Small Molecule Inhibition of the Autophagy Kinase ULK1 and Identification of ULK1 Substrates

Egan, Daniel F.; Chun, Matthew G. H.; Vamos, Mitchell; Zou, Haixia; Rong, Juan; Miller, Chad J.; Lou, Hua; Dhanya, Raveendra-Panickar; Yang, Changjun; Sheffler, Douglas J.; Teriete, Peter; Asara, John M.; Turk, Benjamin E.; Cosford, Nicholas D. P.; Shaw, Reuben J.

doi:10.1016/j.molcel.2015.05.031

Cited by 574 publications

(662 citation statements)

References 46 publications

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“…These targets include vps34, a class III PI3Kinase specific for autophagy, ULK1, the apical kinase for autophagy, and AMPK1, a kinase that activates ULK1. 34,35 However, because of the current lack of a small-molecule inhibitor of Atg7, we used RNA interference to suppress Atg7 and to illustrate proof of concept for this study. Our results show that Atg7 knockdown in AML cells results in a proapoptotic phenotype with increased chemosensitivity that translates to improved overall survival in vivo.…”

Section: Discussionmentioning

confidence: 99%

Atg7 suppression enhances chemotherapeutic agent sensitivity and overcomes stroma-mediated chemoresistance in acute myeloid leukemia

Piya¹,

Kornblau²,

Ruvolo³

et al. 2016

Blood

104

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• Atg7 expression is associated with shorter remission duration in AML.• Atg7 inhibition is a proapoptotic phenotype and enhances sensitivity to chemotherapy.Autophagy is a cellular adaptive mechanism to stress, including that induced by chemotherapeutic agents. Reverse phase protein array suggested that high expression of the essential autophagy-related protein, Atg7, was associated with shorter remission in newly diagnosed acute myeloid leukemia (AML) patient samples, indicating a role in chemoresistance. Knockdown of Atg7 in AML cells using short hairpin RNA markedly increased apoptosis and DNA damage following treatment with cytarabine and idarubicin. Interestingly, coculture of AML cells with stromal cells increased autophagy and chemoresistance in the AML cells exposed to chemotherapeutic agents, and this was reversed following Atg7 knockdown. This effect was further enhanced by concomitant knockdown of Atg7 in both AML and stromal cells. These findings strongly suggest that Atg7, and likely microenvironment autophagy in general, plays an important role in AML chemoresistance. Mechanistic studies revealed that Atg7 knockdown induced a proapoptotic phenotype in AML cells, which was manifested by an increased NOXA expression at the transcriptional level. Finally, in a mouse model of human leukemia, Atg7 knockdown extended overall survival after chemotherapy. Thus, the inhibition of Atg7 appears to be a valid strategy to enhance chemosensitivity, and it could indeed improve outcomes in AML therapy. (Blood. 2016;128(9):1260-1269

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Section: Discussionmentioning

confidence: 99%

Atg7 suppression enhances chemotherapeutic agent sensitivity and overcomes stroma-mediated chemoresistance in acute myeloid leukemia

Piya¹,

Kornblau²,

Ruvolo³

et al. 2016

Blood

104

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“…SBI-0206965 was recently identified as a selective small molecule inhibitor of ULK1 (51). Addition of the ULK1 inhibitor at a concentration used in the prior study (51) led to a decrease in IFN-␤ expression in WT MEFs treated with DMXAA. Thus, these data indicate that ULK1 is unlikely to be a negative regulator of STING-dependent signaling (Fig.…”

Section: The Ulk1 Inhibitor Sbi-0206965 Represses Activation Of Stingmentioning

confidence: 97%

AMP-activated Kinase (AMPK) Promotes Innate Immunity and Antiviral Defense through Modulation of Stimulator of Interferon Genes (STING) Signaling

