Bivatuzumab

CD44 functions as a major hyaluronan receptor on most cell types, with roles in cell adhesion, migration, proliferation, differentiation, and survival. The CD44 gene comprises 20 exons, with alternative splicing producing many different isoforms. CD44 variant isoforms exhibit tissue-specific expression patterns and have been studied as therapeutic targets for several cancers; therefore, anti-CD44 monoclonal antibodies (mAbs) are useful for investigating CD44 expression in various cancers. Previously, we established an anti-CD44 mAb, C 44 Mab-46 (IgG 1 , κ), by immunizing mice with the CD44v3–10 ectodomain. Although C 44 Mab-46 recognized all CD44 isoforms, the binding epitope of C 44 Mab-46 has not been determined. In this study, we first checked the reactivity of C 44 Mab-46 to several CD44v3–10 deletion mutants such as dN79, dN124, dN147, and dN224. We found the N-terminus of the C 44 Mab-46-binding epitope between residues 147 and 224 of CD44v3–10. We next investigated this epitope using a novel mapping system: RIEDL insertion for epitope mapping (REMAP) method. We constructed 31 CD44 standard (CD44s) mutants where the RIEDL tag was inserted into the expected epitope region in CD44s. We observed that the C 44 Mab-46 epitope constituted five amino acids: 174 -TDDDV- 178 of CD44s. Thus, the REMAP method could be used to determine mAb binding epitopes for membrane proteins.

Acute promyelocytic leukemia (APL) was considered to be one of the most lethal forms of leukemia in adults before the introduction of the vitamin A metabolite all-trans retinoic acid (ATRA). Surprisingly, it has been confirmed that FICZ (6-Formylindolo (3, 2-b) carbazole) enhances ATRA-induced differentiation. Moreover, a number of studies have demonstrated that anti CD44 monoclonal antibody (mAb) induces to bring back differentiation blockage the leukemic stem cells. The level of differentiation markers including CD11b and CD11c in NB4 cells was assessed by flow cytometry. The induction of apoptosis was also evaluated. We estimated the induction potential of a triple compound of ATRA-FICZ, anti-CD44 maps. The cells showed the gradually increased expression levels of CD11b and CD11c. A mixture of a "CD44 mAb, ATRA and FICZ effectively promoted granulocytic maturation resulting in increased rates of apoptosis. The differences in expression of CD11b and CD11c at 5 μg/ml and 10 μg/ml were significant. These phenomena were highest at 10 μg/ml CD44 mAb concentrations. Synergistic induction differentiation and apoptosis of APL cells by using a co-treatment with novel triple compound are more effective for eradicating blasts and controlling the metastasis. Our results show that the addition of anti-CD44 mAb improves "ATRA-FICZ"-induced differentiation and has potential to reduce usual chemotherapy based treatments. Taken together, this compound may lead to novel clinical applications of differentiation-based approaches for APL and other types of leukemia. Further clinical studies would be recommended to clarify the clinical efficacy.