Use of a lentivirus/VSV pseudotype virus for highly efficient genetic redirection of human peripheral blood lymphocytes.
Genetic redirection of lymphocytes that have been engineered to recognize antigens other than those originally programmed by their rearranged germlines is a potentially powerful immunotherapeutic tool. The rationale for the protocol described here is that many cancers and persistent or latent viruses have developed strikingly similar mechanisms of evading attack by host immunity that can often be overcome by redirection of host lymphocytes using chimeric T-cell receptor (chTCR) genes. However, for human peripheral blood lymphocytes (PBLs), this is generally regarded as a technically demanding procedure with unacceptably low efficiency using either contemporary transfection methods or retroviral transduction. One of the main difficulties with retroviruses is their reliance on rapidly dividing cells for integration of their genomes carrying the desired chTCR. Here we describe a highly efficient protocol that uses a lentivirus/vesicular stomatitis virus pseudotyped virus to engineer CD3/CD28-stimulated human peripheral blood cells (i.e., primarily T cells), with near 100% efficiency.
Simmons
Anthony
A
Division of Virology, Children's Hospital, 301 University Boulevard, Galveston, Texas 77555-0372, USA. ansimmon@utmb.edu
Jantz
Kristina
K
D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D008214:Lymphocytes / Q000276:immunology*; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics*; D014759:Viral Envelope Proteins / Q000235:genetics
D018106:CD28 Antigens; D017252:CD3 Complex; D014759:Viral Envelope Proteins