VEGFR2 (IC50 = 6.5 nM); VEGFR3 (IC50 = 15 nM); EphB2 (IC50 = 24 nM); VEGFR1 (IC50 = 30 nM); PDGFRα (IC50 = 40 nM)

Tivozanib HCl Hydrate prevents VEGFR-1, VEGFR-2, and VEGFR-3 from being phosphorylated [2]. AV-951 is a novel derivative of urea and quinoline. AV-951 prevents endothelial cell proliferation and the VEGF-dependent activation of mitogen-activated protein kinases.[1]
KRN951 is a novel tyrosine kinase inhibitor for VEGFRs with antitumor angiogenesis and antigrowth activities. KRN951 potently inhibited VEGF-induced VEGFR-2 phosphorylation in endothelial cells at in vitro subnanomolar IC50 values (IC50 = 0.16 nmol/L). It also inhibited ligand-induced phosphorylation of platelet-derived growth factor receptor-beta (PDGFR-beta) and c-Kit (IC50 = 1.72 and 1.63 nmol/L, respectively). KRN951 blocked VEGF-dependent, but not VEGF-independent, activation of mitogen-activated protein kinases and proliferation of endothelial cells. In addition, it inhibited VEGF-mediated migration of human umbilical vein endothelial cells.[1]

Tivozanib hydrochloride hydrate (1 mg/kg; oral; 14 days) can suppress the development of CNV lesions and significantly resolve established CNV, reducing the affected area by 67.7% .
Studies conducted in vivo demonstrate that AV-951, particularly when administered orally at a dose of 1 mg/kg, also reduces micro vessel density and suppresses VEGFR2 phosphorylation levels in tumor xenografts. In athymic rats, AV-951 almost completely inhibits the growth of tumor xenografts (TGI>85%).[1] Another study using a peritoneal disseminated tumor model in rats demonstrates that AV-951 can extend the survival of the tumor-bearing rats up to 53.5 days after the MST. When applied to various human tumor xenografts, such as lung, breast, colon, ovarian, pancreatic, and prostate cancer, AV-951 exhibits antitumor activity.[2]

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Following p.o. administration to athymic rats, KRN951 decreased the microvessel density within tumor xenografts and attenuated VEGFR-2 phosphorylation levels in tumor endothelium. It also displayed antitumor activity against a wide variety of human tumor xenografts, including lung, breast, colon, ovarian, pancreas, and prostate cancer. Furthermore, dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) analysis revealed that a significant reduction in tumor vascular hyperpermeability was closely associated with the antitumor activity of KRN951. These findings suggest that KRN951 is a highly potent, p.o. active antiangiogenesis and antitumor agent and that DCE-MRI would be useful in detecting early responses to KRN951 in a clinical setting. KRN951 is currently in phase I clinical development for the treatment of patients with advanced cancer.[1]

The IC50 values of AV-951 against various recombinant receptor and nonreceptor tyrosine kinases, such as VEGFR1, VEGFR2, VEGFR3, c-Kit, PDGFRβ, Flt-3, and FGFR1, are ascertained by conducting cell-free kinase assays in quadruplicate using 1 μM ATP.

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Kinase selectivity. [1]
Cell-free kinase assays were done in quadruplicate with 1 μmol/L ATP to determine the IC50 values of Tivozanib (AV951; KRN-951) against a variety of recombinant receptor and nonreceptor tyrosine kinases. Recombinant enzymes were obtained from ProQinase GmbH. [1]
Cell-based assays were done to determine the ability of Tivozanib (AV951; KRN-951) to inhibit ligand-dependent phosphorylation of receptor tyrosine kinases as described previously. Briefly, the cells were starved overnight in appropriate basic medium containing 0.5% fetal bovine serum (FBS). Following the addition of KRN951 or 0.1% DMSO, the cells were incubated for 1 hour and then stimulated with the cognate ligand at 37°C. Receptor phosphorylation was induced for 5 minutes except for VEGFR-3 (10 minutes), c-Met (10 minutes), and c-Kit (15 minutes). All the ligands used in the assays were human recombinant proteins, except for VEGF-C, a rat recombinant protein. Following cell lysis, receptors were immunoprecipitated with appropriate antibodies and subjected to immunoblotting with phosphotyrosine. Quantification of the blots and calculation of IC50 values were carried out as described previously. [1]

