Inhibition of mutant KrasG12D -initiated murine pancreatic carcinoma growth by a dual c-Raf and soluble epoxide hydrolase inhibitor t -CUPM

Abstract

Mutant Kras and chronic pancreatitis are the most common pathologic events involved in human pancreatic cancer. It has been demonstrated that c-Raf is responsible for transmitting signals from mutant Ras to its downstream signals including MEK-ERK and for initiating carcinogenesis. The soluble epoxide hydrolase (sEH), a pro-inflammatory enzyme, generally inactivates anti-inflammatory and anti-pain epoxyeicosatrienoic acids (EETs). Herein, we have synthesized a novel compound of trans-4-{4-[3-(4-chloro-3-trifluoromethyl-phenyl)-ureido]-cyclohexyloxy}-pyridine-2-carboxylic acid methylamide (t-CUPM) via modifying the central phenyl ring of sorafenib and confirmed its dual inhibition of sEH and c-Raf by recombinant kinase activity assay. Pharmacokinetic analysis revealed that oral dosing of t-CUPM resulted in the higher blood levels than that of sorafenib throughout the complete time course (48 h). The effect of t-CUPM on the inhibition of mutant KrasG12D–initiated murine pancreatic cancer cell growth was determined using the mouse pancreatic carcinoma cell model obtained from LSL-KrasG12D/Pdx1-Cre mice and showed that t-CUPM significantly inhibited this murine pancreatic carcinoma cell growth both in vitro and in mice in vivo. Inhibition of mutant Kras-transmitted phosphorylations of cRAF/MEK/ERK was demonstrated in these pancreatic cancer cells using Western blot assay and immunohistochemical approach. Modulation of oxylipin profile, particularly increased EETs/DHET ratio by sEH inhibition, was observed in mice treated with t-CUPM. These results indicate that t-CUPM is a highly potential agent to treat pancreatic cancer via simultaneously targeting c-Raf and sEH.

Inhibition of Pancreatic Carcinoma Growth Through Enhancing ω-3 Epoxy Polyunsaturated Fatty Acid Profile by Inhibition of Soluble Epoxide Hydrolase

“…Following our previously described methods (32,33), freshly harvested tumor tissues were extracted for cell lysates using ice-cold RIPA lysis buffer (Santa Cruz Biotechnology, Santa Cruz, CA, USA) and total protein concentrations were measured using the Bradford reagent (Thermo Scientific, Chicago, IL, USA). The tissue lysate (30 μg protein/lane) was separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred onto a polyvinylidene fluoride membrane.…”

Epoxy-Oxylipins and Soluble Epoxide Hydrolase Metabolic Pathway as Targets for NSAID-Induced Gastroenteropathy and Inflammation-Associated Carcinogenesis

“…In the development of chronic pancreatitis aberrant ARA metabolism partially mediated by sEH acts to promote inflammation with a decrease of the anti-inflammatory EETs. Inhibition of sEH has been demonstrated as a successful strategy in murine models to inhibit chronic pancreatitis and the development of pancreatic intraepithelial neoplasms (PanIN) using trans-4-{4-[3-(4-chloro-3-trifluoromethyl-phenyl)-ureido]-cyclohexyloxy}-pyridine-2-carboxylic acid methylamide (t-CUPM) in KRAS-mutant mice (Liao et al, 2016a; Liao et al, 2016b). t-CUPM is a novel molecule with dual inhibition of sEH and RAF1 proto-oncogene serine/threonine kinase (c-RAF).…”

“…sEH inhibitors have been shown to robustly decrease sEH activity with little to no toxicity in animal models. Further, these compounds have proven effective to 1) decrease GI ulcers induced by non-steroidal anti-inflammatory drugs (NSAIDs) (Goswami et al, 2016; Goswami et al, 2017; Yang, 2018), 2) prevent carcinogenesis in murine models of colorectal and pancreatic tumors (Liao et al, 2016a; Liao et al, 2016b; Wang et al, 2018b; Yang, 2018), and 3) decrease chronic inflammation in mouse models for both colitis and pancreatitis (Zhang et al, 2012; Zhang et al, 2013b; Zhang et al, 2013c; Goswami et al, 2016; Liao et al, 2016a; Goswami et al, 2017; Wang et al, 2018b; Yang, 2018).…”

Modulation of mitochondrial dysfunction and endoplasmic reticulum stress are key mechanisms for the wide-ranging actions of epoxy fatty acids and soluble epoxide hydrolase inhibitors

“…sEH is also a contributor to pancreas exocrine functions and is implicated in chronic and acute pancreatitis. Indeed, simultaneous pharmacological inhibition of sEH and RAF1 proto-oncogene serine/threonine kinase (c-RAF) reduces the severity of chronic pancreatitis [112]. Moreover, sEH deficient mice exhibit attenuated chemical-induced (cerulein- and arginine-induced) acute pancreatitis [42].…”