Molecular methods for detection and quantification of myeloma cells after bone marrow transplantation: comparison between real-time quantitative and nested PCR
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Molecular methods for detection and quantification of myeloma cells after bone marrow transplantation: comparison between real-time quantitative and nested PCR. / Tögel, F; Kröger, N; Korioth, F; Fehse, B; Zander, A R.
In: J HEMATOTH STEM CELL, Vol. 11, No. 6, 12.2002, p. 971-976.Research output: SCORING: Contribution to journal › Short publication › Research › peer-review
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TY - JOUR
T1 - Molecular methods for detection and quantification of myeloma cells after bone marrow transplantation: comparison between real-time quantitative and nested PCR
AU - Tögel, F
AU - Kröger, N
AU - Korioth, F
AU - Fehse, B
AU - Zander, A R
PY - 2002/12
Y1 - 2002/12
N2 - Multiple myeloma is characterized by malignant plasma cell-infiltration of bone marrow. Treatment with high-dose therapy results in a high rate of clinical remissions, but almost all patients ultimately relapse. Clinical staging and detection of relapse are limited in sensitivity. Therefore, we established molecular methods based on the highly clone-specific CDR regions of the immunoglobulin VH locus for sensitive and specific detection of residual myeloma cells after bone marrow transplantation. VDJ rearrangements were identified using a set of VH primers and a JH primer. Clone-specific rearrangements were detected by comparison with germ-line sequences. With the nested PCR approach, first-round amplification with the consensus primers was done followed by second amplification with myeloma-specific primers. The real-time quantitative PCR was performed using a myeloma-specific forward primer in combination with a JH consensus TaqMan probe and reverse primer. Sensitivity was tested using dilutions of myeloma cell lines into mononuclear cells. Nested PCR had a sensitivity of 10(-6) and TaqMan PCR of 10(-4) to 10(-5). Specificity was determined by testing different cell lines and patients' probes. These results were confirmed by follow up of 2 patients after allogeneic transplantation with dose-reduced conditioning. Molecular methods are very sensitive and specific tools for follow up of myeloma patients after allogeneic transplantation. By using the quantitative approach, it is possible to see kinetics of bone marrow tumor load, which can be used to guide therapeutic decisions like donor leukocyte infusions (DLI).
AB - Multiple myeloma is characterized by malignant plasma cell-infiltration of bone marrow. Treatment with high-dose therapy results in a high rate of clinical remissions, but almost all patients ultimately relapse. Clinical staging and detection of relapse are limited in sensitivity. Therefore, we established molecular methods based on the highly clone-specific CDR regions of the immunoglobulin VH locus for sensitive and specific detection of residual myeloma cells after bone marrow transplantation. VDJ rearrangements were identified using a set of VH primers and a JH primer. Clone-specific rearrangements were detected by comparison with germ-line sequences. With the nested PCR approach, first-round amplification with the consensus primers was done followed by second amplification with myeloma-specific primers. The real-time quantitative PCR was performed using a myeloma-specific forward primer in combination with a JH consensus TaqMan probe and reverse primer. Sensitivity was tested using dilutions of myeloma cell lines into mononuclear cells. Nested PCR had a sensitivity of 10(-6) and TaqMan PCR of 10(-4) to 10(-5). Specificity was determined by testing different cell lines and patients' probes. These results were confirmed by follow up of 2 patients after allogeneic transplantation with dose-reduced conditioning. Molecular methods are very sensitive and specific tools for follow up of myeloma patients after allogeneic transplantation. By using the quantitative approach, it is possible to see kinetics of bone marrow tumor load, which can be used to guide therapeutic decisions like donor leukocyte infusions (DLI).
KW - Bone Marrow Transplantation
KW - Cell Count
KW - Follow-Up Studies
KW - Humans
KW - Molecular Diagnostic Techniques/standards
KW - Multiple Myeloma/diagnosis
KW - Neoplasm, Residual/diagnosis
KW - Polymerase Chain Reaction/methods
KW - Recurrence
KW - Reference Standards
KW - Sensitivity and Specificity
KW - Tumor Cells, Cultured
U2 - 10.1089/152581602321080637
DO - 10.1089/152581602321080637
M3 - Short publication
C2 - 12590712
VL - 11
SP - 971
EP - 976
JO - J HEMATOTH STEM CELL
JF - J HEMATOTH STEM CELL
SN - 1525-8165
IS - 6
ER -
