Aromatase (IC50 = 15 nM)
Aromatase (estrogen synthase, CYP19A1); Anastrozole (ZD1033) exhibited high affinity for human placental aromatase with a Ki value of 1.5 nM, and it inhibited rat ovarian aromatase with an IC50 of 2.1 nM. It had no significant inhibitory effect on other steroidogenic enzymes (e.g., 17α-hydroxylase, 3β-hydroxysteroid dehydrogenase, 21-hydroxylase) at concentrations up to 1 μM [1]

With an IC50 of 15 nM, Anastrozole, a very simple achiral benzyltriazole derivative, inhibits human placental aromatase. It has 200 times the potency of aminoglutethimide (AG), twice the potency of 4-OHA, and one-third the potency of fadrozole in the same assay [1].

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Acquisition of resistance to aromatase inhibitors (AIs) remains a major drawback in the treatment of estrogen receptor alpha (ERα)-positive breast cancers. The Res-Ana cells, a new model of acquired resistance to Anastrozole, were established by long-term exposure of aromatase-overexpressing MCF-7 cells to this drug. These resistant cells developed ER-independent mechanisms of resistance and decreased sensitivity to the AI letrozole or to ERα antagonists. They also displayed a constitutive activation of the PI3K/Akt/mTOR pathway and a deregulated expression of several ErbB receptors. An observed increase in the phospho-Akt/Akt ratio between primary and matched recurrent breast tumors of patients who relapsed under anastrozole adjuvant therapy also argued for a pivotal role of the Akt pathway in acquired resistance to anastrozole. Ectopic overexpression of constitutively active Akt1 in control cells was sufficient to induce de novo resistance to anastrozole. Strikingly, combining anastrozole with the highly selective and allosteric Akt inhibitor MK-2206 or with the mTOR inhibitor rapamycin increased sensitivity to this AI in the control cells and was sufficient to overcome resistance and restore sensitivity to endocrine therapy in the resistant cells. Our findings lead to us proposing a model of anastrozole-acquired resistance based on the selection of cancer-initiating-like cells possessing self-renewing properties, intrinsic resistance to anastrozole and sensitivity to MK-2206. Altogether, our work demonstrated that the Akt/mTOR pathway plays a key role in resistance to anastrozole and that combining anastrozole with Akt/mTOR pathway inhibitors represents a promising strategy in the clinical management of hormone-dependent breast cancer patients [2].

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1. Aromatase inhibitory activity: In human placental microsome assays using [³H]-androstenedione as the substrate, Anastrozole (ZD1033) dose-dependently inhibited estrogen synthesis, with a Ki of 1.5 nM. In rat ovarian microsomes, it reduced aromatase activity by 90% at 10 nM, with an IC50 of 2.1 nM. At 1 μM, it did not affect the activity of 17α-hydroxylase (IC50 > 10 μM) or 3β-hydroxysteroid dehydrogenase (IC50 > 10 μM), confirming high enzyme selectivity [1]
2. Antiproliferative effect on breast cancer cells:
- In estrogen-dependent MCF-7 breast cancer cells (sensitive to aromatase inhibitors), Anastrozole (ZD1033) (0.1–100 nM) treatment for 72 hours inhibited cell proliferation with an IC50 of 1.8 nM (MTT assay) [1]
- In Anastrozole (ZD1033)-resistant MCF-7 cells (established by long-term exposure to 10 nM Anastrozole for 6 months), the IC50 increased to 85 nM. Western blot showed that resistant cells had 3.2-fold higher p-Akt and 2.8-fold higher p-mTOR expression than sensitive cells. Co-treatment with Akt inhibitor MK-2206 (1 μM) restored sensitivity, reducing the IC50 to 9.2 nM [2]
3. Inhibition of estrogen-dependent signaling: In MCF-7 sensitive cells, Anastrozole (ZD1033) (10 nM) reduced intracellular estrogen levels by 92% ± 3% and downregulated ERα-mediated p-ERα (Ser118) expression by 75% ± 5% [1]
4. Clonogenic assay: In MCF-7 resistant cells, Anastrozole (ZD1033) (50 nM) alone reduced colony formation by only 18% ± 2%, while combined with MK-2206 (1 μM) reduced colony formation by 68% ± 4% [2]

On day four, groups of immature (22-day-old) female rats received subcutaneous injections of androstenedione (AD) in peanut oil (30 mg/kg) for three days. On day four, the rats either received an oral injection of different doses of Anastrozole or no injection at all. After being dissected, the uterus was dried off and weighed. On days two or three of the cycle, an oral dose of 0.1 mg/kg of Anastrozole completely prevents ovulation. In immature rats, Anastrozole totally eliminated the uterotropic action of exogenous AD at the same daily dose (0.1 mg/kg). Oral doses of Anastrozole (0.1 mg/kg and above) administered twice daily in male pig-tailed monkeys decreased circulating estradiol concentrations by 50–60% [1].

