2006 Annual Meeting
ABSTRACT 1
Recurrent Spontaneous Abortion and Skewed X-inactivation: Is There an Association?By Invitation
W. Allen Hogge, M.D.
Tracy L. Prosen, M.D. M.Sc.
Mark C. Lanasa, M.D.
Heather A. Huber
Matthew F. Reeves, M.D.
OBJECTIVE: The purpose of this study was to determine: a) whether there is an association between skewed X-inactivation and recurrent spontaneous abortion in a large well-defined sample of women with recurrent loss, and b) whether women having an aneuploid spontaneous abortion have a higher rate of extremely skewed X-inactivation than patients with chromosomally normal abortion.
METHODS: X-chromosome inactivation patterns were compared in five groups of women. Group I consisted of 418 women with 2 or more spontaneous losses. In Group II, there were 349 subjects from infertility practices recruited at the time of a positive serum betahCG. Group III women (N= 81) were recruited at the time of an ultrasound diagnosis of an embryonic demise or an empty gestational sac. Groups IV and V were healthy pregnant subjects previously enrolled in another study to determine the incidence and etiology of pregnancy complications, such as pre-eclampsia and intrauterine growth restriction. Group IV included 114 women in their first pregnancy, while Group V consisted of 101 women with varied pregnancy histories, but without recurrent pregnancy loss.
RESULTS: The rate of extreme skewing (90% or greater) in Group I (RSA) was 7.7%, and not statistically different from any of the other groups, except Group IV (1.0%, p = 0.01). In Group II the incidence of extreme skewing was not significantly different between those with successful pregnancies (5.2%), and those with spontaneous abortion (3.1%). Of the 46 patients with chromosomally abnormal spontaneous abortion material, only one patient had extreme skewing, and the karyotype of the abortion was 69, XXX. The incidence of X-inactivation skewing of 90% or greater in all groups was no different whether there had been at least one livebirth (5.1%) or no previous livebirths (9.5%, p > 0.05). There was no difference if there had been 2 (6.4%), 3 (8.4%) or 4 or more losses (8.4%) in the incidence of extreme skewing (p > 0.05). When age and skewing of 90% or greater are compared, subjects with extreme skewing have a mean age of 2 years older than those without extreme skewing (p<0.05).
CONCLUSION: Skewed X-inactivation is not associated with recurrent spontaneous abortion, but is associated with increasing maternal age.
ABSTRACT 2
GPR30: A Novel Indicator of Poor Survival for Endometrial CarcinomaBy Invitation
Harriet O. Smith, M.D.
Kimberly K. Leslie, M.D.
Meenakshi Singh, M.D.
Clifford R. Qualls, Ph.D.
Chetana M. Revankar, Ph.D.
Nancy E. Joste, M.D.
Eric R. Prossnitz, Ph.D.
OBJECTIVE: To evaluate the relationship between GPR30, classical steroidal receptor expression (ER/PR), and clinical outcome in patients with endometrial carcinoma.
STUDY DESIGN: Immunohistochemistry was used to investigate the expression of GPR30, estrogen (ER), progesterone (PR), epidermal growth factor (EGFR) receptors and Ki-67 in 47 consecutive consenting patients with endometrial carcinoma diagnosed between 1997 and 2001. Results were correlated with clinical and pathologic predictors of adverse outcome, and survival.
RESULTS: GPR30 correlated positively with EGFR (p = 0.0036), but negatively with PR (p = 0.04) expression. GPR30 overexpression occurred more frequently in tumors with deep myometrial invasion, high grade, biologically aggressive histological subtypes, and advanced stage. In patients with GPR30 overexpression, survival was significantly poorer (65.2% vs. 100%, p = 0.005).
CONCLUSION: GPR30 represents an alternative estrogenresponsive receptor that is overexpressed in tumors where estrogen and progesterone receptors are down regulated, and in high-risk endometrial cancer patients with lower survival rates.
