Post-transplantation tumour load in bone marrow, as assessed by quantitative ASO-PCR, is a prognostic parameter in multiple myeloma.

High-dose therapy (HDT) and autologous transplantation prolongs remission duration and survival in multiple myeloma (MM), but relapse still occurs at a median of 2 years post-HDT. In order to investigate whether the number of residual tumour cells in the bone marrow (BM) after transplantation can predict the duration of response, a quantitative allele-specific oligonucleotide polymerase chain reaction (qASO-PCR) assay was used to measure tumour load in BM at 3-6 months post-HDT in 67 patients. The method of maximally selected log-rank statistics was used to test for the existence of a cut-off value in the BM tumour load data set. A cut-off value with respect to progression-free survival (PFS) was identified (P = 0.001). The estimated threshold for placing patients into a "good" or "bad" prognostic group was 0.015% (n = 22 and 38 respectively) with a median PFS of 64 months vs. 16. Multivariate analysis showed grouping by PCR result to be an independent prognostic factor for PFS (estimated hazard ratio after shrinkage, 3.91). This study identifies for the first time a threshold of the post-HDT tumour load with prognostic significance for PFS in MM. Quantitative molecular assessment thus may help to identify those patients who are in need of further treatment early after autologous transplantation.

Myeloma clonotypic B cells are hampered in their ability to undergo B-cell differentiation in vitro.

In the peripheral blood (PB) of multiple myeloma (MM) patients, clonotypic B cells are present that express the identical V(D)J rearrangements as the malignant plasma cells in the bone marrow. In the present study, the proliferative capacity of clonotypic B cells from MM patients (n = 10) and the ability to differentiate in vitro was determined using the CD40-culturing system. For six patients, the presence of clonotypic B cells expressing variant immunoglobulin (Ig) isotypes was assessed by Ig isotype-specific allele-specific oligonucleotide reverse transcription polymerase chain reaction (ASO-RT-PCR) after culturing with CD40L and interleukin 4 (IL-4). In three out of six patients, clonotypic B cells expressing variant isotypes were detected both before and after culturing. The ability of clonotypic B cells to undergo B-cell differentiation was studied by abrogating CD40 signalling accompanied by IL-10 and IL-2 stimulation, enhancing differentiation towards Ig-secreting cells. The numbers of clonotypic B cells were determined by quantitative ASO-PCR. An increase in cell number was observed upon CD40L and IL-4 stimulation, whereas the relative number of clonotypic B cells was unaltered. In contrast, upon B-cell differentiation the relative number of clonotypic B cells decreased. In conclusion, clonotypic B cells can be cultured and isolated in vitro using the CD40 system. Clonotypic B cells responded to CD40 triggering in a similar fashion as to non-clonotypic normal B cells. However, the ability of clonotypic B cells to undergo in vitro activation and differentiation into Ig-secreting cells is hampered.