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Kim Y, Yeuni Y, Heo HJ, Kim ES, Myung K, Baryawno N, Kim YH, Oh CK. Solute carrier family 2 member 2 (glucose transporter 2): a common factor of hepatocyte and hepatocellular carcinoma differentiation. PLoS One 2025; 20:e0321020. [PMID: 40279337 PMCID: PMC12026939 DOI: 10.1371/journal.pone.0321020] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2024] [Accepted: 02/27/2025] [Indexed: 04/27/2025] Open
Abstract
GLUT2 (SLC2A2), a vital glucose transporter in liver, pancreas, and kidney tissues, regulates blood glucose levels and energy metabolism. Beyond its metabolic role, SLC2A2 contributes to cell differentiation and metabolic adaptation during embryogenesis and tissue regeneration. Despite its significance, the role of SLC2A2 in liver differentiation and hepatocellular carcinoma (HCC) remains underexplored. This study investigated SLC2A2's role in liver differentiation using in silico, in vitro, and in vivo approaches. Analysis of GEO datasets (GSE132606, GSE25417, GSE67848) and TCGA HCC data revealed that while SLC2A2 expression decreases with HCC progression, stemness-associated genes, including SOX2 and POU5F1, are upregulated. Zebrafish embryos injected with SLC2A2-targeting morpholino exhibited reduced expression of the liver differentiation marker fabp10a without significantly altering the hepatoblast marker hhex. In HepG2 cells, SLC2A2 knockdown increased stemness and IGF1R pathway markers, indicating a shift toward less differentiated states. These findings suggest that SLC2A2 supports liver differentiation by regulating glucose metabolism and suppressing pathways associated with stemness and malignancy. Targeting SLC2A2 may serve as a promising therapeutic strategy for liver-related diseases, particularly HCC, by addressing its dual role in differentiation and tumor progression. Further mechanistic studies are warranted to fully elucidate these processes.
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Affiliation(s)
- Yejin Kim
- Department of Convergence Medical Sciences, School of Medicine, Pusan National University, Yangsan, Republic of Korea
| | - Yu Yeuni
- Biomedical research institute, School of Medicine, Pusan National University, Yangsan, Republic of Korea
| | - Hye Jin Heo
- Department of Anatomy, School of Medicine, Pusan National University, Yangsan, Republic of Korea
| | - Eun Sun Kim
- Center for Genomic Integrity, Institute for Basic Science, Ulsan, Republic of Korea
| | - Kyungjae Myung
- Center for Genomic Integrity, Institute for Basic Science, Ulsan, Republic of Korea
| | - Ninib Baryawno
- Childhood Cancer Research Unit, Department of Women’s and Children’s Health, Karolinska Institutet, Stockholm, Sweden
| | - Yun Hak Kim
- Department of Anatomy, School of Medicine, Pusan National University, Yangsan, Republic of Korea
- Department of Biomedical Informatics, School of Medicine, Pusan National University, Yangsan, Republic of Korea
| | - Chang-Kyu Oh
- Department of Convergence Medical Sciences, School of Medicine, Pusan National University, Yangsan, Republic of Korea
- Department of Biochemistry, School of Medicine, Pusan National University, Yangsan, Republic of Korea
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Mehta D, Rajput K, Jain D, Bajaj A, Dasgupta U. Unveiling the Role of Mechanistic Target of Rapamycin Kinase (MTOR) Signaling in Cancer Progression and the Emergence of MTOR Inhibitors as Therapeutic Strategies. ACS Pharmacol Transl Sci 2024; 7:3758-3779. [PMID: 39698262 PMCID: PMC11650738 DOI: 10.1021/acsptsci.4c00530] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2024] [Revised: 11/08/2024] [Accepted: 11/18/2024] [Indexed: 12/20/2024]
Abstract
The mechanistic target of rapamycin kinase (MTOR) is pivotal for cell growth, metabolism, and survival. It functions through two distinct complexes, mechanistic TORC1 and mechanistic TORC2 (mTORC1 and mTORC2). These complexes function in the development and progression of cancer by regulating different cellular processes, such as protein synthesis, lipid metabolism, and glucose homeostasis. The mTORC1 complex senses nutrients and initiates proliferative signals, and mTORC2 is crucial for cell survival and cytoskeletal rearrangements. mTORC1 and mTORC2 have therefore emerged as potential targets for cancer treatment. Several mTOR inhibitors, including rapamycin and its analogs (rapalogs), primarily target mTORC1 and are effective for specific cancer types. However, these inhibitors often lead to resistance and limited long-term advantages due to the activation of survival pathways through feedback mechanisms. Researchers have created next-generation inhibitors targeting mTORC1 and mTORC2 and dual PI3K/mTOR inhibitors to address these difficulties. These inhibitors demonstrate enhanced anti-tumor effects by simultaneously disrupting multiple signaling pathways and show promise for improved and long-lasting therapies. However, development of resistance and adverse side effects remain a significant obstacle. Recent additions known as RapaLinks have emerged as a boon to counter drug-resistant cancer cells, as they are more potent and provide a more comprehensive blockade of mTOR signaling pathways. This Review combines current research findings and clinical insights to enhance our understanding of the crucial role of mTOR signaling in cancer biology and highlights the evolution of mTOR inhibitors as promising therapeutic approaches.
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Affiliation(s)
- Devashish Mehta
- Amity
Institute of Integrative Sciences and Health, Amity University Haryana, Panchgaon, Manesar, Gurgaon-122413, Haryana, India
| | - Kajal Rajput
- Amity
Institute of Integrative Sciences and Health, Amity University Haryana, Panchgaon, Manesar, Gurgaon-122413, Haryana, India
| | - Dolly Jain
- Laboratory
of Nanotechnology and Chemical Biology, Regional Centre for Biotechnology, NCR Biotech Science Cluster, 3rd Milestone Faridabad-Gurgaon
Expressway, Faridabad-121001, Haryana, India
| | - Avinash Bajaj
- Laboratory
of Nanotechnology and Chemical Biology, Regional Centre for Biotechnology, NCR Biotech Science Cluster, 3rd Milestone Faridabad-Gurgaon
Expressway, Faridabad-121001, Haryana, India
| | - Ujjaini Dasgupta
- Amity
Institute of Integrative Sciences and Health, Amity University Haryana, Panchgaon, Manesar, Gurgaon-122413, Haryana, India
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Cervantes-Villagrana RD, García-Jiménez I, Vázquez-Prado J. Guanine nucleotide exchange factors for Rho GTPases (RhoGEFs) as oncogenic effectors and strategic therapeutic targets in metastatic cancer. Cell Signal 2023; 109:110749. [PMID: 37290677 DOI: 10.1016/j.cellsig.2023.110749] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2023] [Revised: 05/11/2023] [Accepted: 06/01/2023] [Indexed: 06/10/2023]
Abstract
Metastatic cancer cells dynamically adjust their shape to adhere, invade, migrate, and expand to generate secondary tumors. Inherent to these processes is the constant assembly and disassembly of cytoskeletal supramolecular structures. The subcellular places where cytoskeletal polymers are built and reorganized are defined by the activation of Rho GTPases. These molecular switches directly respond to signaling cascades integrated by Rho guanine nucleotide exchange factors (RhoGEFs), which are sophisticated multidomain proteins that control morphological behavior of cancer and stromal cells in response to cell-cell interactions, tumor-secreted factors and actions of oncogenic proteins within the tumor microenvironment. Stromal cells, including fibroblasts, immune and endothelial cells, and even projections of neuronal cells, adjust their shapes and move into growing tumoral masses, building tumor-induced structures that eventually serve as metastatic routes. Here we review the role of RhoGEFs in metastatic cancer. They are highly diverse proteins with common catalytic modules that select among a variety of homologous Rho GTPases enabling them to load GTP, acquiring an active conformation that stimulates effectors controlling actin cytoskeleton remodeling. Therefore, due to their strategic position in oncogenic signaling cascades, and their structural diversity flanking common catalytic modules, RhoGEFs possess unique characteristics that make them conceptual targets of antimetastatic precision therapies. Preclinical proof of concept, demonstrating the antimetastatic effect of inhibiting either expression or activity of βPix (ARHGEF7), P-Rex1, Vav1, ARHGEF17, and Dock1, among others, is emerging.
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Collins SE, Wiegand ME, Werner AN, Brown IN, Mundo MI, Swango DJ, Mouneimne G, Charest PG. Ras-mediated activation of mTORC2 promotes breast epithelial cell migration and invasion. Mol Biol Cell 2023; 34:ar9. [PMID: 36542482 PMCID: PMC9930525 DOI: 10.1091/mbc.e22-06-0236] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Revised: 12/07/2022] [Accepted: 12/15/2022] [Indexed: 12/24/2022] Open
Abstract
We previously identified the mechanistic target of rapamycin complex 2 (mTORC2) as an effector of Ras for the control of directed cell migration in Dictyostelium. Recently, the Ras-mediated regulation of mTORC2 was found to be conserved in mammalian cells, and mTORC2 was shown to be an effector of oncogenic Ras. Interestingly, mTORC2 has been linked to cancer cell migration, and particularly in breast cancer. Here, we investigated the role of Ras in promoting the migration and invasion of breast cancer cells through mTORC2. We observed that both Ras and mTORC2 promote the migration of different breast cancer cells and breast cancer cell models. Using HER2 and oncogenic Ras-transformed breast epithelial MCF10A cells, we found that both wild-type Ras and oncogenic Ras promote mTORC2 activation and an mTORC2-dependent migration and invasion in these breast cancer models. We further observed that, whereas oncogenic Ras-transformed MCF10A cells display uncontrolled cell proliferation and invasion, disruption of mTORC2 leads to loss of invasiveness only. Together, our findings suggest that, whereas the Ras-mediated activation of mTORC2 is expected to play a minor role in breast tumor formation, the Ras-mTORC2 pathway plays an important role in promoting the migration and invasion of breast cancer cells.
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Affiliation(s)
- Shannon E. Collins
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
| | - Mollie E. Wiegand
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
| | - Alyssa N. Werner
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
| | - Isabella N. Brown
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
| | - Mary I. Mundo
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
| | - Douglas J. Swango
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
| | - Ghassan Mouneimne
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721
| | - Pascale G. Charest
- Department of Molecular and Cellular Biology, University of Arizona, Tucson, AZ 85721
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Heo HJ, Park Y, Lee JH, Kim Y, Kim EK, Kim GH, Yu Y, Park SY, Seo HB, Pak K, Goh TS, Park S, Oh SO, Kwon W, Kim YH. Clinical big-data-based design of GLUT2-targeted carbon nanodots for accurate diagnosis of hepatocellular carcinoma. NANOSCALE 2022; 14:17053-17064. [PMID: 36367284 DOI: 10.1039/d2nr04238j] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/16/2023]
Abstract
Despite advances in diagnostic and therapeutic methods, the prognosis of patients with hepatocellular carcinoma (HCC) remains poor due to the delay in diagnosis. Herein, we aimed to discover a highly sensitive and specific biomarker for HCC based on genomic big data analysis and create an HCC-targeted imaging probe using carbon nanodots (CNDs) as contrast agents. In genomic analysis, we selected glucose transporter 2 (GLUT2) as a potential imaging target for HCC. We confirmed the target suitability by immunohisto-chemistry tests of 339 patient samples, where 81.1% of the patients exhibited underexpression of GLUT2, i.e., higher GLUT2 intensity in non-tumor tissues than in tumor tissues. To visualize GLUT2, we conjugated CNDs with glucosamine (GLN) as a targeting ligand to yield glucosamine-labeled CNDs (GLN-CNDs). A series of in vitro and in vivo experiments were conducted on GLUT2-modified HepG2 cells to confirm the specificity of the GLN-CNDs. Since the GLUT2 expression is higher in hepatocytes than in HCC cells, the GLUT2-targeted contrast agent is highly attached to normal cells. However, it is possible to produce images in the same form as the images obtained with a cancer cell-targeted contrast agent by inverting color scaling. Our results indicate that GLUT2 is a promising target for HCC and that GLN-CNDs may potentially be used as targeted imaging probes for diagnosing HCC.
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Affiliation(s)
- Hye Jin Heo
- Department of Anatomy, School of Medicine, Pusan National University, Yangsan 50612, Republic of Korea.
| | - Yoonsang Park
- Institute of Advanced Materials and Systems, Sookmyung Women's University, Seoul 04310, Republic of Korea.
- Nano Convergence Technology Research Center, Korea Electronics Technology Institute (KETI), Seongnam 13509, Republic of Korea
| | - Jung Hee Lee
- Department of Pathology, School of Medicine, Pusan National University, Yangsan 50612, Republic of Korea
| | - Yujin Kim
- Department of Chemical and Biological Engineering, Sookmyung Women's University, Seoul 04310, Republic of Korea
| | - Eun Kyoung Kim
- Department of Anatomy, School of Medicine, Pusan National University, Yangsan 50612, Republic of Korea.
| | - Ga Hyun Kim
- Interdisciplinary Program of Genomic Data Science, Pusan National University, Yangsan 50612, Republic of Korea
| | - Yeuni Yu
- Biomedical Research Institute, Pusan National University Hospital, Yangsan 50612, Republic of Korea.
| | - So Youn Park
- Gene & Cell Therapy Research Center for Vessel-associated Diseases, Pusan National University, Yangsan 50612, Republic of Korea
| | - Hie Bum Seo
- Department of Radiology, School of Medicine, Pusan National University, Pusan National University Hospital, Yangsan 50612, Republic of Korea
| | - Kyoungjune Pak
- Biomedical Research Institute, Pusan National University Hospital, Yangsan 50612, Republic of Korea.
- Department of Nuclear Medicine, Pusan National University Hospital, Yangsan 50612, Republic of Korea
| | - Tae Sik Goh
- Biomedical Research Institute, Pusan National University Hospital, Yangsan 50612, Republic of Korea.
- Department of Orthopaedic Surgery, Pusan National University Hospital, Yangsan 50612, Republic of Korea
| | - Sehyeon Park
- Department of Chemical and Biological Engineering, Sookmyung Women's University, Seoul 04310, Republic of Korea
| | - Sae-Ock Oh
- Department of Anatomy, School of Medicine, Pusan National University, Yangsan 50612, Republic of Korea.
| | - Woosung Kwon
- Institute of Advanced Materials and Systems, Sookmyung Women's University, Seoul 04310, Republic of Korea.
- Department of Chemical and Biological Engineering, Sookmyung Women's University, Seoul 04310, Republic of Korea
| | - Yun Hak Kim
- Department of Anatomy, School of Medicine, Pusan National University, Yangsan 50612, Republic of Korea.
- Biomedical Research Institute, Pusan National University Hospital, Yangsan 50612, Republic of Korea.
