DNA Methylation on N6-Adenine in C. elegans.

In mammalian cells, DNA methylation on the fifth position of cytosine (5mC) plays an important role as an epigenetic mark. However, DNA methylation was considered to be absent in C. elegans because of the lack of detectable 5mC, as well as homologs of the cytosine DNA methyltransferases. Here, using multiple approaches, we demonstrate the presence of adenine N(6)-methylation (6mA) in C. elegans DNA. We further demonstrate that this modification increases trans-generationally in a paradigm of epigenetic inheritance. Importantly, we identify a DNA demethylase, NMAD-1, and a potential DNA methyltransferase, DAMT-1, which regulate 6mA levels and crosstalk between methylations of histone H3K4 and adenines and control the epigenetic inheritance of phenotypes associated with the loss of the H3K4me2 demethylase spr-5. Together, these data identify a DNA modification in C. elegans and raise the exciting possibility that 6mA may be a carrier of heritable epigenetic information in eukaryotes.

Nivolumab Combination Therapy in Advanced Esophageal Squamous-Cell Carcinoma.

BACKGROUND: First-line chemotherapy for advanced esophageal squamous-cell carcinoma results in poor outcomes. The monoclonal antibody nivolumab has shown an overall survival benefit over chemotherapy in previously treated patients with advanced esophageal squamous-cell carcinoma. METHODS: In this open-label, phase 3 trial, we randomly assigned adults with previously untreated, unresectable advanced, recurrent, or metastatic esophageal squamous-cell carcinoma in a 1:1:1 ratio to receive nivolumab plus chemotherapy, nivolumab plus the monoclonal antibody ipilimumab, or chemotherapy. The primary end points were overall survival and progression-free survival, as determined by blinded independent central review. Hierarchical testing was performed first in patients with tumor-cell programmed death ligand 1 (PD-L1) expression of 1% or greater and then in the overall population (all randomly assigned patients). RESULTS: A total of 970 patients underwent randomization. At a 13-month minimum follow-up, overall survival was significantly longer with nivolumab plus chemotherapy than with chemotherapy alone, both among patients with tumor-cell PD-L1 expression of 1% or greater (median, 15.4 vs. 9.1 months; hazard ratio, 0.54; 99.5% confidence interval [CI], 0.37 to 0.80; P<0.001) and in the overall population (median, 13.2 vs. 10.7 months; hazard ratio, 0.74; 99.1% CI, 0.58 to 0.96; P = 0.002). Overall survival was also significantly longer with nivolumab plus ipilimumab than with chemotherapy among patients with tumor-cell PD-L1 expression of 1% or greater (median, 13.7 vs. 9.1 months; hazard ratio, 0.64; 98.6% CI, 0.46 to 0.90; P = 0.001) and in the overall population (median, 12.7 vs. 10.7 months; hazard ratio, 0.78; 98.2% CI, 0.62 to 0.98; P = 0.01). Among patients with tumor-cell PD-L1 expression of 1% or greater, a significant progression-free survival benefit was also seen with nivolumab plus chemotherapy over chemotherapy alone (hazard ratio for disease progression or death, 0.65; 98.5% CI, 0.46 to 0.92; P = 0.002) but not with nivolumab plus ipilimumab as compared with chemotherapy. The incidence of treatment-related adverse events of grade 3 or 4 was 47% with nivolumab plus chemotherapy, 32% with nivolumab plus ipilimumab, and 36% with chemotherapy alone. CONCLUSIONS: Both first-line treatment with nivolumab plus chemotherapy and first-line treatment with nivolumab plus ipilimumab resulted in significantly longer overall survival than chemotherapy alone in patients with advanced esophageal squamous-cell carcinoma, with no new safety signals identified. (Funded by Bristol Myers Squibb and Ono Pharmaceutical; CheckMate 648 ClinicalTrials.gov number, NCT03143153.).

Bintrafusp alfa and chemotherapy as first-line treatment in biliary tract cancer: A randomized phase 2/3 trial.

