The formation of bone marrow megakaryocytes is efficiently inhibited by angrelide (IC50=26 nM) [1]. One special medication that prevents bone marrow megakaryocytopoiesis is anagrelide. Only megakaryocyte proliferation was suppressed by angrelide (0.05, 0.3, and 1 µM), but not that of non-megakaryocytic cells. The fact that anagrelide causes cell proliferation to relocate to non-megakaryocytic compartments suggests that it does not have cytotoxic properties [2]. Although anagrelide only exhibits mild action in the GIST48 cell line, it can cause cytotoxicity in the GIST882 cell line at submicromolar concentrations (IC50= 16 nM) [3]. In vitro, anagrelide can decrease the growth of gastrointestinal stromal tumor (GIST) cells and increase apoptosis [3].

Anagrelide is efficacious in xenograft mouse models of GIST with KIT exon 9 mutations, which may represent treatment problems because these GISTs require large daily doses of imatinib [3]. Anagrelide suppresses GIST development in patient-derived mice xenograft models. Anagrelide has therapeutic potential in the treatment of GIST. Anagrelide exhibits anticancer efficacy in gastrointestinal stromal tumor xenograft models [3]. Anagrelide (5 mg/kg/bid) suppresses or decreases tumor growth in GIST2B, GIST9, and GIST882 models [3].

Cell proliferation assay[2]
Cell Types: megakaryocytes and non-megakaryocytes
Tested Concentrations: 0.05, 0.3, 1 µM
Incubation Duration: 12 days
Experimental Results: Inhibition of megakaryocyte growth only at each concentration tested

Animal/Disease Models: Adult female athymic mouse GIST2B, GIST3, GIST9, GIST882 model [3]
Doses: 5 mg/kg/bid
Route of Administration: 5 mg/kg/bid or combined with anagrelide and imatinib ( given simultaneously) dose and schedule as a single agent); for 10 days
Experimental Results: Three of the four models (GIST2B, GIST9, GIST882) inhibited or diminished tumor growth. The most potent effect was observed in the GIST2B model, which contains the KIT exon 9 mutation leading to the p.A502_Y503 duplication. The tumor volume of this model diminished by 68% after 10 days of treatment.

Absorption, Distribution and Excretion
Following oral administration, the bioavailability of anagrelide is approximately 70%. Given on an empty stomach, the Cmax is reached within 1 hour (Tmax) of administration. Co-administration with food slightly lowers the Cmax and increases the AUC, but not to a clinically significant extent.
Following metabolism, urinary excretion of metabolites appears to be the primary means of anagrelide elimination. Less than 1% of an administered dose is recovered in the urine as unchanged parent drug, while approximately 3% and 16-20% of the administered dose is recovered as 3-hydroxy anagrelide and RL603, respectively.
Following oral administration of 14C-anagrelide in people, more than 70% of radioactivity was recovered in urine. The available plasma concentration time data at steady state in patients showed that anagrelide does not accumulate in plasma after repeated administration.
The volume of distribution (VolD) is 12 L/kg of body weight.
Limited data indicate probable dose linearly between doses of 500 mg (0.5 mg) and 2 mg. Bioavailability was found to be modestly reduced by an average of 13.8% when anagrelide was administered after food.
The peak plasma level was lowered by an average of 45% and delayed by 2 hours ... when a 0.5 mg dose of anagrelide was taken after food ...
For more Absorption, Distribution and Excretion (Complete) data for ANAGRELIDE (6 total), please visit the HSDB record page.

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Metabolism / Metabolites
Anagrelide is extensively metabolized, primarily in the liver by cytochrome P450 1A2 (CYP1A2), into two major metabolites: 6,7-dichloro-3-hydroxy-1,5 dihydro-imidazo[2,1-b]quinazolin-2-one (3-hydroxy anagrelide) and 2-amino-5,6-dichloro-3,4,-dihydroquinazoline (RL603). The 3-hydroxy metabolite is considered pharmacologically active and carries a similar potency and efficacy in regards to its platelet-lowering effects, but inhibits PDE3 with a potency 40x greater than that of the parent drug.
Anagrelide is extensively metabolized; less than 1% is recovered unchanged in the urine.

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Biological Half-Life
The t_1/2 of anagrelide and its active metabolite, 3-hydroxy anagrelide, are approximately 1.5 hours and 2.5 hours, respectively.
At fasting and at a dose of 0.5 mg, the plasma half-life is 1.3 hours ... .
Elimination: Plasma 1.3 hours (at a dose of 0.5 mg while fasting). Note: Plasma half life was found to be increased (to 1.8 hours) when anagrelide was taken after food. Steady state plasma concentration measurements show no accumulation of anagrelide in plasma with repeated administration.

