The present investigation was performed to elucidate the role of
purine nucleotides as potential indicators of chemosensitivity of malignant
tumors.
Drug-sensitive (s) and -resistant (r) tumor cell lines grown as monolayers (s: T47D, MCF-7 wild-type; r: NCI/ADR-RES, MCF-7/MDR) or as multicellular spheroids (T47D; NCI/ADR-RES) were exposed to 0.1, 1.0, and 10.0 microM
Doxorubicin for up to 24 h.
Purine nucleotides were assayed using HPLC and with some selected spheroids using imaging bioluminescence. The data show that in the time frame of the experiments reproducible and statistically significant changes in the
nucleotides only occur at the highest
drug concentration investigated. Under these conditions and using monolayer cultures,
Doxorubicin caused a significant increase in
ATP and
GTP in sensitive but not in resistant
cancer cells. Consequently, this differential change may be exploited for
drug sensitivity testing in vitro.
Doxorubicin exposure to spheroids was associated with significant increases in
ATP and
GTP in both sensitive and resistant variants. However, the kinetic of the changes in
GTP was largely different between T47D and NCI/ADR-RES spheroids with a long-lasting, almost 3-fold elevation and a smaller, relatively short transient increase in
GTP, respectively. Supplementing experiments with
Doxorubicin treatment under inhibition of oxidative phosphorylation with
Oligomycin abolished the
drug-induced
ATP and
GTP peaks at persistent increases in
ADP and
AMP. Assuming that the spheroids may represent the in vivo situation to a better degree than monolayer cultures, experimental in vivo studies should clarify whether kinetic changes in
GTP could be used as differential markers for the chemosensitivity of solid
tumors. The experiments using
Oligomycin support the hypothesis that
purine nucleotides may be recycled from
DNA fragments that result from the interaction of the
drug with the
DNA strands.