The membrane-proximal external region (MPER) of HIV-1 Env represents a critical target for broadly neutralizing antibodies (bnAbs) due to its conservation and functional importance. However, MPER-targeting bnAbs recognize composite epitopes comprising peptide and viral membrane lipid components, creating an inherent tension between viral neutralization efficacy and polyreactivity. 10E8-class antibodies exhibit high neutralization potency with low polyreactivity, whereas 4E10-class antibodies show comparably broad neutralization but higher polyreactivity, underscoring the need to understand the structural basis of this distinction. We therefore determined crystal structures of DH511.1 (memory B cell-derived), DH511.12P (plasma cell-derived), and VRC42.01 in complex with MPER peptide and phosphatidic acid, along with a cryo-EM reconstruction of DH511.2 bound to membrane-embedded Env. Through integrative analysis taking into account previously determined structures of other MPER bnAbs, we reveal two distinct lipid recognition strategies. Groove-mediated binders, including 10E8 and DH511, engage lipids through antibody-membrane interface grooves with distinct geometries and angular approaches to the membrane. In contrast, heavy chain-mediated binders, including 4E10, PGZL1, and VRC42, utilize positively charged CDR H1 patches for direct lipid headgroup recognition. Importantly, DH511 lineage members exhibited differential cardiolipin polyreactivity linked to their maturation stage. PGZL1 and VRC42.01 employ weaker positive patches at lipid-binding sites than 4E10, and PGZL1 additionally introduces a CDR H3-mediated negative patch that creates electrostatic repulsion with negatively charged lipid headgroups, thereby limiting nonspecific interactions. These findings provide a structural framework for understanding how MPER bnAbs balance lipid binding with specificity and inform immunogen design for inducing safe and effective neutralizing responses.
Link prediction is important across biological, social, and technological networks, but many methods either require domain-specific node attributes, do not scale to large graphs, or miss higher-order topology. We present BLANT-Predict, a topology-only framework that uses sampled graphlets and orbit-pair frequencies to rank likely missing edges. Across 12 real-world networks (up to about 1 million nodes and 3 million edges), we compare against 13 baseline methods and observe higher precision with strong scalability. Beyond standard k-fold cross-validation, we evaluate predictions against future out-of-sample network snapshots to better reflect real deployment. BLANT-Predict maintains superior precision in these future tests, indicating practical value for predicting previously unobserved links.
Stroke remains a leading cause of mortality worldwide, demanding rapid, accurate diagnosis; distinguishing stroke types matters because treatments differ significantly. Using a recently developed, non-invasive brain scanner integrating a 16-antenna radio-frequency array, we present a deep-learning model that distinguishes the two main stroke types. We employ masked autoencoder-based self-supervised learning and supervised contrastive strategies to improve data efficiency and robustness with limited labeled clinical data. In the current test cohort, the system achieved 92% sensitivity and 85% specificity for hemorrhagic versus non-hemorrhagic detection, and 95% sensitivity and 80% specificity for ischemic versus non-ischemic cases. Beyond classification, the model showed patterns consistent with an ordering of relative dielectric permittivity across conditions (hemorrhagic, ischemic, mimic, and healthy). While not representing direct measurement of tissue dielectric properties, these observations provide insight into relative dielectric differences captured by the RF measurements. These findings support the potential of RF-based measurement-domain analysis to more reliable differentiation between stroke subtypes.
Isothermal nucleic acid amplification offers advantages over qPCR for decentralized diagnostics but remains constrained by rigid primer design, detection complexity, and equipment dependence. Recombinase-based systems operate at lower temperatures than other isothermal methods but require long primers and auxiliary enzymatic processing for detection. Here we introduce Annexing Isothermal Nucleotide Amplification (ANINA), a probe-guided recombinase-based framework integrating amplification and detection at ambient temperatures within a single lyophilized reaction. An Annexing Probe recruits short primers through spatial annexation, enabling efficient amplification at 25 °C within 30 minutes and direct lateral flow detection without auxiliary enzymatic processing. We show this framework detects attomolar WSSV and EBV DNA in abundant host gDNA, supports target-specific detection of DNA and RNA viruses and bacteria in contrived matrices, enables quantitative real-time detection comparable to qPCR, and facilitates a fully equipment-free 45-minute sample-to-answer workflow that detects early viral infection in a natural host model with qPCR-level sensitivity and improved performance over a commercial antigen test.
