Shorter duration of venetoclax administration to 14 days has same efficacy and better safety profile in treatment of acute myeloid leukemia.

Venetoclax (VEN) is now widely used in the treatment of acute myelogenous leukemia (AML) in elderly patients who are not eligible for intensive remission induction therapy. Prolonged myelosuppression, increased incidence of infection, and long duration of hospital stay were major concerns for VEN treatment cases, and we thought that shortening the duration of VEN administration during induction therapy might solve these problems. Thirteen newly diagnosed AML patients who underwent VEN+azacitidine (AZA) induction therapy from March 2021 to June 2022 at Kushiro Rosai Hospital were analyzed retrospectively. The median age was 79 (range, 68-86) years, and 8 of the patients (61.5%) were classified as high risk according to the ELN 2017 risk stratification. Eight patients received VEN for 14 days (VEN14 group), and 5 patients received VEN for 28 days (VEN28 group). The composite complete remission (CRc) rate was 76.9% in total, and the CRc rates in the VEN14 and VEN28 groups were almost the same (75.0% and 80.0%, respectively). The median overall survival (OS) was not reached in the VEN14 group and was 254 days in the VEN28 group. The median event-free survival (EFS) was not reached in the VEN14 group and was 178 days in the VEN28 group. The VEN14 group might have a possibility to reduce febrile neutropenia (37.5% vs. 80%) and reduce the duration of hospital stay (median, 21.5 vs. 31 days) compared with the VEN28 group. VEN14 produced the same CRc rate and survival rate, safer profile, and shorter duration of hospital stay than VEN28.

Dominant-negative type of IKZF1 deletion showed a favorable prognosis in adult B-cell acute lymphoblastic leukemia.

IKZF1 deletion is a recurrent genomic alteration in B-cell acute lymphoblastic leukemia (B-ALL) and is divided into dominant-negative (DN) and loss of function (LOF) deletions. The prognostic impact of each deletion has not been fully elucidated. We retrospectively analyzed 117 patients with adult B-ALL including 60 patients with BCR::ABL1-positive B-ALL and 57 patients with BCR::ABL1-negative B-ALL by the fluorescence in situ hybridization (FISH) method for IKZF1 deletion and multiplex PCR for the 4 most common IKZF1 deletions (∆4-7, ∆2-7, ∆2-8, and ∆4-8). Samples, in which IKZF1 deletion was detected by FISH but a specific type of deletion was not identified by the PCR, were categorized as "other." Patients were classified into a DN group that had at least 1 allele of ∆4-7 (n = 23), LOF and other group (n = 40), and wildtype group (n = 54). DN type IKZF1 deletions were found in 33.3% of BCR::ABL1-positive cases and 5.2% of BCR::ABL1-negative cases. LOF and other type IKZF1 deletions were found in 43.4% of BCR::ABL1-positive cases and 24.6% of BCR::ABL1-negative cases. Patients with the DN group showed significantly higher overall survival (OS) than that of the LOF and other and WT groups (P = 0.011). Multivariate analysis including age, WBC counts, complex karyotype, and DN type IKZF1 deletion showed that the DN type of IKZF1 deletion (HR = 0.22, P = 0.013) had a positive impact and age ≥ 65 (HR = 1.92, P = 0.029) had a negative impact on OS. The prognostic impact of IKZF1 deletion depends on the type of deletion and DN type of IKZF1 deletion showed better prognosis in adult B-ALL patients.Clinical trial registration This study was part of a prospective observational study (Hokkaido Leukemia Net, UMIN000048611). It was conducted in compliance with ethical principles based on the Helsinki Declaration and was approved by the institutional review board of Hokkaido University Hospital (#015-0344).

Feasibility of a modified search, coagulation, and clipping method with and without the use of polyglycolic acid sheets and fibrin glue for preventing delayed bleeding after gastric endoscopic submucosal dissection.

