- Email: [email protected]
Versican and prostaglandin-endoperoxide synthase 2 (PTSG2) gene expression in cumulus cells as a complement of preimplantation genetic testing: better outcomes for in vitro fertilization pregnancy
(age), gonadotropin doses (dose/day, F(hMG)), days of stimulation (days) , estradiol/mature oocyte on day of trigger (E2), number of mature oocytes retrieved (MII), number of embryos biopsied (Bx) and incidence of euploidy (determined using TE biopsy and aCGH) and physician of record (MD) were selected for analysis. Differences in COH parameters were examined using analysis of variance (ANOVA). The number of euploid embryos (#Eup) per donor cycle was examined for associations with COH parameters using multiple regression of COH parameters versus log(#Eup). Parameters were considered significant if their regression coefficient (slope) was significantly different than 0. RESULTS: There were significant differences between donors grouped by physician when considering age, dose/day, F(hMG), days , E2, MII, euploidy rate and #Eup (ANOVA, each test P< 0.01). Using multiple regression, #Eup was significantly associated with MII, days and dose/day (Eq. 1). A greater #Eup was associated with more MII retrieved, fewer days of stimulation and lower dose/day. Age, F(hMG), and E2 were not significantly associated with #Eup. The magnitude of these parameter’s effects on #Eup were different. MII had the greatest effect whereas dose/day and days were roughly comparable to each other but less than MII. CONCLUSIONS: There were significant differences between COH characteristics for donors treated by different physicians indicating that donors were not treated similarly by the physicians. The associations of MII, days and dose/ day with log(#Eup) suggest that #EuR is increased by greater MII, fewer days and decreased dose/day. It remains unclear whether these relationships are causal or merely indicative of the ease of stimulation for donors. Regression Equation log(#Eup) ¼ 0.016 X MII - 0.017 X days 0.00031 X dose/day + 0.78
Eq. 1
P-473 Wednesday, November 1, 2017 ASSESSMENT OF FERTILITY CLINIC WEBSITES ON PREIMPLANTATION GENETIC SCREENING (PGS) AND PREIMPLANTATION GENETIC DIAGNOSIS (PGS). N. Joshi,a,b T. Zore,a,b S. B. Schon,c P. Masson,d J. L. Chan.b aObstetrics and Gynecology, University of California Los Angeles, Los Angeles, CA; bReproductive Endocrinology and Infertility, Cedars-Sinai Medical Center, Los Angeles, CA; cUniversity of Michigan, Ann Arbor, MI; dUniversity of Pennsylvania, Philadelphia, PA. OBJECTIVE: In recent years, improvements in PGS/PGD have led to high implantation rates.1 We sought to determine whether Society of Assisted Reproductive Technology (SART) member clinics mention PGS/PGD on their websites and investigate associations between practice location, volume and type with PGS/PGD availability and reported techniques. DESIGN: Cross-sectional evaluation. MATERIALS AND METHODS: Between 3/2017-4/2017, 389 SART member fertility clinics were systematically examined by two separate reviewers. Websites were surveyed for: size/type of practice, location, whether PGS or PGD were mentioned, whether aneuploidy screening or testing for single gene disorders were discussed, whether sex selection was offered, whether the issue of mosaicism was discussed, the day of biopsy, and the type of platform used for PGS. Chi-squared tests were used to evaluate the differences between the groups. RESULTS: Of the 376 websites evaluated, 79% were private practice and 21% were academic. 30% of clinics performed >500 cycles a year and 70% performed <500 cycles a year. 85% reported PGS/PGD availability, with the majority describing the benefits of PGS/PGD for aneuploidy screening (80%) and testing for single gene diseases (89%). 31% mentioned sex selection as an indication for PGS. Larger practices (>500 cycles/year) were more likely to report PGS/PGD availability (98% vs 79%, p<0.01) and describe aneuploidy screening (87% vs 74%, p¼0.01) as well as single gene disorders (95% vs 86%, p¼0.01) than smaller practices (<500 cycles). Private practices were more likely than academic practices to describe PGS for aneuploidy screening (83% vs 67%, p¼0.01) or sex selection (37% vs 6%, p<0.01). Of the clinics that listed the day of embryo biopsy for PGS/PGD, 14% described day 3 biopsies, 32% described day 5 biopsies, and 8% described both. Most did not describe the platform used for PGS; 8% described array comparative genomic hybridization (aCGH), 6% described fluorescence in-situ hybridization (FISH), 5% described next-generation sequencing (NGS), and 4% described multiple technologies. Only 7% described the possibility of mosaic results. Practice location had no impact on any of these parameters.
