IMMUNOTHERAPY OF LIVER METASTASES OF HUMAN GASTRIC-CARCINOMA WITH INTERLEUKIN 2-ACTIVATED NATURAL-KILLER-CELLS

To better understand the events occurring during immunotherapy of live r metastases with effector cells, we have developed a clinically relev ant animal model in which both effector-tumor cell interactions and su rvival can be evaluated. A cell line of human gastric carcinoma (HR) m etastatic to the liver has been established from a patient's liver bio psy. HR cells (10 x 10(6)) injected intrasplenically metastasize into the liver of immunosuppressed nude mice, with micrometastases detectab le histologically by day 4 and macrometastases by day 7. The animals s ubsequently develop ascites and die between days 30 and 40 after tumor injection. To investigate early metastatic events in the liver, HR ce lls were transduced with a plasmid containing both the lacZ gene under the control of the CMV promoter and NeoR gene. Transfectants selected for neomycin resistance were lacZ gene positive and stained blue in t he presence of a beta-galactosidase substrate, -bromo-4-chloro-3-indol yl-beta-D-galactopyranoside (X- Gal). These transfectants (HRLZ) remai ned lacZ gene positive for at least 25 passages in vitro. Injected int rasplenically, an HRLZ clone grew inva sively in nude mice and formed liver metastases comparably to parental tumor cells. The number and lo calization of blue X-Gal positive tumor cells were followed in liver t issues of animals sacrificed at various times, from 1 h to 28 days pos tinjection of HRLZ cells. HRLZ cells were seen in liver blood vessels and sinusoids within 1 h after injection, and the progressive growth o f micrometastases and macrometastases could be followed with precision by X-Gal staining. On day 3 after injection of HRLZ cells, numerous m icrometastases were established containing 12-16 tumor cells. When the se 3-day established HRLZ micrometastases were treated by the intraspl enic infusion of interleukin 2 (IL2)-activated human natural killer (N K) cells selected by IL2-induced adherence to plastic (A-NK) and syste mic IL2, nearly all liver micrometastases were eliminated within 24 h after a single transfer of A-NK cells (P < 0.001). This xenogeneic mod el was also used for adoptive immunotherapy of 7-day established liver macrometastases with human A-NK cells injected intrasplenically and e xogenous IL2 given i.p.. A significant decrease in the number of hepat ic metastases and the weight of livers (P < 0.003) in comparison with those of control mice was observed. Survival of mice treated with a si ngle dose of A-NK cells plus exogenous IL2 (60,000 international units /mouse twice daily for 5 days) was significantly increased (P = 0.001) as compared to control mice treated with Hanks' balanced salt solutio n or IL2 alone. In this xenogeneic model of liver metastasis, human A- NK cells were remarkably effective in eliminating both micro- and macr ometastases and significantly prolonging survival of mice treated loco regionally with adoptive immunotherapy and IL2.