BACKGROUND: Methods have been developed for preventing delayed bleeding (DB) after gastric endoscopic submucosal dissection (GESD). However, none of the methods can completely prevent DB. We hypothesized that DB could be prevented by a modified search, coagulation, and clipping (MSCC) method for patients at low risk for DB and by combining the use of polyglycolic acid sheets and fibrin glue with the MSCC method (PMSCC method) for patients at high risk for DB (antibleeding [ABI] strategy). This study assessed the technical feasibility of this novel strategy. METHOD: We investigated 123 lesions in 121 consecutive patients who underwent GESD in Kushiro Rosai Hospital between April 2018 and January 2020. The decision for continuation or cessation of antithrombotic agents was based on the Guidelines for Gastroenterological Endoscopy in Patients Undergoing Antithrombotic Treatment. RESULTS: Oral antithrombotic agents were administered to 28 patients (22.8%). The en bloc R0 resection rate was 98.4%. The MSCC method and the PMSCC method for preventing DB were performed in 114 and 9 lesions, respectively. The median time of the MSCC method was 16 min, and the median speed (the resection area divided by the time of method used) was 3.6 cm2/10 min. The median time of the PMSCC method was 59 min, and the median speed was 1.3 cm2/10 min. The only delayed procedural adverse event was DB in 1 (0.8%) of the 123 lesions. CONCLUSIONS: The ABI strategy is feasible for preventing DB both in patients at low risk and in those at high risk for DB after GESD, whereas the PMSCC method may be necessary for reduction of time.

Early response and safety of atezolizumab plus bevacizumab for unresectable hepatocellular carcinoma in patients who do not meet IMbrave150 eligibility criteria.

AIM: A clinical trial (IMbrave150) indicated the efficacy and safety of atezolizumab plus bevacizumab for patients with unresectable hepatocellular carcinoma (HCC). In this study, we evaluated this therapeutic combination in a real-world setting, with a focus on patients who did not meet the IMbrave150 eligibility criteria. METHODS: In this multicenter study, patients with unresectable HCC treated with atezolizumab plus bevacizumab between October 2020 and May 2021 were screened. In patients who did not meet IMbrave150 eligibility criteria, treatment responses and safety at 6 and 12 weeks were evaluated. RESULTS: Atezolizumab plus bevacizumab was initiated in 64 patients, including 46 patients (71.9%) who did not meet IMbrave150 eligibility criteria. Most of these patients had a history of systemic therapy (44/46). The objective response rate and disease control rate observed using Response Evaluation Criteria in Solid Tumors 1.1 were 5.2% and 82.8% at 6 weeks and 10.0% and 84.0% at 12 weeks, respectively; these rates were similar between patients who met and did not meet the IMbrave150 criteria. Ten patients experienced progressive disease (PD) at 6 weeks. Portal vein tumor thrombosis was significantly associated with PD (p = 0.039); none of the 15 patients with hepatitis B virus-related HCC experienced PD (p = 0.050). The most common adverse events of grade 3 or higher were aspartate aminotransferase elevation (n = 8, 13.8%) and the safety profile was similar between patients who met and did not meet the IMbrave150 criteria. CONCLUSION: Most patients treated with atezolizumab plus bevacizumab did not meet the IMbrave150 criteria; however, the combination therapy showed good safety and efficacy at the early treatment phase.

Validation and comparison of prognostic values of GNRI, PNI, and CONUT in newly diagnosed diffuse large B cell lymphoma.

Diffuse large B cell lymphoma (DLBCL) is the most common type of aggressive non-Hodgkin lymphoma. Emerging evidence indicates that poor nutritional status determined with nutritional indices such as geriatric nutritional risk index (GNRI), prognostic nutritional index (PNI), and controlling nutritional status score (CONUT) was associated with poor prognosis of DLBCL. We conducted this multicenter retrospective study to validate and compare prognostic values of the three indices in 615 newly diagnosed DLBCL patients. The overall survival (OS) in patients with poor nutritional status determined with each of these nutritional indices were significantly inferior compared with that in those without nutritional risks (5-year OS in patients with GNRI < 95.7 and GNRI ≥ 95.7 were 56.4% and 83.5%, P < 0.001; PNI < 42.4 and PNI ≥ 42.4 were 56.1% and 81.0%, P < 0.001; CONUT > 4 and CONUT ≤ 4 were 53.1% and 77.1%, P < 0.001). GNRI and CONUT were independent prognostic predictors for OS (GNRI < 95.7, hazard ratio [HR] 1.83, 95% confidence interval [CI] 1.22-2.74, P = 0.0032; CONUT > 4, HR 1.53, 95% CI 1.05-2.23, P = 0.028) after multivariate analyses. Nutritional status determined with GNRI affected OS more strongly in the patients with nongerminal center B cell-like (nonGCB) DLBCL compared with that in those with GCB-type DLBCL. In conclusion, baseline poor nutritional status determined based on GNRI or CONUT was an independent risk factor of newly diagnosed DLBCL, and GNRI was also useful as an independent prognostic factor for patients with nonGCB-type DLBCL.

