Human cancer cells xenografts to assess the efficacy of granulysin-based therapeutics.

Abstract

9-kDa Granulysin is a protein present in the granules of human activated cytotoxic T lymphocytes and natural killer cells. It has been shown to exert cytolytic activity against a wide variety of microbes: bacteria, fungi, yeast and protozoa. Recombinant isolated granulysin is also capable of inducing tumor cell death, so it could be used as an anti-tumor therapy. Our group has developed granulysin-based immunotoxins in order to target granulysin to tumor cells in vivo. We describe in this chapter the suitable animal model used for testing these immunotoxins against human tumor cells in preclinical assays. This method consists in the xenotransplantation of a given number of human tumor cells subcutaneously in nude mice of the Swiss nu/nu strain or homozygous for the nude gene. Nude mice are immune-deficient, with a very reduced number of T cells, being unable to reject efficiently allo- or xeno-transplanted tissues. Using this approach we follow tumor growth in the different experimental conditions assayed, in the presence or absence of treatment with granulysin alone or with the tumor-directed immunotoxins. We also estimate in this model the possible adverse effects of the treatment in the absence of tumor development. Finally, after sacrifice of the experimentation animals, we use several immunohistochemical assays to study the effect of the treatment on the tumors and the presence of apoptotic cell death markers in the tumor tissue.