Fig 1.
DNA damage as the critical step in Pt-induced cell death.
(A) The major pathways of platinum (Pt) drug-induced cell death. After administration, cellular uptake and efflux determines the intracellular accumulation of Pt agents, which can be inactivated by the intracellular thiol-containing molecules. Eventually, Pt agents induce DNA damage, including drug-DNA adducts, which triggers cell cycle arrest and DNA repair. DNA adduct formation and repair determines the fate of cells, although other factors also play important roles, such as pro- and anti-apoptotic proteins. (B) Diagram showing the formation of carboplatin- and oxaliplatin-DNA adducts and the positions of the radiocarbon labels on each drug used for this study in order to enable quantification of drug-DNA adduct formation and repair by accelerator mass spectrometry.
Table 1.
Comparison of drug IC50 values for 5637 and 5637R cells.
Fig 2.
(A-B) Comparison of cell uptake and efflux. A. cell uptake of oxaliplatin. 5637R cells had decreased cell uptake. B: 5637 and 5637R had similar cell efflux rates. (C-D) Oxaliplatin and carboplatin cellular efflux differences between the two cell lines were not statistically significant.
Fig 3.
Drug inactivation by cellular glutathione.
(A) Comparison of carboplatin-DNA adduct formation between 5637 and 5637R. (B) Comparison and correlation of IC50 values with carboplatin-adduct AUC, adduct levels four hours after dosing and DNA repair. (C) Comparison of cell uptake and efflux of carboplatin between 5637 and 5637R cells.
Fig 4.
Oxaliplatin- and carboplatin-DNA adduct formation and repair.
Comparison of oxaliplatin- and carboplatin-DNA adduct formation between 5637 and 5637R cells. The chemoresistant 5637R cells had higher oxaliplatin-DNA adduct levels at all time points compared to more treatment sensitive 5637 cells.
Table 2.
Oxaliplatin-DNA adduct formation and repair.
Table 3.
Chemoresistance-associated gene expression levels in 5637 and 5637R cells.
Fig 5.
RNAseq and qRT-PCR show similar trends in gene expression levels for selected resistance-realated genes.
Four genes (TSPAN7, AKR1C2, AKR1C1, and CYR61) have increased levels in 5637R cells and two genes (HTRA1 and AQP3) have decreased levels in the resistant cells. (A) Fold changes in chemoresistance gene levels relative to the 5637 parental cells as determined by RNA-seq. (B) Fold change in chemoresistance gene transcript levels relative to the 5637 parental cells as determined by qRT-PCR.