Diagnosis and Management of Poor Ovarian Reserve : Evidence & Practice

Invited Lecture by Dr Sujoy Dasgupta in a CME organized by Mankind Zesteva Division held at kolkata on 4 March, 2023

1. Diagnosis and Management of Poor Ovarian Reserve : Evidence & Practice Dr Sujoy Dasgupta MBBS (Gold Medalist, Hons ) MS (OBGY- Gold Medalist) DNB (New Delhi) MRCOG (London) Advanced ART Course for Clinicians (NUHS, Singapore ) M Sc, Sexual and Reproductive Medicine (South Wales, UK) Consultant: Reproductive Medicine, Genome Fertility Centre, Kolkata Managing Committee Member, BOGS, 2022-23 Executive Committee Member, ISAR Bengal, 2022-24 Clinical Examiner, MRCOG Part 3 Examination Winner, Prof Geoffrey Chamberlain Award, RCOG World Congress, London, 2019

2. Boring discussion Endocrinology Physiology Biochemistry Pharmacology Statistics IVF

3. Declining ovarian reserve is THE RULE

4. Declining ovarian reserve is THE RULE Exogenous FSH

5. Terminology Ovarian Reserve- Quantity and quality of remaining oocytes present in both the ovaries at a given age Ovarian Responsiveness- Number of oocytes developed/ retrieved after COH Ovarian Ageing- Decline in quantity and quality of ovarian activity because of individualized rhythm of the “biological clock”

6. Ovarian Reserve Tests Commonly Used Age AMH AFC FSH Others Inhibin B Ovarian Volume Ovarian blood flow (Doppler) Dynamic Tests

7. Female Age- Most valuable parameter

8. Day3 FSH FSH >15 IU/L + E2 >75 pg/ml (>200 pmol /L) Cycle dependent Inter-cycle variation Single abnormal FSH- not reliable <40 yr of age

9. Antral Follicle Count (AFC) Day 2-4 TVS- 2-9 mm follicles Cut off 5-15 Direct measure of cohort of follicles capable of responding to the stimulation Observer and machine dependent Problematic in presence of ovarian mass

10. Anti-M ű llerian Hormone (AMH)  Paracrine control, independent of HPO endocrine feedback Correlates with AFC, FSH, Inhibin B, E2 Higher sensitivity and specificity than conventional markers Declines earlier than FSH rise

11. Minimum variability of AMH

12. Intra-Class Coefficient Fanchin R, Taieb J, Lozano DH, Ducot B, Frydman , R, Bouyer J. High reproducibility of serum anti- Mullerian hormone measurements suggests a multistaged follicular secretion and strengthens its role in the assessment of ovarian follicular status. Hum Reprod 2005; 20(4): 923–7.

13. Factors affecting AMH

14. Problem with AMH Dispute on lower and higher level Poor correlation with oocyte quality and chance of conception

15. Low AMH ≠ Donor oocyte The predictive accuracy of AMH for 1-year CLBR in GnRH antagonist treatment cycles was limited and did NOT yield much additional value on top of age. Withholding treatment based on predictors such as age and AMH, or a combination, remains problematic. Hamdine O, Eijkemans MJ, Lentjes EW, Torrance HL, Macklon NS, Fauser BC, Broekmans FJ. Antimüllerian hormone: Prediction of cumulative live birth in gonadotropin -releasing hormone antagonist treatment for in vitro fertilization. Fertil Steril 2015; 104: 891–8.

16. Discordance between ovarian reserve tests Approximately 1/5 patients in clinical practice had discordance in their AFCs and AMH levels (Zhang et al., Reprod Bio Online, 2019) Laboratory vs Machine? Intermediate prognosis

17. AFC vs AMH The faster decline in AMH than in AFC with age Their correlation changes with age. AMH and AFC showed a very low proportion of concordance in the range of expected poor responders according to Bologna cutoffs. The reproducibility for AMH seemed much better than for AFC. Arvis P, Rongières C, Pirrello O, Lehert P. Reliability of AMH and AFC measurements and their correlation: a large multicenter study. J Assist Reprod Genet. 2022 May;39(5):1045-1053. doi : 10.1007/s10815-022-02449-5. Epub 2022 Mar 3. PMID: 35243569; PMCID: PMC9107554.

