MITOCHONDRIAL DNA COPY NUMBER, TELOMERE LENGTH AND DNA METHYLATION IN PERIPHERAL BLOOD OF WOMEN UNDERGOING IN VITRO FERTILIZATION CYCLES AS NEW PREDICTORS OF LIVE BIRTH

Background: The peripheral biomarker of female reproductive biological age proposed so far (i.e., mitochondrial DNA copy number (mt-DNAcn), telomere length (TL) and DNA methylation (DNAm) in peripheral blood are promising. Unfortunately, the results of studies aimed at investigating their predictive capacity are conflicting. The conduction of a prospective study including women at the beginning of their natural pregnancy seeking, at the present state of knowledge, goes beyond the possibilities offered by even the most favorable settings. In this context, assisted reproductive technology (ART) has emerged as the most reliable study model. Importantly, considering the absence of non-invasive predictors of ART success, the identification of a reliable biomarker would also have positive implications for the determination of the ART risk-benefit ratio and, in a public health perspective, for the rational allocation of economic resources. Objective: To evaluate whether mt-DNAcn, TL or epigenetic age estimators based on DNAm pattern (biological age, epigenetic age acceleration, LINE-1 methylation rate), could be considered reliable predictors of in vitro fertilization (IVF) success in terms of live birth rate (LBR). Design: Prospective cohort study Setting: Infertility Unit of the Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Italy. Patients: 181 women aged 37-39 years who underwent IVF at a single center between January 2017 and December 2018. Interventions: On the day of recruitment, blood samples were collected, and genomic DNA was isolated from white blood cells. TL, mt-DNAcn and DNAm assessment was performed using quantitative real-time polymerase chain reaction (qPCR). Biological age (DNAm age) was computed as the algorithm based on methylation pattern of five genes. Epigenetic age acceleration was estimated from the residuals of the linear model of epigenetic age regressed on chronological age. Long Interspersed Nuclear Elements-1 (LINE-1) methylation pattern was used as a surrogate for global DNA methylation. Main outcome Measures: This study investigated whether peripheral TL, mt-DNAcn and DNAm could predict live birth in IVF cycles. Results: TL, mt-DNAcn and LINE-1 methylation were not associated with IVF success. Conversely, DNAm age resulted significantly lower in women who had a live birth compared to women who did not (36.1 ± 4.2 and 37.3 ± 3.3 years, respectively, p=0.04). For DNAm age, odds ratio (OR) for live birth per year of age was 0.90 (95%CI: 0.82-0.99, p=0.036) after adjusting for FSH and antral follicle count (AFC) and 0.90 (95%CI: 0.82-0.99, p=0.028) after adjusting also for number of oocytes retrieved. A significant association also emerged for epigenetic age acceleration after adjustments (OR=0.91, 95%CI: 0.83-1.00, p=0.048). Conclusion: DNAm age is associated with IVF success but the magnitude of this association is insufficient to claim a clinical use. However, our findings are promising and warrant further investigation. Assessment of biological age using different epigenetic clocks or focusing on different tissues may reveal new predictors of IVF success.