Activated natural killer cell tumor retention and cytokine production in colon tumor using a tissue-isolated model

Abstract

Background: Adoptively transferred activated natural killer (A-NK) cells are capable of selectively infiltrating solid tumors, but only at low efficiency when administered systemically. It is unclear if human A-NK cells can be retained in tumor tissue and, if so, what is their action. We investigated intratumor A-NK cell retention and in situ cytokine production, using an xenogeneic ex vivo tissue-isolated tumor model, which permits direct intraarterial infusion.

Materials and methods: Human colon adenocarcinoma (HT-29) was implanted in the ovarian fat pad of nude rats. The tumors were perfused ex vivo 25 to 30 days postimplant with a known number of cells, and the effluent was collected over time. The number of human A-NK cells and cell surface antigen expression of cells infused and exiting the tumor were calculated, using cell counts and flow cytometry, respectively. Frozen sections were stained with Giemsa and also immunostained for the presence of interleukin-2, -4, and -10, tumor necrosis factor alpha (TNF-alpha), and interferon.

Results: Six perfusions with 8 x 10(6) A-NK cells were performed. The mean number of infused A-NK cells that remained in the tumor at the completion of perfusion was 4.74 x 10(6) (59.2%). No differences were noted in cellular phenotype between the infused cells and the cells exiting the tumor: expression of the markers CD45 (97.5% vs 94. 5%), CD14 (0 vs 0), CD3 (3.83% vs 2.83%), and CD56 (86% vs 83%) was unchanged, P > 0.05. Microscopic examination of tumor sections showed tumor surrounded by A-NK cells, with some tumor nests infiltrated by A-NK cells. In situ immunopositivity for interleukin-2 (2/6), interleukin-4 (3/6), interleukin-10 (2/6), and TNF-alpha (2/6) specimens was observed. Immunostaining for interferon-gamma was negative. Conclusions. The retention of A-NK cells in the transplanted human colon tumor tissue was found to be efficient (59.2 %) in this model. Although perfusion time was limited, A-NK cells were able to infiltrate the tumor and initiate cytokine production.