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Research and Development, NV Organon, Oss, The Netherlands
(Received December 1, 2006; accepted April 5, 2007)
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Abstract |
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-hydroxytibolone, 3ß-hydroxytibolone, and 
4-tibolone), monosulfated (3-sulfate,17ß-hydroxytibolone and 3ß-sulfate,17ß-hydroxytibolone), and disulfated (3,17ß-disulfated-tibolone and 3ß,17ßS-disulfated-tibolone) metabolites were measured by validated gas chromatography with mass spectrometry and liquid chromatography with tandem mass spectrometry. Detection limits were 0.1 to 0.5 ng/ml (plasma) and 0.5 to 2 ng/g (tissues). In brain tissues, estrogenic 3-hydroxytibolone was predominant with 3 to 8 times higher levels than in plasma; levels of sulfated metabolites were low. In vaginal tissues, major nonsulfated metabolites were 3-hydroxytibolone and the androgenic/progestagenic 4-tibolone; disulfated metabolites were predominant. Remarkably high levels of monosulfated metabolites were found in the proximal vagina. In endometrium, myometrium, and mammary glands, levels of 3-hydroxymetabolites were low and those of sulfated metabolites were high (about 98% disulfated). 4-Tibolone/3-hydroxytibolone ratios were 2 to 3 in endometrium, about equal in breast and proximal vagina, and 0.1 in plasma and brain. It is concluded that tibolone metabolites show a unique tissue-specific distribution pattern explaining the tissue effects in monkeys and the clinical effects in postmenopausal women.
; Kenemans and Speroff, 2005; Landgren et al., 2005; Swanson et al., 2006). It does not act as an estrogen on endometrium, resulting in a low incidence of vaginal bleeding (Morris et al., 1999). It has little effects on breast as shown by no increase in mammographic density or tenderness (Lundström et al., 2002). Thus, it differs from estrogen-progestagen combinations. These effects of tibolone on different tissues are explained by the hypothesis that it is a selective tissue estrogenic activity regulator (STEAR).
In postmenopausal women, 3-hydroxytibolone (3OH-tib) is the predominant nonsulfated, free metabolite in blood, followed by 3ß-hydroxytibolone (3ßOH-tib) (for structures, see Fig. 1). Both 3-hydroxymetabolites bind to estrogen receptors (de Gooyer et al., 2003; Kloosterboer, 2004) and are responsible for the estrogenic activity. Serum levels of tibolone and the 4-isomer of tibolone (4-tib), which bind to progestagenic and androgenic receptors (de Gooyer et al., 2003), are low and become undetectable after 4 to 6 h (Timmer and Houwing, 2002; Timmer et al., 2002). More than 75% of the metabolites in blood are sulfated (Vos et al., 2002). Tibolone and its metabolites have no antagonistic effect on estrogen receptors, rendering it different from selective estrogen receptor modulators (Kloosterboer, 2004). To explain why tibolone is estrogenic in some tissues but not in others, it was postulated that tibolone and its active metabolites are converted into sulfated metabolites and/or into the androgenic/progestagenic 4-tib, as has been shown in vitro (Kloosterboer, 2004). Tibolone can be converted to 4-tib in endometrial explants (Tang et al., 1993); enzymes of the aldoketoreductase family (AKR1C) are able to reduce tibolone to 3-hydroxymetabolites (Steckelbroeck et al., 2006) and back again (Schatz et al., 2005). In addition, 3OH-tib and 3ßOH-tib are substrates for sulfotransferases (SULT) expressed in different tissues (Chetrite et al., 1999a; Falany et al., 2004); SULT2A1 has been reported to be the major endogenous enzyme responsible for sulfation of tibolone metabolites in postmenopausal human tissues (Wang et al., 2006). Although sulfated metabolites are not active at receptor levels, sulfatases are able to readily convert 3-monosulfated (mono-S), but not 17-sulfated, steroids into active estrogenic metabolites (de Gooyer et al., 2001). In breast cell lines, tibolone and its nonsulfated and sulfated metabolites inhibit sulfatases (Chetrite et al., 1997, 1999a; Purohit et al., 2002). In addition, one study showed that tibolone and its metabolites inhibit sulfatase in endometrium-derived cell lines, whereas no inhibition is found in a bone-derived cell line (de Gooyer et al., 2001). These tissue-specific differences in the balance between sulfation and desulfation may contribute to tissue-selective effects of tibolone.
