Anti-CD48 monoclonal antibody, clone OX-78

We have evaluated the antitumor activity of a murine antibody (IgG2a) against the leukocyte antigen CD48. CD48 is expressed on T and B lymphocytes, monocytes, and a wide range of lymphoid malignancies, To assess the therapeutic potential of an anti-CD48 antibody, me established a reproducible model of human B-cell (Raji) leukemia/lymphoma in C.B17/scid mice, where untreated mice develop hind leg paralysis due to tumor engraftment, Using this model, the murine anti-CD48 antibody HuLy-m3 was shown to mediate a strong in vivo antitumor effect, Long-term survival (>1 year) of scid mice was obtained after treatment with three 200-mu g i.v. doses of anti-CD48 antibody on days 0, 2, and 4 after i.v. injection of tumor cells, In contrast, mice treated with an isotype control antibody developed hind leg paralysis after 34 +/- 3 days, A strong antitumor response was still observed when a dose of 20 mu g of HuLy-m3 antibody was used, During preclinical investigations, we also examined a number of properties of the CD48 antigen, CD48 is present at high levels on the surface of T and B cells, but most (>95%) CD34-positive cells do not express CD48. Anti-CD48 antibodies are maintained on the surface of antigen-positive cells for extended periods (>24 h), These properties suggest that anti-CD48 antibodies may be useful in the treatment of a number of diseases including lymphoid leukemias and lymphomas.

Lipid rafts are cholesterol- and glycosphingolipid-rich plasma membrane microdomains, which control signal transduction, cellular contacts, pathogen recognition, and internalization processes. Their stability/lifetime, heterogeneity remained still controversial, mostly due to the high diversity of raft markers and cellular models. The correspondence of the rafts of living cells to liquid ordered (Lo) domains of model membranes and the effect of modulating rafts on the structural dynamics of their bulk membrane environment are also yet unresolved questions. Spatial overlap of various lipid and protein raft markers on live cells was studied by confocal laser scanning microscopy, while fluorescence polarization of DiIC18(3) and Bodipy-phosphatidylcholine was imaged with differential polarization CLSM (DP-CLSM). Mobility of the dil probe under different conditions was assessed by fluorescence correlation spectroscopic (FCS). GM1 gangliosides highly colocalized with GPI-linked protein markers of rafts and a new anti -cholesterol antibody (AC8) in various immune cells. On the same cells., albeit not fully excluded from rafts, diI colocalized much less with raft markers of both lipid and protein nature, suggesting the Lo membrane regions are not equivalents to lipid rafts. The DP-CLSM technique was capable of imaging probe orientation and heterogeneity of polarization in the plasma membrane of live cells, reflecting differences in lipid order/packing. This property-in accordance with dil mobility assessed by FCS-was sensitive to modulation of rafts either through their lipids or proteins. Our complex imaging analysis demonstrated that two lipid probes-G(M1) and a new anti-cholesterol antibody-equivocally label the membrane rafts on a variety of cell types, while some raft-associated proteins (MHC-II, CD48, CD59, or CD90) do not colocalize with each other. This indicates the compositional heterogeneity of rafts. Usefulness of the DP-CLSM technique in imaging immune cell surface, in terms of lipid order/packing heterogeneities, was also shown together with its sensitivity to monitor biological modulation of lipid rafts. (c) 2007 International Society for Analytical Cytology.