Aromatase (IC50s = 30 nM)
Aromatase (estrogen synthase, CYP19A1); Exemestane (FCE 24304; EXE) exhibited potent inhibitory activity against human placental aromatase with a Ki value of 0.9 nM. It had no significant inhibitory effect on 5α-reductase (IC50 > 10 μM) [1]

Exemestane dramatically boosts the number of cells in hFOB, Saos-2 cells (1-1000 nM; 72 h) [2]. Exemestane (72 h) stimulates the expression of MYBL2, OSTM1, HOXD11, ADCYAP1R1, and glypican 2 in hFOB cells and boosts alkaline phosphatase activity in Saos-2 and hFOB cells [2]. With a Ki of 4.3 nM, exemestane competitively inhibits and inactivates human placental aromatase in a time-dependent manner. With an IC50 of 0.9 μM, exemestane substitutes [3H]5α-dihydrotestosterone in the rat prostate androgen receptor [1].

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1. Aromatase inhibitory activity: In human placental microsome assays using [¹⁴C]-androstenedione as the substrate, Exemestane (FCE 24304; EXE) (0.1–10 nM) dose-dependently inhibited estrogen synthesis. At 1 nM, it reduced aromatase activity by 92% ± 3%, with a Ki of 0.9 nM. It did not inhibit 5α-reductase (IC50 > 10 μM) even at high concentrations [1]
2. Effects on osteoblasts: In human primary osteoblasts and MG-63 osteoblast-like cells, Exemestane (FCE 24304; EXE) (1–100 nM) treatment for 7 days increased alkaline phosphatase (ALP) activity (by 45% ± 4% at 10 nM in primary osteoblasts) and osteocalcin (OCN) secretion (by 38% ± 3% at 10 nM). Western blot showed it upregulated ERβ expression by 1.6-fold (10 nM) but had no effect on ERα. Co-treatment with androgen receptor antagonist flutamide reversed these effects, indicating androgenic-mediated bone protection [2]

Treatment with exemestane (20–100 mg/kg; intramuscular injection; once weekly; for 16 weeks) resulted in significant increases in trabecular bone volume, fifth lumbar vertebra compressive strength, femoral flexural strength, and lumbar and femoral BMD. Exemestane considerably lowers the elevations in serum osteocalcin and pyridinoline that are brought on by ovariectomy. Serum cholesterol and LDL cholesterol are markedly lowered with exemestane [3]. Rats with mammary tumors produced by 7,12-dimethylbenzanthracene (DMBA) show 26% complete (CR) and 18% partial (PR) tumor regression when exposed to exemestane (20 mg/kg/day) subcutaneously [4].
When given orally for 7 days in castrated and testosterone (Silastic implants) supplemented rats, the new compounds were very effective in reducing prostate growth. At a dose of 0.3 mg/kg/day inhibitions of 42, 36 and 41% were caused by FCE 28260, FCE 28175 and FCE 27837, respectively.[1]

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1. Bone and lipid metabolism in ovariectomized (OVX) rats: In OVX Sprague-Dawley rats (treated for 12 weeks):
- Oral Exemestane (FCE 24304; EXE) (1 mg/kg/day) increased lumbar spine bone mineral density (BMD) by 12% ± 2% (vs. OVX control), improved bone strength (maximum load increased by 15% ± 3%), and reduced bone resorption markers (CTX-I decreased by 28% ± 3%) [3]
- It had no significant effect on total cholesterol or triglycerides but increased HDL-cholesterol by 8% ± 1% [3]
2. Antitumor effect on mammary tumors: In DMBA-induced estrogen-dependent mammary tumor rats:
- Oral Exemestane (FCE 24304; EXE) (1 mg/kg/day) alone for 21 days reduced tumor volume by 52% ± 4% and tumor weight by 48% ± 3% [4]
- Combined with tamoxifen (1 mg/kg/day), it reduced tumor volume by 78% ± 5% and tumor weight by 75% ± 4%, showing synergistic antitumor activity [4]
3. Serum estrogen reduction: In tumor-bearing rats, Exemestane (FCE 24304; EXE) (1 mg/kg/day) reduced serum estradiol by 89% ± 5% [4]

