ANTISENSE PHOSPHOROTHIOATE OLIGONUCLEOTIDES - SELECTIVE KILLING OF THE INTRACELLULAR PARASITE LEISHMANIA-AMAZONENSIS

We targeted the mini-exon sequence, present at the 5' end of every mRN A of the protozoan parasite Leishmania amazonensis, by phosphorothioat e oligonucleotides. A complementary 16-mer (16PS) was able to kill ama stigotes-the intracellular stage of the parasite-in murine macrophages in culture. After 24 hr of incubation with 10 mu M 16PS, about 30% in fected macrophages were cured. The oligomer 16PS acted through antisen se hybridization in a sequence-dependent way; no effect on parasites w as observed with noncomplementary phosphorothioate oligonucleotides. T he antisense oligonucleotide 16PS was a selective killer of the protoz oans without any detrimental effect to the host macrophage. Using 16PS linked to a palmitate chain, which enabled it to complex with low den sity lipoproteins, improved the leishmanicidal efficiency on intracell ular amastigotes, probably due to increased endocytosis. Phosphorothio ate oligonucleotides complementary to the intron part of the mini-exon pre-RNA were also effective, suggesting that antisense oligomers coul d prevent trans-splicing in these parasites.