Mol Ther Nucleic Acids
· 2026 Jun · PMID 42100557
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Autologous cell therapies for knee osteoarthritis, including bone marrow aspirate concentrate (BMAC) and stromal vascular fraction (SVF), have advanced clinically despite an incomplete understanding of their mechanisms....Autologous cell therapies for knee osteoarthritis, including bone marrow aspirate concentrate (BMAC) and stromal vascular fraction (SVF), have advanced clinically despite an incomplete understanding of their mechanisms. Chatterjee et al. apply single-cell transcriptomics to multicenter trial of stem cell therapy for osteoarthritis (MILES) trial samples, revealing that site-to-site technical variability in BMAC composition overwhelms biological signals associated with treatment response. This finding mandates urgent standardization of harvest protocols before personalized genomic profiling can be meaningfully implemented. While responder/non-responder differences were subtle, pathway enrichment analyses and cell-cell communication inferences suggest that therapeutic outcomes may depend more on dynamic post-injection interactions than on baseline cellular states. The study establishes a foundational framework for evidence-based quality control in cellular orthopedics.
Vazaios K, Tallon-Cobos AC, van Oosterhout LPJ
… +11 more, Waranecki P, Cornel AM, Dautzenberg NMM, van Hoesel M, Forbes C, Nierkens S, Kemp V, Hoeben RC, van der Lugt J, Calkoen FG, Hulleman E
Mol Ther Oncol
· 2026 Jun · PMID 42100144
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Despite the success of CAR (chimeric antigen receptor) T cells in hematological malignancies, their effectiveness against solid or brain tumors, such as pediatric diffuse midline gliomas (DMGs), is limited. CAR T cell su...Despite the success of CAR (chimeric antigen receptor) T cells in hematological malignancies, their effectiveness against solid or brain tumors, such as pediatric diffuse midline gliomas (DMGs), is limited. CAR T cell success is hampered by factors including immunosuppression from DMGs and their surrounding tumor microenvironment (TME). Oncolytic viruses (OVs) can reverse this immunosuppression, suggesting a potential combination with CAR T cells. Here, we show that infection with Goravir adenovirus and R124 reovirus induced DMG cell lysis ( = 6 cultures), with minimal effect on the viability of B7H3- or GD2-targeted CAR T cells, even at high virus concentrations. In addition, RNA sequencing of infected tumor cells revealed altered gene expression in cell cycle and antiviral response pathways. Furthermore, co-cultures of CAR T cells with OV-infected DMGs enhanced CAR T-specific anti-tumor killing in 14 out of 24 cases. The successful combinations exhibited enhanced cytokine and chemokine release, coupled with an increased cytotoxic phenotype. These findings highlight the benefit of DMG pre-infection with OVs to boost CAR T cell activity and suggest that immune stimulation is a key driver of enhanced combination responses.
Man Z, Asano S, Kakutani T
… +3 more, Kiyoshi A, Hino K, Ikeda A
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42095135
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Werner syndrome (WS) is a rare autosomal recessive progeroid disorder caused by biallelic mutations in and is frequently complicated by refractory skin ulcers for which no effective therapy exists. We developed WRN-108,...Werner syndrome (WS) is a rare autosomal recessive progeroid disorder caused by biallelic mutations in and is frequently complicated by refractory skin ulcers for which no effective therapy exists. We developed WRN-108, a splice-switching antisense oligonucleotide (ASO) designed to induce exon 27 skipping and restore the open reading frame (ORF) disrupted by the most common mutation in Japanese WS patients, c.3139-1G>C, which leads to exon 26 skipping. In WS patient-derived fibroblasts, WRN-108 efficiently induced exon 27 skipping, restored WRN protein expression, and re-established its nuclear localization. Treatment improved cell proliferation and reduced senescence-associated markers, G-quadruplex accumulation, and γH2AX signaling, consistent with partial restoration of WRN-dependent genome maintenance functions. Topical administration in a rat skin wound model resulted in effective dermal penetration and sustained tissue retention. In a cynomolgus monkey wound model, WRN-108 induced exon 27 skipping in the skin and achieved dermal exposure without local toxicity. Short-term toxicity studies in mice and miniature pigs further confirmed its favorable tolerability. These findings provide preclinical evidence that ASO-mediated exon skipping can restore WRN function, highlighting the translational potential of WRN-108 as a therapeutic approach for refractory skin ulcers in WS patients harboring the c.3139-1G>C mutation.
