Detection of minimal residual disease by polymerase chain reaction in b cell malignancies

Abstract

It was the aim of this study to examine the prognostic value of the detection of minimal residual disease (MRD), with the help of the polymerase chain reaction (PCR), in patients with non-Hodgkin's lymphoma (NHL) and multiple myeloma (MM) who underwent sequential high-dose therapy with peripheral blood progenitor cell (PBPC) support, and in patients with acute myeloid leukemia (AML) of the subclass M4Eo who underwent high-dose consolidation therapy. Basis for the application of a PCR assay in these disease entities are the following specific gene rearrangements: the t(14;18) translocation in a high percentage of NHL, the clonal rearrangement of the Ig heavy chain locus resulting in a unique complementary determining region 3 (CDR3) for MM region and the inversion 16 characteristic for the M4Eo subclass of AML. Before the G-CSF-supported cytotoxic chemotherapy was given, 65% of the 52 patients with low- and intermediate-grade NHL enrolled into the study had PCR+ bone marrow (BM) and/or peripheral blood (PB) samples. The majority of patients (29 of 52) were autografted with a PCR+ transplant. The proportion of harvests containing t(14;18)+ cells was two-fold less in patients mobilized in first remission than in those with a history of previous treatment failure. This was also reflected when examining the B cell contents of the harvests measured as CD19+ cells with a 3.3-fold smaller proportion of CD19+ cells in leukapheresis (LP) products of patients mobilized in first remission. Patients who received a PCR- transplant are in remission and remained PCR- in BM and PB samples post-transplantation. Conversion to PCR-negativity in BM and PB samples post-transplantation was observed in 11 of 19 patients who were also in remission. In contrast, 6 of 29 patients who were autografted with PCR+ products relapsed, while 4 of them presented with PCR- samples on several occasions post-transplantation. In patients with MM, the assessment of MRD in PBPC harvests was based on the CDR3 regions of the Ig heavy chain locus as a marker for clonality. The great majority of LP products (17 out of 19) contained tumor cells. To prove positive enrichment procedures for the elimination of tumor cells, CD34+ and CD19+ cell fractions obtained from LP samples in an experimental setting via preparative flow cytometry were analyzed for MRD resulting in PCR-negativity for all CD34+ fractions. The results of the four patients with AML M4Eo and inversion 16 are preliminary, with a tendency of persistence of PCR-positivity after finishing the high-dose consolidation therapy. In one case, recurrence of disease was accompanied by an increase of the signal strength in the PCR assay. Longer follow-up periods are necessary to determine the prognostic value of these PCR findings in the different disease entities.

Mixed chimerism in the B cell lineage is a rapid and sensitive indicator of minimal residual disease in bone marrow transplant recipients with pre-B cell acute lymphoblastic leukemia

“…16,[37][38][39] Lately, MC detection of Ficoll-Hypaque density gradientseparated cell lineages has been shown to permit detailed monitoring of changes in donor/recipient cell dynamics in specific lineages following donor lymphocyte infusion (DLI) in a small number of CML patients. 40 Ramirez and co-workers 34 used Ficoll-gradient separated mononuclear vs granulocytic cell populations in patients with ALL, in an attempt to detect MC in the cell population specifically containing the leukemic clone.…”

Analysis of circulating tumor cells in patients with multiple myeloma during the course of high-dose therapy with peripheral blood stem cell transplantation

“…17,18 Briefly, cDNA was synthesized using Oligo d(T) 16 and 2 g of total RNA from a BM sample taken prior to HDT. Reverse-transcription was performed with Moloney murine leukemia virus reverse transcriptase (Perkin Elmer, Weiterstadt, Germany) at 42°C for 15 min.…”

Rituximab purging and maintenance combined with auto-SCT: long-term molecular remissions and prolonged hypogammaglobulinemia in relapsed follicular lymphoma

“…In addition, BM and PBSC harvests from patients with B-cell malignancies are recognized to frequently contain malignant cells. [18][19][20][21] Studies employing genetic markers have implicated these graft contaminants as a potential source of relapse post-auto-SCT. 22,23 Strategies to decrease FL relapse post-auto-SCT must reduce or eliminate both of these sources of lymphoma cells.…”