Anwar S, Hay S, Moriyama H
… +3 more, Mir F, Maruyama R, Yokota T
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42327837
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Fibrodysplasia ossificans progressiva (FOP) is an ultra-rare disorder caused by gain-of-function mutations in , most commonly c.617G>A (R206H), leading to progressive heterotopic ossification. In this study, we developed...Fibrodysplasia ossificans progressiva (FOP) is an ultra-rare disorder caused by gain-of-function mutations in , most commonly c.617G>A (R206H), leading to progressive heterotopic ossification. In this study, we developed novel antisense gapmers selectively targeting the mutant transcript while sparing the wild-type allele. We engineered locked nucleic acid (LNA) and 2'-O-methoxyethyl (MOE) gapmers incorporating a single 2'-O-methyl (2'OMe) modification at gap position 2. This is hypothesized to synergize with the wild-type sequence mismatch to restrict RNase H1 cleavage, limiting wild-type degradation while preserving mutant target engagement. In FOP patient-derived fibroblasts carrying the endogenous mutation and in murine-derived C2C12 cells ectopically expressing constructs, 2'OMe-modified gapmers demonstrated robust and preferential suppression of at both RNA and protein levels. Gapmer treatment also reduced osteogenic differentiation, as shown by decreased alkaline phosphatase and Alizarin Red S staining, and lower expression of osteogenic markers. In wild-type mice, 2'OMe modification was associated with higher apparent gapmer levels in skeletal muscle and tendon and lower hepatic and renal stress marker readouts. These findings provide preliminary proof-of-concept that a single-base chemical modification can modulate allele selectivity and biodistribution of gapmers targeting . Further studies in disease-relevant FOP models will be needed to establish therapeutic efficacy and long-term safety.
Gene delivery for neurodegenerative cerebral disorders faces formidable structural and practical challenges. The blood-brain, blood- cerebrospinal fluid (CSF), and arachnoid barriers tightly regulate molecular traffic, r...Gene delivery for neurodegenerative cerebral disorders faces formidable structural and practical challenges. The blood-brain, blood- cerebrospinal fluid (CSF), and arachnoid barriers tightly regulate molecular traffic, restrict paracellular diffusion, and actively clear xenobiotics, limiting brain penetration and retention of large molecules and nucleic acid therapeutics. Additional barriers include heterogeneous and diffuse pathology, the need for precise anatomical targeting, vector dose-limiting toxicities, pre-existing and therapy-induced immunity to viral capsids, and procedural risks of neurosurgical or intrathecal administration. These constraints have slowed translation, reflected by the small number of approved central nervous system (CNS)-directed gene therapies. Against this backdrop, a diverse therapeutic landscape has emerged. In vivo strategies are dominated by adeno-associated virus (AAV)-9 and AAV2 vectors delivered intravenously, intrathecally (including intracisternal and intracerebroventricular routes), or via image-guided intraparenchymal and intraputaminal infusions, alongside intrathecal antisense oligonucleotides and RNA interference therapeutics. Concurrently, emerging approaches, including engineered AAV capsids, receptor- and transporter-mediated transcytosis, nanoparticle platforms, and focused ultrasound with microbubbles, have demonstrated compelling yet preclinical proof-of-concept. Future progress will likely depend on convergent advances in machine learning-guided capsid, more controllable blood-brain barrier modulation, rational route selection tailored to disease topology, and optimized ex vivo and cell-mediated delivery strategies. These innovations could enable a more predictable therapeutic paradigm for cerebral neurodegeneration.
