Biology subjects

Walter, V.

Publications and source records attributed to Walter, V..

10 recordsLinked to original sources

bioRxiv2026

High immune receptor clonality in melanoma-draining lymph nodes associates with immune dysfunction and poor survival

Tumor-draining lymph nodes (tdLNs) are critical hubs of anti-tumor immunity but are also vulnerable to tumor-mediated immunosuppression. We analyzed T cell and B cell receptor (TCR/BCR) repertoires and transcriptomes from sentinel and non-sentinel lymph nodes of patients with melanoma from a historical pre-immune checkpoint inhibitor cohort (1994-2002) and an independent contemporary validation cohort (2022-2024). Melanoma-positive lymph nodes exhibited increased immune receptor clonality compared with tumor-free nodes. While average clonality showed no consistent association with outcome, the presence of extreme high-clonality outliers in individual lymph nodes was strongly associated with poor melanoma-specific survival. These outliers were characterized by a loss of lymphocyte-related genes and activation markers, an enrichment of melanocytic transcripts, and the suppression of immune signaling pathways, consistent with local immune dysfunction. Increased clonality was confined to lymph nodes and not observed in the peripheral blood. T cell responses to melanocyte differentiation antigens were infre-quently shared between lymph nodes and peripheral blood, highlighting immune compartmentalization.

immunology↗

bioRxiv2025

BNIP3-mTOR Signaling Mediates Resistance to MET Inhibition in Glioblastoma

Glioblastoma (GBM) is an aggressive primary brain malignancy with poor prognosis due to rapid progression, extensive invasiveness, and intrinsic resistance to standard therapies. Aberrant activation of receptor tyrosine kinases (RTKs), particularly MET, drives tumor proliferation, invasion, and therapy resistance. Here, we show that MET inhibition with crizotinib induces senescence and mitochondrial dysfunction in glioma-initiating cells (GICs), in part via downregulation of the mitochondrial protein BNIP3. However, BNIP3 downregulation activates mTOR signaling, enabling adaptive resistance. Targeting mTOR with everolimus in combination with crizotinib synergistically enhances anti-tumor effects, inducing apoptosis, senescence, and necroptosis, and significantly reducing cell viability and sphere-forming capacity. In orthotopic GBM xenograft models, this combination, particularly in a sequential regimen, markedly prolongs survival without overt toxicity. Our findings identify a BNIP3-mTOR signaling axis as a critical mediator of resistance to MET inhibition and provide a mechanistic rationale for combined MET and mTOR targeting as a promising therapeutic strategy in GBM. Statement of Translational RelevanceGlioblastoma (GBM) remains a highly aggressive and treatment-resistant brain tumor with limited therapeutic options. Our study identifies a BNIP3-mTOR signaling axis as a key mediator of resistance to MET inhibition. We show for the first time that combined MET and mTOR inhibition exhibits synergistic effects against GBM in vitro and in vivo. This combination prolongs survival without overt toxicity, providing a strong preclinical rationale for clinical evaluation in GBM patients with high MET expression and offering a promising strategy to overcome adaptive resistance.

cancer biology↗

bioRxiv2025

PD-1 regulates CD4+ T cell-mediated CD8+ T cell responses in the brain to balance viral control and neuroinflammation

Programmed cell death protein 1 (PD-1) is expressed by T cells during progressive multifocal leukoencephalopathy (PML), a life-threatening brain disease caused by the human-only JC polyomavirus. Why PD-1 blockade finds variable success in PML patients is unclear. Brain CD4+ and CD8+ T cells are PD-1high during mouse polyomavirus (MuPyV) encephalitis. Here, we show that PD-1 loss during MuPyV infection acts in a brain-autonomous manner to increase the magnitude of brain-infiltrating CD4+ and CD8+ T cells and the function of virus-specific CD8+ T cells; in concert, brain virus levels decline and neuroinflammation increases. Deletion of PD-1 in CD4+ T cells, but not CD8+ T cells, recapitulates effects of global PD-1 loss. Single-cell RNA sequencing shows that PD-1-deficient CD8+ T cells cluster as effectors while transcripts associated with proliferation and function are upregulated with loss of PD-1. Thus, CD4+ T cell-intrinsic PD-1 signaling balances antiviral defense against neural injury during polyomavirus CNS infection.

immunology↗

bioRxiv2023

Loss of STIM2 in colorectal cancer drives growth and metastasis through metabolic reprogramming and PERK-ATF4 endoplasmic reticulum stress pathway

