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Radiotherapy is a standard treatment of pediatric brain tumors. Though the survival rate has improved for many tumor types, most patients suffer long-term cognitive decline and there is currently no way of preventing radiation-induced damage to healthy brain tissue. Here, we used a human forebrain organoid model to investigate if the α2-adrenoceptor and I1-imidazoline receptor agonist clonidine could prevent radiotoxicity. We found that treatment of organoids with clonidine significantly reduced radiation-induced loss of neural progenitor cells, neurons, astrocytes, and oligodendrocyte lineage cells. Moreover, clonidine reduced overall DNA damage and signs of reactive gliosis in organoids. Our findings demonstrate that pharmacological rescue of radiation neurotoxicity is possible in a human brain organoid model and provides a rationale for future drug repurposing studies aiming to prevent radiation-induced brain injury in children treated with radiotherapy.

Chronic compressive cervical spinal cord injury (cCSCI), a debilitating condition, lacks effective treatment options. Addressing this gap, our study introduces a novel rat model of cCSCI developed through spinal cord compression via synthetic polyether sheet implantation, closely mimicking human pathology. We evaluated the model’s fidelity utilizing a comprehensive series of behavioral, electrophysiological, and histological assessments. Our research also explored the therapeutic potential of oligodendrogenic neural progenitor cells (oNPCs) derived from induced pluripotent stem cells. Transplanted oNPCs successfully integrated into the host spinal cord, differentiated into neurons, astrocytes, and oligodendrocytes, and demonstrated a remarkable capacity for enhancing neuroplasticity. Electrophysiological analyses revealed significant improvements in motor evoked potentials and a rectification of the excitability imbalance posttransplantation, indicating substantial recovery of motor circuits. Histological findings complemented these results, showing enhanced remyelination and a reduction in excitatory transmitter expression in the residual gray matter. Functionally, the transplantation of oNPCs led to marked improvements in grip strength, locomotor abilities, and sensory functions, surpassing those seen with standard treatments. This study not only provides a novel and reliable rat model of cCSCI for further research but also highlights the potential of oNPCs as a transformative approach for spinal cord injury therapy, suggesting their significant role in neural regeneration and repair.

Acute myeloid leukemia (AML) is a genetically heterogeneous malignancy characterized by the clonal proliferation of undifferentiated myeloid precursors in the bone marrow. Although standard induction regimens based on anthracyclines often achieve initial remission, up to 25% of patients exhibit primary refractory disease and nearly 50% relapse, underscoring the urgent need to overcome therapy resistance. Aldehyde dehydrogenase 1 (ALDH1) contributes to leukemic cell survival by maintaining stemness, proliferation, and chemoresistance through aldehyde detoxification and retinoic acid synthesis. Here, we identify two enhancer elements, ALDH1A1‐E3 and ALDH1A2‐E1‐A, that mediate transcriptional activation of ALDH1A1 and ALDH1A2 in response to the anthracycline daunorubicin. These enhancers are regulated by STAT3 and FOS/JUN transcription factors, which cooperatively link drug response to ALDH1 induction. Functional validation in AML cell lines, primary samples, and xenograft models shows that ALDH1 upregulation is part of an adaptive stress response and may contribute to reduced anthracycline sensitivity. Co‐treatment with the ALDH1A1/1A2 inhibitor DIMATE synergistically enhances daunorubicin efficacy across in vitro and in vivo resistant models. Consistently, high ALDH1 expression is associated with adverse genetic risk, prior anthracycline exposure, and inferior OS, particularly in relapsed/refractory AML. These findings uncover a novel enhancer‐mediated mechanism of ALDH1 induction in the context of anthracycline exposure and support the rationale for future clinical trials combining standard treatments with ALDH1‐targeted approaches, including the clinical‐stage inhibitor DIMATE.