BACKGROUND AND AIMS: We compared the safety and efficacy of bintrafusp alfa (BA) in combination with gemcitabine+cisplatin (GemCis), to those of GemCis alone, in patients with biliary tract cancer. APPROACH AND RESULTS: This randomized, double-blind, placebo-controlled, adaptive design phase 2/3 trial (NCT04066491) included adults who are treatment-naive with locally advanced/metastatic biliary tract cancer. Patients (N = 297) were randomized to receive an IV infusion of BA (2400 mg once/3 wk) plus GemCis (gemcitabine 1000 mg/m 2 +cisplatin 25 mg/m 2 on days 1 and 8/3 wk; 8 cycles) (BA group, n = 148) or placebo+GemCis (placebo group, n = 149). The primary end point was overall survival (OS). For adaptation analysis (phase 2-phase 3; data cutoff: May 20, 2021), efficacy was assessed in the first 150 patients who were antibiotic-naive when 80 progression-free survival events had occurred and ≥ 19 weeks of follow-up had been completed (BA, n = 73; placebo, n = 77). Median OS (95% CI) for the BA (11.5 mo [9.3-not estimable]) and placebo (11.5 mo [10.0-not estimable]) groups was comparable (hazard ration 1.23 [95% CI 0.66-2.28]; p = 0.7394); OS data maturity was 27.2% (41 events/151 patients). The most common grade ≥3 treatment-related adverse event was anemia (BA, 26.0%; placebo, 22.8%). Bleeding adverse events were reported more frequently in the BA group (28.8%) versus the placebo group (7.4%). Deaths within 60 days of the first dose were reported in 7.5% and 1.3% of patients in the BA and placebo groups, respectively. CONCLUSIONS: BA+GemCis did not provide a clinically meaningful benefit compared with GemCis alone as first-line treatment for biliary tract cancer, and the study was discontinued early (terminated: August 20, 2021).

Sarcopenia and myosteatosis are associated with survival in patients receiving immunotherapy for advanced hepatocellular carcinoma.

OBJECTIVES: To investigate the association of sarcopenia, myosteatosis, and sarcopenic obesity with survival outcomes among patients who underwent immunotherapy for advanced hepatocellular carcinoma (HCC). METHODS: In this retrospective analysis, patients who initiated immunotherapy for advanced HCC were enrolled. Sarcopenia and myosteatosis were evaluated on pretreatment CT at L3 level by skeletal muscle index and mean muscle attenuation using predefined cutoff values. Sarcopenic obesity was defined as concurrent sarcopenia and body mass index > 25 kg/m2. The log-rank test and the Cox proportional hazards model were used to compare overall survival (OS) and progression-free survival (PFS). RESULTS: A total of 138 patients was included (discovery cohort n = 111, validation cohort n = 27). In the discovery cohort, patients with sarcopenia exhibited significantly poorer PFS (p = 0.048) and OS (p = 0.002) than patients without sarcopenia. Patients with myosteatosis exhibited significantly poorer PFS (p < 0.001) and OS (p < 0.001) than patients without myosteatosis. Patients with sarcopenic obesity compared to patients without sarcopenic obesity exhibited significantly poorer OS (p = 0.006) but not PFS (p = 0.31). In multivariate analysis adjusting for patient demographics, tumor extent, and liver function reserve, myosteatosis remained an independent predictor of poor PFS (p = 0.014) and OS (p = 0.007); sarcopenia remained an independent predictor for poor OS (p = 0.007). The prediction models for survival outcomes built by the discovery cohort showed similar performance in the validation cohort. CONCLUSIONS: Sarcopenia and myosteatosis are independent prognostic factors in patients who received immunotherapy for advanced HCC. KEY POINTS: • Sarcopenia and myosteatosis can be evaluated by CT at L3 level. • Sarcopenia, myosteatosis, and sarcopenic obesity were associated with poor survival outcomes in patients who underwent immunotherapy for advanced HCC. • Myosteatosis was an independent predictor of PFS and OS, and sarcopenia was independent for OS in these patients.

RNA m6A methylation regulates the ultraviolet-induced DNA damage response.

