Total synthesis and antileishmanial activity of the natural occurring acetylenic fatty acids 6-heptadecynoic acid and 6-icosynoic acid

Abstract

The first total syntheses of the naturally occurring acetylenic fatty acids-6-heptadecynoic acid (59% overall yield) and 6-icosynoic acid (34% overall yield)-was accomplished in four steps. Using the same synthetic sequence the naturally occurring fatty acids (6Z)-heptadecenoic acid (46% overall yield) and (6Z)-icosenoic acid (27% overall yield) were also synthesized. The Delta(6) acetylenic fatty acids displayed good antiprotozoal activity towards Leishmania donovani promastigotes (EC(50) = 1-6 microg/mL), but the 6-icosynoic acid was the most effective in the series. In addition, the (6Z)-icosenoic acid was a much better antiprotozoal compound (EC(50) = 5-6 microg/mL) than the (6Z)-heptadecenoic acid (EC(50) > 25 microg/mL). The saturated fatty acids n-heptadecanoic acid and n-eicosanoic acid were not effective towards L. donovani, indicating that the Delta(6) unsaturation in these fatty acids is necessary for leishmanicidal activity. In addition, both the 6-icosynoic acid and the (6Z)-icosenoic acid were inhibitors of the Leishmania DNA topoisomerase IB enzyme (EC(50's) = 36-49 microM), a possible intracellular target for these compounds. This is the first study assessing fatty acids as inhibitors of the Leishmania DNA topoisomerase IB enzyme.

Fig. 1

Inhibition of the relaxation activity of recombinant LdTopIB by the 6-icosynoic acid (1b) (top) and the 6-heptadecynoic acid (1a) (bottom). One unit of recombinant LdTop1 was assayed in a plasmid DNA relaxation assay for 30 min at 37 °C (as described under “Materials and Methods”) in the presence of 12.5 to 1000 μM compounds 1a and 1b. Reaction products were resolved in agarose gel and subsequently visualized by ethidium bromide staining. The relative position of the negatively supercoiled DNA substrate is indicated by Sc, N is the nicked DNA, whereas the ladder of relaxed DNA topoisomer bands is labeled R. Reactions were stopped with a mixture of 1 % SDS and 6.1 μg of proteinase K. Lane 1 contains 0.2 μg of pHOT plasmid DNA and lane 2 is a relaxed marker.

Fig. 2

Inhibition of the relaxation activity of recombinant LdTopIB by the (6Z)-icosenoic acid (6b) (top) and the (6Z)-heptadecenoic acid (6a) (bottom). One unit of recombinant LdTop1 was assayed in a plasmid DNA relaxation assay for 30 min at 37°C (as described under “Materials and Methods”) in the presence of 12.5 to 1000 μM compounds 6a and 6b. Reaction products were resolved in agarose gel and subsequently visualized by ethidium bromide staining. The relative position of the negatively supercoiled DNA substrate is indicated by Sc, N is the nicked DNA, whereas the ladder of relaxed DNA topoisomer bands is labeled R. Reactions were stopped with a mixture of 1% SDS and 6.1 μg of proteinase K. Lane 1 contains 0.2 μg of pHOT plasmid DNA and lane 2 is a relaxed marker.

Fig. 3

Plots of relaxation activities (%) of the LdTOPIB enzyme vs. concentration (μM) of the acetylenic fatty acids 1a and 1b, and the olefinic fatty acids 6a and 6b. The EC₅₀ values are shown at the top of each graph.

Scheme 1

Synthesis of the acetylenic acids 1a and 1b and the olefinic acids 6a and 6b.