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From the Clinical Research Centers |
Center on Aging (K.M.P., A.M.K., C.U.) and Division of Biostatistics, Department of Community Medicine (M.K.), University of Connecticut Health Center, Farmington, Connecticut 06030-5215
Address all correspondence and requests for reprints to: Dr. K. M. Prestwood, Center on Aging, University of Connecticut Health Center, Farmington, Connecticut 06030-5215.
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Abstract |
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All markers of bone resorption significantly decreased at 12 weeks on treatment compared with placebo and returned toward baseline at 12 weeks posttreatment. Two markers of bone formation, bone alkaline phosphatase and N-terminal procollagen peptides, significantly decreased 12 weeks posttreatment, but the decrease in osteocalcin varied with time and estrogen dose. Based on equivalence testing, the response of markers of bone turnover to therapy with 0.25 mg/day was similar to that seen with 1.0 mg/day. Serum estradiol increased compared with baseline in all treatment groups and compared with placebo in the two higher dose groups. Breast tenderness, bleeding, and endometrial changes were significantly less frequent in the 0.25 mg/day and placebo groups compared with the higher dose groups.
We conclude that low dose of estrogen (0.25 mg/day 17ß-estradiol) reduced bone turnover to a similar degree as that seen with usual replacement therapy (1.0 mg/day 17ß-estradiol), but had a side effect profile similar to that of placebo. In our study additional increases in estradiol levels, as seen with 0.5 and 1.0 mg/day 17ß-estradiol treatment, resulted in more side effects without evidence of additional benefit to bone. These data suggest that 0.25 mg/day 17ß-estradiol may be an effective and tolerable agent for the treatment of osteoporosis in older women.
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Healthy, community-living women over 65 yr of age were recruited from the greater Hartford area to participate in the study. Exclusion criteria for enrollment were 1) diseases that affect bone metabolism such as primary hyperparathyroidism, Paget’s disease, osteomalacia, or multiple myeloma; 2) medications that affect bone metabolism, including glucocorticoids, anticonvulsants, and methotrexate; 3) use of estrogen or calcitonin within the past 6 months; 4) ever use of bisphosphonates or fluoride; 5) history of breast or endometrial cancer within the past 5 yr; 6) baseline endometrial thickness more than 5 mm; 6) recent deep venous thrombosis or other thrombo-embolic event within 6 months of enrollment; and 7) BMD t-score less than -4, symptomatic vertebral fracture within the past year, or past history of hip fracture. We recruited women from the three largest racial/ethnic groups in the greater Hartford area. White women were recruited primarily by newspaper advertisement or by letter of invitation from existing databases. Black and Hispanic women were recruited through community presentations on osteoporosis and invitation to have complimentary BMD screening.
Study protocol
The institutional review board at University of Connecticut Health Center approved the study, and all women gave written informed consent before screening evaluation. Eligible women were stratified by racial/ethnic group and then randomly assigned to a 12-week course of one of four treatment groups: 1.0, 0., or 0.25 mg micronized E2, or placebo. All women also received 1300 mg elemental calcium (given as citrate) with 1000 IU vitamin D/day throughout the study. The primary outcome was bone turnover, as estimated by serum and urinary biochemical markers collected at baseline, 6 and 12 weeks on treatment, and 6 and 12 weeks posttreatment. PTH, E2, estrone, and sex hormone-binding globulin (SHBG) were also measured in serum collected at baseline, 12 weeks on treatment, and 12 weeks posttreatment. Dietary calcium intake was estimated by 4-day food diary, and physical activity was determined using the Physical Activity Scale for the Elderly (9) every 12 weeks. Side effects, including breast tenderness, fluid retention, bloating, and headache, were assessed at each visit by questionnaire using a Likert scale (0 = none, 1 = mild, 2 = moderate, 3 = severe). Bleeding was assessed by diary in which women recorded the number of days of vaginal bleeding or spotting. Endometrial thickness was measured to the closest 1 mm by transvaginal ultrasound (General Electric, Milwaukee, WI) at baseline and 12 weeks on treatment. If the second measurement was more than 5 mm, an additional transvaginal ultrasound was completed at the end of the study. The BMDs of the proximal femur, lumbar spine, and total body were measured by dual energy x-ray absorptiometry (IQ or L, Lunar Corp., Madison, WI) at baseline only.
