The cost-effectiveness of zolbetuximab in CLDN18.2-positive gastric or gastroesophageal junction adenocarcinoma.

Aim: To estimate the cost-effectiveness of zolbetuximab plus capecitabine/oxaliplatin (CAPOX) in CLDN18.2-positive, HER2-negative, mG/GEJ adenocarcinoma from the perspective of Chinese payers.Materials & methods: A partitioned survival model was developed to assess the costs, quality-adjusted life years (QALYs) and incremental cost-effectiveness ratios (ICER) of zolbetuximab plus CAPOX versus placebo plus CAPOX. Sensitivity analyses were performed to test the robustness of model.Results: Zolbetuximab plus CAPOX gained an additional cost of $91,551 and an extra health benefit of 0.24 QALY over placebo plus CAPOX, producing an ICER of $388,186/QALY, which exceeded the willingness-to-pay threshold of $38,223/QALY. Sensitivity analysis shows that the model was generally robust.Conclusion: Zolbetuximab plus CAPOX would not be a cost-effective first-line treatment regimen in CLDN18.2-positive, HER2-negative, mG/GEJ adenocarcinoma in China.

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Safety and Pharmacokinetic Assessment of the FIC CLDN18.2/4-1BB Bispecific Antibody in Rhesus Monkeys.

Gastric cancer is one of the most common cancers worldwide, particularly in China, with over half a million new cases and over 400 thousand deaths in 2022. Zolbetuximab, a first-in-class investigational monoclonal antibody (mAb) targeting tumor-associated antigen CLDN18.2 which is highly expressed on gastric cancer cells, was recently reported to meet the primary endpoint in Phase III trial as first-line treatment in CLDN18.2 positive and HER2-negative gastric cancers. In the present study, we developed a humanized bispecific antibody (bsAb) CLDN18.2/4-1BB named PM1032. PM1032 activates immune cells via CLDN18.2 mediated crosslinking of 4-1BB, a potent stimulator of T/NK cells. It induced strong immunological memory in multiple tumor-bearing animal models, indicating significant potential as an effective treatment for CLDN18.2 positive cancers such as gastric cancer. Since liver and gastrointestinal (GI) related toxicities were reported in 4-1BB and CLDN18.2 targeting programs during the clinical development, respectively, extensive pharmacokinetics (PK) and safety profile characterization of PM1032 was performed in rhesus monkeys. PM1032 had a half-life comparable to a conventional IgG1 mAb, and serum drug concentration increased in a dose-dependent pattern. Furthermore, PM1032 was generally well tolerated, with no significant abnormalities observed in toxicity studies, including the liver and stomach. In summary, PM1032 demonstrated good PK and an exceptional safety profile in rhesus monkeys supporting further investigation in clinical studies.

Spatially Resolved Single-Cell Assessment of Pancreatic Cancer Expression Subtypes Reveals Co-expressor Phenotypes and Extensive Intratumoral Heterogeneity.

Pancreatic ductal adenocarcinoma (PDAC) has been classified into classical and basal-like transcriptional subtypes by bulk RNA measurements. However, recent work has uncovered greater complexity to transcriptional subtypes than was initially appreciated using bulk RNA expression profiling. To provide a deeper understanding of PDAC subtypes, we developed a multiplex immunofluorescence (mIF) pipeline that quantifies protein expression of six PDAC subtype markers (CLDN18.2, TFF1, GATA6, KRT17, KRT5, and S100A2) and permits spatially resolved, single-cell interrogation of pancreatic tumors from resection specimens and core needle biopsies. Both primary and metastatic tumors displayed striking intratumoral subtype heterogeneity that was associated with patient outcomes, existed at the scale of individual glands, and was significantly reduced in patient-derived organoid cultures. Tumor cells co-expressing classical and basal markers were present in > 90% of tumors, existed on a basal-classical polarization continuum, and were enriched in tumors containing a greater admixture of basal and classical cell populations. Cell-cell neighbor analyses within tumor glands further suggested that co-expressor cells may represent an intermediate state between expression subtype poles. The extensive intratumoral heterogeneity identified through this clinically applicable mIF pipeline may inform prognosis and treatment selection for patients with PDAC. SIGNIFICANCE: A high-throughput pipeline using multiplex immunofluorescence in pancreatic cancer reveals striking expression subtype intratumoral heterogeneity with implications for therapy selection and identifies co-expressor cells that may serve as intermediates during subtype switching.

Clostridium perfringens beta2 toxin induced in vitro oxidative damage and its toxic assessment in porcine small intestinal epithelial cell lines.