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Although the mechanisms involved in the tissue-selective effects of tibolone have been shown in vitro, it is not known whether these mechanisms play a role in vivo. Confirmation by measuring tissue levels of tibolone and its nonsulfated and sulfated metabolites is not yet available. To obtain these tissue levels, six ovariectomized cynomolgus monkeys were treated p.o. with 0.5 mg/kg/day tibolone, and the tissue levels were analyzed. These monkeys have been selected in view of their marked similarity to humans, as found in a previous study with monkeys fed an atherogenic diet, showing protection by tibolone against bone loss, without stimulation of breast and endometrium (Clarkson et al., 2002, 2004; Cline et al., 2002). The dose of 0.5 mg/kg/day was chosen to be able to measure tissue levels of tibolone metabolites. This study presents the levels of tibolone and its nonsulfated, 3-mono-S, and 3- and 17ß-disulfated (di-S) metabolites in efficacy (brain, vagina), safety (uterus, breast), and control (heart) tissues, and in plasma from ovariectomized monkeys treated with 0.5 mg/kg/day tibolone for 36 days.
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Materials and Methods |
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). After at least 1 month postovariectomy, the monkeys received a single dose, and after a washout of 1 week, they received repeated daily p.o. doses of tibolone (Org OD14; 0.5 mg/kg/day) from days 8 through 44 via nasogastric gavage. Blood, urine, and feces were collected for 7 days after the doses on days 1 and 36 for the assessment of the pharmacokinetic parameters of the sulfated and nonsulfated tibolone metabolites. These results are reported elsewhere (Verheul et al., 2007). At day 44, one monkey was necropsied at each of the following times: 1, 1.25, 2.25, 4, 6, and 24 h after the final dose. This design was chosen to establish concentration-time curves in plasma and tissues, rather than to determine the concentrations at one time point in six animals. Blood was collected into tubes containing K3EDTA and centrifuged for 10 min at 2000g within 1 h, and plasma was stored below –20°C until analysis. The following tissues were washed free of blood and stored at –20°C until further processing and analysis: brain (cortex, hypothalamus, brainstem, cerebellum, hippocampus, midbrain, corpus callosum, and corpus striatum), vagina (distal and proximal), uterus (endometrium and myometrium), breast (mammary glands and fat), and heart.
Assay Methods. The assay procedures for the determination of tibolone and its nonsulfated and sulfated metabolites in plasma and tissue homogenates have been described elsewhere (Verheul et al., 2007).
After weighing, tissues were homogenized with internal standards in 70% ethanol for 5 to 15 min using an ultraturrax. The homogenates were divided into aliquots and stored for at least 24 h before further sample processing. The aliquots were centrifugated; the supernatant was evaporated, redissolved in water, and analyzed. In one aliquot, tibolone, 3OH-tib, and 3ßOH-tib were determined by gas chromatographic and mass spectrometric procedures (ABL, Assen, The Netherlands). The second aliquot was subjected to protein precipitation for analysis of the di-S metabolites: 3S,17ßS-tib and 3ßS,17ßS-tib. Liquid-liquid extraction was performed on the third aliquot; 4-tib was determined in the organic phase; and the mono-S metabolites (3S,17ßOH-tib and 3ßS,17ßOH-tib) were extracted by solid-phase extraction from the water phase. All the analytes were determined by liquid chromatography followed by tandem mass spectrometry procedures (Xendo, Groningen, The Netherlands). It was decided to develop assays for the 3-mono-S metabolites only because 1) the supply of most tissues was very limited; 2) the levels of the 17ß-mono-S metabolites were shown in a pilot experiment to be very low in myometrium and liver; and 3) the 17ß-mono-S metabolites cannot readily be reconverted to the nonsulfated metabolites again (Goldzieher et al., 1988; de Gooyer et al., 2001; Takanashi et al., 2003; Simoncini et al., 2004). In a previous study (Vos et al., 2002) with radiolabeled tibolone, 17ß-mono-S metabolites constituted less than 10% of the radiolabel administered. We have also determined the 3- and 3ß-di-S metabolites because their levels were very high in the pilot study, and these di-S metabolites were considered as end-product for sulfation.
