Molecular Therapy最新文献

Living cell therapy for immune regulation could potentially achieve long-lasting effects on inhibiting graft-versus-host disease (GVHD), a life-threatening complication after allogeneic hematopoietic cell transplantation (allo-HCT). However, novel immune regulatory cells with better persistence and sustained immune modulatory function are needed. Here, we demonstrate that engineering human T cells to co-express IFN-α2 and PD-L1 (named αp-T cells) rendered them a potent capacity to inhibit xenogeneic GVHD without impairing anti-leukemia in immunodeficient mice. These αp-T cells maintained stable expression of PD-L1 and IFN-α and potent immunosuppressive effects in peripheral tissues during GVHD control. CD4⁺ αp-T cells activated transcriptional programs that promoted effector differentiation, exhaustion, and proliferation inhibition in both themselves and their treated conventional T (conv-T) cells, ultimately reducing GVHD. These CD4⁺ αp-T cells created a synergistic regulatory loop that modulated conv-T cell responses, in which IFN-α suppressed expansion and survival of activated conv-T cells, promoted their differentiation into PD-1⁺TIM3⁺ exhaustion-like T cells, and sensitized them to PD-L1-mediated suppression. We further verified these regulatory effects of murine CD4⁺ αp-T cells on reducing GVHD in immunocompetent mice of allo-HCT. Thus, αp-T cells may represent a novel translational strategy to improve the safety and efficacy of allo-HCT and inhibit other inflammatory disorders in a broad context.

Identifying essential human microRNAs (miRNAs) remains challenging due to scarce experimental labels. Traditional domain adaptation methods for cross-species knowledge transfer have significant limitations in capturing intricate data features and handling high-dimensional input data. To address these limitations, we propose DeepHEM, a deep domain-adversarial learning framework for predicting human miRNA essentiality. During training stage, a multi-modal feature extractor is designed to capture comprehensive miRNA representations from sequence data, inherent properties, and miRNA-target gene interactions. A multi-loss optimization module is then employed to align feature distributions across two domains, learn domain-invariant features, and ensure robust classification performance. For prediction, the trained feature extractor processes target domain data to generate feature representations, which are then classified by a label predictor. DeepHEM's innovations lie in comprehensive multi-modal feature extraction and reinforced cross-domain feature alignment. Biological correlation analysis reveals significant correlation and consistent trends between predicted essentiality scores and biological indicators. Comparative evaluations against three domain adaptation methods, along with correlation analysis involving biological indicators, demonstrate DeepHEM's superiority. Ablation studies confirm the effectiveness of each individual component. Additionally, a case study confirms 8 of the top 10 predictions with experimental literature. In summary, DeepHEM provides an effective cross-species transfer framework for predicting human miRNA essentiality.

Thermal ablation (TA) (e.g., radiofrequency ablation [RFA]) is a critical curative therapy for hepatocellular carcinoma (HCC). However, the high recurrence rate of HCC after ablation remains a major clinical challenge. Hyperactive mRNA translation mediated by aberrant RNA modifications is vital for tumor heat stress adaptation. One of the most prevalent rRNA modifications is 18S rRNA N⁶-methyladenosine (m⁶A) modification catalyzed by methyltransferase 5 (METTL5); however, the role of METTL5-mediated rRNA modification in HCC recurrence after RFA remains unknown. Here, we found that the levels of METTL5 and 18S rRNA m⁶A modification were significantly upregulated in post-RFA recurrent HCC and were further verified by insufficient RFA (iRFA) models in vitro and in vivo. Four mouse models, together with functional cell experiments, showed that METTL5 promoted HCC progression under heat stress. Mechanically, we demonstrated that heat-mediated METTL5 upregulation enhanced the PEX16 translation that promoted peroxisomal biogenesis and β-oxidation of very long-chain fatty acid, which promoted mitochondrial respiration to mediate HCC progression. Our study reveals the novel mechanistic insights in HCC recurrence after iRFA, demonstrating the important role of heat stress-mediated tumor metabolism adaptation by mRNA translation in HCC development. These findings identify METTL5 as a potential therapeutic target to prevent and treat HCC recurrence after TA.

Multiple lines of genetic and neuropathological evidence strongly implicate α-synuclein (αSyn) as a causal factor in synucleinopathies like Parkinson disease (PD), with numerous studies supporting the therapeutic concept of αSyn reduction as a disease-modifying strategy. Artificial miRNAs can be durably expressed from adeno-associated viral (AAV) vectors and have shown increasing promise as a highly specific modality to reduce levels of target mRNA. This modality is therefore well suited for the long-term treatment of PD and related diseases. Here, we assessed the therapeutic potential of the partial reduction of αSyn by AAV-mediated artificial microRNA (amiRNA) expression by using the pre-formed fibril model of α-synucleinopathy to induce the seeding and spreading of αSyn pathology in wild-type animals. We demonstrate that a 50% decrease in endogenous SNCA mRNA levels is sufficient to fully block the spread of pathogenic αSyn protein and prevent the loss of dopaminergic neurons in the substantia nigra. To support clinical translation, human-specific amiRNAs were designed and evaluated in vitro and in vivo. Concatenation of amiRNAs was evaluated to optimize the potency of individual guide sequences. Several candidates had excellent expression, processing, and SNCA reduction, supporting the continued development of αSyn-lowering therapeutics as a safe and efficacious approach for treating patients with synucleinopathies.

Non-invasive imaging of gene expression remains a critical challenge in gene therapy. Here, we report the first in vivo demonstration of a genetically encoded chemical exchange saturation transfer (CEST) MRI reporter expressed from a recombinant adeno-associated virus (rAAV). The genetically encoded peptide-based reporter superCESTide enables detection of transgene expression without the need for exogenous contrast agents. Mice were systemically administered rAAV encoding superCESTide and fluorescence reporter tdTomato, and they were evaluated using CEST MRI, fluorescence imaging, and reverse transcription-PCR. CEST MRI revealed dose-dependent signal enhancement in the liver, with a strong correlation between CEST contrast and superCESTide expression (r = 0.75, p < 0.05). Mice receiving the highest dose (1.4 × 10¹³ vector genomes/kg) exhibited the strongest MTR_asym signal (mean = 0.0225), and histogram analyses showed distinct expression patterns. These results establish CEST MRI as a powerful, non-invasive method for monitoring rAAV-mediated transgene expression in vivo, opening new possibilities for imaging-guided gene therapy.

Engineered bacterial systems have been widely utilized as versatile biotherapeutic platforms in disease intervention, yet their therapeutic efficacy remains limited. Recent advancements in synthetic biology have enabled the systematic deconstruction of bacteria through rational computational modeling and modular-genetic circuit design, thereby overcoming intrinsic functional limitations. These engineered bacteria represent a promising therapeutic strategy characterized by enhanced spatiotemporal precision, biosafety profiles, and scalable production. Moreover, the advent of sophisticated genetic engineering technologies has facilitated the expansion of bacterial therapeutic applications into new areas, including vaccine development and tumor-microenvironment modulation. Here, we systematically examine recent advances in the development and testing of engineered bacteria across different organs on the basis of tissue specificity. We further summarize the therapeutic roles of engineered bacteria in tumor treatment and metabolic-disorder management. Additionally, we outline advanced technologies for engineered bacterial development, emphasizing their critical roles in optimizing the overall clinical efficacy of therapeutic agents after system-level optimization.