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Anti-CD40 antibody binding modulates human multiple myeloma clonogenicity in vitro
1Cancer Immunology Research Laboratory, Charles A. Sammons Cancer Center, Baylor University Medical Center, Dallas, TX 75246.
Insights
CD40 is expressed on some multiple myeloma cells, enhancing their growth. This study found CD40+ myeloma cells can increase colony formation independently of the IL-6 pathway.
Area of Science:
- Immunology
- Hematology
- Oncology
Background:
- Ligand binding to the B-cell antigen CD40 promotes B-cell growth and IL-6 secretion, forming the CD40/IL-6 loop.
- Interleukin-6 (IL-6) plays a critical role in the autocrine and paracrine growth of human multiple myeloma (MM) cells.
Purpose of the Study:
- To investigate CD40 expression in plasma cell dyscrasias (PCDs), including multiple myeloma (MM).
- To characterize the activity of the IL-6/CD40 loop in MM by examining CD40 binding effects on MM clonogenic colony (MCC) formation.
- To elucidate the interrelationship between CD40 activation and IL-6 production in PCD cell lines.
Main Methods:
- Bone marrow (BM) biopsies from patients with various PCDs were analyzed for CD40 expression on plasmacytoid cells.
- The effect of CD40 monoclonal antibody (MoAb) G28-5 binding on MCC formation was assessed in CD40+ and CD40- PCD BM cultures.
- PCD cell lines (RPMI 8226, ARH77, U266) were used to study CD40 expression, response to anti-CD40 stimulation, and IL-6 production/mRNA levels.
Main Results:
- CD40 was expressed on plasmacytoid cells in 21 of 28 PCD BM biopsies, including MM, Waldenstrom's macroglobulinemia, plasma cell leukemia, and monoclonal gammopathy of undetermined significance.
- G28-5 binding significantly increased MCC formation in 11 of 17 CD40+ PCD BM cultures, with responsiveness observed in MM, WM, PCL, and MGUS.
- Cell-line studies indicated that anti-CD40 stimulation increased MCC formation independently of IL-6, with no detectable IL-6 secretion or increased IL-6 mRNA transcription.
Conclusions:
- CD40 is expressed on a subset of human myeloma cells within various PCDs.
- CD40 engagement can enhance MM clonogenic colony formation.
- The CD40-mediated regulation of MM clonogenic colony formation may occur independently of the IL-6 pathway.
Abstract:
Ligand binding of the B-cell lineage antigen CD40 enhances growth and interleukin-6 (IL-6) secretion in human B cells (the CD40/IL-6 loop). IL-6 has an autocrine and paracrine role in human multiple myeloma (MM) cell growth. With the use of the CD40 monoclonal antibody (MoAb) G28-5, we examined CD40 expression and the effect of CD40 binding on MM clonogenic colony (MCC) formation to characterize the IL-6/CD40 loop activity in MM. CD40 was expressed on plasmacytoid cells in 21 of 28 plasma cell dyscrasia (PCD) bone marrow (BM) biopsies tested (10 of 14 MM, 2 of 2 Waldenstrom's macroglobulinemia [WM], 2 of 2 plasma cell leukemia [PCL], 6 of 8 monoclonal gammopathy of undetermined significance [MGUS], and 1 of 2 primary amyloidosis [AL]). G28-5 binding increased MCCs by 35% to 150% in 11 of 17 CD40+ PCD BM cultures, but did not affect MCC formation in CD40- specimens or normal BM colony forming units (CFU-GEMM, CFU-GM, BFU-E). Responsive cultures originated from BM of patients with MM (2 of 5 cases tested), WM (2 of 2), PCL (2 of 2), and MGUS (5 of 6). CD40-responsiveness was not significantly inhibited by the presence of an anti-IL-6 MoAb (2 of 2 MGUS cultures tested), and did not correlate with the capacity to respond to IL-6 stimulation (n = 17, P > .05) or a detectable level of endogenous IL-6 (n = 15, P > .05). Additional studies were performed with PCD cell lines to characterize the interrelationship of CD40 activation and IL-6 production. Fifty percent to greater than 95% of cells from the RPMI 8226 and ARH77 lines expressed CD40, whereas 6% of U266 cells were CD40+. For RPMI 8226, ARH-77, and U266 cells, the increased MCC formation after anti-CD40 stimulation was not affected by the presence of an anti-IL-6 neutralizing MoAb and was not accompanied by detectable IL-6 secretion. There was no apparent increase in IL-6 mRNA transcription following G28-5 treatment of U266 or RPMI 8226 cells. Our observations indicate that CD40 is expressed in a subset of human myeloma cells present in various PCDs. Cell-line studies suggest that the CD40+ myeloma cell may regulate MM clonogenic colony formation without activating the IL-6 pathway.