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Abstract

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Abstract:  Ospemifene (Ophena™) is a new selective oestrogen receptor modulator currently in phase III clinical development for treatment of post-menopausal vulvar and vaginal atrophy. In the present study, we examined the pharmacokinetics, toxicity, and DNA adduct forming potential of ospemifene in the liver and endometrium of rhesus macaques following single and subchronic dosing schedules to better understand the potential toxicologic effects of ospemifene. During single weekly dosing, six macaques were administered 35 mg/kg/week ospemifene orally for 3 weeks. Pharmacokinetics, haematologic toxicity, uterotrophic effects and serum cholesterol levels were monitored. Additionally, two animals were subchronically dosed with 60 mg ospemifene for 9 weeks, followed by 12 mg/day for 3 weeks. Serum cholesterol and pharmacokinetics were monitored, and serial liver and endometrial biopsies were collected during and after treatment to evaluate DNA adduct formation. Following single weekly dosing, no significant haematologic toxicities or uterotrophic effects associated with ospemifene were observed. Peak absorption was 4–5 hr, and the elimination half-life was approximately 22 hr. Serum low-density lipoprotein and triglyceride levels trended lower while no other effects on serum lipids were observed. Subchronic dosing resulted in no haematologic toxicity, a lowering of low-density lipoprotein and triglyceride levels, and an increase in high-density lipoprotein levels that were reversed following cessation of dosing. No clinically relevant uterine or endometrial effects were observed, and no DNA adducts were detected in the liver or endometrial biopsies. The results of our pilot study show that ospemifene may lack genotoxic and toxic effects while having a favourable pharmacokinetic profile.

Ospemifene (Ophena™) is a new selective oestrogen receptor modulator (SERM) that is currently in late phase III clinical development for the treatment of vulvar and vaginal atrophy (VVA) in post-menopausal women. Results from phases I and II clinical studies have shown that ospemifene is generally safe and well tolerated, has pharmacokinetics suitable for once daily dosing, that it has positive effects on markers of bone turnover similar to raloxifene, and that it does not adversely affect climacteric symptoms, vascular markers or quality of life [1–5]. No safety concerns have been raised in any of the clinical trials completed to date.

Similar to other SERMs like tamoxifen and raloxifene, ospemifene has tissue-specific oestrogenic or anti-oestrogenic effects: it acts as an anti-oestrogen in the breast [6–9], has neutral to weak oestrogen agonist effects in the endometrium [5,10,11], and is predominantly oestrogenic in the bone [2,3,6]. As a triphenylethylene derivative, ospemifene shares some similarities with tamoxifen, but there are also some important differences. Unlike tamoxifen, which acts as an oestrogen agonist in the endometrium [12], ospemifene has been shown to have neutral to weak oestrogen agonist effects in the endometrium in clinical trials completed to date [5,10,11]. Although tamoxifen has also been shown to produce liver cancer in rats, this effect may be due to species-specific metabolic activation of tamoxifen to genotoxic metabolites [13,14] as tamoxifen has not been associated with liver cancer in human beings. Previous studies have shown ospemifene and toremifene, both chlorinated triphenylethylenes, to be less genotoxic compared to tamoxifen [15,16]. The most notable difference between ospemifene and the other SERMs is ospemifene's oestrogenic effects in the vagina, a property that appears to be unique to ospemifene among the known SERMs [5,10,11]. Based on the results of phases I and II clinical trials, which showed the positive effects of ospemifene on the vagina, a phase III trial to study the effects of ospemifene on moderate to severe VVA in post-menopausal women was initiated. Positive results from the phase III trial were announced in January 2008 (unpublished data).

As ospemifene may be an important new endocrine agent for the treatment of VVA in otherwise healthy post-menopausal women, it is essential to rule out any potentially adverse effects of ospemifene, particularly with respect to carcinogenic potential. Therefore, we examined the pharmacokinetics, toxicity, effects on serum cholesterol, and the ability of ospemifene to form DNA adducts in the liver and endometrium in the rhesus macaque monkey, in an effort to better understand the potential toxicologic effects of ospemifene.

Materials and Methods

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Drugs.  Toremifene citrate (Fareston®), molecular weight 598.1, and metabolites of tamoxifen and toremifene were provided by the Orion Corporation, Orion-Pharma (Espoo, Finland). Ospemifene (Ophena™), molecular weight 378.9, was provided by Hormos Medical Ltd. (Turku, Finland).