Gas chromatography with mass spectrometry procedures have been validated for human serum, and liquid chromatography/tandem mass spectrometry procedures have been validated for human serum, myometrium, and breast tissue with regard to selectivity, sensitivity, calibration curves, accuracy, precision, stability, dilution, and carryover. The procedures have been used for monkey plasma and tissues without further validation because no monkey control tissue was available for validation. Detection limits were 0.1 to 0.5 ng/ml (plasma) and 0.5 to 2 ng/g (tissues). Analytes in monkey plasma and tissues were determined with acceptable precision (coefficient of variation <20% for overall, within batch, and between batch) and accuracy (bias <20%), except for the mono-S metabolites. Quality control samples for the mono-S metabolites showed that the bias was >20% for low and medium concentrations, resulting in a maximal 60% overestimation. Because mono-S metabolites contributed little to the metabolite patterns, potential overestimation at lower levels was accepted.
For metabolites with concentrations outside the calibration range, a "best estimate" of the concentration is given, provided that the peak exceeded the background by at least 3-fold; if lower, a best estimate of "0" was assigned.
Calculations. The area under the concentration-time curve from 0 to 24 h (AUC0–24), maximum plasma concentration (Cmax), and time to maximum plasma concentration (Tmax) were calculated using WinNonlin version 4.1 on SAS version 9.1.2 (SAS Institute, Cary, NC). Missing values were estimated by intrapolation from adjacent time points. Plasma and tissue ratios and percentages of metabolites within tissues have been calculated using AUC.
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Results |
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OH-tib; the AUC of the other estrogenic metabolite, 3ßOH-tib, was about 20-fold lower. The progestagenic/androgenic metabolite, 4-tib, had about an 8-fold lower AUC than 3OH-tib. The predominant sulfated metabolite was 3S,17ßS-tib; considerable levels of 3ßS,17ßS-tib were present; and levels of mono-S metabolites were low. Peak levels of the free and di-S metabolites were reached after 1 and 1.25 h, respectively. Plasma concentrations of all the metabolites decreased with time (Fig. 2, A and B). The plasma pharmacokinetic parameters of tibolone metabolites at necropsy, based on one monkey per time point, are comparable with the pharmacokinetic results obtained at day 36, based on six monkeys per time point (Verheul et al., 2007), e.g., the AUC at day 36 were 0.1, 69, 5, 14, 127, 3922, 43, and 1801 for tibolone, 3OH-tib, 3ßOH-tib, 4-tib, 3S,17ßOH-tib, 3S,17ßS-tib, 3ßS,17ßOH-tib, and 3ßS,17ßS-tib, respectively. The concentration-time curves at necropsy and at day 36 are superimposable. This indicates that the plasma results at necropsy obtained in a single monkey per time point are a good reflection of the results obtained in six animals.
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Tissues. The AUC of the metabolites in the heart muscle were <3% of those in plasma, indicating that the tissues had been adequately washed free of blood. Tibolone levels were undetectable in tissues and are not presented in tables and figures.