The endoplasmic reticulum (ER) stores large amounts of calcium (Ca2+), and the controlled release of ER Ca2+ regulates a myriad of cellular functions. Although altered ER Ca2+ homeostasis is known to induce ER stress, the mechanisms by which ER Ca2+ imbalance activate ER stress pathways are poorly understood. Stromal-interacting molecules STIM1 and STIM2 are two structurally homologous ER-resident Ca2+ sensors that synergistically regulate Ca2+ influx into the cytosol through Orai Ca2+ channels for subsequent signaling to transcription and ER Ca2+ refilling. Here, we demonstrate that reduced STIM2, but not STIM1, in colorectal cancer (CRC) is associated with poor patient prognosis. Loss of STIM2 causes SERCA2-dependent increase in ER Ca2+, increased protein translation and transcriptional and metabolic rewiring supporting increased tumor size, invasion, and metastasis. Mechanistically, STIM2 loss activates cMyc and the PERK/ATF4 branch of ER stress in an Orai-independent manner. Therefore, STIM2 and PERK/ATF4 could be exploited for prognosis or in targeted therapies to inhibit CRC tumor growth and metastasis. HighlightsO_LISTIM2 regulates ER Ca2+ homeostasis independently of Orai and SOCE. C_LIO_LISTIM2 downregulation in colorectal cancer cells causes enhanced ER Ca2+ and is associated with poor patient prognosis. C_LIO_LISTIM2 downregulation induces PERK/ATF4 dependent ER stress in colorectal cancer. C_LIO_LIIncreased ER stress drives colorectal cancer metabolic reprogramming, growth, and metastasis. C_LI

cancer biology↗

bioRxiv2023

Alternate splice variants of the mitochondrial fission protein DNM1L/Drp1 regulate mitochondrial dynamics and cell fate in ovarian cancer.

Aberrant mitochondrial fission/fusion dynamics have been reported in cancer cells. While post translational modifications are known regulators of the mitochondrial fission/fusion machinery, we show that alternative splice variants of the fission protein Drp1 (DNM1L) have specific and unique roles in cancer, adding to the complexity of mitochondrial fission/fusion regulation in tumor cells. Ovarian cancer specimens express an alternative splice transcript variant of Drp1 lacking exon 16 of the variable domain, and high expression of this splice variant relative to other transcripts is associated with poor patient outcome. Unlike the full-length variant, expression of Drp1 lacking exon 16 leads to decreased association of Drp1 to mitochondrial fission sites, more fused mitochondrial networks, enhanced respiration, and TCA cycle metabolites, and is associated with a more metastatic phenotype in vitro and in vivo. These pro-tumorigenic effects can also be inhibited by specific siRNA-mediated inhibition of the endogenously expressed transcript lacking exon 16. Moreover, lack of exon 16 abrogates mitochondrial fission in response to pro-apoptotic stimuli and leads to decreased sensitivity to chemotherapeutics. These data emphasize the significance of the pathophysiological consequences of Drp1 alternative splicing and divergent functions of Drp1 splice variants, and strongly warrant consideration of Drp1 splicing in future studies.

cancer biology↗

bioRxiv2023

Urothelium-specific expression of mutationally activated Pik3ca initiates early lesions of non-invasive bladder cancer

Despite the fact that [~]70% of bladder cancers are non-invasive and have high recurrence rates, early stage disease is understudied. The relative lack of models to validate the contribution of molecular drivers of bladder tumorigenesis is a significant issue. While mutations in PIK3CA are frequent in human bladder cancer, an in vivo model for understanding their contribution to bladder tumorigenesis is unavailable. Therefore, a Upk2-Cre/Pik3caH1047R mouse model expressing one or two R26-Pik3caH1047R alleles in a urothelium-specific manner was created. Pik3caH1047R functionality was confirmed by quantifying Akt phosphorylation and mice were characterized by assessing urothelial thickness, nuclear atypia, and expression of luminal and basal markers at 6 and 12 months of age. At 6 months, Pik3caH1047R mice developed increased urothelial thickness and nuclear atypia, however, at 12 months, Pik3caH1047R mice did not exhibit progressive disease. Immunohistochemistry shows urothelium maintained luminal differentiation characterized by high Foxa1 and Ppar{gamma} expression. In addition, mice were subjected to low-dose carcinogen exposure (N-Butyl-N-(4-hydroxybutyl)nitrosamine (BBN)). Surprisingly, Pik3caH1047R mice exhibited no significant differences after exposure relative to mice without exposure. Furthermore, ssGSEA analysis of invasive human tumors showed those with mutant PIK3CA do not exhibit significantly increased PI3K/AKT pathway activity compared to wildtype PIK3CA tumors. Overall, these data suggest that Pik3caH1047R can elicit early tumorigenic changes in the urothelium, but progression to invasion may require additional genetic alterations.

