HTS for agents that increase recognition of tumors by cytotoxic lymphocytes

Haggerty, Timothy

Abstract

The ultimate aim of this proposal is to design improved therapies for malignant melanoma. This proposal will promote the development of a High Throughout Screening (HTS) system for assessing cell mediated cytotoxicity against human tumor cells, which is a critical component of tumor immune therapy. We specifically propose to apply HTS for the identification of agents that enhance cytotoxicity against melanomas, by augmenting the immune recognition of the melanoma target cells which is essential for lysis to occur. Screening of molecular libraries will be focused on changes in the tumor target cells, but will score all agents which positively modulate any aspect of cytotoxic recognition and effector mechanisms. The identification of additional agents that enhance immune-recognition is of great potential benefit for immunotherapy, and there is accordingly a need to find as many such molecules as possible to obtain the best candidates for therapeutic purposes. The detailed HTS assay for development will use cytotoxic T lymphocytes that recognize the melanoma antigens Melan-A/MART-1 and gp100 as they are presented by HLA-A2 in A375 cells. For the cytotoxicity assay we have chosen to use A375 tumor target lines transfected with Enhanced Green Fluorescence Protein (EGFP) that will be used to measure the lysis of target cells. These tumor cells cannot be lysed by specific T lymphocytes unless a specific treatment results in enhanced T cell recognition of the tumor targets. We propose to develop a test to rapidly evaluate a chemical's ability to increase the body's immune sytem in its capacity to recognize and kill melanoma tumor cells. The goal of these investigations is to find new drugs that will improve cancer treatment by helping the body to kill its own tumor. While this project is targeted at the destruction of melanoma tumor cells, the High Throughput Screening system we propose represents a new approach to drug development. When successfully implemented, this High Throughput Screen can be used to identify drugs that will help in the immune destruction of viruses and tumors, while helping to prevent transplant rejection and autoimmune diseases. ? ? ?