HER2 (IC50 = 8 nM)[1];
HER2 (ErbB2)：Irbinitinib (ARRY-380; ONT-380; Tucatinib) is a selective inhibitor of HER2 with an IC₅₀ of 8 nM for HER2 kinase activity, showing >1,000-fold selectivity over EGFR (IC₅₀ > 10,000 nM). [2][3]

Tucatinib hemiethanolate is roughly 500 times more selective for HER2 than EGFR in cell-based experiments, and it exhibits nanomolar activity against purified HER2 enzyme. In comparison to EGFR, tucatinib preferentially inhibits the receptor tyrosine kinase HER2[1]. In HER2 overexpressing cell lines, tutatinib inhibits the growth and phosphorylation of HER2 as well as its downstream effector, Akt. Tucatinib might be able to stop HER2 signaling in EGFR-overexpressing cell lines by weakly inhibiting phosphorylation and proliferation, without having the negative side effects of EGFR inhibition[1].

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- Antiproliferative activity：Irbinitinib inhibits the growth of HER2+ breast cancer cell lines (BT-474, SK-BR-3) with IC₅₀ values of 12–25 nM in MTT assays. It has minimal effect on HER2-negative cell lines (MCF-7, IC₅₀ > 1,000 nM). [2][3]
- HER2 signaling inhibition：In SK-BR-3 cells, Irbinitinib (100 nM, 4 hours) reduces phosphorylation of HER2 (Tyr1248), AKT (Ser473), and ERK1/2 (Thr202/Tyr204) by 80–90% as measured by Western blot. It also downregulates cyclin D1 and upregulates cleaved caspase-3, indicating induction of apoptosis. [3]

When each medication is dosed at the maximum-tolerated dose, tutunitinib hemiethanolate (ONT-380 hemiethanolate, 200 mg/kg/d) exhibits a survival benefit[1]. The amount of brain pErbB2 is significantly reduced (80%) by tucacitabine and its active metabolite[2]. Tucatinib (ARRY-380) hemiethanolate exhibits a noteworthy tumor growth inhibition (TGI; 50% at 50 mg/kg/d and 96% at 100 mg/kg/d) in response to dose, with several partial regressions (>50% reduction from baseline size) observed in 9/12 animals at the higher dose level. When combined with trastuzumab, tutatinib (50 mg/kg/d) exhibits a 98% TGI with two partial regressions and complete regressions in 9 out of 12 animals[3].

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- Tumor growth inhibition in xenografts：In BT-474 human breast cancer xenografts in nude mice, oral Irbinitinib (50 mg/kg, twice daily) reduces tumor volume by 60–70% after 21 days. Combination with RP-56976 (a PI3K inhibitor) enhances efficacy, achieving 85% tumor growth inhibition. [3]
- Clinical activity in HER2+ MBC：In a Phase 1 expansion cohort, Irbinitinib (300 mg twice daily) shows objective response rates (ORR) of 33% in patients with HER2+ metastatic breast cancer (MBC), with a median duration of response (DOR) of 7.5 months. [1]

Irbinitinib, formerly known as ARRY-380 and ONT-380 or Tucatinib, is a potent and selective small molecule inhibitor of HER2 with IC50 value of 8 nM, it is equally potent against truncated p95-HER2, and is 500-fold more selective for HER2 versus EGFR. Irbinitinib acts by blocking the proliferation and phosphorylation of HER2 and its downstream effector, Akt. By contrast, in the EGFR overexpressing cell lines, it weakly inhibits phosphorylation and proliferation, demonstrating that Irbinitinib may have potential to block HER2 signaling without causing the toxicities of EGFR inhibition. Therefore, it has the potential to be used as an anticancer agent.

ONT-380 has nanomolar activity against purified HER2 enzyme and is approximately 500-fold selective for HER2 versus EGFR in cell-based assays. In the EGFR overexpressing cell lines, it weakly inhibits phosphorylation and proliferation, demonstrating that Irbinitinib may have potential to block HER2 signaling without causing the toxicities of EGFR inhibition.