Streptococci are prevalent in animal and human microbiomes. These organisms produce a vast array of small peptides that modulate complex functions within the cell such as quorum sensing, virulence, and metabolism. Transcriptional regulators are central to this process, of which Rgg transcriptional regulators hold prominence in streptococci. These systems are controlled by peptides known as SHPs (short hydrophobic peptides) and LCPs (leaderless communication peptides). Also known as Rgg/SHP quorum sensing (QS) systems, they are ubiquitous across streptococcal species and regulate cellular competence, metabolic programs, virulence, and facilitate colonization of host species. It has been recently demonstrated that Rgg/SHP QS systems can also regulate the production of natural products known as RaS-RiPPs ( Radical S -adenosylmethionine enzyme Ribosomally translated and Post-translationally modified Peptides). RaS-RiPPs are widespread in streptococci with sixteen current subfamilies. Some of these natural products possess inhibitory properties while others' functions are currently unknown. We provide here a review of Rgg/SHP systems within streptococci, the complexities and characterized functions of RaS-RiPPs, as well as the connection between Rgg/SHP and RaS-RiPPs. We also provide a brief overview of competence in streptococci, given the relevance of these systems to peptide signaling in this genus.
The user-operated audiometry (UAud) project aims at introducing an automated system for user-operated audiometric testing into everyday clinical practice. Here, we focus on the Audible Contrast Threshold (ACT) test, which has been proposed as a language-independent alternative to aided speech-in-noise tests. Five distinct user-operated ACT (U-ACT) test candidates were evaluated in terms of performance, reliability, and usability against two established test benchmarks. The study involved 28 participants with diverse hearing and cognitive abilities. The results demonstrated that the primary factor influencing the reliability of test results was the type of task. In terms of usability, the participants reported positive experience with all test candidates, with the Yes/No task resulting in the lowest perceived difficulty and the best understanding. Overall, a test candidate with a custom adaptive procedure inspired by audiologists' experiences with ACT was chosen for the UAud protocol as a user-operated test of hearing-in-noise perception.
Mutations in KRAS are a dominant driver of pancreatic ductal adenocarcinoma (PDAC), with about 50% of patients presenting with KRASG12D mutations. Small molecule inhibitors targeting KRASG12D suppress PDAC; however, the contribution of the tumor microenvironment (TME) to the sustained efficacy of KRASG12D inhibition and mechanisms of resistance to KRASG12D suppression remain to be elucidated. Here, integrated spatial transcriptomics, single-cell RNA sequencing, and CODEX-based spatial proteomics analyses of PDAC mouse models uncover that while KRASG12D inhibition by MRTX1133 initially increases CD11c+ cells and T cell infiltration proximal to cancer cells, long-term treatment results in reversal of the immune responses leading to resistance promoted by multiprotein mediator complex associated kinase CDK8. CDK8 imparts this resistance via induction of CXCL2 chemokine secretion, inhibition of FAS expression, and remodeling of the TME to promote immune evasion. Targeting CDK8 by itself or in combination with αCTLA-4 immunotherapy overcomes resistance to KRASG12D inhibition. We also provide evidence of CDK8 upregulation in PDX tumors resistant to inhibitors selective for RAS(ON) and RASG12D(ON): daraxonrasib and zoldonrasib, respectively, highlighting a common KRAS vulnerability node for TME resistance.
