RECOMBINANT THYROTROPIN CONTAINING A BETA-SUBUNIT CHIMERA WITH THE HUMAN CHORIONIC GONADOTROPIN-BETA CARBOXY-TERMINUS IS BIOLOGICALLY-ACTIVE, WITH A PROLONGED PLASMA HALF-LIFE - ROLE OF CARBOHYDRATE IN BIOACTIVITY AND METABOLIC-CLEARANCE

Recombinant TSH is now successfully being used in clinical studies of thyroid cancer. Because of its therapeutic potential, we have construc ted a longer acting analog of TSH by fusing the carboxy-terminal exten sion peptide (CTEP) of hCG beta onto TSH beta. When coexpressed either with alpha-subunit complementary DNA or a minigene in African green m onkey (COS-7) and human embryonic kidney (293) cells, the chimera was fully bioactive in vitro and exhibited enhanced in vivo potency associ ated with a prolonged plasma half-life. The addition of 25 amino acids with 4 O-linked oligosaccharide chains did not affect the assembly an d secretion of chimeric TSH. Wild-type (WT) and chimeric TSH secreted by COS-7 and 293 cells displayed wide differences in their plasma half -lives, presumably due to the presence of terminal sialic acid and SO4 on their oligosaccharide chains, respectively. Chimeric and WT TSH se creted by both cell lines demonstrated similar bioactivity in cAMP pro duction, with some differences in [H-3]thymidine incorporation. Chimer ic TSH appears to be more effective in COS-7 cells than in 293 cells, as judged by growth assay. COS-7-produced chimeric TSH showed the maxi mum increase in half-life, indicating the importance of sialic acid in prolonging half-life and in vivo potency. Sulfation of both subunits, predominantly beta and to a lesser extent alpha, appears to be respon sible at least in part for the increased metabolic clearance of WT and chimeric TSH secreted by 293 cells. Apart from its therapeutic potent ial, chimeric TSH produced in various cell lines can be used as a tool to delineate the roles of sulfate and sialic acid in the in vivo clea rance and, thereby, the in vivo bioactivity.