A novel cDNA-uPA/SCID/Rag2-/- /Jak3-/- mouse model for hepatitis virus infection and reconstruction of human immune system.

Although human hepatocyte-transplanted immunodeficient mice support infection with hepatitis viruses, these mice fail to develop viral hepatitis due to the lack of an adaptive immune system. In this study, we generated new immunodeficiency cDNA-urokinase-type plasminogen activator (uPA)/SCID/Rag2-/- /Jak3-/- mice and established a mouse model with both a humanized liver and immune system. Transplantation of human hepatocytes with human leukocyte antigen (HLA)-A24 resulted in establishment of a highly replaced liver in cDNA-uPA/SCID/Rag2-/- /Jak3-/- mice. These mice were successfully infected with hepatitis B virus (HBV) and hepatitis C virus (HCV) for a prolonged period and facilitate analysis of the effect of anti-HCV drugs. Administration of peripheral blood mononuclear cells (PBMCs) obtained from an HLA-A24 donor resulted in establishment of 22.6%-81.3% human CD45-positive mononuclear cell chimerism in liver-infiltrating cells without causing graft-versus-host disease in cDNA-uPA/SCID/Rag2-/- /Jak3-/- mice without human hepatocyte transplantation. When mice were transplanted with human hepatocytes and then administered HLA-A24-positive human PBMCs, an alloimmune response between transplanted human hepatocytes and PBMCs occurred, with production of transplanted hepatocyte-specific anti-HLA antibody. In conclusion, we succeeded in establishing a humanized liver/immune system characterized by an allo-reaction between transplanted human immune cells and human liver using a novel cDNA-uPA/SCID/Rag2-/- /Jak3-/- mouse. This mouse model can be used to generate a chronic hepatitis mouse model with a human immune system with application not only to hepatitis virus virology but also to investigation of the pathology of post-transplantation liver rejection.

The presence of vessels encapsulating tumor clusters is associated with an immunosuppressive tumor microenvironment in hepatocellular carcinoma.

Recently, a distinct vascular pattern in hepatocellular carcinoma (HCC) called vessels encapsulating tumor-forming clusters (VETC) has received attention because of its association with poor prognosis. However, little is known about the mechanism by which VETC promotes an aggressive phenotype at the molecular level. In our study, the association between differences in stepwise signal intensity in the HB phase and molecular subtypes and somatic mutations associated with the immune microenvironment were investigated using the International Cancer Genome Consortium (ICGC) cohort (66 patients). To our knowledge, this is the first study to analyze the molecular patterns of VETC using RNA-Seq data. The VETC+ HCC group showed significantly lower overall survival and higher cumulative incidence of extrahepatic metastasis after curative hepatic resection than the VETC- HCC group. The VETC+ group exhibited molecular features indicative of lower immune activation than the VETC- group, suggesting that tumor cells in the VETC+ group were more likely to escape from the immune response, which could lead to the shorter OS (Overall survival) and higher risk of metastasis. On the other hand, gene expression levels of fibroblast growth factor receptors were upregulated in VETC+ HCC, suggesting that VETC+ HCC might benefit from lenvatinib treatment. Our results demonstrate that VETC+ HCC was associated with the suppression of tumor immune responses at the molecular level.

CTL-associated and NK cell-associated immune responses induce different HBV DNA reduction patterns in chronic hepatitis B patients.

The activation of hepatitis B virus (HBV)-related hepatitis is associated with both natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). We analyzed the association between the immune response and changes in the proportion of Pre-S deletion variants. We quantified Pre-S deleted HBV (HBV-del) and wild-type HBV (HBV-wt) DNA levels in sera obtained from HBV-infected mice and chronic hepatitis B patients. In chronic hepatitis B patients, the HBV-del proportion usually increased during or after ALT elevation but did not occur during all ALT elevations. To clarify this difference in the immunological responses, we performed in vivo analyses using HBV-infected human hepatocyte chimeric mice. Although HBV-del proportions did not change in mice with NK cell-associated hepatitis or in mice treated with entecavir, the proportions sharply increased in mice with CTL-associated hepatitis. Furthermore, the number of patients in which HBV-del proportions were greater than 5% was significantly higher in chronic hepatitis B patients than in asymptomatic carriers (P = 0.023). We identified associations between virological response in chronic hepatitis B patients and two different immune responses. The proportion of HBV-del variants could be a useful biomarker for distinguishing between chronic hepatitis and asymptomatic carriers.

