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Lainscsek X, Kong W, Rütgen BC, Beck J, Brenig B, Nolte I, Murua Escobar H, Taher L. Transcriptomic profiling in canine B-cell lymphoma supports a synergistic effect of BTK and PI3K inhibitors. Front Vet Sci 2025; 12:1577028. [PMID: 40351764 PMCID: PMC12063356 DOI: 10.3389/fvets.2025.1577028] [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: 02/14/2025] [Accepted: 03/31/2025] [Indexed: 05/14/2025] Open
Abstract
Introduction B-cell receptor (BCR) signaling has revealed itself as a critical pathway in the pathogenesis of B-cell lymphoma. Within this pathway, the inhibition of Bruton's tyrosine kinase (BTK) or Phosphoinositide 3-kinases (PI3Ks) alone presents encouraging efficacy in the treatment of certain both canine and human hematological malignancies. Methods Here we characterized the effects of the BTK inhibitor Ibrutinib and the PI3K inhibitor AS-605240 as single and combined agents in the canine pre-clinical diffuse large B cell lymphoma (DLBCL) model CLBL-1 by assaying cell proliferation and metabolic activity, and performing RNA-seq to measure gene expression changes. Results We found 2,336 differentially expressed genes (DEGs) across all treatment types and time points relative to the control. The largest number of DEGs were induced by the combination of Ibrutinib and AS-605240. These genes were involved in adaptive immune response, leukotriene D4 metabolic and terms related to regulation of GTP and GTPase mediated signal transduction. Weighted gene co-expression network analysis (WGCNA) detected nine gene modules, five of which were associated with treatment response. Eighteen-percent of genes within these modules were also differentially expressed. Notably, we observed one module that was exclusively associated with the combined treatment whose gene members were related to cellular metabolism, homeostasis signaling, and protein synthesis and regulation. Conclusion Narrowing in on highly connected genes of modules associated with treatment response with large fold changes across treatments which play roles in the main targeted pathways identified PAG1, PRKAR2A, ACACA, FOS, and PRKCA as potential primary candidates of the synergistic treatment effect.
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Affiliation(s)
- Xenia Lainscsek
- Institute of Biomedical Informatics, Graz University of Technology, Graz, Austria
| | - Weibo Kong
- Clinic for Hematology, Oncology and Palliative Care, Rostock University Medical Center, University of Rostock, Rostock, Germany
| | - Barbara C. Rütgen
- Department for Pathobiology, Clinical Pathology, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Julia Beck
- Chronix Biomedical GmbH, Göttingen, Germany
| | - Bertram Brenig
- Institute of Veterinary Medicine, University of Göttingen, Göttingen, Germany
| | - Ingo Nolte
- Small Animal Clinic, University of Veterinary Medicine Hannover, Hannover, Germany
| | - Hugo Murua Escobar
- Clinic for Hematology, Oncology and Palliative Care, Rostock University Medical Center, University of Rostock, Rostock, Germany
- Institute of Medical Genetics, Rostock University Medical Center, University of Rostock, Rostock, Germany
| | - Leila Taher
- Institute of Biomedical Informatics, Graz University of Technology, Graz, Austria
- Institute for Biostatistics and Informatics in Medicine and Ageing Research, Rostock University Medical Center, University of Rostock, Rostock, Germany
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Wang Z, Guo Y, Li K, Huo Y, Wang S, Dong S, Ma M. Targeting the PI3K/mTOR pathway in idiopathic pulmonary fibrosis: Advances and therapeutic potential. Bioorg Med Chem 2024; 115:117908. [PMID: 39471771 DOI: 10.1016/j.bmc.2024.117908] [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: 07/18/2024] [Revised: 09/02/2024] [Accepted: 09/03/2024] [Indexed: 11/01/2024]
Abstract
Idiopathic pulmonary fibrosis (IPF) is a progressive, fatal lung disease characterized by irreversible tissue scarring, leading to severe respiratory dysfunction. Despite current treatments with the drugs Pirfenidone and Nintedanib, effective management of IPF remains inadequate due to limited therapeutic benefits and significant side effects. This review focuses on the phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) signaling pathway, a critical regulator of cellular processes linked to fibrosis, such as fibroblast proliferation, inflammation, and epithelial-mesenchymal transition (EMT). We discuss recent advances in understanding the role of the PI3K/mTOR pathway in IPF pathogenesis and highlight emerging therapies targeting this pathway. The review compiles evidence from both preclinical and clinical studies, suggesting that PI3K/mTOR inhibitors may offer new hope for IPF treatment by modulating fibrosis and improving patient outcomes. Moreover, it outlines the potential for these inhibitors to be developed into effective, personalized treatment options, underscoring the importance of further research to explore their efficacy and safety profiles comprehensively.
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Affiliation(s)
- Zhengyang Wang
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai 200062, China
| | - Yanzhi Guo
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai 200062, China
| | - Kaiyin Li
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai 200062, China
| | - Yan Huo
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai 200062, China
| | - Shuyan Wang
- Department of Anesthesiology, Tongren Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200336, China
| | - Suzhen Dong
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai 200062, China.
| | - Mingliang Ma
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai 200062, China; Wenzhou Key Laboratory of Biophysics, Wenzhou Institute, University of Chinese Academy of Sciences, Wenzhou, Zhejiang 325000, China.
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Ma Y, Sender S, Sekora A, Kong W, Bauer P, Ameziane N, Al-Ali R, Krake S, Radefeldt M, Weiss FU, Lerch MM, Parveen A, Zechner D, Junghanss C, Murua Escobar H. The Inhibitory Response to PI3K/AKT Pathway Inhibitors MK-2206 and Buparlisib Is Related to Genetic Differences in Pancreatic Ductal Adenocarcinoma Cell Lines. Int J Mol Sci 2022; 23:4295. [PMID: 35457111 PMCID: PMC9029322 DOI: 10.3390/ijms23084295] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2022] [Revised: 04/08/2022] [Accepted: 04/11/2022] [Indexed: 01/23/2023] Open
Abstract
The aberrant activation of the phosphoinositide 3-kinase (PI3K)/ protein kinase B (AKT) pathway is common in pancreatic ductal adenocarcinomas (PDAC). The application of inhibitors against PI3K and AKT has been considered as a therapeutic option. We investigated PDAC cell lines exposed to increasing concentrations of MK-2206 (an AKT1/2/3 inhibitor) and Buparlisib (a pan-PI3K inhibitor). Cell proliferation, metabolic activity, biomass, and apoptosis/necrosis were evaluated. Further, whole-exome sequencing (WES) and RNA sequencing (RNA-seq) were performed to analyze the recurrent aberrations and expression profiles of the inhibitor target genes and the genes frequently mutated in PDAC (Kirsten rat sarcoma virus (KRAS), Tumor protein p53 (TP53)). MK-2206 and Buparlisib demonstrated pronounced cytotoxic effects and limited cell-line-specific effects in cell death induction. WES revealed two sequence variants within the direct target genes (PIK3CA c.1143C > G in Colo357 and PIK3CD c.2480C > G in Capan-1), but a direct link to the Buparlisib response was not observed. RNA-seq demonstrated that the expression level of the inhibitor target genes did not affect the efficacy of the corresponding inhibitors. Moreover, increased resistance to MK-2206 was observed in the analyzed cell lines carrying a KRAS variant. Further, increased resistance to both inhibitors was observed in SU.86.86 carrying two TP53 missense variants. Additionally, the presence of the PIK3CA c.1143C > G in KRAS-variant-carrying cell lines was observed to correlate with increased sensitivity to Buparlisib. In conclusion, the present study reveals the distinct antitumor effects of PI3K/AKT pathway inhibitors against PDAC cell lines. Aberrations in specific target genes, as well as KRAS and TP53, individually or together, affect the efficacy of the two PI3K/AKT pathway inhibitors.
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Affiliation(s)
- Yixuan Ma
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
| | - Sina Sender
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
| | - Anett Sekora
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
| | - Weibo Kong
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
- Institute of Muscle Biology and Growth, Research Institute for Farm Animal Biology (FBN), 18196 Dummerstorf, Germany
| | - Peter Bauer
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
- CENTOGENE GmbH, 18057 Rostock, Germany; (N.A.); (R.A.-A.); (S.K.); (M.R.)
| | - Najim Ameziane
- CENTOGENE GmbH, 18057 Rostock, Germany; (N.A.); (R.A.-A.); (S.K.); (M.R.)
- Arcensus GmbH, 18055 Rostock, Germany
| | - Ruslan Al-Ali
- CENTOGENE GmbH, 18057 Rostock, Germany; (N.A.); (R.A.-A.); (S.K.); (M.R.)
| | - Susann Krake
- CENTOGENE GmbH, 18057 Rostock, Germany; (N.A.); (R.A.-A.); (S.K.); (M.R.)
| | - Mandy Radefeldt
- CENTOGENE GmbH, 18057 Rostock, Germany; (N.A.); (R.A.-A.); (S.K.); (M.R.)
| | - Frank Ulrich Weiss
- Department of Medicine A, University Medicine, University of Greifswald, 17475 Greifswald, Germany; (F.U.W.); (M.M.L.)
| | - Markus M. Lerch
- Department of Medicine A, University Medicine, University of Greifswald, 17475 Greifswald, Germany; (F.U.W.); (M.M.L.)
- LMU Munich University Hospital, 81377 Munich, Germany
| | - Alisha Parveen
- Institute for Experimental Surgery, University of Rostock, 18057 Rostock, Germany; (A.P.); (D.Z.)
| | - Dietmar Zechner
- Institute for Experimental Surgery, University of Rostock, 18057 Rostock, Germany; (A.P.); (D.Z.)
| | - Christian Junghanss
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
| | - Hugo Murua Escobar
- Department of Medicine Clinic III, Hematology, Oncology and Palliative Medicine, Rostock University Medical Center, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany; (Y.M.); (S.S.); (A.S.); (W.K.); (P.B.); (C.J.)
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Class I PI3K Biology. Curr Top Microbiol Immunol 2022; 436:3-49. [DOI: 10.1007/978-3-031-06566-8_1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
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Evans JM, Parker HG, Rutteman GR, Plassais J, Grinwis GCM, Harris AC, Lana SE, Ostrander EA. Multi-omics approach identifies germline regulatory variants associated with hematopoietic malignancies in retriever dog breeds. PLoS Genet 2021; 17:e1009543. [PMID: 33983928 PMCID: PMC8118335 DOI: 10.1371/journal.pgen.1009543] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2021] [Accepted: 04/12/2021] [Indexed: 12/18/2022] Open
Abstract
Histiocytic sarcoma is an aggressive hematopoietic malignancy of mature tissue histiocytes with a poorly understood etiology in humans. A histologically and clinically similar counterpart affects flat-coated retrievers (FCRs) at unusually high frequency, with 20% developing the lethal disease. The similar clinical presentation combined with the closed population structure of dogs, leading to high genetic homogeneity, makes dogs an excellent model for genetic studies of cancer susceptibility. To determine the genetic risk factors underlying histiocytic sarcoma in FCRs, we conducted multiple genome-wide association studies (GWASs), identifying two loci that confer significant risk on canine chromosomes (CFA) 5 (Pwald = 4.83x10-9) and 19 (Pwald = 2.25x10-7). We subsequently undertook a multi-omics approach that has been largely unexplored in the canine model to interrogate these regions, generating whole genome, transcriptome, and chromatin immunoprecipitation sequencing. These data highlight the PI3K pathway gene PIK3R6 on CFA5, and proximal candidate regulatory variants that are strongly associated with histiocytic sarcoma and predicted to impact transcription factor binding. The CFA5 association colocalizes with susceptibility loci for two hematopoietic malignancies, hemangiosarcoma and B-cell lymphoma, in the closely related golden retriever breed, revealing the risk contribution this single locus makes to multiple hematological cancers. By comparison, the CFA19 locus is unique to the FCR and harbors risk alleles associated with upregulation of TNFAIP6, which itself affects cell migration and metastasis. Together, these loci explain ~35% of disease risk, an exceptionally high value that demonstrates the advantages of domestic dogs for complex trait mapping and genetic studies of cancer susceptibility. We have identified two regions of the canine genome that explain a striking 35% of risk for developing histiocytic sarcoma in FCRs. The disease is uniformly lethal, affects 20% of FCRs, and parallels a cancer of the same name in humans. Both regions harbor genes involved in cell migration and cancer-related pathways. The first includes variants in regulatory regions at the tumor suppressor PIK3R6 locus that are strongly associated with histiocytic sarcoma and likely confer risk for other hematopoietic cancers. FCRs with risk alleles at the second locus demonstrate increased expression of TNFAIP6, which correlates with poor prognosis in multiple human cancers. In identifying genomic differences between affected and unaffected dogs, we advance our understanding of both canine and human health biology and set the stage for the development of diagnostic and therapeutic strategies.
