sjg-136 and Leukemia--Lymphocytic--Chronic--B-Cell

In this study, we set out to establish whether fludarabine could enhance the DNA interstrand crosslinking capacity of SJG-136 in primary human chronic lymphocytic leukaemia (CLL) cells and thereby offer a rationale for its clinical use in combination with SJG-136. SJG-136 rapidly induced DNA crosslinking in primary CLL cells which was concentration-dependent. Further, the level of crosslinking correlated with sensitivity to SJG-136-induced apoptosis (P=0.001) and higher levels of crosslinking were induced by the combination of SJG-136 and fludarabine (P=0.002). All of the samples tested (n=40) demonstrated synergy between SJG-136 and fludarabine (mean combination index (CI)=0.54+/-0.2) and this was even retained in samples derived from patients with fludarabine resistance (mean CI=0.62+/-0.3). Transcription of the excision repair enzyme, ERCC1, was consistently increased (20/20) in response to SJG-136 (P<0.0001). In contrast, fludarabine suppressed ERCC1 transcription (P=0.04) and inhibited SJG-136-induced ERCC1 transcription when used in combination (P=0.001). Importantly, the ability of fludarabine to suppress ERCC1 transcription correlated with the degree of synergy observed between SJG-136 and fludarabine (r(2)=0.28; P=0.017) offering a mechanistic rationale for the synergistic interaction. The data presented here provides a clear indication that this combination of drugs may have clinical utility as salvage therapy in drug-resistant CLL.

Topics: Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Benzodiazepinones; Cross-Linking Reagents; DNA; DNA Repair; DNA-Binding Proteins; Drug Synergism; Endonucleases; Female; Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Male; Middle Aged; Pyrroles; Transcription, Genetic; Tumor Cells, Cultured; Vidarabine

SJG-136 (NSC 694501) is a novel DNA cross-linking agent that binds in a sequence-selective manner in the minor groove of the DNA helix. It is structurally novel compared with other clinically used DNA cross-linking agents and has exhibited a unique multilog differential pattern of activity in the NCI 60-cell line screen (i.e., is COMPARE negative to other cross-linking agents). Given this profile, we undertook a preclinical evaluation of SJG-136 in primary tumor cells derived from 34 B-cell chronic lymphocytic leukemia (B-CLL) patients. SJG-136 induced apoptosis in all of the B-CLL samples tested with a mean LD50 value (the concentration of drug required to kill 50% of the cells) of 9.06 nmol/L. Its cytotoxicity was undiminished in B-CLL cells derived from patients treated previously, those with unmutated VH genes, and those with p53 mutations (P=0.17; P=0.63; P=0.42, respectively). SJG-136-induced apoptosis was associated with the activation of caspase-3 that could be partially abrogated by the caspase-9 inhibitor Z-LEHD-FMK. Furthermore, SJG-136 did not trigger the phosphorylation of p53 or the up-regulation of GADD45 expression in B-CLL cells whereas the cross-linking agent chlorambucil elicited both of these effects. This suggests that SJG-136 cross-linking adducts are not subject to p53-mediated DNA excision repair mechanisms in B-CLL cells. Taken together, these data demonstrate a novel mechanism of action for SJG-136 that appears to circumvent the effects of poor prognostic markers. This unique cytotoxicity profile warrants further investigation and supports the evaluation of this agent in Phase I clinical trials for patients with B-CLL.

Topics: Apoptosis; B-Lymphocytes; Benzodiazepines; Benzodiazepinones; Case-Control Studies; Caspase 3; Caspases; Cross-Linking Reagents; DNA Damage; DNA Repair; Enzyme Activation; Humans; K562 Cells; Leukemia, Lymphocytic, Chronic, B-Cell; Pyrroles; T-Lymphocytes; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Vidarabine