Defining metabolic flexibility in hair follicle stem cell induced squamous cell carcinoma.

We previously showed that inhibition of glycolysis in cutaneous squamous cell carcinoma (SCC)-initiating cells had no effect on tumorigenesis, despite the perceived requirement of the Warburg effect, which was thought to drive carcinogenesis. Instead, these SCCs were metabolically flexible and sustained growth through glutaminolysis, another metabolic process frequently implicated to fuel tumorigenesis in various cancers. Here, we focused on glutaminolysis and genetically blocked this process through glutaminase (GLS) deletion in SCC cells of origin. Genetic deletion of GLS had little effect on tumorigenesis due to the up-regulated lactate consumption and utilization for the TCA cycle, providing further evidence of metabolic flexibility. We went on to show that posttranscriptional regulation of nutrient transporters appears to mediate metabolic flexibility in this SCC model. To define the limits of this flexibility, we genetically blocked both glycolysis and glutaminolysis simultaneously and found the abrogation of both of these carbon utilization pathways was enough to prevent both papilloma and frank carcinoma.

Targeting Asparagine Metabolism in Well-Differentiated/Dedifferentiated Liposarcoma.

BACKGROUND: mTORC1 activity is dependent on the presence of micronutrients, including Asparagine (Asn), to promote anabolic cell signaling in many cancers. We hypothesized that targeting Asn metabolism would inhibit tumor growth by reducing mTORC1 activity in well-differentiated (WD)/dedifferentiated (DD) liposarcoma (LPS). METHODS: Human tumor metabolomic analysis was utilized to compare abundance of Asn in WD vs. DD LPS. Gene set enrichment analysis (GSEA) compared relative expression among metabolic pathways upregulated in DD vs. WD LPS. Proliferation assays were performed for LPS cell lines and organoid models by using the combination treatment of electron transport chain (ETC) inhibitors with Asn-free media. 13C-Glucose-labeling metabolomics evaluated the effects of combination treatment on nucleotide synthesis. Murine xenograft models were used to assess the effects of ETC inhibition combined with PEGylated L-Asparaginase (PEG-Asnase) on tumor growth and mTORC1 signaling. RESULTS: Asn was enriched in DD LPS compared to WD LPS. GSEA indicated that mTORC1 signaling was upregulated in DD LPS. Within available LPS cell lines and organoid models, the combination of ETC inhibition with Asn-free media resulted in reduced cell proliferation. Combination treatment inhibited nucleotide synthesis and promoted cell cycle arrest. In vivo, the combination of ETC inhibition with PEG-Asnase restricted tumor growth. CONCLUSIONS: Asn enrichment and mTORC1 upregulation are important factors contributing to WD/DD LPS tumor progression. Effective targeting strategies require limiting access to extracellular Asn and inhibition of de novo synthesis mechanisms. The combination of PEG-Asnase with ETC inhibition is an effective therapy to restrict tumor growth in WD/DD LPS.

Lessons Learned from Cancer Metabolism for Physiology and Disease.

Tumor cells divide rapidly and dramatically alter their metabolism to meet biosynthetic and bioenergetic needs. Through studying the aberrant metabolism of cancer cells, other contexts in which metabolism drives cell state transitions become apparent. In this work, we will discuss how principles established by the field of cancer metabolism have led to discoveries in the contexts of physiology and tissue injury, mammalian embryonic development, and virus infection. We present specific examples of findings from each of these fields that have been shaped by the study of cancer metabolism. We also discuss the next important scientific questions facing these subject areas collectively. Altogether, these examples demonstrate that the study of "cancer metabolism" is indeed the study of cell metabolism in the context of a tumor, and undoubtedly discoveries from each of the fields discussed here will continue to build on each other in the future.

Acute activation of adipocyte lipolysis reveals dynamic lipid remodeling of the hepatic lipidome.