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Affiliation(s)
- Jacquelyn M. Evans
- Cancer Genetics and Comparative Genomics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Heidi G. Parker
- Cancer Genetics and Comparative Genomics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Gerard R. Rutteman
- Department of Clinical Sciences, division Internal Medicine of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands
| | - Jocelyn Plassais
- Cancer Genetics and Comparative Genomics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Guy C. M. Grinwis
- Department Biomedical Health Sciences, division Pathology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands
| | - Alexander C. Harris
- Cancer Genetics and Comparative Genomics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Susan E. Lana
- College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
| | - Elaine A. Ostrander
- Cancer Genetics and Comparative Genomics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America
- * E-mail:
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6
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Yang C, Xu W, Gong J, Liu Z, Cui D. Novel somatic alterations underlie Chinese papillary thyroid carcinoma. Cancer Biomark 2020; 27:445-460. [PMID: 32065787 DOI: 10.3233/cbm-191200] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
To characterize the somatic alterations of papillary thyroid carcinomas (PTC) in Chinese patients, we performed the next-generation-sequencing (NGS) study of the tumor-normal pairs of DNA and RNA samples extracted from 16 Chinese PTC patients. The whole genome sequencing (WGS) and transcriptome sequencing (RNA-seq) were conducted for 6 patients who were either current or former smokers and the whole exome sequencing (WES) and RNA-seq were conducted for another 10 patients who were never smokers. The NGS data were analyzed to identify somatic alteration events that may underlie PTC in Chinese patients. We identified a number of PTC driver genes harboring somatic driver mutations with significant functional impact such as COL11A1, TP53, PLXNA4, UBA1, AHNAK, CSMD2 and TTLL5 etc. Significant driver pathways underlying PTC were found, namely, the metabolic pathway, the pathway in cancer, the olfactory transduction pathway and the calcium signaling pathway. In addition, this study revealed genes with significant somatic copy number aberrations and corresponding somatic gene expression changes in PTC tumors, the most promising ones being BRD9, TRIP13, FZD3, and TFDP1 etc. We also identified several structural variants of PTCs, especially the novel in-frame fusion proteins such as TRNAU1AP-RCC1, RAB3GAP1-R3HDM1, and ENAH-ZSWIM5. Our study provided a list of novel PTC candidate genes with somatic alterations that may function as biomarkers for PTC in Chinese patients. The follow-up mechanism studies may be conducted based on the findings from this study.
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Affiliation(s)
- Chuanjia Yang
- Department of General Surgery, Shengjing Hospital of China Medical University, Shenyang, Liaoning, China
| | - Weixue Xu
- Department of General Surgery, Shengjing Hospital of China Medical University, Shenyang, Liaoning, China
| | - Jian Gong
- Department of Clinical Pharmacy, School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, Shenyang, Liaoning, China
| | - Zhen Liu
- Department of General Surgery, Shengjing Hospital of China Medical University, Shenyang, Liaoning, China
| | - Dongxu Cui
- Department of General Surgery, Shengjing Hospital of China Medical University, Shenyang, Liaoning, China
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Zhang M, Jang H, Nussinov R. Structural Features that Distinguish Inactive and Active PI3K Lipid Kinases. J Mol Biol 2020; 432:5849-5859. [PMID: 32918948 PMCID: PMC8916166 DOI: 10.1016/j.jmb.2020.09.002] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2020] [Revised: 08/10/2020] [Accepted: 09/02/2020] [Indexed: 12/25/2022]
Abstract
PI3K lipid kinases signal through the PI3K/Akt pathway, regulating cell growth and proliferation. While the structural features that distinguish between the active and inactive states of protein kinases are well established, that has not been the case for lipid kinases, and neither was the structural mechanism controlling the switch between the two states. Class I PI3Ks are obligate heterodimers with catalytic and regulatory subunits. Here, we analyze PI3K crystal structures. Structures with the nSH2 (inactive state) are featured by collapsed activation loop (a-loop) and an IN kinase domain helix 11 (kα11). In the active state, the a-loop is extended and kα11 in the OUT conformation. Our analysis suggests that the nSH2 domain in the regulatory subunit regulates activation, catalysis and autoinhibition through the a-loop. Inhibition, activation and catalytic scenarios are shared by class IA PI3Ks; the activation is mimicked by oncogenic mutations and the inhibition offers an allosteric inhibitor strategy.
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Affiliation(s)
- Mingzhen Zhang
- Computational Structural Biology Section, Frederick National Laboratory for Cancer Research, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | - Hyunbum Jang
- Computational Structural Biology Section, Frederick National Laboratory for Cancer Research, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | - Ruth Nussinov
- Computational Structural Biology Section, Frederick National Laboratory for Cancer Research, National Cancer Institute at Frederick, Frederick, MD 21702, USA; Department of Human Molecular Genetics and Biochemistry, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
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Rynkiewicz NK, Anderson KE, Suire S, Collins DM, Karanasios E, Vadas O, Williams R, Oxley D, Clark J, Stephens LR, Hawkins PT. Gβγ is a direct regulator of endogenous p101/p110γ and p84/p110γ PI3Kγ complexes in mouse neutrophils. Sci Signal 2020; 13:13/656/eaaz4003. [PMID: 33144519 DOI: 10.1126/scisignal.aaz4003] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
The PI3Kγ isoform is activated by Gi-coupled GPCRs in myeloid cells, but the extent to which the two endogenous complexes of PI3Kγ, p101/p110γ and p84/p110γ, receive direct regulation through Gβγ or indirect regulation through RAS and the sufficiency of those inputs is controversial or unclear. We generated mice with point mutations that prevent Gβγ binding to p110γ (RK552DD) or to p101 (VVKR777AAAA) and investigated the effects of these mutations in primary neutrophils and in mouse models of neutrophilic inflammation. Loss of Gβγ binding to p110γ substantially reduced the activation of both p101/p110γ and p84/p110γ in neutrophils by various GPCR agonists. Loss of Gβγ binding to p101 caused more variable effects, depending on both the agonist and cellular response, with the biggest reductions seen in PIP3 production by primary neutrophils in response to LTB4 and MIP-2 and in the migration of neutrophils during thioglycolate-induced peritonitis or MIP2-induced ear pouch inflammation. We also observed that p101VVKR777AAAA neutrophils showed enhanced p84-dependent ROS responses to fMLP and C5a, suggesting that competition may exist between p101/p110γ and p84/p110γ for Gβγ subunits downstream of GPCR activation. GPCRs did not activate p110γ in neutrophils from mice lacking both the p101 and p84 regulatory subunits, indicating that RAS binding to p110γ is insufficient to support GPCR activation in this cell type. These findings define a direct role for Gβγ subunits in activating both of the endogenous PI3Kγ complexes and indicate that the regulatory PI3Kγ subunit biases activation toward different GPCRs.
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Affiliation(s)
- Natalie K Rynkiewicz
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Karen E Anderson
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Sabine Suire
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Daniel M Collins
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Eleftherios Karanasios
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Oscar Vadas
- MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Francis Crick Avenue, Cambridge CB2 0QH, UK
| | - Roger Williams
- MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Francis Crick Avenue, Cambridge CB2 0QH, UK
| | - David Oxley
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Jonathan Clark
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
| | - Len R Stephens
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK.
| | - Phillip T Hawkins
- Signalling Programme, Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK.
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Varied functions of immune checkpoints during cancer metastasis. Cancer Immunol Immunother 2020; 70:569-588. [PMID: 32902664 PMCID: PMC7907026 DOI: 10.1007/s00262-020-02717-2] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2020] [Accepted: 08/31/2020] [Indexed: 12/14/2022]
Abstract
Immune checkpoints comprise diverse receptors and ligands including costimulatory and inhibitory molecules, which play monumental roles in regulating the immune system. Immune checkpoints retain key potentials in maintaining the immune system homeostasis and hindering the malignancy development and autoimmunity. The expression of inhibitory immune checkpoints delineates an increase in a plethora of metastatic tumors and the inhibition of these immune checkpoints can be followed by promising results. On the other hand, the stimulation of costimulatory immune checkpoints can restrain the metastasis originating from diverse tumors. From the review above, key findings emerged regarding potential functions of inhibitory and costimulatory immune checkpoints targeting the metastatic cascade and point towards novel potential Achilles’ heels of cancer that might be exploited therapeutically in the future.
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10
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Shorning BY, Dass MS, Smalley MJ, Pearson HB. The PI3K-AKT-mTOR Pathway and Prostate Cancer: At the Crossroads of AR, MAPK, and WNT Signaling. Int J Mol Sci 2020; 21:E4507. [PMID: 32630372 PMCID: PMC7350257 DOI: 10.3390/ijms21124507] [Citation(s) in RCA: 374] [Impact Index Per Article: 74.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2020] [Revised: 06/22/2020] [Accepted: 06/22/2020] [Indexed: 12/13/2022] Open
Abstract
Oncogenic activation of the phosphatidylinositol-3-kinase (PI3K), protein kinase B (PKB/AKT), and mammalian target of rapamycin (mTOR) pathway is a frequent event in prostate cancer that facilitates tumor formation, disease progression and therapeutic resistance. Recent discoveries indicate that the complex crosstalk between the PI3K-AKT-mTOR pathway and multiple interacting cell signaling cascades can further promote prostate cancer progression and influence the sensitivity of prostate cancer cells to PI3K-AKT-mTOR-targeted therapies being explored in the clinic, as well as standard treatment approaches such as androgen-deprivation therapy (ADT). However, the full extent of the PI3K-AKT-mTOR signaling network during prostate tumorigenesis, invasive progression and disease recurrence remains to be determined. In this review, we outline the emerging diversity of the genetic alterations that lead to activated PI3K-AKT-mTOR signaling in prostate cancer, and discuss new mechanistic insights into the interplay between the PI3K-AKT-mTOR pathway and several key interacting oncogenic signaling cascades that can cooperate to facilitate prostate cancer growth and drug-resistance, specifically the androgen receptor (AR), mitogen-activated protein kinase (MAPK), and WNT signaling cascades. Ultimately, deepening our understanding of the broader PI3K-AKT-mTOR signaling network is crucial to aid patient stratification for PI3K-AKT-mTOR pathway-directed therapies, and to discover new therapeutic approaches for prostate cancer that improve patient outcome.
