Macropinocytosis confers resistance to therapies targeting cancer anabolism.

Macropinocytic cancer cells scavenge amino acids from extracellular proteins. Here, we show that consuming necrotic cell debris via macropinocytosis (necrocytosis) offers additional anabolic benefits. A click chemistry-based flux assay reveals that necrocytosis provides not only amino acids, but sugars, fatty acids and nucleotides for biosynthesis, conferring resistance to therapies targeting anabolic pathways. Indeed, necrotic cell debris allow macropinocytic breast and prostate cancer cells to proliferate, despite fatty acid synthase inhibition. Standard therapies such as gemcitabine, 5-fluorouracil (5-FU), doxorubicin and gamma-irradiation directly or indirectly target nucleotide biosynthesis, creating stress that is relieved by scavenged nucleotides. Strikingly, necrotic debris also render macropinocytic, but not non-macropinocytic, pancreas and breast cancer cells resistant to these treatments. Selective, genetic inhibition of macropinocytosis confirms that necrocytosis both supports tumor growth and limits the effectiveness of 5-FU in vivo. Therefore, this study establishes necrocytosis as a mechanism for drug resistance

Macropinocytosis fuels cancer cell growth and drug resistance

To support their accelerated non-homeostatic proliferation, cancer cells depends on access to extracellular nutrients derived from the tumor vasculature. However, dysfunctional tumor vasculature and high interstitial pressure limit tumor perfusion leaving many tumor cells nutrient-deprived. To bypass this nutrient limitation, RAS driven cancer cells recycle their own nutrients via autophagy or scavenge macromolecules from the tumor microenvironment (TME) via macropinocytosis. Oncogenic mutations in RAS promote macropinocytosis, a bulk scavenging process through which components in the TME can be non-specifically acquired. When macropinosomes fuse with lysosomes, degradative enzymes release nutrients that can support cancer cell growth and proliferation. We report macropinocytosis is not just a RAS driven phenotype but a cancer phenotype. Lipid PIP3 generated by the PI3K pathway is necessary for closure of macropinosomes in most contexts. We identified prostate and breast cancers with hyper-activation of the PI3K pathway perform macropinocytosis in an AMPK dependent manner. AMPK mediated macropinocytosis is necessary for proliferation of PTEN-deficient cells in low nutrients but not autophagy. Because scavenging allows cells to acquire nutrients from cell extrinsic sources, it is likely to play a greater role in promoting tumor growth than cell-autonomous autophagy, which leads to atrophy. Necrosis is observed in most solid tumors; necrotic cell debris is selectively scavenged via macropinocytosis. Nutrient-deprived macropinocytic breast cancer cells acquired not just amino acids, but also sugars, lipids, and nucleotides from necrotic cell debris (necrocytosis) suggesting macropinocytosis plays a larger role than previously appreciated in supporting cancer anabolism. In addition, necrocytosis also conferred resistance to various therapies that target de novo nucleotide synthesis, fatty acid synthesis as well standard of care genotoxic therapies resulting in the first demonstration that scavenging confers drug resistance. Together, these studies suggest that macropinocytosis inhibitors could have therapeutic value in many cancer patients, particularly in combination with conventional therapies

Macropinocytosis fuels cancer cell growth and drug resistance

To support their accelerated non-homeostatic proliferation, cancer cells depends on access to extracellular nutrients derived from the tumor vasculature. However, dysfunctional tumor vasculature and high interstitial pressure limit tumor perfusion leaving many tumor cells nutrient-deprived. To bypass this nutrient limitation, RAS driven cancer cells recycle their own nutrients via autophagy or scavenge macromolecules from the tumor microenvironment (TME) via macropinocytosis. Oncogenic mutations in RAS promote macropinocytosis, a bulk scavenging process through which components in the TME can be non-specifically acquired. When macropinosomes fuse with lysosomes, degradative enzymes release nutrients that can support cancer cell growth and proliferation. We report macropinocytosis is not just a RAS driven phenotype but a cancer phenotype. Lipid PIP3 generated by the PI3K pathway is necessary for closure of macropinosomes in most contexts. We identified prostate and breast cancers with hyper-activation of the PI3K pathway perform macropinocytosis in an AMPK dependent manner. AMPK mediated macropinocytosis is necessary for proliferation of PTEN-deficient cells in low nutrients but not autophagy. Because scavenging allows cells to acquire nutrients from cell extrinsic sources, it is likely to play a greater role in promoting tumor growth than cell-autonomous autophagy, which leads to atrophy. Necrosis is observed in most solid tumors; necrotic cell debris is selectively scavenged via macropinocytosis. Nutrient-deprived macropinocytic breast cancer cells acquired not just amino acids, but also sugars, lipids, and nucleotides from necrotic cell debris (necrocytosis) suggesting macropinocytosis plays a larger role than previously appreciated in supporting cancer anabolism. In addition, necrocytosis also conferred resistance to various therapies that target de novo nucleotide synthesis, fatty acid synthesis as well standard of care genotoxic therapies resulting in the first demonstration that scavenging confers drug resistance. Together, these studies suggest that macropinocytosis inhibitors could have therapeutic value in many cancer patients, particularly in combination with conventional therapies

Starving PTEN-deficient prostate cancer cells thrive under nutrient stress by scavenging corpses for their supper.

