Ann Kim

A pilot cohort study of granulocyte colony-stimulating factor in the treatment of unresponsive thin endometrium resistant to standard therapies

STUDY QUESTION Is thin endometrium unresponsive to standard treatments expandable by intrauterine perfusion with granulocyte colony-stimulating factor (G-CSF)? SUMMARY ANSWER This cohort study is supportive of the effectiveness of G-CSF in expanding chronically unresponsive endometria. WHAT IS KNOWN ALREADY In a previous small case series, we reported the successful off-label use of G-CSF in four consecutive patients, who had previously failed to expand their endometria beyond 6.9 mm with the use of standard treatments. STUDY DESIGN, SIZE AND DURATION In a prospective observational cohort pilot study over 18 months, we described 21 consecutive infertile women with endometria <7 mm on the day of hCG administration in their first IVF cycles at our center. All previous cycles using traditional treatments with estradiol, sildenafil citrate (Viagra™) and/or beta-blockers had been unsuccessful. G-CSF (Nupogen™) was administered per intrauterine catheter by slow infusion before noon on the day of hCG administration. If the endometrium had not reached at least a 7-mm within 48h, a second infusion was given following oocyte retrieval. Primary and secondary main outcomes were an increase in endometrial thickness and clinical pregnancy, respectively. Endometrial thickness was assessed by vaginal ultrasound at the most expanded area of the endometrial stripe. PARTICIPANTS/MATERIALS, SETTINGS AND METHOD This study was uncontrolled, each patient serving as her own control in a prospective evaluation of endometrial thickness. The mean ± SD age of the cohort was 40.5 ± 6.6 years, gravidity was 1.8 ± 2.1 (range 0-7) and parity was 0.4 ± 1.1 (range 0-4); 76.2% of women had, based on age-specific FSH and anti-Müllerian hormone, an objective diagnosis of diminished ovarian reserve and had failed 2.0 ± 2.1 prior IVF cycles elsewhere. MAIN RESULTS AND THE ROLE OF CHANCE With 5.2 ± 1.9 days between G-CSF perfusions and embryo transfers, endometrial thickness increased from 6.4 ± 1.4 to 9.3 ± 2.1 mm (P < 0.001). The Δ in change was 2.9 ± 2.0 mm, and did not vary between conception and non-conception cycles. A 19.1% ongoing clinical pregnancy rate was observed, excluding one ectopic pregnancy. LIMITATIONS AND REASONS FOR CAUTION Small sample size (but a highly selected patient population) in an uncontrolled cohort study and in unselected first IVF cycles at our center. WIDER IMPLICATIONS OF THE FINDINGS This pilot study supports the utility of G-CSF in the treatment of chronically thin endometrium and suggests that such treatment will, in very adversely affected patients, result in low but very reasonable clinical pregnancy rates. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by the Foundation for Reproductive Medicine, New York, New York, USA, a not-for-profit research foundation and intramural grants from the Center for Human Reproduction (CHR)-New York. N.G. and D.H.B. are members of the board of the Foundation for Reproductive Medicine. N.G. is owner of CHR-New York, where the study was conducted. N.G. and D.H.B. have been recipients of research awards, travel grants and speaker honoraria from various pharmaceutical and medical device companies. None of these companies was, however, in any way associated with the materials and the manuscript presented here. N.G. and D.H.B. are listed as co-inventors on a number of awarded and still pending U.S. patents, none related to the materials presented here. N.G. is on the board of a medically related company, not in any way associated with the data presented here.

