A series of diclofenac N-derivatives (2, 4, 6, 8c, 9c, 10a-c) were synthesized in order to test their anti-cancer and anti-inflammatory effects. The anticarcinogen activity has been assayed against three cancer cell lines: HT29, human colon cancer cells; Hep-G2, human hepatic cells; and B16-F10, murine melanoma cells. First, we determined the cytotoxicity of the different compounds, finding that the most effective compound was compound 8c against all cell lines and both compounds 4 and 6 in human Hep-G2 and HT29 cell lines. Compounds 4 and 8c were selected for the percentage of apoptosis determination, cell cycle distribution, and mitochondrial membrane potential measure because these products presented the lowest IC₅₀ values in two of the three cancer cell lines assayed (B16-F10 and HepG2), and were two of the three products with lowest IC₅₀ in HT29 cell line. Moreover, the percentages of apoptosis induction were determined for compounds 4 and 8c, showing that the highest values were between 30 to 60%. Next, the effects of these two compounds were observed on the cellular cycle, resulting in an increase in the cell population in G2/M cell cycle phase after treatment with product 8c, whereas compound 4 increased the cells in phase G0/G1, by possible differentiation process induction. Finally, to determine the possible apoptosis mechanism triggered by these compounds, mitochondrial potential was evaluated, indicating the possible activation of extrinsic apoptotic mechanism. On the other hand, we studied the anti-inflammatory effects of these diclofenac (DCF) derivatives on lipopolysaccharide (LPS) activated RAW 264.7 macrophages-monocytes murine cells by inhibition of nitric oxide (NO) production. As a first step, we determined the cytotoxicity of the synthesized compounds, as well as DCF, against these cells. Then, sub-cytotoxic concentrations were used to determine NO release at different incubation times. The greatest anti-inflammatory effect was observed for products 2, 4, 8c, 10a, 10b, and 9c at 20 µg·mL⁻¹ concentration after 48 h of treatment, with inhibition of produced NO between 60 to 75%, and a concentration that reduces to the 50% the production of NO (IC_{50 NO}) between 2.5 to 25 times lower than that of DCF. In this work, we synthesized and determined for the first time the anti-cancer and anti-inflammatory potential of eight diclofenac N-derivatives. In agreement with the recent evidences suggesting that inflammation may contribute to all states of tumorigenesis, the development of these new derivatives capable of inducing apoptosis and anti-inflammatory effects at very low concentrations represent new effective therapeutic strategies against these diseases.

Natural products have always played an important role in pharmacotherapy, helping to control pathophysiological processes associated with human disease. Thus, natural products such as oleanolic acid (OA), a pentacyclic triterpene that has demonstrated important activities in several disease models, are in high demand. The relevant properties of this compound have motivated re-searchers to search for new analogues and derivatives using the OA as a scaffold to which new functional groups have been added or modifications have been realized. OA and its derivatives have been shown to be effective in the treatment of inflammatory processes, triggered by chronic diseases or bacterial and viral infections. OA and its derivatives have also been found to be effective in diabetic disorders, a group of common endocrine diseases characterized by hyperglycemia that can affect several organs, including the liver and brain. This group of compounds has been reported to exhibit significant bioactivity against cancer processes in vitro and in vivo. In this review, we summarize the bioactive properties of OA and its derivatives as anti-inflammatory, anti-bacterial, antiviral, anti-diabetic, hepatoprotective, neuroprotective, and anticancer agents.

Hepatocellular carcinoma (HCC) is the most common type of liver cancer. Our recent studies have shown that the diamine-(PEG)ylated oleanolic acid (OADP) has strong anti-tumor effects in HCCs. In this study, we evaluated the anti-tumor mechanisms of OADP in the HepG2 HCC cell line. The cytotoxicity results showed that HepG2 cell viability was markedly reduced, with a very low 50% of cell growth inhibitory concentration (IC₅₀, 0.14 µg/mL). We then investigated the anti-tumor mechanisms of OADP in HepG2 cells. The flow-cytometry analysis was used to evaluate cell apoptosis, indicating that 74–95% of cells were apoptotic. OADP caused cell cycle arrest in the G0/G1 phase and the loss of the mitochondrial membrane potential (MMP). Western blot analysis was performed to assess the expression levels of key proteins associated with the underlying molecular mechanism. The results showed the clear upregulation of caspase-8, caspase-9, caspase-3, Bak, p21, and p53, accompanied by the downregulation of Bcl-2. Similar results were obtained by the cotreatment with OADP and the c-Jun N-terminal kinase (JNK) inhibitor SP600125. Agents such as OADP, which are capable of activating extrinsic and intrinsic apoptotic pathways, may represent potential HCC cancer therapies.

Recent evidence has shown that inflammation can contribute to all tumorigenic states. We have investigated the anti-inflammatory effects of a diamine-PEGylated derivative of oleanolic acid (OADP), in vitro and in vivo with inflammation models. In addition, we have determined the sub-cytotoxic concentrations for anti-inflammatory assays of OADP in RAW 264.7 cells. The inflammatory process began with incubation with lipopolysaccharide (LPS). Nitric oxide production levels were also determined, exceeding 75% inhibition of NO for a concentration of 1 µg/mL of OADP. Cell-cycle analysis showed a reversal of the arrest in the G0/G1 phase in LPS-stimulated RAW 264.7 cells. Furthermore, through Western blot analysis, we have determined the probable molecular mechanism activated by OADP; the inhibition of the expression of cytokines such as TNF-α, IL-1β, iNOS, and COX-2; and the blocking of p-IκBα production in LPS-stimulated RAW 264.7 cells. Finally, we have analyzed the anti-inflammatory action of OADP in a mouse acute ear edema, in male BL/6J mice treated with OADP and tetradecanoyl phorbol acetate (TPA). Treatment with OADP induced greater suppression of edema and decreased the ear thickness 14% more than diclofenac. The development of new derivatives such as OADP with powerful anti-inflammatory effects could represent an effective therapeutic strategy against inflammation and tumorigenic processes.

We have designed and synthesized two novel cobalt coordination compounds using bumetanide (bum) and indomethacin (ind) therapeutic agents. The anti-inflammatory effects of cobalt metal complexes with ind and bum were assayed in lipopolysaccharide stimulated RAW 264.7 macrophages by inhibition of nitric oxide production. Firstly, we determined the cytotoxicity and the anti-inflammatory potential of the cobalt compounds and ind and bum ligands in RAW 264.7 cells. Indomethacin-based metal complex was able to inhibit the NO production up to 35% in a concentration-dependent manner without showing cytotoxicity, showing around 6–37 times more effective than indomethacin. Cell cycle analysis showed that the inhibition of NO production was accompanied by a reversion of the differentiation processes in LPS-stimulated RAW 264.7 cells, due to a decreased of cell percentage in G0/G1 phase, with the corresponding increase in the number of cells in S phase. These two materials have mononuclear structures and show slow relaxation of magnetization. Moreover, both compounds show anti-diabetic activity with low in vitro cell toxicities. The formation of metal complexes with bioactive ligands is a new and promising strategy to find new compounds with high and enhanced biochemical properties and promises to be a field of great interest.