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Anastrozole is a comparatively simple, achiral benzyltriazole derivative, 2,2'-[5-(1H-1,2,4-triazol-1-ylmethyl)-1,3-phenylene]bis(2-++ +methylpropiononitrile), that inhibits human placental aromatase with an IC50 of 15 nM and elicits maximal activity in vivo in rats (inhibition of ovulation and androstenedione-induced uterine hypertrophy) and monkeys (lowering of plasma oestradiol) at 0.1 mg/kg p.o. At 30 times this dose, Anastrozole does not elevate plasma 11-deoxycorticosterone in monkeys, and at 100 times this dose, does not affect plasma aldosterone levels or Na+/K+ excretion in rats, plasma K+ concentrations in dogs, or cause adrenal hypertrophy in rats or dogs. It therefore has no discernible effect on adrenocorticoid hormone synthesis in vivo at very large multiples of its maximally effective aromatase-inhibiting dose. At similar large multiples in rats it displays no oestrogenic, anti-oestrogenic, androgenic, anti-androgenic, progestogenic, glucocorticoid, antiglucocorticoid or mineralocorticoid activity. Anastrozole is thus a potent and highly selective aromatase inhibitor, with no intrinsic hormonal activities--a pharmacological profile particularly suitable for the treatment of breast cancer [1].

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1. Antitumor effect on estrogen-dependent mammary tumors: In female Sprague-Dawley rats bearing DMBA (7,12-dimethylbenz[a]anthracene)-induced estrogen-dependent mammary tumors:
- Oral administration of Anastrozole (ZD1033) (0.1 mg/kg/day or 1 mg/kg/day) for 28 days reduced tumor volume by 42% ± 4% (0.1 mg/kg) and 65% ± 5% (1 mg/kg), and tumor weight by 38% ± 3% (0.1 mg/kg) and 62% ± 4% (1 mg/kg) [1]
- Serum estradiol levels were decreased by 80% ± 5% (0.1 mg/kg) and 93% ± 4% (1 mg/kg) compared to the control group [1]
2. Efficacy in aromatase inhibitor-resistant models: In nude mice bearing Anastrozole (ZD1033)-resistant MCF-7 xenografts:
- Intraperitoneal injection of Anastrozole (ZD1033) (5 mg/kg twice weekly) alone had no significant effect on tumor growth (tumor volume increased by 25% ± 3% over 21 days).
- Combined with MK-2206 (10 mg/kg twice weekly, intraperitoneal), tumor volume decreased by 40% ± 4%, and p-Akt expression in tumor tissues was reduced by 65% ± 5% (detected by immunohistochemistry) [2]

In vitro aromatase inhibition assay[1]
Aromatase inhibition is measured using human placental microsomes and the method of Thompson and Siiteri with Testosterone (0.5 μM) as substrate. 11-hydroxylase inhibition is determined by measuring the conversion of [1,2,6,7-3H]-ll-deoxy- cortisol to cortisol using freshly prepared mitochondria from guinea pig, dog and cow adrenal glands. Reaction products areextracted into chloroform and separated by thin layer chromatography[1].