Emergence of drug resistance-associated variants and changes in serum lipid profiles in sofosbuvir plus ledipasvir-treated chronic hepatitis C patients.

Combination of sofosbuvir plus ledipasvir therapy has been expected to enhance sustained virological response (SVR) rates in hepatitis C virus (HCV) genotype 1 chronic infected patients. We analyzed the emergence of drug resistance-associated variants (RAVs) in treatment failure and changes in lipid profiles in sofosbuvir/ledipasvir-treated patients. A total of 176 patients with chronic HCV genotype 1 infection without decompensated liver cirrhosis were treated with sofosbuvir/ledipasvir for 12 weeks. NS5A and NS5B RAVs were determined by either Invader assay or direct sequencing. Serum lipid-related markers were measured at the start of treatment and at week 4 in patients who received sofosbuvir/ledipasvir and ombitasvir/paritaprevir/ritonavir therapies. SVR was achieved in 94.9% (167 out of 176) of patients. SVR12 rate was 97.1% for patietns with low frequncy (<25%) of baseline NS5A RAVs, but 82.8% for patients with high frequency (>75%) of NS5A RAVs. In multivariate regression analysis, higher albumin (odds ratio [OR] = 0.020 for presence; P = 0.007), and NS5A-L31/Y93 RAVs with a population frequency <75% (OR = 29.860 for presence; P = 0.023) were identified as significant independent predictors for SVR12. NS5A-Y93H substitutions were detected in all nine treatment failures at HCV relapse, and three out of six patients with NS5A inhibitor-naïve patients achieved additional NS5A RAVs. Serum low-density lipoprotein cholesterol and apolipoprotein B levels were significantly elevated at week 4 in sofosbuvir/ledipasvir-treated patients. These elevations were greater than in ombitasvir/paritaprevir/ritonavir-treated patients. In conclusion, NS5A multi-RAVs are likely to develop in patients who fail to respond to sofosbuvir/ledipasvir therapy. Inhibition of HCV replication with sofosbuvir might affect lipid metabolism.

A case of complete response to radiotherapy combined with durvalumab and tremelimumab in a patient with unknown primary hepatocellular carcinoma arising in the lumbar spine.

A 58-year-old man visited an orthopedic clinic complaining of pain in his right lower back and numbness in his lower limbs for one month. Imaging tests revealed a tumorous lesion from the left side of the second lumbar vertebra to the paraspinal muscles. CT-guided biopsy of the tumor was performed, and immunostaining results diagnosed hepatocellular carcinoma (HCC). Although the liver showed signs of chronic liver damage, no primary tumor was found within the liver or in other organs. Blood tests showed negative hepatitis virus markers for both HBV and HCV. The tumor markers AFP, AFP-L3, and DCP were high. Because he developed spinal cord compression syndrome due to a lumbar tumor, radiation therapy and denosumab administration were performed. Subsequently, systemic therapy with durvalumab plus tremelimumab was started. In the year following the start of treatment, the tumor has shrunk, and no new lesions have been observed. Tumor markers have also decreased. We have experienced a case of HCC in the lumbar spine without a primary tumor in the liver. This is a very rare case, and the combination therapy with durvalumab and tremelimumab resulted in a complete response, which we consider to be a valuable case.

Peripheral T Cell Subpopulations as a Potential Surrogate Biomarker during Atezolizumab plus Bevacizumab Treatment for Hepatocellular Carcinoma.

The therapeutic benefits of the immunotherapeutic combination of atezolizumab and bevacizumab (Atez/Bev) in hepatocellular carcinoma (HCC) vary. Therapeutic biomarkers might help improve outcomes for HCC patients receiving Atez/Bev therapy. The role of systemic immune profiles in HCC progression also remains unclear. This study aimed to evaluate the status and dynamics of peripheral T cell subpopulations in HCC patients receiving Atez/Bev treatment and to explore biomarkers predictive of a therapeutic response. We enrolled 83 unresectable advanced HCC patients who commenced Atez/Bev treatment at our hospital between October 2020 and June 2022. Peripheral T cell subpopulations in peripheral blood mononuclear cells at baseline and 3 weeks post-treatment were investigated using flow cytometry and compared with those in control samples from 18 healthy individuals. We retrospectively analyzed the association between peripheral T cell subpopulation profiles and clinical outcomes. Baseline peripheral T cell subpopulations could be profiled in 70 patients with sufficient cell counts, among whom 3-week subpopulations could be evaluated in 51 patients. Multivariate analysis showed that a high baseline proportion of CD8+ central memory T (TCM) cells was independently associated with longer progression-free survival (PFS). Further, overall survival (OS) was significantly prolonged in patients with increased CD8+ effector memory T (TEM) cell proportions. In conclusion, TCM proportion at baseline might be a good indicator of the efficacy of Atez/Bev therapy. Furthermore, observation of increasing TEM proportions might be an early predictor of the potential clinical benefits of treatment.

