IVF IVF 2.0: 2.0: Ena Enabling ling compr comprehensiv ehensive genetic enetic testing in in Repr productiv ductive Medicine Medicine
The Endome ometri trial al Facto ctor: : EM EMMA, MA, ALICE ICE and ER ERA Carlos Simon MD. PhD. Professor Ob/Gyn, University of Valencia. Spain Adjunct Clinical Professor Ob/Gyn. Baylor College,USA Head of the Scientific Advisory Board, Igenomix
Carlos Simon Disclosure Founder & Head of Scientific Advisory Board, Igenomix, S.L. Editorial Editor: External Scientific Advisor:
ART remains inefficient Globally, live birth rates in ART range between 25% and 30% per initiated cycle The most fundamental reason why IVF treatments are not successful is because transfer of good quality embryos or even euploid embryos into the endometrial cavity does not lead to pregnancy. Malizia B et al., N Engl J Med. 2009 Adamson et al., Fertil Steril. 2011
The contribution of the embryonic and endometrial factor Embryo Aneuploidies Drosophila 0.01 % Mouse 0.01 % Human 20 – 100 % Implantation Rate (IR) Natural cycle 35 % Euploid embryos 60 % (Rubio et al. RCT Fertil Steril 2017 ) Rodents 95 % Rabbits 96% The main difference between humans and rodents lies in The endometrial/decidual control in human implantation versus The embryo control in rodent implantation (embryonic diapause)
Personalized Medicine “Variability is the law of life , as no two faces are the same, so no two bodies are alike, and no two individuals react alike, and behave alike under the abnormal conditions which we know as disease. ” Sir William Osler (1849-1919) H. Sapiens 150,000 y
Days 8 - 9 Days 4 - 5 Days 4 - 5 Days 4 - 5 Days 6 - 7
The Maternal Contribution Learning objetives Materno-Embryonic Crosstalk Endometrial Microbiota Endometrial Receptivity Moreno et, al. AJOG 2016 Vilella et, al. Development 2015 Diaz-Gimeno et, al. F&S 2011 Moreno et, al. AJOG 2018 Balaguer et, al. MHR 2018 Ruiz-Alonso et, al. F&S 2013 Decidualization Decidualization Garrido-Gomez et. al, HR 2014 Von Grothusen et. al, HR 2018 Garrido-Gomez et. al, JCEM 2011 Wang W. et al. Cell. 2018 in press Garrido-Gomez et. al, Development 2017 Garrido-Gomez et. Al, PNAS 2017
Endometrial microbiome Materno-embryonic crosstalk Endometrial microbiota Endometrial receptivity Moreno I. et al., AJOG 2016 Moreno I. et al., AJOG 2018 Decidualization Residents, Tourists, or Invaders?
Molecular assessment of endometrial microbiota by NGS gDNA PURIFICATION ENDOMETRIAL/VAGINAL ASPIRATION 16S rRNA gene BARCODED BACTERIAL 16S rRNA PCR DATA ANALYSIS & TAXONOMICAL ASSIGNMENT SEQUENCING
Average bacterial communities in endometrial and vaginal microbiota of fertile subjects In fertile and asymptomatic women: The endometrial microbiota is similar to that on the vagina. It is mainly composed of Lactobacillus species . Significant differences are found between endometrial and vaginal microbiota in 20% of these women. Some bacterial genera are only found in endometrial samples. Endometrium Vagina Moreno et al., Am J Obstet Gynecol 2016
Low abundance of Lactobacillus in endometrium is associated with poor reproductive IVF outcomes BMI: body mass index; LDM: Lactobacillus -dominated microbiota; NLDM: non- Lactobacillus -dominated microbiota; *Chi Square (χ² test) and Student’s t -test were performed; *p-value<0.05; § : Voluntary termination of pregnancy. Moreno et al., Am J Obstet Gynecol 2016
Prospective Multicenter Observational MICrobiome study Register: 03330444 First release: Nov 6, 2017 Last Update: Mar 25, 2019 Status: Active. Not recruiting First IRB/EC approval May 29, 2017 Last IRB/EC approval March 27, 2018 FPFI August 4, 2017 LPLI February 26, 2019 Recruitment length 1.5 years (finished) Study length 2.5 years
MIC study: 452 patients were enrolled at 13 participant sites from 8 countries worldwide
OBJECTIVE: We sought to develop a molecular diagnostic method for CE equivalent to the 3 classic methods together, overcoming the bias of using any of them alone. 16S METAGENOMICS RT-PCR 16S bacterial library Specific amplification of CE pathogens Enterobacteria Escherichia coli Enterococcus faecalis Staphylococcus spp. Streptococcus spp. Gardnerella vaginalis Neisseria gonorrhoeae Chlamydia trachomatis Ureaplasma urealyticum Mycoplasma hominis MICROBIOME PROFILE
