Novel Biomarker Catalyst Lab In Vitro Fertilization - IVF sLHCGR - - PowerPoint PPT Presentation

Novel Biomarker Catalyst Lab

In Vitro Fertilization - IVF sLHCGR & LH-sLHCGR ELISA

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In Vitro Fertilization

 IVF is a treatment that is offered to couples with unexplained fertility, endometriosis or the female has blocked or an absence of fallopian tubes  All treatment centers use different drug protocols, the following example is a standard long cycle  IVF Stage 1 – Stop normal activity of ovaries  IVF Stage 2 – Stimulate ovaries / ovarian induction  IVF Stage 3 – Follicle count  IVF Stage 4 – Egg collection  IVF Stage 5 – Embryo transfer to the uterus

*Medical condition in which cells from the lining of the uterus appear and flourish outside the uterine cavity ** Two tubes leading from the ovaries into the uterus

 Diagnostics - Before starting the procedure  Determining factors for egg quality are  Day-3 FSH (Follicle Stimulating Hormone), high FSH (LH:FSH ratio) predicts poor egg quality  Ovarian reserve by AMH (Anti-Mullerian Hormone) and by antral follicular count (AFC). Usually, low AMH means poor oocyte reserve & high AMH indicates polycystic ovary (risk of ovarian hyperstimulation)  AMH is also useful for determining the correct dose of fertility drugs  Drugs  Nasal spray or subcutaneous injection of GnRH (Gonandotrophin releasing Hormone) for approx. two weeks  GnRH temporarily stops the normal activity of the ovary so that ovulation does not occur when the ovaries are stimulated  Diagnostics – After drugs  Ultra sound scan to ensure that the ovaries are inactive

IVF Stage 1 - Stop normal activity

• vary

 Drugs  A second drug will be introduced, FSH (Follicle Stimulating Hormone) by subcutaneous injection, on a daily basis  FSH stimulates the ovaries to produce multiple follicles  Diagnostics  The progress will be assessed by ultra sound scans 3-4 times during this stage  E2 (Estradiol) and FSH are measured 3-4 times during this period  Direct correlation between the E2 value and the number of eggs/follicles  The physician uses the E2 and ultrasound result to determine if the treatment is going well  If a patient is hyper-stimulated and develops too many eggs, the cycle is usually cancelled

IVF Stage 2 – Stimulate ovaries / Ovarian induction

 The follicles will be counted and measured by echography. A size of approx.18mm indicates mature egg may be present (not all follicles will contain eggs)  Depending on the number and size of the follicles, egg collection will be scheduled

IVF Stage 3 – Follicle Count

 Drugs  If the ovaries have responded well, injection with a third drug, hCG will be administered 

• Approx. 36 hours prior to the

scheduled egg collection  hCG helps to mature the eggs present and release the eggs in the follicles for the egg collection  Diagnostics  On the day of the hCG trigger, the E2/follicle ratio should be approx. 100- 200 and the Progesterone level should be 0.8 ng/mL

 The egg collection (oocyte retrieval) performed under general anesthetic or sedation  Vaginal probe with a needle attached to it and is passed through the vaginal wall into each

• vary under ultra sound guidance

 The follicles are individually drained and embryologist checks the follicular fluid for eggs.  Once the eggs have been retrieved they will be left to rest in an incubator  Later that day they will then be mixed with a high concentration of prepared sperm. They will then be checked the next day for fertilization  The embryos are checked on day 2/3 of

• development. If they are progressing as

expected then it may be recommended to aim for a blastocyst transfer on day 5

IVF Stage 4 – Egg collection

 Discussion of the number and quality of embryos to be replaced will take place  A fine catheter containing the embryo/ embryos is passed through the cervix and deposited in the uterus. This is performed under ultra sound guidance

IVF Stage 5 – Embryo replacement

 Drugs  Progesterone is administered to maintain the thickness of the lining of the uterus to aid implantation  Taken in the form of intra-muscular injection or suppository prior to the day of the embryo transfer and continued until the pregnancy test  Diagnostics  If successful, the hCG test will be positive on the 14th day after egg retrieval  The hCG doubles every 48 hours and is monitored throughout the pregnancy

 OHSS – Ovarian Hyper Stimulation  Complication from fertility medication, in particular hCG used as a trigger in Stage 3  Presence of multiple luteinized cysts within the ovaries  Classification  Mild – Ovaries are enlarged (5-12 cm), accumulation of ascites, abdominal pain, nausea and diarrhea  Severe – Hemoconcentration, thrombosis and distention, oliguria, pleural effusion and respiratory distress.  Complications of OHSS  Ovarian torsion, ovarian rupture, thrombophlebitis and renal insufficiency  About 5% of the treated patients may encounter moderate to severe OHSS

