FGFR1 (IC50 = 0.9 nM); FGFR2 (IC50 = 1.4 nM); FGFR3 (IC50 = 1 nM); FGFR4 (IC50 = 60 nM)

Infigratinib is given to athymic nude mice that have had RT112/luc1 tumors implanted subcutaneously. The administration options include an intravenous bolus of 5 mg/kg in NMP/PEG200 (1:9, v/v) or an oral gavage of a suspension in PEG300/D5W (2:1, v/v) at a dose of 20 mg/kg. According to the pertinent pharmacokinetic (PK) parameters, infigratinib has a 32% oral bioavailability in this investigation. Following intravenous administration, infigratinib exhibits a high volume of distribution (26 L/kg) due to its quick distribution from the vascular compartment into the peripheral tissues. With a clearance of 3.3 L/h/kg (61% of liver blood flow), the plasma level is high. Based on AUC, the ratio of tumor to plasma following oral dosing is found to be 10[1]. The growth of endometrial cancer xenograft models with FGFR2 mutations is markedly inhibited by infigratinib (30 mg/kg)[2].

The purified GST-fusion FGFR3-K650E kinase domain phosphorylates a synthetic substrate in the presence of radiolabeled ATP to measure the enzymatic kinase activity. Enzyme activities are determined by combining 10 μL of the corresponding substrate mixture (peptidic substrate, ATP, and [γ³³P]ATP) with 10 μL of a 3-fold concentrated Infigratinib solution or control. The assay buffer is mixed with 10 μL of a concentrated enzyme solution three times over to start the reactions. The following are the assay components' final concentrations: 0.5 μM ATP (γ-[³³P]-ATP 0.1 μCi), 3 mM MnCl₂, 3 mM MgCl₂, 1 mM DTT, 250 μg/mL PEG 20000, 2 μg/mL poly(EY) 4:1, 1% DMSO, and 10 ng of GST-FGFR3-K650E were added. The assay is performed using the filter binding (FB) method in 96-well plates for 10 minutes at room temperature and 30 μL of final volume, which includes the components mentioned above. The following measurement is used to determine the amount of ³³P incorporated into the polypeptidic substrates after 20 μL of 125 mM EDTA is added to stop the enzymatic reactions: Using a disconnected vacuum source, mount the Immobilon-PVDF membranes on a vacuum manifold and transfer 30 μL of the stopped reaction mixture onto them after they have been soaked in methanol for 5 minutes, rinsed with water, and soaked in 0.5% H₃PO₄ for 5 minutes. Following spotting, each well is rinsed with 200 μL of 0.5% H₃PO₄ and vacuumed. After extracting the free membranes, they are shaker-washed four times with 1% H₃PO₄ and once with ethanol. After desiccating the membranes, 10 μL of a scintillation fluid are added per well. In the end, the plates are sealed and counted using a microplate scintillation counter. By using linear regression analysis to determine the percentage inhibition of NVP-BGJ398[1], IC50 values are determined.

RPMI-1640 medium supplemented with 10% FBS, 4.5 g/L glucose, 1.5 g/L sodium bicarbonate, and Pen/Strep is used to cultivate mouse BaF3 cell lines. Twice a week, cells are passed through. A Luciferase bioluminescent assay is used to evaluate compound-mediated inhibition of BaF3 cell proliferation and viability. Using a μFill liquid dispenser, exponentially growing BaF3 or BaF3 Tel-TK cells are seeded at 50 μL/well into 384-well plates (4250 cells/well) in fresh medium. After being serially diluted in DMSO, infigratinib is arranged in a 384-well polypropylene plate. The plates were then incubated at 37°C (5% CO₂) for 48 hours after 50 nL of the compound was transferred into them using the pintool transfer device. Next, add 25 μL of Bright-Glo, and use an Analyst-GT to measure the luminescence. A logistic fit of the percent cell viability as a function of the logarithm of inhibitor concentration is generated using specialized curve-fitting software. The concentration of a compound required to lower cell viability to 50% of a DMSO control is known as the IC50 value[1].

Mice: HsdNpa female: Athymic Nude-nu mice are employed. Infigratinib is taken orally for 12 straight days at doses of 10 and 30 mg/kg/qd. It is prepared as a suspension in PEG300/D5W (2:1, v/v). ANOVA is used to analyze the tumor and body weight data, and Dunnett's test is used to compare the treatment group to the control group post hoc. For intragroup comparison, the post hoc Tukey test is employed. To perform statistical analysis, use GraphPad Prism 4.02. One calculates the T/C (%) value as a measure of efficacy.
Rats: Woman in the nude 6 to 9-week-old Rowett rats are utilized. The tumor-bearing rats (n=8) receive infigratinib intraperitoneally (gavage) once a day for 20 days at doses of 5, 10, and 15 mg/kg/qd (free base equivalents). The drug is prepared as a solution in acetic acid-acetate buffer pH 4.6/PEG300 (1:1, v/v). It uses five milliliters per kilogram. The formula for determining tumor volumes is length×width×height×π/6, which can be measured using calipers. Antitumor activity is represented as T/C (%), which is calculated as (mean change in tumor volume of treated animals / mean change in tumor volume of control animals)×100. One calculates regressions (%).