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Affiliation(s)
| | | | | | - Helen B. Pearson
- The European Cancer Stem Cell Research Institute, Cardiff University, Hadyn Ellis Building, Maindy Road, Cardiff CF24 4HQ, Wales, UK; (B.Y.S.); (M.S.D.); (M.J.S.)
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Ni H, Chai P, Yu J, Xing Y, Wang S, Fan J, Ge S, Wang Y, Jia R, Fan X. LncRNA CANT1 suppresses retinoblastoma progression by repellinghistone methyltransferase in PI3Kγ promoter. Cell Death Dis 2020; 11:306. [PMID: 32366932 PMCID: PMC7198571 DOI: 10.1038/s41419-020-2524-y] [Citation(s) in RCA: 28] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2020] [Revised: 04/12/2020] [Accepted: 04/14/2020] [Indexed: 02/07/2023]
Abstract
Retinoblastoma (RB) is the most common malignant intraocular tumor of childhood. Recent studies have shown that long noncoding RNAs (lncRNAs), which are longer than 200 bp and without protein-coding ability, are key regulators of tumorigenesis. However, the role of lncRNAs in retinoblastoma remains to be elucidated. In this study, we found that the expression of lncRNA CASC15-New-Transcript 1 (CANT1) was significantly downregulated in RB. Notably, overexpression of CANT1 significantly inhibited RB growth both in vitro and in vivo. Furthermore, lncRNA CANT1, which was mainly located in the nucleus, occupied the promoter of phosphoinositide 3-kinase gamma (PI3Kγ) and blocked histone methyltransferase hSET1 from binding to the PI3Kγ promoter, thus abolishing hSET1-mediated histone H3K4 trimethylation of the PI3Kγ promoter and inhibiting PI3Kγ expression. Furthermore, we found that silencing PI3Kγ either by lncRNA CANT1 overexpression or by PI3Kγ siRNA, reduced the activity of PI3K/Akt signaling and suppressed RB tumorigenesis. In summary, lncRNA CANT1 acts as a suppressor of RB progression by blocking gene-specific histone methyltransferase recruitment. These findings outline a new CANT1 modulation mechanism and provide an alternative option for the RB treatment.
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Affiliation(s)
- Hongyan Ni
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Peiwei Chai
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Jie Yu
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Yue Xing
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Shaoyun Wang
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Jiayan Fan
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Shengfang Ge
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011.,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011
| | - Yefei Wang
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011. .,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011.
| | - Renbing Jia
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011. .,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011.
| | - Xianqun Fan
- Department of Ophthalmology, Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai, China, 200011. .,Shanghai Key Laboratory of Orbital Diseases and Ocular Oncology, Shanghai, China, 200011.
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12
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Rathinaswamy MK, Burke JE. Class I phosphoinositide 3-kinase (PI3K) regulatory subunits and their roles in signaling and disease. Adv Biol Regul 2019; 75:100657. [PMID: 31611073 DOI: 10.1016/j.jbior.2019.100657] [Citation(s) in RCA: 76] [Impact Index Per Article: 12.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2019] [Revised: 09/23/2019] [Accepted: 09/25/2019] [Indexed: 02/06/2023]
Abstract
The Class I phosphoinositide 3-kinases (PI3Ks) are a group of heterodimeric lipid kinases that regulate crucial cellular processes including proliferation, survival, growth, and metabolism. The diversity in functions controlled by the various catalytic isoforms (p110α, p110β, p110δ, and p110γ) depends on their abilities to be activated by distinct stimuli such as receptor tyrosine kinases (RTKs), G-protein coupled receptors (GPCRs), and the Ras family of small G-proteins. A major factor determining the ability of each p110 enzyme to be activated is the presence of regulatory binding partners. Given the overwhelming evidence for the involvement of PI3Ks in diseases such as cancer, inflammation, immunodeficiency and diabetes, an understanding of how these regulatory proteins influence PI3K function is essential. This article highlights research deciphering the role of regulatory subunits in PI3K signaling and their involvement in human disease.
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Affiliation(s)
- Manoj K Rathinaswamy
- Department of Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, V8W 2Y2, Canada
| | - John E Burke
- Department of Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia, V8W 2Y2, Canada.
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13
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Function, Regulation and Biological Roles of PI3Kγ Variants. Biomolecules 2019; 9:biom9090427. [PMID: 31480354 PMCID: PMC6770443 DOI: 10.3390/biom9090427] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/23/2019] [Revised: 08/14/2019] [Accepted: 08/15/2019] [Indexed: 12/19/2022] Open
Abstract
Phosphatidylinositide 3-kinase (PI3K) γ is the only class IB PI3K member playing significant roles in the G-protein-dependent regulation of cell signaling in health and disease. Originally found in the immune system, increasing evidence suggest a wide array of functions in the whole organism. PI3Kγ occur as two different heterodimeric variants: PI3Kγ (p87) and PI3Kγ (p101), which share the same p110γ catalytic subunit but differ in their associated non-catalytic subunit. Here we concentrate on specific PI3Kγ features including its regulation and biological functions. In particular, the roles of its non-catalytic subunits serving as the main regulators determining specificity of class IB PI3Kγ enzymes are highlighted.
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14
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Buchanan CM, Lee KL, Shepherd PR. For Better or Worse: The Potential for Dose Limiting the On-Target Toxicity of PI 3-Kinase Inhibitors. Biomolecules 2019; 9:biom9090402. [PMID: 31443495 PMCID: PMC6770514 DOI: 10.3390/biom9090402] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2019] [Revised: 08/15/2019] [Accepted: 08/21/2019] [Indexed: 02/07/2023] Open
Abstract
The hyper-activation of the phosphoinositide (PI) 3-kinase signaling pathway is a hallmark of many cancers and overgrowth syndromes, and as a result, there has been intense interest in the development of drugs that target the various isoforms of PI 3-kinase. Given the key role PI 3-kinases play in many normal cell functions, there is significant potential for the disruption of essential cellular functions by PI 3-kinase inhibitors in normal tissues; so-called on-target drug toxicity. It is, therefore, no surprise that progress within the clinical development of PI 3-kinase inhibitors as single-agent anti-cancer therapies has been slowed by the difficulty of identifying a therapeutic window. The aim of this review is to place the cellular, tissue and whole-body effects of PI 3-kinase inhibition in the context of understanding the potential for dose limiting on-target toxicities and to introduce possible strategies to overcome these.
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Affiliation(s)
- Christina M Buchanan
- Department of Molecular Medicine and Pathology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand
- Maurice Wilkins Centre for Molecular Biodiscovery, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand
| | - Kate L Lee
- Department of Molecular Medicine and Pathology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand
- Maurice Wilkins Centre for Molecular Biodiscovery, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand
| | - Peter R Shepherd
- Department of Molecular Medicine and Pathology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand.
- Maurice Wilkins Centre for Molecular Biodiscovery, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand.
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15
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Erami Z, Heitz S, Bresnick AR, Backer JM. PI3Kβ links integrin activation and PI(3,4)P 2 production during invadopodial maturation. Mol Biol Cell 2019; 30:2367-2376. [PMID: 31318314 PMCID: PMC6741064 DOI: 10.1091/mbc.e19-03-0182] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2019] [Revised: 06/17/2019] [Accepted: 07/01/2019] [Indexed: 11/17/2022] Open
Abstract
The invasion of tumor cells from the primary tumor is mediated by invadopodia, actin-rich protrusive organelles that secrete matrix metalloproteases and degrade the extracellular matrix. This coupling between protrusive activity and matrix degradation facilitates tumor invasion. We previously reported that the PI3Kβ isoform of PI 3-kinase, which is regulated by both receptor tyrosine kinases and G protein-coupled receptors, is required for invasion and gelatin degradation in breast cancer cells. We have now defined the mechanism by which PI3Kβ regulates invadopodia. We find that PI3Kβ is specifically activated downstream from integrins, and is required for integrin-stimulated spreading and haptotaxis as well as integrin-stimulated invadopodia formation. Surprisingly, these integrin-stimulated and PI3Kβ-dependent responses require the production of PI(3,4)P2 by the phosphoinositide 5'-phosphatase SHIP2. Thus, integrin activation of PI3Kβ is coupled to the SHIP2-dependent production of PI(3,4)P2, which regulates the recruitment of PH domain-containing scaffolds such as lamellipodin to invadopodia. These findings provide novel mechanistic insight into the role of PI3Kβ in the regulation of invadopodia in breast cancer cells.
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Affiliation(s)
- Zahra Erami
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, NY 10461
| | - Samantha Heitz
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, NY 10461
| | - Anne R. Bresnick
- Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461
| | - Jonathan M. Backer
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, NY 10461
- Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461
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16
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Noorolyai S, Shajari N, Baghbani E, Sadreddini S, Baradaran B. The relation between PI3K/AKT signalling pathway and cancer. Gene 2019; 698:120-128. [PMID: 30849534 DOI: 10.1016/j.gene.2019.02.076] [Citation(s) in RCA: 393] [Impact Index Per Article: 65.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2018] [Revised: 02/08/2019] [Accepted: 02/17/2019] [Indexed: 12/19/2022]
Abstract
Phosphatidylinositol 3-kinases (PI3Ks) are crucial coordinators of intracellular signalling in response to the extracellular stimulators. Hyperactivation of PI3K signalling cascades is one among the most ordinary events in human cancers. Focusing on the PI3K pathway remains both a chance and a challenge for cancer therapy. The high recurrence of phosphoinositide 3-kinase (PI3K) pathway adjustments in cancer has led to a surge in the progression of PI3K inhibitors. Recent developments incorporate a re-assessment of the oncogenic mechanisms behind PI3K pathway modifications. Receptor tyrosine kinases upstream of PI3K, the p110a catalytic fractional unit of PI3K, the downstream kinase, AKT, and therefore the negative regulator, PTEN, are all often altered in cancer. In this review, we consider about the phosphoinositide 3-kinases family and mechanisms of PI3K-Akt stimulation in cancer.