Complications during IVF

 PCOS – Polycystic Ovarian Syndrome  Multiple cysts in the ovary. These “cysts” are actually immature follicles, not cysts  One of the most common female endocrine disorders and produces symptoms in 5-10% of women of reproductive age (12-45 years)  One of the leading causes of female infertility  Majority of patients with PCOS have insulin resistance and/or are obese  30% of PCOS women can not be diagnosed with ultrasound

 Medical conditions such as diabetes and thyroid disorders

Causes of Infertility

Novel Biomarker LH-sLHCGR

hCG LH sLHCGR sLHCGR sLHCGR LH hCG

Two hormones One receptor LH & hCG have the same receptor: LHCGR

 Women who produced < 7 oocytes or > 15 oocytes had low concentrations of sLHCGR and a good IVF outcome (i.e. pregnancy)  A high level of sLHCGR was indicative of a poor IVF outcome (i.e. not pregnant)  An intermediate number of oocytes (8-14) the levels of sLHCGR did not appear to affect pregnancy  LH-LHCGR is usually undetectable in women with recurrent miscarriage prior to (pre-treatment) and following implantation  Pre-treatment serum sLHCGR/LH-sLHCGR and LH levels could provide an indication

• f functional LH levels that would allow the adjustment of hormone dose prior to
• varian induction (Stage 2)

 This could be an important step towards avoidance of OHSS, particularly for patients whose AMH levels do not correlate with high oocyte yield and potential OHSS  New tests could be useful in avoiding OHSS and may help circumvent a situation where all embryos need to be frozen  If used before uterine transfer of the embryo, these assays may also identify those women who may benefit from short-term supplementation with hCG in order to firmly establish the pregnancy

Potential sLHCGR/LH-LHCGR ELISA

 Europe leads the world in ART (Assisted Reproductive Technology), initiating approx. 71% of all reported ART cycles (not including Asia)  In 2009 there were reported 537.000 treatment cycles from 33 European countries and compares with 142.000 cycles from the US and 57.000 from Australia and New Zealand  France - 75.000 cycles  Germany – 68.000 cycles  Spain – 54.000 cycles  UK – 54.000 cycles  Italy – 52.000 cycles  Sweden – 17.000 cycles  Denmark – 14.000 cycles  Most active countries in the world are the USA and Japan  Annual growth of 5-10% over the last few years in the developed countries  Worldwide approx. 1,5 million ART cycles each year. 500.000 of these ART cycles are conventional IVF treatments

Market IVF

Source: European Society of Human Reproduction and Embryology

AMH versus sLHCGR/LH-sLHCGR

AMH sLHCGR/LH-sLHCGR

Ovarian reserve assessment AMH low or high -> reduced oocyte yields Ovarian reserve assessment Not an indicator of ovarian reserve Responsiveness to IVF No correlation between AMH and the embryo implantation potential Responsiveness to IVF Women who produced < 7 or > 15 oocytes had low concentrations of sLHCGR and a good IVF outcome (i.e. pregnancy). Women with undetectable pre- treatment sLHCGR tend to miscarry. PCOS AMH 3x higher in case of PCOS PCOS LH-sLHCGR in combination with LH, FSH & AMH helps to identify patients with PCOS. AMH fails to predict PCOS in a significant number of women. In this group of patients with normal AMH, low-to-undetectable pre- treatment sLHCGR can identify women susceptible OHSS. OHSS High AMH -> Risk of OHSS; in only in about half of the cases OHSS LH-sLHCGR in combination with LH/FSH ratio could be useful in avoiding OHSS Hormone drug dose Women with very high pre-treatment AMH could be sensitive to hormonal stimulation Hormone drug dose Pre-treatment serum sLHCGR/LH-sLHCGR and LH levels would allow the adjustment of hormone dose prior to

• varian induction

 Quantitative ELISAs for serum soluble LHCGR and hCG- LHCGR complex: potential diagnostics in first trimester pregnancy screening for stillbirth, Down’s syndrome, preterm delivery and preeclampsia - Reproductive Biology and Endocrinology 2012 – Anne E Chambers et al.  Circulating LH/hCG receptor (sLHCGR) may identify pre- treatment IVF patients at risk of OHSS and poor implantation - Reproductive Biology and Endocrinology 2011 – Anne E Chambers et al.  Microvesicle-mediated release of soluble LH/hCG receptor (LHCGR) from transfected cells and placenta explants - Reproductive Biology and Endocrinology 2011 – Anne E Chambers et al.

References

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Novel Biomarker Catalyst Lab

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