Effects of ablating protein complexes in cancer as surrogate for drug treatment

Future cancer therapy promises to be improved by development of drugs that target mutant proteins, rather than non-specific treatments such as chemotherapy. Biologists need to know how proteins in cancer cells send signals that influence the growth characteristics of the tumours. The development of reagents to interfere with particular functions of cancer-related proteins gives us tools to look at complex cellular processes and potentially uncover new therapy targets. Our development of small protein inhibitors of cancer proteins has been used by us to focus on two proteins (called RAS and LMO2) which are important in a large number of human cancers. We will use our inhibitors to study responses in cells and find out which molecules are affected using a new technology that gives a read-out of all the molecules in a cancer cell. The outcome of this work will be the identification of proteins involved or altered in cancer and important as targets for new drugs.

Validation of cancer therapy targets is a major goal in molecular and cell biology studies as is the development of cancer drugs against these validated targets. The mutational spectrum in human cancer is critical to the development and maintenance of overt disease. While mutant proteins are tumour-specific, making them attractive drug targets, many are inside the cell and involved in protein complexes with various functions, ranging from transcription to signal transduction. Our work on the RAS and LMO2 oncogenic proteins, using antibody fragments to block their respective protein complexes, showed that their protein-protein interactions are necessary for the tumourigenicity of those tumours in which mutant protein (in the case of RAS) or aberrant expression (in the case of LMO2) occurs. The new research planned is to examine the physiological consequences of interference with RAS-effector interaction in lung cancer and acute myeloid leukaemia with KRAS or NRAS mutation respectively and interference with the LMO2 transcriptional complex in T cell acute leukaemia and diffuse large B cell lymphoma. The method to be used is to generate cell lines expressing competitive amounts of single antibody domain (intracellular antibody) fragments targeted to RAS or LMO2 and to study the transcriptomes using new generation RNA sequencing to generate digital data sets of mRNA species. This will provide a molecular definition of genes expressed and the relative levels of expression in the presence or absence of functional RAS or LMO2 complexes. Further, in characterising tumourigenicity of cell lines expressing anti-RAS single domain antibodies, we can evaluate the significance of blockading RAS-effector interactions in lung cancers with mutation of the epidermal growth factor receptor but with non-mutated RAS or acute myeloid leukaemias with FLT3 mutation but non-mutated RAS. The outcome of this work will be the identification of molecules involved or altered in cellular responses after ablating specific functions of RAS or LMO2. These molecules will be of biological relevance and importance as accessory therapy targets.