ERA. Endometrial Receptivity Analysis. Patented since
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1 Endometrial Receptivity Analysis Patented since ERA
2 ERA analyzes by NGS the expression of 236 genes in order to determine the personalized window of implantation for each patient. 01
3 ERA Test The ERA test consists of: NGS analysis of the expression of 236 genes involved in endometrial receptivity A molecular tool based on gene expression used to determine when the endometrium is receptive. An informatic predictor that analyzes the gene expression data and classifies the endometrium as Receptive" or "Non Receptive with a sensitivity of and a specificity of Patented in 2009: PCT/ES 2009/
4 PIONEERS IN REPRODUCTIVE GENETICS Scientific Support: IGENOMIX Publications The development of this tool was published in the paper by Díaz-Gimeno et al, 2011 (Fertil Steril Jan;95(1):50-60, 60.e1-15). Furthermore, its effectiveness and consistency was demonstrated in the paper by Díaz-Gimeno et al, 2013 (Fertil Steril Feb;99(2):508-17). Finally, its clinical applicability to patients with implantation failure was confirmed in publications by Ruiz-Alonso et al, 2013 (Fertil Steril Sep;100(3):818-24) and Ruiz-Alonso et al, 2014 (Hum Reprod Jun;29(6):1244-7). A randomised study is currently in progress to assess its applicability to patients without any prior assisted reproduction treatment. (ClinicalTrials.gov Identifier:NCT ). 03
5 PIONEERS IN REPRODUCTIVE GENETICS Scientific Support Receptive Pre-receptive Hierarchical clustering LH+3 LH+1 LH+5 LH+7 LH+9 IGENOMIX Publication: Díaz-Gimeno P. et al. - Fertil Steril 2011; 95(1):
6 Diagnostic Value Endometrial Evaluation: Diagnose the state of the endometrium during the Window of Implantation and determine the time frame for embryo transfer. In a blinded study, ERA classifies endometrial receptivity better than Noyes criteria: Kappa value Pathologist 1 (P1) Pathologist 2 (P2) P1 vs P2 ERA ( ) ( ) ( ) ( ) Kappa value: Good Concordance Very Good Concordance IGENOMIX Publication: Díaz-Gimeno P. et al. - Fertil Steril 2013; 99 (2):
7 PIONEERS IN REPRODUCTIVE GENETICS For whom? Recurrent implantation failure patients: - 2 or more implantation failures with own embryos or 1 with ovum donation. - Good quality embryos. Patients with morphologically normal endometrium: - ERA after intervention in the case of an congenital uterine anormalities. Patients with normal, atrophic or hipertrophic endometrium - In case of atrophic or hipertrophic, it has to be consistent for all the cycles in the patient. 06
8 PIONEERS IN REPRODUCTIVE GENETICS Biopsy Hormone replacement therapy Patient will start with estradiol from the 1st or 2nd day of the menstrual cycle. Ultrasound assesment will be performed between 7 to 10 days after start the estradiol administration. When a trilamminar endometrium >6mm is reached with a serum progesterone <1ng/ml, the progesterone intake can start. Progesterone will be administered for five full days (120 hours). Natural cycle hcg either recombinant or urinary will be administered according routine parameters in a natural cycle (follicle size > 17 mm). The biopsy will be taken 7 days (168 hours) after the hcg triggering. Controlled Ovarian Stimulation Biopsy can be taken on the next natural or HRT cycle. E2Days P4Days P+0 P+1 P+2 P+3 P+4 P+5 Ultrasound: 6mm; Triple layer P4 <1ng/ml Biopsy 07
