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Antisense oligonucleotides inhibit intercellular adhesion molecule 1 expression by two distinct mechanisms.
M Y Chiang1, H Chan, M A Zounes
1Department of Molecular and Cellular Biology and Medicinal Chemistry, ISIS Pharmaceuticals, Carlsbad, California 92008.
The Journal of Biological Chemistry
|September 25, 1991
Summary
Two antisense oligonucleotides, ISIS 1570 and ISIS 1939, effectively inhibit intercellular adhesion molecule 1 (ICAM-1) expression post-transcriptionally. ISIS 1939 reduces ICAM-1 mRNA via RNase H, while ISIS 1570 inhibits ICAM-1 function through a different mechanism.
Area of Science:
- Molecular Biology
- Immunology
- Cell Biology
Background:
- Intercellular adhesion molecule 1 (ICAM-1) is a cell surface glycoprotein crucial for leukocyte emigration from vasculature.
- ICAM-1 expression is upregulated by inflammatory cytokines like interleukin-1 beta, tumor necrosis factor alpha, and interferon gamma in endothelial and carcinoma cells.
Purpose of the Study:
- To investigate the efficacy of phosphorothioate antisense oligonucleotides in inhibiting ICAM-1 expression.
- To elucidate the mechanisms of action for two specific oligonucleotides, ISIS 1570 and ISIS 1939, targeting ICAM-1 mRNA.
Main Methods:
- Utilized enzyme-linked immunosorbent assay (ELISA) to measure ICAM-1 expression.
- Employed immunoprecipitation of radiolabeled proteins to confirm specific inhibition.
- Assessed ICAM-1 mRNA levels and gene transcription rates.
- Investigated RNase H-dependent and -independent mechanisms using 2'-O-Methyl phosphorothioate analogs.
- Evaluated the impact of oligonucleotide treatment on HL-60 cell adherence to endothelial cells.
Main Results:
- Both ISIS 1570 and ISIS 1939 significantly inhibited ICAM-1 expression post-transcriptionally without affecting gene transcription.
- ISIS 1939 reduced ICAM-1 mRNA levels, suggesting an RNase H-dependent mechanism.
- ISIS 1570 inhibited ICAM-1 expression and function without altering mRNA levels, indicating a distinct mechanism.
- Oligonucleotide activity was sequence-specific and blocked by sense strand hybridization.
- Both oligonucleotides effectively reduced HL-60 cell adherence to endothelial cell monolayers.
Conclusions:
- Antisense oligonucleotides targeting ICAM-1 mRNA offer a viable strategy for modulating ICAM-1-mediated cellular interactions.
- ISIS 1939 acts via RNase H-dependent degradation of ICAM-1 mRNA, while ISIS 1570 employs a different post-transcriptional mechanism.
- The inhibition of ICAM-1 expression by these oligonucleotides has functional consequences, reducing leukocyte adherence.