HER1 (IC50 = 7 nM); HER2 (IC50 = 2 nM); HER4 (IC50 = 4 nM)
Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) potently inhibits epidermal growth factor receptor (EGFR) tyrosine kinase with an IC₅₀ of 27 nM and ErbB2 (HER2) tyrosine kinase with an IC₅₀ of 18 nM [2]
It shows no significant inhibitory activity against VEGFR2, PDGFRβ, or c-Src (IC₅₀ > 1000 nM) [2]

In vitro activity: ARRY334543 behaves as a reversible ATP-competitive inhibitor with nanomolar potency (Ki=1 nM) both in vitro and in cell-based proliferation assays using A431 and BT-474 cells. ARRY334543 is an AKT pathway inhibitor in cells that contains active ErbB-2 receptors. ARRY334543 suppresses ErbB-2 and ErbB-1 phosphorylation in human cancer cells, such as BT-474 with IC50 of 43 nM) and A431 with IC50 of 36 nM) that overexpress ErbB-2 and ErbB-1, respectively

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Kinase Assay: Varlitinib (ARRY-334543; ASLAN001) is a novel, potent, reversible, small molecule pan-EGFR inhibitor with IC50s of 7, 2, 4 nM for HER1, HER2 and HER4, respectively.

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Cell Assay: In cell-based assays using tumor cells that over-express EGFR (A431) or ErbB-2 (BT474), Varlitinib (ARRY-334543) potently inhibits substrate phosphorylation. Varlitinib is shown to be highly selective for EGFR/ErbB-2, and does not show any significant activity when screened against a panel of 104 kinases

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Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) dose-dependently inhibited the proliferation of EGFR/ErbB2-overexpressing tumor cell lines, including SK-BR-3 (breast cancer, IC₅₀ = 0.12 μM), BT474 (breast cancer, IC₅₀ = 0.15 μM), and HepG2 (hepatocellular carcinoma, IC₅₀ = 0.2 μM). It blocked EGFR/ErbB2 phosphorylation and downstream AKT/ERK1/2 signaling at concentrations ≥ 0.2 μM [2]
In patient-derived hepatocellular carcinoma (HCC) cells, the drug (0.1-0.5 μM) suppressed cell viability by 40%-60% and induced apoptosis, as evidenced by increased cleaved caspase-3 and PARP expression [1]
It inhibited the clonogenicity of ErbB2-amplified breast cancer cells (IC₅₀ = 0.08 μM) and reduced the expression of anti-apoptotic protein Bcl-2 [2]

Varlitinib (ARRY-334543) treatment significantly slows the growth of tumors; full tumor regression is seen at dosages of 100 mg/kg twice daily. The phosphorylation of HER1-3, RAS/RAF/MEK/MAPK, p70S6K, S6 ribosomal, 4EBP1, Cdk-2, Cdc-2, and retinoblastoma is significantly inhibited after five days of Varlitinib treatment. Treatment with varlitinib causes survivin to significantly decrease while Caspase 3 cleavage products increase simultaneously[1]. Varlitinib (ARRY-334543), when taken orally twice a day for 21 days, significantly inhibits the growth of A431-derived tumors in murine xenograft models at doses of 25, 50, and 100 mg/kg[2].

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Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) significantly inhibited tumor growth in patient-derived HCC xenograft models. Oral administration of 40 mg/kg/day for 28 days reduced tumor volume by ~65% compared to the control group, and intratumoral EGFR/ErbB2 phosphorylation was downregulated by ~70% [1]
In nude mice bearing SK-BR-3 xenografts, the drug (50 mg/kg/day, oral for 21 days) achieved a tumor growth inhibition rate of 72% and prolonged median survival by 35% [2]
It exhibited good tumor penetration, with a tumor-to-plasma concentration ratio of 2.3 at 4 hours post-administration [2]

Varlitinib (ARRY-334543; ASLAN001) is a novel, potent, reversible, small molecule pan-EGFR inhibitor with IC50s of 7, 2, 4 nM for HER1, HER2 and HER4, respectively.

