RESUMEN
Metabolic reprogramming is a necessary component of oncogenesis and cancer progression that solid tumors undergo when their growth outstrips local nutrient supply. The supply of lipids such as cholesterol and fatty acids is required for continued tumor cell proliferation, and oncogenic mutations stimulate de novo lipogenesis to support tumor growth. Sterol regulatory element-binding protein (SREBP) transcription factors control cellular lipid homeostasis by activating genes required for lipid synthesis and uptake. SREBPs have been implicated in the progression of multiple cancers, including brain, breast, colon, liver, and prostate. However, the role the SREBP pathway and its central regulator SREBP cleavage activating protein (SCAP) in pancreatic ductal adenocarcinoma (PDAC) has not been studied in detail. Here, we demonstrated that pancreas-specific knockout of Scap has no effect on mouse pancreas development or function, allowing for examination of the role for Scap in the murine KPC model of PDAC. Notably, heterozygous loss of Scap prolonged survival in KPC mice, and homozygous loss of Scap impaired PDAC tumor progression. Using subcutaneous and orthotopic xenograft models, we showed that S CAP is required for human PDAC tumor growth. Mechanistically, chemical or genetic inhibition of the SREBP pathway prevented PDAC cell growth under low serum conditions due to a lack of lipid supply. Highlighting the clinical importance of this pathway, the SREBP pathway is broadly required for cancer cell growth, SREBP target genes are upregulated in human PDAC tumors, and increased expression of SREBP targets genes is associated with poor survival in PDAC patients. Collectively, these results demonstrate that SCAP and the SREBP pathway activity are essential for PDAC cell and tumor growth in vitro and in vivo , identifying SCAP as a potential therapeutic target for PDAC. SIGNIFICANCE: Our findings demonstrate that SREBP pathway activation is a critical part of the metabolic reprogramming that occurs in PDAC development and progression. Therefore, targeting the SREBP pathway has significant therapeutic potential.
RESUMEN
Sterol regulatory element-binding proteins (SREBPs) are a family of membrane-bound transcription factors that regulate the uptake and synthesis of cholesterol and fatty acids in mammalian cells. SREBP cleavage-activating protein (SCAP) is an endoplasmic reticulum (ER) protein that binds newly synthesized SREBP, retaining it in the ER where SREBP is inactive. SCAP binds cholesterol and functions as the cholesterol sensor in this regulatory system. Under low cholesterol conditions, SCAP escorts SREBP from the ER to the Golgi apparatus where two proteases sequentially cleave and activate SREBP. Given their central importance in maintaining cellular lipid homeostasis, other mechanisms exist to regulate SREBP activity, such as control of protein synthesis and degradation. Here, we describe methods to assay ER-to-Golgi transport of SCAP in vitro using immunofluorescence microscopy and two different cell systems, Chinese hamster ovary (CHO) cells stably expressing hamster GFP-SCAP and human HeLa cells transiently expressing human GFP-SCAP. These methods will permit investigators to determine if cellular perturbations act by affecting the ER-to-Golgi transport of SCAP.
Asunto(s)
Péptidos y Proteínas de Señalización Intracelular , Esteroles , Cricetinae , Animales , Humanos , Esteroles/metabolismo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Células CHO , Células HeLa , Cricetulus , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Aparato de Golgi/metabolismo , Retículo Endoplásmico/metabolismo , Colesterol/metabolismo , Microscopía Fluorescente , Transporte de ProteínasRESUMEN
Purpose: The goal of this study is to enhance the efficacy of imipridones, a novel class of AKT/ERK inhibitors that displayed limited therapeutic efficacy against glioblastoma (GBM).Experimental Design: Gene set enrichment, LC/MS, and extracellular flux analyses were used to determine the mechanism of action of novel imipridone compounds, ONC206 and ONC212. Orthotopic patient-derived xenografts were utilized to evaluate therapeutic potency.Results: Imipridones reduce the proliferation of patient-derived xenograft and stem-like glioblastoma cell cultures in vitro and in multiple xenograft models in vivo ONC212 displayed the highest potency. High levels of c-myc predict susceptibility to growth inhibition and apoptosis induction by imipridones and increased host survival in orthotopic patient-derived xenografts. As early as 1 hour, imipridones elicit on-target inhibition, followed by dephosphorylation of GSK3ß at serine 9. GSK3ß promotes phosphorylation of c-myc at threonine 58 and enhances its proteasomal degradation. Moreover, inhibition of c-myc by BRD4 antagonists sensitizes for imipridone-induced apoptosis in stem-like GBM cells in vitro and in vivo Imipridones affect energy metabolism by suppressing both glycolysis and oxidative phosphorylation, which is accompanied by a compensatory activation of the serine-one carbon-glycine (SOG) pathway, involving the transcription factor ATF4. Interference with the SOG pathway through novel inhibitors of PHGDH results in synergistic cell death induction in vitro and in vivo Conclusions: These results suggest that c-myc expression predicts therapeutic responses to imipridones and that imipridones lead to suppression of tumor cell energy metabolism, eliciting unique metabolic vulnerabilities that can be exploited for clinical relevant drug combination therapies. Clin Cancer Res; 24(21); 5392-406. ©2018 AACR.