Compositions of a peptide targeting system for treating cancer
This invention describes a protein nanoparticle system for targeting siRNA or other drugs into tumors. The basis of the protein system is elastin-like peptides that self-assemble once exposed to the nucleic acid of the siRNA. Specific targeting peptides are fused to the core ELP structure by standard genetic engineering techniques. These targeting peptides confer specific binding of the nanoparticle to receptors on the surface of tumor cells and allow for uptake of the nanoparticle into the tumor cells.
1. An isolated polypeptide capable of self-assembling with a siRNA molecule into a nanoparticle in solution, the isolated polypeptide comprising:
a) an assembly domain (AD),
b) a cell targeting domain (CTD) comprising the amino acid sequence of any one of SEQ ID NO:1-18 or SEQ ID NO: 20-27, the CDT being positioned at the N-terminal end of the isolated polypeptide; and
c) a nucleic acid binding domain (NBD) capable of binding to an siRNA molecule, the NBD including a cationic domain, the NBD being positioned at the C-terminal end of the isolated polypeptide.
2. The polypeptide of claim 1 , wherein the NBD comprises the amino acid sequence of SEQ ID NO:58 or SEQ ID NO:59.
3. The polypeptide of claim 1 , wherein the AD comprises an elastin-like polypeptide (ELP).
4. An isolated cDNA polynucleotide comprising a nucleotide sequence that encodes the polypeptide of claim 1 .
5. The isolated cDNA polynucleotide of claim 4 , wherein the nucleotide sequence comprises any one of SEQ ID NO:28-54.
6. A recombinant expression construct comprising the polynucleotide of claim 4 .
7. An isolated recombinant host cell comprising the polynucleotide of claim 4 .
8. A method of targeting a siRNA molecule to a target cell, comprising:
obtaining one or more polypeptides, each comprising an assembly domain (AD), a nucleotide binding domain (NBD), and a cell targeting domain (CTD);
contacting the siRNA molecule with the NBD of one or more of the polypeptides;
assembling the AD of one or more polypeptides to generate an assembled polypeptide;
contacting the assembled polypeptide with the target cell; and
contacting the CTD of the assembled polypeptide with a biological molecule present outside of the target cell,
wherein one or more of the one or more polypeptides is the polypeptide of claim 1 .
9. The method of claim 8 , wherein the cell comprises a tumor cell.
10. The method of claim 9 , wherein the tumor cell is a human bladder cell, a human breast cell, a human colon cell, a human liver cell, a human lung cell, a human neuroblastoma cell, a human ovarian cell, a human pancreatic cell, a human prostate cell, or a human skin cell.
11. The isolated polypeptide of claim 1 , wherein the cationic domain of the NBD comprises an oligolysine or an oligoarginine.
12. The polypeptide of claim 1 , wherein the nucleic acid binding domain (NBD) comprises the amino acid sequence (Val-Gly-Lys K -Gly), wherein K is an integer greater than 4, or the amino acid sequence (Val-Gly-Arg R -Gly), wherein R is an integer greater than 4.
13. A nanoparticle comprising one or more polypeptides according to claim 1 and one or more siRNA molecules, the nanoparticle having an exterior and an interior, wherein
the one or more siRNA molecules are disposed at the interior of the nanoparticle and are complexed to one or more of the NBDs of the one or more polypeptides,
the AD of each of the one or more polypeptides is assembled with one or more of the ADs of the other polypeptides, the ADs of the one or more polypeptides surrounding the one or more siRNA molecules, and
the CTD of the one or more are disposed at the exterior of the nanoparticle.
14. A population of a plurality of nanoparticles according to claim 12 , wherein the nanoparticles are substantially monodisperse.
15. A population according to claim 14 , wherein the nanoparticles have an average size less than 100 nm.
16. A population according to claim 15 , wherein the AD of each of the one or more polypeptides comprises an elastin-like polypeptide (ELP), and the cationic domain of the NBD comprises an oligolysine or an oligoargninine.