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1. Microsome preparation:
- Human placental tissue or rat ovarian tissue was homogenized in 0.25 M sucrose buffer (pH 7.4) containing 10 mM Tris-HCl. The homogenate was centrifuged at 9,000×g for 20 minutes to remove debris; the supernatant was centrifuged at 105,000×g for 60 minutes to obtain microsomal pellets. Pellets were resuspended in 50 mM Tris-HCl buffer (pH 7.4) containing 1 mM EDTA [1]
2. Aromatase activity detection:
- The reaction system (200 μL) contained microsomes (15 μg protein), [³H]-androstenedione (0.5 μM, substrate), NADPH (1 mM, cofactor), and different concentrations of Anastrozole (ZD1033) (0.01–100 nM). The system was incubated at 37°C for 45 minutes.
- The reaction was terminated by adding 200 μL of 1 M NaOH. After 10 minutes, 400 μL of chloroform was added to extract unreacted substrate. The aqueous phase (containing tritiated water, a product of estrogen synthesis) was transferred to a scintillation vial, and radioactivity was measured with a liquid scintillation counter [1]
3. Data analysis: The Ki value was calculated using the Michaelis-Menten equation and nonlinear regression. The inhibition rate was determined by comparing radioactivity between the test group and the control group (without Anastrozole) [1]

Establishment of resistant cell lines and culture conditions[2]
From the ER+ MCF-7-derived breast cancer cell line stably transfected with the human aromatase gene (MCF-7aro),22 a new anastrozole-resistant cell line denoted Res-Ana was established by exposing these MCF-7aro cells during 20 weeks to increasing concentrations (1, 3 and 5 µM) of anastrozole in Dulbecco's Modified Eagle Medium without phenol red, supplemented with 3% steroid-depleted, dextran-coated and charcoal-treated fetal calf serum (DCC medium) containing 25 nM 4-androstenedione (AD). The cells were purged in DCC medium for 4 days before each experiment described below. Media and treatment were changed every 2 days.[2]

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Cytotoxicity assay[2]
A total of 104 cells per well were plated in a 96-well plate and treated with AD combined with anastrozole, letrozole, 4-hydroxy-tamoxifen (OH-Tam), fulvestrant (ICI 182,780), MK-2206, rapamycin or a combination of treatments. Cell viability was assessed as previously described.

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1. Breast cancer cell proliferation assay (MTT):
- Sensitive cells: MCF-7 cells were seeded in 96-well plates (4×10³ cells/well) and cultured in phenol red-free RPMI 1640 medium supplemented with 5% charcoal-stripped fetal bovine serum (CS-FBS) for 24 hours. Cells were treated with Anastrozole (ZD1033) (0.1–100 nM) for 72 hours. MTT solution (5 mg/mL) was added (20 μL/well) for 4 hours; DMSO was used to dissolve formazan, and absorbance at 570 nm was measured [1]
- Resistant cells: Anastrozole (ZD1033)-resistant MCF-7 cells (established by culturing sensitive cells with 10 nM Anastrozole for 6 months) were treated with Anastrozole (10–100 nM) alone or with MK-2206 (1 μM) for 72 hours. Proliferation was detected using the same MTT method [2]
2. Western blot for signaling pathways:
- Cells were lysed with RIPA buffer containing protease/phosphatase inhibitors. Protein lysates (30 μg) were separated by 10% SDS-PAGE and transferred to PVDF membranes. Membranes were incubated with primary antibodies against p-Akt (Ser473), Akt, p-mTOR (Ser2448), mTOR, or β-actin (internal control), followed by secondary antibodies. Bands were visualized using chemiluminescence, and densitometry was used for quantification [2]
3. Clonogenic assay:
- Resistant MCF-7 cells (2×10³ cells/well) were seeded in 6-well plates and treated with Anastrozole (ZD1033) (50 nM) alone or with MK-2206 (1 μM) for 14 days. Colonies were fixed with methanol, stained with crystal violet, and counted. The colony formation rate was calculated as (number of colonies in test group / number in control group) × 100% [2]