Our recent work demonstrates that inactivating mutations in phosphatase and tensin homolog (PTEN) are sufficient to drive macropinocytosis in the context of AMP-activated protein kinase (AMPK) activation. Given that blocking macropinocytosis limits PTEN-deficient prostate tumor growth, AMPK or phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) inhibitors could have therapeutic value in castration-resistant prostate cancer patients, particularly when used in combination with standard of care therapies. Abbreviations: ATG5: autophagy related 5; NHE: Na(+)/H(+) exchanger; PAK1: p21-activated kinase 1; PI3K: phosphatidylinositol-4,5-bisphosphate 3-kinase; PIP3: phosphatidylinositol (3,4,5)-trisphosphate; PIP2: phosphatidylinositol (4,5)-bisphosphate; RAC1: Rac family small GTPase 1

Nutrient scavenging in cancer.

While cancer cell proliferation depends on access to extracellular nutrients, inadequate tumour perfusion means that glucose, amino acids and lipids are often in short supply. To overcome this obstacle to growth, cancer cells utilize multiple scavenging strategies, obtaining macromolecules from the microenvironment and breaking them down in the lysosome to produce substrates for ATP generation and anabolism. Recent studies have revealed four scavenging pathways that support cancer cell proliferation in low-nutrient environments: scavenging of extracellular matrix proteins via integrins, receptor-mediated albumin uptake and catabolism, macropinocytic consumption of multiple components of the tumour microenvironment and the engulfment and degradation of entire live cells via entosis. New evidence suggests that blocking these pathways alone or in combination could provide substantial benefits to patients with incurable solid tumours. Both US Food and Drug Administration (FDA)-approved drugs and several agents in preclinical or clinical development shut down individual or multiple scavenging pathways. These therapies may increase the extent and durability of tumour growth inhibition and/or prevent the development of resistance when used in combination with existing treatments. This Review summarizes the evidence suggesting that scavenging pathways drive tumour growth, highlights recent advances that define the oncogenic signal transduction pathways that regulate scavenging and considers the benefits and detriments of therapeutic strategies targeting scavenging that are currently under development

Starving PTEN-deficient prostate cancer cells thrive under nutrient stress by scavenging corpses for their supper.

Our recent work demonstrates that inactivating mutations in phosphatase and tensin homolog (PTEN) are sufficient to drive macropinocytosis in the context of AMP-activated protein kinase (AMPK) activation. Given that blocking macropinocytosis limits PTEN-deficient prostate tumor growth, AMPK or phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) inhibitors could have therapeutic value in castration-resistant prostate cancer patients, particularly when used in combination with standard of care therapies. Abbreviations: ATG5: autophagy related 5; NHE: Na(+)/H(+) exchanger; PAK1: p21-activated kinase 1; PI3K: phosphatidylinositol-4,5-bisphosphate 3-kinase; PIP3: phosphatidylinositol (3,4,5)-trisphosphate; PIP2: phosphatidylinositol (4,5)-bisphosphate; RAC1: Rac family small GTPase 1

Characterization of the Zebrafish Homolog of Zipper Interacting Protein Kinase

Zipper-interacting protein kinase (ZIPK) is a conserved vertebrate-specific regulator of actomyosin contractility in smooth muscle and non-muscle cells. Murine ZIPK has undergone an unusual divergence in sequence and regulation compared to other ZIPK orthologs. In humans, subcellular localization is controlled by phosphorylation of threonines 299 and 300. In contrast, ZIPK subcellular localization in mouse and rat is controlled by interaction with PAR-4. We carried out a comparative biochemical characterization of the regulation of the zebrafish ortholog of ZIPK. Like the human orthologs zebrafish ZIPK undergoes nucleocytoplasmic-shuttling and is abundant in the cytoplasm, unlike the primarily nuclear rat ZIPK. Rat ZIPK, but not human or zebrafish ZIPK, interacts with zebrafish PAR-4. Mutation of the conserved residues required for activation of the mammalian orthologs abrogated activity of the zebrafish ZIPK. In contrast to the human ortholog, mutation of threonine 299 and 300 in the zebrafish ZIPK has no effect on the activity or subcellular localization. Thus, we found that zebrafish ZIPK functions in a manner most similar to the human ZIPK and quite distinct from murine orthologs, yet the regulation of subcellular localization is not conserved