Knowledge of the human genetic contribution to the risk of complications from influenza is limited. This study assessed the association between human single-nucleotide polymorphisms (SNPs) and disease progression in individuals with influenza. A targeted analysis of 10 SNPs with prior evidence in COVID-19 and a genome-wide association study (GWAS) were used to assess associations between SNPs and disease progression in two multinational cohorts with suspected or laboratory-confirmed influenza: a hospitalized cohort (n = 1634) and a pooled cohort of hospitalized and outpatients (n = 3469). Disease progression was defined as prolonged hospitalization (> 28 days), progression to mechanical ventilation, admittance to intensive care unit, or death (for hospitalized individuals) or progression to hospitalization or death (for outpatients). Disease progression was observed in 9.1% of hospitalized patients and 2.2% of outpatients. Age was a significant risk factor for disease progression, with 20% increased odds of disease progression per 10-year increase in age (OR: 1.20, 95%CI: 1.08-1.33, p < 0.001). Disease progression rates also differed by continent (p < 0.0001). Targeted SNP analyses did not identify significant associations with disease progression; however, the strength of associations was most pronounced in sensitivity analyses for the pooled cohort in individuals < 65 years old. GWAS analyses did not identify significant common SNP associations in either the hospitalized or pooled cohorts, nor in sensitivity analysis of (1) individuals with laboratory-confirmed influenza and (2) those aged < 65 years. In a geographically diverse cohort of individuals with influenza, the genetic links to disease progression only started to become evident in the sensitivity analyses, mainly when looking at younger individuals. The power to detect associations was limited by the rate of disease progression and heterogeneity in phenotypes of the individuals studied, and therefore, additional studies focused on the role of genetics in influenza disease progression are needed.
Breast cancer is the most commonly diagnosed cancer globally. The potential role of oncogenic viruses, particularly Human Betaretrovirus (HBRV, formerly MMTV-LV/HMTV), in the pathogenesis of breast cancer has been a subject of research for decades. However, studies investigating this association have produced conflicting results. This systematic review and meta-analysis aim to assess the prevalence of HBRV in breast cancer cases and evaluate its potential association with breast cancer. A systematic literature search was conducted in MEDLINE, Web of Science, Scopus, and EMBASE following PRISMA guidelines. Studies assessing HBRV prevalence in breast cancer patients and case-control studies investigating its association with breast cancer risk were included. The Newcastle-Ottawa Scale (NOS) was used to evaluate study quality, and meta-analysis was performed using RevMan 5.1. Heterogeneity was assessed using the I² statistic, and subgroup analyses were conducted based on detection methods, sample types, and geographic regions. The literature search identified a total of 45 studies that were deemed suitable for inclusion in the systematic review. 26 studies were used in the subsequent meta-analysis. The initial meta-analysis revealed a significant association between HBRV and breast cancer (OR = 4.92, 95% CI: 4.00-6.04, p < 0.00001) but exhibited high heterogeneity (I² = 82%). After excluding an outlier, heterogeneity was significantly reduced (I² = 22%), with a revised OR of 11.95 (95% CI: 8.78-16.25, p < 0.00001 ). Subgroup analysis demonstrated variation in detection methods, with Nested PCR (OR = 19.15) and Frozen tissue samples (OR = 18.00) showing the strongest associations. Geographic analysis indicated the highest odds in North America (OR = 24.75), followed by Europe (OR = 15.02). This meta-analysis suggests strong epidemiological evidence supporting an association between HBRV infection and human breast cancer, and is consistent with a possible etiological role. However, variability in study methodologies and geographic differences warrant further investigation through standardized, large-scale studies to confirm these findings and explore potential mechanisms of viral oncogenesis in breast cancer.
Transgenic mouse models expressing predefined T-cell receptors (TCRs) have been instrumental in advancing our understanding of T-cell biology. However, these traditional models rely on random genomic insertion of large constructs, require labor-intensive embryo manipulation, and frequently result in aberrant TCR expression and phenotypes. These limitations render traditional models insufficient to meet the mounting demands for rapid and precise model systems to evaluate TCR specificities. In this study, we developed a streamlined method that uses adeno-associated virus (AAV) and CRISPR/Cas9-mediated genome editing to precisely integrate pre-rearranged TCRα/β sequences into the mouse TCRβ (Trb) locus, enabling the rapid generation of TCR knock-in mice with physiological TCR expression and functional T-cell differentiation upon antigenic challenge. This approach bypasses the need for screening multiple founders for faithful TCR expression, enhancing the versatility and utility of monoclonal TCR mice in basic immunology and preclinical research, such as in the fields of cancer immunotherapy and vaccine development.
