Interleukin-2 gene-transduced human leukemic cells induce major histocompatibility complex-restricted and -unrestricted anti-leukemic effectors in mixed lymphocyte-tumor cultures

Abstract

To explore the feasibility of designing vaccination protocols in acute leukemia patients with cytokine gene-transduced leukemic cells, we studied in vitro the growth potential of human leukemic cells transduced with the interleukin-2 (IL-2), IL-7, or IL-7 plus IL-2 genes, as well as the capacity of generating both autologous and allogeneic cytotoxic lymphocytes directed against the parental cells. A lymphoblastic T-cell line, ST4, obtained from a patient in long-lasting complete remission, was retrovirally engineered with the IL-2, IL-7, and IL-7 plus IL-2 genes; in addition, clones releasing different amounts of the cytokines were obtained by limiting dilution. Mixed lymphocyte-tumor cultures (MLTCs) were set up with parental or transduced leukemic cells as stimulators and with autologous or allogeneic lymphocytes as responders. When nonirradiated ST4 parental cells or clones producing <50 international units (IU)/mL/10⁶ cells/72 hours of IL-2 were used as stimulators, leukemic overgrowth was observed in MLTCs within 16 days of culture. When clones producing >80 IU/mL/10⁶ cells/72 hours of IL-2 were used as stimulators, the proliferation of leukemic cells was blocked and the transduced leukemic cells were completely cleared from the cultures by day 16; repeated restimulations with IL-2-producing leukemic cells were required to sustain long-term lymphocyte survival. On the contrary, when IL-7- or IL-7-IL-2-producing cells were used as stimulators, only a delay in leukemic cell overgrowth was observed, and lymphocytes were completely cleared from the cultures after day 60. IL-7 production by the different clones ranged between 11 and 36 ng/mL/10⁶ cells/72 hours, whereas the highest IL-2-producing IL-7-IL-2 clone released 50 IU/mL/10⁶ cells/72 hours of IL-2. When the stimulator efficacy of the highest IL-2-producing clone (ST4/IL-2#A7) was compared with that of exogenous IL-2 plus parental cells, a 7-fold higher amount of exogenous IL-2 was required to achieve the same results obtained with IL-2-producing leukemic cells. Autologous and allogeneic long-term MLTCs (up to 35 days) with ST4/IL-2#A7 as the stimulator were capable of generating cytotoxic effectors equally endowed with both major histocompatibility complex (MHC) class I-unrestricted and -restricted activity against parental ST4 cells. By day 18 of both autologous and allogeneic cultures, a substantial proportion of CD56⁺ cells was consistently recorded; this was coupled to a predominantly MHC-unrestricted cytotoxic activity directed against parental ST4 cells. CD56⁺ cells decreased considerably at the end of the different MLTCs, together with the unrestricted cytotoxic activity. At this time, >50% of the cells were CD8⁺, and 55% of the activity could be blocked by an anti-MHC class I monoclonal antibody. The results of this study demonstrate that IL-2 gene-transduced human acute leukemia cells cocultured with both autologous and allogeneic lymphocytes are capable of inducing a strong MHC-unrestricted anti-leukemic activity and subsequently “educating” MHC class I-restricted anti-leukemic effectors. The evidence that the immunogenic potential of human leukemic blasts can be boosted after transfer of the IL-2 gene suggests that the possibility of using leukemic cells engineered to release IL-2 as a therapeutic vaccine needs to be explored further.