Patient-derived Stem Cells Research Articles

SFX-01 is therapeutic against myeloproliferative disorders caused by activating mutations in Shp2.

Activating mutations of Src homology-2 domain-containing protein tyrosine phosphatase-2 (Shp2) cause multiple childhood conditions for which there is an unmet therapeutic need, including juvenile myelomonocytic leukemia (JMML) and Noonan syndrome. SFX-01, an α-cyclodextrin-stabilized sulforaphane complex currently in clinical development, covalently adducts cysteine residues. Using unbiased proteomics, its protein targets were identified, including Shp2. SFX-01 induced an inhibitory dithiolethione modification at the Shp2 active site cysteine. Importantly, in a transgenic mouse model of human Noonan syndrome with hyperactive D61G Shp2, SFX-01 concomitantly normalized their phosphatase activity and myeloid cell count. Furthermore, SFX-01 also attenuated JMML human patient-derived hematopoietic stem cell proliferation that was linked to STAT1 signaling and decreased cyclin D1 expression, resulting in cell-cycle arrest. We conclude that SFX-01 is an activating mutant Shp2 inhibitor and may offer beneficial effects in patients with JMML or Noonan syndrome.

A novel discovery platform for targeted drug repurposing: application for psychiatric disorders.

A novel discovery platform for targeted drug repurposing: application for psychiatric disorders.

Recent advances on modeling retinal disease: Towards efficient gene/drug therapy.

Recent advances on modeling retinal disease: Towards efficient gene/drug therapy.

Preclinical evaluation of lentiviral gene therapy for adenosine deaminase 2 deficiency (DADA2): engraftment efficiency and biodistribution in humanised NBSGW mice.

Adenosine deaminase type 2 deficiency (DADA2) is caused by bi-allelic loss-of-function mutations in ADA2. While anti-TNF therapy is effective for the autoinflamatory and vasculitic components of the disease it does not correct marrow failure or immunodeficiency. Allogeneic stem cell transplantation (HSCT) offers a potential cure but is limited by challenges such as graft-versus-host-disease and donor availability. We previously demonstrated that lentiviral-mediated ADA2 gene therapy could restore ADA2 enzyme activity in patient-derived cells, correct macrophage inflammatory activation and reduce endothelial activation in vitro. Here, we evaluated the biodistribution and engraftment potential of lentivirally transduced healthy donor and patient-derived haematopoietic stem cells (HSC) in vivo using a humanised NBSGW mouse model. Transduced healthy HSC retained multilineage differentiation and engraftment capacity, without functional impairment. PCR analysis confirmed the absence of viral integration in non-haematopoietic organs, and histology showed no abnormal tissue changes, underscoring the safety and precision of this approach. In DADA2 patient-derived HSC, ADA2 transduction restored protein expression and enzyme activity, supporting improved cellular function and enhanced engraftment potential. These findings provide a strong foundation for advancing ADA2 gene therapy as a therapeutic strategy for DADA2, bringing it closer to clinical application.

Stem cell repair strategies for epilepsy.

Epilepsy is a serious neurological disorder; however, the effectiveness of current medications is often suboptimal. Recently, stem cell technology has demonstrated remarkable therapeutic potential in addressing various neurological diseases, igniting interest in its applicability for epilepsy treatment. This comprehensive review summarizes different therapeutic approaches utilizing various types of stem cells. Preclinical experiments have explored the use and potential therapeutic effects of mesenchymal stem cells, including genetically modified variants. Clinical trials involving patient-derived mesenchymal stem cells have shown promising results, with reductions in the frequency of epileptic seizures and improvements in neurological, cognitive, and motor functions reported. Another promising therapeutic strategy involves neural stem cells. These cells can be cultured outside the body and directed to differentiate into specific cell types. The transplant of neural stem cells has the potential to replace lost inhibitory interneurons, providing a novel treatment avenue for epilepsy. Embryonic stem cells are characterized by their significant capacity for self-renewal and their ability to differentiate into any type of somatic cell. In epilepsy treatment, embryonic stem cells can serve three primary functions: neuron regeneration, the maintenance of cellular homeostasis, and restorative activity. One notable strategy involves differentiating embryonic stem cells into γ-aminobutyric acidergic neurons for transplantation into lesion sites. This approach is currently undergoing clinical trials and could be a breakthrough in the treatment of refractory epilepsy. Induced pluripotent stem cells share the same genetic background as the donor, thereby reducing the risk of immune rejection and addressing ethical concerns. However, research on induced pluripotent stem cell therapy remains in the preclinical stage. Despite the promise of stem cell therapies for epilepsy, several limitations must be addressed. Safety concerns persist, including issues such as tumor formation, and the low survival rate of transplanted cells remains a significant challenge. Additionally, the high cost of these treatments may be prohibitive for some patients. In summary, stem cell therapy shows considerable promise in managing epilepsy, but further research is needed to overcome its existing limitations and enhance its clinical applicability.

