ERα (IC50 = 26 nM); ERβ (IC50 = 99 nM)[1]
Bazedoxifene targets estrogen receptors ERα and ERβ. It exhibits agonist activity in bone, the cardiovascular system, and the CNS, while exhibiting antagonist activity in breast and endometrial tissues. It binds to ERα with IC50 values ranging from 0.6 to 26 nM and to ERβ with IC50 values ranging from 85 to 99 nM. It also inhibits IL-6/GP130 interactions.

The GP130 D1 domain is bound by the small molecule GP130 inhibitor benzedoxifene [1]. When IL-6 and IL-11 trigger STAT3 phosphorylation in the GP130/STAT3 pathway signaling, benzedoxifene blocks this process [1]. In human pancreatic cancer cells, zedoxifene (10 μM–20 μM; 2 hours) suppresses the phosphorylation of STAT3 caused by cytokines [2]. Human pancreatic cancer cells undergo apoptosis when exposed to bezoxifene (5–20 μM) over night [2]. STAT3 nuclear translocation caused by IL-6 is inhibited by benzedoxifene [2]. By blocking GP130, benzedoxifene stops pancreatic cancer cells from migrating [2].

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In vitro, bazedoxifene binds to both ERα (IC50 values of 0.6-26 nM) and ERβ (IC50 values of 85-99 nM). It exhibits agonist activity in bone and cardiovascular cells while exhibiting antagonist activity in breast and endometrial cells. It also acts as an inhibitor of IL-6/GP130 protein-protein interactions.

In an in vivo mouse model, benzedoxifene (5 mg/kg; ir; once daily for 18 days) suppresses the formation of Capan-1 tumors [2].

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In vivo, bazedoxifene is used for the prevention and treatment of postmenopausal osteoporosis. It is orally active and penetrates the blood-brain barrier. It has been studied for its potential benefits in pancreatic cancer via IL-6/GP130 inhibition. It effectively increases bone mineral density and reduces fracture risk in postmenopausal women.

Ligand binding[1]
Interaction of bazedoxifene acetate (BZA) with human ERα and ERβ was assessed with a solid phase competitive radioligand binding assay using [3H]-17β- estradiol as previously described.

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STAT3 DNA binding assay[2]
BxPC-3 cells were seeded in a 10-cm plate and treated with Bazedoxifene (5–10 μmol/L) or DMSO for 24 hours. The Nuclear Extract Kit was used to prepare cell nuclear extracts following the manufacturer's protocol. Nuclear extracts were analyzed for STAT3 DNA–binding activity using a STAT3 DNA binding ELISA kit (Active Motif) with an ELISA-based method. Absorbance was read at 450 nm.

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STATs phosphorylation induced by cytokines or growth factors[2]
PANC-1, AsPC-1, and HPAF-II pancreatic cancer cells were seeded in 10-cm plates and allowed to adhere overnight. The following night, the cells were serum starved. The cells were then left untreated or were treated with Bazedoxifene (5–20 μmol/L) or DMSO. After 2 hours, the untreated and Bazedoxifene-treated cells were stimulated by IL6 (50 ng/mL), IL11 (50 ng/mL), OSM (50 ng/mL), or INFγ (50 ng/mL) for 30 minutes. The cells were harvested and analyzed by Western blot analysis for p-STAT3Y705 or p-STAT1Y701.

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In vitro receptor binding assays for bazedoxifene typically involve competitive radioligand binding using membrane preparations expressing human ERα or ERβ. Membranes are incubated with radiolabeled estradiol and varying concentrations of bazedoxifene. IC50 values are calculated from displacement curves. Functional activity (agonist vs. antagonist) is assessed by measuring reporter gene expression or downstream signaling.

Western Blot Analysis[2]
Cell Types: AsPC-1 Cell
Tested Concentrations: 10 μM, 20 μM
Incubation Duration: 2 hrs (hours)
Experimental Results: Inhibition of STAT3 phosphorylation induced by IL-6, IL-11 or OSM (50 ng/mL).

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Apoptosis analysis [2]
Cell Types: Capan-1 cells, BxPC-3 cells, HPAF-II cells, HPAC cells
Tested Concentrations: 10 μM, 20 μM (Capan-1); 5μM, 10μM (BxPC-3); 10 μM, 20 μM (HPAF-II); 10 μM, 15 μM (HPAC)
Incubation Duration: Overnight
Experimental Results: Induction of apoptosis.

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Cell-based assays for bazedoxifene involve culturing estrogen receptor-positive cell lines (e.g., MCF-7 breast cancer cells) in appropriate media. Cells are treated with bazedoxifene at concentrations ranging from 0.1 nM to 10 µM for 24-72 hours. Cell proliferation is assessed by MTT or BrdU incorporation. Estrogen-responsive gene expression is measured by qPCR or reporter assays. IL-6/GP130 inhibition is evaluated in relevant cell lines.

