A novel approach to immunomodulation of frozen human bone marrow with interleukin-2 for clinical application.
Interleukin-2 (IL-2) activation of fresh or frozen bone marrow (BM) in vitro generates killer cells with potent anti-tumor effect both in vitro and in vivo. The IL-2-activated BM (ABM) retains the capacity to reconstitute the hematopoietic system in an autologous bone marrow transplantation (ABMT) setting. The killer cells lose their cytotoxicity if the ABM undergoes the procedures of freezing and thawing. Therefore, for clinical application, the ABM has to be generated after thawing a frozen stock of BM before ABMT. The thawed BM cells are fragile and may undergo lysis, resulting in clump formation and cell loss. The frozen autograft also contains components of cryoprotectant mixture whose effects on the generation of ABM have not been defined. The present studies have been carried out to optimize a technique of handling the frozen BM for immunomodulation with IL-2 for 24 h at 37 degrees C prior to ABMT, with minimal loss of cells. IL-2-activation of BM was carried out in bags containing serum free medium which were designed to permit gaseous exchange. Addition of deoxyribonuclease (DNAse) (100 micrograms/ml of BM concentrate) immediately after thawing and the presence of heparin (20 units/ml) in the medium completely abrogated immediate or delayed clumping of cells. The presence of DNAse and/or heparin during in vitro culture did not affect the cell viability, cytotoxicity against tumor cells or the progenitor cell activity of the ABM; all these functions were well maintained even when BM was placed in culture immediately after thawing (without washing). There was no microbial contamination.(ABSTRACT TRUNCATED AT 250 WORDS)