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Ling LJ, Li MD, Lu JW, Zhang F, Pan F, Su Y, Myatt L, Wang WS, Sun K, Ying H. Induction of epithelial cell senescence by SERPINE1 derived from fibroblasts in the amnion at parturition. Mech Ageing Dev 2025; 225:112053. [PMID: 40132749 DOI: 10.1016/j.mad.2025.112053] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2024] [Revised: 02/28/2025] [Accepted: 03/17/2025] [Indexed: 03/27/2025]
Abstract
Senescence of amnion epithelial cells not only disrupts the fetal membrane structure, but also becomes a source of proinflammatory signals contributing to membrane inflammation at parturition. However, the trigger initiating their senescence awaits identification. In this study, we found that SERPINE1 abundance was significantly increased in the amnion at parturition, where SERPINE1 was found predominantly expressed in amnion fibroblasts. SERPINE1 from amnion fibroblasts induced amnion epithelial cell senescence by causing vitronectin shedding from the cells thereby interrupting the association of vitronectin with integrin subunit αV, which led to the inhibition of the cell survival-associated focal adhesion pathway. In turn, proinflammatory cytokines such as interleukin-1β from senescent amnion epithelial cells enhanced SERPINE1 expression in amnion fibroblasts, thus forming a feed-forward loop between SERPINE1 production in amnion fibroblasts and epithelial cell senescence at parturition. Studies in the pregnant mice showed that intra-amniotic injection of SERPINE1 induced preterm birth with increased cellular senescence in the fetal membranes, which could be reversed by co-administration of vitronectin. Our findings indicate that SERPINE1 derived from amnion fibroblasts participates in the induction of amnion epithelial cell senescence at parturition. Intervening in the interaction of SERPINE1 with vitronectin may have therapeutic benefit in the treatment of preterm birth.
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Affiliation(s)
- Li-Jun Ling
- Department of Obstetrics, Shanghai First Maternity and Infant Hospital, School of Medicine, Tongji University, Shanghai, PR China; Shanghai Key Laboratory of Maternal Fetal Medicine, Shanghai Institute of Maternal-Fetal Medicine and Gynecologic Oncology, Shanghai, PR China
| | - Meng-Die Li
- Center for Reproductive Medicine, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China; Shanghai Key Laboratory for Assisted Reproduction and Reproductive Genetics, Shanghai, PR China
| | - Jiang-Wen Lu
- Center for Reproductive Medicine, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China; Shanghai Key Laboratory for Assisted Reproduction and Reproductive Genetics, Shanghai, PR China
| | - Fan Zhang
- Center for Reproductive Medicine, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China; Shanghai Key Laboratory for Assisted Reproduction and Reproductive Genetics, Shanghai, PR China
| | - Fan Pan
- Center for Reproductive Medicine, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China; Shanghai Key Laboratory for Assisted Reproduction and Reproductive Genetics, Shanghai, PR China
| | - Yao Su
- Department of Obstetrics, Shanghai First Maternity and Infant Hospital, School of Medicine, Tongji University, Shanghai, PR China; Shanghai Key Laboratory of Maternal Fetal Medicine, Shanghai Institute of Maternal-Fetal Medicine and Gynecologic Oncology, Shanghai, PR China
| | - Leslie Myatt
- Department of Obstetrics and Gynecology, Oregon Health & Science University, Portland, OR, USA
| | - Wang-Sheng Wang
- Center for Reproductive Medicine, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China; Shanghai Key Laboratory for Assisted Reproduction and Reproductive Genetics, Shanghai, PR China.
| | - Kang Sun
- Center for Reproductive Medicine, Ren Ji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China; Shanghai Key Laboratory for Assisted Reproduction and Reproductive Genetics, Shanghai, PR China.
| | - Hao Ying
- Department of Obstetrics, Shanghai First Maternity and Infant Hospital, School of Medicine, Tongji University, Shanghai, PR China; Shanghai Key Laboratory of Maternal Fetal Medicine, Shanghai Institute of Maternal-Fetal Medicine and Gynecologic Oncology, Shanghai, PR China.
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Katoh K. Signal Transduction Mechanisms of Focal Adhesions: Src and FAK-Mediated Cell Response. FRONT BIOSCI-LANDMRK 2024; 29:392. [PMID: 39614431 DOI: 10.31083/j.fbl2911392] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2024] [Revised: 08/02/2024] [Accepted: 08/05/2024] [Indexed: 12/01/2024]
Abstract
Cell-to-substrate adhesion sites, also known as focal adhesion sites (FAs), are complexes of different proteins on the cell surface. FAs play important roles in communication between cells and the extracellular matrix (ECM), leading to signal transduction involving different proteins that ultimately produce the cell response. This cell response involves cell adhesion, migration, motility, cell survival, and cell proliferation. The most important component of FAs are integrins. Integrins are transmembrane proteins that receive signals from the ECM and communicate them to the cytoplasm, thus activating several downstream proteins in a signaling cascade. Cellular Proto-oncogene tyrosine-protein kinase Src (c-Src) and focal adhesion kinase (FAK) are non-receptor tyrosine kinases that functionally interact to promote crucial roles in FAs. c-Src is a tyrosine kinase, activated by autophosphorylation and, in turn, activates another important protein, FAK. Activated FAK directly interacts with the cytoplasmic domain of integrin and activates other FA proteins by attaching to them. These proteins activated by FAK then activate other downstream pathways such as mitogen-activated protein kinase (MAPK) and Akt pathways involved in cell proliferation, migration, and cell survival. Src can induce detachment of FAK from the integrin to increase the focal adhesion turnover. As a result, the Src-FAK complex in FAs is critical for cell adhesion and survival mechanisms. Overexpression of FA proteins has been linked to a variety of pathological disorders, including cancers, growth retardation, and bone deformities. FAK and Src are overexpressed in various cancers. This review, which focuses on the roles of two important signaling proteins, c-Src and FAK, attempts to provide a thorough and up-to-date examination of the signal transduction mechanisms mediated by focal adhesions. The author also described that FAK and Src may serve as potential targets for future therapies against diseases associated with their overexpression, such as certain types of cancer.
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Affiliation(s)
- Kazuo Katoh
- Laboratory of Human Anatomy and Cell Biology, Faculty of Health Sciences, Tsukuba University of Technology, 305-8521 Tsukuba, Japan
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3
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Feng Z, Wei W, Wang S, Li X, Zhao L, Wan G, Hu R, Yu L. A novel selective FAK inhibitor E2 inhibits ovarian cancer metastasis and growth by inducing cytotoxic autophagy. Biochem Pharmacol 2024; 229:116461. [PMID: 39102992 DOI: 10.1016/j.bcp.2024.116461] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2024] [Revised: 07/28/2024] [Accepted: 08/01/2024] [Indexed: 08/07/2024]
Abstract
Ovarian cancer (OC) is the deadliest form of the gynecologic malignancies and effective therapeutic drugs are urgently needed. Focal adhesion kinase (FAK) is overexpressed in various solid tumors, and could serve as a potential biomarker of ovarian cancer. However, there are no launched drugs targeting FAK. Hence, the development of the novel FAK inhibitors is an emerging approach for the treatment of ovarian cancer. In this work, we characterized a selective FAK inhibitor E2, with a high inhibitory potency toward FAK. Moreover, E2 had cytotoxic, anti-invasion and anti-migration activity on ovarian cancer cells. Mechanistically, after treatment with E2, FAK downstream signaling cascades (e.g., Src and AKT) were suppressed, thus resulting in the ovarian cancer cell arrest at G0/G1 phase and the induction of cytotoxic autophagy. In addition, E2 attenuated the tumor growth of PA-1 and ES-2 ovarian cancer subcutaneous xenografts, as well as suppressed peritoneal metastasis of OVCAR3-luc. Furthermore, E2 exhibited favorable pharmacokinetic properties. Altogether, these findings demonstrate that E2 is a selective FAK inhibitor with potent anti-ovarian cancer activities both in vivo and in vitro, offering new possibilities for OC treatment strategies.
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Affiliation(s)
- Zhanzhan Feng
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Wei Wei
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Shirui Wang
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Xiao Li
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Lifeng Zhao
- Sichuan Industrial Institute of Antibiotics, Chengdu University, Chengdu 610106, China
| | - Guoquan Wan
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Rong Hu
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China
| | - Luoting Yu
- Department of Biotherapy, Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China.
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Thomasy SM, Leonard BC, Greiner MA, Skeie JM, Raghunathan VK. Squishy matters - Corneal mechanobiology in health and disease. Prog Retin Eye Res 2024; 99:101234. [PMID: 38176611 PMCID: PMC11193890 DOI: 10.1016/j.preteyeres.2023.101234] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2023] [Revised: 12/22/2023] [Accepted: 12/27/2023] [Indexed: 01/06/2024]
Abstract
The cornea, as a dynamic and responsive tissue, constantly interacts with mechanical forces in order to maintain its structural integrity, barrier function, transparency and refractive power. Cells within the cornea sense and respond to various mechanical forces that fundamentally regulate their morphology and fate in development, homeostasis and pathophysiology. Corneal cells also dynamically regulate their extracellular matrix (ECM) with ensuing cell-ECM crosstalk as the matrix serves as a dynamic signaling reservoir providing biophysical and biochemical cues to corneal cells. Here we provide an overview of mechanotransduction signaling pathways then delve into the recent advances in corneal mechanobiology, focusing on the interplay between mechanical forces and responses of the corneal epithelial, stromal, and endothelial cells. We also identify species-specific differences in corneal biomechanics and mechanotransduction to facilitate identification of optimal animal models to study corneal wound healing, disease, and novel therapeutic interventions. Finally, we identify key knowledge gaps and therapeutic opportunities in corneal mechanobiology that are pressing for the research community to address especially pertinent within the domains of limbal stem cell deficiency, keratoconus and Fuchs' endothelial corneal dystrophy. By furthering our understanding corneal mechanobiology, we can contextualize discoveries regarding corneal diseases as well as innovative treatments for them.
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Affiliation(s)
- Sara M Thomasy
- Department of Surgical and Radiological Sciences, School of Veterinary Medicine, University of California - Davis, Davis, CA, United States; Department of Ophthalmology & Vision Science, School of Medicine, University of California - Davis, Davis, CA, United States; California National Primate Research Center, Davis, CA, United States.
| | - Brian C Leonard
- Department of Surgical and Radiological Sciences, School of Veterinary Medicine, University of California - Davis, Davis, CA, United States; Department of Ophthalmology & Vision Science, School of Medicine, University of California - Davis, Davis, CA, United States
| | - Mark A Greiner
- Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IA, United States; Iowa Lions Eye Bank, Coralville, IA, United States
| | - Jessica M Skeie
- Department of Ophthalmology and Visual Sciences, Carver College of Medicine, University of Iowa, Iowa City, IA, United States; Iowa Lions Eye Bank, Coralville, IA, United States
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Ng WH, Soo KC, Huynh H. Vinorelbine Improves the Efficacy of Sorafenib against Hepatocellular Carcinoma: A Promising Therapeutic Approach. Int J Mol Sci 2024; 25:1563. [PMID: 38338842 PMCID: PMC10855313 DOI: 10.3390/ijms25031563] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2023] [Revised: 01/22/2024] [Accepted: 01/25/2024] [Indexed: 02/12/2024] Open
Abstract
Hepatocellular carcinoma (HCC) is a leading global cause of cancer-related mortality. Despite the widespread adoption of sorafenib as the standard HCC treatment, its efficacy is constrained, frequently encountering resistance. To augment the effectiveness of sorafenib, this study investigated the synergy of sorafenib and vinorelbine using 22 HCC patient-derived xenograft (PDX) models. In this study, mice bearing HCC tumors were treated with the vehicle, sorafenib (15 mg/kg), vinorelbine (3 mg/kg), and sorafenib-vinorelbine combination (Sora/Vino). Rigorous monitoring of the tumor growth and side effects coupled with comprehensive histological and molecular analyses was conducted. The overall survival (OS) of mice bearing HCC orthotopic tumors was also assessed. Our data showed a notable 86.4% response rate to Sora/Vino, surpassing rates of 31.8% for sorafenib and 9.1% for vinorelbine monotherapies. Sora/Vino significantly inhibited tumor growth, prolonged OS of mice bearing HCC orthotopic tumors (p < 0.01), attenuated tumor cell proliferation and angiogenesis, and enhanced necrosis and apoptosis. The combination therapy effectively suppressed the focal adhesion kinase (FAK) pathway, which is a pivotal player in cell proliferation, tumor angiogenesis, survival, and metastasis. The noteworthy antitumor activity in 22 HCC PDX models positions Sora/Vino as a promising candidate for early-phase clinical trials, leveraging the established use of sorafenib and vinorelbine in HCC and other cancers.
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Affiliation(s)
- Wai Har Ng
- Laboratory of Molecular Endocrinology, National Cancer Centre Singapore, Singapore 168583, Singapore;
| | - Khee Chee Soo
- Division of Surgery and Surgical Oncology, National Cancer Centre Singapore, Singapore 168583, Singapore
| | - Hung Huynh
- Laboratory of Molecular Endocrinology, National Cancer Centre Singapore, Singapore 168583, Singapore;
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Tan X, Yan Y, Song B, Zhu S, Mei Q, Wu K. Focal adhesion kinase: from biological functions to therapeutic strategies. Exp Hematol Oncol 2023; 12:83. [PMID: 37749625 PMCID: PMC10519103 DOI: 10.1186/s40164-023-00446-7] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2023] [Accepted: 09/11/2023] [Indexed: 09/27/2023] Open
Abstract
Focal adhesion kinase (FAK), a nonreceptor cytoplasmic tyrosine kinase, is a vital participant in primary cellular functions, such as proliferation, survival, migration, and invasion. In addition, FAK regulates cancer stem cell activities and contributes to the formation of the tumor microenvironment (TME). Importantly, increased FAK expression and activity are strongly associated with unfavorable clinical outcomes and metastatic characteristics in numerous tumors. In vitro and in vivo studies have demonstrated that modulating FAK activity by application of FAK inhibitors alone or in combination treatment regimens could be effective for cancer therapy. Based on these findings, several agents targeting FAK have been exploited in diverse preclinical tumor models. This article briefly describes the structure and function of FAK, as well as research progress on FAK inhibitors in combination therapies. We also discuss the challenges and future directions regarding anti-FAK combination therapies.
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Affiliation(s)
- Ximin Tan
- Department of Oncology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Yuheng Yan
- Department of Oncology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Bin Song
- Cancer Center, Shanxi Bethune Hospital, Shanxi Academy of Medical Science, Tongji Shanxi Hospital, Third Hospital of Shanxi Medical University, Taiyuan, 030032, China
| | - Shuangli Zhu
- Department of Oncology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Qi Mei
- Cancer Center, Shanxi Bethune Hospital, Shanxi Academy of Medical Science, Tongji Shanxi Hospital, Third Hospital of Shanxi Medical University, Taiyuan, 030032, China.
| | - Kongming Wu
- Cancer Center, Shanxi Bethune Hospital, Shanxi Academy of Medical Science, Tongji Shanxi Hospital, Third Hospital of Shanxi Medical University, Taiyuan, 030032, China.
