Faculty Bibliography

PURPOSE/OBJECTIVE:To investigate the differences concerning post-thawing/warming follicle survival, DNA damage and apoptosis in human ovarian tissues cryopreserved by slow freezing, open, or closed vitrification methods. METHODS:A total of 50 pieces of 5 × 5 × 1 mm ovarian cortical pieces were harvested (5 donor ovaries; mean age 31 ± 6.62 years). From each donor, one cortical piece was used as baseline; the remaining were randomly assigned to slow freezing (SF), vitrification using open device (VF-open), or closed device (VF-closed) groups. After 8-10 weeks of cryostorage, tissues were evaluated 4 h after thawing/warming. Histological analysis was evaluated for follicle survival (primordial and primary follicle densities) by H&E staining. The percentages of primordial and primary follicles with DNA double-strand breaks (γH2AX) and apoptotic cell death pathway activation (AC3) were immunohistochemically assessed. Data were analysed using one-way ANOVA and LSD post hoc comparison. RESULTS:Compared to the baseline, primordial follicle (pdf) densities significantly declined in all cryopreserved groups (SF, VF-open, and VF-closed, P < 0.05). However, the total and non-apoptotic pdf densities were similar among SF, VF-open, and VF-closed. SF and VF with either open or closed devices did not increase the percentages of primordial or primary follicles with DNA double-strand breaks (DSBs) or apoptosis compared to the baseline or among the freezing methods in the present study. CONCLUSION/CONCLUSIONS:Based on the intact primordial follicle survival, DNA damage, and apoptosis rates after thawing/warming, SF vs VF with either open or newly developed closed devices appear to be comparable.

Ovarian tissue cryopreservation and autotransplantation can restore ovarian endocrine function and fertility and recently were changed from experimental to fertility preservation procedures for medical indications by the American Society of Reproductive Medicine. Such advances have resulted in discussions around the utility of ovarian cryopreservation in healthy women to preserve fertility and delay menopause or as a hormone replacement approach. Such 'elective' use of ovarian tissue cryopreservation requires a risk-benefit assessment. Here, we review evidence for and against the utility of ovarian tissue harvesting in healthy women, scrutinize recent and needed advances to enhance the feasibility of such an approach, and provide practice and future research guidelines.

PURPOSE/OBJECTIVE:) pathogenic variants are associated with decreased ovarian reserve. MATERIALS AND METHODS/METHODS:four centers studied those affected with breast cancer (n = 161) and one studied unaffected individuals (n = 89). The data were adjusted for the center, age, body mass index, smoking, and oral contraceptive pill use before the final analysis. Anti-Müllerian hormone (AMH) levels in affected women were drawn before presystemic therapy. RESULTS:= .64). CONCLUSION/CONCLUSIONS:, have lower serum AMH levels compared with controls. They may need to be preferentially counseled about the possibility of shortened reproductive lifespan because of diminished ovarian reserve.

PURPOSE/OBJECTIVE:To determine the longitudinal impact of adjuvant chemotherapy and tamoxifen-only treatments on the reproductive potential of women with breast cancer by using a sensitive ovarian reserve marker anti-Mullerian hormone (AMH) as a surrogate. METHODS:One-hundred-and-forty-two women with a primary diagnosis of breast cancer were prospectively followed with serum AMH assessments before the initiation, and 12, 18 and 24 months after the completion of adjuvant chemotherapy or the start of tamoxifen-only treatment. The chemotherapy regimens were classified into Anthracycline-Cyclophosphamide-based (AC-based) and Cyclophosphamide-Methotrexate + 5-Fluorouracil (CMF). Longitudinal data were analyzed by mixed effects model for treatment effects over time, adjusting for baseline age and BMI. RESULTS:Both chemotherapy regimens resulted in significant decline in ovarian reserve compared to the tamoxifen-only treatment (p < 0.0001 either regimen vs. tamoxifen for overall trend). AMH levels sharply declined at 12 months but did not show a significant recovery from 12 to 18 and 18 to 24 months after the completion of AC-based or CMF regimens. The degree of decline did not differ between the two chemotherapy groups (p = 0.53). In contrast, tamoxifen-only treatment did not significantly alter the age-adjusted serum AMH levels over the 24-month follow up. Likewise, the use of adjuvant tamoxifen following AC-based regimens did not affect AMH recovery. CONCLUSIONS:Both AC-based regimens and CMF significantly compromise ovarian reserve, without a recovery beyond 12 months post-chemotherapy. In contrast, tamoxifen-only treatment does not seem to alter ovarian reserve. These data indicate that the commonly used chemotherapy regimens but not the hormonal therapy compromise future reproductive potential.

Among the investigated mechanisms of chemotherapy-induced damage to human primordial follicle reserve are induction of DNA double-strand breaks (DSBs) and resultant apoptotic death, stromal-microvascular damage and follicle activation. Accumulating basic and translational evidence suggests that acute exposure to gonadotoxic chemotherapeutics, such as cyclophosphamide or doxorubicin, induces DNA DSBs and triggers apoptotic death of primordial follicle oocytes within 12-24 h, resulting in the massive loss of ovarian reserve. Evidence also indicates that chemotherapeutic agents can cause microvascular and stromal damage, induce hypoxia and indirectly affect ovarian reserve. While it is possible that the acute reduction of the primordial follicle reserve by massive apoptotic losses may result in delayed activation of some primordial follicles, this is unlikely to be a predominant mechanism of loss in humans. Here, we review these mechanisms of chemotherapy-induced ovarian reserve depletion and the potential reasons for the discrepancies among the studies. Based on the current literature, we propose an integrated hypothesis that explains both the acute and delayed chemotherapy-induced loss of primordial follicle reserve in the human ovary.