Adipose tissue is the site of long-term energy storage. During the fasting state, exercise, and cold exposure, the white adipose tissue mobilizes energy for peripheral tissues through lipolysis. The mobilization of lipids from white adipose tissue to the liver can lead to excess triglyceride accumulation and fatty liver disease. Although the white adipose tissue is known to release free fatty acids, a comprehensive analysis of lipids mobilized from white adipocytes in vivo has not been completed. In these studies, we provide a comprehensive quantitative analysis of the adipocyte-secreted lipidome and show that there is interorgan crosstalk with liver. Our analysis identifies multiple lipid classes released by adipocytes in response to activation of lipolysis. Time-dependent analysis of the serum lipidome showed that free fatty acids increase within 30 min of ß3-adrenergic receptor activation and subsequently decrease, followed by a rise in serum triglycerides, liver triglycerides, and several ceramide species. The triglyceride composition of liver is enriched for linoleic acid despite higher concentrations of palmitate in the blood. To further validate that these findings were a specific consequence of lipolysis, we generated mice with conditional deletion of adipose tissue triglyceride lipase exclusively in adipocytes. This loss of in vivo adipocyte lipolysis prevented the rise in serum free fatty acids and hepatic triglycerides. Furthermore, conditioned media from adipocytes promotes lipid remodeling in hepatocytes with concomitant changes in genes/pathways mediating lipid utilization. Together, these data highlight critical role of adipocyte lipolysis in interorgan crosstalk between adipocytes and liver.

Atlas of fetal metabolism during mid-to-late gestation and diabetic pregnancy.

Mounting evidence suggests metabolism instructs stem cell fate decisions. However, how fetal metabolism changes during development and how altered maternal metabolism shapes fetal metabolism remain unexplored. We present a descriptive atlas of in vivo fetal murine metabolism during mid-to-late gestation in normal and diabetic pregnancy. Using 13C-glucose and liquid chromatography-mass spectrometry (LC-MS), we profiled the metabolism of fetal brains, hearts, livers, and placentas harvested from pregnant dams between embryonic days (E)10.5 and 18.5. Our analysis revealed metabolic features specific to a hyperglycemic environment and signatures that may denote developmental transitions during euglycemic development. We observed sorbitol accumulation in fetal tissues and altered neurotransmitter levels in fetal brains isolated from hyperglycemic dams. Tracing 13C-glucose revealed disparate fetal nutrient sourcing depending on maternal glycemic states. Regardless of glycemic state, histidine-derived metabolites accumulated in late-stage fetal tissues. Our rich dataset presents a comprehensive overview of in vivo fetal tissue metabolism and alterations due to maternal hyperglycemia.

Endothelial Jagged1 levels and distribution are post-transcriptionally controlled by ZFP36 decay proteins.

Vascular morphogenesis requires a delicate gradient of Notch signaling controlled, in part, by the distribution of ligands (Dll4 and Jagged1). How Jagged1 (JAG1) expression is compartmentalized in the vascular plexus remains unclear. Here, we show that Jag1 mRNA is a direct target of zinc-finger protein 36 (ZFP36), an RNA-binding protein involved in mRNA decay that we find robustly induced by vascular endothelial growth factor (VEGF). Endothelial cells lacking ZFP36 display high levels of JAG1 and increase angiogenic sprouting in vitro. Furthermore, mice lacking Zfp36 in endothelial cells display mispatterned and increased levels of JAG1 in the developing retinal vascular plexus. Abnormal levels of JAG1 at the sprouting front alters NOTCH1 signaling, increasing the number of tip cells, a phenotype that is rescued by imposing haploinsufficiency of Jag1. Our findings reveal an important feedforward loop whereby VEGF stimulates ZFP36, consequently suppressing Jag1 to enable adequate levels of Notch signaling during sprouting angiogenesis.

MYC is a regulator of androgen receptor inhibition-induced metabolic requirements in prostate cancer.

