HUMAN MDR-1 PROTEIN OVEREXPRESSION DELAYS THE APOPTOTIC CASCADE IN CHINESE-HAMSTER OVARY FIBROBLASTS

Several laboratories have reported that overexpression of the multidru g resistance (MDR) protein is associated with intracellular alkaliniza tion, and several investigators have reported that cells induced to un dergo programmed cell death (apoptosis) acidify quite significantly. B ecause it is difficult to fully explain the resistance to apoptosis-in ducing chemotherapeutic drugs that is exhibited by MDR tumor cells sol ely via altered drug transport alone [Hoffman et al. (1996) J. Gen. Ph ysiol. 108, 295-313], we have investigated whether overexpression of t he hu MDR 1 protein alters progression of the apoptotic cascade. LR73 fibroblasts induced to undergo apoptosis either via treatment with the chemotherapeutic drug colchicine or by serum withdrawal exhibit cellu lar volume changes, intracellular acidification, nuclear condensation, and chromosomal digestion (''ladder formation''), characteristic of a poptosis, in a temporally well-defined pattern. However, multidrug res istant LR73/20E or LR73/27 hu MDR 1 transfectants recently created in our laboratory without selection on chemotherapeutic drug are signific antly delayed in the onset of apoptosis as defined by the above criter ia, regardless of whether apoptosis is induced by colchicine treatment or by serum withdrawal. Thus, the delay cannot simply be due to the w ell-known ability of MDR protein overexpression to lower chemotherapeu tic drug accumulation in MDR cells. LR73/27V500 ''selectants'' exhibit ing similar levels of MDR protein overexpression but higher multidrug resistance due to selection with the chemotherapeutic drag vincristine , exhibit a slightly longer delay in the progression of apoptosis. Nor mal apoptotic cascade kinetics are partially restored by pre-treatment of the MDR cells with the MDR protein inhibitor verapamil. Untransfec ted LR73 cells not expressing MDR protein but elevated in pH(i) via ma nipulation of CO2/HCO3- as described [Hoffman et al. (1996) J. Gen. Ph ysiol. 108, 295-313] are inhibited in DNA ladder formation, similar to LR73/hu MDR 1 transfectants. These results uncover an additional mech anism whereby MDR protein overexpression may promote the survival of t umor cells and further support the notion that in some systems intrace llular acidification may be either causal or permissive for proper pro gression of the apoptotic cascade.