Animals.  All rhesus macaques used in the studies described below were purchased and housed at the California National Primate Research Center (CNPRC). The CNPRC is an organized research unit of the University of California at Davis, which is an Association for Assessment and Accreditation of Laboratory Animal Care accredited institution. All animal studies were conducted under a protocol approved by the University of California at Davis Animal Use and Care Administrative Advisory Committee, and all animals were individually housed in compliance with established standards of the Federal Animal Welfare Act and the Institute of Laboratory Animal Resources’Guide for the Care and Use of Laboratory Animals. The animals were fed Purina Certified Primate Diet (LabDiet® 5048, PMI® Nutrition International, St. Louis, MO, USA), with fruit and other supplements given according to CNPRC environmental enrichment standard operating procedures. Drinking water was provided ad libitum by automatic limit devices. Water quality and purity were monitored according to established CNPRC standard operating procedures. When necessary (e.g. during biopsies and ultrasound examinations), animals were given ketamine anaesthesia. Telazol was administered at the discretion of CNPRC veterinarians. Following the completion of our studies, all animals were returned to the general population following an adequate recovery period.

Single weekly dosing.  Six female rhesus macaque monkeys (Macaca mulatta) (#19560, #17164, #22194, #22449, #20645 and #22971) ranging from 9–17 years of age were used. Ospemifene pharmacokinetics, haematologic toxicity, uterotrophic effects and serum cholesterol levels were monitored. The study was designed so that each animal could serve as its own control. Each monkey was administered ospemifene at a dose of 35 mg/kg once a week for 3 weeks. The animals initially weighed between 5.7 and 10.0 kg and were reweighed before each dose (see table 1 for detailed weight data). The drug was suspended in 20 ml of a 2% (w/v) hydroxypropylmethylcellulose (Sigma-Aldrich, St. Louis, MO, USA) in saline solution and given orally via nasogastric tube while the macaques remained fully conscious. All animals were fasted for 12 hr prior to each dose. The first dose was timed to coincide with the beginning of the menstrual cycle in each animal. Blood specimens (3 ml) for ospemifene pharmacokinetics were collected in green-top Vacutainer® blood collection tubes (Becton Dickinson, Franklin Lakes, NJ, USA) containing sodium heparin at the following time-points: pre-dose #1, 1, 2, 3, 4, 5, 6, 12, 24, 48 and 72 hr post-dose #1. Blood specimens were also collected pre-dose #2, pre-dose #3 and 7 days post-dose #3. For serum lipid analysis and complete blood counts, additional blood specimens (2 ml for each assay) were collected in red-top Vacutainer® clotting tubes (Becton Dickinson) pre-dose #1, pre-dose #2, pre-dose #3, 7 days post-dose #3 and, for complete blood count only, 2 weeks post-dose #3. A fecal specimen was collected from each animal 72 hr post-dose #1. Uterine ultrasound scans were performed prior to dose #1 (baseline), immediately before dose #3, and then 7 days post-dose #3. An endometrial punch biopsy was taken at 48 hr post-dose #3, along with a matching blood specimen, to determine ospemifene drug levels.

Table 1.  Animal weights prior to each dose in the single weekly dose study.
Time-pointWeight (kg)
#19560#17164#22194#22449#20645#22971
Pre-dose 15.668.6410.047.096.596.65
Pre-dose 25.538.8410.027.266.066.62
Pre-dose 35.358.47 9.506.635.686.36

Sample handling, processing and storage. All blood samples drawn for serum lipid analysis were analysed for low-density lipoprotein (LDL), high-density lipoprotein (HDL), total cholesterol and triglycerides. After examining the sample data from the first macaque (#19560), it was determined that additional blood should be drawn at 3, 5, 10 and 14 days post-dose #3, so that serum cholesterol levels could be more closely followed. Serum required for lipid analysis was isolated by centrifugation (2000 ×g for 15 min.) after 30–60 min. clotting time and sent to the Clinical Nutrition and Metabolism Lipid Laboratory at the University of California, Davis for processing. Serum samples were analysed for lipid content according to standard operating procedures following Good Clinical Practice guidelines. Blood specimens for ospemifene pharmacokinetics were immediately centrifuged, and the plasma was removed and frozen at approximately –20° until extracted and analysed by high-performance liquid chromatography (HPLC) as described below. Blood drawn for complete blood counts was processed on site in the CNPRC haematology laboratory. Endometrial biopsies and fecal specimens were also stored at approximately –20° until they could be processed for HPLC analysis.