18. Zhang, Y., Xu , Y., Xue , Q.  et al.  Discordance between antral follicle counts and anti- Müllerian hormone levels in women undergoing in vitro fertilization.  Reprod Biol Endocrinol  17, 51 (2019). https://doi.org/10.1186/s12958-019-0497-4 AFC being better than the AMH level for predicting POR AFC should be preferred in the prediction of ovarian response, to ultimately develop an optimal individualized COH protocol.

19. Ovarian Reserve Quantitative Poor response to COH Qualitative No test can predict Age

20. Qualitative Ovarian Reserve

21. Age and Euploidy Capalbo A et al.. Hum Reprod Update 2017; 23:706–722 .

22. Dynamic tests for Ovarian reserve CC challenge test EFORT Previous response to ovarian stimulation using sufficient dose of FSH

23. Ovarian Response Ovarian reserve = f ( recruitable follicles) Follicular sensitivity to FSH = f (FSH receptors) Exogenous Gonadotropin = f ( pharmaco - kinetics and dynamics)

24. IVF is stressful for Fertility Physicians

25. How many oocytes should be retrieved? Law YJ, Zhang N, Kolibianakis EM, Costello MF, Keller E, Chambers GM, Venetis CA. Is there an optimal number of oocytes retrieved at which live birth rates or cumulative live birth rates per aspiration are maximized after ART? A systematic review. Reprod Biomed Online. 2021 Jan;42(1):83-104. doi : 10.1016/j.rbmo.2020.10.008. Epub 2020 Oct 22. PMID: 33390313.

26. Number oocytes ≈ CLBR Law et al., Reprod Biomed Online. 2021

27. Defining Poor Responders

28. A young woman with a poor response >>> older woman with a poor response Female age and number of oocytes retrieved in particular will modulate the chances for pregnancy in current and subsequent cycles. Poor Responders ≠ Same Oudendijk JF, Yarde F, Eijkemans MJ, Broekmans FJ, Broer SL. The poor responder in IVF: is the prognosis always poor?: a systematic review. Hum Reprod Update. 2012 Jan-Feb;18(1):1-11. doi : 10.1093/ humupd /dmr037. Epub 2011 Oct 10. PMID: 21987525.

29. ESHRE Bologna: One size does NOT fit for all

30. Poseidon (Patient-Oriented Strategies Encompassing IndividualizeD Oocyte Number)

31. Front. Endocrinol . 10:387. doi : 10.3389/fendo.2019.00387 POSEIDON

32. Young women Older women Good Ovarian Reserve Group 1 Group 2 Poor Ovarian Reserve Group 3 Group 4

33. Good Ovarian Reserve Group 1 Group 2 Poor Ovarian Reserve Group 3 Group 4 Unexpected poor response Expected poor response Asynchrony Gn Receptor gene polymorphism Low dose Gn (like high BMI) Less potent Gn Unexplained Ovarian aging Ovarian insult (surgery) Genetic ( Karyo , FMR1)

34. Young women Older women Good Ovarian Reserve Group 1 Group 2 Poor Ovarian Reserve Group 3 Group 4 Low risk of Aneuploidy High risk of Aneuploidy

35. CLBR depends on POSEIDON group

36. Personalized Treatment Haahr , T.; Dosouto , C.; Alviggi , C.; Esteves , S.C.; Humaidan , P. Management Strategies for POSEIDON Groups 3 and 4.  Front. Endocrinol .   2019 ,   11 , 99. doi:10.3389/fendo.2019.00614. 

37. Personalized Treatment Haahr , T.; Dosouto , C.; Alviggi , C.; Esteves , S.C.; Humaidan , P. Management Strategies for POSEIDON Groups 3 and 4.  Front. Endocrinol .   2019 ,   11 , 99. doi:10.3389/fendo.2019.00614. 