Picht S, Farrera-Sal M, Hiller AL
… +14 more, Brumhard S, Klaas AM, Schulenberg S, Friedrich R, Scholz C, Hemmerling L, Simon DN, Krönke G, Pichon C, Sander LE, Volk HD, Gossen M, Schmueck-Henneresse M, Drzeniek NM
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42095134
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Chimeric antigen receptor (CAR) T cells offer a promising strategy for the treatment of autoimmune diseases. However, clinical translation is limited by the high cost, complexity, and poor scalability of current manufact...Chimeric antigen receptor (CAR) T cells offer a promising strategy for the treatment of autoimmune diseases. However, clinical translation is limited by the high cost, complexity, and poor scalability of current manufacturing, restricting broad patient access and persisting safety concerns including insertional mutagenesis risk from integrating vectors and uncontrolled long-term CAR T cell persistence. -transcribed (IVT) mRNA enables transient, non-integrating CAR expression with improved safety and scalability, making it particularly suited for non-malignant indications where prolonged persistence may not be required. Here, we systematically compare two IVT mRNA delivery platforms, electroporation and lipid nanoparticles (LNPs), for transient CAR T cell engineering in primary human T cells using single-cell transcriptomics and functional cell assays. We show that electroporation yields higher transfection efficiency and more sustained CAR surface expression, whereas LNP delivery reduces stress- and senescence-related transcriptional signatures as well as exhaustion marker expression, while enhancing antigen-driven activation, chemotactic responses, and cytotoxic function. Our comparative analysis highlights that the mode of mRNA delivery is associated with distinct transcriptional signatures and functional properties of CAR T cells, providing a framework to guide future development of mRNA-based approaches. These insights support LNP-mediated delivery as a functionally favorable strategy for transient CAR T cell engineering in autoimmune disease and beyond.
Choi MJ, So EY, Akosman B
… +7 more, Lee YE, Raufi AG, Reginato AM, Chen CC, Lawler SE, Wong ET, Liang OD
Mol Ther Oncol
· 2026 Jun · PMID 42094336
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Recent clinical trials show that CAR T cell therapies can initially blunt tumor growth in patients with glioblastoma (GBM). However, therapeutic efficacy remains limited by the immunosuppressive tumor microenvironment an...Recent clinical trials show that CAR T cell therapies can initially blunt tumor growth in patients with glioblastoma (GBM). However, therapeutic efficacy remains limited by the immunosuppressive tumor microenvironment and restricted immune cell trafficking across the blood-brain barrier (BBB). To counteract these challenges, we have utilized the oncolytic adenovirus (OV) Ad5-Δ24-RGD as a platform to overexpress a bispecific T cell engager (BiTE) targeting both CD3 on T cells and the GBM-specific tumor associated antigen IL-13Rα2. We first demonstrated that OV-BiTE can significantly increase the recruitment of T cells to GBM, both and . Moreover, OV-BiTE treatment also markedly enhanced CAR T cell infiltration and tumor cytotoxicity in a GBM-BBB spheroid model, possibly through downregulation of endothelial junction protein expression. We then showed that intratumoral injection of OV-BiTE followed by infusion of combined EGFR and EGFRvIII CAR T cells was more effective than OV-BiTE supplemented with either CAR T therapy alone and led to significant tumor reduction in a GBM xenograft mouse model. In conclusion, our multimodal OV-BiTE plus CAR T cell immunotherapy is capable of overcoming the immunosuppressive tumor microenvironment and GBM resistance to treatment.
Rhodiola rosea L. is a traditional Chinese medicine used clinically to treat pulmonary fibrosis. Due to the presence of multicomponents in Rhodiola rosea L., the mechanism of its anti-fibrotic effect remains unclear. Her...Rhodiola rosea L. is a traditional Chinese medicine used clinically to treat pulmonary fibrosis. Due to the presence of multicomponents in Rhodiola rosea L., the mechanism of its anti-fibrotic effect remains unclear. Here, we identified that herbacetin from Rhodiola rosea L. is an active molecule with anti-fibrotic effect in two mouse models of bleomycin-induced pulmonary fibrosis and unilateral ureteral obstruction-induced renal fibrosis. We also uncovered the molecular mechanism through which herbacetin exerted its anti-fibrotic effect by inhibiting transforming growth factor beta (TGF-β)/Smad3 signaling. We found that herbacetin could bind to the transforming growth factor beta receptor 2 (TGFBR2) and induced its degradation via the lysosomal pathway following K48-linked ubiquitination, thereby suppressing TGF-β/Smad3 signaling. Thus, herbacetin is a novel anti-fibrotic agent and exerts its anti-fibrotic effect on pulmonary and renal fibrosis by targeting TGFBR2 for degradation.
