Abstract
Introduction: Melphalan reacts extensively with cellular DNA. DNA monoadducts (MA) and interstrand cross-links (ICL) are the main events occurring after drug exposure. Subsequently, complex pathways contribute to repair of DNA lesions. We have previously showed that individual repairing capacity in p53 gene in vivo varied up to 16-fold among pts with MM receiving HDM, while increased DNA damage and slower repairing capacity correlated with improved clinical outcome (Dimopoulos et al, JCO 2005). We examine here if measurement of gene-specific DNA damage formation and repair after in vitro exposure of peripheral blood lymphocytes (PBL) to melphalan correlates with in vivo DNA damage and repair after exposure to HDM and if in vitro findings correlate with subsequent clinical outcome.
Methods: Gene-specific MA and ICL formation and repair in the p53 gene were measured in PBL from MM pts, candidates for HDM (200mg/m2) and ASCT following in vitro exposure to 10 μg/ml melphalan for 1 h at 37°C. Measurements were performed 0, 2, 8 and 24 hours after in vitro exposure to melphalan. The same measurements were performed in vivo at 0, 2, 8 and 24 hours after treatment with HDM as previously described. Individual amounts of each type of DNA adducts over time (0–24h) were assessed by the area under the curve (AUC) during the whole experiment. Response after HDM was assessed according to the EBMT criteria. Measurements of DNA adducts after in vivo and in vitro treatment were correlated by the correlation-coefficient method.
Results: So far, in 25 pts in vitro measurements have been performed and in 15 pts in vivo measurements and correlations with clinical outcome were made as well. One pt was treated in relapse, 3 patients had primary refractory disease and 11 were in remission. Individual kinetics of melphalan-induced DNA damage formation and repair varied remarkably among patients both for the in vivo and in vitro measurements. A strong correlation between in vivo and in vitro measurements was found (p≤0.02 for all measurements). Patients were separated into 2 groups. Responders i.e. patients who achieved CR or PR (n=10) after HDM and non responders i.e. pts who were rated as SD or PD (n=5). A significant correlation of clinical response with p53 gene-specific damage formation and repair was found in both in vitro and in vivo data.
AUC (adducts/106 nucleotides x h) Total adducts (mean) Interstrand crosslinks (mean) Monoadducts (mean) Responders 919 +/− 215 303,9 +/− 108 616 +/− 154 In vitro Non responders 495 +/− 220 161,3 +/− 80,8 336 +/− 154 p-value 0,003 0,02 0,006 Responders 257 +/− 76 29,4 +/− 8,5 229 +/− 68 In vivo Non responders 122 +/− 91 13,8 +/− 4,6 112,8 +/− 82 p-value 0,009 0,002 0,01
Conclusion: Our results suggests that individuals with slower repairing capacity of the in vitro melphalan-induced p53 damage in peripheral blood lymphocytes have improved clinical outcome following subsequent treatment with HDM. We believe that our ongoing study may help select patients with MM who are more likely to benefit from HDM.
Repair of cis-platinum-DNA adducts by ABC excinuclease in vivo and in vitro.