Feasibility of a new ligation using the double-loop clips technique without an adhesive agent for ulceration after endoscopic submucosal dissection of the colon (with video).

BACKGROUND AND AIMS: Several ligation techniques for ulceration after endoscopic submucosal dissection (ESD) have been reported, but none have been established for clinical use because of technical complexity and the need for expensive equipment. Therefore, the technical feasibility of a new ligation method using the double-loop clips (D-L clips) technique without an adhesive agent for ulceration after ESD of the colon was assessed. METHODS: Among 35 patients who underwent ESD of the colon in Kushiro Rosai Hospital between April 2019 and September 2019, 26 patients who underwent ligation using the D-L clips technique for the post-ESD ulcer bed were included in this retrospective study. Continuation or cessation of antithrombotic agents was based on the Guidelines for Gastroenterological Endoscopy in Patients Undergoing Antithrombotic Treatment. RESULTS: The rate of en bloc R0 resection was 97.1%, the median length of the resected specimen was 3.2 cm (interquartile range [IQR], 2.8-3.8 cm), and the complete ligation rate was 88.5% (23 of 26). Excluding patients with lesion sites in the rectum below the peritoneal reflection, the complete ligation rate was 95.5% (21 of 22). The median duration of the ligation procedure was 20 minutes (IQR, 16-24 minutes). The only delayed procedural adverse event was post-ESD coagulation syndrome in 1 patient. Incomplete ligation was significantly more frequent in patients with lesion sites in the inferior rectal valve/anal verge area (P = .0269). CONCLUSIONS: Ligation using the D-L clips technique without an adhesive agent is feasible for closing ulceration after ESD of the colon, whereas other techniques may be necessary for lesions in the rectum below the peritoneal reflection.

Lenvatinib in patients with unresectable hepatocellular carcinoma who do not meet the REFLECT trial eligibility criteria.

AIM: This study aimed to determine the efficacy and safety of lenvatinib for patients with unresectable hepatocellular carcinoma (HCC) who did not meet REFLECT eligibility criteria (phase 3 clinical trial). METHODS: In this multicenter retrospective study, patients with unresectable HCC treated with lenvatinib between 2018 and 2019 and had adequate clinical data were included. Objective response rate, progression-free-survival (PFS) and safety were evaluated according to meeting or not meeting the REFLECT eligibility criteria and according to the criteria of the REFLECT trial. RESULTS: Of the 105 patients included, 61% (64 of 105) did not meet the REFLECT eligibility criteria. Safety and median PFS of lenvatinib were similar between the patients who did and those who did not meet the criteria. Among the patients who did not meet the criteria, 28, 27, 14, six, seven and five had a history of tyrosine kinase inhibitor (TKI) treatment, Child-Pugh score B, HCC in ≥50% of the liver, reduced platelet count, bile duct invasion and main portal vein invasion, respectively. The efficacy and safety of lenvatinib for patients with or without Child-Pugh-score B or HCC in ≥50% of the liver were similar. Although treatment outcome was not significantly different, patients with TKI treatment history tended to have longer median PFS, whereas those with main portal vein invasion tended to have shorter median PFS. CONCLUSION: Lenvatinib was effective for patients who did not meet the REFLECT inclusion criteria. However, the treatment outcome may vary according to several factors, such as a history of TKI treatment and tumor invasion.

Clinicopathological features of HCV-positive splenic diffuse large B cell lymphoma.

The hepatitis C virus (HCV) is a single-stranded RNA virus which is thought to be involved in the onset of B cell lymphoma. HCV-positive diffuse large B cell lymphoma (DLBCL) has been reported to clinically manifest in extranodal lesions (e.g., in the liver, spleen, and stomach). Here, we investigated HCV-positive and -negative primary splenic DLBCL (p-spDLBCL) and non-primary splenic DLBCL (ordinary DLBCL). Furthermore, to examine HCV lymphomagenesis, RNA in situ hybridization (ISH), RT-PCR (reverse-transcription polymerase chain reaction), and NS3 immunostaining of HCV viral nonstructural proteins were performed. HCV-positive p-spDLBCL patients presented fewer B symptoms (asymptomatic) and better performance status, with elevated presence of splenic macronodular lesions and more germinal center B cell (GCB) sub-group cases than HCV-negative p-spDLBCL patients. However, HCV-positive ordinary DLBCL patients were found to have more non-GCB sub-group cases than HCV-negative ordinary DLBCL patients. HCV-positive DLBCL patients showed 20.6% (7/34) NS3 positivity, 16.7% (1/6) HCV-RNA in situ positivity, and 22.2% (2/9) detection of HCV-RNA in tumor tissue by RT-PCR. Splenic samples were found to have a higher frequency of HCV detection than lymph node samples, thus suggesting that HCV may be closely related to lymphomagenesis, especially in splenic lymphoma.