Novel therapies are urgently needed for breast cancer. While our previous work investigating the use of oncolytic viruses against the disease demonstrated its promise, heterogeneous responses were observed. Interestingly...Novel therapies are urgently needed for breast cancer. While our previous work investigating the use of oncolytic viruses against the disease demonstrated its promise, heterogeneous responses were observed. Interestingly, breast cancer is classified into subtypes based on the expression of hormone receptors. We noted that triple-negative breast cancers (TNBCs), which lack the expression of the estrogen (E2) receptor (ER) were resistant to the oncolytic vesicular stomatitis virus (oVSV), whereas ER cancers were particularly sensitive. We found that E2 stimulation enhanced virus infection, while E2-antagonizing endocrine therapies counteract the virus. Further mechanistic studies revealed a defect of nuclear factor (NF)-κB activation with E2 stimulation. With the goal of enhancing oncolytic virotherapy independently of E2, we designed a treatment strategy that combines oVSV with the NF-κB inhibitors IKK16 and sulfasalazine and found that both drugs improved virus replication in TNBC cell lines and patient samples. Our data uncover a novel impact of E2 on oncolytic virotherapy and demonstrate that combining virus treatment with NF-κB inhibitors recapitulates the E2-mediated virus enhancement. Importantly, our treatment combinations could be beneficial to all breast cancers, including TNBCs, as well as other breast cancers that do not respond to E2.
Lu F, Wu Z, Chi S
… +12 more, Li J, Ponek A, Liou C, Ma Q, Bonde D, Pavlaki N, Sweat ME, Shafaattalab S, Tharani Y, Prondzynski M, Bezzerides VJ, Pu WT
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited arrhythmia caused by pathogenic variants in RYR2 that increase its diastolic Ca release. Current standard of care is insufficient to protect pa...Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited arrhythmia caused by pathogenic variants in RYR2 that increase its diastolic Ca release. Current standard of care is insufficient to protect patients from ventricular tachycardia (VT) and sudden death. We identified RYR2 interaction with a subset of MYBPC3 in cardiomyocytes and validated this interaction by immunostaining, proximity ligation assay, and co-immunoprecipitation. Adeno-associated virus serotype 9 (AAV9)-mediated MYBPC3 overexpression reduced VT in Ryr2 mice, a model of CPVT. We identified a C-terminal fragment of MYBPC3, C6C10Δ, that is sufficient for RYR2 interaction and anti-arrhythmic activity. Recombinant C6C10Δ reduced the probability of open RYR2-R176Q channels in lipid bilayer single-channel recordings, and virally delivered C6C10Δ reduced VT inducibility and duration in the CPVT mouse model and in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) harboring several different CPVT-causing RYR2 variants. In wild-type mice, MyoAAV4E-C6C10Δ did not detectably affect cardiac morphology or systolic function. However, a proportion of MyoAAV4E-C6C10Δ-treated wild-type mice exhibited inducible VT, suggesting pro-arrhythmic potential that warrants further study. Together, these findings demonstrate that a subset of MYBPC3 interacts with RYR2 and that this interaction may be therapeutically exploited to develop CPVT gene therapy.