- Cancer Center, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China.
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Chen Y, Jin L, Ma Y, Liu Y, Zhu Q, Huang Y, Feng W. BACH1 promotes lung adenocarcinoma cell metastasis through transcriptional activation of ITGA2. Cancer Sci 2023; 114:3568-3582. [PMID: 37311571 PMCID: PMC10475762 DOI: 10.1111/cas.15884] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2023] [Revised: 05/24/2023] [Accepted: 05/31/2023] [Indexed: 06/15/2023] Open
Abstract
BACH1 plays an important role in promoting cancer. This study aims to further verify the relationship between the expression level of BACH1 in lung adenocarcinoma prognosis, as well as the influence of BACH1 expression on lung adenocarcinoma and the potential mechanism. The expression level of BACH1 in lung adenocarcinoma and its relationship with prognosis was evaluated by lung adenocarcinoma tissue microarray analysis combined with bioinformatics approaches. Gene knockdown and overexpression were used to investigate the functions and molecular mechanisms of BACH1 in lung adenocarcinoma cells. The regulatory downstream pathways and target genes of BACH1 in lung adenocarcinoma cells were explored by bioinformatics and RNA sequencing data analysis, real-time PCR, western blot analysis, and cell immunofluorescence and cell adhesion assays. Chromatin immunoprecipitation and dual-luciferase reporter assays were carried out to verify the target gene binding site. In the present study, BACH1 is abnormally highly expressed in lung adenocarcinoma tissues, and high BACH1 expression is negatively correlated with patient prognosis. BACH1 promotes the migration and invasion of lung adenocarcinoma cells. Mechanistically, BACH1 directly binds to the upstream sequence of the ITGA2 promoter to promote ITGA2 expression, and the BACH1-ITGA2 axis is involved in cytoskeletal regulation in lung adenocarcinoma cells by activating the FAK-RAC1-PAK signaling pathway. Our results indicated that BACH1 positively regulates the expression of ITGA2 through a transcriptional mechanism, thereby activating the FAK-RAC1-PAK signaling pathway to participate in the formation of the cytoskeleton in tumor cells and then promoting the migration and invasion of tumor cells.
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Affiliation(s)
- Yingji Chen
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
| | - Longyu Jin
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
| | - Yuchao Ma
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
| | - Yicai Liu
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
| | - Qianjun Zhu
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
| | - Yu Huang
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
| | - Wei Feng
- Department of Cardiothoracic SurgeryThird Xiangya Hospital of Central South UniversityChangshaChina
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Fard D, Testa E, Panzeri V, Rizzolio S, Bianchetti G, Napolitano V, Masciarelli S, Fazi F, Maulucci G, Scicchitano BM, Sette C, Viscomi MT, Tamagnone L. SEMA6C: a novel adhesion-independent FAK and YAP activator, required for cancer cell viability and growth. Cell Mol Life Sci 2023; 80:111. [PMID: 37002363 PMCID: PMC10066115 DOI: 10.1007/s00018-023-04756-1] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2022] [Revised: 02/22/2023] [Accepted: 03/11/2023] [Indexed: 04/03/2023]
Abstract
Transmembrane semaphorins are signaling molecules, controlling axonal wiring and embryo development, which are increasingly implicated in human diseases. Semaphorin 6C (Sema6C) is a poorly understood family member and its functional role is still unclear. Upon targeting Sema6C expression in a range of cancer cells, we observed dramatic growth suppression, decreased ERK phosphorylation, upregulation of cell cycle inhibitor proteins p21, p27 and p53, and the onset of cell senescence, associated with activation of autophagy. These data are consistent with a fundamental requirement for Sema6C to support viability and growth in cancer cells. Mechanistically, we unveiled a novel signaling pathway elicited by Sema6C, and dependent on its intracellular domain, mediated by tyrosine kinases c-Abl and Focal Adhesion Kinase (FAK). Sema6C was found in complex with c-Abl, and induced its phosphorylation, which in turn led to FAK activation, independent of cell-matrix adhesion. Sema6C-induced FAK activity was furthermore responsible for increased nuclear localization of YAP transcriptional regulator. Moreover, Sema6C conferred YAP signaling-dependent long-term cancer cell survival upon nutrient deprivation. In conclusion, our findings demonstrate that Sema6C elicits a cancer promoting-signaling pathway sustaining cell viability and self-renewal, independent of growth factors and nutrients availability.
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Affiliation(s)
- Damon Fard
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy
| | - Erika Testa
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy
| | - Valentina Panzeri
- Department of Neuroscience, Università Cattolica del Sacro Cuore, Rome, Italy
| | | | - Giada Bianchetti
- Department of Neuroscience, Università Cattolica del Sacro Cuore, Rome, Italy
| | - Virginia Napolitano
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy
| | - Silvia Masciarelli
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy
- Department of Anatomical, Histological, Forensic and Orthopaedic Sciences, Section of Histology and Medical Embryology, Sapienza University of Rome, Rome, Italy
| | - Francesco Fazi
- Department of Anatomical, Histological, Forensic and Orthopaedic Sciences, Section of Histology and Medical Embryology, Sapienza University of Rome, Rome, Italy
| | - Giuseppe Maulucci
- Department of Neuroscience, Università Cattolica del Sacro Cuore, Rome, Italy
- Fondazione Policlinico Gemelli-IRCCS, Rome, Italy
| | - Bianca Maria Scicchitano
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy
- Fondazione Policlinico Gemelli-IRCCS, Rome, Italy
| | - Claudio Sette
- Department of Neuroscience, Università Cattolica del Sacro Cuore, Rome, Italy
- Fondazione Policlinico Gemelli-IRCCS, Rome, Italy
| | - Maria Teresa Viscomi
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy
- Fondazione Policlinico Gemelli-IRCCS, Rome, Italy
| | - Luca Tamagnone
- Department of Life Sciences and Public Health, Università Cattolica del Sacro Cuore, Rome, Italy.
- Fondazione Policlinico Gemelli-IRCCS, Rome, Italy.
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Nishinakagawa T, Hazekawa M, Hosokawa M, Ishibashi D. RCAS1 increases cell morphological changes in murine fibroblasts by reducing p38 phosphorylation. Mol Med Rep 2023; 27:62. [PMID: 36734265 PMCID: PMC9926866 DOI: 10.3892/mmr.2023.12949] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2022] [Accepted: 12/09/2022] [Indexed: 02/04/2023] Open
Abstract
Receptor‑binding cancer antigen expressed on SiSo cells (RCAS1) is a tumor‑associated antigen that is expressed in a number of human malignancies. RCAS1 acts as a ligand for a putative RCAS1 receptor that is present on various human cells including T and B lymphocytes and natural killer cells, in which it induces cell growth inhibition and apoptosis. It has been suggested that RCAS1 might serve an important role in tumor cell evasion from the host immune system. In fact, RCAS1 expression is related to malignant characteristics including tumor size, invasion depth, clinical stage and poor overall survival. The authors previously established doxycycline‑induced RCAS1 overexpression murine fibroblast L cells to analyze the biological functions of RCAS1 and reported that its expression inhibited cell cycle progression via the downregulation of cyclin D3, which subsequently induced apoptosis. Additionally, it was found that RCAS1 expression induced cell morphological changes prior to caspase‑mediated apoptosis. Thus, the present study examined signaling pathways associated with changes in cell morphology that were induced by RCAS1 expression. The data showed that increased RCAS1 expression caused a reduction in actin stress fibers and decreased cofilin phosphorylation. Recent studies have shown that p38 signaling regulates actin polymerization. The data the present study showed that increased RCAS1 expression significantly decreased p38 phosphorylation.
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Affiliation(s)
- Takuya Nishinakagawa
- Department of Immunological and Molecular Pharmacology, Faculty of Pharmaceutical Science, Fukuoka University, Fukuoka 814-0180, Japan,Correspondence to: Dr Takuya Nishinakagawa or Professor Daisuke Ishibashi, Department of Immunological and Molecular Pharmacology, Faculty of Pharmaceutical Science, Fukuoka University, 8-19-1 Nanakuma, Jonan-ku, Fukuoka 814-0180, Japan, E-mail:
| | - Mai Hazekawa
- Department of Immunological and Molecular Pharmacology, Faculty of Pharmaceutical Science, Fukuoka University, Fukuoka 814-0180, Japan
| | - Masato Hosokawa
- Department of Immunological and Molecular Pharmacology, Faculty of Pharmaceutical Science, Fukuoka University, Fukuoka 814-0180, Japan
| | - Daisuke Ishibashi
- Department of Immunological and Molecular Pharmacology, Faculty of Pharmaceutical Science, Fukuoka University, Fukuoka 814-0180, Japan,Correspondence to: Dr Takuya Nishinakagawa or Professor Daisuke Ishibashi, Department of Immunological and Molecular Pharmacology, Faculty of Pharmaceutical Science, Fukuoka University, 8-19-1 Nanakuma, Jonan-ku, Fukuoka 814-0180, Japan, E-mail:
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10
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Zhang Z, Li J, Jiao S, Han G, Zhu J, Liu T. Functional and clinical characteristics of focal adhesion kinases in cancer progression. Front Cell Dev Biol 2022; 10:1040311. [PMID: 36407100 PMCID: PMC9666724 DOI: 10.3389/fcell.2022.1040311] [Citation(s) in RCA: 18] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2022] [Accepted: 10/18/2022] [Indexed: 11/07/2022] Open
Abstract
Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase and an adaptor protein that primarily regulates adhesion signaling and cell migration. FAK promotes cell survival in response to stress. Increasing evidence has shown that at the pathological level, FAK is highly expressed in multiple tumors in several systems (including lung, liver, gastric, and colorectal cancers) and correlates with tumor aggressiveness and patient prognosis. At the molecular level, FAK promotes tumor progression mainly by altering survival signals, invasive capacity, epithelial-mesenchymal transition, the tumor microenvironment, the Warburg effect, and stemness of tumor cells. Many effective drugs have been developed based on the comprehensive role of FAK in tumor cells. In addition, its potential as a tumor marker cannot be ignored. Here, we discuss the pathological and pre-clinical evidence of the role of FAK in cancer development; we hope that these findings will assist in FAK-based clinical studies.
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Affiliation(s)
- Zhaoyu Zhang
- Department of Gastrointestinal Nutrition and Hernia Surgery, The Second Hospital of Jilin University, Changchun, Jilin, China
| | - Jinlong Li
- Department of Gastrointestinal Nutrition and Hernia Surgery, The Second Hospital of Jilin University, Changchun, Jilin, China
| | - Simin Jiao
- Department of Gastrointestinal Nutrition and Hernia Surgery, The Second Hospital of Jilin University, Changchun, Jilin, China
| | - Guangda Han
- Department of Gastrointestinal Nutrition and Hernia Surgery, The Second Hospital of Jilin University, Changchun, Jilin, China
| | - Jiaming Zhu
- Department of Surgical Oncology and General Surgery, The First Affiliated Hospital of China Medical University, Shenyang, Liaoning, China
| | - Tianzhou Liu
- Department of Gastrointestinal Nutrition and Hernia Surgery, The Second Hospital of Jilin University, Changchun, Jilin, China
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11
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Ulyanova T, Cherone JM, Sova P, Papayannopoulou T. α4-Integrin deficiency in human CD34+ cells engenders precocious erythroid differentiation but inhibits enucleation. Exp Hematol 2022; 108:16-25. [DOI: 10.1016/j.exphem.2022.01.007] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2021] [Revised: 01/21/2022] [Accepted: 01/25/2022] [Indexed: 11/26/2022]
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Microglial Cytokines Induce Invasiveness and Proliferation of Human Glioblastoma through Pyk2 and FAK Activation. Cancers (Basel) 2021; 13:cancers13246160. [PMID: 34944779 PMCID: PMC8699228 DOI: 10.3390/cancers13246160] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2021] [Revised: 11/29/2021] [Accepted: 12/03/2021] [Indexed: 12/25/2022] Open
Abstract
Simple Summary Microglia infiltrate most gliomas and have been demonstrated to promote tumor growth, invasion, and treatment resistance. To develop improved treatment methods, that take into consideration the supporting role of microglia in tumor progression, the functional and mechanistic pathways of glioma–microglia interactions need to be identified and experimentally dissected. Our recent studies and literature reports revealed the overexpression of Pyk2 and FAK in glioblastomas. Pyk2 and FAK signaling pathways have been shown to regulate migration and proliferation in glioma cells, including microglia-promoted glioma cell migration. However, the specific factors released by microglia that modulate Pyk2 and FAK to promote glioma invasiveness and proliferation are poorly understood. The aim of this study was to identify key microglia-derived signaling molecules that induce the activation of Pyk2- and FAK-dependent glioma cell proliferation and invasiveness. Abstract Glioblastoma is the most aggressive brain tumor in adults. Multiple lines of evidence suggest that microglia create a microenvironment favoring glioma invasion and proliferation. Our previous studies and literature reports indicated the involvement of focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (Pyk2) in glioma cell proliferation and invasion, stimulated by tumor-infiltrating microglia. However, the specific microglia-released factors that modulate Pyk2 and FAK signaling in glioma cells are unknown. In this study, 20 human glioblastoma specimens were evaluated with the use of RT-PCR and western blotting. A Pierson correlation test demonstrated a correlation (0.6–1.0) between the gene expression levels for platelet-derived growth factor β(PDGFβ), stromal-derived factor 1α (SDF-1α), IL-6, IL-8, and epidermal growth factor (EGF) in tumor-purified microglia and levels of p-Pyk2 (Y579/Y580) and p-FAK(Y925) in glioma cells. siRNA knockdown against Pyk2 or FAK in three primary glioblastoma cell lines, developed from the investigated specimens, in combination with the cytokine receptor inhibitors gefitinib (1 μM), DMPQ (200 nM), and burixafor (1 μM) identified EGF, PDGFβ, and SDF-1α as key extracellular factors in the Pyk2- and FAK-dependent activation of invadopodia formation and the migration of glioma cells. EGF and IL-6 were identified as regulators of the Pyk2- and FAK-dependent activation of cell viability and mitosis.
