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Increased oocyte cohort size at retrieval for in vitro fertilization is associated with a higher proportion of aneuploid embryos
cases. 34 deliveries are obtained while 7 pregnancies are still ongoing. According to female age, clinical pregnancy rate drops from 34.7% to 22.2% in young (<37) and old age (R37) groups, respectively. Miscarriage rates are as follows: 19 % and 37.5%. In 13 cases Talassemia:9;WAS:1,ALD:1,ALL:1;Histiocytosis:1) successful aHSCT transplantation from the HLA compatible healthy siblings were perfomed and 18 cases are waiting for transplantation. CONCLUSIONS: The demand for the preimplantation HLA typing with or without a single gene disorder is increasing tremendously since the method is promising and an effective therapeutic tool for affected sibling. Our results show successful diagnosis of single gene disorders and application of HLA typing on the embryos. Despite the lower probability of finding suitable embryos for embryo transfer, acceptable pregnancy rates were observed in young females with good ovarian reserve. Testing for age- related aneuploidy applied in preimplantation HLA typing combined with PGD may improve the reproductive outcome. Supported by: None.
P-598 DOES OVARIAN STIMULATION PROTOCOL FOR IVF AND NUMBER OF OOCYTES RETRIEVED AFFECT EMBRYOS’ ANEUPLOIDY RATE? I. Tur-Kaspa, A. Bernal, N. Tkachenko, A. Kuliev, Z. Zlatopolsky, Y. Verlinsky. Institute for Human Reproduction (IHR), And Reproductive Genetics Institute, Chicago, IL; Reproductive Genetics Institute, Chicago, IL. OBJECTIVE: PGD results significantly decrease the number of embryos available for transfer by 30-70%. Thus, optimizing ovarian stimulation for IVF with PGD may improve outcome. We investigated if the ovarian stimulation protocol used for IVF and the number of oocytes retrieved affect embryos’ aneuploidy rate. DESIGN: Retrospective study. MATERIALS AND METHODS: 251 consecutive cycles of women age 22–47 years who underwent IVF/ICSI with PGD for aneuploidy were reviewed. Before treatment, patients’ ovarian reserve were estimated based on age, antral follicle count and day-3 FSH level, and, accordingly, changes were implemented in the stimulation protocol or in medication dosage to try to safely improve oocyte yield (to 10-15 oocytes). The data was analyzed by age, number of oocytes retrieved, and type of ovarian stimulation protocol used: long mid-luteal Lupron, antagonist, or Micro-dose Lupron. The primary endpoints were embryos’ aneuploidy rate and clinical pregnancy rate (þFHT). RESULTS: 1905 oocytes/embryos of women age < 43 years old were tested. Within the age groups of < 35 and 35-42 years old, the aneuploidy rates were similar and independent of the number of oocytes retrieved, % 7, 8-15, 16-20, or R 21 oocytes: 53%, 56%, 54%, 56% and 69%, 69%, 72%, 68%, respectively. The percent of cycles with ET increased with the increase number of oocytes retrieved. Within these age groups, comparable aneuploidy rates were obtained with the different stimulation protocols: 58%, 52%, 52% and 68%, 73%, 71% for the Lupron, antagonist, and Micro-dose protocols, respectively. The clinical pregnancy rates at age < 35 were 41%, 37%, 44% and at 35-42 years 26%, 33%, 35% for the patients with Lupron, antagonist, Micro-dose protocols, respectively. CONCLUSIONS: The ovarian stimulation protocol used for IVF and the number of oocytes retrieved had no effect on oocytes/embryos’ aneuploidy rate. Individually adjusted ovarian stimulation based on estimated ovarian reserve for patients undergoing IVF-PGD may improve outcome. Prospective, randomized trials are needed. Maximizing oocyte yield seems to increase the chance to generate a sufficient number of chromosomally normal embryos for transfer. Supported by: None.
