Explore the vast range of titles available.

Elevated levels of the gut microbe-derived metabolite trimethylamine N -oxide (TMAO) are associated with cardiometabolic disease risk. However, the mechanism(s) linking TMAO production to human disease are incompletely understood. Initiation of the metaorganismal TMAO pathway begins when dietary choline and related metabolites are converted to trimethylamine (TMA) by gut bacteria. Gut microbe-derived TMA can then be further oxidized by host flavin-containing monooxygenases to generate TMAO. Previously, we showed that drugs lowering both TMA and TMAO protect mice against obesity via rewiring of host circadian rhythms (Schugar et al., 2022). Although most mechanistic studies in the literature have focused on the metabolic end product TMAO, here we have instead tested whether the primary metabolite TMA alters host metabolic homeostasis and circadian rhythms via trace amine-associated receptor 5 (TAAR5). Remarkably, mice lacking the host TMA receptor ( Taar5 −/ − ) have altered circadian rhythms in gene expression, metabolic hormones, gut microbiome composition, and diverse behaviors. Also, mice genetically lacking bacterial TMA production or host TMA oxidation have altered circadian rhythms. These results provide new insights into diet–microbe–host interactions relevant to cardiometabolic disease and implicate gut bacterial production of TMA and the host receptor that senses TMA (TAAR5) in the physiologic regulation of circadian rhythms in mice.

Epigenetic dysregulation, including accumulation of Histone H3 lysine 27 acetylation (H3K27ac), is a hallmark of pVHL-deficient clear cell Renal Cell Carcinomas (ccRCCs). Using an in vivo positive selection ORF screen in poorly tumorigenic pVHL-proficient cells and mechanistic studies in pVHL-deficient cells, we discovered that the aspartate (Asp) and glutamate (Glu) transporter, SLC1A1/EAAT3, is a metabolic dependency in ccRCC. pVHL loss promotes Hypoxia Inducible Factor (HIF)-independent SLC1A1 expression via H3K27ac dysregulation. SLC1A1 inactivation, genetically or pharmacologically, depletes Asp/Glu-derived metabolites (e.g., Tricarboxylic acid cycle and nucleotide intermediates), impedes ccRCC growth, and sensitizes ccRCCs to anti-metabolite drugs (e.g., glutaminase blockers). In human tumors, higher SLC1A1 expression is associated with reduced immune infiltration, oncogenic metabolic programs, and advanced stage/metastatic disease. Finally, in ccRCC animal models, SLC1A1 inactivation diminishes lung metastasis and the outgrowth of established renal tumors. Altogether, our studies credential SLC1A1 as an actionable, HIF-independent, metabolic dependency in pVHL-deficient ccRCCs. Clear cell renal cell carcinoma (ccRCC) bears the hallmark loss of VHL but remains incurable. Here, the authors identify the SLC1A1 dicarboxylic amino acid transporter as an actionable, oncogenic, HIF-independent, metabolic dependency in VHL-deficient ccRCCs.

Glioblastoma, IDH-wild type (GBM) is the most common and lethal malignant primary brain tumor. Standard of care includes surgery, radiotherapy, and chemotherapy with the DNA alkylating agent temozolomide (TMZ). Despite these intensive efforts, current GBM therapy remains mainly palliative with only modest improvement achieved in overall survival. With regards to radiotherapy, GBM is ranked as one of the most radioresistant tumor types. In this study, we wanted to investigate if enriching cells in the most radiosensitive cell cycle phase, mitosis, could improve localized radiotherapy for GBM. To achieve cell cycle arrest in mitosis we used ispinesib, a small molecule inhibitor to the mitotic kinesin, KIF11. Cell culture studies validated that ispinesib radiosensitized patient-derived GBM cells. In vivo, we validated that ispinesib increased the fraction of tumor cells arrested in mitosis as well as increased apoptosis. Critical for the translation of this approach, we validated that combination therapy with ispinesib and irradiation led to the greatest increase in survival over either monotherapy alone. Our data highlight KIF11 inhibition in combination with radiotherapy as a new combinatorial approach that reduces the overall radioresistance of GBM and which can readily be moved into clinical trials.