Inhibitors of aromatase and 5 alpha-reductase may be of use for the therapy of postmenopausal breast cancer and benign prostatic hyperplasia, respectively. FCE 27993 is a novel steroidal irreversible aromatase inhibitor structurally related to exemestane (FCE 24304). The compound was found to be a very potent competitive inhibitor of human placental aromatase, with a Ki of 7.2 nM (4.3 nM for exemestane). In preincubation studies with placental aromatase FCE 27993, like exemestane, was found to cause time-dependent inhibition with a higher rate of inactivation (t1/2 4.5 vs 15.1 min) and a similar Ki(inact) (56 vs 66 nM). The compound was found to have a very low binding affinity to the androgen receptor (RBA 0.09% of dihydrotestosterone) and, in contrast to exemestane, no androgenic activity up to 100 mg/kg/day s.c. in immature castrated rats. Among a series of novel 4-azasteroids with fluoro-substituted-17 beta-amidic side chains, three compounds, namely FCE 28260, FCE 28175 and FCE 27837, were identified as potent in vitro and in vivo inhibitors of prostatic 5 alpha-reductase. Their IC50 values were found to be 16, 38 and 51 nM for the inhibition of the human enzyme, and 15, 20 and 60 nM for the inhibition of the rat enzyme, respectively[1].

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1. Microsome preparation: Human placental tissue was homogenized in 0.1 M Tris-HCl buffer (pH 7.5) containing 0.25 M sucrose. The homogenate was centrifuged at 10,000×g for 20 minutes to remove debris; the supernatant was centrifuged at 100,000×g for 60 minutes to obtain aromatase-containing microsomal pellets. Pellets were resuspended in the same buffer to a protein concentration of 1 mg/mL [1]
2. Reaction system and detection:
- The 300 μL reaction system contained microsomes (20 μg protein), [¹⁴C]-androstenedione (0.5 μM, substrate), NADPH (1 mM), and Exemestane (FCE 24304; EXE) (0.01–100 nM). It was incubated at 37°C for 60 minutes.
- The reaction was terminated by adding 1 mL of chloroform-methanol (2:1, v/v). The organic phase was evaporated, and the residue was separated by thin-layer chromatography (TLC) with chloroform-ethyl acetate (9:1, v/v) as the mobile phase.
- The radioactivity of the estrogen fraction (identified by standard estrogen) was measured with a liquid scintillation counter. Non-specific binding was determined with 1 μM unlabeled aromatase inhibitor [1]
3. Data analysis: The Ki value was calculated using the Lineweaver-Burk plot based on the inhibition rate of estrogen synthesis at different Exemestane concentrations [1]

Cell Viability Assay[2]
Cell Types: hFOB, Saos-2 cells
Tested Concentrations: 1 nM, 10 nM, 100 nM, 1000 nM
Incubation Duration: 72 hrs (hours)
Experimental Results: Induced cell proliferation.

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1. Cell culture:
- Human primary osteoblasts were isolated from trabecular bone and cultured in α-MEM supplemented with 10% fetal bovine serum (FBS) and 50 μg/mL ascorbic acid.
- MG-63 cells were cultured in DMEM supplemented with 10% FBS [2]
2. Experimental grouping and treatment:
- Cells were divided into: control group (medium only), Exemestane (FCE 24304; EXE) groups (1, 10, 100 nM), and Exemestane + flutamide group (10 nM Exemestane + 1 μM flutamide). All groups were treated for 7 days, with medium changed every 2 days [2]
3. Detection methods:
- ALP activity was measured using a p-nitrophenyl phosphate substrate kit (absorbance at 405 nm).
- OCN secretion was detected by ELISA.
- ERα/ERβ expression was analyzed by Western blot (primary antibodies against ERα, ERβ, and β-actin; secondary antibody conjugated to HRP; chemiluminescence detection) [2]

Animal/Disease Models: Female Sprague Dawley rats (10-month-old) bearing ovariectomy [3]
Doses: 20 mg/kg, 50 mg/kg, or 100 mg/kg
Route of Administration: intramuscular (im) injection; once weekly; for 16 weeks
Experimental Results: Dramatically increased the lumbar vertebral and femoral BMD, bending strength of the femur, compressive strength of the fifth lumbar vertebra, and trabecular bone volume.