Efficacy Tissues. Brain. Tibolone metabolite patterns were qualitatively comparable in all the brain tissues measured, but quantitative, regional differences were found as shown by the AUC (Table 2). The hypothalamic concentration-time curve is shown in Fig. 3A as a representative brain tissue. The predominant free metabolite in brain tissues was the estrogenic 3OH-tib, with peak levels at 1 to 2.25 h and decreasing to 10 to 20% at 24 h (Fig. 3A). The predominant sulfated metabolite in most brain regions was 3S,17ßS-tib, with peak levels later and lower (3–20 times) than the corresponding 3OH-tib. AUC of 3ßS,17ßS-tib were lower than those of 3OH-tib (Table 2), and AUC of mono-S metabolites were relatively high. Levels of all the sulfated metabolites decreased with time and were less than 1 ng/g at 24 h (data not shown). Assuming that plasma levels (ng/ml) and tissue levels (ng/g) may be compared, AUC in most brain areas were higher for 3OH-tib, 3ßOH-tib, and 4-tib than in plasma. In contrast, AUC of di-S metabolites in most brain tissues were considerably lower, ranging from 1/25 times in midbrain to <1/100 times in other brain regions.
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Vagina. The predominant free metabolite in distal vagina was 3OH-tib. In proximal vagina, the androgenic/progestagenic 4-tib metabolite had the largest AUC of the three nonsulfated metabolites (Table 2), although the difference with 3OH-tib is not large. In both distal and proximal vaginal tissues, 3S,17ßS-tib was the predominant inactive sulfated metabolite, followed by 3ßS,17ßS-tib (Table 2). The Tmax were 4 and 1 to 2.5 h for di-S metabolites and for mono-S and free metabolites, respectively. AUC of mono-S metabolites were considerably (6 times) higher in proximal vagina. Compared with plasma, AUC in vagina tissues were lower, except for 3ßOH-tib and 4-tib and for mono-S metabolites (proximal vagina only). Metabolite patterns in vaginal tissues are different from the metabolite patterns in brain and plasma.
Safety Tissues. Uterus. The AUC of the estrogenic metabolite, 3OH-tib, in endometrium was 8-fold higher than that of 3ßOH-tib (Table 3; Fig. 3B). After their peak at 1 h, both decreased rapidly. In contrast, 4-tib levels initially increased for about 4 h and became higher than those of 3OH-tib and 3ßOH-tib (Fig. 3B). The predominant sulfated metabolite in endometrium was 3S,17ßS-tib, whereas the mono-S metabolites in endometrium were 40- to 50-fold lower than those of the di-S. The metabolite pattern in myometrium was comparable with the endometrial pattern. Compared with plasma, AUC of free and sulfated metabolites were lower in uterine tissues, except those of 4-tib, which were considerably (8- to 10-fold) higher. Compared with brain and vagina, the AUC of di-S and 4-tib in uterus were higher. The metabolite pattern in uterus is different from those in plasma, brain, and vagina.
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Breast. The predominant metabolites in glandular breast tissue were di-S metabolites (Table 3) with peak levels at 4 h. AUC of 3OH-tib and 3ßOH-tib were 30 times and >100 times lower (Table 3). The AUC of 4-tib was comparable with that of 3OH-tib. As in the endometrium, levels of 3OH-tib (Tmax = 1 h) in the breast rapidly decreased, whereas those of 4-tib (Tmax = 2.25 h) appeared to decrease later in time. Compared with fat breast tissue, AUC of mono-S and di-S metabolites in glandular tissue were higher (1.5-fold), whereas those of free metabolites were lower (0.5-fold). Compared with plasma, AUC in glandular and fat tissue of S-metabolites were lower (3-fold); those of 3OH-tib and 3ßOH-tib were comparable; and those of 4-tib were higher. Compared with vaginal tissue, in particular proximal vagina, the AUC of the mono-S in breast tissues were lower. In both breast tissues and uterus, the AUC of the di-S metabolites were large, but the AUC of the 4-tib were lower in mammary glands and higher in mammary fat tissues. The metabolite pattern in breast tissues is different from those in the efficacy tissues, brain, and vagina and from that in uterus.