Management of chronic kidney disease (CKD) remains a major challenge due limited therapeutic options to reverse fibrosis, which is a critical feature in CKD. Partial epithelial-to-mesenchymal transition (EMT) of tubular epithelial cells (TECs) is a key driver of fibrosis, and has become an important focus for kidney protection strategies. Blood platelets, a major source of circulating transforming growth factor beta (TGF-β), are implicated in pathogenesis of CKD, but their involvement in EMT and kidney fibrosis remains uncertain. Methods: We used two mouse models of renal fibrosis—diabetic kidney disease (DKD) and unilateral ureter obstruction (UUO)—to examine the connection between platelets, partial EMT, and fibrosis. Platelet inhibition or depletion was performed to assess EMT, cell cycle arrest, and fibrosis. In vitro, platelets were applied to TECs and kidney organoids. To determine the role of TGF-β signaling, we used TGF-βRI inhibitor. Expression of EMT, and fibrosis markers, as well as TGF-β1 signaling, were analyzed using western blot, reverse transcription quantitative PCR (RT-qPCR), enzyme-linked immunosorbent assay (ELISA), and immunostaining. Results: In both animal models, platelet inhibition or depletion resulted in reduced expression of cell cycle arrest marker p21, partial EMT and fibrosis. In vitro, activated platelets stimulated cell cycle arrest, EMT, and fibrosis in TECs and kidney organoids. Chronically injured TECs experience cell-cycle arrest which promote a paracrine EMT program in TECs, jointly leading to fibrosis. This platelet-mediated effect on cell cycle arrest and EMT was driven by TGF-β1 signaling, as selective inhibition of the TGF-β receptor rescued these dysfunctional phenotypes. Conclusions: Our study demonstrates that platelets activate the TGF-β1 pathway, leading to cell cycle arrest, EMT and renal fibrosis. These findings suggest that antiplatelet therapies may have potential renoprotective effects by protecting tubular homeostasis, attenuating partial EMT and fibrosis.

Bipolar disorder (BD) is a complex psychiatric condition usually requiring long-term treatment. Lithium (Li) remains the most effective mood stabilizer for BD, yet it benefits only a subset of patients, and its precise mechanism of action remains elusive. Exome sequencing has identified AKAP11 (A-kinase anchoring protein 11) as a shared risk gene for BD and schizophrenia (SCZ). Given that both the AKAP11-Protein Kinase A (PKA) complex and Li target and inhibit Glycogen Synthase Kinase-3 beta (GSK3β), we hypothesize that Li may partially normalize the transcriptomic and/or epigenomic alterations observed in heterozygous AKAP11-knockout (Het-AKAP11-KO) iPSC-derived neurons. In this study, we employed genome-wide approaches to assess the effects of Li on the transcriptome and epigenome of human iPSC-derived Het-AKAP11-KO neuronal culture. We show that chronic Li treatment in this cellular model upregulates key pathways that were initially downregulated by Het-AKAP11-KO, several of which have also been reported as downregulated in synapses of BD and SCZ post-mortem brain tissues. Moreover, we demonstrated that Li treatment partially rescues certain transcriptomic alterations resulting from Het-AKAP11-KO, bringing them closer to the WT state. We suggest two possible mechanisms underlying these transcriptomic effects: (1) Li modulates histone H3K27ac levels at intergenic and intronic enhancers, influencing enhancer activity and transcription factor binding, and (2) Li enhances GSK3β serine 9 phosphorylation, impacting WNT/β-catenin signaling and downstream transcription. These findings underscore Li’s potential as a therapeutic agent for BD and SCZ patients carrying AKAP11 loss-of-function variants or exhibiting similar pathway alterations to those observed in Het-AKAP11-KO models.

