title: Robotic Mammosphere Assay for High-Throughput Screening in Triple-Negative Breast Cancer.
abstract: In order to identify novel treatment principles specifically affecting cancer stem cells in triple-negative breast cancer, we have developed a high-throughput screening method based on the mammosphere and anoikis resistance assays allowing us to screen compounds using a functional readout. The assay was validated against manual protocols and through the use of positive controls, such as the response to hypoxia and treatment with the known cancer stem cell-targeting compound salinomycin. Manual and robotic procedures were compared and produced similar results in cell handling, cell cultures, and counting techniques, with no statistically significant difference produced from either method. The variance between samples processed manually versus robotically was no greater than 0.012, while Levene's test of significance was 0.2, indicating no significant difference between mammosphere data produced manually or robotically. Through the screening of 989 FDA-approved drugs and a follow-up screen assessing the antineoplastic subgroup, we have identified three therapeutic compounds with the ability to modulate the breast cancer stem cell fraction in the triple-negative breast cancer cell line MDA-MB-231, highlighting their potential usage as stem cell-specific adjuvant treatments.
authors.affiliation: 1 Sahlgrenska Cancer Center, Göteborgs Universitet, Göteborg, Sweden.
authors.forename: P A
authors.initials: PA
authors.lastname: Fitzpatrick
authors.affiliation: 1 Sahlgrenska Cancer Center, Göteborgs Universitet, Göteborg, Sweden.
authors.forename: N
authors.initials: N
authors.lastname: Akrap
authors.affiliation: 1 Sahlgrenska Cancer Center, Göteborgs Universitet, Göteborg, Sweden.
authors.forename: E M V
authors.initials: EMV
authors.lastname: Söderberg
authors.affiliation: 2 Faculty of Life Sciences, Manchester University, Manchester, UK.
authors.forename: H
authors.initials: H
authors.lastname: Harrison
authors.affiliation: 3 Drug Discovery Unit, CRUK Manchester Institute, Manchester, UK.
authors.forename: G J
authors.initials: GJ
authors.lastname: Thomson
authors.affiliation: 1 Sahlgrenska Cancer Center, Göteborgs Universitet, Göteborg, Sweden.
authors.forename: G
authors.initials: G
authors.lastname: Landberg
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D005260:Female; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D014411:Neoplastic Stem Cells / Q000473:pathology; D011714:Pyrans / Q000494:pharmacology; D065287:Robotic Surgical Procedures / Q000379:methods; D064726:Triple Negative Breast Neoplasms / Q000188:drug therapy / Q000473:pathology*; D000095662:Polyether Polyketides
chemical_list: D000970:Antineoplastic Agents; D011714:Pyrans; C010327:salinomycin; D000095662:Polyether Polyketides
keywords: cancer and cancer drugs; cell-based assays; high-content screening; stem cells