The synthesis and biological activity of nitrogen containing chalcones and analogues
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Malaria is a global health problem, with an estimated 300–500 million new clinical cases and 1–2 million fatalities reported annually. Almost 90% of the incidences of malaria and deaths from the disease occur in sub-Saharan Africa. Since malaria affects mainly the poor, it is not profitable for pharmaceutical companies to develop new treatments; thus malaria is classified as a „neglected‟ or „orphan disease‟. Malaria in humans, transmitted by female Anopheles mosquitoes, is caused by four species of Plasmodium, of which P. vivax is the most common malaria parasite, while the most severe form of malaria is caused by P. falciparum. These protists, the targets of antimalarial drugs, gradually develop resistance to malarial drugs. For example, quinine from the bark of the cinchona tree was the first drug against malaria. When it became obsolete in the 1940s it was replaced by chloroquine (CQ), a synthetic analogue, which is also reaching a stage of obsolescence. Many of the subsequently developed drugs are structurally related to chloroquine; thus they are or will soon be ineffective. Artemisinin, developed from an ancient Chinese herbal medication for fever, is the latest antimalarial drug of choice. It is structurally unrelated to aminoquinoline, but resistance has already been observed in East Asia. Derivatives – including artesunate, dihydroartemisinin, artemether and arteether – are often used in combination with other antimalarial compounds to increase the half-life of a drug and delay the development of resistance to it. Because flavonoids are not detected in human blood after oral administration (i.e. are not bioavailable), it is difficult to explain the plethora of biological activities and beneficial dietary effects reported for flavonoids. The low bioavailability of polyphenols is explained by their poor absorption in the intestines and their rapid enzymatic degradation in blood. Furthermore, according to the Lipinski rules, most commercially available drugs contain nitrogen as hydrogen bond acceptors. We thus hypothesized that the introduction of nitrogen and the removal of as many OH groups as possible from flavonoids would enhance the bioactivity and the bioavailability and, in turn, lead to new drug leads. Since chalcones are easily synthesized from readily available commercial reagents and only a ew reports of bioactive nitrogen-containing chalcones have been published, we embarked on a project to synthesize nitrogen-containing chalcones and analogues and test their bioactivity for malaria and cancer. We used the Mannich reaction to introduce nitrogen as an aminoalkyl moiety. Initial results with our first-generation aminoalkylated chalcones supported our hypothesis by showing moderate to good activity against malaria and cancer. Medicinal chemistry predicts that the enone moiety is responsible for both high toxicity and low bioavailability. Upon replacement of the enone moiety with a propyl moiety, via catalytic hydrogenation, the bioactivity of the resulting aminoalkylated diarylpropanes was increased about a hundredfold. Consequently, we launched a programme to synthesize a wide array of aminoalkylated diarylpropane analogues, not only to enhance the bioactivity, but also to reduce the toxicity and increase the bioavailability. The Mannich reaction requires at least one aromatic OH group on one of the aromatic rings; therefore all our analogues are phenols. A total of 56 compounds were synthesized, characterized and tested for bioactivity. A smaller number was tested for toxicity and four were tested in in vivo mice models for bioavailability. These tests were outsourced to the University of Cape Town. The analogues synthesized by us included compounds with different amine groups (for example piperidine, pyrrolidine, morpholine, 1-methylpiperazine, 1-ethylpiperazine, and dimethylamine), different A-ring substituents (F, Br, methyl, ethyl, butanyl, propanyl, CF3, NH2 etc.) including compounds with furan and thiophene A-rings, as well as other compounds, including a diarylethane and an analogue with an aminoalkyl group on both the A- and B-ring. Toxicity was seldom a constraint and most of our compounds demonstrated high selectivity indices (in excess of 7000). Most of these compounds conform to the Lipinski rules. The first compound we tested showed bioavailability of 3%. We attributed this to first pass metabolism and attempted to protect the aromatic OH group ortho to the aminoalkyl group, via a prodrug strategy. However, this OH resisted ether and ester formation, probably due to a hydrogen bond to the aminoalkyl amine group (via a stable six-membered ring). Subsequently, we established that substituents on the A-ring increase the bioavailability. The analogue with a CF3 group on the A-ring has a bioavailability of 25%, placing it within the range of some commercially available drugs. It is not clear whether this is due to enhanced lipophilicity (CLogP) or whether a large substituent on the A-ring can protect a B-ring OH from enzymatic degradation. Work is in progress to test carbamates and other prodrugs and analogues with larger substituents on the A-ring. Some of our compounds indicated promising activity against TK-10 (renal), UACC-62 (melanoma) and MCF-7 (breast) cancer cell lines. The best result was a TGI value of 2.11 against melanoma, which is smaller than the parthenolide TGI value of 4.47. We believe that this thesis lays the foundations for an antimalarial drug with good bioavailability and low toxicity, which will potentially be cheap to manufacture. Since our compounds are totally unrelated to existing antimalarial compounds, resistance is not a problem, as indicated by the good activity of these compounds against chloroquine-resistant malaria strains (Dd2 and K1).