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Previously published digital autoradiography of H-3-labeled capecitabine reveals a near-uniform distribution of activity throughout a murine pancreatic model. This is in contrast both to C-14-labeled gemcitabine, and established expectations, as the dense stroma of pancreatic cancer is understood to inhibit drug penetration. Capecitabine is a pro-drug for 5 FU. The positioning of the radiolabel on capecitabine leaves open the possibility that much of the autoradiographic signal is generated by nontoxic compounds. Studies were performed on tumors derived via organoid culture from a murine KPC tumor. As before, we performed autoradiography comparing H-3 capecitabine to the gemcitabine analog F-18-FAC. The metabolism of capecitabine in this model was studied through LC-MS of tumor tissue. The autoradiographs confirmed that the H-3 label from capecitabine was much more uniformly distributed through the tumor than the F-18 from the gemcitabine analog. LC-MS revealed that approximately 75% of the molar mass of capecitabine had been converted into 5 FU or pre-5 FU compounds. The remainder had been converted into nontoxic species. Therapeutically relevant capecitabine metabolites achieve a relatively even distribution in this pancreatic cancer model, in contrast to the gemcitabine analog F-18-FAC. In a human xenograft model, (BxPC3), the H-3 label from capecitabine was also uniformly spread across the tumor autoradiographs. However, at 2 h post-administration the metabolism of capecitabine had proceeded further and the bulk of the agent was in the form of nontoxic species.

These data identify a host pathway by which VPS29 and associated factors control the endosomal environment in a manner that influences susceptibility to viral infection. This pathway could serve as a pharmaceutical target for intervention in zoonotic viral diseases, including those caused by coronaviruses, influenza viruses, and filoviruses, all of which are pandemic threats. Emerging zoonotic viral pathogens threaten global health, and there is an urgent need to discover host and viral determinants influencing infection. We performed a loss-of-function genome-wide CRISPR screen in a human lung cell line using HCoV-OC43, a human betacoronavirus. One candidate gene, VPS29, a component of the retromer complex, was required for infection by HCoV-OC43, SARS-CoV-2, other endemic- and pandemic-threat coronaviruses, as well as ebolavirus. Notably, we observed a heightened requirement for VPS29 by the recently described Omicron variant of SARS-CoV-2 compared to the ancestral variant. However, VPS29 deficiency had no effect on certain other viruses that enter cells via endosomes and had an opposing, enhancing effect on influenza A virus infection. Deficiency in VPS29 or other retromer components caused changes in endosome morphology and acidity and attenuated the activity of endosomal proteases. These changes in endosome properties caused incoming coronavirus, but not influenza virus particles, to become entrapped therein. Overall, these data show how host regulation of endosome characteristics can influence cellular susceptibility to viral infection and identify a host pathway that could serve as a pharmaceutical target for intervention in zoonotic viral diseases. IMPORTANCE These data identify a host pathway by which VPS29 and associated factors control the endosomal environment in a manner that influences susceptibility to viral infection. This pathway could serve as a pharmaceutical target for intervention in zoonotic viral diseases, including those caused by coronaviruses, influenza viruses, and filoviruses, all of which are pandemic threats. Our findings show how host regulation of endosome characteristics can influence viral susceptibility in both a positive and negative manner.

Globally, autosomal recessive IFNAR1 deficiency is a rare inborn error of immunity underlying susceptibility to live attenuated vaccine and wild-type viruses. We report seven children from five unrelated kindreds of western Polynesian ancestry who suffered from severe viral diseases. All the patients are homozygous for the same nonsense IFNAR1 variant (p.GLu386*). This allele encodes a truncated protein that is absent from the cell surface and is loss-of-function. The fibroblasts of the patients do not respond to type I IFNs (IFN-alpha 2, IFN-omega, or IFN-beta). Remarkably, this IFNAR1 variant has a minor allele frequency >1% in Samoa and is also observed in the Cook, Society, Marquesas, and Austral islands, as well as Fiji, whereas it is extremely rare or absent in the other populations tested, including those of the Pacific region. Inherited IFNAR1 deficiency should be considered in individuals of Polynesian ancestry with severe viral illnesses.

