Halimane Derivatives from Plectranthus ornatus Codd. as Novel Anti-cancer Agents.

The Plectranthus genus is often cited for its medicinal properties. Plectranthus ornatus Codd. is traditionally used in Africa for the treatment of gastric and liver diseases and their leaves are used for their antibiotic action. The main constituent of P. ornatus is the halimane compound, 11 R∗-acetoxyhalima-5,13E-dien-15-oic acid (Hal), described for its antimicrobial and anticancer properties. The objective of this work was to improve the activity of the halimane lead molecule. Further physiochemical characterisation was performed on Hal. To the best of our knowledge, this work constitutes the first published data of the absolute configurations by SCXRD and thermal stability of Hal. Using Hal, reactions with different amines were carried out to afford novel semi-synthetic derivatives and their structural elucidation was completed. The cytotoxicity of the derivatives was assessed against three leukaemia cancer cell lines (CCRF-CEM, K562 and HL-60). The antioxidant activity was investigated using H2O2-induced HGF-1 cells and their anti-inflammatory activity was studied using RT-PCR and ELISA. Our data showed that amide derivatives of Hal presented moderate cytotoxicity and more potent activity when compared to the parent molecule, giving insight into the SAR of Hal. The derivatives also displayed protection against oxidative damage to DNA. Finally, the derivatives possessed anti-inflammatory properties at the level of gene and protein expression for the cytokines IL-1ß, TNF-α and IL-6, induced by LPS in normal HGF-1 cells. Overall, our study provides useful insight into the enhanced biological activities of semi-synthetic Hal derivatives, as a starting point for novel drug formulations in cancer therapy.

Discovery of a Necroptosis Inhibitor Improving Dopaminergic Neuronal Loss after MPTP Exposure in Mice.

Parkinson's disease (PD) is the second most common neurodegenerative disorder, mainly characterized by motor deficits correlated with progressive dopaminergic neuronal loss in the substantia nigra pars compacta (SN). Necroptosis is a caspase-independent form of regulated cell death mediated by the concerted action of receptor-interacting protein 3 (RIP3) and the pseudokinase mixed lineage domain-like protein (MLKL). It is also usually dependent on RIP1 kinase activity, influenced by further cellular clues. Importantly, necroptosis appears to be strongly linked to several neurodegenerative diseases, including PD. Here, we aimed at identifying novel chemical inhibitors of necroptosis in a PD-mimicking model, by conducting a two-step screening. Firstly, we phenotypically screened a library of 31 small molecules using a cellular model of necroptosis and, thereafter, the hit compound effect was validated in vivo in a sub-acute 1-methyl-1-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP) PD-related mouse model. From the initial compounds, we identified one hit-Oxa12-that strongly inhibited necroptosis induced by the pan-caspase inhibitor zVAD-fmk in the BV2 murine microglia cell line. More importantly, mice exposed to MPTP and further treated with Oxa12 showed protection against MPTP-induced dopaminergic neuronal loss in the SN and striatum. In conclusion, we identified Oxa12 as a hit compound that represents a new chemotype to tackle necroptosis. Oxa12 displays in vivo effects, making this compound a drug candidate for further optimization to attenuate PD pathogenesis.

Synthesis and Biological Evaluation of Hybrid 1,5- and 2,5-Disubstituted Indoles as Potentially New Antitubercular Agents.

BACKGROUND: Tuberculosis (TB) is the second leading cause of mortality worldwide being a highly contagious and insidious illness caused by Mycobacterium tuberculosis, Mtb. Additionally, the emergence of multidrug-resistant and extensively drug-resistant strains of Mtb, together with significant levels of co-infection with HIV and TB (HIV/TB) make the search for new antitubercular drugs urgent and challenging. METHODS: This work was based on the hypothesis that an active compound could be obtained if substituents present in some other active compounds were attached on a core of an important structure, in this case the indole scaffold, thus generating a hybrid compound. A QSAR-oriented design based on classification and regression models along with the estimation of physicochemical and biological properties have also been used to assist in the selection of compounds. Chosen compounds were synthesized using various synthetic procedures and evaluated against M. tuberculosis H37Rv strain. RESULTS: Selected compounds possess substituents at positions C5, C2 and N1 of the indole ring. The substituents involve p-halophenyl, pyridyl, benzyloxy and benzylamine groups. Four compounds were synthesised using suitable synthetic procedures to attain the desired substitution at the indole core. From these, three compounds are new and have been fully characterized, and tested in vitro against the H37Rv ATCC27294T Mtb strain, using isoniazid as a control. One of them, compound 2, with the pyridyl group at N1, has an experimental log (1/MIC) very close to 5 and can be considered as being (weakly) active. In fact, it is more active than 64% of all indole molecules in our data sets of experimental results from literature. The most active indole in this data sets has log (1/MIC)=5.93 with only 6 compounds with log (1/MIC) above 5.5. CONCLUSION: Despite the lower activity found for the tested compounds, when compared to other reported indole-derivatives, these structures, which rely on a hybrid design concept, may constitute interesting scaffolds to prepare a new family of TB inhibitors with improved activity.

