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BEZ235-Mediated PI3K/mTOR dual inhibition improves ovarian follicle survival in a preclinical model
Jules Bindels1, Laëtitia Bernet1, Marlyne Squatrito1
1Laboratory of Biology of Tumor and Development, GIGA-Cancer, Université de Liège, 4000, Liège, Belgium.
Background:
Follicular loss after ovarian tissue cryopreservation and autotransplantation (OTCTP) remains a major challenge due to follicle activation and ischemia. We evaluated BEZ235, a dual PI3K/mTOR inhibitor, as a strategy to improve follicle survival in a preclinical model. Its effects were evaluated during ovarian culture, cryopreservation, and transplantation, including the potential benefit of post-grafting VEGF/G-CSF injections.
Methods:
Murine ovaries, organotipically cultured with chemotherapeutic treatment (4-HC, 2 µM), with or without supplementation with BEZ235 (1 µM), rapamycin (1 µM), LY294002 (25 µM), or AMH (200 ng/ml) were used to evaluate follicle activation. For cryopreservation studies, those inhibitors were added to the freezing medium, and pathways activation were assessed via Western blot. In vivo, ovaries cryopreserved with or without BEZ235 or rapamycin were autotransplanted under the kidney capsule of mice. A subset of mice received intraperitoneal VEGF/G-CSF injections for five days post-transplantation. Follicle quantification, proliferation and activation marker assessment, and fibrosis evaluation were performed three weeks post-grafting.
Results:
In vitro, BEZ235 significantly counteracted chemotherapy-induced activation of both Akt and mTOR pathways, whereas rapamycin and LY29400 inhibited only mTOR or Akt, respectively. Similarly, during cryopreservation, only BEZ235 significantly reduced activation of both pathways. AMH did not enhance BEZ235's protective effects. In vivo, ovaries slow-frozen with BEZ235 retained a higher percentage of primordial follicles and showed reduced follicle proliferation and activation compared to both control and rapamycin three weeks after transplantation. Additionally, post-grafting injection of VEGF/G-CSF did not further enhance follicle preservation or reduce fibrosis.
Conclusion:
Dual inhibition of PI3K/mTOR with BEZ235 provides superior protection of the primordial follicle pool by maintaining follicle dormancy, in both in vitro and in vivo models. These findings highlight BEZ235's potential to enhance OTCTP outcomes, extend graft longevity and improve fertility preservation strategies in women.
Insights
Dual PI3K/mTOR inhibition with BEZ235 protects primordial follicles during ovarian cryopreservation and transplantation. This strategy maintains follicle dormancy, improving fertility preservation outcomes.
Area of Science:
- Reproductive biology and cryobiology research.
- Preclinical models for fertility preservation.
Background:
- Follicular loss is a significant challenge in ovarian tissue cryopreservation and autotransplantation (OTCTP).
- Follicle activation and ischemia contribute to graft failure post-OTCTP.
Purpose of the Study:
- To evaluate BEZ235, a dual PI3K/mTOR inhibitor, for improving follicle survival during OTCTP.
- To assess the effects of BEZ235 in vitro, during cryopreservation, and in vivo post-transplantation.
Main Methods:
- Murine ovaries were cultured with chemotherapy and inhibitors (BEZ235, rapamycin, LY294002, AMH).
- Ovaries were cryopreserved with inhibitors, and pathway activation was assessed.
- Cryopreserved ovaries were autotransplanted, with some receiving VEGF/G-CSF injections post-transplantation.
Main Results:
- BEZ235 inhibited chemotherapy-induced Akt and mTOR activation in vitro.
- BEZ235 reduced pathway activation during cryopreservation.
- BEZ235-treated ovaries retained more primordial follicles and showed less activation post-transplantation; VEGF/G-CSF had no additional benefit.
Conclusions:
- Dual PI3K/mTOR inhibition with BEZ235 effectively protects the primordial follicle pool.
- BEZ235 maintains follicle dormancy, enhancing OTCTP outcomes and graft longevity.
- BEZ235 shows potential for improving fertility preservation strategies.