- Department of Biomedical Informatics, School of Medicine, Pusan National University, Yangsan 50612, Republic of Korea
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Codenotti S, Zizioli D, Mignani L, Rezzola S, Tabellini G, Parolini S, Giacomini A, Asperti M, Poli M, Mandracchia D, Vezzoli M, Bernardi S, Russo D, Mitola S, Monti E, Triggiani L, Tomasini D, Gastaldello S, Cassandri M, Rota R, Marampon F, Fanzani A. Hyperactive Akt1 Signaling Increases Tumor Progression and DNA Repair in Embryonal Rhabdomyosarcoma RD Line and Confers Susceptibility to Glycolysis and Mevalonate Pathway Inhibitors. Cells 2022; 11:cells11182859. [PMID: 36139434 PMCID: PMC9497225 DOI: 10.3390/cells11182859] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2022] [Revised: 09/06/2022] [Accepted: 09/09/2022] [Indexed: 11/18/2022] Open
Abstract
In pediatric rhabdomyosarcoma (RMS), elevated Akt signaling is associated with increased malignancy. Here, we report that expression of a constitutively active, myristoylated form of Akt1 (myrAkt1) in human RMS RD cells led to hyperactivation of the mammalian target of rapamycin (mTOR)/70-kDa ribosomal protein S6 kinase (p70S6K) pathway, resulting in the loss of both MyoD and myogenic capacity, and an increase of Ki67 expression due to high cell mitosis. MyrAkt1 signaling increased migratory and invasive cell traits, as detected by wound healing, zymography, and xenograft zebrafish assays, and promoted repair of DNA damage after radiotherapy and doxorubicin treatments, as revealed by nuclear detection of phosphorylated H2A histone family member X (γH2AX) through activation of DNA-dependent protein kinase (DNA-PK). Treatment with synthetic inhibitors of phosphatidylinositol-3-kinase (PI3K) and Akt was sufficient to completely revert the aggressive cell phenotype, while the mTOR inhibitor rapamycin failed to block cell dissemination. Furthermore, we found that pronounced Akt1 signaling increased the susceptibility to cell apoptosis after treatments with 2-deoxy-D-glucose (2-DG) and lovastatin, enzymatic inhibitors of hexokinase, and 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGCR), especially in combination with radiotherapy and doxorubicin. In conclusion, these data suggest that restriction of glucose metabolism and the mevalonate pathway, in combination with standard therapy, may increase therapy success in RMS tumors characterized by a dysregulated Akt signaling.
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Affiliation(s)
- Silvia Codenotti
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Daniela Zizioli
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Luca Mignani
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Sara Rezzola
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Giovanna Tabellini
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Silvia Parolini
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Arianna Giacomini
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Michela Asperti
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Maura Poli
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Delia Mandracchia
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Marika Vezzoli
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Simona Bernardi
- Department of Clinical and Experimental Sciences, ASST Spedali Civili di Brescia, University of Brescia, 25123 Brescia, Italy
| | - Domenico Russo
- Department of Clinical and Experimental Sciences, ASST Spedali Civili di Brescia, University of Brescia, 25123 Brescia, Italy
| | - Stefania Mitola
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Eugenio Monti
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
| | - Luca Triggiani
- Radiation Oncology Department, ASST Spedali Civili di Brescia, University of Brescia, 25123 Brescia, Italy
| | - Davide Tomasini
- Radiation Oncology Department, ASST Spedali Civili di Brescia, University of Brescia, 25123 Brescia, Italy
| | - Stefano Gastaldello
- Department of Physiology and Pharmacology, Karolinska Institutet, 17177 Stockholm, Sweden
- Precision Medicine Research Center, School of Pharmacy, Binzhou Medical University, Laishan District, Guanhai Road 346, Yantai 264003, China
| | - Matteo Cassandri
- Department of Hematology and Oncology, Cell and Gene Therapy, Bambino Gesù Children’s Hospital, IRCCS, 00165 Rome, Italy
- Department of Radiotherapy, Policlinico Umberto I, “Sapienza” University of Rome, 00161 Rome, Italy
| | - Rossella Rota
- Department of Hematology and Oncology, Cell and Gene Therapy, Bambino Gesù Children’s Hospital, IRCCS, 00165 Rome, Italy
| | - Francesco Marampon
- Department of Radiotherapy, Policlinico Umberto I, “Sapienza” University of Rome, 00161 Rome, Italy
| | - Alessandro Fanzani
- Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
- Correspondence: ; Tel.: +39-030-3717567
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Ha JM, Jin SY, Lee HS, Kum HJ, Vafaeinik F, Ha HK, Song SH, Kim CD, Bae SS. Akt1-dependent expression of angiopoietin 1 and 2 in vascular smooth muscle cells leads to vascular stabilization. EXPERIMENTAL & MOLECULAR MEDICINE 2022; 54:1133-1145. [PMID: 35931736 PMCID: PMC9440121 DOI: 10.1038/s12276-022-00819-8] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/07/2022] [Revised: 05/23/2022] [Accepted: 05/25/2022] [Indexed: 11/09/2022]
Abstract
Retinal angiogenesis was delayed in VSMC-specific Akt1-deficient mice (Akt1∆SMC) but not in Akt2∆SMC mice. The proliferation of ECs, recruitment of pericytes, and coverage of VSMCs to the endothelium were defective in Akt1∆SMC. The silencing of Akt1 in VSMCs led to the downregulation of angiopoietin 1 (Ang1) and the upregulation of Ang2. The activation of Notch3 in VSMCs was significantly reduced in the retinas of Akt1∆SMC mice. Silencing Akt1 suppressed the activation of Notch3. Moreover, the silencing of Notch3 downregulated Ang1, whereas the overexpression of Notch3 intracellular domain (NICD3) enhanced Ang1 expression. The nuclear localization and transcriptional activity of yes-associated protein (YAP) were affected by the expression level of Akt1. Silencing YAP downregulated Ang2 expression, whereas overexpression of YAP showed the opposite results. Ang1 antibody and Ang2 suppressed endothelial sprouting of wild-type aortic tissues, whereas the Ang2 antibody and Ang1 facilitated the endothelial sprouting of aortic tissues from Akt1∆SMC mice. Finally, severe hemorrhage was observed in Akt1∆SMC mice, which was further facilitated under streptozotocin (STZ)-induced diabetic conditions. Therefore, the Akt1-Notch3/YAP-Ang1/2 signaling cascade in VSMCs might play an essential role in the paracrine regulation of endothelial function.
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Affiliation(s)
- Jung Min Ha
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Seo Yeon Jin
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Hye Sun Lee
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Hye Jin Kum
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Farzaneh Vafaeinik
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Hong Koo Ha
- Department of Urology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Sang Heon Song
- Department of Internal Medicine, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Chi Dae Kim
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea
| | - Sun Sik Bae
- Gene and Cell Therapy Research Center for Vessel-Associated Disease, Medical Research Institute, and Department of Pharmacology, Pusan National University School of Medicine, Yangsan, 50612, Republic of Korea.
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8
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Beltrán-Navarro YM, Reyes-Cruz G, Vázquez-Prado J. P-Rex1 Signaling Hub in Lower Grade Glioma Patients, Found by In Silico Data Mining, Correlates With Reduced Survival and Augmented Immune Tumor Microenvironment. Front Oncol 2022; 12:922025. [PMID: 35875157 PMCID: PMC9300953 DOI: 10.3389/fonc.2022.922025] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2022] [Accepted: 06/02/2022] [Indexed: 11/21/2022] Open
Abstract
Systematic analysis of tumor transcriptomes, combined with deep genome sequencing and detailed clinical assessment of hundreds of patients, constitutes a powerful strategy aimed to identify potential biomarkers and therapeutic targets to guide personalized treatments. Oncogenic signaling cascades are integrated by multidomain effector proteins such as P-Rex1, a guanine nucleotide exchange factor for the Rac GTPase (RacGEF), known to promote metastatic dissemination of cancer cells. We hypothesized that patients with high P-Rex1 expression and reduced survival might be characterized by a particular set of signaling proteins co-expressed with this effector of cell migration as a central component of a putative signaling hub indicative of poor prognosis. High P-Rex1 expression correlated with reduced survival of TCGA Lower Grade Glioma (LGG) patients. Thus, guided by PREX1 expression, we searched for signaling partners of this RacGEF by applying a systematic unbiased in silico data mining strategy. We identified 30 putative signaling partners that also correlated with reduced patient survival. These included GPCRs such as CXCR3, GPR82, FZD6, as well as MAP3K1, MAP2K3, NEK8, DYRK3 and RPS6KA3 kinases, and PTPN2 and PTPN22 phosphatases, among other transcripts of signaling proteins and phospho-substrates. This PREX1 signaling hub signature correlated with increased risk of shorter survival of LGG patients from independent datasets and coincided with immune and endothelial transcriptomic signatures, indicating that myeloid infiltration and tumor angiogenesis might contribute to worsen brain tumor pathology. In conclusion, P-Rex1 and its putative signaling partners in LGG are indicative of a signaling landscape of the tumor microenvironment that correlates with poor prognosis and might guide the characterization of signaling targets leading the eventual development of immunotherapeutic strategies.
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Affiliation(s)
| | | | - José Vázquez-Prado
- Department of Pharmacology, Cinvestav-IPN, Mexico City, Mexico
- *Correspondence: José Vázquez-Prado,
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9
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Ren J, Lu X, Hall G, Privratsky JR, Robson MJ, Blakely RD, Crowley SD. IL-1 receptor signaling in podocytes limits susceptibility to glomerular damage. Am J Physiol Renal Physiol 2022; 322:F164-F174. [PMID: 34894725 PMCID: PMC8782651 DOI: 10.1152/ajprenal.00353.2021] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2021] [Revised: 11/22/2021] [Accepted: 12/07/2021] [Indexed: 02/03/2023] Open
Abstract
Interleukin (IL)-1 receptor type 1 (IL-1R1) activation triggers a proinflammatory signaling cascade that can exacerbate kidney injury. However, the functions of podocyte IL-1R1 in glomerular disease remain unclear. To study the role of IL-1R1 signaling in podocytes, we selectively ablated podocyte IL-1R1 in mice (PKO mice). We then subjected PKO mice and wild-type controls to two glomerular injury models: nephrotoxic serum (NTS)- and adriamycin-induced nephropathy. Surprisingly, we found that IL-1R1 activation in podocytes limited albuminuria and podocyte injury during NTS- and adriamycin-induced nephropathy. Moreover, deletion of IL-1R1 in podocytes drove podocyte apoptosis and glomerular injury through diminishing Akt activation. Activation of Akt signaling abrogated the differences in albuminuria and podocyte injury between wild-type and PKO mice during NTS. Thus, IL-1R1 signaling in podocytes limits susceptibility to glomerular injury via an Akt-dependent signaling pathway. These data identify an unexpected protective role for IL-1R1 signaling in podocytes in the pathogenesis of glomerular disease.NEW & NOTEWORTHY The present study establishes that activation of the receptor for interleukin-1 limits susceptibility to damage to the kidney glomerulus in preclinical mouse models by stimulating Akt signaling cascades inside the podocyte.
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Affiliation(s)
- Jiafa Ren
- Division of Nephrology, Department of Medicine, Durham Veterans Affairs and Duke University Medical Centers, Durham, North Carolina
- Department of Nephrology, The First Affiliated Hospital of Nanjing Medical University, Nanjing Medical University, Nanjing, People's Republic of China
| | - Xiaohan Lu
- Division of Nephrology, Department of Medicine, Durham Veterans Affairs and Duke University Medical Centers, Durham, North Carolina
| | - Gentzon Hall
- Division of Nephrology, Department of Medicine, Durham Veterans Affairs and Duke University Medical Centers, Durham, North Carolina
| | - Jamie R Privratsky
- Department of Anesthesiology, Durham Veterans Affairs and Duke University Medical Centers, Durham, North Carolina
| | - Matthew J Robson
- Division of Pharmaceutical Sciences, James L. Winkle College of Pharmacy, University of Cincinnati, Cincinnati, Ohio
| | - Randy D Blakely
- Department of Biomedical Science, Charles E. Schmidt College of Medicine and FAU Brain Institute, Jupiter, Florida
| | - Steven D Crowley
- Division of Nephrology, Department of Medicine, Durham Veterans Affairs and Duke University Medical Centers, Durham, North Carolina
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10
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Fujita S, Horitani E, Miyashita Y, Fujita Y, Fukui K, Kamada Y, Mineo I, Asano Y, Iwahashi H, Kozawa J, Shimomura I. Whole-exome sequencing analysis of a Japanese patient with hyperinsulinemia and liver dysfunction. J Endocr Soc 2022; 6:bvac008. [PMID: 35187381 PMCID: PMC8852682 DOI: 10.1210/jendso/bvac008] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/16/2021] [Indexed: 11/30/2022] Open
Abstract
Hyperinsulinemia is often observed in obese subjects because of insulin resistance, but it may occur in nonobese subjects with unknown etiology. A 72-year-old man was admitted to our hospital for the examination of hyperinsulinemia, reactive hypoglycemia, and liver dysfunction. The patient’s body mass index was 23.7 kg/m2, but he had an elevated visceral fat area (125 cm2). His laboratory data showed mildly elevated liver enzymes, whereas plasma fasting glucose and serum insulin levels were 91 mg/dL and 52.3 μU/mL, respectively. In a 75-g oral glucose tolerance test, the serum insulin level reached the highest value of 1124 μU/mL at 180 minutes. There was no obvious etiology except for mild liver steatosis shown by liver biopsy. We suspected genetic abnormalities related to hyperinsulinemia. We performed whole-exome sequencing (WES) analyses and identified a heterozygous nonsense variant p.R924X in the insulin receptor (INSR) gene, a novel heterozygous missense variant p.V416M in the AKT1 gene, and a novel hemizygous missense variant p.R310Q in the PHKA2 gene, which is the causative gene of hepatic injury as glycogen storage disease type IX. It was speculated that the INSR gene variant, in addition to visceral fat accumulation, was the main cause of hyperinsulinemia and reactive hypoglycemia, and the remaining 2 variants were also partly responsible for hyperinsulinemia. WES analysis revealed candidate gene variants of hyperinsulinemia and hepatic-type glycogenosis. Thus, WES analysis may be a useful tool for clarifying the etiology when unexplained genetic pathophysiological conditions are suspected.
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Affiliation(s)
- Shingo Fujita
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Emi Horitani
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Yohei Miyashita
- Department of Legal Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Yukari Fujita
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
- Department of Community Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Kenji Fukui
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Yoshihiro Kamada
- Department of Advanced Metabolic Hepatology, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Ikuo Mineo
- Diabetes Center, Toyonaka Municipal Hospital, 4-14-1 Shibahara, Toyonaka, Osaka, 560-8565, Japan
| | - Yoshihiro Asano
- Department of Cardiovascular Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Hiromi Iwahashi
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
- Diabetes Center, Toyonaka Municipal Hospital, 4-14-1 Shibahara, Toyonaka, Osaka, 560-8565, Japan
- Department of Diabetes Care Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Junji Kozawa
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
- Department of Diabetes Care Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
| | - Iichiro Shimomura
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2-B5 Yamada-oka, Suita, Osaka, 565-0871, Japan
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11
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Joechle K, Guenzle J, Hellerbrand C, Strnad P, Cramer T, Neumann UP, Lang SA. Role of mammalian target of rapamycin complex 2 in primary and secondary liver cancer. World J Gastrointest Oncol 2021; 13:1632-1647. [PMID: 34853640 PMCID: PMC8603445 DOI: 10.4251/wjgo.v13.i11.1632] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/25/2021] [Revised: 04/30/2021] [Accepted: 08/16/2021] [Indexed: 02/06/2023] Open
Abstract
The mammalian target of rapamycin (mTOR) acts in two structurally and functionally distinct protein complexes, mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). Upon deregulation, activated mTOR signaling is associated with multiple processes involved in tumor growth and metastasis. Compared with mTORC1, much less is known about mTORC2 in cancer, mainly because of the unavailability of a selective inhibitor. However, existing data suggest that mTORC2 with its two distinct subunits Rictor and mSin1 might play a more important role than assumed so far. It is one of the key effectors of the PI3K/AKT/mTOR pathway and stimulates cell growth, cell survival, metabolism, and cytoskeletal organization. It is not only implicated in tumor progression, metastasis, and the tumor microenvironment but also in resistance to therapy. Rictor, the central subunit of mTORC2, was found to be upregulated in different kinds of cancers and is associated with advanced tumor stages and a bad prognosis. Moreover, AKT, the main downstream regulator of mTORC2/Rictor, is one of the most highly activated proteins in cancer. Primary and secondary liver cancer are major problems for current cancer therapy due to the lack of specific medical treatment, emphasizing the need for further therapeutic options. This review, therefore, summarizes the role of mTORC2/Rictor in cancer, with special focus on primary liver cancer but also on liver metastases.