Chen Y, Jiang Z, Jin S
… +16 more, Liu M, Chen M, Kuo TC, Zhou K, Pu L, Chen M, Chen S, Li X, Chen AS, Xie J, Zhang H, Wang Q, Xu J, Sheng J, Huang Y, Chen G
Mol Ther Oncol
· 2026 Jun · PMID 42088619
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Glioblastoma(GBM), a highly aggressive primary brain tumor characterized by rapid progression, frequent recurrence, and limited clinical options, remains one of the most lethal central nervous system malignancies. Here,...Glioblastoma(GBM), a highly aggressive primary brain tumor characterized by rapid progression, frequent recurrence, and limited clinical options, remains one of the most lethal central nervous system malignancies. Here, we report a gene therapy strategy to treat glioma utilizing NeuroD1, a neurogenic transcription factor with demonstrated capacity to reprogram both glial cells and GBM cells into neuronal lineages. We developed a self-complementary adeno-associated virus (scAAV) vector, scAAV6-NeuroD1, and evaluated its therapeutic potential across and GBM models, including multiple GBM cell lines, patient-derived organoids, and orthotopic models in immunodeficient mice. Our findings reveal that scAAV6-NeuroD1 preferentially infects glioma cells and induces dual therapeutic effects by simultaneously inhibiting glioma cell proliferation and inducing neuronal reprogramming. Importantly, scAAV6-NeuroD1-treated mice with orthotopic GBM transplants exhibited reduced tumor burden, infiltration of T cells into the glioma, attenuated tumor-induced body weight loss, and dose-dependent survival extension. Analysis of published patient datasets further revealed that high NeuroD1 expression level correlates with improved overall survival and lower tumor malignancy grade. Together, these findings position scAAV6-NeuroD1 as a promising therapeutic candidate, potentially redefining the therapeutic landscape for GBM.
Wang J, Sun Y, Starzyk J
… +7 more, Wang F, Dong X, Shan R, He X, Xie K, Xie G, Wu H
Mol Ther Oncol
· 2026 Jun · PMID 42088618
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Natural killer (NK) cells hold promise for adoptive cell therapy due to their innate cytotoxicity. Early clinical trials confirm their safety and efficacy in cancer and autoimmune disease treatment. Engineering NK cells...Natural killer (NK) cells hold promise for adoptive cell therapy due to their innate cytotoxicity. Early clinical trials confirm their safety and efficacy in cancer and autoimmune disease treatment. Engineering NK cells with chimeric antigen receptors (CARs) enhances target specificity and facilitates their development as off-the-shelf allogeneic therapies. However, both viral and non-viral engineering methods of NK cells present challenges. Here, we introduce locus integrated CAR killer (CLICK), a novel non-viral approach using a mini-circular single-stranded DNA genome editing system. CLICK enables efficient integration of CD19CAR sequences into the metabolic checkpoint locus, simultaneously disrupting and driving stable, progressively increasing CAR expression in peripheral blood-derived NK cells. CLICK-engineered CAR-NK cells exhibit potent cytotoxicity, enhanced anti-tumor activity and , extended persistence, and reduced exhaustion. Together, these findings highlight CLICK as a highly efficient and versatile platform for non-viral CAR-NK cell engineering, offering a scalable approach for next-generation allogeneic immune cell therapies.
Elliot SJ, Grimaldo S, Catanuto P
… +7 more, Pereira-Simon S, Xia X, Civettini G, Shahzeidi S, Pastar I, Tomic-Canic M, Glassberg MK
Mol Ther Nucleic Acids
· 2026 Jun · PMID 42088451
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Mesenchymal stem cells (MSCs) deliver their effects via paracrine signaling, including the release of extracellular vesicles (EVs), which transfer microRNAs (miRNAs) and mRNAs to recipient cells. Aging alters exosome com...Mesenchymal stem cells (MSCs) deliver their effects via paracrine signaling, including the release of extracellular vesicles (EVs), which transfer microRNAs (miRNAs) and mRNAs to recipient cells. Aging alters exosome composition and function, raising questions about donor age, the use of autologous vs. allogeneic donors, and exosome dose and dosing frequency for exosome-based therapies in fibrotic lung disease. To address these gaps, we investigated how exosomes from younger (28-39 years, adult-exo) and older (58-66 years) donors influence fibrosis and tissue repair. Exosomes were delivered intravenously to 18-months-old male C57BL/6 mice on day 12 post-bleomycin (BLM), with lung injury evaluated on day 21 by histologic, molecular, mitochondrial, and telomere analyses. A human skin injury model was used to quantitate exosome-mediated wound healing. Adult-exo reduced lung injury and accelerated skin wound closure. RNA sequencing (RNA-seq) revealed that adult-exo carried elevated antifibrotic miRNAs, such as let-7. Pathway enrichment linked this cargo to extracellular matrix (ECM) remodeling, mitochondrial function, and senescence. Adult-exo treatment also downregulated fibrosis- and senescence-associated miRNAs, such as miR-34, in lung tissue. experiments confirmed alterations in multiple downstream pathways, mediating reparative effects. These findings underscore age-dependent differences in stem cell-derived exosomes and demonstrate that synergistic miRNA cargo drives maximal efficacy.