Direct-acting Antiviral Therapy Improves the Outcome of Chronic Hepatitis C/intermediate-stage B Hepatocellular Carcinoma Patients

Background/Aim: The benefit of direct-acting antiviral therapy (DAA) in chronic hepatitis C (CHC) infected patients who received curative treatment for early-stage hepatocellular carcinoma (HCC) is well known, but is unclear for intermediate stage HCC. Patients and Methods: CHC patients with Barcelona Clinic Liver Cancer (BCLC) stage B HCC receiving chemoembolization were identified. Univariate, multivariate analyses, and Kaplan–Meier curve were used to identify factors associated with survival outcomes. Results: Among 113 included patients, the median survival of DAA treated group (n=14) and non-treated group (n=99) were 40.1 months and 22.9 months, respectively. Multivariate analysis showed that Eastern Cooperative Oncology Group (ECOG) score, DAA, and serum albumin were key independent factors associated with overall survival. Moreover, the time-to-complete remission (TTCR) was improved in the DAA treated group. Conclusion: ECOG, DAA, and serum albumin were prognostic factors for CHC/intermediate-stage HCC patients. DAA was also a beneficial factor for TTCR.

Primary liver cancer ranks sixth in cancer incidence (841,000 new cases) and fourth in cancer-related mortality (781,000 cases) globally. Hepatocellular carcinoma (HCC) is the most common primary liver cancer and comprises over 80% of all liver tumors. Several known risk factors are linked to development of HCC including chronic viral infection with either hepatitis B virus (HBV) or hepatitis C virus (HCV), heavy alcohol intake, type 2 diabetes, obesity/fatty liver disease, aflatoxin contaminated foodstuffs, hereditary hemochromatosis, alpha1-antitrypsin deficiency, autoimmune hepatitis, some porphyrias, and Wilson's disease (2). These diseases also lead to hepatic fibrosis and cirrhosis. Furthermore, pre-existing cirrhosis is observed in over 80% of HCC patients.
The Barcelona Clinic Liver Cancer (BCLC) staging classification demonstrates the best predictive efficacy for HCC survival and is used as the universal practice guideline to determine the HCC treatment strategy. For BCLC stage B patients, transcatheter arterial chemoembolization (TACE) is one of the common treatments.
Chronic hepatitis C (CHC) infection is the second most common cause of HCC globally, which accounts for 10~25% of cases in Taiwan, and is the most common cause of HCC in Western countries. There are over 184 million people infected with HCV worldwide. CHC is associated with 20~30-fold increased risk for HCC. For HCV related cirrhosis, the annual incidence of HCC rate is around 3-5% and the 5-year cumulative risk ranges from 5~30%. Studies have shown that liver related morbidity and mortality can be reduced after CHC patients have achieved sustained viral response (SVR). As medicine advances, HCV treatment has evolved from pegylated interferon (Peg-IFN) to direct-acting antiviral (DAA) combination regimens which can achieve over 95% SVR with a low number of adverse events. Current guidelines recommend antiviral treatment for all CHC  Doxorubicin 20-60 mg (depending on the tumor size) mixed with 5-20 ml of lipiodol emulsion was infused through the target artery. Further, Gelfoam injection for target artery embolization was performed until no tumor stain was seen on angiography.
TACE treatment response and follow-up. Multiphase CT/MRI was performed 6-10 weeks after each TACE treatment. Treatment response was evaluated using the modified Response Evaluation Criteria in Solid Tumors (mRECIST). Patients were followed until the time of death or loss to follow-up. For patient with complete response after TACE, serum alpha-fetoprotein (AFP) and abdominal ultrasound were monitored every 3 months. In the presence of newly suspected lesions or elevated AFP, further CT/MRI were performed for diagnosis of HCC recurrence.
DAA treatment. Depending on the DAA availability in Taiwan, different regimens were used. Achieving SVR was defined as undetectable HCV RNA at 12 weeks after completing DAA treatment.
Statistical analysis. Statistical analyses were performed using IBM SPSS statistic software, version 23 (IBM Corporation, Armonk, NY, USA). For the descriptive statistic, categorical data were expressed in number and percentage, whereas the continuous data were presented in mean with standard deviation or median with range. Median overall survival (OS) was calculated in months. Kaplan-Meier curve was used to evaluate the survival curves. Relevant baseline variables were analyzed using Cox regression hazard model to identify the factors associating with OS, time-to-complete response (TTCR), and progression-free survival (PFS). Statistically significant variables (p<0.05) in univariate analysis were further included in the multivariate analysis.