Hypoandrogenism in association with diminished functional ovarian reserve

STUDY QUESTION Is diminished functional ovarian reserve (DFOR) associated with low androgen levels? SUMMARY ANSWER Low androgen levels are associated with DFOR at all ages. WHAT IS KNOWN ALREADY Androgen supplementation via dehydroepiandrosterone (DHEA) has been reported to improve functional ovarian reserve (FOR); pregnancy rates in IVF cycles are associated with how well DHEA converts to testosterone (T); and androgen effects through the androgen receptor have been demonstrated in mice to beneficially affect early stages of follicle maturation. STUDY DESIGN, SIZE, DURATION In a controlled cohort study we investigated consecutive women presenting to our center with two forms of DFOR, premature ovarian aging/occult primary ovarian insufficiency (POA/OPOI) and physiologic diminished ovarian reserve (DOR). As controls for POA/OPOI patients, infertile women with normal age-specific FOR were recruited. PARTICIPANTS/MATERIALS, SETTINGS, METHODS The study involved 140 women with POA/OPOI, defined as age <38 years and abnormally low FOR by age-specific FSH and/or anti-Müllerian hormone (AMH), 166 women with DOR, defined as women age >40 years. Forty-nine control patients <38 years demonstrated normal FOR by FSH and/or AMH. In all three patient groups dehydroepiandrosterone (DHEA), DHEA-sulfate (DHEAS), total testosterone (TT) and free testosterone (FT) at the time of initial presentation to our fertility center were assessed. In a small subgroup of women early morning cortisol levels were also assessed. MAIN RESULTS AND THE ROLE OF CHANCE DHEAS marginally varied between the three groups (P = 0.04), with older women with DOR actually demonstrating higher levels than controls (P = 0.03). TT differed between the three groups more profoundly (P = 0.005), with women with POA/OPOI demonstrating significantly lower levels than controls (P = 0.009). Adjustment for body mass index, age and race in principle maintained observed differences in TT between groups, while adjustment for FMR1 (fragile X mental retardation 1) genotypes/sub-genotypes eliminated all differences. All three patient groups demonstrated low morning cortisol levels. LIMITATIONS, REASONS FOR CAUTION While results support lower androgen levels in women with DOR, and even more so in women with POA/OPOI, presented data should be viewed as preliminary, considering the known variability of androgen levels and the small number of women in whom morning cortisol levels were available. WIDER IMPLICATIONS OF THE FINDINGS Especially at young ages DFOR appears associated with significant hypoandrogenism (low T) in comparison with young control patients with normal FOR, raising the question whether this hypoandrogenism originates in adrenals or ovaries. POA/OPOI, thus, phenotypically mimics the polycystic ovary syndrome, where similar questions arise, though in regard to hyperandrogenism. STUDY FUNDING/COMPETING INTEREST(S) This research was supported by the Foundation for Reproductive Medicine, a not-for-profit medical research foundation and intramural funds from the Center for Human Reproduction. N.G. and D.H.B are members of the Board of the Foundation for Reproductive Medicine. N.G., A.W. and D.H.B. received research support, lecture fees and travel support from a variety of pharmaceutical and medical device companies, none in any way related to the issues discussed in this manuscript. N.G. and D.H.B. are listed as co-inventors on two, already granted US user patents, which claim therapeutic benefits from DHEA supplementation in women with DFOR and DOR: both authors are also listed on additional pending patents in regard to DHEA supplementation and on pending patents, claiming diagnostic and therapeutic benefits from the determination of CGG repeats on the FMR1 gene. N.G. is the owner of the Center for Human Reproduction, where this research was performed.

Differences in ovarian aging patterns between races are associated with ovarian genotypes and sub-genotypes of the FMR1 gene

BackgroundOvarian aging patterns differ between races, and appear to affect fertility treatment outcomes. What causes these differences is, however, unknown. Variations in ovarian aging patterns have recently been associated with specific ovarian genotypes and sub-genotypes of the FMR1 gene. We, therefore, attempted to determine differences in how functional ovarian reserve (FOR) changes with advancing age between races, and whether changes are associated with differences in distribution of ovarian genotypes and sub-genotypes of the FMR1 gene.MethodsWe determined in association with in vitro fertilization (IVF) FOR in 62 young Caucasian, African and Asian oocyte donors and 536 older infertility patients of all three races, based on follicle stimulating hormone (FSH), anti-Müllerian hormone (AMH) and oocyte yields, and investigated whether differences between races are associated with differences in distribution of FMR1 genotypes and sub-genotypes.ResultsChanges in distribution of mean FSH, AMH and oocyte yields between young donors and older infertility patients were significant (all P < 0.001). Donors did not demonstrate significant differences between races in AMH and FSH but demonstrated significant differences in oocyte yields [F(2,59) = 4.22, P = 0.019]: Specifically, African donors demonstrated larger oocyte yields than Caucasians (P = 0.008) and Asians (P = 0.022). In patients, AMH levels differed significantly between races [F (2,533) = 4.25, P = 0.015]. Holm-Sidak post-hoc comparisons demonstrated that Caucasians demonstrated lower AMH in comparison to Asians (P = 0.007). Percentages of FMR1 genotypes and sub-genotypes in patients varied significantly between races, with Asians demonstrating fewer het-norm/low sub-genotypes than Caucasians and Africans (P = 0.012).ConclusionFOR changes in different races at different rates, and appears to parallel ovarian FMR1 genotypes and sub-genotype distributions. Differences in ovarian aging between races may, therefore, be FMR1-associated.