Biochemical markers of bone turnover and hormone measurements
Serum and urine samples were collected between 0700–0930 h after a 10- to 12-h fast. Serum and urine samples were divided into 0.5-mL aliquots and immediately stored at -80 C. Bone marker assays were run in duplicate after one thaw; all samples for an individual were assayed using the same kit. All bone marker assays, except total deoxypyridinoline cross-links, were performed at the Core Laboratory of the General Clinical Research Center at the University of Connecticut Health Center. Markers of bone formation were osteocalcin (OC), bone alkaline phosphatase (BAP), and N-terminal procollagen peptides (PINP). BAP was measured by enzyme-linked immunosorbent assay (ELISA; Metra Biosystems, Mountain View, CA), OC by immunoradiometric assay (Nichols Institute Diagnostics, San Juan Capistrano, CA), and PINP by RIA (Orion Diagnostica, Inc., Finland). The percent within-run coefficient of variation in the Core Laboratory is 5% for BAP, 4.6% for OC, and 6% for PINP.
Markers of bone resorption were urinary cross-linked N-telopeptides (NTx) and C-telopeptides of type I collagen (CTx) and total deoxypyridinoline cross-links (Dpyr). NTx and CTx were measured by ELISA [Ostex International, Inc. (Seattle, WA), Osteometer A/S (Copenhagen, Denmark), and Metra Biosystems, respectively]. Intraassay variability was 7.6% for NTx and 4.4% for CTx. Total Dpyr was measured by high pressure liquid chromatography in the laboratory of Dr. Markus Seibel, University of Heidelberg (Heidelberg, Germany), with an intraassay variability of less than 10%.
Assays for PTH, E2, estrone, and SHBG were completed in the General Clinical Research Center Core Laboratory. Intact PTH was measured by ELISA, E2, and estrone by RIA, and SHBG by immunoradiometric assay using kits from Diagnostics Systems Laboratories, Inc. (Webster, TX). The intraassay variability for these studies in the General Clinical Research Center laboratory is less than 10%. The detection limit of the E2 assay is 2 pg/mL.
Statistical analysis
Baseline and clinical characteristics were reported using means and SDs stratified by treatment group. One-way ANOVA was used to test the difference in baseline characteristics between the treatment groups.
For each study participant, we calculated the percent change in biochemical markers of bone turnover and hormones from baseline to 12 weeks and from baseline to 24 weeks. We compared the percent change in each estrogen treatment group to the percent change in the placebo group using one-way ANOVA, obtaining a point estimate of the difference in percent change. The point estimates are reported with the 95% confidence intervals and a P value for the test that the difference in percent change between each treatment group and placebo is zero. The same analyses were repeated for the absolute change from baseline to 12 weeks and from baseline to 24 weeks. We checked the biochemical markers and hormones for normality of distribution and for the impact of outliers. When there were outliers greater than 3 SD from the mean, these outliers were removed, and the analysis was repeated to check for the robustness of the results.
Using one-way ANOVA, we also tested for differences in the response of each biochemical marker to treatment with the three doses of E2. We then estimated the equivalence of the treatment response with 0.25 and 0.5 mg/day compared with 1.0 mg/day calculating point estimates of the mean difference in response (ß weights) and one-sided 95% confidence intervals for each marker at each time point.
We also examined the changes in markers of bone turnover over time. Using the baseline measurement as a standard, we calculated the percent change in biochemical markers of bone turnover from baseline to subsequent time points at 6, 12, 18, and 24 weeks, respectively. These point estimates were reported with 95% confidence intervals for the three treatment groups as well as the placebo group.
