title: ABCG2 is a potential marker of tumor-initiating cells in breast cancer.
abstract: The existence of tumor-initiating cells (TICs) within solid tumors has been hypothesized to explain tumor heterogeneity and resistance to cancer therapy. In breast cancer, the expression of CD44 and CD24 and the activity of aldehyde dehydrogenase 1 (ALDH1) can be used to selectively isolate a cell population enriched in TICs. However, the ideal marker to identify TICs has not been established. The aim of this study was to evaluate the expression of novel potential markers for TIC in breast carcinoma. We prospectively analyzed the expression of CD44, CD24, ABCG2, and CXCR4, and the activity of ALDH1 by using flow cytometry in 48 invasive ductal carcinomas from locally advanced and metastatic breast cancer patients who were administered primary chemotherapy. A mammosphere assay was employed in 30 samples. The relationship among flow cytometric analyses, ABCG2 gene expression, and clinical and pathological responses to therapy was analyzed. The GSE32646 database was analyzed in silico to identify genes associated with tumors with low and high ABCG2 expression. We observed that the presence of ABCG2(+) cells within the primary tumor was the only marker to predict the formation of mammospheres in vitro (R (2) = 0.15, p = 0.029). Quantitative polymerase chain reaction (qPCR) revealed a positive correlation between ABCG2 expression and the presence of ABCG2(+) cells within the primary tumor. The expression of ABCG2 was predictive of the response to neoadjuvant chemotherapy in our experiments and in the GSE32646 dataset (p = 0.04 and p = 0.002, respectively). The in silico analysis demonstrated that ABCG2(Up) breast cancer samples have a slower cell cycle and a higher expression of membrane proteins but a greater potential for chromosomal instability, metastasis, immune evasion, and resistance to hypoxia. Such genetic characteristics are compatible with highly aggressive and resistant tumors. Our results support the hypothesis that the presence of ABCG2(+) cells in breast carcinomas is a marker of resistance to chemotherapy, and based on in vitro assays and the genetic profile, we show, for the first time, that ABCG2 protein can be used as an independent marker for TIC identification in breast cancer.
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. resicchieri@usp.br.
authors.forename: Renata Danielle
authors.initials: RD
authors.lastname: Sicchieri
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. willian.silveira@usp.br.|Institut de Recherche en Cancérologie de Montpellier, Université Montpellier INSERM U896, F-34298, Montpellier, France. willian.silveira@usp.br.
authors.forename: Willian Abraham
authors.initials: WA
authors.lastname: da Silveira
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. laramouro@usp.br.
authors.forename: Larissa Raquel Mouro
authors.initials: LR
authors.lastname: Mandarano
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. tatimendes@usp.br.
authors.forename: Tatiane Mendes Gonçalves
authors.initials: TM
authors.lastname: de Oliveira
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. hhacarra@fmrp.usp.br.
authors.forename: Hélio Humberto Angotti
authors.initials: HH
authors.lastname: Carrara
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. muglia@convex.com.br.
authors.forename: Valdair Francisco
authors.initials: VF
authors.lastname: Muglia
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. jmandrade@usp.br.
authors.forename: Jurandyr Moreira
authors.initials: JM
authors.lastname: de Andrade
authors.affiliation: Department of Gynecology and Obstetrics - Breast Disease Division, Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. dtiezzi@usp.br.|CISBi - Center for Integrative Systems Biology - Ribeirão Preto Medical School, University of São Paulo, Av. Bandeirantes, 3900 - Monte Alegre, Ribeirão Preto, São Paulo, Brazil, 14048-900. dtiezzi@usp.br.
authors.forename: Daniel Guimarães
authors.initials: DG
authors.lastname: Tiezzi
mesh_terms: D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis* / Q000235:genetics; D000328:Adult; D000368:Aged; D000080924:Aldehyde Dehydrogenase 1 Family; D014408:Biomarkers, Tumor / Q000096:biosynthesis* / Q000235:genetics; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D051927:CD24 Antigen / Q000096:biosynthesis; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D018960:Hyaluronan Receptors / Q000096:biosynthesis; D007527:Isoenzymes / Q000096:biosynthesis; D008875:Middle Aged; D020360:Neoadjuvant Therapy; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D014411:Neoplastic Stem Cells / Q000473:pathology*; D019718:Receptors, CXCR4 / Q000096:biosynthesis; D050697:Retinal Dehydrogenase / Q000096:biosynthesis
chemical_list: C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D014408:Biomarkers, Tumor; D051927:CD24 Antigen; C527276:CXCR4 protein, human; D018960:Hyaluronan Receptors; D007527:Isoenzymes; D009363:Neoplasm Proteins; D019718:Receptors, CXCR4; D000080924:Aldehyde Dehydrogenase 1 Family; C510223:ALDH1A1 protein, human; D050697:Retinal Dehydrogenase
keywords: Breast cancer; Chemotherapy; Resistance to treatment; Tumor-initiating cells