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Human embryo transfer11

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Human embryo transfer11

  1. 1. Key Success Factors For a Successful IVF Laboratory Gloria Calderón PhD IVI-Barcelona gloria.calderon@ivi.es www.ivi.es
  2. 2. Factors Affecting the Success of ART Diagnosis and therapeutic guidance Ovarian stimulation Oocyte retrieval IVF procedures Embryo transfer Luteal phase treatment TEAMWORK
  3. 3. Why? Laboratory improvements affects directly in the ART programs results. Program results are mostly affected by laboratory variations Rackowsky et al. 11th World Congress on IVF and Human Reproductive Genetics. Sydney, Australia 1999.
  4. 4. Quality Control 1. Everybody must follow standard operating protocols 2. Pay attention to every single detail 3. Internal tests passed by laboratory staff every 6 months. 4. Daily monitoring of results 5. Stable culture system to control: A. Temperature B. pH C. Osmolarity
  5. 5. Human resources 2000 cycles/year 6 embryologists 10 technicians 1035 cycles/year 13 (10) embryologists 800 cycles/year 5 embryologists 6 technicians
  6. 6. ENOUGH TIME FOR EVERY CYCLE Optimal ratio of laboratory staff to ensure high attention during the procedures CONCENTRATION Each sample must be handled individually. RESPONSABILITY For the correct application of the protocols in the laboratory during ART procedures
  7. 7. Air Quality Influence Cohen et al., 1997 Hum Reprod 12:1742-1749
  8. 8. Air Quality Influence
  9. 9. What might be used in the near future? • Morphology assessment of human embryo • Time-Lapse (embryo kinetics) • Cryopreservation (Vitrification) • PGS (on demand) • Transfer at the Blastocyst stage routinely (SET) • “Omics”
  10. 10. What we are used to - Embryo Prediction Model: Static Morphology Number of blastomeres with visible Number of blastomeres Symmetry single nucleus Degree of fragmentation Multinucleation The Istanbul consensus workshop on embryo assessment: proceedings of an expert meeting Alpha Scientists in Reproductive Medicine and ESHRE Special Interest Group of Embryology; Human Reproduction 2011; doi: 10.1093/humrep/der037
  11. 11. Historic and Current Approaches Day 1 Day 2 Day 3 Day 4 Day 5 Oocyte morphology: Dimorphism: cytoplasm, zona, PB’s Fertilization Checks: 2PN Assessments – Z scoring Early cleavage/ Morphologic evaluation Genetic screen ( PGD); metabolic evaluation, morphologic evaluation; extended culture decision Signals of compaction Selection blastocyst for transfer, embryo cryopreservation, PGD Day 0
  12. 12. Post-Modern lab: Time-lapse Assessment
  13. 13. Exact Timing, Developmental Events, Kinetics • cc1=t2: First cell cycle • cc2=t3-t2: Second cell cycle, duration of period as 2 cells • s2=t4-t3: Synchrony in division from 3 to 4 blastomere embryo 1 2 4 8 Meseguer et al. Hum Reprod, 2011;10:2658.
  14. 14. Embryo Development until day 3 Stage using EmbryoScope
  15. 15. Development Anomalies on day 2
  16. 16. PGS using Embryoscope
  17. 17. Embryo Development until Blastocyst Stage using EmbryoScope
  18. 18. Cryopreservation in ART Advantages • Great flexibility in infertility treatments • Makes feasible postponement of embryo transfer (ET) for future cycles (OHSS, other multiple reasons) • Contributes to decrease the incidence of multiple pregnancies (SET) • Contributes to the overall success of a single stimulation cycle by increasing the cumulative pregnancy rate
  19. 19. Slow Freezing Vitrification Physiological solution Cryoprotectant solution Vitrification solution Before cooling During cooling In LN2 Ice seeding Slow cooling Rapid cooling Ultra rapid 0.3ºC/min cooling 200,000ºC/min Cryopreservation Techniques
  20. 20. The Post-Modern Lab: Standard Cleavage Stage/Blastocyst Vitrification Vitrification appears to be associated with significantly higher post-thawing survival rates than slow freezing. Loutradi et al. Fertil Steril 2008;90:186.