Okada H, Nio K, Ohno N
… +15 more, Masuo Y, Hirokawa T, Shimakami T, Miyati T, Takamura H, Ohta T, Kitao A, Kobayashi S, Gabata T, Matsui O, Kato Y, Seiki M, Honda M, Kaneko S, Yamashita T
Mol Ther Oncol
· 2026 Jun · PMID 42318058
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Activation of tumor suppressors represents an attractive strategy for cancer treatment. Hepatocyte nuclear factor 4 alpha (HNF4A) functions as a tumor suppressor in the liver by inhibiting hepatocyte proliferation; howev...Activation of tumor suppressors represents an attractive strategy for cancer treatment. Hepatocyte nuclear factor 4 alpha (HNF4A) functions as a tumor suppressor in the liver by inhibiting hepatocyte proliferation; however, no effective agonists have been identified. Here, we aimed to identify novel ligands for HNF4A and evaluate their role in hepatocarcinogenesis. We identified polyprenoic acid (PA, peretinoin) as the first ligand capable of directly binding HNF4A. Reanalysis of prior clinical trial data revealed that PA suppressed the progression of dysplastic nodules (DNs) to hepatocellular carcinoma (HCC), while showing no effect on local HCC recurrence after initial treatment. Consistently, PA inhibited DN growth but not established HCC in Pdgf-C transgenic mice. Mechanistically, PA selectively bound to the HNF4A P1 isoform, enhancing its transcriptional activity and upregulating hepatocyte maturation markers (ALB, TTR, and SLCO1B3), while suppressing alpha-fetoprotein expression driven by the HNF4A P2 isoform. Importantly, hepatocyte-specific knockdown or lipid-nanoparticle-mediated siRNA abrogated the protective effects of PA. These findings establish HNF4A as a pharmacologically controllable tumor suppressor and highlight PA-like compounds as promising agents for preventing liver carcinogenesis.
Taguchi S, Fukuhara H, Cai S
… +7 more, Li XK, Naito A, Kakutani S, Takeshima Y, Miyakawa J, Kume H, Todo T
Mol Ther Oncol
· 2026 Jun · PMID 42318057
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Oncolytic herpes simplex virus type 1 (HSV-1), including marketing-approved G47Δ, elicits antitumor immunity via facilitating antigen presentation during immune-mediated clearance of infected tumor cells. Dendritic cell...Oncolytic herpes simplex virus type 1 (HSV-1), including marketing-approved G47Δ, elicits antitumor immunity via facilitating antigen presentation during immune-mediated clearance of infected tumor cells. Dendritic cell (DC) therapy should therefore enhance oncolytic HSV-1 efficacy, but its clinical application is hindered by the need to generate autologous DCs. Induced pluripotent stem cell (iPSC) technologies offer a scalable and standardized source of DCs. Here, we investigate the therapeutic potential of combining iPSC-derived DCs (iPSDCs) with an interleukin-12-expressing oncolytic HSV-1, T-mfIL12. iPSDCs, generated from a murine embryonic fibroblast-derived iPSC line, possess characteristics and functions comparable to bone marrow-derived DCs (BMDCs), including costimulatory molecule expression, antigen uptake capacity, and the ability to stimulate CD8 and CD4 T cells isolated from ovalbumin-specific transgenic mice both and . In a syngeneic murine bladder cancer model, iPSDCs markedly enhanced the efficacy of intratumorally inoculated T-mfIL12, eliciting augmented immune responses including intratumoral T cell infiltration, tumor-responsive T cell activation, and maturation of DCs, similarly to BMDCs. The results indicate that iPSDCs can serve as an effective and practical alternative to conventional DCs in oncolytic virus immunotherapy. The combination of T-mfIL12 with iPSDCs offers a promising and clinically applicable cancer treatment strategy.
