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Anti-sense phosphorothioate oligonucleotides have both specific and non-specific effects on cells containing human
1International Center for Genetic Engineering and Biotechnology, Trieste, Italy.
Abstract:
A range of specific nuclease resistant phosphorothioate oligodeoxynucleotides (S-oligos) complementary to mRNA of human papillomavirus type 16 (HPV16), were tested for their ability to inhibit cell proliferation and to alter the level of HPV-specific mRNA and proteins in CaSki cells, a human cervical carcinoma cell line containing HPV16 DNA. Only certain of the S-oligos to the viral upstream regulatory region (URR) and the early viral open reading frames (ORF), E6 and E7, were found to display any activity on the cells. These S-oligos were found to exhibit potent anti-proliferative activity at concentrations between 0.25 microM and 20 microM, inhibiting the uptake of [3H]-thymidine into CaSki cells by up to 90% at higher concentrations. The rate of synthesis of E6 and E7 proteins and the steady state level of the E7 protein however remained largely unchanged. E7 protein exhibited a greater decrease in phosphorylation in the presence of only one of the antisense oligos. Other S-oligos including a random sequence, unmodified sequences or O-methylphosphonate modified oligos, had no specific effect on the cells. The results imply that the anti-sense S-oligonucleotides had both specific anti-HPV16 and other non-specific effects on cell proliferation and synthesis of virally encoded proteins.
Insights
Specific phosphorothioate oligodeoxynucleotides (S-oligos) targeting human papillomavirus type 16 (HPV16) showed potent anti-proliferative effects on cervical cancer cells. However, they did not significantly alter HPV-specific E6 and E7 protein levels.
Area of Science:
- Molecular Biology
- Virology
- Oncology
Background:
- Human papillomavirus type 16 (HPV16) is a major cause of cervical cancer.
- HPV16 oncoproteins E6 and E7 play crucial roles in viral oncogenesis.
- Antisense oligodeoxynucleotides offer a potential therapeutic strategy for HPV-associated cancers.
Purpose of the Study:
- To evaluate the efficacy of specific nuclease-resistant phosphorothioate oligodeoxynucleotides (S-oligos) against HPV16.
- To determine the impact of these S-oligos on HPV16-infected cervical carcinoma cell proliferation and viral gene expression.
Main Methods:
- Synthesis and testing of various S-oligos complementary to HPV16 mRNA.
- Assessing the effect of S-oligos on CaSki cell proliferation using [3H]-thymidine uptake.
- Analyzing the levels and phosphorylation of HPV16 E6 and E7 proteins.
Main Results:
- Certain S-oligos targeting the HPV16 upstream regulatory region (URR) and E6/E7 open reading frames (ORFs) demonstrated potent anti-proliferative activity (up to 90% inhibition).
- These active S-oligos did not significantly alter the synthesis rate or steady-state levels of E6 and E7 proteins.
- One S-oligo notably decreased E7 protein phosphorylation, while control oligos showed no specific effects.
Conclusions:
- Antisense S-oligonucleotides can specifically inhibit HPV16-infected cell proliferation.
- The anti-proliferative effects may involve mechanisms beyond direct modulation of E6 and E7 protein levels.
- Further research is warranted to explore the non-specific effects and optimize S-oligo-based therapies for HPV-related cancers.