Interferon alpha 2b decreases hepatic micrometastasis in a murine model of ocular melanoma by activation of intrinsic hepatic natural killer cells.
OBJECTIVE
To investigate in a murine model the mechanism by which micrometastatic melanoma, which spreads from the eye to the liver, is controlled by interferon (IFN)-alpha 2b.


METHODS
Major histocompatibility (MHC) class I antigen (H-2, all haplotypes) expression in three murine melanoma cell lines (Queens, B16LS9, B16F10) was determined by flow cytometric immunophenotyping. The cell lines were heterotopically inoculated into the posterior compartments (PCs) of C57Bl/6 mice, and the mice were given intraperitoneal (IP) injections of IFN-alpha 2b or PBS for 1 or 4 days before enucleation at 7 days after inoculation. Groups of mice were made NK deficient or depleted with subcutaneous (s.c.) injection of anti-asialo GM1. The mice were killed at 28 days or 56 days (survival experiment) after inoculation, and the number of hepatic micrometastases was histologically determined. NK cells were isolated from the spleen and liver at necropsy, and propidium iodide labeled target-specific cytolysis was determined by flow cytometry. The micrometastases were evaluated for apoptosis and proliferation with TUNEL and MIB1 immunostaining, respectively, and TUNEL-to-MIB1 ratios were determined. Hepatic NK cells were immunostained with CD49b.


RESULTS
MHC class I antigen was expressed in the three cell lines in the order of Queens < B16LS9 < B16F10. All cell lines grew, were confined to the PC, and formed hepatic micrometastases. A decrease in micrometastases, an increase in target-specific cytolysis, and an increase in survival correlated with decreased HLA class I expression by the melanoma cells. The IFN-alpha 2b treatment resulted in a boost of intrinsic hepatic NK cells, demonstrated in NK-deficient but not NK-depleted mice. The treatment effect corresponded to increased apoptosis (TUNEL)-proliferation (MIB1) ratios in the micrometastases. Immunostaining demonstrated an increased number of intrahepatic NK cells associated with the micrometastases in treated groups.


CONCLUSIONS
Neoadjuvant IFN-alpha 2b results in decreased hepatic micrometastasis and increased survival time through increased intrinsic hepatic NK cell-mediated tumor apoptosis in a murine model of metastatic ocular melanoma.
Yang
Hua
H
Department of Ophthalmology, Emory University, Atlanta, Georgia 30322, USA.
Dithmar
Stefan
S
Grossniklaus
Hans E
HE
D000818:Animals; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D017209:Apoptosis; D003602:Cytotoxicity, Immunologic; D004195:Disease Models, Animal; D005260:Female; D005434:Flow Cytometry; D005677:G(M1) Ganglioside / Q000494:pharmacology; D015395:Histocompatibility Antigens Class I / Q000276:immunology; D016130:Immunophenotyping; D020287:In Situ Nick-End Labeling; D007274:Injections, Intraperitoneal; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000627:therapeutic use*; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology*; D008099:Liver / Q000276:immunology; D008114:Liver Neoplasms, Experimental / Q000188:drug therapy* / Q000276:immunology / Q000556:secondary; D008213:Lymphocyte Activation; D008212:Lymphocyte Depletion; D008546:Melanoma, Experimental / Q000188:drug therapy* / Q000276:immunology / Q000556:secondary; D051379:Mice; D008810:Mice, Inbred C57BL; D011994:Recombinant Proteins; D013154:Spleen / Q000276:immunology; D014604:Uveal Neoplasms / Q000188:drug therapy* / Q000276:immunology / Q000473:pathology
D000970:Antineoplastic Agents; D015395:Histocompatibility Antigens Class I; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D005677:G(M1) Ganglioside; C018835:asialo GM1 ganglioside