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Cellular distribution of phosphorothioate oligonucleotide following intravenous administration in mice
1Hybridon, Inc., Cambridge, MA 02139, USA.
Abstract:
Oligonucleotides are promising therapeutic agents for the prevention or treatment of a variety of diseases. The therapeutic potential of oligonucleotide therapy depends greatly on the bioavailability of oligonucleotides to their target cells and organs. We previously reported the pharmacokinetics and distribution of phosphorothioate oligonucleotide in mice using [35S]-labeled oligonucleotide ([35S]-oligo). To extend this study, we administered 30 mg/kg of fluorescent-labeled oligonucleotide (FITC-oligo) to mice and examined oligonucleotide distribution by measuring the fluorescence intensity in various cells and tissues using flow cytometry. Following FITC-oligo administration, fluorescence was detected in all the tissues examined. In terms of the fluorescent intensity, accumulation was greatest in liver and kidney, intermediate in spleen and bone marrow, and very low in peripheral blood mononuclear cells (PBMC). At 4 hours after administration, the level of oligonucleotide uptake in PBMC, spleen lymphocytes, and bone marrow cells revealed the following pattern: monocytes/macrophages > B cells > T cells. Confocal microscopy detected intracellular fluorescence in PBMC prepared under the same conditions as those for flow cytometry. These studies provide a rationale for designing cell targets for antisense therapeutics.
Insights
Fluorescent-labeled oligonucleotides distribute widely in mice, accumulating most in the liver and kidney. This study details oligonucleotide biodistribution and cellular uptake patterns, informing future antisense therapeutic design.
Area of Science:
- Pharmacology
- Molecular Biology
- Biotechnology
Background:
- Oligonucleotides show therapeutic promise for various diseases.
- Effective oligonucleotide therapy relies on reaching target cells and organs.
- Previous studies established phosphorothioate oligonucleotide pharmacokinetics in mice.
Purpose of the Study:
- To investigate the biodistribution and cellular uptake of fluorescent-labeled oligonucleotides (FITC-oligo) in mice.
- To extend previous pharmacokinetic studies using a different labeling method.
- To provide a basis for designing targeted antisense therapeutics.
Main Methods:
- Administration of 30 mg/kg fluorescent-labeled oligonucleotide (FITC-oligo) to mice.
- Quantification of fluorescence intensity in various tissues and cells using flow cytometry.
- Confocal microscopy to visualize intracellular fluorescence in peripheral blood mononuclear cells (PBMC).
Main Results:
- FITC-oligo was detected in all examined tissues.
- Highest accumulation observed in liver and kidney, followed by spleen and bone marrow.
- Peripheral blood mononuclear cells (PBMC) showed very low accumulation.
- Uptake in PBMC, spleen lymphocytes, and bone marrow cells followed the pattern: monocytes/macrophages > B cells > T cells.
- Intracellular fluorescence was confirmed in PBMC via confocal microscopy.
Conclusions:
- Oligonucleotide distribution is tissue-specific, with significant uptake in the liver and kidney.
- Cellular uptake varies among immune cell types, with monocytes/macrophages showing the highest uptake.
- These findings support the rationale for developing cell-targeted antisense therapeutics.