Cell proliferation and survival require the faithful maintenance and propagation of genetic information, which are threatened by the ubiquitous sources of DNA damage present intracellularly and in the external environment. A system of DNA repair, called the DNA damage response, detects and repairs damaged DNA and prevents cell division until the repair is complete. Here we report that methylation at the 6 position of adenosine (m6A) in RNA is rapidly (within 2 min) and transiently induced at DNA damage sites in response to ultraviolet irradiation. This modification occurs on numerous poly(A)+ transcripts and is regulated by the methyltransferase METTL3 (methyltransferase-like 3) and the demethylase FTO (fat mass and obesity-associated protein). In the absence of METTL3 catalytic activity, cells showed delayed repair of ultraviolet-induced cyclobutane pyrimidine adducts and elevated sensitivity to ultraviolet, demonstrating the importance of m6A in the ultraviolet-responsive DNA damage response. Multiple DNA polymerases are involved in the ultraviolet response, some of which resynthesize DNA after the lesion has been excised by the nucleotide excision repair pathway, while others participate in trans-lesion synthesis to allow replication past damaged lesions in S phase. DNA polymerase κ (Pol κ), which has been implicated in both nucleotide excision repair and trans-lesion synthesis, required the catalytic activity of METTL3 for immediate localization to ultraviolet-induced DNA damage sites. Importantly, Pol κ overexpression qualitatively suppressed the cyclobutane pyrimidine removal defect associated with METTL3 loss. Thus, we have uncovered a novel function for RNA m6A modification in the ultraviolet-induced DNA damage response, and our findings collectively support a model in which m6A RNA serves as a beacon for the selective, rapid recruitment of Pol κ to damage sites to facilitate repair and cell survival.

Corrigendum: RNA m6A methylation regulates the ultraviolet-induced DNA damage response.

This corrects the article DOI: 10.1038/nature21671.

Chyloperitoneum signifying late bowel obstruction following gastric clipping with proximal jejunal bypass: A case report.

Chyloperitoneum (CP) is a rare complication after bariatric surgery. We present a 37-year-old female with CP caused by a bowel volvulus following a gastric clipping with proximal jejunal bypass for morbid obesity. An abdominal CT image of a mesenteric swirl sign and abnormal triglyceride level of ascites fluid can confirm the diagnosis. In this patient, laparoscopy demonstrated dilated lymphatic ducts caused by a bowel volvulus resulting in the exudation of chylous fluid into the peritoneal cavity. After the reduction of bowel volvulus, she made an uneventful recovery with complete resolution of the chylous ascites. The presence of CP could indicate a situation of small bowel obstruction in patients with a history of bariatric surgery.

Ramucirumab for Patients with Advanced Hepatocellular Carcinoma and Elevated Alpha Fetoprotein Following Non-Sorafenib Systemic Therapy: An Expansion Cohort of REACH-2.

BACKGROUND: Ramucirumab is indicated for patients with advanced hepatocellular carcinoma (HCC) and α-fetoprotein (AFP) ≥400 ng/mL following sorafenib. Here, we prospectively studied ramucirumab following non-sorafenib systemic therapies. MATERIALS AND METHODS: This open-label, non-comparative cohort of REACH-2 enrolled patients with advanced HCC, Child-Pugh class-A liver disease, and AFP ≥400 ng/mL who had received 1-2 lines of therapy, excluding sorafenib or chemotherapy. Ramucirumab was administered 8 mg/kg intravenously Q2W. The primary endpoint was safety. Secondary endpoints were overall survival, progression-free survival, objective response rate (RECIST v1.1), time to progression, pharmacokinetics, and patient-reported outcomes. Final analysis occurred after all enrolled patients completed ≥3 treatment cycles or discontinued treatment. RESULTS: Between April 27, 2018, and March 29, 2021, 47 patients were treated at 21 investigative sites in Asia, Europe, and USA. The most frequently reported grade ≥3 adverse events, regardless of causality, were hypertension (11%), proteinuria (6%), hyponatremia (6%), and AST increased (6%). Two patients died from adverse events (myocardial infarction and upper gastrointestinal hemorrhage), deemed related to treatment. Median progression-free survival, time to progression, and overall survival were 1.7 months, 2.8 months, and 8.7 months, respectively. The objective response rate was 10.6% with a median duration response of 8.3 months. Median time to deterioration in FHSI-8 total score was 4.4 months. CONCLUSION: Ramucirumab demonstrated consistent and meaningful clinical activity with no new safety signals following non-sorafenib therapies in patients with advanced HCC and AFP ≥400 ng/mL. This represents one of the first sequencing studies for patients with advanced HCC not treated with sorafenib.

Cyclin-Dependent Kinase 9 Inhibition as a Potential Treatment for Hepatocellular Carcinoma.

