PDGFR; WT KIT (IC50 = 4 nM); D816H KIT (IC50 = 5 nM); V654A KIT (IC50 = 8 nM); D816V KIT (IC50 = 14 nM)

c-Kit-IN-1 is an inhibitor of c-Kit and c-Met, taken from patent 2010051373A1, compound example 45, IC50 <200 nM. c-Kit-IN-1 also suppresses KDR, PDGFR α and β, with IC50 of <2 μM, <10 μM and <10 μM respectively[1].

In the GIST T1 xenograft model, DCC-2618 administration at 50 mg/kg results in an ED90 for KIT phosphorylation inhibition, which corresponds to an EC90 concentration of roughly 470 ng/mL. This oral dose causes nearly total tumor stasis when taken twice a day. In a KIT exon 17 N822K AML xenograft model and a patient-derived xenograft (PDX) GIST expressing KIT exon 11 delW557K558/exon 17 Y823D, this dosage of DCC-2618 results in tumor regressions[1]. DCC-2618 inhibits PDGFRA- and KIT-driven tumor growth in xenograft studies, including KIT exon 17 mutants present in AML (N822K), GIST (Y823D), and mastocytosis (D816V) models[3].

In order to assess KIT and BTK signaling, ROSAKIT WT, ROSAKIT D816V, HMC-1.1, and HMC-1.2 cells were incubated for 4 hours at 37°C in either control medium or DCC-2618 (0.5–5 μM). Western blotting was done essentially according to other instructions. In order to assess the downstream signaling pathways of KIT, HMC-1.1, HMC-1.2, ROSAKIT WT, and ROSAKIT D816V cells were initially pre-cultured for an entire night in Iscove-modified Dulbecco medium that was devoid of stem cell factor and fetal calf serum. Then, for 90 minutes at 37°C, DCC-2618 (0.001–10 μM) was applied to 106 cells from each line. Following the course of treatment, ROSAKIT WT cells were stimulated for 10 minutes at room temperature using 10% of the supernatants of Chinese hamster ovary cells transfected with the murine scf (kl) gene (CHO-KL). Western blotting was then carried out essentially in the same manner as previously mentioned.

In order to assess KIT and BTK signaling, HMC-1.1, HMC-1.2, ROSA (KIT WT), and ROSA (KIT D816V) cells are incubated for 4 hours at 37°C in either control medium or DCC-2618 (0.5–5 μM). One method used is western blotting.

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Ripretinib (DCC-2618) was tested for inhibition of KIT isoforms using a standard PK/LDH coupled spectrophotometric assay. CHO cells were transiently transfected to express mutant KIT or PDGFRα constructs. Transfected cells were treated with a range of DCC-2618 and levels of phosphorylated KIT or PDGFRα in cell lysates were determined by ELISA or western blot. Cell proliferation of several cell lines was measured using the fluorescent dye resazurin. Experiments were performed in triplicate.[1]

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Western blotting[2]
For evaluation of KIT and BTK signaling, HMC-1.1, HMC-1.2, ROSAKIT WT and ROSAKIT D816V cells were incubated in control medium or in Ripretinib (DCC-2618) (0.5–5 μM) for 4 h at 37°C. Western blotting was performed essentially as described elsewhere. For evaluation of downstream signaling pathways of KIT, HMC-1.1, HMC-1.2, ROSAKIT WT and ROSAKIT D816V cells were first pre- incubated overnight in Iscove modified Dulbecco medium devoid of fetal calf serum and of stem cell factor. Cells (106) from each line were then treated with DCC-2618 (0.001–10 μM) for 90 min at 37°C. At the end of the treatment, ROSAKIT WT cells were stimulated with stem cell factor-containing supernatants (10%) of Chinese hamster ovary cells transfected with the murine scf (kl) gene (CHO-KL) at room temperature for 10 min. Thereafter, Western blotting was performed essentially as described previously.

xenograft models (mice)
100 mg/kg/day or 25 mg/kg/day or 50 mg/kg BID
oral

Absorption
Ripretinib is absorbed from the gastrointestinal tract, with a peak time (Tmax) of 4 hours. Steady-state plasma concentrations are reached within 14 days.
Excretion
34% of ripretinib is excreted in feces, and 0.2% in urine.
Volume of Distribution
The mean volume of distribution of ripretinib is 307 liters.
Clearance
The mean apparent clearance of ripretinib is 15.3 liters/hour.