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Recombinant EGFR and ErbB2 kinase domains were individually incubated with serial dilutions of Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) (0.001-100 μM) in kinase buffer containing ATP and a specific peptide substrate. The reaction was conducted at 37°C for 60 minutes, and phosphorylated substrates were detected using a radiometric assay. Inhibition rates were calculated by comparing radioactivity with vehicle controls, and IC₅₀ values were derived from dose-response curves [2]
To assess selectivity, recombinant VEGFR2, PDGFRβ, and c-Src kinase domains were tested using the same protocol. Reaction conditions were identical, and IC₅₀ values were determined to confirm preferential targeting of EGFR and ErbB2 [2]

Varlitinib (ARRY-334543) potently inhibits substrate phosphorylation in cell-based assays with tumor cells that overexpress EGFR (A431) or ErbB-2 (BT474). When tested against a panel of 104 kinases, varlitinib is found to be highly selective for EGFR/ErbB-2 and to exhibit no discernible activity.

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SK-BR-3, BT474, HepG2, and patient-derived HCC cells were seeded in 96-well plates at 5×10³ cells/well and treated with Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) (0.05-1 μM) for 72 hours. Cell viability was measured using a tetrazolium-based assay to calculate IC₅₀ values [1,2]
For Western blot analysis, cells were treated with 0.1-0.3 μM drug for 24 hours, lysed, and probed with antibodies against phosphorylated EGFR, ErbB2, AKT, ERK1/2, cleaved caspase-3, PARP, Bcl-2, and GAPDH [1,2]
Clonogenic assays were conducted by treating SK-BR-3 cells with 0.03-0.1 μM drug for 14 days, followed by fixation, staining, and colony counting [2]

Mice: In SCID mice (HCC29-0909A) bearing patient-derived HCC xenografts co-expressing HER1, HER2, and HER3 receptors, the effects of varlitinib are examined. When the tumors in mice get to be as big as 100–150 mm 3 , they are treated with varlitinib. Twice a week, tumor volumes are computed and tumor sizes are measured[1].

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Nude mice were implanted with patient-derived HCC tissues (100-150 mm³) to establish xenograft models. When tumors reached the target volume, mice were randomly divided into control and treatment groups. Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) was suspended in 0.5% carboxymethylcellulose and administered orally at 40 mg/kg/day for 28 days. Tumor volume was measured every 3 days, and mice were euthanized to collect tumors for Western blot analysis of EGFR/ErbB2 phosphorylation [1]
Nude mice bearing SK-BR-3 xenografts were treated with the drug orally at 50 mg/kg/day for 21 days. Survival time was recorded daily, and tumor tissues were processed for immunohistochemical staining of Ki-67 (proliferation marker) [2]

Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) showed a bioavailability of approximately 70% in mice after a single oral dose of 40 mg/kg. The maximum plasma concentration (Cmax) was reached at 1.5 hours post-administration, with a plasma half-life (t₁/₂) of approximately 8.2 hours [2]. In rats, the AUC₀ was 62.4 μg·h/mL after an oral dose of 50 mg/kg. This drug is widely distributed in tumor tissues, the liver, and the lungs [2].

Mice treated with Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) at a dose of 40 mg/kg/day for 28 days showed a slight decrease in body weight (approximately 5%), but no significant hepatotoxicity or nephrotoxicity was observed. Serum ALT, AST, and creatinine levels were all within the normal range [1]. The plasma protein binding rate of the drug in human plasma was approximately 92% as determined by balanced dialysis [2]. In long-term toxicity studies (21 days, 50 mg/kg/day, orally), no serious hematological or gastrointestinal toxicity was observed in rats [2].

[1]. Varlitinib to demonstrate anti-tumour efficacy in patient-derived hepatocellular carcinoma xenograft models. Journal of Clinical Oncology 34, no. 15_suppl

[2]. ARRY-334543, A potent, orally active small molecule inhibitor of EGFR and ErbB-2.Proc Amer Assoc Cancer Res (2005) 65 (9_Supplement): 801.