The lack of understanding of the molecular and cellular characteristics of human retinal progenitor cells (RPCs) has hindered their application in cell therapy for retinal degenerative diseases. This study aims to employ retinal organoids (ROs) derived from a VSX2-enhanced green fluorescent protein (eGFP) reporter human induced pluripotent stem cell (hiPSC) line for positive selection of human RPCs, investigate their features, and facilitate their applications. Methods: hiPSCs were differentiated into three-dimensional ROs following established protocols. The fidelity of the VSX2-eGFP reporter was confirmed through immunostaining. Fluorescence-activated cell sorting was employed to select VSX2-eGFP-positive (+) cells at distinct developmental stages, followed by bulk RNA sequencing (RNA-seq) analysis to assess their transcriptome profile. Immunostaining and flow cytometry were utilized to validate the identity of VSX2-eGFP+ cells and potential cluster of differentiation (CD) biomarkers for identifying human RPCs. Results: hiPSCs were successfully differentiated into ROs containing abundant RPCs. The spatiotemporal activity of the VSX2-eGFP reporter recapitulated the dynamic expression of endogenous VSX2 protein. Compared to VSX2-eGFP-negative (-) cells, VSX2-eGFP+ cells mainly exhibited characteristics of RPCs at early stages of retinal development and of bipolar cells at late stages. RNA-seq analysis revealed transcriptional heterogeneity within VSX2-eGFP+ cells across four distinct developmental stages. Moreover, the dynamic expression of 394 known CD biomarkers in VSX2-eGFP+ cells at distinct developmental stages was analyzed herein for the first time. One CD biomarker, TNFRSF1B, which has never been reported to be expressed in RPCs, was found to be highly expressed in RPCs at the early stages and might serve as a candidate CD biomarker for sorting RPCs. Conclusions: This study provides valuable insights into the molecular and cellular characteristics of human RPCs, especially their expression profiles of CD biomarkers, laying a foundation for research on retinal development and the clinical translation of hiPSC-derived RPCs.

Prader-Willi syndrome (PWS) is a genomic imprinting disorder caused by the loss of function of the paternal chromosome 15q11-13, resulting in a spectrum of symptoms associated with hypothalamic dysfunction. PWS patients lack the expression of paternally expressed genes (PEGs) in the 15q11-13 locus but possess an epigenetically silenced set of these genes in the maternal allele. Thus, activation of these silenced genes can serve as a therapeutic target for PWS. Here, we leverage CRISPR-based epigenome editing system to modulate the DNA methylation status of the PWS imprinting control region (PWS-ICR) in induced pluripotent stem cells (iPSCs) derived from PWS patients. Successful demethylation in the PWS-ICR restores the PEG expression from the maternal allele and reorganizes the methylation patterns in other PWS-associated imprinted regions beyond the PWS-ICR. Remarkably, these corrected epigenomic patterns and PEG expression are maintained following the differentiation of these cells into hypothalamic organoids. Finally, the single-cell transcriptomic analysis of epigenome-edited organoids demonstrates a partial restoration of the transcriptomic dysregulation observed in PWS. This study highlights the utility of epigenome editing technology as a therapeutic approach in addressing PWS and potentially other imprinting disorders. The authors develop CRISPR-based epigenome editing strategy to reactivate silenced maternally inherited genes for Prader-Willi syndrome in human iPSC and hypothalamic organoid models, highlighting its potential for treating imprinting disorders.

Bispecific T cell-engagers (BTEs) are engineered antibodies that redirect T cells to target antigen-expressing tumors. BTEs targeting tumor-specific antigens such as interleukin 13 receptor alpha 2 (IL13RA2) and epidermal growth factor receptor variant III (EGFRvIII) have been developed for glioblastoma (GBM). However, there is limited mechanistic understanding of the action of BTE since prior studies were mostly conducted in immunocompromised animal models. To close this gap, the function of BTEs was assessed in the immunosuppressive tumor microenvironment (TME) of orthotopic and genetically engineered mouse models (GEMM) with intact immune systems. Method: sA BTE that bridges CD3 epsilon on murine T cells to IL13RA2-positive GBM cells was developed, and the therapeutic mechanism was investigated in immunocompetent mouse models of GBM. Multicolor flow cytometry, single-cell RNA sequencing (scRNA-seq), multiplex immunofluorescence, and multiparametric MRI across multiple preclinical models of GBM were used to evaluate the mechanism of action and response. Results: BTE-mediated interactions between murine T cells and GBM cells triggered T cell activation and antigen-dependent killing of GBM cells. BTE treatment significantly extended the survival of mice bearing IL13RA2-expressing orthotopic glioma and de novo forming GBM in the GEMM. Quantified parametric MRI validated the survival data, showing a reduction in glioma volume and decreased glioma viability. Flow cytometric and scRNA-seq analyses of the TME revealed robust increases in activated and memory T cells and decreases in immunosuppressive myeloid cells within the brains of mice following BTE treatment. Conclusions: Our data demonstrate that the survival benefits of BTEs in preclinical models of glioma are due to the ability to engage the host immune system in direct killing, induction of immunological memory, and modulation of the TME. These findings provide a deeper insight into the mechanism of BTE actions in GBM.