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Metabolism/Metabolites
Ripretinib is primarily metabolized by CYP3A subfamily enzymes. The formation of its active metabolite DP-5439 is also involved by CYP2D6 and CYP2E1.

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Biological Half-Life
The mean half-life of ripretinib is 14.8 hours.

Hepatotoxicity
In premarketing placebo-controlled clinical trials in patients with refractory and extensively treated gastrointestinal stromal tumors (GIST), 13% of patients in the ripretinib treatment group experienced elevated alanine aminotransferase (ALT), compared to 5% in the placebo group. ALT elevations were generally transient and mild, with only 1% of treated patients experiencing ALT elevations exceeding 5 times the upper limit of normal (ULN), requiring no dose adjustment or discontinuation. Elevated bilirubin was reported in 22% of patients in the ripretinib treatment group, compared to only 7.5% in the placebo group. Bilirubin elevations were transient and mild, but their timing, severity, and whether the bilirubin was bound or unbound (direct or indirect) were not characterized. No clinically significant liver injury, liver failure, or death due to liver injury has been reported in the open-label and controlled trials supporting ripretinib approval. Since ripretinib's approval in the US and Europe, no clinically significant cases of liver injury related to ripretinib treatment have been reported.
Likelihood Score: E (Unlikely to be a clinically significant cause of liver injury).

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Effects during pregnancy and lactation
◉ Overview of use during lactation
There is currently no information regarding the use of ripretinib during lactation. Because ripretinib and its metabolites bind to plasma proteins at a rate exceeding 99%, the concentration in breast milk may be low. However, they have a long half-life. The manufacturer recommends that mothers should not breastfeed during ripretinib treatment and for one week after the last dose.
◉ Effects on breastfed infants
As of the revision date, no relevant published information was found.
◉ Effects on lactation and breast milk
As of the revision date, no relevant published information was found.

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Protein binding
Ripretinib binds to albumin and α1-acid glycoprotein at a rate exceeding 99%.

[1]. Cancer Res (2015) 75 (15_Supplement): 2690.

[2]. Haematologica . 2018 May;103(5):799-809.

[3]. AACR Annual Meeting. 2018: Abstract 3925; Poster Section 39, Board 5.

Ripretinib is a kinase inhibitor used to treat advanced gastrointestinal stromal tumors (GIST) that have not responded to other kinase inhibitors, such as sunitinib and imatinib. Ripretinib, also known as Qinlock, is manufactured by Deciphera Pharmaceuticals and was first approved by the U.S. Food and Drug Administration (FDA) on May 15, 2020. It was the first approved fourth-line treatment for patients who have received at least three other kinase inhibitors. Ripretinib is a kinase inhibitor. Its mechanism of action is as a stem cell factor (KIT) receptor inhibitor, a platelet-derived growth factor α receptor inhibitor, a cytochrome P450 2C8 inhibitor, a P-glycoprotein inhibitor, and a breast cancer resistance protein inhibitor. Ripretinib is a multi-kinase inhibitor used to treat refractory advanced GIST. A small number of patients receiving ripretinib may experience elevated serum transaminases, but there have been no reports of clinically significant liver damage with jaundice due to ripretinib use.
Ripretinib is an orally bioavailable switch-pocket control inhibitor that inhibits both wild-type and mutant forms of tumor-associated antigen (TAA) mast cell/stem cell factor receptor (SCFR) KIT and platelet-derived growth factor receptor α (PDGFR-α; PDGFRα), exhibiting potential antitumor activity. After oral administration, ripretinib specifically targets and binds to the switch-pocket binding sites of both wild-type and mutant KIT and PDGFRα, thereby preventing the conversion of these kinases from their inactive to active conformations and inactivating both wild-type and mutant forms. This blocks KIT/PDGFRα-mediated tumor cell signaling and inhibits KIT/PDGFRα-driven cancer proliferation. DCC-2618 also inhibits a variety of other kinases, including vascular endothelial growth factor receptor type 2 (VEGFR2; KDR), angiopoietin-1 receptor (TIE2; TEK), PDGFR-β, and macrophage colony-stimulating factor 1 receptor (FMS; CSF1R), thereby further inhibiting tumor cell growth. KIT and PDGFRα are tyrosine kinase receptors, upregulated or mutated in various cancer cell types; mutated forms play a key role in the regulation of tumor cell proliferation and chemotherapy resistance.