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Márquez-Ortiz RA, Contreras-Zárate MJ, Tesic V, Alvarez-Eraso KLF, Kwak G, Littrell Z, Costello JC, Sreekanth V, Ormond DR, Karam SD, Kabos P, Cittelly DM. IL13Rα2 Promotes Proliferation and Outgrowth of Breast Cancer Brain Metastases. Clin Cancer Res 2021; 27:6209-6221. [PMID: 34544797 PMCID: PMC8595859 DOI: 10.1158/1078-0432.ccr-21-0361] [Citation(s) in RCA: 18] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2021] [Revised: 08/05/2021] [Accepted: 09/16/2021] [Indexed: 01/09/2023]
Abstract
PURPOSE The survival of women with brain metastases (BM) from breast cancer remains very poor, with over 80% dying within a year of their diagnosis. Here, we define the function of IL13Rα2 in outgrowth of breast cancer brain metastases (BCBM) in vitro and in vivo, and postulate IL13Rα2 as a suitable therapeutic target for BM. EXPERIMENTAL DESIGN We performed IHC staining of IL13Rα2 in BCBM to define its prognostic value. Using inducible shRNAs in TNBC and HER2+ breast-brain metastatic models, we assessed IL13Rα2 function in vitro and in vivo. We performed RNAseq and functional studies to define the molecular mechanisms underlying IL13Rα2 function in BCBM. RESULTS High IL13Rα2 expression in BCBM predicted worse survival after BM diagnoses. IL13Rα2 was essential for cancer-cell survival, promoting proliferation while repressing invasion. IL13Rα2 KD resulted in FAK downregulation, repression of cell cycle and proliferation mediators, and upregulation of Ephrin B1 signaling. Ephrin-B1 (i) promoted invasion of BC cells in vitro, (ii) marked micrometastasis and invasive fronts in BCBM, and (iii) predicted shorter disease-free survival and BM-free survival (BMFS) in breast primary tumors known to metastasize to the brain. In experimental metastases models, which bypass early tumor invasion, downregulation of IL13Rα2 before or after tumor seeding and brain intravasation decreased BMs, suggesting that IL13Rα2 and the promotion of a proliferative phenotype is critical to BM progression. CONCLUSIONS Non-genomic phenotypic adaptations at metastatic sites are critical to BM progression and patients' prognosis. This study opens the road to use IL13Rα2 targeting as a therapeutic strategy for BM.
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Affiliation(s)
| | | | - Vesna Tesic
- Department of Pathology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | | | - Gina Kwak
- Department of Pathology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - Zachary Littrell
- Department of Pathology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - James C Costello
- Department of Pharmacology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - Varsha Sreekanth
- Department of Pharmacology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - D Ryan Ormond
- Department of Neurosurgery, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - Sana D Karam
- Department of Radiation Oncology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - Peter Kabos
- Department of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, Colorado
| | - Diana M Cittelly
- Department of Pathology, University of Colorado, Anschutz Medical Campus, Aurora, Colorado.
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Buhrmann C, Brockmueller A, Harsha C, Kunnumakkara AB, Kubatka P, Aggarwal BB, Shakibaei M. Evidence That Tumor Microenvironment Initiates Epithelial-To-Mesenchymal Transition and Calebin A can Suppress it in Colorectal Cancer Cells. Front Pharmacol 2021; 12:699842. [PMID: 34276382 PMCID: PMC8283792 DOI: 10.3389/fphar.2021.699842] [Citation(s) in RCA: 22] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2021] [Accepted: 06/23/2021] [Indexed: 12/12/2022] Open
Abstract
Background: Tumor microenvironment (TME) has a pivotal impact on tumor progression, and epithelial-mesenchymal transition (EMT) is an extremely crucial initial event in the metastatic process in colorectal cancer (CRC) that is not yet fully understood. Calebin A (an ingredient in Curcuma longa) has been shown to repress CRC tumor growth. However, whether Calebin A is able to abrogate TME-induced EMT in CRC was investigated based on the underlying pathways. Methods: CRC cell lines (HCT116, RKO) were exposed with Calebin A and/or a FAK inhibitor, cytochalasin D (CD) to investigate the action of Calebin A in TME-induced EMT-related tumor progression. Results: TME induced viability, proliferation, and increased invasiveness in 3D-alginate CRC cultures. In addition, TME stimulated stabilization of the master EMT-related transcription factor (Slug), which was accompanied by changes in the expression patterns of EMT-associated biomarkers. Moreover, TME resulted in stimulation of NF-κB, TGF-β1, and FAK signaling pathways. However, these effects were dramatically reduced by Calebin A, comparable to FAK inhibitor or CD. Finally, TME induced a functional association between NF-κB and Slug, suggesting that a synergistic interaction between the two transcription factors is required for initiation of EMT and tumor cell invasion, whereas Calebin A strongly inhibited this binding and subsequent CRC cell migration. Conclusion: We propose for the first time that Calebin A modulates TME-induced EMT in CRC cells, at least partially through the NF-κB/Slug axis, TGF-β1, and FAK signaling. Thus, Calebin A appears to be a potential agent for the prevention and management of CRC.
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Affiliation(s)
- Constanze Buhrmann
- Musculoskeletal Research Group and Tumor Biology, Chair of Vegetative Anatomy, Faculty of Medicine, Institute of Anatomy, Ludwig-Maximilian-University Munich, Munich, Germany.,Faculty of Medicine, Institute of Anatomy and Cell Biology, University of Augsburg, Augsburg, Germany
| | - Aranka Brockmueller
- Musculoskeletal Research Group and Tumor Biology, Chair of Vegetative Anatomy, Faculty of Medicine, Institute of Anatomy, Ludwig-Maximilian-University Munich, Munich, Germany
| | - Choudhary Harsha
- Cancer Biology Laboratory and DBT-AIST International Center for Translational and Environmental Research (DAICENTER), Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, India
| | - Ajaikumar B Kunnumakkara
- Cancer Biology Laboratory and DBT-AIST International Center for Translational and Environmental Research (DAICENTER), Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, India
| | - Peter Kubatka
- Department of Medical Biology, Jessenius Faculty of Medicine, Comenius University in Bratislava, Martin, Slovakia
| | | | - Mehdi Shakibaei
- Musculoskeletal Research Group and Tumor Biology, Chair of Vegetative Anatomy, Faculty of Medicine, Institute of Anatomy, Ludwig-Maximilian-University Munich, Munich, Germany
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15
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Adorno-Cruz V, Hoffmann AD, Liu X, Dashzeveg NK, Taftaf R, Wray B, Keri RA, Liu H. ITGA2 promotes expression of ACLY and CCND1 in enhancing breast cancer stemness and metastasis. Genes Dis 2021; 8:493-508. [PMID: 34179312 PMCID: PMC8209312 DOI: 10.1016/j.gendis.2020.01.015] [Citation(s) in RCA: 41] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/14/2019] [Revised: 01/22/2020] [Accepted: 01/26/2020] [Indexed: 02/07/2023] Open
Abstract
Cancer metastasis is largely incurable and accounts for 90% of breast cancer deaths, especially for the aggressive basal-like or triple negative breast cancer (TNBC). Combining patient database analyses and functional studies, we examined the association of integrin family members with clinical outcomes as well as their connection with previously identified microRNA regulators of metastasis, such as miR-206 that inhibits stemness and metastasis of TNBC. Here we report that the integrin receptor CD49b-encoding ITGA2, a direct target of miR-206, promotes breast cancer stemness and metastasis. ITGA2 knockdown suppressed self-renewal related mammosphere formation and pluripotency marker expression, inhibited cell cycling, compromised migration and invasion, and therefore decreased lung metastasis of breast cancer. ITGA2 overexpression reversed miR-206-caused cell cycle arrest in G1. RNA sequencing analyses revealed that ITGA2 knockdown inhibits genes related to cell cycle regulation and lipid metabolism, including CCND1 and ACLY as representative targets, respectively. Knockdown of CCND1 or ACLY inhibits mammosphere formation of breast cancer cells. Overexpression of CCND1 rescues the phenotype of ITGA2 knockdown-induced cell cycle arrest. ACLY-encoded ATP citrate lyase is essential to maintain cellular acetyl-CoA levels. CCND1 knockdown further mimics ITGA2 knockdown in abolishing lung colonization of breast cancer cells. We identified that the low levels of miR-206 as well as high expression levels of ITGA2, ACLY and CCND1 are associated with an unfavorable relapse-free survival of the patients with estrogen receptor-negative or high grade breast cancer, especially basal-like or TNBC, possibly serving as potential biomarkers of cancer stemness and therapeutic targets of breast cancer metastasis.
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Affiliation(s)
- Valery Adorno-Cruz
- Department of Pharmacology, Case Western Reserve University, Cleveland, OH 11318, USA
- Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
| | - Andrew D. Hoffmann
- Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
| | - Xia Liu
- Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
- University of Kentucky, College of Medicine, Lexington, KY 40536, USA
| | - Nurmaa K. Dashzeveg
- Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
| | - Rokana Taftaf
- Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
| | - Brian Wray
- Bioinformatic Core, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
| | - Ruth A. Keri
- Department of Pharmacology, Case Western Reserve University, Cleveland, OH 11318, USA
- Department of Genetics and Genome Sciences, The Division of General Medical Sciences-Oncology, Case Western Reserve University, Cleveland, OH 11318, USA
| | - Huiping Liu
- Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
- Department of Medicine, The Division of Hematology and Oncology, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
- Lurie Comprehensive Cancer Center, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA
- Department of Pathology and Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, OH 11318, USA
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Targeting RGD-binding integrins as an integrative therapy for diabetic retinopathy and neovascular age-related macular degeneration. Prog Retin Eye Res 2021; 85:100966. [PMID: 33775825 DOI: 10.1016/j.preteyeres.2021.100966] [Citation(s) in RCA: 35] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2021] [Revised: 03/15/2021] [Accepted: 03/19/2021] [Indexed: 12/14/2022]
Abstract
Integrins are a class of transmembrane receptors that are involved in a wide range of biological functions. Dysregulation of integrins has been implicated in many pathological processes and consequently, they are attractive therapeutic targets. In the ophthalmology arena, there is extensive evidence suggesting that integrins play an important role in diabetic retinopathy (DR), age-related macular degeneration (AMD), glaucoma, dry eye disease and retinal vein occlusion. For example, there is extensive evidence that arginyl-glycyl-aspartic acid (Arg-Gly-Asp; RGD)-binding integrins are involved in key disease hallmarks of DR and neovascular AMD (nvAMD), specifically inflammation, vascular leakage, angiogenesis and fibrosis. Based on such evidence, drugs that engage integrin-linked pathways have received attention for their potential to block all these vision-threatening pathways. This review focuses on the pathophysiological role that RGD-binding integrins can have in complex multifactorial retinal disorders like DR, diabetic macular edema (DME) and nvAMD, which are leading causes of blindness in developed countries. Special emphasis will be given on how RGD-binding integrins can modulate the intricate molecular pathways and regulate the underlying pathological mechanisms. For instance, the interplay between integrins and key molecular players such as growth factors, cytokines and enzymes will be summarized. In addition, recent clinical advances linked to targeting RGD-binding integrins in the context of DME and nvAMD will be discussed alongside future potential for limiting progression of these diseases.
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17
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Paul MR, Pan TC, Pant DK, Shih NN, Chen Y, Harvey KL, Solomon A, Lieberman D, Morrissette JJ, Soucier-Ernst D, Goodman NG, Stavropoulos SW, Maxwell KN, Clark C, Belka GK, Feldman M, DeMichele A, Chodosh LA. Genomic landscape of metastatic breast cancer identifies preferentially dysregulated pathways and targets. J Clin Invest 2020; 130:4252-4265. [PMID: 32657779 PMCID: PMC7410083 DOI: 10.1172/jci129941] [Citation(s) in RCA: 50] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2019] [Accepted: 05/05/2020] [Indexed: 12/21/2022] Open
Abstract
Nearly all breast cancer deaths result from metastatic disease. Despite this, the genomic events that drive metastatic recurrence are poorly understood. We performed whole-exome and shallow whole-genome sequencing to identify genes and pathways preferentially mutated or copy-number altered in metastases compared with the paired primary tumors from which they arose. Seven genes were preferentially mutated in metastases - MYLK, PEAK1, SLC2A4RG, EVC2, XIRP2, PALB2, and ESR1 - 5 of which are not significantly mutated in any type of human primary cancer. Four regions were preferentially copy-number altered: loss of STK11 and CDKN2A/B, as well as gain of PTK6 and the membrane-bound progesterone receptor, PAQR8. PAQR8 gain was mutually exclusive with mutations in the nuclear estrogen and progesterone receptors, suggesting a role in treatment resistance. Several pathways were preferentially mutated or altered in metastases, including mTOR, CDK/RB, cAMP/PKA, WNT, HKMT, and focal adhesion. Immunohistochemical analyses revealed that metastases preferentially inactivate pRB, upregulate the mTORC1 and WNT signaling pathways, and exhibit nuclear localization of activated PKA. Our findings identify multiple therapeutic targets in metastatic recurrence that are not significantly mutated in primary cancers, implicate membrane progesterone signaling and nuclear PKA in metastatic recurrence, and provide genomic bases for the efficacy of mTORC1, CDK4/6, and PARP inhibitors in metastatic breast cancer.
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Affiliation(s)
- Matt R. Paul
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | - Tien-chi Pan
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | - Dhruv K. Pant
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | - Natalie N.C. Shih
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Pathology and Laboratory Medicine
| | - Yan Chen
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | - Kyra L. Harvey
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | - Aaron Solomon
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | | | | | - Danielle Soucier-Ernst
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Medicine
| | - Noah G. Goodman
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Medicine
| | - S. William Stavropoulos
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Radiology, and
| | - Kara N. Maxwell
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Medicine
| | - Candace Clark
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Medicine
| | - George K. Belka
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
| | - Michael Feldman
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Pathology and Laboratory Medicine
| | - Angela DeMichele
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Department of Medicine
- Department of Biostatistics, Epidemiology and Informatics, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Lewis A. Chodosh
- Secondary Prevention through Surveillance and Intervention (2-PREVENT) Translational Center of Excellence
- Abramson Family Cancer Research Institute
- Department of Cancer Biology
- Department of Medicine
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18
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Miller AE, Hu P, Barker TH. Feeling Things Out: Bidirectional Signaling of the Cell-ECM Interface, Implications in the Mechanobiology of Cell Spreading, Migration, Proliferation, and Differentiation. Adv Healthc Mater 2020; 9:e1901445. [PMID: 32037719 PMCID: PMC7274903 DOI: 10.1002/adhm.201901445] [Citation(s) in RCA: 57] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2019] [Revised: 01/10/2020] [Indexed: 12/16/2022]
Abstract
Biophysical cues stemming from the extracellular environment are rapidly transduced into discernible chemical messages (mechanotransduction) that direct cellular activities-placing the extracellular matrix (ECM) as a potent regulator of cell behavior. Dynamic reciprocity between the cell and its associated matrix is essential to the maintenance of tissue homeostasis and dysregulation of both ECM mechanical signaling, via pathological ECM turnover, and internal mechanotransduction pathways contribute to disease progression. This review covers the current understandings of the key modes of signaling used by both the cell and ECM to coregulate one another. By taking an outside-in approach, the inherent complexities and regulatory processes at each level of signaling (ECM, plasma membrane, focal adhesion, and cytoplasm) are captured to give a comprehensive picture of the internal and external mechanoregulatory environment. Specific emphasis is placed on the focal adhesion complex which acts as a central hub of mechanical signaling, regulating cell spreading, migration, proliferation, and differentiation. In addition, a wealth of available knowledge on mechanotransduction is curated to generate an integrated signaling network encompassing the central components of the focal adhesion, cytoplasm and nucleus that act in concert to promote durotaxis, proliferation, and differentiation in a stiffness-dependent manner.