We compared side effects in the placebo, 0.25 mg/day, and 0.5 mg/day
groups, respectively, to those in the group receiving standard
treatment (1.0 mg/day). We also compared side effects in each treatment
group to those in the placebo group. One-way ANOVA was used to examine
differences in endometrial thickness and 
2
tests for all other factors. All analyses were performed using SPSS
version 8.0 (SPSS, Inc., Chicago, IL).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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). Of the women randomized, 26 women
were black, 16 were Hispanic, and 64 were white. Baseline demographics
and clinical characteristics of the women who were enrolled in the
study are summarized in Table 1. There
were no significant differences in these baseline factors among the
treatment groups, except that CTx was lower in the 1.0 mg/day group
than in the placebo group. Overall, 34% of women had had a
hysterectomy, 28% had used ERT in the past, 32% had a history of past
fracture, and 21% and 12% were currently taking thiazide diuretics or
thyroid hormone replacement, respectively. The distribution of these
variables across treatment groups was not significantly different. Mean
BMD t-scores were -1.33 for femoral neck, -0.94 for lumbar
spine, and -0.61 for total body. Overall, 5 women with BMD
t-scores less than -2.5, who were eligible for osteoporosis
treatment, were randomly assigned to placebo. Fourteen women who were
randomized to treatment did not complete all study visits (Fig. 1). The
dropout rate was slightly higher in the 1.0 mg/day group than in the
placebo and 0.25 mg/day groups. Compliance, as estimated by pill count,
for treatment and calcium plus vitamin D supplementation was, on the
average, 90% (range, 33–109%) and 84% (range, 28–110%),
respectively, and was similar across treatment groups.
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The biochemical markers of bone turnover were normally
distributed. The percent decrease in NTx and CTx, both markers of bone
resorption, was significantly different from placebo in all treatment
groups at 12 weeks on treatment (Table 2). For total Dpyr, the percent decrease
was not significantly different from the placebo value in any treatment
group until we removed total Dpyr outliers (n = 3), then the
percent decrease was significantly different from placebo in the 0.25
and 1.0 mg/day groups (Table 2). If we log-transformed the total Dpyr
values, the results remained the same. When comparing percent changes
from baseline to 24 weeks of the study, CTx values remained
significantly different from placebo in all
E2-treated groups. We obtained similar results
for the difference in absolute changes in marker values between each
E2 treatment group and the placebo group as we
did in percent change (data not shown).
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compared with baseline values. NTx and Dpyr levels increased toward
baseline values in all E2-treated groups 12 weeks
after termination of E2 and were not
significantly different from baseline at this time point. CTx values,
however, remained significantly lower when comparing 24 weeks to
baseline. Although resorption markers increased slightly over time in
the placebo group, the changes were not statistically significant.
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). The percent decrease from baseline to 12 weeks in
BAP was significantly different from the placebo value in the 1.0 and
0.5 mg E2 groups and for all treatment groups at
24 weeks. PINP decreased in all E2 groups
compared with placebo at 12 and 24 weeks of the study. Changes in OC
compared with placebo were variable. When just considering change over
time, BAP decreased approximately 20% from baseline at week 24. For
PINP, the mean percent decrease from baseline was 28–36% at 24 weeks
of the study (Fig. 2). OC decreased compared with baseline in all
groups at week 24. Hormone levels
Baseline mean levels of sex hormones, SHBG, and PTH were similar
across groups (Table 1). Mean E2 and estrone
levels increased in all treatment groups compared with baseline, but
only in the 1.0 and 0.5 mg groups was the difference in increase
significant compared with placebo (Table 2). The mean serum
E2 level was 28 pg/mL (350% increase compared
with baseline) after 12 weeks of treatment with 0.25 mg/day
E2. Although these changes were not significantly
different from placebo, two thirds of women receiving 0.25 mg/day
E2 responded to treatment with at least a 100%
increase in E2 levels. The percent change in
E2 levels and the percent change in CTx or NTx
were inversely correlated in the 0.25 mg treatment group (r =
-0.543; P = 0.006 and r = -0.418;
P = 0.042, respectively). In the placebo group, 12
women had a decrease and 14 had an increase in serum
E2 levels. Of the women in the placebo group who
demonstrated increased E2 levels, only 1 had more
than a 50% increase. As shown in Table 2, serum estrone levels
demonstrated increases similar to those in serum
E2 with treatment. SHBG increased compared with
baseline in all treatment groups, but not compared with placebo. PTH
did not change in any of the study groups.
