Novel Binding Partners of PBF in Thyroid Tumourigenesis
Lead Research Organisation:
University of Birmingham
Department Name: Clinical and Experimental Medicine
Abstract
Thyroid cancer is a relatively common cancer whose incidence is rising. Whilst numerous genes have well characterised roles in thyroid cancer, new therapeutic targets are vital to improved treatment success. We have identified a new gene which is highly expressed in thyroid tumours and which can cause cells to become cancerous. Most proteins work co-operatively, binding to other proteins within cells and influencing their functions. We have found that our protein of interest, PBF, binds to 3 other proteins. We now wish to widen this by considering all possible binding partners of PBF in thyroid cells through a technique called mass spectrometry. Following this, we would validate these findings and then examine precisely what effect such interactions have on some of the known functions of PBF. In terms of outcomes, we would hope to identify 3 to 5 novel proteins, which we would then be able to examine in human thyroid tumours, to see if they show any association with cancer outcome. We may thus identify new proteins which can be targeted in thyroid cancer, given that up to 30% of tumours recur, causing significant complications and death.
Technical Summary
BACKGROUND
Thyroid cancer is the 8th most common cancer, and its incidence is rising. We have shown that PTTG Binding Factor (PBF) is over-expressed in differentiated thyroid cancer and induces tumours in nude mice. PBF was initially speculated to be a putative surface glycoprotein. Our recent published and unpublished data have delineated a role for PBF in the binding of 3 proteins - p53, NIS and MCT8 - and the regulation of their function.
AIMS AND OBJECTIVES
Given that PBF is a transforming gene which is over-expressed in thyroid cancer and binds other proteins in thyroid cells, we now wish to (i) search for other potential binding partners of PBF; (ii) examine their impact on PBF function, and (iii) discern their expression and clinical associations in thyroid tumours. Our hypothesis is that PBF interacts with numerous proteins at the cell membrane and internally, and that oncogenic expression of PBF in human cancer disrupts normal cellular processes in addition to those we have already discovered.
DESIGN AND METHODOLOGY
Our central model is differentiated thyroid cancer. We will use K1 papillary thyroid cells which retain thyroid follicular cell differentiation, are amenable to transfection, and have low endogenous PBF expression. K1 cells will be transfected with our existing HA-tagged PBF and control vectors, and immunoprecipitations carried out. Subsequently, mass spectrometry will be carried out using our recently installed state-of-the-art mass spectrometers. Positive interactions will be investigated through pull-downs and co-IPs. Validated novel interactions will be assessed functionally by manipulating expression of interacting partners (transfection and siRNA) in thyroid cells and examining 3 known end-points of PBF action. Finally, expression of novel binding partners will be determined in our existing and expanding cohort of 80 normal and tumourous thyroid specimens, with gene expression related to markers of clinical outcome.
SCIENTIFIC AND MEDICAL OPPORTUNITIES
Thyroid cancer is the commonest endocrine malignancy. We are the only group currently publishing on PBF, and we have well-validated antibodies, mutants, vectors, expression assays, siRNAs and biological readouts for the gene. Our recent papers on PBF have been published in Oncogene and the Journal of Cell Science, and we currently have a paper under revision with Cancer Research. PBF thus remains a relatively uncharacterised gene which is highly publishable, and which we lead on.
PBF has been implicated in the initiation and promotion of transformed cell growth. This project will identify proteins which bind to PBF in thyroid cells, and may therefore identify novel interactions and pathways which could be amenable to commercial exploitation as therapeutic modalities in thyroid cancer.
Thyroid cancer is the 8th most common cancer, and its incidence is rising. We have shown that PTTG Binding Factor (PBF) is over-expressed in differentiated thyroid cancer and induces tumours in nude mice. PBF was initially speculated to be a putative surface glycoprotein. Our recent published and unpublished data have delineated a role for PBF in the binding of 3 proteins - p53, NIS and MCT8 - and the regulation of their function.
AIMS AND OBJECTIVES
Given that PBF is a transforming gene which is over-expressed in thyroid cancer and binds other proteins in thyroid cells, we now wish to (i) search for other potential binding partners of PBF; (ii) examine their impact on PBF function, and (iii) discern their expression and clinical associations in thyroid tumours. Our hypothesis is that PBF interacts with numerous proteins at the cell membrane and internally, and that oncogenic expression of PBF in human cancer disrupts normal cellular processes in addition to those we have already discovered.
DESIGN AND METHODOLOGY
Our central model is differentiated thyroid cancer. We will use K1 papillary thyroid cells which retain thyroid follicular cell differentiation, are amenable to transfection, and have low endogenous PBF expression. K1 cells will be transfected with our existing HA-tagged PBF and control vectors, and immunoprecipitations carried out. Subsequently, mass spectrometry will be carried out using our recently installed state-of-the-art mass spectrometers. Positive interactions will be investigated through pull-downs and co-IPs. Validated novel interactions will be assessed functionally by manipulating expression of interacting partners (transfection and siRNA) in thyroid cells and examining 3 known end-points of PBF action. Finally, expression of novel binding partners will be determined in our existing and expanding cohort of 80 normal and tumourous thyroid specimens, with gene expression related to markers of clinical outcome.
SCIENTIFIC AND MEDICAL OPPORTUNITIES
Thyroid cancer is the commonest endocrine malignancy. We are the only group currently publishing on PBF, and we have well-validated antibodies, mutants, vectors, expression assays, siRNAs and biological readouts for the gene. Our recent papers on PBF have been published in Oncogene and the Journal of Cell Science, and we currently have a paper under revision with Cancer Research. PBF thus remains a relatively uncharacterised gene which is highly publishable, and which we lead on.
PBF has been implicated in the initiation and promotion of transformed cell growth. This project will identify proteins which bind to PBF in thyroid cells, and may therefore identify novel interactions and pathways which could be amenable to commercial exploitation as therapeutic modalities in thyroid cancer.