Recombinant AAV gene therapies deliver durable clinical benefit but face immune-mediated challenges that vary by vector, dose, route, and patient. Gulve and colleagues synthesize the clinical manifestations, temporal patterns, and mechanisms of rAAV immunogenicity, highlighting risk assessment and emerging mitigation strategies to support safer, more effective gene therapy development.

Infusion-related reactions (IRRs) can occur with AAV gene therapy. IRRs were observed in two participants who received AAV8 gene therapy and in one who received AAV9 gene therapy. AAV gene therapy IRRs may be rate-related, possibly driven by complement activation. In our studies, a slow, staged infusion mitigated additional IRRs.

Transduction efficiency is a critical determinant of CAR T-cell manufacturing quality. Analysis of 204 clinical CAR T-cell products revealed that transduction efficiency is shaped primarily by manufacturing workflows and protocol-dependent starting material composition. Higher transduction efficiency was associated with early memory-like cellular states, providing insights into optimizing CAR T-cell.

Automated ÄKTA-connected zonal-rotor CsCl ultracentrifugation enables high-resolution AAV polishing at 1.65-L scale. The workflow enriched full capsids and resolved empty and intermediate populations in AAV8-X and AAV9-Y. The AAV8-X intermediate fraction retained about a half of qPCR signal of equimolar full capsid but produced minimal mRNA expression potency.

COL17A1 nonsense mutations generate premature termination codons, reducing collagen XVII through truncated protein production and/or nonsense-mediated decay (NMD), and causing junctional epidermolysis bullosa. Translational readthrough drug offers a potential approach to nonsense mutations suppression. Different COL17A1 nonsense mutations showed distinct responses to readthrough drugs and NMD inhibitors, supporting personalized therapeutic strategies.

CRISPR hybrid guides containing RNA and DNA nucleotides (chRDNA) enhance Cas nuclease specificity by reducing off-target editing. Intravenous administration of LNP-encapsulated Cas12a mRNA with a chRDNA guide in mice led to dose-responsive hepatic genome editing without detectable off-target editing and near elimination of the targeted plasma protein for one year.

Immunization with a nucleoside-modified mRNA-LNP vaccine encoding the conserved malaria antigen E140 induced durable humoral immunity in mice and generated invasion-blocking antibodies against Plasmodium vivax. The results highlight E140 as a promising candidate for next-generation malaria vaccines.

Circular RNAs (circRNAs) have emerged as promising next-generation therapeutic platforms due to their exceptional stability, prolonged protein expression, and low immunogenicity. This review highlights advances in enzymatic, chemical, and bio-orthogonal circRNA synthesis, compares their advantages and limitations, and discusses emerging applications in vaccines, cancer therapy, and precision medicine.

Yuzaki and colleagues identify biallelic CBLN1 variants as a cause of early-onset hereditary ataxia and show that loss of extracellular CBLN1 disrupts cerebellar synapses. Astrocyte-targeted AAV delivery restores synaptic CBLN1 and rescues circuit and motor dysfunction, establishing extracellular synaptic organizer replacement as a therapeutic strategy.

Blancafort and colleagues describe a non-viral polymeric system for the delivery of dCas9-KRAB silencers as ribonucleoprotein (RNP) payloads for EWSR1-FLI1 repression. They demonstrate highly efficient RNP delivery and robust silencing of EWSR1-FLI1 in both cell line and patient-derived xenografts of Ewing sarcoma, accompanied by potent anti-tumor effects.

Ouabain triggers cochlear spiral ganglion neuron (SGN) ferroptosis and hearing loss via SCD1 downregulation. MITF directly activates Scd1 transcription, and the MITF–SCD1 axis mitigates SGN ferroptosis and hearing impairment in ototoxic ouabain and cisplatin models, revealing a lipid metabolic vulnerability and therapeutic target for sensorineural hearing loss.

This study identifies the interleukin-2-inducible T cell Kinase (ITK)-mediated phosphorylation of Y218 in the CD28 cytoplasmic domain as key for CAR-T cell function and demonstrates that engineering a synthetic ITK-binding motif into the CAR enhances IL-2 production and antitumor efficacy in vivo.

Utilizing medicinal chemistry approach, Datta and colleagues demonstrate that EZH2-EGFR dual targeting halts protein translation in TNBC and inhibits tumor growth and metastasis in preclinical animal models.

The efficacy of vaccines remains suboptimal in many settings, underscoring the need for new strategies. Baydemir and colleagues show that modulation of acetyl-CoA metabolism reshapes metabolic and epigenetic programs underlying Bacille Calmette-Guérin-induced trained immunity, enhancing cellular innate immune responses and identifying immunometabolic targeting as a promising approach to improve vaccine efficacy.

AAV gene therapies are powerful but often limited by inefficient or unwanted tissue delivery. This study maps host genes that help or hinder AAV transduction, revealing that transiently tuning these factors can reshape vector biodistribution, offering a new strategy to improve gene therapy precision.

This study characterizes a GalNAc-C5 small interfering RNA with potent in vitro and in vivo activity. Single subcutaneous dosing sustains long-term C5 suppression in cynomolgus monkeys with IgA nephropathy, outperforming Nefecon in blocking glomerular complement deposition, supporting its standalone or combinational clinical application.

Liu and colleagues develop a dendritic cell vaccine loaded with the gut commensals Bacteroides fragilis (DC-Bf), which triggers CD8+ T cell-dependent antitumor immune responses via MHC-I-mediated cross-presentation and interleukin-12 secretion, and enhances the efficacy of PD-1 antibody by improving the immunosuppressive tumor microenvironment and diversifying the T cell receptor repertoire.

Liu and colleagues developed a robust in cellulo plasmid DNA replication system in human cells for replicating plasmid-borne adeno-associated virus (AAV) Rep/Cap genes during recombinant AAV (rAAV) production. This new approach not only enables a 10- to 20-fold plasmid reduction to significantly lower manufacturing costs but also substantially enhances rAAV potency, titer, and purity.

This study developed a multi-layer inducible RNA switch that achieves transient expression of gene-delivery vectors in hepatic and non-hepatic tissues. As an exemplary application, this RNA switch triggers pulsive expression of gene editors that reduces the off-target effects and immunotoxicity of gene editing.

NFAT3-regulated extracellular vesicles deliver a combinatorial miRNA signature that suppresses proliferation and invasion in aggressive breast and pancreatic cancer models. This study identifies the underlying molecular programs targeted by the miRNA combination and supports extracellular vesicles as a promising platform for multi-target RNA-based cancer therapy.