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Affiliation(s)
- Saeed Noorolyai
- Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Neda Shajari
- Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Elham Baghbani
- Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Sanam Sadreddini
- Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Behzad Baradaran
- Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
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17
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Oo AKK, Calle AS, Nair N, Mahmud H, Vaidyanath A, Yamauchi J, Khayrani AC, Du J, Alam MJ, Seno A, Mizutani A, Murakami H, Iwasaki Y, Chen L, Kasai T, Seno M. Up-Regulation of PI 3-Kinases and the Activation of PI3K-Akt Signaling Pathway in Cancer Stem-Like Cells Through DNA Hypomethylation Mediated by the Cancer Microenvironment. Transl Oncol 2018; 11:653-663. [PMID: 29621663 PMCID: PMC6054593 DOI: 10.1016/j.tranon.2018.03.001] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2018] [Revised: 03/09/2018] [Accepted: 03/12/2018] [Indexed: 12/18/2022] Open
Abstract
Previously, we have succeeded in converting induced pluripotent stem cells (iPSCs) into cancer stem cells (CSCs) by treating the iPSCs with conditioned medium of Lewis lung carcinoma (LLC) cells. The converted CSCs, named miPS-LLCcm cells, exhibited the self-renewal, differentiation potential, and potential to form malignant tumors with metastasis. In this study, we further characterized miPS-LLCcm cells both in vivo and in vitro. The tumors formed by subcutaneous injection showed the structures with pathophysiological features consisting of undifferentiated and malignant phenotypes generally found in adenocarcinoma. Metastasis in the lung was also observed as nodule structures. Excising from the tumors, primary cultured cells from the tumor and the nodule showed self-renewal, differentiation potential as well as tumor forming ability, which are the essential characters of CSCs. We then characterized the epigenetic regulation occurring in the CSCs. By comparing the DNA methylation level of CG rich regions, the differentially methylated regions (DMRs) were evaluated in all stages of CSCs when compared with the parental iPSCs. In DMRs, hypomethylation was found superior to hypermethylation in the miPS-LLCcm cells and its derivatives. The hypo- and hypermethylated genes were used to nominate KEGG pathways related with CSC. As a result, several categories were defined in the KEGG pathways from which most related with cancers, significant and high expression of components was PI3K-AKT signaling pathway. Simultaneously, the AKT activation was also confirmed in the CSCs. The PI3K-Akt signaling pathway should be an important pathway for the CSCs established by the treatment with conditioned medium of LLC cells.
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Affiliation(s)
- Aung Ko Ko Oo
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan; Department of Biotechnology, Mandalay Technological University, Mandalay, Myanmar.
| | - Anna Sanchez Calle
- Division of Molecular and Cellular Medicine, National Cancer Center Research Institute, Tokyo 104-0045, Japan.
| | - Neha Nair
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Hafizah Mahmud
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Arun Vaidyanath
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Junya Yamauchi
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Aprilliana Cahya Khayrani
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Juan Du
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Md Jahangir Alam
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Akimasa Seno
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Akifumi Mizutani
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Hiroshi Murakami
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Yoshiaki Iwasaki
- Department of Gastroenterology and Hepatology, Graduate School of Medicine, Okayama University, Okayama 700-8558, Japan.
| | - Ling Chen
- Department of Pathology, Tianjin Central Hospital of Gynecology Obstetrics, Tianjin 300100, People's Republic of China.
| | - Tomonari Kasai
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
| | - Masaharu Seno
- Department of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan.
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18
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Ali AY, Wu X, Eissa N, Hou S, Ghia JE, Murooka TT, Banerji V, Johnston JB, Lin F, Gibson SB, Marshall AJ. Distinct roles for phosphoinositide 3-kinases γ and δ in malignant B cell migration. Leukemia 2018; 32:1958-1969. [PMID: 29479062 PMCID: PMC6127087 DOI: 10.1038/s41375-018-0012-5] [Citation(s) in RCA: 39] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2017] [Revised: 12/13/2017] [Accepted: 12/15/2017] [Indexed: 12/19/2022]
Abstract
The PI 3-kinases (PI3K) are essential mediators of chemokine receptor signaling necessary for migration of chronic lymphocytic leukemia (CLL) cells and their interaction with tissue-resident stromal cells. While the PI3Kδ-specific inhibitor idelalisib shows efficacy in treatment of CLL and other B cell malignancies, the function of PI3Kγ has not been extensively studied in B cells. Here, we assess whether PI3Kγ has non-redundant functions in CLL migration and adhesion to stromal cells. We observed that pharmaceutical PI3Kγ inhibition with CZC24832 significantly impaired CLL cell migration, while dual PI3Kδ/γ inhibitor duvelisib had a greater impact than single isoform-selective inhibitors. Knockdown of PI3Kγ reduced migration of CLL cells and cell lines. Expression of the PI3Kγ subunits increased in CLL cells in response to CD40L/IL-4, whereas BCR cross-linking had no effect. Overexpression of PI3Kγ subunits enhanced cell migration in response to SDF1α/CXCL12, with the strongest effect observed within ZAP70 + CLL samples. Microscopic tracking of cell migration within chemokine gradients revealed that PI3Kγ functions in gradient sensing and impacts cell morphology and F-actin polarization. PI3Kγ inhibition also reduced CLL adhesion to stromal cells to a similar extent as idelalisib. These findings provide the first evidence that PI3Kγ has unique functions in malignant B cells.
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Affiliation(s)
- Ahmed Y Ali
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada.,Research Institute in Oncology and Hematology, CancerCare Manitoba, 675 McDermot Ave., Winnipeg, MB, R3E 0V9, Canada
| | - Xun Wu
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada
| | - Nour Eissa
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada
| | - Sen Hou
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada
| | - Jean-Eric Ghia
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada.,Department of Internal Medicine, Section of Gastroenterology, University of Manitoba, 820 Sherbrooke St., Winnipeg, MB, R3A 1R9, Canada
| | - Thomas T Murooka
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada.,Department of Medical Microbiology and Infectious Diseases, University of Manitoba, 745 Bannatyne Ave., Winnipeg, MB, R3E 0J9, Canada
| | - Versha Banerji
- Research Institute in Oncology and Hematology, CancerCare Manitoba, 675 McDermot Ave., Winnipeg, MB, R3E 0V9, Canada.,Department of Biochemistry and Medical Genetics, University of Manitoba, 745 Bannatyne Ave., Winnipeg, MB, R3E 0J9, Canada
| | - James B Johnston
- Research Institute in Oncology and Hematology, CancerCare Manitoba, 675 McDermot Ave., Winnipeg, MB, R3E 0V9, Canada
| | - Francis Lin
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada.,Department of Physics and Astronomy, University of Manitoba, Allen Building, Winnipeg, MB, R3T 2N2, Canada
| | - Spencer B Gibson
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada.,Research Institute in Oncology and Hematology, CancerCare Manitoba, 675 McDermot Ave., Winnipeg, MB, R3E 0V9, Canada.,Department of Biochemistry and Medical Genetics, University of Manitoba, 745 Bannatyne Ave., Winnipeg, MB, R3E 0J9, Canada
| | - Aaron J Marshall
- Department of Immunology, University of Manitoba, 750 McDermot Avenue, Winnipeg, MB, R3E 0T5, Canada. .,Research Institute in Oncology and Hematology, CancerCare Manitoba, 675 McDermot Ave., Winnipeg, MB, R3E 0V9, Canada. .,Department of Biochemistry and Medical Genetics, University of Manitoba, 745 Bannatyne Ave., Winnipeg, MB, R3E 0J9, Canada.
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19
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Rac1-stimulated macropinocytosis enhances Gβγ activation of PI3Kβ. Biochem J 2017; 474:3903-3914. [PMID: 29046393 DOI: 10.1042/bcj20170279] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2017] [Revised: 10/05/2017] [Accepted: 10/10/2017] [Indexed: 12/30/2022]
Abstract
Phosphoinositide 3-kinases (PI 3-kinases) are regulated by a diverse range of upstream activators, including receptor tyrosine kinases (RTKs), G-protein-coupled receptors (GPCRs), and small GTPases from the Ras, Rho and Rab families. For the Class IA PI 3-kinase PI3Kβ, two mechanisms for GPCR-mediated regulation have been described: direct binding of Gβγ subunits to the C2-helical domain linker of p110β, and Dock180/Elmo1-mediated activation of Rac1, which binds to the Ras-Binding Domain of p110β. We now show that the integration of these dual pathways is unexpectedly complex. In breast cancer cells, expression of constitutively activated Rac1 (CA-Rac1) along with either GPCR stimulation or expression of Gβγ led to an additive PI3Kβ-dependent activation of Akt. Whereas CA-Rac1-mediated activation of Akt was blocked in cells expressing a mutated PI3Kβ that cannot bind Gβγ, Gβγ and GPCR-mediated activation of Akt was preserved when Rac1 binding to PI3Kβ was blocked. Surprisingly, PI3Kβ-dependent CA-Rac1 signaling to Akt was still seen in cells expressing a mutant p110β that cannot bind Rac1. Instead of directly binding to PI3Kβ, CA-Rac1 acts by enhancing Gβγ coupling to PI3Kβ, as CA-Rac1-mediated Akt activation was blocked by inhibitors of Gβγ. Cells expressing CA-Rac1 exhibited a robust induction of macropinocytosis, and inhibitors of macropinocytosis blocked the activation of Akt by CA-Rac1 or lysophosphatidic acid. Our data suggest that Rac1 can potentiate the activation of PI3Kβ by GPCRs through an indirect mechanism, by driving the formation of macropinosomes that serve as signaling platforms for Gβγ coupling to PI3Kβ.
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20
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Scaffolding Function of PI3Kgamma Emerges from Enzyme's Shadow. J Mol Biol 2017; 429:763-772. [PMID: 28179187 DOI: 10.1016/j.jmb.2017.01.023] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2016] [Revised: 01/12/2017] [Accepted: 01/31/2017] [Indexed: 11/20/2022]
Abstract
Traditionally, an enzyme is a protein that mediates biochemical action by binding to the substrate and by catalyzing the reaction that translates external cues into biological responses. Sequential dissemination of information from one enzyme to another facilitates signal transduction in biological systems providing for feed-forward and feed-back mechanisms. Given this viewpoint, an enzyme without its catalytic activity is generally considered to be an inert organizational protein without catalytic function and has classically been termed as pseudo-enzymes. However, pseudo-enzymes still have biological function albeit non-enzymatic like serving as a chaperone protein or an interactive platform between proteins. In this regard, majority of the studies have focused solely on the catalytic role of enzymes in biological function, overlooking the potentially critical non-enzymatic roles. Increasing evidence from recent studies implicate that the scaffolding function of enzymes could be as important in signal transduction as its catalytic activity, which is an antithesis to the definition of enzymes. Recognition of non-enzymatic functions could be critical, as these unappreciated roles may hold clues to the ineffectiveness of kinase inhibitors in pathology, which is characteristically associated with increased enzyme expression. Using an established enzyme phosphoinositide 3-kinase γ, we discuss the insights obtained from the scaffolding function and how this non-canonical role could contribute to/alter the outcomes in pathology like cancer and heart failure. Also, we hope that with this review, we provide a forum and a starting point to discuss the idea that catalytic function alone may not account for all the actions observed with increased expression of the enzyme.
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21
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Vázquez-Prado J, Bracho-Valdés I, Cervantes-Villagrana RD, Reyes-Cruz G. Gβγ Pathways in Cell Polarity and Migration Linked to Oncogenic GPCR Signaling: Potential Relevance in Tumor Microenvironment. Mol Pharmacol 2016; 90:573-586. [PMID: 27638873 DOI: 10.1124/mol.116.105338] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2016] [Accepted: 09/14/2016] [Indexed: 02/14/2025] Open
Abstract
Cancer cells and stroma cells in tumors secrete chemotactic agonists that exacerbate invasive behavior, promote tumor-induced angiogenesis, and recruit protumoral bone marrow-derived cells. In response to shallow gradients of chemotactic stimuli recognized by G protein-coupled receptors (GPCRs), Gβγ-dependent signaling cascades contribute to specifying the spatiotemporal assembly of cytoskeletal structures that can dynamically alter cell morphology. This sophisticated process is intrinsically linked to the activation of Rho GTPases and their cytoskeletal-remodeling effectors. Thus, Rho guanine nucleotide exchange factors, the activators of these molecular switches, and their upstream signaling partners are considered participants of tumor progression. Specifically, phosphoinositide-3 kinases (class I PI3Ks, β and γ) and P-Rex1, a Rac-specific guanine nucleotide exchange factor, are fundamental Gβγ effectors in the pathways controlling directionally persistent motility. In addition, GPCR-dependent chemotactic responses often involve endosomal trafficking of signaling proteins; coincidently, endosomes serve as signaling platforms for Gβγ In preclinical murine models of cancer, inhibition of Gβγ attenuates tumor growth, whereas in cancer patients, aberrant overexpression of chemotactic Gβγ effectors and recently identified mutations in Gβ correlate with poor clinical outcome. Here we discuss emerging paradigms of Gβγ signaling in cancer, which are essential for chemotactic cell migration and represent novel opportunities to develop pathway-specific pharmacologic treatments.