Animal/Disease Models: 6weeks old female athymic nude mice [2]
Doses: 5 mg/kg
Route of Administration: po (oral gavage), daily, for 18 days.
Experimental Results: Inhibited the growth of pancreatic cancer xenograft tumors and induced tumor cell apoptosis.

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Vasomotor instability (hot flush)[1]
Ovariectomized female (60 d) rats were obtained after surgery. The surgeries were performed minimally 7 d before initiation of any experiment. Vehicle and ethinyl estradiol (0.3 mg/kg) were included in each replicate. Bazedoxifene was administered orally in a saline, Tween-80, methylcellulose vehicle. A detailed description of methodology for evaluating vasomotor instability in rats has been published (21). Briefly, compound treatment (17β-estradiol, ethinyl estradiol, or bazedoxifene) is initiated, and on the third day of treatment each animal receives a morphine pellet sc. This is followed by two more pellets on the fifth day of treatment. On the eighth day, a thermistor is taped to the animal’s tail to measure tail skin temperature for 15 min (to obtain baseline temperature) followed by a sc injection of naloxone (1 mg/kg). Tail skin temperature readings continue for 1 h after naloxone injection.

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All animal studies were conducted in accordance with the principles and standard procedures approved by IACUC of the Research Institute at Nationwide Children's Hospital. Capan-1 (3 × 106) and HPAF-II (3 × 106) cells in Matrigel were injected subcutaneously into the both side of flank area of 6-week-old female athymic nude mice which were purchased from Harlan. After Capan-1 tumor development, which was 1 week after initial implantation, mice were divided into two treatment groups consisting of four mice (tumors: n = 8): DMSO vehicle control and gavage injection of Bazedoxifene (5 mg/kg/d). Mice bearing HPAF-II tumor were irrigated with Bazedoxifene(5 mg/kg/d) and/or injected via abdomen with paclitaxel (15 mg/kg, 2/w). Tumor growth was determined by measured the length (L) and width (W) of the tumor every other day with a caliper, and tumor volume was calculated on the basis of the following formula: volume = 0.52 × LW2. After 21 days of treatment, tumors were harvested, snap-frozen in dry ice, and stored at −80°C. Tumors tissue homogenates were lysed and separated by SDS-PAGE to examine the expression of STAT3 phosphorylation, P-ERK1/2, P-AKT (Ser473), and cleaved caspase-3.[2]

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In vivo animal experiments for bazedoxifene typically involve administration to ovariectomized rat models of osteoporosis via oral gavage. Bone mineral density is measured by DEXA. Bone strength is assessed by mechanical testing. Uterine weight is measured to evaluate estrogenic effects in the endometrium. Anti-tumor efficacy is evaluated in xenograft mouse models of pancreatic cancer.

Absorption, Distribution and Excretion
Bazedoxifene is rapidly absorbed, reaching peak concentration in approximately 2 hours. Plasma concentrations increase linearly with single doses from 0.5 mg to 120 mg and multiple daily doses from 1 mg to 80 mg. The absolute bioavailability of Bazedoxifene is approximately 6%. The primary route of excretion for radiolabeled Bazedoxifene is feces, with less than 1% excreted in urine. The volume of distribution after intravenous administration of 3 mg Bazedoxifene is 14.7 ± 3.9 L/kg. The apparent oral clearance of Bazedoxifene is approximately 4 to 5 L/h/kg. Metabolism/Metabolites Glucuronization is the primary metabolic pathway. After oral administration, Bazedoxifene is metabolized by UDP-glucuronyltransferases (UGTs) to Bazedoxifene-4'-glucuronide (M4) and Bazedoxifene-5-glucuronide (M5). Almost no cytochrome P450-mediated metabolism was observed. The concentration of this glucuronide in plasma was approximately 10 times that of the unmetabolized active substance.

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Biological half-life
Approximately 30 hours.

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Bazedoxifene (molecular weight 470.60, formula C30H34N2O3) is orally active and penetrates the blood-brain barrier. It is metabolized in the liver and excreted via bile and urine. It has a long half-life supporting once-daily dosing. Detailed pharmacokinetic parameters are available in clinical pharmacology literature. It is available as a combination product with conjugated estrogens for osteoporosis treatment.

Protein Binding
98-99%.

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Bazedoxifene is generally well-tolerated. Common side effects include hot flashes, leg cramps, and gastrointestinal effects. It has a favorable endometrial and breast safety profile compared to other SERMs due to its antagonist activity in these tissues. Comprehensive toxicological evaluation has been conducted in preclinical and clinical studies. It is approved for postmenopausal osteoporosis.

[1]. Bazedoxifene acetate: a selective estrogen receptor modulator with improved selectivity. Endocrinology. 2005 Sep;146(9):3999-4008.

[2]. Bazedoxifene as a Novel GP130 Inhibitor for Pancreatic Cancer Therapy. Mol Cancer Ther. 2016 Nov; 15(11): 2609–2619.

