Boskovic, P. | Search bioRxiv

bioRxiv2026

A framework for reparative CAR T engineering in the CNS

Chimeric antigen receptor (CAR) T cells have shown remarkable therapeutic promise in hematological malignancies and, more recently, in autoimmunity. They also hold considerable potential for neurodegenerative diseases and CNS injury, where the therapeutic objective shifts from cell depletion to immune modulation and tissue repair. Using ischemic stroke as proof-of-concept, we engineered MOG-targeting CAR T cells to dissect how distinct CAR designs shape the CNS microenvironment. CD4/CD8 CAR T cells (a mixture of CD4 and CD8 subsets) proliferated robustly and efficiently infiltrated the ischemic hemisphere, but induced broad immune recruitment and exacerbated neuroinflammation. In contrast, CD4 restricted CAR T cells markedly reduced immune infiltration, reprogrammed microglia, and minimized inflammatory activation. We engineered CD4 CAR T cells to secrete brain-derived neurotrophic factor (BDNF) to determine whether they could be redirected toward a reparative, non-cytotoxic phenotype. CD4 BDNF-CAR T cells further attenuated inflammation, reduced immune infiltration, and promoted the expansion of regulatory T cells. CD4 BDNF-CAR T treated mice showed significantly improved gait performance following stroke. Together, these findings establish a cellular framework and outline principles for engineering reparative CAR T platforms for neurological diseases.

immunology↗

bioRxiv2023

BCAT1 regulates glioblastoma cell plasticity and contributes to immunosuppression

Glioblastoma is the most common malignant brain tumor in adults. Cellular plasticity and the poorly differentiated features result in a fast relapse of the tumors following treatment. Moreover, the immunosuppressive microenvironment proved to be a major obstacle to immunotherapeutic approaches. Branched-chain amino acid transaminase 1 (BCAT1) is a metabolic enzyme that converts branched-chain amino acids into branched-chain keto acids, depleting cellular -ketoglutarate and producing glutamate. BCAT1 is expressed in and drives the growth of glioblastoma and other cancers. Here we show that low-BCAT1 expression correlates with differentiated glioblastoma subtypes and its knockout (KO) results in a differentiated phenotype in human and mouse glioblastoma cells. Consistent with these observations, Bcat1-KO mouse glioblastoma cells were highly susceptible to serum-induced differentiation in vitro. The transition to a differentiated cell state was linked to the increased activity of TET demethylases and the hypomethylation and activation of neuronal differentiation genes. Orthotopic tumor injection into immunocompetent mice demonstrated that the brain microenvironment is sufficient to induce differentiation of Bcat1-KO tumors in vivo. In addition, the knockout of Bcat1 attenuated immunosuppression, allowing for an extensive infiltration of CD8+ cytotoxic T-cells and complete abrogation of tumor growth. Additional analysis in immunodeficient hosts revealed that both Bcat1-KO-induced differentiation and immunomodulation contribute to the long-term suppression of tumor growth. In summary, our study demonstrates that BCAT1 promotes glioblastoma growth by blocking tumor cell differentiation and sustaining an immunosuppressive microenvironment. These findings suggest novel modes limiting glioblastoma phenotypic plasticity and therapeutic failure through targeting BCAT1. Importance of the studyHigh expression of BCAT1 occurs in many tumor entities and is related to aggressiveness, proliferation and invasion of tumor cells. In this study, we show that its expression is crucial for the continuous growth of glioblastoma cells by preventing their differentiation. Furthermore, we show that the expression of BCAT1 modulates the tumor immune microenvironment, suppressing the CD8 T-cell response. BCAT1 knockout causes glioblastoma cell differentiation and a persistent CD8 T-cell response, which is sufficient to abrogate tumor growth and prolong survival in in vivo immunocompetent and immunodeficient models, respectively. Our findings consolidate BCAT1 as a major player in glioblastoma and highlight its importance as a potential future target of research in this and other tumor entities. Key PointsO_LIBCAT1 expression maintains poorly differentiated features of glioblastoma cells and provides resistance to differentiation. C_LIO_LIExpression of BCAT1 in glioblastoma cells contributes to the immunosuppressive features of the tumor. C_LI

cancer biology↗

bioRxiv2020

A novel neural stem cell-derived immunocompetent mouse model of glioblastoma for preclinical studies

Glioblastomas are the most lethal tumors affecting the central nervous system in adults. Simple and inexpensive syngeneic in vivo models that closely mirror human glioblastoma, including interactions between tumor and immune cells, are urgently needed for deciphering glioma biology and developing more effective treatments. Here, we generated glioblastoma cell lines by repeated in-vivo passaging of cells isolated from a neural stem cell-specific Pten/p53 double-knockout genetic mouse brain tumor model. Transcriptome and genome analyses of the cell lines revealed molecular heterogeneity comparable to that observed in human glioblastoma. Upon orthotopic transplantation into syngeneic hosts they formed high-grade gliomas that faithfully recapitulated the histopathological features, invasiveness and myeloid cell infiltration characteristic of human glioblastoma. These features make our cell lines unique and useful tools to study multiple aspects of glioblastoma pathomechanism and test novel treatments, especially immunotherapies in syngeneic preclinical models.

cancer biology↗