ABSTRACT 3
Identifying the Safety Profile of a Novel Infectivity Enhanced Conditionally Replicative Adenovirus (CRAd), Ad524-RGD, in Anticipation of a Phase I Trial for Recurrent Ovarian CancerBy Invitation
Ronald D. Alvarez, M.D.
John G. Page, Ph.D.
Baohong Tian
Karen Schweikart, Ph.D.
Joseph Tomaszewski, Ph.D.
Ray Harris, Ph.D.
Trevor Broadt
Judith Polley-Nelson, Ph.D.
Minghui Wang
Sharmila Makhija, M.D.
Rose Aurigemma, Ph.D.
David T. Curiel, M.D., Ph.D.
OBJECTIVE: To evaluate the biodistribution and toxicity of the tropism modified infectivity enhanced CRAd, Ad524-RGD.
METHODS: : Cohorts of cotton rats were treated intravenously (IV) or intraperitoneally (IP) for 3 consecutive days with 5x108 - 5x1011 particles/kg of Ad524-RGD or controls and sacrificed on Day 8, 17, or 56. For biodistribution studies, tissue samples from 14 organ sites and whole blood were evaluated for the presence of virus using quantitative PCR analysis. For toxicity experiments, tissue from over 30 organ sites and blood were obtained for assessment of virus tissue and biochemical effects.
RESULTS: Ad524-RGD was noted in tested samples at D8 and D17 in both IV and IP treated animals with clearance by D56. There were lower copies of vector noted in blood and liver specimens of IP treated animals. Mild peritonitis histopathologic findings were noted in rats treated IP with Ad524-RGD; pathologic findings did not significantly vary with dose, over time or in comparison to that noted in Ad524 treated animals.
CONCLUSION: These studies provide critical insights regarding Ad524-RGD dosing and anticipated toxicity for a planned clinical trial for ovarian cancer.
ABSTRACT 4
Stromal Cells of Endometriosis Fail to Produce Paracrine Factors That Induce Epithelial 17b-Hydroxysteroid Dehydrogenase Type 2 Gene and Its Transcriptional Regulator Sp1: a Mechanism for Defective Estradiol MetabolismBy Invitation
Serdar E. Bulun. M.D., Ph.D.
You-Hong Cheng, M.D., Ph.D.
Ayse Imir, M.D.
Veysel Fenkci, M.D.
Bertan Yilmaz, M.S.
INTRODUCTION: 17b-Hydroxsteroid dehydrogenase type 2 (HSD2) enzyme is responsible for inactivating the biologically potent estrogen estradiol by catalyzing its conversion to estrone in endometrium. We demonstrated recently a stromal progesterone receptor (PR)-mediated paracrine mechanism responsible for induction of epithelial 17ß-HSD2 expression. Paracrine factors secreted from stromal cells induce the production of the transcription factor Sp1 that binds to the promoter of the 17ß-HSD2 gene to regulate its expression. In contrast to eutopic endometrium, epithelial cells of endometriosis do not express 17ß-HSD2 in vivo in response to progesterone resulting in accumulation of estradiol in this ectopic tissue. The molecular and cellular mechanisms for 17b-HSD2 deficiency in endometriosis are not known.
STUDY DESIGN: We studied herein the differential regulation of epithelial 17ß-HSD2 expression and its promoter activity by paracrine factors from primary cultures of human eutopic endometrial stromal cells (EUS) and primary cultures of human endometriotic stromal cells (EOS).
RESULTS: The addition of conditioned medium (CM) from R5020 (progesterone agonist)-pretreated EUS but not from EOS significantly stimulated 17ß-HSD2 mRNA levels and promoter activity in Ishikawa endometrial epithelial cancer cells. Sp1 level was significantly upregulated by the addition of CM from R5020 pre-treated EUS but not from R5020 pre-treated EOS. Moreover, chromatin immunoprecipitation PCR assay showed that CM from R5020 pre-treated EUS but not from EOS enhanced binding of Sp1 to the 17ß-HSD2 promoter region.
