Gene Delivery to Human B-Precursor Acute Lymphoblastic Leukemia Cells

Abstract

Autologous leukemia cells engineered to express immune-stimulating molecules may be used to elicit antileukemia immune responses. Gene delivery to human B-precursor acute lymphoblastic leukemia (ALL) cells was investigated using the enhanced green fluorescent protein (EGFP) as a reporter gene, measured by flow cytometry. Transfection of the Nalm-6 and Reh B-precursor ALL leukemia cell lines with an expression plasmid was investigated using lipofection, electroporation, and a polycationic compound. Only the liposomal compound Cellfectin showed significant gene transfer (3.9% to 12% for Nalm-6 cells and 3.1% to 5% for Reh cells). Transduction with gibbon-ape leukemia virus pseudotyped Moloney murine leukemia virus (MoMuLV)-based retrovirus vectors was investigated in various settings. Cocultivation of ALL cell lines with packaging cell lines showed the highest transduction efficiency for retroviral gene transfer (40.1% to 87.5% for Nalm-6 cells and 0.3% to 9% for Reh cells), followed by transduction with viral supernatant on the recombinant fibronectin fragment CH-296 (13% to 35.5% for Nalm-6 cells and 0.4% to 6% Reh cells), transduction on human bone marrow stroma monolayers (3.2% to 13.3% for Nalm-6 cells and 0% to 0.2% Reh cells), and in suspension with protamine sulfate (0.7% to 3.1% for Nalm-6 cells and 0% for Reh cells). Transduction of both Nalm-6 and Reh cells with human immunodeficiency virus–type 1 (HIV-1)–based lentiviral vectors pseudotyped with the vesicular stomatitis virus-G envelope produced the best gene transfer efficiency, transducing greater than 90% of both cell lines. Gene delivery into primary human B-precursor ALL cells from patients was then investigated using MoMuLV-based retrovirus vectors and HIV-1–based lentivirus vectors. Both vectors transduced the primary B-precursor ALL cells with high efficiencies. These studies may be applied for investigating gene delivery into primary human B-precursor ALL cells to be used for immunotherapy.