GNE-617, a specific NAMPT inhibitor(IC50=5 nM), shows potency in xenograft models of cancer.

NAMPT:5 nM

在A549细胞中，GNE-617抑制NAMPT（IC50=18.9 nM）。

在大鼠中，GNE-617 hydrochloride (每日一次给药)和GNE-875(每日两次给药)与GMX-1778(每日两次给药)相比，在相似的暴露量和给药持续时间下，都显示出更严重的视网膜毒性。使用GNE-617、GNE-618和GMX-1778的小鼠有效性研究旨在评估化合物的有效性，并机会性地用于评估小鼠的视网膜毒性。观察到GNE-617和GMX-1778具有NAMPTi视网膜毒性；然而，GNE-617和GMX-1778之间不同的研究持续时间，不允许直接比较视网膜毒性。

For RNA interference (RNAi), A549 cells are plated at 1,500 cells per well in 96-well plates, allowed to adhere for 24 hours, and transfected with 25 nM siRNA oligonucleotide using Dharmafect 4. Transfected cells are treated with the indicated concentrations of GNE-617 (0.1, 1 , 10 , 100 , and 1000 nM) for 72 hours and viability is evaluated with CellTiter-Glo. Lysates for detection of NAPRT1 protein are collected 72 hours after transfection of 1 million A549 cells in 10 cm dishes. For NAPRT1 re-expression, RERF-LC-MS cells are transfected with pCMV6-AC.NAPRT1 and empty vector pCMV6-AC using Amaxa Nucleofector technology and selected with Geneticin

GNE-617 is dissolved in DMSO and stored, and then diluted with appropriate media before use.Cells are grown in RPMI-1640 medium supplemented with 10% FBS and 2 mM glutamine and passaged not more than 20 times after thawing. To determine the IC50 values and nicotinic acid rescue status, cells are treated with nine point dose titrations of GNE-617 with or without 10 μM nicotinic acid. At 96 hours post-drug addition, the GNE-617-treated cells are evaluated using CyQUANT Direct Cell Proliferation Assay followed by CellTiter-Glo Luminescent Cell Viability Assay quantified with a Wallac EnVision 2104 Multilabel Reader. IC50 values are calculated using XLfit 5.1. To examine the protein level, cells are lysed in ice-cold radioimmunoprecipitation assay buffer, run on SDS-PAGE (4%-12% Bis-Tris), and evaluated by Western blotting using antibodies directed against NAPRT1 and β-actin

Male na?ve Sprague Dawley rats are administered once daily via oral gavage with GNE-617( 30?mg/kg), formulated as a solution in the vehicle of 60% polyethylene glycol (PEG 400)/10% ethanol/30% 5% dextrose in water (D5W) .