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Mitogen-activated protein kinase activation. [1]
This was evaluated as described previously. Briefly, HUVECs were starved for 16 hours in a basic medium (EBM-2) containing 0.5% FBS. Following incubation with Tivozanib (AV951; KRN-951) for 1 hour, HUVECs were stimulated with 50 ng/mL VEGF, 25 ng/mL basic fibroblast growth factor or 20 ng/mL EGF. Cell lysates were subjected to SDS-PAGE followed by immunoblotting of phosphorylated MAPKs with phosphorylated p44/42 mitogen-activated protein kinase (MAPK) antibody.[1]

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Permeability assay and detection of VEGFR‐2 phosphorylation.[2]
The effects of malignant ascites on endothelial cell permeability and VEGFR‐2 phosphorylation, and the inhibitory activity of Tivozanib (AV951; KRN-951) on these effects, were evaluated. Ascites samples taken from a vehicle‐treated peritoneal disseminated tumor model on day 25 were pooled and the resulting supernatant was used in these assays. We examined propidium iodide uptake as a measure of permeability in an in vitro assay. VEGFR‐2 phosphorylation was determined by western blotting. For the permeability assay, HUVEC at 90% confluence in culture were serum starved overnight in a basic medium (EBM‐2) containing 0.5% fetal bovine serum. These cells were then washed with phosphate‐buffered saline, and malignant ascites and a 10‐nM concentration of Tivozanib (AV951; KRN-951) were added to the culture plates. Medium alone or 50 ng/mL VEGF without ascites served as the internal controls. After 7 h incubation, the cells were harvested and treated with propidium iodide (1 µg/mL), and subjected to FACS analysis. Permeability was assessed by measuring the uptake of propidium iodide by the HUVEC. For western blotting, HUVEC were treated in the same way except that the stimulation time with ascites of 10 min. Following cell lysis, VEGFR proteins were immunoprecipitated with an anti‐VEGFR‐2 antibody and then subjected to immunoblotting with an antiphosphotyrosine antibody, as described previously.[2]

The ability of AV-951 to inhibit ligand-dependent phosphorylation of tyrosine kinase receptors is assessed using assays based on human umbilical vein endothelial cells (HUVEC) and normal human dermal fibroblasts. In the proper basic medium with 0.5% fetal bovine serum (FBS), the cells are starved for the duration of the next day. The cells are stimulated with the cognate ligand at 37 °C after being incubated for an hour with either AV-951 or 0.1% DMSO. With the exception of VEGFR3, c-Met, and c-Kit, which are induced for 10 and 15 minutes, respectively, receptor phosphorylation lasts for five minutes. VEGF-C, a rat recombinant protein, is the only ligand utilized in the assays that is not a human recombinant protein. After cell lysis, receptors are phosphotyrosine-immunoblotted after being immunoprecipitated with the proper antibodies. Both the blot quantification and IC50 value computation are completed.

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Endothelial cell proliferation. [1]
HUVECs were seeded in M-199 containing 5% FBS in collagen-coated 96-well plates at a density of 4,000 cells/200 μL/well. After 24 hours, Tivozanib (AV951/KRN-951) was added followed by 20 ng/mL VEGF or 10 ng/mL bFGF, and the cells were cultured for 72 hours. [3H]thymidine (1 μCi/mL) was added and the cells were cultured for a further 12 hours. Cells were then harvested and their radioactivity was measured with a Liquid Scintillation Counter. [1]

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Chemotaxis assay. [1]
HUVEC migration was assessed using 96-well microchamber plates. Cells were starved for 5 hours in EBM-2 containing 0.1% bovine serum albumin (BSA). Then, cells were harvested, resuspended in EBM-2 containing 0.1% BSA, and placed in the upper chamber. Cell migration was initiated by placing medium containing 10 ng/mL VEGF, 0.1% FBS, and 0.1% BSA to the bottom chamber. When indicated, Tivozanib (AV951/KRN-951) was added to both the upper and lower chambers. After 22 hours of incubation, cells were stained with 4 μg/mL calcein AM in HBSS. Fluorescence in the cells that had migrated through the pores of the fluorescence blocking membrane was directly measured through the bottom of the chambers in a fluorescence plate reader at excitation/emission wavelengths of 485/530 nm. [1]

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Cytotoxicity assays. [1]
These assays were done as described previously. Briefly, cells were seeded in 96-well plates and cultured in medium containing 10% FBS. Tivozanib (AV951/KRN-951) was added ∼24 hours after the start of culture and the cells were then incubated for 72 hours. WST-1 reagent was used for the detection of cell viability.[1]

Mice: The athymic rats receive a subcutaneous injection of cancer cells in their right flank. Tumors up to 1,500 mm3 are surgically removed, and smaller pieces (20–30 mg) are s.c. implanted into the right flank of rats exposed to radiation. Beginning on day zero of randomization, oral administration of KRN951 (0.2 or 1 mg/kg) or the vehicle is administered. Using Vernier calipers, tumor volume is measured and computed twice a week.

