title: Primary breast cancer cell culture yields intra-tumor heterogeneous subpopulations expressing exclusive patterns of receptor tyrosine kinases.
abstract: It has become evident that intra-tumor heterogeneity of breast cancer impact on several biological processes such as proliferation, migration, cell death and also might contribute to chemotherapy resistance. The expression of Receptor Tyrosine Kinases (RTKs) has not been analyzed in the context of intra-tumor heterogeneity in a primary breast cancer cell culture. Several subpopulations were isolated from the MBCDF (M serial-breast cancer ductal F line) primary breast cancer cells and were successfully maintained in culture and divided in two groups according to their morphology and RTKs expression pattern, and correlated with biological processes like proliferation, migration, anchorage-independent cell growth, and resistance to cytotoxic chemotherapy drugs and tyrosine kinase inhibitors (TKIs).
  
  
  
  Subpopulations were isolated from MBCDF primary breast cancer cell culture by limiting dilution. RTKs and hormone receptors were examined by Western blot. Proliferation was measure by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT assay). Cell viability was evaluated by Crystal Violet. Migration was assessed using Boyden chambers. Anchorage-independent cell growth was evaluated by colony formation in soft agar.
  
  
  
  Several subpopulations were isolated from the MBCDF breast cancer cells that were divided into two groups according to their morphology. Analysis of RTKs expression pattern showed that HER1, HER3, c-Met and VEGFR2 were expressed exclusively in cells from group 1, but not in cells from group 2. PDGFR was expressed only in cells from group 2, but not in cells from group 1. HER2, HER4, c-Kit, IGF1-R were expressed in all subpopulations. Biological processes correlated with the RTKs expression pattern. Group 2 subpopulations present the highest rate of cell proliferation, migration and anchorage-independent cell growth. Analysis of susceptibility to chemotherapy drugs and TKIs showed that only Paclitaxel and Imatinib behaved differently between groups. Group 1-cells were resistant to both Paclitaxel and Imatinib.
  
  
  
  We demonstrated that subpopulations from MBCDF primary cell culture could be divided into two groups according to their morphology and a RTKs excluding-expression pattern. The differences observed in RTKs expression correlate with the biological characteristics and chemoresistance of each group. These results suggest that intra-tumor heterogeneity contributes to generate groups of subpopulations with a more aggressive phenotype within the tumor.
authors.affiliation: Unidad de Bioquímica, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico.
authors.forename: José
authors.initials: J
authors.lastname: Esparza-López
authors.affiliation: Unidad de Bioquímica, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico.
authors.forename: Pier A
authors.initials: PA
authors.lastname: Ramos-Elías
authors.affiliation: Unidad de Bioquímica, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico.
authors.forename: Andrea
authors.initials: A
authors.lastname: Castro-Sánchez
authors.affiliation: Departamento de Biología Molecular y Biotecnología, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Circuito Escolar S/N, Ciudad Universitaria, Delegación Coyoacán, CP 04500, Distrito Federal, Mexico.
authors.forename: Leticia
authors.initials: L
authors.lastname: Rocha-Zavaleta
authors.affiliation: Hospital Ángeles del Pedregal, Camino a Santa Teresa # 1055, México, CP 10700, Distrito Federal, Mexico.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Escobar-Arriaga
authors.affiliation: Unidad de Bioquímica, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico.|Departamento de Medicina Genómica y Toxicología Ambiental, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Circuito Escolar S/N, Ciudad Universitaria, Delegación Coyoacán, CP 04500, Distrito Federal, Mexico.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Zentella-Dehesa
authors.affiliation: Departamento de Hemato-Oncología, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico.
authors.forename: Eucario
authors.initials: E
authors.lastname: León-Rodríguez
authors.affiliation: Departamento de Cirugía, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico.
authors.forename: Heriberto
authors.initials: H
authors.lastname: Medina-Franco
authors.affiliation: Unidad de Bioquímica, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Vasco de Quiroga 15, Belisario Domínguez Sección XVI, Delegación Tlalpan, CP 14080, Distrito Federal, Mexico. maria.ibarras@incmnsz.mx.
authors.forename: María de Jesus
authors.initials: Mde J
authors.lastname: Ibarra-Sánchez
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D019008:Drug Resistance, Neoplasm; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D018740:Genetic Heterogeneity; D006801:Humans; D000068877:Imatinib Mesylate / Q000494:pharmacology*; D017239:Paclitaxel / Q000494:pharmacology*; D061251:Primary Cell Culture / Q000379:methods*; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
chemical_list: D000068877:Imatinib Mesylate; D020794:Receptor Protein-Tyrosine Kinases; D017239:Paclitaxel
keywords: Breast cancer; Intra-tumor heterogeneity; PDGFR; Receptor tyrosine kinases; Tyrosine kinase inhibitor