Autologous Stem Cell Transplantation Induces a Phenotypical Shift From CMP to GMP Progenitors, Reduces Clonogenic Potential and Enhances in Vitro and In Vivo Cycling Activity Defined by ¹⁸f-FLT PET Scan.

Abstract 4473

Autologous stem cell transplantation (ASCT) is a frequently applied treatment modality for patients with multiple myeloma (MM) and relapsing lymphoma. However, patients treated for relapsing disease post-ASCT demonstrate a reduced tolerance to chemotherapy, even those patients that have shown an adequate engraftment. These observations suggest long-term effects of the transplantation procedure on the bone marrow (BM) capacity. In the present study we analyzed in more detail the hematopoietic stem cell/progenitor defects in de BM of patients post-ASCT. BM cells from patients 6-9 months post-ASCT (n=16 including 5 patients with MM and 11 patients with relapsing lymphoma) were obtained after informed consent. At the moment of investigation patients displayed normal peripheral blood cell counts with a mean Hb level of 7.6 mmol/l, a mean leukocyte count of 6.9 × 10⁹/l, a mean granulocyte count of 4.4 × 10⁹/l and a mean platelet count of 202 × 10⁹/l. CD34+ cells were isolated from the BM material and analyzed by flowcytometry for progenitor subsets. Common myeloid progenitors (CMP) were defined as CD34+CD110-CD45RA-, granulocyte-macrophage progenitors (GMP) as CD34+CD110-CD45RA+ and megakaryocyte-erythroid progenitors (MEP) as CD34+CD110+CD45RA-. A phenotypical shift was observed from CMP (mean percentage 3.7% (95%CI 0.7-6.9) vs. 19.4% (95%CI 11.3-27.6), p=0.001) to GMP (mean percentage 51.8% (95%CI 39.6-64.2) vs. 27.6% (95%CI 19.6-35.5), p=0.01) in patients 6-9 months post-ASCT compared to CD34+ cells from healthy controls (n=7). No distinct difference in progenitor subsets was observed between MM and lymphoma patients. Comparable results were obtained in a limited number of patients at a later time point of follow-up (24 months post-ASCT). To further characterize the CD34+ cell fraction post-ASCT, progenitor frequencies were determined in CFC assays. A significant decrease in CFC frequency per 10³ plated CD34+ cells was observed compared to normal CD34+ BM cells (109 ± 52 vs. 185 ± 56, p=0.008). The decrease in CFC frequency from post-ASCT CD34+ cells was not due to an impaired proliferative activity. Cell cycle analysis revealed a significantly higher fraction of CD34+ cells post-ASCT in G2/S phase (mean 29% (95%CI 19.1-38.4) vs. 14% (95%CI 1.6-25.7), p=0.03) and a reduced percentage of cells in G1 phase (mean 67% (95%CI 56.4-78.2) vs. 86% (95%CI 73.9-98.4), p=0.03) compared to normal CD34+ cells. It appeared that especially the GMP fraction of post-ASCT CD34+ cells displayed a higher cycling activity (35% (95%CI 27.7-43.0) in G2/S phase vs. 19% (95%CI 13.0-25.7), p=0.03) whereas no significant differences for the MEP and CMP fractions were observed. The increased in vitro cycling activity could be confirmed in vivo by performing ¹⁸F-FLT PET scans in 10 patients post-ASCT. The results indicated a significant increase in standard uptake values (SUV) measured in the BM compartment. The mean SUV of the left and right area were compared to normal controls (n=12) and measured at different locations of the BM compartment including femur (3.6 ± 0.7 vs. 1.2 ± 0.5, p<0.001), crista (3.9 ± 0.6 vs. 2.2 ± 0.5, p<0.001) and spine (4.7 ± 2.5 vs. 3.9 ± 0.7, p<0.005). In addition, a significant expansion of the BM compartment of patients post-ASCT was noticed compared to normal controls. In summary, the results of this study demonstrate that ASCT results in long-term defects of the hematopoietic compartment characterized by a shift in progenitor composition from CMP to GMP, a reduced in vitro colony frequency and a higher in vitro and in vivo cycling activity of CD34+ BM cells of patients 6-9 months post-ASCT.