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- Proliferation and signaling assay： 1. HER2+ breast cancer cells (BT-474, SK-BR-3) are seeded in 96-well plates and treated with Irbinitinib (0.1–1,000 nM) for 72 hours. 2. Cell viability is measured by MTT assay to determine IC₅₀ values. 3. For signaling analysis, cells are treated with 100 nM Irbinitinib for 1–24 hours, lysed, and proteins (p-HER2, p-AKT, p-ERK) are detected by Western blot. [2][3]

Animal/Disease Models: Female nude mice[3].
Doses: 200 mg/kg/d.
Route of Administration: PO, daily.
Experimental Results: demonstrated anti-tumor activity and benefited survival.

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- Breast cancer xenograft model： 1. Female nude mice are implanted with BT-474 cells (5×10⁶) subcutaneously. 2. When tumors reach 100 mm³, mice are randomized to receive Irbinitinib (25–100 mg/kg, oral, twice daily) alone or with RP-56976 (20 mg/kg, oral, daily) for 21 days. 3. Tumor volume is measured twice weekly; at study end, tumors are analyzed for p-HER2 and Ki-67 expression by immunohistochemistry. [3]

Human pharmacokinetics: In the Phase I clinical trial, the steady-state peak plasma concentration (Cmax) of ibritinib (300 mg twice daily) was 2.8 μg/mL, the half-life was 8.5 hours, and the oral bioavailability was approximately 32%. It is mainly metabolized by CYP2C8 and CYP3A4. [1]

- Clinical toxicity: In the Phase I clinical trial, common adverse events (AEs) included diarrhea (45%), fatigue (30%), and nausea (25%); Grade 3 or higher adverse events were rare (10%), and no dose-limiting hepatotoxicity was observed. Plasma protein binding was >98%. [1]

[1]. Phase 1 Study of ONT-380, a HER2 Inhibitor, in Patients with HER2+ Advanced Solid Tumors, with an Expansion Cohort in HER2+ Metastatic Breast Cancer (MBC). Clin Cancer Res. 2017 Jan 4. pii: clincanres.1496.2016.

[2]. Abstract: In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 852. doi:1538-7445.AM2012-852.

[3]. In Vivo Activity of ARRY-380, a Potent, Small Molecule Inhibitor of ErbB2 in Combination with RP-56976. Cancer Research.