Prantner

Perkins

Vogel

2017

Journal of Biological Chemistry

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Edited by Luke O'NeillThe host protein Stimulator of Interferon Genes (STING) has been shown to be essential for recognition of both viral and intracellular bacterial pathogens, but its regulation remains unclear. Previously, we reported that mitochondrial membrane potential regulates STING-dependent IFN-␤ induction independently of ATP synthesis. Because mitochondrial membrane potential controls calcium homeostasis, and AMP-activated protein kinase (AMPK) is regulated, in part, by intracellular calcium, we postulated that AMPK participates in STING activation; however, its role has yet to be been defined. Addition of an intracellular calcium chelator or an AMPK inhibitor to either mouse macrophages or mouse embryonic fibroblasts (MEFs) suppressed IFN-␤ and TNF-␣ induction following stimulation with the STING-dependent ligand 5,6-dimethyl xanthnone-4-acetic acid (DMXAA). These pharmacological findings were corroborated by showing that MEFs lacking AMPK activity also failed to up-regulate IFN-␤ and TNF-␣ after treatment with DMXAA or the natural STING ligand cyclic GMP-AMP (cGAMP). As a result, AMPK-deficient MEFs exhibit impaired control of vesicular stomatitis virus (VSV), a virus sensed by STING that can cause an influenza-like illness in humans. This impairment could be overcome by pretreatment of AMPK-deficient MEFs with type I IFN, illustrating that de novo production of IFN-␤ in response to VSV plays a key role in antiviral defense during infection. Loss of AMPK also led to dephosphorylation at Ser-555 of the known STING regulator, UNC-51-like kinase 1 (ULK1). However, ULK1-deficient cells responded normally to DMXAA, indicating that AMPK promotes STING-dependent signaling independent of ULK1 in mouse cells.Pathogen sensing by innate immune cells is essential for host defense in response to invading microorganisms. Recognition by pattern recognition receptors typically activates signal transduction pathways, leading to up-regulation of cytokines like TNF-␣ and IFN-␤. Although IFN-␤ has a well established role in defense against viral infection, intracellular bacterial infections also induce this cytokine. In the past decade, our understanding of the molecular basis for these signaling pathways has expanded greatly and it has been discovered that there is some overlap between proteins responsible for recognition of bacteria and virus alike. One particular example is the mitochondrial and endoplasmic reticulum resident protein stimulator of interferon genes (STING) 2 (1-3). STING has been shown to be crucial for recognition of both DNA viruses (4 -6) and intracellular bacterial pathogens (7-12). STING senses DNA viruses indirectly, requiring the host enzyme cyclic GMP-AMP synthase (cGAS) to convert the viral double stranded DNA into the compound cGAMP (13-15). This metazoan second messenger, in turn, binds with high affinity to STING dimers, changing the protein conformation (16), such that it leads to activation of the kinase TBK1 and subsequent phosphorylation of the transcription factor IRF3, which is piv...

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“…Several groups have developed ATP-competitive inhibitors of ULK1 kinase (the apical kinase important for initiating autophagosome formation) that block autophagy in cells in vitro (Egan et al 2015;Lazarus and Shokat 2015;Petherick et al 2015). The most characterized of these started with a FAK inhibitor that potently inhibited ULK1 function (Russell et al 2013;Egan et al 2015).…”

Section: Ulk1mentioning

confidence: 99%

Recent insights into the function of autophagy in cancer

2016

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Macroautophagy (referred to here as autophagy) is induced by starvation to capture and degrade intracellular proteins and organelles in lysosomes, which recycles intracellular components to sustain metabolism and survival. Autophagy also plays a major homeostatic role in controlling protein and organelle quality and quantity. Dysfunctional autophagy contributes to many diseases. In cancer, autophagy can be neutral, tumor-suppressive, or tumorpromoting in different contexts. Large-scale genomic analysis of human cancers indicates that the loss or mutation of core autophagy genes is uncommon, whereas oncogenic events that activate autophagy and lysosomal biogenesis have been identified. Autophagic flux, however, is difficult to measure in human tumor samples, making functional assessment of autophagy problematic in a clinical setting. Autophagy impacts cellular metabolism, the proteome, and organelle numbers and quality, which alter cell functions in diverse ways. Moreover, autophagy influences the interaction between the tumor and the host by promoting stress adaptation and suppressing activation of innate and adaptive immune responses. Additionally, autophagy can promote a cross-talk between the tumor and the stroma, which can support tumor growth, particularly in a nutrient-limited microenvironment. Thus, the role of autophagy in cancer is determined by nutrient availability, microenvironment stress, and the presence of an immune system. Here we discuss recent developments in the role of autophagy in cancer, in particular how autophagy can promote cancer through suppressing p53 and preventing energy crisis, cell death, senescence, and an anti-tumor immune response.The core autophagy machinery is encoded by autophagyrelated (ATG) genes, of which there are now >30 (Ktistakis and Tooze 2016). These genes and their protein products are regulated by numerous upstream signals, including nutrient availability, stressors, defective organelles, and pathogenic conditions. ATG gene products control the formation of the hallmark double-membrane vesicle, the autophagosome. Other genes encode cargo receptors that direct the cargo to the forming autophagosome. Control of the trafficking machinery then orchestrates fusion of the cargo-laden autophagosomes with lysosomes. Degradative enzymes, contributed by lysosomes, break down the cargo, and the products are exported into the cytoplasm, where they are recycled into metabolic and biosynthetic pathways (Rabinowitz and White 2010;Guo et al. 2016). Under normal nutrient conditions, autophagy functions at a constitutive, low basal level to maintain protein and organelle quality, quantity, and functionality. The ATG genes and autophagy are dramatically induced by starvation and other stressors, which enable cellular and organismal survival. Up-regulation of autophagy is a means to survive nutrient stress, which is conserved from yeast to mammals. More recently, there is a growing appreciation of the role played by ATG genes in modulating intracellular trafficking, endocytosis, exoc...

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Small Molecule Inhibition of the Autophagy Kinase ULK1 and Identification of ULK1 Substrates

Cited by 574 publications

References 46 publications

Atg7 suppression enhances chemotherapeutic agent sensitivity and overcomes stroma-mediated chemoresistance in acute myeloid leukemia

Atg7 suppression enhances chemotherapeutic agent sensitivity and overcomes stroma-mediated chemoresistance in acute myeloid leukemia

AMP-activated Kinase (AMPK) Promotes Innate Immunity and Antiviral Defense through Modulation of Stimulator of Interferon Genes (STING) Signaling

Recent insights into the function of autophagy in cancer

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