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Drug Indications
Repretinib is indicated for the treatment of adult patients with advanced gastrointestinal stromal tumors (GIST) who have previously received at least three kinase inhibitors (including imatinib).
Zinlock is indicated for the treatment of adult patients with advanced gastrointestinal stromal tumors (GIST) who have previously received three or more kinase inhibitors (including imatinib).

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Pharmacodynamics
Ripretinib, as a broad-spectrum kinase inhibitor, is a kinase inhibitor that inhibits multiple gene mutations, thereby prolonging progression-free survival in patients with advanced gastrointestinal stromal tumors (GIST). It is also effective against mutations resistant to chemotherapy with other kinase inhibitors, such as imatinib. Ripretinib carries the risk of causing cardiac dysfunction and new primary skin malignancies. Therefore, measuring cardiac ejection fraction before and during treatment, as well as regular dermatological evaluations, are crucial.

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Mechanism of Action
Protein kinases play an important role in cellular function, and their dysregulation can lead to carcinogenesis. Ripretinib inhibits multiple protein kinases, including wild-type and mutant platelet-derived growth factor receptor A (PDGFRA) and KIT, which are pathogenic factors in most gastrointestinal stromal tumors (GISTs). In vitro studies have shown that ripretinib inhibits the PDGFRB, BRAF, VEGF, and TIE2 genes. Ripretinib binds to the KIT and PDGFRA receptors; KIT mutations are located in exons 9, 11, 13, 14, 17, and 18, and PDGFRA mutations are located in exons 12, 14, and 18. The "switch pocket" of a protein kinase normally binds to the activation loop, acting as a kinase "switch." Ripretinib has a unique dual mechanism of action; it binds to both the kinase's switch pocket and the activation loop, thereby shutting down the kinase and its ability to cause dysregulation of cell growth. DCC-2618 inhibits multiple KIT mutants with nanomolar potency: wild-type (IC50 4 nM), V654A (8 nM), T670I (18 nM), D816H (5 nM), and D816V (14 nM). In CHO cells transiently transfected with single-mutant and double-mutant (primary/secondary) KIT mutants, DCC-2618 effectively inhibits KIT exon 17, exon 9/13, exon 9/14, and exon 9/17 mutants, as well as KIT exon 11/17 mutants, including primary or secondary mutations in exon 17 such as D816V, D816G, D820A, D820E, D820Y, N822K, N822Y, N822H, and Y823D. DCC-2618 inhibits phosphorylation of wild-type KIT in the MO7e cell line (IC50 36 nM). DCC-2618 effectively inhibits KIT activation in human gastrointestinal stromal tumor (GIST) cell lines, including GIST T1 (exon 11 deletion, IC50 2 nM), GIST 430 (exon 11 deletion/exon 13 V654A, IC50 7 nM), and GIST 48 (exon 11 V560D/exon 17 D820A, IC50 53 nM). In the mouse mast cell proliferation P815 cell line expressing the exon 17 D816Y mutation, DCC-2618 effectively inhibits cell proliferation (IC50 2 nM). In in vivo experiments, in a GIST T1 xenograft model, a dose of 50 mg/kg of DCC-2618 inhibited KIT phosphorylation, with an ED90 value of approximately 470 ng/mL. This dose, administered orally twice daily, almost completely halted tumor growth. This dose of DCC-2618 also induced tumor regression in a patient-derived xenograft (PDX) model of gastrointestinal stromal tumor (GIST) expressing the KIT exon 11 delW557K558/exon 17 Y823D mutation, and in an acute myeloid leukemia (AML) xenograft model expressing the KIT exon 17 N822K mutation. Conclusion: DCC-2618 is a potent inhibitor of both single and double mutations in KIT, characterized by a pairing of primary exon 9 or exon 11 mutations with secondary mutations in exons 13, 14, or 17. DCC-2618 inhibits exon 17 mutations, including the D816V mutation, which is not inhibited by currently marketed KIT inhibitors. DCC-2618 has the potential to treat KIT mutation-driven cancers, including gastrointestinal stromal tumors, systemic mastocytosis, acute myeloid leukemia, or melanoma. DCC-2618 has been selected for formal IND application for clinical development. [1] Systemic mastocytosis is a complex disease characterized by the abnormal proliferation and accumulation of neoplastic mast cells in various organs. Most patients carry the D816V mutation variant of the KIT gene, which confers resistance to imatinib. The clinical problems of systemic mastocytosis stem from mediator-related symptoms and/or organ destruction caused by the malignant proliferation of neoplastic mast cells and/or other myeloid cells in various organ systems. DCC-2618 is a spectrum-selective pan-KIT and PDGFRA inhibitor that blocks the KIT D816V mutation as well as a number of other kinase targets associated with systemic mastocytosis. We found that DCC-2618 inhibited the proliferation and survival of various human mast cell lines (HMC-1, ROSA, MCPV-1) and primary neoplastic mast cells obtained from patients with advanced systemic mastocytosis (SMD) (IC50 <1 μM). Furthermore, DCC-2618 reduced the growth and survival of primary neoplastic eosinophils isolated from patients with SMD or eosinophilic leukemia, leukemic mononuclear cells isolated from patients with chronic myelomonocytic leukemia with or without SMD, and primitive cells isolated from patients with acute myeloid leukemia. In addition, the study found that DCC-2618 inhibited endothelial cell proliferation, suggesting its potential role in SMD-related angiogenesis. Finally, the study found that DCC-2618 downregulated IgE-mediated histamine release from basophils and trypsin release from mast cells. In summary, DCC-2618 can inhibit the growth, survival, and activation of various cell types associated with advanced systemic mastocytosis. Currently, the efficacy of DCC-2618 in patients with advanced systemic mastocytosis is being investigated in clinical trials. [2]

