The Effect of Novel Oleanolic Acid Oximes Conjugated with Indomethacin on the Nrf2-ARE And NF-κB Signaling Pathways in Normal Hepatocytes and Human Hepatocellular Cancer Cells

Abstract

Nrf2 and NF-κB play a key role in inflammation-driven cancers. Conjugation of anti-inflammatory drugs with oleanolic acid oxime (OAO) may enhance their therapeutic potential as a result of downregulation of these pathways. Novel OAO derivatives conjugated with indomethacin (IND) were synthesized, and their effect on the activation and expression of Nrf2 and NF-κB in HepG2 hepatoma cells and THLE-2 immortalized normal hepatocytes was evaluated in relation to cell cycle arrest and apoptosis. Treatment with OAO–IND conjugates reduced the activation of Nrf2 and NF-κB and the expression of their active forms in HepG2 cells, while in normal hepatocytes, the activation of Nrf2 was increased and NF-κB diminished. Compounds 3d, 3-indomethacinoxyiminoolean-12-en-28-oic acid morpholide, and 3c, 3-indomethacinoxyiminoolean-12-en-28-oic acid benzyl ester, were the most efficient. In THLE-2 cells, as opposed to HepG2 cells, the expressions of SOD-1 and NQO1 were significantly enhanced after treatment with these compounds. The COX-2 expression was diminished in both cell lines. OAO–IND derivatives affected the cell cycle arrest at G2/M, leading to increased apoptosis and increased number of resting HepG2 cells. Therefore, the conjugation of IND with OAO derivatives may preserve cancer cells against chemoresistance through the inhibition of the Nrf2-ARE pathway and NF-κB and, at the same time, exert a chemopreventive effect in normal hepatocytes.

Conjugation of Diclofenac with Novel Oleanolic Acid Derivatives Modulate Nrf2 and NF-κB Activity in Hepatic Cancer Cells and Normal Hepatocytes Leading to Enhancement of Its Therapeutic and Chemopreventive Potential

“…On the other hand, the relationship between ERK and Nrf2 activation was described [ 37 ]. While in the case of the OAO derivatives alone, both mechanisms, i.e., the down-regulation of Keap1 and activation of ERK, might contribute to Nrf2 activation and cytotoxicity, their conjugation with DCL or indomethacin [ 11 ] results in the opposite effect. Therefore, the mechanism of Nrf2 inhibition by the DCL–OAO derivative hybrids in HCC cells requires further study.…”

“…In contrast, in normal hepatocytes, activation of Nrf2 increased, and that of NF-κB was diminished. Hence, conjugation of IND with OAO derivatives may preserve cancer cells against chemoresistance by inhibiting the Nrf2-ARE and NF-κB pathways, whilst at the same time exerting a chemopreventive impact in normal hepatocytes [ 11 ].…”

Anti-Cancer Potential of Synthetic Oleanolic Acid Derivatives and Their Conjugates with NSAIDs

“…OAOs themselves, particularly OAO substituted with morpholide, may be considered therapeutic agents, which may support conventional treatment strategy [ 55 ]. In contrast to OAO conjugates with ASP, OAO hybrids with indomethacin and diclofenac reduced activation and expression, not only NF-κB, but also Nrf2 in hepatoma cells, while in normal cells, increased activation of Nrf2 was still observed ( Figure 3 and Figure 4 ) [ 39 , 62 ].…”

“… OAO-IND and OAO-DCL exert different effects on the Nrf2 pathway in normal hepatocytes and HCC cells. OAO–IND conjugates increase the activation of Nrf2 in normal hepatocytes by 25–55%, but inhibit its activation in HCC cells (by 30–40%) [ 62 ]. Similarly, OAO–DCL conjugates enhance the activation of Nrf2 in normal hepatocytes by 22–46% and inhibit activation of Nrf2 in HCC cells by 21–55% [ 39 ].…”

“… OAO–NSAIDs conjugates reduce NF-κB activation and expression of COX-2 in normal hepatocytes and HCC cells. OAO–NSAIDs inhibit activation of NF-κB in normal hepatocytes and in HCC cells by ~50–60% and ~40–50%, respectively [ 35 , 39 , 62 ]. Figure was created with BioRender.com by the authors.…”