The aryl hydrocarbon receptor as an antitumor target of synthetic curcuminoids in colorectal cancer

Abstract

BACKGROUND Curcumin has proven to be a potent antitumor agent in both preclinical and clinical models of colorectal cancer (CRC). It has also been identified as a ligand of the transcription factor known as the aryl hydrocarbon receptor (AHR). Our lab has identified the AHR as a mechanism which contributes to both tumorigenesis in a mouse model of inflammatory CRC as well an apoptotic target in vitro. Curcumin’s role as an AHR ligand may modulate its effects to induce colon cancer cell death, and this role may be enhanced via structural modification of the curcumin backbone. We sought to determine if the two piperidone analogs of curcumin, RL66 and RL118, exhibit more robust antitumor actions than their parent compound in the context of colorectal cancer in vitro. Moreover, to ascertain the ability of curcumin, RL66, and RL118 to activate the aryl hydrocarbon receptor and evaluate if this activation has any effect on CRC cell death. MATERIALS AND METHODS DLD1, HCT116, LS513, and RKO colon cell lines were propagated in vitro. Natural curcumin was obtained commercially, while RL66 and RL118 were synthesized and characterized de novo. Multi-well fluorescent/luminescent signal detection was utilized to simultaneously ascertain cell viability, cell cytonecrosis, and relative amounts of apoptotic activity. AHR activity was measured with a dual luciferase reporter gene system. Stable expression of siRNA interference was established in the HCT116 cell lines to create AHR “knock down” cell lines. RESULTS Both RL66 and RL118 proved to be more potent antitumor agents than their parent compound curcumin in all cell lines tested. The majority of this cell death was due to induction of apoptosis, which occurred earlier and to a greater degree following RL66 and RL118 treatment as opposed to curcumin. Also, RL66 and RL118 were found to be activators of AHR, and a portion of their ability to cause cell death was dependent on this induction. Curcumin was found unable to activate the AHR, and levels of AHR mRNA did not change its effects on cell death. CONCLUSION Piperidone analogs of curcumin exhibited enhanced antitumor effects in vitro as opposed to their parent compound. Even more, this enhanced cell death profile may be partially attributed to the ability of these compounds to activate the AHR. Further study of synthetic curcumin analogs as chemopreventives and chemoadjuncts in CRC is warranted. Also, more generally, the AHR may represent a potential putative target for novel anticancer agents for CRC.

2,3,7,8‑tetrachlorodibenzo‑p‑dioxin suppresses the growth of human colorectal cancer cells in vitro: Implication of the aryl hydrocarbon receptor signaling

“…To investigate the mechanism of action of TCDD, it was first demonstrated that the AHR and CYP1A1 proteins are present in RKO cells, consistent with previous studies, demonstrating that these mRNAs are expressed in these cells in vitro ( 34 , 35 ). In the present study, TCDD treatment was demonstrated to be caused a reduction of AHR levels and an elevation of CYP1A1 levels in the cytosol, including endoplasmic reticulum of RKO cells.…”

“…To characterize the mechanism of TCDD action, and determine whether or not TCDD treatment regulates the levels of key transcription factors, western blot analysis was used. AHR and CYP1A1 mRNAs were previously reported to be expressed in RKO cells in vitro ( 34 , 35 ). It was demonstrated that the levels of AHR and CYP1A1 were altered by TCDD in RKO cells ( Fig.…”

“…The AHR is expressed and characterized in human colon adenocarcinoma cells, including RKO cells ( 33 - 35 ), and has been demonstrated to regulate the expression levels of CYP1A1 ( 36 ) and CYP1A2 ( 37 ) in colorectal cancer cells in vitro . The role of AHR thus has been reported in colon cancer cells ( 38 ).…”

Untargeted Metabolomics Reveals Intestinal Pathogenesis and Self-Repair in Rabbits Fed an Antibiotic-Free Diet

“…Jiang et al [37] believe that suppression of the gut microbiome by addition of antibiotics in feed is likely the cause for the reduced level of phenylalanine metabolites, tryptophan metabolites from the serotonin pathway, and the secondary bile acids. These metabolites play an important role in regulating the expression of inflammation-related genes [38], enhancing the epithelial cell barrier [39], and inhibiting the growth of CRC cells in an aryl hydrocarbon receptor (AHR)-dependent manner [40]. These results are supported by several in vitro and in vivo studies [41].…”

Alteration of fecal tryptophan metabolism correlates with shifted microbiota and may be involved in pathogenesis of colorectal cancer

“…In addition to being excreted in the stool and used for protein synthesis, gut Trp is catabolized mainly along the following three pathways[ 15 ]: (1) Trp indole pathway: Trp in feces can be metabolized into indole and indole derivatives by gut microorganisms, such as Clostridium sporogenes and E. coli [ 16 ] , Achromobacter liquefaciens, Bacteroides spp [ 17 ] , and Bifidobacterium spp [ 18 ]. Indoles, including indole, indole-3-acid-acetic (IAA), skatole (3-methylindole), indole-3-propionic acid (IPA), indole-3-aldehyde (IALD), indole-3-acetaldehyde (IAALD), and so on[ 11 ], play an important role in modulating the expression of inflammation-related genes[ 19 ], strengthening the epithelial cell barrier[ 20 ], and inhibiting the growth of CRC cells[ 21 ] in an aryl hydrocarbon receptor (AHR)-dependent way; (2) Kynurenine (KYN) pathway (KP): Most Trp in tissue is metabolized to KYN along the KP, which is regulated by the main rate-limited enzymes, including indoleamine 2,3-dioxygenase 1 (IDO1) in almost all tissue and Trp 2,3-dioxygenase in the liver[ 22 ]. Intestinal bacteria can indirectly participate in the KP by affecting the activity of IDO or the concentration of other metabolites[ 23 ]; and (3) Serotonin pathway (SP): Trp can be converted to 5-hydroxytryptophan (5-HTP) by Trp hydroxylase, followed by the decarboxylation of 5-HTP through the activation of aromatic L-amino acid decarboxylase[ 15 ].…”