Hippo pathway effectors are associated with glioma patient survival, control cell proliferation and sterol metabolism through TEAD3.

Glioblastomas represent the most common and lethal primary brain tumors in the world. Despite therapeutic advances during the last two decades, patient prognosis remains very poor. The Hippo signaling pathway effectors YAP/TAZ-TEADs play a crucial role in tumor progression and represent promising therapeutic targets in gliomas. In this study, we identified and investigated the clinical and biological significance of TEAD transcription factors. Through comprehensive analyses of TCGA glioma data and patient samples, we identified TEAD3-4 transcription factors as robust prognostic markers of patient outcome. Using up to five different patient-derived glioblastoma stem cell cultures, we confirmed the preferential expression and activation of TEAD3-4 along with their transcriptional coactivators YAP/TAZ. Pharmacological inhibition of YAP/TAZ-TEAD interaction by Verteporfin significantly decreased tumor cell growth, whereas specific inhibition of TEAD3 did not impact cell proliferation but affected sterol/cholesterol biosynthetic and metabolic processes. This study contributes to a better understanding of the role of Hippo effectors in glioblastoma pathophysiology. These transcription factors, particularly TEAD3, could potentially serve as therapeutic targets, especially considering recent data on cholesterol homeostasis in glioblastomas.

Combining prelamin A accumulation and oxidative stress: A strategy to target glioblastoma.

Combining prelamin A accumulation and oxidative stress: A strategy to target glioblastoma.

Durable HTT silencing using non-evolved dCas9 epigenome editors in patient-derived cells.

Durable HTT silencing using non-evolved dCas9 epigenome editors in patient-derived cells.

The Development and Characterisation of A Porcine Large Intestinal Biological Scaffold by Perfusion Decellularisation

The rising incidence of colorectal cancer and ulcerative colitis underscores an urgent need for regenerative solutions to address functional deficits after colectomy. However, the creation of clinically applicable large intestine scaffolds remains underdeveloped. Here, we report the successful generation and thorough characterisation of transplantable-sized porcine large intestinal scaffolds via perfusion decellularisation. This method effectively preserved extracellular matrix (ECM) structural and biochemical integrity while minimising immunogenicity through cellular component removal. Crucially, native vasculature remained intact, confirmed by histology, DNA quantification, and high-resolution CT angiography. Despite efficient decellularisation, challenges including residual nucleic acids, ECM heterogeneity, and partial microvascular occlusion were noted, echoing ongoing limitations in engineered, perfusable, full-thickness scaffolds. In vivo implantation demonstrated favourable biocompatibility and host integration; however, thrombosis occurred due to the lack of pre-seeded cells, emphasising the necessity of recellularisation for functional perfusion prior to implantation. This study addresses significant field limitations, presenting the first reproducible approach for structurally intact, perfusable, full-thickness large intestinal scaffolds of transplantable dimensions. Our innovations offer a strong foundation for future integration of patient-derived cells, stem cells, and organoids, progressing toward clinically viable, scalable, tissue-engineered large intestine constructs, from xenogeneic sources, relevant for regenerative medicine, disease modelling, and pharmacological screening.

Importin-alpha transports Norrin to the nucleus to promote proliferation and Notch signaling in glioblastoma stem cells.

Norrin, a secreted protein encoded by NDP gene, is recognized for its established role as a paracrine canonical Frizzled-4/Wnt ligand that mediates angiogenesis and barrier function in the brain. However, emerging evidence suggests that Norrin possesses Frizzled-4-independent functions, notably impacting Notch activation and proliferation of cancer stem cells. We conducted a BioID protein-proximity screen to identify Norrin-interacting proteins. Surprisingly, a significant proportion of the proteins we identified were nuclear. Through comprehensive tagging and proximity ligation assays, we demonstrate that Norrin is transported to the nucleus through KPNA2 (member of the Importin-alpha family). Subsequently, we demonstrate that KPNA2 loss of function in patient-derived primary glioblastoma stem cells results in a nuclear to cytoplasmic shift of Norrin distribution, and a complete abrogation of its function in stimulating Notch signaling and cellular proliferation. These results indicate that Norrin is actively transported into the nucleus to regulate vital signaling pathways and cellular functions.