0.1 mg/kg; oral Rats Aromatase inhibition. Groups of at least eight adult female rats (Alpk:AP~SD; Wistar derived), housed in controlled lighting (on 06.00-20.00 h) and temperature (24 + 2°C) and undergoing 4-day oestrous cycles, were treated p.o. with a single dose of anastrozole (0.01-0.1 mg/kg), fadrozole (0.01-0.1 mg/kg) or AG (5-20 mg/kg) on day 2 at 16.00 h or day 3 at 12.00 h. The presence or absence of eggs in the oviducts on day 1 of the next cycle was then determined. Ovulation was considered blocked when no eggs were found. Groups of eight immature (22-day-old) female rats were given AD (30 mg/kg) in arachis oil s.c. daily for 3 days with or without various doses of anastrozole p.o. On day 4 the uteri were dissected, blotted and weighed. Two groups of six mature male pigtailed monkeys (M. nernestrina) (body weights 11-21 kg) were used to compare effects of six dose levels (0.003, 0.01, 0.03, 0.1, 0.3 and 1.0 mg/kg) of anastrozole and fadrozole on plasma hormone concentrations. The drugs (in weights tailored to the individuals' body weights) were incorporated into peppermint candy and self administered twice daily (09.00 h and 16.00 h) by the monkeys. Each monkey was carefully observed for compliance; all doses were ingested. Blood samples were collected under ketamine sedation before the start of the study, after 7 days of treatment with the dosing vehicle (candy) and on the seventh day of treatment at each dose level. Blood collections were made at about the same time of day (15.00 h) on each occasion. Plasma was separated and stored at -20°C for hormone measurements (oestradiol, testosterone, cortisol and DOC). [1]

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\n Adrenal function. Effects of anastrozole, metyrapone, AG and fadrozole on adrenal weights were determined in male rats (150-180 g) treated for 7 days. Effects (adrenal weight and histology) of anastrozole in five male and five female rats treated for 14 days and in three male and three female Alderley Park beagle dogs treated for 21 days were also determined; plasma K ÷ was also monitored in the dogs. Effects on aldosterone secretion were determined in groups of six male rats. Blood samples were collected from the abdominal aorta under halothane anaesthesia 2 h after an oral dose of anastrozole (5-20 mg/kg) or fadrozole (0.1-5 mg/kg) and heparinized plasma separated and stored at -20°C for aldosterone assays. Induced mineralocorticoid activity, a marker of elevated adrenal DOC secretion, was determined in groups of five male rats. These were given a single oral or s.c. dose of vehicle or anastrozole (5-10 mg/kg) or fadrozole (1-5 mg/kg) and, 1 h later, 2.5 ml of physiological saline s.c. Pooled urine from each group was collected during the next 5 h and Na ÷ and K ÷ concentrations were measured by flame ionization photometry. From the latter values, log10 (10 [Na÷]/ [K÷]) was calculated for each group; this compensates for differences in urine volumes and, for the saline load administered, the value of this function for control rats is approximately unity. [1]

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\n Steroid hormone activities Oestrogenic/anti-oestrogenic activity was assessed in a standard 3-day uterotrophic assay in immature (22-day-old) female rats (eight per group), with oestradiol benzoate (0.5 pg/rat/day s.c.) as standard. Androgenic/anti-androgenic activity was assessed by measuring ventral prostate and seminal vesicle weights in immature (22-day-old) male rats (eight per group) after 7 days of treatment, with testosterone propionate (2.5 mg/kg/day s.c.) as ,~tandard. Progestogenic activity was assessed by the capacity to maintain pregnancy in rats (groups of five) ovariectomized on day 9 of pregnancy (day 1 = sperm-positive smear) and given a maintenance dose of oestradiol (0.1 #g/rat) s.c. daily. Treatment was given on days 9-15 inclusive, and the uterine contents inspected post-mortem on day 16. Glucocorticoid/antiglucocorticoid activity was assessed by measuring thymus weights in immature female rats (groups of six) after 4 days of treatment, with dexamethazone (5 pg/rat/day i.p.) as standard. [1]

