Prospective, Open-Label, and Three-Arm Investigator-Initiated Study to Compare the Efficacy and Safety of Three Estradiol Treatment Protocols for Endometrial Preparation in Frozen Embryo Transfer (FET) Cycles

Background Hormone Replacement Therapy (HRT) is a widely used method for endometrial preparation in Frozen Embryo Transfer (FET) cycles. Although multiple estradiol formulations and routes of administration exist, limited data directly compare oral estradiol hemihydrate, vaginal estradiol hemihydrate, and oral estradiol valerate. This study evaluated the efficacy and safety of these three estradiol regimens for endometrial preparation in an assisted reproduction setting. Objectives To compare endometrial outcomes, serum estradiol levels, and pregnancy outcomes among three estradiol treatment protocols, oral estradiol hemihydrate, vaginal estradiol hemihydrate, and oral estradiol valerate, in HRT‑based FET cycles. Methods This prospective, open‑label, three‑arm investigator‑initiated study was conducted at a single center and included 133 women aged 25-42 years undergoing HRT‑FET cycles. Participants received one of the following regimens: oral estradiol hemihydrate, vaginal estradiol hemihydrate, or oral estradiol valerate. Endometrial thickness, endometrial volume, and serum estradiol levels on progesterone‑start day were measured. Clinical outcomes included serum β-human chorionic gonadotropin (β‑hCG) positivity and clinical pregnancy rates. Data were analyzed using descriptive and comparative statistics. Results The mean daily estradiol dose was significantly lower with vaginal estradiol hemihydrate compared to both oral arms (p<0.001). Endometrial thickness and volume showed no significant differences among groups. Mean serum estradiol levels were highest with vaginal estradiol hemihydrate (701.63 pg/mL), significantly exceeding levels with oral estradiol hemihydrate (331.84 pg/mL; p=0.042). Serum β‑hCG positivity rates were 58.33%, 47.62%, and 40.47% (p=0.375) and clinical pregnancy rates were comparable at 44.0%, 28.0%, and 22.7% (p=0.168) in oral estradiol hemihydrate, vaginal estradiol hemihydrate, and oral estradiol valerate groups, respectively. Conclusions Estradiol hemihydrate, whether administered orally or vaginally, demonstrated non‑inferior efficacy to oral estradiol valerate for endometrial preparation in FET cycles. Vaginal estradiol hemihydrate achieved adequate endometrial development at significantly lower doses overcoming the barriers of first pass metabolism and minimizing the safety concerns of thrombo-embolism.