Ratios. We calculated various ratios and percentages to characterize and compare the qualitative metabolite profiles independent of the actual levels and to illustrate differences in metabolite patterns in different tissues (Tables 2 and 3). In brain tissues, percentages of free, nonsulfated metabolites were high, in contrast to plasma and all the nonbrain tissues. Percentages of mono-S metabolites, which can be back-converted to active estrogenic metabolites, were less than 10%, except in most brain tissues and proximal vagina. This indicates that sulfated metabolites in plasma, distal vagina, uterus, and breast predominantly (>95%) occur in the di-S form. The difference in percentage of mono-S/total-S between proximal (39%) and distal (9%) vagina was remarkable. In all the tissues and plasma, 3OH-tib was predominant over 3ßOH-tib, with ratios ranging from >20 in brain to about 2 in vagina. The ratio [(tib +4-tib) / (3OH-tib + 3ßOH-tib)] favored the estrogenic (3OH-tib + 3ßOH-tib) metabolites in plasma and all the brain tissues, was about equal in the vagina (proximal and distal), breast (glands and fat), and myometrium, and was clearly toward the progestagenic and androgenic (tibolone +4-tib) metabolites in endometrium. These ratios illustrate that tibolone has unique metabolite patterns in plasma and tissues.
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Discussion |
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OH-tib, whereas levels of tibolone and 4-tib are low and rapidly decrease over time. A quantitative difference from humans is the ratio of 3OH-tib/3ßOH-tib in plasma (2- to 35-fold versus a constant 3-fold in humans). This may be attributed to differences in species, formulation, or design (e.g., dose). The 0.5-mg/kg/day dose was used to enhance the chance to detect metabolites in plasma and tissues and also at longer intervals. Based on body weight, this is 10- to 15-fold, and on calorie intake, 2- to 3-fold higher than the human dose (Clarkson et al., 2002; Cline et al., 2002).
Enzymes involved in tibolone's metabolism were described previously. Tibolone is rapidly hydrolyzed by AKR1C isoenzymes (Steckelbroeck et al., 2006). The liver-specific AKR1C4 preferentially forms 3OH-tib, accounting for the predominance of 3OH metabolites in plasma. AKR1C1 and AKR1C2 preferentially forming 3ßOH-tib were shown in tissues resulting in 3ßOH-tib predominance in tissues in vitro. However, our data show that 3OH-tib is predominant in monkey tissues, suggesting that in vitro data should be extrapolated with caution to in vivo. The high degree of sulfation in plasma may be explained by the SULT2A1 enzyme in the stomach, liver, and intestine (Wang et al., 2006), which is able to convert tibolone and its metabolites to monosulfates and disulfates (Falany et al., 2004). Sulfation renders compounds inactive at receptors. Sulfation at the 17-position seems to be irreversible (de Gooyer et al., 2001) as shown in rats with double-labeled estradiol sulfate (Takanashi et al., 2003). Two studies suggest, however, that 17-sulfation may, to some degree, be reversible (Goldzieher et al., 1988; Simoncini et al., 2004).
Both AUC and concentration-time curves of tibolone metabolites in plasma at necropsy from one monkey per time point were similar to AUC and concentration-time curves obtained on day 36, based on the mean of six monkeys. Thus, it would appear that—despite single measurements at necropsy—these results are a good reflection of the actual situation. The time-dependent concentration-time curves in tissues also support this conclusion.
