Pazopanib HCl is a receptor tyrosine kinase inhibitor that targets multiple kinases, including VEGFR1, VEGFR2, VEGFR3, PDGFR, FGFR, c-Kit, and c-Fms, with IC50s 10nM, 30nM, 47nM, 84nM, 74nM, 140nM, and 146nM respectively [1-3]. Pazopanib inhibits both tumor growth and angiogenesis through suppressing these targets. In preclinical studies, it has shown anti-tumor activity against several human tumor xenografts, including renal, prostate, colon, lung, melanoma, head and neck, and breast cancer [4]. It also showed desirable pharmacokinetics and oral bioavailability in animal models [4].
Pazopanib has been approved for advanced/metastatic renal cell carcinoma and advanced soft tissue sarcomas by multiple regulatory administrations worldwide, including FDA, EMA, MHRA and TGA. In the clinical trial for soft tissue sarcomas, Pazopanib improved median progression-free survival (PFS) to 4.6 months compared to 1.6 months for patients receiving placebo [5]. In the trial for renal cell carcinoma, Pazopanib increased the median PFS from 4.2 monthe (placebo) to 9.2 months [5]. The most common adverse effect of Pazopanib were diarrhea, hypertension, hair color change, nausea, fatigue, anorexia, and vomiting [6].
参考文献:
1.?Verweij J, Sleijfer S. Pazopanib, a new therapy for metastatic soft tissue sarcoma. Expert Opin Pharmacother 2013; 14: 929-935.
2.?Pick AM, Nystrom KK. Pazopanib for the treatment of metastatic renal cell carcinoma. Clin Ther 2012; 34: 511-520.
3.?Bukowski RM, Yasothan U, Kirkpatrick P. Pazopanib. Nat Rev Drug Discov 2010; 9: 17-18.
4.?Sonpavde G, Hutson TE. Pazopanib: a novel multitargeted tyrosine kinase inhibitor. Curr Oncol Rep 2007; 9: 115-119.
5.?http://www.cancer.gov/cancertopics/druginfo/fda-pazopanibhydrochloride
6.?http://www.gsksource.com/gskprm/en/US/adirect/gskprm?cmd=ProductDetailPage&product_id=1336067580985&featureKey=603422
Kinase experiment: | VEGFR enzyme assays are initiated by the addition of 10 μL of activated VEGFR2 kinase solution [final concentration, 1 nM enzyme in 0.1 M 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), pH 7.5, containing 0.1 mg/mL bovine serum albumin (BSA), 300 μM dithiothreitol (DTT)] to 10 μL substrate solution [final concentration, 360 nM peptide, (biotin-aminohexyl-EEEEYFELVAKKKK-NH2), 75 μM ATP, 10 μM MgCl2], and 1 μL of titrated compound (Pazopanib) in DMSO. Plates are incubated at room temperature for 60 min, and then the reaction is quenched by the addition of 20 μL of 100 mM ethylene diamine tetraacetic acid (EDTA). After quenching, 20 μL HTRF reagents (final concentration, 15 nM Streptavidin-linked allophycocyanin, 1 nM Europium-labeled antiphosphotyrosine antibody diluted in 0.1 mg/mL BSA, 0.1 M HEPES, pH 7.5) is added and the plates incubated for a minimum of 10 min. The fluorescence at 665 nM is measured with a plate reader[1]. |
Cell experiment: | The effect of Pazopanib on cell proliferation is measured using 5-bromo-2-deoxyuridine (BrdU) incorporation method using commercially available kits. HUVEC is seeded in medium containing 5% fetal bovine serum (FBS) in type 1 collagen coated 96-well plates and incubated overnight at 37°C, 5% CO2. The medium is aspirated from the cells, and various concentrations of Pazopanib in serum-free medium are added to each well. After 30 min, either VEGF (10 ng/mL) or bFGF (0.3 ng/mL) is added to the wells. Cells are incubated for an additional 72 h and BrdU (10 μM) is added during the last 18 to 24 h of incubation. At the end of incubation, BrdU incorporation in cells is measured by ELISA. Data are fitted with a curve described by the equation, y=Vmax(1?(x/(K+x))), where K is equal to the IC50[1]. |
Animal experiment: | Mice[1] Tumors are initiated by injection of tumor cell suspension in 8?12 week old nude mice. When tumors reach a volume of 100?200 mm3, mice are randomized and divided into groups of eight. Pazopanib is administered once or twice daily at 10, 30, or 100 mg/kg. Animals are euthanized by inhalation of CO2 at the completion of the study. Tumor volume is measured twice weekly by calipers, using the equation: tumor volume (mm3)=(length×width2)/2. Results are routinely reported as % inhibition=1?(average growth of the drug treated population/average growth of vehicle treated control population). Rats[2] Male Brown-Norway (BN; pigmented) rats weighing 200 to 250 g are acclimatized for at least two days prior to any experimental procedure. After overnight fasting for 12-16 h, an intraperitoneal injection of 30 mg/mL solution of Streptozotocin in 10 mM citrate buffer (pH 4.5) is administered (60 mg/kg body weight) to induce diabetes. After 3-4 h of Streptozotocin injection, animals are put on a regular diet and 24 h after Streptozotocin injection, blood sample (5-10 μL) is collected via tail vein. The blood glucose levels in the animals are determined with a glucose monitor. Animals with blood glucose levels greater than 250 mg/dL are considered diabetic. The animals are divided into three groups. Group 1: Healthy (n=12), Group 2: Diabetic (n=12) and Group 3: Diabetic+Treatment (n=12). Treatment is started immediately after diabetes induction. Both eyes are dosed twice daily for 30 days with 0.5 % w/v Pazopanib suspension (10 μL volume in each eye) and animals in all groups are sacrificed on day 31, 16-17 h after last dose on day 30. |
参考文献: [1]. Harris PA, et al. Discovery of 5-[[4-[(2,3-dimethyl-2H-indazol-6-yl)methylamino]-2-pyrimidinyl]amino]-2-methyl-benzenesulfonamide (Pazopanib), a novel and potent vascular endothelial growth factor receptor inhibitor. J Med Chem. 2008, 51(15), 4632-4640. | |
动态评分
0.0