CONCLUSION: Taken together, a stromal cell defect in endometriosis appears to be responsible for the lack of progesterone-mediated secretion of factors that induce Sp1 production and its binding activity to the 17ßb-HSD2 promoter for expression of this gene in epithelial cells. This may be the molecular basis for clinically observed progesterone resistance in endometriosis.
ABSTRACT 5
Predicting Gestational Diabetes: Choosing the Optimal Early Serum MarkerBy Invitation
Jeffrey L. Ecker, M.D.
Karen V. Smirnakis, M.D., Ph.D., M.P.H.
Alicia Plati, B.A.
Myles Wolf, M.S., M.M.Sc.
Ravi Thadhani, M.D., M.P.H.
OBJECTIVE: Serum markers measured early in pregnancy have been associated with the later diagnosis of gestational diabetes (GDM). To select an optimal early marker, we prospectively compared 3 serum markers examined simultaneously in a single cohort.
STUDY DESIGN: A nested case-control design was used to evaluate the association of sex hormone binding globulin (SHBG), high sensitive C-reactive protein (hsCRP) and measures of fasting glucose and insulin (homeostasis assessment model, HOMA) obtained in the late first trimester and early second trimester of pregnancy with the diagnosis of GDM. Multivariate modeling and log likelihood ratios were used to identify the optimal biomarker associated with GDM.
RESULTS: In both first and second trimester samples, SHBG was lower and hsCRP higher among women who subsequently developed GDM. Similarly an elevated second trimester HOMA was associated with GDM. Multivariate analysis suggested that SHBG measured from non-fasting first trimester sera was the best predictor of GDM in our population.
CONCLUSION: Among three biomarkers examined prospectively, first trimester non-fasting SHBG appeared to be the optimal marker to predict subsequent gestational diabetes mellitus.
ABSTRACT 6
Identification of Proteins within the Nuclear Factor-kappa ß Transcriptional Complex Including Estrogen Receptor-aBy Invitation
Kimberly K. Leslie, M.D.
Irv Feldman, B.S.
Gerald M. Feldman, Ph.D.
Charlotte Mobarak, Ph.D.
OBJECTIVE: The objective of these studies was to identify regulatory proteins that participate in the NF-?B transcriptional complex.
STUDY DESIGN: Electromobility gel shift assays (EMSAs) were performed to identify known or suspected proteins interacting with the NF-ÎB DNA response element. Unknown interacting proteins were identified using mass spectrometry. As estrogen receptors (ER) were found to be interacting proteins with NF-?B on DNA, reporter gene assays were carried out to assess how this interaction affected estrogen-mediated transcriptional activity.
RESULTS: ER and the p65 subunit of NF-κB co-localized when p65 was bound to the NF-κB DNA response element and when ER was bound to the estrogen response element. This interaction was inhibitory for ER transcriptional activity. Sequencing of proteins bound to the NF-κB/DNA complex identified DNA and RNA modifying enzymes, scaffolding proteins, heat shock proteins, and elements of the nuclear matrix.
CONCLUSION: Together, these factors comprise the molecular machine that controls NF-κB transcription.
ABSTRACT 7
The Effect of Over-expression of SFlt-1 on Blood Pressure, and the Occurrence of Other Manifestations of Preeclampsia, in Unrestrained Conscious Pregnant MiceBy Invitation
George R. Saade, M.D.
Fang Lu
Monica Longo, M.D., Ph.D.
Esther Tamayo
William Maner
Ayman Al-Hendy, M.D., Ph.D.
Garland D. Anderson, M.D.
Gary D. V. Hankins, M.D.
OBJECTIVE: It has been shown that the level of soluble fms-like tyrosine kinase1 (sFlt-1) is elevated in gravidas destined to develop preeclampsia, and a role for sFlt-1 in its pathogenesis has been suggested. Our objective was to determine the effect of overexpression of sFlt-1 on blood pressure and occurrence of other manifestations of preeclampsia in pregnant mice.