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CONCLUSIONS: While most practices describe PGS/PGD on their websites, reported indications for PGS/PGD varies from type and size of practice. Interestingly, 14% of clinics describe day 3 embryo biopsies, despite recent data demonstrating reduced implantation rates2 and 6% describe FISH despite the availability of technologies associated with increased aneuploidy detection rates. While an increasing number of patients use the Internet as their primary source of medical information, this information is often inconsistent and outdated. References: 1. Fiorentino F, Bono S, Biricik A, Nuccitelli A, Cotroneo E, Cottone G, Kokocinski F, Michel CE, Minasi MG, Greco E. Application of nextgeneration sequencing technology for comprehensive aneuploidy screening of blastocysts in clinical preimplantation genetic screening cycles. Hum Reprod. 2014 Dec;29(12):2802-13. 2. Scott RT Jr, Upham KM, Forman EJ, Zhao T, Treff NR. Cleavage-stage biopsy significantly impairs human embryonic implantation potential while blastocyst biopsy does not: a randomized and paired clinical trial. Fertil Steril. 2013 Sep;100(3):624-30. P-474 Wednesday, November 1, 2017 VERSICAN AND PROSTAGLANDIN-ENDOPEROXIDE SYNTHASE 2 (PTSG2) GENE EXPRESSION IN CUMULUS CELLS AS A COMPLEMENT OF PREIMPLANTATION GENETIC TESTING: BETTER OUTCOMES FOR IN VITRO FERTILIZATION PREGNANCY. E. Lopez-Bayghen,a E. Schaeffer,b G. M. Ortiz Olivera,c J. Pedraza,d A. Ocampo-Barcenas.e aToxicology, Cinvestav-IPN, Mexico, Mexico; bGenetics and Molecular Biology, Cinvestav-IPN, Mexico City, Mexico; cLidmol and Reproductive Medicine, Instituto de Infertilidad y Genetica, Ingenes Mexico, Ciudad de Mexico, Mexico; d FIV Lab, Instituto de Infertilidad y Genetica, Ingenes Mexico, Mexico, Mexico; e Lidmol, Instituto de Infertilidad y Genetica, Ingenes Mexico, Mexico, Mexico. OBJECTIVE: To determine if a combination of PTGS2-VCAN-L19 expression score in cumulus cells (CCs), plus the evaluation of genomic integrity with Preimplantation Genetic Testing (PGT) methods can be used together to predict implantation outcomes. DESIGN: Prospective study with 30 infertile patients attending Ingenes Institute for an ART procedure with a signed informed consent. Patients were subjected to controlled ovarian stimulation protocol with GnRH. In previous work, we developed a formula using the expression values of PTGS2 and VCAN, the PTGS2-VCAN-L19 (PVL) index score to predict fertilization and implantation outcomes. For embryo selection, CCs gene expression profiles (PVL index) were combined with Comparative Genomic Hybridization (CGH) and in other hand, we use Whole Genome Amplification (WGA) as template source for PCR as a rapid, inexpensive, and highly sensitive option for genotyping Trisomies 21 and 18. MATERIALS AND METHODS: CCs were obtained during oocyte retrieval and RNA was isolated using Trizol per manufacturer’s suggestions. The expression of PTGS2, VCAN, and L19 was measured by qPCR and the PVL index [(PTGS2+VCAN)*L19normalized] was determined. Embryo biopsy was performed at day 5. Trophectoderm cells were retrieved and used for WGA, which was then analyzed by CGH or PCR. WGA provided sufficient DNA to determine Trisomy 18 and 21 by examining the Threshold Cycle (CT) difference for RPL17 and TTC3 genes, respectively. After 5 days in culture, a single embryo presenting no chromosomopathies (CGH) or trisomy (PCR), with high morphological score and PVL score R58 was transferred. Pregnancy was confirmed by bhCG and the presence of a fetal heartbeat. RESULTS: Patient’s age was 36.5 years’ average. BMI was 24.24. For embryo selection, the combination of high morphological score, a PVL score equal or higher than 58 and absence of aneuploidies determined via CGH or WGA-genotyping were used as criteria. 95% of transfers performed under this criterion favored one embryo transfer pregnancies. The simultaneous detection of PVL index in CC from individual oocytes combined with CGH or PCR genotyping works as a suitable system that can improve embryo selection in IVF. CONCLUSIONS: This tool will allow to improve the embryonic selection increasing the rate of pregnancy and decreasing the probability of implanting embryos with most common viable trisomies. This system is intended to reduce the implantation failure and miscarriage rates. Reference: 1. Huang Z, Wells D. The human oocyte and cumulus cells relationship: new insights from the cumulus cell transcriptome. Molecular human reproduction. 2010 Oct;16(10):715-25. PubMed PMID: 20435609. Supported by: Proinnova Conacyt.