Absorption, Distribution and Excretion
Absorption
Mean (%CV) Cmax is 282.5 ng/mL (54%) and AUC0-24h is 3780 ngxh/mL (59%) for infigratinib. Infigratinib Cmax and AUC increase more than proportionally across the dose range of 5 to 150 mg and steady state is achieved within 15 days. At steady state, median time to achieve peak infigratinib plasma concentration (Tmax) is six hours, with a range between two and seven hours. Mean (%CV) Cmax is 42.1 ng/mL (65%) for BHS697 and 15.7 ng/mL (92%) for CQM157. Mean (%CV) AUC0-24h is 717 ngxh/mL (55%) for BHS697 and 428 ngxh/mL (72%) for CQM157. In healthy subjects, a high-fat and high-calorie meal increased AUCinf of infigratinib by 80%-120% and Cmax by 60%-80%. The median Tmax also shifted from four hours to six hours. A low-fat low-calorie meal increased the mean AUCinf of infigratinib by 70% and Cmax by 90%/

Route of Elimination
Following administration of a single oral dose of radiolabeled infigratinib in healthy subjects, approximately 77% of the dose was recovered in feces, where 3.4% of the dose was in the unchanged parent form. About 7.2% was recovered in urine with 1.9% of the dose was unchanged.

Volume of Distribution
At steady state, the geometric mean (CV%) apparent volume of distribution of infigratinib was 1600 L (33%). In rats receiving a single oral dose, infigratinib had brain-to-plasma concentration ratios (based on AUC0-inf) of 0.682.

Clearance
The geometric mean (CV%) total apparent clearance (CL/F) of infigratinib was 33.1 L/h (59%) at steady state.

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Metabolism / Metabolites
According to _in vitro_ findings, about 94% of infigratinib is metabolized by CYP3A4 and about 6% of the drug is metabolized by flavin-containing monooxygenase 3 (FMO3). About 38% of the dose is circulating parent drug in the plasma and BHS697 and CQM157 are two major metabolites of infigratinib that are each found at >10% of the dose. They are pharmacologically active, with BHS697 representing about 16% to 33% of the overall pharmacological activity of infigratinib and CQM157 contributing to about 9% to 12%. BHS697 undergoes further metabolism mediated by CYP3A4 and CQM157 is metabolized through both Phase I and Phase II biotransformation pathways. The exact metabolic pathways and the structure of BHS697 and CQM157 are not fully characterized.

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Biological Half-Life
The geometric mean (CV%) terminal half-life of infigratinib was 33.5 h (39%) at steady state.

Hepatotoxicity
In the open label clinical trials of infigratinib for advanced or metastatic cholangiocarcinoma, adverse events were common and led to dose interruptions in 64%, dose reductions in 60% of patients, and permanent discontinuations in 15% largely for hyperphosphatemia, infections and sepsis rather than liver injury. In preregistration trials in 108 patients, ALT elevations arose in 51% and to above 5 times ULN in 6%. The elevations were typically self-limited and resolved rapidly with or without dose adjustments. No patients developed clinically apparent liver injury or jaundice. Since its approval, there have been no reports clinically apparent liver injury attributed to infigratinib. However, the total clinical experience with its use has been limited and the frequency of serum aminotransferase elevations during therapy suggest that clinically significant liver injury may occur.
Likelihood score: E* (unproven but possible, rare cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Infigratinib is no longer marketed in the US. No information is available on the clinical use of infigratinib during breastfeeding. Because infigratinib is 96.8% bound to plasma proteins, the amount in milk is likely to be low. However, because of its potential toxicity in the breastfed infant and its half-life of 33.5 hours, the manufacturer recommends that breastfeeding be discontinued during infigratinib therapy and for 1 month 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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Protein Binding
Infigratinib is about 96.8% bound to plasma proteins, primarily to lipoprotein. Protein binding is concentration-dependent.

[1]. Discovery of 3-(2,6-dichloro-3,5-dimethoxy-phenyl)-1-{6-[4-(4-ethyl-piperazin-1-yl)-phenylamino]-pyrimidin-4-yl}-1-methyl-urea (NVP-BGJ398), a potent and selective inhibitor of the fibroblast growth factor receptor family of receptor tyrosine kinase. J Med Chem . 2011 Oct 27;54(20):7066-83.