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Affiliation(s)
- José Vázquez-Prado
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
| | - Ismael Bracho-Valdés
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
| | - Rodolfo Daniel Cervantes-Villagrana
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
| | - Guadalupe Reyes-Cruz
- Departments of Pharmacology (J.V.-P., R.D.C.-V.) and Cell Biology (G.R.-C.). CINVESTAV-IPN, Mexico City, and Department of Pharmacology (I.B.-V.), School of Medicine, UABC, Mexicali, B.C., Mexico
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22
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Bai H, Li H, Li W, Gui T, Yang J, Cao D, Shen K. The PI3K/AKT/mTOR pathway is a potential predictor of distinct invasive and migratory capacities in human ovarian cancer cell lines. Oncotarget 2016; 6:25520-32. [PMID: 26267321 PMCID: PMC4694849 DOI: 10.18632/oncotarget.4550] [Citation(s) in RCA: 76] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2015] [Accepted: 07/04/2015] [Indexed: 12/21/2022] Open
Abstract
Objectives To explore the genetic and molecular events that control subclones exhibiting distinct invasive/migratory capacities derived from human epithelial ovarian cancer (EOC) cell line A2780 and SKOV3. Methods Single-cell subclones were isolated and established that were derived from the SKOV3 and A2780 cell lines through limiting dilution methodology. Transwell insert assays and MTT assays were performed to screen and identify the subclones exhibiting the highest and the lowest invasive/migratory capacities, and the selected subclones were renamed as A-H (A2780 high), A-L (A2780 low), S-H (SKOV3 high), and S-L (SKOV3 low). Their biological characteristics were evaluated. RNA-Seq was conducted on the targeted subclones. Results Compared with their corresponding counterparts, A-H/S-H cells exhibited significantly higher invasive/migratory capacities (P < 0.001 and = 0.001, respectively). A-H/S-H cells displayed a clear reduction in doubling time (P = 0.004 and 0.001, respectively), and a significant increase in the percentage of cells in S phase (P = 0.004 and 0.022, respectively). Additionally, the apoptotic rates of A-H/S-H cells were significantly lower than those of A-L/S-L cells (P = 0.002 and 0.026, respectively). At both mRNA and protein levels, caspase-3 and caspase-7 expression were reduced but Bcl-2 expression was increased in A-H/S-H cells. The TrkB (anoikis-related) and Beclin1 (autophagy-related) levels were consistently high and low, respectively, in both A-H/S-H cells. Resistance to chemotherapy in vitro and higher capacities on tumor formation in vivo was presented in both A-H/S-H cells. PI3K/AKT/mTOR pathway components, PIK3CA, PIK3CD, AKT3, ECM1, GPCR, mTOR and PRKCB were increased but that the Nur77 and PTEN were decreased in A-H/S-H cells, identified by RNA-Seq and consistently confirmed by RT-PCR and Western blot analyses. Conclusions Heterogeneous cell subpopulations exhibiting distinct invasive and migratory capacities co-exist within the SKOV3 and A2780 cell lines. PI3K/AKT/mTOR pathway activation is associated with higher invasive and migratory capacities in subpopulations of human ovarian cancer cell lines. Inhibiting this pathway may be useful for the chemoprevention or treatment of EOC.
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Affiliation(s)
- Huimin Bai
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China.,Department of Obstetrics and Gynecology, Beijing Chao-Yang Hospital, China Capital Medical University, Beijing China
| | - Haixia Li
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China
| | - Weihua Li
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China
| | - Ting Gui
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China
| | - Jiaxin Yang
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China
| | - Dongyan Cao
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China
| | - Keng Shen
- Department of Obstetrics and Gynecology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing China
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23
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Diao HY, Shao JG, Bian ZL, Chen L, Ju LL, Zhang Y. Role of phosphoinositide-3 kinase signaling pathways in pathogenesis of acute pancreatitis. Shijie Huaren Xiaohua Zazhi 2016; 24:3002-3008. [DOI: 10.11569/wcjd.v24.i19.3002] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Acute pancreatitis (AP) as a common acute disease poses a great threat to people's health. According to statistics, about one-fifth of cases develop acute respiratory distress syndrome and multiple organ dysfunction, which result in high mortality. The early understanding of the pathogenesis of this disease is limited to an inflammatory response resulting in autodigestion, edema, hemorrhage and necrosis of pancreatic tissue after the abnormal activation of trypsin. In recent years, researchers have focused their research on the role of immune inflammatory response in the pathogenesis of AP. Here we discuss the relationship between the immune inflammation and PI3K signaling pathways in AP.
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24
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Khalil BD, Hsueh C, Cao Y, Abi Saab WF, Wang Y, Condeelis JS, Bresnick AR, Backer JM. GPCR Signaling Mediates Tumor Metastasis via PI3Kβ. Cancer Res 2016; 76:2944-53. [PMID: 27013201 DOI: 10.1158/0008-5472.can-15-1675] [Citation(s) in RCA: 45] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2015] [Accepted: 02/23/2016] [Indexed: 12/16/2022]
Abstract
Inappropriate activation of PI3K signaling has been implicated strongly in human cancer. Although studies on the role of PI3K signaling in breast tumorigenesis and progression have focused most intensively on PI3Kα, a role for PI3Kβ has begun to emerge. The PI3Kβ isoform is unique among class IA PI3K enzymes in that it is activated by both receptor tyrosine kinases and G-protein-coupled receptors (GPCR). In previous work, we identified a mutation that specifically abolishes PI3Kβ binding to Gβγ (p110(526KK-DD)). Expression of this mutant in p110β-silenced breast cancer cells inhibits multiple steps of the metastatic cascade in vitro and in vivo and causes a cell autonomous defect in invadopodial matrix degradation. Our results identify a novel link between GPCRs and PI3Kβ in mediating metastasis, suggesting that disruption of this link might offer a novel therapeutic target to prevent the development of metastatic disease. Cancer Res; 76(10); 2944-53. ©2016 AACR.
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Affiliation(s)
- Bassem D Khalil
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York
| | - Christine Hsueh
- Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York
| | - Yanyan Cao
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York
| | - Widian F Abi Saab
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York
| | - Yarong Wang
- Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York
| | - John S Condeelis
- Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York
| | - Anne R Bresnick
- Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York.
| | - Jonathan M Backer
- Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York. Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York.
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25
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Different inhibition of Gβγ-stimulated class IB phosphoinositide 3-kinase (PI3K) variants by a monoclonal antibody. Specific function of p101 as a Gβγ-dependent regulator of PI3Kγ enzymatic activity. Biochem J 2015; 469:59-69. [PMID: 26173259 DOI: 10.1042/bj20150099] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
Class IB phosphoinositide 3-kinases γ (PI3Kγ) are second-messenger-generating enzymes downstream of signalling cascades triggered by G-protein-coupled receptors (GPCRs). PI3Kγ variants have one catalytic p110γ subunit that can form two different heterodimers by binding to one of a pair of non-catalytic subunits, p87 or p101. Growing experimental data argue for a different regulation of p87-p110γ and p101-p110γ allowing integration into distinct signalling pathways. Pharmacological tools enabling distinct modulation of the two variants are missing. The ability of an anti-p110γ monoclonal antibody [mAb(A)p110γ] to block PI3Kγ enzymatic activity attracted us to characterize this tool in detail using purified proteins. In order to get insight into the antibody-p110γ interface, hydrogen-deuterium exchange coupled to MS (HDX-MS) measurements were performed demonstrating binding of the monoclonal antibody to the C2 domain in p110γ, which was accompanied by conformational changes in the helical domain harbouring the Gβγ-binding site. We then studied the modulation of phospholipid vesicles association of PI3Kγ by the antibody. p87-p110γ showed a significantly reduced Gβγ-mediated phospholipid recruitment as compared with p101-p110γ. Concomitantly, in the presence of mAb(A)p110γ, Gβγ did not bind to p87-p110γ. These data correlated with the ability of the antibody to block Gβγ-stimulated lipid kinase activity of p87-p110γ 30-fold more potently than p101-p110γ. Our data argue for differential regulatory functions of the non-catalytic subunits and a specific Gβγ-dependent regulation of p101 in PI3Kγ activation. In this scenario, we consider the antibody as a valuable tool to dissect the distinct roles of the two PI3Kγ variants downstream of GPCRs.
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26
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Chen S, Li F, Chai H, Tao X, Wang H, Ji A. miR-502 inhibits cell proliferation and tumor growth in hepatocellular carcinoma through suppressing phosphoinositide 3-kinase catalytic subunit gamma. Biochem Biophys Res Commun 2015; 464:500-5. [PMID: 26163264 DOI: 10.1016/j.bbrc.2015.06.168] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2015] [Accepted: 06/30/2015] [Indexed: 01/06/2023]
Abstract
MicroRNAs (miRNAs) play a key role in carcinogenesis and tumor progression in hepatocellular carcinoma (HCC). In the present study, we demonstrated that miR-502 significantly inhibits HCC cell proliferation in vitro and tumor growth in vivo. G1/S cell cycle arrest and apoptosis of HCC cells were induced by miR-502. Phosphoinositide 3-kinase catalytic subunit gamma (PIK3CG) was identified as a direct downstream target of miR-502 in HCC cells. Notably, overexpression of PIK3CG reversed the inhibitory effects of miR-502 in HCC cells. Our findings suggest that miR-502 functions as a tumor suppressor in HCC via inhibition of PI3KCG, supporting its utility as a promising therapeutic gene target for this tumor type.
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Affiliation(s)
- Suling Chen
- Department of Infectious Disease, Heping Hospital Attached to Changzhi Medical College, Changzhi 046000, China.
| | - Fang Li
- Department of Infectious Disease, Heping Hospital Attached to Changzhi Medical College, Changzhi 046000, China
| | - Haiyun Chai
- Department of Infectious Disease, Heping Hospital Attached to Changzhi Medical College, Changzhi 046000, China
| | - Xin Tao
- Department of Infectious Disease, Heping Hospital Attached to Changzhi Medical College, Changzhi 046000, China
| | - Haili Wang
- Department of Hematology, Heping Hospital Attached to Changzhi Medical College, Changzhi 046000, China
| | - Aifang Ji
- Central Laboratory, Heping Hospital Attached to Changzhi Medical College, Changzhi 046000, China
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27
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Stat1 stimulates cap-independent mRNA translation to inhibit cell proliferation and promote survival in response to antitumor drugs. Proc Natl Acad Sci U S A 2015; 112:E2149-55. [PMID: 25870277 DOI: 10.1073/pnas.1420671112] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
The signal transducer and activator of transcription 1 (Stat1) functions as a tumor suppressor via immune regulatory and cell-autonomous pathways. Herein, we report a previously unidentified cell-autonomous Stat1 function, which is its ability to exhibit both antiproliferative and prosurvival properties by facilitating translation of mRNAs encoding for the cyclin-dependent kinase inhibitor p27(Kip1) and antiapoptotic proteins X-linked inhibitor of apoptosis and B-cell lymphoma xl. Translation of the select mRNAs requires the transcriptional function of Stat1, resulting in the up-regulation of the p110γ subunit of phosphoinositide 3-kinase (PI3K) class IB and increased expression of the translational repressor translation initiation factor 4E (eIF4E)-binding protein 1 (4EBP1). Increased PI3Kγ signaling promotes the degradation of the eIF4A inhibitor programmed cell death protein 4, which favors the cap-independent translation of the select mRNAs under conditions of general inhibition of protein synthesis by up-regulated eIF4E-binding protein 1. As such, Stat1 inhibits cell proliferation but also renders cells increasingly resistant to antiproliferative effects of pharmacological inhibitors of PI3K and/or mammalian target of rapamycin. Stat1 also protects Ras-transformed cells from the genotoxic effects of doxorubicin in culture and immune-deficient mice. Our findings demonstrate an important role of mRNA translation in the cell-autonomous Stat1 functions, with implications in tumor growth and treatment with chemotherapeutic drugs.