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Affiliation(s)
- Katharina Joechle
- Department of General, Visceral and Transplantation Surgery, University Hospital Rheinisch-Westfälisch Technische Hochschule Aachen, Aachen 52074, Germany
| | - Jessica Guenzle
- Department of General and Visceral Surgery, Medical Center-University of Freiburg, Faculty of Medicine, University of Freiburg, Freiburg 79106, Germany
| | - Claus Hellerbrand
- Institute of Biochemistry, Friedrich-Alexander University Erlangen-Nürnberg, Erlangen 91054, Germany
| | - Pavel Strnad
- Department of Internal Medicine III, University Hospital Rheinisch-Westfälisch Technische Hochschule Aachen, Aachen 52074, Germany
| | - Thorsten Cramer
- Department of General, Visceral and Transplantation Surgery, University Hospital Rheinisch-Westfälisch Technische Hochschule Aachen, Aachen 52074, Germany
| | - Ulf Peter Neumann
- Department of General, Visceral and Transplantation Surgery, University Hospital Rheinisch-Westfälisch Technische Hochschule Aachen, Aachen 52074, Germany
| | - Sven Arke Lang
- Department of General, Visceral and Transplantation Surgery, University Hospital Rheinisch-Westfälisch Technische Hochschule Aachen, Aachen 52074, Germany
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12
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Rocconi RP, Wilhite AM, Schambeau L, Scalici J, Pannell L, Finan MA. A novel proteomic-based screening method for ovarian cancer using cervicovaginal fluids: A window into the abdomen. Gynecol Oncol 2021; 164:181-186. [PMID: 34756750 DOI: 10.1016/j.ygyno.2021.10.083] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2021] [Revised: 10/19/2021] [Accepted: 10/20/2021] [Indexed: 11/25/2022]
Abstract
OBJECTIVE Our objective is to develop a site-specific proteomic-based screening test for ovarian cancer(OC) using the mucus of the cervix and vagina and evaluate a potential means for home testing. METHODS Cervicovaginal fluid samples were obtained from ovarian cancer and normal control patients for LC-mass spectrometry(MS) proteomic evaluation. Statistical modeling determined the protein panel with the highest penetrance across ovarian cancer samples. A subcohort of patients consented to provide self-collected vaginal samples at home with questionnaire on feasibility. Cohen's kappa methodology was utilized to determine agreement between physician-collected and patient-collected samples. RESULTS A total of 83 consecutive patient samples were collected prospectively (33 ovarian cancer & 50 controls). Thirty patients consented for self-collection. Using LC-MS, 30 peptides demonstrated independent statistical significance for detecting ovarian cancer. Using statistical modeling, the protein panel that determined the best predictor for detecting OC formed a "fingerprint" consisting of 5 proteins: serine proteinase inhibitor A1; periplakin; profilin1; apolipoprotein A1; and thymosin beta4-like protein. These peptides demonstrated a significant increase probability of detecting ovarian cancer with the ROC curve having an AUC of 0.86 (p = 0.00001). Physician-collected and patient-collected specimens demonstrated moderate agreement with kappa average of 0.6 with upper bound of 0.75. CONCLUSIONS Using novel site-specific collection methods, we identified an OC "fingerprint" with adequate sensitivity and specificity to warrant further evaluation in a larger cohort. Agreement of physician-collected and patient-collected samples were encouraging and could improve access to screening with a home self-collection if this screening test is validated in future studies.
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Affiliation(s)
- Rodney P Rocconi
- University of South Alabama Mitchell Cancer Institute, Mobile, AL, United States of America.
| | - Annelise M Wilhite
- University of South Alabama Mitchell Cancer Institute, Mobile, AL, United States of America
| | - Lindsay Schambeau
- University of South Alabama Mitchell Cancer Institute, Mobile, AL, United States of America
| | - Jennifer Scalici
- University of South Alabama Mitchell Cancer Institute, Mobile, AL, United States of America
| | - Lewis Pannell
- University of South Alabama Mitchell Cancer Institute, Mobile, AL, United States of America
| | - Michael A Finan
- University of South Alabama Mitchell Cancer Institute, Mobile, AL, United States of America
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13
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Merckaert T, Zwaenepoel O, Gevaert K, Gettemans J. An AKT2-specific nanobody that targets the hydrophobic motif induces cell cycle arrest, autophagy and loss of focal adhesions in MDA-MB-231 cells. Biomed Pharmacother 2020; 133:111055. [PMID: 33378961 DOI: 10.1016/j.biopha.2020.111055] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2020] [Revised: 10/21/2020] [Accepted: 11/19/2020] [Indexed: 02/06/2023] Open
Abstract
The AKT kinase family is a high-profile target for cancer therapy. Despite their high degree of homology the three AKT isoforms (AKT1, AKT2 and AKT3) are non-redundant and can even have opposing functions. Small-molecule AKT inhibitors affect all three isoforms which severely limits their usefulness as research tool or therapeutic. Using AKT2-specific nanobodies we examined the function of endogenous AKT2 in breast cancer cells. Two AKT2 nanobodies (Nb8 and Nb9) modulate AKT2 and reduce MDA-MB-231 cell viability/proliferation. Nb8 binds the AKT2 hydrophobic motif and reduces IGF-1-induced phosphorylation of this site. This nanobody also affects the phosphorylation and/or expression levels of a wide range of proteins downstream of AKT, resulting in a G0/G1 cell cycle arrest, the induction of autophagy, a reduction in focal adhesion count and loss of stress fibers. While cell cycle progression is likely to be regulated by more than one isoform, our results indicate that both the effects on autophagy and the cytoskeleton are specific to AKT2. By using an isoform-specific nanobody we were able to map a part of the AKT2 pathway. Our results confirm AKT2 and the hydrophobic motif as targets for cancer therapy. Nb8 can be used as a research tool to study AKT2 signalling events and aid in the design of an AKT2-specific inhibitor.
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Affiliation(s)
- Tijs Merckaert
- Department of Biomolecular Medicine, Faculty of Medicine and Health Sciences, Ghent University, Tech Lane Ghent Science Park 75, 9052 Ghent, Belgium; VIB-UGent Center for Medical Biotechnology, Tech Lane Ghent Science Park 75, 9052 Ghent, Belgium.
| | - Olivier Zwaenepoel
- Department of Biomolecular Medicine, Faculty of Medicine and Health Sciences, Ghent University, Tech Lane Ghent Science Park 75, 9052 Ghent, Belgium.
| | - Kris Gevaert
- Department of Biomolecular Medicine, Faculty of Medicine and Health Sciences, Ghent University, Tech Lane Ghent Science Park 75, 9052 Ghent, Belgium; VIB-UGent Center for Medical Biotechnology, Tech Lane Ghent Science Park 75, 9052 Ghent, Belgium.
| | - Jan Gettemans
- Department of Biomolecular Medicine, Faculty of Medicine and Health Sciences, Ghent University, Tech Lane Ghent Science Park 75, 9052 Ghent, Belgium.
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14
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Liu MM, Ma RH, Ni ZJ, Thakur K, Cespedes-Acuña CL, Jiang L, Wei ZJ. Apigenin 7-O-glucoside promotes cell apoptosis through the PTEN/PI3K/AKT pathway and inhibits cell migration in cervical cancer HeLa cells. Food Chem Toxicol 2020; 146:111843. [PMID: 33152472 DOI: 10.1016/j.fct.2020.111843] [Citation(s) in RCA: 77] [Impact Index Per Article: 15.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2020] [Revised: 10/25/2020] [Accepted: 10/28/2020] [Indexed: 02/08/2023]
Abstract
Epidemiologic evidence promote the inclusion of flavones in diet due to their inhibitory effects on certain types of cancers, particularly in women. Among the naturally occurring plant flavonoids, Apigenin 7-O-glucoside (AGL) is endowed with anti-inflammatory, anti-oxidant, and anti-cancer activities. However, its mechanism of action on cervical cancer, the fourth largest cancer in women, has not yet been clarified. In the current study, we have determined the effect of AGL on human cervical cancer cells and studied its molecular mechanism against cervical cancer. The results showed that AGL inhibited the proliferation of HeLa cells (IC50 was 47.26 μM at 48 h) by inducing apoptosis. Furthermore, AGL treatment caused G0/G1 phase arrest, reduced mitochondrial membrane potential (MMP), and upgraded intracellular ROS production. AGL could promote the release of cytochrome c by regulating Bcl-2 family proteins, and then activated caspase 9/3 to promote cell apoptosis. Moreover, AGL treatment promoted the expression of p16 INK4A, while inhibited the expression of Cyclin A/D/E and CDK2/6. At the same time in HeLa cells treated with AGL, the PTEN/PI3K/AKT pathway was inhibited in a concentration-dependent manner, and cell migration was also impeded correspondingly through the matrix metalloproteinase 2 and 9. Our study may provide a new research direction for harnessing the novel natural compounds in cervical cancer treatment.
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Affiliation(s)
- Miao-Miao Liu
- School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009, People's Republic of China.
| | - Run-Hui Ma
- School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009, People's Republic of China; Collaborative Innovation Center for Food Production and Safety, School of Biological Science and Engineering, North Minzu University, Yinchuan, 750021, People's Republic of China.
| | - Zhi-Jing Ni
- School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009, People's Republic of China; Collaborative Innovation Center for Food Production and Safety, School of Biological Science and Engineering, North Minzu University, Yinchuan, 750021, People's Republic of China.
| | - Kiran Thakur
- School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009, People's Republic of China; Collaborative Innovation Center for Food Production and Safety, School of Biological Science and Engineering, North Minzu University, Yinchuan, 750021, People's Republic of China.
| | | | - Li Jiang
- School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009, People's Republic of China.
| | - Zhao-Jun Wei
- School of Food and Biological Engineering, Hefei University of Technology, Hefei, 230009, People's Republic of China; Collaborative Innovation Center for Food Production and Safety, School of Biological Science and Engineering, North Minzu University, Yinchuan, 750021, People's Republic of China.
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15
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Camara AB, Brandao IA. The Role of Vitamin D and Sunlight Incidence in Cancer. Anticancer Agents Med Chem 2020; 19:1418-1436. [PMID: 30864510 DOI: 10.2174/1389557519666190312123212] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2018] [Revised: 11/19/2018] [Accepted: 02/13/2019] [Indexed: 12/17/2022]
Abstract
BACKGROUND Vitamin D (VD) deficiency affects individuals of different ages in many countries. VD deficiency may be related to several diseases, including cancer. OBJECTIVE This study aimed to review the relationship between VD deficiency and cancer. METHODS We describe the proteins involved in cancer pathogenesis and how those proteins can be influenced by VD deficiency. We also investigated a relationship between cancer death rate and solar radiation. RESULTS We found an increased bladder cancer, breast cancer, colon-rectum cancer, lung cancer, oesophagus cancer, oral cancer, ovary cancer, pancreas cancer, skin cancer and stomach cancer death rate in countries with low sunlight. It was also observed that amyloid precursor protein, ryanodine receptor, mammalian target of rapamycin complex 1, and receptor for advanced glycation end products are associated with a worse prognosis in cancer. While the Klotho protein and VD receptor are associated with a better prognosis in the disease. Nfr2 is associated with both worse and better prognosis in cancer. CONCLUSION The literature suggests that VD deficiency might be involved in cancer progression. According to sunlight data, we can conclude that countries with low average sunlight have high cancers death rate. New studies involving transcriptional and genomic data in combination with VD measurement in long-term experiments are required to establish new relationships between VD and cancer.
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Affiliation(s)
- Alice B Camara
- Department of Biophysics and Pharmacology, Bioscience Center, Federal University of Rio Grande do Norte, 59064-741, Natal/RN, Brazil
| | - Igor A Brandao
- Metrópole Digital Institute, Federal University of Rio Grande do Norte, 59078-970, Natal/RN, Brazil
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16
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Lone MUD, Miyan J, Asif M, Malik SA, Dubey P, Singh V, Singh K, Mitra K, Pandey D, Haq W, Amita H, Singh PK, Kiess W, Kaessner F, Garten A, Bhadauria S. Direct physical interaction of active Ras with mSIN1 regulates mTORC2 signaling. BMC Cancer 2019; 19:1236. [PMID: 31856761 PMCID: PMC6921532 DOI: 10.1186/s12885-019-6422-6] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2019] [Accepted: 12/02/2019] [Indexed: 11/14/2022] Open
Abstract
Background The mechanistic (or mammalian) target of rapamycin (mTOR), a Ser/Thr kinase, associates with different subunits forming two functionally distinct complexes, mTORC1 and mTORC2, regulating a diverse set of cellular functions in response to growth factors, cellular energy levels, and nutrients. The mechanisms regulating mTORC1 activity are well characterized; regulation of mTORC2 activity, however, remains obscure. While studies conducted in Dictyostelium suggest a possible role of Ras protein as a potential upstream regulator of mTORC2, definitive studies delineating the underlying molecular mechanisms, particularly in mammalian cells, are still lacking. Methods Protein levels were measured by Western blotting and kinase activity of mTORC2 was analyzed by in vitro kinase assay. In situ Proximity ligation assay (PLA) and co-immunoprecipitation assay was performed to detect protein-protein interaction. Protein localization was investigated by immunofluorescence and subcellular fractionation while cellular function of mTORC2 was assessed by assaying extent of cell migration and invasion. Results Here, we present experimental evidence in support of the role of Ras activation as an upstream regulatory switch governing mTORC2 signaling in mammalian cancer cells. We report that active Ras through its interaction with mSIN1 accounts for mTORC2 activation, while disruption of this interaction by genetic means or via peptide-based competitive hindrance, impedes mTORC2 signaling. Conclusions Our study defines the regulatory role played by Ras during mTORC2 signaling in mammalian cells and highlights the importance of Ras-mSIN1 interaction in the assembly of functionally intact mTORC2.
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Affiliation(s)
- Mehraj-U-Din Lone
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Javed Miyan
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India.,Academy of Scientific and Innovative Research (AcSIR), New Delhi, 110025, India
| | - Mohammad Asif
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Showkat A Malik
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Parul Dubey
- Department of Surgical Oncology, King George Medical University, Lucknow, Uttar Pradesh, 226003, India
| | - Varsha Singh
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Kavita Singh
- Electron Microscopy Unit, Sophisticated Analytical Instrumentation Facility, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Kalyan Mitra
- Academy of Scientific and Innovative Research (AcSIR), New Delhi, 110025, India.,Electron Microscopy Unit, Sophisticated Analytical Instrumentation Facility, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Deepali Pandey
- Medicinal and Process Chemistry Division, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Wahajul Haq
- Medicinal and Process Chemistry Division, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Himanshi Amita
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Prince Kumar Singh
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India
| | - Wieland Kiess
- Center for Pediatric Research Leipzig, University Hospital for Children and Adolescents, Faculty of Medicine, University of Leipzig, Leipzig, Germany
| | - Franziska Kaessner
- Center for Pediatric Research Leipzig, University Hospital for Children and Adolescents, Faculty of Medicine, University of Leipzig, Leipzig, Germany
| | - Antje Garten
- Center for Pediatric Research Leipzig, University Hospital for Children and Adolescents, Faculty of Medicine, University of Leipzig, Leipzig, Germany.,Institute of Metabolism and Systems Research, University of Birmingham, Birmingham, UK
| | - Smrati Bhadauria
- Division of Toxicology and Experimental Medicine, Central Drug Research Institute (CSIR), Lucknow, Uttar Pradesh, 226031, India. .,Academy of Scientific and Innovative Research (AcSIR), New Delhi, 110025, India.
