Synthetic membrane-receiver complexes
Compositions comprising synthetic membrane-receiver complexes, methods of generating synthetic membrane-receiver complexes, and methods of treating or preventing diseases, disorders or conditions therewith.
1. A method of making an erythroid cell comprising at least 1,000 copies of an exogenous polypeptide comprising phenylalanine ammonia lyase (PAL), comprising:
i) introducing an exogenous nucleic acid encoding the exogenous polypeptide into an erythroid cell precursor; and
ii) culturing the erythroid cell precursor to provide the erythroid cell comprising the exogenous polypeptide,
wherein the erythroid cell comprising said exogenous polypeptide is not a hypotonically loaded cell.
2. The method of claim 1 , wherein the erythroid cell comprises at least 10,000 copies of the exogenous polypeptide.
3. The method of claim 1 , wherein the erythroid cell comprises at least 25,000 copies of the exogenous polypeptide.
4. The method of claim 1 , wherein the erythroid cell comprises at least 50,000 copies of the exogenous polypeptide.
5. The method of claim 1 , wherein the erythroid cell further comprises a second exogenous polypeptide which is surface-localized and comprises a phenylalanine transporter.
6. The method of claim 1 , wherein the erythroid cell is a reticulocyte.
7. The method of claim 1 , wherein the erythroid cell is a mature erythrocyte.
8. The method of claim 1 , wherein the erythroid cell lacks A and B antigens.
9. The method of claim 1 , wherein the erythroid cell comprises fetal hemoglobin.
10. The method of claim 1 , wherein the erythroid cell exhibits substantially the same osmotic membrane fragility as an isolated, unmodified, cultured or uncultured erythroid cell.
11. The method of claim 1 , wherein the exogenous polypeptide is glycosylated.
12. The method of claim 1 , wherein the exogenous polypeptide does not substantially diffuse out of the erythroid cell.
13. The method of claim 1 , wherein the exogenous nucleic acid comprises DNA.
14. The method of claim 1 , wherein the introducing step comprises transduction, transfection, or electroporation.
15. The method of claim 14 , wherein transfection comprises transfection with nanoparticles.
16. The method of claim 1 , wherein the culturing step comprises expanding the erythroid cell precursor by at least 20,000-fold in culture.
17. The method of claim 1 , which produces a population of erythroid cells wherein at least 70% of the cells are enucleated and comprise the exogenous polypeptide.
18. The method of claim 1 , wherein the exogenous polypeptide is not fused to an endogenous polypeptide.
19. The method of claim 1 , wherein the exogenous polypeptide is intracellular.
20. The method of claim 1 , wherein the exogenous polypeptide consists essentially of PAL.
21. The method of claim 1 , wherein the exogenous polypeptide consists of PAL.
22. The method of claim 1 , wherein the erythroid cell precursor comprises a CD34+ hematopoietic stem cell.
23. The method of claim 1 , wherein the erythroid cell precursor is a human cell.
24. The method of claim 1 , wherein the exogenous nucleic acid comprises RNA.
25. The method of claim 1 , wherein the erythroid cell comprising 1,000 copies of the exogenous polypeptide is an enucleated erythroid cell.