The retromer mediates protein trafficking through recycling cargo from endosomes to the trans-Golgi network in eukaryotes. However, the role of such trafficking events during pathogen-host interaction remains unclear. Here, we report that the cargo-recognition complex (MoVps35, MoVps26 and MoVps29) of the retromer is essential for appressorium-mediated host penetration by Magnaporthe oryzae, the causal pathogen of the blast disease in rice. Loss of retromer function blocked glycogen distribution and turnover of lipid bodies, delayed nuclear degeneration and reduced turgor during appressorial development. Cytological observation revealed dynamic MoVps35-GFP foci co-localized with autophagy-related protein RFP-MoAtg8 at the periphery of autolysosomes. Furthermore, RFP-MoAtg8 interacted with MoVps35-GFP in vivo, RFP-MoAtg8 was mislocalized to the vacuole and failed to recycle from the autolysosome in the absence of the retromer function, leading to impaired biogenesis of autophagosomes. We therefore conclude that retromer is essential for autophagy-dependent plant infection by the rice blast fungus.

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The epidermal growth factor receptor (EGFR) and the insulin-like growth factor-1 receptor (IGF-1R) play critical roles in tumor growth, providing a strong rationale for the combined inhibition of IGF-1R and EGFR signaling in cancer therapy. We describe the design, affinity maturation, in vitro and in vivo characterization of the bispecific anti-IGF-1R/EGFR antibody XGFR*. XGFR* is based on the bispecific IgG antibody XGFR, which enabled heterodimerization of an IGF-1R binding scFab heavy chain with an EGFR-binding light and heavy chain by the “knobs-into-holes” technology. XGFR* is optimized for monovalent binding of human EGFR and IGF-1R with increased binding affinity for IGF-1R due to affinity maturation and highly improved protein stability to oxidative and thermal stress. It bears an afucosylated Fc-portion for optimal induction of antibody-dependent cell-mediated cytotoxicity (ADCC). Stable Chinese hamster ovary cell clones with production yields of 2–3 g/L were generated, allowing for large scale production of the bispecific antibody. XGFR* potently inhibits EGFR- and IGF-1R-dependent receptor phosphorylation, reduces tumor cell proliferation in cells with heterogeneous levels of IGF-1R and EGFR receptor expression and induces strong ADCC in vitro. A comparison of pancreatic and colorectal cancer lines demonstrated superior responsiveness to XGFR*-mediated signaling and tumor growth inhibition in pancreatic cancers that frequently show a high degree of IGF-1R/EGFR co-expression. XGFR* showed potent anti-tumoral efficacy in the orthotopic MiaPaCa-2 pancreatic xenograft model, resulting in nearly complete tumor growth inhibition with significant number of tumor remissions. In summary, the bispecific anti-IGF-1R/EGFR antibody XGFR* combines potent signaling and tumor growth inhibition with enhanced ADCC induction and represents a clinical development candidate for the treatment of pancreatic cancer.

ASGPR is a candidate receptor for HBV that mediates further steps of HBV entry.

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Background and AimsNon-alcoholic fatty liver disease (NAFLD) is a major chronic liver disease worldwide, and non-alcoholic steatohepatitis (NASH) is one of its pathological subtypes. The pathogenesis of NASH has not yet been fully elucidated. The purpose of this study was to identify the hub genes and pathways involved in NASH using bioinformatics methods. The hub genes were confirmed in human and animal models.Materials and MethodsThree Gene Expression Omnibus (GEO) datasets (GSE48452, GSE58979, and GSE151158) of NASH patients and healthy controls were included in the study. We used GEO2R to identify differentially expressed genes (DEGs) between NASH patients and healthy controls. Functional enrichment analyses were then performed to explore the potential functions and pathways of the DEGs. In all DEGs, only two genes were highly expressed in NASH patients throughout the three datasets; these two genes, SPP1 and CXCL9, were further studied. Serum and liver tissues from NASH patients and healthy controls were collected. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were measured in NASH patients and healthy controls. Liver tissues were stained with hematoxylin and eosin. Immunohistochemical staining was used to evaluate the expression levels of the two genes in liver tissues. Male C57BL/6J mice were fed a methionine choline-deficient (MCD) diet for 8 weeks, after which serum ALT and AST levels were measured and liver tissues were stained.ResultsSPP1 and CXCL9 were the hub genes detected in the three datasets. “Lipid metabolism,” “inflammatory response,” and “lymphocyte activation” were the most significant biological functions in GSE48452, GSE58979, and GSE151158, respectively. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the toll-like receptor signaling pathway was significantly enriched in NASH patients. Serum ALT and AST levels were significantly increased in NASH patients compared to healthy controls. Liver tissues had more serious steatosis, hepatocyte ballooning degeneration, and lobular inflammatory infiltration, and the expression of SPP1 and CXCL9 in liver cells was significantly upregulated in NASH patients compared to healthy controls. MCD diet mice were consistent with NASH patients.ConclusionSPP1 and CXCL9 may play important roles in NASH pathogenesis and could be potential therapeutic targets and biomarkers of NASH in the future. Further experimental studies are needed to confirm our results.