matory cytokines, maximal MMP-3 secretion was observed. Addition of E2to the medium and subsequent challenge with pro-inflammatory cytokinesresulted in partial reduction of MMP-3 secretion. Replacement of E2 and Pinto the medium reduced MMP-3 secretion to baseline. Similarly, E2 plusRA addition to endometrial stromal cells reduced MMP-3 expression to nearbasal levels.

Conclusions: Endometrium in the absence of steroids, as seen in post-menopausal atrophic endometrium, responds to a destabilizing challengewith excessive secretion of proteolytic matrix metalloproteinase. Repletionwith estrogen may be only partially effective in restoring endometrialstability as represented by reduced MMP-3 production in isolated stromalcells, whereas exposure to E2 and P results in maximal interference of MMPproduction. Interestingly, addition of E2 and RA also restores endometrialstability. These findings may explain the clinical condition of postmeno-pausal uterine bleeding in the presence of atrophic endometrium and ininstances when hormone replacement therapy is unbalanced in favor ofestrogen rather than progestogen. Achieving optimal endometrial stabilityappears to require the presence of both estrogen and progesterone.

Supported by: Supported through cooperative agreement (U54-HD-37321) as part of the Specialized Cooperative Centers Program in Repro-duction Research.

P-135

Micellar nanoparticles: A new topical emulsion for systemic delivery ofestradiol and testosterone. Marvin Heuer, Craig Wright. Novavax, Co-lumbia, MD; Novavax, Rockville, MD.

Objective: To design a topical oil-and-water—based drug delivery sys-tem containing low concentrations of ethanol (�10%) that safely andeffectively delivers ethanol-soluble drugs systemically.

Design: A retrospective overview of formulation development and pre-clinical testing in rabbits and rhesus monkeys.

Materials/Methods: A nanoemulsion was produced by injecting waterinto a mixture of oil, surfactant, and either estradiol or testosterone (solu-bilized in ethanol). The preferred ratio of the pre-mixed materials to wateris 7:3 on a volume/volume basis. The delivery system is described in USPatent #5,629,021 (assignee: Novavax, Inc., Columbia, Md).

Results: The final products have the consistency and properties of acosmetic—like lotion. Each gram of the EstrasorbTM product contains 2.5mg of estradiol and each gram of the Androsorb� product contains 31.25 mgof testosterone. The resulting emulsions have mean particle sizes �1micron. The products are stable at 30°C for 24 months. Preclinical topicaltesting of Estrasorb and Androsorb in single-dose rhesus monkey and/orrabbit models resulted in significant systemic delivery of estradiol andtestosterone, respectively, with acceptable preclinical safety profiles.

Conclusions: A novel topical emulsion drug delivery system has beendeveloped for ethanol-soluble drugs like estradiol and testosterone. Theseproducts are safe and deliver drugs systemically after a single topicalapplication in our animal models. Both products are candidates for humanclinical trials. This delivery system may also be suitable for other ethanol-soluble drugs, such as other steroids, anti-inflammatory agents, and centralnervous system-active drugs.

Supported by: Novavax, Inc.

P-136

Considering heterogeneity in a power analysis exemplified by ovarianaging. Kristel van Asselt, Helen Kok, Peter Pearson, Egbert te Velde,Diederick Grobbee, Lodewijk Sandkuijl. Univ Medical Ctr Utrecht, Utre-cht, Netherlands; Leiden Univ Medical Ctr, Leiden, Netherlands.

Objective: Age at menopause is probably under influence of variousgenes. The range of normal menopausal age is between 40–60 year with amean of 50–51 years. Age at menopause is not normally distributed with askewness to the left side. Women with premature ovarian failure (POF),defined as cessation of cycles before the arbitrary age of 40 years, could beresponsible for the skewness. A number of deletions and translocations inthe long arm of the X-chromosome have been described in women withPOF. We plan to perform a genome scan to search for genes responsible forthe normal range of age at menopause. Non-parametric linkage analysesapproaches focus on identical by descent (IBD) allele sharing between

relatives at a locus. Relatives with a similar trait value, like menopausal age,are expected to share more alleles IBD at a marker locus linked to a locuscontributing to the trait of interest than they would share under Mendelianindependent segregation. We calculated the power of the sib pairs in ourstudy.

Design: simulation for powercalculation for a double proband sib pairanalysis

Materials/Methods: We collected sisters with extreme concordant anddiscordant ages at menopause, defined as age at menopause before 46 andafter 54 years. The problem could arise that in the lower tail of thedistribution a mixture of women with monogenetic POF causes and womenwith a normal early menopause, probably caused by more genes, arecollected. To estimate the power of such a sib pair analysis we have used aprocedure in which both discordant pairs from the ends of the variation andconcordant pairs from the upper tail of the distribution were used. By doingso we hope to avoid mapping strong POF genes and we are now dealingwith only the normally distributed upper tail of the distribution of meno-pausal age. The IBD probabilities were calculated with the method of Rischand Zhang using the known menopausal ages, the mean menopausal age andthe corresponding standard deviation of a population based cohort. TheseIBD probabilities were used with simulated marker data in a Haseman-Elston model.

Results: Results of the power of 158 sib pairs are given in the table.

Results of the power calculation by Haseman-Elston

HeritabilityNumberof loci

allelefrequency

Powerp �� 0.01

Powerp �� 0.001

Probabilityto detectat least

one locus

80% 2 0.1 92% 70% 99%80% 2 0.25 87% 60% 98%80% 3 0.1 59% 24% 93%80% 3 0.25 46% 17% 85%60% 2 0.1 68% 35% 90%60% 2 0.25 58% 25% 83%60% 3 0.1 34% 10% 72%60% 3 0.25 25% 7% 58%

Conclusions: The dataset is large enough to perform a genome scan tofind regions that might contain genes determining menopausal age.

Supported by: Grants from the University Medical Center Utrecht andOrganon Akzo Nobel.

P-137

Identification and functional characterization of disease-associatedgenes in premature ovarian failure. Andreas Schroer, Wolfgang Kupker,Klaus Diedrich. Medicine Universitat zu Lubeck, Lubeck, Germany.

Objective: Premature ovarian failure (POF) is a heterogeneus conditionand can be defined as secondary amenorrhea, hypoestrogenism and elevatedgonadotropins occurring before the age of 40. The prevalence of earlyovarian failure is 1–2 % of women in the general population. POF can beassociated to autoimmune disorders, ovarian surgery, chemo-radiotherapyor to systemic diseases like galactosaemia. Among genetic causes, X mono-somy as in Turner syndrome or X deletions and translocations and muta-tions of some single genes are known to be responsible for several syn-dromic forms of POF. However, in most cases the aetiology of POF is stillunknown. Familial cases of POF and results from twin studies have pro-vided convincing evidence that genetic factors are important in determingPOF. The prognosis with respect to a pregnancy is unfavourable and thereis no evidence that any treatment can enhance the pregnancy rate of thispatients

Design: 1. Determination of expression profiles: To determine relevantgene expression patterns and single candidate genes causing POF, we usehigh-density oligonucleotide array technology. Targets for hybridization areisolated from primary ovarian cells from well-characterized patients andprobands we see in the clinic. 2. Validation of relevant expression profilesand single genes: The clinical role of the new potential genetic factors willbe validated in clinical trials within clinical networks. 3. Functional genom-ics aspect: The role of the identified genetic alterations will be functionally

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