The aim of our study was to induce in vitro a secondary response to a C57BL/6 Friend leukemia (FBL-3) by C57BL/6 lymphoid cells, to document their enhanced anti-tumor cytotoxicity in vitro and to study their anti-tumor effect in vivo by tumor neutralization (Winn assay) and tumor therapy of progressive FBL-3 in C57BL/6 mice. Spleen cells (Ci) from mice immunized once in vivo with FBL-3 were cultured for 5 days with either x-irradiated FBL-3 (CiFx), C57BL/6 spleen cells (CiCx), BALB/c spleen cells (CiBx) or without any x-irradiated cells (Ci-cult.) and their activity subsequently was assayed in vitro and in vivo. CiFx cells were markedly more cytotoxic in vitro by the 51Cr release assay than were CiCx or CiBx and were most effective in tumor neutralization (Winn assay). For adoptive immunotherapy, mice given lethal (104) FBL-3 i.p. on day 0 received the cultured spleen cells on day 1. Treatment with 5 × 106 CiFx cured 14 of 14 mice whereas 19 of 20 mice given 5 × 106 CiCx or CiBx died with tumor. For adoptive chemoimmunotherapy of advanced leukemia, mice were given 107 FBL-3 i.p. on day 0. On day 5, they received cyclophosphamide (CY) plus cultured spleen cells. Untreated mice died by day 14. CY alone prolonged survival but all mice died within 4 weeks. Treatment with CY plus 5 × 106 Ci-cult. or CiBx was no more effective than CY alone—40 of 40 mice died with tumor. However, CY plus 5 × 106 CiFx cured 16 of 22 mice. Thus, cells generated in a secondary response in vitro were not only cytotoxic in vitro and effective in tumor neutralization, but were also curative in immunotherapy of an early leukemia and in chemoimmunotherapy of an advanced leukemia.

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This work was supported by Grants CA10777 and CA 05231 from the National Cancer Institute, National Institutes of Health.

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