Lymphokine-activated Killer Activity in Long-Term Cultures with Anti-CD3 plus Interleukin 2: Identification and Isolation of Effector Subsets1

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 lymphokine-activated killer (LAR) function. Phenotypic analysis has shown these cells to be natural killer cells, i.e., CD16+ 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 (11.2).These culture conditions repeatedly resulted in a several hundred fold 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 ¡nterleukin1/3, ß-, or -y-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 + 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, III 60 and Daudi. Further dissection of the subpopulations developing LAK activity demonstrated that, in addition to CD16*,CD3~ cells, CD3+, CD4-,CD8cells and Leu 19+,CD3-,CD16cells also developed high LAK activity in long-term cultures with OKT3 + IL2. Further, long-term culture with OKT3 + IL2 induced increases in the numbers not only of CD3*,CD4-,CD8cells 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.