Faculty Bibliography

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Cancer-associated fibroblasts (CAFs) are a key component of the tumour microenvironment with diverse functions, including matrix deposition and remodelling, extensive reciprocal signalling interactions with cancer cells and crosstalk with infiltrating leukocytes. As such, they are a potential target for optimizing therapeutic strategies against cancer. However, many challenges are present in ongoing attempts to modulate CAFs for therapeutic benefit. These include limitations in our understanding of the origin of CAFs and heterogeneity in CAF function, with it being desirable to retain some antitumorigenic functions. On the basis of a meeting of experts in the field of CAF biology, we summarize in this Consensus Statement our current knowledge and present a framework for advancing our understanding of this critical cell type within the tumour microenvironment.

Background/UNASSIGNED:Metabolic programs in cancer cells are influenced by genotype and the tissue of origin. We have previously shown that central carbon metabolism is rewired in pancreatic ductal adenocarcinoma (PDA) to support proliferation through a glutamate oxaloacetate transaminase 1 (GOT1)-dependent pathway. Methods/UNASSIGNED:test (unpaired, two-tailed) between all PDA metabolites and CRC metabolites. Results/UNASSIGNED:While PDA exhibits profound growth inhibition upon GOT1 knockdown, we found CRC to be insensitive. In PDA, but not CRC, GOT1 inhibition disrupted glycolysis, nucleotide metabolism, and redox homeostasis. These insights were leveraged in PDA, where we demonstrate that radiotherapy potently enhanced the effect of GOT1 inhibition on tumor growth. Conclusions/UNASSIGNED:Taken together, these results illustrate the role of tissue type in dictating metabolic dependencies and provide new insights for targeting metabolism to treat PDA.

The most frequently mutated oncogene in cancer is KRAS, which uses alternative fourth exons to generate two gene products (KRAS4A and KRAS4B) that differ only in their C-terminal membrane-targeting region¹. Because oncogenic mutations occur in exons 2 or 3, two constitutively active KRAS proteins-each capable of transforming cells-are encoded when KRAS is activated by mutation². No functional distinctions among the splice variants have so far been established. Oncogenic KRAS alters the metabolism of tumour cells³ in several ways, including increased glucose uptake and glycolysis even in the presence of abundant oxygen⁴ (the Warburg effect). Whereas these metabolic effects of oncogenic KRAS have been explained by transcriptional upregulation of glucose transporters and glycolytic enzymes^3-5, it is not known whether there is direct regulation of metabolic enzymes. Here we report a direct, GTP-dependent interaction between KRAS4A and hexokinase 1 (HK1) that alters the activity of the kinase, and thereby establish that HK1 is an effector of KRAS4A. This interaction is unique to KRAS4A because the palmitoylation-depalmitoylation cycle of this RAS isoform enables colocalization with HK1 on the outer mitochondrial membrane. The expression of KRAS4A in cancer may drive unique metabolic vulnerabilities that can be exploited therapeutically.

Pancreatic ductal adenocarcinoma is an aggressive cancer with limited treatment options¹. Approximately 10% of cases exhibit familial predisposition, but causative genes are not known in most families². We perform whole-genome sequence analysis in a family with multiple cases of pancreatic ductal adenocarcinoma and identify a germline truncating mutation in the member of the RAS oncogene family-like 3 (RABL3) gene. Heterozygous rabl3 mutant zebrafish show increased susceptibility to cancer formation. Transcriptomic and mass spectrometry approaches implicate RABL3 in RAS pathway regulation and identify an interaction with RAP1GDS1 (SmgGDS), a chaperone regulating prenylation of RAS GTPases³. Indeed, the truncated mutant RABL3 protein accelerates KRAS prenylation and requires RAS proteins to promote cell proliferation. Finally, evidence in patient cohorts with developmental disorders implicates germline RABL3 mutations in RASopathy syndromes. Our studies identify RABL3 mutations as a target for genetic testing in cancer families and uncover a mechanism for dysregulated RAS activity in development and cancer.

Autophagy, a multistep lysosomal degradation pathway that supports nutrient recycling and metabolic adaptation, has been implicated as a process that regulates cancer. Although autophagy induction may limit the development of tumors, evidence in mouse models demonstrates that autophagy inhibition can limit the growth of established tumors and improve response to cancer therapeutics. Certain cancer genotypes may be especially prone to autophagy inhibition. Different strategies for autophagy modulation may be needed depending on the cancer context. Here, we review new advances in the molecular control of autophagy, the role of selective autophagy in cancer, and the role of autophagy within the tumor microenvironment and tumor immunity. We also highlight clinical efforts to repurpose lysosomal inhibitors, such as hydroxychloroquine, as anticancer agents that block autophagy, as well as the development of more potent and specific autophagy inhibitors for cancer treatment, and review future directions for autophagy research.Significance: Autophagy plays a complex role in cancer, but autophagy inhibition may be an effective therapeutic strategy in advanced cancer. A deeper understanding of autophagy within the tumor microenvironment has enabled the development of novel inhibitors and clinical trial strategies. Challenges and opportunities remain to identify patients most likely to benefit from this approach.

Ncoa4 mediates autophagic degradation of ferritin, the cytosolic iron storage complex, to maintain intracellular iron homeostasis. Recent evidence also supports a role for Ncoa4 in systemic iron homeostasis and erythropoiesis. However, the specific contribution and temporal importance of Ncoa4-mediated ferritinophagy in regulating systemic iron homeostasis and erythropoiesis is unclear. Here, we show that Ncoa4 has a critical role in basal systemic iron homeostasis and both cell autonomous and non-autonomous roles in murine erythropoiesis. Using an inducible murine model of Ncoa4 knockout, acute systemic disruption of Ncoa4 impaired systemic iron homeostasis leading to tissue ferritin and iron accumulation, a decrease in serum iron, and anemia. Mice acutely depleted of Ncoa4 engaged the Hif2α-erythropoietin system to compensate for anemia. Mice with targeted deletion of Ncoa4 specifically in the erythroid compartment developed a pronounced anemia in the immediate postnatal stage, a mild hypochromic microcytic anemia at adult stages, and were more sensitive to hemolysis with higher requirements for the Hif2α-Epo axis and extra-medullary erythropoiesis during recovery. These studies demonstrate the importance of Ncoa4-mediated ferritinophagy as a regulator of systemic iron homeostasis and define the relative cell autonomous and non-autonomous contributions of Ncoa4 in supporting erythropoiesis in vivo.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by KRAS- and autophagy-dependent tumorigenic growth, but the role of KRAS in supporting autophagy has not been established. We show that, to our surprise, suppression of KRAS increased autophagic flux, as did pharmacological inhibition of its effector ERK MAPK. Furthermore, we demonstrate that either KRAS suppression or ERK inhibition decreased both glycolytic and mitochondrial functions. We speculated that ERK inhibition might thus enhance PDAC dependence on autophagy, in part by impairing other KRAS- or ERK-driven metabolic processes. Accordingly, we found that the autophagy inhibitor chloroquine and genetic or pharmacologic inhibition of specific autophagy regulators synergistically enhanced the ability of ERK inhibitors to mediate antitumor activity in KRAS-driven PDAC. We conclude that combinations of pharmacologic inhibitors that concurrently block both ERK MAPK and autophagic processes that are upregulated in response to ERK inhibition may be effective treatments for PDAC.