ALBERT

Description: Recent immunophenotypic analysis has shown that the heterogeneous expression of the adhesion molecule VLA-5 classifies myeloma cells into VLA-5+ mature and VLA-5- immature subpopulations. To further clarify the two myeloma subpopulations, we generated a monoclonal antibody, MPC- 1, by immunizing mice with an adherent human myeloma cell line, KMS-5. The MPC-1 antibody recognized a 48-Kd surface antigen on KMS-5 but not on U-266, a nonadherent human myeloma cell line. Specificity characterization showed that MPC-1 antigen was expressed on mature myeloma cells, normal plasma cells, and mature B cells, whereas pre-B cells and germinal center B cells lacked its expression. Monocytes and a human bone marrow stromal cell line, KM102, also expressed this antigen. Two subclones of MPC-1+ VLA-5+ (KMS-5Ad) and MPC-1-VLA-5+ (KMS- 5NAd) were separated from the KMS-5 cell line. The KMS-5NAd adhered to KM102 more tightly than did the KMS-5NAd, and the U-266 (MPC-1-VLA-5-) displayed almost no adherence to the KM102. The adhesion of the KMS-5Ad was partially inhibited by the MPC-1 antibody. These results, taken together, suggest that the MPC-1 antigen serves as a differentiation marker for B-lineage cells, including plasma cells, and may function as an adhesion molecule involved in the interaction of mature myeloma cells with bone marrow stromal cells.

Description: With regard to the expression of adhesion molecules, human myeloma cells freshly isolated from bone marrow were heterogeneous. By two- color analysis with anti-VLA-5 antibody (PE staining) and FITC-labeled anti-CD38 antibody, we found all myeloma cells located at CD38-strong positive (CD38++) fraction and identified two subpopulations among these myeloma cells: CD38++ VLA-5-(VLA-5-) myeloma cells and CD38++ VLA- 5+ (VLA-5+) myeloma cells. To clarify the biologic character of these two subpopulations, the morphology, in vitro proliferative activity and in vitro M-protein secretion were examined in each fraction isolated by the purification procedure or a cell sorter. Morphologic examination showed that VLA-5- myeloma cells were mostly immature or plasmablastic and VLA-5+ cells were mature myeloma cells. Furthermore, VLA-5- myeloma cells proliferated markedly in vitro and responded to interleukin 6 (IL- 6), a growth factor for myeloma cells, while VLA-5+ myeloma cells showed very low uptakes of 3H-thymidine and no responses to IL-6 but secreted higher amounts of M-protein (immunoglobulin) in vitro significantly. Therefore, we could clarify here heterogeneity of human myeloma cells in the bone marrow with regard to the expression of VLA- 5, one of integrin adhesion molecules; VLA-5- myeloma cells were proliferative immature cells and VLA-5+ cells were mature myeloma cells.

Description: We have recently shown that two-color analysis with fluorescein isothiocyanate (FITC)-anti-CD38 antibody could clearly distinguish myeloma cells (plasma cells) from other hematopoietic cells in the bone marrow. Myeloma cells (plasma cells) alone were located at CD38strong positive (++) fractions. To further distinguish normal plasma cells from mature myeloma cells phenotypically, we examined immunophenotypes of normal plasma cells and myeloma cells by two-color flow cytometry with FITC-anti-CD38 antibody and phycoerythrin staining with antibody to VLA-4, MPC-1, CD44, CD56, CD19, CD20, CD24, or CD10. Normal plasma cells were all VLA-4+VLA-5+MPC-1+CD44+ CD19+CD56- in the bone marrows from seven healthy donors, tonsils from four patients with chronic tonsillitis, a spleen from one patient with idiopathic thrombocytopenic purpura, and lymph nodes from two patients with chronic lymphadenitis, respectively. On the other hand, mature myeloma cells (12 of 20 cases), VLA-4+VLA-5+MPC-1+, were all CD19- and most of them CD56+, and there were no myeloma cells with the CD19+CD56- phenotype in the 20 cases of myelomas we tested. Thus, as for the expression of CD19 and CD56, normal plasma cells from various tissues are all CD19+CD56-, whereas no myeloma cells have the CD19+CD56- phenotype. According to this finding, we investigated the expression of CD19 and CD56 on plasma cells (CD38++ fractions) in monoclonal gammopathy of undetermined significance (MGUS). Both CD19+CD56- and CD19-DC56+ plasma cells were found in all five cases of MGUS we tested, suggesting that MGUS consists of phenotypically normal plasma cells and myeloma cells. Therefore, it is reasoned that phenotypic analysis of plasma cells with anti-CD19 and anti-CD56 antibodies can distinguish normal plasma cells from malignant plasma cells (myeloma cells), and can detect malignant plasma cells even in MGUS or premyeloma states.

