Residual malignant and normal plasma cells shortly after high dose melphalan and stem cell transplantation. Highlight of a putative therapeutic window in Multiple Myeloma?

Abstract

Multiple Myeloma (MM) is an incurable malignant plasma cell disorder. We have evaluated the counts of Multiple Myeloma Cells (MMCs) and normal plasma cells (N-PCs), seven days after high-dose melphalan (HDM) and autologous stem transplantation (ASCT). Two third of patients had detectable minimal residual disease (MRD+) (71.7 MMCs/µL) after induction treatment with dexamethasone and proteasome inhibitor. MMC counts were reduced by 92% (P ≤ .05) but not eradicated 7 days after HDM+ASCT. Post-HDM+ASCT MMCs were viable and bathed in a burst of MMC growth factors, linked with post-HDM aplasia. In one third of patients (MRD- patients), MMCs were not detectable after induction treatment and remained undetectable after HDM+ASCT. Major difference between MRD- and MRD+ patients is that N-PC counts were increased 3 fold (P〈.05) by HDM+ASCT in MRD- patients, but were unaffected in MRD+ patients. Possible explanation could be that clearance of MMCs in MRD- patients makes more niches available for N-PCs. Thus, MMCs are not fully eradicated shortly after HDM, are bathed in high concentrations of MMC growth factors in an almost desert BM, are viable in short-term culture, which suggests providing additional therapies shortly after HDM to kill resistant MMCs before full repair of lesions.

Figure 1. CONSORT diagram of patients with previously-untreated multiple myeloma in Montpellier University Hospital, showing number of patients, treatments delivered, and outcome

MRD, minimal residual disease; HDM, high dose melphalan; ASCT, autologous hematopoietic stem cells transplantation; MFC, multiparameter flow cytometry.

Figure 2. Assessment of Multiple Myeloma Cells and Normal Plasma cells in representative patients with Multiple Myeloma before and after high dose melphalan

Using 7 color-multiparameter flow cytometry, multiple myeloma cells (MMCs) and normal plasma cells (N-PCs) were assessed in bone marrow or peripheral blood samples of patients after induction treatment (1 day before high dose melphalan, HDM), 7 days after HDM and autologous hematopoietic stem cell transplantation (ASCT), 3 months after HDM. MMCs and N-PCs were also measured in the thawed stem cell leukapheresis product grafted to the patients. (A) MMC and N-PC evaluation in a representative patient with positive residual disease after induction treatment (MRD⁺). MMCs were identified on the basis of aberrant CD56 expression and monoclonal lambda light chain expression. Data are the dotplots of CD56 and Lambda light chain expressions. MMCs were undetectable 7 days and 3 months after HDM+ASCT in the peripheral blood. (B) Bone marrow cells from one representative MRD⁺ patient out of 3 were harvested 7 days after HDM+ASCT and cultured for 6 days with 2 ng/mL of IL-6. Before culture, bone marrow cells contained CD38^high CD56⁺ MMCs (70% of CD38^high cells) and CD38^highCD56⁻ N-PCs. After a 6-day, the CD38^high CD56⁺ MMCs were viable, being Annexin V⁻ 7AAD⁻. (C) N-PC evaluation and lack of MMCs in a representative patient with negative residual disease after induction treatment. MMCs of this patient aberrantly expressed CD56 and CD200 at diagnosis. After induction treatment, only N-PCs could be detected in the bone marrow, peripheral blood and leukapheresis product.

Figure 3. Cell communication proteins in bone marrow plasma before and 7 days after high dose melphalan (HDM) and autologous stem cell transplantation (ASCT)

Paired bone marrow plasmas from 17 patients were collected 1 day before and 7 days after HDM+ASCT (respectively black circles and open circles) and assayed for cell communication proteins using Luminex methodology or ELISA. Data are the concentrations of IL-6, BAFF, CCL2, VEGF, IL-8, IL-15, IL-5, G-CSF, CCL3, CCL5 and IL-12 medullary plasma concentrations measured for each of the 17 patients. Horizontal bars represent the median value. The * indicates that the data before and after HDM are significantly different (P ≤ 0.05) using a paired t-test (when normality test passed) or Wilcoxon signed rank test (when normality test failed).