The impact of androgen metabolism and FMR1 genotypes on pregnancy potential in women with dehydroepiandrosterone (DHEA) supplementation

BACKGROUND For decades androgens have been considered detrimental to follicle maturation. Animal studies now suggest that they are essential for normal folliculogenesis. Especially in women with premature ovarian aging (POA), recent IVF data in humans are supportive. The literature also suggests an association between recently reported ovarian genotypes of the FMR1 gene and ovarian aging patterns. We, therefore, attempted to determine a potential difference in androgen concentrations and androgen interactions in women with POA who do or do not become pregnant while undergoing androgen supplementation, and whether androgen concentrations and pregnancy chances are affected by FMR1 genotypes. METHODS We longitudinally assessed androgen metabolism in 91 women with POA, following pre-supplementation with micronized dehydroepiandrosterone (DHEA) prior to IVF. IVF outcomes were assessed based on androgen levels and ovarian FMR1 genotypes. RESULTS The mean age of the women was 39.8 ± 4.4 years; the clinical pregnancy rate was 25.3%. Total androgen concentrations were not associated with pregnancy; however, in women with abnormal FMR1 genotypes, but not those with the normal genotype, free testosterone significantly affected clinical pregnancy potential (β = 1.101, SE ± 0.508, P = 0.03). At the start of the IVF cycle, interactions of DHEA with total and free testosterone also significantly affected subsequent pregnancy rates (β = -0.058, SE ± 0.023, P = 0.01 and β = -0.496, SE ± 0.197, P = 0.012). CONCLUSIONS Androgen interactions significantly influence IVF pregnancy rates in women with POA, with the impact of total androgens on cycle outcomes varying according to FMR1 genotypes. These observations suggest that the effectiveness of androgen supplementation in women with POA varies based on FMR1 genotypes, and defines androgen deficiency as a subset of diminished ovarian reserve.

The impact in older women of ovarian FMR1 genotypes and sub-genotypes on ovarian reserve.

We recently associated ovarian FMR1genotypes and sub-genotypes with distinct ovarian aging patterns. How they impact older females is, however, unknown. We, therefore, investigated 217 consecutive first in vitro fertilization (IVF) cycles in women >40 assessing oocyte yields, stratified for better (anti-Müllerian hormone, AMH >1.05 ng/mL) or poorer (AMH≤1.05 ng/mL) functional reserve (FOR)). Mean age was 42.4±2.0 years, mean AMH 0.76±0.92 ng/mL and mean oocyte yield 5.3±5.4. Overall, and in women with better FOR, FMR1 did not affect oocyte yields. With poorer FOR (AMH≤1.05 ng/mL) women with het-norm/high, however, demonstrated higher oocyte yields (5.0±3.8) than those with het-norm/low sub-genotype 3.1±2.5; P = 0.03), confirmed after log conversion. Known associated with low FOR at young age, het-norm/high, thus, appears to preserve FOR into older age, and both het sub-genotypes appear to expand female reproductive lifespan into opposite directions.

BRCA1/2 Mutations Appear Embryo-Lethal Unless Rescued by Low (CGG n<26) FMR1 Sub-Genotypes: Explanation for the “BRCA Paradox”?

