title: Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway.
abstract: Energy metabolism plasticity enables stemness programs during the reprogramming of somatic cells to an induced pluripotent stem cell (iPSC) state. This relationship may introduce a new era in the understanding of Warburg's theory on the metabolic origin of cancer at the level of cancer stem cells (CSCs). Here, we used Yamanaka's stem cell technology in an attempt to create stable CSC research lines in which to dissect the transcriptional control of mTOR--the master switch of cellular catabolism and anabolism--in CSC-like states. The rare colonies with iPSC-like morphology, obtained following the viral transduction of the Oct4, Sox2, Klf4, and c-Myc (OSKM) stemness factors into MCF-7 luminal-like breast cancer cells (MCF-7/Rep), demonstrated an intermediate state between cancer cells and bona fide iPSCs. MCF-7/Rep cells notably overexpressed SOX2 and stage-specific embryonic antigen (SSEA)-4 proteins; however, other stemness-related markers (OCT4, NANOG, SSEA-1, TRA-1-60, and TRA-1-81) were found at low to moderate levels. The transcriptional analyses of OSKM factors confirmed the strong but unique reactivation of the endogenous Sox2 stemness gene accompanied by the silencing of the exogenous Sox2 transgene in MCF-7/Rep cells. Some but not all MCF-7/Rep cells acquired strong alkaline phosphatase (AP) activity compared with MCF-7 parental cells. SOX2-overexpressing MCF-7/Rep cells contained drastically higher percentages of CD44(+) and ALDEFLUOR-stained ALDH(bright) cells than MCF-7 parental cells. The overlap between differentially expressed mTOR signaling-related genes in 3 different SOX2-overexpressing CSC-like cell lines revealed a notable downregulation of 3 genes, PRKAA1 (which codes for the catalytic α 1 subunit of AMPK), DDIT4/REDD1 (a stress response gene that operates as a negative regulator of mTOR), and DEPTOR (a naturally occurring endogenous inhibitor of mTOR activity). The insulin-receptor gene (INSR) was differentially upregulated in MCF-7/Rep cells. Consistent with the downregulation of AMPK expression, immunoblotting procedures confirmed upregulation of p70S6K and increased phosphorylation of mTOR in Sox2-overexpressing CSC-like cell populations. Using an in vitro model of the de novo generation of CSC-like states through the nuclear reprogramming of an established breast cancer cell line, we reveal that the transcriptional suppression of mTOR repressors is an intrinsic process occurring during the acquisition of CSC-like properties by differentiated populations of luminal-like breast cancer cells. This approach may provide a new path for obtaining information about preventing the appearance of CSCs through the modulation of the AMPK/mTOR pathway.
authors: Metabolism & Cancer Group; Translational Research Laboratory; Catalan Institute of Oncology-Girona (ICO-Girona); Girona, Spain; Molecular Oncology; Girona Biomedical Research Institute (IDIBGI); Girona, Spain.
authors: Bruna
authors: B
authors: Corominas-Faja
authors: Sílvia
authors: S
authors: Cufí
authors: Cristina
authors: C
authors: Oliveras-Ferraros
authors: Elisabet
authors: E
authors: Cuyàs
authors: Eugeni
authors: E
authors: López-Bonet
authors: Ruth
authors: R
authors: Lupu
authors: Tomás
authors: T
authors: Alarcón
authors: Luciano
authors: L
authors: Vellon
authors: Juan Manuel
authors: JM
authors: Iglesias
authors: Olatz
authors: O
authors: Leis
authors: Ángel G
authors: ÁG
authors: Martín
authors: Alejandro
authors: A
authors: Vazquez-Martin
authors: Javier A
authors: JA
authors: Menendez
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D001943:Breast Neoplasms; D065150:Cellular Reprogramming; D015536:Down-Regulation; D054890:Fatty Acid Synthase, Type I / Q000378:metabolism; D005260:Female; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D016547:Kinesins / Q000235:genetics / Q000378:metabolism; D000090062:Kruppel-Like Factor 4; D061986:MCF-7 Cells; D014411:Neoplastic Stem Cells / Q000166:cytology* / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D011972:Receptor, Insulin / Q000235:genetics / Q000378:metabolism; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation; D015854:Up-Regulation
chemical_list: C469991:DDIT4 protein, human; D047908:Intracellular Signaling Peptides and Proteins; C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D050814:Octamer Transcription Factor-3; D016271:Proto-Oncogene Proteins c-myc; D034741:RNA, Small Interfering; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; D014157:Transcription Factors; C518729:FASN protein, human; D054890:Fatty Acid Synthase, Type I; C556139:DEPTOR protein, human; C546842:MTOR protein, human; D011972:Receptor, Insulin; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; C526180:PRKAA1 protein, human; C484228:KIF4A protein, human; D016547:Kinesins
keywords: AMPK; SOX2; breast cancer; cancer stem cells; mTOR; reprogramming
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