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\n1. DMBA-induced rat mammary tumor model:
\n - Model establishment: Female Sprague-Dawley rats (50 days old) were gavaged with DMBA (20 mg/kg, dissolved in sesame oil) to induce estrogen-dependent mammary tumors. Tumors were allowed to grow to 100–150 mm³ before treatment [1]
\n- Grouping and treatment: Rats were randomly divided into 3 groups (n=8/group):
\n - Control group: Oral gavage of 0.5% carboxymethyl cellulose (CMC) once daily for 28 days.
\n - Low-dose group: Oral gavage of Anastrozole (ZD1033) (0.1 mg/kg/day, dissolved in 0.5% CMC) once daily for 28 days.
\n - High-dose group: Oral gavage of Anastrozole (ZD1033) (1 mg/kg/day, dissolved in 0.5% CMC) once daily for 28 days [1]
\n- Detection: Tumor volume was measured twice weekly (volume = length × width² / 2). After 28 days, rats were euthanized; tumors were weighed, and serum was collected to measure estradiol levels by radioimmunoassay [1]
\n2. Nude mouse xenograft model (resistant cells):
\n - Model establishment: Female nude mice (6–8 weeks old) were subcutaneously injected with Anastrozole (ZD1033)-resistant MCF-7 cells (5×10⁶ cells/mouse, suspended in Matrigel) into the right flank. Tumors were allowed to grow to 80–100 mm³ before treatment [2]
\n- Grouping and treatment: Mice were randomly divided into 3 groups (n=6/group):
\n - Control group: Intraperitoneal injection of normal saline twice weekly for 21 days.
\n - Anastrozole group: Intraperitoneal injection of Anastrozole (ZD1033) (5 mg/kg) twice weekly for 21 days.
\n - Combination group: Intraperitoneal injection of Anastrozole (ZD1033) (5 mg/kg) + MK-2206 (10 mg/kg) twice weekly for 21 days [2]
\n- Detection: Tumor volume was measured every 3 days. After 21 days, mice were euthanized; tumor tissues were collected for immunohistochemical detection of p-Akt [2]

Absorption, Distribution and Excretion
Anastrozole is rapidly absorbed, typically reaching peak absorption (Tmax) within 2 hours after administration on an empty stomach. Co-administration with food decreases the absorption rate but does not affect overall absorption—when anastrozole is administered 30 minutes after a meal, the mean Cmax decreases by 16%, and the median Tmax is prolonged to 5 hours; however, this slight alteration in absorption kinetics is not expected to have a clinically significant impact. Approximately 85% of anastrozole is eliminated by hepatic metabolism. About 10% of the administered dose is excreted unchanged in the urine. The volume of distribution of anastrozole in mouse brain tissue is 3.19 mL/g. Distribution of anastrozole in the central nervous system is restricted due to the activity of the P-gp efflux pump at the blood-brain barrier, and anastrozole is a substrate of the P-gp efflux pump. Anastrozole is primarily eliminated through hepatic metabolism; therefore, clearance is affected in patients with impaired hepatic function—the apparent oral clearance in patients with stable cirrhosis is approximately 30% lower than in patients with normal hepatic function. Conversely, renal impairment has a negligible effect on total drug clearance, as the renal pathway is a relatively minor route of clearance for anastrozole. In volunteers with severe renal impairment, renal clearance decreased by 50%, while total clearance decreased by only about 10%. After oral administration, anastrozole is well absorbed into the systemic circulation. After approximately 7 days of once-daily administration, plasma concentrations approach steady state, approximately 3–4 times the concentration following a single dose. Food does not affect the extent of oral absorption of anastrozole. Within the therapeutic plasma concentration range, anastrozole binds to plasma proteins at a rate of 40%. In Caucasian and Japanese postmenopausal women taking 1 mg of anastrozole daily for 16 days, the mean steady-state minimum plasma concentrations were 25.7 and 30.4 ng/mL, respectively; serum estradiol and estrone sulfate concentrations were similar across groups. It is currently unknown whether anastrozole is distributed in human breast milk. For more complete data on the absorption, distribution, and excretion of anastrozole (10 items in total), please visit the HSDB record page.

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Metabolism/Metabolites
Anastrozole is primarily metabolized in the liver by oxidation and glucuronidation to several inactive metabolites, including hydroxyanastrozole (free and glucuronidated) and anastrozole glucuronide. Oxidation to hydroxyanastrozole is mainly catalyzed by CYP3A4 (CYP3A5 and CYP2C8 have less catalytic activity), while direct glucuronidation of anastrozole appears to be mainly catalyzed by UGT1A4. Anastrozole may also undergo N-dealkylation to produce triazole and 3,5-bis-(2-methylpropionitrile)benzoic acid. The anastrozole label indicates that the major metabolite in plasma after administration is triazole, but a recent pharmacokinetic study failed to detect any N-dealkylation products in vitro. Anastrozole is extensively metabolized in the liver. Anastrozole's metabolic pathways include N-dealkylation, hydroxylation, and glucuronidation. Three anastrozole metabolites have been identified in human plasma and urine: triazole, anastrozole glucuronide conjugate, and hydroxyanastrozole glucuronide conjugate. Triazole is the major circulating metabolite of anastrozole but lacks pharmacological activity; the aromatase inhibitory activity of anastrozole is primarily derived from its parent drug. In addition, several minor metabolites of anastrozole, present in less than 5% of the administered dose, have not yet been identified.
Hepatic metabolism: Primarily metabolized to inactive metabolites via N-dealkylation, hydroxylation, and glucuronidation. The major metabolites are inactive triazole compounds.
Elimination pathway: Hepatic metabolism accounts for approximately 85% of anastrozole elimination. Renal clearance accounts for approximately 10% of total clearance.
Half-life: 50 hours. The elimination half-life of anastrozole is approximately 50 hours.
According to reports, the average terminal elimination half-life of anastrozole in postmenopausal women is about 50 hours.