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Affiliation(s)
- Andrew E Miller
- Department of Biomedical Engineering, University of Virginia, 415 Lane Rd. MR5 1225, Charlottesville, VA, 22903, USA
| | - Ping Hu
- Department of Biomedical Engineering, University of Virginia, 415 Lane Rd. MR5 1225, Charlottesville, VA, 22903, USA
| | - Thomas H Barker
- Department of Biomedical Engineering, University of Virginia, 415 Lane Rd. MR5 1225, Charlottesville, VA, 22903, USA
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19
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Hung IC, Chen TM, Lin JP, Tai YL, Shen TL, Lee SJ. Wnt5b integrates Fak1a to mediate gastrulation cell movements via Rac1 and Cdc42. Open Biol 2020; 10:190273. [PMID: 32097584 PMCID: PMC7058935 DOI: 10.1098/rsob.190273] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Focal adhesion kinase (FAK) mediates vital cellular pathways during development. Despite its necessity, how FAK regulates and integrates with other signals during early embryogenesis remains poorly understood. We found that the loss of Fak1a impaired epiboly, convergent extension and hypoblast cell migration in zebrafish embryos. We also observed a clear disturbance in cortical actin at the blastoderm margin and distribution of yolk syncytial nuclei. In addition, we investigated a possible link between Fak1a and a well-known gastrulation regulator, Wnt5b, and revealed that the overexpression of fak1a or wnt5b could cross-rescue convergence defects induced by a wnt5b or fak1a antisense morpholino (MO), respectively. Wnt5b and Fak1a were shown to converge in regulating Rac1 and Cdc42, which could synergistically rescue wnt5b and fak1a morphant phenotypes. Furthermore, we generated several alleles of fak1a mutants using CRISPR/Cas9, but those mutants only revealed mild gastrulation defects. However, injection of a subthreshold level of the wnt5b MO induced severe gastrulation defects in fak1a mutants, which suggested that the upregulated expression of wnt5b might complement the loss of Fak1a. Collectively, we demonstrated that a functional interaction between Wnt and FAK signalling mediates gastrulation cell movements via the possible regulation of Rac1 and Cdc42 and subsequent actin dynamics.
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Affiliation(s)
- I-Chen Hung
- Department of Life Science, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan
| | - Tsung-Ming Chen
- Department of Life Science, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan.,Department of Plant Pathology and Microbiology, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan.,Department and Graduate Institute of Aquaculture, National Kaohsiung Marine University, Kaohsiung, Taiwan
| | - Jing-Ping Lin
- Department of Plant Pathology and Microbiology, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan
| | - Yu-Ling Tai
- Department of Plant Pathology and Microbiology, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan
| | - Tang-Long Shen
- Department of Plant Pathology and Microbiology, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan.,Center for Biotechnology, National Taiwan University, Taipei, Taiwan
| | - Shyh-Jye Lee
- Department of Life Science, National Taiwan University, No. 1, Roosevelt Road, Section 4, Taipei 10617, Taiwan.,Research Center for Developmental Biology and Regenerative Medicine, National Taiwan University, Taipei, Taiwan.,Center for Biotechnology, National Taiwan University, Taipei, Taiwan.,Center for Systems Biology, National Taiwan University, Taipei, Taiwan
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20
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Aboubakar Nana F, Vanderputten M, Ocak S. Role of Focal Adhesion Kinase in Small-Cell Lung Cancer and Its Potential as a Therapeutic Target. Cancers (Basel) 2019; 11:E1683. [PMID: 31671774 PMCID: PMC6895835 DOI: 10.3390/cancers11111683] [Citation(s) in RCA: 31] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2019] [Revised: 10/22/2019] [Accepted: 10/24/2019] [Indexed: 02/07/2023] Open
Abstract
Small-cell lung cancer (SCLC) represents 15% of all lung cancers and it is clinically the most aggressive type, being characterized by a tendency for early metastasis, with two-thirds of the patients diagnosed with an extensive stage (ES) disease and a five-year overall survival (OS) as low as 5%. There are still no effective targeted therapies in SCLC despite improved understanding of the molecular steps leading to SCLC development and progression these last years. After four decades, the only modest improvement in OS of patients suffering from ES-SCLC has recently been shown in a trial combining atezolizumab, an anti-PD-L1 immune checkpoint inhibitor, with carboplatin and etoposide, chemotherapy agents. This highlights the need to pursue research efforts in this field. Focal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase that is overexpressed and activated in several cancers, including SCLC, and contributing to cancer progression and metastasis through its important role in cell proliferation, survival, adhesion, spreading, migration, and invasion. FAK also plays a role in tumor immune evasion, epithelial-mesenchymal transition, DNA damage repair, radioresistance, and regulation of cancer stem cells. FAK is of particular interest in SCLC, being known for its aggressiveness. The inhibition of FAK in SCLC cell lines demonstrated significative decrease in cell proliferation, invasion, and migration, and induced cell cycle arrest and apoptosis. In this review, we will focus on the role of FAK in cancer cells and their microenvironment, and its potential as a therapeutic target in SCLC.
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Affiliation(s)
- Frank Aboubakar Nana
- Institut de Recherche Expérimentale et Clinique (IREC), Pôle de Pneumologie, ORL et Dermatologie (PNEU), Université catholique de Louvain (UCLouvain), 1200 Brussels, Belgium.
- Division of Pneumology, Cliniques Universitaires St-Luc, UCL, 1200 Brussels, Belgium.
| | - Marie Vanderputten
- Institut de Recherche Expérimentale et Clinique (IREC), Pôle de Pneumologie, ORL et Dermatologie (PNEU), Université catholique de Louvain (UCLouvain), 1200 Brussels, Belgium.
| | - Sebahat Ocak
- Institut de Recherche Expérimentale et Clinique (IREC), Pôle de Pneumologie, ORL et Dermatologie (PNEU), Université catholique de Louvain (UCLouvain), 1200 Brussels, Belgium.
- Division of Pneumology, CHU UCL Namur (Godinne Site), UCL, 5530 Yvoir, Belgium.
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21
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Stafman LL, Williams AP, Marayati R, Aye JM, Markert HR, Garner EF, Quinn CH, Lallani SB, Stewart JE, Yoon KJ, Whelan K, Beierle EA. Focal Adhesion Kinase Inhibition Contributes to Tumor Cell Survival and Motility in Neuroblastoma Patient-Derived Xenografts. Sci Rep 2019; 9:13259. [PMID: 31519958 PMCID: PMC6744403 DOI: 10.1038/s41598-019-49853-z] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2019] [Accepted: 08/24/2019] [Indexed: 12/29/2022] Open
Abstract
Patient-derived xenografts (PDXs) provide an opportunity to evaluate the effects of therapies in an environment that more closely resembles the human condition than that seen with long-term passage cell lines. In the current studies, we investigated the effects of FAK inhibition on two neuroblastoma PDXs in vitro. Cells were treated with two small molecule inhibitors of FAK, PF-573,228 (PF) and 1,2,4,5-benzentetraamine tetrahydrochloride (Y15). Following FAK inhibition, cell survival and proliferation decreased significantly and cell cycle arrest was seen in both cell lines. Migration and invasion assays were used to determine the effect of FAK inhibition on cell motility, which decreased significantly in both cell lines in the presence of either inhibitor. Finally, tumor cell stemness following FAK inhibition was evaluated with extreme limiting dilution assays as well as with immunoblotting and quantitative real-time PCR for the expression of stem cell markers. FAK inhibition decreased formation of tumorspheres and resulted in a corresponding decrease in established stem cell markers. FAK inhibition decreased many characteristics of the malignant phenotype, including cancer stem cell like features in neuroblastoma PDXs, making FAK a candidate for further investigation as a potential target for neuroblastoma therapy.
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Affiliation(s)
- Laura L Stafman
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Adele P Williams
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Raoud Marayati
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Jamie M Aye
- Division of Pediatric Hematology Oncology, Department of Pediatrics, University of Alabama, Birmingham, AL, 35233, USA
| | - Hooper R Markert
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Evan F Garner
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Colin H Quinn
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Shoeb B Lallani
- Department of Cell Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, AL, 35233, USA
| | - Jerry E Stewart
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA
| | - Karina J Yoon
- Department of Pharmacology and Toxicology, University of Alabama, Birmingham, AL, 35233, USA
| | - Kimberly Whelan
- Division of Pediatric Hematology Oncology, Department of Pediatrics, University of Alabama, Birmingham, AL, 35233, USA
| | - Elizabeth A Beierle
- Division of Pediatric Surgery, Department of Surgery, University of Alabama, Birmingham, AL, 35205, USA.
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22
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Zhang C, Stockwell SR, Elbanna M, Ketteler R, Freeman J, Al-Lazikani B, Eccles S, De Haven Brandon A, Raynaud F, Hayes A, Clarke PA, Workman P, Mittnacht S. Signalling involving MET and FAK supports cell division independent of the activity of the cell cycle-regulating CDK4/6 kinases. Oncogene 2019; 38:5905-5920. [PMID: 31296956 PMCID: PMC6756076 DOI: 10.1038/s41388-019-0850-2] [Citation(s) in RCA: 25] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2019] [Revised: 05/07/2019] [Accepted: 05/13/2019] [Indexed: 12/23/2022]
Abstract
Deregulation of cyclin-dependent kinases 4 and 6 (CDK4/6) is highly prevalent in cancer; yet, inhibitors against these kinases are currently used only in restricted tumour contexts. The extent to which cancers depend on CDK4/6 and the mechanisms that may undermine such dependency are poorly understood. Here, we report that signalling engaging the MET proto-oncogene receptor tyrosine kinase/focal adhesion kinase (FAK) axis leads to CDK4/6-independent CDK2 activation, involving as critical mechanistic events loss of the CDKI p21CIP1 and gain of its regulator, the ubiquitin ligase subunit SKP2. Combined inhibition of MET/FAK and CDK4/6 eliminates the proliferation capacity of cancer cells in culture, and enhances tumour growth inhibition in vivo. Activation of the MET/FAK axis is known to arise through cancer extrinsic and intrinsic cues. Our work predicts that such cues support cell division independent of the activity of the cell cycle-regulating CDK4/6 kinases and identifies MET/FAK as a tractable route to broaden the utility of CDK4/6 inhibitor-based therapies in the clinic.
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Affiliation(s)
- Chi Zhang
- UCL Cancer Institute, University College London, London, WC1E 6DD, UK
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Simon R Stockwell
- UCL Cancer Institute, University College London, London, WC1E 6DD, UK
| | - May Elbanna
- UCL Cancer Institute, University College London, London, WC1E 6DD, UK
| | - Robin Ketteler
- MRC Laboratory for Molecular Cell Biology, University College London, London, WC1E 6BT, UK
| | - Jamie Freeman
- MRC Laboratory for Molecular Cell Biology, University College London, London, WC1E 6BT, UK
| | - Bissan Al-Lazikani
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Suzanne Eccles
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Alexis De Haven Brandon
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Florence Raynaud
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Angela Hayes
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Paul A Clarke
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK
| | - Paul Workman
- Cancer Research UK Cancer Therapeutics Unit at The Institute of Cancer Research, London, SM2 5NG, UK.
| | - Sibylle Mittnacht
- UCL Cancer Institute, University College London, London, WC1E 6DD, UK.
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23
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Yeo MS, Subhash VV, Suda K, Balcıoğlu HE, Zhou S, Thuya WL, Loh XY, Jammula S, Peethala PC, Tan SH, Xie C, Wong FY, Ladoux B, Ito Y, Yang H, Goh BC, Wang L, Yong WP. FBXW5 Promotes Tumorigenesis and Metastasis in Gastric Cancer via Activation of the FAK-Src Signaling Pathway. Cancers (Basel) 2019; 11:cancers11060836. [PMID: 31213005 PMCID: PMC6627937 DOI: 10.3390/cancers11060836] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2019] [Revised: 06/10/2019] [Accepted: 06/14/2019] [Indexed: 12/26/2022] Open
Abstract
F-box/WD repeat-containing protein 5 (FBXW5) is a member of the FBXW subclass of F-box proteins. Despite its known function as a component of the Skp1-Cullin-F-box (SCF) ubiquitin ligase complex, the role of FBXW5 in gastric cancer tumorigenesis and metastasis has not been investigated. The present study investigates the role of FBXW5 in tumorigenesis and metastasis, as well as the regulation of key signaling pathways in gastric cancer; using in-vitro FBXW5 knockdown/overexpression cell line and in-vivo models. In-vitro knockdown of FBXW5 results in a decrease in cell proliferation and cell cycle progression, with a concomitant increase in cell apoptosis and caspase-3 activity. Furthermore, knockdown of FBXW5 also leads to a down regulation in cell migration and adhesion, characterized by a reduction in actin polymerization, focal adhesion turnover and traction forces. This study also delineates the mechanistic role of FBXW5 in oncogenic signaling as its inhibition down regulates RhoA-ROCK 1 (Rho-associated protein kinase 1) and focal adhesion kinase (FAK) signaling cascades. Overexpression of FBXW5 promotes in-vivo tumor growth, whereas its inhibition down regulates in-vivo tumor metastasis. When considered together, our study identifies the novel oncogenic role of FBXW5 in gastric cancer and draws further interest regarding its clinical utility as a potential therapeutic target.
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Affiliation(s)
- Mei Shi Yeo
- Department of Haematology-Oncology, National University Hospital of Singapore, Singapore 119228, Singapore.
| | - Vinod Vijay Subhash
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
- Lowy Cancer Research Centre, University of New South Wales, Sydney 20152, Australia.
| | - Kazuto Suda
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Hayri Emrah Balcıoğlu
- Mechanobiology Institute, National University of Singapore, Singapore 117411, Singapore.
| | - Siqin Zhou
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Win Lwin Thuya
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Xin Yi Loh
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Sriganesh Jammula
- Cancer Research UK Cambridge Institute, University of Cambridge, CB2 0RE Cambridge, UK.
| | - Praveen C Peethala
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Shi Hui Tan
- Department of Haematology-Oncology, National University Hospital of Singapore, Singapore 119228, Singapore.
| | - Chen Xie
- Department of Haematology-Oncology, National University Hospital of Singapore, Singapore 119228, Singapore.
| | - Foong Ying Wong
- Department of Haematology-Oncology, National University Hospital of Singapore, Singapore 119228, Singapore.
| | - Benoit Ladoux
- Mechanobiology Institute, National University of Singapore, Singapore 117411, Singapore.