Results
Baseline characteristics of patients. A total of 113 CHC patients who were also diagnosed as HCC BCLC stage B, were included in the study. The basic characteristics of patients are listed in Table I. The mean age was 67.2±9.3 years, and 76 of the patients were males. The group was predominantly cirrhotic Child-Pugh A (103/113, 91.2%). Sixty patients had tumor number ≥3, largest tumor size mean of 3.2±1.7 cm. All patients had received ongoing TACEs for HCC as the standard of care according the BCLC guideline, 14 patients also received DAA treatment for HCV eradication.
Subgroup characteristics. We subdivided the 113 patients into two groups, DAA treatment group and non-treatment group (Table I). When the demographic findings of the two groups were compared, the DAA treatment group had statistically significant better Eastern Cooperative Oncology Group (ECOG) performance status, albumin, aspartate aminotransferase (AST), alanine aminotransferase (ALT). Notably, none of the DAAtreated patients had a history of alcoholism. However, they had significantly more numbers of HCC.
CHC DAA treatments and the overall survival. Out of 113 patients, 14 patients had received various DAA treatments with durations ranging from 8~24 weeks. Two started DAA prior to TACE treatments, and the rest were treated by DAA in parallel with TACE treatment. Thirteen achieved SVR12 (13/14, 92.9%). Only patient number two had HCV reactivation. The clinical and virological data are listed in Table II. The overall survival in the DAA-treated patients was significantly longer than that in untreated patients as shown in the Kaplan-Meier curve (p=0.005; Figure 1). The median survival was 22.9 months in the untreated CHC/HCC patients and 40.2 months in the DAA-treated patients, respectively (Table I).    We also found that DAA therapy has a positive effect on TTCR. In contrary, male sex, higher bilirubin, ALT, and WBC were associated with poor TACE treatment response. In patients who had previous HCC treatments, higher AFP and WBC were indicators of poor progression-free survival. Similar to data recorded in various types of malignancy and diseases, it is not surprising that performance status (PS) and hypoalbuminemia are significant indicators of survival outcomes. Patients who have worse PS tend to be frail and are less likely to tolerate rigorous anti-cancer therapy. They are prone to have less favorable outcomes than patients with better PS. Thus, this is probably also the reason that PS is one of the three assessing components for the BCLC HCC staging. Serum albumin is not only an indicator of nutrition status, it is also a liver function parameter that is included in several scoring criteria such as the Child-Pugh Scoring system and Albumin-Bilirubin (ALBI). Our univariate and multivariate analyses showed comparable results.

