Fusion protein and use thereof
| Title: | Fusion protein and use thereof |
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| Patent Number: | 8,481,492 |
| Publication Date: | July 09, 2013 |
| Appl. No: | 12/997659 |
| Application Filed: | June 15, 2009 |
| Abstract: | The invention relates to a fusion protein and a method for the generation of the fusion protein of the invention. Further, the invention relates to the use of the fusion protein of the invention for the generation of induced pluripotent cells. Moreover, the invention relates to a composition comprising at least one fusion protein of the invention. |
| Inventors: | Edenhofer, Frank (Erftstadt, DE); Hadenfeld, Manal (Cologne, DE); Peitz, Michael (Bonn, DE); Thier, Marc-Christian (Bad-Munstereifel, DE) |
| Assignees: | Life & Brain GmbH (Bonn, DE) |
| Claim: | 1. A Fusion protein comprising: a) a protein transduction domain (PTD) comprising 6 to 12 basic amino acids; and b) at least one transcription factor, wherein said transcription factor is a stem cell factor. |
| Claim: | 2. The fusion protein of claim 1 , wherein the PTD further comprises hydrophobic amino acids. |
| Claim: | 3. The fusion protein of claim 1 , wherein the PTD further comprises at least one of TAT, Penetratin, HSV-VP22, Transportan, K-FGF, Oligoarginine, Arg-9, and peptides consisting of combinations of Arginine and Lysine residues. |
| Claim: | 4. The fusion protein of claim 1 , wherein the PTD is comprised of no more than 20 amino acids. |
| Claim: | 5. The fusion protein of claim 1 , further comprising a nuclear localization signal (NLS). |
| Claim: | 6. The fusion protein of claim 1 , wherein the fusion protein comprises an artificial transactivation domain (ATAD). |
| Claim: | 7. The fusion protein of claim 1 , further comprising a linker selected from the group consisting of Proline, Glycine, Alanine, Leucine, Glutamic acid, Threonine, and Serine. |
| Claim: | 8. The fusion protein of claim 1 , further comprising at least one end group selected from the group consisting of Alanine and Glycine. |
| Claim: | 9. The fusion protein of claim 1 , said fusion protein comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. |
| Claim: | 10. A composition comprising the fusion protein of claim 1 and at least one stabilizer. |
| Claim: | 11. The composition of claim 10 , wherein the stabilizer is a lipid rich serum. |
| Claim: | 12. A method for generating induced pluripotent cells, the method comprising the step of: a. contacting at least one fusion protein as defined in claim 1 with at least one target cell. |
| Claim: | 13. The method of claim 12 , wherein the step of contacting is carried out for a duration of from 6 hours to 60 days. |
| Claim: | 14. The method of claim 12 , wherein the at least one target cells is selected from the group consisting of embryonic stem cells, adult stem cells, and somatic cells of mammals. |
| Claim: | 15. The fusion protein of claim 1 , wherein the stem cell factor is selected from the group comprising Oct4, Sox2, nanog, klf4, lin28, hTERT, myc, and SV40. |
| Claim: | 16. The method of claim 14 , wherein the cells are derived from at least one of the group consisting of cells of flies, worms, and lower eukaryotes. |
| Claim: | 17. The method of claim 16 , wherein the flies are D. melanogaster , the worms are C. elegans , and the lower eukaryotes are yeasts. |
| Claim: | 18. The method of claim 17 , wherein the yeasts are S. cerevisiae. |
| Current U.S. Class: | 514/165 |
| Patent References Cited: | 2005-110565 April 2005 WO 2006/113181 October 2006 WO 2008/105630 September 2008 WO 2009/032456 March 2009 WO 2009/067757 June 2009 |
| Other References: | Krosl et al., Nature Medicine, 9(11), 1428-1432, 2003. cited by examiner Bosnali Manal, et. al.; “Generation of transducible versions of transcription factors Oct4 and Sox2”, Biological Chemistry, Walter De Gruyter GMBH & Co., Berlin, DE, vol. 389, No. 7, Jul. 1, 2008, pp. 851-861, XP009118395; ISSN: 1431-6730. cited by applicant Heng Boon Chin, et al., “Induced Pluripotent Stem Cells (iPSC)—can direct delivery of transcription factors into the cytosol overcome the perils of permanent genetic modification?”, Minimally Invasive Therapy & Allied Technologies: Official Journal of the Society for Minimally Invasive Therapy; Mitat, Informa Healthcare, London, vol. 17, No. 5, Jan. 1, 2008, pp. 326-328, XP009118397; ISSN: 1365-2931. cited by applicant J. Krosl, et al., “In vitro expansion of hematopoietic stem cells by recombinant TAT-HOXB4 protein”, Nature Medicine, Nature Publishing Group, New York, NY, vol. 9, No. 11, Nov. 1, 2003, pp. 1428-1432, XP003016542, ISSN: 1078-8956. cited by applicant Landry, Josette-Renée, et al., “Runx genes are direct targets of Scl/Tall in the yolk sac and fetal liver.”, Blood, Mar. 15, 2008, vol. 111, No. 6, pp. 3005-3014, XP002545673; ISSN: 0006-4971. cited by applicant Bian Jing, et al., “Effect of cell-based intercellular delivery of transcription factor GATA4 on ischemic cardiomyopathy.”, Circulation Research, Jun. 8, 2007, vol. 100, No. 11, pp. 1626-1633, XP002545674, ISSN: 1524-4571. cited by applicant Heng B C, et al., “Incorporating protein transduction domains (PTD) within intracellular proteins associated with the ‘stemness’ phenotype. Novel use of such recombinant ‘fusion’ proteins to overcome current limitations of applying autologous adult stem cells in regenerative medicine?”, Medical Hypotheses, Eden Press, Penrith, US, vol. 64, No. 5, Jan. 1, 2005, pp. 992-996, XP004952000, ISSN: 0306-9877. cited by applicant Heng B C, et al., “Incorporating protein transduction domains (PTD) within recombinant ‘fusion’ transcription factors. A novel strategy for directing stem cell differentiation?”, Biomedicine and Pharmacotherapy, Elsevier, Paris, FR, vol. 59, No. 3, Apr. 1, 2005; pp. 132-134, XP004861305, ISSN: 0753-3322. cited by applicant |
| Primary Examiner: | Monshipouri, Maryam |
| Attorney, Agent or Firm: | Faegre Baker Daniels LLP |
| Accession Number: | edspgr.08481492 |
| Database: | USPTO Patent Grants |
| Language: | English |
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