Lymphokine-activated Killer Activity in Long-Term Cultures with Anti-CD3 plus Interleukin 2: Identification and Isolation of Effector Subsets

Peripheral blood lymphocytes cultured in recombinant interleukin 2 during 3 to 5 days (short-term cultures) develop the ability to lyse natural killer-resistant tumor lines and fresh tumor cells, i.e., express lympho-kine-activated killer (LAK) function. Phenotypic analysis has shown these cells to be natural killer cells, i.e., CD 16+ and/or Leu 19+ cells. CD3+, CD16+ T-cells, instead, develop very low LAK function in these cultures. We recently reported the development of long-term (up to 21 days) cultured cells with LAK activity by stimulation with OKT3 + interleukin 2 (IL2). These culture conditions repeatedly resulted in a several hundredfold expansion in cell number. Specific LAK activity on Day 14 of culture was comparable to that of 3-day LAK cultures and could be further enhanced by the addition of interleukin 1α,α or γ-interferon. Total LAK activity was greatly increased in OKT3 + IL2 cultures over that found in short-term cultures. Isolation of effectors mediating LAK function in long-term cultures stimulated with OKT3 4 IL2 showed that both CD3+, CD16-cells and CD16+,CD3-cells tested on Day 14 of culture expressed equivalent levels of LAK activity as shown by lysis of natural killer-resistant targets, HL60 and Daudi. Further dissection of the subpopulations developing LAK activity demonstrated that, in addition to CD16+CD3+cells, CD3+CD4-CD8-cells and Leu 19+CD3-CD16-cells also developed high LAK activity in long-term cultures with OKT3 + IL2. Further, long-term culture with OKT3 4+ IL2 induced increases in the numbers not only of CD3+, CD4+, CD8+ cells but also of CD16+, CD3+ and Leu 19+CD3-CD16-cells. Although there is a significant increase in the number of CD3+, CD8+ cells, neither these, nor the CD3+, CD4+ cells, mediate LAK activity to the same extent as the populations mentioned above.