PURPOSE: Composite cyclin-dependent kinase (CDK) inhibition has shown potential as a treatment for hepatocellular carcinoma (HCC) in preclinical studies. We tested whether the specific inhibition of CDK9 was effective against HCC. METHODS: The effects of two specific CDK9 inhibitors, BAY1143572 and AZD4573, in HCC cell lines were examined. We tested the in vivo efficacy of CDK9 inhibition in mouse xenograft models of HuH7 human HCC cells and in an orthotopic model of BNL mouse HCC cells. Overexpression and knockdown of CDK9 were performed to confirm the efficacy of CDK9 inhibition. RESULTS: CDK9 inhibitors exhibited potent antiproliferative activities in HCC cells regardless of the levels of c-myc expression while inhibiting the downstream signals of CDK9, such as the phosphorylation of RNA polymerase II. These 2 CDK9 inhibitors induced apoptosis in HCC cells and reduced the expression of antiapoptotic proteins such as myeloid cell leukemia-1 and survivin. In the xenograft studies, mice receiving either CDK9 inhibitor exhibited significantly slower tumor growth than did the mice receiving vehicles. In the orthotopic model, the HCC growth in mice receiving a CDK9 inhibitor also tended to be slower than that in the control group. Overexpression of CDK9 in HuH7 cells reduced the efficacy of both CDK9 inhibitors. Knockdown of CDK9 expression reduced the proliferative activities of HCC cells. CONCLUSION: We demonstrated the in vitro and in vivo activity of CDK9 inhibition on multiple HCC cell lines. Our data support further clinical development of CDK9 inhibitors as a treatment for HCC.

ICOS-Positive Regulatory T Cells in Hepatocellular Carcinoma: The Perspective from Digital Pathology Analysis.

INTRODUCTION: Inducible co-stimulator (ICOS), an important co-stimulatory receptor on effector T cells (Teffs), may also contribute to tumor growth due to its high expression on regulatory T cells (Tregs). This study explored the clinical significance of ICOS-expressing Tregs in hepatocellular carcinoma (HCC). METHODS: Tumor tissues from HCC patients who received curative hepatectomy were obtained at a referral center. Dual immunohistochemistry was performed to evaluate the expression of ICOS and Foxp3. The cell densities and proximities between stained cells in regions of interest were measured by digital pathology and the associations with clinical outcome were analyzed. RESULTS: A total of 142 patients (male:female = 112: 30, median age of 61.0 years) were enrolled. Among them, 87 (61.3%) had chronic hepatitis B virus infection and 33 (23.2%) had chronic hepatitis C infection. Low α-fetoprotein level (<20 ng/mL) and early-stage were significantly associated with improved overall survival (OS). The density of ICOS+Foxp3+ cells and the ratio of ICOS+Foxp3+/total Foxp3+ cells were significantly higher (p < 0.001) in the tumor center than in the peritumor area. Patients with a high density of ICOS+Foxp3+ cells or a high ratio of ICOS+Foxp3+/total Foxp3+ cells in the tumor center trended to have a shorter OS. A shorter distance between ICOS+Foxp3+ cells and ICOS+Foxp3- cells (likely Teffs) in the tumor center was significantly associated with a shorter OS (p = 0.030), suggesting active immunosuppression of ICOS+ Tregs on ICOS+ Teffs. CONCLUSION: An increased abundance of ICOS+ Tregs in the tumor center in comparison to the peritumor area indicates a strong immunosuppressive tumor microenvironment of HCC. A high proportion of ICOS+Foxp3+ cells and a shorter distance between ICOS+ Tregs and other ICOS+ cells were associated with a poor OS, suggesting that depleting ICOS+ Tregs might provide clinical benefit for patients with HCC.

Association of annexin A10 expression with poor prognosis of intrahepatic cholangiocarcinoma.