Ospemifene subchronic dosing.  Two additional female rhesus macaque monkeys (#20630 and #20590), both aged 16 years and 8 months, and weighing 5.8 and 6.6 kg, respectively, were used to conduct a 12-week subchronic dosing study of ospemifene. The purpose of this study was to determine whether or not subchronic ospemifene treatment results in the formation of DNA adducts in the liver and endometrium, and whether the drug has an effect on serum lipids. So that these animals could serve as their own controls, pre-treatment liver and endometrial biopsies were taken, along with 3-ml blood specimens for pharmacokinetic analysis, and 2-ml specimens for serum lipid analysis, complete blood counts and chemistry panels. Each animal was administered 60 mg/day ospemifene (approximately 9.7 mg/kg) for the first 9 weeks of treatment. For the remaining 3 weeks of treatment, the dose was reduced to 12 mg/day. The drug was suspended in 5 ml peanut oil and given orally via nasogastric tube while the animals were conscious. Additional 3-ml blood specimens were collected once a week throughout the study for drug level analysis. As in the previous study, 2-ml blood specimens were taken simultaneously and placed in clotting tubes to provide serum for lipid analysis. Additional liver and endometrial punch biopsies were taken after 3 and 12 weeks of dosing and stored at approximately –70°. Blood was collected post-treatment for complete blood counts and chemistry panels. Two months after the final dose, liver and endometrial biopsies were again collected. Biopsy tissues were used in DNA adduct studies as described below.

HPLC analysis of ospemifene.  Ospemifene was quantified using an HPLC bioanalytical method essentially as described [17]. Briefly, the assay system consisted of a Beckman (Fullerton, CA, USA) Model 320 gradient liquid chromatograph, two Model 110A pumps, and a Model 420 controller. The system was equipped with a Beckman Ultrasphere 5-µm reverse phase C18 ODS 4.6 mm × 250 mm column and 100-µl injection loop connected to a Rheodyne injector (Rohnert Park, CA, USA). Fluorescence of photochemically activated ospemifene and toremifene (used as the internal standard) was detected with a Linear Instruments Model LC305 fluorescence detector (Thermo Finnigan, San Jose, CA, USA) set at an excitation wavelength of 266 nm and an emission wavelength of 370 nm. Retention times and peak heights were recorded using a Hewlett-Packard (Corvallis, OR, USA) Model 3394 integrator.

DNA modification, post-labelling and thin layer chromatography.  

DNA adduct formation in vitro Rhesus macaque spleen DNA was purified by standard methods [18]. DNA (250 µg) was reacted with 100 µg ospemifene in 0.5 ml 20 mM Tris, pH 7.9, 30% ethanol in a rotating incubator at 37° for 64 hr. After extraction with ether and ethanol precipitation, the DNA pellet was dissolved in water and digested with micrococcal nuclease and spleen phosphodiesterase overnight essentially as described [19]. An aliquot (approximately 10–15 µg DNA) was treated with 2 units (U) nuclease P1 (Boehringer Mannheim, Indianapolis, IN, USA) at room temperature for 30 min and labelled with 95 µCi 32P ATP (Amersham Life Science Inc., Arlington Heights, IL, USA) in a final volume of 20 µl. Approximately 2–10 µl were spotted on a PEI-cellulose sheet (Macherey-Nagel, purchased from Bodman, Aston, PA, USA) and thin layer chromatography (TLC) was performed essentially as described [15]. Adduct spots were detected by exposure of Kodak Biomax film (Eastman Kodak Company, Rochester, NY, USA) with intensifying screen for 20 hr at approximately –70°. Autoradiographs were superimposed onto the TLC plates to mark the position of a red marker dye from the isotope solution relative to the adduct spots. As a positive control, monkey DNA was reacted with tamoxifen metabolite Bx (4-hydroxy-N-desmethyl-tamoxifen), a compound known to reproducibly form DNA adducts [15,20].

