In this webinar session, Evi Timotheou, BcS, MSc, Senior Clinical Embryologist, Lab Director at Assisting Nature, Thessaloniki, Greece, has talked about the indications for Pre-implantation Genetic Testing (PGT-A)
. Evi has presented 5 different IVF cases and explained all the steps taken that allowed the couples to achieve a pregnancy.
The history of PGT
Evi started her presentation by introducing a brief history of genetic testing. Since 1990, genetic testing worldwide has increased, and indications for its use have expanded. Originally, the goal of PGD was to detect and eliminate the embryos that contained monogenic X linked disorders.
Nowadays, PGS and PGD have expanding roles in reproductive medicine. According to The International Glossary on Infertility and Fertility Care of 2017, Pre-implantation genetic testing is a test performed to analyse the DNA from oocytes or embryos for determining genetic abnormalities. The names have been divided, PGS is now called PGT-A (Preimplantation Genetic Testing for Aneuploidies), while PGD is now called PGT-M (Preimplantation genetic testing for monogenic/single gene defects) and PGT-SR (Preimplantation Genetic Testing for Chromosomal Structural Rearrangements).
The most common indications for the use of PGT-A are:
- advanced maternal age (AMA)
- repeated implantation failure (RIF)
- repeated miscarriages
- severe male factor
- changes in the parental karyotype
Around 40% of cases where PGT-A is indicated are because of advanced maternal age. There are different biopsy methods. There is Polar Body Biopsy, Blastomere Biopsy, Trophectoderm Biopsy. Throughout the years, different genetic techniques were used that improved chromosomal analysis. In 1990, FISH PCR was applied, and after 2008, other techniques like aCGH (Array Comparative Genomic Hybridization) were mostly used. Finally, in 2014, with the Next-Generation Sequencing (NGS), a new era began.
On the graph shown where FISH, which was the first method used, we can observe a reduction in implantation and an increase in aneuploidy with advanced material age even though only a few chromosomes were tested. The transition from PGS 0.1 (FISH), which has only 9 chromosomes, to the transition of PGS 2.0 where all 24 chromosomes were analysed using Comprehensive Chromosome Screening (CCS). Numerous studies were presented about the ideal method of biopsy and analysis for each indication.
Nowadays, with the new screening techniques, we can observe and categorize the embryos as normal, mosaic and abnormal. According to The Preimplantation Genetic Diagnosis International Society (PGDIS), less than 20% of aneuploidy in an embryo is considered normal. If it is between 20 to 80%, they are considered mosaic, and more than 80% is categorized as aneuploid and is not suitable for transfer.
Mosaicism is a state in which there is more than one karyotypically distinct cell population arising from a single embryo. It’s a huge area, and it’s not easily covered.
IVF with PGT-A – real IVF patients cases
- a 40-year-old woman, no pregnancy before, 2x IUIs, 2x failed IVF cycles which ended in miscarriage
The first woman was 40 years old with a regular cycle. She didn’t have any pregnancies before. Her partner had normal sperm analysis, they had 2 previous IVF cycles in other centres. She started with 2 Intrauterine Inseminations (IUI) and had negative results both times. Then, the couple proceeded with the first IVF, 6 mature oocytes were retrieved, 5 fertilized, fresh 2 day-5 embryos were transferred, she had a positive result that ended in a spontaneous abortion. Then she had a second IVF cycle, another 6 oocytes were retrieved and fertilized, she transferred 2 day-4 embryos that ended in a biochemical pregnancy.
When she came to our clinic (Assisting Nature), we decided to do IVF with PGT-A and freeze all embryos because when we perform NGS we have to freeze them first, and then we wait for the results. We suggested doing a hysteroscopy, and the doctor suggested a long agonist protocol. We got again 6 oocytes, and 5 were fertilized, we had 5 blastocysts for biopsy. We do the file, the blastocyst, the biopsy. The results showed 4 aneuploid embryos and 1 euploid. Therefore, we transferred that 1 euploid embryo and the result was positive. A baby girl was born at 39 weeks.
- a 41-year-old with irregular cycles, repeated miscarriages, chronic endometritis, no previous IVF cycles
The patient was 41 with irregular cycles. She had natural pregnancies before, the first one was a 9-week pregnancy, but she lost the baby. And the second, she had an 8-week pregnancy loss and a 3rd biochemical pregnancy. In 2019, she did an endometrial biopsy, and it was discovered that she had chronic endometriosis. Her partner had a normal sperm analysis, they didn’t have any previous IVF cycles.