Oxygen is required for most multi-cellular life; however, this was not always the case. But with oxygen’s utilization in aerobic respiration, came mechanisms to deal with its scarcity (hypoxia). Indeed, the ability of cells to sense and respond to changing oxygen concentrations is critical to the physiology of all aerobic life. Responses to hypoxia are broad and organized at both organismal (hyperventilation, increased cardiac output) and cellular (reduced energy expenditure, changes in metabolism, dedifferentiation) levels. Importantly, when cellular responses go awry, they can contribute to the development of cancer.Spurred by separate reports that tumors contained regions of hypoxia and that anaerobic cells were resistant to ionizing radiation, research began to understand the clinical implications of hypoxia. We now know hypoxia associates with worse disease prognosis across solid tumors and is recognized as a critical component of the tumor microenvironment. Notably, all solid tumors contain regions of hypoxia, resultant of poor vascularization, responsible for driving many hallmarks of cancer. Hypoxia-inducible factor (HIF) transcription factors, which are understood to drive the majority of known cellular adaptations to hypoxia, are a mainstay for therapeutic development. Yet, HIF targeted interventions are insufficient to cure disease. Importantly, accumulating evidence within the last decade, has demonstrated the relevance of HIF-independent hypoxia-induced cellular adaptations to oncogenesis. Herein, our aim is to identify these mechanisms.Chapter 1 will discuss how we leveraged epigenetic and transcriptomic data to identify the SLC1A1 Asp/Glu transporter as the HIF-independent dependency in clear cell Renal Cell Carcinoma (ccRCC). Chapter 2 will discuss how we leveraged cancer lineage-specific gene dependencies to that ccRCCs are particularly dependent on the BCL-XL anti-apoptotic protein, but in a HIF-independent manner. Chapter 3 will discuss how we leveraged pharmacological screens using a stemness reporter system to identify the glucocorticoid receptor as a HIF-independent regulator of hypoxia-induced epithelial-to-mesenchymal transition in triple-negative breast cancer. Finally, chapter 4 will describe our future directions. Together, these findings add to a larger body of work demonstrating how hypoxia can drive oncogenesis independent of HIF and may explain why perturbing HIF alone does not always achieve durable anti-tumor responses.

Abstract Background Epigenetic dysregulation, including accumulation of Histone H3 lysine 27 acetylation (H3K27ac), is a hallmark of pVHL-deficient clear cell Renal Cell Carcinomas (ccRCCs). H3K27ac is associated with transcriptional activation and its accumulation at cis-regulatory elements (eg, promoters and enhancers/super-enhancers) marks key oncogenes and regulators of cellular identity in many cancers. In ccRCC, specific alterations in H3K27ac have been linked to tumorigenesis and metastatic progression. Importantly, these earlier studies largely relied on the HIF2α-dependent 786-O cells (or their metastatic derivatives), perhaps, missing the importance of HIF-independent epigenetic programs. Altogether, we hypothesized that H3K27ac marks critical genes in pVHL-deficient ccRCCs that sustain tumorigenic and metastatic programs via both HIF-dependent and independent mechanisms. Methods Using an in vivo positive selection ORF screen in poorly tumorigenic pVHL-proficient cells and cell-based mechanistic studies in pVHL-deficient cells, we discovered that the aspartate (Asp) and glutamate (Glu) transporter, SLC1A1/EAAT3, is a metabolic oncogenic dependency in ccRCC. Results pVHL loss promotes HIF-independent SLC1A1 expression via H3K27ac dysregulation. SLC1A1 inactivation, using either genetic or pharmacological approaches, depletes Asp/Glu-derived metabolites [eg, Tricarboxylic acid (TCA) cycle and nucleotide intermediates], impedes ccRCC growth, and sensitizes ccRCCs to anti-metabolite drugs (eg, glutaminase blockers). In human tumors, higher SLC1A1 expression is associated with reduced immune infiltration, oncogenic metabolic programs, and advanced stage/metastatic disease. Finally, in ccRCC animal models, SLC1A1 inactivation diminishes lung metastasis and the outgrowth of established renal tumors. Conclusions Altogether, our studies credential SLC1A1 as a novel, actionable, HIF-independent, metabolic dependency in pVHL-deficient ccRCCs.

Elevated levels of the gut microbe-derived metabolite trimethylamine N -oxide (TMAO) are associated with cardiometabolic disease risk. However, the mechanism(s) linking TMAO production to human disease are incompletely understood. Initiation of the metaorganismal TMAO pathway begins when dietary choline and related metabolites are converted to trimethylamine (TMA) by gut bacteria. Gut microbe-derived TMA can then be further oxidized by host flavin-containing monooxygenases to generate TMAO. Previously, we showed that drugs lowering both TMA and TMAO protect mice against obesity via rewiring of host circadian rhythms (Schugar et al., 2022). Although most mechanistic studies in the literature have focused on the metabolic end product TMAO, here we have instead tested whether the primary metabolite TMA alters host metabolic homeostasis and circadian rhythms via trace amine-associated receptor 5 (TAAR5). Remarkably, mice lacking the host TMA receptor ( Taar5 −/ − ) have altered circadian rhythms in gene expression, metabolic hormones, gut microbiome composition, and diverse behaviors. Also, mice genetically lacking bacterial TMA production or host TMA oxidation have altered circadian rhythms. These results provide new insights into diet–microbe–host interactions relevant to cardiometabolic disease and implicate gut bacterial production of TMA and the host receptor that senses TMA (TAAR5) in the physiologic regulation of circadian rhythms in mice.

