Complexation of nucleic acids with disulfide-crosslinked cationic components for transfection and immunostimulation
The present invention is directed to a polymeric carrier cargo complex, comprising as a cargo at least one nucleic acid (molecule) and disulfide-crosslinked cationic components as a (preferably non-toxic and non-immunogenic) polymeric carrier. The inventive polymeric carrier cargo complex allows for both efficient transfection of nucleic acids into cells in vivo and in vitro and/or for induction of an (innate and/or adaptive) immune response, preferably dependent on the nucleic acid to be transported as a cargo. The present invention also provides, pharmaceutical compositions, particularly vaccines and adjuvants, comprising the inventive polymeric carrier cargo complex and optionally an antigen, as well as the use of such the inventive polymeric carrier cargo complex and optionally an antigen for transfecting a cell, a tissue or an organism, for (gene-)therapeutic purposes as disclosed herein, and/or as an immunostimulating agent or adjuvant, e.g. for eliciting an immune response for the treatment or prophylaxis of diseases as mentioned above. Finally, the invention relates to kits containing the inventive polymeric carrier cargo complex and/or the inventive pharmaceutical composition, adjuvant or vaccine in one or more parts of the kit.
1. A method of stimulating, enhancing or increasing TNFα and/or INFα expression in a subject in need thereof comprising administering to the subject a polymeric carrier cargo complex comprising:
(a) as a carrier a polymeric carrier formed by disulfide-crosslinked cationic peptides or proteins selected from the group consisting of CR 12 C (SEQ ID NO: 6), CR 9 C (SEQ ID NO: 3), CR 7 C (SEQ ID NO: 1), CH 6 R 4 H 6 C (SEQ ID NO: 388), CHK 7 HC (SEQ ID NO: 389) and CH 3 R 4 H 3 C (SEQ ID NO: 390), said polymeric carrier comprising at least 2 of said cationic peptides or proteins which are disulfide crosslinked, and
(b) as a cargo at least one nucleic acid molecule,
thereby stimulating, enhancing or increasing TNFα and/or INFα expression in the subject.
2. The method of claim 1 , wherein the at least one nucleic acid molecule is an RNA.
3. The method of claim 2 , wherein the RNA is a GU rich RNA.
4. The method of claim 1 , wherein the at least one nucleic acid molecule is an immunostimulatory nucleic acid.
5. The method of claim 1 , wherein the at least one nucleic acid molecule is an immunostimulatory RNA (is RNA) or an mRNA.
6. The method of claim 1 , wherein the nitrogen/phosphate (N/P) ratio of the cationic components of the polymeric carrier to the at least one nucleic acid molecule is in the range of 0.1-20.
7. The method of claim 6 , wherein the nitrogen/phosphate (N/P) ratio of the cationic components of the polymeric carrier to the at least one nucleic acid molecule is in the range of 0.1-5.
8. The method of claim 7 , wherein the nitrogen/phosphate (N/P) ratio of the cationic components of the polymeric carrier to the at least one nucleic acid molecule is in the range of 0.1-1.
9. The method of claim 1 , wherein the polymeric carrier further comprises functional peptides or proteins.
10. The method of claim 9 , wherein the functional peptides or proteins are peptide or protein antigens or antigen epitopes.
11. The method of claim 1 , wherein the polymeric carrier further comprises a ligand.
12. The method of claim 11 , wherein the ligand is mannose.
13. The method of claim 1 , wherein disulfide-bonds are formed by the N- and C-terminal cysteine residues in the cationic peptides or proteins.
14. The method of claim 1 , wherein the subject has a cancer, a cardiovascular disease, an infectious disease, an autoimmune disease or an allergy.
15. The method of claim 1 , wherein the polymeric carrier cargo complex does not comprise a polyethylene glycol (PEG) polymer chain.
16. The method of claim 1 , wherein the cationic peptides or proteins are CR 12 C (SEQ ID NO: 6), CR 9 C (SEQ ID NO: 3), CR 7 C (SEQ ID NO: 1), CH 6 R 4 H 6 C (SEQ ID NO: 388), or CH 3 R 4 H 3 C (SEQ ID NO: 390).