Association of Hepatobiliary Phase of Gadoxetic-Acid-Enhanced MRI Imaging with Immune Microenvironment and Response to Atezolizumab Plus Bevacizumab Treatment.

It has been reported that high intensity in the hepatobiliary (HB) phase of Gd-EOB-DTPA-enhanced MRI (EOB-MRI) is associated with an immune-cold microenvironment in HCC. The aim of this study is to reveal whether non-high-intensity HCCs are homogeneous with respect to the immune microenvironment and to investigate the predictive ability of EOB-MRI for the response to atezolizumab + bevacizumab therapy (Atezo/Bev). The association between differences in stepwise signal intensity of HB phase and molecular subtypes and somatic mutations associated with the immune microenvironment was investigated in 65 HCC patients (cohort 1). The association between EOB-MRI and the therapeutic effect of Atezo/Bev was evaluated in the Atezo/Bev cohort (60 patients in cohort 2). The proportion of HCCs having CTNNB1 mutations and classified as Chiang CTNNB1 and Hoshida S3 was high in the high-intensity HB-phase group. Infiltration of tumor-associated macrophages (TAM) and regulatory T-lymphocytes (Treg) was characteristic of the high-intensity and low-intensity groups, respectively. Although EOB-MRI could not predict the response to Atezo/Bev treatment, our results demonstrate that EOB-MRI could serve as a surrogate marker predicting the immune microenvironment. This suggests that Atezo/Bev treatment can be selected regardless of signal intensity in the EOB-MRI HB phase.

A case with life-threatening secondary sclerosing cholangitis caused by nivolumab.

Immune checkpoint inhibitor (ICI) therapy has potent anti-cancer effects but is associated with immune-related adverse events (irAEs). We present a case who developed secondary sclerosing cholangitis following treatment with nivolumab for non-small cell lung cancer who did not respond to immunosuppressive treatments and died of liver failure. A 75 year-old male with lung cancer who had been treated with nivolumab for non-small cell lung cancer developed Grade 3 liver injury with significant elevation of hepatobiliary enzymes. Magnetic resonance cholangiopancreatography (MRCP) revealed diffuse dilatation of the common bile duct and multifocal stenosis with prestenotic dilatation from the perihilar to intrahepatic bile duct, consistent with sclerosing cholangitis. Histological findings represented an infiltration of mainly CD8-positive T cells around the bile ducts in the liver. Despite treatments with ursodeoxycholic acid, prednisolone, and mycophenolate mofetil, the sclerosing cholangitis did not improve, and the patient died due to liver failure and aggravation of lung cancer. These findings suggest that immune checkpoint inhibitors may lead to resistance to immunosuppressive treatment as well as pose a risk of life-threatening sclerosing cholangitis.

Real-word efficacy of sofosbuvir, velpatasvir plus ribavirin therapy for chronic hepatitis patients who failed to prior DAA therapy with NS5A-P32 deletion mutated HCV infection.

The hepatitis C virus (HCV) NS5A-P32 deletion (P32del) confers potent resistance to NS5A inhibitors. Chronic hepatitis C patients in whom NS5A-P32del variants had emerged during prior direct-acting antiviral (DAA) therapy with an NS5A inhibitor show poor response to DAA retreatment. Here, we report three patients with HCV NS5A-P32del infection who were treated with sofosbuvir, velpatasvir plus ribavirin (SOF/VEL + RBV) in a real-world setting. The patients developed HCV NS5A-P32del, L31F + P32del, or L31V + P32del variants following failure of daclatasvir plus asunaprevir (DCV/ASV) therapy. One of the patients failed to respond to subsequent DCV/ASV and beclabuvir therapy, and the remaining two patients failed to respond to subsequent glecaprevir and pibrentasvir therapy. All three patients completed 24-week SOF/VEL + RBV therapy. Serum HCV RNA became negative at the end of the therapy in all three patients. Two patients with NS5A-P32del and NS5A-L31F + P32del achieved sustained virological response 12 weeks after completion of treatment (SVR12), but HCV relapsed in the remaining NS5A-L13V + P32del patient. Direct sequence analysis detected no additional variants within either the NS5A or NS5B regions at the time of relapse. In conclusion, three patients with prior NS5A-P32del-associated DAA treatment failure received 24 weeks of SOF/VEL + RBV therapy, and two of the patients achieved SVR12.