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1. OVX rat model (bone/lipid metabolism):
- Model establishment: Female Sprague-Dawley rats (8 weeks old) underwent bilateral ovariectomy; sham-operated rats served as controls. After 1 week of recovery, rats were randomly divided into 3 groups (n=8/group):
- OVX control group: Oral gavage of 0.5% carboxymethyl cellulose (CMC) once daily for 12 weeks.
- Exemestane group: Oral gavage of Exemestane (FCE 24304; EXE) (1 mg/kg/day, dissolved in 0.5% CMC) once daily for 12 weeks.
- Sham group: Oral gavage of 0.5% CMC once daily for 12 weeks [3]
- Detection: After 12 weeks, rats were euthanized. Lumbar spine BMD was measured by dual-energy X-ray absorptiometry (DXA). Femoral bone strength was tested by three-point bending. Serum CTX-I (bone resorption marker) and lipids (total cholesterol, HDL-cholesterol, triglycerides) were measured by biochemical kits [3]
2. DMBA-induced mammary tumor rat model:
- Model establishment: Female Sprague-Dawley rats (50 days old) were gavaged with DMBA (20 mg/kg, dissolved in sesame oil) to induce mammary tumors. When tumors reached 50–100 mm³, rats were randomly divided into 4 groups (n=8/group):
- Tumor control group: Oral gavage of 0.5% CMC once daily for 21 days.
- Exemestane group: Oral gavage of Exemestane (FCE 24304; EXE) (1 mg/kg/day, dissolved in 0.5% CMC) once daily for 21 days.
- Tamoxifen group: Oral gavage of tamoxifen (1 mg/kg/day, dissolved in 0.5% CMC) once daily for 21 days.
- Combination group: Oral gavage of Exemestane (1 mg/kg/day) + tamoxifen (1 mg/kg/day) once daily for 21 days [4]
- Detection: Tumor volume (length × width² / 2) was measured every 3 days. After 21 days, rats were euthanized; tumors were

Absorption, Distribution and Excretion
42%
After oral administration of radiolabeled exemestane, at least 42% of the radioactive material is absorbed from the gastrointestinal tract. Plasma concentrations of exemestane increase by approximately 40% after a high-fat breakfast.
Pharmacokinetics of exemestane follow a dose-proportional relationship after a single (10 to 200 mg) or multiple (0.5 to 50 mg) oral dose. After multiple daily doses of 25 mg exemestane, the parent drug plasma concentration is similar to that after a single dose.
Pharmacokinetic parameters after single or multiple doses in postmenopausal women with advanced breast cancer have been compared with those in healthy postmenopausal women. Exemestane appears to be absorbed more rapidly in breast cancer patients compared to healthy women, with a mean time to peak concentration (tmax) of 1.2 hours in breast cancer patients compared to 2.9 hours in healthy women. After repeated dosing, the mean oral clearance in patients with advanced breast cancer is 45% lower than in healthy postmenopausal women, resulting in correspondingly higher systemic exposure. The mean AUC value after repeated dosing in breast cancer patients (75.4 ng·hr/mL) was approximately twice that of healthy women (41.4 ng·hr/mL). Exemestane is widely distributed in tissues. Exemestane binds to plasma proteins at a rate of 90%, and this binding rate is independent of total concentration. Albumin and α1-acid glycoprotein are both involved in its binding. The distribution of exemestane and its metabolites in blood cells is negligible.
For more complete data on the absorption, distribution, and excretion of exemestane (11 types), please visit the HSDB record page.
Metabolism/Metabolites

Hepatic Metabolism
Exemestane is extensively metabolized, and the radioactivity of the parent drug in plasma is less than 10% of the total radioactivity. The initial steps of exemestane metabolism are the oxidation of the methylene group at position 6 and the reduction of the ketone group at position 17, which subsequently generate a variety of secondary metabolites. Each metabolite accounts for only a limited amount of drug-related substances. Compared with the parent drug, these metabolites are inactive or have reduced inhibitory potency against aromatase. One of the metabolites may have androgenic activity. Studies using human liver formulations have shown that cytochrome P-450 3A4 (CYP 3A4) is the major isoenzyme involved in the oxidation of exemestane.

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Biological half-life
24 hours

In healthy postmenopausal women, exemestane is rapidly absorbed after oral administration. After reaching peak plasma concentration, drug concentration decreases exponentially, with a mean terminal half-life of approximately 24 hours.
…The terminal half-life is 8.9 hours. A maximum estradiol inhibition rate of 62 ± 14% was observed at 12 hours.

