Methods of making and using a cell penetrating peptide for enhanced delivery of nucleic acids, proteins, drugs, and adenovirus to tissues and cells, and compositions and kits
A peptide-POD with ability to penetrate and deliver fluorophores, siRNA, DNA and quantum dots to cells in culture and retinal and ocular tissues in vivo is provided herein. POD couples to adenovirus vectors, enhancing tropism for certain cells, potentially providing a safer and more efficacious method to deliver molecules to ocular and other tissues in vivo. POD constructs are therapeutic delivery vehicles for treating cells and tissues, including ocular cells and tissues suffering from retinal degeneration.
1. A method for delivery of a therapeutic agent to a cell or a tissue or for transduction into the cell or cells of the tissue, the method comprising:
providing the therapeutic agent conjugated or linked to at least one copy of an amino acid sequence comprising: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, or SEQ ID NO: 17, wherein the amino acid sequence is effective for delivery to the cell or transduction into the cell of the therapeutic agent; and contacting the cell or tissue with the therapeutic agent conjugated or linked to the amino acid sequence, wherein the therapeutic agent is delivered to or transduced into the cell or the cells of the tissue, wherein the cell or the tissue is ocular.
2. The method according to claim 1 , wherein the cell or the tissue is in culture.
3. The method according to claim 1 , wherein the cell or the tissue is in vivo.
4. A method for delivery of a therapeutic agent to a cell or a tissue or for transduction into the cell or cells of the tissue, the method comprising:
providing the therapeutic agent conjugated or linked to at least one copy of an amino acid sequence selected from the group consisting of: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, or SEQ ID NO: 17, wherein the amino acid sequence is effective for delivery to the cell or transduction into the cell of the therapeutic agent;
formulating the conjugated or linked therapeutic agent as a medicament for diagnosing, prognosing, or treating a condition in a mammalian subject; and
contacting the cell or tissue with the therapeutic agent conjugated or linked to the amino acid sequence, wherein the therapeutic agent is delivered to or transduced into the cell or the cells of the tissue, wherein contacting the cell or the tissue comprises administering the therapeutic agent conjugated or linked to the amino acid sequence to the subject by an intravitreal route.
5. A method for delivery of a therapeutic agent into cells or tissue of an eye, the method comprising:
providing a PEGylated peptide for overall delivery (POD) comprising at least one copy of an amino acid sequence comprising: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, or SEQ ID NO: 17, wherein the amino acid sequence is effective for delivery to or transduction into the cells of the therapeutic agent;
mixing the PEGylated POD and the therapeutic agent, thereby forming a PEGylated POD-therapeutic agent complex; and
contacting the cells or tissue with the complex, wherein the therapeutic agent is transduced into the cells or the tissue of the eye; and,
observing reduced retinal degeneration in the cells or the tissue of the eye in comparison to uncontacted cells.
6. The method according to claim 5 , wherein contacting the cells or the tissue of the eye comprises contacting in vivo.
7. The method according to claim 5 , wherein contacting the cells or the tissue of the eye comprises contacting in vitro.
8. The method according to claim 5 , wherein prior to mixing, the method further comprises preparing a cDNA nucleotide sequence encoding the therapeutic agent amino acid sequence.
9. The method according to claim 8 , wherein the cDNA nucleotide sequence encodes a protein that is a neurotrophic factor selected from a group consisting of glial cell line-derived neurotrophic Factor (GDNF).
10. The method according to claim 5 , wherein observing reduced retinal degeneration further comprises determining eye function by electroretinography.