BACKGROUND: Annexin A10 expression influences the prognosis of several gastrointestinal cancers. We explored the association of annexin A10 expression with the overall survival (OS) of patients who underwent curative surgery for cholangiocarcinoma. METHODS: Patients who underwent curative surgery for cholangiocarcinoma (except gallbladder cancer) and had pathological stage T1-3N0M0 disease were enrolled. Annexin A10 expression was examined by performing immunohistochemical staining. Patient demographics and survival outcome data were retrieved from medical records. RESULTS: In total, 185 patients were enrolled. The primary tumor location was intrahepatic and extrahepatic (including the perihilar region) for 89% and 11% of patients, respectively. Positive annexin A10 staining was detected for 61 (33%) patients and associated with extrahepatic or perihilar cholangiocarcinoma (p = 0.001) and lower histological grade (p < 0.001). Patients with positive annexin A10 staining exhibited significantly poorer survival relative to patients with negative staining results (median OS, 2.5 vs. 4.9 years, p = 0.025). In the multivariate analysis adjusting for age, sex, tumor location, tumor grade, hepatitis infection, and disease stage, positive annexin A10 remained an independent predictor of poor OS (hazard ratio 1.572, p = 0.034). In the subgroup analysis, the association between annexin A10 and prognosis was restricted to intrahepatic cholangiocarcinoma. Among patients with intrahepatic cholangiocarcinoma, patients with positive annexin A10 staining exhibited significantly poorer survival compared with patients with negative annexin A10 staining (median OS, 2.3 vs. 4.9 years, p = 0.008). CONCLUSION: Positive annexin A10 expression was associated with poor prognosis of intrahepatic cholangiocarcinoma.

Bevacizumab and atezolizumab as first-line therapy for advanced hepatocellular carcinoma: A Taiwanese subgroup analysis on efficacy and safety.

BACKGROUND: The combination of bevacizumab and atezolizumab has been established as a standard first-line systemic treatment for unresectable hepatocellular carcinoma (HCC). We examined the treatment outcomes of patients in Taiwan who received the combination in 2 pivotal clinical trials. METHODS: All patients who resided in Taiwan, were enrolled in the IMbrave150 and GO30140 studies, and received bevacizumab and atezolizumab as the first-line systemic therapy for unresectable HCC were included. We extracted and pooled anonymous raw data from the study records. RESULTS: We enrolled 40 patients, with the median age of 62.5 years; 36 (90%) had Barcelona Clinic Liver Cancer stage C disease. The response rate was 37.5%, including 3 (7.5%) complete responses. The disease control rate was 85%. The median duration of response was 21.4 months (95% confidence interval [CI], 16.6-not estimable). The median progression-free survival (PFS) and overall survival (OS) were 8.6 (95% CI, 5.6-18.6) and 24.9 months (95% CI, 14.2-not estimable), respectively. The most common adverse events of all grades were proteinuria (50%) and hypertension (37.5%), the median onset of which were 157 and 127 days, respectively. Bevacizumab and atezolizumab treatment had to be interrupted in 20 (50%) and 13 (32.5%) patients, respectively. Among patients whose treatment duration was ≥6 months, 50% of them had to skip bevacizumab, but no signal of poorer PFS or OS was observed. CONCLUSION: In Taiwanese patients with advanced HCC, the efficacy and safety outcomes of bevacizumab and atezolizumab treatment were generally consistent with the global intent-to-treat populations.

Deleterious alterations of DNA damage response and repair genes and clinical benefit to anti-PD-1 therapy in esophageal squamous cell carcinoma.

BACKGROUND: Although DNA damage response and repair (DDR) gene alteration has been demonstrated as a biomarker for anti-PD-1 therapy in several cancer types, its role in esophageal squamous cell carcinoma (ESCC) is unknown. METHODS: Patients with advanced ESCC treated with anti-PD-1-based immunotherapy were enrolled. Tumor response was evaluated according to RECIST 1.1. Archival ESCC tissues were analyzed using FoundationOne CDx. Deleterious alterations, defined by loss of function, of DDR genes were correlated with patient survival by Cox proportional hazards model. The prognostic significance of deleterious alterations of DDR genes in The Cancer Genome Atlas (TCGA)-ESCC cohort was explored. RESULTS: Forty-three patients were enrolled. The objective response rate (ORR) was 19%. The median tumor mutational burden was 4 mutations/Mb (0-20); none of the tumors were microsatellite instable. Compared with patients with wild-type or other alterations of DDR genes (N = 35, 81%), those with deleterious alterations of DDR genes (N = 8, 19%) had a higher ORR (38 vs. 14%), longer median progression-free survival (4.1 vs. 2.0 months), and significantly longer median overall survival (OS; 27.7 vs. 6.1 months, P = 0.011). In multivariate analysis, harboring deleterious alterations of DDR genes was a favorable prognostic factor for OS (HR = 0.31 [95% CI: 0.11-0.91], P = 0.033). In the TCGA-ESCC cohort, the presence of deleterious alterations of DDR genes was not a favorable prognostic factor. CONCLUSIONS: Deleterious alterations of DDR genes may be associated with improved prognosis and efficacy of anti-PD-1 therapy in patients with advanced ESCC.