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17
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Feng L, Xia B, Tian BF, Lu GB. MiR-152 influences osteoporosis through regulation of osteoblast differentiation by targeting RICTOR. PHARMACEUTICAL BIOLOGY 2019; 57:586-594. [PMID: 31492082 PMCID: PMC6747012 DOI: 10.1080/13880209.2019.1657153] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/15/2023]
Abstract
Context: Evidence suggests that microRNA (miRNA) regulate gene expression and bone tissue homoeostasis of osteoporosis. MiR-152 has found to be abnormally expressed in osteoporosis, but its role in osteoblast differentiation has not been elucidated. Objective: To understand the potential mechanism of miR-152 in osteoblast differentiation via regulation of RICTOR. Materials and methods: The expression of miR-152 and RICTOR were tested in ovariectomized rat models of osteoporosis. Primary osteoblasts and MC3T -E1 cells were assigned into four groups, namely Control, miR-152 inhibitor, miR-control and miR-152 inhibitor + siRICTOR groups. qRT PCR and Western blot were performed to detect the expressions of miR-152 and RICTOR, respectively. MTT assay was used to evaluate cell viability, and ALP activity determination and mineralization analyses were also conducted. Results: In ovariectomy-induced osteoporotic rats, miR-152 (3.06 ± 0.35) in femoral tissues increased significantly, while RICTOR (0.31 ± 0.04) decreased. Compared with Control group, miR-152 inhibitor group presented appreciable reduction of miR-152 in primary osteoblasts and MC3T3-E1 cells, as well as remarkable increases in RICTOR, p-Akt(s473)/Akt ratio, and osteogenesis-related genes, with enhanced cell viability, ALP activity and mineralization. In comparison with cells in the miR-152 inhibitor group, those in the miR-152 inhibitor + siRICTOR group had no observable difference in miR-152, but were dramatically up-regulated in RICTOR, as well as the corresponding opposite tendencies of other factors. Conclusion: Inhibiting miR-152 promoted osteoblasts differentiation and alleviated osteoporosis by up-regulating RICTOR. Therefore, miR-152 may be an essential mediator of osteoblast differentiation and a new therapeutic strategy for osteoporosis.
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Affiliation(s)
- Li Feng
- Department of Traumatic Orthopedics, Jining No. 1 People’s Hospital, Jining, China
| | - Bo Xia
- Department of Traumatic Orthopedics, Jining No. 1 People’s Hospital, Jining, China
| | - Bao-Fang Tian
- Department of Traumatic Orthopedics, Jining No. 1 People’s Hospital, Jining, China
| | - Gong-Biao Lu
- Department of Spine Surgery, Jining No, 1 People’s Hospital, Jining, China
- CONTACT Gong-Biao Lu Department of Spine Surgery, Jining No. 1 People’s Hospital, No.6, Jiankang Road, Jining, Shandong 272011, China
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18
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Kobliakov VA. The Mechanisms of Regulation of Aerobic Glycolysis (Warburg Effect) by Oncoproteins in Carcinogenesis. BIOCHEMISTRY (MOSCOW) 2019; 84:1117-1128. [DOI: 10.1134/s0006297919100018] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
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19
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Zhang S, Qian G, Zhang QQ, Yao Y, Wang D, Chen ZG, Wang LJ, Chen M, Sun SY. mTORC2 Suppresses GSK3-Dependent Snail Degradation to Positively Regulate Cancer Cell Invasion and Metastasis. Cancer Res 2019; 79:3725-3736. [PMID: 31142514 DOI: 10.1158/0008-5472.can-19-0180] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2019] [Revised: 04/23/2019] [Accepted: 05/21/2019] [Indexed: 12/11/2022]
Abstract
mTOR complex 1 (mTORC1) positively regulates cell invasion and metastasis by enhancing translation of Snail. A connection between mTOR complex 2 (mTORC2) and cell invasion and metastasis has also been suggested, yet the underlying biology or mechanism is largely unknown and thus is the focus of this study. Inhibition of mTOR with both mTOR inhibitors and knockdown of key components of mTORC, including rictor, Sin1, and raptor, decreased Snail protein levels. Inhibition of mTOR enhanced the rate of Snail degradation, which could be rescued by inhibition of the proteasome. Critically, inhibition of mTORC2 (by knocking down rictor) but not mTORC1 (by knocking down raptor) enhanced Snail degradation. Therefore, only mTORC2 inhibition induces Snail proteasomal degradation, resulting in eventual Snail reduction. Interestingly, inhibition of GSK3 but not SCF/β-TrCP rescued the Snail reduction induced by mTOR inhibitors, suggesting GSK3-dependent, but SCF/β-TrCP-independent proteasomal degradation of Snail. Accordingly, mTOR inhibitors elevated E-cadherin levels and suppressed cancer cell migration and invasion in vitro and metastasis in vivo. Collectively, this study reveals that mTORC2 positively regulates Snail stability to control cell invasion and metastasis. SIGNIFICANCE: These findings delineate a new regulation mechanism of Snail, an important master regulator of epithelial-mesenchymal transition and invasion in cancers.
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Affiliation(s)
- Shuo Zhang
- First Affiliated Hospital of Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi, P.R. China
- Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia
| | - Guoqing Qian
- Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia
| | - Qian-Qian Zhang
- Vascular Biology Research Institute, School of Basic Science, Guangdong Pharmaceutical University, Guangzhou, Guangdong, P.R. China
| | - Yuying Yao
- Vascular Biology Research Institute, School of Basic Science, Guangdong Pharmaceutical University, Guangzhou, Guangdong, P.R. China
| | - Dongsheng Wang
- Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia
| | - Zhuo G Chen
- Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia
| | - Li-Jing Wang
- Vascular Biology Research Institute, School of Basic Science, Guangdong Pharmaceutical University, Guangzhou, Guangdong, P.R. China
| | - Mingwei Chen
- First Affiliated Hospital of Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi, P.R. China
| | - Shi-Yong Sun
- Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia.
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20
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Alwhaibi A, Verma A, Adil MS, Somanath PR. The unconventional role of Akt1 in the advanced cancers and in diabetes-promoted carcinogenesis. Pharmacol Res 2019; 145:104270. [PMID: 31078742 PMCID: PMC6659399 DOI: 10.1016/j.phrs.2019.104270] [Citation(s) in RCA: 59] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/28/2018] [Revised: 04/03/2019] [Accepted: 05/08/2019] [Indexed: 12/15/2022]
Abstract
Decades of research have elucidated the critical role of Akt isoforms in cancer as pro-tumorigenic and metastatic regulators through their specific effects on the cancer cells, tumor endothelial cells and the stromal cells. The pro-cancerous role of Akt isoforms through enhanced cell proliferation and suppression of apoptosis in cancer cells and the cells in the tumor microenvironment is considered a dogma. Intriguingly, studies also indicate that the Akt pathway is essential to protect the endothelial-barrier and prevent aberrant vascular permeability, which is also integral to tumor perfusion and metastasis. To complicate this further, a flurry of recent reports strongly indicates the metastasis suppressive role of Akt, Akt1 in particular in various cancer types. These reports emanated from different laboratories have elegantly demonstrated the paradoxical effect of Akt1 on cancer cell epithelial-to-mesenchymal transition, invasion, tumor endothelial-barrier disruption, and cancer metastasis. Here, we emphasize on the specific role of Akt1 in mediating tumor cell-vasculature reciprocity during the advanced stages of cancers and discuss how Akt1 differentially regulates cancer metastasis through mechanisms distinct from its pro-tumorigenic effects. Since Akt is integral for insulin signaling, endothelial function, and metabolic regulation, we also attempt to shed some light on the specific effects of diabetes in modulating Akt pathway in the promotion of tumor growth and metastasis.
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Affiliation(s)
- Abdulrahman Alwhaibi
- Program in Clinical and Experimental Therapeutics, College of Pharmacy, University of Georgia and the Charlie Norwood VA Medical Center, Augusta, GA, USA
| | - Arti Verma
- Program in Clinical and Experimental Therapeutics, College of Pharmacy, University of Georgia and the Charlie Norwood VA Medical Center, Augusta, GA, USA
| | - Mir S Adil
- Program in Clinical and Experimental Therapeutics, College of Pharmacy, University of Georgia and the Charlie Norwood VA Medical Center, Augusta, GA, USA
| | - Payaningal R Somanath
- Program in Clinical and Experimental Therapeutics, College of Pharmacy, University of Georgia and the Charlie Norwood VA Medical Center, Augusta, GA, USA; Department of Medicine, Vascular Biology Center and Cancer Center, Augusta University, USA.
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21
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Zhang Q, Zhang Y, Chen Y, Qian J, Zhang X, Yu K. A Novel mTORC1/2 Inhibitor (MTI-31) Inhibits Tumor Growth, Epithelial-Mesenchymal Transition, Metastases, and Improves Antitumor Immunity in Preclinical Models of Lung Cancer. Clin Cancer Res 2019; 25:3630-3642. [PMID: 30796032 DOI: 10.1158/1078-0432.ccr-18-2548] [Citation(s) in RCA: 49] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2018] [Revised: 12/21/2018] [Accepted: 02/15/2019] [Indexed: 11/16/2022]
Abstract
PURPOSE We aimed to investigate efficacy and mechanism of MTI-31 (LXI-15029), a novel mTORC1/mTORC2 inhibitor currently in human trial (NCT03125746), in non-small cell lung cancer (NSCLC) models of multiple driver mutations and tyrosine kinase inhibitor (TKI)-resistance. EXPERIMENTAL DESIGN Gene depletion, inhibitor treatment, immunological, flow cytometry, cellular, and animal studies were performed to determine in vitro and in vivo efficacy in NSCLC models of driver mutations and elucidate roles by mTOR complexes in regulating migration, epithelial-mesenchymal transition (EMT), metastasis, intracranial tumor growth, and immune-escape. RESULTS MTI-31 potently inhibited cell proliferation (IC50 <1 μmol/L) and in vivo tumor growth in multiple NSCLC models of EGFR/T790M, EML4-ALK, c-Met, or KRAS (MED <10 mg/kg). In EGFR-mutant and/or EML4-ALK-driven NSCLC, MTI-31 or disruption of mTORC2 reduced cell migration, hematogenous metastasis to the lung, and abrogated morphological and functional traits of EMT. Disruption of mTORC2 inhibited EGFR/T790M-positive tumor growth in mouse brain and prolonged animal survival correlating a diminished tumor angiogenesis and recruitment of IBA1+ microglia/macrophages in tumor microenvironment. MTI-31 also suppressed programmed death ligand 1 (PD-L1) in EGFR- and ALK-driven NSCLC, mediated in part by mTORC2/AKT/GSK3β-dependent proteasomal degradation. Depletion of mTOR protein or disruption of mTOR complexes profoundly downregulated PD-L1 and alleviated apoptosis in Jurkat T and primary human T cells in a tumor-T cell coculture system. CONCLUSIONS Our results highlight mTOR as a multifaceted regulator of tumor growth, metastasis, and immune-escape in EGFR/ALK-mutant and TKI-resistant NSCLC cells. The newly characterized mechanisms mediated by the rapamycin-resistant mTORC2 warrant clinical investigation of mTORC1/mTORC2 inhibitors in patients with lung cancer.
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Affiliation(s)
- Qianwen Zhang
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Yan Zhang
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Yaqing Chen
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Jianchang Qian
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Xuesai Zhang
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Ker Yu
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China.
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22
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Adame-García SR, Cervantes-Villagrana RD, Orduña-Castillo LB, Del Rio JC, Gutkind JS, Reyes-Cruz G, Taylor SS, Vázquez-Prado J. cAMP-dependent activation of the Rac guanine exchange factor P-REX1 by type I protein kinase A (PKA) regulatory subunits. J Biol Chem 2019; 294:2232-2246. [PMID: 30530493 PMCID: PMC6378977 DOI: 10.1074/jbc.ra118.006691] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2018] [Revised: 11/29/2018] [Indexed: 12/22/2022] Open
Abstract
Regulatory subunits of protein kinase A (PKA) inhibit its kinase subunits. Intriguingly, their potential as cAMP-dependent signal transducers remains uncharacterized. We recently reported that type I PKA regulatory subunits (RIα) interact with phosphatidylinositol 3,4,5-trisphosphate-dependent Rac exchange factor 1 (P-REX1), a chemotactic Rac guanine exchange factor (RacGEF). Because P-REX1 is known to be phosphorylated and inhibited by PKA, its interaction with RIα suggests that PKA regulatory and catalytic subunits may fine-tune P-REX1 activity or those of its target pools. Here, we tested whether RIα acts as a cAMP-dependent factor promoting P-REX1-mediated Rac activation and cell migration. We observed that Gs-coupled EP2 receptors indeed promote endothelial cell migration via RIα-activated P-REX1. Expression of the P-REX1-PDZ1 domain prevented RIα/P-REX1 interaction, P-REX1 activation, and EP2-dependent cell migration, and P-REX1 silencing abrogated RIα-dependent Rac activation. RIα-specific cAMP analogs activated P-REX1, but lost this activity in RIα-knockdown cells, and cAMP pulldown assays revealed that P-REX1 preferentially interacts with free RIα. Moreover, purified RIα directly activated P-REX1 in vitro We also found that the RIα CNB-B domain is critical for the interaction with P-REX1, which was increased in RIα mutants, such as the acrodysostosis-associated mutant, that activate P-REX1 at basal cAMP levels. RIα and Cα PKA subunits targeted distinct P-REX1 molecules, indicated by an absence of phosphorylation in the active fraction of P-REX1. This was in contrast to the inactive fraction in which phosphorylated P-REX1 was present, suggesting co-existence of dual stimulatory and inhibitory effects. We conclude that PKA's regulatory subunits are cAMP-dependent signal transducers.
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Affiliation(s)
| | | | | | | | | | | | - Susan S Taylor
- the Departments of Pharmacology
- Chemistry and
- Biochemistry, University of California San Diego, La Jolla, California 92093
| | - José Vázquez-Prado
- Pharmacology, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico and
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23
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Chen J, Liu K, Liu Y, Wang X, Zhang Z. Targeting mTORC1/2 with OSI-027 inhibits proliferation and migration of keloid keratinocytes. Exp Dermatol 2019; 28:270-275. [PMID: 30650200 DOI: 10.1111/exd.13882] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2018] [Revised: 01/02/2019] [Accepted: 01/11/2019] [Indexed: 12/14/2022]
Abstract
Keloid is a dermal proliferative disorder characterized by the excessive proliferation and migration of keratinocytes and fibroblasts. Over-activation of the serine/threonine protein kinase, mammalian target of rapamycin (mTOR), plays a pivotal role in the process. Here, we show that both mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2) were hyper-activated in keloid-derived primary keratinocytes. Further, OSI-027, an mTOR kinase inhibitor, potently inhibited proliferation and migration of keloid keratinocytes. At the molecular level, OSI-027 disrupted the assembly of mTORC1 (mTOR-Raptor) and mTORC2 (mTOR-Rictor-mLST8). Further, OSI-027 almost completely blocked the phosphorylation of the mTORC1 substrates, S6K1, S6 and 4EBP1, and the mTORC2 substrate, AKT, at Ser-473. The OSI-027 treatment of keloid keratinocytes showed more effectively inhibited cell proliferation and migration compared to the mTORC1 inhibitor, rapamycin. Moreover, restoring mTORC1 activation by the introduction of the constitutively active S6K1 only partly alleviated OSI-027-induced inhibition of keloid keratinocytes. Notably, mTOR2 inhibition by Rictor siRNAs also inhibited keloid keratinocyte proliferation and migration, but less efficiently than OSI-027. Together, our results imply that concurrent targeting of mTORC1/2 by OSI-027 potently inhibits the proliferation and the migration of keloid keratinocytes. Thus, OSI-027 may have translational value for the treatment of keloid.