P-599 INCREASED OOCYTE COHORT SIZE AT RETRIEVAL FOR IN VITRO FERTILIZATION IS ASSOCIATED WITH A HIGHER PROPORTION OF ANEUPLOID EMBRYOS. A. Kittai, A. Benner, R. Pen, K. Richter, W. G. Kearns. Shady Grove Center for Preimplantation Genetics, Rockville, MD; Shady Grove Fertility Reproductive Science Center, Rockville, MD.
FERTILITY & STERILITYÒ
OBJECTIVE: Evaluate potential risk factors for aneuploidy among embryos of patients undergoing in vitro fertilization (IVF) with preimplantation genetic screening (PGS). DESIGN: Retrospective review of results of PGS among 339 consecutive infertility patients undergoing autologous in vitro fertilization from Jan 2005Dec 2007. MATERIALS AND METHODS: Multi-color fluorescence in situ hybridization was used to determine aneuploidy for chromosomes 13, 14, 15, 16, 17, 18, 21, 22, X and Y. Potential risk factors for aneuploidy in embryos were investigated by simple linear and stepwise multiple regression. Independent variables included in the analysis were maternal and paternal age, oocytes per retrieval, total stimulation medication dosage (IU FSH), medication dosage per retrieved oocyte, and number of days of stimulation. RESULTS: Simple linear regression analyses indicated significant increases in the percentage of tested embryos that were chromosomally abnormal with both increasing maternal age (p¼0.003) and increasing paternal age (p¼0.02). There was a trend that showed the proportion of abnormal embryos increasing with an increasing number of oocytes per retrieval as well, although it was not significant (p¼0.1). Total stimulation medication dosage per cycle, medication dosage per oocyte retrieved, and length of stimulation were unrelated to aneuploidy rates with p-values of 0.97, 0.73 and 0.58 respectively. Maternal age was correlated with paternal age (r ¼ 0.63, p < 0.0001) and oocyte number (r ¼ 0.16, p¼0.0025). In the stepwise multiple regression analysis, both maternal age (p¼0.001) and the number of oocytes retrieved (p¼0.029) were significant predictors of the percentage of abnormal embryos, while none of the other examined variables showed any significance to chromosome copy-number (p > 0.4 for all). The percentage of aneuploid embryos increased with both increasing maternal age (0.9% per year) and increasing numbers of oocytes retrieved (0.3% per oocyte). CONCLUSIONS: Increased retrieved oocytes and higher maternal age are independent risk factors for aneuploidy. The apparent relationship between paternal age and aneuploidy is the result of a strong positive correlation between maternal and paternal age and is not a direct effect of paternal age, as shown by the stepwise multiple regression analysis. The relationships between aneuploidy and both maternal age and retrieved oocytes are clearer in multiple regression models incorporating both due to the negative correlation between these two variables. Supported by: None.
P-600 24-HOUR DETECTION OF ANEUPLOIDY ACROSS 24 CHROMOSOMES IN 100 BLASTOMERES WITH COMPUTED CONFIDENCES. D. S. Johnson, J. Salzman, C. Cinnioglu, M. Alper, D. Potter, M. Rabinowitz. Gene Security Network, Redwood City, CA; Boston IVF, Waltham, MA; Huntington Reproductive Center, Laguna Hills, CA. OBJECTIVE: Current technologies for measuring genome-wide copy number in single cells use intensities from microarray probes to compute quantitative differences in probe intensities between chromosomes. Extreme preferential amplification in whole genome amplification (WGA) makes it difficult to assign statistical confidences with such approaches. Our goal was to develop and validate a statistical technology that uses genetic data from the mother and the father, together with the noisy measurements of embryonic DNA, to determine copy numbers across all 24 chromosomes, from a single cell, within 24 hours, with reliability of each call for the cell explicitly computed and typically