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Clonally expanded CD4+ T cells harbouring rebound-competent HIV persist lifelong during antiretroviral therapy1-5. Latency is considered the principal barrier to viral eradication and has resisted pharmacological reversal6,7, yet sustained immune pressure appears to erode reservoirs8-15. Recent advances have yielded glimpses into exceptionally rare reservoir-harbouring cells, implicating prosurvival properties in persistence16-18. Here we isolate and characterize authentic reservoir clones (ARCs) that robustly proliferate and accumulate while producing infectious virus, without overtly succumbing to cytopathicity. At any moment, only small fractions of ARCs expressed HIV proteins, a state associated with conserved host transcriptional programs but remarkably refractory to potent T cell stimulation. Nevertheless, sustained co-culture with a CD8+ cytotoxic T lymphocyte clone substantially culled proliferating ARCs, revealing time-integrated vulnerability to immune pressure. The corresponding ex vivo CD8+ T cell response was poorly cytotoxic, and in vivo erosion of ARCs occurred only slowly. A regulatory T cell ARC displayed pronounced cell-intrinsic resistance to cytotoxic T cells - a longstanding hypothesis now directly demonstrated - linked to low oxidative stress and reversed with deferoxamine19, a hypoxic stress inducer and FDA-approved therapeutic. Overall, we provide insights into the vulnerabilities of reservoir clones to potent, sustained cytotoxic T cell pressure and highlight intrinsic resistance pathways as actionable therapeutic targets, opening opportunities for advancing immune-based HIV cure strategies.
Pathologic tumor response and changes in the tumor microenvironment (TME) predict outcomes to immune checkpoint inhibitors, but are understudied in rare tumors. We investigated whether artificial intelligence (AI)-powered analyses of pretreatment and on-treatment biopsies may inform treatment outcomes to pembrolizumab. We evaluated 256 baseline and 248 on-treatment biopsies from 84 patients with rare tumors (10 cohorts) in a phase II pembrolizumab trial. Intratumoral tumor-infiltrating lymphocyte (iTIL) density and tumor content (TC) were assessed on H&E-stained slides using a deep learning-based analyzer (Lunit SCOPE IO). Baseline iTIL and changes in iTIL and TC were correlated with progression-free survival (PFS) and overall survival (OS). Multiplex immunofluorescence was performed in 27 paired samples to assess TME changes. In the high-iTIL tumor group, a baseline iTIL of ≥60 cells/mm2 was associated with favorable PFS (HR 0.49, 95% CI 0.25 to 0.99, p=0.046) and higher CD8+ and CD8+PD-1+ and lower FoxP3+CD8+PD-1+ T-cell density. However, this association with PFS was not seen in the overall cohort (HR 0.62, 95% CI 0.37 to 1.06, p=0.082). In paired biopsies, on-treatment increase in iTIL showed a trend toward improved PFS (HR 0.64, 95% CI 0.40 to 1.06, p=0.084) and was significantly associated with improved OS (HR 0.55, 95% CI 0.35 to 1.01, p=0.037). This increase was also associated with reduced spatial distance between CD8+ immune and tumor cells. Decreased TC during treatment was significantly associated with prolonged PFS and OS (PFS: HR 0.51, p=0.019; OS: HR 0.54, p=0.042). The combination of increased iTIL and decreased TC was significantly associated with better PFS (HR 0.36, p=0.009) and OS (HR 0.36, p=0.029). AI-powered assessment of the TME before and during treatment may help inform treatment outcomes to pembrolizumab in patients with rare tumors. NCT02721732.
