title: Epithelial-to-mesenchymal transition (EMT) confers primary resistance to trastuzumab (Herceptin).
abstract: The rate of inherent resistance to single-agent trastuzumab in HER2-overexpressing metastatic breast carcinomas is impressive at above 70%. Unfortunately, little is known regarding the distinctive genetic signatures that could predict trastuzumab refractoriness ab initio. The epithelial-to-mesenchymal transition (EMT) molecular features, HER2 expression status and primary responses to trastuzumab were explored in the public Lawrence Berkeley Laboratory (LBL) Breast Cancer Collection. Lentivirus-delivered small hairpin RNAs were employed to reduce specifically and stably the expression of EMT transcription factors in trastuzumab-refractory basal/HER2+ cells. Cell proliferation assays and pre-clinical nude mice xenograft-based studies were performed to assess the contribution of specific EMT transcription factors to inherent trastuzumab resistance. Primary sensitivity to trastuzumab was restricted to the SLUG/SNAIL2-negative subset of luminal/HER2+ cell lines, whereas all of the SLUG/SNAIL2-positive basal/HER2+ cell lines exhibited an inherent resistance to trastuzumab. The specific knockdown of SLUG/SNAIL2 suppressed the stem-related CD44+CD24(-/low) mesenchymal immunophenotype by transcriptionally upregulating the luminal epithelial marker CD24 in basal/HER2+ cells. Basal/HER2+ cells gained sensitivity to the growth-inhibitory effects of trastuzumab following SLUG/SNAIL2 gene depletion, which induced the expression of the mesenchymal-to-epithelial transition (MET) genes involved in promoting an epithelial phenotype. The isolation of CD44+CD24(-/low) mesenchymal cells by magnetic-activated cell sorting (MACS) confirmed their intrinsic unresponsiveness to trastuzumab. A reduction in tumor growth and dramatic gain in sensitivity to trastuzumab in vivo were confirmed when the SLUG/SNAIL2 knockdown basal/HER2+ cells were injected into nude mice. HER2 overexpression in a basal, rather than in a luminal molecular background, results in a basal/HER2+ breast cancer subtype that is intrinsically resistant to trastuzumab. EMT transcription factors might induce an enhanced phenotypic plasticity that would allow basal/HER2+ breast cancer cells to "enter" into and "exit" dynamically from trastuzumab-responsive stem cell-like states. The systematic determination of SLUG/SNAIL2 as a stem/CD44+CD24(-/low) cell-associated protein may improve the therapeutic management of HER2+ breast carcinomas.
authors.affiliation: Metabolism & Cancer Group, Translational Research Laboratory, Catalan Institute of Oncology, Girona, Catalonia, Spain.
authors.forename: Cristina
authors.initials: C
authors.lastname: Oliveras-Ferraros
authors.forename: Bruna
authors.initials: B
authors.lastname: Corominas-Faja
authors.forename: Sílvia
authors.initials: S
authors.lastname: Cufí
authors.forename: Alejandro
authors.initials: A
authors.lastname: Vazquez-Martin
authors.forename: Begoña
authors.initials: B
authors.lastname: Martin-Castillo
authors.forename: Juan Manuel
authors.initials: JM
authors.lastname: Iglesias
authors.forename: Eugeni
authors.initials: E
authors.lastname: López-Bonet
authors.forename: Ángel G
authors.initials: ÁG
authors.lastname: Martin
authors.forename: Javier A
authors.initials: JA
authors.lastname: Menendez
mesh_terms: D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000633:toxicity*; D000970:Antineoplastic Agents / Q000633:toxicity*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D051927:CD24 Antigen / Q000378:metabolism; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D058750:Epithelial-Mesenchymal Transition; D005260:Female; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D016130:Immunophenotyping; D051379:Mice; D008819:Mice, Nude; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014183:Transplantation, Heterologous; D000068878:Trastuzumab
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D051927:CD24 Antigen; D018960:Hyaluronan Receptors; D018719:Erb-b2 Receptor Tyrosine Kinases; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; D000068878:Trastuzumab; C000604407:SNAI1 protein, human; C000604429:Snai1 protein, mouse; C000604449:Snai2 protein, mouse