Successful retreatment with 12 weeks of glecaprevir and pibrentasvir for a genotype 2a HCV-infected hemodialysis patient who failed to respond to 8 weeks of prior glecaprevir and pibrentasvir therapy.

Although NS3/4 protease inhibitor glecaprevir (GLE) plus NS5A inhibitor pibrentasvir (PIB) therapy has a high efficacy for chronic hepatitis C virus (HCV)-infected patients with hemodialysis, some patients fail to respond to the therapy. Here, we report a hemodialysis genotype 2 HCV-infected patient who achieved sustained virological response (SVR) by 12 weeks of GLE/PIB therapy after failing to respond to 8 weeks of GLE/PIB therapy. A 44-year-old man with chronic genotype 2a HCV-infection without any evidence of cirrhosis and who was undergoing hemodialysis received GLE/PIB therapy. He completed 8 weeks of therapy, but his serum HCV relapsed after the end of therapy. No resistance-associated substitutions were detected in the NS3 region, but NS5A-C92C/S was detected by direct sequence analysis prior to the start of therapy and subsequently shifted to NS5A-C92S at the time of HCV relapse. Four months after initial GLE/PIB therapy, he started a 12-week course of GLE/PIB retreatment. Serum HCV RNA level became and remained undetectable during the therapy and never relapsed after the end of the treatment. Finally, he succeeded in achieving sustained virological response following 12 weeks of GLE/PIB retreatment.

PNPLA3 and HLA-DQB1 polymorphisms are associated with hepatocellular carcinoma after hepatitis C virus eradication.

BACKGROUND: Even though both interferon (IFN)-based and direct-acting antiviral (DAA) therapies against hepatitis C virus (HCV) reduce the risk of hepatocellular carcinoma (HCC), post-sustained virological response (SVR) patients remain at elevated risk of HCC. METHODS: A total of 4620 patients who achieved SVR were enrolled in this retrospective cohort study. After excluding patients who had a history of HCC or developed HCC within 1 year and whose follow-up period was less than 1 year and who were positive for HBsAg, we investigated the association between clinical characteristics and HCC development after SVR in the remaining 3771 patients. RESULTS: Median observation period was 41 months. We confirmed known risk factors. In addition, we found that PNPLA3 and HLA-DQB1 polymorphisms were associated with HCC after SVR. Finally, we propose an estimation model for the incidence of HCC after SVR. Based on gender, FIB-4 index, AFP, and PNPLA3 polymorphism, about 18% of all patients were classified as having high risk, with a cumulative incidence rate (CIR) at 5 years of 16.5%. Another 17% were classified as having moderate risk with a CIR of 7.6%. The remaining 65% showed a CIR of 0.5%. The effect of PNPLA3 polymorphism might be more pronounced in patients with lower body mass index (BMI) and without diabetes mellitus compared to those with higher BMI and diabetes mellitus. CONCLUSIONS: We demonstrated that PNPLA3 and HLA-DQB1 polymorphisms were associated with HCC after SVR. These findings might be useful to inform risk stratification for HCC surveillance after SVR.

Tumor Fibroblast Growth Factor Receptor 4 Level Predicts the Efficacy of Lenvatinib in Patients With Advanced Hepatocellular Carcinoma.

OBJECTIVES: Biomarkers for optimizing the outcome of treatment with lenvatinib in patients with advanced hepatocellular carcinoma remain to be established despite intensive and comprehensive genomic research. Lenvatinib is characterized by its prominent inhibitory potency for fibroblast growth factor receptor (FGFR) 4 compared with earlier tyrosine kinase inhibitors. Thus, in this study, we focused on simplified quantification of FGFR4 in tumors as a potential predictive indicator. METHODS: According to The Cancer Genome Atlas data set curation, FGFR4 messenger RNA is broadly overexpressed in hepatocellular carcinoma in the absence of gene alteration. Gene set enrichment analysis revealed that the aggressiveness of the tumor was closely related to the FGFR4 level. To confirm the relationship between the benefits of lenvatinib and tumor addiction to the FGFR4 pathway, we analyzed protein levels in tumors and peripheral blood obtained from 57 prospectively registered patients treated with lenvatinib. RESULTS: Positive immunohistochemistry (>10% of tumor cells) for FGFR4 in biopsy samples before treatment was associated with a longer progression-free survival (2.5 vs 5.5 months, P = 0.01) and a favorable objective response rate (31% vs 81%, P = 0.006). By contrast, the concentration of soluble FGFR4 in peripheral blood as measured by an enzyme-linked immunosorbent assay was not associated with survival outcomes, because its fluctuations reflect hepatic fibrosis. Additional RNA sequencing analysis using archival surgical specimens (n = 90) suggested that alternative RNA splicing of FGFR4 in cancer may also explain this discrepancy. DISCUSSION: The tumor FGFR4 level was an independent predictor of response to lenvatinib.