Adeno-associated virus (AAV) vector-based liver gene therapy for inherited diseases has demonstrated efficacy in clinical trials in adults. However, its application to pediatric patients is limited by loss of AAV genomes during hepatocyte proliferation, compromising long-term benefits. Additionally, the anti-AAV immune responses induced after the initial AAV-administration preclude vector re-dosing. One key driver of this immune response is mTOR-dependent activation of dendritic cells. Inhibition of this pathway with rapamycin can promote immune tolerance. We assessed the safety and efficacy of rapamycin-loaded synthetic nanoparticles (ImmTOR) in juvenile OTCSpf-Ash mice, a model of ornithine transcarbamylase (OTC) deficiency (OTCD). Treatment of postnatal day 30 mice with ImmTOR alone transiently activated autophagy and reduced urinary orotic acid levels. Treatment with a liver-specific AAV8 vector encoding codon-optimized human OTC cDNA normalized urinary orotic acid levels and significantly increased ureagenesis, OTC protein expression, and enzyme activity compared with untreated controls. AAV-vector co-administration with ImmTOR prevented IgM and IgG formation and induced a dose-dependent reduction of anti-AAV neutralizing antibodies, consistent with modulation of the humoral immune response. These findings suggest that ImmTOR can mitigate humoral immune responses to AAV vectors in OTCSpf-Ash mice and may enable AAV vector re-administration, although further optimization is required for clinical translation.
REM sleep behavior disorder (RBD) is a robust prodromal marker of α-synucleinopathies: idiopathic RBD carries a 10-15-year phenoconversion risk of 80-90% to Parkinson's disease (PD) and related disorders. In major depressive disorder (MDD), comorbid RBD marks a subgroup at elevated prodromal PD risk, yet is frequently missed in psychiatric practice. Here, we developed a multimodal AI framework to detect comorbid RBD in MDD. From a clinical cohort of 329 patients, we obtained 261 video clips in 31 patients during reading and spontaneous-speech tasks, including 19 patients with MDD-RBD and 12 demographically and medication-matched MDD-only controls that, to our knowledge, formed the largest cohort of its kind worldwide. We used a dual-stream multimodal model that learned facial dynamics from video and vocal features from speech, and then combined both signals to predict comorbid RBD. In 5-fold cross-validation, our best model achieved 80.5% accuracy and 0.848 F1-score. Explainability analysis highlighted lower-face tension and variability, together with brow lowering, as candidate biomarkers requiring further validation. Predicted risk correlated with RBDQ score (r = 0.53, p = 0.005) and weakly with UPDRS motor score (r = 0.34, p = 0.067). Out-of-distribution evaluation showed broadly similar patterns, supporting the promise of multimodal AI for predicting RBD in MDD and identifying interpretable potential digital markers of prodromal synucleinopathy.
The complement system is essential for distinguishing self-tissue from foreign threats; however, complement activation on biomedical surfaces such as implants, transplants, or drug-delivery systems may lead to severe thromboinflammatory complications. Unlike systemic complement inhibitors, surface-targeted strategies remain scarce, despite their potential advantages regarding safety and efficacy. Recent years have seen the emergence of diverse surface-targeting approaches that impair different mechanisms underlying complement-mediated complications. These strategies, ranging from surface coatings to inhibitors targeting complement-tagged surfaces, are progressing from conceptual development to clinical application. In this review, we provide a comprehensive overview of complement-activation mechanisms on biomedical surfaces, highlight ongoing clinical investigations, and discuss a broad spectrum of emerging approaches to prevent adverse complement activation on biomedical interfaces.
α-Synucleinopathies are devastating neurodegenerative diseases characterized by pathological accumulation of a neuronal protein, α-synuclein (αSyn). Lowering soluble αSyn levels is a promising therapeutic strategy to limit aggregation and neurotoxicity, but directly targeting this protein is hindered by its intrinsically disordered structure and other factors, such as its conformational heterogeneity and intracellular drug delivery barriers. Consequently, increasing attention has been directed toward targeting the SNCA transcript, which encodes αSyn. Here, we developed phosphorodiamidate morpholino oligonucleotide (PMO)-based RNA-degrading chimeras (RDCs) that selectively bind the 5' untranslated region of SNCA messenger RNA (mRNA) and recruit RNase L for targeted RNA degradation. Through the systematic evaluation of nine RDCs, we identified and optimized 4-D1, which effectively reduced SNCA mRNA and αSyn protein expression in HEK293T cells in an RNase L-dependent manner. 4-D1 lowered SNCA transcript and αSyn protein levels in both primary cortical neurons from humanized SNCA mice and in human induced pluripotent stem cell-derived cortical neurons. This reduction prevented prion-like seeding induced by patient-derived αSyn fibrils and protected neurons from fibril-induced cytotoxicity. Finally, in vivo studies confirmed the efficacy of 4-D1 in reducing αSyn mRNA expression in humanized SNCA mice. These findings indicate that PMO-based RDCs may represent a promising therapeutic modality for α-synucleinopathies.
