SEPARATION OF RESISTANCE TO ANTITUMOR DIARYLSULFONYLUREA AGENTS FROM COLLATERAL SENSITIVITY TO MITOCHONDRIAL TOXINS

Compared with parental GC(3)/c1 human colon adenocarcinoma cells, whic h are diarylsulfonylurea (DSU)-sensitive cells, the DSU-resistant clon e LYC5 demonstrates 4.2-, 12.8-, and 5.3-fold increase in sensitivity to the mitochondrial toxins rotenone, antimycin, and oligomycin, respe ctively. Studies with hybrids formed by fusion of parental GC(3)/c1 ce lls with LYC5 cells have indicated that resistance to antitumor DSUs a nd collateral sensitivity to mitochondrial toxins are recessive and th erefore potentially linked. To examine this, we transfected a cDNA lib rary from GC(3)/c1 cells, constructed in pcDNA3, into LYC5 cells. G418 -resistant colonies were selected and further selected in a single ste p for resistance to rotenone (100 nM). Individual colonies (designated T5LR) were expanded and tested for sensitivity to mitochondrial toxin s, antitumor DSU agents (LY195779 and LY186391) that demonstrate a 45- 50-fold differential potency against GC(3)/c1, LYC5 cells, and the ant imitotic agent vincristine. Results demonstrate that resistance to mit ochondrial toxins rotenone, antimycin, and oligomycin can be transferr ed without conferring a DSU-sensitive phenotype. Furthermore, in T5LR clones, resistance to mitochondrial toxins was not associated with inc reased resistance to vincristine or increased P-glycoprotein expressio n, supporting the contention that resistance to these agents is indepe ndent of P-glycoprotein. Southern blot analysis of T5LR clones demonst rated unique integration sites for the neomycin phosphotransferase gen e into genomic DNA in clones 4 and 9, indicating independent derivatio n. Analysis of clones 4, 6, and 9 with the use of polymerase chain rea ction demonstrated a cDNA insert of similar to 1.0 kilobase.