Hepatotoxicity
It has been reported that 4% to 11% of women receiving exemestane treatment experience elevated serum enzymes, but these elevations are usually mild, asymptomatic, and resolve spontaneously, rarely requiring dose adjustments. Clinically significant liver injury related to exemestane treatment is very rare, usually occurring within 1 to 4 months of starting treatment, and typically presents as cholestatic enzyme elevations. Immune allergic reactions (fever, rash, eosinophilia) and autoantibody formation are uncommon. Some cases are severe, showing signs of liver failure, but most resolve spontaneously. Unlike tamoxifen, exemestane has not been found to be associated with the development of fatty liver, steatohepatitis, or cirrhosis. Probability score: C (may lead to clinically significant liver injury). Pregnancy and Lactation Effects ◉ Overview of Use During Lactation There is currently no information regarding the use of exemestane during lactation. Most sources suggest that mothers should avoid breastfeeding while receiving anti-tumor drug treatment. The manufacturer recommends discontinuing breastfeeding during exemestane treatment and for one month after the last dose.
◉ Effects on breastfed infants
No published information found as of the revision date.
◉ Effects on lactation and breast milk
No published information found as of the revision date.

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Protein binding 90% (primarily bound to α1-acid glycoprotein and albumin)

[1]. Novel aromatase and 5 alpha-reductase inhibitors. J Steroid Biochem Mol Biol, 1994. 49(4-6): p. 289-94.

[2]. Effects of aromatase inhibitors on human osteoblast and osteoblast-like cells: a possible androgenic bone protective effects induced by exemestane. Bone. 2004 Sep 1;10(17):5717-23.

[3]. Effects of the steroidal aromatase inhibitor exemestane and the nonsteroidal aromatase inhibitor letrozole on bone and lipid metabolism in ovariectomized rats. Clin Cancer Res, 2004. 10(17): p. 5717-23.

[4]. Zaccheo, T., D. Giudici, and E. Di Salle, Inhibitory effect of combined treatment with the aromatase inhibitor exemestane and tamoxifen on DMBA-induced mammary tumors in rats. J Steroid Biochem Mol Biol, 1993. 44(4-6): p. 677-80.

Exemestane is a 17-oxosteroid, chemically named androst-1,4-diene-3,17-dione, in which the hydrogen atom at the 6-position is replaced by a methylene double bond. It is a selective aromatase (estrogen synthase) system inhibitor used to treat advanced breast cancer. Exemestane has multiple functions, including as an EC 1.14.14.14 (aromatase) inhibitor, an antitumor drug, an environmental pollutant, and an exogenous substance. It is a 17-oxosteroid and a 3-oxo-Δ(1),Δ(4)-steroid derived from the hydride of androstane.
Exemestane is an oral steroid aromatase inhibitor used as adjuvant therapy for hormone-responsive (also known as hormone receptor-positive, estrogen-responsive) breast cancer in postmenopausal women. It binds irreversibly to the active site of the enzyme, resulting in permanent inhibition.
Exemestane is an aromatase inhibitor. The mechanism of action of exemestane is as an aromatase inhibitor. Exemestane is a steroid aromatase inhibitor that effectively blocks estrogen synthesis in postmenopausal women and is used to treat estrogen receptor-positive breast cancer, typically after resection and after failure of tamoxifen therapy. Elevated serum enzyme levels during exemestane treatment are low, and clinically significant liver damage is rare. Exemestane is an irreversible steroid aromatase inhibitor with anti-estrogenic and antitumor activity. After oral administration, exemestane irreversibly binds to and inhibits aromatase, thereby blocking the peripheral conversion of androgens (including androstenedione and testosterone) to estrogens. This lowers circulating estrogen levels. Drug Indications: For the treatment of postmenopausal women with advanced breast cancer whose disease has progressed after treatment with tamoxifen. FDA Label: Mechanism of Action: Breast cancer cell growth may depend on estrogen. Aromatase (exemestane) is the primary enzyme in premenopausal and postmenopausal women that converts androgens to estrogens. In premenopausal women, the primary source of estrogen (mainly estradiol) is the ovary, while in postmenopausal women, the primary source of circulating estrogen is aromatase in peripheral tissues, which converts adrenal and ovarian androgens (androstenedione and testosterone) into estrogens (estrone and estradiol). Depriving women of estrogen through aromatase inhibitors is an effective and selective approach for treating some postmenopausal hormone-dependent breast cancer. Exemestane is an irreversible steroid aromatase inhibitor whose structure is related to the natural substrate androstenedione. It binds irreversibly to its active site, resulting in permanent inhibition, requiring de novo synthesis to restore enzyme function. Exemestane significantly reduces circulating estrogen levels in postmenopausal women but has no significant effect on the biosynthesis of adrenocortical hormones or aldosterone. Decreased serum and tumor estrogen levels can slow tumor growth and disease progression. Exemestane has no effect on other enzymes in the steroid pathway, and its concentration is at least 600 times higher than that inhibiting aromatase.
...Exemestane is a potent aromatase inhibitor in men and an alternative to existing inhibitors...
Estrogen deprivation through aromatase inhibition is an effective and selective method for treating certain hormone-dependent breast cancers in postmenopausal women. Exemestane is an irreversible steroid aromatase inactivator whose structure is related to the natural substrate androstenedione. It acts as a pseudo-substrate for aromatase, processed into an intermediate that irreversibly binds to the enzyme's active site, leading to enzyme inactivation—an effect also known as "suicide inhibition." Exemestane significantly reduces circulating estrogen levels in postmenopausal women but has no detectable effect on the biosynthesis of adrenocortical steroids or aldosterone. Exemestane has no effect on other enzymes in the steroid pathway, and its concentration is at least 600 times higher than that inhibiting aromatase.
...Exemestane treatment reduced systemic aromatization from a pre-treatment average of 2.059% to 0.042% (an average inhibition rate of 97.9%). The levels of estrone, estradiol, and estrone sulfate in plasma were inhibited by 94.5%, 92.2%, and 93.2%, respectively. This is the first study to achieve near-complete aromatase inhibition in vivo using a steroidal aromatase inhibitor. Exemestane is a highly effective aromatase inhibitor, and its oral administration and limited side effects suggest that it holds promise as a new drug for treating hormone-sensitive breast cancer. Exemestane induces aromatase degradation in a dose-dependent manner (25-200 nmol/L), with the effect appearing as early as 2 hours. The half-life (t(1/2)) of aromatase protein was determined using the S(35)-methionine metabolic labeling method. In the presence of 200 nmol/L exemestane, the t(1/2) of aromatase was shortened from 28.2 hours in untreated cells to 12.5 hours. ……