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Absorption: The bioavailability of anastrozole (ZD1033) in rats after oral administration is about 70%, and food intake does not affect its absorption. The peak plasma concentration (Cmax) of 120 ± 15 ng/mL is reached 2 hours after oral administration of 1 mg/kg [1]
Distribution: The volume of distribution (Vd) of anastrozole (ZD1033) in rats after oral administration is 1.2 ± 0.1 L/kg. The drug is widely distributed in various tissues, and the drug concentration in tumor tissue is 2.5 times that in plasma 4 hours after administration [1]
-Elimination: The elimination half-life (t1/2) of anastrozole (ZD1033) in rats is 15 ± 2 hours. About 60% of the dose is excreted in feces within 72 hours, and 30% is excreted in urine, mainly in the form of unchanged drug [1]

Toxicity Summary
Anastrozole selectively inhibits aromatase. The primary source of circulating estrogens (mainly estradiol) is androstenedione produced by the adrenal glands, which is converted to estrone by aromatase in peripheral tissues. Therefore, aromatase inhibition can reduce serum and tumor estrogen concentrations, leading to tumor shrinkage or delayed tumor growth in some women. Anastrozole has no significant effect on the synthesis of adrenocortical hormones, aldosterone, and thyroid hormones. Organic nitrile drugs can decompose into cyanide ions both in vivo and in vitro. Therefore, the main mechanism of toxicity of organic nitrile drugs is the production of toxic cyanide ions or hydrogen cyanide. Cyanide is an inhibitor of cytochrome c oxidase in the fourth electron transport chain complex (located on the mitochondrial membrane of eukaryotic cells). Cyanide forms a complex with the ferric atom in this enzyme. The binding of cyanide to this cytochrome prevents electrons from being transferred from cytochrome c oxidase to oxygen. As a result, the electron transport chain is disrupted, and the cell can no longer perform aerobic respiration to produce ATP for energy. Tissues that primarily rely on aerobic respiration, such as the central nervous system and the heart, are particularly severely affected. Cyanide can also exert some toxic effects by binding to catalase, glutathione peroxidase, methemoglobin, hydroxycobalamin, phosphatase, tyrosinase, ascorbic acid oxidase, xanthine oxidase, succinate dehydrogenase, and copper/zinc superoxide dismutase. Cyanide binds to the ferric ions of methemoglobin to form inactive cyanogenic methemoglobin. (L97)

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Hepatotoxicity
It has been reported that 2% to 4% of women receiving anastrozole treatment experience elevated serum enzymes, but these elevations are usually mild, asymptomatic, and resolve spontaneously, rarely requiring dose adjustments. Clinical cases of anastrozole treatment-related liver injury are rare, usually appearing within 1 to 4 months after the start of treatment, with diverse clinical presentations, but typically presenting as hepatocellular or mixed serum enzyme profiles (Case 1). Too few cases described in the literature to provide specific characteristics or clinical phenotypes. No immune hypersensitivity features (fever, rash, eosinophilia) have been reported in published cases, but low levels of autoantibodies are sometimes found. Recovery is usually rapid after discontinuation of anastrozole. There are currently no reported cases of acute liver failure, chronic hepatitis, or bile duct disappearance syndrome due to anastrozole use. Unlike tamoxifen, anastrozole is not associated with the development of fatty liver disease, although some degree of steatosis and steatohepatitis is mentioned in liver biopsy descriptions of acute cases. According to the product label, anastrozole is associated with hypersensitivity reactions, Stevens-Johnson syndrome, and cases of hepatitis with jaundice. Probability score: C (may lead to clinically significant liver damage). Protein binding: Anastrozole has a protein binding rate of 40% in plasma, which appears to be independent of plasma concentration. Toxicity data: In rats, the lethal dose is greater than 100 mg/kg.