38. Stimulation regimes for POR High dose Gn No benefits >300 IU FSH GnRH Agonist protocols Long protocol Oversuppression Short protocol (Flare-up) Theoretically less suppression Microdose flare up protocol Cost-friendly Ultra-short protocol Not much benefit Cessation/ Stop protocol No additional advantage GnRH Antagonist protocols Flexible vs fixed protocol Most widely used GnRH Agonist-antagonist protocol No t much benefit Natural cycle Not recommended Modified natural cycle Not recommended Mild stimulation protocol CC/ letrozole + Gn + Antag No additional benefit Luteal phase stimulation No additional benefit Dual stimulation Needs further studies

39. Try differently than before Every cycle is different

40. Every cycle is different- The "Waves" concept Baerwald AR, Adams GP, Pierson RA. Ovarian antral folliculogenesis during the human menstrual cycle: a review. Hum Reprod Update. 2012 Jan-Feb;18(1):73-91. doi : 10.1093/ humupd /dmr039. Epub 2011 Nov 8. PMID: 22068695.

41. ESHRE, 2019 GnRH antagonists and GnRH agonists are equally recommended for predicted poor responders. Clomiphene citrate alone or in combination with gonadotrophins , and gonadotropin stimulation alone are equally recommended for predicted poor responders. The addition of letrozole to gonadotropins in stimulation protocols is probably not recommended for predicted poor responders. A gonadotropin dose higher than 300 IU is not recommended for predicted poor responders. The use of modified natural cycle is probably not recommended over conventional ovarian stimulation for predicted poor responders. Random-start ovarian stimulation is probably not recommended for the general IVF/ICSI population. Late luteal phase start of gonadotropins is probably not recommended for poor responders. Early luteal phase start of gonadotropins is probably not recommended for normal and poor responders. Luteal phase stimulation could be used in non-transfer cycles. Double stimulation in poor responders should only be used in the context of clinical research.

42. WHO, 2017

43. Role of adjuvants

44. Androgen

45. Two-Cell, Two- Gonadotrophin Theory

46. Hum Reprod Update , Volume 22, Issue 6, 20 November 2016, Pages 709–724, https:// doi.org/10.1093/humupd/dmw027 Two Triangle Theory

47. Androgen works before FSH

48. Site of action of Androgen

49. Low androgen = Low IVF success Frattarelli JL, Peterson EH. Effect of androgen levels on in vitro fertilization cycles. Fertil Steril . 2004 Jun;81(6):1713-4. doi : 10.1016/j.fertnstert.2003.11.032. PMID: 15193506.

50. Barbieri RL , Sluss PM, Powers RD, McShane PM, Vitonis A, Ginsburg E, Cramer DC. Association of body mass index, age, and cigarette smoking with serum testosterone levels in cycling women undergoing in vitro fertilization. Fertil Steril . 2005 Feb;83(2):302-8. doi : 10.1016/j.fertnstert.2004.07.956. PMID : 15705366. Serum T decreased significantly throughout the 4 th decade of life (P<.03). Serum T correlated positively with pre- hCG serum E2 and number of oocytes retrieved. However, serum T did NOT significantly influence fertilization or pregnancy rates.

51. Strategies to increase androgen T or DHEA supplementation before IVF Blocking intraovarian androgen conversion with the use of an aromatase enzyme inhibitor Stimulating theca cells with r- hLH prior to r- hFSH stimulation in the long agonist protocol

52. Testosterone Author Year Definition of POR No. of patients Dose Duration Stimulation protocol Primary outcome Massin et al. (29) 2006 * Previous POR (Peak E2<1200pg/mL and ≤5 oocytes) and D3 FSH > 12 IU/L or E2 > 70pg/mL or Inhibin B <45ng/mL 49 10 mg/d 15-20 d NR Total number of retrieved oocytes Fábregues et al. (30) 2009 Previous POR and 31-39y 62 20 ug /kg/d 5 d Long GnRH agonist Incidence of low responders Kim et al. (31) 2011 Previous cycle with ≤3 oocytes retrieved despite high Gn dose 110 12.5 mg/d 21 d GnRH antagonist Number of MII oocytes retrieved Kim et al. (32) 2014 Previous cycle with ≤3 oocytes retrieved despite high Gn dose 120 12.5 mg/d I1: 14 d/ I2: 21 d/ I3: 28 d GnRH antagonist Number of MII oocytes retrieved Marzal Escriva ́ et al. (33) 2015 ≥2: ≥38y, AFC ≤6, FSH ≥10 IU/L, AMH ≤5pg/mL AND ≤4 follicles of ≥16 mm on the day of trigger or E2 ≤500 pg /mL on the day of trigger or ≤ 4 MII 66 20 ug /kg/d 7 d GnRH antagonist Number of MII oocytes retrieved Bosdou et al. (34) 2016 Bologna criteria 50 10 mg/d 21 d Long GnRH agonist Total number of retrieved oocytes Saharkhiz et al. (35) 2018 * Bologna criteria 48 25 mg/d During COS GnRH antagonist NR Front Endocrinol (Lausanne). 2021; 12: 653857 Effect on follicular development was reported with transdermal testosterone 20 μ g/kg/d obtained with 1.25mg/d gel application or 2.5mg/d patch Higher dose usage above physiological range, question the potential benefit (or harm) of using high doses of testosterone

