| Background: Several therapeutic approaches have been used to treat multiple myeloma (MM), including thalidomide and intensified high-dose chemotherapy regiments followed by autologous peripheral blood stem cell transplantation (APBSCT). Although these approaches have resulted in an increased number of patients achieving complete remissions, most patients still relapse within a few years after transplantation. Relapse is presumed to result from the persistence of small numbers of MM cells that cannot be detected using standard staging criteria. Polymerase chain reaction (PCR)-based studies have been used to determine the clinical significance of minimal residual disease (MRD) after intensive procedures. The most suitable clonal marker for MRD evaluation in MM is the clonal immunoglobulin H (IgH) rearrangement. This marker has been used extensively to examine the clinical significance of MRD detection in a variety of B-cell malignancies, including MM. Several studies have addressed quantitative analysis of MRD using the IgH rearrangement in MM, most often using end-point quantitation of PCR products and limiting dilution methods. The traditional quantitative and semi quantitative PCR approaches of end-product DNA analysis and limiting dilution strategies show a limited dynamic range, or give no reproducible results because extensive post-PCR processing is required of because potential bias occurs during late post exponential phases of PCR amplification.Recently, the introduction of hybridization probes, TaqMan probes and molecular beacon technology and the development of analyticalThermal cyclers have allowed development of extremely powerful systems for quantitation. The limitation of these approaches is that they require design of patient-specific probes. The high cost of each patient-specific probe severely limits such an approach for clinical studies involving large numbers of patients.In the present study, we used SYBR Green I dye and consensus primers for amplification of IgH rearrangement by real-time quantitative PCR in the peripheral blood of patients with MM treated with thalidomide or high-dose chemotherapy followed by APBSCT and assessed the utility of this approach in detecting the MRD of MM patients.Objective: To investigate the effect of real-time quantitative PCR with SYBR Green I dye and consensus primers detects IgH rearrangement in the peripheral blood of patients with MM treated with thalidomide or high-dose chemotherapy followed by APBSCT and assess the utility of this approach in detecting the MRD of patients with MM. So we can predict patients' outcomes and assess response to novel treatment strategies.Methods: Real-time quantitative PCR were performed to quantitate the IgH rearrangement on 26 patients with MM treated with thalidomide and 8 patients with MM treated with high-dose chemotherapy followed by APBSCT and used SYBE Green I dye and consensus IgH rearrangement primers.Results: The copies of IgH rearrangement are 3028 655.59, 43.4 24.85 and 7467 611.25, 619 271.2 in the patients with MM of completeremission group and partial responses group. They are 61531 527.02, 6575 1227.13 in the non responses group. Moreover, they have the positive relation with the proportion of plasma cells of the bone marrow. We obtained the same results from the patients with MM treated with high-dose chemotherapy followed by APBSCT. The copies are 3108 347.68 and 594 220.07 and they have the positive relation with proportion of plasma cells. IgH rearrangement of a patient was detected continuously and got the results parallelism with the clinical significance.Conclusion: The real-time quantitative PCR using SYBR Green I dye and consensus IgH rearrangement primers provides a feasible, accurate and reproducible method for evaluating MRD in MM and possibly in other differentiated B-cell malignancies. This technique can be used to assess tumor depletion and patient outcomes or response to novel treatment strategies.
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