Characterization of the CD30L binding domain on the human CD30 molecule using anti-CD30 antibodies

CD30 and its counter-receptor CD30 ligand (CD30L) are members of the TNF-receptor/TNF alpha superfamily and function to regulate lymphocyte survival and differentiation. Several monoclonal antibodies (MAbs) have been developed against CD30 and, based on mutual inhibition assays, are grouped into three nonoverlapping serologic clusters. However, the relationship between the epitopes recognized by the antibodies comprising each cluster and the binding domain for CD30L is not known. Using a soluble CD30L/CD8 alpha chimeric protein, we assessed the ability of anti-CD30 MAb to inhibit the binding of CD30L to CD30 expressed by the CD30+ Karpas 299 cell line, CD30L binding by CD30 is blocked by MAb that recognize epitopes belonging to cluster Group A (like Ber-HZ, Ber-HS, and HRS-4) as well as cluster Group C (like HeFi-1 and M44), Cluster Group B antibodies, including M67 and Ki-l, do not affect CD30L binding to CD30, The pattern of CD30L binding inhibition shows only limited correspondence to the functional capacity of some anti-CD30 MAb to trigger CD30 signaling. Finally, we demonstrate that the anti-CD30L MAb M81 also completely inhibits CD30/CD30L interaction. This information is useful for applying these MAbs in functional studies to further investigate the CD30/CD30L system and for designing assays for soluble CD30L.