CRYSTAL-STRUCTURES OF A RAT ANTI-CD52 (CAMPATH-1) THERAPEUTIC ANTIBODY FAB FRAGMENT AND ITS HUMANIZED COUNTERPART

The CAMPATH-1 family of antibodies are able systematically to lyse hum an lymphocytes with human complement by targeting the small cell-surfa ce glycoprotein CD52, commonly called the CAMPATH-1 antigen. These ant ibodies have been used clinically for several years, providing therapy for patients with a variety of immunologically mediated diseases. We report here the first X-ray crystallographic analyses of a Fab fragmen t from a rat antibody, the original therapeutic monoclonal CAMPATH-1G and its humanized counterpart CAMPATH-1H, into which the six complemen tarity-determining regions of the rat antibody have been introduced. T hese structures have been refined at 2.6 Angstrom and 3.25 Angstrom re solution, respectively. The V-L domains of adjacent molecules of CAMPA TH-1H form a symmetric dimer within the crystals with an inter-molecul ar extended beta-sheet as seen in light chain dimers of the kappa clas s. Crystals of CAMPATH-1G have translational pseudo-symmetry. Within t he antibody-combining sites, which are dominated by the protrusion of Lys(H52b) and Lys(H53) from hypervariable loop H2, the charge distribu tion and overall integrity are highly conserved, but large changes in the position of loop H1 are observed and an altered conformation of lo op H2. The major determinants of this are framework residues H71 and H 24, whose identity differs in these two antibodies. These structures p rovide a detailed structural insight into the transplantation of an in tact antibody-combining site between a rodent and a human framework, a nd provide an increased understanding of the specificity and antigen a ffinity of this pair of CAMPATH-1 antibodies for CD52. This study form s the structural basis for future modification and design of more effe ctive antibodies to this important antigen. (C) 1998 Academic Press.