Abstract
CD300f, a member of the CD300 family of immunoregulatory molecules, is capable of signalling through association with both SHP-1 phosphatase and the p85α subunit of phosphoinositide 3-kinase. On normal cells, CD300f is expressed on monocytes and dendritic cells in the blood and bone marrow. CD300f is expressed on myeloid derived cell lines and Acute Myeloid Leukaemias (AMLs) and is an acknowledged candidate for antibody targeting of AML (
Modra CJ et al. Blood 2006;108:225B-225B
and Zhao X et al Blood 2007;110:531a–532a
). We have generated a monoclonal antibody, MMRI- 23, specific for the extracellular domain of CD300f. MMRI-23 immunoprecipitates a 57kd protein from the myeloid derived cell lines HEL, THP-1 and U937 and this protein binds to a polyclonal antibody to CD300f (LMIR3) in Western blot analysis. Binding of MMRI-23 to myeloid cells was blocked by the CD300f recombinant proteins or the polyclonal CD300f antibody. The MMRI-23 mAb was used as a surrogate ligand to study the functional consequences of crosslinking CD300f on normal monocytes and myeloid derived cell lines. Purified peripheral blood monocytes were cultured for 18 hours in the presence of immobilised MMRI-23 or control mAb. MMRI-23 binding was not altered by activation of peripheral blood monocytes but crosslinking monocytes with MMRI- 23 induced downregulation of CD14 and CD33. There was significant inhibition of IL-6 but not IL-1β, IL-8 or TNFα secretion. Crosslinking monocytes or the U937 cell line with MMRI-23 increased specific chemotaxis towards CXCL12 (SDF-1). This increased migration index following MMRI-23 crosslinking was reversed in the presence of wortmannin indicating that MMRI-23 induces signalling through phosphoinositide 3-kinase. The effect of crosslinking did not enhance CXCR4 upregulation but did induce localization of CD300f and CXCR4 to the lipid rafts in myeloid cell line, U937. Thus CD300f plays a significant role in the regulation of monocyte migration to CXCR4. As CD300f is upregulated on around 70% of AMLs, this regulation of homing has major implications for the treatment of AML.Disclosures: No relevant conflicts of interest to declare.
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2008, The American Society of Hematology
2008
