Defining a protein hydroxylase tumour suppressor pathway

Normal cell growth and survival is tightly controlled by multiple overlapping biochemical pathways that work together to ensure that cells only divide at the appropriate time and only survive in the correct environment. When these fail-safe mechanisms go wrong the uncontrolled cell division that results can initiate the early stages of cancer development. These so called 'tumour suppressor' pathways are often inactivated in cancer due to mutations in the DNA of specific genes.

We have identified a gene that when experimentally mutated promotes tumourigenesis in cell models. Consistent with a role in human cancer this gene is also mutated in human tumour samples. This new tumour suppressor gene, called 'MINA', encodes a protein with enzyme activity. Through this activity MINA modifies other proteins with oxygen in a process called 'hydroxylation'. We propose that MINA normally helps promote healthy cell division and survival by hydroxylating proteins that control the expression, or 'transcription', of key cancer-associated genes.

In this proposal we aim to characterise the role and regulation of MINA in cancer. We will first investigate whether MINA is inactivated by mechanisms other than gene mutation. Specifically, we will ask whether the MINA protein is itself modified, and how this modification switches off its enzyme activity in cancer cells. Then we will use a combination of biochemical and genomic approaches to identify the pathways by which MINA activity suppresses cancer. We will use 'mass spectrometry', a biophysical technique that measure the mass of protein fragments, to identify new proteins that MINA hydroxylates, and a DNA sequencing technique called 'ChIP-seq' to identify gene transcription regulated by MINA. Finally, we will validate our findings by measuring our biomarkers of MINA regulation and targets in several hundred human tumour samples.

Through these approaches we hope to define a novel tumour suppressor pathway involving a hydroxylase enzyme. Because this class of enzyme is not widely appreciated to play an important role in cancer, the work will stimulate further research in this important area. Furthermore, by discovering the key players in this new tumour suppressor pathway the work may identify new therapeutic opportunities for cancer treatments. Importantly, because MINA has important but poorly characterised roles in other disease-associated pathways, particularly in the immune system, the findings will also be of interest to a wide variety of other medical researchers.

Deregulated protein modifications have been widely implicated in the pathogenesis of major human diseases. The discovery and characterisation of novel post-translational modifications (PTM) has opened new avenues of biomedical research and stimulated new therapeutic opportunities.

Hydroxylation is an under-studied but emerging PTM. Pioneering work on collagen and oxygen sensing highlighted the potential importance of hydroxylation in physiology and pathology, and related oxygenases have since been implicated in a variety of important disease-associated processes. However, several members of the hydroxylase family are poorly characterised and the extent and functions of protein hydroxylation in disease remain unclear.

We previously reported that a nuclear JmjC protein called 'Myc-Induced Nuclear Antigen' (MINA) is a histidine hydroxylase. Our functional studies indicate that MINA has tumour suppressor (TS) activity, consistent with the observation that it is mutated and inactivated in a variety of tumour types, including gastrointestinal cancers. However, a detailed understanding of the extent of MINA deregulation in cancer requires knowledge of other potential mechanisms of inactivation. Furthermore, the downstream consequences of MINA inhibition are not known. Here we propose to address these limitations by characterising the regulation of MINA by an oncogenic kinase and identifying novel cancer-relevant substrates and gene targets involved in MINA-dependent transcriptional control. Importantly, the findings will be extended into an independent cohort of several hundred gastric tumour samples in a combined approach involving targeted resequencing, gene expression analyses and histopathology.

The work will improve our understanding of protein hydroxylation and its role in fundamental cellular processes, define a novel tumour suppressor pathway, and highlight new therapeutic opportunities for the treatment of cancer and other diseases.

The work proposed here aims to significantly advance our understanding of a poorly characterised protein modification (hydroxylation) by defining its role in regulating transcriptional control in a novel tumour suppressor pathway. The work will deliver significant scientific advances that will benefit academic and industrial researchers working on cancer and other important diseases. Therefore, the project has the potential to deliver impact by enhancing health and in the future to lead to the development of novel therapeutic opportunities.

The areas in which this proposal will provide meaningful impact are:

1) Academic
The work will engage a number of international academic beneficiaries, including; fundamental cell biology researchers, researchers and clinicians with an interest in mechanisms of Aurora Kinase A-mediated tumourigenesis and gastrointestinal cancer, and immunologists investigating the role of pro-inflammatory T-cells in autoimmune disease (including our collaborator, Mark Bix).

2) Commercial
The identification of novel chemotherapeutic targets has classically followed the discovery and characterisation of oncogene signalling pathways and tumour suppressor genes. The work proposed here will be important to the pharmaceutical industry for two reasons. Firstly, it will highlight the risk of small molecule inhibition of the MINA hydroxylase in cancer (which is being promoted by some academic groups). Secondly, it will define a novel tumour suppressor pathway that will help identify new therapeutic opportunities. By delivering a very high level of scientific proof the work will provide a strong foundation for successful drug discovery.

Beyond cancer MINA is attracting interest as a therapeutic target in autoimmune disease due to an important role in transcriptional regulation of the TH17 differentiation program. Through the proposed work identifying the transcriptional targets of MINA the findings will be of importance to the commercial sector in this area.

3) Socio-economic
Cancer is a major worldwide societal and economic burden that is increasing due to a growing and ageing population. Several major cancers are still associated with poor survival rates, due to a variety of factors including poor treatment options, resistance to targeted therapies and heterogeneity complicating molecular classification. The work proposed here uses gastrointestinal cancers as a model to define a novel tumour suppressor pathway. Therefore, the findings will likely benefit patients in the future and also potentially reduce public costs and improve the quality of health care.

4) Training and education
This proposal involves combined biochemical, cell biology, genomic and pathology approaches that will provide exceptional training opportunities for the next generation of young scientists. This applies not only to the staff directly involved in the project, but also interns and undergraduate, MSc and PhD students that the Coleman group regularly host.