Xue W, Wang Y, Smirnova AV
… +4 more, Malakhov PA, Pustovalova M, Kuzmin DV, Leonov S
Mol Ther Oncol
· 2026 Jun · PMID 42318056
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The TGF-β signaling pathway has both tumor-suppressive and metastasis-promoting effects in cancer, yet the molecular determinants governing this switch remain unclear. Here, we investigated the miR-16-1-3p/MDM2/p53 axis...The TGF-β signaling pathway has both tumor-suppressive and metastasis-promoting effects in cancer, yet the molecular determinants governing this switch remain unclear. Here, we investigated the miR-16-1-3p/MDM2/p53 axis as a potential regulator of TGF-β/Smad signaling output in osteosarcoma. miR-16-1-3p overexpression alone markedly reduced the proliferative and clonogenic potential of U2OS cells, while its combination with TGF-β treatment strongly enhanced G1-phase arrest and nearly abolished tumor growth capacity. miR-16-1-3p inhibited TGF-β-induced actin remodeling and epithelial-mesenchymal transition (EMT)-associated changes. While TGF-β promoted both 2D and 3D migration, miR-16-1-3p overexpression, alone or combined with TGF-β, counteracted its pro-migratory effects. Mechanistically, miR-16-1-3p reduced MDM2 expression and stabilized p53, which was associated with enhanced p21 induction and suppression of proliferative responses under TGF-β stimulation. Co-administration of TGF-β and miR-16-1-3p markedly increased cisplatin sensitivity in wild-type U2OS cells and reduced tumor nodule volume, Ki67 expression, and metastasis in the chicken chorioallantoic membrane model. Collectively, these findings suggest that miR-16-1-3p biases TGF-β signaling output toward anti-growth responses, while attenuating pro-migratory effects through MDM2 inhibition and p53 stabilization, providing a mechanistic rationale for improving therapeutic responses in osteosarcoma.
Metabolic dysfunction-associated steatotic liver disease (MASLD) is increasingly recognized as an independent risk factor for cardiovascular disease (CVD), yet the underlying molecular pathways mediating liver-to-vascula...Metabolic dysfunction-associated steatotic liver disease (MASLD) is increasingly recognized as an independent risk factor for cardiovascular disease (CVD), yet the underlying molecular pathways mediating liver-to-vasculature communication remain largely unidentified. Here, we show that extracellular vesicles (EVs) derived from hepatocytes contribute to endothelial dysfunction and atherogenesis. Treatment with EVs derived from palmitic acid (PA)-treated hepatocytes induced inflammation and endothelial dysfunction in human endothelial cells. miR-30b-5p was identified as a candidate cargo, and its elevation was confirmed by qPCR in EVs from PA-treated hepatocytes and Western-diet-induced steatotic livers. ELOVL5 (elongation of very long chain fatty acid protein 5), a key enzyme involved in fatty acid elongation, was identified as a direct target of miR-30b-5p. Overexpression of miR-30b-5p or knockdown of Elovl5 inhibited the elongation of polyunsaturated fatty acids (PUFAs), leading to endothelial inflammation, which was rescued by PUFA supplementation. In vivo, overexpression of miR-30b-5p accelerated atherogenesis, whereas its inhibition ameliorated vascular lesions. miR-30b-5p levels in human serum-derived EVs positively correlated with the early-stage MASLD indices. Our findings identify the EV-derived miR-30b-5p/ELOVL5 axis as a novel mechanistic link between MASLD and CVD. miR-30b-5p may function as both a biomarker and a therapeutic target for early intervention of CVD among patients with MASLD.
Pseudomonas aeruginosa (P.a) is a Gram-negative opportunistic pathogen that poses a major global health threat, particularly in immunocompromised individuals, patients with cystic fibrosis, and those with burn injuries o...Pseudomonas aeruginosa (P.a) is a Gram-negative opportunistic pathogen that poses a major global health threat, particularly in immunocompromised individuals, patients with cystic fibrosis, and those with burn injuries or ventilator-associated pneumonia. Despite intense efforts, no vaccine is available for human use. In this context, live attenuated vaccines represent a promising but underexplored approach, offering the potential to elicit robust, long-lasting, and multifaceted immune responses including inducing trained immunity. Here, we sub-cultured ΔLasB-PAO1 (a P.a strain we have previously shown to have reduced virulence) in artificial sputum medium, a culture medium mimicking cystic fibrosis (CF) sputum in which bacteria often show auxotrophy. We showed that such strain ("V" for vaccine) was auxotrophic, less virulent, and had characteristics of "CF-like strains." Crucially, "V" induced both local (IgA) and systemic humoral responses as well as memory Th17 immune responses, and could, when administered in the lung, but not intra-muscularly, protect mice against a lethal PAO1-WT infection. Overall, the present study demonstrates that our vaccine formulation, in addition to providing an advantageous auxotrophic phenotype adapted to the CF setting, was efficient, when given mucosally and was associated with a pathway involving Th17 responses and secretory IgA, a critical barrier that neutralizes pathogens before tissue invasion.