9 Biopsy and Storage Biopsy Prepare and Label the cryotube either with the Patient s initials, DOB and date of biopsy or with the Patient s initials and MRN. Immediately after the biopsy is introduced into the cryotube, shake it vigorously for 10 seconds. The amount of tissue should reach the white line on the cryotube in order to preserve the genetic material. Please do not exceed this amount. Storage Immediately place the biopsy sample in a fridge (4-8 C/39-46ºF) for at least 4 hours. This preserved sample, in the original cryotube, can then be shipped at room temperature. Alternatively, the samples can be kept in the fridge for 3 weeks or frozen at -20 C/-4ºF (recommended) until the moment of shipment. The sample can then be shipped at room temperature (<35 C/95ºF). For shipment during the summer, it is recommended to add an ice pack. 08
10 Shipment Samples are sent at room temperature accompanied with the patient's informed consent and a completed requisition form. Samples shipped at room temperature should reach us in a maximum of 4 to 5 days. For more instructions regarding sample packaging, please contact us via Confirm shipping by accessing the following link: 09
11 Results Interpretation of results After the analysis, there are two possible results: Receptive (R): The gene expression profile is similar to that of a normal receptive endometrium. You are recommended to proceed with the embryo transfer in the same conditions, type of cycle and day in which the endometrial biopsy has been performed. Non Receptive (NR): The gene expression profile is similar to an endometrium out of the receptive phase. We recommend taking a new endometrial biopsy to validate a displacement of the window of implantation and to guide the personalized embryo transfer (pet). 10
12 Results Displacement of the WOI Personalization of the embryo transfer Embryo Embryo Day 3 Day 5 Keep developing Implantation Endometrium Pre-Receptive Receptive Post-Receptive HRT CYCLE P+0 P+5 Patient Receptive at P+4 P+0 P+1 P+2 P+3 P+4 P+5 P+6 P+7 P+... Stardard WOI (ERA P+5: R) Delayed WOI (ERA P+5: PRE) Advanced WOI (ERA P+5: POST) Patient Receptive at P+7 Patient Receptive at P+5 Start Progesterone Start Progesterone P+0 P+1 P+2 P+3 P+4 P+5 P+6 P+7 P+8 P+... P+0 P+1 P+2 P+3 P+4 P+5 P+6 P+7 P+8 P+... Pre-receptive Start Progesterone Endometrium s state Receptive Post-receptive 11
13 PIONEERS IN REPRODUCTIVE GENETICS Results ERA decision tree P+5 R: Receptive NR: Non receptive pet: Personalized embryo transfer ERA Test P+4 P+7 Diagnosis R Non-analyzed samples 2nd ERA Test NR Diagnosis pet R NR Treatment evaluation Gene profile analysis Invalid RNA (3%), potential causes: - High temperatures (>35º) - Sample size too large - Too much blood or mucus Insufficient RNA (2.5%), potential causes: - Sample size too small - Too much blood or mucus NO CHARGE Non informative (1%) 12
14 Clinical outcome Updated results 01/01/2017
15 Results 20,000 patients 71.4% Receptive 28.6% Non Receptive 2.4% Proliferative 85% Pre-receptive 12.6% Post-receptive 54 countries More than 600 clinics 14