Although HPV infection is obligatory for almost all cases of cervical cancer (CC), other risk factors can promote the progression of cervical cancer. In this context, the expression of human endogenous retroviruses (HERVs) in the development of CC has been investigated. In this study, the expression status of HERV-E env transcripts was analyzed in 111 cervical biopsies, including 35 cervical cancer samples, 20 precancerous lesions, and 56 normal samples. Real-time PCR with specific primers was used to quantify the relative expression of HERV-E env, HPV 16 and 18 E6/E7 genes, and GAPDH as a normalization control. Our results indicated an increase in the expression of HERV-E env, and the difference was statistically significant in the cancer group compared to the precancerous group (1.5-fold change) (P = 0.031). In HPV 16 or 18-infected patients, a higher mean value of HERV-E env mRNA was also found in the cancer group than in the precancerous group. ROC curve analysis showed a significant difference in env expression between precancerous and cancerous lesions in all patients analyzed (P = 0.015) and in a group of patients infected with HPV 16 or 18 genotypes (P = 0.023). In addition, there was a positive correlation between the higher expression of HERV-E env mRNA with E7 (R = 0.34, P = 0.016) and age (R = 0.35, p = 0.016) in HPV 16-infected patients. In conclusion, our study found a possible association between HERV-E env expression and cervical cancer, as HERV-E is actively transcribed during the progression of cervical lesions. Future studies on the potential interaction of HERV-E env with HPV 16 E7 oncoprotein are likely to elucidate common signaling pathways in the progression of cervical cancer and other HPV-related malignancies.
The SARS-CoV-2 main protease (Mpro) remains a prime antiviral target because its inhibition halts viral replication. To probe how subtle atomic changes influence drug performance, we carried out a systematic halogen scan on a potent ketoamide scaffold, replacing a single fluorine with chlorine, bromine, or iodine. Enzymatic assays revealed that the F- and Cl-substituted analogues inhibit Mpro at nanomolar levels, whereas Br and I variants are 10- to 20-fold weaker. Cell-based antiviral tests mirrored this trend, yet uptake studies showed the opposite: iodine markedly enhances intracellular accumulation. High-resolution X-ray structures (1.6-1.8 Å) explain the dichotomy: small halogens fit snugly in the S1' σ-hole pocket, maximizing hydrogen-bond geometry, while bulkier atoms distort binding but create a lipophilic patch that boosts permeability. These data yield the first fluorine-to-iodine structure-activity map for SARS-CoV-2 Mpro inhibitors. These findings highlight the critical role of halogen selection in antiviral inhibitor design.
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Joubert syndrome (JS) is a rare genetic disorder characterized by developmental abnormalities, particularly in the brainstem and cerebellar vermis, alongside multisystem manifestations such as kidney and liver anomalies, polydactyly, cleft lip or palate, and tongue defects. The underlying ciliopathy causing JS may also contribute to gastrointestinal symptoms and immune dysregulation via Wnt signaling and impaired epithelial maintenance. Dysmotility including Hirschsprung disease has been documented at increased rates in JS and in other ciliopathies. Our case series highlights how JS patients frequently exhibit feeding intolerance, vomiting, and poor growth, which may raise suspicion for an underlying gastrointestinal condition, such as eosinophilic esophagitis (EoE). Gastrointestinal symptoms often overlap with other chronic issues, delaying diagnosis and treatment, which can affect long-term outcomes. These cases underscore the importance of thorough evaluations, including endoscopy, to investigate persistent symptoms suggestive of eosinophilic gastrointestinal diseases (EGIDs)/EoE. Such vigilance promotes early targeted therapies, improves quality of life, and decreases the risk of complications including formation of esophageal strictures.
Walter Reed Army Institute of Research-Armed Forces Research Institute of Medical Sciences provides medical readiness training in the Indo-Pacific. While executing Thailand rotations, it is developing similar training opportunities in the Philippines. Increased funding and personnel support ensure that this program supports the force health protection readiness of military health professionals.