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28
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Zhang S, Chung WC, Wu G, Egan SE, Miele L, Xu K. Manic fringe promotes a claudin-low breast cancer phenotype through notch-mediated PIK3CG induction. Cancer Res 2015; 75:1936-43. [PMID: 25808869 DOI: 10.1158/0008-5472.can-14-3303] [Citation(s) in RCA: 55] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2014] [Accepted: 02/25/2015] [Indexed: 12/13/2022]
Abstract
Claudin-low breast cancer (CLBC) is a poor prognosis disease biologically characterized by stemness and mesenchymal features. These tumors disproportionately affect younger patients and women with African ancestry, causing significant morbidity and mortality, and no effective targeted therapy exists at present. CLBC is thought to originate from mammary stem cells, but little is known on how or why these tumors express a stable epithelial-to-mesenchymal transition phenotype, or what are the driving forces of this disease. Here, we report that Manic Fringe (Mfng), which encodes an O-fucosylpeptide 3-β-N-acetylglucosaminyltransferase known to modify EGF repeats in the Notch extracellular domain, is highly expressed in CLBC and functions as an oncogene in this context. We show that Mfng modulates Notch activation in human and mouse CLBC cell lines, as well as in mouse mammary gland. Mfng silencing in CLBC cell lines reduced cell migration, tumorsphere formation, and in vivo tumorigenicity associated with a decrease in the stem-like cell population. Mfng deletion in the Lfng(flox/flox);MMTV-Cre mouse model, in which one-third of mammary tumors resemble human CLBC, caused a tumor subtype shift away from CLBC. We identified the phosphoinositide kinase Pik3cg as a direct transcriptional target of Mfng-facilitated RBPJκ-dependent Notch signaling. Indeed, pharmacologic inhibition of PI3Kγ in CLBC cell lines blocked migration and tumorsphere formation. Taken together, our results define Mfng as an oncogene acting through Notch-mediated induction of Pik3cg. Furthermore, they suggest that targeting PI3Kγ may prove beneficial for the treatment of CLBC subtype.
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Affiliation(s)
- Shubing Zhang
- Cancer Institute, University of Mississippi Medical Center, Jackson, Mississippi. State Key Laboratory of Medical Genetics and School of Life Sciences, Central South University, Changsha, Hunan, China
| | - Wen-Cheng Chung
- Cancer Institute, University of Mississippi Medical Center, Jackson, Mississippi
| | - Guanming Wu
- Department of Medical Informatics and Clinical Epidemiology and Oregon Clinical and Translational Research Institute, Oregon Health and Science University, Portland, Oregon
| | - Sean E Egan
- Program in Developmental and Stem Cell Biology, The Hospital for Sick Children and Department of Molecular Genetics, University of Toronto, Toronto, Canada
| | - Lucio Miele
- Stanley S. Scott Cancer Center, Louisiana State University Health Sciences Center and Louisiana Cancer Research Center, New Orleans, Louisiana
| | - Keli Xu
- Cancer Institute, University of Mississippi Medical Center, Jackson, Mississippi. Department of Neurobiology and Anatomical Sciences, University of Mississippi Medical Center, Jackson, Mississippi.
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29
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Turvey ME, Klingler-Hoffmann M, Hoffmann P, McColl SR. p84 forms a negative regulatory complex with p110γ to control PI3Kγ signalling during cell migration. Immunol Cell Biol 2015; 93:735-43. [PMID: 25753393 DOI: 10.1038/icb.2015.35] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2014] [Revised: 02/23/2015] [Accepted: 02/23/2015] [Indexed: 02/05/2023]
Abstract
Phosphoinositide 3-kinase γ (PI3Kγ) consists of the catalytic subunit p110γ that forms a mutually exclusive heterodimer with one of the two adaptor subunits, p101 or p84. Although activation of PI3Kγ is necessary for cell migration downstream of G-protein-coupled receptor engagement, particularly within the immune system, aberrant PI3Kγ signalling has been associated with transformation, increased migration and the progression of multiple cancer types. Regulation of PI3Kγ signal activation and duration is critical to controlling and maintaining coordinated cellular migration; however, the mechanistic basis for this is not well understood. We have recently demonstrated that, in contrast to the tumour-promoting potential of p110γ and p101, p84 possesses tumour-suppressor activity, suggesting a negative regulatory role within PI3Kγ signalling. The present study investigated the role of p84 phosphorylation in the context of PI3Kγ signalling, cell migration and p84-mediated tumour suppression. Two putative phosphorylation sites were characterised within p84, Ser358 and Thr607. Expression of wild-type p84 reduced the oncogenic potential of MDA.MB.231 cells and inhibited metastatic lung colonisation in vivo, effects that were dependent on Thr607. Furthermore, loss of Thr607 enhanced migration of MDA.MB.231 cells in vitro and prevented the induction of p84/p110γ dimers. The dimerisation of wild-type p84 with p110γ was not detected at the plasma membrane, indicating an inhibitory interaction preventing PI3Kγ lipid-kinase activity. In contrast, Ser358 phosphorylation was not determined to be critical for p84 activity in the context of migration. Our findings suggest that p84 binding to p110γ may represent a novel negative feedback signal that terminates PI3Kγ activity.
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Affiliation(s)
- Michelle E Turvey
- Chemokine Biology Laboratory, School of Molecular and Biomedical Sciences, The University of Adelaide and Centre for Molecular Pathology, Adelaide, South Australia, Australia
| | | | - Peter Hoffmann
- Adelaide Proteomics Centre, The University of Adelaide, Adelaide, South Australia, Australia
| | - Shaun R McColl
- Chemokine Biology Laboratory, School of Molecular and Biomedical Sciences, The University of Adelaide and Centre for Molecular Pathology, Adelaide, South Australia, Australia
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30
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Deladeriere A, Gambardella L, Pan D, Anderson KE, Hawkins PT, Stephens LR. The regulatory subunits of PI3Kγ control distinct neutrophil responses. Sci Signal 2015; 8:ra8. [DOI: 10.1126/scisignal.2005564] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/02/2022]
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31
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Burke JE, Williams RL. Synergy in activating class I PI3Ks. Trends Biochem Sci 2015; 40:88-100. [PMID: 25573003 DOI: 10.1016/j.tibs.2014.12.003] [Citation(s) in RCA: 161] [Impact Index Per Article: 16.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2014] [Revised: 12/03/2014] [Accepted: 12/05/2014] [Indexed: 12/20/2022]
Abstract
The class I phosphoinositide 3-kinases (PI3Ks) are lipid kinases that transduce a host of cellular signals and regulate a broad range of essential functions including growth, proliferation, and migration. As such, PI3Ks have pivotal roles in diseases such as cancer, diabetes, primary immune disorders, and inflammation. These enzymes are activated downstream of numerous activating stimuli including receptor tyrosine kinases, G protein-coupled receptors (GPCRs), and the Ras superfamily of small G proteins. A major challenge is to decipher how each PI3K isoform is able to successfully synergize these inputs into their intended signaling function. This article highlights recent progress in characterizing the molecular mechanisms of PI3K isoform-specific activation pathways, as well as novel roles for PI3Ks in human diseases, specifically cancer and immune diseases.
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Affiliation(s)
- John E Burke
- Department of Biochemistry and Microbiology, University of Victoria, 3800 Finnerty Drive, Victoria BC, V8P 5C2, Canada.
| | - Roger L Williams
- Medical Research Council (MRC) Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge, CB2 0QH, UK
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32
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Lupia E, Pigozzi L, Goffi A, Hirsch E, Montrucchio G. Role of phosphoinositide 3-kinase in the pathogenesis of acute pancreatitis. World J Gastroenterol 2014; 20:15190-15199. [PMID: 25386068 PMCID: PMC4223253 DOI: 10.3748/wjg.v20.i41.15190] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/01/2014] [Revised: 06/12/2014] [Accepted: 07/22/2014] [Indexed: 02/06/2023] Open
Abstract
A large body of experimental and clinical data supports the notion that inflammation in acute pancreatitis has a crucial role in the pathogenesis of local and systemic damage and is a major determinant of clinical severity. Thus, research has recently focused on molecules that can regulate the inflammatory processes, such as phosphoinositide 3-kinases (PI3Ks), a family of lipid and protein kinases involved in intracellular signal transduction. Studies using genetic ablation or pharmacologic inhibitors of different PI3K isoforms, in particular the class I PI3Kδ and PI3Kγ, have contributed to a greater understanding of the roles of these kinases in the modulation of inflammatory and immune responses. Recent data suggest that PI3Ks are also involved in the pathogenesis of acute pancreatitis. Activation of the PI3K signaling pathway, and in particular of the class IB PI3Kγ isoform, has a significant role in those events which are necessary for the initiation of acute pancreatic injury, namely calcium signaling alteration, trypsinogen activation, and nuclear factor-κB transcription. Moreover, PI3Kγ is instrumental in modulating acinar cell apoptosis, and regulating local neutrophil infiltration and systemic inflammatory responses during the course of experimental acute pancreatitis. The availability of PI3K inhibitors selective for specific isoforms may provide new valuable therapeutic strategies to improve the clinical course of this disease. This article presents a brief summary of PI3K structure and function, and highlights recent advances that implicate PI3Ks in the pathogenesis of acute pancreatitis.