- Institut Jacques Monod, Centre National de la Recherche Scientifique, CNRS UMR 7592, Université Paris-Diderot, CEDEX 13, 75205 Paris, France.
| | - Yoshiaki Ito
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Henry Yang
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Boon Cher Goh
- Department of Haematology-Oncology, National University Hospital of Singapore, Singapore 119228, Singapore.
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Lingzhi Wang
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
| | - Wei Peng Yong
- Department of Haematology-Oncology, National University Hospital of Singapore, Singapore 119228, Singapore.
- Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.
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Al-Ghabkari A, Qasrawi DO, Alshehri M, Narendran A. Focal adhesion kinase (FAK) phosphorylation is a key regulator of embryonal rhabdomyosarcoma (ERMS) cell viability and migration. J Cancer Res Clin Oncol 2019; 145:1461-1469. [PMID: 31006845 DOI: 10.1007/s00432-019-02913-3] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2019] [Accepted: 04/02/2019] [Indexed: 02/07/2023]
Abstract
BACKGROUND Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in children. Pathogenesis of RMS is associated with aggressive growth pattern and increased risk of morbidity and mortality. There are two main subtypes or RMS: embryonal and alveolar. The embryonal type is characterized by distinct molecular aberrations, including alterations in the activity of certain protein kinases. Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that plays a vital role in focal adhesion (FA) assembly to promote cytoskeleton dynamics and regulation of cell motility. It is regulated by multiple phosphorylation sites: tyrosine 397, Tyr 576/577, and Tyr 925. Tyrosine 397 is the autophosphorylation site that regulates FAK localization at the cell periphery to facilitate the assembly and formation of the FA complex. The kinase activity of FAK is mediated by the phosphorylation of Tyr 576/577 within the kinase domain activation loop. Aberrations of FAK phosphorylation have been linked to the pathogenesis of different types of cancers. In this regard, pY397 upregulation is linked to increase ERMS cell motility, invasion, and tumorigenesis. METHODS In this study, we have used an established human embryonal muscle rhabdomyosarcoma cell line RD as a model to examine FAK phosphorylation profiles to characterize its role in the pathogenies of RMS. RESULTS Our findings revealed a significant increase of FAK phosphorylation at pY397 in RD cells compared to control cells (hTERT). On the other hand, Tyr 576/577 phosphorylation levels in RD cells displayed a pronounced reduction. Our data showed that Y925 residue exhibited no detectable change. The in vitro analysis showed that the FAK inhibitor, PF-562271 led to G1 cell-cycle arrest induced cell death (IC50, ~ 12 µM) compared to controls. Importantly, immunostaining analyses displayed a noticeable reduction of Y397 phosphorylation following PF-562271 treatment. Our data also showed that PF-562271 suppressed RD cell migration in a dose-dependent manner associated with a reduction in Y397 phosphorylation. CONCLUSIONS The data presented herein indicate that targeting FAK phosphorylation at distinct sites is a promising strategy in future treatment approaches for defined subgroups of rhabdomyosarcoma.
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Affiliation(s)
- Abdulhameed Al-Ghabkari
- Department of Biochemistry and Molecular Biology, Arnie Charbonneau Cancer Institute, Cumming School of Medicine, University of Calgary, 3280 Hospital Drive NW, Calgary, AB, T2N 4Z6, Canada.
| | - Deema O Qasrawi
- Department of Pathology and Laboratory Medicine, Cumming School of Medicine, University of Calgary, 3280 Hospital Drive NW, Calgary, AB, T2N 4N1, Canada
| | - Mana Alshehri
- Department of Biochemistry and Molecular Biology, Arnie Charbonneau Cancer Institute, Cumming School of Medicine, University of Calgary, 3280 Hospital Drive NW, Calgary, AB, T2N 4Z6, Canada
- King Abdullah International Medical Research Center (KAIMRC), Riyadh, Saudi Arabia
| | - Aru Narendran
- Department of Biochemistry and Molecular Biology, Arnie Charbonneau Cancer Institute, Cumming School of Medicine, University of Calgary, 3280 Hospital Drive NW, Calgary, AB, T2N 4Z6, Canada
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25
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Sun A, Hao J, Yu L, Lahiri SK, Yang W, Lin Q, Zhao J. Regulation of Krüppel-like factor 8 by the NEDD4 E3 ubiquitin ligase. Am J Transl Res 2019; 11:1521-1530. [PMID: 30972179 PMCID: PMC6456509] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2018] [Accepted: 12/18/2018] [Indexed: 06/09/2023]
Abstract
Krüppel-like factor 8 (KLF8) plays many important roles in various diseases, especially cancer. Previous studies have shown that KLF8 is regulated by ubiquitylation. The molecular mechanism underlying this posttranslational modification of KLF8, however, has not been investigated. Reported here is our identification of the neural precursor cell expressed, developmentally down-regulated 4 (NEDD4) as the E3 ubiquitin ligase for this modification. By co-immunoprecipitation and ubiquitylation assays, we determined that KLF8 interacts with NEDD4 and is ubiquitylated by NEDD4. By site-directed mutagenesis and pharmacological inhibition of MEK, we found that the ubiquitylation of KLF8 by NEDD4 depends upon the phosphorylation of KLF8 at serine 48 by ERK. Cycloheximide chase analysis, target gene promoter reporter assay and fluorescent staining indicated that NEDD4 plays a critical role in promoting the stability and transcriptional activity of KLF8 in the nucleus. Taken together, this work identified NEDD4 as a novel E3 ubiquitin ligase for KLF8 that provides insights into targeting the KLF8-NEDD4 axis to treat various types of cancer associated with overexpression of both proteins.
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Affiliation(s)
- Aiqin Sun
- Burnett School of Biomedical Sciences University of Central Florida College of MedicineOrlando, FL 32827, USA
- School of Medicine Jiangsu UniversityZhenjiang, China
| | - Jie Hao
- Burnett School of Biomedical Sciences University of Central Florida College of MedicineOrlando, FL 32827, USA
| | - Lin Yu
- Burnett School of Biomedical Sciences University of Central Florida College of MedicineOrlando, FL 32827, USA
| | - Satadru K Lahiri
- Burnett School of Biomedical Sciences University of Central Florida College of MedicineOrlando, FL 32827, USA
- Cardiovascular Research Institute and Department of Molecular Physiology and Biophysics, Baylor College of MedicineHouston, TX 77030, USA
| | - Wannian Yang
- School of Medicine Jiangsu UniversityZhenjiang, China
| | - Qiong Lin
- School of Medicine Jiangsu UniversityZhenjiang, China
| | - Jihe Zhao
- Burnett School of Biomedical Sciences University of Central Florida College of MedicineOrlando, FL 32827, USA
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26
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Chuang HH, Wang PH, Niu SW, Zhen YY, Huang MS, Hsiao M, Yang CJ. Inhibition of FAK Signaling Elicits Lamin A/C-Associated Nuclear Deformity and Cellular Senescence. Front Oncol 2019; 9:22. [PMID: 30761269 PMCID: PMC6363943 DOI: 10.3389/fonc.2019.00022] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2018] [Accepted: 01/08/2019] [Indexed: 01/07/2023] Open
Abstract
Focal adhesion kinase (FAK) is a non-receptor kinase that facilitates tumor aggressiveness. The effects of FAK inhibition include arresting proliferation, limiting metastasis, and inhibiting angiogenesis. PF-573228 is an ATP-competitive inhibitor of FAK. Treating lung cancer cells with PF-573228 resulted in FAK inactivation and changes in the expressions of lamin A/C and nuclear deformity. Since lamin A/C downregulation or deficiency was associated with cellular senescence, the senescence-associated β-galactosidase (SA-β-gal) assay was used to investigate whether PF-573228 treatment drove cellular senescence, which showed more SA-β-gal-positive cells in culture. p53 is known to play a pivotal role in mediating the progression of cellular senescence, and the PF-573228-treated lung cancer cells resulted in a higher p53 expression level. Subsequently, the FAK depletion in lung cancer cells was employed to confirm the role of FAK inhibition on cellular senescence. FAK depletion and pharmacological inhibition of lung cancer cells elicited similar patterns of cellular senescence, lamin A/C downregulation, and p53 upregulation, implying that FAK signaling is associated with the expression of p53 and the maintenance of lamin A/C levels to shape regular nuclear morphology and manage anti-senescence. Conversely, FAK inactivation led to p53 upregulation, disorganization of the nuclear matrix, and consequently cellular senescence. Our data suggest a new FAK signaling pathway, in that abolishing FAK signaling can activate the senescence program in cells. Triggering cellular senescence could be a new therapeutic approach to limit tumor growth.
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Affiliation(s)
- Hsiang-Hao Chuang
- Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Pei-Hui Wang
- Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Sheng-Wen Niu
- Division of Nephrology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Yen-Yi Zhen
- Division of Nephrology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Ming-Shyan Huang
- Department of Internal Medicine, E-Da Cancer Hospital, School of Medicine, I-Shou University, Kaohsiung, Taiwan
| | - Michael Hsiao
- Genomics Research Center, Academia Sinica, Taipei, Taiwan.,Department of Biochemistry, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Chih-Jen Yang
- Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.,Department of Internal Medicine, Kaohsiung Municipal Ta-Tung Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.,Department of Respiratory Therapy, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
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27
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Baghdadi MB, Firmino J, Soni K, Evano B, Di Girolamo D, Mourikis P, Castel D, Tajbakhsh S. Notch-Induced miR-708 Antagonizes Satellite Cell Migration and Maintains Quiescence. Cell Stem Cell 2018; 23:859-868.e5. [PMID: 30416072 DOI: 10.1016/j.stem.2018.09.017] [Citation(s) in RCA: 75] [Impact Index Per Article: 10.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/25/2017] [Revised: 07/18/2018] [Accepted: 09/27/2018] [Indexed: 12/31/2022]
Abstract
Critical features of stem cells include anchoring within a niche and activation upon injury. Notch signaling maintains skeletal muscle satellite (stem) cell quiescence by inhibiting differentiation and inducing expression of extracellular components of the niche. However, the complete spectrum of how Notch safeguards quiescence is not well understood. Here, we perform Notch ChIP-sequencing and small RNA sequencing in satellite cells and identify the Notch-induced microRNA-708, which is a mirtron that is highly expressed in quiescent cells and sharply downregulated in activated cells. We employ in vivo and ex vivo functional studies, in addition to live imaging, to show that miR-708 regulates quiescence and self-renewal by antagonizing cell migration through targeting the transcripts of the focal-adhesion-associated protein Tensin3. Therefore, this study identifies a Notch-miR708-Tensin3 axis and suggests that Notch signaling can regulate satellite cell quiescence and transition to the activation state through dynamic regulation of the migratory machinery.
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Affiliation(s)
- Meryem B Baghdadi
- Stem Cells and Development, Department of Developmental & Stem Cell Biology, Institut Pasteur, Paris 75015, France; CNRS UMR 3738, Institut Pasteur, Paris 75015, France; Sorbonne Universités, UPMC, University of Paris 06, IFD-ED 515, Paris 75252, France
| | - Joao Firmino
- Bioimaging and Optics platform (BIOP), School of Life Sciences, Swiss Federal Institute of Technology (EPFL), Lausanne, Switzerland
| | - Kartik Soni
- Stem Cells and Development, Department of Developmental & Stem Cell Biology, Institut Pasteur, Paris 75015, France; CNRS UMR 3738, Institut Pasteur, Paris 75015, France
| | - Brendan Evano
- Stem Cells and Development, Department of Developmental & Stem Cell Biology, Institut Pasteur, Paris 75015, France; CNRS UMR 3738, Institut Pasteur, Paris 75015, France
| | - Daniela Di Girolamo
- Stem Cells and Development, Department of Developmental & Stem Cell Biology, Institut Pasteur, Paris 75015, France; Dipartimento di Medicina Clinica e Chirurgica, Università degli Studi di Napoli Frederico II, 80131 Naples, Italy
| | | | - David Castel
- UMR8203 "Vectorologie et Thérapeutiques Anticancéreuses," CNRS, Gustave Roussy, Université Paris-Sud, Université Paris-Saclay, 94805 Villejuif, France; Département de Cancérologie de l'Enfant et de l'Adolescent, Gustave Roussy, Université Paris-Sud, Université Paris-Saclay, 94805 Villejuif, France
| | - Shahragim Tajbakhsh
- Stem Cells and Development, Department of Developmental & Stem Cell Biology, Institut Pasteur, Paris 75015, France; CNRS UMR 3738, Institut Pasteur, Paris 75015, France.
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28
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Cerium Oxide Nanoparticles Sensitize Pancreatic Cancer to Radiation Therapy through Oxidative Activation of the JNK Apoptotic Pathway. Cancers (Basel) 2018. [DOI: 10.10.3390/cancers10090303] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
Side effects of radiation therapy (RT) remain the most challenging issue for pancreatic cancer treatment. Cerium oxide nanoparticles (CONPs) are currently being tested in pre-clinical trials as an adjuvant to sensitize pancreatic cancer cells to RT and protect normal tissues from the harmful side effects. CONPs were not able to significantly affect RT-induced DNA damage in cancer cells, thereby ruling out sensitization through increased mitotic catastrophe. However, activation of c-Jun terminal kinase (JNK), a key driver of RT-induced apoptosis, was significantly enhanced by co-treatment with CONPs and RT in pancreatic cancer cells in vitro and human pancreatic tumors in nude mice in vivo compared to CONPs or RT treatment alone. Further, CONP-driven increase in RT-induced JNK activity was associated with a marked increase in Caspase 3/7 activation, indicative of apoptosis. We have previously shown that CONPs increase reactive oxygen species (ROS) production in cancer cells. ROS has been shown to drive the oxidation of thioredoxin 1 (TRX1) which results in the activation of apoptosis signaling kinase 1 (ASK1). The increase in ASK1 activation following the co-treatment with CONPs followed by RT suggests that the increased JNK activation is the result of increased TRX1 oxidation. The ability of CONPs to sensitize pancreatic cancer cells to RT was mitigated when the TRX1 oxidation was prevented by mutagenesis of a cysteine residue or when the JNK activation was blocked by an inhibitor. Taken together, these data demonstrate an important mechanism for CONPs in specifically killing cancer cells and provide novel insights into the utilization of CONPs as a radiosensitizer and therapeutic agent for pancreatic cancer.