Comparison of intracellular responses between HBV genotype A and C infection in human hepatocyte chimeric mice.

BACKGROUND AND AIMS: The clinical course and responsiveness to antiviral treatments differs among hepatitis B virus (HBV) genotypes. However, the cause of these differences is unclear. In the present study, we compared mRNA expression profiles in human hepatocyte chimeric mice infected with HBV genotypes A and C. METHODS: Fifteen chimeric mice were prepared and divided into the following three groups: uninfected control mice, HBV genotype A-infected mice, and HBV genotype C-infected mice. Human hepatocytes were collected from these mouse livers and gene expression analyses were performed using next-generation RNA sequencing. RESULTS: Although similar pathways were influenced by HBV infection, including inflammation mediated by chemokine and cytokine signaling, p53, and integrin signaling pathways, expression levels of up-regulated genes by HBV genotype A or C infection were quite different. In HBV genotype A-infected hepatocytes, 172 genes, including KRT23 and C10orf54, were significantly more highly expressed than in HBV genotype C-infected cells, whereas 10 genes, including SPX and IER3, were expressed at significantly lower levels. Genes associated with the p53 pathway and the inflammation mediated by chemokine and cytokine signaling pathway were more highly expressed in cells with HBV genotype A infection, whereas genes associated with CCKR signaling map and oxidative stress response were more highly expressed in cells with HBV genotype C infection. CONCLUSION: Several differences in gene expression with respect to HBV genotype A and C infection were detected in human hepatocytes. These differences might be associated with genotypic difference in the clinical course or responsiveness to treatment.

Involvement of Porphyromonas gingivalis in the progression of non-alcoholic fatty liver disease.

BACKGROUND AND AIMS: The risk factors in the progression of nonalcoholic fatty liver disease (NAFLD) have not been fully clarified. Porphyromonas gingivalis (P.g) has been considered to be a confounding risk factor for systemic diseases. We aimed to evaluate the effect of P.g infection on risk of progression to NASH. METHODS: (1) Serum IgG antibody titers against P.g fimbriae (fimA) in 200 biopsy-proven NAFLD patients were measured by ELISA and compared with histological findings. (2) C57BL/6J mice were fed a control diet (CD) or high-fat diet (HFD) with or without P.g-odontogenic infection and analyzed histologically. Mouse livers were analyzed using CE-TOFMS and LC-TOFMS. RESULTS: (1) A significant correlation between fibrosis progression and antibody titers against P.g possessing fimA type 4 was identified (P = 0.0081). Multivariate analysis identified older age and type 4 P.g-positivity as risk factors for advanced fibrosis. (2) Fibrosis and steatosis were more severe in HFD P.g(+) mice compared with HFD P.g(-) mice. In metabolome analysis, fatty acid metabolism was significantly disrupted with HFD in P.g-infected mouse livers. Monounsaturated/saturated fatty acid ratios were significantly higher in the HFD P.g(+) group than in the HFD P.g(-) group (P < 0.05). Moreover, expression levels of SCD1 and ELOVL6 were significantly reduced. CONCLUSIONS: These results suggest that P.g infection is an important risk factor for pathological progression in NAFLD. Increase in the monounsaturated/saturated fatty acid ratio may be an important change that facilitates progression of NAFLD.

Efficacy and safety of ledipasvir/sofosbuvir with ribavirin in chronic hepatitis C patients who failed daclatasvir/asunaprevir therapy: pilot study.