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33
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Falasca M, Maffucci T. Targeting p110gamma in gastrointestinal cancers: attack on multiple fronts. Front Physiol 2014; 5:391. [PMID: 25360116 PMCID: PMC4197894 DOI: 10.3389/fphys.2014.00391] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2014] [Accepted: 09/21/2014] [Indexed: 12/12/2022] Open
Abstract
Phosphoinositide 3-kinases (PI3Ks) regulate several cellular functions that are critical for cancer progression and development, including cell survival, proliferation and migration. Three classes of PI3Ks exist with the class I PI3K encompassing four isoforms of the catalytic subunit known as p110α, p110β, p110γ, and p110δ. Although for many years attention has been mainly focused on p110α recent evidence supports the conclusion that p110β, p110γ, and p110δ can also have a role in cancer. Amongst these, accumulating evidence now indicates that p110γ is involved in several cellular processes associated with cancer and indeed this specific isoform has emerged as a novel important player in cancer progression. Studies from our laboratory have identified a specific overexpression of p110γ in human pancreatic ductal adenocarcinoma (PDAC) and in hepatocellular carcinoma (HCC) tissues compared to their normal counterparts. Our data have further established that selective inhibition of p110γ is able to block PDAC and HCC cell proliferation, strongly suggesting that pharmacological inhibition of this enzyme can directly affect growth of these tumors. Furthermore, increasing evidence suggests that p110γ plays also a key role in the interactions between cancer cells and tumor microenvironment and in particular in tumor-associated immune response. It has also been reported that p110γ can regulate invasion of myeloid cells into tumors and tumor angiogenesis. Finally p110γ has also been directly involved in regulation of cancer cell migration. Taken together these data indicate that p110γ plays multiple roles in regulation of several processes that are critical for tumor progression and metastasis. This review will discuss the role of p110γ in gastrointestinal tumor development and progression and how targeting this enzyme might represent a way to target very aggressive tumors such as pancreatic and liver cancer on multiple fronts.
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Affiliation(s)
- Marco Falasca
- Inositide Signalling Group, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London London, UK
| | - Tania Maffucci
- Inositide Signalling Group, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London London, UK
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The atypical chemokine receptor CCX-CKR regulates metastasis of mammary carcinoma via an effect on EMT. Immunol Cell Biol 2014; 92:815-24. [PMID: 25027038 DOI: 10.1038/icb.2014.58] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2013] [Revised: 05/16/2014] [Accepted: 06/06/2014] [Indexed: 01/21/2023]
Abstract
Over the last decade, the significance of the homeostatic CC chemokine receptor-7 and its ligands CC chemokine ligand-19 (CCL19) and CCL21, in various types of cancer, particularly mammary carcinoma, has been highlighted. The chemokine receptor CCX-CKR is a high-affinity receptor for these chemokine ligands but rather than inducing classical downstream signalling events promoting migration, it instead sequesters and targets its ligands for degradation, and appears to function as a regulator of the bioavailability of these chemokines in vivo. Therefore, in this study, we tested the hypothesis that local regulation of chemokine levels by CCX-CKR expressed on tumours alters tumour growth and metastasis in vivo. Expression of CCX-CKR on 4T1.2 mouse mammary carcinoma cells inhibited orthotopic tumour growth. However, this effect could not be correlated with chemokine scavenging in vivo and was not mediated by host adaptive immunity. Conversely, expression of CCX-CKR on 4T1.2 cells resulted in enhanced spontaneous metastasis and haematogenous metastasis in vivo. In vitro characterisation of the tumourigenicity of CCX-CKR-expressing 4T1.2 cells suggested accelerated epithelial-mesenchymal transition (EMT) revealed by their more invasive and motile character, lower adherence to the extracellular matrix and to each other, and greater resistance to anoikis. Further analysis of CCX-CKR-expressing 4T1.2 cells also revealed that transforming growth factor (TGF)-β1 expression was increased both at mRNA and protein levels leading to enhanced autocrine phosphorylation of Smad 2/3 in these cells. Together, our data show a novel function for the chemokine receptor CCX-CKR as a regulator of TGF-β1 expression and the EMT in breast cancer cells.
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35
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Edros R, McDonnell S, Al-Rubeai M. The relationship between mTOR signalling pathway and recombinant antibody productivity in CHO cell lines. BMC Biotechnol 2014; 14:15. [PMID: 24533650 PMCID: PMC3937030 DOI: 10.1186/1472-6750-14-15] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2014] [Accepted: 02/05/2014] [Indexed: 11/20/2022] Open
Abstract
Background High recombinant protein productivity in mammalian cell lines is often associated with phenotypic changes in protein content, energy metabolism, and cell growth, but the key determinants that regulate productivity are still not clearly understood. The mammalian target of rapamycin (mTOR) signalling pathway has emerged as a central regulator for many cellular processes including cell growth, apoptosis, metabolism, and protein synthesis. This role of this pathway changes in response to diverse environmental cues and allows the upstream proteins that respond directly to extracellular signals (such as nutrient availability, energy status, and physical stresses) to communicate with downstream effectors which, in turn, regulate various essential cellular processes. Results In this study, we have performed a transcriptomic analysis using a pathway-focused polymerase chain reaction (PCR) array to compare the expression of 84 target genes related to the mTOR signalling in two recombinant CHO cell lines with a 17.4-fold difference in specific monoclonal antibody productivity (qp). Eight differentially expressed genes that exhibited more than a 1.5-fold change were identified. Pik3cd (encoding the Class 1A catalytic subunit of phosphatidylinositol 3-kinase [PI3K]) was the most differentially expressed gene having a 71.3-fold higher level of expression in the high producer cell line than in the low producer. The difference in the gene’s transcription levels was confirmed at the protein level by examining expression of p110δ. Conclusion Expression of p110δ correlated with specific productivity (qp) across six different CHO cell lines, with a range of expression levels from 3 to 51 pg/cell/day, suggesting that p110δ may be a key factor in regulating productivity in recombinant cell lines.
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Affiliation(s)
| | | | - Mohamed Al-Rubeai
- School of Chemical and Bioprocess Engineering and Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Dublin, Ireland.
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Edling CE, Selvaggi F, Ghonaim R, Maffucci T, Falasca M. Caffeine and the analog CGS 15943 inhibit cancer cell growth by targeting the phosphoinositide 3-kinase/Akt pathway. Cancer Biol Ther 2014; 15:524-32. [PMID: 24521981 DOI: 10.4161/cbt.28018] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Caffeine is a naturally occurring methylxanthine that acts as a non-selective adenosine receptor antagonist. Epidemiological studies demonstrated habitual coffee drinking to be significantly associated with liver cancer survival. We aimed to investigate the effects of caffeine and its analog CGS 15943 on hepatocellular carcinoma (HCC) and pancreatic cancer adenocarcinoma (PDAC). We demonstrate that caffeine and CGS 15943 block proliferation in HCC and PDAC cell lines by inhibiting the PI3K/Akt pathway. Importantly a kinase profiling assay reveals that CGS 15943 targets specifically the catalytic subunit of the class IB PI3K isoform (p110γ). These data give mechanistic insight into the action of caffeine and its analogs and they identify these compounds as promising lead compounds to develop drugs that can specifically target this PI3K isoform whose key role in cancer progression is emerging.
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Affiliation(s)
- Charlotte E Edling
- Queen Mary University of London; Blizard Institute; Barts and The London School of Medicine and Dentistry; Inositide Signalling Group; London, UK
| | - Federico Selvaggi
- Queen Mary University of London; Blizard Institute; Barts and The London School of Medicine and Dentistry; Inositide Signalling Group; London, UK
| | - Ragheda Ghonaim
- Queen Mary University of London; Blizard Institute; Barts and The London School of Medicine and Dentistry; Inositide Signalling Group; London, UK
| | - Tania Maffucci
- Queen Mary University of London; Blizard Institute; Barts and The London School of Medicine and Dentistry; Inositide Signalling Group; London, UK
| | - Marco Falasca
- Queen Mary University of London; Blizard Institute; Barts and The London School of Medicine and Dentistry; Inositide Signalling Group; London, UK
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Molecular determinants of PI3Kγ-mediated activation downstream of G-protein-coupled receptors (GPCRs). Proc Natl Acad Sci U S A 2013; 110:18862-7. [PMID: 24190998 DOI: 10.1073/pnas.1304801110] [Citation(s) in RCA: 109] [Impact Index Per Article: 9.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Phosphoinositide 3-kinase gamma (PI3Kγ) has profound roles downstream of G-protein-coupled receptors in inflammation, cardiac function, and tumor progression. To gain insight into how the enzyme's activity is shaped by association with its p101 adaptor subunit, lipid membranes, and Gβγ heterodimers, we mapped these regulatory interactions using hydrogen-deuterium exchange mass spectrometry. We identify residues in both the p110γ and p101 subunits that contribute critical interactions with Gβγ heterodimers, leading to PI3Kγ activation. Mutating Gβγ-interaction sites of either p110γ or p101 ablates G-protein-coupled receptor-mediated signaling to p110γ/p101 in cells and severely affects chemotaxis and cell transformation induced by PI3Kγ overexpression. Hydrogen-deuterium exchange mass spectrometry shows that association with the p101 regulatory subunit causes substantial protection of the RBD-C2 linker as well as the helical domain of p110γ. Lipid interaction massively exposes that same helical site, which is then stabilized by Gβγ. Membrane-elicited conformational change of the helical domain could help prepare the enzyme for Gβγ binding. Our studies and others identify the helical domain of the class I PI3Ks as a hub for diverse regulatory interactions that include the p101, p87 (also known as p84), and p85 adaptor subunits; Rab5 and Gβγ heterodimers; and the β-adrenergic receptor kinase.
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Shymanets A, Prajwal, Bucher K, Beer-Hammer S, Harteneck C, Nürnberg B. p87 and p101 subunits are distinct regulators determining class IB phosphoinositide 3-kinase (PI3K) specificity. J Biol Chem 2013; 288:31059-68. [PMID: 24014027 DOI: 10.1074/jbc.m113.508234] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Class IB phosphoinositide 3-kinase γ (PI3Kγ) comprises a single catalytic p110γ subunit, which binds to two non-catalytic subunits, p87 or p101, and controls a plethora of fundamental cellular responses. The non-catalytic subunits are assumed to be redundant adaptors for Gβγ enabling G-protein-coupled receptor-mediated regulation of PI3Kγ. Growing experimental data provide contradictory evidence. To elucidate the roles of the non-catalytic subunits in determining the specificity of PI3Kγ, we tested the impact of p87 and p101 in heterodimeric p87-p110γ and p101-p110γ complexes on the modulation of PI3Kγ activity in vitro and in living cells. RT-PCR, biochemical, and imaging data provide four lines of evidence: (i) specific expression patterns of p87 and p101, (ii) up-regulation of p101, providing the basis to consider p87 as a protein forming a constitutively and p101 as a protein forming an inducibly expressed PI3Kγ, (iii) differences in basal and stimulated enzymatic activities, and (iv) differences in complex stability, all indicating apparent diversity within class IB PI3Kγ. In conclusion, expression and activities of PI3Kγ are modified differently by p87 and p101 in vitro and in living cells, arguing for specific regulatory roles of the non-catalytic subunits in the differentiation of PI3Kγ signaling pathways.
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Affiliation(s)
- Aliaksei Shymanets
- From the Department of Pharmacology and Experimental Therapy, Institute of Experimental and Clinical Pharmacology and Toxicology, Eberhard Karls University Hospitals and Clinics and Interfaculty Centre of Pharmacogenomics and Pharmaceutical Research, University of Tübingen, 72074 Tübingen, Germany
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39
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Affiliation(s)
- Len Stephens
- The Babraham Institute, Cambridge, United Kingdom.