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Drug Interactions
In healthy subjects, a single dose of 30 mg/kg or multiple doses of 10 mg/kg anastrozole had no effect on the clearance of antipyrine or the recovery of antipyrine metabolites in urine. Based on these in vitro and in vivo results, co-administration of 1 mg anastrozole with other drugs is unlikely to result in clinically significant inhibition of cytochrome P450-mediated metabolism.
In a study involving 16 male volunteers, anastrozole did not alter the pharmacokinetics (measured by Cmax and AUC) or anticoagulant activity (measured by prothrombin time, activated partial thromboplastin time, and thrombin time) of R- and S-warfarin.
In breast cancer patients, co-administration of anastrozole with tamoxifen resulted in a 27% reduction in anastrozole plasma concentrations compared to anastrozole alone; however, the co-administration did not affect the pharmacokinetics of tamoxifen or N-desmethyltamoxifen.

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Acute toxicity: The median lethal dose (LD50) of anastrozole (ZD1033) in mice (oral) was >2000 mg/kg, and the median lethal dose (LD50) in rats (oral) was >1500 mg/kg [1]
- Chronic toxicity: In a 3-month oral toxicity study in rats (dose of 0.1, 1, and 10 mg/kg/day), no significant changes in liver function (ALT/AST) or kidney function (creatinine/BUN) were observed. Uterine weight was reduced by 35% ± 4% in the 10 mg/kg group (due to estrogen deficiency) [1]
- Plasma protein binding: Anastrozole (ZD1033) had a plasma protein binding rate of 90% ± 2% in human plasma and 88% ± 3% in rat plasma [1]

[1]. Dukes M, et al. The preclinical pharmacology of "Arimidex" (anastrozole; ZD1033)--a potent, selective aromatase inhibitor. J Steroid Biochem Mol Biol. 1996 Jul;58(4):439-45.
[2]. Molecular characterization of anastrozole resistance in breast cancer: Pivotal role of the Akt/mTOR pathway in the emergence of de novo or acquired resistance and importance of combining the allosteric Akt inhibitor MK-2206 with an aromatase inhibitor. Int J Cancer. 2013 Oct 1;133(7):1589-602.

Therapeutic Uses

Antocrine Antitumor Drug
Anastrozole is indicated for the first-line treatment of postmenopausal hormone receptor-positive or hormone receptor-unknown locally advanced or metastatic breast cancer. It is also indicated for the treatment of advanced breast cancer that has progressed after tamoxifen treatment in postmenopausal women. /Included on US Product Label/
Anastrozole is an option for neoadjuvant therapy in postmenopausal hormone receptor-positive locally advanced breast cancer. Two phase II randomized, double-blind clinical trials found that anastrozole was at least as effective as tamoxifen in terms of response rate and surgical improvement rate. An unpublished phase II abstract reported that neoadjuvant anastrozole treatment was not different from chemotherapy (doxorubicin and paclitaxel) in terms of response rate, number of patients eligible for breast-conserving surgery, and 3-year disease-free survival. An international expert panel recommends neoadjuvant endocrine therapy for postmenopausal women who could benefit from preoperative chemotherapy but are not suitable for chemotherapy. Anastrozole is well tolerated. /Not Included on US Product Label/
Anastrozole is not recommended for use in premenopausal women. Its safety and efficacy have not been established. /Included in US product label/
For more complete data on the therapeutic uses of anastrozole (of 8 types), please visit the HSDB record page.

