Analysis of B-lymphoid malignancies using allele-specific polymerase chain reaction: a technique for sequential quantitation of residual disease

Abstract

The junctional sequences corresponding to the complementarity determining region 3 (CDR3) of rearranged heavy chain Ig genes can provide allele-specific markers in the detection of B-lymphoid malignancies. Consensus oligonucleotide primers were used to amplify CDR3 regions of rearranged heavy chain alleles in clinical samples from myeloma, acute lymphocytic leukemia, and chronic lymphocytic leukemia patients. From the sequence of the amplified products, allele-specific primers were synthesized and used directly in polymerase chain reaction (PCR) amplification to detect the malignant clone. This method was both highly specific and sensitive to 1 malignant B-cell in a background of 10(5) normal cells. In addition, parameters that affect the linearity of PCR detection were determined and, by using titrations of malignant target cells to generate standard curves, quantitations of residual malignancies were determined. The application of this method is shown in an analysis of myeloma patients whose marrows were analyzed sequentially during therapy. Allele-specific oligonucleotide-PCR provided a rapid, highly specific and quantitative measure of residual disease, even in patients with clinical parameters indicating complete remission.

CD34 selections from myeloma peripheral blood cell autografts contain residual tumour cells due to impurity, not to CD34+ myeloma cells

“…Least squares was used to fit a linear regression equation for ln (OD) as a function of ln (tumour fraction) for each patient. The number of tumour cells in patient samples and its 95% Scheffe's confidence interval was computed using this patient-specific linear regression equation (Billadeau et al, 1991).…”

Multiple Myeloma: An Overview in 1996

“…When the M-protein is not detected in the serum and urine with immunofixation and the bone marrow contains no identifiable myeloma cells with immunofluorescence, the patient still frequently has relapse with myeloma of the same isotype that was present initially. Oligonucleotide primers to amplify regions of rearranged heavy-chain alleles with polymerase chain reaction can detect one myeloma cell in 100,000 cells [33].…”

Circulating plasma cells in multiple myeloma: characterization and correlation with disease stage

“…Consensus primer techniques, sensitive to 10 ¹3 -10 ¹4 , can detect the clone in approximately 30-50% of patients (Chiu et al, 1989;Owen et al, 1996a,b;Fend et al, 1993). More sensitive patient-specific PCR techniques (ASO-PCR), sensitive to 10 ¹5 -10 ¹6 (Billadeau et al, 1991), detect a clonal IgH rearrangement in 90-100% of patients (Corradini et al, 1993;Billadeau et al, 1992). These results suggest that the circulating myeloma cells are a minor population, representing fewer than 0·1% of the peripheral blood cells in at least half of the patients at presentation.…”