16 Clinical Outcome in RIF Patients and Controls No. of patients Age (years) Nº OF RECEPTIVE ERA /TOTAL ANALYZED (%) No. of previous failed cycles Total Patients with pet after RECEPTIVE ERA Implantation rate after 1 ST pet (%) Pregnancy rate after 1 ST pet (%) Biochemical pregnancies (%) Clinical abortions (%) Ovum donation patients/total (%) Patients with pet after RECEPTIVE ERA Implantation rate after 1 ST pet (%) Pregnancy rate after 1 ST pet (%) Biochemical pregnancies (%) Clinical abortions (%) IVF/ICSI patients/total (%) Patients with pet after RECEPTIVE ERA Implantation rate after 1 ST pet (%) Pregnancy rate after 1 ST pet (%) Biochemical pregnancies (%) Clinical abortions (%) RIF ± /85 (74.1) 4.8 ± /56 (33.9) 15/29 (51.7) 3/15 (20.0) 0/15 (0.0) 22/63 (34.9) 16 12/33 (36.4) 9/16 (56.2) 2/9 (22.2) 0/9 (0.0) 41/63 (65.1) 13 7/23 (30.4) 6/13 (46.2) 1/6 (16.7) 0/6 (0.0) Control ± /25 (88.0) 0.5 ± /20 (55.0) 9/11 (81.8) 2/9 (22.2) 0/9 (0.0) 13/22 (59.1) 8 7/15 (46.7) 6/8 (75.0) 1/6 (16.7) 0/6 (0.0) 9/22 (40.9) 3 4/5 (80.0) 3/3 (100.0) 1/3 (33.3) 0/3 (0.0) Nº OF NON RECEPTIVE ERA /TOTAL ANALYZED (%) No. of previous failed cycles Ovum donation patients/total (%) IVF/ICSI patients/total (%) 22/85 (25.9) 5.0 ± /22 (50.0) 11/22 (50.0) 3/25 (12.0) 0.3 ± 0.6 2/3 (66.6) 1/3 (33.3) *pet Personalized Embryo Transfer IGENOMIX Publication: Ruiz-Alonso M. et al. - Fertil Steril 2013; 100(3):
17 Clinical Outcome in ERA Receptive Patients pet outcome after receptive ERA in patients with RIF (n=310) PR IR First Embryo Transfer outcome after a RECEPTIVE ERA Test Rate (%) Months after ERA Test Number of patients Implantation rate (%) Pregnancy rate (%) ,9 43,1 44,6 45,8 58,3 48,6 54,9 59,8 55,3 53,3 73,3 70,0 16
18 PIONEERS IN REPRODUCTIVE GENETICS Clinical Outcome in Non Receptive ERA Patients Dating the WOI of the Non Receptive samples The predictor and clustering analysis indicate that Non Receptive samples classify as Pre-Receptive or Post-Receptive and therefore personalization of the WOI with a second ERA is recommended. 17
19 Clinical Outcome in Non Receptive ERA Patients Example of case 6272 Case 6272 P+5 Case 6272 P+7 Proliferative Pre-Receptive Receptive Post-Recep 18
20 Clinical Outcome in ERA Non Receptive Patients : RIF and controls No. of patients No. of previous failed cycles RIF Patients No. of previous failed cycles Control Patients ERA Prediction Pre-receptive Post-receptive 2 nd ERA at the specified day (P+4;P+6;P+7;LH+9) a Months between 1 st and 2 nd ERA 2 nd ERA Receptive at the specified day Patients with pet b after 2 nd RECEPTIVE ERA Months between 2 nd RECEPTIVE ERA and pet Implantation rate using pet Pregnancy rate using pet Biochemical pregnancies (%) Clinical abortions (%) Non Receptive ± ±0.6 21/25 (84.0) 4/25 (16.0) ± ±0.7 5/13 (38.5) 4/8 (50.0) 0/4 (0.0) 0/4 (0.0) IGENOMIX Publication: Ruiz-Alonso M. et al. - Fertil Steril 2013; 100(3): a The specified day for the 2 nd ERA was calculated by ERA Test according to the gene profile of each sample. b pet implies to do the embryo transfer under the same conditions in which a Receptive ERA was obtained. In those patients was under the same conditions in which the 2 nd ERA was performed. 19