Hepatotoxicity
In preregistration studies, anagrelide was not associated with serum enzyme elevations or with episodes of clinically apparent liver injury. Since its approval, there has been a single published abstract reporting progressive, ultimately fatal cholestasis after liver transplantation and use of anagrelide, but there have been no other published reports of anagrelide hepatotoxicity in the literature. In large, long term follow up studies there have been occasional instances of transient serum enzyme elevations without jaundice or symptoms. The product label for anagrelide mentions abnormal enzymes as an adverse event but not clinically apparent liver injury, hepatitis or jaundice. However, the general clinical experience with anagrelide has been limited.
Likelihood score: E* (unlikely, but suspected rare cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the use of anagrelide during breastfeeding. The manufacturer recommends that the drug not be used during breastfeeding and for 1 week after the last dose.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Interactions
Bioavailability studies evaluating possible interactions between anagrelide and other drugs have not been conducted. The most common medications used concomitantly with anagrelide have been aspirin, acetaminophen, furosemide, iron, ranitidine, hydroxyurea, and allopurinol. The most frequently used concomitant cardiac medication has been digoxin. Although drug-to-drug interaction studies have not been conducted, there is no clinical evidence to suggest that anagrelide interacts with any of these compounds.
A case report has suggested that sucralfate may interfere with anagrelide absorption.

[1]. Comparison of the biological activities of Anagrelide and its major metabolites in haematopoietic cell cultures. Br J Pharmacol. 2005 Oct;146(3):324-32.

[2]. Comparison between Anagrelide and hydroxycarbamide in their activities against haematopoietic progenitor cell growth and differentiation: selectivity of Anagrelide for the megakaryocytic lineage. Leukemia. 2006 Jun;20(6):1117-22.

[3]. Anagrelide for Gastrointestinal Stromal Tumor. Clin Cancer Res. 2019 Mar 1;25(5):1676-1687.

Therapeutic Uses
Antithrombocythemic
Anagrelide is indicated for reduction of elevated platelet counts and of the risk of thrombosis, as well as for amelioration of symptoms, in patients with thrombocythemia secondary to myeloproliferative disorders. /Included in US product labeling/

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Drug Warnings
While most reported adverse events during anagrelide therapy have been mild in intensity and have decreased in frequency with continued therapy, serious adverse events were reported in patients with myeloproliferative diseases of varying etiology. These include the following: congestive heart failure, myocardial infarction, cardiomyopathy, cardiomegaly, complete heart block, atrial fibrillation, cerebrovascular accident, pericarditis, pulmonary infiltrates, pulmonary fibrosis, pulmonary hypertension, pancreatitis, gastric/duodenal ulceration, and seizure.
The most frequently reported adverse reactions to anagrelide (in 5% or greater of 942 patients with myeloproliferative disease) in clinical trials were: headache 43.5%; palpitations 26.1%; diarrhea 25.7%; asthenia 23.1%; edema, other 20.6%; nausea 17.1%; abdominal pain 16.4%; dizziness 15.4%; pain, other 15.0%; dyspnea 11.9%; flatulence 10.2%; vomiting 9.7%; fever 8.9%; peripheral edema 8.5%; rash, including urticaria 8.3%; chest pain 7.8%; anorexia 7.7%; tachycardia 7.5%; pharyngitis 6.8%; malaise 6.4%; cough 6.3%; paresthesia 5.9%; back pain 5.9%; pruritus 5.5%; dyspepsia 5.2%.
It is recommended that patients with renal insufficiency (creatinine 2 mg/dL or greater) receive anagrelide when the potential benefits of therapy outweigh the potential risks. Monitor patients closely for signs of renal toxicity while receiving anagrelide.
Use with caution in patients with known or suspected heart disease only if the potential benefits outweigh the potential risks. Because of the positive inotropic effects and side effects of anagrelide, a pretreatment cardiovascular examination is recommended along with careful monitoring during treatment. Therapeutic doses of anagrelide may cause cardiovascular effects, including vasodilation, tachycardia, palpitations, and congestive heart failure.
For more Drug Warnings (Complete) data for ANAGRELIDE (11 total), please visit the HSDB record page.

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Pharmacodynamics
Anagrelide decreases platelet counts by suppressing transcription factors necessary for the synthesis and maturation of platelet-producing cells. The drug itself appears to have a relatively short residence time in the body necessitating twice or four times daily dosing. However, given that the pharmacological effect of anagrelide therapy is reliant on a gradual suppression of platelet-producing cells, it may take 7 to 14 days for its administration to be reflected in reduced platelet counts - for this reason any changes to anagrelide doses should not exceed 0.5 mg/day in any one week. Evidence from animal studies suggests anagrelide may impair female fertility. Female patients of reproductive age should be advised of the potential for adverse effects on fertility prior to initiating therapy.

C10H7N3OCL2

256.08808

254.996

68475-42-3

Anagrelide hydrochloride;58579-51-4;Anagrelide hydrochloride monohydrate;823178-43-4

135409400

Typically exists as solid at room temperature

1.8±0.1 g/cm3

376.5±52.0 °C at 760 mmHg

280 °C

181.5±30.7 °C

0.0±0.9 mmHg at 25°C

1.791

1.96

1

2

0

16

360

0

O=C1N=C2NC3=C(C(Cl)=C(Cl)C=C3)CN2C1

OTBXOEAOVRKTNQ-UHFFFAOYSA-N

InChI=1S/C10H7Cl2N3O/c11-6-1-2-7-5(9(6)12)3-15-4-8(16)14-10(15)13-7/h1-2H,3-4H2,(H,13,14,16)

6,7-dichloro-3,5-dihydro-1H-imidazo[2,1-b]quinazolin-2-one

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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