Genetically engineered dendritic cells (DC) presenting specific antigens to T cells may be of great interest for immunotherapy. For this reason, the production of transgene‐expressing DC derived from CD34+ cells transduced either shortly after ex vivo purification or during their differentiation into DC were evaluated.

CD34+ cells were transduced with lentivectors encoding for GFP before or after 21 days of culture with FLT3‐ligand, thrombopoietin and stem cell factor and induction into DC with GM‐CSF+IL‐4 (G4) or G4+TNF (GT4). GFP and DC‐specific marker expression was assessed by flow cytometry, and allostimulatory capacity was evaluated on GFP+ and GFP− sorted cells.

Immature (G4‐induced) DC obtained from amplified CD34+ cells were transducible by lentiviral vectors while mature (GT4‐induced) DC were rather refractory. Moreover, since differentiated DC did not proliferate, large quantities of vectors were required to generate transgene‐expressing cells with this protocol. In contrast, greater numbers of both immature and mature GFP−expressing DC were obtained with CD34+ cells exposed to lentivector shortly after purification. By the time of DC induction, GFP+ cells had increased by approximately 170‐fold. After DC induction with G4, 32% of CD1a+, HLA‐DR+, or CD40+ cells expressed GFP. CD1a+E‐cadherin+GFP+ Langerhans‐like DC were also obtained. Incubation with TNF induced mature CD83+GFP+ DC that displayed a higher allostimulatory capacity than cells induced with G4 alone.

The transduction of a small number of CD34+ cells with minimal doses of lentivector may allow for the production of a large number of DC expressing selected antigens useful for immunotherapy. Copyright © 2001 John Wiley & Sons, Ltd.