in excess of 99%. DESIGN: Prospective, randomized, and blinded. MATERIALS AND METHODS: We implemented a Bayesian algorithm that uses data from the parents and noisy measurements from single cells to compute explicit confidences of copy number calls in the single cells. The technology is called Parental SupportTM (PS). We measured copy number on 50 single cells with known chromosome copy numbers. One of the families included a trisomy 21 child, and the remainder of the children was euploid. We also measured copy number on 100 single blastomeres. Genotyping was performed across hundreds of thousands of loci using Illumina 370CNV-Duo microarrays and a modified 24-hour protocol. RESULTS: On tissues where true chromosome copy numbers were known, PS correctly called copy number on 24 chromosomes in each of 50 single cells, for a total of 1200 correct calls, with explicitly computed statistical confidences typically in excess of 99%. These chromosome copy
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P-598 DOES OVARIAN STIMULATION PROTOCOL FOR IVF AND NUMBER OF OOCYTES RETRIEVED AFFECT EMBRYOS’ ANEUPLOIDY RATE? I. Tur-Kaspa, A. Bernal, N. Tkachenko, A. Kuliev, Z. Zlatopolsky, Y. Verlinsky. Institute for Human Reproduction (IHR), And Reproductive Genetics Institute, Chicago, IL; Reproductive Genetics Institute, Chicago, IL. OBJECTIVE: PGD results significantly decrease the number of embryos available for transfer by 30-70%. Thus, optimizing ovarian stimulation for IVF with PGD may improve outcome. We investigated if the ovarian stimulation protocol used for IVF and the number of oocytes retrieved affect embryos’ aneuploidy rate. DESIGN: Retrospective study. MATERIALS AND METHODS: 251 consecutive cycles of women age 22–47 years who underwent IVF/ICSI with PGD for aneuploidy were reviewed. Before treatment, patients’ ovarian reserve were estimated based on age, antral follicle count and day-3 FSH level, and, accordingly, changes were implemented in the stimulation protocol or in medication dosage to try to safely improve oocyte yield (to 10-15 oocytes). The data was analyzed by age, number of oocytes retrieved, and type of ovarian stimulation protocol used: long mid-luteal Lupron, antagonist, or Micro-dose Lupron. The primary endpoints were embryos’ aneuploidy rate and clinical pregnancy rate (þFHT). RESULTS: 1905 oocytes/embryos of women age < 43 years old were tested. Within the age groups of < 35 and 35-42 years old, the aneuploidy rates were similar and independent of the number of oocytes retrieved, % 7, 8-15, 16-20, or R 21 oocytes: 53%, 56%, 54%, 56% and 69%, 69%, 72%, 68%, respectively. The percent of cycles with ET increased with the increase number of oocytes retrieved. Within these age groups, comparable aneuploidy rates were obtained with the different stimulation protocols: 58%, 52%, 52% and 68%, 73%, 71% for the Lupron, antagonist, and Micro-dose protocols, respectively. The clinical pregnancy rates at age < 35 were 41%, 37%, 44% and at 35-42 years 26%, 33%, 35% for the patients with Lupron, antagonist, Micro-dose protocols, respectively. CONCLUSIONS: The ovarian stimulation protocol used for IVF and the number of oocytes retrieved had no effect on oocytes/embryos’ aneuploidy rate. Individually adjusted ovarian stimulation based on estimated ovarian reserve for patients undergoing IVF-PGD may improve outcome. Prospective, randomized trials are needed. Maximizing oocyte yield seems to increase the chance to generate a sufficient number of chromosomally normal embryos for transfer. Supported by: None.
P-599 INCREASED OOCYTE COHORT SIZE AT RETRIEVAL FOR IN VITRO FERTILIZATION IS ASSOCIATED WITH A HIGHER PROPORTION OF ANEUPLOID EMBRYOS. A. Kittai, A. Benner, R. Pen, K. Richter, W. G. Kearns. Shady Grove Center for Preimplantation Genetics, Rockville, MD; Shady Grove Fertility Reproductive Science Center, Rockville, MD.