Molecular microbiological diagnosis of chronic endometritis ASSESSED FOR ELIGIBILITY (n=113) Not DNA for RT-PCR diagnosis (n=18) MOLECULAR DIAGNOSIS OF CE (n=95) Excluded for uncomplete clinical diagnosis: Non informative (n=9) Only 1 classical diagnosis method (n=3) Only 2 classical diagnosis methods (n=18) CE DIAGNOSED BY 3 CLASSICAL METHODS (n=65) Concordant CE diagnosis by Discordant CE diagnosis by the 3 classical methods the 3 classical methods (n=13) (n=52) Discordant Discordant Discordant histology+hysteroscopy histology+microbial hysteroscopy+microbial with microbial culture culture with hysteroscopy culture with histology (n=14) (n=17) (n=21) Moreno et al., AJOG 2018
Chronic endometritis Comparison of 3 classical methods: Histology vs Hysteroscopy vs Microbial culture (n=65) POS NEG HIS 25 40 HYS 63 2 CUL 34 31 Total: 122 73 Accuracy (13/65): 20.00% Concordant: 12 1 Molecular microbiology vs Concordant classical methods (n=13) Negative POS NEG Positive Concordant 12 1 HIS+HYS+CUL Molecular 9 4 (PCR) Total: 21 5 Concordant: 9 1 Accuracy (10/13): 76.92%
New diagnostic posibilities HUMAN RNA ERA ENDOMETRIAL TISSUE 16S BACTERIAL DNA RT-PCR METAGENOMICS Specific amplification of 16S bacterial library CE pathogens Enterobacteria MICROBIOME PROFILE Escherichia coli Enterococcus faecalis Staphylococcus spp. Streptococcus spp. Gardnerella vaginalis Neisseria gonorrhoeae Chlamydia trachomatis Ureaplasma urealyticum Mycoplasma hominis
Take home message: ENDOMETRIAL MICROBIOTA The uterine cavity is not a sterile site. Endometrial microbiota is not a carry-over from the vagina. The percentage of Lactobacillus together with specific pathogens are significant variables to predict reproductive success. The molecular diagnosis of pathogens causing CE is equivalent to using histology, hysteroscopy and microbial culture together
The Maternal Contribution Endometrial receptivity Materno-embryonic crosstalk Endometrial microbiota Endometrial receptivity Decidualization Plasma membrane transformation
Molecular medicine: human endometrial transcriptome Riesewijk et al., 2003 (HG-U133 2.0) WOI Ponnapalam et al., 2004 (Home-made array) menstrual cycle Talbi et al., 2005 (HG-U133 2.0) menstrual cycle WOI Díaz-Gimeno et al., Fertil Steril 2011
ERA classifies the molecular receptivity status of the endometrium Pre-Receptive Receptive Post-Receptive
The symphony of sync ncroniz onizati tion on P P +1 P +2 P +3 P +4 P +5 P +6 P +7 P +8 P +9 Epithelial PR Progesterone
Personalized embryo transfer (pET) as a treatment for RIF of endometrial origen pET ET P+3 P+5 P+7 LH+5 LH+7 LH+9
2014 Clinical Management of Endometrial Receptivity Semin Reprod Med.32(5):410-4 Timing the window of implantation by nucleolar channel system prevalence matches the accuracy of the ERA Publi lica catio tions 2014 Fertility and Sterility. 102(5):1477-81 endometrial receptivity array Cold Spring Harb Perspect Med. 2015 Human Endometrial Transcriptomics: Implications for Embryonic Implantation 5(7):a022996 YEAR TITLE JOURNAL Current Opinion in Obstetrics & 2015 Understanding and improving endometrial receptivity Gynecology. 27(3):187-92 ReproductiveBioMedicine Online. 2015 Is endometrial receptivity transcriptomics affected in women with endometriosis? A pilot study 31(5):647-54 2016 Diagnosis of endometrial-factor infertility: current approaches and new avenues for research Geburtshilfe Frauenheilkd.76(6): 699-703 Does an increased body mass index affect endometrial gene expression patterns in infertile patients? A functional 2017 Fertility and Sterility.107(3):740-748.e2 genomics analysis 2017 Endometrial function: facts, urban legends, and an eye to the future Fertility and Sterility. 108(1):4-8 Implantation failure of endometrial origin: it is not pathology, but our failure to synchronize the developing embryo 2017 Fertility and Sterility. 108(1):15-18 with a receptive endometrium 2017 Meta-signature of human endometrial receptivity: a meta-analysis and validation study of transcriptomic biomarkers Scientific Reports.7(1):10077 Window of implantation transcriptomic stratification reveals different endometrial subsignatures associated with live 2017 Fertility and Sterility. 108(4):703-710.e3 birth and biochemical pregnancy Current Opinion in Obstetrics and 2018 Implantation failure of endometrial origin: what is new? Gynecology. PMID: 29889670 Journal of Assisted Reproduction and 2018 Inter-cycle consistency versus test compliance in endometrial receptivity analysis test Genetics. 2018 May 26. PMID: 29804174.
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