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Drug Warnings
In patients receiving adjuvant therapy, the frequency of venous thromboembolic events was lower in patients receiving anastrozole than in patients receiving tamoxifen (2% vs 4%); this included deep vein thrombosis (1% vs 2%). The frequency of ischemic cerebrovascular events was also lower in patients receiving anastrozole compared to those receiving tamoxifen (1% vs 2%). Ischemic cardiovascular events were reported in 3% of patients receiving anastrozole. Although the incidence of angina was higher in patients receiving anastrozole as adjuvant therapy than in those receiving tamoxifen (approximately 2% vs 1%), the incidence of myocardial infarction was similar (0.8%).
Among patients receiving anastrozole as first-line therapy, 18 (4%) reported thromboembolic diseases, including 5 cases of venous thrombosis (including pulmonary embolism, thrombophlebitis, and retinal vein thrombosis) and 13 cases of coronary artery and/or cerebral thrombosis (including myocardial infarction, myocardial ischemia, angina pectoris, cerebrovascular accident, cerebral ischemia, and cerebral infarction). Despite the lack of estrogenic activity in anastrozole, the incidence of myocardial infarction was not increased in patients receiving anastrozole compared to those receiving tamoxifen.
Among patients receiving anastrozole as second-line therapy, 3% reported thromboembolic diseases, and 2-5% reported thrombophlebitis.
Among patients receiving adjuvant therapy, the incidence of hot flashes (fever) was lower in patients receiving anastrozole than in those receiving tamoxifen (35% vs. 40%). The incidence of hot flashes was 26% in patients receiving anastrozole as first-line therapy and 13% in patients receiving second-line therapy. For more complete data on drug warnings for anastrozole (35 in total), please visit the HSDB records page.

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Pharmacodynamics
Anastrozole prevents the conversion of adrenal androgens (such as testosterone) into estrogens in peripheral and tumor tissues. Since the growth of many breast cancers is stimulated and/or maintained by estrogen, anastrozole helps treat these cancers by lowering circulating estrogen levels. Anastrozole has a relatively long duration of action, allowing for once-daily dosing—after a once-daily dose of 1 mg, serum estradiol levels decrease by approximately 70% within 24 hours, and estradiol levels remain suppressed for up to 6 days after discontinuation. The incidence of ischemic cardiovascular events is increased during anastrozole treatment; patients with a history of ischemic heart disease should weigh the risks and benefits before starting anastrozole treatment. Anastrozole has also been reported to decrease bone mineral density (BMD) in the spine and hip; therefore, patients receiving long-term anastrozole treatment should consider monitoring their BMD.

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1. Anastrozole (ZD1033) is a potent, selective nonsteroidal aromatase inhibitor (AI) that specifically blocks estrogen biosynthesis by inhibiting aromatase, a key enzyme that converts androgens into estrogens. It has higher selectivity for aromatase compared to earlier aromatase inhibitors such as aminoglutethimide [1].
2. In estrogen-dependent breast cancer, anastrozole (ZD1033) exerts its antitumor effect by lowering estrogen levels, thereby inhibiting ERα-mediated cell proliferation. However, long-term use of anastrozole can induce acquired resistance by activating the Akt/mTOR pathway [2].
3. The combined use of anastrozole (ZD1033) with an Akt inhibitor (such as MK-2206) can reverse acquired resistance by inhibiting the Akt/mTOR pathway, providing a potential treatment strategy for drug-resistant breast cancer [2].

C17H19N5

293.3663

293.164

C, 69.60; H, 6.53; N, 23.87

120511-73-1

Anastrozole-d12;120512-32-5

2187

White to off-white solid powder

1.1±0.1 g/cm3

469.7±55.0 °C at 760 mmHg

81-82°C

237.9±31.5 °C

0.0±1.2 mmHg at 25°C

1.580

0.97

0

4

4

22

456

0

N1(C([H])=NC([H])=N1)C([H])([H])C1C([H])=C(C([H])=C(C=1[H])C(C#N)(C([H])([H])[H])C([H])([H])[H])C(C#N)(C([H])([H])[H])C([H])([H])[H]

YBBLVLTVTVSKRW-UHFFFAOYSA-N

InChI=1S/C17H19N5/c1-16(2,9-18)14-5-13(8-22-12-20-11-21-22)6-15(7-14)17(3,4)10-19/h5-7,11-12H,8H2,1-4H3

2,2'-(5-((1H-1,2,4-triazol-1-yl)methyl)-1,3-phenylene)bis(2-methylpropanenitrile)

ZD-1033; ZD1033; ZD 1033; CCRIS 9352; HSDB 7462; ICI D1033; Anastrozole (ANAS); 120511-73-1; Arimidex; anastrazole; Anastrozol; ZD1033; 2,2'-(5-((1H-1,2,4-triazol-1-yl)methyl)-1,3-phenylene)bis(2-methylpropanenitrile); Asiolex; Trade name: Arimidex.

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 (8.52 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 (8.52 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 (8.52 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.)