Method of targeted gene delivery using viral vectors
Methods and compositions are provided for delivering a polynucleotide encoding a gene of interest to a target cell using a virus. The virus envelope comprises a cell-specific binding determinant that recognizes and binds to a component on the target cell surface, leading to endocytosis of the virus. A separate fusogenic molecule is also present on the envelope and facilitates delivery of the polynucleotide across the membrane and into the cytosol of the target cell. The methods and related compositions can be used for treating patients having suffering from a wide range of conditions, including infection, such as HIV; cancers, such as non-Hodgkin's lymphoma and breast cancer; and hematological disorders, such as severe combined immunodeficiency.
1. A method of delivering a polynucleotide encoding an antigen of interest to a dendritic cell, the method comprising:
infecting the dendritic cell with a recombinant lentivirus, wherein the recombinant lentivirus comprises:
(i) the polynucleotide encoding the antigen of interest and wherein the polynucleotide is operably linked to an expression control sequence, and wherein the polynucleotide further comprises an R sequence and an U5 sequence from a 5′ lentiviral long terminal repeat (LTR), and a self-inactivating lentiviral 3′ LTR, and
(ii) a viral envelope comprising a fusogenic molecule which is a Sindbis virus E1 glycoprotein, and a cell-specific binding determinant which is an anti-DEC-205 antibody that (a) targets dendritic cells, (b) is separate from the fusogenic molecule, and (c) is incorporated into the envelope of the recombinant lentivirus.
2. The method of claim 1 , wherein the antibody comprises the light and the heavy constant chain regions of human IgG1.
3. The method of claim 1 , wherein the cell-specific binding determinant further comprises immunoglobulin alpha and immunoglobulin beta.
4. The method of claim 1 , wherein the antibody is a single chain antibody.
5. The method of claim 4 , wherein the single chain antibody is fused with a transmembrane domain from another protein.
6. The method of claim 1 , wherein the R sequence and the U5 sequence are from HIV.
7. The method of claim 1 wherein the self-inactivating 3′ LTR comprises a U3 element with a deletion of its enhancer sequence.
8. The method of claim 7 , wherein the self-inactivating 3′ LTR is a modified HIV 3′ LTR.
9. The method of claim 1 , wherein the recombinant lentivirus is pseudotyped.
10. The method of claim 1 , wherein the antigen of interest is a tumor antigen.
11. The method of claim 1 , wherein the envelope comprises a Sindbis virus E2 glycoprotein.
12. The method of claim 1 , wherein the recombinant lentivirus further comprises a stimulatory molecule.