Research projects

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Evaluation of combination therapies targeting DNA damage repair signalling pathways in acute myeloid leukaemia

Supervisors: heather.jorgensen@glasgow.ac.uk, helen.wheadon@glasgow.ac.uk, xu.huang@glasgow.ac.uk

Project description: 

Acute myeloid leukaemia (AML) is an aggressive cancer affecting mostly adult and elderly patients. It has a very poor 5-year survival of <20% in the UK. Oncogene driven genomic instability leads to accumulation of DNA damage; this is a key and common phenomenon in AML cells, that could be therapeutically targeted. Targeted inhibitor efficacy as single agents in clinical trials has been limited, partly due to the activation of alternative compensatory DNA damage response (DDR) pathways therefore rational combination strategies may be more appropriate.

We previously established a family of histone demethylases as critical and selective oncogenic factors in AML. Genetic knockdown or pharmacological inhibition of family members was sufficient to induce apoptosis in a broad spectrum of human AML cell lines and primary patient blasts with no effect on normal haematopoiesis, indicating leukaemia cells are more sensitive to inhibition thereby offering a potential therapeutic window.

We hypothesise that a combination treatment of DDRi with histone demethylase inhibitor may result in enhanced cytotoxic effects in human AML cells. Our preliminary data indeed show promising synergistic lethality activity with this combination in human AML cell line suspension culture. In this project we wish to further evaluate histone demethylase inhibitors as single agents or in combination with DDRi in primary patient blasts to inform future clinical trial design.

Methodology:

We have a collection of individual primary AML patient samples and normal human bone marrow cells in our Glasgow biobank. An in vitro co-culture system has been established in our laboratory that mimics in vivo bone marrow microenvironment, that has been demonstrated to be a reproducible and reliable system for assessing clonal function and drug efficacy in primary AML cells. Functional assays including cell proliferation assay, cell apoptosis assay and colony formation assays will be performed. These data will be correlated with gene expression in each patient sample.

Aims:

1. To validate the efficacy of histone demethylase inhibitor monotherapy or in combination with DDRi in stratified AML patient blasts.
2. To evaluate biomarker expression pattern in AML patient samples following single or combination treatment.

References:

1. ME Massett, et al (2021): A KDM4A-PAF-1-mediated epigenomic network is essential for acute myeloid leukemia cell self-renewal and survival. Cell Death Dis 12(6):573. doi: 10.1038/s41419-021-03738-0.
2. L Monaghan, et al (2019): The emerging role of H3K9me3 as a potential therapeutic target in acute myeloid leukaemia. Front Oncol doi.org/10.3389/fonc.2019.00705

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Investigating the role of autophagy and mitochondrial function in leukaemic stem cells

Supervisors: Vignir.Helgason@Glasgow.ac.uk, Eric.Kalkman@Glasgsow.ac.uk

Our lab is interested in biological processes that contribute to drug resistance in myeloid leukaemias, with particular focus on leukaemic stem cells (LSCs).

Introduction:

Chronic myeloid leukaemia (CML) is caused by a reciprocal chromosomal translocation within a haemopoietic stem cell. This leads to transcription of BCR-ABL, a constitutively active tyrosine kinase that is necessary to induce CML.

The development of the tyrosine kinase inhibitor (TKI) imatinib significantly improved the life expectancy of CML patients; however, we have shown that disease persistence is caused by the remarkable ability of CML LSCs to survive, despite complete BCR-ABL inhibition mediated by TKI treatment1,2. Acute myeloid leukaemia (AML) is a more heterogeneous, involving different disease-causing genetic mutations. First line treatment for AML patients consists of chemotherapy, aiming at inducing remission. Generally, five-year survival rate in AML remains at a dismal 20%.

Activating internal tandem duplication mutations in FLT3 (FLT3-ITD), detected in about 20% of AML, represents driver mutations and a valid therapeutic target in AMLFLT-ITD. However, although new FLT3 inhibitors have begun to show promising clinical activity it is unlikely that they will have durable effects as single agents In recent years there has been resurgence in interest in autophagy, energy metabolism and mitochondria function as a possible area for development of novel anti-cancer agents.