Equivalence testing
There were no statistically significant differences in any marker
response to treatment with 0.25 mg/day compared with 1.0 mg/day. When
comparing different doses, however, we were more interested in the
equivalence of the doses rather than in testing for differences between
doses. Table 3 provides the mean absolute
difference between the response of each marker to 0.25 mg/day compared
with 1.0 mg/day E2 treatment with the one-sided
95% confidence intervals. For all markers the difference in response
between doses was quite small. For example, CTx decreased 95.4
µg/mol·L after 12 weeks of treatment 0.25 mg/day and 61.5
µg/mmol·L with 1.0 mg/day. The point estimate of the absolute mean
difference between the response to the two doses is -34 µg/mmol·L
with a one-sided 95% confidence interval of (-
,31) µg/mmol·L
(Table 3). Thus, with 95% confidence we know that the response of CTx
to 0.25 mg/day is at most 31 µg/mmol·L less than it is to 1.0
mg/day. This difference is quite small when one considers the range of
baseline CTx values for women in the study (29–935 µg/mmol·L).
Results for markers were similar when comparing 0.5 to 1.0 mg/day and
when looking at percent differences rather than absolute differences
(data not shown). There were significant differences in the response of
E2 and estrone levels, but not SHBG levels, to
treatment with 1.0 and 0.5 mg/day compared with 0.25 mg/day. The
equivalence data for 0.25 mg/day compared with 1.0 mg/day is provided
in Table 3.
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Breast tenderness and episodes of bleeding were significantly more
frequent in the 1.0 and 0.5 mg groups than in the other treatment
groups (Table 4). Overall, 12 women
reported bleeding or spotting during the study; 11 of these women were
taking either 0.5 or 1.0 mg/day ERT. Most women (8 of 12) who reported
bleeding also had increased endometrial thickness of more than 5 mm at
week 12 of the study. Baseline mean endometrial thickness did not
differ by treatment group (Table 1). Mean endometrial thickness
increased in the 1.0 and 0.5 mg/day groups compared with baseline; the
mean increase in the 1.0 mg group was significantly different compared
with those in the placebo and 0.25 mg groups. Fluid retention was
higher in the 0.5 mg/day group than in the other groups. Other
symptoms, such as bloating and headache, were not significantly
different among the 4 treatment groups.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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In younger postmenopausal women, the use of lower doses of estrogen may not be as effective in preventing bone loss. Lindsay et al. (10), in an early study, examined the effects of four doses of CEE (0.15, 0.3, 0.625, and 1.25 mg/day) on bone density, as measured by single photon absorptiometry of the metacarpal, in women within 5 yr of menopause (mean age, 49 yr). In this analysis, only women who received the two higher doses of CEE had decreased bone resorption and maintenance of bone mineral content over the ensuing 2 yr. Ettinger, in two separate studies of women (average age, 50 and 53 yr) demonstrated a protective effect of 0.3 mg/day CEE plus calcium and 0.5 mg/day E2 plus calcium on spine bone density in early postmenopausal women compared with calcium alone (11, 12). Genant et al. (13) examined the effects of three doses of esterified estrogens (0.3, 0.625, and 1.0 mg/day) on hip and spine bone density in women within 4 yr past menopause (mean age, 51 yr). Women who received the lowest dose of estrogen had increases in hip and spine bone densities compared with those given placebo, although the increase in women receiving 1.25 mg/day was greater than that in the 0.3 and 0.625 mg/day groups. In the above studies of younger postmenopausal women, there was a dose-response effect of ERT on bone; however, many women dropped out of the studies due to inadequate control of menopausal symptoms. In older women far beyond menopause, who rarely experience menopausal symptoms and tend to have lower rates of bone loss than women in the early menopausal years, lower doses of estrogen may provide adequate protection for bone. As we did not find an ineffective dose of estrogen in our population, it may be possible to use even lower doses of estrogen to prevent bone loss in older women.