Bazedoxifene is a phenyl indole compound. It is a third-generation selective estrogen receptor modulator (SERM) developed by Pfizer after its acquisition of Wyeth Pharmaceuticals. In late 2013, Pfizer received approval to include Bazedoxifene as a component of the combination drug DUAVEE for the prevention (not treatment) of postmenopausal osteoporosis. The drug is approved as monotherapy in the EU (marketed in Italy and Spain) and Japan. In 2013, a combination formulation containing conjugated estrogen and Bazedoxifene was approved by the US Food and Drug Administration (FDA) for the treatment of moderate to severe vasomotor symptoms associated with menopause and for the prevention of postmenopausal osteoporosis in women. Bazedoxifene is an estrogen agonist/antagonist. Its mechanism of action is as a selective estrogen receptor modulator. Bazedoxifene is an indole derivative, belonging to the third-generation selective estrogen receptor modulator (SERM), and has potential antitumor activity. After administration, pazedoxifen specifically binds to estrogen receptors in sensitive tissues, including the liver, bone, breast, and endometrium. The resulting ligand-receptor complex translocates to the cell nucleus and, depending on the tissue type, promotes or inhibits the transcription of estrogen-regulated genes. In uterine and breast tissues, pazedoxifen acts as an estrogen antagonist, blocking the proliferative effects of estrogen binding to ER-positive cells in these tissues. In lipid metabolism, pazedoxifen acts as an estrogen agonist, thereby lowering total cholesterol and low-density lipoprotein cholesterol (LDL-C) levels. In bone, it reduces bone resorption and bone turnover and increases bone mineral density.

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Drug Indications
Indications for use in women with a uterus for the following conditions, either alone or in combination with conjugated estrogen: - Treatment of moderate to severe vasomotor symptoms associated with menopause - Prevention of postmenopausal osteoporosis
FDA Label
Conpiride is indicated for the treatment of osteoporosis in postmenopausal women at increased risk of fracture. It has been shown to significantly reduce the incidence of vertebral fractures; however, its efficacy in treating hip fractures has not been established. When choosing buprofen or other therapies (including estrogen) for specific postmenopausal women, menopausal symptoms, effects on uterine and breast tissue, and cardiovascular risks and benefits should be considered.

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Mechanism of Action
Bazedoxifene belongs to a class of compounds called selective estrogen receptor modulators (SERMs). Depending on cell and tissue type and target genes, Bazedoxifene can act as either an estrogen receptor agonist or an antagonist. Bazedoxifene reduces bone resorption and lowers biochemical markers of bone turnover to premenopausal levels. These effects on bone remodeling can increase bone mineral density (BMD), thereby reducing the risk of fracture. In uterine and breast tissue, Bazedoxifene primarily functions as an estrogen receptor antagonist.

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Bazedoxifene (CAS#: 198481-32-2) is a non-steroidal, orally active SERM. It targets ERα and ERβ. It is used for postmenopausal osteoporosis. It also inhibits IL-6/GP130 interactions and is studied for pancreatic cancer. Molecular weight: 470.60, formula: C30H34N2O3.

C30H34N2O3

470.602568149567

470.256

C, 76.57; H, 7.28; N, 5.95; O, 10.20

198481-32-2

Bazedoxifene acetate;198481-33-3;Bazedoxifene hydrochloride;198480-56-7;Bazedoxifene-d4;1133695-49-4;Bazedoxifene-d4 acetate;1795027-71-2

154257

White to yellow solid powder

1.2±0.1 g/cm3

694.4±55.0 °C at 760 mmHg

373.8±31.5 °C

0.0±2.3 mmHg at 25°C

1.622

6.59

2

4

7

35

623

0

O(C1C=CC(=CC=1)CN1C2C=CC(=CC=2C(C)=C1C1C=CC(=CC=1)O)O)CCN1CCCCCC1

UCJGJABZCDBEDK-UHFFFAOYSA-N

InChI=1S/C30H34N2O3/c1-22-28-20-26(34)12-15-29(28)32(30(22)24-8-10-25(33)11-9-24)21-23-6-13-27(14-7-23)35-19-18-31-16-4-2-3-5-17-31/h6-15,20,33-34H,2-5,16-19,21H2,1H3

1-(4-(2-(azepan-1-yl)ethoxy)benzyl)-2-(4-hydroxyphenyl)-3-methyl-1H-indol-5-ol

WAY-140424; WAY140424; WAY 140424; TSE 424; Bazedoxifene [INN]; 1H-Indol-5-ol, 1-[[4-[2-(hexahydro-1H-azepin-1-yl)ethoxy]phenyl]methyl]-2-(4-hydroxyphenyl)-3-methyl-; Bazedoxifene free base; Q16TT9C5BK; 1-(4-(2-(azepan-1-yl)ethoxy)benzyl)-2-(4-hydroxyphenyl)-3-methyl-1H-indol-5-ol; TSE424; TSE-424 Viviant.

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)

DMSO : ~100 mg/mL (~212.49 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.31 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 (5.31 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 (5.31 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.)