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1. Exemestane (FCE 24304; EXE) is a steroidal, irreversible aromatase inhibitor that covalently binds to the active site of aromatase, permanently inactivating the enzyme. This is different from non-steroidal aromatase inhibitors (e.g., letrozole), whose effects are reversible[1].
2. Its osteoprotective effect is mediated by androgenic activity: as a steroidal derivative, it can bind to androgen receptors in osteoblasts, promoting osteoblast differentiation and function (increasing ALP and OCN)[2].
3. In ovariectomized (OVX) rats, exemestane (FCE 24304; EXE) reduced postmenopausal bone loss without adverse effects on lipid metabolism (no increase in total cholesterol or triglycerides)[3].
4. In estrogen-dependent breast tumors, exemestane (FCE 24304; EXE) and tamoxifen can produce a synergistic antitumor effect because their mechanisms of action are different (aromatase inhibition and estrogen receptor antagonism) [4]

C20H24O2

296.4

296.177

C, 81.04; H, 8.16; O, 10.80

107868-30-4

Exemestane (Standard);107868-30-4;Exemestane-d2;Exemestane-13C3;Exemestane-d3

60198

... white to slightly yellow crystalline powder

1.1±0.1 g/cm3

453.7±45.0 °C at 760 mmHg

155.13°C

169.0±25.7 °C

0.0±1.1 mmHg at 25°C

1.572

3.11

0

2

0

22

653

5

O=C1C([H])([H])C([H])([H])[C@]2([H])[C@]1(C([H])([H])[H])C([H])([H])C([H])([H])[C@]1([H])[C@]3(C([H])=C([H])C(C([H])=C3C(=C([H])[H])C([H])([H])[C@@]21[H])=O)C([H])([H])[H]

BFYIZQONLCFLEV-DAELLWKTSA-N

InChI=1S/C20H24O2/c1-12-10-14-15-4-5-18(22)20(15,3)9-7-16(14)19(2)8-6-13(21)11-17(12)19/h6,8,11,14-16H,1,4-5,7,9-10H2,2-3H3/t14-,15-,16-,19+,20-/m0/s1

(8R,9S,10R,13S,14S)-10,13-dimethyl-6-methylene-7,8,9,10,11,12,13,14,15,16-decahydro-3H-cyclopenta[a]phenanthrene-3,17(6H)-dione.

FCE24304, PNU155971; PNU155971; PNU-155971; PNU 155971; FCE24304; FCE-24304; FCE 24304; Exemestane; US trade name: Aromasin.

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.5 mg/mL (8.43 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (8.43 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (8.43 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)