P-411 Ceratonia siliqua extract protects the ovarian reserve in a breast cancer mouse model treated with high-dose Cyclophosphamide

Abstract

Study question Can Ceratonia siliqua extract (CSE) mitigate cyclophosphamide (CPA)-induced ovarian damage in a breast cancer mouse model undergoing chemotherapy, without compromising the efficacy of tumor treatment? Summary answer Yes, Ceratonia siliqua extract, a potent antioxidant, protected the ovarian reserve by modulating Bax/Bcl-2-mediated apoptosis, while enhancing the anti-tumor effects of cyclophosphamide without adverse effects. What is known already Breast cancer is the most common cancer in women worldwide. Cytotoxic chemotherapy can lead to premature ovarian insufficiency and reduced ovarian reserve in survivors. While ART-based approaches effectively restore fertility, they fall short in long-term ovarian function maintenance. Infertility-related side effects can significantly impact survivors' quality of life. Antioxidant systems and redox homeostasis may rescue impaired AMH production in damaged ovarian follicles, maintaining reserves and preventing exhaustion. Ceratonia siliqua extract, a potent antioxidant, selectively protects normal cells from alkylating agent-induced damage, while exerting pro-apoptotic effects on cancer cells. Its success in preserving male fertility suggests potential for female fertility preservation. Study design, size, duration A controlled experimental study was conducted over five weeks using 6-week-old female BALB/c mice (n = 20). Mice were divided into 4 groups: Control, CSE-only, CPA-only, and CSE+CPA (n = 5 per group). An orthotopic 4T1 murine breast cancer model was established (tumor diameter = <5 mm). CSE (800 mg/kg) was administered orally for seven days before CPA chemotherapy and continued for 14 days after the final CPA dose. CPA (150 mg/kg) was given in two doses, eight hours apart. Participants/materials, setting, methods Mice were sacrificed at two time points: day 14 and day 21 post-CPA administration. On day 14, two mice were sacrificed for sample collection: blood serum for AMH measurement (ELISA), ovaries for histology and Bax/Bcl-2 qRT-PCR. Remaining mice underwent ovarian stimulation for IVF and sample collection on day 21. Blood serum and ovarian tissues were collected. Right ovarian tissue was used for histological follicle count, and left ovaries for qRT-PCR quantification of Bax/Bcl-2 expression levels. Main results and the role of chance CPA treatment significantly reduced ovarian reserve, as evidenced by a lower primordial follicle count (P < 0.05) and an increased number of growing follicles (P < 0.05), indicating enhanced follicular activation. Serum AMH levels were significantly higher in the CSE+CPA group compared to CPA-only (P < 0.05), suggesting a protective effect on granulosa cells. The number of MII oocytes in CSE+CPA mice was 2.6 times higher than in the CPA-only group (P < 0.01), with improved fertilization and blastocyst development rates (P < 0.05).Gene expression analysis showed increased Bax and decreased Bcl-2 expression in CPA-treated ovaries. In contrast, CSE+CPA treatment resulted in significantly downregulated Bax (P < 0.001) and upregulated Bcl-2 (P < 0.01), indicating reduced apoptosis. Tumor growth analysis demonstrated significant reduction across all experimental groups compared to the control group (CSE P < 0.05, CPA P < 0.01, CSE+CPA P < 0.001); however, there was no significant difference between CPA and CSE+CPA groups, confirming that CSE did not interfere with CPA’s anti-tumor effects. These findings suggest that CSE supplementation effectively mitigates CPA-induced ovarian damage, potentially through apoptosis regulation, and improves fertility outcomes without compromising cancer treatment efficacy. Limitations, reasons for caution Findings are based on a 4T1 murine breast cancer model. Cyclophosphamide doses used human equivalent calculations and an 8-hour interval to comply with 3R’s and ethical guidelines. Ceratonia siliqua extract dosage was derived from published studies. Further research is required to validate these results and explore underlying mechanisms. Wider implications of the findings Ceratonia siliqua mitigates cyclophosphamide-induced ovarian damage in mice, suggesting a promising, non-invasive strategy to protect ovarian reserve and fertility in young cancer patients. It may provide a novel fertility-preserving approach alongside chemotherapy without compromising efficacy. Further research on its safety, accessibility, and antioxidant properties is warranted. Trial registration number No