53. Dehydroandrosterone (DHEA) 75 mg/day 3 months Orally well absorbed Converted to DHEAS in intestinal wall and liver Interferes with progesterone assay Side effects- acne, hirsutism - minimum Contraindication- PCOS and androgen excess

54. Nagels HE, Rishworth JR, Siristatidis CS, Kroon B. Androgens ( dehydroepiandrosterone or testosterone) for women undergoing assisted reproduction. Cochrane Database Syst Rev. 2015 Nov 26;(11):CD009749. doi : 10.1002/14651858.CD009749.pub2. PMID: 26608695. In women identified as poor responders undergoing ART, pre-treatment with DHEA or testosterone may be associated with improved live birth rates. The overall quality of the evidence is moderate . There is insufficient evidence to draw any conclusions about the safety of either androgen. Definitive conclusions regarding the clinical role of either androgen awaits evidence from further well-designed studies.

55. The pooled analysis showed that the clinical pregnancy rates were increased significantly in DOR patients who were pre-treated with DHEA (OR=1.47, 95% CI: 1.09-1.99) However, it is worth noting that when data were restricted to RCTs , there was a non-significant difference in the clinical pregnancy rate (OR=1.08, 95% CI: 0.67-1.73). No differences were found in the number of oocytes retrieved, the cancellation rate of IVF cycles and the miscarriage rate between the cases and controls (WMD= -0.69, 95% CI: -2.18-0.81; OR=0.74, 95% CI: 0.51-1.08; OR=0.34, 95% CI: 0.10-1.24). We concluded that DHEA supplementation in DOR patients might improve the pregnancy outcomes. To further confirm this effect, more randomized controlled trials with large sample sizes are needed.

56. No significant differences in AFC, ovarian response to a standard low dose of gonadotrophin stimulation and number of oocytes obtained were detected in anticipated normal responders receiving 12 weeks of DHEA prior to IVF treatment relative to placebo.

57. Role of DHEA on improving the ovarian reserve markers in DOR : Findings from an Indian study A significant increase (p<0.05) in the serum AMH in all age groups (35, 36–38 and >38 years) Significant rise in peak estradiol level on the day of hCG administration (p<0.05) Significant reduction (p<0.05) in Day 2 FSH in all age groups Thus, DHEA has a significant effect in improving the ovarian reserve in poor responders with previously failed IVF cycles It can help in enhancing clinical pregnancy rate in patients with DOR AMH, Antimullerian hormone; AFC, antral follicle count; FSH, follicle-stimulating hormone; DHEA, dehydroepiandrosterone; hCG , human chorionic gonadotrophin Impact of treatment on AMH and FSH Singh N, Zangmo R, Kumar S, et al. A prospective study on role of dehydroepiandrosterone (DHEA) on improving the ovarian reserve markers in infertile patients with poor ovarian reserve. Gynecol Endocrinol. 2013 Nov;29(11):989-92.

58. Higher LBR in women with DOR with DHEA pretreatment: An Indian study DHEA is found to be effective in achieving spontaneous or IVF pregnancy in patients with DOR Chatterjee S, Chaudhuri R, Chowdhury RG, et al. Infertile Women with Diminished Ovarian Reserve have more Live Births Following Dehydroepiandrosterone Pre-Treatment. J Reprod Med Gynecol Obstet. 2019 ; 4:020. DOR, Diminished ovarian reserve; IVF, in vitro fertilization; DHEA, Dehydroepiandrosterone; LBR, Live birth rate

59. Because of the uncertainty of published data , we suggest that well-designed multicentre RCTs are required to provide more insight on the effectiveness of DHEA. The absence of significant side effects should NOT be considered as an argument to support DHEA treatment.

