MUNICH – Vitrification, quick freezing oocytes or embryos with concentrated cyroprotectant and no ice crystal formation to better preserve viability, has been around clinically some 15 years, but now is gathering steam and may have already helped downshift the rate of triplets or higher pregnancies for women undergoing assisted reproduction.

While vitrification of all embryos fertilized in vitro has not proven superior to fresh embryo transfers and may even be inferior for older women whose eggs may not be as forgiving of cryopreservation, its use has grown. It offers the unique advantage over fresh embryos of easier coordination between the in vitro fertilization process and subsequent transfer to the woman who will carry the pregnancy. Vitrification has also made possible true oocyte banking, be it autologous for fertility preservation, or for oocyte donation.

"It is now possible to freeze embryos with a 90% survival rate; with slow freezing there was 50%-60% survival. Now we have a technique that is really good, and it may cause a paradigm shift toward elective freezing," said Dr. Anja Pinborg, a professor of ob.gyn. at Hvidovre (Denmark) Hospital.

Most clinicians still use the approach of placing an embryo during the same cycle when the oocyte was harvested, "the way it’s always been done, but at many clinics now they see that with vitrification they can get pregnancy rates that are similar to or even better than fresh embryo transfer. I think they will go more and more to frozen embryos," Dr. Pinborg said in an interview during the annual meeting of the European Society of Human Reproduction and Embryology.

For now the trend is specific for women younger than 35 years. Safety and efficacy of vitrification for embryos made from oocytes taken from older women has not yet been studied, cautioned Dr. Pinborg and others.

"All we know about cryopreservation is from young women with young oocytes. We don’t know what will happen with oocytes from women who are 41 or 42, who are the types of patients I see," said Dr. Richard J. Paulson, professor of ob.gyn. and chief of the division of reproductive endocrinology and infertility at the University of Southern California and director of USC Fertility in Los Angeles. "Vitrification is so good and embryos come out of it so well that the benefit from improved endometrial receptivity outweighs any change in embryo quality. The timing coordinates so much better."

The biggest impact of vitrification for embryo freezing may be its ability to successfully cryopreserve blastocysts. "We will see many more programs going to 5-day embryos" for implantation, and this will allow preimplantation genetic diagnosis (PGD), predicted Dr. Paulson during an interview at the meeting. "It looks like you can do pretty good trophectoderm biopsy on a vitrified blastocyst, so you can place only chromosomally normal embryos. That will reduce miscarriages." So far, only a handful of groups have presented case reports using this approach, with "very good numbers," he said.

Vitrification’s impact on embryo transfer number

Despite lingering questions about vitrification safety and limited knowledge of its efficacy for embryos begun from oocytes from older women, the most recent worldwide and European data on patterns of assisted reproductive technology show increases in frozen embryo use and decreases in triplets or greater multiples.

Worldwide, frozen embryo transfers grew from 10% of all in vitro fertilization or sperm injection procedures in 1991 to 28% in 2010, the most recent year with worldwide data available. Transfers of one or two embryos grew from 20% of all in vitro procedures done in North America in 1998 to 68% in 2010. In Australia in 2010, the rate of one- or two-embryo transfers stood at 98%.

"The trend is for more frozen transfers, and for reducing multiple rates. I think that is a very positive development," said Dr. David Adamson, a reproductive endocrinologist who practices in Palo Alto, Calif., and presented at the meeting worldwide data collected by the International Committee Monitoring Assisted Reproductive Technologies (ICMART). "The pregnancy rate is continuing to go up, but the multiple rate is dropping. We hope that will continue," he said in an interview.

"There is no question that because vitrification works so much better than slow freezing, clinics have more confidence to put in fewer embryos and still have viability," he said. "We’ll see more frozen embryo transfers in the next few years. When you decrease the number of embryos transferred, you increase the likelihood that you’ll need to follow with a frozen embryo transfer, but now you have higher-quality embryos."

