CHIMERIC HUMAN-MOUSE IGG ANTIBODIES WITH SHUFFLED CONSTANT-REGION EXONS DEMONSTRATE THAT MULTIPLE DOMAINS CONTRIBUTE TO IN-VIVO HALF-LIFE

Structural features that determine the differing rates of immunoglobul in catabolism are of great relevence to the engineering of immunologic ally active reagents. Sequences in the C(H)2 and C(H)3 region of IgG h ave been shown to regulate the rate of clearance through their interac tion with FcRn, In an attempt to probe additional structural features that regulate antibody half-life, we have investigated two families of chimeric antibodies, composed of identical murine heavy and light ant idansyl variable regions joined to human kappa light-chains and wild-t ype or shuffled human IgG heavy-chain constant regions. These antibodi es were iodinated, and their clearance was studied in severe combined immunodeficient mice hosts by whole-body radioactivity measurements. C learances of the wildtype and recombinant antibodies were biphasic. In a panel of immunoglobulins derived from IgG, and IgG,, as successive domains were varied from gamma(2) to gamma(3), beta-phase half-life gr adually decreased from 337.0 h to 70.6 h. Statistical analysis suggest ed that the composition of each of the three domains affected half-lif e, and no single region of the molecule by itself determined the rate of clearance. In the second panel of immunoglobulins derived from IgG( 1) and IgG(4), the construct with the amino terminus portion of the mo lecule derived from IgG(4), joined within the C(H)2 domain to the COOH terminus portion of IgG,, had a half-life paradoxically greater than either IgG(1) or IgG(4) (P < 0.012). All four IgG(1)/IgG(4) constructs demonstrated presence of the concentration catabolism phenomenon, whi ch is a unique hallmark of immunoglobulin catabolism. The contribution of all three constant region domains to immunoglobulin half-life may be due to distant conformational effects in addition to direct binding to protective receptors, and emphasizes the importance of distant seq uences on the rate of immunoglobulin catabolism. Interesting possibili ties regarding mechanisms controlling immunoglobulin metabolism are ra ised by the hybrid gamma(4)/gamma(1), molecule with a half-life greate r than either parental immunoglobulin. Understanding the relationships between the structure of these molecules and their clearance rate wil l further our ability to produce immunoglobulins with improved pharmac okinetic properties.