Analysis of DNA adducts generated in vitro Frozen liver and endometrial biopsies (2–10 mg tissue from animal #20590) were lysed overnight with proteinase K at 55° and extracted using the Qiagen tissue kit (Qiagen Inc., Santa Clarita, CA, USA). The DNA was precipitated with ethanol, dissolved in 10 µl H2O and quantified by a dot blot procedure described elsewhere. The DNA was digested with 0.25 U micrococcal nuclease and 2 mU spleen phosphodiesterase (total volume 12 µl) overnight and 4 µl were subjected to the nuclease P1 and labelling procedure described above. An aliquot of DNA (approximately 1 µg) was analysed by TLC as described above.

Statistics.  The data are expressed as means ± standard deviations where appropriate. Statistical significance of differences between groups was assessed by two-tailed Student's t-test. Statistical calculations were performed using Microsoft Excel® 2004 software (Microsoft, Redmond, WA, USA). P-values of <0.05 were interpreted as being statistically significant.

Results

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Single weekly dosing.

The pharmacokinetics of ospemifene were examined in six female rhesus macaque monkeys following an oral dose of 35 mg/kg once a week for 3 weeks. All animals were weighed prior to each dose (table 1). Figure 1 shows the ospemifene plasma concentrations in each animal up to 72 hr following the initial dose. Peak oral absorption was observed between 4–5 hr (n = 6), with Cmax reaching an average (± standard deviation) of 194 ± 188 nM (74 ± 71 ng/ml) (n = 6). The average (± standard deviation) half-life of elimination (first-order kinetics) was 22.4 ± 3.8 hr (n = 4), which is similar to that observed in phase I clinical trials [1]. An average (± standard deviation) ospemifene concentration of 2.4 ± 1.7 nmoles/g (n = 6) was observed in the endometrial biopsies. Two major metabolites of ospemifene, 4-hydroxyospemifene and carboxyospemifene [9], were detected in the plasma and endometrium (4-hydroxyospemifene only) of each study animal. HPLC analysis of fecal specimens revealed very high concentrations (2.8 nmoles/g) of the parent drug 12 hr after dose #2 in macaque #22449. This compares to a level of 0.2 nmoles/g 6–8 hr after dose #2. An even higher concentration (14.1 nmoles/g) was observed 12 hr following dose #2 in macaque #20645. These data indicate that fecal elimination plays an important role in the metabolism of ospemifene in macaques. At the completion of the study, there was no evidence of haematologic toxicity in any of the six study animals.

image

Figure 1. Ospemifene plasma concentrations in the single weekly dose study. Concentrations were determined by HPLC up to 72 hr following the first 35-mg/kg dose in six rhesus macaques. The vertical axis is scaled logarithmically.

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The six adult, cycling female macaques were examined sonographically prior to (1–3 days pre-treatment), during (1–2 days prior to dose #3), and following (4–5 days after dose #3; day of biopsy) oral treatment with 35 mg/kg ospemifene. Assessments of uterine size, shape, contour, position and texture, as well as appearance of the uterine cavity and thickness of the endometrium, were included in these evaluations. As shown in table 2, there were no significant changes in uterine volume or endometrial thickness during the course of the study. In summary, no significant uterine or endometrial effects of ospemifene treatment were noted in the six animals studied at the time-points evaluated, which is consistent with the results observed in phases I and II clinical trials [5,10,11].

Table 2.  Average uterine volume and endometrial thickness (± standard deviation) in rhesus macaque monkeys following treatment with 35 mg/kg/week ospemifene for 3 weeks (n = 6).
Time-pointUterine volume (cm3)Endometrial thickness (mm)
  • 1

    P = 0.12.

  • 2

    P > 0.05.

Baseline13.3 ± 1.92.7 ± 1.0
Pre-dose 313.6 ± 2.83.6 ± 0.81
4–5 days post-dose 312.9 ± 2.93.8 ± 0.62

Table 3 shows total serum cholesterol, LDL, HDL and triglyceride levels in the six macaques following administration of ospemifene at 35 mg/kg/week for 3 weeks. Although none of these differences were statistically significant (P < 0.05), the data do suggest a trend towards reduction in total cholesterol and LDL and a slight increase in triglycerides. In this study, HDL did not change significantly from baseline.

