Biology subjects

Garg, A. D.

Publications and source records attributed to Garg, A. D..

6 recordsLinked to original sources

bioRxiv2025

Distinct tumor immune microenvironmental (TIME) landscapes drive divergent immunotherapy responses in glioblastoma

BackgroundImmunotherapies have improved outcomes in many cancers but show limited efficacy in glioblastoma (GBM). This study aimed to determine whether immunotherapy could be tailored to GBM by functional subtyping of vascular-immune landscapes. MethodsWe employed single-cell RNA sequencing, multiplex immunohistochemistry to characterize three distinct TIME subtypes in human and murine GBMs. We evaluated responses to combination of anti-angiogenic immunomodulating therapies (CD40 Ag, anti-PDL1, PI3K{gamma}/{delta} inhibition) in orthotopic syngeneic GBM mouse models. ResultsWe identified three distinct functional TIME subtypes with unique vascular-immune landscapes in human and murine GBM: TIME-low (immune-low/deserted, leaky vasculature), TIME-med (intermediate immune-infiltration, angiogenic), and TIME-high (heavily infiltrated with immunosuppressive myeloid cells and dysfunctional T cells). Representative mouse models of TIME-GBMs responded in subtype specific ways to anti-angiogenic immunomodulating therapies. TIME-low GBMs enhanced T-cell activity but relapsed due to emerging myeloid immunosuppression, concomitant with mesenchymal transition. TIME-med displayed the most immune-activated, yet angiogenic phenotype, and showed overall good responses to various anti-angiogenic immunomodulating therapies. TIME-high GBMs were mostly non-responsive but improved when the myeloid-cell PI3K{gamma} was targeted. However, CD40 agonist treatment, expected to enhance APC function, unexpectedly worsened survival by promoting angiogenesis and heightening immunosuppression, leading to dysfunctional T cells and reduced NK cell recruitment, and subsequent enhanced tumor propagation. ConclusionsOur study reveals three GBM TIME subtypes with distinct vascular-immune landscapes that require tailored therapies. TIME-med tumors are predicted to respond best to immunotherapies, TIME-low tumors show transient effects with anti-angiogenic immunomodulating therapies, while TIME-high tumors, due to their profound immunosuppression, can even have worse outcomes. Key pointsO_LIThree TIME subtypes were identified in GBM with distinct vascular-immune landscapes C_LIO_LITIME subtypes show divergent immunotherapy responses C_LIO_LITIME classification supports personalized treatment strategy for GBM immunotherapy C_LI Importance of the StudyThis study advances glioblastoma immunotherapy by providing the first comprehensive single-cell characterization of TIME subtypes, moving beyond bulk RNA-sequencing to reveal detailed functional states of immune cells. We establish clinically relevant murine models that recapitulate human TIME subtypes, enabling preclinical testing of TIME-targeted therapies. Our findings identify TIME-low GBM as immune deserted and TIME-med tumors as the most immunotherapy-responsive subtype that should be prioritized for clinical selection. We found that high immune infiltration correlates with non-responsiveness and even unexpected detrimental effects with CD40 agonist treatment in TIME-high tumors--critical information given ongoing clinical trials. Identifying distinct immunosuppressive mechanisms across TIME subtypes and differential treatment responses provides a framework for personalized immunotherapy selection. The immediate translational impact of this work highlights the importance of TIME classification for treatment stratification and the urgent need to consider TIME status in clinical trial design, potentially explaining variable patient responses in previous GBM immunotherapy trials.

cancer biology↗

bioRxiv2025

The molecular landscape of polycystic kidneys is marked by common alterations in purine metabolism

Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited kidney disease. Cysts develop through dedifferentiation of tubular epithelial cells, but the sequence of molecular events and their relative importance remain unclear. To address this gap in knowledge, 40 cysts from 4 ADPKD kidneys and 4 microcystic tissues were mapped on transcriptomic and histological level. Cyst were heterogenous and we identified 6 cystic subclusters with 2 deviations from the main trajectory, dependent on the rate of interstitial remodeling, inflammation and dedifferentiation. Loss of proximal tubular marker gene expression was more pronounced compared to those of other tubular segments. Altered expression of metabolic pathways was consistent among the cysts, which was further analyzed in human and mouse cell lines. Purine metabolism was similarly altered in all ADPKD cell lines, and its modulation with azathioprine suppressed cyst formation in vitro. In conclusion, by focusing on common altered pathways in cysts and cell models, we have identified purine metabolism as a novel potential target in ADPKD.

molecular biology↗

bioRxiv2023

A lymph node-to-tumour PD-L1+macrophage circuit antagonizes dendritic cell immunotherapy

Immune-checkpoint blockers (ICB) provide limited benefit against T cell-depleted tumours, calling for therapeutic innovation. Here, we aimed at designing a new type of dendritic cell (DC) vaccine by unbiased computational integration of multi-omics data from cancer patients. In a first attempt, a DC vaccine designed to present tumor antigens from cancer cells succumbing to immunogenic cancer cell death (ICD) and to elicit high type I interferon (IFN) responses failed to induce the regression of mouse tumors lacking T cell infiltrates. In lymph nodes (LNs), instead of activating CD4+ and CD8+T cells, DCs stimulated immunosuppressive PD-L1+LN-associated macrophages (LAMs) via a type I IFN response. Moreover, DC vaccines of this type stimulated pre-existing, T cell-suppressive, PD-L1+tumour-associated macrophages (TAMs). This created a T cell-suppressive circuit of PD-L1+macrophages, spanning across LNs and tumours. Accordingly, DC vaccines synergised with PD-L1 blockade to deplete PD-L1+macrophages, suppress myeloid inflammation affecting the tumor bed and draining lymph nodes, and de-inhibit effector/stem-like memory T cells, eventually causing tumour regression. The synergistic interaction between the DC vaccine and PD-L1 blockade was lost when DCs were manipulated to lose Ifnar1or Ccr7 or when macrophages were depleted. Interestingly, clinical DC vaccines also potentiated lymphocyte-suppressive PD-L1+TAMs in patients bearing T cell-depleted tumours. Altogether, our results reveal the existence of a novel PD-L1+LAM/TAM-driven immunosuppressive pathway that can be elicited by DC vaccines, yet can be subverted for improving the outcome of immunotherapy.