ARRY-380 is an orally effective, potent small-molecule tyrosine kinase inhibitor targeting ErbB2. This compound is a reversible ATP-competitive inhibitor, exhibiting nanomolar inhibitory activity against ErbB2 in both in vitro and cellular experiments. It possesses excellent in vivo and in vitro pharmacokinetic/ADME properties and demonstrates superior activity in various mouse tumor models, including breast cancer (BT-474, MDA-MB-453), ovarian cancer (SKOV-3), and gastric cancer (N87) models. In this paper, we demonstrate the superior monotherapy activity of ARRY-380 in breast and ovarian cancer models, as well as its efficacy in combination with trastuzumab, docetaxel, or bevacizumab. In the BT-474 study, tumor fragments were implanted into female SCID beige mice. In the SKOV-3 tumor study, cells were subcutaneously inoculated into the flanks of female nude mice. Animals received treatments including: oral (PO) ARRY-380 at doses up to 200 mg/kg/day; and/or intraperitoneal (IP) trastuzumab at 20 mg/kg every 3 days or weekly; and/or intravenous (IV) docetaxel at 10 mg/kg every 3 days; and/or intraperitoneal (IP) bevacizumab at 10 mg/kg every 4 days for a total of 3 times. Tumor size was measured periodically, and some animals were monitored for up to 90 days to determine tumor-free survival. In the BT-474 model, ARRY-380 demonstrated significant dose-related tumor growth inhibition (TGI; 50% at 50 mg/kg/day and 96% at 100 mg/kg/day), with 9 out of 12 animals in the high-dose group experiencing multiple partial regressions (a reduction of more than 50% from baseline size). One complete remission was observed in the high-dose group. The TGI of trastuzumab alone was 45%, but no tumor regression was observed. Treatment with ARRY-380 (50 mg/kg/day) in combination with trastuzumab achieved a tumor growth inhibition rate (TGI) of 98%, with complete tumor regression in 9 out of 12 animals and partial tumor regression in 2. At a combined dose of 100 mg/kg/day, the TGI reached 100%, and all animals achieved complete remission. Docetaxel monotherapy achieved a TGI of 55%, with no tumor regression observed. In combination with ARRY-380 (50 mg/kg/day), the TGI was 81%, and partial regression was observed in 5 animals. In the SKOV-3 model, ARRY-380 demonstrated significant dose-dependent tumor growth inhibition (TGI: 39% at 50 mg/kg twice daily; 96% at 100 mg/kg twice daily), with partial regression (more than 50% reduction in size from baseline) observed in all animals in the high-dose group. Bevacizumab monotherapy achieved a 55% tumor growth inhibition rate (TGI), but no tumor regression was observed. ARRY-380 (50 mg/kg twice daily) in combination with bevacizumab achieved an 80% TGI, with 7 out of 8 animals achieving partial remission and 1 animal achieving stable disease. This study demonstrates that ARRY-380 exhibits excellent monotherapy activity in both the BT-474 human breast cancer xenograft model and the SKOV-3 human ovarian cancer model. Furthermore, ARRY-380 showed synergistic effects and good tolerability when combined with trastuzumab, docetaxel, and bevacizumab. ARRY-380 has entered a Phase I clinical trial for patients with advanced cancer. [3] Mechanism of action: Ibitinib selectively binds to the HER2 kinase domain, inhibiting autophosphorylation and downstream PI3K/AKT and MAPK pathways, thereby inhibiting the proliferation of HER2-positive tumor cells and inducing their apoptosis. [2][3] Indications: It has been studied for HER2-positive advanced solid tumors, with a focus on metastatic breast cancer (MBC), including brain metastases due to blood-brain barrier penetration. [1][3]

C26H24N8O2.1/2C2H6O

503.57

1006.446

1429755-56-5

Tucatinib;937263-43-9

162623678

White to off-white solid powder

5

17

12

75

799

0

CCO.CC1=C(C=CC(=C1)NC2=NC=NC3=C2C=C(C=C3)NC4=NC(CO4)(C)C)OC5=CC6=NC=NN6C=C5.CC1=C(C=CC(=C1)NC2=NC=NC3=C2C=C(C=C3)NC4=NC(CO4)(C)C)OC5=CC6=NC=NN6C=C5

UGAFQGGOUGJBMF-UHFFFAOYSA-N

InChI=1S/2C26H24N8O2.C2H6O/c2*1-16-10-17(5-7-22(16)36-19-8-9-34-23(12-19)28-15-30-34)31-24-20-11-18(4-6-21(20)27-14-29-24)32-25-33-26(2,3)13-35-25;1-2-3/h2*4-12,14-15H,13H2,1-3H3,(H,32,33)(H,27,29,31);3H,2H2,1H3

6-N-(4,4-dimethyl-5H-1,3-oxazol-2-yl)-4-N-[3-methyl-4-([1,2,4]triazolo[1,5-a]pyridin-7-yloxy)phenyl]quinazoline-4,6-diamine hemiethanolate

Irbinitinib hemiethanolate; ONT380; Tucatinib hemiethanolate; 1429755-56-5; CDZ29KT2FT; UNII-CDZ29KT2FT; Tucatinib ethanolate (JAN); orb1693558; Tucatinib hemiethanolate [WHO-DD]; ARRY-380 hemiethanolate; ARRY380; ONT-380 hemiethanolate

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: 125 mg/mL (248.23 mM)
H2O: < 0.1 mg/mL

Solubility in Formulation 1: 2.08 mg/mL (4.13 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
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 2: 2.08 mg/mL (4.13 mM) 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; 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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Solubility in Formulation 3: ≥ 2.08 mg/mL (4.13 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 20.8 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.)