Effects of synthetic oligonucleotides on human complement and coagulation

Area of Science:

• Biochemistry
• Pharmacology
• Immunology

Background:

• Oligodeoxynucleotide phosphorothioates (PS-oligos) are investigated for gene expression inhibition.
• Preclinical studies noted complement and coagulation effects with high-dose PS-oligos in monkeys.

Purpose of the Study:

• To investigate the in vitro effects of PS-oligos on human blood clotting and complement activity.
• To determine the chemical properties of PS-oligos responsible for these effects.

Main Methods:

• In vitro assays using normal human plasma and serum.
• Measurement of clotting times (aPTT, PT, TT) and hemolytic complement activity.
• Assessment of complement fragment C4d levels.
• Inhibition studies using protamine sulfate.
• Comparison of oligonucleotides with varied internucleotide linkages, sequences, and structures.

Main Results:

• PS-oligos caused concentration-dependent prolongations in clotting times and reduced hemolytic complement activity.
• Activated partial thromboplastin time was more sensitive to PS-oligo inhibition than PT or TT.
• Reduced hemolysis correlated with increased C4d levels.
• Protamine sulfate inhibited PS-oligo effects, indicating involvement of charged internucleotide linkages.
• Effects were linked to internucleotide linkages, not nucleotide sequence.

Conclusions:

• PS-oligos directly inhibit human blood coagulation and complement activation in vitro.
• The phosphorothioate backbone's charged linkages are responsible for these activities.
• Modified oligonucleotides with altered linkages may offer safer antisense therapeutics.