Cell culture production of a recombinant bovine follicle-stimulating hormone (rbFSH) and a hyperglycosylated variant

VILLARRAZA, CARLOS JAVIER; TARDIVO, MARÍA BELÉN; ANTUÑA, SEBASTIÁN; DÍAZ, PABLO U.; ORTEGA, HUGO H.; PRIETO, CLAUDIO; CEAGLIO, NATALIA

Santa Fe

Simposio; IX Simposio Latinoamericano de Tecnología de Cultivos Celulares - IX SLATCC 2022; 2022

Centro Biotecnológico del Litoral - FBCB - UNL

Porcine follicle-stimulating hormone (pFSH) is a glycoprotein hormone used to induce superovulation in cows. The only product available on the market consists of partially purified pFSH preparations from pituitary glands comprising many disadvantages, including batch to batch variations, possible contamination with luteinizing hormone and an extremely short circulating half-life. For these reasons, the development of a recombinant bovine FSH (rbFSH) represents a powerful strategy to substitute the pituitary-derived preparations. The aim of this work was to develop a biotechnological process for the production of rbFSH and a hyperglycosylated long-lasting variant (rbFSH-LD) both fused to a His-tag and expressed in suspension CHO-K1 (sCHO-K1) cells cultured in serum-free medium (SFM).Cell lines were generated through lentiviral transgenesis of sCHO-K1 cells. After adaptation to SFM, cells were cultured in a one-liter stirred tank bioreactor for 18 days. rbFSH and rbFSH-LD proteins were purified from culture supernatants by immobilized metal affinity chromatography (IMAC). Both proteins eluted with high purity levels (>95%) with traces of free  and  subunits as main contaminants. The biopotency of purified hormones was evaluated in female rats through the Steelman and Pohley bioassay using a recombinant human FSH (rhFSH) as a reference preparation. These preliminary assays showed that both hormones display FSH activity in experimental animals. Moreover, the specific biological activity of rbFSH-LD was similar to the one obtained for non-modified rbFSH, suggesting that the addition of new glycosylation sites to the molecule did not alter the hormone´s function.