Targeting the Cargo Receptor TMED9 as a Therapeutic Strategy Against Brain Tumors

Glioblastoma is one of the most aggressive and lethal forms of brain cancer, with limited therapeutic options and poor patient prognosis. Recent research has identified the TMED family of proteins as key regulators of tumor progression and aggressiveness across multiple cancer types. TMED members are cargo receptors expressed within the early secretory pathway and involved in bidirectional traffic of various proteins including EGFR, TGF-ɑ and WNT. In this study, we explored the therapeutic potential of genetic and pharmacologic inhibition of the cargo receptor TMED9 in glial tumor models. Our findings demonstrate that TMED9 expression is upregulated in glioma and that this upregulation is associated with poor patient survival. Using patient-derived glioma tumor cells, we demonstrate that TMED9 is highly expressed in the cancer stem cell population and that this upregulation promotes the cells’ self-renewal and migration. This is the first time, to the best of our knowledge, that TMED9 has been shown to play a major role in the function and tumorigenesis of brain tumor cancer stem cells. BRD4780, a small molecule that targets TMED9, effectively reduced TMED9 abundance, resulting in decreased viability, migration and stemness of patient-derived glioma stem cells. Moreover, BRD4780 mitigated the proliferation and migration of differentiated glioma tumor cells. When applied together with temozolomide, an established glioblastoma treatment, BRD4780 elicited an enhanced anti-tumor response. Lastly, to demonstrate the broad applicability of our findings, we targeted TMED9 in pediatric glioma cells and showed efficient inhibition of various oncogenic functions. Collectively, our study identifies TMED9 inhibition as a promising therapeutic approach that impairs the tumorigenesis and aggressiveness of brain tumors, with high efficacy against the tumor stem cell population. The effectiveness of TMED9 targeting in different tumor cell populations, the potential of combining this strategy with established therapies and the broad applicability of this approach to multiple cancer types highlight the significance of these findings.

Quantitative Proteomics and Phosphoproteomics Analysis of Patient-Derived Ovarian Cancer Stem Cells.

High-grade serous ovarian carcinoma (HGSOC) is the deadliest gynecologic cancer. Key to the progression and ultimate lethality of this subtype is the intra-tumoral heterogeneity, which is defined as the coexistence of different cell types and populations within a single tumor. Among those, ovarian cancer stem cells (OCSCs) are a distinct subpopulation of tumor cells endowed with stem-like properties, which can survive current standard therapies, resulting in tumor recurrence. Here, we generated ex vivo primary OCSC-enriched three-dimensional (3D) spheres from 10 distinct treatment naive patient-derived adherent (2D) cultures. We used state-of-the-art quantitative mass spectrometry to characterize the molecular events associated with OCSCs by analyzing their proteome and phosphoproteome. Our data revealed a stemness-related protein signature, shared within a heterogeneous patient cohort, which correlates with chemo-refractoriness in a clinical proteomics dataset. Moreover, we identified targetable deregulated kinases and aberrant PDGF receptor activation in OCSCs. Pharmacological inhibition of PDGFR in adherent OC cells reduced the stemness potential, measured by sphere formation assay. Overall, we provide a valuable resource to identify new OCSC markers and putative targets for OCSC-directed therapies.

Using a brain-like endothelial cell differentiation to characterize the CS79iBRCA-n2 BRCA1 mutated patient derived stem cell line.

BRCA1/2 genes are considered tumor suppressor genes and help repair damaged DNA. Pathogenic germline mutations of BRCA1/2 genes are the most common hereditary cause of breast cancer and ovarian cancer. It has been established that BRCA1 mutations increase the risk of brain metastasis compared to the BRCA1 wildtype, and once metastasis occurs to the brain the disease is considered uncurable. The blood-brain barrier (BBB) is essential for maintaining and regulating homeostasis of the central nervous system and is composed of highly specialized brain endothelial cells. Using a human induced pluripotent stem cell (hiPSC) based model, we characterized an hiPSC line from an invasive cancer patient harboring a BRCA1 mutation. This patient-derived hiPSC line can be utilized to study BBB properties as after differentiation into brain-like endothelial cells (BECs), BECs derived from this line express BBB markers such as tight junction proteins, and functional efflux transporters. Future application of patient-derived stem cell models could provide a platform to discover genetic predispositions to BBB disruption in individuals with BRCA1 mutations, as well as the potential molecular mechanisms contributing to brain metastasis.