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29
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Wason MS, Lu H, Yu L, Lahiri SK, Mukherjee D, Shen C, Das S, Seal S, Zhao J. Cerium Oxide Nanoparticles Sensitize Pancreatic Cancer to Radiation Therapy through Oxidative Activation of the JNK Apoptotic Pathway. Cancers (Basel) 2018; 10:cancers10090303. [PMID: 30200491 PMCID: PMC6162528 DOI: 10.3390/cancers10090303] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/10/2018] [Revised: 08/30/2018] [Accepted: 08/30/2018] [Indexed: 12/11/2022] Open
Abstract
Side effects of radiation therapy (RT) remain the most challenging issue for pancreatic cancer treatment. Cerium oxide nanoparticles (CONPs) are currently being tested in pre-clinical trials as an adjuvant to sensitize pancreatic cancer cells to RT and protect normal tissues from the harmful side effects. CONPs were not able to significantly affect RT-induced DNA damage in cancer cells, thereby ruling out sensitization through increased mitotic catastrophe. However, activation of c-Jun terminal kinase (JNK), a key driver of RT-induced apoptosis, was significantly enhanced by co-treatment with CONPs and RT in pancreatic cancer cells in vitro and human pancreatic tumors in nude mice in vivo compared to CONPs or RT treatment alone. Further, CONP-driven increase in RT-induced JNK activity was associated with a marked increase in Caspase 3/7 activation, indicative of apoptosis. We have previously shown that CONPs increase reactive oxygen species (ROS) production in cancer cells. ROS has been shown to drive the oxidation of thioredoxin 1 (TRX1) which results in the activation of apoptosis signaling kinase 1 (ASK1). The increase in ASK1 activation following the co-treatment with CONPs followed by RT suggests that the increased JNK activation is the result of increased TRX1 oxidation. The ability of CONPs to sensitize pancreatic cancer cells to RT was mitigated when the TRX1 oxidation was prevented by mutagenesis of a cysteine residue or when the JNK activation was blocked by an inhibitor. Taken together, these data demonstrate an important mechanism for CONPs in specifically killing cancer cells and provide novel insights into the utilization of CONPs as a radiosensitizer and therapeutic agent for pancreatic cancer.
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Affiliation(s)
- Melissa S Wason
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
| | - Heng Lu
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
- Department of Surgery, Miller School of Medicine, University of Miami, Miami, FL 33136, USA.
| | - Lin Yu
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
| | - Satadru K Lahiri
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
- Cardiovascular Research Institute, Department of Molecular Physiology & Biophysics, Baylor College of Medicine, Houston, TX 77030, USA.
| | - Debarati Mukherjee
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
- Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, NC 27710, USA.
| | - Chao Shen
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
- Department of Microbiology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China.
| | - Soumen Das
- Department of Mechanical, Materials and Aerospace Engineering, Advanced Materials Processing and Analysis Center; Nanoscience and Nanotechnology Center, University of Central Florida, Orlando, FL 32816, USA.
| | - Sudipta Seal
- Department of Mechanical, Materials and Aerospace Engineering, Advanced Materials Processing and Analysis Center; Nanoscience and Nanotechnology Center, University of Central Florida, Orlando, FL 32816, USA.
| | - Jihe Zhao
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL 32827, USA.
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Giannini M, Primerano C, Berger L, Giannaccini M, Wang Z, Landi E, Cuschieri A, Dente L, Signore G, Raffa V. Nano-topography: Quicksand for cell cycle progression? NANOMEDICINE-NANOTECHNOLOGY BIOLOGY AND MEDICINE 2018; 14:2656-2665. [PMID: 30010000 DOI: 10.1016/j.nano.2018.07.002] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/15/2018] [Revised: 06/26/2018] [Accepted: 07/01/2018] [Indexed: 01/01/2023]
Abstract
The 3-D spatial and mechanical features of nano-topography can create alternative environments, which influence cellular response. In this paper, murine fibroblast cells were grown on surfaces characterized by protruding nanotubes. Cells cultured on such nano-structured surface exhibit stronger cellular adhesion compared to control groups, but despite the fact that stronger adhesion is generally believed to promote cell cycle progression, the time cells spend in G1 phase is doubled. This apparent contradiction is solved by confocal microscopy analysis, which shows that the nano-topography inhibits actin stress fiber formation. In turn, this impairs RhoA activation, which is required to suppress the inhibition of cell cycle progression imposed by p21/p27. This finding suggests that the generation of stress fibers, required to impose the homeostatic intracellular tension, rather than cell adhesion/spreading is the limiting factor for cell cycle progression. Indeed, nano-topography could represent a unique tool to inhibit proliferation in adherent well-spread cells.
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Affiliation(s)
| | | | - Liron Berger
- Department of Biology, Università di Pisa, Pisa, Italy.
| | | | - Zhigang Wang
- Institute for Medical Science and Technology, University of Dundee, Dundee, United Kingdom.
| | - Elena Landi
- Department of Biology, Università di Pisa, Pisa, Italy.
| | - Alfred Cuschieri
- Institute for Medical Science and Technology, University of Dundee, Dundee, United Kingdom.
| | - Luciana Dente
- Department of Biology, Università di Pisa, Pisa, Italy.
| | - Giovanni Signore
- Center for Nanotechnology Innovation @NEST, Istituto Italiano di Tecnologia, Pisa, Italy; NEST, Scuola Normale Superiore, and Istituto Nanoscienze-CNR, Pisa, Italy.
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Liu H, Liu J, Zhang T, Li L, Wang J, Han C, He H. The Gene Ontology Differs in Bursa of Fabricius Between Two Breeds of Ducks Post Hatching by Enriching the Differentially Expressed Genes. BRAZILIAN JOURNAL OF POULTRY SCIENCE 2018. [DOI: 10.1590/1806-9061-2016-0416] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Affiliation(s)
- H Liu
- Sichuan Agricultural University, China
| | - J Liu
- Sichuan Agricultural University, China
| | - T Zhang
- Sichuan Agricultural University, China
| | - L Li
- Sichuan Agricultural University, China
| | - J Wang
- Sichuan Agricultural University, China
| | - C Han
- Sichuan Agricultural University, China
| | - H He
- Sichuan Agricultural University, China
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Mukherjee D, Lu H, Yu L, He C, Lahiri SK, Li T, Zhao J. Krüppel-like factor 8 activates the transcription of C-X-C cytokine receptor type 4 to promote breast cancer cell invasion, transendothelial migration and metastasis. Oncotarget 2018; 7:23552-68. [PMID: 26993780 PMCID: PMC5029647 DOI: 10.18632/oncotarget.8083] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2015] [Accepted: 02/25/2016] [Indexed: 02/04/2023] Open
Abstract
Krüppel-like factor 8 (KLF8) has been strongly implicated in breast cancer metastasis. However, the underlying mechanisms remain largely unknown. Here we report a novel signaling from KLF8 to C-X-C cytokine receptor type 4 (CXCR4) in breast cancer. Overexpression of KLF8 in MCF-10A cells induced CXCR4 expression at both mRNA and protein levels, as determined by quantitative real-time PCR and immunoblotting. This induction was well correlated with increased Boyden chamber migration, matrigel invasion and transendothelial migration (TEM) of the cells towards the ligand CXCL12. On the other hand, knockdown of KLF8 in MDA-MB-231 cells reduced CXCR4 expression associated with decreased cell migration, invasion and TEM towards CXCL12. Histological and database mining analyses of independent cohorts of patient tissue microarrays revealed a correlation of aberrant co-elevation of KLF8 and CXCR4 with metastatic potential. Promoter analysis indicated that KLF8 directly binds and activates the human CXCR4 gene promoter. Interestingly, a CXCR4-dependent activation of focal adhesion kinase (FAK), a known upregulator of KLF8, was highly induced by CXCL12 treatment in KLF8-overexpressing, but not KLF8 deficient cells. This activation of FAK in turn induced a further increase in KLF8 expression. Xenograft studies showed that overexpression of CXCR4, but not a dominant-negative mutant of it, in the MDA-MB-231 cells prevented the invasive growth of primary tumor and lung metastasis from inhibition by knockdown of KLF8. These results collectively suggest a critical role for a previously unidentified feed-forward signaling wheel made of KLF8, CXCR4 and FAK in promoting breast cancer metastasis and shed new light on potentially more effective anti-cancer strategies.
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Affiliation(s)
- Debarati Mukherjee
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL, USA
| | - Heng Lu
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL, USA
| | - Lin Yu
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL, USA
| | - Chunjiang He
- School of Basic Medical Sciences, Wuhan University, Wuhan, China
| | - Satadru K Lahiri
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL, USA
| | - Tianshu Li
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL, USA.,Current address: Cleveland Clinic, Cleveland, OH, USA
| | - Jihe Zhao
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine, Orlando, FL, USA
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Kumar V, Soni UK, Maurya VK, Singh K, Jha RK. Integrin beta8 (ITGB8) activates VAV-RAC1 signaling via FAK in the acquisition of endometrial epithelial cell receptivity for blastocyst implantation. Sci Rep 2017; 7:1885. [PMID: 28507287 PMCID: PMC5432530 DOI: 10.1038/s41598-017-01764-7] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2016] [Accepted: 04/06/2017] [Indexed: 12/02/2022] Open
Abstract
Integrin beta8 (ITGB8) is involved in the endometrial receptivity. The blastocyst first interacts with the luminal endometrial epithelial cells during its implantation; therefore, we have investigated the signaling of ITGB8 via FAK and VAV-RAC1 in the endometrial epithelial cells. Integrin beta8 was found elevated in epithelial cells at late-pre-receptive (day4, 1600 h) and receptive (day5, 0500 h) stages of endometrial receptivity period in the mouse. Integrins downstream molecule FAK has demonstrated an increased expression and phosphorylation (Y397) in the endometrium as well as in the isolated endometrial epithelial cells during receptive and post-receptive stages. Integrin beta8 can functionally interact with FAK, VAV and RAC1 as the levels of phosphorylated-FAK, and VAV along with the RAC-GTP form was reduced after ITGB8 knockdown in the endometrial epithelial cells and uterus. Further, VAV and RAC1 were seen poorly active in the absence of FAK activity, suggesting a crosstalk of ITGB8 and FAK for VAV and RAC1 activation in the endometrial epithelial cells. Silencing of ITGB8 expression and inhibition of FAK activity in the Ishikawa cells rendered poor attachment of JAr spheroids. In conclusion, ITGB8 activates VAV-RAC1 signaling axis via FAK to facilitate the endometrial epithelial cell receptivity for the attachment of blastocyst.
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Affiliation(s)
- Vijay Kumar
- Division of Endocrinology, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, U.P., India
| | - Upendra Kumar Soni
- Division of Endocrinology, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, U.P., India
| | - Vineet Kumar Maurya
- Division of Endocrinology, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, U.P., India
| | - Kiran Singh
- Department of Molecular & Human Genetics, Banaras Hindu University (BHU), Varanasi, UP, India
| | - Rajesh Kumar Jha
- Division of Endocrinology, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, U.P., India.
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Lemler DJ, Lynch ML, Tesfay L, Deng Z, Paul BT, Wang X, Hegde P, Manz DH, Torti SV, Torti FM. DCYTB is a predictor of outcome in breast cancer that functions via iron-independent mechanisms. Breast Cancer Res 2017; 19:25. [PMID: 28270217 PMCID: PMC5341190 DOI: 10.1186/s13058-017-0814-9] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2016] [Accepted: 02/09/2017] [Indexed: 02/08/2023] Open
Abstract
BACKGROUND Duodenal cytochrome b (DCYTB) is a ferrireductase that functions together with divalent metal transporter 1 (DMT1) to mediate dietary iron reduction and uptake in the duodenum. DCYTB is also a member of a 16-gene iron regulatory gene signature (IRGS) that predicts metastasis-free survival in breast cancer patients. To better understand the relationship between DCYTB and breast cancer, we explored in detail the prognostic significance and molecular function of DCYTB in breast cancer. METHODS The prognostic significance of DCYTB expression was evaluated using publicly available microarray data. Signaling Pathway Impact Analysis (SPIA) of microarray data was used to identify potential novel functions of DCYTB. The role of DCYTB was assessed using immunohistochemistry and measurements of iron uptake, iron metabolism, and FAK signaling. RESULTS High DCYTB expression was associated with prolonged survival in two large independent cohorts, together totaling 1610 patients (cohort #1, p = 1.6e-11, n = 741; cohort #2, p = 1.2e-05, n = 869; log-rank test) as well as in the Gene expression-based Outcome for Breast cancer Online (GOBO) cohort (p < 1.0e-05, n = 1379). High DCYTB expression was also associated with increased survival in homogeneously treated groups of patients who received either tamoxifen or chemotherapy. Immunohistochemistry revealed that DCYTB is localized on the plasma membrane of breast epithelial cells, and that expression is dramatically reduced in high-grade tumors. Surprisingly, neither overexpression nor knockdown of DCYTB affected levels of ferritin H, transferrin receptor, labile iron or total cellular iron in breast cancer cells. Because SPIA pathway analysis of patient microarray data revealed an association between DCYTB and the focal adhesion pathway, we examined the influence of DCYTB on FAK activation in breast cancer cells. These experiments reveal that DCYTB reduces adhesion and activation of focal adhesion kinase (FAK) and its adapter protein paxillin. CONCLUSIONS DCYTB is an important predictor of outcome and is associated with response to therapy in breast cancer patients. DCYTB does not affect intracellular iron in breast cancer cells. Instead, DCYTB may retard cancer progression by reducing activation of FAK, a kinase that plays a central role in tumor cell adhesion and metastasis.
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Affiliation(s)
- David J. Lemler
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT 06030 USA
- Present address: Department of Molecular Biomedical Sciences, North Carolina State University, CVM Research Building 474, Raleigh, NC 27695 USA
| | - Miranda L. Lynch
- Center for Quantitative Medicine, University of Connecticut Health Center, Farmington, CT 06030 USA
- Present address: Statistical Sciences Group CCS-6, Los Alamos National Laboratory, Los Alamos, NM 87545 USA
| | - Lia Tesfay
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - Zhiyong Deng
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - Bibbin T. Paul
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - Xiaohong Wang
- Department of Pathology, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - Poornima Hegde
- Department of Pathology, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - David H. Manz
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT 06030 USA
- School of Dental Medicine, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - Suzy V. Torti
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT 06030 USA
| | - Frank M. Torti
- Department of Medicine, University of Connecticut Health Center, Farmington, CT 06030 USA
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Griffin M, Ibrahim A, Seifalian A, Butler P, Kalaskar D, Ferretti P. Chemical group-dependent plasma polymerisation preferentially directs adipose stem cell differentiation towards osteogenic or chondrogenic lineages. Acta Biomater 2017; 50:450-461. [PMID: 27956359 PMCID: PMC5331891 DOI: 10.1016/j.actbio.2016.12.016] [Citation(s) in RCA: 49] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2016] [Revised: 11/25/2016] [Accepted: 12/07/2016] [Indexed: 12/17/2022]
Abstract
Human adipose derived stem cells (ADSCs) are being explored for the repair of craniofacial defects due to their multi-differentiation potential and ease of isolation and expansion. Crucial to using ADSCs for craniofacial repair is the availability of materials with appropriate biomechanical properties that can support their differentiation into bone and cartilage. We tested the hypothesis that different modifications of chemical groups on the surface of a nanocomposite polymer could increase human ADSC adhesion and selectively enhance their osteogenic and chondrogenic differentiation. We show that the COOH modification significantly promoted initial cell adhesion and proliferation over 14 days compared to NH2 surfaces. Expression of focal adhesion kinase and vinculin was enhanced after plasma surface polymerisation at 24 h. The COOH modification significantly enhanced chondrogenic differentiation as indicated by up-regulation of aggrecan and collagen II transcripts. In contrast, NH2 group functionalised scaffolds promoted osteogenic differentiation with significantly enhanced expression of collagen I, alkaline phosphatase and osteocalcin both at the gene and protein level. Finally, chorioallantoic membrane grafting demonstrated that both NH2 and COOH functionalised scaffolds seeded with ADSCs were biocompatible and supported vessel ingrowth apparently to a greater degree than unmodified scaffolds. In summary, our study shows the ability to direct ADSC chondrogenic and osteogenic differentiation by deposition of different chemical groups through plasma surface polymerisation. Hence this approach could be used to selectively enhance bone or cartilage formation before implantation in vivo to repair skeletal defects. Statement of Significance Human adipose derived stem cells (hADSCs) are an exciting stem cell source for regenerative medicine due to their plentiful supply and ease of isolation. However, the optimal environmental cues to direct stem cells towards certain lineages change have to has not been identified. We have shown that by modifying the surface of the scaffold with specific chemical groups using plasma surface polymerisation techniques we can control ADSCs differentiation. This study shows that ADSCs can be differentiated towards osteogenic and chondrogenic lineages on amine (NH2) and carboxyl (COOH) modified scaffolds respectively. Plasma polymerisation can be easily applied to other biomaterial surfaces to direct stem cell differentiation for the regeneration of bone and cartilage.