Table 3.  Average serum cholesterol and triglyceride levels (± standard deviation) in rhesus macaque monkeys following single weekly dosing of ospemifene at 35 mg/kg for three weeks (n = 6).
Time-pointAnimal #Total cholesterol (mg/dl)LDL (mg/dl)HDL (mg/dl)Triglycerides (mg/dl)
  • HDL, high-density lipoprotein; LDL, low-density lipoprotein. 1P = 0.12.

  • 2

    P = 0.27.

  • 3

    P = 0.31.

  • 4

    P = 0.10.

Pre-dose 11956016684 7441
229711888410021
2219413856 7629
1716415883 6740
206451827710024
2244914459 8024
Average163 ± 2074 ± 1383 ± 1430 ± 9
Pre-dose 219560176887282
22971178849022
22194138547640
17164154856043
20645118367631
22449153529627
Average153 ± 2367 ± 2278 ± 1341 ± 22
Pre-dose 319560167 827740
22971171 789111
22194130 517235
171641821056561
20645133 666224
22449181 888823
Average161 ± 2378 ± 1976 ± 1232 ± 17
7 days post-dose 3195601636462No data
22971144548333
22194139706045
17164145765665
20645119486722
22449148637932
Average143 ± 1463 ± 1068 ± 1139 ± 17
14 days post-dose 319560No dataNo dataNo dataNo data
22971161787542
22194134527060
17164160856077
20645138646831
22449120199625
Average143 ± 18160 ± 26274 ± 14347 ± 214

Ospemifene subchronic dosing.

Following 9 weeks of oral ospemifene therapy at 60 mg/day and 3 weeks of oral therapy at 12 mg/day, macaques #20630 and #20590 showed no evidence of haematologic toxicities and had essentially normal blood chemistries. No clinically relevant effects of ospemifene treatment on the uterus or endometrium were observed. Steady state plasma drug concentrations during the first 9 weeks averaged 47.5 ± 28.0 nM. These levels dropped to an average of 25.1 ± 10.7 nM after the dose was reduced to 12 mg/day in the last 3 weeks of the study.

Serum cholesterol.

Following subchronic dosing of ospemifene at 60 mg/day for 9 weeks and 12 mg/day for 3 weeks in macaques #20590 and #20630, definite trends towards decreasing total cholesterol, LDL and triglycerides, and increasing HDL, were seen. Total cholesterol went from a pre-dose average of 157 mg/dl to 147 mg/dl at 9 weeks, when the dose was reduced to 12 mg/day. At 12 weeks, the average total cholesterol declined further to 140 mg/dl. Similarly, average serum LDL dropped from 67 mg/dl pre-dose to 61 mg/dl at 9 weeks, and at 12 weeks the level reached 45 mg/dl. Triglyceride levels fell from a pre-dose average of 108 mg/dl to an average of 76 mg/dl at 9 weeks, and then began to rebound by 12 weeks, when the average triglyceride level was 89 mg/dl. Interestingly, average serum HDL levels increased slightly from 69 mg/dl pre-dosing to 71 mg/dl at 9 weeks and then to 77 mg/dl at 12 weeks. These trends were reversed 2 months after cessation of dosing, with total cholesterol, LDL, HDL and triglycerides returning to near pre-treatment levels.

DNA adduct formation.

When incubated with monkey DNA in vitro, ospemifene did not form DNA adducts (data not shown). In contrast, several DNA adduct spots were observed with tamoxifen metabolite Bx [15,20]. These adducts were used as control for the labelling and TLC conditions. Biopsies from one of the animals in the subchronic dosing study were extracted and sufficient DNA was obtained for analysis of DNA adducts by 32P post-labelling and TLC. As shown in figs 2 and 3, no adducts were observed in either the liver or endometrial biopsies, respectively, after a single dose or subchronic dosing with ospemifene. If adducts were present, labelled spots would be apparent close to the non-radioactive marker (m). Instead, the pattern of radioactivity is the same in biopsies taken before treatment with ospemifene as in biopsies taking during and after treatment, and adduct-like spots (endogenous adducts) are confined to an area near the origin (upper left corner). The positive control (C) shows the location of adducts produced by tamoxifen metabolite Bx. In a second animal, ospemifene drug concentrations in liver and endometrial biopsy tissue were also analysed by HPLC. At week 3, the ospemifene concentration in liver reached 7.05 nmoles/g, falling to 3.41 nmoles/g 2 months after the final dose. Drug concentrations in the endometrium reached 1.39 nmoles/g at week 3, falling to undetectable levels 2 months after the last dose. The ospemifene metabolites 4-hydroxyospemifene and carboxyospemifene (liver only) were also detected in these tissues.

image

Figure 2. DNA adduct analysis of liver biopsies as determined by 32P-postlabeling thin layer chromatography. L0 is before treatment, L1 after a single dose, L2 after 3 weeks of daily dosing, L3 after 12 weeks of dosing, and L4 2 months post-treatment. C represents positive control (monkey DNA reacted with tamoxifen metabolite Bx). The origin is in the upper left corner, and m denotes the location of a non-radioactive marker dye.