Multiplexed phosphoproteomics of low cell numbers using SPARCE

Understanding cellular diversity and disease mechanisms requires a global analysis of proteins and their modifications. While next-generation sequencing has advanced our understanding of cellular heterogeneity, it fails to capture downstream signalling networks. Ultrasensitive mass spectrometry-based proteomics enables unbiased protein-level analysis of low cell numbers, down to single cells. However, phosphoproteomics remains limited to high-input samples due to sample losses and poor reaction efficiencies associated with processing low cell numbers. Isobaric stable isotope labelling is a promising approach for reproducible and accurate quantification of low abundant phosphopeptides. Here, we introduce SPARCE (Streamlined Phosphoproteomic Analysis of Rare CElls) for multiplexed phosphoproteomic analysis of low cell numbers. SPARCE integrates cell isolation, water-based lysis, on-tip TMT labelling, and phosphopeptide enrichment. SPARCE outperforms traditional methods by enhancing labelling efficiency and phosphoproteome coverage. To demonstrate the utility of SPARCE, we analysed four patient-derived glioblastoma stem cell lines, reliably quantifying phosphosite changes from 1000 FACS-sorted cells. This workflow expands the possibilities for signalling analysis of rare cell populations.

Drug Combinations Targeting FAK and MEK Overcomes Tumor Heterogeneity in Glioblastoma.

Background/Objectives: Glioblastoma (GBM) is an aggressive brain tumor with limited treatment options and poor prognosis, largely owing to its heterogeneity and the involvement of multiple intracellular signaling pathways that contribute to drug resistance. While recent advancements in targeted drug combination therapies, such as dabrafenib and trametinib, show promise for certain GBM subgroups, identifying effective drug combinations across the broader GBM population remains a challenge. Integrin-mediated signaling, particularly through Focal Adhesion Kinase (FAK), plays a pivotal role in GBM pathogenesis and invasion, making it a potential therapeutic target and component of future drug combination strategies. Methods: In this study, we utilized a chemogenomic screening approach to identify synergistic drug combinations that target FAK in glioblastoma. We initially employed a CRISPR-engineered GBM model to assess the effects of FAK depletion and subsequently discovered that combining FAK inhibitors such as VS4718 with MEK inhibitors, particularly trametinib, demonstrated synergistic effects. This potent combination was validated using various 2D and 3D assays, including cell viability/apoptosis assessment, synergistic analysis, cellular imaging, and target engagement assays. This combination also effectively inhibited spheroid growth and invasion across a diverse panel of patient-derived GBM stem cells. Molecular mechanisms underlying these effects include suppression of multiple kinase signaling pathways and enhanced apoptosis, elucidated using Reverse-Phase Protein Array (RPPA) profiling and Western blot validation. Result: In vivo, combination therapy significantly reduced the tumor volume in orthotopic transplantation models. Conclusions: These findings suggest that the combination of FAK and MEK inhibitors represents a promising therapeutic strategy to overcome the challenges of GBM treatment.

Abstract 6878: Ag5: a novel quantum cluster therapy targeting mitochondria and antioxidant systems in glioblastoma

