Jeonghun Hana,1, Yukyung Junb,c,1, So Hyun Kimd, Hong-Hoa Hoanga, Yeonjoo Jungc, Suyeon Kimb,c, Jaesang Kimb,c, Robert H. Austine,2, Sanghyuk Leeb,c,2, and Sungsu Parka,2
aSchool of Mechanical Engineering, Sungkyunkwan University, Suwon 16419, Korea;
bDepartment of Life Science, Ewha Womans University, Seoul 03760, Korea;
cEwha Research Center for Systems Biology, Ewha Womans University, Seoul 03760, Korea;
dCenter for BioMicrosystems, Brain Science Institute, Korea Institute of Science and Technology, Seoul 0202792, Korea;
eDepartment of Physics, Princeton University, Princeton, NJ 08544
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.
microhabitats, cancer, doxorubicin, evolution, resistance
1J.H. and Y. Jun contributed equally to this work.
2To whom correspondence may be addressed.
Author contributions: R.H.A., S.L., and S.P. designed research; J.H., S.H.K., H.-H.H., Y. Jung, and S.K. performed experimental works; R.H.A. and S.P. contributed new reagents/analytic tools; Y. Jun analyzed data; and J.K., R.H.A., S.L., and S.P. wrote the paper.