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Tumor xenograft models. Athymic rats (RH-rnu/rnu) were used. Twenty-four hours after whole body irradiation with a γ-source (7 Gy, Co60), cancer cells were s.c. inoculated into the right flank of the rats. Once established, tumors of ∼1,500 mm3 were surgically excised and smaller tumor fragments (20-30 mg) were s.c. implanted in the right flank of irradiated rats. Oral administration of Tivozanib (AV951/KRN-951) (0.2 or 1 mg/kg) or vehicle was initiated at the day of randomization (day 0). Tumor volume was measured twice weekly with Vernier calipers, and calculated as (length × width2) × 0.5. Relative tumor volume (RTV) was calculated by the formula: RTV at day x = tumor volume at day x / tumor volume at day 0. Percentage tumor growth inhibition (TGI%) was calculated as described previously. Statistical analysis of RTV was done using the unpaired t test.DCE-MRI. Athymic rats (RH-rnu/rnu) were s.c. implanted with fresh Calu-6 tumor fragments. Rats were randomized when the tumors reached a volume of 274 to 287 mm3 (day −1). Once-daily p.o. administration of Tivozanib (AV951/KRN-951) or vehicle was initiated the day after randomization (day 0) and continued for 2 weeks (days 0-13).MRI experiments were carried out at 1.5 T on a whole body magnet equipped with a flexible receiver coil (circularly polarized). DCE-MRI acquisitions were done on day −1 (before the start of treatment), day 2, day 13, and day 21. On days 2 and 13, the rats were imaged 4 hours after p.o. administration of Tivozanib (AV951/KRN-951). The rat tail vein was cannulated for contrast agent injection before placing the animals in the magnet. During the experiment, rats were anesthetized by an i.m. injection of a mixture of ketamine and xylazine (2/1, v/v, 70 and 15 mg/kg, respectively). The anesthetized rats were placed in a cradle supine position inside the resonator. The exact position of the rats was assessed by a scout imaging sequence. [1]

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Measurement of tumor vessel diameter. During the MRI study, an additional three groups of Calu-6 tumor-bearing rats (RH-rnu/rnu, three rats per group) were used for measurement of tumor vessel diameter using the fluorescent dye H33342 (24). Rats were treated with Tivozanib (AV951/KRN-951) (0.2 or 1 mg/kg) or vehicle for 14 days (from day 0 to day 13) and were sacrificed 1 minute after the i.v. injection of H33342 (20 mg/kg) at day 13. The tumors were removed and 10 μm cryosections prepared from five levels of each tumor separated by at least 200 μm. Tumor sections were studied under UV illumination using a Nikon epifluorescence microscope to identify blood vessels with a surrounding halo of fluorescent H33342-labeled cells. The lumen enclosed by the halos was measured as the vessel diameter using Win ROOF software. Statistical analysis was done using the Mann-Whitney test.[1]

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Histologic analysis of smooth muscle actin–positive pericyte coverage of tumor vessels. Calu-6 tumor xenografts were established in athymic rats by s.c. implantation of cells. Rats were randomized when tumor volumes reached an average of 273 to 275 mm3 and then treated p.o. with Tivozanib (AV951/KRN-951) or vehicle for 2 weeks. Immunofluorescence staining of pericytes in tumors was done with Cy3-conjugated monoclonal anti-α-smooth muscle actin antibody following the staining of endothelial cells with anti-CD31 antibody. Tissue images were captured digitally at ×100 magnification with LSM 510 systems. Six fields per section (0.8489 mm2 each) were randomly analyzed, excluding peripheral surrounding connective tissues and central necrotic tissues. The number of CD31-positive objects and those surrounded by the region positive for α-smooth muscle actin were quantified using Win ROOF software after blind-coding the histology slides to avoid operator bias.[1]