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Affiliation(s)
- Jun Chen
- Department of Dermatology and Dermatologic Surgery, Shanghai Ninth People's Hospital, Affiliated to Shanghai Jiaotong University School of Medicine, Center for Specialty Strategy Research of Shanghai, JiaoTong University China Hospital Development Institute, Shanghai, China
| | - Ke Liu
- Department of Dermatology and Dermatologic Surgery, Shanghai Ninth People's Hospital, Affiliated to Shanghai Jiaotong University School of Medicine, Center for Specialty Strategy Research of Shanghai, JiaoTong University China Hospital Development Institute, Shanghai, China
| | - Yang Liu
- Department of Dermatology and Dermatologic Surgery, Shanghai Ninth People's Hospital, Affiliated to Shanghai Jiaotong University School of Medicine, Center for Specialty Strategy Research of Shanghai, JiaoTong University China Hospital Development Institute, Shanghai, China
| | - Xue Wang
- Department of Dermatology and Dermatologic Surgery, Shanghai Ninth People's Hospital, Affiliated to Shanghai Jiaotong University School of Medicine, Center for Specialty Strategy Research of Shanghai, JiaoTong University China Hospital Development Institute, Shanghai, China
| | - Zhen Zhang
- Department of Dermatology and Dermatologic Surgery, Shanghai Ninth People's Hospital, Affiliated to Shanghai Jiaotong University School of Medicine, Center for Specialty Strategy Research of Shanghai, JiaoTong University China Hospital Development Institute, Shanghai, China
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24
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Cervantes-Villagrana RD, Adame-García SR, García-Jiménez I, Color-Aparicio VM, Beltrán-Navarro YM, König GM, Kostenis E, Reyes-Cruz G, Gutkind JS, Vázquez-Prado J. Gβγ signaling to the chemotactic effector P-REX1 and mammalian cell migration is directly regulated by Gα q and Gα 13 proteins. J Biol Chem 2018; 294:531-546. [PMID: 30446620 DOI: 10.1074/jbc.ra118.006254] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2018] [Revised: 11/12/2018] [Indexed: 11/06/2022] Open
Abstract
G protein-coupled receptors stimulate Rho guanine nucleotide exchange factors that promote mammalian cell migration. Rac and Rho GTPases exert opposing effects on cell morphology and are stimulated downstream of Gβγ and Gα12/13 or Gαq, respectively. These Gα subunits might in turn favor Rho pathways by preventing Gβγ signaling to Rac. Here, we investigated whether Gβγ signaling to phosphatidylinositol 3,4,5-trisphosphate-dependent Rac exchange factor 1 (P-REX1), a key Gβγ chemotactic effector, is directly controlled by Rho-activating Gα subunits. We show that pharmacological inhibition of Gαq makes P-REX1 activation by Gq/Gi-coupled lysophosphatidic acid receptors more effective. Moreover, chemogenetic control of Gi and Gq by designer receptors exclusively activated by designer drugs (DREADDs) confirmed that Gi differentially activates P-REX1. GTPase-deficient GαqQL and Gα13QL variants formed stable complexes with Gβγ, impairing its interaction with P-REX1. The N-terminal regions of these variants were essential for stable interaction with Gβγ. Pulldown assays revealed that chimeric Gα13-i2QL interacts with Gβγ unlike to Gαi2-13QL, the reciprocal chimera, which similarly to Gαi2QL could not interact with Gβγ. Moreover, Gβγ was part of tetrameric Gβγ-GαqQL-RGS2 and Gβγ-Gα13-i2QL-RGS4 complexes, whereas Gα13QL dissociated from Gβγ to interact with the PDZ-RhoGEF-RGS domain. Consistent with an integrated response, Gβγ and AKT kinase were associated with active SDF-1/CXCL12-stimulated P-REX1. This pathway was inhibited by GαqQL and Gα13QL, which also prevented CXCR4-dependent cell migration. We conclude that a coordinated mechanism prioritizes Gαq- and Gα13-mediated signaling to Rho over a Gβγ-dependent Rac pathway, attributed to heterotrimeric Gi proteins.
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Affiliation(s)
| | - Sendi Rafael Adame-García
- Cell Biology, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - Irving García-Jiménez
- Cell Biology, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | | | | | - Gabriele M König
- the University of Bonn, Institute of Pharmaceutical Biology, 53115 Bonn, Germany, and
| | - Evi Kostenis
- the University of Bonn, Institute of Pharmaceutical Biology, 53115 Bonn, Germany, and
| | - Guadalupe Reyes-Cruz
- Cell Biology, Center for Research and Advanced Studies of the National Polytechnic Institute (CINVESTAV-IPN), 07360 Mexico City, Mexico
| | - J Silvio Gutkind
- the Moores Cancer Center and Department of Pharmacology, University of California, San Diego, La Jolla, California 92093
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25
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P-Rex1 is dispensable for Erk activation and mitogenesis in breast cancer. Oncotarget 2018; 9:28612-28624. [PMID: 29983884 PMCID: PMC6033363 DOI: 10.18632/oncotarget.25584] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2017] [Accepted: 05/19/2018] [Indexed: 12/21/2022] Open
Abstract
Phosphatidylinositol-3,4,5-Trisphosphate Dependent Rac Exchange Factor 1 (P-Rex1) is a key mediator of growth factor-induced activation of Rac1, a small GTP-binding protein widely implicated in actin cytoskeleton reorganization. This Guanine nucleotide Exchange Factor (GEF) is overexpressed in human luminal breast cancer, and its expression associates with disease progression, metastatic dissemination and poor outcome. Despite the established contribution of P-Rex1 to Rac activation and cell locomotion, whether this Rac-GEF has any relevant role in mitogenesis has been a subject of controversy. To tackle the discrepancies among various reports, we carried out an exhaustive analysis of the potential involvement of P-Rex1 on the activation of the mitogenic Erk pathway. Using a range of luminal breast cancer cellular models, we unequivocally showed that silencing P-Rex1 (transiently, stably, using multiple siRNA sequences) had no effect on the phospho-Erk response upon stimulation with growth factors (EGF, heregulin, IGF-I) or a GPCR ligand (SDF-1). The lack of involvement of P-Rex1 in Erk activation was confirmed at the single cell level using a fluorescent biosensor of Erk kinase activity. Depletion of P-Rex1 from breast cancer cells failed to affect cell cycle progression, cyclin D1 induction, Akt activation and apoptotic responses. In addition, mammary-specific P-Rex1 transgenic mice (MMTV-P-Rex1) did not show any obvious hyperproliferative phenotype. Therefore, despite its crucial role in Rac1 activation and cell motility, P-Rex1 is dispensable for mitogenic or survival responses in breast cancer cells.
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26
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Lamanuzzi A, Saltarella I, Desantis V, Frassanito MA, Leone P, Racanelli V, Nico B, Ribatti D, Ditonno P, Prete M, Solimando AG, Dammacco F, Vacca A, Ria R. Inhibition of mTOR complex 2 restrains tumor angiogenesis in multiple myeloma. Oncotarget 2018; 9:20563-20577. [PMID: 29755672 PMCID: PMC5945497 DOI: 10.18632/oncotarget.25003] [Citation(s) in RCA: 37] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2017] [Accepted: 03/13/2018] [Indexed: 01/27/2023] Open
Abstract
The mammalian Target of Rapamycin (mTOR) is an intracellular serine/threonine kinase that mediates intracellular metabolism, cell survival and actin rearrangement. mTOR is made of two independent complexes, mTORC1 and mTORC2, activated by the scaffold proteins RAPTOR and RICTOR, respectively. The activation of mTORC1 triggers protein synthesis and autophagy inhibition, while mTORC2 activation promotes progression, survival, actin reorganization, and drug resistance through AKT hyper-phosphorylation on Ser473. Due to the mTOR pivotal role in the survival of tumor cells, we evaluated its activation in endothelial cells (ECs) from 20 patients with monoclonal gammopathy of undetermined significance (MGUS) and 47 patients with multiple myeloma (MM), and its involvement in angiogenesis. MM-ECs showed a significantly higher expression of mTOR and RICTOR than MGUS-ECs. These data were supported by the higher activation of mTORC2 downstream effectors, suggesting a major role of mTORC2 in the angiogenic switch to MM. Specific inhibition of mTOR activity through siRNA targeting RICTOR and dual mTOR inhibitor PP242 reduced the MM-ECs angiogenic functions, including cell migration, chemotaxis, adhesion, invasion, in vitro angiogenesis on Matrigel®, and cytoskeleton reorganization. In addition, PP242 treatment showed anti-angiogenic effects in vivo in the Chick Chorioallantoic Membrane (CAM) and Matrigel® plug assays. PP242 exhibited a synergistic effect with lenalidomide and bortezomib, suggesting that mTOR inhibition can enhance the anti-angiogenic effect of these drugs. Data to be shown indicate that mTORC2 is involved in MM angiogenesis, and suggest that the dual mTOR inhibitor PP242 may be useful for the anti-angiogenic management of MM patients.
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Affiliation(s)
- Aurelia Lamanuzzi
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Ilaria Saltarella
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Vanessa Desantis
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Maria Antonia Frassanito
- Department of Biomedical Sciences and Human Oncology, General Pathology Unit, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Patrizia Leone
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Vito Racanelli
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Beatrice Nico
- Department of Basic Medical Sciences, Neurosciences, and Sensory Organs, Section of Human Anatomy and Histology, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Domenico Ribatti
- Department of Basic Medical Sciences, Neurosciences, and Sensory Organs, Section of Human Anatomy and Histology, University of Bari Aldo Moro Medical School, Bari, Italy.,National Cancer Institute Giovanni Paolo II, Bari, Italy
| | | | - Marcella Prete
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Antonio Giovanni Solimando
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Francesco Dammacco
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Angelo Vacca
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
| | - Roberto Ria
- Department of Biomedical Sciences and Human Oncology, Internal Medicine Unit G. Baccelli, University of Bari Aldo Moro Medical School, Bari, Italy
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27
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Li N, Li Y, Gao X, Yu Z, Pan W, Tang B. Multiplexed gene silencing in living cells and in vivo using a DNAzymes-CoOOH nanocomposite. Chem Commun (Camb) 2018; 53:4962-4965. [PMID: 28422198 DOI: 10.1039/c7cc00822h] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
We demonstrate a novel DNAzymes-based nanocomposite that can simultaneously silence three types of genes in living cells and in vivo. The synergetic strategy for silencing three different genes can significantly enhance the knockdown efficacy and effectively inhibit the cancer cells' progression.
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Affiliation(s)
- Na Li
- College of Chemistry, Chemical Engineering and Materials Science, Collaborative Innovation Center of Functionalized Probes for Chemical Imaging in Universities of Shandong, Key Laboratory of Molecular and Nano Probes, Ministry of Education, Institute of Molecular and Nano Science, Shandong Normal University, Jinan 250014, P. R. China.
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28
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A Boolean network of the crosstalk between IGF and Wnt signaling in aging satellite cells. PLoS One 2018; 13:e0195126. [PMID: 29596489 PMCID: PMC5875862 DOI: 10.1371/journal.pone.0195126] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2017] [Accepted: 03/16/2018] [Indexed: 12/29/2022] Open
Abstract
Aging is a complex biological process, which determines the life span of an organism. Insulin-like growth factor (IGF) and Wnt signaling pathways govern the process of aging. Both pathways share common downstream targets that allow competitive crosstalk between these branches. Of note, a shift from IGF to Wnt signaling has been observed during aging of satellite cells. Biological regulatory networks necessary to recreate aging have not yet been discovered. Here, we established a mathematical in silico model that robustly recapitulates the crosstalk between IGF and Wnt signaling. Strikingly, it predicts critical nodes following a shift from IGF to Wnt signaling. These findings indicate that this shift might cause age-related diseases.
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29
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Montero JC, Seoane S, García-Alonso S, Pandiella A. Multisite phosphorylation of P-Rex1 by protein kinase C. Oncotarget 2018; 7:77937-77949. [PMID: 27788493 PMCID: PMC5363633 DOI: 10.18632/oncotarget.12846] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2016] [Accepted: 10/12/2016] [Indexed: 02/07/2023] Open
Abstract
P-Rex proteins are guanine nucleotide exchange factors (GEFs) that act on the Rho/Rac family of GTP binding proteins. The activity of P-Rex proteins is regulated by several extracellular stimuli. In fact, activation of growth factor receptors has been reported to activate a phosphorylation/dephosphorylation cycle of P-Rex1. Such cycle includes dephosphorylation of serines 313 and 319 which negatively regulate the GEF activity of P-Rex1, together with phosphorylation of serines 605 and 1169 which favour P-Rex1 GEF activity. However, the kinases that regulate phosphorylation at these different regulatory sites are largely unknown. Here we have investigated the potential regulatory action of several kinases on the phosphorylation of P-Rex1 at S313, S319, S605 and S1169. We show that activation of protein kinase C (PKC) caused phosphorylation of S313, S319 and S1169. Activation of growth factor receptors induced phosphorylation of S1169 through a mechanism that was independent of PKC, indicating that distinct kinases and mechanisms control the phosphorylation of P-Rex1 at different regulatory serines. Genetic and biochemical studies confirmed that the PKC isoform PKCδ was able to directly phosphorylate P-Rex1 at S313. Functional studies using cells with very low endogenous P-Rex1 expression, transfected with wild type P-Rex1 or a mutant form in which S313 was substituted by alanine, indicated that phosphorylation at that residue negatively regulated P-Rex1 exchange activity. We suggest that control of P-Rex1 activity depends on a highly dynamic interplay among distinct signalling routes and its multisite phosphorylation is controlled by the action of different kinases.
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Affiliation(s)
- Juan Carlos Montero
- Instituto de Biología Molecular y Celular del Cáncer, CSIC-Universidad de Salamanca, Spain
| | - Samuel Seoane
- Instituto de Biología Molecular y Celular del Cáncer, CSIC-Universidad de Salamanca, Spain
| | - Sara García-Alonso
- Instituto de Biología Molecular y Celular del Cáncer, CSIC-Universidad de Salamanca, Spain
| | - Atanasio Pandiella
- Instituto de Biología Molecular y Celular del Cáncer, CSIC-Universidad de Salamanca, Spain
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30
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Masters TA, Tumbarello DA, Chibalina MV, Buss F. MYO6 Regulates Spatial Organization of Signaling Endosomes Driving AKT Activation and Actin Dynamics. Cell Rep 2018; 19:2088-2101. [PMID: 28591580 PMCID: PMC5469940 DOI: 10.1016/j.celrep.2017.05.048] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2016] [Revised: 04/05/2017] [Accepted: 05/12/2017] [Indexed: 02/06/2023] Open
Abstract
APPL1- and RAB5-positive signaling endosomes play a crucial role in the activation of AKT in response to extracellular stimuli. Myosin VI (MYO6) and two of its cargo adaptor proteins, GIPC and TOM1/TOM1L2, localize to these peripheral endosomes and mediate endosome association with cortical actin filaments. Loss of MYO6 leads to the displacement of these endosomes from the cell cortex and accumulation in the perinuclear space. Depletion of this myosin not only affects endosome positioning, but also induces actin and lipid remodeling consistent with endosome maturation, including accumulation of F-actin and the endosomal lipid PI(3)P. These processes acutely perturb endosome function, as both AKT phosphorylation and RAC-dependent membrane ruffling were markedly reduced by depletion of either APPL1 or MYO6. These results place MYO6 and its binding partners at a central nexus in cellular signaling linking actin dynamics at the cell surface and endosomal signaling in the cell cortex.