BRCA1/2 mutations and recently described constitutional FMR1 genotypes have, independently, been associated with prematurely diminished ovarian reserve. Whether they interrelate in distribution, and whether observed effects of BRCA1/2 and FMR1 on ovaries are independent of each other, is unknown. In a prospective comparative cohort study, we, therefore, investigated the distribution of constitutional FMR1 genotypes, normal (norm), heterozygous (het) and homozygous (hom), and of their respective sub-genotypes (high/low), in 99 BRCA1/2 mutation-positive women and 410 female controls to determine whether distribution patterns differed between study and control patients. In contrast to controls, BRCA1/2 carriers demonstrated almost complete absence of all constitutional FMR1 genotypes except for sub-genotypes with low (CGG n<26) alleles. Cross tabulation between BRCA1/2-positive patients and controls confirmed significant group membership, related to FMR1 distribution (P<0.0001). These results offer as most likely explanation the conclusion that BRCA1/2 mutations are embryo-lethal, unless rescued by low (CGG n<26) FMR1 sub-genotypes, present in approximately one quarter of all women. Women with low FMR1 sub-genotypes, therefore, should reflect increased BRCA1/2-associated cancer risks, while the remaining approximately 75 percent should face almost no such risks. If confirmed, this observation offers opportunities for more efficient and less costly BRCA1/2 cancer screening. The study also suggests that previously reported risk towards prematurely diminished ovarian reserve in association with BRCA mutations is FMR1-mediated, and offers a possible explanation for the so-called “BRCA paradox” by raising the possibility that the widely perceived BRCA1/2-associated tumor risk is actually FMR1-mediated.

Age at menarche: a predictor of diminished ovarian function?

OBJECTIVE To investigate whether age at menarche is associated with functional ovarian reserve (FOR) later in life. DESIGN Retrospective cohort study. SETTING Fertility center. PATIENT(S) Five hundred and two infertile women. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Levels of menarcheal age, antimüllerian hormone (AMH), follicle-stimulating hormone (FSH), and functional ovarian reserve. RESULT(S) The mean age of the patients was 38.9 ± 4.9 years, and their mean level of AMH was 1.4 ± 2.0 ng/mL and of FSH was 10.7 ± 6.1 mIU/mL. Their current age-specific diminished functional ovarian reserve (DFOR) was statistically significantly associated with early menarche, defined as age <13 years. Logistic regression analysis, adjusting for race, affirmed the higher likelihood of early age at menarche in infertile patients with DFOR. When women with DFOR were grouped into quartiles, early menarche (<25th percentile) was associated with statistically significantly higher DFOR risk than late menarche (>75th percentile). CONCLUSION(S) This study demonstrates a statistically significant impact of age at menarche on DFOR risk later in life among infertile women. The occurrence of menarche may relate to follicular pool size and/or speed of follicle recruitment, which in turn is predictive of occurrence of DFOR later in life.

Does hormonal contraception prior to in vitro fertilization (IVF) negatively affect oocyte yields? - A pilot study

BackgroundAs oral contraceptives (OCs) suppress anti-Müllerian hormone (AMH), and hormonal contraceptives (HCs), likely, suppress functional ovarian reserve, this study was initiated to determine whether HC affect oocyte yields.MethodsWe investigated in a retrospective cohort study 43 oocyte donors in 71 in vitro fertilization (IVF) cycles, evaluating anti-Müllerian hormone (AMH) and oocyte yields as reflections of functional ovarian reserve (OR). In 25 IVF cycles egg donors were on HC within one month prior to IVF, and in 46 cycles they were not. Donors, based on their HCs, were further subdivided into 12 with less, and 13 with more androgenic progestins.ResultsWhile the three groups did not differ in age, age at menarche, BMI and AMH, oocyte yields among donors who utilized estrane- and gonane-derived (higher androgenic) HCs were lower 11.3 (95% CI 8.3 – 14.3) than either donors using no HCs 16.6 (95% CI 14.7 -18.4) (P < 0.05) or those using anti-androgenic HCs 19.0 (95% CI 12.2-25.8) (P< 0.01). Significance was maintained after adjustments for the donor age and total FSH dose used in ovulation induction.ConclusionsEven in young oocyte donors, high androgenic OC exposure appears to suppress functional ovarian reserve and oocyte yields. Since OCs are often routinely used in preparation for IVF, such practice may require reevaluation. Especially in women with diminished ovarian reserve OCs, and especially high androgenic progestin HCs, should, likely, be avoided.