Genome-wide genetic screens have identified cellular dependencies in many cancers. Using the Broad Institutes Achilles shRNA screening dataset, we mined for targetable dependencies by cell lineage. Our studies identified a strong dependency on BCL2L1, which encodes the BCL-XL anti-apoptotic protein, in a subset of kidney cancer cells. Genetic and pharmacological inactivation of BCL-XL, but not the related anti-apoptotic proteins BCL-2, led to fitness defects in renal cancer cells, and also sensitized them to chemotherapeutics. Neither BCL-XL levels (absolute or normalized to BCL-2) nor the status of the VHL gene, which is frequently mutated in kidney cancer, predicted BCL-XL dependence. Transcriptional profiling, however, identified a BCL-XL dependency mRNA signature, which included elevated mesenchymal gene expression in BCL-XL dependent cells. Promoting mesenchymal transition increased BCL-XL dependence; whereas, conversion to a more differentiated state overcame BCL-XL dependence in kidney cancer cells. The BCL-XL dependency mRNA signature was observed in almost a third of human clear cell Renal Cell Carcinomas (ccRCCs), which were also associated with worse clinical outcomes. Finally, an orally bioavailable BCL-XL inhibitor, A-1331852, showed anti-tumor efficacy in vivo. Altogether, our studies uncovered an unexpected link between cancer cell state and dependence on the anti-apoptotic BCL-XL protein and justify further testing on BCL-XL blockade as a potential way to target a clinically aggressive subset of human kidney cancers. Competing Interest Statement The authors have declared no competing interest.

The BCL-X anti-apoptotic protein is a clear cell Renal Cell Carcinoma (ccRCC) dependency; however, the mechanism of this dependence and its relevance in other aggressive kidney cancer contexts, including metastatic and/or rare RCC subtypes [e.g., Fumarate Hydratase (FH)-deficient and sarcomatoid RCCs], is unknown. Computational predictions, using a machine learning model trained on the human RCC TCGA dataset, and cell-based validations, confirmed BCL-X dependence in all RCC subtypes. Remarkably, cell state changes, 'anoikis' programs, inflammatory state, and metabolic perturbations (e.g., fumarate production in FH-deficient RCCs) independently conferred increased BCL-X dependence. Correlation studies revealed that increased AMPK isoform 2 ( ) expression is a kidney-specific biomarker of BCL-X dependence. Indeed, pharmacological AMPK activation sensitized RCCs to BCL-X blockade. Finally, using functional studies, we developed a multivariate model that accurately predicted BCL-X dependence in RCC. Our studies offer biomarkers for patient stratification and credential BCL-X as a subtype agnostic vulnerability in difficult-to-treat RCCs.

Elevated levels of the gut microbe-derived metabolite trimethylamine N-oxide (TMAO) are associated with cardiometabolic disease risk. However, the mechanism(s) linking TMAO production to human disease are incompletely understood. Initiation of the metaorganismal TMAO pathway begins when dietary choline and related metabolites are converted to trimethylamine (TMA) by gut bacteria. Gut microbe-derived TMA can then be further oxidized by host flavin-containing monooxygenases to generate TMAO. Previously, we showed that drugs lowering both TMA and TMAO protect mice against obesity via rewiring of host circadian rhythms. Although most mechanistic studies in the literature have focused on the metabolic end product TMAO, here we have instead tested whether the primary metabolite TMA alters host metabolic homeostasis and circadian rhythms via trace amine-associated receptor 5 (TAAR5). Remarkably, mice lacking the host TMA receptor ( ) have altered circadian rhythms in gene expression, metabolic hormones, gut microbiome composition, and diverse behaviors. Also, mice genetically lacking bacterial TMA production or host TMA oxidation have altered circadian rhythms. These results provide new insights into diet-microbe-host interactions relevant to cardiometabolic disease, and implicate gut bacterial production of TMA and the host receptor that senses TMA (TAAR5) in the physiologic regulation of circadian rhythms in mice.

Although strong evidence links the gut microbiome to metabolic disease, the mechanisms linking microbiota to hormonal and metabolic responses to food are not well understood. After a meal, gut bacteria produce a wide array of small molecule, protein, and lipid metabolites originating from bacterial sources. Annotating physiological function to select gut microbe-derived metabolites is critical to understanding diet-microbe-host interactions, and to developing microbiome-inspired therapies to improve human health. Here, we have investigated the role of a poorly annotated class of gut microbiome-derived lipids called N-acyl amides in postprandial metabolic physiology. Here we show both bacterial overproduction and provision of exogenous N-acyl amides reorganize host hormone-driven metabolic transition after a meal. Moreover, N-acyl amides exert broad effects on the meal- and circadian-related reorganization of gene expression, metabolic hormones, and gut microbiome composition. Collectively, these results demonstrate that microbiota-derived N-acyl amides play a physiologic role in postprandial metabolic homeostasis in the host.