The plan was again to do IVF with PGT-A and freeze the embryos. Hysteroscopy was performed before embryo transfer. classical antagonist protocol was done. We retrieved only 3 oocytes, but all 3 were fertilized. We got 2 blastocysts, and with the result, we had 1 euploid and 1 aneuploid. We transferred the euploid embryo and the couple got a positive result with a live birth at 38 weeks.
- a 42-year-old, partner had frozen sperm – non-Hodgkin lymphoma chemotherapy, previous failed IVF cycles, repeated miscarriages
The couple where a woman was 42 and her partner was 47. The partner had preserved sperm before he underwent chemotherapy for non-Hodgkin’s lymphoma. They had previous IUIs cycles in other centres that were negative, and they had an IVF cycle with 7 oocytes, from which 7 were fertilized, and they had 6 blastocysts. They proceeded with 2 blastocysts and had the first embryo transfer. The result was positive, but ended in an 11-week pregnancy loss. Then they proceeded with the frozen embryos, again she had a positive pregnancy test, but a 6-week pregnancy loss and then the last 2 frozen embryos were transferred, but she had a biochemical pregnancy.
When they came to our clinic, we suggested PGT-A because of her age, it was normal to proceed with a PGT-A. We planned IVF with classical antagonist protocol. In our clinic, we had 6 oocytes, 6 were fertilized and we had 4 blastocysts. We only got 1 euploid embryo, the rest were aneuploid. We proceeded with the embryo transfer, and the result was positive, and we had an ongoing pregnancy. Now, she is at 31 weeks, and she is well.
- a 42-year-old with previous 3 failed IVF cycles, normal sperm analysis
The couple had a pregnancy through natural conception and a 2-week pregnancy loss. Then, they proceeded with IVF cycles because of her age, she had 3 consecutive IVF cycles in other centres. Firstly, she had 5 mature oocytes, 4 were fertilized with 2 blastocysts, she had an embryo transfer, but with a negative result. Then she had another IVF cycle with 4 oocytes, 2 fertilised, and 1 blastocyst was transferred, which resulted in a negative. In the last embryo transfer, she had 4 oocytes, 3 fertilized, and they got 2 blastocysts, resulting in a biochemical pregnancy.
We planned the protocol and PGT-A. We proceeded with the antagonist protocol, she had the first IVF cycle where we retrieved 4 mature oocytes, 4 fertilized, 2 blastocysts were biopsied, but unfortunately, we didn’t have any normal embryos. All blastocysts were abnormal. They proceeded with a second IVF cycle where 5 mature oocytes were retrieved, 4 fertilized, and we got 3 blastocysts. This time, they had 1 euploid embryo and 2 aneuploid embryos, and she had an embryo transfer. She had a positive result and an ongoing pregnancy, currently at 13 weeks of pregnancy.
- a 44-year-old woman, 2 natural conceptions with biochemical pregnancies, previous 2 failed IVF cycles
A 44-years-old woman with irregular cycles and normal sperm analysis. She had natural conceptions with 2 biochemical pregnancies, and then she started the IVF cycles. Firstly, she had 5 mature oocytes, 4 were fertilized, 2 blastocysts with a fresh embryo transfer, and she had a negative result. Secondly, she had another cycle with 4 mature oocytes, 2 were fertilized, 1 blastocyst was transferred, and she had a biochemical pregnancy.
Once again, we planned PGT-A, hysteroscopy, appropriate protocol for her. The doctor also discussed the egg donation option if there were no euploid embryos. In her first IVF cycles at our clinic, we got 4 mature oocytes, 3 were fertilized, 2 blastocysts were biopsied, but the result showed no euploid embryos. They had a second IVF cycle with 4 mature oocytes, all 4 were fertilized with 3 blastocysts. Unfortunately, all embryos were abnormal.
The couple decided to proceed with the last 3rd attempt, 3 oocytes, 2 fertilized, and we got 1 blastocyst. The result was also an abnormal embryo. Finally, the couple decided not to proceed with other IVF cycles and proceeded with an oocyte donation program.
PGT-A – success rates
On the graph shown, in the last 3 years in our clinic (Assisting Nature), between 2018-2020, the pregnancy rate is 90%. The clinical pregnancy rate is 85%, and the live birth rate is 70%. In most cases, a single embryo transfer was performed. In 65%, it was a single embryo transfer, and we can see the live birth rates are very high if somebody proceeds with the PGT-A method.