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Affiliation(s)
- Thomas A Masters
- Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK
| | - David A Tumbarello
- Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK
| | - Margarita V Chibalina
- Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK
| | - Folma Buss
- Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK.
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Qian J, Chen Y, Meng T, Ma L, Meng L, Wang X, Yu T, Zask A, Shen J, Yu K. Molecular regulation of apoptotic machinery and lipid metabolism by mTORC1/mTORC2 dual inhibitors in preclinical models of HER2+/PIK3CAmut breast cancer. Oncotarget 2018; 7:67071-67086. [PMID: 27563814 PMCID: PMC5341858 DOI: 10.18632/oncotarget.11490] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2016] [Accepted: 08/09/2016] [Indexed: 11/25/2022] Open
Abstract
The mechanistic target of rapamycin (mTOR) is a rational target for cancer treatment. While the mTORC1-selective rapalogs have shown significant benefits in the clinic, antitumor response may be further improved by inhibiting both mTORC1 and mTORC2. Herein, we established target profile of a novel mTOR kinase inhibitor (mTOR-KI) MTI-31 and employed it to study new therapeutic mechanism in breast cancer. MTI-31 demonstrated a potent mTOR binding affinity with >5000 fold selectivity over the related PI3K family isoforms. MTI-31 inhibited mTORC1- and mTORC2 function at ≤120 nM in cellular assays or 5 mg/kg orally in tumor-bearing mice. In a panel of breast cancer lines, the antitumor efficacy of MTI-31 was dependent on HER2+ and/or PIK3CAmut (HER2+/PIK3CAmut) status of the tumors and required mTORC2-specific modulation of Bim, MCL-1 and GSK3. Inactivation of Bim or GSK3 each attenuated apoptotic death resulting in mTOR-KI resistance. The antitumor response also required a suppression of lipid metabolism in therapy-sensitive tumors. Treatment with MTI-31 or AZD8055 substantially reduced lipogenesis and acetyl-CoA homeostasis, which was mechanistically linked to a blockade of mTORC2-dependent glucose-to-lipid conversion rate. We also found that the basal levels of carnitine palmitoyltransferase 1A and lipid catabolism were elevated in HER2+/PIK3CAmut breast cells and were inhibited upon mTOR-KI treatment. A CPT1A inhibitor etomoxir mimicked MTI-31 action in selective downregulation of cellular lipid catabolism. Co-treatments with MTI-31 and etomoxir enhanced the suppression of cyclin D1, c-Myc and cell growth in HER2+/PIK3CAmut tumors. These new mechanistic findings provide a rationale for targeting mTORC1 and mTORC2 in HER2+/PIK3CAmut breast cancer.
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Affiliation(s)
- Jianchang Qian
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Yaqing Chen
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Tao Meng
- Department of Medicinal Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
| | - Lanping Ma
- Department of Medicinal Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
| | - Lanfang Meng
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
| | - Xin Wang
- Department of Medicinal Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
| | - Ting Yu
- Department of Medicinal Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
| | - Arie Zask
- Department of Biological Sciences, Columbia University, New York, NY, USA
| | - Jingkang Shen
- Department of Medicinal Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
| | - Ker Yu
- Department of Pharmacology, Fudan University School of Pharmacy, Shanghai, China
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32
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Lawson CD, Ridley AJ. Rho GTPase signaling complexes in cell migration and invasion. J Cell Biol 2018; 217:447-457. [PMID: 29233866 PMCID: PMC5800797 DOI: 10.1083/jcb.201612069] [Citation(s) in RCA: 357] [Impact Index Per Article: 51.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2017] [Revised: 10/23/2017] [Accepted: 11/17/2017] [Indexed: 12/02/2022] Open
Abstract
Cell migration is dependent on the dynamic formation and disassembly of actin filament-based structures, including lamellipodia, filopodia, invadopodia, and membrane blebs, as well as on cell-cell and cell-extracellular matrix adhesions. These processes all involve Rho family small guanosine triphosphatases (GTPases), which are regulated by the opposing actions of guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). Rho GTPase activity needs to be precisely tuned at distinct cellular locations to enable cells to move in response to different environments and stimuli. In this review, we focus on the ability of RhoGEFs and RhoGAPs to form complexes with diverse binding partners, and describe how this influences their ability to control localized GTPase activity in the context of migration and invasion.
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Affiliation(s)
- Campbell D Lawson
- Randall Centre for Cell and Molecular Biophysics, King's College London, London, England, UK
| | - Anne J Ridley
- Randall Centre for Cell and Molecular Biophysics, King's College London, London, England, UK
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33
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Masui K, Kato Y, Sawada T, Mischel PS, Shibata N. Molecular and Genetic Determinants of Glioma Cell Invasion. Int J Mol Sci 2017; 18:E2609. [PMID: 29207533 PMCID: PMC5751212 DOI: 10.3390/ijms18122609] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2017] [Revised: 11/27/2017] [Accepted: 12/02/2017] [Indexed: 12/21/2022] Open
Abstract
A diffusely invasive nature is a major obstacle in treating a malignant brain tumor, "diffuse glioma", which prevents neurooncologists from surgically removing the tumor cells even in combination with chemotherapy and radiation. Recently updated classification of diffuse gliomas based on distinct genetic and epigenetic features has culminated in a multilayered diagnostic approach to combine histologic phenotypes and molecular genotypes in an integrated diagnosis. However, it is still a work in progress to decipher how the genetic aberrations contribute to the aggressive nature of gliomas including their highly invasive capacity. Here we depict a set of recent discoveries involving molecular genetic determinants of the infiltrating nature of glioma cells, especially focusing on genetic mutations in receptor tyrosine kinase pathways and metabolic reprogramming downstream of common cancer mutations. The specific biology of glioma cell invasion provides an opportunity to explore the genotype-phenotype correlation in cancer and develop novel glioma-specific therapeutic strategies for this devastating disease.
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Affiliation(s)
- Kenta Masui
- Department of Pathology, Tokyo Women's Medical University, Tokyo 162-8666, Japan.
| | - Yoichiro Kato
- Department of Pathology, Tokyo Women's Medical University, Tokyo 162-8666, Japan.
| | - Tatsuo Sawada
- Department of Pathology, Tokyo Women's Medical University, Tokyo 162-8666, Japan.
| | - Paul S Mischel
- Ludwig Institute for Cancer Research, University of California San Diego, La Jolla, CA 92093, USA.
| | - Noriyuki Shibata
- Department of Pathology, Tokyo Women's Medical University, Tokyo 162-8666, Japan.
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34
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May CD, Landers SM, Bolshakov S, Ma X, Ingram DR, Kivlin CM, Watson KL, Sannaa GAA, Bhalla AD, Wang WL, Lazar AJ, Torres KE. Co-targeting PI3K, mTOR, and IGF1R with small molecule inhibitors for treating undifferentiated pleomorphic sarcoma. Cancer Biol Ther 2017; 18:816-826. [PMID: 29099264 PMCID: PMC5678691 DOI: 10.1080/15384047.2017.1373230] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Undifferentiated pleomorphic sarcomas (UPSs) are aggressive mesenchymal malignancies with no definitive cell of origin or specific recurrent genetic hallmarks. These tumors are largely chemoresistant; thus, identification of potential therapeutic targets is necessary to improve patient outcome. Previous studies demonstrated that high expression of activated protein kinase B (AKT) in patients with UPS corresponds to poor disease-specific survival. Here, we demonstrate that inhibiting phosphatidylinositol-3-kinase/mammalian target of rapamycin (PI3K/mTOR) signaling using a small molecule inhibitor reduced UPS cell proliferation and motility and xenograft growth; however, increased phosphorylation of insulin-like growth factor 1 receptor (IGF1R) indicated the potential for adaptive resistance following treatment through compensatory receptor activation. Co-treatment with a dual PI3K/mTOR inhibitor and an anti-IGF1R kinase inhibitor reduced in vivo tumor growth rates despite a lack of antiproliferative effects in vitro. Moreover, this combination treatment significantly decreased UPS cell migration and invasion, which is linked to changes in p27 subcellular localization. Our results demonstrate that targeted inhibition of multiple components of the IGF1R/PI3K/mTOR pathway was more efficacious than single-agent therapy and suggest that co-targeting this pathway could be a beneficial therapeutic strategy for patients with UPS.
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Affiliation(s)
- Caitlin D May
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA.,b The University of Texas Health Science Center at Houston , Graduate School of Biomedical Sciences , Houston , TX , USA
| | - Sharon M Landers
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Svetlana Bolshakov
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - XiaoYan Ma
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Davis R Ingram
- c Department of Pathology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Christine M Kivlin
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA.,b The University of Texas Health Science Center at Houston , Graduate School of Biomedical Sciences , Houston , TX , USA
| | - Kelsey L Watson
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Ghadah A Al Sannaa
- c Department of Pathology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Angela D Bhalla
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Wei-Lien Wang
- c Department of Pathology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Alexander J Lazar
- b The University of Texas Health Science Center at Houston , Graduate School of Biomedical Sciences , Houston , TX , USA.,c Department of Pathology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA
| | - Keila E Torres
- a Department of Surgical Oncology , The University of Texas MD Anderson Cancer Center , Houston , TX , USA.,b The University of Texas Health Science Center at Houston , Graduate School of Biomedical Sciences , Houston , TX , USA
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Kim YY, Jee HJ, Um JH, Kim YM, Bae SS, Yun J. Cooperation between p21 and Akt is required for p53-dependent cellular senescence. Aging Cell 2017; 16:1094-1103. [PMID: 28691365 PMCID: PMC5595696 DOI: 10.1111/acel.12639] [Citation(s) in RCA: 77] [Impact Index Per Article: 9.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 06/04/2017] [Indexed: 12/30/2022] Open
Abstract
Cellular senescence has been implicated in normal aging, tissue homeostasis, and tumor suppression. Although p53 has been shown to be a central mediator of cellular senescence, the signaling pathway by which it induces senescence remains incompletely understood. In this study, we have shown that both Akt and p21 are required to induce cellular senescence in response to p53 expression. In a p53-induced senescence model, we found that Akt activation was essential for inducing a cellular senescence phenotype. Surprisingly, Akt inhibition did not abolish p53-induced cell cycle arrest, but it suppressed the increase in intracellular reactive oxygen species (ROS) levels. The results of the cell cycle and morphological analysis suggest that p53 induced quiescence, not senescence, following Akt inhibition. Conversely, the inhibition of p21 induction abolished cell cycle arrest but did not affect the p53-induced increase in ROS levels. Additionally, p21 and Akt separately controlled cell cycle arrest and ROS levels, respectively, during H-Ras-induced senescence in human normal fibroblasts. The mechanistic analysis revealed that Akt increased ROS levels through NOX4 induction, and increased Akt-dependent NF-κB binding to the NOX4 promoter is responsible for NOX4 induction upon p53 expression. We further showed that Akt activation upon p53 expression is mediated by mammalian target of rapamycin complex 2. In addition, p53-mediated IL6 and IL8 induction was abrogated by Akt inhibition, suggesting that Akt activation is also required for the senescence-associated secretory phenotype. Collectively, these results suggest that p53 simultaneously controls multiple pathways to induce cellular senescence through p21 and Akt.
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Affiliation(s)
- Young Yeon Kim
- Peripheral Neuropathy Research Center; College of Medicine; Dong-A University; Busan 49201 Korea
- Department of Biochemistry; College of Medicine; Dong-A University; Busan 49201 Korea
| | - Hye Jin Jee
- Peripheral Neuropathy Research Center; College of Medicine; Dong-A University; Busan 49201 Korea
- Department of Biochemistry; College of Medicine; Dong-A University; Busan 49201 Korea
| | - Jee-Hyun Um
- Peripheral Neuropathy Research Center; College of Medicine; Dong-A University; Busan 49201 Korea
- Department of Biochemistry; College of Medicine; Dong-A University; Busan 49201 Korea
| | - Young Mi Kim
- Peripheral Neuropathy Research Center; College of Medicine; Dong-A University; Busan 49201 Korea
- Department of Biochemistry; College of Medicine; Dong-A University; Busan 49201 Korea
| | - Sun Sik Bae
- Department of Pharmacology; School of Medicine; Pusan National University; Yangsan-si 602-739 Korea
| | - Jeanho Yun
- Peripheral Neuropathy Research Center; College of Medicine; Dong-A University; Busan 49201 Korea
- Department of Biochemistry; College of Medicine; Dong-A University; Busan 49201 Korea
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36
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AKT/PKB Signaling: Navigating the Network. Cell 2017; 169:381-405. [PMID: 28431241 DOI: 10.1016/j.cell.2017.04.001] [Citation(s) in RCA: 2593] [Impact Index Per Article: 324.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2017] [Revised: 03/29/2017] [Accepted: 03/31/2017] [Indexed: 12/14/2022]
Abstract
The Ser and Thr kinase AKT, also known as protein kinase B (PKB), was discovered 25 years ago and has been the focus of tens of thousands of studies in diverse fields of biology and medicine. There have been many advances in our knowledge of the upstream regulatory inputs into AKT, key multifunctional downstream signaling nodes (GSK3, FoxO, mTORC1), which greatly expand the functional repertoire of AKT, and the complex circuitry of this dynamically branching and looping signaling network that is ubiquitous to nearly every cell in our body. Mouse and human genetic studies have also revealed physiological roles for the AKT network in nearly every organ system. Our comprehension of AKT regulation and functions is particularly important given the consequences of AKT dysfunction in diverse pathological settings, including developmental and overgrowth syndromes, cancer, cardiovascular disease, insulin resistance and type 2 diabetes, inflammatory and autoimmune disorders, and neurological disorders. There has also been much progress in developing AKT-selective small molecule inhibitors. Improved understanding of the molecular wiring of the AKT signaling network continues to make an impact that cuts across most disciplines of the biomedical sciences.
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37
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Naruse T, Yanamoto S, Okuyama K, Yamashita K, Omori K, Nakao Y, Yamada SI, Umeda M. Therapeutic implication of mTORC2 in oral squamous cell carcinoma. Oral Oncol 2016; 65:23-32. [PMID: 28109464 DOI: 10.1016/j.oraloncology.2016.12.012] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/26/2016] [Revised: 11/09/2016] [Accepted: 12/14/2016] [Indexed: 12/16/2022]
Abstract
The aim of the present study was to clarify the association of mTORC2 expression with the cancer progression and the anti-tumor effects of Torin-1 alone and combined treatment with Cetuximab in OSCC cells. The expressions of Rictor and SGK1 were immunohistochemically evaluated and the relationships between the expressions of molecular markers and clinicopathological factors were determined. Moreover, OSCC cells were treated with Torin-1, Cetuximab or combined agents, and anti-tumor effects of OSCC cells were examined in vitro and in vivo. Rictor and SGK1 expressions were significantly associated with tumor stage and pattern of invasion in OSCC sections (P<0.05 and P<0.01, respectively). Treatment of OSCC cell lines with Torin-1 resulted in dose and time-dependent inhibition of proliferation with decrease of phosphorylation on downstream molecules. Combined treatment with Torin-1 and Cetuximab resulted in enhanced anti-tumor effects in vitro compared with either agent alone. Furthermore, treatment of mice bearing OSCC xenografts with Torin-1 and Cetuximab also demonstrated a remarked growth inhibition of tumor volumes. The results suggested that new regimens of systemic therapy combined with Cetuximab and Torin-1 may be useful for very advanced OSCC patients.