Clostridium perfringens beta2 (CPB2), a key virulence factor, is produced by C. perfringens type C that is the main pathogenic microorganism causing diarrhea in piglets. However, little is known concerning the toxic damage effect of CPB2 on intestinal cells of piglets. In present study, CPB2 toxin obtained by genetic recombination technology was evaluated for its cytotoxicity property using the intestinal porcine epithelial (IPEC-J2) cells, which aims to attempt to understand and explain its mechanism of action in porcine small intestinal epithelial cells. IPEC-J2 cells were treated with different concentrations of CPB2 toxin (5, 10, 20, 30, 40, and 50 µg/mL), and MTT assay results showed that the cell viability of CPB2-treated IPEC-J2 cells decreased in a dose-dependent manner. Lactate dehydrogenase (LDH) assay results revealed that CPB2 significantly increased the LDH release, relative to the control. The expression of tumor necrosis factor α (TNF-α) and interleukin 8 (IL-8) gradually increased, while the expression of interleukin 10 (IL-10) gradually decreased in IPEC-J2 cells with increasing concentration of CPB2 (10-30 µg/mL), as analyzed by quantitative real-time PCR (RT-qPCR). Also, CPB2 increased the content of intracellular reactive oxygen species (ROS) and decreased mitochondrial membrane potential (ΔΨm) of IPEC-J2 cells. Western blot and immunofluorescence results demonstrate that CPB2 decreased the expression of zonula occludens (ZO-1), claudin12 (CLDN12) and occludin (OCLN) in IPEC-J2 cells. In addition, CPB2 increased Bax expression, and inhibited Bcl-2 and Bcl-xL expression, as measured by Western blot. Considering all of these findings, it was concluded that CPB2 toxin shows significant cytotoxicity, cell growth inhibition and increase in cell permeability in IPEC-J2 cells in a concentration-dependent manner, thus leading to abnormal cell apoptosis and functions in porcine small intestinal epithelial cells.

A novel scoring system for gastric cancer risk assessment based on the expression of three CLIP4 DNA methylation-associated genes.

Gastric cancer (GC) is the fifth most common cancer and the third leading cause of cancer-associated mortality worldwide. In the current study, comprehensive bioinformatic analyses were performed to develop a novel scoring system for GC risk assessment based on CAP-Gly domain containing linker protein family member 4 (CLIP4) DNA methylation status. Two GC datasets with methylation sequencing information and mRNA expression profiling were downloaded from the The Cancer Genome Atlas and Gene Expression Omnibus databases. Differentially expressed genes (DEGs) between the CLIP4 hypermethylation and CLIP4 hypomethylation groups were screened using the limma package in R 3.3.1, and survival analysis of these DEGs was performed using the survival package. A risk scoring system was established via regression factor-weighted gene expression based on linear combination to screen the most important genes associated with CLIP4 methylation and prognosis. Genes associated with high/low-risk value were selected using the limma package. Functional enrichment analysis of the top 500 DEGs that positively and negatively associated with risk values was performed using DAVID 6.8 online and the gene set enrichment analysis (GSEA) software. In total, 35 genes were identified to be that significantly associated with prognosis and CLIP4 DNA methylation, and three prognostic signature genes, claudin-11 (CLDN11), apolipoprotein D (APOD), and chordin like 1 (CHRDL1), were used to establish a risk assessment system. The prognostic scoring system exhibited efficiency in classifying patients with different prognoses, where the low-risk groups had significantly longer overall survival times than those in the high-risk groups. CLDN11, APOD and CHRDL1 exhibited reduced expression in the hypermethylation and low-risk groups compare with the hypomethylation and high-risk groups, respectively. Multivariate Cox analysis indicated that risk value could be used as an independent prognostic factor. In functional analysis, six functional gene ontology terms and five GSEA pathways were associated with CLDN11, APOD and CHRDL1. The results established the credibility of the scoring system in this study. Additionally, these three genes, which were significantly associated with CLIP4 DNA methylation and GC risk assessment, were identified as potential prognostic biomarkers.

Population genetics and comparative genetics of CLDN1, a gene involved in hepatitis C virus entry.

The claudin-1 gene (CLDN1) is a member of a family of genes that encodes proteins found in tight junctions and it has recently been implicated as one of several receptors for late stage binding of hepatitis C virus (HCV). Exploration of the population genetics of this gene could be informative, especially in the investigation of a possible genetic contribution to HCV infection. Comparison to a highly similar gene, claudin-7 (CLDN7) could provide insight into the recent molecular evolution of CLDN1. Mean interspecies conservation score was 0.11 (SD 0.28) for CLDN1 and 0.31 (SD 0.43) for CLDN7. Re-sequence analysis was performed across all exons and evolutionarily conserved regions in CLDN1 (13 kb in total) and CLDN7 (2 kb in total) in 204 chromosomes drawn from the SNP500Cancer resource of four self-described ethnic groups in the US. For CLDN1, 133 SNPs were identified as well as 8 indels and an AC repeat length polymorphism. For CLDN7, 5 SNPs were identified. Assessment of nucleotide diversity (including F(st), theta and pi statistics) did not show evidence for recent positive or negative selection in either gene. The pattern of linkage disequilibrium was determined for each group and there is substantial difference for common SNPS (>5%) between populations as well as genes, further supporting the absence of signatures of recent selection.