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Pharmacokinetic analysis of Tivozanib (AV951/KRN-951). Athymic rats (F344/N JcL-rnu, four females per group) received Tivozanib (AV951/KRN-951) p.o. and blood samples were collected from their tail vein at predetermined intervals up to 72 hours postdose. An appropriate amount of internal standard material, KRN633, was added to each serum sample. Serum samples were deproteinated with acetonitrile and supernatants were analyzed by high-performance liquid chromatography–tandem mass spectrometry. Pharmacokinetic variables were calculated by noncompartmental analysis. The serum concentration of Tivozanib (AV951/KRN-951) at steady state after repeated p.o. administrations of a 0.2 mg/kg dose was simulated as described previously [1]

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Tivozanib (AV951/KRN-951) was suspended in vehicle (0.5% methyl cellulose in distilled water) and stored at 4°C. Fresh solutions were prepared weekly.[2]
Experimental design.  Rats inoculated with RCN‐9 cells were assigned randomly to three groups and given daily oral doses of Tivozanib (AV951/KRN-951) (1 or 3 mg/kg) or a 0.5% methylcellulose vehicle control. These treatments commenced at 4 or 14 days after tumor transplantation, and continued for 10 or 11 days, respectively. At the end of the treatment periods, the rats were killed and tumor progression was evaluated. Ascites were also collected and their volumes were measured. Each transparent window in the mesentery surrounded by fatty tissue was observed microscopically. The percentages of the mesenteric windows with a vasculature and the number of tumor nodules with or without a vasculature on the mesenteric windows were then counted. [2]
In a subsequent survival study, rats inoculated with RCN‐9 cells were assigned randomly to vehicle‐treated or 1 mg/kg Tivozanib (AV951/KRN-951)‐treated groups (n = 10 per group). Separate treatments then commenced from the day of tumor inoculation or at 14 days after this transplantation. The results were plotted using Kaplan–Meier methods and the differences in survival were analyzed by log‐rank test. A P‐value of <0.05 was considered statistically significant. [2]

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Tumor vessel imaging.  RCN‐9 cell‐inoculated rats with and without Tivozanib (AV951/KRN-951) treatment were anesthetized and injected intravenously with fluorescein isothiocyanate‐labeled dextran (molecular weight 200 000). After the animals had been killed, the mesenteries were fixed with 4% paraformaldehyde and placed on glass slides. The vasculature associated with each mesenteric window was then photographed microscopically. The number of vessel joints and paths (as vessel bifurcation characteristics), the areas and lengths of vessels (as the angiogenesis density), and the tortuosity of the vasculatures were recorded objectively and evaluated quantitatively using an angiogenesis image analyzer (Kurabo, Osaka, Japan). Four rats were used in these experiments from each group and 12–15 different fields from each animal were analyzed. [2]

Absorption, Distribution and Excretion
The median time to peak concentration (Tmax) of tevozanib is 10 hours, but it can range from 3 to 24 hours. A pharmacokinetic study in 8 healthy subjects showed that the Cmax and AUC of radiolabeled tevozanib were 12.1 ± 5.67 ng/mL and 1084 ± 417.0 ng·h/mL, respectively. Steady-state concentrations of tevozanib are reached at concentrations 6–7 times the normal dose. Tevozanib is primarily excreted in feces. Following oral administration of 1.34 mg of radiolabeled tevozanib to healthy volunteers, 79% of the administered dose was found in feces (26% of which was the unchanged drug), and 12% was found only as metabolites in urine. The apparent volume of distribution (V/F) of tevozanib is 123 L.
The apparent clearance (CL/F) of tevozanib is approximately 0.75 L/h.

Metabolism/Metabolites

Tevozanib is primarily metabolized via CYP3A4. Following oral administration of 1.34 mg of radiolabeled tevozanib to healthy volunteers, 90% of the radiopharmaceutical detected in serum was unmetabolized tevozanib.

Biological Half-Life

According to prescribing information, the half-life of tevozanib is approximately 111 hours. Clinical study data show a half-life of 4–5 days.

Hepatotoxicity
In published pre-registration clinical trials of tevozanib, the incidence of elevated serum ALT or AST levels ranged from 10% to 29%, with 1% to 4% of treated patients experiencing ALT or AST levels exceeding 5 times the upper limit of normal (ULN). Some clinical trials have reported cases of clinically significant liver injury, including death from liver failure, but all cases were attributed to liver metastases or other underlying liver diseases. Since tevozanib's approval and wider clinical application, no clinically significant liver injury or liver failure has been reported, but its clinical use remains limited. Probability score: E (Unproven but suspected cause of clinically significant liver injury). Protein Binding In vitro studies have shown that tevozanib is primarily bound to albumin, with a binding rate ≥99%.

