title: Rapid emergence and mechanisms of resistance by U87 glioblastoma cells to doxorubicin in an in vitro tumor microfluidic ecology.
abstract: In vitro prediction of the probable rapid emergence of resistance to a drug in tumors could act to winnow out potential candidates for further costly development. We have developed a microfluidic device consisting of ∼500 hexagonal microcompartments that provides a complex ecology with wide ranges of drug and nutrient gradients and local populations. This ecology of a fragmented metapopulation induced the drug resistance in stage IV U87 glioblastoma cells to doxorubicin in 7 d. Exome and transcriptome sequencing of the resistant cells identified mutations and differentially expressed genes. Gene ontology and pathway analyses of the genes identified showed that they were functionally relevant to the established mechanisms of doxorubicin action. Specifically, we identified (i) a frame-shift insertion in the filamin-A gene, which regulates the influx and efflux of topoisomerase II poisons; (ii) the overexpression of aldo-keto reductase enzymes, which convert doxorubicin into doxorubicinol; and (iii) activation of NF-κB via alterations in the nucleotide-binding oligomerization domain (NOD)-like receptor signaling pathway from mutations in three genes (CARD6, NSD1, and NLRP13) and the overexpression of inflammatory cytokines. Functional experiments support the in silico analyses and, together, demonstrate the effects of these genetic changes. Our findings suggest that, given the rapid evolution of resistance and the focused response, this technology could act as a rapid screening modality for genetic aberrations leading to resistance to chemotherapy as well as counter selection of drugs unlikely to be successful ultimately.
authors.affiliation: School of Mechanical Engineering, Sungkyunkwan University, Suwon 16419, Korea.
authors.forename: Jeonghun
authors.initials: J
authors.lastname: Han
authors.affiliation: Department of Life Science, Ewha Womans University, Seoul 03760, Korea.|Ewha Research Center for Systems Biology, Ewha Womans University, Seoul 03760, Korea.
authors.forename: Yukyung
authors.initials: Y
authors.lastname: Jun
authors.affiliation: Center for BioMicrosystems, Brain Science Institute, Korea Institute of Science and Technology, Seoul 0202792, Korea.
authors.forename: So Hyun
authors.initials: SH
authors.lastname: Kim
authors.affiliation: School of Mechanical Engineering, Sungkyunkwan University, Suwon 16419, Korea.
authors.forename: Hong-Hoa
authors.initials: HH
authors.lastname: Hoang
authors.affiliation: Ewha Research Center for Systems Biology, Ewha Womans University, Seoul 03760, Korea.
authors.forename: Yeonjoo
authors.initials: Y
authors.lastname: Jung
authors.affiliation: Department of Life Science, Ewha Womans University, Seoul 03760, Korea.|Ewha Research Center for Systems Biology, Ewha Womans University, Seoul 03760, Korea.
authors.forename: Suyeon
authors.initials: S
authors.lastname: Kim
authors.affiliation: Department of Life Science, Ewha Womans University, Seoul 03760, Korea.|Ewha Research Center for Systems Biology, Ewha Womans University, Seoul 03760, Korea.
authors.forename: Jaesang
authors.initials: J
authors.lastname: Kim
authors.affiliation: Department of Physics, Princeton University, Princeton, NJ 08544 nanopark@skku.edu sanghyuk@ewha.ac.kr austin@princeton.edu.
authors.forename: Robert H
authors.initials: RH
authors.lastname: Austin
authors.affiliation: Department of Life Science, Ewha Womans University, Seoul 03760, Korea; nanopark@skku.edu sanghyuk@ewha.ac.kr austin@princeton.edu.|Ewha Research Center for Systems Biology, Ewha Womans University, Seoul 03760, Korea.
authors.forename: Sanghyuk
authors.initials: S
authors.lastname: Lee
authors.affiliation: School of Mechanical Engineering, Sungkyunkwan University, Suwon 16419, Korea; nanopark@skku.edu sanghyuk@ewha.ac.kr austin@princeton.edu.
authors.forename: Sungsu
authors.initials: S
authors.lastname: Park
mesh_terms: D000074408:Aldo-Keto Reductases / Q000235:genetics / Q000378:metabolism; D000903:Antibiotics, Antineoplastic / Q000493:pharmacokinetics / Q000494:pharmacology*; D001932:Brain Neoplasms / Q000188:drug therapy* / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D019020:Directed Molecular Evolution; D004317:Doxorubicin / Q000493:pharmacokinetics / Q000494:pharmacology*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D064448:Filamins / Q000235:genetics / Q000378:metabolism; D005909:Glioblastoma / Q000188:drug therapy* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D056656:Lab-On-A-Chip Devices; D046210:Microfluidic Analytical Techniques / Q000379:methods; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D000903:Antibiotics, Antineoplastic; D064448:Filamins; D016328:NF-kappa B; D004317:Doxorubicin; D000074408:Aldo-Keto Reductases
keywords: cancer; doxorubicin; evolution; microhabitats; resistance