  21. 21. The clinical pregnancy rate has doubled with the introduction of vitrification Tulandi, 2008; Cao et al., 2009; Smith et al., 2010 Efficiency in donation program not compromised with vitrification (RCT) Cobo et al., 2007; 2011; Nagy et al., 2007 Prospective randomized study with own sibling oocytes demonstrates the lab efficiency of the technique (RCT) Rienzi et al., 2010 Cumulative ongoing pregnancy rate with oocyte vitrification without embryo selection in a standard infertility program is comparable to what is obtained with embryo cryopreservation Ubaldi et al., 2010; Schoolcraft et al., 2009; Chian et al., 2008; Kim et al., 2010; Garcia et al., 2011 The Post-Modern Lab: Standard Oocyte Vitrification
  22. 22. Fresh oocytes 37ºC Donors oocytes Vitrified oocytes 2 hours After OPU Warming 1 hour after + ICSI 2 hours after warming
  23. 23. Oocyte Vitrification: Clinical Evidence Cobo et al. Fertil Steril 2011;96:277.
  24. 24. IVF Lab Will Resolve PGS Controversy • Mastenbroek et al. N Engl J Med. 2007;357:9-17. – 408 women 35-41 years: 206 PGS (434 IVF cycles), 202 Control (402 IVF cycles). – Ongoing pregnancy rate: PGS 25% vs Control 37%. – Live-birth rate: PGS 24% vs Control 35%. – PGS significantly reduced the rates of ongoing pregnancies (viable intrauterine pregnancy after 12 weeks of gestation) and live births after IVF in women of advanced maternal age. • Hardarson et al. Hum Reprod. 2008;23:2806-2812. – 118 women ≥38 years: 56 PGS, 53 Control (Halted accrual at interim analysis) – Clinical pregnancy rate: PGS 8.9% vs Control 24.5% (P=0.039). – Concluded study provided evidence against using PGS for AMA patients when performing IVF. • Schoolcraft et al. Fertil Steril. 2009;92:157-62. – 62 women: 32 PGS, mean age 38.3 years, 30 Control, mean age 38.2 years. – Spontaneous abortion rate: PGS 25.9% vs Control 32.3% – Delivery rate: PGS 78% vs Control 67.7% – In infertile AMA patients PGS did not significantly improve outcome parameters, but a trend toward a decrease in spontaneous abortion rate with higher delivery rate was observed.
  25. 25. PGS: RCT Advanced Maternal age: 41-44 yrs (2009- 2011) Inclusion criteria: • Age range: 41-44 years • Number MII oocytes: ≥6 MII in a single fresh cycle or in two cycles (fresh+vitrified) • Without chromosomal abnormalities in previous pregnancies • < 2 previous miscarriages • < 3 previous IVF/ICSI cycles 232 patients informed 25 rejected 90 patients Blastocyst 24 did not meet inclusion criteria Rubio et al., Poster ESHRE 2012 93 patients PGS 183 patients enrolled
  26. 26. RCT Advanced Maternal Age 41-44 yrs (2009-2011) Blastocyst PGS P-value No. of cycles 90 93 Mean Age (SD) 41.7 (0.9) 41.8 (0.9) ---- No. of transfers (%) 73 (82.9) 70 (75.3) ---- % Abnormal embryos ---- 69.7 ---- Mean embryos transferred (SD) 2.0 (0.8) 1.6 (0.6) ---- Ongoing PR/transfer (%) 14/74 (18.9) 30/70 (42.8) p=0.0021 OR 3.214, CI [1.518-6.806] Ongoing PR/retrieval (%) 14/90 (15.5) 30/93 (32.3) p=0.0099 OR 2.585, CI [1.262-5.295] Ongoing implantation rate (%) 20/152 (13.1) 40/114 (35.1) P<0.0001 OR 3.568, CI [1.943-6.551] *Two-sides Fisher’s test