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He P, Liu D, Zhang B, Zhou G, Su X, Wang Y, Li D, Yang X. Hepatitis B Virus X Protein Reduces Podocyte Adhesion via Downregulation of α3β1 Integrin. Cell Physiol Biochem 2017; 41:689-700. [PMID: 28214836 DOI: 10.1159/000458428] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2016] [Accepted: 12/13/2016] [Indexed: 01/27/2023] Open
Abstract
BACKGROUND/AIMS Hepatitis B virus (HBV)-associated glomerulonephritis (HBV-GN) is characterized by a reduced number of podocytes due to apoptosis and shedding from the basement membrane. However, the pathological mechanism of HBV-GN is unclear. We previously showed that hepatitis B virus X protein (HBx) promotes apoptosis in tubular epithelial cells. In this study, we transfected podocytes with HBx and examined the effects on adhesion and apoptosis of these cells. METHODS Podocytes were transfected with pc-DNA3.1 (+)-HBx. One control group was not transfected and another control group was transfected with empty plasmids. Podocyte adhesion was assessed by a fluorescence assay, apoptosis was measured by flow cytometry and fluorescence microscopy, and expression of α3β1 integrin was determined by western blotting and the reverse transcription polymerase chain reaction (RT-PCR). Activity of caspase-8 was measured by a spectrophotometric assay. RESULTS Relative to controls, podocytes with pc-DNA3.1(+)-HBx had reduced cell adhesion, increased apoptosis, reduced expression of α3β1 integrin, and increased caspase-8 activity. β1 integrin blockage reduced podocyte adhesion, but increased apoptosis and caspase-8 activity. Treatment of transfected podocytes with a caspase-8 inhibitor (Z-IETD-FMK) had no effect on the HBx-mediated integrin downregulation and reduced podocyte adhesion, suggesting that α3β1 integrin downregulaton is sufficient to alter cell adhesion. CONCLUSIONS Our in vitro results indicate that HBx reduced podocyte adhesion and expression of α3β1 integrin, and increased apoptosis. Moreover, HBx-mediated downregulation of α3β1 integrin expression is sufficient to reduce podocyte adhesion. HBx-induced apoptosis of podocytes may contribute to HBV-GN.
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Wang L, Lu L. Ultraviolet Irradiation-Induced Volume Alteration of Corneal Epithelial Cells. Invest Ophthalmol Vis Sci 2016; 57:6747-6756. [PMID: 27978555 PMCID: PMC5172162 DOI: 10.1167/iovs.16-19763] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
Purpose The purpose of the study is to understand how extracellular stresses, such as ultraviolet (UV) irradiation, affect corneal epithelial cells. Cell volume changes, damage to corneal epithelial integrity, and cellular responses were assessed after exposure to UVC stresses. Methods Primary human and rabbit corneal epithelial cells were exposed to UVC light in culture conditions. Ultraviolet C irradiation–induced changes in cell size and volume were measured by real-time microscopy and self-quenching of the fluorescent dye calcein, respectively. The effects of UVC irradiation on Src and focal adhesion kinase (FAK) phosphorylation and FAK-dependent integrin signaling were detected by ELISA, immunoblotting, and immunostaining. Results Ultraviolet C irradiation induced both size and volume shifts in human and rabbit corneal epithelial cells. Ultraviolet C irradiation-induced decrease of cell volume elicited activation of Src and FAK, characterized by increased phosphorylations of SrcY416, FAKY397, and FAKY925. In addition, immunostaining studies showed UVC irradiation–induced increases in phosphorylation of FAK and formation of integrin β5 clustering. Application of Kv channel blockers, including 4-aminopyridine (4-AP), α-DTX, and depressing substance-1 (BDS-1), effectively suppressed UVC irradiation–induced cell volume changes, and subsequently inhibited UVC irradiation–induced phosphorylation of Src/FAK, and formation of integrin β5 clustering, suggesting UVC irradiation–induced volume changes and Src/FAK activation. Hyperosmotic pressure–induced volume decreases were measured in comparison with effects of UVC irradiation on volume and Src/FAK activation. However, Kv channel blocker, 4-AP, had no effect on hyperosmotic pressure–induced responses. Conclusions The present study demonstrates that UVC irradiation–induced decreases in cell volume lead to Src/FAK activation due to a rapid loss of K ions through membrane Kv channels.
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Affiliation(s)
- Ling Wang
- Division of Molecular Medicine, Department of Medicine, School of Medicine University of California Los Angeles, Torrance, California, United States
| | - Luo Lu
- Division of Molecular Medicine, Department of Medicine, School of Medicine University of California Los Angeles, Torrance, California, United States
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Luo J, Liu J, Liu H, Zhang T, Wang J, He H, Han C. Enrichment and verification of differentially expressed miRNAs in bursa of Fabricius in two breeds of duck. ASIAN-AUSTRALASIAN JOURNAL OF ANIMAL SCIENCES 2016; 30:920-929. [PMID: 27660025 PMCID: PMC5495669 DOI: 10.5713/ajas.16.0325] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/25/2016] [Revised: 06/06/2016] [Accepted: 09/15/2016] [Indexed: 12/03/2022]
Abstract
Objective The bursa of Fabricius (BF) is a central humoral immune organ belonging specifically to avians. Recent studies had suggested that miRNAs were active regulators involved in the immune processes. This study was to investigate the possible differences of the BF at miRNA level between two genetically disparate duck breeds. Methods Using Illumina next-generation sequencing, the miRNAs libraries of ducks were established. Results The results showed that there were 66 differentially expressed miRNAs and 28 novel miRNAs in bursa. A set of abundant miRNAs (i.e., let-7, miR-146a-5p, miR-21-5p, miR-17~92) which are involved in immunity and disease were detected and the predicted target genes of the novel miRNAs were associated with duck high anti-adversity ability. By gene ontology analysis and enriching KEGG pathway, the targets of differential expressed miRNAs were mainly involved in immunity and disease, supporting that there were differences in the BF immune functions between the two duck breeds. In addition, the metabolic pathway had the maximum enriched target genes and some enriched pathways that were related to cell cycle, protein synthesis, cell proliferation and apoptosis. It indicted that the difference of metabolism may be one of the reasons leading the immune difference between the BF of two duck breeds. Conclusion This data lists the main differences in the BF at miRNAs level between two genetically disparate duck breeds and lays a foundation to carry out molecular assisted breeding of poultry in the future.
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Affiliation(s)
- Jun Luo
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
| | - Junying Liu
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
| | - Hehe Liu
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
| | - Tao Zhang
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
| | - Jiwen Wang
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
| | - Hua He
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
| | - Chunchun Han
- Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu 611130, China
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Roy-Luzarraga M, Hodivala-Dilke K. Molecular Pathways: Endothelial Cell FAK-A Target for Cancer Treatment. Clin Cancer Res 2016; 22:3718-24. [PMID: 27262114 PMCID: PMC5386133 DOI: 10.1158/1078-0432.ccr-14-2021] [Citation(s) in RCA: 80] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2016] [Accepted: 05/13/2016] [Indexed: 01/28/2023]
Abstract
The nonreceptor protein tyrosine kinase, focal adhesion kinase (FAK, also known as PTK2), is a key mediator of signal transduction downstream of integrins and growth factor receptors in a variety of cells, including endothelial cells. FAK is upregulated in several advanced-stage solid tumors and has been described to promote tumor progression and metastasis through effects on both tumor cells and stromal cells. This observation has led to the development of several FAK inhibitors, some of which have entered clinical trials (GSK2256098, VS-4718, VS-6062, VS-6063, and BI853520). Resistance to chemotherapy is a serious limitation of cancer treatment and, until recently, most studies were restricted to tumor cells, excluding the possible roles performed by the tumor microenvironment. A recent report identified endothelial cell FAK (EC-FAK) as a major regulator of chemosensitivity. By dysregulating endothelial cell-derived paracrine (also known as angiocrine) signals, loss of FAK solely in the endothelial cell compartment is able to induce chemosensitization to DNA-damaging therapies in the malignant cell compartment and thereby reduce tumor growth. Herein, we summarize the roles of EC-FAK in cancer and development and review the status of FAK-targeting anticancer strategies. Clin Cancer Res; 22(15); 3718-24. ©2016 AACR.
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Affiliation(s)
- Marina Roy-Luzarraga
- Adhesion and Angiogenesis Laboratory, Centre for Tumor Biology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Kairbaan Hodivala-Dilke
- Adhesion and Angiogenesis Laboratory, Centre for Tumor Biology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom.
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Identification of epidermal growth factor receptor and its inhibitory microRNA141 as novel targets of Krüppel-like factor 8 in breast cancer. Oncotarget 2016; 6:21428-42. [PMID: 26025929 PMCID: PMC4673276 DOI: 10.18632/oncotarget.4077] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/17/2015] [Accepted: 05/14/2015] [Indexed: 01/22/2023] Open
Abstract
Krüppel-like factor 8 (KLF8) is a dual transcriptional factor critical for breast cancer progression. Epidermal growth factor receptor (EGFR) is frequently overexpressed in aggressive such as triple-negative breast cancer and associated with poor clinical outcomes. Here we report a novel KLF8-EGFR signaling axis in breast cancer. We identified a highly correlated co-overexpression between KLF8 and EGFR in invasive breast cancer cells and patient tumor samples. Overexpression of KLF8 in the non-tumorigenic MCF-10A cells induced the expression of EGFR, whereas knockdown of KLF8 from the MDA-MB-231 cells decreased it. Promoter activation and binding assays indicated that KLF8 promotes the EGFR expression by directly binding its gene promoter. We also revealed that KLF8 directly represses the promoter of miR141 and miR141 targets the 3′-untranslational region of EGFR transcript to inhibit EGFR translation. Treatment with the EGFR inhibitor AG1478 or overexpression of miR141 blocked the activity of ERK downstream of EGFR and inhibited KLF8-depndent cell invasiveness, proliferation and viability in cell culture and invasive growth and lung metastasis in nude mice. Conversely, overexpression of an inhibitory sponge of miR141 led to the opposite phenotypes. Taken together, these findings demonstrate a novel KLF8 to miR141/EGFR signaling pathway potentially crucial for breast cancer malignancy.
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Lahiri SK, Lu H, Mukherjee D, Yu L, Zhao J. ERK2 phosphorylates Krüppel-like factor 8 protein at serine 48 to maintain its stability. Am J Cancer Res 2016; 6:910-923. [PMID: 27293988 PMCID: PMC4889709] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2016] [Accepted: 02/22/2016] [Indexed: 06/06/2023] Open
Abstract
Krüppel-like factor 8 (KLF8) plays important roles in cancer and is strictly regulated by various post-translational modifications such as sumoylation, acetylation, ubiquitylation and PARylation. Here we report a novel phosphorylation of KLF8 by ERK2 responsible and critical for the stability of KLF8 protein. The full-length KLF8 protein displays a doublet in SDS-PAGE gel. The upper band of the doublet, however, disappeared when the N-terminal 50 amino acids were deleted. In its full-length the upper band disappeared upon phosphatase treatment or mutation of the serine 48 (S48) to alanine whereas the lower band was lost when the S48 was mutated to aspartic acid that mimics phosphorylated S48. These results suggest that S48 phosphorylation is responsible for the motility up-shift of KLF8 protein. Pharmacological and genetic manipulations of various potential kinases identified ERK2 as the likely one that phosphorylates KLF8 at S48. Functional studies indicated that this phosphorylation is crucial for protecting KLF8 protein from degradation in the nucleus and promoting cell migration. Taken together, this study identifies a novel mechanism of phosphorylation critical for KLF8 protein stabilization and function.
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Affiliation(s)
- Satadru K Lahiri
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine Orlando, Florida 32827
| | - Heng Lu
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine Orlando, Florida 32827
| | - Debarati Mukherjee
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine Orlando, Florida 32827
| | - Lin Yu
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine Orlando, Florida 32827
| | - Jihe Zhao
- Burnett School of Biomedical Sciences University of Central Florida College of Medicine Orlando, Florida 32827
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Direito I, Madeira A, Brito MA, Soveral G. Aquaporin-5: from structure to function and dysfunction in cancer. Cell Mol Life Sci 2016; 73:1623-40. [PMID: 26837927 PMCID: PMC11108570 DOI: 10.1007/s00018-016-2142-0] [Citation(s) in RCA: 74] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/17/2015] [Revised: 12/29/2015] [Accepted: 01/18/2016] [Indexed: 01/05/2023]
Abstract
Aquaporins, a highly conserved group of membrane proteins, are involved in the bidirectional transfer of water and small solutes across cell membranes taking part in many biological functions all over the human body. In view of the wide range of cancer malignancies in which aquaporin-5 (AQP5) has been detected, an increasing interest in its implication in carcinogenesis has emerged. Recent publications suggest that this isoform may enhance cancer cell proliferation, migration and survival in a variety of malignancies, with strong evidences pointing to AQP5 as a promising drug target and as a novel biomarker for cancer aggressiveness with high translational potential for therapeutics and diagnostics. This review addresses the structural and functional features of AQP5, detailing its tissue distribution and functions in human body, its expression pattern in a variety of tumors, and highlighting the underlying mechanisms involved in carcinogenesis. Finally, the actual progress of AQP5 research, implications in cancer biology and potential for cancer detection and prognosis are discussed.
