consuming either glcose (glc) or glutamic acid (glu). Its energy metabolism was studied in vivo using 31P NMR spectroscopy coupled with a circulating fermentation system (CFS). CFS enabled us to validate directly the cellular dependency on carbon sources and changes in biomolecules produced according to alterations in the cellular energetic status. For the most efficient XMP production, the glutamic acid and glcose molar ratios (glu/glc) in the medium were adjusted to a molar ratio of 0.31. The 31P NMR illustrated the two distinct phases of the cellular energetic status due to the availability of the substrates from the medium. In the earlier phase, both glc and glu were utilized, resulting in average ATP and ADP concentrations in cells of 0.50 +/- 0.17 micro mol.g-1 of dry cell weight (DCW) and an undetermined level, respectively. The ADP concentration in the later phase increased to 2.15 +/- 1.30 micro mol.g-1 of DCW, while the ATP concentration decreased to an undetectable level in association with a remarkable decrease in XMP production. This decrease in the XMP-producing ability was associated with an increase in production of the by-product hypoxanthine. Because glu was found to be consumed completely during the earlier phase, glc was the only available substrate in the later phases. These findings by in vivo NMR indicate that changes in the carbon metabolism profoundly affect XMP production by C. ammoniagenes.