Vol. 108, No. 3, Supplement, September 2017
Eq. 1
P-473 Wednesday, November 1, 2017 ASSESSMENT OF FERTILITY CLINIC WEBSITES ON PREIMPLANTATION GENETIC SCREENING (PGS) AND PREIMPLANTATION GENETIC DIAGNOSIS (PGS). N. Joshi,a,b T. Zore,a,b S. B. Schon,c P. Masson,d J. L. Chan.b aObstetrics and Gynecology, University of California Los Angeles, Los Angeles, CA; bReproductive Endocrinology and Infertility, Cedars-Sinai Medical Center, Los Angeles, CA; cUniversity of Michigan, Ann Arbor, MI; dUniversity of Pennsylvania, Philadelphia, PA. OBJECTIVE: In recent years, improvements in PGS/PGD have led to high implantation rates.1 We sought to determine whether Society of Assisted Reproductive Technology (SART) member clinics mention PGS/PGD on their websites and investigate associations between practice location, volume and type with PGS/PGD availability and reported techniques. DESIGN: Cross-sectional evaluation. MATERIALS AND METHODS: Between 3/2017-4/2017, 389 SART member fertility clinics were systematically examined by two separate reviewers. Websites were surveyed for: size/type of practice, location, whether PGS or PGD were mentioned, whether aneuploidy screening or testing for single gene disorders were discussed, whether sex selection was offered, whether the issue of mosaicism was discussed, the day of biopsy, and the type of platform used for PGS. Chi-squared tests were used to evaluate the differences between the groups. RESULTS: Of the 376 websites evaluated, 79% were private practice and 21% were academic. 30% of clinics performed >500 cycles a year and 70% performed <500 cycles a year. 85% reported PGS/PGD availability, with the majority describing the benefits of PGS/PGD for aneuploidy screening (80%) and testing for single gene diseases (89%). 31% mentioned sex selection as an indication for PGS. Larger practices (>500 cycles/year) were more likely to report PGS/PGD availability (98% vs 79%, p<0.01) and describe aneuploidy screening (87% vs 74%, p¼0.01) as well as single gene disorders (95% vs 86%, p¼0.01) than smaller practices (<500 cycles). Private practices were more likely than academic practices to describe PGS for aneuploidy screening (83% vs 67%, p¼0.01) or sex selection (37% vs 6%, p<0.01). Of the clinics that listed the day of embryo biopsy for PGS/PGD, 14% described day 3 biopsies, 32% described day 5 biopsies, and 8% described both. Most did not describe the platform used for PGS; 8% described array comparative genomic hybridization (aCGH), 6% described fluorescence in-situ hybridization (FISH), 5% described next-generation sequencing (NGS), and 4% described multiple technologies. Only 7% described the possibility of mosaic results. Practice location had no impact on any of these parameters.
