Poor cellular accumulation of therapeutic nucleic acids is a limiting step of wide application of nucleic acids as therapeutics. Various approaches for nucleic acids delivery are used such as electroporation, viral and chemical vectors. However, electroporation has relatively high cell mortality (29%) and viral vectors may lead to mutagenesis and carcinogenesis as a result of viral genome integration into DNA of host cells and undesired immune responses. Non-viral delivery systems, such as cationic liposomes, demonstrated high efficiency of intracellular delivery of nucleic acids along with low immunogenicity and the simplicity of their synthesis that makes very promising for the purpose of gene therapy.
Antitumor vaccines on the base of ex vivo generated dendritic cells (DCs) pulsed with nucleic acids encoding tumor-associated antigens (TAA) are perspective technique to treat cancer due to the ability of modified DCs to trigger tumor-specific cytotoxic T cell responses, prevent systemic metastasis and tumor relapse. Nevertheless, one of the main impediments in this therapy is poor uptake of nucleic acids by DC.
We studied the ability of liposomes on the base of novel cholesterol based cationic lipids and lipid-helper DOPE (dioleoylphosphatidylethanolamine) to deliver DNA (plasmid pEGFP encoding enhanced green fluorescent protein) and RNA (total RNA isolated from BHK-IR780 cells expressing endogenously EGFP) into murine immature DC and their bone marrow progenitors.
The cationic lipids contain one (X2, S1, S2 and S3) or two (2X3) cholesterol residues or long-chain hydrocarbon substituent (2D3) linked with spermine. Data show that liposomes 2X3-DOPE, 2D3-DOPE, X2-DOPE and S2-DOPE display high transfection efficiency with respect to DNA (30-47% of DC progenitors and up to 57% of immature DC were transfected) and RNA (up to 57% of cells were transfected). The studied lipids exhibited an efficiency of DNA and RNA delivery in DCs several times higher in comparison with Lipofectamine 2000. DCs loaded with mRNAs encoding a set of tumor-associated antigens using liposomes 2X3-DOPE or 2D3-DOPE caused 3-5 fold suppression of metastasis number in a model of murine B16 melanoma in vivo.
The obtained data show that the novel spermine-based polycationic lipids are efficient tool for nucleic acids delivery.