Abstract Despite extensive efforts from drug developers, physicians, and patients, the prognosis for glioblastoma (GBM) remains one of the poorest in adult oncology. Current oncology drug development often revisits established technologies, with few truly novel treatments emerging to tackle challenging diseases like GBM. ARJUNA Therapeutics is breaking new ground with quantum cluster technology, creating a unique family of small molecules. The first of these, Ag5, consists of five silver atoms (a quantum cluster) with demonstrated drug-like qualities, including the ability to cross the blood-brain barrier. Ag5 penetrates cells and specifically catalyzes the irreversible inactivation of both glutathione and thioredoxin antioxidant systems in cancer cells, rapidly depleting mitochondrial glutathione and inducing cell death with minimal off-target effects at therapeutic doses. In collaboration with ARJUNA, the Laboratory for Precision Cancer Medicine at KU Leuven, Belgium, screened 20 patient-derived glioma stem cell lines from adult GBM tumors. Ag5 demonstrated cell-killing effectiveness at submicromolar concentrations in approximately 50% of these models, showing activity in both newly diagnosed and recurrent tumor models, as well as in cells from both the core and infiltrating regions. Importantly, tumor cell lines taken from individual patients at various disease stages retained their Ag5 sensitivity, even as they acquired resistance to radiotherapy and/or TMZ. Biomarkers defining sensitivity and resistance to Ag5 in patient-derived GBM lines have been identified, supported by promising preclinical in vivo data. Ag5 has the potential to benefit a significant proportion of GBM patients. A clinical phase 0 neuro-oncology trial at University Hospitals Leuven, Belgium, is planned to test both the biomarker and therapeutic hypothesis for Ag5 in adult GBM patients. Citation Format: Jayesh Telang, Marleen Derweduwe, Vanesa Porto, Blanca Dominguez, Ross Breckenridge, Martin Treder, Fernando Dominguez, Annelies Claeys, Isabel Aguirre Alcolea, Frederik De Smet. Ag5: a novel quantum cluster therapy targeting mitochondria and antioxidant systems in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6878.

Abstract 5614: Advancing treatment for brain tumors and metastases with brain permeable SHP2 inhibitor I-1000233

Background: SHP2, a non-receptor protein tyrosine phosphatase encoded by PTPN11, plays a pivotal role as a downstream effector of receptor tyrosine kinases (RTKs) in the MAPK signaling cascade. We report the discovery of I-1000233, a potent and selective SHP2 allosteric inhibitor that is orally bioavailable and penetrates the blood-brain barrier (BBB). This compound holds great potential for treating primary brain tumors and brain metastases associated with dysregulated RTK/RAS/ERK signaling pathways. Experimental Procedures: Through iterative optimization of the physicochemical and pharmacological properties of a lead compound series, we identified I-1000233 as a SHP2 inhibitor with excellent drug-like characteristics. Preclinical studies included in vitro assays to assess potency, in vivo pharmacokinetics to confirm BBB penetration, and proof-of-concept studies in orthotopic xenograft models of glioblastoma and brain metastases. Combination studies were conducted using other brain-penetrant agents targeting the MAPK pathway. Additionally, patient-derived glioblastoma stem cell (GSC) models were collected to refine a responder identification (ID) strategy for potential application in personalized medicine. Results: I-1000233 demonstrated potent in vitro inhibition of SHP2 activity (IC50 = 2 nM) with prolonged target residence time. In vivo, the compound exhibited strong anti-tumor activity in orthotopic glioblastoma and brain metastasis models, both as a monotherapy and in combination with other brain-permeable inhibitors, underscoring its versatility and potential for combination treatment regimens. Moreover, I-1000233 showed efficacy not only in targeting tumor cells bearing EGFR/RAS/MAPK mutations but also in modulating the tumor microenvironment (TME), enhancing the overall anti-tumor response. Further studies using patient-derived GSC models were conducted to identify biomarkers predictive of response, facilitating the development of precision medicine approaches for SHP2-targeted therapies. Conclusions: I-1000233 is a potent, selective, and orally available SHP2 inhibitor with demonstrated efficacy in preclinical models of primary brain tumors and metastases. Its ability to penetrate the BBB, modulate the tumor microenvironment, and provide combinatory potential with other agents highlights its promise as a therapeutic candidate for treating SHP2-responsive brain cancers. Citation Format: Francesca Puca, Alina Ciammaichella, Danilo Fabbrini, Martina Nibbio, Antonino Cucinotta, Giulio Auciello, Francesco Scalabrì, Christian Montalbetti, Alessia Petrocchi, Alessandro Carugo, Carlo Toniatti. Advancing treatment for brain tumors and metastases with brain permeable SHP2 inhibitor I-1000233 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5614.