Recent studies reported the presence of pre-existing autoantibodies (auto-Abs) neutralizing type I interferons (IFNs) in at least 15% of patients with critical COVID-19 pneumonia. In one study, these auto-Abs were found in almost 20% of deceased patients across all ages. We aimed to assess the prevalence and clinical impact of the auto-Abs to type I IFNs in the Seine-Saint-Denis district, which was one of the most affected areas by COVID-19 in France during the first wave. We tested for the presence of auto-Abs neutralizing type I IFNs in a cohort of patients admitted for critical COVID-19 pneumonia during the first wave in the spring of 2020 in the medicine departments at Robert Ballanger Hospital, Aulnay sous Bois. We found circulating auto-Abs that neutralized 100 pg/mL IFN-alpha 2 and/or IFN-omega in the plasma (diluted 1/10) of 7.9% (11 of 139) of the patients hospitalized for critical COVID-19. The presence of neutralizing auto-Abs was associated with an increased risk of mortality, as these auto-Abs were detected in 21% of patients who died from COVID-19 pneumonia. Deceased patients with and without auto-Abs did not present overt clinical differences. These results confirm both the importance of type I IFN immunity in host defense against SARS-CoV-2 infection and the usefulness of detection of auto-Abs neutralizing type I IFNs in the management of patients.

Background. Acute rejection (AR) and recurrent hepatitis C virus (R-HCV) are significant complications in liver allograft recipients. Noninvasive diagnosis of intragraft pathologies may improve their management. Methods. We performed small RNA sequencing and microRNA (miRNA) microarray profiling of RNA from sera matched to liver allograft biopsies from patients with nonimmune, nonviral (NINV) native liver disease. Absolute levels of informative miRNAs in 91 sera matched to 91 liver allograft biopsies were quantified using customized real-time quantitative PCR (RT-qPCR) assays: 30 biopsy-matched sera from 26 unique NINV patients and 61 biopsy-matched sera from 41 unique R-HCV patients. The association between biopsy diagnosis and miRNA abundance was analyzed by logistic regression and calculating the area under the receiver operating characteristic curve. Results. Nine miRNAs-miR-22, miR-34a, miR-122, miR-148a, miR-192, miR-193b, miR-194, miR-210, and miR-885-5p-were identified by both sRNA-seq and TLDA to be associated with NINV-AR. Logistic regression analysis of absolute levels of miRNAs and goodness-of-fit of predictors identified a linear combination of miR-34a + miR-210 (P < 0.0001) as the best statistical model and miR-122 + miR-210 (P < 0.0001) as the best model that included miR-122. A different linear combination of miR-34a + miR-210 (P < 0.0001) was the best model for discriminating NINV-AR from R-HCV with intragraft inflammation, and miR-34a + miR-122 (P < 0.0001) was the best model for discriminating NINV-AR from R-HCV with intragraft fibrosis. Conclusions. Circulating levels of miRNAs, quantified using customized RT-qPCR assays, may offer a rapid and noninvasive means of diagnosing AR in human liver allografts and for discriminating AR from intragraft inflammation or fibrosis due to R-HCV.

Interferon signaling mediates resistance to immune checkpoint blockade therapy, but the underlying mechanisms are poorly understood. In this issue of Immunity, Cucolo et al. identify RIPK1 as an interferon-stimulated gene with potent effects on cell extrinsic and intrinsic immunotherapy resistance.

Hydrocephalus, characterized by cerebral ventricular dilatation, is routinely attributed to primary defects in cerebrospinal fluid (CSF) homeostasis. This fosters CSF shunting as the leading reason for brain surgery in children despite considerable disease heterogeneity. In this study, by integrating human brain transcriptomics with whole-exome sequencing of 483 patients with congenital hydrocephalus (CH), we found convergence of CH risk genes in embryonic neuroepithelial stem cells. Of all CH risk genes, TRIM71/lin-41 harbors the most de novo mutations and is most specifically expressed in neuroepithelial cells. Mice harboring neuroepithelial cell-specific Trim71 deletion or CH-specific Trim71 mutation exhibit prenatal hydrocephalus. CH mutations disrupt TRIM71 binding to its RNA targets, causing premature neuroepithelial cell differentiation and reduced neurogenesis. Cortical hypoplasia leads to a hypercompliant cortex and secondary ventricular enlargement without primary defects in CSF circulation. These data highlight the importance of precisely regulated neuroepithelial cell fate for normal brain-CSF biomechanics and support a clinically relevant neuroprogenitor-based paradigm of CH. Duy and Weise et al. combined human functional integrative genomics with mouse experimental biology to reveal a neuroprogenitor-based genetic subtype of human hydrocephalus with defective neurogenesis and altered brain-fluid biomechanics.