Regulatory RNA inhibition in human phagocytes using cell-penetrating PNA oligonucleotides

Here, we determined the ability of peptide nucleic acid (PNA) oligomers, coupled to different cell-penetrating peptides (CPPs), to interfere in regulatory RNA circuits of human blood-derived leukocytes. Using RNA-seq, FACS and confocal microscopy we identified octaarginin as a CPP enabling PNA delivery and sequence-dependent RNA inhibition in blood-derived myeloid cells at nanomolar concentration. At 200 nM, an R8-PNA targeting immune-regulatory microRNA-155 was delivered into nearly 100 % of human macrophages within 24 hours without apparent cytotoxicity, and globally de-repressed microRNA-155 target-mRNAs. This was not observed when coupling the PNA inhibitor to a K3 instead of the R8 peptide. We suggest that CPP choice is a fundamental success-determining factor for therapeutic RNA-inhibition in human myeloid leukocytes. Expression profiles of poly(A)-RNA from WT and microRNA-155 deficient U937 monocytes and scrambled or anti-miR-155 PNA inhibitor treated primary human macrophages. Both cell types were treated with LPS, before recording the consequences of miR-155 deficiency by RNA-seq. Two replicates were recorded, each replicate representing an equimolar pool of RNA from two (U937) or three (primary macrophages) independent experiments).