Doxorubicin is an
anthracycline anticancer agent that functions primarily by inhibiting
topoisomerase II, but also forms covalent
DNA adducts depending on the cellular availability of
formaldehyde. The combination of
formaldehyde-releasing
prodrugs (such as
AN-9) with
doxorubicin has been shown to result in synergistic
doxorubicin-DNA adduct formation and synergistic apoptosis in HL-60 leukemic cells, offering the potential for lower concentrations of
doxorubicin to be used clinically in order to minimize side-effects. However, the overexpression of Bcl-2 confers resistance to
doxorubicin/
AN-9 DNA adduct forming treatments, thus limiting the therapeutic potential of this
drug combination. The small molecule inhibitor,
ABT-737, which binds to and inhibits Bcl-2, Bcl-xL and Bcl-w, was used in combination with
doxorubicin/
AN-9 treatments to overcome resistance to
doxorubicin-DNA adducts in Bcl-2 overexpressing HL-60 cells (HL-60/Bcl-2). The combination treatment of
doxorubicin and
AN-9 (and all single agent controls) failed to induce an apoptotic response in HL-60/Bcl-2 cells, however, the addition of low nanomolar (sub-lethal) concentrations of
ABT-737 was able to greatly increase apoptosis levels. Various control compounds were used to demonstrate that the mechanism of cell kill in response to the 'triple treatment' (
doxorubicin,
AN-9 and ABT-737) is dependent on
DNA adduct formation. Therefore, the ability of
ABT-737 to inhibit Bcl-2 renders previously resistant HL-60
cancer cells highly sensitive to
doxorubicin-DNA adducts, leading to a classical apoptotic response. In conclusion, the data obtained provides promising evidence that the anticancer activity of
doxorubicin-DNA adducts can be substantially enhanced in Bcl-2 overexpressing
cancers with the use of the small molecule Bcl-2 inhibitor,
ABT-737.