[2]. Activity of the fibroblast growth factor receptor inhibitors dovitinib (TKI258) and NVP-BGJ398 in human endometrial cancer cells. Mol Cancer Ther. 2013 May;12(5):632-42.

Infigratinib Phosphate is the phosphate salt form of infigratinib, an orally bioavailable pan-inhibitor of human fibroblast growth factor receptors (FGFRs) with potential antiangiogenic and antineoplastic activities. Upon administration, infigratinib selectively binds to and inhibits the activities of FGFRs, which may result in the inhibition of angiogenesis and cell proliferation, and the induction of cell death in tumors with activating FGFR amplifications, mutations, or fusions. FGFRs are a family of receptor tyrosine kinases that are involved in tumor cell differentiation and proliferation, tumor angiogenesis, and tumor cell survival. Activating FGFR amplifications, mutations, or fusions occur in various cancer cell types.
See also: Infigratinib (has active moiety).

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A novel series of N-aryl-N'-pyrimidin-4-yl ureas has been optimized to afford potent and selective inhibitors of the fibroblast growth factor receptor tyrosine kinases 1, 2, and 3 by rationally designing the substitution pattern of the aryl ring. On the basis of its in vitro profile, compound 1h (NVP-BGJ398) was selected for in vivo evaluation and showed significant antitumor activity in RT112 bladder cancer xenografts models overexpressing wild-type FGFR3. These results support the potential therapeutic use of 1h as a new anticancer agent.[1]

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The recent identification of activating fibroblast growth factor receptor 2 (FGFR2) mutations in endometrial cancer has generated an opportunity for a novel target-based therapy. Here, we explore the therapeutic potential of 2 FGFR inhibitors, the multikinase inhibitor dovitinib (TKI258) and the more selective FGFR inhibitor NVP-BGJ398 for the treatment of endometrial cancer. We examined the effects of both inhibitors on tumor cell growth, FGFR2 signaling, cell cycle, and apoptosis using a panel of 20 molecularly characterized human endometrial cancer cell lines. Anchorage-independent growth was studied using soft agar assays. In vivo studies were conducted using endometrial cancer xenograft models. Cell lines with activating FGFR2 mutations (S252W, N550K) were more sensitive to dovitinib or NVP-BGJ398 when compared with their FGFR2 wild-type counterparts (P = 0.073 and P = 0.021, respectively). Both agents inhibited FGFR2 signaling, induced cell-cycle arrest, and significantly increased apoptosis in FGFR2-mutant lines. In vitro, dovitinib and NVP-BGJ398 were both potent at inhibiting cell growth of FGFR2-mutant endometrial cancer cells, but the activity of dovitinib was less restricted to FGFR2-mutant lines when compared with NVP-BGJ398. In vivo, dovitinib and NVP-BGJ398 significantly inhibited the growth of FGFR2-mutated endometrial cancer xenograft models. In addition, dovitinib showed significant antitumor activity in FGFR2 wild-type endometrial cancer xenograft models including complete tumor regressions in a long-term in vivo study. Dovitinib and NVP-BGJ398 warrant further clinical evaluation in patients with FGFR2-mutated endometrial cancer. Dovitinib may have antitumor activity in endometrial cancer beyond FGFR2-mutated cases and may permit greater flexibility in patient selection.[2]

C26H34CL2N7O7P

658.47

657.163

C, 47.43; H, 5.20; Cl, 10.77; N, 14.89; O, 17.01; P, 4.70

1310746-10-1

Infigratinib;872511-34-7

56669626

White to off-white solid powder

4.574

5

12

8

43

773

0

ClC1C(=C([H])C(=C(C=1N([H])C(N(C([H])([H])[H])C1C([H])=C(N=C([H])N=1)N([H])C1C([H])=C([H])C(=C([H])C=1[H])N1C([H])([H])C([H])([H])N(C([H])([H])C([H])([H])[H])C([H])([H])C1([H])[H])=O)Cl)OC([H])([H])[H])OC([H])([H])[H].P(=O)(O[H])(O[H])O[H]

GUQNHCGYHLSITB-UHFFFAOYSA-N

InChI=1S/C26H31Cl2N7O3.H3O4P/c1-5-34-10-12-35(13-11-34)18-8-6-17(7-9-18)31-21-15-22(30-16-29-21)33(2)26(36)32-25-23(27)19(37-3)14-20(38-4)24(25)28;1-5(2,3)4/h6-9,14-16H,5,10-13H2,1-4H3,(H,32,36)(H,29,30,31);(H3,1,2,3,4)

3-(2,6-dichloro-3,5-dimethoxyphenyl)-1-[6-[4-(4-ethylpiperazin-1-yl)anilino]pyrimidin-4-yl]-1-methylurea;phosphoric acid

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, avoid exposure to moisture.

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