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Affiliation(s)
- Inês Direito
- Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, 1649-003, Lisbon, Portugal
- Department of Biochemistry and Human Biology, Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal
| | - Ana Madeira
- Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, 1649-003, Lisbon, Portugal
- Department of Biochemistry and Human Biology, Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal
| | - Maria Alexandra Brito
- Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, 1649-003, Lisbon, Portugal
- Department of Biochemistry and Human Biology, Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal
| | - Graça Soveral
- Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, 1649-003, Lisbon, Portugal.
- Department of Biochemistry and Human Biology, Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
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HER2 drives Mucin-like 1 to control proliferation in breast cancer cells. Oncogene 2016; 35:4225-34. [PMID: 26725324 PMCID: PMC4996539 DOI: 10.1038/onc.2015.487] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2015] [Revised: 10/22/2015] [Accepted: 11/14/2015] [Indexed: 01/09/2023]
Abstract
Mucin-like 1 (MUCL1) was first identified as a breast-specific gene over a decade
ago. Based on its highly restricted mRNA expression in breast tissue and
continued expression during breast tumorigenesis and progression, MUCL1 is an
attractive tumor-associated antigen and a potential therapeutic target. However,
very little is known about the cellular location, biological functions and
regulation of the MUCL1 protein, which will have a major impact on its
druggability. Here we describe our efforts to fully characterize the cellular
localization of MUCL1, investigate its regulation by key breast cancer oncogenes
such as human epidermal growth factor receptor 2 (HER2) and discover its
functional roles in breast cancer. Although some mucins are membrane bound, our
data indicate that MUCL1 is secreted by some breast cancer cells, whereas others
only express high levels of intracellular MUCL1. MUCL1 expression is highest in
HER2-amplified breast tumors and inhibiting HER2 activity in tumor cells
resulted in a decreased MUCL1 expression. In-depth investigation demonstrated
that phosphoinositide3-kinase/Akt pathway, but not Ras/MEK pathway,
controls MUCL1 expression downstream of HER2. Phenotypic assays revealed a
strong dependence of HER2-positive cells on MUCL1 for cell proliferation. We
further identified the mechanism by which MUCL1 regulates cell growth. Knockdown
of MUCL1 induced a G1/S phase arrest concomitant with decreased cyclin D and
increased p21 and p27 levels. Finally, we investigated the impact of MUCL1 loss
on kinase signaling pathways in breast cancer cells through phospho-kinase array
profiling. MUCL1 silencing abrogated phospho-focal adhesion kinase (FAK), Jun
NH2-terminal kinase (JNK) and c-Jun signals, but not
extracellular signal-regulated kinase or Akt pathway activities, thereby
pointing to FAK/JNK pathway as the downstream effector of MUCL1 signaling.
We are the first to identify an important role for MUCL1 in the proliferation of
breast cancer cells, probably mediated via the FAK/JNK signaling pathway.
Taken together, these data suggest a potential utility for therapeutic targeting
of this protein in breast cancer.
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Webber PJ, Park C, Qui M, Ramalingam SS, Khuri FR, Fu H, Du Y. Combination of heat shock protein 90 and focal adhesion kinase inhibitors synergistically inhibits the growth of non-small cell lung cancer cells. Oncoscience 2015; 2:765-776. [PMID: 26501082 PMCID: PMC4606010 DOI: 10.18632/oncoscience.245] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2015] [Accepted: 09/12/2015] [Indexed: 02/07/2023] Open
Abstract
Discovery of effective drug combinations is a promising strategy to improve patient survival. This study explores the impact of heat shock protein 90 (Hsp90) inhibition in combination with focal adhesion kinase (FAK) inhibitor on the growth of non-small cell lung cancer cells (NSCLC cells). Our data show that 17-N-Allylamino-17-demethoxygeldanamycin (17-AAG), a well-studied Hsp90 inhibitor, synergized with FAK inhibitor, PF-573228, on the growth inhibition of NSCLC cells. This combination effect was confirmed using additional chemically distinct Hsp90 inhibitor, STA-9090, which is currently undergoing phase 3 clinical evaluation. Co-treatment of NSCLC cells with Hsp90 and FAK inhibitors significantly enhanced the inhibition on long-term colony formation compared to that with single agent. Inhibition of FAK exacerbated the G2 cell cycle arrest and annexin-V apoptotic staining induced by 17-AAG. Further mechanistic studies revealed that the combination of Hsp90 and FAK inhibitors reduced the activity of canonical proliferative and survival Akt-mTOR signaling, and increased pro-apoptotic caspase activation. Interestingly, FAK inhibition alone induced feedback activation of pro-survival Erk signaling, which was abrogated by co-treatment with Hsp90 inhibitors. Both Hsp90 and FAK inhibitors are undergoing clinical evaluation. Our studies suggest the tandem of Hsp90 and FAK inhibitors may provide an effective treatment option for NSCLC patients.
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Affiliation(s)
- Philip J Webber
- Department of Pharmacology, Emory University, Atlanta, GA, USA
| | - Chanhee Park
- Department of Pharmacology, Emory University, Atlanta, GA, USA
| | - Min Qui
- Department of Pharmacology, Emory University, Atlanta, GA, USA.,Emory Chemical Biology Discovery Center, Emory University School of Medicine, Emory University, Atlanta, GA, USA
| | - Suresh S Ramalingam
- Department of Hematology and Medical Oncology, Winship Cancer Institute, Emory University, Atlanta, GA, USA
| | - Fadlo R Khuri
- Department of Hematology and Medical Oncology, Winship Cancer Institute, Emory University, Atlanta, GA, USA
| | - Haian Fu
- Department of Pharmacology, Emory University, Atlanta, GA, USA.,Emory Chemical Biology Discovery Center, Emory University School of Medicine, Emory University, Atlanta, GA, USA.,Department of Hematology and Medical Oncology, Winship Cancer Institute, Emory University, Atlanta, GA, USA
| | - Yuhong Du
- Department of Pharmacology, Emory University, Atlanta, GA, USA.,Emory Chemical Biology Discovery Center, Emory University School of Medicine, Emory University, Atlanta, GA, USA
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Zhang H, Yang S, Masako N, Lee DJ, Cooper LF, Ko CC. Proliferation of preosteoblasts on TiO 2 nanotubes is FAK/RhoA related. RSC Adv 2015; 5:38117-38124. [PMID: 26090090 PMCID: PMC4467958 DOI: 10.1039/c4ra16803h] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
Topographies promote surface-dependent behaviors which may positively influence peri-implant bone healing. In this study the topological effects of TiO2 nanotubes (TNTs) on aspects of preosteoblast behavior was investigated. Specifically, we hypothesize that TNTs can influence cell proliferation of preosteoblasts through cell adhesion and related modulation of FAK and RhoA. By culturing MC3T3-E1 cells on TNTs with different diameters (40nm and 150nm diameters), topography-dependent modulation in cell morphology and cell growth were observed. The average spreading area of the cell on Flat Ti, 40nm TNTs and 150nm TNTs were respectively 2176.05 μm2, 1510.44 μm2 and 800.72 μm2. Proliferation increased among cells cultured on the 150nm TNTs (28.6%) compared with on Flat Ti (17.06%). The expression of FAK was 86.2% down regulated superimposition of TNTs topography. RhoA expression only slightly decreased (45.9%). Increasing TNT diameter enhanced initial adherent cell growth, which was relevant to the increased RhoA-to-FAK ratio in the cell. Increased TNT diameter was associated with higher ratio and greater proliferation in the first 24 hours. These findings not only support our hypothesis, but suggest that RhoA might be critically involved in TNTs mediated cell proliferation. Future investigation using functional gain and loss of RhoA may further reveal its mechanism.
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Affiliation(s)
- He Zhang
- Chongqing Key Laboratory of Oral Diseases and Biomedical Sciences, Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Hi gher Education, College of Stomatology, Chongqing Medical University, Chongqing 401147, China
| | - Sheng Yang
- Chongqing Key Laboratory of Oral Diseases and Biomedical Sciences, Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Hi gher Education, College of Stomatology, Chongqing Medical University, Chongqing 401147, China
| | - Nagasawa Masako
- Division of Bioprosthodontics, Department of Oral Science, Niigata University Graduate School of Medical and Dental Sciences, Niigata 951-8514, Japan ; NC Oral Health Institute, School of Dentistry, University of North Carolina, CB #7454, Chapel Hill, NC 27599, USA
| | - Dong Joon Lee
- NC Oral Health Institute, School of Dentistry, University of North Carolina, CB #7454, Chapel Hill, NC 27599, USA
| | - Lyndon F Cooper
- NC Oral Health Institute, School of Dentistry, University of North Carolina, CB #7454, Chapel Hill, NC 27599, USA ; Department of Prosthodontics, School of Dentistry, University of North Carolina, CB #7450, Chapel Hill, NC 27599-7450, USA
| | - Ching-Chang Ko
- NC Oral Health Institute, School of Dentistry, University of North Carolina, CB #7454, Chapel Hill, NC 27599, USA ; Department of Orthodontics, School of Dentistry, University of North Carolina, CB #7450, Chapel Hill, NC 27599-7450, USA
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Cellular processes involved in human epidermal cells exposed to extremely low frequency electric fields. Cell Signal 2015; 27:889-98. [DOI: 10.1016/j.cellsig.2015.02.007] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2014] [Accepted: 02/08/2015] [Indexed: 01/18/2023]
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Emerging roles of focal adhesion kinase in cancer. BIOMED RESEARCH INTERNATIONAL 2015; 2015:690690. [PMID: 25918719 PMCID: PMC4396139 DOI: 10.1155/2015/690690] [Citation(s) in RCA: 153] [Impact Index Per Article: 15.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/16/2015] [Accepted: 02/20/2015] [Indexed: 12/13/2022]
Abstract
Focal adhesion kinase (FAK) is a cytoplasmic nonreceptor tyrosine kinase that enables activation by growth factor receptors or integrins in various types of human cancers. The kinase-dependent and kinase-independent scaffolding functions of FAK modulate the authentic signaling and fundamental functions not only in cancer cells but also in tumor microenvironment to facilitate cancer progression and metastasis. The overexpression and activation of FAK are usually investigated in primary or metastatic cancers and correlated with the poor clinical outcome, highlighting FAK as a potential prognostic marker and anticancer target. Small molecule inhibitors targeting FAK kinase activity or FAK-scaffolding functions impair cancer development in preclinical or clinical trials. In this review, we give an overview for FAK signaling in cancer cells as well as tumor microenvironment that provides new strategies for the invention of cancer development and malignancy.
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48
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Integrin-mediated adhesion and mechano-sensing in cutaneous wound healing. Cell Tissue Res 2014; 360:571-82. [DOI: 10.1007/s00441-014-2064-9] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2014] [Accepted: 11/11/2014] [Indexed: 12/30/2022]
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Kasi RAP, Moi CS, Kien YW, Yian KR, Chin NW, Yen NK, Ponnudurai G, Fong SH. Para-phenylenediamine-induces apoptosis via a pathway dependent on PTK-Ras-Raf-JNK activation but independent of the PI3K/Akt pathway in NRK-52E cells. Mol Med Rep 2014; 11:2262-8. [PMID: 25411820 DOI: 10.3892/mmr.2014.2979] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2013] [Accepted: 08/08/2014] [Indexed: 11/06/2022] Open
Abstract
para‑Phenylenediamine (p‑PD) is a potential carcinogen, and widely used in marketed hair dye formulations. In the present study, the role of the protein tyrosine kinase (PTK)/Ras/Raf/c‑Jun N‑terminal kinase (JNK) and phosphoinositide 3‑kinase (PI3k)/protein kinase B (Akt) pathways on the growth of NRK‑52E cells was investigated. The results demonstrated that p‑PD reduced cell viability in a dose‑dependent manner. The cell death due to apoptosis was confirmed by cell cycle analysis and an Annexin‑V‑fluorescein isothiocyanate binding assay. Subsequent to staining with 2',7'‑dichlorofluorescin diacetate, the treated cells demonstrated a significant increase in reactive oxygen species (ROS) generation compared with the controls. The effects of p‑PD on the signalling pathways were analysed by western blotting. p‑PD‑treated cells exhibited an upregulated phospho‑stress‑activated protein kinase/JNK protein expression level and downregulated Ras and Raf protein expression levels; however, Akt, Bcl‑2, Bcl‑XL and Bad protein expression levels were not significantly altered compared with the control. In conclusion, p‑PD induced apoptosis by a PTK/Ras/Raf/JNK‑dependent pathway and was independent of the PI3K/Akt pathway in NRK‑52E cells.
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Affiliation(s)
- Reena A P Kasi
- Department of Human Biology, Cells and Molecules, International Medical University, Kuala Lumpur 57000, Malaysia
| | - Chye Soi Moi
- Department of Human Biology, Cells and Molecules, International Medical University, Kuala Lumpur 57000, Malaysia
| | - Yip Wai Kien
- Department of Pathology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Serdang, Selangor 43400, Malaysia
| | - Koh Rhun Yian
- Department of Human Biology, Cells and Molecules, International Medical University, Kuala Lumpur 57000, Malaysia
| | - Ng Wei Chin
- Department of Human Biology, Cells and Molecules, International Medical University, Kuala Lumpur 57000, Malaysia
| | - Ng Khuen Yen
- Jeffrey Cheah School of Medicine and Health Sciences, Monash University Sunway Campus, Bandar Sunway, Selangor 47500, Malaysia
| | - Gnanajothy Ponnudurai
- Department of Human Biology, Cells and Molecules, International Medical University, Kuala Lumpur 57000, Malaysia
| | - Seow Heng Fong
- Department of Pathology, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Serdang, Selangor 43400, Malaysia
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Abstract
In cancer dormancy, residual tumor cells persist in a patient with no apparent clinical symptoms, only to potentially become clinically relevant at a later date. In prostate cancer (PCa), the primary tumor is often removed and many patients experience a prolonged period (>5 years) with no evidence of disease before recurrence. These characteristics make PCa an excellent candidate for the study of tumor cell dormancy. However, the mechanisms that constitute PCa dormancy have not been clearly defined. Additionally, the definition of tumor cell dormancy varies in the literature. Therefore, we have separated tumor cell dormancy in this review into three categories: (a) micrometastatic dormancy--a group of tumor cells that cannot increase in number due to a restrictive proliferation/apoptosis equilibrium. (b) Angiogenic dormancy--a group of tumor cells that cannot expand beyond the formation of a micrometastasis due to a lack of angiogenic potential. (c) Conditional dormancy--an individual cell or a very small number of cells that cannot proliferate without the appropriate cues from the microenvironment, but do not require angiogenesis to do so. This review aims to identify currently known markers, mechanisms, and models of tumor dormancy, in particular as they relate to PCa, and highlight current opportunities for advancement in our understanding of clinical cancer dormancy.
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