Kinetic selection of HPV 16 E6/E7-directed antisense nucleic acids: anti-proliferative effects on HPV 16-transformed cells

The E6/E7-coding sequences of the human papillomavirus type 16 (HPV 16) wer e probed for kinetic accessibility in vitro by pools of catalytic antisense RNA. Only long-chain complementary RNA and very few antisense sequences wi th a 3' portion complementary to a 10 nt window within unspliced and splice d EG-coding target sequences showed fast annealing with k(ass) values of up to 10(4) M(-1)s(-1) indicating that the majority of E6/E7 RNA sequences ar e unfavourable targets for antisense inhibitors and ribozymes. Fast-anneali ng antisense oligodeoxyribonucleotides directed against the window of 10 nt inhibited cell proliferation of HPV 16-transformed SiHa cells but not slow -annealing antisense species. Antisense RNA of several hundred nucleotides in length also showed significant antiproliferative activity. Biological ef fects of antisense oligodeoxyribonucleotides were specific for the antisens e sequence, could only be found in HPV-positive but not in HPV-negative cel l lines, and were related to decreased levels of E7 protein and E6/E7-speci fic transcripts. This work suggests that HPV 16 E7/E6 sequences exhibit a l ow accessibility for antisense oligonucleotides. This can be overcome, howe ver, by exploiting the relationship between fast annealing of antisense spe cies and their increased efficacy in human cells.