immunology↗

bioRxiv2021

Distinct cDC subsets co-operate in CD40 agonist response while suppressive microenvironments and lack of antigens subvert efficacy

Agonistic CD40 therapy has shown to inhibit cancer progression, but only in a fraction of patients. Hence, understanding the cancer cell-intrinsic and microenvironmental determinants of CD40 therapy response is crucial to identify responsive patient populations and design efficient combination treatments. Here, we showed that the therapeutic efficacy of CD40 in responder melanoma tumours, relied on pre-existing cDC1-primed CD8+ T cells, however cDC1s were dispensable after CD40 administration. Surprisingly, in response to CD40 the abundance of activated cDCs, potentially derived from cDC2s increased, thereby further activating antitumour CD8+ T cells. Hence, distinct cDC subsets are required to induce CD40 responses. By contrast, lung tumours, characterised by a high abundance of macrophages, were resistant to CD40 therapy. Combining CD40 therapy with macrophage depletion led to tumour growth inhibition only in the presence of strong neoantigens. Accordingly, treatment with immunogenic cell-death inducing chemotherapy sensitised non-immunogenic tumours to CD40 therapy.

cancer biology↗

bioRxiv2021

Immunogenomic, single-cell and spatial dissection of CD8+T cell exhaustion reveals critical determinants of cancer immunotherapy

Tumoural-CD8+T cells exhibit exhausted or dysfunctional states. Contrary to immunotherapy-responsive exhausted-CD8+T cells, the clinical features of dysfunctional-CD8+T cells are disputed. Hence, we conducted large-scale multi-omics and multi-dimensional mapping of CD8+T cell-states across multiple cancer patient-cohorts. This identified tumour-specific continuum of CD8+T cell-states across 6 human cancers, partly imprinted by organ-specific immuno-modulatory niches. Herein, melanoma and glioblastoma enriched prototypical exhausted (CD8+TEXT) and severely-dysfunctional (CD8+TSDF) states, respectively. Contrary to CD8+TEXT, CD8+TSDF displayed transcriptomic and epigenetic effector/cytolytic dysfunctions, and dysregulated effector/memory single-cell trajectories, culminating into maladaptive prodeath stress and cell-cycle defects. Suboptimal antigen-priming underscored CD8+TSDF, which was distinct from immune-checkpoints "rich" CD8+TEXT, reflecting chronic antigen-stimulation. Continuum variation also existed on tumour spatial-level, with convergent (CD8+TEXT-supportive vascular regions) and divergent features (dysfunctional CD4+T::CD8+TSDFcell-to-cell interactions) between melanoma and glioblastoma. Globally, IFN{gamma}-IL2 disparities, paucity of intra-tumoural CD4+/CD8+T cells, and myeloid TGF{beta}/wound healing responses, distinguished CD8+TSDF-landscape. Within immuno-oncology clinical-trials, anti-PD1 immunotherapy failed to "reinvigorate" CD8+TSDF-landscape, and instead facilitated effector-dysfunction and TGF{beta}/wound healing. However, cellular immunotherapies (dendritic cell-vaccines, adoptive T-cell therapy) ameliorated assorted CD8+TSDF-landscape disparities, highlighting a roadmap for anti-glioblastoma multimodal-immunotherapy. Collectively, our study comprehensively expands clinical-knowledge on CD8+T cell-exhaustion and suggests that tumour-specific, pre-existing CD8+TEXT/TSDF-states, determine immunotherapy-responses.

immunology↗

bioRxiv2020

The IRE1 Inhibitor KIRA6 Curtails The Inflammatory Trait Of Immunogenic Anticancer Treatments By Targeting HSP60 Independent Of IRE1

Cellular stress evoked by immunogenic anticancer treatments engaging the unfolded protein response (UPR) can elicit inflammation with conflicting therapeutic outcomes. To define cell-autonomous mechanisms coupling the UPR to molecular mediators of inflammation, we profiled the transcriptome of cancer cells responding to immunogenic or weakly immunogenic-treatments. Bioinformatics-driven pathway analysis indicated that immunogenic treatments instigated NF-{kappa}B/AP-1-inflammatory pathways, which were abolished by the IRE1-kinase inhibitor KIRA6. Cell-free fractions of chemotherapy and KIRA6 co-treated cancer cells were deprived of pro-inflammatory/chemoattractant factors and failed to mobilize innate immune cells. Strikingly, these potent KIRA6 anti-inflammatory effects were found to be independent of IRE1. Generation of a KIRA6-clickable photoaffinity probe, mass spectrometry and co-immunoprecipitation analysis identified cytosolic HSP60 as a KIRA6 off-target in the NF-{kappa}B pathway. In sum, our study unravels that inflammation evoked by immunogenic treatments is curtailed by KIRA6 independently of IRE1 and further suggests great caution in interpreting the anti-inflammatory action of IRE1 chemical inhibitors.

cancer biology↗