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image

Figure 3. DNA adduct analysis of endometrial tissues from the same animal as shown in fig. 2. E0 is before treatment, E1 after a single dose, E2 after 3 weeks of daily dosing, E3 after 12 weeks of dosing, and E4 2 months post-treatment. C represents positive control (monkey DNA reacted with tamoxifen metabolite Bx). The origin is in the upper left corner, and m denotes the location of a non-radioactive marker dye.

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Discussion

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Ospemifene is a new SERM currently being developed for the treatment of post-menopausal VVA, a condition that adversely affects the quality of life for approximately 40% of post-menopausal women [21,22]. Currently, available therapies for VVA include hormone replacement therapy, transvaginal oestrogen products, and non-hormonal lubricants and moisturizers [23]; however, there are limitations associated with each of these treatments [24–26]. Thus, there is an unmet need for safe and effective therapies for VVA.

In the present pilot study, we found no evidence of ospemifene-DNA adduct formation in vitro, and no ospemifene–DNA adducts were detected in vivo in liver or endometrial tissue during or after ospemifene therapy in rhesus macaques, suggesting that ospemifene has a low genotoxic potential similar to toremifene, in agreement with our previous studies [15,20]. Additionally, ospemifene was found to have no significant effects on either uterine volume or endometrial thickness in our study, which is in agreement with clinical data showing that ospemifene has neutral to weak oestrogen agonist effects in the endometrium [5,10,11], in contrast to tamoxifen.

We have also presented data showing that ospemifene has pharmacokinetics well suited for once daily dosing, did not cause any acute or chronic toxicity, and had positive effects on serum cholesterol. The elimination half-life calculated for ospemifene in rhesus macaques (approximately 22.5 hr) was found to be very similar to that found in human beings in phase I studies [1]. High levels of ospemifene were found in fecal specimens, suggesting that fecal elimination is important in ospemifene metabolism in macaques. Not surprisingly, the major route of elimination in human beings has also been found to be through the feces (unpublished data). Although no significant changes in serum total cholesterol, LDL, HDL or triglycerides were observed in the single weekly dose study, this may have been due to the small sample size. Interestingly, serum total cholesterol, LDL and triglycerides trended downward while HDL levels trended upwards in the 12-week subchronic dosing study; however, because only two macaques were included in this study, no definite conclusions can be reached. Nevertheless, positive downward trends in total cholesterol and LDL levels were seen in both the single weekly dose study and the subchronic dosing study, which is in agreement with results observed clinically [4,11].

In conclusion, we have shown in this pilot study that ospemifene does not cause the formation of DNA adducts in the liver or endometrium, suggesting a low genotoxic potential, has ideal pharmacokinetics for once daily dosing, and has no acute or chronic toxicity in rhesus macaque monkeys. These results, coupled with the findings of clinical trials published to date, suggest that ospemifene is a safe and effective therapy for VVA in post-menopausal women.

Acknowledgments

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

This work was supported by grants from Hormos Medical Ltd. (Finland), the Orion Corporation, Orion-Pharma (Finland), Hibbard E. Williams Research Fund (University of California at Davis), the California National Primate Research Center National Institutes of Health grant RR00169 and the UC Davis Cancer Center (NIH grant 2P30 CA93373). The authors would like to thank Sarah Davis and David Robb of the California National Primate Research Center Research Services for their technical expertise in handling and dosing the rhesus macaques used in our studies.

Dr. M. DeGregorio has a potential financial interest in the development of ospemifene. Drs. G. Wurz and U. Hellmann-Blumberg declare no other potential conflicts of interest that would prejudice the impartiality of this scientific work.

References

  1. Top of page
  2. Abstract
  3. Materials and Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References
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