A novel long-acting recombinant follicle-stimulating hormone with hyperglycosylation exhibits improved pharmacokinetic and bioactivity on promoting follicle growth.

Follicle-stimulating hormone (FSH) plays an indispensable role in fertility. Although Recombinant human FSH (rh-FSH) and long-acting FSH analogs, such as FSH-CTP (Elonva), have been used to treat infertility for many years. The need for multiple injections remains inconvenient and uncomfortable. Therefore, long-acting FSH with optimized metabolic properties and bioactivity is urgently required. In this study, we designed a new long-acting FSH by fusing mucin domain II of CD164 to the β-subunit of FSH, which was then combined with the α-subunit, generating FSH164. SDS-PAGE and a series of liquid chromatography analyses showed the changed physicochemical properties, in which the MW of FSH164 was 2-fold greater than that of rh-FSH, and exceeded the MW of Elonva. Glycosylation analysis via ultra-performance liquid chromatography (UPLC) demonstrated an abundance of O-glycans and N-glycans, with sialic acid in the extremities. In vitro experiments suggested that FSH164 improved efficacy in addressing the attenuation of downstream signaling activation associated with sustained-action FSH modifications compared with Elonva. Pharmacokinetic results suggested that, compared with Elonva, FSH164 had a 1.5-fold longer half-life in rats. Furthermore, increased activities in terms of ovary weight gain and superovulations were verified in rats and mice. In conclusion, the designed hyperglycosylated FSH164 exhibits a prolonged metabolic duration, with a single dose of FSH164 possessing bioactivity comparable to that of multiple rh-FSH injections, suggesting that it has great potential as a therapy for infertility.