Bueloni B, Fusco M, Fiore E
… +12 more, Malvicini M, Cantero MJ, Lameroli L, Casadei M, Ruiz BM, Mercogliano F, Santamaría E, Atorrasagasti C, Argemi J, Canbay A, Bayo J, Mazzolini G
Mol Ther Oncol
· 2026 Jun · PMID 42306568
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Therapeutic options for hepatocellular carcinoma (HCC) remain scarce and limited by resistance, underscoring the need for novel approaches. The lysine methyltransferase SMYD2 is overexpressed in several cancers and regul...Therapeutic options for hepatocellular carcinoma (HCC) remain scarce and limited by resistance, underscoring the need for novel approaches. The lysine methyltransferase SMYD2 is overexpressed in several cancers and regulates oncogenic signaling through histone and non-histone substrates. However, its role in HCC is incompletely defined. We integrated transcriptomic analyses of patient samples with assays and orthotopic murine models to evaluate the therapeutic potential of SMYD2 inhibition in HCC. was enriched in intratumoral regions and associated with immunosuppressive and Wnt/β-catenin-driven programs. , SMYD2 pharmacological inhibition with AZ-505 reduced tumor burden and induced pro-inflammatory transcriptional changes. The antitumor activity of AZ-505 was also evident in Wnt/β-catenin-active tumors, which further displayed reduced expression of proliferative and immunosuppressive signatures. , SMYD2 inhibition decreased tumor-cell Wnt ligand transcription and shifted macrophage responses toward reduced expression, in a pattern consistent with reduced Wnt/β-catenin-associated signaling. Finally, AZ-505 synergized with anti-PD-1 in Wnt/β-catenin-active tumors, expanding cytotoxic T cell and antigen-presenting populations. Transcriptomic analyses of combination-treated tumors showed suppression of Wnt/β-catenin- and TGF-β-pathway-associated signatures alongside activation of immune-stimulatory responses. These results position SMYD2 as a dual regulator of tumor growth and immune suppression, and as a promising target to overcome ICI resistance in HCC.
Zhang L, Pham L, Steele MB
… +4 more, Jenks N, Nace R, Russell SJ, Peng KW
Mol Ther Oncol
· 2026 Jun · PMID 42306567
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Oncolytic viruses (OVs) encoding interferon beta (IFN-β) are under clinical evaluation. IFN-β is an attractive transgene due to its antiproliferative and immunomodulatory properties; however, the systemic effects of sust...Oncolytic viruses (OVs) encoding interferon beta (IFN-β) are under clinical evaluation. IFN-β is an attractive transgene due to its antiproliferative and immunomodulatory properties; however, the systemic effects of sustained IFN-β expression remain poorly defined. Here, we demonstrate that prolonged circulating IFN-β induces dose-dependent toxicity in murine models. Persistent IFN-β exposure led to elevated transaminases, thrombocytopenia, lymphopenia, and reduced hemoglobin. Histopathologic analysis revealed sinusoidal endothelial sloughing, microvascular coagulation with microthrombi, and hepatocellular vacuolation and degeneration, along with bone marrow hemorrhage and cell death. These toxicities were absent in type I interferon receptor knockout mice. Importantly, termination of IFN-β production via inducible caspase-9 in adeno-associated vector-transduced cells reversed toxicity and normalized serum transaminases. In Balb/c mice bearing MPC-11 myeloma tumors which are highly permissive to IFN-β-expressing OVs, treatment induced rapid tumor lysis, elevated plasma IFN-β, and mortality. Importantly, co-administration of ruxolitinib, a clinically approved JAK1/2 inhibitor, dampened IFN-β signaling and rescued mice from lethal toxicity. Collectively, these findings define the pathophysiological consequences of sustained systemic IFN-β exposure and identify ruxolitinib as a potential mitigation strategy to manage IFN-β-mediated toxicity during OV treatment.
