Cholesterol-functionalized DNA/RNA heteroduplexes cross the blood-brain barrier and knock down genes in the rodent CNS

Achieving regulation of endogenous gene expression in the central nervous system (CNS) with antisense oligonucleotides (ASOs) administered systemically would facilitate the development of ASO-based therapies for neurological diseases. We demonstrate that DNA/RNA heteroduplex oligonucleotides (HDOs) conjugated to cholesterol or alpha-tocopherol at the 5' end of the RNA strand reach the CNS after subcutaneous or intravenous administration in mice and rats. The HDOs distribute throughout the brain, spinal cord and peripheral tissues and suppress the expression of four target genes by up to 90% in the CNS, whereas single-stranded ASOs conjugated to cholesterol have limited activity. Gene knockdown was observed in major CNS cell types and was greatest in neurons and microglial cells. Side effects, such as thrombocytopenia and focal brain necrosis, were limited by using subcutaneous delivery or by dividing intravenous injections. By crossing the blood-brain barrier more effectively, cholesterol-conjugated HDOs may overcome the limited efficacy of ASOs targeting the CNS without requiring intrathecal administration.

Automated summary

This study demonstrates that DNA/RNA heteroduplex oligonucleotides (HDOs) conjugated to cholesterol or alpha-tocopherol can regulate endogenous gene expression in the central nervous system when administered systemically, with limited side effects. The HDOs effectively distribute throughout the CNS and suppress the expression of target genes by up to 90%, showing potential for ASO-based therapies for neurological diseases.