Serum alpha-fetoprotein and clinical outcomes in patients with advanced hepatocellular carcinoma treated with ramucirumab.

BACKGROUND: Post hoc analyses assessed the prognostic and predictive value of baseline alpha-fetoprotein (AFP), as well as clinical outcomes by AFP response or progression, during treatment in two placebo-controlled trials (REACH, REACH-2). METHODS: Serum AFP was measured at baseline and every three cycles. The prognostic and predictive value of baseline AFP was assessed by Cox regression models and Subpopulation Treatment Effect Pattern Plot method. Associations between AFP (≥ 20% increase) and radiographic progression and efficacy were assessed. RESULTS: Baseline AFP was confirmed as a continuous (REACH, REACH-2; p < 0.0001) and dichotomous (≥400 vs. <400 ng/ml; REACH, p < 0.01) prognostic factor, and was predictive for ramucirumab survival benefit in REACH (p = 0.0042 continuous; p < 0.0001 dichotomous). Time to AFP (hazard ratio [HR] 0.513; p < 0.0001) and radiographic (HR 0.549; p < 0.0001) progression favoured ramucirumab. Association between AFP and radiographic progression was shown for up to 6 (odds ratio [OR] 5.1; p < 0.0001) and 6-12 weeks (OR 1.8; p = 0.0065). AFP response was higher with ramucirumab vs. placebo (p < 0.0001). Survival was longer in patients with an AFP response than patients without (13.6 vs. 5.6 months, HR 0.451; 95% confidence interval, 0.354-0.574; p < 0.0001). CONCLUSIONS: AFP is an important prognostic factor and a predictive biomarker for ramucirumab survival benefit. AFP ≥ 400 ng/ml is an appropriate selection criterion for ramucirumab. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, REACH (NCT01140347) and REACH-2 (NCT02435433).

An Underdiagnosed Hypothyroidism and Its Clinical Significance in Patients with Advanced Hepatocellular Carcinoma.

BACKGROUND: Many systemic therapies for advanced hepatocellular carcinoma (HCC) may cause hypothyroidism; however, in these patients, hypothyroidism prevalence before therapy and its prognostic impact remain unclear. MATERIALS AND METHODS: We previously established a prospective cohort of patients who received sorafenib as first-line therapy for advanced HCC. No patients had been clinically diagnosed with hypothyroidism before or during sorafenib treatment. We retrospectively determined the levels of thyrotropin and free thyroxine before initiation of systemic therapy. Hypothyroidism was defined as thyrotropin level higher than the upper limit of the normal range. Among patients with hypothyroidism, free thyroxine level less than the lower normal range was defined as overt hypothyroidism, and free thyroxine level within the normal range was defined as subclinical hypothyroidism. RESULTS: In total, 79 patients were enrolled; of them, 16 (20%) had hypothyroidism (overt hypothyroidism, 10; subclinical hypothyroidism, 6). Patients with hypothyroidism, compared with those without hypothyroidism, were more likely to be older than 65 years (56% vs. 29%, p = .037), have a serum α-fetoprotein level of >400 ng/mL (81% vs. 52%, p = .037), and have a significantly poorer overall survival (OS; median, 5.5 vs. 11.6 months, p = .043). After adjusting for other potential prognostic factors, hypothyroidism remained an independent predictor for poorer OS (hazard ratio, 2.53, p = .018). Patients with overt hypothyroidism and subclinical hypothyroidism exhibited similarly poor OS (p = .768). CONCLUSION: Underdiagnosis of hypothyroidism in patients with advanced HCC was common. Hypothyroidism, whether overt or subclinical, is associated with poor prognosis of advanced HCC. IMPLICATIONS FOR PRACTICE: The results of this study showed the underdiagnosis of hypothyroidism in patients with advanced hepatocellular carcinoma (HCC) and its influence on prognosis. These findings implied the importance of thyroid function check before initiation of systemic therapy for patients with advanced HCC.