Jo S, Shim WC, Park S
… +13 more, Kweon T, Ryu WJ, Hwang Y, Kim MW, Ahn JH, Lee YJ, Lee SH, Won D, Nam EJ, Han JW, Kim TI, Park JS, Park HS
Mol Ther Oncol
· 2026 Jun · PMID 42306566
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Germline mutations in homologous recombination-related genes significantly increase the lifetime risk of breast, ovarian, and other cancers; thus, the concept of BRCAness has been extensively studied. In this study, we a...Germline mutations in homologous recombination-related genes significantly increase the lifetime risk of breast, ovarian, and other cancers; thus, the concept of BRCAness has been extensively studied. In this study, we aimed to investigate the association between germline mutations and breast cancer in clinical and experimental settings. Germline pathogenic or likely pathogenic variants in were identified through multigene panel testing in the PLEASANT studies. Fourteen cases of breast cancer with germline pathogenic or likely pathogenic variants in were identified. Compared to 1,446 wild-type cases of breast cancer, they appeared to be more aggressive. The proportion of patients with triple-negative breast cancer (TNBC) was higher and larger tumors (>2 cm) were more common. RAD51D-deficient tumors showed better pathological complete responses, defined as ypT0 ypN0, to neoadjuvant chemotherapy. Based on clinical findings, we analyzed the impact of RAD51D-deficiency on drug sensitivity to identify targetable vulnerabilities in RAD51D-deficient tumors. These TNBC cells were significantly more sensitive to cisplatin than wild-type TNBC cells, consistent with our clinical data. In conclusion, RAD51D-deficient tumors were shown to have a phenotype similar to that of BRCA1-deficient tumors, and further investigation of responses to DNA-damaging agents, particularly platinum-based chemotherapy, is warranted.
Naoe Y, Fujimoto N, Makita Y
… +3 more, Li D, Inukai N, Hotta A
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42306087
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Ensuring the safety of CRISPR-Cas9 genome editing requires comprehensive assessment of off-target mutagenesis, particularly for therapeutic applications under regulatory review. DNA-free lipid nanoparticle (LNP) delivery...Ensuring the safety of CRISPR-Cas9 genome editing requires comprehensive assessment of off-target mutagenesis, particularly for therapeutic applications under regulatory review. DNA-free lipid nanoparticle (LNP) delivery is expected to minimize insertional risks compared to adeno-associated virus (AAV) vectors, but experimental validation has been limited. Here, on-target amplicon sequencing in mouse muscle demonstrated insertions largely derived from host genomic sequences, with no detectable AAV or transgene integration. Benchmarking 13 prediction tools identified Cas-OFFinder as the most sensitive, though precision remained low, and correlation with CIRCLE-seq data was modest. To strengthen off-target detection, we employed karyotypically normal human iPSCs and developed an "indel cluster" method using high-depth whole-genome sequencing, enabling discrimination between clustered genome editing-induced indels and background variants. Integration of predictions, CIRCLE-seq cleavage sites, and WGS clusters yielded 11 high-confidence off-target candidates, predominantly associated with one gRNA. While sensitivity was maximized, reproducibility across conditions remained limited, and many candidate sites overlapped repetitive or low-mappability regions. Our multilayered framework demonstrates both the utility and current limitations of off-target risk assessment, providing a practical foundation for the safety evaluation of genome editing therapies.