Our results show that tibolone metabolites have unique patterns in various tissues. Brain tissues contain high levels of 3-hydroxymetabolites and low levels of di-S metabolites and a considerable percentage of mono-S metabolites (Fig. 4). One explanation may be that only more lipophilic, nonsulfated metabolites can pass the blood-brain barrier. However, this would not explain the higher levels of free metabolites in brain compared with plasma. Recently, organic anion transporter proteins have been found in various tissues, including the brain (Kullak-Ublick et al., 1998; Steckelbroeck et al., 2004). These proteins can transport 3-sulfated steroids, such as estrone sulfate and dehydroepiandrosterone sulfate, across membranes. Assuming that these organic anion transporter proteins described in human temporal lobe are expressed in monkey brain and that they act on other 3-sulfated steroids, it would explain the relatively high levels of the 3-mono-S tibolone metabolites in brain tissues. Mono-S metabolites entering the brain can then readily be desulfated by sulfatases (Goldzieher et al., 1988; de Gooyer et al., 2001; Takanashi et al., 2003), leading to receptor-active 3OH-tib and 3ßOH-tib. Thus, the 3-mono-S metabolites can serve as a reservoir for estrogenic activity. These two mechanisms, transporters for 3-mono-S steroids and local presence of sulfatases, can explain high levels of free and mono-S metabolites in brain tissues. The regional differences in brain mono-S and di-S metabolite levels may be explained by a site-specific balance between SULT and sulfatase activity; for example, the lack of activity and mRNA expression of SULT in the temporal lobe (cortex) of human brain (Kullak-Ublick et al., 1998) may account for the low di-S levels in the cortex, whereas different SULT activities as shown in different brain tissues in rats (Aldred and Waring, 1999) may explain regional differences in sulfation. The region-specific expression of SULT4A1 in human brain further supports this notion (Liyou et al., 2003), when SULT4A1 can also sulfate tibolone and its metabolites. Irrespective of these mechanisms, our data show that estrogenic 3OH-tib and 3ßOH-tib are predominant in brain, including the hypothalamus, which is primarily involved in control of hot flushes (Stearns et al., 2002). This estrogenic metabolite pattern is in line with the observed estrogenic effects on the brain, i.e., a reduction in hot flushes as shown in ovariectomized monkeys (Jelinek et al., 1984) and postmenopausal women (Landgren et al., 2005). However, estrogenic effects of tibolone on the pituitary seem not to be very strong because tibolone only partly reduces follicle-stimulating hormone levels in postmenopausal women (Doeren et al., 2001) and cynomolgus monkeys (Gibbs et al., 2002), in contrast to continuous combined regimens resulting in near-complete suppression. Unfortunately, the pituitary was too small for assessment of hormone levels.
Compared with the brain, the proximal vagina has a different metabolite pattern (Fig. 4) with higher progestagen/estrogen ratios. The vaginal balance is only slightly toward estrogenic metabolites, but this is apparently sufficient for favorable vaginal effects of tibolone (Kloosterboer, 2004). Interestingly, in the proximal vagina, the percentage of mono-S metabolites is high (39%) compared with the distal vagina. These high levels of 3-mono-S metabolites in the proximal vagina can be desulfated to active estrogenic hydroxy metabolites and may thus serve as an estrogenic reservoir. The androgenic/progestagenic 4-tib is likely not to interfere with tibolone's estrogenic effects on the vagina: in rats (de Gooyer et al., 2003) progestagens do not interfere with estrogenic responses in vagina, and in early postmenopausal women addition of medroxyprogesterone acetate does not prevent estrogenic effects of conjugated equine estrogens on the vagina maturation index (Utian et al., 2001). The androgenic effects of 4-tib may also contribute to vaginal lubrication (Traish et al., 2002). These observations provide an explanation for favorable effects of tibolone on vaginal atrophy and dryness observed in postmenopausal women (Morris et al., 1999; Kloosterboer, 2004; Kenemans and Speroff, 2005).
Compared with efficacy tissues, metabolite patterns in safety tissues were different. In contrast to the brain, tibolone metabolites in the mammary glands were almost completely (98%) in the di-S form (Fig. 4), implying that the balance is toward sulfation. This is supported by in vitro data. Falany et al. (2004) have shown that of the three major human SULT, SULT2B1b is expressed in breast tissue, and that tibolone, 3OH-tib, and 3ßOH-tib are substrates. In addition, tibolone and its metabolites inhibit sulfatase activity in breast cell lines (de Gooyer et al., 2001). These combined mechanisms shift the system toward sulfation. The AUC of the androgenic/progestagenic 4-tib metabolite are higher in breast tissues than in plasma and the brain, and the AUC-based 4-tib/3-OH-tib ratios in mammary glands and fat are approximately 1, suggesting no androgenic/progestagenic or estrogenic predominance. Recently, Stute et al. (2006) showed that in monkeys the amount of breast sulfatase activity is lower after 2 years of tibolone treatment in tissues with a high fat content.