Response to Immune Checkpoint Inhibitors in Recurrent or Metastatic Esophageal Squamous Cell Carcinoma May Be Affected by Tumor Sites.

INTRODUCTION: Heterogeneous tumor response has been reported in cancer patients treated with immune checkpoint inhibitors (ICIs). This study investigated whether the tumor site is associated with the response to ICIs in patients with recurrent or metastatic esophageal squamous cell carcinoma (ESCC). METHODS: Patients with ESCC who had measurable tumors in the liver, lung, or lymph node (LN) according to the response evaluation criteria in solid tumors (RECIST) 1.1 and received ICIs at 2 medical centers in Taiwan were enrolled. In addition to RECIST 1.1, tumor responses were determined per individual organ basis according to organ-specific criteria modified from RECIST 1.1. Fisher test or χ2 test was used for statistical analysis. RESULTS: In total, 37 patients were enrolled. The overall response rate per RECIST 1.1 was 13.5%. Measurable tumors in the LN, lung, and liver were observed in 26, 17, and 13 patients, respectively. The organ-specific response rates were 26.9%, 29.4%, and 15.4% for the LN, lung, and liver tumors, respectively (p = 0.05). The organ-specific disease control rates were 69.2%, 52.9%, and 21.1% for the LN, lung, and liver tumors, respectively (p = 0.024). Five (27.8%) among 18 patients harboring at least 2 involved organs had heterogeneous tumor response. CONCLUSION: The response and disease control to ICIs may differ in ESCC tumors located at different metastatic sites, with a lesser likelihood of response and disease control in metastatic liver tumors than in tumors located at the LNs and lung.

White shrimp Litopenaeus vannamei that have received mixtures of heat-killed and formalin-inactivated Vibrio alginolyticus and V. harveyi exhibit recall memory and show increased phagocytosis and resistance to Vibrio infection.

Heat-killed Vibrio alginolyticus (HVa), formalin-inactivated V. alginolyticus (FVa), heat-killed Vibrio harveyi (HVh), formalin-inactivated V. harveyi (FVh), live V. alginolyticus (LVa), and live V. harveyi (LVh) were used in this study. White shrimp Litopenaeus vannamei receiving two mixtures (HVa + FVa) or four mixtures (HVa + FVa + HVh + FVh) served as primary exposure, and shrimp receiving LVa or LVh afterward served as secondary exposure. Shrimp receiving marine saline and then receiving either LVa or LVh served as controls. Phagocytic activity and clearance efficiency were examined in shrimp that received two mixtures after 1-8 weeks and then received LVa. Both the phagocytic activity and clearance efficiency of shrimp receiving two mixtures were significantly higher than in control shrimp after 1-8 weeks. In another experiment, phagocytic activity and clearance efficiency were examined in shrimp that received four mixtures after 1-8 weeks and then received LVa and LVh, respectively. The phagocytic activity of shrimp receiving four mixtures was significantly higher than in control shrimp after 1-8 weeks post exposure to LVa and LVh. The clearance efficiency of shrimp receiving four mixtures was significantly higher than in control shrimp after 1-6 weeks post exposure to LVa, and 1-7 weeks post exposure to LVh. In the other experiment, the survival rate of shrimp that received four mixtures after five weeks were challenged with LVa at 6.4 × 107 colony-forming units (cfu) shrimp-1 and LVh at 4.4 × 106 cfu shrimp-1. Shrimp that received marine saline for five weeks and then challenged with LVa and LVh at a same dose served as challenged controls. The survival rate of shrimp that received four mixtures was significantly higher (90%) than that of control shrimp (67%), and significantly higher (73%) than that of control shrimp (53%) after 3-7 days post challenge with LVa and LVh. It is concluded that the mixtures have feature of adjuvant and antigen, and shrimp receiving mixtures of heat-killed and formalin-inactivated V. alginolyticus and V. harveyi even after 5-8 weeks exhibit memory recall and show increased phagocytosis and resistance to Vibrio infections.

Solving the deficit of cancer pain management skills by education programs.