Orthohantaviruses (hantaviruses) are emerging rodent-borne pathogens that can cause severe human disease. They are present on multiple continents and are responsible for thousands of human cases per year. Despite this, no licenced therapeutics are available, vaccines for most strains are lacking, and the immunological response to infection is poorly characterised. This study aimed to analyse the humoral immune response to Puumala virus (PUUV) infection to inform future studies focussing on the production of therapeutic monoclonal antibodies and vaccination strategies. Serum was obtained from a cohort of 24 patients hospitalised with PUUV infection at four time points, covering the early acute, late acute, early convalescent, and late convalescent stages of the disease. The humoral immune responses at each time point were quantified, and cross-binding, cross-neutralising antibody responses were investigated. Serum cytokine levels were also interrogated, and expression was correlated with humoral outputs. PUUV infection elicited a robust anti-PUUV neutralising antibody response. However, cross-reactive antibodies that were capable of binding diverse hantaviruses were also induced in late convalescence. Modulations in the abundance of IgG subclasses were evident following infection, with significant differences present months after infection. This study demonstrates that broadly reactive anti-hantavirus antibodies are produced in response to Old-World hantavirus infection, but predominantly months after recovery. As this is concomitant with changes in IgG subtypes, our results suggest that PUUV infection promotes prolonged class-switching and somatic hypermutation, favouring conserved epitopes long after exposure. Work at Mount Sinai was supported by Institutional Funds, work at the Medical University of Vienna was supported by Institutional Funds. The study was supported in part by the Styrian government, Austria (project no. ABT12-106729/2022-13) and the Austrian Science Fund (FWF) (number J 4737-B).
A central problem in achieving vaccine-based protection against viral infections is eliciting antibodies that are resilient to viral variation. Successive waves of SARS-CoV-2 infection during the COVID19 pandemic were driven by variants that acquired resistance to neutralizing antibodies elicited by prior SARS-CoV-2 variants. To the extent that serum neutralization breadth occurs in individuals with multiple exposures to SARS-CoV-2 antigens, we and others find that it is largely comprised of antibodies that target the variable receptor binding domain (RBD), rather than more conserved spike protein domains. By designing synthetic dimeric RBD immunogens we show that limiting divergence in heterodimeric components favors the generation of cross-reactive B cells and antibodies. We thus devised a vaccine approach based on a two-dose immunization with a pool of five overlapping heterodimeric synthetic RBD variants. Collectively, the RBD heterodimer pool was designed to cover 10% sequence variation and elicited greater antibody cross-reactivity and neutralization breadth than homodimers or heterodimers with highly divergent components. Using an unconventional 'prospective' challenge model in mice, we demonstrate the effectiveness of the RBD heterodimer pool in inducing antibody responses that attenuate infection by future SARS-CoV-2 variants, as well as protection in a challenge model based on a chimeric vesicular stomatitis virus bearing a spike protein from SARS-CoV-1.
Nanoparticle immunogens excel at rapidly inducing high levels of circulating antibodies and are being deployed as part of several novel vaccines. However, their ability to elicit memory B cell responses is less well understood. Here, we compared serologic and memory B cell responses after prime boost vaccination with either SARS-CoV-2 Wuhan-Hu-1 mRNA vaccine, or protein nanoparticles: SARS-CoV-2 B.1.351 homotypic containing a single receptor binding domain (RBD); (homotypic beta) or a combination of different Sarbecovirus RBDs (mosaic 8b), respectively. The memory B cells elicited by the 3 vaccine regimens showed closely related antibody sequences, similar levels of somatic mutation, and clonal diversity. The breadth of serologic responses elicited by the mosaic nanoparticles was comparable to the homotypic nanoparticle and superior to the mRNA vaccine for some mismatched strains. However, serum neutralizing titers to SARS-CoV-2 were highest after mRNA vaccination. The three vaccines elicited memory B cells that produced antibodies specific to a broad range of epitopes on the RBD that differed in a way that may reflect epitope masking. Monoclonal antibodies derived from memory B cells elicited by the mosaic 8b nanoparticle showed greater breadth against a panel of SARS-CoV-2 variants and SARS-CoV.
Twice-daily dosing of Dolutegravir is approved for adults with HIV and integrase strand transfer inhibitor resistance, but not for children. Population pharmacokinetic modeling and simulations identified a weight-tiered twice-daily dose for children, predicted to yield dolutegravir exposures within the therapeutic window and provide similar efficacy and safety as seen in adults with HIV-1 and first-generation integrase strand transfer inhibitor resistance.