  27. 27. PGS Results IVI – Barcelona Nb Oocytes (X ± DS) 1906 (9.5 ± 5.6) Nb Biopsied E (%) 1358 (91.3) Nb no informatives(%) 29 (2.2) Nb abnormal (%) 879 (68.6) % Transfers 64.2 E Transferred (X ± DS) 1.63 ± 0.8 % + BHCG 68.8 % Clinical pregnancy 64.3 % Implantation 54.2 % Miscarriage 17.2 % Twin pregnancies 33.3 % Triplets 1.9 Period: 06/2010-07/2011 Nb=242 X Age= 39.29 ± 3.7 ALL INDICATIONS Key factor for PGS success:  Highly trained team  Low O2 tension incubators  Stable culture system  Culture medium up to Day 5 (LifeGlobal ® Media)  Protein support (LifeGlobal ® LGPS) Calderón G. ALPHA 2012, London
  28. 28. IVI-BARCELONA Results: Prospective, Randomized Trial May-July 2011 Global Global TOTAL p Nb of cycles 50 42 X Age 35,48 ± 0,13 36,71 ± 0,6 NS (0,07) X oocytes 9,44 ± 1,65 8,76 ± 1,31 NS Nb cells D2 3,99 ± 0,97 3,81 ± 1,09 NS % fragments D2 6,96 ± 3,68 6,69 ± 3,09 NS Nb cells D3 7,96 ± 1,65 7,32 ± 1,67 p 0,02 % fragments D3 6,76 ± 3,88 6,48 ± 3,88 NS Good Quality Embryos 55,16% (187/339) 53,11% (128/241) NS Nb Transfers 48 40 X Embryos Transferred 2 ± 0,16 2,08 ± 0,18 NS X Embryos Frozen 1,82 ± 0,86 1,07 ± 0,6 NS Cancellation Rate 4% (2/50) 5% (2/42) NS Implantation Rate 34% 31% NS Pregnancy Rate 50% (24/48) 45% (18/40) NS Multiple Preg Rate 29,17% (7/24) 33,33 (6/18) NS Miscarriage Rate 4,17% (1/24) 5,55% (1/18) NS
  29. 29. ‘OMICS’ Technologies Genome DNA Transcriptome ≈25,000 Genes RNA Proteome Metabolome Proteome Metabolites ≈1,000,000 Proteins Chromosomes ≈2500 metabolites Transcription Translation 23 pairs
  30. 30. Post-Modern Lab: Future Directions in ‘Omics’ • Development comprehensive Omic database • “Grand Unification” of transcriptomic/ proteomic and metabolomic technologies Assou et al. 2011 HRU.
  31. 31. Single Embryo Transfer The ideal situation to avoid multiple pregnancies would be to perform SET routinely when: 1. Couple meets indication, basically woman’s age 2. Perfect culture system till blastocyst stage 3. Large cohort of good quality embryos 4. Extremely good cryopreservation program 5. Controlled randomized trials have demonstrated cumulative pregnancy rates of SET+ FET are equal to DET
  32. 32. Single Embryo Transfer
  33. 33. Single Embryo Transfer Conclusion: eSET is associated with decrease risk of PTB and LBW compared with DET but higher risk of PTB when compared with naturally conceived singletons. Grady et al. Fertil Steril 2012;97:324-31
  34. 34. Conclusion The post-modern laboratory will have:  Improved techniques to evaluate embryos - Time-lapsed morphology assessments with computerized algorithms linked to clinical outcomes  Optimized cryopreservation (vitrification) techniques to with higher survival, pregnancies and implantation rates  Understands value of PGS  Utilizes and unifies all data from ‘Omics’ to help more accurate embryo selection  Allows for SET – maximizing child outcome  Unify criteria in quality control and quality assurance
  35. 35. Gracias por su atención Thank you for your attention IVI-Barcelona Ronda General Mitre 14, 08017 Barcelona Tel. +3493 206 3000 – Fax. +3493 205 34 28 ivibarcelona@ivi.es

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