e288
ASRM Abstracts
CONCLUSIONS: While most practices describe PGS/PGD on their websites, reported indications for PGS/PGD varies from type and size of practice. Interestingly, 14% of clinics describe day 3 embryo biopsies, despite recent data demonstrating reduced implantation rates2 and 6% describe FISH despite the availability of technologies associated with increased aneuploidy detection rates. While an increasing number of patients use the Internet as their primary source of medical information, this information is often inconsistent and outdated. References: 1. Fiorentino F, Bono S, Biricik A, Nuccitelli A, Cotroneo E, Cottone G, Kokocinski F, Michel CE, Minasi MG, Greco E. Application of nextgeneration sequencing technology for comprehensive aneuploidy screening of blastocysts in clinical preimplantation genetic screening cycles. Hum Reprod. 2014 Dec;29(12):2802-13. 2. Scott RT Jr, Upham KM, Forman EJ, Zhao T, Treff NR. Cleavage-stage biopsy significantly impairs human embryonic implantation potential while blastocyst biopsy does not: a randomized and paired clinical trial. Fertil Steril. 2013 Sep;100(3):624-30. P-474 Wednesday, November 1, 2017 VERSICAN AND PROSTAGLANDIN-ENDOPEROXIDE SYNTHASE 2 (PTSG2) GENE EXPRESSION IN CUMULUS CELLS AS A COMPLEMENT OF PREIMPLANTATION GENETIC TESTING: BETTER OUTCOMES FOR IN VITRO FERTILIZATION PREGNANCY. E. Lopez-Bayghen,a E. Schaeffer,b G. M. Ortiz Olivera,c J. Pedraza,d A. Ocampo-Barcenas.e aToxicology, Cinvestav-IPN, Mexico, Mexico; bGenetics and Molecular Biology, Cinvestav-IPN, Mexico City, Mexico; cLidmol and Reproductive Medicine, Instituto de Infertilidad y Genetica, Ingenes Mexico, Ciudad de Mexico, Mexico; d FIV Lab, Instituto de Infertilidad y Genetica, Ingenes Mexico, Mexico, Mexico; e Lidmol, Instituto de Infertilidad y Genetica, Ingenes Mexico, Mexico, Mexico. OBJECTIVE: To determine if a combination of PTGS2-VCAN-L19 expression score in cumulus cells (CCs), plus the evaluation of genomic integrity with Preimplantation Genetic Testing (PGT) methods can be used together to predict implantation outcomes. DESIGN: Prospective study with 30 infertile patients attending Ingenes Institute for an ART procedure with a signed informed consent. Patients were subjected to controlled ovarian stimulation protocol with GnRH. In previous work, we developed a formula using the expression values of PTGS2 and VCAN, the PTGS2-VCAN-L19 (PVL) index score to predict fertilization and implantation outcomes. For embryo selection, CCs gene expression profiles (PVL index) were combined with Comparative Genomic Hybridization (CGH) and in other hand, we use Whole Genome Amplification (WGA) as template source for PCR as a rapid, inexpensive, and highly sensitive option for genotyping Trisomies 21 and 18. MATERIALS AND METHODS: CCs were obtained during oocyte retrieval and RNA was isolated using Trizol per manufacturer’s suggestions. The expression of PTGS2, VCAN, and L19 was measured by qPCR and the PVL index [(PTGS2+VCAN)*L19normalized] was determined. Embryo biopsy was performed at day 5. Trophectoderm cells were retrieved and used for WGA, which was then analyzed by CGH or PCR. WGA provided sufficient DNA to determine Trisomy 18 and 21 by examining the Threshold Cycle (CT) difference for RPL17 and TTC3 genes, respectively. After 5 days in culture, a single embryo presenting no chromosomopathies (CGH) or trisomy (PCR), with high morphological score and PVL score R58 was transferred. Pregnancy was confirmed by bhCG and the presence of a fetal heartbeat. RESULTS: Patient’s age was 36.5 years’ average. BMI was 24.24. For embryo selection, the combination of high morphological score, a PVL score equal or higher than 58 and absence of aneuploidies determined via CGH or WGA-genotyping were used as criteria. 95% of transfers performed under this criterion favored one embryo transfer pregnancies. The simultaneous detection of PVL index in CC from individual oocytes combined with CGH or PCR genotyping works as a suitable system that can improve embryo selection in IVF. CONCLUSIONS: This tool will allow to improve the embryonic selection increasing the rate of pregnancy and decreasing the probability of implanting embryos with most common viable trisomies. This system is intended to reduce the implantation failure and miscarriage rates. Reference: 1. Huang Z, Wells D. The human oocyte and cumulus cells relationship: new insights from the cumulus cell transcriptome. Molecular human reproduction. 2010 Oct;16(10):715-25. PubMed PMID: 20435609. Supported by: Proinnova Conacyt.
Vol. 108, No. 3, Supplement, September 2017