BACKGROUND: Suboptimal cancer pain management is a worldwide problem. We examined whether an educational program on cancer pain management implemented during training could benefit primary care physicians. METHODS: We enrolled all the primary care physicians who visited the oncology ward at a medical center for the first time. Educational classes on cancer pain management were conducted. The participants' abilities in cancer pain management were measured in a pretest before the classes and approximately 2 weeks later in the first posttest. The second posttest was conducted on participants who visited the oncology ward again. All 3 tests had the same set of questions and were scored on a scale of 0 to 100. RESULTS: In total, 247 participants were enrolled. Less than 10% of them considered their previous education on cancer pain management adequate. The test scores increased significantly from the pretest to the first posttest (mean 65.6 vs. 89.7, p < 0.001). The participants' self-reported cancer pain management abilities, on a scale of 0 to 100, also improved significantly (mean 57.8 vs. 75.5, p < 0.001). The pretest scores were not associated with the participants' self-reported abilities or their perceptions about the adequacy of previous training on cancer pain management. The mean score on the second posttest, conducted 234.5 days after the program, on an average, remained similar to that of the first posttest (p = 0.254). CONCLUSION: A specific educational program on cancer pain management provided to primary care physicians improved their pain management skills substantially, with persistent effects.

Ash2l interacts with Oct4-stemness circuitry to promote super-enhancer-driven pluripotency network.

Pluripotency and cell fates can be modulated through the regulation of super-enhancers; however, the underlying mechanisms are unclear. Here, we showed a novel mechanism in which Ash2l directly binds to super-enhancers of several stemness genes to regulate pluripotency and self-renewal in pluripotent stem cells. Ash2l recruits Oct4/Sox2/Nanog (OSN) to form Ash2l/OSN complex at the super-enhancers of Jarid2, Nanog, Sox2 and Oct4, and further drives enhancer activation, upregulation of stemness genes, and maintains the pluripotent circuitry. Ash2l knockdown abrogates the OSN recruitment to all super-enhancers and further hinders the enhancer activation. In addition, CRISPRi/dCas9-mediated blocking of Ash2l-binding motifs at these super-enhancers also prevents OSN recruitment and enhancer activation, validating that Ash2l directly binds to super-enhancers and initiates the pluripotency network. Transfection of Ash2l with W118A mutation to disrupt Ash2l-Oct4 interaction fails to rescue Ash2l-driven enhancer activation and pluripotent gene upregulation in Ash2l-depleted pluripotent stem cells. Together, our data demonstrated Ash2l formed an enhancer-bound Ash2l/OSN complex that can drive enhancer activation, govern pluripotency network and stemness circuitry.

The unique characteristic in peripheral immune cells in patients with advanced hepatocellular carcinoma.

BACKGROUND/PURPOSE: Recent progress in cancer immunology provides more insight in immune evasion of cancer cells. Cancer cells may achieve immune evasion through several ways including ineffective antigen presentation, T cell checkpoint utilization, immunosuppressive cytokines secretion and immunosuppressive cells recruitment. However, few literatures mentioned about the change of peripheral blood immune cells in advanced hepatocellular carcinoma (HCC) patients. To answer this question, we initiated a pilot study through detailed flow cytometry. METHODS: We enrolled patients with advanced HCC patients who had informed consent to the collection of their peripheral blood. We also recruited healthy individuals for the control group. Using flow cytometry, we analyzed lymphocyte subclasses and the PD-1 or PD-L1 positivity of immune cells in peripheral blood from HCC patients and healthy individuals. RESULTS: Twenty-four HCC patients were enrolled and twenty healthy individuals were enrolled. Most of the HCC patients were HBV carrier (58.3%), and the mean age was 61 years old. Among 55 immune cell parameters we examined in peripheral blood, 16 were significantly different between advanced HCC patients and healthy individuals by univariate analysis. Multivariate analysis was then conducted by fitting logistic regression model and showed that CD69-CD25- Naïve CD4αßT cell percentage and dendritic cell percentage can reasonably predict the advanced HCC status from peripheral blood. By our regression model, the adjusted generalized R2 = 0.918 and the estimated area under the Receiver Operating Characteristic (ROC) curve was 0.99. CONCLUSION: CD69-CD25- Naïve CD4αßT cell percentage and dendritic cell percentage in peripheral blood are highly correlated with the advanced HCC status. The change may result from immune evasion initiated by hepatocellular carcinoma cells and further investigation is warranted. Validation study is ongoing and this mechanism may be utilized to treat advanced HCC patient in the future.