The effect of intraportal mannitol on the short-term in vivo distribution of radiolabelled A-LAK cells in rats

The effectiveness of adoptive immunotherapy of cancer using LAK cells and IL-2 depends on the accumulation of transferred effector cells at the tumour sites. In vitro LAK cell have been demonstrated to have broad cytotoxic activity to a wide variety of tumour cells in a non-major-histocompatibility complex-restricted manner, and independent of the presence of tumour specific antigens. LAK cells have now been used effectively in a small number of human trials. The effective delivery of these cells to the tumour site in vivo is one of the main aspects of this type of immunotherapy that requires further investigation. Conventional systemic infusion has shown a limited migration pattern of LAK cells. In this study the degree of entrapment of LAK cells in organs following local infusion has been determined. The large size and rigidity of LAK cells may be one mechanism which restricts the distribution of these cells. In addition, in this study the effect of mannitol, a hyperosmotic agent which increases the space between vascular endothelial cells, on the uptake of intraportal LAK cells into liver has been determined. A-LAK cells are obtained by culture of N.W.P. lymphocytes with IL-2. In this study A-LAK cells were characterised by typical morphologic appearance, cell surface phenotype, and cytotoxic specificity. Purified A-LAK cells were morphologically large granular lymphocytes, 67%-90% of which showed the surface marker phenotype of NK/LGL cells (OX8). The population contained few pan-T cells (only 4.0-6.5% of cells expressing OX19). No B cell surface marker lg was detectable in the A-LAK cell population. These cells showed high ability to lyse YAC-1 and P-815 cultured tumour target cells in 4h 51Cr-release cytotoxic assays. At an Effector:Target ratio of 40:1 A-LAK cells lysed 70% P815 and 100% YAC-1 cells. After labelling A-LAK cells with 51Cr, the effect of intraportal infussion of 30% mannitol on the distribution of intraportally infused A-LAK cells in liver was studied. The trafficking studies were carried out in three groups. In Group 1, 51Cr labelled A-LAK cells were systemically infused through the tail vein of rats as a control group. In Group 2 and Group 3, A-LAK cells were infused into syngeneic rats through the portal vein without or following prior portal infusion of 30% mannitol. Two hours after LAK cell administration the rats were sacrificed and the radioactivity in liver, lung, spleen, blood, MLN, kidney and brain were measured to determine the distribution of A-LAK cells to these organs. The results showed that intraportal mannitol was associated with an increased percentage of LAK cells in the liver compared with regionally infused LAK cells without mannitol (54% vs 24%; P<0.0005). The administration of intraportal mannitol was also associated with increased distribution of A-LAK cells into the brain (0.26% vs 0.08%; P<0.05) and MLN (0.05% vs 0.02%; P<0.05). There was no significant increase in uptake of A-LAK cells in lung (8.39% vs 6.01 %), spleen (1.00% vs 0.98%), or kidney (1.44% vs 1.78%) following intraportal mannitol. There was no significant increase of the A-LAK cell distribution to the liver by regional infusion without mannitol (24% vs 16%). Systemic injection gave greater A-LAK uptake into lungs (15.65%) than portal injection (6.05%, P=0.05). There was no significant differences of cell distribution in spleen, MLN, kidney and brain by these two routes of infusion. These results showed that mannitol has the effect of increasing the distribution of LAK cells into the liver. Augmentation of LAK cell accumulation in the liver may help to enhance the tumouricidal activity of these cells to hepatic metastases. The results of this thesis suggest area_s for further investigation into the effect of mannitol and other agents on A-LAK cell uptake into the liver following regional infusion.