Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, State University of New York, 352 Kapoor Hall, Buffalo, NY, 14214, USA.
AAPS J. 2019 Jan 7;21(2):13. doi: 10.1208/s12248-018-0279-5.
AR-C155858 and AZD3965, pyrrole pyrimidine derivatives, represent potent monocarboxylate transporter 1 (MCT1) inhibitors, with potential immunomodulatory and chemotherapeutic properties. Currently, there is limited information on the inhibitory properties of this new class of MCT1 inhibitors. The purpose of this study was to characterize the concentration- and time-dependent inhibition of L-lactate transport and the membrane permeability properties of AR-C155858 and AZD3965 in the murine 4T1 breast tumor cells that express MCT1. Our results demonstrated time-dependent inhibition of L-lactate uptake by AR-C155858 and AZD3965 with maximal inhibition occurring after a 5-min pre-incubation period and prolonged inhibition. Following removal of AR-C155858 or AZD3965 from the incubation buffer, inhibition of L-lactate uptake was only fully reversed after 3 and 12 h, respectively, indicating that these inhibitors are slowly reversible. The uptake of AR-C155858 was concentration-dependent in 4T1 cells, whereas the uptake of AZD3965 exhibited no concentration dependence over the range of concentrations examined. The uptake kinetics of AR-C155858 was best fitted to a Michaelis-Menten equation with a diffusional clearance component, P (K = 0.399 ± 0.067 μM, V = 4.79 ± 0.58 pmol/mg/min, and P = 0.330 ± 0.088 μL/mg/min). AR-C155858 uptake, but not AZD3965 uptake, was significantly inhibited by alpha-cyano-4-hydroxycinnamic acid, a known nonspecific inhibitor of MCTs 1, 2, and 4. AR-C155858 demonstrated a trend toward higher uptake at lower pH, a characteristic of proton-dependent MCT1. These findings provide evidence that AR-C155858 and AZD3965 exert slowly reversible inhibition of MCT1-mediated L-lactate uptake in 4T1 cells, with AR-C155858 representing a potential substrate of MCT1.
AR-C155858 和 AZD3965 是吡咯嘧啶衍生物，是有效的单羧酸转运蛋白 1（MCT1）抑制剂，具有潜在的免疫调节和化疗特性。目前，关于这一新类 MCT1 抑制剂的抑制特性的信息有限。本研究旨在研究 AR-C155858 和 AZD3965 对表达 MCT1 的小鼠 4T1 乳腺癌细胞中 L-乳酸转运的浓度和时间依赖性抑制作用以及膜通透性特性。我们的结果表明，AR-C155858 和 AZD3965 对 L-乳酸摄取具有时间依赖性抑制作用，最大抑制作用发生在 5 分钟预孵育期后，并持续抑制。当从孵育缓冲液中去除 AR-C155858 或 AZD3965 后，L-乳酸摄取的抑制作用仅在分别去除 3 小时和 12 小时后才完全逆转，表明这些抑制剂是缓慢可逆的。AR-C155858 在 4T1 细胞中的摄取呈浓度依赖性，而 AZD3965 的摄取在研究的浓度范围内没有浓度依赖性。AR-C155858 的摄取动力学最适合米氏方程，具有扩散清除率成分，P（K=0.399±0.067μM，V=4.79±0.58pmol/mg/min，和 P=0.330±0.088μL/mg/min）。AR-C155858 的摄取，但不是 AZD3965 的摄取，被已知的非特异性 MCT1、2 和 4 抑制剂α-氰基-4-羟基肉桂酸显著抑制。AR-C155858 显示出在较低 pH 值下摄取增加的趋势，这是质子依赖型 MCT1 的特征。这些发现提供了证据，表明 AR-C155858 和 AZD3965 在 4T1 细胞中对 MCT1 介导的 L-乳酸摄取具有缓慢可逆的抑制作用，AR-C155858 是 MCT1 的潜在底物。
