A high density cell culture system for generation of human lymphokine-activated killer (LAK) cells for clinical use in adoptive immunotherapy

Abstract

A high density cell culture system has been developed for large-scale production of lymphokine-activated killer (LAK) cells from peripheral blood lymphocytes (PBLs) of malignant tumor patients. The system consists of a culture bag, which has two compartments separated by a semipermeable membrane, and an external rotator. The system allows for a long-term, at least 4 weeks, culture of LAK cells at high cell density in the inner compartment. The collected PBLs were first divided between the two culture bags and cultured without harvesting for 7-10 days to obtain LAK cells. Half of the LAK cells from each bag was administered to patients twice a week for clinical trials. Culture of the remaining half was continued following addition of a fresh culture medium. LAK cells were transferred to patients alternatively from each bag for the following 2-3 weeks. The total number of LAK cells administered amounted to 3.9-9.8 (mean 5.8) times more than the PBLs collected by leukapheresis (n = 10). The 5 x 10(6)/ml of PBLS of the initial concentration reached a maximum of 2 x 10(7)/ml. Our system does not need for a CO2 incubator. Cytotoxicity of the LAK cells was evaluated in 4 hr 51Cr release assays. Mean cytotoxicity at maximum cell density was 95.4 +/- 3.2% against ONS-12 (a human glioma cell) and 84.8 +/- 3.0% against Daudi cells (n = 10), but gradually decreased to about 50% at the end of fourth week of the culture period. Cell viability of the LAK cells was normally over 80% through the entire culture period.(ABSTRACT TRUNCATED AT 250 WORDS)