METHODS: At day 8 of gestation CD-1 mice were randomly allocated to injection of an adenovirus carrying sFlt-1 (109 PFU; sFlt-1 group), adenovirus carrying the murine IgG2· Fc fragment (109 PFU; mFc group used as a control for the virus), or saline (100µl; saline group). At day 10 of gestation, blood pressure (BP) catheters were inserted through the left carotid artery into the aortic arch and tunneled to a telemetric transmitter. BP was monitored continuously in the conscious unrestrained animals until day 18. Blood was collected from the pregnant mice at different gestational times and plasma sFlt-1 was measured by ELISA. Pups and placentae were weighed, and maternal platelet counts determined at sacrifice on day 18.
RESULTS: Plasma levels of sFlt-1 increased significantly in the sFlt-1 mice and were significantly higher than the 2 control groups. The mean BP in the sFlt-1 mice was significantly higher on days 17 and 18 of gestation compared with the mFc and saline groups. The timecourse of BP rise mirrored that of the sFlt-1 levels. The average pup weight, placental weight, and maternal platelet counts were significantly lower in the sFlt-1 group compared with the controls.
CONCLUSION: sFlt-1 induces hypertension and fetal growth restriction in pregnant mice, supporting its hypothesized role in the pathogenesis of preeclampsia. This animal model minimizes the need for manipulation or administration of various compounds to induce the condition.
ABSTRACT 8
Correlation of ACTH-Stimulated Steroid Levels between Women with Polycystic Ovary Syndrome and Their SistersBy Invitation
Ricardo Azziz, M.D., M.B.A., M.P.H.
Mark O. Goodarzi, M.D., Ph.D.
Xiuqing Guo, Ph.D.
OBJECTIVE: The purpose of this study was to assess the sibling correlation of ACTH-stimulated steroid hormone levels between probands with polycystic ovary syndrome (PCOS) and their sisters.
STUDY DESIGN: Twenty-four women with PCOS and 25 of their sisters underwent an ACTH stimulation test with measurement of the steroids DHEA, androstenedione, and cortisol 60 minutes later. The 60-minute values were used to calculate sister-sister correlations by regression analyses.
RESULTS: The 60-minute androstenedione and the cortisol values were significantly correlated between siblings (r=0.43, p=0.033 and r=0.50, p=0.009, respectively); a similar trend was observed for the 60-minute DHEA values (r=0.31, p=0.13).
CONCLUSION: Women with PCOS and their sisters have significantly correlated levels of ACTH-stimulated steroids, supporting a genetic basis of the adrenal androgen excess observed in PCOS.
ABSTRACT 9
Increased Leukocyte Adhesion to Vascular Endothelium in Preeclampsia is Inhibited by AntioxidantsBy Invitation
Scott W. Kauma, M.D.
Seongho Ryu, M.S.
Alison R Huppmann, M.D.
Nirmala Sambangi, M.D.
Peter Takacs, M.D.
OBJECTIVES: To test the hypothesis that plasma from women with preeclampsia increases leukocyte adhesion to vascular endothelial cells and antioxidants inhibit this effect.
STUDY DESIGN: Plasma from 12 severe preeclamptic women and from 12 normal pregnant women was tested in an in vitro leukocyteendothelial adhesion assay in the presence or absence of vitamin E, vitamin C, or N-acetyl-cysteine.
RESULTS: Preeclamptic plasma significantly increased monocyte (U937 cells) and T-cell (Jurkat) adhesion to human umbilical vein (HUVEC) and microvascular endothelial cells compared to normal pregnant plasma. The antioxidants vitamin E, vitamin C, and Nacetyl-cysteine significantly inhibited monocyte adhesion to HUVEC in the presence of preeclamptic but not normal pregnant plasma. Increased adhesion in response to preeclamptic plasma was not mediated through a PKC mechanism, as the PKC inhibitor GFX had no effect on adhesion in the presence of preeclamptic plasma.
