(329) Repurposing Drugs for Fertility Preservation

Marie-Charlotte Meinsohn^1,2, Josephine Cleverdon^1,2, Alana Mermin-Bunnell^1,2,3, David Pepin^1,2
1Pediatric Surgical Research Laboratories, Massachusetts General Hospital, Boston MA, USA
²Department of Surgery, Harvard Medical School, Boston, MA, USA
³Harvard-MIT Program in Health Sciences and Technology, Institute for Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, MA, USA

Abstract Text:

Women are born with a finite number of primordial follicles, forming their ovarian reserve. Natural depletion of this reserve leads to menopause around age 51, but chemotherapy can accelerate this process, posing a significant concern for young female cancer patients of reproductive age, including pediatric cases. Chemotherapy-induced depletion of the ovarian reserve can result in primary ovarian insufficiency (POI) and premature menopause, subsequently leading to cardiovascular, neurological, and bone health complications due to the loss of ovarian hormones. Current fertility preservation strategies, such as oocyte or embryo cryopreservation and ovarian cortex tissue freezing, offer reproductive options but do not prevent POI or restore long-term hormonal function. This underscores the need for a fertoprotective approach that safeguards both fertility and hormonal health in chemotherapy-treated patients.

Chemotherapy damages the ovarian reserve through two main mechanisms: inducing apoptosis in growing follicles and overactivating dormant primordial follicles, leading to their premature depletion. Understanding the regulation of follicle quiescence and activation is crucial for developing protective therapies. Anti-Müllerian hormone (AMH), a key regulator of ovarian function, is the only known hormone capable of inhibiting follicle activation. Studies have shown that recombinant AMH administration protects ovarian follicles in mice exposed to chemotherapeutic agents such as doxorubicin, cyclophosphamide, and carboplatin.

Our research aims to repurpose existing drugs that activate the AMHR2 pathway to preserve ovarian function. In a drug repurposing screen, we identified CYC116, an aurora kinase inhibitor, as a small-molecule agonist of AMHR2. CYC116 replicates AMH activity in vitro and suppresses follicle activation in vivo. We hypothesize that pharmacological modulation of the AMH pathway using CYC116 is a viable strategy to protect the ovarian reserve in chemotherapy-treated patients of reproductive age.

To validate this hypothesis, we conducted in vitro experiments using primary mouse granulosa cells, as well as immortalized rat (GFHSR17) and human (HGL5) granulosa cells. Cells were pretreated with CYC116 or AMH for 24h before a 24h exposure to doxorubicin. Results showed a significant reduction in double-strand DNA breaks (γH2AX-positive cells) and improved cell viability (CellTiter-Glo assay) in cells co-treated with CYC116 or AMH compared to chemotherapy alone. For in vivo validation, we used a prepubertal mouse model to mimic pediatric patients. Mice received doxorubicin (3 mg/kg) weekly for two weeks and were treated with either AMH (0.75 mg/kg, twice daily) or CYC116 (50 mg/kg, daily), starting one day before chemotherapy. Mice were euthanized 24 hours after the second doxorubicin injection, and ovarian follicles were counted.  We expect to show significant preservation of dormant follicles with CYC116 co-treatment, as previously demonstrated with AMH, confirming protection against chemotherapy-induced follicle overactivation.

To conclude, our findings indicate that CYC116 mimics AMH activity and helps preserve the ovarian reserve during chemotherapy by protecting activated granulosa cells against DNA double-strand break and apoptosis and has the potential to prevent overactivation of primordial follicles.