asialo-gm1-ganglioside and Uveal-Neoplasms

Metastases account for 90% of all cancer-related deaths, becoming a therapeutic problem. Approximately 50% of all uveal melanoma (UM) patients will develop metastases, mainly in the liver. Post-mortem analyses of livers from metastatic UM patients showed two different metastatic growth patterns: infiltrative and nodular. The infiltrative pattern exhibits tumor infiltration directly to the hepatic lobule and minimal angiogenesis. The nodular pattern shows clusters of tumor cells around the portal venules that efface the liver parenchyma. We recently demonstrated Natural Killer (NK) cells play a pivotal role in the control of hepatic metastases and the pigment epithelial-derived factor (PEDF) controls angiogenesis in the liver using our established ocular melanoma animal model. In this study we investigated the role of NK cells and PEDF in the development of metastatic growth patterns, as this can contribute to the development of novel therapeutics specific towards each growth pattern.. Our in vivo work showed two distinct metastatic growth patterns, the infiltrative and nodular, recapitulating the post-mortem analyses on human liver tissue. We discovered NK cells control the infiltrative growth. In contrast, PEDF controlled anti-angiogenic responses, showing higher MVD values compared to NK-depleted and WT animals. The myeloid lineage, comprised of monocytes, macrophages, and myeloid-derived suppressor cells, was reduced in the absence of NK cells or PEDF.. Our animal model recapitulates the metastatic growth patterns observed in the human disease. We demonstrated a role for NK cells in the development of the infiltrative growth pattern, and a role for PEDF in the development of the nodular pattern. The understanding of the complexity associated with the metastatic progression has profound clinical implications in the diagnostic and disease-management as we can develop and direct more effective therapies.

Topics: Animals; Antibodies; Cell Line, Tumor; Eye Proteins; Female; G(M1) Ganglioside; Gene Knockout Techniques; Killer Cells, Natural; Liver Neoplasms; Macrophages; Melanoma; Mice; Mice, Inbred C57BL; Monocytes; Myeloid-Derived Suppressor Cells; Neoplasm Transplantation; Nerve Growth Factors; Serpins; Uveal Neoplasms

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.. 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.. 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.. 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.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cytotoxicity, Immunologic; Disease Models, Animal; Female; Flow Cytometry; G(M1) Ganglioside; Histocompatibility Antigens Class I; Immunophenotyping; In Situ Nick-End Labeling; Injections, Intraperitoneal; Interferon alpha-2; Interferon-alpha; Killer Cells, Natural; Liver; Liver Neoplasms, Experimental; Lymphocyte Activation; Lymphocyte Depletion; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Recombinant Proteins; Spleen; Uveal Neoplasms

This study determined whether adenovirus-mediated transfer of the murine interferon-beta (AdCMVIFN-beta) gene protects against liver metastases arising from intraocular melanomas in mice.. A replication-deficient adenovirus vector (AdCMVIFN-beta) was used for the in vivo transfer of the murine IFN-beta gene into intraocular melanoma-bearing mice. AdCMVIFN-beta was injected either intravenously or directly into the intraocular melanomas. The effect of gene transfer on liver metastases was ascertained by histopathologic analysis of the livers and by measuring serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT), which are two enzymes associated with liver metastases in patients with uveal melanoma.. Mice treated with two intratumoral injections of AdCMVIFN-beta had a 68% reduction in metastatic liver lesions (P = 0.016) and a 51% reduction in liver enzyme levels compared with control mice (P = 0.02). However, the antimetastatic effect of AdCMVIFN-beta was not directly attributable to the adenovirus vector or virus-mediated cytolysis of tumor cells. Intravenous treatment with AdCMVIFN-beta resulted in an 86% reduction in the number of metastatic foci in the liver (P = 0.014) and a 61% reduction of serum AST levels compared with mice treated with AdCMVLacZ (P = 0.015). AdCMVIFN-beta treatment produced a sharp increase in the NK cell activity that was demonstrable in vivo and in vivo. In vivo depletion of NK cells by anti-asialo GM1 antibody abrogated the antimetastatic effects of AdCMVIFN-beta.. The results support the feasibility of activation of NK cell function through gene transfer as one possible therapeutic strategy for reducing hepatic metastases of uveal melanomas.

Topics: Adenoviridae; Alanine Transaminase; Animals; Antineoplastic Agents; Aspartate Aminotransferases; Enzyme-Linked Immunosorbent Assay; Female; G(M1) Ganglioside; Gene Transfer Techniques; Genetic Therapy; Genetic Vectors; Interferon-beta; Killer Cells, Natural; Liver Neoplasms, Experimental; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Reverse Transcriptase Polymerase Chain Reaction; Uveal Neoplasms