[Aplastic anemia following drug-induced liver injury].

Aplastic anemia (AA), a hematopoietic disorder characterized by hypocellular bone marrow, is caused by immunologically-mediated hematopoietic stem cell injury. Viral infection is hypothesized as the underlying cause of hepatitis-associated AA, although its mechanism is still unclear. This report describes a case of AA following suspected drug-induced liver injury (DILI). An 18-year-old man developed severe liver dysfunction after taking oral over-the-counter drugs. The patient was diagnosed with suspected DILI based on drug-induced lymphocyte stimulation test and liver biopsy results. Although liver dysfunction improved after a course of steroid pulse therapy and liver supporting therapy, the man gradually developed pancytopenia within 3 months of DILI diagnosis, prompting the diagnosis of AA following DILI. Paroxysmal nocturnal hemoglobinuria-type cells were detected by high-sensitivity flow cytometry. Immunosuppressive therapy with antithymocyte globulin and cyclosporin was administered, with pancytopenia improvement. To the best of our knowledge, this is the first report in the literature with a case of AA following DILI, and we believe it is important for evaluating the pathogenesis of drug-induced and hepatitis-associated AA.

Pretreatment evaluation of fluorescence resonance energy transfer-based drug sensitivity test for patients with chronic myelogenous leukemia treated with dasatinib.

Tyrosine kinase inhibitors (TKI) are used for primary therapy in patients with newly diagnosed CML. However, a reliable method for optimal selection of a TKI from the viewpoint of drug sensitivity of CML cells has not been established. We have developed a FRET-based drug sensitivity test in which a CrkL-derived fluorescent biosensor efficiently quantifies the kinase activity of BCR-ABL of living cells and sensitively evaluates the inhibitory activity of a TKI against BCR-ABL. Here, we validated the utility of the FRET-based drug sensitivity test carried out at diagnosis for predicting the molecular efficacy. Sixty-two patients with newly diagnosed chronic phase CML were enrolled in this study and treated with dasatinib. Bone marrow cells at diagnosis were subjected to FRET analysis. The ΔFRET value was calculated by subtraction of FRET efficiency in the presence of dasatinib from that in the absence of dasatinib. Treatment response was evaluated every 3 months by the BCR-ABL1 International Scale. Based on the ΔFRET value and molecular response, a threshold of the ΔFRET value in the top 10% of FRET efficiency was set to 0.31. Patients with ΔFRET value ≥0.31 had significantly superior molecular responses (MMR at 6 and 9 months and both MR4 and MR4.5 at 6, 9, and 12 months) compared with the responses in patients with ΔFRET value <0.31. These results suggest that the FRET-based drug sensitivity test at diagnosis can predict early and deep molecular responses. This study is registered with UMIN Clinical Trials Registry (UMIN000006358).

Rapid reduction in BCR-ABL1 transcript predicts deep molecular response in dasatinib-treated chronic-phase chronic myeloid leukaemia patients.

OBJECTIVES: We conducted a phase-II study to evaluate the efficacy and safety of dasatinib in patients newly diagnosed with chronic-phase chronic myeloid leukaemia (CML-CP) in Japan (IMIDAS PART 2 study). METHODS: Seventy-nine patients were administered 100 mg dasatinib once daily. We examined pretreatment and post-treatment influences of various factors. The BCR-ABL1 international scale (IS), halving time (HT) and reduction rate of BCR-ABL1 transcript within the initial 1 or 3 months of therapy (RR-BCR-ABL11m,3m ) were the post-treatment factors investigated to predict the molecular response. RESULTS: The estimated major molecular response (MMR), molecular response 4.0 (MR4.0) and molecular response 4.5 (MR4.5) rates were 77.2%, 49.4% and 35.4%, respectively, at 12 months. Grade 3/4 non-haematologic adverse events were infrequent. Multivariate analysis showed that age >65 years was significantly correlated with MR4.0 and MR4.5 (deep molecular response: DMR) at 12 months. All post-treatment factors at 3 months predicted DMR by univariate analysis. However, RR-BCR-ABL13m was the only significant landmark for predicting DMR by multivariate analysis. CONCLUSIONS: Primary treatment of CML-CP with dasatinib enabled early achievement of MMR and DMR, particularly in elderly patients, with high safety. Furthermore, RR-BCR-ABL13m was found to be a more useful predictor of DMR than HT-BCR-ABL1 and BCR-ABL1 IS.