21 Clinical Outcome Published Case Report regarding clinical applicability of the ERA test in a RIF patient Previous ART treatments ART treatments in our center Routine work negative 1. IVF with fresh day-3 ET 2. IVF with fresh day-3 ET 3. IVF with fresh day-5 ET 4. IVF with frozen day-5 ET in a natural cycle 5. OD with day-3 ET in an HRT cycle (P+2) 6. OD with day-3 ET in a natural cycle 7. OD with day-5 ET in an HRT cycle (P+5) INTERVENTION Finding Personalized WOI (ERA Test) ET: Embryo Transfer HRT: Hormonal Replacement Teraphy IVF: In-Vitro Fertilization OD: Ovum Donation WOI: Window of Implantation IGENOMIX Publication: Ruiz-Alonso, et al. Hum Reprod Jun;29(6): OD with day-5 ET in an HRT cycle (P+7) Succesful twin pregnancy 20
22 Clinical Outcome CLINICALOUTCOME Data from a pilot study comparing Frozen Embryo Transfer (FET) to personalized Embryo Transfer (pet) in the same patient cohort. Patients underwent an FET before performing their first ERA test, on a day that was later diagnosed as non-receptive by the ERA. Patients then received a pet on their receptive day, after confirmation from a 2nd ERA biopsy. First attempt Number of patients Source of oocytes Age Total attempts Number of embryos transferred Implantation rate Pregnancy rate On going pregnancy rate Clinical abortion ET pet 17 Ovum donation 40.7 ± ± ± ± ± % 40.0% 19.4% 60.0% 0% 75.0% 71.4% 0% IGENOMIX Publication: Ruiz-Alonso, et al. Hum Reprod Jun;29(6): Biochemical pregnancy 28.6% 25.0% 21
23 Clinical Outcome Data comparing clinical outcome from patients with ET before performing their first ERA test, on a day that was later diagnosed as non-receptive by the ERA, to clinical outcome from patients performing pet on their receptive day according to ERA result. Clinical Outcome IR First Attempt IR Total Attempts PR First Attemp PR Total Attempts OPR First Attempt OPR Total Attempts NR (100) 12% (21/175) 10% (34/332) 20% (20/100) 18% (33/186) 0% (0/20) 0% (0/33) R (400) 45% (312/692) 42% (367/866) 58% (233/400) 56% (278/496) 76% (178/233) 76% (211/278) IR: Implantation rate PR: Pregnancy rate OPR: Ongoing pregnancy rate 22
24 Receptivity and obesity BMI Category Normal ( ) (n=163) Overweight ( ) (n=47) Obese ( 30) (n=11) Age (mean) BMI (mean) % non receptive patients by ERA 25.2% 25.5% 36.4% Lathi R B et al, PCRS. 23
25 Receptivity and endometrial thickness Endometrial thickness (mm) < >12 Receptive 6/4 (42.85%)* 333/431 (77.26%)* 24/37 (64.86%) Non receptive 8/14 (57.14%)* 98/431 (22.73%)* 13/37 (35.13%) P: by Chi-square test IGENOMIX Publication: Valbuena D et al, 2016 ESHRE. 24
26 PIONEERS IN REPRODUCTIVE GENETICS ERA Randomized Controlled Study Prospective Randomized Controlled Study presented at ASRM shows the improvement on Pregnancy Rates and Ongoing Pregnancy Rates. Fresh ET Award Winner Society for Reproductive Endocrinology and Infertility (SREI) Our paper PROSPECTIVE, RANDOMIZED STUDY OF THE ENDOMETRIAL RECEPTIVITY ANALYSIS (ERA ) TEST IN THE INFERTILITY WORKUP TO GUIDE PERSONALIZED EMBRYO TRANSFER VERSUS FRESH TRANSFER OR DEFERRED EMBRYO TRANSFER* has been awarded as Prize Paper by the Society for Reproductive Endocrinology and Infertility, in this year's ASRM Annual Meeting Deferred ET Personalized ET Patients (n) Transfers (n) Preg. Rate/ET (%) 61.7% 60.8% 85.7 %* Implantation Rate (%) 35.3% 41.4% 47.8% Ongoing PR/ET (%) 43.3% 44.6% 55.1% * p=0.003 by Chi-Square test ERA Test Improves Clinical Outcomes 90,0% 80,0% 70,0% 60,0% 50,0% 40,0% 30,0% 20,0% 10,0% 0,0% Implantation Rate Fresh ET Preg. Rate/ET Deferred ET Personalized ET Ongoing PR/ET p=0.003 by Chi-Square test 25
27
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