A similar shift to fewer embryos transferred at a time appeared in the exclusively European registry with data derived from 33 countries that reported complete or partial data to the European IVF Monitoring Program, a project organized by ESHRE. The prevalence of triplet or greater deliveries from in vitro pregnancies fell in Europe from about 4% in 1999 to less than 1% in 2011. By 2011, roughly 28% of IVF or sperm-injected pregnancies involved placement of two embryos, and about 57% involved a single embryo, reported Dr. Markus S. Kupka, an ob.gyn. and reproductive medicine specialist who practices in Hamburg, Germany. Frozen embryos involved 31% of transfers in 2011 in Europe. European data for 2010 were included in the worldwide tallies reported by Dr. Adamson.

Vitrification’s safety

Despite growing use, some questions remain about vitrification’s impact on children, although the concern has not been strong enough to dim enthusiasm.

The most striking safety signal is a consistent, 50% increased rate of babies born following vitrification who are large for gestational age. Dr. Pinborg also cited a possibly increased rate of maternal preeclampsia.

"It’s only a 50% increased risk for large for gestational age with frozen embryo transfer. We should not pay too much attention to this, but it reminds us that when we freeze, it may have an effect on the children" Dr. Pinborg said in an interview.

"We can’t say that vitrified embryos do better than after fresh embryo transfer, but they have an altered risk profile." Frozen embryos have a reduced risk for preterm birth and small for gestational age, compared with fresh embryo transfers, but an increased risk of large for gestational age (LGA). "And the risk is small," stressed Dr. Pinborg. "When you stimulate ovulation, you harvest 10-fold as many oocytes than occur with a normal cycle, and the woman’s estrogen and progesterone levels are very high. So, you could ask: Why should we ever place an embryo during an oocyte harvesting cycle? We did that because we had poor freezing methods, but with vitrification we have a way to use preserved embryos and there is better cycle synchronization."

In addition, seeing increased rates of LGA babies following frozen embryo transfer may result from unadjusted confounding by, for example, the embryo’s age when cryopreservation occurred that could influence epigenetic changes that happen early after fertilization or maternal disorders such as polycystic ovary syndrome, Dr. Paulson said.

Safety aside, vitrification is very effective for assisted reproduction because any decrement in viability is counterbalanced by the advantage of placing embryos squarely during a natural cycle, these experts said.

Vitrification and oocytes

Vitrification does not stop with embryos. The impact it has had on oocyte banking has arguably been even greater.

Creating multiple embryos at one time is problematic, especially in the United States, Dr. Paulson said. "The obvious thing we should do is freeze oocytes. This approach would create another significant advantage. Routine vitrification at the oocyte stage rather than after embryos are made "would make donor oocytes a reality. You would be able to buy frozen oocytes the same way as sperm. Results from good randomized, controlled trials show that outcomes with cryopreserved oocytes are as good as with fresh oocytes, which was never possible with slow freezing."

"Vitrification created the ability to have oocyte banking. It has been a breakthrough in assisted reproductive technology," said Ana Cobo, Ph.D., director of the cryopreservation laboratory at the IVI Foundation in Valencia, Spain. Cryopreserving oocytes was essentially impossible before vitrification became available, she said in an interview. Studies run by her group showed that the vitrification process has no detectable effect on oocyte viability, embryo viability, or the health of the child. It also made oocyte donation much more feasible by eliminating the need for cycle synchrony between the donor and recipient.

The success rate of in vitro fertilization using a donated oocyte is generally much higher than for an autologous oocyte because "we use a highly selected population of young, healthy women with the best quality oocytes. In assisted reproductive technology, oocyte quality means a lot," Dr. Cobo said.

Dr. Pinborg said that she has received research grants from MSD and Ferring. Dr. Paulson said that he has been a speaker on behalf of Ferring and served on advisory boards for Origio and Cooper Surgical. Dr. Adamson said that he had received honoraria from or consulted for Bayer, Glycotype, and Ziva Medical, that he owns stock in Advanced Reproductive Care, and has received research support from Auxogyn and LabCorp. Dr. Kupka said he had no disclosures. Dr. Cobo had no disclosures.

mzoler@frontlinemedcom.com

On Twitter @mitchelzoler

Quick freezing oocytes and embryos transforms assisted reproduction