cancer biology↗

bioRxiv2022

Orai3 and Orai1 are essential for CRAC channel function and metabolic reprogramming in B cells

The essential role of store-operated Ca2+ entry (SOCE) through Ca2+ release-activated Ca2+ (CRAC) channels in T cells is well established. In contrast, the contribution of individual Orai isoforms to SOCE and their downstream signaling functions in B cells are poorly understood. Here, we demonstrate changes in expression of Orai isoforms in response to B cell activation. We show that Orai3 and Orai1 are essential components of native CRAC channels in B cells and are critical for primary B cell proliferation and survival. The combined loss of Orai1 and Orai3 strongly impairs SOCE, nuclear factor for activated T cells (NFAT) activation, mitochondrial respiration, glycolysis, and the metabolic reprogramming of B cells in response to antigenic stimulation. Our results clarify the molecular composition and cellular functions of SOCE in B lymphocytes.

immunology↗

bioRxiv2020

Loss of the Pioneer Factor FOXA1 Results in Genome-wide Epigenetic Reprogramming and activation of Interferon-Response Genes including CD274/PD-L1

Forkhead Box A1 (FOXA1) is a pioneer transcription factor critical in epigenetic regulation of chromatin and cell fate determination. Reduced FOXA1 expression is an independent predictor of poor overall survival in bladder cancer patients. However, the impact of FOXA1 loss on chromatin epigenetics in bladder cancer is unknown. Therefore, we determined the impact of FOXA1 knock out (KO) on epigenetic modification of chromatin and associated gene expression. We identified 8,230 differentially expressed genes following FOXA1 KO. Surprisingly, Gene Set Enrichment Analysis (GSEA) identified IFN[a]/{gamma} gene expression signatures as enriched following FOXA1 KO. FOXA1 KO induced both increased and decreased numbers of histone 3 lysine 27 acetylation (H3K27ac) sites throughout the genome. As expected, the majority of differences in H3K27ac across genomic areas in FOXA1 KO cells is mapped to intergenic and intronic regions where enhancers reside. In addition, a subset of differential H3K27ac levels were also mapped to proximal promoters and within gene bodies. Integrated analysis of RNA/ChIP-seq data shows changes in gene expression that are mirrored by differences in H3K27ac. Motif analysis of DNA sequence enriched for H3K27ac identified significant increases in transcription factor binding motifs including the interferon sensitive response element (ISRE) and interferon response factors such as IRF1. Moreover, we identified increased H3K27ac of regulatory elements as being associated with several upregulated interferon sensitive genes (ISGs) in FOXA1 KO cells, including CD274/PD-L1. Western blotting and Q-RT-PCR confirmed upregulation of CD274/PD-L1 following FOXA1 KO. Analysis of TCGA data confirmed an inverse relationship between FOXA1 and CD274 in bladder cancer, as well as in other cancers. In summary, we provide evidence of widespread epigenetic reprogramming after FOXA1 KO in bladder cancer cells. Additionally, we provide evidence that FOXA1 KO-induced epigenetic changes contribute to activation of a global interferon-dominant expression signature, including the immune checkpoint target CD274/PD-L1 in a cancer cell-intrinsic manner.

cancer biology↗

bioRxiv2020

Colorectal adenocarcinomas downregulate the mitochondrial Na+/Ca2+ exchanger NCLX to drive metastatic spread