Rezek FO, Sanchez-Pintado B, Eden ER
… +8 more, Corral-Serrano JC, Aychoua N, Webster AR, de Guimarães TAC, Carr AF, Michaelides M, Cheetham ME, van der Spuy J
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42306086
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Doyne honeycomb retinal dystrophy is an incurable juvenile macular dystrophy that leads to visual impairment by early to mid-adulthood. It is an autosomal dominant disorder caused by a c.1033C>T, p.(Arg345Trp) variant in...Doyne honeycomb retinal dystrophy is an incurable juvenile macular dystrophy that leads to visual impairment by early to mid-adulthood. It is an autosomal dominant disorder caused by a c.1033C>T, p.(Arg345Trp) variant in , and is characterized by the early onset of extracellular deposition of drusen between the retinal pigment epithelium and underlying Bruch's membrane. In this study, we developed an antisense oligonucleotide approach to target . We reprogrammed patient-derived renal epithelial cells to induced pluripotent stem cells, followed by directed differentiation to retinal pigment epithelium and compared the phenotype to gene-corrected and knockout patient-derived retinal pigment epithelium. In the patient-derived disease model, remodeling of the extracellular matrix (ECM) occurred with progressive accumulation of the drusen-associated proteins apolipoprotein E and collagen IV, in addition to the intracellular accumulation and extracellular deposition of lipids. We developed an allele-specific antisense oligonucleotide which specifically and effectively promoted the clearance of the c.1033C>T transcript in the patient-derived disease model following assisted or gymnotic delivery. Gymnotic delivery rescued remodeling of the ECM, reduced intracellular accumulation of lipids, and cleared extracellular deposits, even after the onset of the disease phenotype, suggesting that this could be a practical and effective therapeutic approach.
Murmann AE, Ebadi M, Patel M
… +14 more, Ewe A, Barajas S, Xiao S, Ko MJ, Lee S, Cai W, Paudel B, Sun L, Bartom ET, Kocherginsky M, Liu Y, Kim DH, Aigner A, Peter ME
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42306085
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RNA interference (RNAi) regulates gene expression through small RNAs that act via Argonaute-containing RNA-induced silencing complexes (RISCs). We previously found that short RNAs with G-rich 6mer seeds (e.g., GGGGGC and...RNA interference (RNAi) regulates gene expression through small RNAs that act via Argonaute-containing RNA-induced silencing complexes (RISCs). We previously found that short RNAs with G-rich 6mer seeds (e.g., GGGGGC and G5C) can kill cells by targeting C-rich 3' UTR seed matches in essential survival genes (SGs), a mechanism termed death induced by survival gene elimination (DISE). To assess therapeutic potential, we systemically delivered two DISE-inducing sRNAs, sG5C and sCAG (based on CAG trinucleotide repeats), using lipopolyplexes (LPPs) composed of low-molecular-weight polyethyleneimines and lipids. In mouse ovarian and prostate cancer models and a rat hepatocellular carcinoma model, LPP-delivered small RNAs (sRNAs) markedly reduced or eliminated tumors without harming normal tissues. Predicted SG targets were engaged in tumors. Transcriptomic analyses across 10 major human cancers showed that many sG5C-targeted SGs are consistently upregulated in tumors and increase with stage, revealing a therapeutic window. These results support LPP-delivered DISE-inducing sRNAs as a promising pan-cancer therapy.
Maximov VV, Shebanov N, Nikitchina N
… +5 more, Rapoport R, Maor Y, Tarassov I, Pines O, Entelis N
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42306084
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Mutations in mitochondrial DNA (mtDNA) are associated with severe human diseases, lacking efficient therapies. Direct correction of mtDNA mutations may offer a cure for such diseases. We propose a novel strategy based on...Mutations in mitochondrial DNA (mtDNA) are associated with severe human diseases, lacking efficient therapies. Direct correction of mtDNA mutations may offer a cure for such diseases. We propose a novel strategy based on double-stranded DNA (dsDNA) oligonucleotide delivery into mitochondria and intrinsic microhomology-mediated end joining (MMEJ) for mtDNA editing. This strategy enables the introduction of multiple predefined nucleotide changes in mtDNA. For this, the presence of MMEJ activity in the human mitochondrial lysates was confirmed. Forty-nine bp DNA oligonucleotide duplexes, fused to an RNA hairpin previously identified as a mitochondrial import signal, were delivered into the mitochondria of cultured human cells. Delivery of these donor dsDNA molecules, homologous to an site of mtDNA and bearing designed nucleotide changes, led to a low but statistically significant introduction of the intended nucleotide changes into mtDNA. Donor dsDNA delivery combined with the CRISPR-mito-AsCas12a system also resulted in a statistically significant number of an expected concomitant change of five nucleotides distributed across a 16 nt site of the mitochondrial genome. The proposed strategy may become an efficient mtDNA editing tool suitable for the correction of near-homoplasmic mutations, such as Leber's hereditary optic neuropathy (LHON)-associated mutations in the gene of mtDNA.
