Significant Changes in Inducible Cytokine Synthesis by Activated Peripheral Blood Leukocytes of Multiple Myeloma Patients Receiving Standard Dose Autologous Stem Cell Transplantation

Abstract 5096

Multiple myeloma (MM) is an incurable cancer characterized by the clonal proliferation of plasma cells within the bone marrow. We previously reported a positive correlation between serum levels of inflammatory cytokines and the severity of the self-reported symptoms in MM patients undergoing high dose chemotherapy (HDC) and autologous stem cell transplantation (auto-SCT). As cytokines produced by activated peripheral blood (PB) leukocytes are likely to increase the severity of self-reported symptoms, we determined the ability of PB CD4⁺ and CD8⁺ T-lymphocytes, and CD14⁺ monocytes to produce cytokines following activation through the T-cell receptor (TCR) with anti-CD3 antibodies and toll-like receptor-4 with lipopolysaccharide (LPS), respectively.

We studied 24 patients (65 yrs ± 7 yrs) with Durie-Salmon stages II/III MM, 15 men and 9 women; 18 Caucasian, 2 Hispanic, and 3 African-American and 1 Asian. All patients were part of an ongoing clinical trial studying high versus standard dose of autologous stem cells (AutoSC) during HDC followed by auto-SCT. All patients received standard high dose melphalan (200 mg/m² in 2 divided doses) as their myeloablative chemotherapy. Fourteen subjects received a standard dose (SD) of AutoSC (4–6×10⁶ CD34+ cells/kg) and 10 patients received a high dose (HD) of AutoSC (10–15×10⁶ CD34+ cells/kg). The ability of TCR-activated CD4⁺ and CD8⁺ T cells to synthesize cytokines (Th1: IL-2, TNF-α, and IFN-γ; Th2: IL-4, and IL-10) and LPS-activated monocytes to synthesize IL-1β, IL-6, IL-8, IL-12, MCP-1, and TNF-a was assessed prior to auto-SCT and thereafter at 2 weeks, 1, 2, 6 and 12 months by multi-color flow cytometry assays.

Friedman test was used to determine the differences in the percentages of cytokine-producing cells by each cohort across time. Wilcoxon tests were used as follow-up tests to determine at which time the differences exist. Mann-Whitney test was used to determine if there were differences in the percentages of leukocytes producing cytokines in patients who received SD or the HD of AuSC.

The SD AuSC cohort had significant changes in the mean ± SEM percentages of TCR-activated CD3⁺CD8⁺ T cells that produced IL-2 and of CD3⁺CD4⁺ T cells that produced IL-10 across time. The increase in percentage of IL-2-producing T cells was followed by a significant decline (when compared with baseline) at 1-month post auto-SCT (14.5% ± 3.7% vs. 4.5% ± 1.3%; P = 0.019). This decline was sustained throughout the 12-month post auto-SCT and was attributable to CD3⁺CD8⁺ T cells (8.6% ± 2.3% vs 1.6% ± 0.4%; P = 0.013). Moreover, the SD cohort had an increase in the percentage of TCR-activated CD3⁺CD4⁺ T cells that synthesized IL-10 at 2 weeks post auto-SCT. Finally, the SD cohort also had a significant increase in the percentage of LPS-activated CD14⁺ monocytes that produced TNF-α at 3 months post auto-SCT when compared with LPS-activated CD14⁺ monocytes at baseline (0.6% ± 0.4% vs. 1.6% ± 0.4%; P = 0.006), and this increase was sustained over the remainder of 12-month period. These dynamic changes in cytokine production were not seen in the HD AuSC group. Moreover, there were no significant differences in the percentages of activated leukocytes synthesizing cytokines at any time point between the SD and HD AuSC groups.

These preliminary results suggest that activated leukocytes from MM patients who received the SD of AuSC had a dysregulated cytokine profile while that of patients who received HD of AuSC did not. These significant changes may be associated with more variations in symptom burden of the SD cohort. Further analyses to correlate these findings with symptom burden are warranted to support this hypothesis.