| Primaquine, an 8-aminoquinoline, has been used in the treatment of malaria for over 40 years. Primaquine therapy remains the drug of choice in the treatment against the exoerythrocytic forms of Plasmodium spp. Unfortunately, methemoglobinemia and hemolytic anemia associated with primaquine therapy have limited its usefulness in many patients. Over the years, there have been many reports of derivatives of primaquine produced by synthesis and biocatalysis in a search for derivatives or metabolites that lack or reduce the degree of the associated methemoglobinemia and hemolytic anemia, increase the potency, and/or are effective in resistant forms of the microorganism.
Bioconversions, using a variety of microorganisms, of primaquine have lead to many unique structural modifications of the primaquine scaffold. In any living organism, the most common of the primaquine metabolites is carboxyprimaquine. It is interestingly to note that the transformation of primaquine to carboxyprimaquine completely eliminates any effective antimalarial activity. A second common microbial metabolite, produced by bioconversion across almost all species of Streptomyces, is the N-acetyl derivative of primaquine. Other transformations that have been reported include dealkylation, N-hydroxylation, aromatic hydroxylation, and conjugation. Studies have also shown that 6-methoxy-8-hydroxyaminoquinolines induce the formation of the undesired methemoglobinemia and hemolytic anemia.
Here, we report our findings of the bioconversion of primaquine by the versatile Beauveria bassiana, which has previously been shown to have a wide array of biotransforming capabilities with numerous substrates.
Even after decades of use, primaquine remains one of the few clinically useful antimalarial agents. Unlike other 8-aminoquinoline antimalarials, primaquine therapy offers the advantage of successfully erradicates the sporozoites, merozoites, and gametes that reside outside of the infected erythrocytes. Unfortunately, metabolism of the primaquine results in the formation of 8-hydroxylaminoquinoline metabolites in humans (Fig. 7), which are well known for their hemolytic activities. Years of synthetic and biosynthetic manipulation of the chemical skeleton of primaquine have failed to produce any compounds that retain the unique antimalarial activity of primaquine, yet are void of the hemolytic-causing metabolites.
Many previous biotransformation experiments have focused on reactions involving alkylation and acetylation of primaquine’s primary amine, the formation of carboxyprimaquine, and hydroxylation of the aromatic ring. Interestingly, not any of these changes to the primaquine structure produces structures with the ability to avoid the formation of 8-hydroxylaminoquinolines during metabolism. We report the identification of a common microbial metabolite, N-acetylprimaquine, as well as the formation of a novel metabolite, N-formylprimaquine. It is the formylated primaquine derivative that provides the structural framework that offers potential avoidance of metabolism to the 8-hydroxylaminoquinolines and its subsequent hemolytic side effects. By blocking the seconadary nitrogen with the formyl moiety, deacetylation on the moieties attached to the nitrogen is likely to be inhibited, or slowed. In addition, the formylated primaquine derivative offers hope of a new structural series of antimalarial, which is of extreme significance as primaquine-resistant Plasmodium strains become more and more common.
In an effort to identify microorganisms capable of N-formilation and N-acetylation of the secondary amine of the 8-aminoquinoline derviatives, we examined microbial transformation methodologies using primaquine as a metabolic substrate in liquid culture of different strains of fungi. Obtaining N-formylated or N-acetylated derivatives in position 8- of primaquine molecule (Fig. 2) as therapeutic materials might be able to help to avoid formation 8-hydroxylaminoquinolines in human organisms and also to eliminate the resistance of Plasmodium for these drugs.
We observed that in the list of all checked strains of fungi only B. bassiana was capable for the formation of the N-formylated derivative at secondary amine’s group and of the N-acetylated derivative at the primary amine’s group of the side chaine of primaquine. The formyl-substituted derivative (N-[4-amino-1-methyl-butyl)-N-(6-methoxy-quinolin-8-yl]- formamide) provides a unique structural characteristic for the examination of a new series of 8-aminoquinolines in the search for new antimalarial chemotherapeutics. |
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