Naphthylisoquinoline alkaloids and their synthetic analogs as potent novel inhibitors against Babesia canis in vitro.

Babesia canis is the predominant and clinically relevant canine Babesia species in Europe. Transmitted by vector ticks, the parasite enters red blood cells and induces a severe, potentially fatal hemolytic anemia. Here, we report on the antibabesial activities of three extracts of the West African tropical plant species Triphyophyllum peltatum (Dioncophyllaceae) and Ancistrocladus abbreviatus (Ancistrocladaceae) and of 13 genuine naphthylisoquinoline alkaloids isolated thereof. Two of the extracts and eight of the alkaloids were found to display strong activities against Babesia canis in vitro. Among the most potent compounds were the C,C-coupled dioncophyllines A (1a) and C (2) and the N,C-linked alkaloids ancistrocladium A (3) and B (4), with half-maximum inhibition concentration (IC50) values of 0.48 µM for 1a, 0.85 µM for 2, 1.90 µM for 3, and 1.23 µM for 4. Structure-activity relationship (SAR) studies on a small library of related genuine analogs and non-natural synthetic derivatives of 1a and 2 revealed the likewise naturally occurring alkaloid N-methyl-7-epi-dioncophylline A (6b) to be the most potent (IC50, 0.14 µM) among the investigated compounds. Although none of the tested naphthylisoquinolines showed 100 % inhibition of parasite infection - as displayed by imidocarb dipropionate (IC50, 0.07 µM), which was used as a positive control - the antibabesial potential of the dioncophyllines A (1a) and C (2) and related compounds such as 6b, its atropo-diastereomer 6a (IC50, 1.45 µM), and 8-O-(p-nitrobenzyl)dioncophylline A (14) (IC50, 0.82 µM) is to be considered as high. The SAR results showed that N-methylation and axial chirality exert a strong impact on the antibabasial activities of the naphthylisoquinolines presented here, whereas dimerization, as in jozimine A2 (5) (IC50, 140 µM), leads to a significant decrease of activity against B. canis. Alkaloids displaying good to high activities against B. canis like the dioncophyllines 1a, 2, 6a, and 6b were found to cause only a small degree of hemolysis (< 0.7 %), whereas compounds with moderate to weak antibabesial activities such as 6-O-methyl-4'-O-demethylancistrocladine (15a) (IC50, 14.0 µM) and its atropo-diastereomer 6-O-methyl-4'-O-demethylhamatine (15b) (IC50, 830 µM) caused a high degree of hemolysis (7.3 % for 15a and 11.2 % for 15b). In this respect, the most effective anti-Babesia naphthylisoquinolines are also the safest ones.

Naphthylisoquinoline alkaloids, validated as hit multistage antiplasmodial natural products.

The discovery and development of multistage antimalarial drugs targeting intra-erythrocytic asexual and sexual Plasmodium falciparum parasites is of utmost importance to achieve the ambitious goal of malaria elimination. Here, we report the validation of naphthylisoquinoline (NIQ) alkaloids and their synthetic analogues as multistage active antimalarial drug candidates. A total of 30 compounds were tested, of which 17 exhibited IC50 values <1 µM against drug-sensitive P. falciparum parasites (NF54 strain); 15 of these retained activity against a panel of drug-resistant strains. These compounds showed low in vitro cytotoxicity against HepG2 cells, with selectivity indices of >10. The tested compounds showed activity in vitro against both early- and late-stage P. falciparum gametocytes while blocking male gamete formation (>70% inhibition of exflagellation at 2 µM). Additionally, five selected compounds were found to have good solubility (≥170 µM in PBS at pH 6.5), while metabolic stability towards human, mouse, and rat microsomes ranged from >90% to >7% after 30 min. Dioncophylline C (2a) emerged as a front runner from the study, displaying activity against both asexual parasites and gametocytes, a lack of cross-resistance to chloroquine, good solubility, and microsomal stability. Overall, this is the first report on the multistage activity of NIQs and their synthetic analogues including gametocytocidal and gametocidal effects induced by this class of compounds.