Yang Y, Qiu Y, Wang K
… +5 more, Liu Y, Sanyal G, Whitford PC, Rouhanifard SH, Xie W
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42306083
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mRNA lipid nanoparticle (mRNA-LNP) technology has emerged as a cornerstone in vaccine development due to its high delivery efficiency, molecular stability, and favorable safety profile. However, rapid and reliable potenc...mRNA lipid nanoparticle (mRNA-LNP) technology has emerged as a cornerstone in vaccine development due to its high delivery efficiency, molecular stability, and favorable safety profile. However, rapid and reliable potency assessment remains challenging because of limited mechanistic understanding of delivery processes and sparse experimental data. To address these gaps, we introduce a mechanism-informed, multi-scale kinetic modularized modeling framework that quantitatively captures the coupled dynamics of mRNA delivery across nanoparticle, cellular, and macroscopic scales. The model incorporates variability in LNP-cell interactions and integrates key determinants, such as dosage, LNP and cell size distributions, cell proliferation, and membrane properties-factors that critically shape delivery efficiency and response heterogeneity. Its cell-based architecture and modular design enable adaptability to diverse delivery systems and physiological contexts. By leveraging advanced multi-omics assays, including single-molecule fluorescent hybridization (smFISH) for single-cell resolution of mRNA and protein expression, our framework provides mechanistically grounded modeling and robust prediction of therapeutic potency, offering a powerful platform for optimizing mRNA-based interventions.
Philosof NR, Walsh APG, Xu B
… +14 more, Watson AMD, Ren S, Hann L, Nathaniel B, Nguyen A, Liu H, Shi P, Bongcaron V, Nguyen H, Noonan J, Avci-Adali M, Peter K, Vidallon MLP, Wang X
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42306082
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Messenger RNA (mRNA) therapeutics offer a promising strategy for treating inflammatory disease by enabling transient local expression of therapeutic proteins. CD39 (ectonucleoside triphosphate diphosphohydrolase-1) hydro...Messenger RNA (mRNA) therapeutics offer a promising strategy for treating inflammatory disease by enabling transient local expression of therapeutic proteins. CD39 (ectonucleoside triphosphate diphosphohydrolase-1) hydrolyzes pro-inflammatory ATP and ADP and plays a central role in localized immune regulation. We developed cationic nanoliposomes (NLps) for the delivery of CD39 mRNA and evaluated them in a murine model of localized inflammation induced by lipopolysaccharide and matrigel. Biodistribution studies showed substantial retention within the Matrigel matrix, with limited systemic distribution at 24 h. CD39 mRNA-NLps significantly reduced cellular infiltration at day 5, with decreased monocyte and macrophage staining via histological analysis. qPCR confirmed sustained local CD39 mRNA, while flow cytometry demonstrated increased CD39 protein staining in matrigel-derived immune cells, and phosphate release assays showed functional ectonucleotidase activity at the inflammatory site. Interleukin (IL)-6 levels were slightly reduced in matrigel extracts following treatment, supporting suppression of local inflammation. Hemocompatibility was confirmed using in human blood and in treated mice, with liver enzymes and histological analysis supporting formulation's safety and favorable tolerability. These findings establish CD39 mRNA-NLps as a safe, effective strategy for a spatially confined mRNA-based approach to regulate local inflammation.