Tumor-derived extracellular vesicles and particles (EVPs) represent a promising analyte class for early cancer detection. However, discrimination of tumor-derived EVPs from those shed by healthy tissues represents a major analytical challenge. This work describes the design and characterization of a novel proximity ligation-based immunoassay which targets tumor-derived EVPs in plasma displaying four colocalized surface biomarkers. We demonstrate the functionality of this approach in cancer cell line-derived EVPs and apply the method to lung adenocarcinoma (LUAD) detection. Importantly, we find that requiring four colocalized cancer-associated biomarkers reduces interference from healthy EVPs versus a three-biomarker design, resulting in strong discrimination performance for several biomarker combinations. Using this approach, we developed a prototype assay for LUAD detection which was evaluated in a case-control study composed of 92 LUAD cases and 290 non-cancer controls. The assay and trained classifier exhibited 48.5% (33/68; 95% confidence interval (CI) 37.1-60.2%) stage I sensitivity, 83.3% (15/18; 95% CI 59.8-94.8%) stage II sensitivity, and 100% (6/6; 95% CI 55.2-100%) stage III/IV sensitivity at 90% specificity. Assay signal was significantly correlated with tumor size and uncorrelated with smoking history. These preliminary results demonstrate the technical feasibility of this platform for early-stage lung cancer detection.
Compared with chimeric antigen receptor (CAR) T cell therapy, antibody-drug conjugates (ADCs) offer distinct advantages. Here, we report on two FGFR4-targeted ADCs, using the high-affinity monoclonal antibody 3A11, the same binder used in a CAR T cell format that is being evaluated at the NCI (NCT06865664) for patients with relapsed/refractory rhabdomyosarcoma (RMS). These ADCs, conjugated to monomethyl auristatin E (MMAE) or an exatecan derivative, are rapidly internalized, causing potent fibroblast growth factor receptor 4 (FGFR4)-dependent cytotoxicity in vitro. In subcutaneous RMS xenograft models, both ADCs demonstrated robust anti-tumor activities, significantly prolonging survival. Notably, exatecan-ADC showed better efficacy, with durable tumor control, in aggressive fusion-negative (FN) RMS559 and fusion-positive (FP) RH4 RMS cell-line-derived xenografts (CDXs) and a patient-derived RMS xenograft (PDX). Furthermore, exatecan-ADC effectively controls tumors in an FGFR4-expressing MDA-MB-453 breast cancer mouse model, eradicating relapsed tumors with retreatment. These findings highlight FGFR4-targeted ADCs as potent therapeutic agents against aggressive FGFR4-expressing malignancies, supporting their further clinical development.
Even with new drugs available, how best to treat unfit adults with acute myeloid leukemia (AML) remains uncertain. In a previous trial in such patients, we found high-dose cytarabine-based therapy with CLAG-M yielded higher response rates but no more toxicity than lower-intensity therapy with dose-attenuated CLAG-M. Here, we conducted a single-institution phase 2 trial (NCT04195945) randomizing 60 adults with untreated AML and medical unfitness with Treatment-Related Mortality (TRM) score of ≥13.1 (68% with ECOG performance status 3-4) 1:1 to standard-dose CPX-351 or CLAG-M. Primary endpoint was 3-month overall survival (OS); key secondary endpoints included overall response rate, rate of measurable residual disease (MRD) negativity, toxicity/mortality rates, and survival estimates. Only CLAG-M met the primary endpoint of ≥63% 3-month OS (70% vs. 60%; P = 0.41), and CLAG-M therapy was associated with a non-significantly higher complete remission (CR) plus CR with incomplete hematologic recovery rate (73% vs. 47%, P = 0.064). Nonetheless, there was no statistically significant difference in relapse-free survival following CLAG-M vs. CPX-351 (median 37.6 vs. 19.9 months; P = 0.80) or OS (median 10.5 vs. 5.8 months; P = 0.76). In patients with proliferative disease, however, OS following CLAG-M was longer (median 18.5 vs. 3.9 months; P = 0.02) suggesting a role for intensive therapy in this patient subset.