Based on the equivalence testing, the effect of 0.25 mg/day E2 is quite similar to that of 1.0 mg/day in decreasing markers of bone turnover. At worst, the difference in treatment response with 0.25 mg/day compared with 1.0 mg/day is 31 µg/mmol·L for CTx, 9 nmol/L BCE·mmol/L for NTx, and 1.2 nmol/L·mmol/L for Dpd. Compared with the range of baseline values of each marker in this population, these differences are small and suggest that use of a lower dose of estrogen may be equally beneficial to bone in older women.
Breast tenderness and bleeding, two side effects of ERT that older women frequently cite as the reason for discontinuation, were markedly diminished in the group receiving 0.25 mg/day E2, compared with the two higher doses of E2. In fact, women receiving the lowest dose of E2 reported no more breast tenderness and only slightly more bleeding than reported by women receiving placebo. Endometrial thickness also increased to a greater extent in women taking the two higher doses compared with women in the placebo and 0.25 mg/day E2 groups. If lower dose ERT results in fewer side effects, then the rate of acceptance of ERT as well as long-term compliance will probably be greater than what has been reported with higher doses of ERT. Further, the use of lower dose estrogen may allow lower doses of progesterone to prevent endometrial hyperplasia or cancer. As progesterone is known to have its own side effects, the use of lower doses of progesterone may further enhance compliance (14).
In determining the minimal estrogen dose that is beneficial to bone in older women, one could potentially use E2 levels as a guide. Three reports from the Study of Osteoporotic Fractures indicate that higher endogenous E2 levels are associated with increased bone density, reduced bone loss, and reduced fracture incidence in women over age 65 yr (15, 16, 17). Stone et al. (15) demonstrated that women with E2 levels less than 5 pg/mL experienced 0.8%/yr bone loss at the hip, whereas those with levels greater than 10 pg/mL experienced only 0.1%/yr loss. In the same study high SHBG levels were associated with increased bone loss at the hip. Ettinger et al. (16) demonstrated that women with E2 levels of 10–25 pg/mL had approximately 5% higher hip BMD compared with women with E2 levels less than 5 pg/mL. Finally, Cummings et al. (17) demonstrated decreased hip and spine fracture risk in women with any detectable level of E2, defined as more than 5 pg/mL. Our data support the concept that moderate increases in E2 levels may result in beneficial effects on bone. In our study serum E2 levels increased in all treatment groups compared with baseline. Women who received 0.25 mg/day ERT had one quarter the mean increase in total E2 levels seen in the 1.0 mg group, yet the changes in bone turnover in the lowest dose group were similar to those in the higher dose groups. The mean total E2 level in the 0.25 mg/day group after 12 weeks on treatment was 28 pg/mL, similar to that found to be protective to bone in the above studies. Moreover, the number of side effects reported with the lowest dose of estrogen in our study was equivalent to that in the placebo group. Thus, the additional increase in E2 levels seen with 1.0 mg/day did not result in further decreases in bone resorption, but did result in more side effects.
The results of this study demonstrate that 0.25 mg/day E2 has effects on bone turnover similar to those seen with 1.0 mg/day with few side effects in women whose mean age is 75 yr. Long-term studies are needed to determine whether the effects on bone metabolism seen with 0.25 mg/day E2 will translate into increases in bone density and decreased fracture incidence. However, it is possible that low dose ERT will be an effective and tolerable option for fracture prevention in older women.
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Acknowledgments |
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Footnotes |
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Received April 25, 2000.
Revised August 10, 2000.
Accepted August 20, 2000.
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References |
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), 0.25 mg/day (
), 0.5 mg/day
(
), and 1.0 mg/day (•).