BACKGROUND: In Japan, daclatasvir (DCV) and asunaprevir (ASV) therapy was the first IFN-free treatment to be approved, and thousands of patients have since been successfully treated, with an SVR rate of around 90%. The converse, however, is that around 10% of patients fail to achieve viral eradication and must be retreated using a different approach. This study is to evaluate treatment efficacy of ledipasvir/sofosbuvir and ribavirin in patients who failed to respond to DCV and ASV therapy. METHODS: Thirty patients were treated with 12 weeks of ledipasvir/sofosbuvir and ribavirin. We evaluated the rate of sustained virological response 12 weeks after the end of treatment (SVR12) and examined the incidence of adverse events during ledipasvir/sofosbuvir and ribavirin treatment. NS5A and NS5B resistance-associated variants (RAVs) in treatment failure cases were examined. RESULTS: The overall SVR12 rate was 86.7% (26/30). Large decreases in mean log10 HCV RNA levels were observed in patients without cirrhosis, and the SVR12 rate for these patients was 100% (12/12). In cases of cirrhosis, SVR12 rate was 72.2% (13/18). The common factors in treatment failure cases were the presence of liver cirrhosis and both NS5A L31M/I and Y93H RAVs. The frequency of RAVs did not change before and after treatment among patients who relapsed. CONCLUSION: Ledipasvir/sofosbuvir with ribavirin is an effective retreatment option for patients with chronic hepatitis C who failed to respond to prior daclatasvir and asunaprevir therapy.

The relationship between HBcrAg and HBV reinfection in HBV related post-liver transplantation patients.

BACKGROUND: Post-transplant hepatitis B virus (HBV) reinfection is one of the major problems facing patients who undergo HBV-related liver transplantation (LT). We analyzed the clinical impact of serum hepatitis B core-related antigen (HBcrAg) on HBV reinfection in post-LT patients with HBV-related liver diseases. METHODS: Serum hepatitis B surface antigen (HBsAg), HBV DNA, and HBcrAg were measured over time in 32 post-LT patients. Twenty-one out of 32 patients had HCC at LT. The effects of HBcrAg, hepatocellular carcinoma (HCC) recurrence, and HBs gene mutation on HBV reinfection and withdrawal from hepatitis B immune globulin (HBIG) were analyzed. RESULTS: Sixteen out of 32 patients (50 %) were positive for HBcrAg even though only six patients were thought to have experienced HBV reinfection based on reappearance of either HBV DNA or HBsAg during a median follow-up time of 75 months. Three of these six patients who became re-infected with HBV experienced HCC recurrence after LT. The HBV DNA reappearance rate was significantly higher in patients with HCC recurrence after LT (p < 0.001). Two HBV re-infected patients without HCC recurrence had HBs gene mutations G145R and G145A, respectively. Anti-HBs antibody development rate by HB vaccination was similar between HBcrAg-positive and negative patients (p = 0.325). CONCLUSIONS: HBV reinfection is more common than is usually considered based on conventional measurement of HBsAg and HBV DNA. HCC recurrence and mutations in the HBV S gene were associated with HBV reinfection after LT.

ITPA polymorphism effects on decrease of hemoglobin during sofosbuvir and ribavirin combination treatment for chronic hepatitis C.

BACKGROUND: Polymorphisms in the inosine triphosphatase (ITPA) gene is associated with anemia induced by peg-interferon (PEG-IFN) plus ribavirin (RBV) treatment for patients with chronic hepatitis C virus (HCV) infection. However, the effect of ITPA polymorphism on sofosbuvir plus RBV treatment is unknown. METHODS: Two hundred and forty-four patients with chronic HCV genotype 2 infection without decompensated liver cirrhosis were treated with sofosbuvir plus RBV for 12 weeks. The effects of ITPA polymorphism on hemoglobin levels and RBV dose reduction and treatment response were analyzed. ITPA (rs1127354) was genotyped using the Invader assay. Multivariate regression analysis was performed to identify factors associated with sustained virological response (SVR). RESULTS: Overall, SVR12 was achieved in 231 (94.7%) patients, based on intention to treat analysis. During the therapy, reduction of hemoglobin levels was significantly greater in ITPA genotype CC patients than CA/AA patients. Therefore, the cumulative proportion of patients with RBV dose reduction was significantly higher and total dose of RBV was significantly lower in patients with CC genotype compared to CA/AA genotypes. SVR12 rates were similar between ITPA genotypes CC and CA/AA (94.7 and 94.4%, respectively, P = 0.933). Multivariate logistic regression analysis identified FIB4 index <3.25 (odds ratio [OR], 9.388 for ≥3.25; P = 0.005) and low body weight (OR, 1.059, for high body weight; P = 0.017) as independent predictors for SVR12. CONCLUSIONS: ITPA polymorphism influences hemoglobin levels and incidence of RBV dose reduction during sofosbuvir plus RBV therapy. However, ITPA genotype CC patients can expect a curative effect equivalent to CA/AA patients for chronic HCV genotype 2 infection.