Indole based cyclooxygenase inhibitors: synthesis, biological evaluation, docking and NMR screening.

The close structural similarity between the two cyclooxygenase (COXs) isoforms and the absence of selective inhibitors without side effects continues to stimulate the development of novel approaches towards selective anti-inflammatory drugs. In the present study a small library of new indolic compounds involving two different substitutions patterns at the indole scaffold was synthesized. In order to establish a relation between the spatial distribution of known functional groups related with inhibitory activity, two substitution patterns were explored: one with substituents at N-1, C-3, C-5 positions and another at C-2, C-3 and C5 positions. Accordingly, indole positions C-5, C-3 and N-1 were substituted with: sulfonamide or methylsulfone at C-5, p-halo-benzyl group at C-3, and an alkyl chain with a trifluoromethyl group at N-1. Alternatively, a p-halo-benzyl group was introduced at C-2, leaving the indolic nitrogen free. Inhibitory studies were performed and the activity results obtained against both COXs isoforms were rationalized based on docking and NMR studies. Docking studies show that dialkyation at C-2 and C-3 favors a binding with an orientation similar to that of the known selective inhibitor SC-558. From the tested compounds, this substitution pattern is correlated with the highest inhibitory activity and selectivity: 70% COX-2 inhibition at 50 µM, and low COX-1 inhibition (18 ± 9%). Additionally, Saturation Transfer Difference NMR experiments reveal different interaction patterns with both COXs isoforms that may be related with different orientations of the sulfonamide group in the binding pocket. Despite the moderated inhibitory activities found, this study represents an innovative approach towards COXs inhibitory activity rationalization and to the design of anti-inflammatory drugs.

A new insight on the hypochlorous acid scavenging mechanism of tryptamine and tryptophan derivatives.

The reaction mechanisms of hypochlorous acid (HOCl) with several tryptophan and tryptamine derivatives, previously reported to scavenge this powerful oxidant, was investigated to determine whether ionic or radical pathways were involved. For this purpose, the reaction of tryptamine and tryptophan derivatives with HOCl was optimized and some compounds were isolated by HPLC and their structures assigned. In order to prevent possible radical reaction pathway, experiments have been carried in the presence of the radical trap TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl). The obtained results show that the reaction mechanisms are influenced by the type of structure and that a complex pathway is involved, in which both ionic and radical mechanisms can occur.

Antioxidant activity of unexplored indole derivatives: synthesis and screening.

The present study envisaged the development of novel antioxidant candidates using the indole scaffold. Several tryptophan and tryptamine derivatives were synthesized, in particular prenylated indole compounds, and their scavenging activity for reactive oxygen species (ROS) and reactive nitrogen species (RNS) was investigated. The library substitution pattern included several alkyl chains at positions N-1, C-2 of the indole nucleus, including prenyl and isopentyl chain, as well as different groups at the side chain (C-3) that allowed the investigation of a possible radical stabilization. The results obtained showed that tryptophan (8), tryptamine (9), N-phthaloyl tryptamine (5) and N-prenyl tryptophan (13) were the most active against peroxyl radical (ROO(•)) with activities higher than Trolox, which was used as control. The scavenging of hypochlorous acid (HOCl) was also evaluated and tryptophan (8) and tryptamine (9) showed IC(50) of 3.50 ± 0.4 and 6.00 ± 0.60 µM, respectively. Significant activity was also found for the N-prenyl tryptophan (13) with an IC(50) of 4.13 ± 0.17 µM and C-2 prenylated derivative (14), with 4.56 ± 0.48 µM. The studies were extended to RNS and best results were obtained against peroxynitrite anion (ONOO(-)) in the presence of NaHCO(3). N-alkylated tryptophan (18) showed a high activity with an IC(50) of 14.0 ± 6.8 µM. The results show that the tested compounds are effective scavengers of ROS and RNS, and suggest that the radical stabilization is strongly dependent on the type of substituents on the indolic moiety and on their relative positions. In addition, the radical dissipation inside the indolic system is mandatory for the observed antioxidant activity.