60. Aromatase inhibition

61. Mitwally MF, Casper RF. Aromatase inhibition improves ovarian response to follicle-stimulating hormone in poor responders. Fertil Steril . 2002 Apr;77(4):776-80. doi : 10.1016/s0015-0282(01)03280-0. PMID : 11937133 .

62. Ozmen B, Sönmezer M, Atabekoglu CS, Olmus H. Use of aromatase inhibitors in poor-responder patients receiving GnRH antagonist protocols. Reprod Biomed Online. 2009 Oct;19(4):478-85. doi : 10.1016/j.rbmo.2009.05.007. PMID : 19909587. In conclusion, adjunctive letrozole administration seems to restore an IVF cycle by decreasing the rate of cycle cancellation and seems to reduce the cost by reducing the total gonadotrophin dosage .

63. Letrozole - Inferior results regarding- days of stimulation mean gonadotropin dose number of mature follicles metaphase II oocytes retrieved serum estradiol level on the day of hCG administration percentage of top and good quality embryos implantation and clinical pregnancy rates total cancellation rate

64. Sunkara SK, Pundir J, Khalaf Y. Effect of androgen supplementation or modulation on ovarian stimulation outcome in poor responders: a meta-analysis. Reprod Biomed Online. 2011 Jun;22(6):545-55. doi : 10.1016/j.rbmo.2011.01.015. Epub 2011 Feb 17. PMID : 21493151.

65. Sunkara SK, Pundir J, Khalaf Y. Effect of androgen supplementation or modulation on ovarian stimulation outcome in poor responders: a meta-analysis. Reprod Biomed Online. 2011 Jun;22(6):545-55. doi : 10.1016/j.rbmo.2011.01.015. Epub 2011 Feb 17. PMID : 21493151.

66. Role of LH in POR

67. LH supplementation useful in older women ONLY Bosch E, Labarta E, Crespo J, Simón C, Remohí J, Pellicer A. Impact of luteinizing hormone administration on gonadotropin-releasing hormone antagonist cycles: an age-adjusted analysis. Fertil Steril. 2011 Mar 1;95(3):1031-6. doi: 10.1016/j.fertnstert.2010.10.021. Epub 2010 Nov 10. PMID: 21067717.

68. Meta-analyses on rLH in POR Significantly higher clinical pregnancy rates (odds ratio: 2.03,  P  = 0.003), implantation rates (odds ratio: 2.62,  P  = 0.004) and number of oocytes retrieved (weight mean differences: 1.98,  P  = 0.03) were observed in hypo-responders supplemented with recombinant LH versus hypo-responders who underwent FSH monotherapy . No differences in terms of mature oocytes or miscarriage rates were found between the two groups. Conforti A, Esteves SC, Di Rella F, Strina I, De Rosa P, Fiorenza A, Zullo F, De Placido G, Alviggi C. The role of recombinant LH in women with hypo-response to controlled ovarian stimulation: a systematic review and meta-analysis. Reprod Biol Endocrinol . 2019 Feb 6;17(1):18. doi : 10.1186/s12958-019-0460-4. Erratum in: Reprod Biol Endocrinol . 2019 Mar 14;17(1):31. PMID: 30728019; PMCID: PMC6366097. rLH supplementation did NOT increase the ongoing pregnancy rate in poor responders (OR 1.30, 95% CI: 0.80, 2.11). Furthermore, there was no significant difference in the number of oocytes retrieved, total dose of rFSH used, total duration of stimulation, number of retrieved metaphase II oocytes and cycle cancellation rate between the study and control groups. Fan W, Li S, Chen Q, Huang Z, Ma Q, Wang Y. Recombinant Luteinizing Hormone supplementation in poor responders undergoing IVF: a systematic review and meta-analysis. Gynecol Endocrinol . 2013 Apr;29(4):278-84. doi : 10.3109/09513590.2012.743016. Epub 2013 Jan 24. PMID: 23347045.

69. hMG vs rLH in POR Number of follicles in ovaries, total number of oocytes or M 2   oocytes and quality of fetuses has no significant differences between two groups (p>0.05). Total number of fetuses were significantly higher in patients who received rFSH + HMG (p=0.02). Fertility outcomes consisted of: live birth rate, chemical pregnancy and clinical pregnancy rate were higher in rFSH + HMG group in comparison to rFSH +r-LH group (p<0.05). Shahrokh Tehraninejad E, Farshbaf Taghinejad M, Hossein Rashidi B, Haghollahi F. Controlled ovarian stimulation with r-FSH plus r-LH vs. HMG plus r-FSH in patients candidate for IVF/ICSI cycles: An RCT. Int J Reprod Biomed. 2017 Jul;15(7):435-440. PMID: 29177245; PMCID: PMC5601935.