CONCLUSION: Severe preeclampsia is associated with increased leukocyte-endothelial adhesion and clinically useful antioxidants can inhibit this effect.
ABSTRACT 10
The Effects of Metformin and Rosiglitazone, Alone and in Combination, on the Ovary and Endometrium in PCOSBy Invitation
Richard S. Legro, M.D.
Richard J. Zaino, M.D.
Laurence M. Demers, Ph.D.
Allen R. Kunselman, M.A.
Carol L. Gnatuk, M.D.
Nancy I. Williams, Sc.D.
William C. Dodson, M.D.
OBJECTIV(S): To examine the effects of metformin and rosiglitazone, alone and in combination, on endometrial histology and ovarian steroid production.
STUDY DESIGN: Randomized open label study of metformin and rosiglitazone in 16 women with PCOS performed at a single academic health center. The study consisted of a 6 week baseline observation period, a 3 month treatment period of single agent therapy (rosiglitazone or metformin) and then a 3 month period of combined therapy.
RESULTS: Abnormal endometrial histology was found in three subjects at baseline, including one case of adenocarcinoma of the endometrium in an asymptomatic subject, who was excluded from further study. The two other abnormal cases (disordered proliferation) resolved with treatment. Three months of single agent therapy showed a benefit of rosiglitazone (N= 9) compared to metformin (N = 6) in terms of reducing circulating unbound testosterone levels (-11.8, 95% CI: -21.7 to -2.0 ng/dL) and 2h glucose (-42.0, 95% CI: -76.2 to -7.8 mg/dL), 2h insulin (-150.4, 95% CI: -272.7 to -28.1 µU/mL) as well as a significant decrease in integrated levels of glucose and insulin by area under the curve analysis, all obtained from oral glucose tolerance testing. Daily urinary progestin to estrogen ratios improved on rosiglitazone compared to metformin therapy (0.08, 95% CI: 0.02 to 0.14). Ovulatory rates tended to improve on both treatments and combined treatments (33%, 30/90 cycles), compared to baseline ovulatory rate (13%, 2/15). Despite six months of therapy alone or in combination, 5 women displayed no evidence of biochemical ovulation by urinary or serum progestin measurements.
CONCLUSION: This study provides preliminary evidence that insulin-sensitizing drugs may have beneficial effects on the endometrium, though the exact mechanism beyond improving ovulatory function is still unknown. In addition, we suggest that rosiglitazone may be more beneficial than metformin therapy on raised insulin and androgen levels in an obese PCOS population. Combined therapy did not demonstrate significant benefit above and beyond single agent therapy.
ABSTRACT 11
The PESSRI Study: Satisfaction Outcomes of a Randomized Crossover Trial of the Ring and Gellhorn PessariesBy Invitation
Geoffrey W. Cundiff, M.D.
Cindy L. Amundsen, M.D.
Alfred E. Bent, M.D.
Kimberly W. Coates, M.D.
Joe Schaffer, M.D.
Kris Strohbehn, M.D.
Victoria Handa, M.D.
OBJECTIVE: The aim of this randomized crossover trial was to compare symptom relief for women using the ring with support and Gellhorn pessaries.
STUDY DESIGN: Subjects were randomized to use each pessary for 3 months. Outcome data included a visual analog satisfaction score, and quality of life questionnaires. Analysis included student's t-test, Wilcoxon Signed-rank test and logistical regression.
RESULTS: Subjects were primarily white, parous, postmenopausal women with a mean age of 61. The median POPQ stage was III. We enrolled 134 subjects and collected 3-month data on 84 ring and 92 Gellhorn subjects. There was a statistically significant change in the majority of the PFDI and PFIQ scales.
CONCLUSION: The ring with support and Gellhorn pessaries are effective and equivalent in relieving symptoms of protrusion and voiding dysfunction.