FERTILITY & STERILITYÒ
OBJECTIVE: Evaluate potential risk factors for aneuploidy among embryos of patients undergoing in vitro fertilization (IVF) with preimplantation genetic screening (PGS). DESIGN: Retrospective review of results of PGS among 339 consecutive infertility patients undergoing autologous in vitro fertilization from Jan 2005Dec 2007. MATERIALS AND METHODS: Multi-color fluorescence in situ hybridization was used to determine aneuploidy for chromosomes 13, 14, 15, 16, 17, 18, 21, 22, X and Y. Potential risk factors for aneuploidy in embryos were investigated by simple linear and stepwise multiple regression. Independent variables included in the analysis were maternal and paternal age, oocytes per retrieval, total stimulation medication dosage (IU FSH), medication dosage per retrieved oocyte, and number of days of stimulation. RESULTS: Simple linear regression analyses indicated significant increases in the percentage of tested embryos that were chromosomally abnormal with both increasing maternal age (p¼0.003) and increasing paternal age (p¼0.02). There was a trend that showed the proportion of abnormal embryos increasing with an increasing number of oocytes per retrieval as well, although it was not significant (p¼0.1). Total stimulation medication dosage per cycle, medication dosage per oocyte retrieved, and length of stimulation were unrelated to aneuploidy rates with p-values of 0.97, 0.73 and 0.58 respectively. Maternal age was correlated with paternal age (r ¼ 0.63, p < 0.0001) and oocyte number (r ¼ 0.16, p¼0.0025). In the stepwise multiple regression analysis, both maternal age (p¼0.001) and the number of oocytes retrieved (p¼0.029) were significant predictors of the percentage of abnormal embryos, while none of the other examined variables showed any significance to chromosome copy-number (p > 0.4 for all). The percentage of aneuploid embryos increased with both increasing maternal age (0.9% per year) and increasing numbers of oocytes retrieved (0.3% per oocyte). CONCLUSIONS: Increased retrieved oocytes and higher maternal age are independent risk factors for aneuploidy. The apparent relationship between paternal age and aneuploidy is the result of a strong positive correlation between maternal and paternal age and is not a direct effect of paternal age, as shown by the stepwise multiple regression analysis. The relationships between aneuploidy and both maternal age and retrieved oocytes are clearer in multiple regression models incorporating both due to the negative correlation between these two variables. Supported by: None.
P-600 24-HOUR DETECTION OF ANEUPLOIDY ACROSS 24 CHROMOSOMES IN 100 BLASTOMERES WITH COMPUTED CONFIDENCES. D. S. Johnson, J. Salzman, C. Cinnioglu, M. Alper, D. Potter, M. Rabinowitz. Gene Security Network, Redwood City, CA; Boston IVF, Waltham, MA; Huntington Reproductive Center, Laguna Hills, CA. OBJECTIVE: Current technologies for measuring genome-wide copy number in single cells use intensities from microarray probes to compute quantitative differences in probe intensities between chromosomes. Extreme preferential amplification in whole genome amplification (WGA) makes it difficult to assign statistical confidences with such approaches. Our goal was to develop and validate a statistical technology that uses genetic data from the mother and the father, together with the noisy measurements of embryonic DNA, to determine copy numbers across all 24 chromosomes, from a single cell, within 24 hours, with reliability of each call for the cell explicitly computed and typically in excess of 99%. DESIGN: Prospective, randomized, and blinded. MATERIALS AND METHODS: We implemented a Bayesian algorithm that uses data from the parents and noisy measurements from single cells to compute explicit confidences of copy number calls in the single cells. The technology is called Parental SupportTM (PS). We measured copy number on 50 single cells with known chromosome copy numbers. One of the families included a trisomy 21 child, and the remainder of the children was euploid. We also measured copy number on 100 single blastomeres. Genotyping was performed across hundreds of thousands of loci using Illumina 370CNV-Duo microarrays and a modified 24-hour protocol. RESULTS: On tissues where true chromosome copy numbers were known, PS correctly called copy number on 24 chromosomes in each of 50 single cells, for a total of 1200 correct calls, with explicitly computed statistical confidences typically in excess of 99%. These chromosome copy
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