Yarla Nagendra Sastri, Pathuri Gopal, Gali Hariprasad, Terzyan Simon, Panneerselvam Janani, Chandrakesan Parthasarathy, Scotti Marcus Tullius, Houchen Courtney, Madka Venkateshwar, Rao Chinthalapally V
Center for Cancer Prevention and Drug Development, Hem-Onc Section, Department of Medicine, Stephenson Cancer Center, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
College of Pharmacy, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
J Inflamm Res. 2020 Dec 31;13:1261-1278. doi: 10.2147/JIR.S286110. eCollection 2020.
Non-steroidal anti-inflammatory drugs, cyclooxygenase (COX)-2 selective inhibitors, have been explored for prevention and treatment of several inflammatory chronic conditions including arthritis, and cancer. However, the long-term use of these drugs is associated with gastrointestinal, renal, and cardiovascular side effects. Later, COX/5-lipoxygenase (5-LOX) dual inhibitors (eg, licofelone) have been developed but did not enter into the market from the clinical trails due to COX-1/2 inhibition-associated side effects. Hence, targeting microsomal prostaglandin E synthase-1 (mPGES-1) and 5-LOX can be an ideal approach while sparing COX-1/2 activities for development of the next generation of anti-inflammatory drugs with better efficacy and safety.
In silico (molecular modelling) studies were used to design a mPGES-1/5-LOX dual inhibitory and COX-1/2 sparing lead molecule licofelone analogue-9 (LFA-9) by modifying the pharmacophore of licofelone. In vitro cell-free enzymatic (mPGES-1, 5-LOX, COX-1/2) assays using fluorometric/colorimetric methods and cell-based assays (LPS-induced PGE, LTB, and PGI productions from macrophages) using ELISA technique, isothermal calorimetry, and circular dichroism techniques were performed to determine the mPGES-1/5-LOX inhibitory efficacy and selectivity. Anti-inflammatory efficacy of LFA-9 was evaluated using a carrageenan (inflammogen)-induced rat paw edema model. Infiltration/expression of CD68 immune cells and TNF-α in paw tissues were evaluated using confocal microscope and immunoblot analysis. Anti-cancer effect of LFA-9 was evaluated using colon spheroids in vitro.
LFA-9 inhibited mPGES-1/5-LOX and their products PGE and LTB, spared COX-1/2 and its product PGI. LFA-9 bound strongly with human mPGES-1/5-LOX enzymes and induced changes in their secondary structure, thereby inhibited their enzymatic activities. LFA-9 inhibited carrageenan-induced inflammation (70.4%) in rats and suppressed CD68 immune cell infiltration ( ≤ 0.0001) and TNF-α expression. LFA-9 suppressed colon tumor stemness (60.2%) in vitro through inhibition of PGE (82%) levels.
Overall study results suggest that LFA-9 is a mPGES-1/5-LOX dual inhibitor and showed anti-inflammatory and colorectal cancer preventive activities, and warranted detailed studies.
非甾体抗炎药，即环氧化酶（COX）-2选择性抑制剂，已被用于探索预防和治疗包括关节炎和癌症在内的多种炎症性慢性病。然而，这些药物的长期使用与胃肠道、肾脏和心血管副作用相关。后来，COX/5-脂氧合酶（5-LOX）双重抑制剂（如利考昔芬）被开发出来，但由于与COX-1/2抑制相关的副作用，未能从临床试验进入市场。因此，靶向微粒体前列腺素E合酶-1（mPGES-1）和5-LOX可能是一种理想的方法，同时保留COX-1/2的活性，以开发出疗效更好、安全性更高的下一代抗炎药物。
通过计算机模拟（分子建模）研究，对利考昔芬的药效团进行修饰，设计出一种mPGES-1/5-LOX双重抑制且保留COX-1/2活性的先导分子利考昔芬类似物-9（LFA-9）。使用荧光/比色法进行体外无细胞酶促（mPGES-1、5-LOX、COX-1/2）测定，并使用ELISA技术、等温滴定量热法和圆二色性技术进行基于细胞的测定（巨噬细胞中脂多糖诱导的PGE、LTB和PGI生成），以确定mPGES-1/5-LOX的抑制效果和选择性。使用角叉菜胶（炎症原）诱导的大鼠足爪水肿模型评估LFA-9的抗炎效果。使用共聚焦显微镜和免疫印迹分析评估足爪组织中CD68免疫细胞和TNF-α的浸润/表达。使用体外结肠球体评估LFA-9的抗癌效果。
LFA-9抑制mPGES-1/5-LOX及其产物PGE和LTB，保留COX-1/2及其产物PGI。LFA-9与人mPGES-1/5-LOX酶紧密结合，并诱导其二级结构发生变化，从而抑制其酶活性。LFA-9抑制角叉菜胶诱导的大鼠炎症（70.4%），并抑制CD68免疫细胞浸润（≤0.0001）和TNF-α表达。LFA-9通过抑制PGE（82%）水平在体外抑制结肠肿瘤干性（60.2%）。
总体研究结果表明，LFA-9是一种mPGES-1/5-LOX双重抑制剂，具有抗炎和预防结直肠癌的活性，值得进行详细研究。