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Affiliation(s)
- Tomofumi Naruse
- Department of Clinical Oral Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan.
| | - Souichi Yanamoto
- Department of Clinical Oral Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan
| | - Kohei Okuyama
- Department of Clinical Oral Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan
| | - Kentaro Yamashita
- Department of Clinical Oral Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan
| | - Keisuke Omori
- Department of Clinical Oral Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan
| | - Yuji Nakao
- Department of Dentistry and Oral Surgery, Shinshu University School of Medicine, Matsumoto, Nagano 390-8621, Japan
| | - Shin-Ichi Yamada
- Department of Dentistry and Oral Surgery, Shinshu University School of Medicine, Matsumoto, Nagano 390-8621, Japan
| | - Masahiro Umeda
- Department of Clinical Oral Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8588, Japan
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38
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Wang Y, Li D, Luo J, Tian G, Zhao LY, Liao D. Intrinsic cellular signaling mechanisms determine the sensitivity of cancer cells to virus-induced apoptosis. Sci Rep 2016; 6:37213. [PMID: 27849011 PMCID: PMC5111159 DOI: 10.1038/srep37213] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2016] [Accepted: 10/26/2016] [Indexed: 12/28/2022] Open
Abstract
Cancer cells of epithelial and mesenchymal phenotypes exhibit different sensitivities to apoptosis stimuli, but the mechanisms underlying this phenomenon remain partly understood. We constructed a novel recombinant adenovirus expressing Ad12 E1A (Ad-E1A12) that can strongly induce apoptosis. Ad-E1A12 infection of epithelial cancer cells displayed dramatic detachment and apoptosis, whereas cancer cells of mesenchymal phenotypes with metastatic propensity were markedly more resistant to this virus. Notably, forced detachment of epithelial cells did not further sensitize them to Ad-E1A12-induced apoptosis, suggesting that cell detachment is a consequence rather than the cause of Ad-E1A12-induced apoptosis. Ad-E1A12 increased phosphorylation of AKT1 and ribosomal protein S6 through independent mechanisms in different cell types. Ad-E1A12–induced AKT1 phosphorylation was PI3K-dependent in epithelial cancer cells, and mTOR-dependent in mesenchymal cancer cells. Epithelial cancer cells upon Ad-E1A12-induced detachment could not sustain AKT activation due to AKT1 degradation, but AKT1 activation was maintained in mesenchymal cancer cells. Expression of epithelial cell-restricted miR-200 family in mesenchymal cells limited mTOR signaling and sensitized them to Ad-E1A12-induced cell killing. Thus, epithelial cancer cells rely on the canonical PI3K-AKT signaling pathway for survival, while mesenchymal cancer cells deploy the PI3K-independent mTORC2-AKT axis in response to strong death stimuli.
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Affiliation(s)
- Yunfei Wang
- Department of Anatomy and Cell Biology, UF Health Cancer Center, UF Genetics Institute, University of Florida College of Medicine, Gainesville, Florida, USA.,Shaanxi Key Laboratory of Agriculture Molecular Biology, Department of Biochemistry and Molecular Biology, College of Life Science, Northwest Agriculture and Forestry University, Yangling, Shaanxi, China
| | - Dawei Li
- Department of Anatomy and Cell Biology, UF Health Cancer Center, UF Genetics Institute, University of Florida College of Medicine, Gainesville, Florida, USA.,Department of Urology, Qilu Hospital, Shandong University, Jinan, Shandong, China
| | - Jian Luo
- Department of Anatomy and Cell Biology, UF Health Cancer Center, UF Genetics Institute, University of Florida College of Medicine, Gainesville, Florida, USA
| | - Guimei Tian
- Department of Anatomy and Cell Biology, UF Health Cancer Center, UF Genetics Institute, University of Florida College of Medicine, Gainesville, Florida, USA
| | - Lisa Y Zhao
- Department of Anatomy and Cell Biology, UF Health Cancer Center, UF Genetics Institute, University of Florida College of Medicine, Gainesville, Florida, USA
| | - Daiqing Liao
- Department of Anatomy and Cell Biology, UF Health Cancer Center, UF Genetics Institute, University of Florida College of Medicine, Gainesville, Florida, USA
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39
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Vázquez-Prado J, Bracho-Valdés I, Cervantes-Villagrana RD, Reyes-Cruz G. Gβγ Pathways in Cell Polarity and Migration Linked to Oncogenic GPCR Signaling: Potential Relevance in Tumor Microenvironment. Mol Pharmacol 2016; 90:573-586. [PMID: 27638873 DOI: 10.1124/mol.116.105338] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2016] [Accepted: 09/14/2016] [Indexed: 02/14/2025] Open
Abstract
Cancer cells and stroma cells in tumors secrete chemotactic agonists that exacerbate invasive behavior, promote tumor-induced angiogenesis, and recruit protumoral bone marrow-derived cells. In response to shallow gradients of chemotactic stimuli recognized by G protein-coupled receptors (GPCRs), Gβγ-dependent signaling cascades contribute to specifying the spatiotemporal assembly of cytoskeletal structures that can dynamically alter cell morphology. This sophisticated process is intrinsically linked to the activation of Rho GTPases and their cytoskeletal-remodeling effectors. Thus, Rho guanine nucleotide exchange factors, the activators of these molecular switches, and their upstream signaling partners are considered participants of tumor progression. Specifically, phosphoinositide-3 kinases (class I PI3Ks, β and γ) and P-Rex1, a Rac-specific guanine nucleotide exchange factor, are fundamental Gβγ effectors in the pathways controlling directionally persistent motility. In addition, GPCR-dependent chemotactic responses often involve endosomal trafficking of signaling proteins; coincidently, endosomes serve as signaling platforms for Gβγ In preclinical murine models of cancer, inhibition of Gβγ attenuates tumor growth, whereas in cancer patients, aberrant overexpression of chemotactic Gβγ effectors and recently identified mutations in Gβ correlate with poor clinical outcome. Here we discuss emerging paradigms of Gβγ signaling in cancer, which are essential for chemotactic cell migration and represent novel opportunities to develop pathway-specific pharmacologic treatments.
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Affiliation(s)
- José Vázquez-Prado
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
| | - Ismael Bracho-Valdés
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
| | - Rodolfo Daniel Cervantes-Villagrana
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
| | - Guadalupe Reyes-Cruz
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
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40
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Liu HJ, Ooms LM, Srijakotre N, Man J, Vieusseux J, Waters JE, Feng Y, Bailey CG, Rasko JEJ, Price JT, Mitchell CA. PtdIns(3,4,5)P3-dependent Rac Exchanger 1 (PREX1) Rac-Guanine Nucleotide Exchange Factor (GEF) Activity Promotes Breast Cancer Cell Proliferation and Tumor Growth via Activation of Extracellular Signal-regulated Kinase 1/2 (ERK1/2) Signaling. J Biol Chem 2016; 291:17258-70. [PMID: 27358402 DOI: 10.1074/jbc.m116.743401] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2016] [Indexed: 12/20/2022] Open
Abstract
PtdIns(3,4,5)P3-dependent Rac exchanger 1 (PREX1) is a Rac-guanine nucleotide exchange factor (GEF) overexpressed in a significant proportion of human breast cancers that integrates signals from upstream ErbB2/3 and CXCR4 membrane surface receptors. However, the PREX1 domains that facilitate its oncogenic activity and downstream signaling are not completely understood. We identify that ERK1/2 MAPK acts downstream of PREX1 and contributes to PREX1-mediated anchorage-independent cell growth. PREX1 overexpression increased but its shRNA knockdown decreased ERK1/2 phosphorylation in response to EGF/IGF-1 stimulation, resulting in induction of the cell cycle regulators cyclin D1 and p21(WAF1/CIP1) PREX1-mediated ERK1/2 phosphorylation, anchorage-independent cell growth, and cell migration were suppressed by inhibition of MEK1/2/ERK1/2 signaling. PREX1 overexpression reduced staurosporine-induced apoptosis whereas its shRNA knockdown promoted apoptosis in response to staurosporine or the anti-estrogen drug tamoxifen. Expression of wild-type but not GEF-inactive PREX1 increased anchorage-independent cell growth. In addition, mouse xenograft studies revealed that expression of wild-type but not GEF-dead PREX1 resulted in the formation of larger tumors that displayed increased phosphorylation of ERK1/2 but not AKT. The impaired anchorage-independent cell growth, apoptosis, and ERK1/2 signaling observed in stable PREX1 knockdown cells was restored by expression of wild-type but not GEF-dead-PREX1. Therefore, PREX1-Rac-GEF activity is critical for PREX1-dependent anchorage-independent cell growth and xenograft tumor growth and may represent a possible therapeutic target for breast cancers that exhibit PREX1 overexpression.
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Affiliation(s)
- Heng-Jia Liu
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia
| | - Lisa M Ooms
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia
| | - Nuthasuda Srijakotre
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia
| | - Joey Man
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia
| | - Jessica Vieusseux
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia
| | - JoAnne E Waters
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia
| | - Yue Feng
- the Centenary Institute of Cancer Medicine and Cell Biology, New South Wales 2050, Australia
| | - Charles G Bailey
- the Centenary Institute of Cancer Medicine and Cell Biology, New South Wales 2050, Australia, Sydney Medical School, University of Sydney, Sydney, New South Wales 2006, Australia
| | - John E J Rasko
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia, Sydney Medical School, University of Sydney, Sydney, New South Wales 2006, Australia, Cell and Molecular Therapies, Royal Prince Alfred Hospital, Camperdown, New South Wales 2050, Australia, and
| | - John T Price
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia, the Centre for Chronic Disease, College of Health and Biomedicine, Victoria University, Victoria 8001, Australia
| | - Christina A Mitchell
- From the Cancer Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Victoria 3800, Australia,
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41
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Daulat AM, Bertucci F, Audebert S, Sergé A, Finetti P, Josselin E, Castellano R, Birnbaum D, Angers S, Borg JP. PRICKLE1 Contributes to Cancer Cell Dissemination through Its Interaction with mTORC2. Dev Cell 2016; 37:311-325. [PMID: 27184734 DOI: 10.1016/j.devcel.2016.04.011] [Citation(s) in RCA: 43] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2015] [Revised: 03/15/2016] [Accepted: 04/18/2016] [Indexed: 12/21/2022]
Abstract
Components of the evolutionarily conserved developmental planar cell polarity (PCP) pathway were recently described to play a prominent role in cancer cell dissemination. However, the molecular mechanisms by which PCP molecules drive the spread of cancer cells remain largely unknown. PRICKLE1 encodes a PCP protein bound to the promigratory serine/threonine kinase MINK1. We identify RICTOR, a member of the mTORC2 complex, as a PRICKLE1-binding partner and show that the integrity of the PRICKLE1-MINK1-RICTOR complex is required for activation of AKT, regulation of focal adhesions, and cancer cell migration. Disruption of the PRICKLE1-RICTOR interaction results in a strong impairment of breast cancer cell dissemination in xenograft assays. Finally, we show that upregulation of PRICKLE1 in basal breast cancers, a subtype characterized by high metastatic potential, is associated with poor metastasis-free survival.
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Affiliation(s)
- Avais M Daulat
- Inserm, U1068, CRCM, Cell Polarity, Cell Signalling and Cancer "Equipe labellisée Ligue Contre le Cancer", Marseille 13009, France; Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France
| | - François Bertucci
- Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France; Inserm, U1068, CRCM, Molecular Oncology "Equipe labellisée Ligue Contre le Cancer", Marseille 13009, France
| | - Stéphane Audebert
- Inserm, U1068, CRCM, Cell Polarity, Cell Signalling and Cancer "Equipe labellisée Ligue Contre le Cancer", Marseille 13009, France; Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France
| | - Arnauld Sergé
- Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France; Inserm, U1068, CRCM, Leuko/Stromal Interactions, Marseille 13009, France
| | - Pascal Finetti
- Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France; Inserm, U1068, CRCM, Molecular Oncology "Equipe labellisée Ligue Contre le Cancer", Marseille 13009, France
| | - Emmanuelle Josselin
- Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France; Inserm, U1068, CRCM, TrGET Platform, Marseille 13009, France
| | - Rémy Castellano
- Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France; Inserm, U1068, CRCM, TrGET Platform, Marseille 13009, France
| | - Daniel Birnbaum
- Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France; Inserm, U1068, CRCM, Molecular Oncology "Equipe labellisée Ligue Contre le Cancer", Marseille 13009, France
| | - Stéphane Angers
- Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, ON M5S3M2, Canada; Department of Biochemistry, Faculty of Medicine, University of Toronto, ON M5S1A8, Canada
| | - Jean-Paul Borg
- Inserm, U1068, CRCM, Cell Polarity, Cell Signalling and Cancer "Equipe labellisée Ligue Contre le Cancer", Marseille 13009, France; Institut Paoli-Calmettes, Marseille 13009, France; Aix-Marseille Université, UM 105, Marseille 13284, France; CNRS, UMR7258, CRCM, Marseille 13009, France.
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Kim M, Kim YY, Jee HJ, Bae SS, Jeong NY, Um JH, Yun J. Akt3 knockdown induces mitochondrial dysfunction in human cancer cells. Acta Biochim Biophys Sin (Shanghai) 2016; 48:447-53. [PMID: 26972278 DOI: 10.1093/abbs/gmw014] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2015] [Accepted: 01/12/2016] [Indexed: 11/15/2022] Open
Abstract
Akt/PKB plays a pivotal role in cell proliferation and survival. However, the isotype-specific roles of Akt in mitochondrial function have not been fully addressed. In this study, we explored the role of Akt in mitochondrial function after stable knockdown of the Akt isoforms in EJ human bladder cancer cells. We found that the mitochondrial mass was significantly increased in the Akt1- and Akt3-knockdown cells, and this increase was accompanied by an increase in TFAM and NRF1. Akt2 knockdown did not cause a similar effect. Interestingly, Akt3 knockdown also led to severe structural defects in the mitochondria, an increase in doxorubicin-induced senescence, and impairment of cell proliferation in galactose medium. Consistent with these observations, the mitochondrial oxygen consumption rate was significantly reduced in the Akt3-knockdown cells. An Akt3 deficiency-induced decrease in mitochondrial respiration was also observed in A549 lung cancer cells. Collectively, these results suggest that the Akt isoforms play distinct roles in mitochondrial function and that Akt3 is critical for proper mitochondrial respiration in human cancer cells.