ABSTRACT 12
Cervical Cancer Prevention: Safety, Acceptability, and Feasibility of a Single-visit Approach in Accra, GhanaBy Invitation
Paul D. Blumenthal, M.D., M.P.H.
Lynne Gaffikin, Dr.P.H.
Sylvia Deganus, M.B., Ch,B.
Robbyn Lewis, M.P.H.
Mark Emerson, M.A.
Sydney Adadevoh, M.D.
for the Ghana Cervicare Group
OBJECTIVE: To assess the safety and acceptability of a single-visit approach to cervical cancer prevention combining visual inspection of the cervix with acetic acid wash (VIA) and cryotherapy in Accra, Ghana.
STUDY DESIGN: Observational. Nine clinicians were trained in VIA and cryotherapy. Over 18 months 3665 women were VIAtested at a community hospital. If positive and eligible, cryotherapy was offered immediately. Treated women were followed-up at 3 months and at one year.
RESULTS: The test-positive rate was 13.2%. Of those eligible, 70.2% and 21% received immediate or delayed treatment, respectively. No major complications were recorded and only 5.5% presented for a perceived problem post-cryotherapy. Over 90% expressed satisfaction with their experience. One year later, the squamocolumnar junction was easily identified and only 2.6% were test positive.
CONCLUSION: A single-visit approach using VIA and cryotherapy proved to be safe, acceptable, and feasible in an urban African setting.
ABSTRACT 13
Oral Contraceptive Discontinuation: Do Side Effects Matter?By Invitation
Carolyn L. Westhoff, M.D.
Stephen Heartwell, Ph.D.
Sharon Edwards, M.D.
Mimi Zieman, M.D.
Gretchen Stuart, M.D.
Carrie Cwiak, M.D.
Anne Davis, M.D.
Tina Robilotto, M.P.H.
Linda Cushman, Ph.D.
Debra Kalmuss, Ph.D.
OBJECTIVES: To assess self-reported side effects in women after starting the OC, and to compare discontinuation rates according to presence or absence of side effects.
STUDY DESIGN: 1716 women aged less than 25 years initiated the OC at 3 publicly funded family planning clinics and completed structured interviews after 3 and 6 months.
RESULTS: Nearly 60% of subjects discontinued the OC by 6 months. Most subjects reported no changes in headaches, weight, moodiness, and sexual satisfaction during the first 3 months of OC use. Subjects with any complaints, especially those with increased headaches or moodiness, were more likely to discontinue the OC prematurely. Nonetheless, most discontinuation occurred for reasons unrelated to side effects.
CONCLUSIONS: Side effects are absent or mild among most OC users, but women with complaints are more likely to discontinue. Side effects are a less important reason for discontinuation than widely believed.
AAOGF ENDOWMENT SCHOLAR LECTURE
The Cervix as a Biomechanical Structure
By Invitation
Michael House, M.D.
OBJECTIVE: To use a structural engineering paradigm to investigate
cervical mechanical function during pregnancy.
STUDY DESIGN:
Cervical Strength: A three-dimensional, constitutive model for the
large-strain, time dependent mechanical behavior of the cervical
stroma was developed. The model captured the global tissue
response, which was controlled by the cooperative contributions of
the following major constituents: collagen fibers and hydrated
glycosaminoglycans.
Cervical Loading: The constitutive model was implemented in a finite
element framework using anatomic geometry derived from pelvic
MRI during pregnancy. The cervix was dynamically loaded using
finite element methods.
Simulations: Three simulations were performed: cervical funneling,
uterine contractions and cerclage support.
RESULTS: 1) Funneling occurred in the setting of a "soft" stroma
under conditions of passive loading. 2) Uterine contractions
augmented a baseline passive load. 3) Cerclage support depended
on the integrity of the tissue and the location of the suture.
CONCLUSION: Investigation of cervical mechanical function during
pregnancy is facilitated by objective methods of structural analysis.