Aicardi-Goutières syndrome (AGS) is a type I interferonopathy presently associated with nine genes. PTPN1 is a negative regulator of the interferon pathway previously associated with chronic inflammation and recently type 1 IFN autoinflammation. Genomic data from undiagnosed individuals with suspected AGS were interrogated for PTPN1 variants, and predicted loss-of-function (pLOF) and damaging missense variants in PTPN1 were sought in two additional academic databases as well as the All of Us database. We identified 13 cases with ultra-rare heterozygous pLOF or highly damaging missense variants in PTPN1. Nine cases were identified in a cohort of 53 individuals (~ 17%) with clinical, imaging and persistent biochemical features of AGS. Median age of onset is 1.75 years (IQR 0.67), significantly later (p< 0.0001) than other AGS genotypes. Four additional cases were identified in academic datasets with variable clinical features suggestive of autoinflammation. Additionally, 49 individuals with ultra-rare, damaging PTPN1 variants were identified in the All of Us database, none had features suggestive of AGS, but autoimmunity was highly prevalent (~21.6%). Our data implicate PTPN1 as a cause of later-onset presentations of AGS within a broader spectrum of autoinflammatory phenotypes. Segregation and biobank data demonstrate reduced penetrance, with carriers being enriched for autoimmune disorders.
Macrophages differentiated with macrophage colony-stimulating factor (M-CSF) (M-Mac) are widely used as an experimental model. Interleukin 27 (IL-27)-polarized M-Mac (27M-Mac) suppresses HIV replication; however, the effects of IL-27 polarization on granulocyte-macrophage colony-stimulating factor (GM-CSF)-induced macrophages (GM-Mac) remain less investigation. Here, we compare multiple functional properties and gene expression profiles of 27M-Mac and IL-27-polarized GM-Mac (27GM-Mac). M-Mac and GM-Mac were generated from monocytes of healthy donors and subsequently treated with IL-27 for three days. HIV replication in 27M-Mac, GM-Mac, and 27GM-Mac was suppressed to nearly 10% of that in M-Mac; however, single-cell RNA sequencing showed that M-Mac clustered with GM-Mac, and 27M-Mac clustered with 27GM-Mac. Expression of CD38 and secretion of CXCL9 and C1q were significantly increased in 27M-Mac and 27GM-Mac compared with M-Mac and GM-Mac. Although CD16 and CD64 expression increased in 27M-Mac and 27GM-Mac relative to their respective controls, phagocytic activity in 27M-Mac and 27GM-Mac was 30% of that in M-Mac. Autophagy was promoted 3.7-fold more strongly in 27M-Mac than in M-Mac, reaching levels comparable to those in GM-Mac and 27GM-Mac. Collectively, these findings indicate that IL-27 polarizes M-Mac and GM-Mac toward transcriptionally and functionally similar subtypes, providing insight into the role of IL-27 in macrophage polarization and plasticity.
Antiretroviral inhibitors that target specific aspects of human immunodeficiency virus 1 (HIV-1) replication have made a huge impact on the management of the global AIDS pandemic and the health and well-being of people that live with HIV. Current maintenance therapies commonly consist of one or two nucleoside reverse transcriptase inhibitors co-formulated with a second-generation integrase strand transfer inhibitor (INSTI), and regimens containing the second-generation INSTI dolutegravir are in wide-use across the globe. INSTIs are active site inhibitors that block HIV-1 DNA integration- more precisely, the ability of the integrase enzyme to join the 3' ends of the viral reverse transcript with host chromosomal DNA. High-level resistance to first-generation INSTIs readily occurs via amino acid substitutions proximal to the integrase active site. Second-generation INSTIs, by contrast, impart comparatively high genetic barriers to the generation of drug resistance. When observed, such changes within integrase tend to confer comparatively low level resistance to both first- and second-generation INSTIs. Moreover, there are several reports of clinical failure to dolutegravir in the absence of known integrase drug resistance mutations. Laboratory-based research has at the same time identified three non-integrase pathways that can lead to dolutegravir resistance, including changes within the viral surface and transmembrane envelope glycoproteins, the nucleocapsid protein, and the genomic RNA 3' polypurine tract that serves to prime plus-strand DNA synthesis during reverse transcription. Each type of change has been shown to confer resistance to dolutegravir through a unique molecular mechanism. Herein, I review the non-integrase changes that are known to occur and the proposed mechanisms that lead to the generation of dolutegravir resistance. The potential for these different types of changes to impact INSTI drug resistance in the clinic is also discussed. The unexpected observation that multiple non-integrase pathways can contribute to the generation of dolutegravir resistance highlights the remarkable plasticity of HIV-1 to circumvent challenge with a highly efficacious small molecule inhibitor. Given its current global use as a frontline anti-HIV inhibitor, this research informs regions of sequence surveillance for the continued safe and efficacious use of dolutegravir-based antiretroviral therapies.
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