P-148 Denudation employing human recombinant hyaluronidase enzyme does not affect fertilization and blastocyst formation rates in sibling oocytes from patients undergoing ART: A prospective randomized study
Authors: Almaguer-Fernandez L.D , Ramirez Dominguez L.B , Jimenez-Medina I , Maldonado-Rosas I , Izquierdo-Martinez M , Sanchez-Encarnacion Y.J , Villar-Muñoz L.G , Simopoulou M , Agarwal A 
Published: June 2022

Study question

Does human recombinant hyaluronidase (Cumulase®) improve fertilization and blastocyst formation rates in sibling human oocytes compared to bovine-derived hyaluronidase?

Summary answer

The use of Cumulase® for oocyte denudation does not appear to affect fertilization or usable blastocyst formation rate in patients undergoing intracytoplasmic sperm injection (ICSI).

What is known already

Bovine-derived hyaluronidase has been widely employed to denude the cumulus-oocyte complexes (COCs) prior to ICSI or oocyte vitrification. Nonetheless, the recent concern for its safety, due to its animal origin, has led to the development of alternatives ascertaining the enzyme’s purity. It has been reported that human recombinant hyaluronidase is not equally effective as its bovine-derived analogue, leading to a longer time required for complete denudation coupled by the subsequent toxicity risk for oocytes. This may result to lower fertilization and blastocyst formation rates. Although there are some studies on IVF outcomes with recombinant hyaluronidase, its use remains rather limited.

Study design, size, duration

This randomized double-blinded clinical trial was conducted at Citmer Reproductive Medicine, in Puebla and Monterrey, Mexico from February to December 2021. The impact of denudation on sibling oocytes from 103 patients employing bovine-derived hyaluronidase (InVitroCare®) or human recombinant hyaluronidase (ICSI Cumulase® (rHuPH20), CooperSurgical®) was evaluated. A total of 1237 oocytes were assigned as follows: The Hyaluronidase Group included 626 oocytes, while the Cumulase® Group included 611 oocytes. Preimplantation Genetic Testing was performed in 18 cycles.

Participants/materials, setting, methods

After retrieval, COCs were incubated for 2-4 h and placed in microdropets with 10 IU/mL bovine-derived hyaluronidase or Cumulase® in culture media GlobalTotal® (LifeGlobal®) for granulosa cells removal. COCs were gently pipetted up and down employing 20 μL and 150 μm tips. Metaphase II oocytes were injected and incubated for 17-20 hours in 8% CO2, 20% O2, 37°C. Normally fertilized zygotes were cultured to blastocyst stage and embryo development was evaluated. Seventy-nine blastocysts were biopsied.

Main results and the role of chance

The mean age of the patients was 32.12±1.11 years old. A total of 506 and 493 metaphase II oocytes from Groups 1 and 2 respectively were subjected to ICSI. Normal fertilization rates for oocytes treated with Hyaluronidase and Cumulase® were 71.34% (n = 361) and 72.41% (n = 357) respectively, with no statistically significant differences between the two groups (p = 0.35).

Usable blastocysts rate (selected for embryo transfer or blastocyst cryopreservation) did not differ with any statistical difference between the bovine-derived hyaluronidase (n = 170, 47.09%) and the Cumulase® treated oocytes (n = 158, 44.25%), (p = 0.44).

Preimplantation genetic testing for aneuploidies (PGT-A) was performed in 18 cycles. Seventy-nine blastocysts were biopsied and tested. Fourty-seven embryos from the bovine-derived hyaluronidase group were biopsied, presenting with a euploidy rate of 51.06% (n = 24), compared to 32 embryos from the recombinant hyaluronidase group with a euploidy rate of 50% (n = 16). Results indicated no statistically significant difference (p = 0.92). These data support that human recombinant hyaluronidase presents with similar efficacy to bovine-derived hyaluronidase. in terms of normal fertilization, embryo development as well as euploidy rates.

Limitations, reasons for caution

The small sample size regarding the euploidy outcome stands as a limitation of the study. Further studies should focus on the effect of recombinant hyaluronidase on genetic anomalies on blastocysts.

Wider implications of the findings

Fertilization, usable blastocyst and euploidy rates did not differ when employing bovine-derived or human recombinant hyaluronidase, rendering the latter a safe and efficient alternative. However, it is important to assess long-term effects, as only a limited number of studies report on perinatal and neonatal outcomes

Trial registration number

25-12-21MTYPUECIT