Toward a Better Understanding of Functional Ovarian Reserve: AMH (AMHo) and FSH (FSHo) Hormone Ratios per Retrieved Oocyte

CONTEXT Functional ovarian reserve (FOR) has recently been demonstrated to differ with ovarian genotypes of the FMR1 gene and is currently routinely determined with anti-Müllerian hormone (AMH) and FSH. Both, however, reflect distinctively different stages of folliculogenesis. OBJECTIVES To better understand how AMH and FSH reflect FOR, we evaluated both hormones in association with in vitro fertilization (IVF) by determining how they associate with oocyte yields, considered the most accurate available measure of FOR. DESIGN AND SETTING Using a series of logistic regressions, we assessed AMH and FSH per oocyte retrieved (AMHo and FSHo) in only first IVF cycles and determined whether at different ages and/or based on ovarian FMR1 genotypes and subgenotypes AMHo and/or FSHo are associated with clinical pregnancy chances in IVF. PATIENTS We investigated 392 consecutive IVF patients of all ages, with among them 60.7% suffering from diminished FOR. INTERVENTIONS Interventions included routine IVF cycles. MAIN OUTCOME MEASURE Clinical pregnancy rate in IVF cycle was assessed. RESULTS FSHo, but not AMHo, was, overall, statistically associated with pregnancy chances in IVF. This association was further limited to women above age 38 yr. FSHo was also significantly associated with pregnancy chances in women with normal FMR1 genotype, although only almost reaching significance with heterozygous-normal/low. Normal-genotype patients also demonstrate significant interaction between FSHo and age in pregnancy outcome, although insignificant for all other FMR1 genotypes and subgenotypes and universally for AMHo. CONCLUSIONS AMHo and FSHo are representative of distinctively different components of FOR, likely influenced by different ovarian FMR1 genotypes and subgenotypes.

Lessons from elective in vitro fertilization (IVF) in, principally, non-infertile women

BackgroundWe here report the first investigation of exclusively elective in vitro fertilization (IVF) cycles in women with no apparent history of infertility. Since IVF outcome in women with infertility are always influenced by underlying causes of infertility, a study on non-infertile women may offer new insights.MethodsWe investigated 88 females without history of infertility in 109 consecutive elective IVF cycles, almost exclusively performed for purposes of preimplantation genetic screening (PGS; i.e., elective gender selection). The following questions were addressed: (i) impact of PGS on IVF pregnancy chances; (ii) impact of transfer of 1 vs. ≥2 embryos on IVF pregnancy chances; (iii) correlation of anti-Müllerian hormone (AMH) levels to embryo ploidy (iv) effect of gonadotropin dosage used in stimulation on available embryos for transfer; and (v) in form of a 1:1 case control study, compared 33 elective PGS cycles with matched control cycles without PGS, performed in couples with either prior tubal ligations and/or severe male factor infertility as indication of IVF.ResultsThe overall clinical pregnancy rate for the group was 36.7%; pregnancy was associated with number of euploid (P = 0.009) and number of embryos transferred (P = 0.001). Odds of pregnancy were 3.4-times higher if ≥4 euploid embryos were produced in comparison to <4 (95% CI 1.2 to 9.2; P = 0.019), and odds of pregnancy were 6.6-times higher if greater than or equal to 2 rather than <1 euploid embryos were transferred (95% CI 2.0 to 21.7; P = 0.002). Increasing AMH (P = 0.001) and gonadotropin dosage used in ovarian stimulation (P = 0.024), was, independently, associated with number of available euploid embryos. Increasing AMH, but not follicle stimulating hormone (FSH), was associated with number of embryos available for biopsy and PGS (P = 0.0001). Implantation rates were 26.4% with PGS and 9.5% without (P = 0.008). Women undergoing PGS, demonstrated 4.58-times higher odds of pregnancy than matched controls (95% CI 1.102 to 19.060, Exp 4.584, P = 0.036).ConclusionsThis study suggests that outcomes of elective IVF cycles may significantly deviate from infertility-associated cycles. Affirming proof of concept for PGS, utilizing day-3 embryo biopsy and fluorescence in-situ hybridization (FISH), both widely held responsible for earlier failures to establish such proof, suggests that the principal cause of prior failures were likely not insufficient laboratory techniques but poor patient selection for PGS. Such a conclusion questions the current reintroduction of PGS with improved techniques and technologies in absence of prior determination of suited patient populations.