[1]. Cancer Res. 2006 Sep 15;66(18):9134-42.

[2]. Cancer Sci. 2008 Mar;99(3):623-30.

Tevozanib hydrochloride is the hydrochloride salt of tevozanib, an orally bioavailable inhibitor of vascular endothelial growth factor receptors (VEGFR) 1, 2, and 3 with potential anti-angiogenic and antitumor activities. Tevozanib binds to and inhibits the activity of VEGFR 1, 2, and 3, thereby inhibiting endothelial cell migration and proliferation, suppressing tumor angiogenesis, and leading to tumor cell death. VEGFR tyrosine kinases are frequently overexpressed in various tumor cell types and play a crucial role in angiogenesis.
See also: Tevozanib (containing the active fraction).
Indications
Fotivda is indicated for the first-line treatment of adult patients with advanced renal cell carcinoma (RCC), and for adult patients with advanced RCC whose disease has progressed after prior cytokine therapy and who have not received VEGFR and mTOR pathway inhibitor therapy. It is used to treat advanced renal cell carcinoma. 1-[2-chloro-4-[(6,7-dimethoxy-4-quinolinyl)oxy]phenyl]-3-(5-methyl-3-isoxazolyl)urea is an aromatic ether. Renal cell carcinoma (RCC) accounts for 3% of cancer cases and is one of the top ten most common cancers in adults. The average age of diagnosis is between 65 and 74 years. Tivozanib, also known as FOTIVDA, is a kinase inhibitor used to treat adult patients with relapsed or refractory advanced renal cell carcinoma (RCC) who have failed prior systemic therapy. It was approved by the FDA on March 10, 2021. Tivozanib is marketed by Aveo Oncology and is a promising treatment for RCC patients who have failed other therapies. Tivozanib is a kinase inhibitor. The mechanism of action of tivozanib is as a tyrosine kinase inhibitor. Tivozanib is a small molecule multi-kinase inhibitor used to treat relapsed or refractory renal cell carcinoma. Transient and mild elevations in serum transaminases are common during tevozanib treatment, but no cases have been reported associated with clinically significant acute liver injury. Tevozanib is an orally bioavailable inhibitor of vascular endothelial growth factor receptors (VEGFR) 1, 2, and 3 with potential anti-angiogenic and antitumor activity. Tevozanib binds to and inhibits the activity of VEGFR 1, 2, and 3, which may lead to suppression of endothelial cell migration and proliferation, inhibition of tumor angiogenesis, and tumor cell death. VEGFR tyrosine kinases are frequently overexpressed in various tumor cell types and play a key role in angiogenesis. See also: tevozanib hydrochloride (salt form); tevozanib anhydrous hydrochloride (its active ingredient). Drug Indications Tevozanib is approved in the United States for the treatment of adult patients with relapsed or refractory renal cell carcinoma who have received two or more prior systemic therapies. In the UK and other countries, tevozarib is approved as a first-line treatment for adult patients with advanced renal cell carcinoma (RCC) whose disease has progressed after prior cytokine therapy and who have not received VEGFR and mTOR pathway inhibitors. Fosetida is also approved as a first-line treatment for adult patients with advanced RCC whose disease has progressed after prior cytokine therapy and who have not received VEGFR and mTOR pathway inhibitors. Treatment of advanced renal cell carcinoma. Mechanism of Action: VHL mutation-HIF upregulation-VEGF transcription is a major pathway in the growth of renal cell carcinoma. Vascular endothelial growth factor receptor (VEGFR receptor) is an important target for tyrosine kinase inhibitors, which can inhibit tumor growth. Tevozarib is a tyrosine kinase inhibitor whose mechanism of action is by inhibiting the phosphorylation of vascular endothelial growth factor receptor (VEGFR)-1, VEGFR-2, and VEGFR-3, and by inhibiting other kinases, such as c-kit and platelet-derived growth factor β (PDGFRβ). The above-mentioned effects can inhibit tumor growth and progression, thereby treating renal cell carcinoma. Vascular endothelial growth factor (VEGF) plays a crucial role in tumor angiogenesis by activating the VEGF receptor (VEGFR) tyrosine kinase, stimulating the