Development and characterization of a long-acting recombinant hFSH agonist.
BACKGROUND
Fusion of the carboxyterminal peptide (CTP) of hCG to FSH results in a follitropin agonist with an extended half-life, presumably due to the four O-oligosaccharides on the CTP. Alternatively, an rhFSH analogue containing additional N-linked carbohydrate is described in this report.


METHODS
A DNA sequence containing two N-oligosaccharide signal sequences was ligated into a vector containing hFSHbeta- and alpha-subunit encoding cDNA, and expressed in CHO-K1 cells. In-vitro bioactivity of the single-chain hormone was assessed in CHO cells expressing the hFSH receptor. Pharmacokinetic values were derived from serial serum assays of the analogue in immature female rats following a single i.v. injection. In-vivo bioactivity was assessed by measuring ovarian weight gain 3 days post-injection.


RESULTS
rhFSH-N2 and native rhFSH induced comparable levels of cAMP in vitro. t(1/2) for native rhFSH, rhFSH-CTP and rhFSH-N2 were 3.7, 7.1 and 7.3 h respectively. Rats receiving rhFSH-N2 had a mean +/- SD ovarian weight 3 days post-i.v. injection (22 +/- 3.6 mg) significantly greater than rats receiving rhFSH and saline (16.7 +/- 1.5 and 15.3 +/- 0.47 mg respectively, P < 0.05).


CONCLUSIONS
rhFSH-N2 has prolonged half-life and increased bioactivity compared with native rhFSH. This rhFSH agonist, and other analogues containing additional N-oligosaccharides may have important clinical applications.
Klein
J
J
Department of Obstetrics and Gynecology and Center for Reproductive Sciences, School of Public Health, Columbia University, New York, NY 10032, USA.
Lobel
L
L
Pollak
S
S
Lustbader
B
B
Ogden
R T
RT
Sauer
M V
MV
Lustbader
J W
JW
D000818:Animals; D016466:CHO Cells; D002999:Clone Cells; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D005260:Female; D005640:Follicle Stimulating Hormone / Q000819:agonists* / Q000235:genetics*; D006207:Half-Life; D006801:Humans; D007275:Injections, Intravenous; D008596:Menotropins / Q000235:genetics*; D009929:Organ Size / Q000187:drug effects; D010053:Ovary / Q000033:anatomy & histology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000737:chemistry* / Q000494:pharmacology*; D013678:Technology, Pharmaceutical; D013997:Time Factors
D011993:Recombinant Fusion Proteins; D008596:Menotropins; D005640:Follicle Stimulating Hormone; D000242:Cyclic AMP