OBJECTIVE:To assess whether woman who have BRCA mutations (WBM) experience more declines in ovarian reserve after chemotherapy treatment, as it induces oocyte death by deoxyribonucleic acid (DNA) damage, and BRCA mutations result in DNA damage repair deficiency. DESIGN/METHODS:Longitudinal cohort study. SETTING/METHODS:Academic centers. PATIENT(S)/METHODS:Of the 235 enrolled, 108 evaluable women with breast cancer were stratified into those never tested (negative family history; n = 35) and those negative (n = 59) or positive (n = 14) for a pathogenic BRCA mutation. INTERVENTION(S)/METHODS:Sera were longitudinally obtained before and 12-24 months after chemotherapy tratment, assayed for antimüllerian hormone (AMH), and adjusted for age at sample collection. MAIN OUTCOME MEASURE(S)/METHODS:Ovarian recovery, defined as the geometric mean of the after chemotherapy age-adjusted AMH levels compared with baseline levels. RESULT(S)/RESULTS:Compared with the controls, the before chemotherapy treatment AMH levels were 24% and 34% lower in those negative or positive for BRCA mutations, consistent with accelerated ovarian aging in WBM. The WBM had a threefold difference in AMH recovery after chemotherapy treatment (1.6%), when compared with BRCA negative (3.7%) and untested/low risk controls (5.2%). Limiting the analysis to the most common regimen, doxorubicin and cyclophosphamide followed by paclitaxel, showed similar results. These findings were mechanistically confirmed in an in vitro mouse oocyte BRCA knockdown bioassay, which showed that BRCA deficiency results in increased oocyte susceptibility to doxorubicin. CONCLUSION(S)/CONCLUSIONS:Women who have pathogenic BRCA mutations are more likely to lose ovarian reserve after chemotherapy treatment, suggesting an emphasis on fertility preservation. Furthermore, our findings generate the hypothesis that DNA repair deficiency is a shared mechanism between aging, infertility, and cancer. CLINICAL TRIAL REGISTRATION NUMBER/BACKGROUND:NCT00823654.

The loss of fertility and early menopause are common after gonadotoxic therapies and radical pelvic surgery. The strategy of ovarian tissue cryopreservation and auto-transplantation was introduced to prevent this significant quality of health issue. Ovarian transplantation with cryopreserved tissue has gone through remarkable evolution in the last 20 years. In this review, we detail the history and evolution of ovarian transplantation with cryopreserved tissue from its origins to the present. Ovarian cryopreservation and transplantation approach was first tested with animal models. The approach was then validated in human ovarian xenografting models before being applied to patients in pioneering clinical studies. The first orthotopic and heterotopic approaches to ovarian transplantation was developed by Oktay et al. who reported the first successful restoration of ovarian function with these approaches beginning in 2000 with first embryo development in 2004. Controversy remains on when the first live birth occurred after orthotopic ovarian transplantation with cryopreserved tissue as the patient was ovulating with elevated progesterone levels in the case reported in 2004; first live birth is likely to be the one reported by Meirow et al. in 2005. Nevertheless, the technique has evolved to reach a level where most recent live birth rates are exceeding 35% and the procedure is no longer considered experimental by many.

Recent data suggest that paternal age can have major impact on reproductive outcomes, and with increased age, there is increased likelihood of chromosomal abnormalities in the sperm. Here, we studied DNA damage and repair as a function of male aging and assessed whether sphingosine-1-phosphate (S1P), a ceramide-induced death inhibitor, can prevent sperm aging by enhancing DNA double-strand breaks (DSB) repair. We observed a significant increase in DNA damage with age and this increase was associated with a decline in the expression of key DNA DSB repair genes in mouse sperm. The haploinsufficiency of BRCA1 male mice sperm showed significantly increased DNA damage and apoptosis, along with decreased chromatin integrity when compared to similar age wild type (WT) mice. Furthermore, haploinsufficiency of BRCA1 male mice had lower sperm count and smaller litter size when crossed with WT females. The resulting embryos had a higher probability of growth arrest and reduced implantation. S1P treatment decreased genotoxic-stress-induced DNA damage in sperm and enhanced the expressions of key DNA repair genes such as BRCA1. Co-treatment with an ATM inhibitor reversed the effects of S1P, implying that the impact of S1P on DNA repair is via the ATM-mediated pathway. Our findings indicate a key role for DNA damage repair mechanism in the maintenance of sperm integrity and suggest that S1P can improve DNA repair in sperm. Further translational studies are warranted to determine the clinical significance of these findings and whether S1P can delay male reproductive aging. There is mounting evidence that sperm quality declines with age, similar to that of the oocyte. However, the reasons behind this decline are poorly understood and there is no medical intervention to improve sperm quality. Our study suggests a strong role for DNA damage repair in maintenance of sperm quality, and for the first time, a potential pharmaceutical approach to prevent sperm aging.