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Chiron D, Martin P, Di Liberto M, Huang X, Ely S, Lannutti BJ, Leonard JP, Mason CE, Chen-Kiang S. Induction of prolonged early G1 arrest by CDK4/CDK6 inhibition reprograms lymphoma cells for durable PI3Kδ inhibition through PIK3IP1. Cell Cycle 2013; 12:1892-900. [PMID: 23676220 PMCID: PMC3735703 DOI: 10.4161/cc.24928] [Citation(s) in RCA: 63] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Phosphatidylinositol-3-kinase (PI3K) signaling is constitutive in most human cancers. Selective inhibition of PI3Kδ (p110δ) by GS-1101 has emerged as a promising therapy in chronic lymphocytic leukemia and indolent lymphomas. In aggressive non-Hodgkin lymphomas such as mantle cell lymphoma (MCL), however, efficacy has been observed, but the extent and duration of tumor control is modest. To determine if tumor killing by GS-1101 is cell cycle-dependent, we show in primary MCL cells by whole-transcriptome sequencing that, despite aberrant expression and recurrent mutations in Cyclin D1, mutations are rare in coding regions of CDK4, RB1 and other genes that control G1-S cell cycle progression or PI3K/AKT signaling. PI3Kδ is the predominant PI3K catalytic subunit expressed, and inhibition by GS-1101 transiently inhibits AKT phosphorylation but not proliferation in MCL cells. Induction of prolonged early G1-arrest (pG1) by selective inhibition of CDK4/CDK6 with PD 0332991 amplifies and sustains PI3Kδ inhibition, which leads to robust apoptosis. Accordingly, inhibition of PI3Kδ induces apoptosis of primary MCL tumor cells once they have ceased to cycle ex vivo, and this killing is enhanced by PD 0332991 inhibition of CDK4/CDK6. PIK3IP1, a negative PI3K regulator, appears to mediate pG1 sensitization to PI3K inhibition; it is markedly reduced in MCL tumor cells compared with normal peripheral B cells, profoundly induced in pG1 and required for pG1 sensitization to GS-1101. Thus, the magnitude and duration of PI3K inhibition and tumor killing by GS-1101 is pG1-dependent, suggesting induction of pG1 by CDK4/CDK6 inhibition as a strategy to sensitize proliferating lymphoma cells to PI3K inhibition.
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Affiliation(s)
- David Chiron
- Department of Pathology and Laboratory Medicine, Weill-Cornell Medical College, New York, NY, USA
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41
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Xie Y, Abel PW, Kirui JK, Deng C, Sharma P, Wolff DW, Toews ML, Tu Y. Identification of upregulated phosphoinositide 3-kinase γ as a target to suppress breast cancer cell migration and invasion. Biochem Pharmacol 2013; 85:1454-62. [PMID: 23500535 PMCID: PMC3637857 DOI: 10.1016/j.bcp.2013.03.001] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2013] [Revised: 02/28/2013] [Accepted: 03/01/2013] [Indexed: 12/31/2022]
Abstract
Metastasis is the major cause of breast cancer mortality. We recently reported that aberrant G-protein coupled receptor (GPCR) signaling promotes breast cancer metastasis by enhancing cancer cell migration and invasion. Phosphatidylinositol 3-kinase γ (PI3Kγ) is specifically activated by GPCRs. The goal of the present study was to determine the role of PI3Kγ in breast cancer cell migration and invasion. Immunohistochemical staining showed that the expression of PI3Kγ protein was significantly increased in invasive human breast carcinoma when compared to adjacent benign breast tissue or ductal carcinoma in situ. PI3Kγ was also detected in metastatic breast cancer cells, but not in normal breast epithelial cell line or in non-metastatic breast cancer cells. In contrast, PI3K isoforms α, β and δ were ubiquitously expressed in these cell lines. Overexpression of recombinant PI3Kγ enhanced the metastatic ability of non-metastatic breast cancer cells. Conversely, migration and invasion of metastatic breast cancer cells were inhibited by a PI3Kγ inhibitor or by siRNA knockdown of PI3Kγ but not by inhibitors or siRNAs of PI3Kα or PI3Kβ. Lamellipodia formation is a key step in cancer metastasis, and PI3Kγ blockade disrupted lamellipodia formation induced by the activation of GPCRs such as CXC chemokine receptor 4 and protease-activated receptor 1, but not by the epidermal growth factor tyrosine kinase receptor. Taken together, these results indicate that upregulated PI3Kγ conveys the metastatic signal initiated by GPCRs in breast cancer cells, and suggest that PI3Kγ may be a novel therapeutic target for development of chemotherapeutic agents to prevent breast cancer metastasis.
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MESH Headings
- Breast Neoplasms/enzymology
- Breast Neoplasms/genetics
- Breast Neoplasms/pathology
- Carcinoma, Ductal/enzymology
- Carcinoma, Ductal/genetics
- Carcinoma, Ductal/pathology
- Cell Line, Tumor
- Cell Movement/drug effects
- Cell Movement/genetics
- Class Ib Phosphatidylinositol 3-Kinase/genetics
- Class Ib Phosphatidylinositol 3-Kinase/metabolism
- Diffusion Chambers, Culture
- Epithelial Cells/cytology
- Female
- Gene Expression Regulation, Neoplastic/drug effects
- Humans
- Neoplasm Invasiveness/genetics
- Neoplasm Invasiveness/pathology
- Phosphoinositide-3 Kinase Inhibitors
- Protein Kinase Inhibitors/pharmacology
- Pseudopodia/drug effects
- Pseudopodia/pathology
- RNA, Small Interfering/genetics
- Receptor, PAR-1/genetics
- Receptor, PAR-1/metabolism
- Receptors, CXCR4/genetics
- Receptors, CXCR4/metabolism
- Signal Transduction/drug effects
- Transfection
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Affiliation(s)
- Yan Xie
- Creighton University School of Medicine, Department of Pharmacology, Omaha, NE 68178
| | - Peter W. Abel
- Creighton University School of Medicine, Department of Pharmacology, Omaha, NE 68178
| | - Joseph K. Kirui
- Creighton University School of Medicine, Department of Pharmacology, Omaha, NE 68178
| | | | | | - Dennis W. Wolff
- Creighton University School of Medicine, Department of Pharmacology, Omaha, NE 68178
| | - Myron L. Toews
- University of Nebraska Medical Center, Department of Pharmacology and Experimental Neuroscience, Omaha, NE 68198
| | - Yaping Tu
- Creighton University School of Medicine, Department of Pharmacology, Omaha, NE 68178
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Niu M, Klingler-Hoffmann M, Brazzatti JA, Forbes B, Akekawatchai C, Hoffmann P, McColl SR. Comparative proteomic analysis implicates eEF2 as a novel target of PI3Kγ in the MDA-MB-231 metastatic breast cancer cell line. Proteome Sci 2013; 11:4. [PMID: 23320409 PMCID: PMC3564858 DOI: 10.1186/1477-5956-11-4] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2012] [Accepted: 12/23/2012] [Indexed: 11/30/2022] Open
Abstract
Background Cancer cell migration is fundamentally required for breast tumour invasion and metastasis. The insulin-like growth factor 1 tyrosine kinase receptor (IGF-1R) and the chemokine G-protein coupled receptor, CXCR4 have been shown to play an important role in breast cancer metastasis. Our previous study has shown that IGF-1R can transactivate CXCR4 via a physical association in the human MDA-MB-231 metastatic breast cancer cell line and that this plays a key role in IGF-I-induced migration of these cells. In the present study we used pharmacological inhibition and RNAi to identify PI3Kγ as an important migration signalling molecule downstream of receptor transactivation in MDA-MB-231 cells. To identify PI3Kγ-regulated proteins upon transactivation of CXCR4 by IGF-I, we undertook a comparative proteomics approach using 2-D- Fluorescence Difference Gel Electrophoresis (DIGE) and identified the proteins by mass spectrometry. Results These experiments identified eukaryotic elongation factor 2 (eEF2) as a novel downstream target of PI3Kγ after activation of the IGF-1R-CXCR4 heterodimer by IGF-I. Further analysis demonstrated that eEF2 is phosphorylated in MDA-MB-231 cells in response to IGF-I and that this is dependent on PI3Kγ activity. Conclusions Our data imply a novel role for PI3Kγ in facilitating cell migration by regulating phosphorylation of eEF2.
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Affiliation(s)
- Meizhi Niu
- School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA 5005, Australia
| | | | - Julie A Brazzatti
- School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA 5005, Australia.,Current address: Immunology Group, Paterson Institute for cancer research, The University of Manchester, Manchester, M20 4BX, England
| | - Briony Forbes
- School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA 5005, Australia
| | - Chareeporn Akekawatchai
- School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA 5005, Australia.,Current address: Department of Medical Technology, Thammasat University, Patumtani, 121212, Thailand
| | - Peter Hoffmann
- School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA 5005, Australia
| | - Shaun R McColl
- School of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA 5005, Australia
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Foster JG, Blunt MD, Carter E, Ward SG. Inhibition of PI3K signaling spurs new therapeutic opportunities in inflammatory/autoimmune diseases and hematological malignancies. Pharmacol Rev 2012; 64:1027-54. [PMID: 23023033 DOI: 10.1124/pr.110.004051] [Citation(s) in RCA: 96] [Impact Index Per Article: 7.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
The phosphoinositide 3-kinase/mammalian target of rapamycin/protein kinase B (PI3K/mTOR/Akt) signaling pathway is central to a plethora of cellular mechanisms in a wide variety of cells including leukocytes. Perturbation of this signaling cascade is implicated in inflammatory and autoimmune disorders as well as hematological malignancies. Proteins within the PI3K/mTOR/Akt pathway therefore represent attractive targets for therapeutic intervention. There has been a remarkable evolution of PI3K inhibitors in the past 20 years from the early chemical tool compounds to drugs that are showing promise as anticancer agents in clinical trials. The use of animal models and pharmacological tools has expanded our knowledge about the contribution of individual class I PI3K isoforms to immune cell function. In addition, class II and III PI3K isoforms are emerging as nonredundant regulators of immune cell signaling revealing potentially novel targets for disease treatment. Further complexity is added to the PI3K/mTOR/Akt pathway by a number of novel signaling inputs and feedback mechanisms. These can present either caveats or opportunities for novel drug targets. Here, we consider recent advances in 1) our understanding of the contribution of individual PI3K isoforms to immune cell function and their relevance to inflammatory/autoimmune diseases as well as lymphoma and 2) development of small molecules with which to inhibit the PI3K pathway. We also consider whether manipulating other proximal elements of the PI3K signaling cascade (such as class II and III PI3Ks or lipid phosphatases) are likely to be successful in fighting off different immune diseases.
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Affiliation(s)
- John G Foster
- Inflammatory Cell Biology Laboratory, Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath, UK.
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Blunt MD, Ward SG. Targeting PI3K isoforms and SHIP in the immune system: new therapeutics for inflammation and leukemia. Curr Opin Pharmacol 2012; 12:444-51. [PMID: 22483603 DOI: 10.1016/j.coph.2012.02.015] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2012] [Accepted: 02/23/2012] [Indexed: 10/28/2022]
Abstract
PI3K is critical for the normal function of the immune system, however dysregulated PI3K mediated signaling has been linked to the development of many immune mediated pathologies. This review describes current progress in the development of isoform-specific PI3K inhibitors that hold promise for the treatment of hematopoietic malignancies as well as for inflammatory and autoimmune diseases. A SH2-domain containing inositol-5-phosphatase (SHIP) is a regulator of PI3K signaling, and is also discussed as a potential drug target for immunomodulation and the treatment of leukemia. Recent progress has been made in the development of small molecule compounds that potently and selectively modulate SHIP activity and hence provide a novel mechanism to alter PI3K mediated signaling.
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Affiliation(s)
- Matthew D Blunt
- Inflammatory Cell Biology Laboratory, Department of Pharmacy and Pharmacology, University of Bath, Claverton Down, Bath, BA2 7AY, UK
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