We recently developed improved protocols for autophagy and metabolic assays in rare LSCs and highlighted mitochondrial oxidative phosphorylation (OXPHOS) as a metabolic dependency in CML LSCs3. Primitive AML cells have also been shown to depend on increased mitochondrial respiration4,5. We will therefore further investigate mitochondrial metabolism and through validation of drug-repurposing screen, identify new clinically applicable drugs that inhibit OXPHOS in CML, and in AML where improved therapy options with acceptable toxicities are urgently needed.

Hypothesis/Aims:

Our working hypothesis is that autophagy and deregulated mitochondrial metabolism in LSCs renders them sensitive to inhibition of the ULK1 autophagy complex and pathways that sustain mitochondrial OXPHOS. Our first aim is to use complementary functional and omic approaches to further assess the dependency of CML/AML LSCs to recycle or oxidise major mitochondrial fuels (objective 1). Our second aim is to test ULK1 inhibitors6 and FDA-approved OXPHOS inhibitors (which we have recently identified through drug-repurposing screening), in combination with TKI treatment, for eradication of CML and AMLFLT3-ITD/TKD LSCs (objective 2).

Deliverables:

This project will therefore promote identification of a core fuel pathway signature of CML/AML LSCs and a set of new potentially selective LSC-specific metabolic drug targets (objective 1). The student will also use state-of-the-art in vitro and in vivo models to test clinically relevant drugs, which will in the longer term, facilitate the translation of our findings into the clinic, with the overall aim for CML and AML LSC eradication.

References:

1. Holyoake, T.L., et al. Immunological reviews. 106-23 (2015).
2. Hamilton, A., et al. Blood. 1501-10 (2012).
3. Kuntz, E.M., et al. Nat Med. 1234-40 (2017).
4. Skrtic, M., et al. Cancer Cell. 674-88 (2011).
5. Lagadinou, E.D., et al. Cell Stem Cell. 329-41 (2013).
6. Ianniciello, A., et al. Sci Transl Med. (2021)

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Understanding and exploiting immunogenic cell death to treat cancer

Supervisors:  stephen.tait@glasgow.ac.uk, Ed.Roberts@glasgow.ac.uk

Project description: 

Cell death both prevents and treats cancer. New anti-cancer therapies that directly target cell death are revolutionising the treatment of cancer. Nevertheless, a major problem to effective cancer treatment is the emergence of treatment resistance. We are interested in killing cancer cells in a way that alerts the immune system to the presence of cancer - in essence harnessing the power and adaptability of tumour immunity to eradicate cancer. Our focus is on mitochondrial apoptosis - we have found that blocking caspase protease activity makes cell death immunogenic.

This PhD project will seek to understand why such caspase-inhibited cell death is immunogenic - both at the level of the dying cell but also in understanding how the immune system responds to the dying cell. We will employ novel approaches to inhibit caspase activity. In short, this exciting project will focus on discovery science with clear translation impact for cancer treatment.

The techniques it will entail will be varied but include CRISPR-Cas9 genome editing, super-resolution microscopy and in vivo modelling of cancer. This will be a collaborative project jointly supervised by Stephen Tait and Ed Roberts. You will join a young, dynamic interdisciplinary research team, based withing the CR-UK Beatson Institute with access to cutting edge technology.

References:

1. Targeting immunogenic cell death in cancer. Ahmed A, Tait SWG. Mol Oncol. 2020 Dec;14(12):2994-3006. doi: 10.1002/1878-0261.12851. Epub 2020 Dec 1.
2. Mitochondrial permeabilization engages NF-κB-dependent anti-tumour activity under caspase deficiency

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Investigation of new therapeutic approaches to combat viral-associated cancer

Supervisor: Dr. Joanna B. Wilson
Email: Joanna.wilson@glasgow.ac.uk

Background: Epstein-Barr virus (EBV) is a human Herpesvirus that is associated with several forms of human cancer. The virus leads to a life long infection, avoiding eradication by the immune system and has evolved intriguing tricks to do this. In the lab we investigate the role of key viral genes in disease processes, their mechanism of action at the molecular level and how they perturbate the immune system. Central to this are the mechanisms by which viral proteins disrupt normal cellular processes. New insights into viral action permit an exploration into novel therapeutic approaches to combat EBV-associated cancer.