Advanced prostate cancers are treated with therapies targeting the androgen receptor (AR) signaling pathway. While many tumors initially respond to AR inhibition, nearly all develop resistance. It is critical to understand how prostate tumor cells respond to AR inhibition in order to exploit therapy-induced phenotypes prior to the outgrowth of treatment-resistant disease. Here, we comprehensively characterize the effects of AR blockade on prostate cancer metabolism using transcriptomics, metabolomics, and bioenergetics approaches. The metabolic response to AR inhibition is defined by reduced glycolysis, robust elongation of mitochondria, and increased reliance on mitochondrial oxidative metabolism. We establish DRP1 activity and MYC signaling as mediators of AR-blockade-induced metabolic phenotypes. Rescuing DRP1 phosphorylation after AR inhibition restores mitochondrial fission, while rescuing MYC restores glycolytic activity and prevents sensitivity to complex I inhibition. Our study provides insight into the regulation of treatment-induced metabolic phenotypes and vulnerabilities in prostate cancer.

Vitamin B6 is governed by the local compartmentalization of metabolic enzymes during growth.

Vitamin B6 is a vital micronutrient across cell types and tissues, and dysregulated B6 levels contribute to human disease. Despite its importance, how B6 vitamer levels are regulated is not well understood. Here, we provide evidence that B6 dynamics are rapidly tuned by precise compartmentation of pyridoxal kinase (PDXK), the rate-limiting B6 enzyme. We show that canonical Wnt rapidly led to the accumulation of inactive B6 by shunting cytosolic PDXK into lysosomes. PDXK was modified with methyl-arginine Degron (MrDegron), a protein tag for lysosomes, which enabled delivery via microautophagy. Hyperactive lysosomes resulted in the continuous degradation of PDXK and B6 deficiency that promoted proliferation in Wnt-driven colorectal cancer (CRC) cells. Pharmacological or genetic disruption of the coordinated MrDegron proteolytic pathway was sufficient to reduce CRC survival in cells and organoid models. In sum, this work contributes to the repertoire of micronutrient-regulated processes that enable cancer cell growth and provides insight into the functional impact of B6 deficiencies for survival.

Nutrient regulation of development and cell fate decisions.

Diet contributes to health at all stages of life, from embryonic development to old age. Nutrients, including vitamins, amino acids, lipids and sugars, have instructive roles in directing cell fate and function, maintaining stem cell populations, tissue homeostasis and alleviating the consequences of aging. This Review highlights recent findings that illuminate how common diets and specific nutrients impact cell fate decisions in healthy and disease contexts. We also draw attention to new models, technologies and resources that help to address outstanding questions in this emerging field and may lead to dietary approaches that promote healthy development and improve disease treatments.

FH Variant Pathogenicity Promotes Purine Salvage Pathway Dependence in Kidney Cancer.

Fumarate accumulation due to loss of fumarate hydratase (FH) drives cellular transformation. Germline FH alterations lead to hereditary leiomyomatosis and renal cell cancer (HLRCC) where patients are predisposed to an aggressive form of kidney cancer. There is an unmet need to classify FH variants by cancer-associated risk. We quantified catalytic efficiencies of 74 variants of uncertain significance. Over half were enzymatically inactive, which is strong evidence of pathogenicity. We next generated a panel of HLRCC cell lines expressing FH variants with a range of catalytic activities, then correlated fumarate levels with metabolic features. We found that fumarate accumulation blocks de novo purine biosynthesis, rendering FH-deficient cells reliant on purine salvage for proliferation. Genetic or pharmacologic inhibition of the purine salvage pathway reduced HLRCC tumor growth in vivo. These findings suggest the pathogenicity of patient-associated FH variants and reveal purine salvage as a targetable vulnerability in FH-deficient tumors. SIGNIFICANCE: This study functionally characterizes patient-associated FH variants with unknown significance for pathogenicity. This study also reveals nucleotide salvage pathways as a targetable feature of FH-deficient cancers, which are shown to be sensitive to the purine salvage pathway inhibitor 6-mercaptopurine. This presents a new rapidly translatable treatment strategy for FH-deficient cancers. This article is featured in Selected Articles from This Issue, p. 1949.