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Affiliation(s)
- Minjee Kim
- Department of Biochemistry, College of Medicine, Dong-A University, Busan 607-714, Republic of Korea Institute of Convergence Bio-Health, Dong-A University, Busan 607-714, Republic of Korea
| | - Young Yeon Kim
- Department of Biochemistry, College of Medicine, Dong-A University, Busan 607-714, Republic of Korea Institute of Convergence Bio-Health, Dong-A University, Busan 607-714, Republic of Korea
| | - Hye Jin Jee
- Department of Biochemistry, College of Medicine, Dong-A University, Busan 607-714, Republic of Korea Institute of Convergence Bio-Health, Dong-A University, Busan 607-714, Republic of Korea
| | - Sun Sik Bae
- Department of Pharmacology, School of Medicine, Pusan National University, Yangsan-si 602-739, Republic of Korea
| | - Na Young Jeong
- Institute of Convergence Bio-Health, Dong-A University, Busan 607-714, Republic of Korea Department of Anatomy and Cell Biology, College of Medicine, Dong-A University, Busan 602-714, Republic of Korea
| | - Jee-Hyun Um
- Department of Biochemistry, College of Medicine, Dong-A University, Busan 607-714, Republic of Korea Institute of Convergence Bio-Health, Dong-A University, Busan 607-714, Republic of Korea
| | - Jeanho Yun
- Department of Biochemistry, College of Medicine, Dong-A University, Busan 607-714, Republic of Korea Institute of Convergence Bio-Health, Dong-A University, Busan 607-714, Republic of Korea
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43
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Kim YH, Baek SH, Kim EK, Ha JM, Jin SY, Lee HS, Ha HK, Song SH, Kim SJ, Shin HK, Yong J, Kim DH, Kim CD, Bae SS. Uncoordinated 51-like kinase 2 signaling pathway regulates epithelial-mesenchymal transition in A549 lung cancer cells. FEBS Lett 2016; 590:1365-74. [DOI: 10.1002/1873-3468.12172] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2015] [Revised: 03/22/2016] [Accepted: 03/29/2016] [Indexed: 01/08/2023]
Affiliation(s)
- Young Hwan Kim
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
| | - Seung Hoon Baek
- Department of Anesthesia and Pain Medicine; Pusan National University Hospital; Yangsan Korea
| | - Eun Kyoung Kim
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
| | - Jung Min Ha
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
| | - Seo Yeon Jin
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
| | - Hye Sun Lee
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
| | - Hong Koo Ha
- Department of Urology; Pusan National University Hospital; Busan Korea
| | - Sang Heon Song
- Department of Internal Medicine; Pusan National University Hospital; Busan Korea
| | - Sun Ja Kim
- Department of Physics; Dong-A University; Busan Korea
| | - Hwa Kyoung Shin
- Department of Anatomy; Pusan National University School of Korean Medicine; Yangsan Korea
| | - Jeongsik Yong
- Department of Biochemistry, Molecular Biology and Biophysics; University of Minnesota Twin Cities; Minneapolis MN USA
| | - Do-Hyung Kim
- Department of Biochemistry, Molecular Biology and Biophysics; University of Minnesota Twin Cities; Minneapolis MN USA
| | - Chi Dae Kim
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
| | - Sun Sik Bae
- Department of Pharmacology; Gene and Cell Therapy Center for Vessel-Associated Disease; Medical Research Institute; Pusan National University School of Medicine; Yangsan Korea
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Ziegler ME, Hatch MMS, Wu N, Muawad SA, Hughes CCW. mTORC2 mediates CXCL12-induced angiogenesis. Angiogenesis 2016; 19:359-71. [PMID: 27106789 DOI: 10.1007/s10456-016-9509-6] [Citation(s) in RCA: 48] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2015] [Accepted: 04/03/2016] [Indexed: 01/26/2023]
Abstract
The chemokine CXCL12, through its receptor CXCR4, positively regulates angiogenesis by promoting endothelial cell (EC) migration and tube formation. However, the relevant downstream signaling pathways in EC have not been defined. Similarly, the upstream activators of mTORC2 signaling in EC are also poorly defined. Here, we demonstrate for the first time that CXCL12 regulation of angiogenesis requires mTORC2 but not mTORC1. We find that CXCR4 signaling activates mTORC2 as indicated by phosphorylation of serine 473 on Akt and does so through a G-protein- and PI3K-dependent pathway. Significantly, independent disruption of the mTOR complexes by drugs or multiple independent siRNAs reveals that mTORC2, but not mTORC1, is required for microvascular sprouting in a 3D in vitro angiogenesis model. Importantly, in a mouse model, both tumor angiogenesis and tumor volume are significantly reduced only when mTORC2 is inhibited. Finally, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), which is a key regulator of glycolytic flux, is required for microvascular sprouting in vitro, and its expression is reduced in vivo when mTORC2 is targeted. Taken together, these findings identify mTORC2 as a critical signaling nexus downstream of CXCL12/CXCR4 that represents a potential link between mTORC2, metabolic regulation, and angiogenesis.
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Affiliation(s)
- Mary E Ziegler
- The Department of Molecular Biology and Biochemistry, University of California Irvine, 3219 McGaugh Hall, Mail Code: 3900, Irvine, CA, 92697, USA
| | - Michaela M S Hatch
- The Department of Molecular Biology and Biochemistry, University of California Irvine, 3219 McGaugh Hall, Mail Code: 3900, Irvine, CA, 92697, USA
| | - Nan Wu
- The Department of Molecular Biology and Biochemistry, University of California Irvine, 3219 McGaugh Hall, Mail Code: 3900, Irvine, CA, 92697, USA
| | - Steven A Muawad
- The Department of Molecular Biology and Biochemistry, University of California Irvine, 3219 McGaugh Hall, Mail Code: 3900, Irvine, CA, 92697, USA
| | - Christopher C W Hughes
- The Department of Molecular Biology and Biochemistry, University of California Irvine, 3219 McGaugh Hall, Mail Code: 3900, Irvine, CA, 92697, USA. .,The Department of Biomedical Engineering, University of California Irvine, Irvine, CA, 92697, USA. .,The Edwards Lifesciences Center for Advanced Cardiovascular Technology, University of California Irvine, Irvine, CA, 92697, USA.
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Regulation of PtdIns(3,4,5)P3/Akt signalling by inositol polyphosphate 5-phosphatases. Biochem Soc Trans 2016; 44:240-52. [DOI: 10.1042/bst20150214] [Citation(s) in RCA: 39] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
The phosphoinositide 3-kinase (PI3K) generated lipid signals, PtdIns(3,4,5)P3 and PtdIns(3,4)P2, are both required for the maximal activation of the serine/threonine kinase proto-oncogene Akt. The inositol polyphosphate 5-phosphatases (5-phosphatases) hydrolyse the 5-position phosphate from the inositol head group of PtdIns(3,4,5)P3 to yield PtdIns(3,4)P2. Extensive work has revealed several 5-phosphatases inhibit PI3K-driven Akt signalling, by decreasing PtdIns(3,4,5)P3 despite increasing cellular levels of PtdIns(3,4)P2. The roles that 5-phosphatases play in suppressing cell proliferation and transformation are slow to emerge; however, the 5-phosphatase PIPP [proline-rich inositol polyphosphate 5-phosphatase; inositol polyphosphate 5-phosphatase (INPP5J)] has recently been identified as a putative tumour suppressor in melanoma and breast cancer and SHIP1 [SH2 (Src homology 2)-containing inositol phosphatase 1] inhibits haematopoietic cell proliferation. INPP5E regulates cilia stability and INPP5E mutations have been implicated ciliopathy syndromes. This review will examine 5-phosphatase regulation of PI3K/Akt signalling, focussing on the role PtdIns(3,4,5)P3 5-phosphatases play in developmental diseases and cancer.
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46
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Chávez-Vargas L, Adame-García SR, Cervantes-Villagrana RD, Castillo-Kauil A, Bruystens JGH, Fukuhara S, Taylor SS, Mochizuki N, Reyes-Cruz G, Vázquez-Prado J. Protein Kinase A (PKA) Type I Interacts with P-Rex1, a Rac Guanine Nucleotide Exchange Factor: EFFECT ON PKA LOCALIZATION AND P-Rex1 SIGNALING. J Biol Chem 2016; 291:6182-99. [PMID: 26797121 DOI: 10.1074/jbc.m115.712216] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2015] [Indexed: 12/15/2022] Open
Abstract
Morphology of migrating cells is regulated by Rho GTPases and fine-tuned by protein interactions and phosphorylation. PKA affects cell migration potentially through spatiotemporal interactions with regulators of Rho GTPases. Here we show that the endogenous regulatory (R) subunit of type I PKA interacts with P-Rex1, a Rac guanine nucleotide exchange factor that integrates chemotactic signals. Type I PKA holoenzyme interacts with P-Rex1 PDZ domains via the CNB B domain of RIα, which when expressed by itself facilitates endothelial cell migration. P-Rex1 activation localizes PKA to the cell periphery, whereas stimulation of PKA phosphorylates P-Rex1 and prevents its activation in cells responding to SDF-1 (stromal cell-derived factor 1). The P-Rex1 DEP1 domain is phosphorylated at Ser-436, which inhibits the DH-PH catalytic cassette by direct interaction. In addition, the P-Rex1 C terminus is indirectly targeted by PKA, promoting inhibitory interactions independently of the DEP1-PDZ2 region. A P-Rex1 S436A mutant construct shows increased RacGEF activity and prevents the inhibitory effect of forskolin on sphingosine 1-phosphate-dependent endothelial cell migration. Altogether, these results support the idea that P-Rex1 contributes to the spatiotemporal localization of type I PKA, which tightly regulates this guanine exchange factor by a multistep mechanism, initiated by interaction with the PDZ domains of P-Rex1 followed by direct phosphorylation at the first DEP domain and putatively indirect regulation of the C terminus, thus promoting inhibitory intramolecular interactions. This reciprocal regulation between PKA and P-Rex1 might represent a key node of integration by which chemotactic signaling is fine-tuned by PKA.
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Affiliation(s)
| | - Sendi Rafael Adame-García
- Cell Biology, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV-IPN), Mexico City, 07360 Mexico
| | | | - Alejandro Castillo-Kauil
- Cell Biology, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV-IPN), Mexico City, 07360 Mexico
| | | | - Shigetomo Fukuhara
- Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute (NCVC), Osaka, 565-8565 Japan, and
| | - Susan S Taylor
- Departments of Chemistry and Biochemistry and Pharmacology, University of California San Diego, La Jolla, California 92093
| | - Naoki Mochizuki
- Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute (NCVC), Osaka, 565-8565 Japan, and
| | - Guadalupe Reyes-Cruz
- Cell Biology, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV-IPN), Mexico City, 07360 Mexico
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Abstract
SUMMARY Stimuli that promote cell migration, such as chemokines, cytokines, and growth factors in metazoans and cyclic AMP in Dictyostelium, activate signaling pathways that control organization of the actin cytoskeleton and adhesion complexes. The Rho-family GTPases are a key convergence point of these pathways. Their effectors include actin regulators such as formins, members of the WASP/WAVE family and the Arp2/3 complex, and the myosin II motor protein. Pathways that link to the Rho GTPases include Ras GTPases, TorC2, and PI3K. Many of the molecules involved form gradients within cells, which define the front and rear of migrating cells, and are also established in related cellular behaviors such as neuronal growth cone extension and cytokinesis. The signaling molecules that regulate migration can be integrated to provide a model of network function. The network displays biochemical excitability seen as spontaneous waves of activation that propagate along the cell cortex. These events coordinate cell movement and can be biased by external cues to bring about directed migration.
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Affiliation(s)
- Peter Devreotes
- Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
| | - Alan Rick Horwitz
- Department of Cell Biology, University of Virginia School of Medicine, Charlottesville, Virginia 22908
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Ha JM, Yun SJ, Kim YW, Jin SY, Lee HS, Song SH, Shin HK, Bae SS. Platelet-derived growth factor regulates vascular smooth muscle phenotype via mammalian target of rapamycin complex 1. Biochem Biophys Res Commun 2015; 464:57-62. [DOI: 10.1016/j.bbrc.2015.05.097] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2015] [Accepted: 05/28/2015] [Indexed: 10/23/2022]
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Yan H, Wu W, Ge H, Li P, Wang Z. Up-Regulation of miR-204 Enhances Anoikis Sensitivity in Epithelial Ovarian Cancer Cell Line Via Brain-Derived Neurotrophic Factor Pathway In Vitro. Int J Gynecol Cancer 2015; 25:944-52. [PMID: 25962115 DOI: 10.1097/igc.0000000000000456] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
OBJECTIVE Genomic loci encoding miR-204, which was predicted to target brain-derived neurotrophic factor (BDNF), were frequently lost in multiple cancer, including epithelial ovarian cancer (EOC). In this study, we aimed to find out the influence of miR-204 expression level on EOC cell anoikis sensitivity and to explore possible mechanisms of this process. METHODS First, we screened EOC cells, which maintain anoikis resistance forming an anoikis pattern. miR-204 expression level and apoptosis were measured, respectively, by quantitative reverse transcriptase polymerase chain reaction and Annexin-V-R-PE/7-amino-actinomycin assay. Then we restored the expression level of miR-204 by transfection with pre-miR-204. miR-204 expression level and apoptosis were measured as before; cell invasion and migration ability were detected by transwell invasion assay and wound-healing assay. The messenger RNA level of BDNF was also detected by quantitative reverse transcriptase polymerase chain reaction; Western blot analysis was performed to assess pAKT expression. RESULTS Expression of miR-204 is significantly down-regulated in an anoikis pattern. Restored expression level of miR-204 enables cells to acquire more sensitivity to anoikis and decrease invasive and metastatic behavior, and also results in BDNF down-expression and inhibits activation of mitochondria-dependent pathway through the PI3K/AKT signaling pathway leading to cancer cell anoikis in EOC cells. CONCLUSIONS miR-204 up-regulation may be linked directly to the sensitivity of EOC cell anoikis by contributing to BDNF down-regulation. Our findings provide a novel mechanism for manipulating miR-204 levels therapeutically to restore anoikis sensitivity.
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Affiliation(s)
- Hongliang Yan
- *Hebei United University, Tangshan, PR China; †Department of Gynaecology, the Affiliated Hospital of the Chinese People's Armed Forces Logistic College, Tianjin, PR China; ‡Healthy Care Center of Women and Children of Tianjin Beichen, Tianjin, PR China; §Biomechanics Laboratory of Orthopaedic Research Institute of Tian Jin Hospital, Tianjin, China; and ∥Hebei United University, Tangshan, PR China
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50
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Lucato CM, Halls ML, Ooms LM, Liu HJ, Mitchell CA, Whisstock JC, Ellisdon AM. The Phosphatidylinositol (3,4,5)-Trisphosphate-dependent Rac Exchanger 1·Ras-related C3 Botulinum Toxin Substrate 1 (P-Rex1·Rac1) Complex Reveals the Basis of Rac1 Activation in Breast Cancer Cells. J Biol Chem 2015; 290:20827-20840. [PMID: 26112412 DOI: 10.1074/jbc.m115.660456] [Citation(s) in RCA: 45] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2015] [Indexed: 12/16/2022] Open
Abstract
The P-Rex (phosphatidylinositol (3,4,5)-trisphosphate (PIP3)-dependent Rac exchanger) family (P-Rex1 and P-Rex2) of the Rho guanine nucleotide exchange factors (Rho GEFs) activate Rac GTPases to regulate cell migration, invasion, and metastasis in several human cancers. The family is unique among Rho GEFs, as their activity is regulated by the synergistic binding of PIP3 and Gβγ at the plasma membrane. However, the molecular mechanism of this family of multi-domain proteins remains unclear. We report the 1.95 Å crystal structure of the catalytic P-Rex1 DH-PH tandem domain in complex with its cognate GTPase, Rac1 (Ras-related C3 botulinum toxin substrate-1). Mutations in the P-Rex1·Rac1 interface revealed a critical role for this complex in signaling downstream of receptor tyrosine kinases and G protein-coupled receptors. The structural data indicated that the PIP3/Gβγ binding sites are on the opposite surface and markedly removed from the Rac1 interface, supporting a model whereby P-Rex1 binding to PIP3 and/or Gβγ releases inhibitory C-terminal domains to expose the Rac1 binding site.
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Affiliation(s)
- Christina M Lucato
- Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, Victoria 3800, Australia
| | - Michelle L Halls
- Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, Victoria 3052, Australia
| | - Lisa M Ooms
- Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia
| | - Heng-Jia Liu
- Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia
| | - Christina A Mitchell
- Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia.
| | - James C Whisstock
- Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, Victoria 3800, Australia.
| | - Andrew M Ellisdon
- Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, Victoria 3800, Australia.
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