Discussion
In the newly established 10 th Annual Asia-Pacific Primary liver cancer Expert (APPLE) consensus, it has been stated that liver function preservation is as important as achieving high objective response for the treatment of intermediatestage of HCC. CHC induced liver inflammation causes deterioration of liver functions and results in limited HCC treatment options. Studies have shown that eradication of HCV can improve liver function, promote fibrosis regression, prevent cirrhosis progression, and reduce risk of liver decompensation, which further result in decreased liverrelated mortality. Current guidelines recommend HCV eradication for all patients except for those with limited life expectancy. The benefits of HCV clearance with DAA therapy after curative treatments have been extensively studied in HCC BLCL stage 0 and A patients. However, data regarding whether anti-HCV treatment should be initiated on intermediate or later stage HCC is currently unknown. Our rationale is that if HCV clearance can be achieved with DAA therapy, liver dysfunction may be reversed, which might further improve survival even in HCC BCLC stage B patients. Our study showed that DAA therapy had a beneficial effect on overall outcome. It is known that TACE is the standard treatment for intermediate stage HCC, whereas systemic therapy is the treatment of choice for those refractory to TACE. Furthermore, a recent randomized multicenter prospective study conducted by Kudo et al. showed that combination treatment with TACE plus sorafenib in unresectable HCC significantly improved progression free survival compared to TACE alone. However, poor liver function and its related complications may hinder eligibility to TACE treatments or systemic therapy. It is important to preserve liver function in these patients to maintain their eligibility for not only subsequent systemic therapy after TACE failure/refractory HCC but also for new combination therapy with TACE and systemic therapy. In this respect, it is strategic and critical to treat HCV with DAA in patients with BCLC stage B HCC. In patients receiving TACE, tumor response based on mRECIST especially initial complete response (CR) can predict a longer survival benefit. Our study revealed that DAA therapy has a positive effect on TTCR. This may be the reason DAA could improve OS and further support the use of DAA in HCV patients with BCLC stage B HCC.
Ng et al. found that female gender tends to have HCC survival advantage over male which is likely due to the differences in tumor nature between genders. On the other hand, underlying differences in gene expression in opposite sex may also play a role. Liu et al. identified that Family with sequence similarity 83 (FAM83D) over-expression was observed significantly more in male and was associated with worse outcomes. These findings may provide possible explanations why the TACE treatment response was worse in male patients in the present study.
Studies have shown that poor liver function reserve, highdose chemotherapeutic agents, and history of multiple embolization procedures increased risk of acute liver failure after TACE. Jeon et al. also demonstrated higher total bilirubin levels increased risk of acute hepatic failure after TACE. In our study, higher bilirubin and ALT levels were associated with decreased response to TACE. Both higher levels of bilirubin and ALT were indicators of poor liver dysfunction. Patients with higher bilirubin and ALT levels may be less tolerant to higher dosage of TACE due to higher risk of post-TACE irreversible liver parenchymal injury or even liver failure, thus affecting treatment outcome and leading to a less favorable response.
Tumor progression from previously treated residual HCC and/or repeated tumor recurrence after TACE may be associated with increased malignant potential of the tumor nature. Sustained liver parenchymal injury from previous locoregional treatments may further adversely affect progression-free survival as was seen in our analysis.
Several reports have identified AFP as an independent factor for HCC recurrence. Chen et al. have identified the pro-oncogenic role of AFP in HCC progression (44). It is therefore not surprising to observe that higher AFP levels are an unfavorable prognostic factor of PFS in the present study. Previously, studies have revealed the association between the pro-tumor and anti-tumor inflammatory effects of immune response with elevated WBC and neutrophil-lymphocyte ration (NLR) in different cancers. In our study, elevated WBC and NLR are both associated with unfavorable outcomes. However, NLR did not reach statistical significance, only WBC was found to be a significant factor for TTCR and PFS.
A previous study reported a lower SVR rate with DAA therapy in active HCC compared to inactive HCC (85.5% vs. 93.7%). In the present study, we achieved a much higher SVR of 92.9% in patients with BCLC stage B HCC. However, this may be due to the fact that we had a very small sample size of 14 patients who received DAA.
Our study had several limitations. First, it was a retrospective evaluation from a single medical center in Taiwan. The study population had little heterogeneity. The fact that the entire study cohort was ethnically Taiwanese could contribute to potential selection bias. This may limit the applicability of the results as it is unclear whether similar results would be observed in different ethnic groups. Second, the sample size of 14 patients who ultimately received DAA therapy was small. This limits the statistical power of the study results. However, this was an anticipated problem as the current guidelines do not support the use of DAA for HCV eradication in HCC BCLC stage B patients. Lastly, of the 14 patients treated by DAA, 12 were Child-Pugh A (85.7%). We could not draw conclusions on the benefits of DAA treatment in HCC BLCL stage B patients who had more advanced liver disease. Currently, Child-Push B/C score is listed as contraindication of some DAA therapies.
In conclusion, we demonstrated that DAA therapy was associated with improved OS and TTCR in CHCintermediate stage HCC. Moreover, we demonstrated that ECOG, higher serum albumin, and DAA therapy are independent predictors of survival in CHC-HCC patients.

Conflicts of Interest
All Authors declare no conflicts of interest regarding this article.