Assessment of ileal epithelial P-glycoprotein dysfunction induced by ischemia/reperfusion using in vivo animal model.

We presented the ischemia/reperfusion (I/R) model which can evaluate changes in P-glycoprotein (P-gp) function induced by lipid peroxidation using surgical-sutures connected with the spring balance. The superior mesenteric artery and vein was occluded by hanging itself using surgical-sutures connected with the spring balance for 60 min (ischemia), followed by reperfusion by cutting of sutures. To determine the hanging force of blood vessel during ischemia, treatment at the hanging force of 50g load, 100g load and 150g load for 60 min was carried out and survival rate was evaluated. Although our 150g load group had some effect on survival, the survival was 100% in the case of 50g and 100g load groups. Thiobarbituric acid-reactive substance (TBA-RS) as an indicator of lipid peroxidation and P-gp expression level after I/R was increased and decreased in a load-dependent manner during ischemia, respectively. Also, the decrease in the level of mdr1a mRNA and function of P-gp by I/R depended on load during ischemia. The changes in TBA-RS, P-gp expression level and P-gp function observed in this study corresponded with our in vitro I/R model reported previously. In conclusion, it was shown that this in vivo I/R model can evaluate the function of P-gp through lipid peroxidation.

Probing effects of pH change on dynamic response of Claudin-2 mediated adhesion using single molecule force spectroscopy.

Claudins belong to a large family of transmembrane proteins that localize at tight junctions (TJs) where they play a central role in regulating paracellular transport of solutes and nutrients across epithelial monolayers. Their ability to regulate the paracellular pathway is highly influenced by changes in extracellular pH. However, the effect of changes in pH on the strength and kinetics of claudin mediated adhesion is poorly understood. Using atomic force microscopy, we characterized the kinetic properties of homophilic trans-interactions between full length recombinant GST tagged Claudin-2 (Cldn2) under different pH conditions. In measurements covering three orders of magnitude change in force loading rate of 10(2)-10(4) pN/s, the Cldn2/Cldn2 force spectrum (i.e., unbinding force versus loading rate) revealed a fast and a slow loading regime that characterized a steep inner activation barrier and a wide outer activation barrier throughout pH range of 4.5-8. Comparing to the neutral condition (pH 6.9), differences in the inner energy barriers for the dissociation of Cldn2/Cldn2 mediated interactions at acidic and alkaline environments were found to be <0.65 k(B)T, which is much lower than the outer dissociation energy barrier (>1.37 k(B)T). The relatively stable interaction of Cldn2/Cldn2 in neutral environment suggests that electrostatic interactions may contribute to the overall adhesion strength of Cldn2 interactions. Our results provide an insight into the changes in the inter-molecular forces and adhesion kinetics of Cldn2 mediated interactions in acidic, neutral and alkaline environments.

Comparison of CT enhancement patterns and histologic features in hepatocellular carcinoma up to 2 cm: assessment of malignant potential with claudin-10 immunohistochemistry.

The purpose of this study was to compare features of computed tomography (CT) and histologic features of hepatocellular carcinoma (HCC) of up to 2 cm in diameter with the immunohistochemical staining pattern of claudin-10, a recently identified prognostic factor for resected HCC. Twelve cases of resected HCC of up to 2 cm in diameter were assessed retrospectively. Preoperative dynamic CT was reviewed, and three enhancement patterns were identified. Type I (n=6) was defined as a pattern of hyperenhancement throughout the tumor in early-phase CT, decreasing to hypoenhancement in late-phase CT (high-low), a finding characteristic of HCC. Type II tumors (n=2) appeared as an area of hyperenhancement in early-phase CT, and an area of hyperenhancement in late-phase CT (low-high). Histologically, type II tumors were considered unusual types, and included combined hepatocellular-cholangiocarcinoma and scirrhous HCC. In type III tumors (n=4), high-low, low-high and low-low components were observed. Type III tumors showed multinodular-type morphology and comprised both moderately and poorly differentiated HCC with marked necrosis and fibrosis. Each nodule was divided by fibrous septa. In three of the four type III cases, recurrence occurred within 6 months after resection. Immunohistochemistry of claudin-10 showed positivity in five of the 12 cases, including four type III cases. Results of this study suggest that type III tumors have greater malignant potential than the other tumor types.