P-621 Rekovelle® vs Gonal-F®: in vitro characterization of intracellular signals induced by the two FSH preparations
C Lazzaretti, L Baschieri, E Mascolo, S Sperduti, C Perri, C Fusco, M Simoni, L CasariniAbstract
Study question
Do Rekovelle® and Gonal-F® modulate preparation-specific signalling?
Summary answer
Gonal-F® and Rekovelle® induce preparation-specific pattern of intracellular signalling.
What is known already
Rekovelle® and Gonal-F® are two commercial preparations of recombinant FSH used in assisted reproduction. These preparations differ in originator cells (PER.C6® cell line of human fetal retinal origin for Rekovlle®; modified Chinese Hamster Ovary cell line for Gonal-F®), likely reflecting glycosylation profiles impacting their mode of action.
Study design, size, duration
3-year in vitro functional comparison of Rekovelle® and Gonal-F®, using the transfected HEK293 cell line expressing FSH receptor (FSHR).
Participants/materials, setting, methods
FSHR-expressing HEK293 cells were treated with increasing concentrations of GonalF® and Rekovelle® (pM-µM) to evaluate receptor-receptor interaction and trafficking, intracellular cyclic adenosine monophosphate (cAMP) and Ca2+ increase by bioimaging techniques, as well as phospho-protein by Western blotting, and genomic effects by CRE-luciferase reporter gene activity.
Main results and the role of chance
Both the preparations induced similar, dose-dependent increase of receptor trafficking through intracellular organelles. These data were obtained by evaluation of the interaction between FSHR and endosomal markers RAB-GTPases (Rab) 5 and 7. Rekovelle® induced more pronounced steroifogenic signals than Gonal-F®, detected as FSHR-FSHR interactions at the cell membrane, cAMP accumulation, cAMP-responsive elements binding protein (CREB) phosphorylation and CRE promoter activity. Interestingly, while cell treatment by Gonal-F® mediated complete dose-response of cAMP, consisting in the continuous increase of the second messenger together with the increase of FSH concentration, Rekovelle® had biphasic action where cAMP increased from 0 to 100 nM although decreased at 1 µM Rekovelle® dose. Moreover, Rekovelle® was 10-times more potent than Gonal-F® in activating intracellular Ca2+ activation. The two preparations induced similar pattern of extracellular signal-regulated kinases 1 and 2 (ERK1/2) phosphorylation. In conclusion, Rekovelle® has higher potency than Gonal-F® in activating cAMP- and Ca2+-dependent intracellular events, potentially resulting in preparation-specific steroidogenic pattern.
Limitations, reasons for caution
Experiments were performed in the transfected HEK293 cell line in vitro and should be confirmed by clinical observations.
Wider implications of the findings
We described different intracellular events mediated by Gonal-F® and Rekovelle® in vitro, likely originated by different glycan profiles and possibly impacting the ovarian response during assisted reproduction.
Trial registration number
Not Applicable