Characterisation of Chromatin Landscapes of Pre-leukaemic and Leukaemic Stem Cells in Core Binding Factor AML and their Response to Epigenetic Therapy

急性髓系白血病(AML)是一种相对常见的血癌，在我作为血液学家的临床工作中占很大比例。虽然强化化疗方案(自20世纪80年代以来变化很小)在大约80%的患者可以实现暂时的临床缓解，5年存活率只有15%，而不需要经历干细胞移植等高风险的程序。因此，一个关键的临床问题是在基因组水平上了解是什么原因导致所有这些患者复发，以期预防。RUNX1是一个在血细胞发育中至关重要的基因，在其研究界被称为“血细胞发育的主要调节因子”。多年来，我们团队一直致力于了解这种基因及其相互作用，我们拥有相当多的专业知识。RUNX1的突变也恰好是AML最常见的缺陷之一。我们的目标是将我们对该基因的专业知识与AML复发的临床问题联系起来，并旨在解决4个关键目标：1)表征白血病前期干细胞化疗是一种相当迟钝的工具，主要杀死分裂最快的细胞。“白血病前干细胞”指的是具有基因变化的血细胞，这些血细胞最终可能会导致癌症，但尚未完全癌变。我们认为这些细胞分裂缓慢，因此对化疗有抵抗力。因此，它们可以在明显处于缓解状态并导致未来复发的患者中存活。我们的目标是从RUNX1突变患者的骨髓中纯化白血病前干细胞。然后，我们将使用最先进的技术，如单细胞RNA测序来了解他们的生物学。2)改善急性髓细胞白血病缓解期的监测实现临床缓解的患者通过观察其疾病特异性基因的拷贝数来监测复发。然而，我们仍然没有好的标志物来预测复发。通过表征白血病前干细胞，这项工作旨在确定可以帮助患者更好地监测的临床相关基因靶点。3)调查Wilms Tumour 1(WT1)基因的作用我们已经从文献中了解到，WT1基因水平高的患者复发很快。WT1传统上被癌症研究人员认为是一种抗癌基因，所以我们的目标是调查为什么它在急性髓细胞白血病中有害。我们将在细胞系上进行实验，以找出WT1在AML基因组中的作用位置以及它与其他分子的相互作用。我们还将有选择地改变基因水平，以确定其在复发白血病中的作用。4)调查表观遗传药物治疗的反应在伯明翰，我们领导了一项II期临床试验(Ravva-目前最大的表观遗传疗法试验)，招募了259名患者，这些患者接受了2种改变基因表达的药物-阿扎替丁和维拉尼坦的联合治疗。然而，目前尚不清楚这些药物在基因组水平上对AML细胞的确切作用。我们旨在通过比较这些药物受益的患者和没有受益的患者的基因表达变化来阐明这一点。这可能会突出特定的基因或信号通路，从而可能产生更持久的临床反应。

Acute Myeloid Leukaemia (AML) is a relatively common blood cancer that forms a significant proportion of my clinical work as a Haematologist. Whilst intensive chemotherapy regimens (which have changed very little since the 1980s(!) can achieve temporary clinical remission in about 80% of patients, 5 year survival is only 15% without going through risky procedures such as a stem cell transplant. Consequently, a key clinical question is to understand on a genomic level what causing all these patients to relapse, with a view to prevention.RUNX1 is a gene that is of utmost importance in blood cell development and has been dubbed in its research community as the "master regulator of blood cell development". We have worked on understanding this gene and its interactions for many years in our group and we have considerable expertise. Mutations in RUNX1 also happen to be one of the commonest defects in AML. We aim to tie our expertise with this gene with the clinical problems of relapse in AML and aim to tackle 4 key objectives:1) Characterise Pre-leukaemic Stem CellsChemotherapy is a rather blunt instrument that kills mainly the fastest-dividing cells. 'Pre-leukaemic stem cells' are blood cells with genetic changes that can eventually give rise to cancer but aren't yet fully cancerous. We propose that these cells are slow-dividing and hence resistant to chemotherapy. They can therefore survive in patients who are apparently in remission and lead to future relapses. We aim to purify pre-leukaemic stem cells from bone marrow of patients with RUNX1 mutations. We will then use state-of-the art technology such as single cell RNA sequencing to understand their biology.2) Improve AML monitoring in RemissionPatients that achieve clinical remission are monitored for relapse by looking at the number of copies of a gene specific to their disease. However we still do not have good markers that would predict relapse. By characterising pre-leukaemic stem cells, this work aims to identify clinically relevant genes targets that can help patients be monitored better.3) Investigate the Role of the Wilms Tumour 1 (WT1) GeneWe already know from the literature that patients that have high levels of the WT1 gene relapse quickly. WT1 has traditionally been thought of by cancer researchers as a gene protective against cancer so we aim to investigate why it harmful in AML. We will perform experiments on cell lines to find out where in the AML genome WT1 acts and other molecules it interacts with. We will also selectively alter levels of the gene in order to determine its role in relapsed leukaemia.4) Investigate Response to Treatment by Epigenetic DrugsAt Birmingham, we have led a phase II clinical trial (RAvVA - currently the largest epigenetic therapies trial of its kind) which recruited 259 patients that were treated with combinations of 2 drugs that alter gene expression - Azacitidine and Vorinostat. However, it is not currently known what exactly these drugs are doing on a genomic level on the AML cells. We aim to elucidate this by comparing changes in gene expression in patients for whom these drugs have benefited compared to those for who they haven't. This can potentially highlight specific genes or signalling pathways to target that might give a more durable clinical response.