INH1 (IBT13131) is a cell-permeable Hec1 inhibitor that specifically disrupts the Hec1/Nek2 interaction.

Hec1:10-21 μM

Intraperitoneal injection of 100 mg/kg INH1 inhibits the growth of mammary tumors in mice with MDA-MB-468 human breast cancer xenografts.

INH1 effectively inhibits the proliferation of human breast cancer cells with a GI50 of 10-21 μM. Additionally, INH1 induces cell-killing activity by disrupting the spindle checkpoint-regulated Hec1/Nek2 pathway.

Binding assays: Surface plasma resonance (SPR) assays are performed at 22.5°C in HBSD buffer [10 mmol/L HEPES, 150 mmol/L NaCl, 0.1% DMSO (pH 7.5)] on Biacore 3000. 6×His-Hec1 and GST-Nek2 are purified. NTA sensor chip or glutathione-modified CM5 chip are used to capture His-Hec1 and GST-Nek2, respectively. The capture level is about 140 to 180 resonance units (RU) at the flow rate of 5 μL/min. For the binding assay, chips are sequentially treated with compounds (1 or 20 μmol/L) and then proteins (50 μg/mL). Retained RUs are recorded and processed (triplicate experiments).

Standard 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays with a 3-d drug treatment procedure are performed to measure the dose-dependent cytotoxicity of INH1 in cultured cells. Triplicate sets are measured and compiled for final data presentation.(Only for Reference)