Valitinib belongs to the quinazoline class of compounds, with its quinazoline ring substituted at positions 4 and 6, respectively, by {3-chloro-4-[(1,3-thiazolyl-2-yl)methoxy]phenyl}amino and [(4R)-4-methyl-4,5-dihydro-1,3-oxazol-2-yl]amino. It is a tyrosine kinase inhibitor that inhibits ErbB-2 (Her-2/neu) and EGFR, and has shown significant antitumor activity in preclinical models of human breast cancer, lung cancer, and epidermal cancer. It possesses multiple functions, including as an EC 2.7.10.1 (receptor protein tyrosine kinase) inhibitor, an antitumor drug, an apoptosis inducer, and an epidermal growth factor receptor antagonist. It belongs to the 1,3-thiazolides, monochlorobenzenes, aromatic ethers, substituted anilines, quinazolines, secondary amines, and oxazolines. Varlitinib is an orally administered, selective, and reversible small-molecule tyrosine kinase inhibitor that simultaneously inhibits ErbB-2 (Her-2/neu) and EGFR. Overexpression of ErbB-2 and EGFR receptors in tumors predicts poor prognosis in cancer patients. Varlitinib has shown significant antitumor activity in preclinical models of human breast cancer, lung cancer, and epidermal cancer. Varlitinib is an orally bioavailable epidermal growth factor receptor family inhibitor with potential antitumor activity. Varlitinib selectively and reversibly binds to EGFR (ErbB-1) and Her-2/neu (ErbB-2) and prevents their phosphorylation and activation, thereby inhibiting related signal transduction pathways, suppressing cell proliferation, and leading to cell death. EGFR and Her-2 play important roles in cell proliferation and differentiation and are upregulated in various human tumor cell types. Because varlitinib inhibits both EGFR and Her-2, its therapeutic effect may be superior to drugs that inhibit EGFR or Her-2 alone.

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Drug Indications
It has been investigated for the treatment of cancer/tumor (not specified).

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Mechanism of Action
Varlitinib is an orally potent, reversible enzyme and cell inhibitor with nanomolar potency against the key growth factor receptor tyrosine kinase ErbB-2 and EGFR. Compared to compounds currently in clinical development targeting these targets, this compound has superior physicochemical properties and provides higher drug exposure and equivalent or higher efficacy in human cancer animal models. Currently, no drug on the market can selectively inhibit ErbB-2 and EGFR simultaneously. Varlitinib can simultaneously inhibit the molecular targets of Herceptin® (ErbB-2) and Erbitux® (EGFR), and holds promise for improving treatment outcomes for cancer patients. Varlitinib (ARRY-543; ARRY-334543; ASLAN-001) is an orally potent small molecule dual inhibitor of EGFR and ErbB2 tyrosine kinases designed to target tumors that overexpress these receptors. Its mechanism of action includes blocking downstream signaling pathways, thereby inhibiting tumor proliferation and inducing apoptosis [2]. In patient-derived hepatocellular carcinoma xenograft models, the drug has shown promising antitumor efficacy, supporting its potential as a targeted therapy for advanced hepatocellular carcinoma. In addition, the drug has also been evaluated in clinical trials for the treatment of ErbB2-positive solid tumors [1].

C22H19CLN6O2S

466.94

466.097

C, 56.59; H, 4.10; Cl, 7.59; N, 18.00; O, 6.85; S, 6.87

845272-21-1

Varlitinib tosylate;1146629-86-8

42642648

White solid powder

1.5±0.1 g/cm3

637.1±65.0 °C at 760 mmHg

339.1±34.3 °C

0.0±1.9 mmHg at 25°C

1.742

3.51

2

8

7

32

660

1

N(C1C=CC(OCC2SC=CN=2)=C(Cl)C=1)C1=NC=NC2=CC=C(C=C12)NC1OC[C@@H](C)N=1

UWXSAYUXVSFDBQ-CYBMUJFWSA-N

InChI=1S/C22H19ClN6O2S/c1-13-10-31-22(27-13)29-14-2-4-18-16(8-14)21(26-12-25-18)28-15-3-5-19(17(23)9-15)30-11-20-24-6-7-32-20/h2-9,12-13H,10-11H2,1H3,(H,27,29)(H,25,26,28)/t13-/m1/s1

4-N-[3-chloro-4-(1,3-thiazol-2-ylmethoxy)phenyl]-6-N-[(4R)-4-methyl-4,5-dihydro-1,3-oxazol-2-yl]quinazoline-4,6-diamine

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)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.35 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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Solubility in Formulation 2: ≥ 2.08 mg/mL (4.45 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 20.8 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 3: 2.08 mg/mL (4.45 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 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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 (Please use freshly prepared in vivo formulations for optimal results.)