After myocardial infarction (MI), quantifying platelet FcɣRIIa (pFCG) stratifies ischemic risk. This study aimed to assess a prognostic biomarker, pFCG, in patients with and without subsequent ischemic and bleeding events. Patients (n = 765) with type 1 MI were enrolled in a prospective noninterventional trial. Inclusion criteria included at least 2 of the following: age ≥65 years, multivessel coronary artery disease, prior MI, chronic kidney disease, and diabetes. Flow cytometry quantified pFCG at a core laboratory. Ischemic endpoints (n = 158) were MI, stroke, and death. Bleeding endpoints (n = 74) were Bleeding Academic Research Consortium 2, 3, and 5. Patients were categorized as no events (n = 562), bleeding events alone (n = 45), single ischemic event (n = 101), bleeding + ischemic events (n = 29), multiple ischemic events (n = 23), and fatal MI (defined as MI combined with death within 2 days, n = 5). Average pFCG (molecules/platelet) in patient groups was 1,491 ± 655 (no events), 1700 ± 740 (single ischemic), 1850 ± 625 (multiple ischemic), and 1880 ± 440 (fatal MI, P < 0.001 for trend). Early ischemic events associated with high pFCG were driven by first ischemic endpoints and were accentuated between 6 months and 1 year by additional ischemic endpoints (HR: 2.7; 95% CI: 1.82-4.02; P < 0.0001). The average expression (molecules/platelet) in patients with bleeding (without ischemia) 1,491 ± 517 was similar to that in patients without ischemia or bleeding. High pFCG is associated with a greater risk of both first and additional ischemic endpoints. The strong association of pFCG with a greater risk of recurrent ischemic events may be useful to inform clinical decision-making. NCT05175261.
Salinity is an escalating agricultural challenge, yet plant microbiomes offer a promising avenue for improving salt tolerance. Nevertheless, most naturally occurring microbes remain unevaluated for plant growth-promoting function, and systematic approaches to uncover salt-tolerance-enhancing plant growth-promoting rhizobacteria (PGPR) are limited. Here, using soybean as a model, we implement a quantitative framework to characterize rhizosphere microbial networks and nominate novel taxa functionally associated with plant salt tolerance. We introduced a salt tolerance index (STI) to quantify plant salt tolerance and normalize performance across heterogeneous natural soil salinity. Metagenomic sequencing and co-occurrence analysis revealed distinct rhizosphere microbiota structures between tolerant and susceptible soybeans. In tolerant soybeans, Pseudomonas dominated as the hub of a highly interconnected network, whereas susceptible accessions showed a fragmented network dominated by Acinetobacter. Correlation analyses identified bacterial taxa positively associated with STI, including documented salt-tolerant PGPR and novel candidates. Greenhouse experiments showed that one candidate, Thalassospira xiamenensis, enhances soybean salt tolerance and reshapes host ion-transport and oxidative-stress gene expression under salinity, validating our screening strategy. Our culture-independent metagenomic association approach reveals host genotype-structured rhizosphere microbial networks underlying salt tolerance and provides an efficient, labor-saving means for high-throughput identification of salt-tolerant PGPR.
Chimeric antigen receptor (CAR) T cell therapy has limited efficacy against solid tumors such as prostate cancer due to the immunosuppressive tumor microenvironment (TME). Combining CAR T cells with existing therapies that remodel the TME and promote endogenous immune responses, such as radiation therapy and chemotherapies, may strengthen antitumor responses. Here, we assessed the potency of combining focal radiotherapy (RT), cyclophosphamide (Cy) preconditioning, and prostate stem cell antigen (PSCA)-CAR T cells against syngeneic prostate cancer models. Focal RT alone increased T cell and dendritic cell infiltration and activation in the irradiated tumor. Furthermore, the combination of all three therapies was critical for enhanced antitumor responses and survival across multiple subcutaneous, bone-metastatic, and multifocal disease models. This combination, in the irradiated TME and tumor-draining lymph nodes (tdLN), led to greater antigen presentation by myeloid cells and endogenous T cell activation and cytotoxicity. Our study demonstrates the potency of combining focal RT with PSCA-CAR T cells, significantly improving therapeutic responses in the irradiated tumor and contributing to a more robust systemic immune response against metastatic burden in prostate cancer.