Synchronous case of follicular lymphoma and Langerhans cell sarcoma in the same lymph node.

Langerhans cell sarcoma (LCS) is a very rare histiocytic and dendritic cell neoplasm originating from Langerhans cells. There are case reports of histiocytic and dendritic cell neoplasms synchronously or sequentially observed in patients with malignant lymphoma. We present a case in which LCS and follicular lymphoma (FL) grade 3a were observed within the same lymph node. A 66-year-old male visited our hospital with a general malaise. Pleural effusion and systemic lymph adenopathy were detected. Biopsy of an inguinal lymph node was performed. The lymph node had regions with follicular structure and regions with acidophilic cytoplasm and large proliferating atypical cells. The tumor cells in the regions with follicular structure showed positivity for CD20 and BCL2 consistent with an FL grade 3a diagnosis. The tumor cells in the regions without follicular structure showed positivity for CD1a and S-100 and were consistent with an LCS diagnosis. Both tumor cells showed the positivity of BCL6 split by FISH. PCR detected IgH clonality in DNA collected from each region, and direct sequence analysis of cells from both tumors detected almost identical amino acid sequences. This finding is important for future research on the development of very rare histiocytic and dendritic cell neoplasms.

[Recurrent invasive pneumococcal disease in a patient with IgG-κ smoldering multiple myeloma].

A 68-year-old female with smoldering multiple myeloma (IgG-κ type) was admitted to the hospital owing to general fatigue, fever, and pain in the right leg. On the day following admission, she developed shock, and a blood culture revealed Streptococcus pneumoniae. She was diagnosed with septic shock and invasive pneumococcal disease (IPD). She received antibiotics and intravenous immunoglobulin and improved after several days. She had a history of recurrent IPD and had received the pneumococcal polysaccharide vaccine 23 (PPSV23) 2 years earlier. Therefore, we inquired with the National Institute of Infectious Diseases if the pneumococcal serotype isolated from her present IPD contained PPSV23. The results showed that her serotype was 19F, a serotype present in PPSV23. We administered pneumococcal conjugate vaccine 13 (PCV13) ; however, she was unable to mount high enough opsonophagocytic assay titers against some serotypes, including 19F. We think she was unable to mount effective humoral immune responses to PPSV23 or PCV13 owing to her underlying disease, smoldering myeloma. It should be considered how IPD can be effectively prevented in patients with multiple myeloma.

Clinicopathological analysis of primary splenic diffuse large B-cell lymphoma.

Splenic infiltration is often seen in diffuse large B-cell lymphoma (DLBCL). However, primary splenic DLBCL is rare and studies on its clinicopathological features are limited. We assessed 66 cases of primary splenic DLBCL and 309 control DLBCL, not otherwise specified. Hepatitis C virus antibody prevalence, B symptoms, poor performance status and CD5 positivity differed significantly between the primary splenic DLBCL and control DLBCL groups. Primary splenic DLBCL cases were classified histopathologically into two groups [white pulp pattern (n = 46), red pulp pattern (n = 20)]. Survival analysis showed no difference in overall survival between the primary splenic DLBCL and the control group, but the former had a more favourable progression-free survival. In the examination of primary splenic DLBCL, the white pulp pattern was statistically associated with a lower performance status (2-4), and a lower CD5 positivity than the red pulp pattern. In the survival analysis, the red pulp pattern demonstrated poorer overall survival. Multivariate analysis of overall survival in primary splenic DLBCL cases identified CD5 positivity as an indicator of poor prognosis. Classifying primary splenic DLBCL into white and red pulp patterns was useful in terms of clinicopathological features and overall survival.

Study protocol of HGCSG1404 SNOW study: a phase I/II trial of combined chemotherapy of S-1, nab-paclitaxel and oxaliplatin administered biweekly to patients with advanced gastric cancer.

