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.

He and colleagues developed a transient, non-gene-editing platform for off-the-shelf allogeneic TCR-T therapy. By conferring tacrolimus resistance characteristics to IL-2/4/7 expanded TCR-T cells that display reduced alloreactivity, He et al. provided a proof-of-concept for an allogeneic TCR-T therapy approach that addresses both manufacturability, and unwanted graft and host alloreactivity concerns.

Todo and colleagues describe the development of IL-12-expressing oncolytic HSV-1 derived from the clinically validated G47Δ platform for malignant melanoma. Intratumoral treatment controls both injected and distant tumors and shows enhanced efficacy with PD-1 blockade. Favorable preclinical safety supports clinical translation and has enabled the ongoing investigator-initiated clinical trial.

In 2012, I had the distinct honor of being part of one of the trailblazing teams leading first-in-human/first-in-child chimeric antigen receptor (CAR) T cell trials in B cell acute lymphoblastic leukemia (B-ALL). Little did I know then that we were only at the tip of the iceberg for use of immunotherapy in pediatric oncology. While the multiple CAR T cell approvals for B cell malignancies, including in pediatric B-ALL, reflect the curative potential in relapsed/refractory disease, what has additionally transpired is a cascade of novel immune-effector cell therapy-based approaches, each iteratively building upon the prior to further extend the therapeutic index of immunotherapy.

Nguyen and colleagues engineered bispecific TCEs targeting p53R273H neoantigens presented by HLA-C*01:02. Machine learning-guided optimization (Boltz-2/EvotProGrad) yielded TCE01Cr3 (Kd = 2.51 nM), a small, high-affinity, specific TCE with mouse serum stability (t1/2 ≈ 40 h), and monotherapy efficacy (> 80% tumor elimination in 3D models) without off-target toxicity or chemotherapy.

Zhang and colleagues engineered the poly(A) tail as a programmable regulatory element, showing that embedding cell-type-specific microRNA target sites directly within it confers robust, position-dependent silencing in off-target tissues while preserving activity in target cells. This strategy offers a new modular tool to enhance mRNA therapeutic safety and precision.

Kis and colleagues report an integrated sequential-batch IVT–oligo-dT process that links RNA synthesis and affinity capture through a shared buffer, enabling direct crude-IVT loading and flowthrough recycling. The workflow improves cap-analog utilization and raw-material efficiency while preserving functional mRNA expression across five cycles.

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.

Et1Ψ-modified mRNA shows UUU-dependent translational sensitivity, but targeted UUU-to-UUC recoding restores protein expression. With this sequence constraint addressed, Et1Ψ reduces early innate immune activation while preserving vaccine-induced humoral, cellular, and neutralizing responses, thereby establishing codon–modification compatibility as a practical design principle for mRNA therapeutics.

Lipid nanoparticles incorporating DOTAP and DOPE enable serial mRNA transfection of primary human T cells with significantly improved viable cell yields compared to standard electroporation. This platform supports iterative functional modifications across multiple rounds of delivery while preserving cell viability, facilitating the manufacturing of complex, multi-payload cell therapies.

Ionizable lipid components of LNP vaccine formulations play an integral role in providing the necessary immune stimulatory signals required for effective anti-tumor T cell responses. This study finds that inclusion of a novel ionizable lipid confers effective tumor control in part through the generation of type I interferons.

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.

(Molecular Therapy 31, 1596–1614; June 2023)

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.

Yuan and colleagues demonstrate that inhaled biodegradable nanosilica activates an alveolar macrophage–NK axis, triggering an IL-12/15/18 triad that programs memory-like NK cells. This non-fibrotic, cell-free strategy suppresses melanoma growth, prevents postsurgical recurrence, and synergizes with anti-PD-1, establishing a robust framework for in vivo NK cell immunotherapy.

Zhang and colleagues reveal that GPX4 serves as a critical regulator of NRF2-mediated otoprotection against aminoglycoside-induced hair cell injury. Their findings identify a GPX4-dependent antioxidant mechanism that enables therapeutic activation of NRF2 and provides new insights into strategies for preventing drug-induced hearing loss.