Melphalan-Induced DNA Damage In Vitro as Predictor for Clinical Outcome in Multiple Myeloma.

Introduction: We have previously shown that measuring in vivo DNA adduct formation/repair in the p53 gene, in a readily accessible tissue such as peripheral blood mononuclear cells (PBMC) provides a non-invasive method for evaluating the effectiveness of high-dose melphalan (HDM) in MM. In the present study, we tested the hypothesis that quantification of p53-specific damage formation and repair induced in PBMC by in vitro melphalan treatment before therapy correlates with the respective data obtained in vivo, ie after administration of HDM. Furthermore, we studied whether this in vitro assay can predict outcome after HDM.

Patients and Methods: Thirty-two MM patients, with measurable disease, candidates for HDM and ASCT were included in the study. Response and progression were assessed according to the EBMT criteria. Prior to treatment with HDM, PBMC from patients were exposed in vitro to melphalan; subsequently, formation and repair of monoadducts and interstrand cross-links in the p53 gene were measured during the first 24 hours. The same studies were performed in PBMC-derived DNA following HDM administration to the same patients as previously reported (Dimopoulos et al, JCO 2005). Spearman’s correlation coefficient was used to assess correlation of in vivo to in vitro parameters in the p53 gene. Clinical response and time to progression were correlated with various molecular end-points using logistic regression and proportional hazards (Cox) models.

Results: Twenty-three patients (72%, responders) achieved complete (n=9) or partial response (n=14). Nine patients (28%, non-responders) did not have tumor reduction after HDM. During the follow-up period (median 15.4 months, range 2.4 to 26 months), 15 patients (47%) experienced disease progression.

Individual in vivo and in vitro values were, in general, highly correlated for all DNA adducts. The strongest correlation was observed for the Area Under the Curve of total adducts (AUC-TA, Spearman correlation coefficient: 0.93).

All in vitro molecular end-points indicative of increased DNA damage and slower repair capacity were predictive of a favorable response to HDM whilst AUC-TA had the highest predictive ability. Using the cut-off value of 736 adducts/10⁶nucleotidesxh for AUC-TA, the predictive value for clinical response to HDM was 100%. Moreover, patients with AUC-TA equal to or greater than this cut-off value had significantly longer time to progression than patients with a lower AUC-TA (hazard ratio 0.19, 95% confidence intervals 0.06 to 0.60).

Conclusion: We found that the extent of p53-specific damage formation/repair in PBMC from MM patients following in vitro exposure to melphalan correlates with the respective results obtained in vivo, i.e. after treatment with HDM, and is of value in predicting clinical response and progression free survival. Thus, this in vitro assay can be used to select those patients with MM who are more likely to benefit from HDM.