BACKGROUND: In Japan, S-1 plus cisplatin (SP) regimen has become a standard therapy for patients with advanced gastric cancer. Moreover, the S-1 plus oxaliplatin regimen is now a standard treatment. Nab-paclitaxel was developed for chemotherapy of gastric cancer in Japanese clinical practice. Nab-paclitaxel, created with albumin-bound paclitaxel particles, has high transferability to tumour tissues and does not cause hypersensitivity reactions because of a different chemical composition compared with docetaxel and paclitaxel. A combination of S-1, nab-paclitaxel and oxaliplatin (which we named 'SNOW regimen') can be a promising triplet therapy for advanced gastric cancer. Although we have to pay attention to chemotherapy-induced neuropathy, we aim to investigate the recommended dose of this regimen in a phase I study. Furthermore, we will investigate its efficacy and toxicity in a phase II study. METHODS: The phase I study is a dose-escalation study using a standard 3 plus 3 design, followed by expansion cohorts. The SNOW regimen involves 28-day cycles with escalated doses of nab-paclitaxel (100-175 mg/m2 on days 1 and 15) and fixed doses of oxaliplatin (65 mg/ m2 on days 1 and 15) and S-1 (80 mg/m2/day on day 1 to 14). The primary endpoints are assessment of dose limiting toxicities and determination of maximum tolerated dose to investigate the recommended dose in the subsequent phase II study. In the phase II study, the primary endpoint is objective response rate. Secondary endpoints are assessment of safety, progression-free survival, disease control rate, overall survival and time to treatment failure. Adverse events were monitored and graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events version 4.0. DISCUSSION: Triplet therapies for advanced gastric cancer patients have been evaluated in clinical trials. The SNOW regimen can be a promising new triplet therapy. TRIAL REGISTRATION: This study is performed at institutes that participate in Hokkaido Gastrointestinal Cancer Study Group (HGCSG) and registered as UMIN000016788 . Registrated 16 March 2015.

Clinicopathological features of primary splenic follicular lymphoma.

Follicular lymphoma (FL) is a low-grade lymphoma that is usually characterized by generalized lymphadenopathy. Extranodal invasion by FL generally involves the bone marrow, skin, and duodenum; splenic infiltration often occurs in the advanced stages. However, primary splenic FL is very rare. Hence, few studies have been performed on splenic FL, and its clinicopathological features have not been established. This study aimed to investigate the clinicopathological features of primary splenic FL, as compared to nodal FL. We analyzed 17 patients diagnosed with primary splenic FL and 153 control patients with systemic FL. Hepatitis C virus (HCV)-positive status was significantly more common in patients with splenic FL than in the control patients (p = 0.02). Ann Arbor stage III or IV (p = 0.0003) and high-risk FLIPI (Follicular Lymphoma International Prognostic Index) (p = 0.03) were significantly less common in patients with splenic FL than in the control patients; however, the overall and progression-free survival curves were not significantly different between the groups. Among the 17 patients with splenic FL, the progression-free survival was significantly worse in patients who underwent splenectomy without receiving postoperative chemotherapy than in those who did (p = 0.03). These results suggest that primary splenic FL should be considered different from systemic FL; accordingly, its management should also be conducted differently.

Prevalence and characteristics of naturally occurring sofosbuvir resistance-associated variants in patients with hepatitis C virus genotype 1b infection.

AIM: Sofosbuvir (SOF), a nucleotide analog pro-drug, targets hepatitis C virus (HCV) NS5B polymerase and shows potential for treating HCV infection, given its high efficacy and good barrier to resistance. However, in addition to the rare resistant-associated variant (RAV) of non-structural protein NS5B S282T, several new potential RAVs of SOF have been reported, especially related to HCV genotype 1b. However, the prevalence and characteristics of these RAVs have not been clarified. METHODS: We analyzed the prevalence of variants in the NS3/NS5A/NS5B regions in 96 patients treated with simeprevir (SMV) combination therapy, and the prevalence of RAVs in patients showing treatment failure was determined by direct- or deep-sequencing methods. Associations between these potential RAVs and clinical factors were also analyzed. RESULTS: Prevalence of NS5B RAV C316N was high (46.9%, 45/96), whereas that of NS5B L159F was relatively low (1.04%, 1/96); however, deep sequencing showed that 30.0% of patients with C316N also had NS5B RAV L159F. Additionally, there was no significant relationship between the existence of potential NS5B and NS5A or NS3 RAVs. However, the presence of NS5B C316N was significantly associated with an HCV core amino acid 91 substitution. No significant difference was detected between each RAV and sustained virological response in simeprevir combination therapy. CONCLUSION: We provide clear evidence of the high prevalence of two potential naturally occurring NS5B RAVs (C316N and L159F) in Japan. It may be important to pay particular attention to these new potential RAVs, especially when using SOF-based therapy in patients with RAVs due to previous direct-acting antiviral therapy failure.