A network-based approach to overcome BCR::ABL1-independent resistance in chronic myeloid leukemia

BackgroundAbout 40% of relapsed or non-responder tumors exhibit therapeutic resistance in the absence of a clear genetic cause, suggesting a pivotal role of intracellular communication. A deeper understanding of signaling pathways rewiring occurring in resistant cells is crucial to propose alternative effective strategies for cancer patients.MethodsTo achieve this goal, we developed a novel multi-step strategy, which integrates high sensitive mass spectrometry-based phosphoproteomics with network-based analysis. This strategy builds context-specific networks recapitulating the signaling rewiring upon drug treatment in therapy-resistant and sensitive cells.ResultsWe applied this strategy to elucidate the BCR::ABL1-independent mechanisms that drive relapse upon therapy discontinuation in chronic myeloid leukemia (CML) patients. We built a signaling map, detailing - from receptor to key phenotypes - the molecular mechanisms implicated in the control of proliferation, DNA damage response and inflammation of therapy-resistant cells. In-depth analysis of this map uncovered novel therapeutic vulnerabilities. Functional validation in patient-derived leukemic stem cells revealed a crucial role of acquired FLT3-dependency and its underlying molecular mechanism.ConclusionsIn conclusion, our study presents a novel generally applicable strategy and the reposition of FLT3, one of the most frequently mutated drivers of acute leukemia, as a potential therapeutic target for CML relapsed patients.

Generation and validation of a novel multitarget small molecule in glioblastoma

The development of multitarget small molecules (MSMs) has emerged as a powerful strategy for the treatment of multifactorial diseases such as cancer. Glioblastoma is the most prevalent and malignant primary brain tumor in adults, which is characterized by poor prognosis and a high heterogeneity. Current standards of treatment present limited effectiveness, as patients develop therapy resistance and recur. In this work, we synthesized and characterized a novel multi-target molecule (named DDI199 or contilistat), which is a polyfunctionalized indole derivative developed by juxtaposing selected pharmacophoric moieties of the parent compounds Contilisant and Vorinostat (SAHA) to act as multifunctional ligands that inhibit histone deacetylases (HDACs), monoamine oxidases (MAOs) and cholinesterases (ChEs), and modulate histamine H3 (H3R) and Sigma 1 Receptor (S1R) receptors. DDI199 exerts high cytotoxic activity in conventional glioblastoma cell lines and patient-derived glioma stem cells in vitro. Importantly, it significantly reduces tumor growth in vivo, both alone and in combination with temozolomide (TMZ). The comparison with SAHA showed higher target specificity and antitumor activity of the new molecule. Transcriptomic and proteomic analyses of patient-derived glioma stem cells revealed a deregulation in cell cycle, DNA remodeling and neurotransmission activity by the treatment with DDI199. In conclusion, our data reveal the efficacy of a novel MSM in glioblastoma pre-clinical setting.

Non-immune targeting of CXCR3 compromises mitochondrial function and suppresses tumor growth in glioblastoma

The chemokine receptor CXCR3 is traditionally recognized for its role in immune cell trafficking. However, emerging evidence suggests that its functions may extend beyond the immune system, particularly in cancer, where its roles remain to be elucidated. In this study, we demonstrated that CXCR3 expression correlates with glioblastoma (GBM) grading, with CXCR3-A isoform being associated with poorer patient prognosis compared to CXCR3-B. Ablation of both CXCR3 isoforms significantly impaired GBM cell proliferation, migration, and tumor growth both in vitro and in immunodeficient mice. To elucidate the mechanistic role of CXCR3, we conducted transcriptomic profiling of tumor xenografts, revealing that CXCR3 depletion would disrupt mitochondrial homeostasis. This was further supported by our findings that CXCR3 would localize to the mitochondrial membrane, and that inhibition of CXCR3 would lead to mitochondrial depolarization and increased reactive oxygen species production. Notably, activation of phosphorylated-STAT3 rescued cell viability in CXCR3-depleted cells, suggesting that CXCR3 may modulate mitochondrial function through a STAT3-dependent mechanism, consistent with the known functional role of STAT3 in maintaining mitochondrial redox balance. Furthermore, treatment with the selective CXCR3 antagonist AMG487 reduced tumor growth and disrupted mitochondrial function in vitro, in vivo, and in patient-derived GBM stem cells. Our findings reveal CXCR3 as a previously unrecognized regulator of mitochondrial function in cancer cells, positioning the CXCR3-mitochondrial signaling axis as a promising therapeutic target for GBM.Chemokine receptors are well-established mediators of inflammatory responses, emerging evidence suggests that these receptors may play roles beyond the immune system. In this study, we have demonstrated that CXCR3 would localize to the mitochondrial membrane and exert a previously unrecognized function in regulating cancer metabolism and mitochondrial function. Figure created using BioRender (https://biorender.com).