Aims: To assess the efficiency of novel treatments in killing viral infected tumour cells and to explore the mechanism of action of such drugs in targeting the function of selected viral proteins

Techniques: The project will involve the use of several molecular biological and genetical techniques to examine protein, DNA, RNA and molecular interactions, as well as immunological and cell culture methods, also high resolution imaging.

References:

1. AlQarni, S., Al-Sheikh, Y., Campbell, D., Drotar, M., Hannigan, A., Boyle, S., Herzyk, P., Kossenkov, A., Armfield, K., Jamieson, L., Bailo, M., Lieberman, P., Tsimbouri, P. and Wilson, J.B. (2018) Lymphomas driven by Epstein-Barr virus nuclear antigen-1 (EBNA1) are dependant upon Mdm2. Oncogene (in the press)
2. Gnanasundram, S.V., Pyndiah, S., Daskalogianni, C., Armfield, K., Nylander, K. , Wilson, J.B. and Fåhraeus,, R. (2017) PI3Kd activates E2F1 synthesis in response to EBNA1-induced mRNA translation stress. Nature Communications 8:2103, doi:10.1038/s41467-017-02282-w Gao, X., Lampraki, E., Al-Khalidi, S., Qureshi, M.A., Desai, R. and Wilson, J.B. (2017) N-acetyl cysteine (NAC) ameliorates Epstein-Barr virus latent membrane protein 1 induced chronic inflammation. PLoS-ONE 12 (12) e0189167
3. Deschamps, T., Quentin, B, Leske, D.M., MacLeod, R., Mompelat, D., Tafforeau, L., Lotteau, V., Baillie, G.S., Gruffat, H., Wilson, J.B. and Manet, E. (2017) Epstein-Barr Virus Nuclear Antigen 1 (EBNA1) interacts with Regulator of Chromosome Condensation (RCC1) dynamically throughout the cell cycle. J. Gen. Virol. 98:251-265 PMID:28284242
4. Hussain, M., Gatherer, D. and Wilson, J.B. (2014) Modelling the structure of full-length Epstein-Barr Virus Nuclear Antigen 1. Virus Genes 49:358-372 PMID: 25011696

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Microenvironment in paediatric and adult acute myeloid leukaemia

Supervisors: Dr Karen Keeshan and Prof Brenda Gibson
Email: karen.keeshan@glasgow.ac.uk

Abstract: Acute myeloid leukaemia (AML) is a genetically and phenotypically heterogeneous disease that is characterized by a block in myeloid differentiation, as well as enhanced proliferation and survival. It affects people of all ages with an incidence of 2-3 per 100 000 per annum in children, increasing to 15 per 100 000 per annum in older adults. The relapse risk for childhood AML remains unacceptably high and relapse is the commonest cause of death. Multiple courses of chemotherapy remain the mainstay of treatment in adult and childhood AML but a ceiling of benefit has been reached and toxicity is significant (Chaudhury et al, 2015). There have been few, if any, new treatments in the past 30 years and there is a pressing need for novel effective therapies in AML.