Description: Recent immunophenotypic analysis has shown that the heterogeneous expression of the adhesion molecule VLA-5 classifies myeloma cells into VLA-5+ mature and VLA-5- immature subpopulations. To further clarify the two myeloma subpopulations, we generated a monoclonal antibody, MPC- 1, by immunizing mice with an adherent human myeloma cell line, KMS-5. The MPC-1 antibody recognized a 48-Kd surface antigen on KMS-5 but not on U-266, a nonadherent human myeloma cell line. Specificity characterization showed that MPC-1 antigen was expressed on mature myeloma cells, normal plasma cells, and mature B cells, whereas pre-B cells and germinal center B cells lacked its expression. Monocytes and a human bone marrow stromal cell line, KM102, also expressed this antigen. Two subclones of MPC-1+ VLA-5+ (KMS-5Ad) and MPC-1-VLA-5+ (KMS- 5NAd) were separated from the KMS-5 cell line. The KMS-5NAd adhered to KM102 more tightly than did the KMS-5NAd, and the U-266 (MPC-1-VLA-5-) displayed almost no adherence to the KM102. The adhesion of the KMS-5Ad was partially inhibited by the MPC-1 antibody. These results, taken together, suggest that the MPC-1 antigen serves as a differentiation marker for B-lineage cells, including plasma cells, and may function as an adhesion molecule involved in the interaction of mature myeloma cells with bone marrow stromal cells.

Description: With regard to the expression of adhesion molecules, human myeloma cells freshly isolated from bone marrow were heterogeneous. By two- color analysis with anti-VLA-5 antibody (PE staining) and FITC-labeled anti-CD38 antibody, we found all myeloma cells located at CD38-strong positive (CD38++) fraction and identified two subpopulations among these myeloma cells: CD38++ VLA-5-(VLA-5-) myeloma cells and CD38++ VLA- 5+ (VLA-5+) myeloma cells. To clarify the biologic character of these two subpopulations, the morphology, in vitro proliferative activity and in vitro M-protein secretion were examined in each fraction isolated by the purification procedure or a cell sorter. Morphologic examination showed that VLA-5- myeloma cells were mostly immature or plasmablastic and VLA-5+ cells were mature myeloma cells. Furthermore, VLA-5- myeloma cells proliferated markedly in vitro and responded to interleukin 6 (IL- 6), a growth factor for myeloma cells, while VLA-5+ myeloma cells showed very low uptakes of 3H-thymidine and no responses to IL-6 but secreted higher amounts of M-protein (immunoglobulin) in vitro significantly. Therefore, we could clarify here heterogeneity of human myeloma cells in the bone marrow with regard to the expression of VLA- 5, one of integrin adhesion molecules; VLA-5- myeloma cells were proliferative immature cells and VLA-5+ cells were mature myeloma cells.

Description: We have recently shown that two-color analysis with fluorescein isothiocyanate (FITC)-anti-CD38 antibody could clearly distinguish myeloma cells (plasma cells) from other hematopoietic cells in the bone marrow. Myeloma cells (plasma cells) alone were located at CD38strong positive (++) fractions. To further distinguish normal plasma cells from mature myeloma cells phenotypically, we examined immunophenotypes of normal plasma cells and myeloma cells by two-color flow cytometry with FITC-anti-CD38 antibody and phycoerythrin staining with antibody to VLA-4, MPC-1, CD44, CD56, CD19, CD20, CD24, or CD10. Normal plasma cells were all VLA-4+VLA-5+MPC-1+CD44+ CD19+CD56- in the bone marrows from seven healthy donors, tonsils from four patients with chronic tonsillitis, a spleen from one patient with idiopathic thrombocytopenic purpura, and lymph nodes from two patients with chronic lymphadenitis, respectively. On the other hand, mature myeloma cells (12 of 20 cases), VLA-4+VLA-5+MPC-1+, were all CD19- and most of them CD56+, and there were no myeloma cells with the CD19+CD56- phenotype in the 20 cases of myelomas we tested. Thus, as for the expression of CD19 and CD56, normal plasma cells from various tissues are all CD19+CD56-, whereas no myeloma cells have the CD19+CD56- phenotype. According to this finding, we investigated the expression of CD19 and CD56 on plasma cells (CD38++ fractions) in monoclonal gammopathy of undetermined significance (MGUS). Both CD19+CD56- and CD19-DC56+ plasma cells were found in all five cases of MGUS we tested, suggesting that MGUS consists of phenotypically normal plasma cells and myeloma cells. Therefore, it is reasoned that phenotypic analysis of plasma cells with anti-CD19 and anti-CD56 antibodies can distinguish normal plasma cells from malignant plasma cells (myeloma cells), and can detect malignant plasma cells even in MGUS or premyeloma states.