Ex vivo development, expansion and in vivo analysis of a novel lineage of dendritic cells
Disclosed herein are new methods of producing a novel line of dendritic cells. The method comprises subjecting a sample of hematopoietic stem/precursor cells to a first feeder culture system that is supplemented with a first set of factors and a second feeder culture system supplemented with a second group of factors. The disclosure also pertains to new cell types that may be used as cancer immunotherapy.
1. A method of producing dendritic cell (DC) progenitor cells, said method comprising:
culturing a population of hematopoietic stem/progenitor cells (HPCs) in a first feeder culture comprising kit ligand (KL), fms-like tyrosine kinase 3 ligand (FL), thrombopoietin (TPO), IL-3, IL-6 and basic fibroblast growth factor (bFGF) under conditions to produce a population of first expanded cells; wherein the first feeder culture does not comprise GM-CSF or IL-15;
culturing said first expanded cells in a second feeder culture comprising KL, FL, TPO, IL-3, IL-6, bFGF, GM-CSF and IL-15 under conditions to produce DC progenitor cells (DCPs).
2. The method of claim 1 , wherein the first feeder culture comprises cells engineered to produce KL, FL, TPO, IL-3, IL-6 and/or bFGF.
3. The method of claim 2 , wherein said first feeder culture comprises cells engineered via a viral vector comprising an expression construct comprising a sequence encoding KL, FL, TPO, IL-3, IL-6 and/or bFGF.
4. The method of claim 1 , wherein the second feeder culture comprises cells engineered to produce KL, FL, TPO, GM-CSF, and/or IL-15.
5. The method of claim 1 , wherein said first expanded cells possess HPC phenotypic characteristics.
6. The method of claim 1 , further comprising culturing said DC progenitor cells in a third culture supplemented with GM-CSF and IL-15 under conditions to produce a population of cells having a phenotype similar to myeloid DCs.
7. The method of claim 6 , wherein the third culture is feeder free.