The treatment of paediatric AML is in essence extrapolated from that of adults with AML. Our previous work (Chaudhury et al, 2015) in addition to recent timely publications (Beerman et al 2015) have questioned the appropriateness of this approach which assumes that a similar aetiology underlies AML in the young and old. There is additional evidence that disease characteristics differ between a paediatric and adult population with AML (Appelbaum 2006; Creutzig et al. 2008). Functional interplay between AML cells and the bone marrow microenvironment is a distinctive characteristic of AML disease. AML cells in the adult bone marrow BM reside in leukaemic niches (Colmone et al 2008) that support leukaemic cell survival and expansion. The importance of the microenvironment in paediatric versus adult AML (fetal liver, cord blood, bone marrow) and its role is disease characteristics has not been well explored. Our lab focuses on the proliferation and self-renewal capabilities of the leukaemic cell and the influence of the leukaemic niche. We hypothesize that the microenvironment influences the initiation, maintenance, and aggressiveness of paediatric and adult AML disease.

Methods & approaches: This project will investigate the role of the microenvironment in AML disease initiation and maintenance. We will focus on genetically distinct subtypes of paediatric and adult using a number of models and approaches including: Bone marrow transduction and transplantation (BMT) murine models: expression of AML oncogenes in viral constructs and using CRISPR/Cas9 gene editing approaches; assessments on disease in vivo; Stromal co-cultures and transcriptional profiling using haematopoietic stem cells; primary AML samples from paediatric and adult patients. The project will also employ flow cytometry, cellular and molecular biology technologies. This PhD studentship offers extensive dual training in both fundamental and translational biology of leukaemia, an environment encompassing clinical and basic researchers, and training opportunities as part of the college graduate program.

References:

1. Chaudhury SS, Morison JK, Gibson BES, Keeshan K. Insights into cell ontogeny, age and acute myeloid leukaemia. Experimental Hematology. 2015 Jun 4.
2. Beerman I, Rossi DJ. Epigenetic Control of Stem Cell Potential during Homeostasis, Aging, and Disease. Cell Stem Cell. 2015 Jun;16(6):613–25.
3. Appelbaum, F.R., 2006. Age and acute myeloid leukemia. Blood, 107(9), pp.3481–3485.
4. Creutzig, U. et al., 2008. Significance of age in acute myeloid leukemia patients younger than 30 years: a common analysis of the pediatric trials AML-BFM 93/98 and the adult trials AMLCG 92/99 and AMLSG HD93/98A. Cancer, 112(3), pp.562–571.
5. Colmone A, Amorim M, Pontier AL, Wang S, Jablonski E, Sipkins DA. Leukemic cells create bone marrow niches that disrupt the behavior of normal hematopoietic progenitor cells. Science. 2008;322(5909):1861-1865.

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SWATH Proteomic analysis to identify putative biomarkers to predict disease related complications involved in Polycythaemia Vera

Supervisors: Dr Helen Wheadon and Professor Mhairi Copland
E-mail address: Helen.Wheadon@glasgow.ac.uk; Mhairi.Copland@glasgow.ac.uk

