Fig. 5

CYP19A1 knockout reverses chemoresistance in CRC cells. (A-F) The OCR of wild-type and CYP19A1 knockout chemoresistant SW480 cells was measured using the Seahorse Mito Stress Test. Basal, ATP-linked, and maximal respiration were assessed by sequential treatment with oligomycin (O), FCCP (F), and rotenone/antimycin A (R/A) in 5-fluorouracil (5FU-R; A, B), irinotecan (IRI-R; C, D), and oxaliplatin (OXA-R; E, F) resistant cells. (G) Mitochondrial complex I activity in 5FU-R, IRI-R, and OXA-R cells was measured using a commercially available assay kit. (H-J) Dose-response curves of parental SW480 cells treated with 5FU, IRI, and OXA in wild-type (WT), CYP19A1 overexpressing (OE), and estradiol-treated (WT + E2) conditions. (K-M) Dose-response curves showing cell viability of 5FU, IRI, and OXA in 5FU-R, IRI-R, and OXA-R cell lines treated with different concentrations of 5FU, IRI, and OXA, respectively. Curves are shown for WT, knockout (KO), and KO cells complemented with constructs expressing WT or mutant (D309N, Y361F) CYP19A1. (N-P) Dose-response curves illustrating cell viability in 5FU-R, IRI-R, and OXA-R lines exposed to varying concentrations of their respective drugs in WT, KO, and KO cells treated with estradiol (KO + E2). (Q-S) Dose-response curves demonstrating cell viability in 5FU-R, IRI-R, and OXA-R lines exposed to varying concentrations of their respective drugs in WT, KO, and KO cells supplemented with mitochondria extracted from WT cells (KO + Mito). Statistical significance of KO groups compared to the WT group was determined using Student’s t-test (*** p < 0.001). Data are presented as mean ± SD. (A-F) n = 6; (G-S) n = 3