70. ESHRE, 2019 The use of recombinant LH ( rLH ) + recombinant FSH ( rFSH ) for ovarian stimulation is probably not recommended over hMG in GnRH agonist protocols with regards to safety.

71. Other adjuvants

72. Coenzyme Q-10 Women in CoQ10 group demonstrated an increased number of retrieved oocytes (4, IQR 2–5), higher fertilization rate and more high-quality embryos (1, IQR 0–2); p < 0.05 Significantly fewer women treated with CoQ10 had canceled embryo transfer due to poor embryo development than controls Significantly more women from the treatment group had available cryopreserved embryos The clinical pregnancy and live birth rates per embryo transfer and per one complete stimulation cycle tended to be higher in CoQ10 group but did not achieve statistical significance Xu Y, Nisenblat V, Lu C, et al. Pretreatment with coenzyme Q10 improves ovarian response and embryo quality in low-prognosis young women with decreased ovarian reserve: a randomized controlled trial. Reprod Biol Endocrinol . 2018 Mar 27;16(1):29.

73. Coenzyme Q10 concentration and its association with embryo morphokinetics and pregnancy rate in assisted reproductive techniques Follicular fluid CoQ10 levels were significantly higher in grades A and B than in grades C and D embryos (p < 0.05) The concentration of CoQ10 levels was significantly higher in pregnant women (p < 0.05) CoQ10 levels were significantly higher in the follicular fluid of embryos that had optimal cc2 (p < 0.05) as shown in the Table High follicular fluid CoQ10 level is associated with optimal embryo morphokinetic parameters and higher pregnancy rates Akarsu S, Gode F, Isik AZ, et al. The association between coenzyme Q10 concentrations in follicular fluid with embryo morphokinetics and pregnancy rate in assisted reproductive techniques. J Assist Reprod Genet. 2017 May;34(5):599-605.

74. Growth Hormone Cozzolino M, Cecchino GN, Troiano G, Romanelli C. Growth hormone cotreatment for poor responders undergoing in vitro fertilization cycles: a systematic review and meta-analysis. Fertil Steril . 2020 Jul;114(1):97-109. doi : 10.1016/j.fertnstert.2020.03.007. Epub 2020 Jun 16. PMID: 32553470. Needs further studies

75. International guidelines on Adjuvants

76. The available evidence does not support the routine use of DHEA as an adjuvant in IVF cycles. The use of DHEA in this setting cannot therefore be supported Do not use growth hormone or dehydroepiandrosterone (DHEA) as adjuvant treatment in IVF protocols

77. Consider DHEA/ Testosterone GH

78. No Good quality evidence

79. Laboratory options in POR ICSI for all ? PGT for all? Ideal day of transfer-D3/D5? Fresh vs frozen ET? Oocyte pooling and embryo banking? There is insufficient evidence to support

80. In vitro Activation (IVA) of oocytes - Science Fiction?

81. Mitochondrial Transfer

82. Intraovarian PRP Intra-ovarian autologous PRP infusion increases the ovarian reserve parameters resulting in increased mature oocyte yield, fertilization rate , as well as the formation of good-quality embryos . There is a great need for future, high-quality randomized controlled trials  to estimate its efficacy in terms of clinical pregnancy and live birth rate. Also, there is a need to identify an optimum level of serum AMH or another marker of ovarian reserve for the success of intra-ovarian PRP infusion and identify the subpopulation that would get the most benefit from PRP.

83. Social egg freezing- race against time? Often perceived (and promoted) as a form of insurance Success rates will be limited in women who are already in their mid–late 30s The significant costs associated with the procedure and subsequent egg storage Low fecundity rate

84. Donor oocyte Repeated IVF failure High FSH or low AMH ( how high is high and how low is low) Counselling ART Law, 2022

85. Conclusion

86. Conclusion

87. Conclusion

88. Conclusion

89. Conclusion

90. Conclusion

91. Thank you