Anti-CD72 monoclonal antibody, clone 20-2.E3

Background: CD72 is an inhibitory co-receptor expressed on B cells. We previously demonstrated significant association of the polymorphism of the CD72 gene with susceptibility to human systemic lupus erythematosus (SLE) in individuals carrying a SLE-susceptible FCGR2B genotype (FCGR2B-232Thr/Thr). The human CD72 locus generates a splicing isoform that lacks exon 8 (CD72 Delta ex8) as well as full-length CD72 (CD72fl), and the CD72 polymorphism regulates exon 8 skipping. Results: Here we demonstrated that individuals carrying the disease-protective CD72 genotype exhibit significantly lower serum immunoglobulin levels than do individuals carrying other CD72 genotypes (P < 0.05). Although expression level of CD72fl in the peripheral blood B cells was similar regardless of CD72 genotype, the protein level of CD72 Delta ex8 was increased in individuals carrying the disease-protective CD72 genotype, suggesting a crucial role of CD72 Delta ex8 in regulation of antibody production. By expressing these human CD72 isoforms in mouse cell lines, we further demonstrated that CD72 Delta ex8 is accumulated in endoplasmic reticulum (ER) and fails to regulate BCR signaling whereas human CD72fl is efficiently transported to the cell surface and inhibits signaling through the B cell antigen receptor (BCR), as is the case for mouse CD72. Conclusion: Human CD72 polymorphism appears to regulate antibody production as well as susceptibility to SLE by regulating expression of ER-localizing CD72 Delta ex8.

CD72 has been reported to regulate BCR-mediated signals both positively and negatively. SHP-1 and Grb2 bind, respectively, to ITIM1 and ITIM2 of CD72. We generated transformed B cell lines with an immature phenotype following J2 virus infection of splenocytes from CD72(-/-) and wild-type (Wt) mice. The transformed lines were infected with retroviral vectors carrying Tyr (Y) to Phe (F) substitutions in the ITIM sequences (ITIM1 mutated: Y7/F; ITIM2 mutated: Y39/F; and both ITIM mutated: Y7,39/F). Crosslinking of the BCR induced growth inhibition in transfectants expressing Wt CD72, but this response was less sensitive in transfectants with Y7,39/F. The Y7/F transfectants demonstrated the least sensitive response. We were not able to obtain transfectants with Y39/F, suggesting that CD72 associated with SHP-1, but not with Grb2, delivers a strong negative signal. Pre-ligation of CD72, which induces dephosphorylation of the molecule, partially rescued the Wt transfectants from growth inhibition, leading to a growth response profile similar to that of Y7,39/F transfectants. These results suggest that ITIM1/SHP-1 delivers a very strong negative signal that is down-modulated by signals through ITIM2/Grb2, leading to delivery of an attenuated negative signal. Thus, preligation of CD72 results in the manifestation of an ostensible positive signal.