Despite the established role of mitochondria in tumorigenesis, the molecular mechanisms by which mitochondrial Ca2+ (mtCa2+) signaling regulates tumor growth and metastasis remain unknown. The crucial role of mtCa2+ in tumorigenesis is highlighted by the altered expression of proteins mediating mtCa2+ uptake and extrusion in cancer cells. Here, we demonstrate that expression of the mitochondrial Na+/Ca2+ exchanger NCLX (SLC8B1) is decreased in colorectal tumors and is associated with advanced-stage disease in patients. We reveal that downregulation of NCLX leads to mtCa2+ overload, mitochondrial depolarization, mitophagy, and reduced tumor size. Concomitantly, NCLX downregulation drives metastatic spread, chemoresistance, the expression of epithelial-to-mesenchymal transition (EMT), hypoxia, and stem cell pathways. Mechanistically, mtCa2+ overload leads to an increase in mitochondrial reactive oxygen species (mtROS) which activates HIF1 signaling supporting the metastatic behavior of tumor cells lacking NCLX. Our results reveal that loss of NCLX expression is a novel driver of metastatic progression, indicating that control of mtCa2+ levels is a novel therapeutic approach in metastatic colorectal cancer. HighlightsO_LIThe expression of NCLX is decreased in colorectal tumors and is associated with advanced-stage disease in patients. C_LIO_LINCLX plays a dichotomous role in colorectal tumor growth and metastasis. C_LIO_LINCLX downregulation causes mitophagy and reduced colorectal cancer tumor growth. C_LIO_LINCLX downregulation induces stemness, chemoresistance and metastasis through mtCa2+/ROS/HIF1 signaling axis. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=189 HEIGHT=200 SRC="FIGDIR/small/083071v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@30360dorg.highwire.dtl.DTLVardef@12ca72borg.highwire.dtl.DTLVardef@9328ddorg.highwire.dtl.DTLVardef@15ffcdb_HPS_FORMAT_FIGEXP M_FIG C_FIG SignificanceMitochondrial Ca2+ (mtCa2+) homeostasis is essential for cellular metabolism and growth and plays a critical role in cancer progression. mtCa2+ uptake is mediated by an inner membrane protein complex containing the mitochondrial Ca2+ uniporter (MCU). mtCa2+ uptake by the MCU is followed by a [~]100-fold slower mtCa2+ extrusion mediated by the inner mitochondrial membrane ion transporter, the mitochondrial Na+/Ca2+ exchanger NCLX. Because NCLX is a slower transporter than the MCU, it is a crucial rate-limiting factor of mtCa2+ homeostasis that cannot easily be compensated by another Ca2+ transport mechanism. This represents the first study investigating the role of NCLX in tumorigenesis and metastasis. We demonstrate for the first time that colorectal cancers exhibit loss of NCLX expression and that this is associated with advanced-stage disease. Intriguingly, decreased NCLX function has a dichotomous role in colorectal cancer. Thus, we reveal that NCLX loss leads to reduced primary tumor growth and overall tumor burden in vivo. Yet, the consequential increases in mtCa2+ elicit pro-survival, hypoxic and gene transcription pathways that enhance metastatic progression. This dichotomy is a well-established feature of chemoresistant and recurrent tumor cells including cancer stem cells. Moreover, the downstream changes elicited by NCLX loss are reminiscent of mesenchymal colorectal cancer subtypes that display poor patient survival. Our data indicate that the demonstrated changes to the mtCa2+/mtROS/HIF1 signaling axis elicited through the loss of NCLX are a key adaptation and driver of metastatic colorectal cancer.

cancer biology↗

bioRxiv2018

Repression of Transcription Factor AP-2 Alpha by Peroxisome Proliferator Activated Receptor Gamma Reveals a Novel Transcriptional Circuit in basal-squamous Bladder Cancer

The discovery of bladder cancer transcriptional subtypes provides an opportunity to identify high risk patients, and tailor disease management. Recent studies suggest tumor heterogeneity contributes to \"plasticity\" of molecular subtype during progression and following treatment. Nonetheless, the transcriptional drivers of the aggressive basal-squamous subtype remain unidentified. As PPAR{gamma} has been repeatedly implicated in the luminal subtype of bladder cancer, we hypothesized inactivation of this transcriptional master regulator during progression results in increased expression of basal-squamous specific transcription factors (TFs) which act to drive aggressive behavior. We initiated a pharmacologic and RNA-seq-based screen to identify PPAR{gamma}-repressed, basal-squamous specific TFs. Hierarchical clustering of RNA-seq data following treatment of a panel of human bladder cancer cell lines with a PPAR{gamma} agonist identified a number of TFs regulated by PPAR{gamma} activation, several of which are implicated in urothelial and squamous differentiation. One PPAR{gamma}-repressed TF implicated in squamous differentiation identified is Transcription Factor Activating Protein 2 alpha (TFAP2A). We show TFAP2A and its paralog TFAP2C are overexpressed in basal-squamous bladder cancer and in squamous areas of cystectomy samples, and that overexpression is associated with increased lymph node metastasis and distant recurrence, respectively. Biochemical analysis confirmed the ability of PPAR{gamma} activation to repress TFAP2A, while PPAR{gamma} antagonist studies indicate the requirement of a functional receptor. In vivo tissue recombination studies show TFAP2A and TFAP2C promote tumor growth in line with the aggressive nature of basal-squamous bladder cancer. Our findings suggest PPAR{gamma} inactivation, as well as TFAP2A and TFAP2C overexpression cooperate with other TFs to promote the basal-squamous transition.

cancer biology↗