Unravelling signatures of clonal response, resistance and evolution of high-risk essential thrombocythaemia at single-cell resolution

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In recent years, substantial advances in genetic testing have transformed our ability to carry out analysis of the entire human DNA code (human genome) and its mirroring RNA sequence (transcriptome), a surrogate for gene expression, from minute amounts of genetic material. This has propelled our ability to analyse single-cells at a genome/transcriptome wide level, which is now possible with unprecedented scale and resolution. These techniques are now poised for clinical use in precision medicine to improve diagnosis, risk stratification, disease monitoring and drug discovery. However, translating this powerful new technology through to direct clinical application requires integration of clinical and scientific expertise as well as evidence of how such an approach might benefit patients in a specific disease area. Ideally, this should be assessed within the context of a prospective clinical trial. This is the overarching problem/challenge that we aim to address in this research i.e. to provide proof of principle of the clinical utility of single-cell genomic analysis in a specific disease with an unmet need: high-risk essential thrombocythaemia (ET). ET is a form of chronic myeloproliferative neoplasm (MPN) characterised by a high platelet count, increased risk of thrombosis, bleeding and progression to aggressive blood cancers such as myelofibrosis (MF) and acute myeloid leukaemia (AML). MPNs arise from the earliest form of a blood cell, a stem cell, in the bone marrow which acquires a genetic change, leading to changes in gene expression which promotes uninhibited cell growth. Patients with ET have a variable disease course, with approximately 20% of high-risk patients developing resistance to standard first line therapy with hydroxycarbamide (HC). HC-resistant ET patients have an increased risk of disease progression to MF or AML and significantly reduced overall survival (26% at 10 years). Current therapies prevent bleeding or thrombosis through reduction of blood cells but none have shown ability to prevent disease progression. Ruxolitinib, a JAK inhibitor, is the first targeted treatment approved in MPNs in MF and polycythaemia vera (PV). It has been evaluated in high-risk HC-resistant or intolerant ET in the MAJIC study with variable responses observed and in a proportion, disease progression occurred on this treatment. This project aims to apply single-cell technology to understand the cellular and molecular differences in patients with HC-resistant/intolerant ET on the MAJIC study and in doing so, determine mechanisms for ruxolitinib response (or lack of response) and disease progression. Firstly, I will compare clinical information of these patients with mutation testing of over 30 genes associated with MPNs to understand if these influence ruxolitinib response and disease progression. Guided by these results, I will then carry out extensive single-cell genomic analysis of mutated bone marrow stem and progenitor cells of subgroups of patients to understand whether gene expression differences between mutated and non-mutated stem cells at a single-cell level identifies mechanisms influencing responses and disease progression. There is increasing evidence to support that the bone marrow environment is abnormal in blood cancers and may influence the disease course. Therefore, I will also study non-mutated stem cells in HC-resistant/intolerant ET patients using single-cell analysis to determine if increased number of these cells or their altered gene expression may be associated with response to treatment, disease progression or side effects of the treatment.This clinical research training fellowship will provide a state-of-the-art training in the application of single-cell genomics in clinical medicine. We anticipate a direct, tangible impact on patient outcomes through better approaches to diagnose, monitor (identify biomarkers of early relapse) and treat high-risk ET.