The potential stimulatory estrogenic effects of tibolone on the breast may be attenuated by the androgenic effects of 4-tib because physiological doses of testosterone inhibit estrogen-induced mammary epithelial proliferation in ovariectomized monkeys (Dimitrakakis et al., 2003). These observations suggest antiestrogenic effects of tibolone on breast tissue, in line with its neutral effects on mammographic density and its low incidence of breast tenderness in postmenopausal women (Lundström et al., 2002; Hofling et al., 2005).
In the endometrium, the percentage of sulfated metabolites is also high (>97%) (Fig. 4), indicating that comparable mechanisms may be operational to those in the breast. SULT1E1 is expressed in endometrium and modulated by progestagenic activity (Falany and Falany, 1996; Chetrite et al., 1999a,b), and sulfatase activity in endometrial tissue-derived cell lines is inhibited by tibolone and its nonsulfated and sulfated metabolites (de Gooyer et al., 2001). In addition to the high sulfation rate, levels of 4-tib in endometrium increase after about 1 h and remain high, exceeding levels of 3OH-tib and 3ßOH-tib after about 4 h. This increase in 4-tib at times when plasma 4-tib is undetectable strongly suggests local formation from the 3-hydroxy metabolites, possibly by AKR1C enzymes, as shown in vitro (Schatz et al., 2005; Steckelbroeck et al., 2006). The (tib +4-tib)/(3OH + 3ßOH-tib) ratios in the endometrium are toward the androgenic/progestagenic 4-tib. The presence of 4-tib is supported by the generation of progestagen-sensitive factors in endometrial cells (Tang et al., 1993; Schatz et al., 2005). It appears that at least two mechanisms preventing stimulation are operational in the endometrium: a high degree of sulfation and the presence of the progestagenic metabolite 4-tib. This corroborates with the lack of endometrial stimulation after tibolone treatment observed in cynomolgus monkeys (Cline et al., 2002) and in clinical studies (Kloosterboer, 2004).
In conclusion, the different tibolone metabolite patterns in plasma and tissues of cynomolgus monkeys are the result of multiple mechanisms, including metabolism and enzymatic inactivation or activation, confirming that tibolone is a selective tissue estrogen activity regulator. The observed patterns explain the clinical effects in postmenopausal women.
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ABBREVIATIONS: 3/ßOH-tib, 3/ß-hydroxytibolone; 4-tib, 4-isomer of tibolone; AKR1C, aldoketoreductase 1C family; SULT, sulfotransferase(s); mono-S, monosulfated; di-S, disulfated; AUC, area(s) under the curve.
Address correspondence to: H. A. M. Verheul, NV Organon, KR6419, Molenstraat 110, 5320BH Oss, The Netherlands. E-mail: herman.verheul{at}organon.com
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/3ß-hydroxysteroid dehydrogenase activities of the four human isoforms of the aldo-keto reductase (AKR)1C subfamily. J Pharmacol Exp Ther 316: 1300–1309.This article has been cited by other articles:
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  H. A. M. Verheul, C. J. Timmer, M. L. P. S. van Iersel, L. P. C. Delbressine, and H. J. Kloosterboer Pharmacokinetic Parameters of Tibolone and Metabolites in Plasma, Urine, Feces, and Bile from Ovariectomized Cynomolgus Monkeys after a Single Dose or Multiple Doses of Tibolone Drug Metab. Dispos., July 1, 2007; 35(7): 1112 - 1118. [Abstract] [Full Text] [PDF]
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) of (3