Elongases are essential enzymes in the biosynthesis of sex pheromones in many lepidopteran species, where together with desaturases they determine pheromone precursor carbon skeletons and contribute to species-specific chemical communication. Despite their presumed importance, no fatty acyl elongase involved in lepidopteran sex pheromone biosynthesis has been functionally characterized to date. The rice leaffolder, Cnaphalocrocis medinalis, utilizes a blend of C18 monounsaturated aldehydes and alcohols as its sex pheromone, implying a critical elongation step from C16 precursors. Here, transcriptome analysis of the pheromone gland identified 45 candidate genes potentially involved in pheromone biosynthesis. Functional assays in Nicotiana benthamiana established a desaturation-elongation pathway in which the Δ11 desaturase Cmed070400 produces (Z)-11-hexadecenoic acid, which serves as the substrate for elongation. Multiple elongases, including Cmed092440, Cmed063180 and Cmed153360 catalyzed the conversion to (Z)-13-octadecenoic acid, the direct precursor of the major pheromone component. These findings provide the first experimental evidence for elongase-mediated formation of C18 pheromone precursors in C. medinalis. The identification of a minimal set of functionally active enzymes enables reconstruction of this pathway in plant systems, offering a basis for sustainable production of pheromone precursors for pest management applications.
Therapeutic vaccines represent a potential strategy for HIV-1 control without antiretroviral therapy (ART). We report a first-in-human study on ICVAX™, a PD-1-enhanced HIV-1 therapeutic DNA vaccine. This NMPA-approved, randomized, double-blind, placebo-controlled, dose-escalation phase I trial (NCT06253533) was conducted at Shenzhen Third People's Hospital, China. Adults with HIV-1 aged 18-50 years on ART were sequentially allocated to 1, 2, or 4 mg dose groups of 15, each randomized 12:3 to ICVAX or placebo. Participants received intramuscular injections with electroporation at Weeks 0, 4, 8, 12, and 36. Primary safety, secondary immunological, and exploratory virological outcomes were recorded. All 45 participants completed the trial, with 12 per ICVAX cohort and 9 in pooled Placebo. Treatment-related adverse events occurred in 100% of placebo and 83.3% of ICVAX recipients, all mild and transient, predominantly local reactions. A ≥2-fold increase of peak ELISpot T-cell responses over baseline was achieved by 75%, 75%, and 58.3% of low-dose, mid-dose, and high-dose recipients, versus 33.3% for Placebo. ICVAX recipients also showed low-frequency Gag-specific T cells expressing CD107a and/or effector cytokines. ICVAX was well-tolerated and elicited robust antigen-specific T-cell responses in ART-suppressed people with HIV-1. Its efficacy will be assessed in future studies.
Geroprotection aims at extending healthspan by delaying age-associated pathologies. Polyamines including spermine and spermidine are interconvertible metabolites whose longevity-promoting effects have traditionally been attributed to autophagy induction. In addition, recent evidence identifies spermine as an endogenous Fe2+ chelator that suppresses ferroptosis, thereby complementing the autophagy-inducing activity of spermidine. Indeed, spermidine inhibits EP300 acetyltransferase activity and supports hypusination-dependent activation of TFEB, both leading to autophagy. However, enhanced autophagic flux may increase susceptibility to ferroptosis through ferritinophagy and lipid remodeling. In parallel, polyamine catabolism generates H2O2 and acrolein, both of which facilitate lipid peroxidation and ferroptotic demise. The discovery that spermine directly chelates redox-active Fe2+ closes a conceptual gap by explaining how polyamine supplementation can promote longevity while avoiding excessive ferroptotic cell loss. Multiple lines of evidence including metabolomics, isotope tracing, cell-free lipid peroxidation systems, Fe2+-binding biophysics, mass spectrometry, Raman spectroscopy, nuclear magnetic resonance and disease models demonstrate that spermine limits labile iron and ferroptosis. Together, these findings support a unified model in which spermidine-driven autophagy and spermine-mediated ferroptosis inhibition cooperate to preserve tissue homeostasis and healthspan.