pro-angiogenic signaling pathway in endothelial cells. Therefore, VEGFR is a highly attractive target in cancer therapy. In this study, we discovered that the quinoline-urea derivative KRN951 is a novel VEGFR tyrosine kinase inhibitor with anti-tumor angiogenesis and anti-tumor growth activities. KRN951 effectively inhibited VEGF-induced VEGFR-2 phosphorylation in endothelial cells with a sub-nanomolar IC50 value (IC50 = 0.16 nmol/L). It also inhibited ligand-induced phosphorylation of platelet-derived growth factor receptor β (PDGFR-β) and c-Kit (IC50 values of 1.72 and 1.63 nmol/L, respectively). KRN951 blocked VEGF-dependent mitogen-activated protein kinase activation and endothelial cell proliferation, but had no effect on VEGF-independent mitogen-activated protein kinase activation and endothelial cell proliferation. Furthermore, it inhibited VEGF-mediated migration of human umbilical vein endothelial cells. Following oral administration to athymic rats, KRN951 reduced microvessel density and attenuated VEGFR-2 phosphorylation levels in tumor endothelial cells. It also demonstrated antitumor activity in various human tumor xenograft models, including lung cancer, breast cancer, colon cancer, ovarian cancer, pancreatic cancer, and prostate cancer. Moreover, dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) analysis showed that a significant reduction in tumor vascular permeability was closely associated with the antitumor activity of KRN951. These findings suggest that KRN951 is a potent drug with anti-angiogenic and antitumor activity, and that DCE-MRI can be used to detect the early efficacy of KRN951 in a clinical setting. KRN951 is currently in Phase I clinical development for the treatment of patients with advanced cancer. [1] We evaluated the antitumor efficacy of KRN951 (a novel vascular endothelial growth factor receptor tyrosine kinase inhibitor) using a rat colon cancer RCN-9 homology model. In this model, tumor cells were transplanted into the peritoneal cavity of F344 rats. KRN951 treatment initiated on day 4 post-transplantation (day 4) significantly inhibited tumor-induced angiogenesis, formation of tumor nodules within the mesenteric window, and accumulation of malignant ascites. Furthermore, KRN951 treatment initiated on day 14 (when angiogenesis and malignant ascites had largely formed) led to the regression of abnormal neovascularization and significant disappearance of malignant ascites by the end of the study. Quantitative analysis of mesenteric window vascular structures showed that KRN951 not only regressed tumor-induced neovascularization but also restored it to normal. Compared with the carrier control group, continuous daily administration of KRN951 significantly prolonged the survival of rats with early and late-stage tumors. Our current findings suggest that KRN951 inhibits the progression of intraperitoneal colon cancer and prolongs patient survival by suppressing tumor angiogenesis, ascites formation, and tumor spread. Furthermore, these studies clearly demonstrate that KRN951 has a therapeutic effect on established abdominal tumors, including the regression and normalization of tumor angiogenesis. Therefore, our findings suggest that KRN951 has great potential as a future treatment for peritoneal cancer with ascites. [2]

C22H22CL2N4O6

509.339283466339

508.092

682745-41-1

Tivozanib;475108-18-0; 682745-40-0 (hydrate); 682745-41-1 (HCl hydrate)

11547978

White to off-white solid powder

5.811

4

8

6

34

631

0

RQXMKRRBJITKRN-UHFFFAOYSA-N

InChI=1S/C22H19ClN4O5.ClH.H2O/c1-12-8-21(27-32-12)26-22(28)25-16-5-4-13(9-15(16)23)31-18-6-7-24-17-11-20(30-3)19(29-2)10-14(17)18;;/h4-11H,1-3H3,(H2,25,26,27,28);1H;1H2

1-[2-chloro-4-(6,7-dimethoxyquinolin-4-yl)oxyphenyl]-3-(5-methyl-1,2-oxazol-3-yl)urea;hydrate;hydrochloride

TIVOZANIB HYDROCHLORIDE; 682745-41-1; Tivozanib HCl hydrate; Tivozanib hydrochloride hydrate; UNII-8A9H4VK35Z; fotivda; 8A9H4VK35Z; Tivozanib hydrochloride [USAN];

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

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

DMSO : ~250 mg/mL (~490.83 mM)

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.)