C26H21F2N5O3

489.4735

489.161

1225278-16-9

1442472-39-0

46208890

White to off-white solid powder

1.40±0.1 g/cm3 (20 ºC 760 Torr)

5.632

2

7

7

36

794

0

WWOXKWLDMLMYQY-UHFFFAOYSA-N

InChI=1S/C26H21F2N5O3/c1-33-15-16(14-30-33)21-11-18(7-10-29-21)36-23-13-19(27)22(12-20(23)28)32-25(35)26(8-9-26)24(34)31-17-5-3-2-4-6-17/h2-7,10-15H,8-9H2,1H3,(H,31,34)(H,32,35)

1-N'-[2,5-difluoro-4-[2-(1-methylpyrazol-4-yl)pyridin-4-yl]oxyphenyl]-1-N-phenylcyclopropane-1,1-dicarboxamide

1225278-16-9; c-Kit-IN-1; 1-N'-[2,5-difluoro-4-[2-(1-methylpyrazol-4-yl)pyridin-4-yl]oxyphenyl]-1-N-phenylcyclopropane-1,1-dicarboxamide; N-(2,5-Difluoro-4-((2-(1-methyl-1H-pyrazol-4-yl)pyridin-4-yl)oxy)phenyl)-N-phenylcyclopropane-1,1-dicarboxamide; N'1-(2,5-DIFLUORO-4-{[2-(1-METHYLPYRAZOL-4-YL)PYRIDIN-4-YL]OXY}PHENYL)-N1-PHENYLCYCLOPROPANE-1,1-DICARBOXAMIDE; PDGFR inhibitor 1; SCHEMBL2450218; CHEMBL4303619;

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 (~204.30 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.11 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 (5.11 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 (5.11 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.)