Abstract: Polycythaemia Vera (PV) is a haemopoietic stem cell disease, with ~95% of patients having the V617F JAK2 mutation, which results in constitutive activation of the JAK/STAT signalling pathway, leading to increased myelopoiesis and high erythrocyte counts. Patients also suffer from symptoms related to chronic inflammation, which is thought to explain, in part, the increased risk of thrombosis. Chronic inflammation is well recognised as a driver of atherosclerotic plaque formation and a risk for development of arterial and venous thromboembolism in patients with autoimmune conditions and other chronic medical illnesses, but the evidence for this in PV is only just starting to become available. It is known that an elevated white cell count in PV is associated with an increase in arterial events, and this appears to relate to inappropriate activation of the haemopoietic cells resulting in a pro-coagulation phenotype, however there is much that remains unclear about what drives these symptoms. One avenue worth exploring is the role of monocytes in PV related thrombotic and cardiovascular complications. Intermediate monocyte frequency has recently been identified as a positive predictor of cardiovascular events playing a significant role in inflammation through high secretion of TNFa and IL-1b.  Intermediate monocytes also express CCR2 and CCR5, which are critical for monocyte homing and trans-endothelial migration into atherosclerotic plaques. Platelet activation and blood coagulation is also initiated at the site of injury by monocyte/macrophage secreted exosomes which contain tissue factor and P-selectin glycoprotein ligand-1 (PSGL-1) on their surface. Work from the Wheadon/Copland laboratory has recently identified significantly higher intermediate monocyte frequency in PV patients compared to normal aged matched control donors and high levels of pro-inflammatory cytokines/chemokines in PV patient serum. Recent interest has focused on the role that exosomes play in immune regulation, especially in neoplasms. Exosomes are homogenous membrane vesicles (40-150nm diameter), derived from the exocytosis of intraluminal vesicles and released into the extracellular space when fused to the plasma membrane. The majority of cells including haemopoietic cells release exosomes, with malignancy specific exosomes, expressing cell of origin cluster of differentiation (CD) antigens identified in leukaemia and PV. Exosomes are important for the extracellular transfer of proteins, lipids, mRNAs, microRNAs and DNAs to neighbouring cells where they can alter the function of the recipient cell. Tumour-derived exosomes (TEX) are released both locally and into the circulation to interact with a variety of cell types, including immune cells. TEX have been shown to alter immunoregulatory mechanisms such as; antigen presentation, immune activation, immune suppression, immune surveillance and cell communication. It is known that erythrocytes retain their microRNA content during maturation and that they secrete exosomes, however their functional role is still not clearly defined. One could postulate that the exosomes act as immunosuppressive signals given that erythrocyte homeostasis requires the constant removal of damaged or aged erythrocytes (2x10⁶ per second)via phagocytes within the liver, spleen or lymph nodes. How erythrocyte exosomes modulate immune responses and whether this is altered in PV patients still remains to be investigated, as does the role of monocyte and erythrocyte derived TEX in PV related complications namely chronic inflammation and thrombotic events.

Hypothesis/research question: Are tumour-derived exosomes involved in the chronic inflammatory and pro-coagulation symptoms experienced by Polycythaemia Vera patients?

Aims/objectives:

1. Mathematical evaluation to determine cells of origin of exosomes in PV patients and normal aged matched donor plasma.
2. SWATH profiling to identify novel biomarkers involved in PV pathophysiology
3. Evaluate Ruxolitinib, hydroxycarbamide (HC), statins and aspirin treatment on TEX secretion & TEX effects on the inflammasome and coagulation.

References:

1. Hasselbalch, H.C., Perspectives on chronic inflammation in essential thrombocythemia, polycythemia vera, and myelofibrosis: is chronic inflammation a trigger and driver of clonal evolution and development of accelerated atherosclerosis and second cancer? Blood, 2012. 119(14): p. 3219-25.
2. Hulsmans, M. and P. Holvoet, MicroRNA-containing microvesicles regulating inflammation in association with atherosclerotic disease. Cardiovasc Res, 2013. 100(1): p. 7-18.
3. Stansfield, B.K. and D.A. Ingram, Clinical significance of monocyte heterogeneity. Clin Transl Med, 2015. 4: p. 5.
4. Caivano, A., et al., High serum levels of extracellular vesicles expressing malignancy-related markers are released in patients with various types of hematological neoplastic disorders. Tumour Biol, 2015.
5. Greening, D.W., et al., Exosomes and their roles in immune regulation and cancer. Semin Cell Dev Biol, 2015. 40: p. 72-81. 

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Investigation of BET and MDM2 inhibitors as a candidate novel combination therapy for AML

Supervisors: Professor Mhairi CoplandE-mail: p.adams@beatson.gla.ac.uk; Mhairi.Copland@glasgow.ac.uk 

Abstract: In 2015, there will be approximately 2400 new cases of AML in the United Kingdom (https://www.hmrn.org/statistics/survival). After diagnosis, five year survival is currently ~15.5%. Therefore, there remains a critical requirement for novel therapies for AML. Bromodomain and extra-terminal domain (BET) inhibitors are emerging as exciting therapeutic agents for hematopoietic malignancies, including AML [1].  Pharmacological inhibition of BET bromodomains targets malignant cells by preventing reading of acetylated lysine residues, thus disrupting chromatin-mediated signal transduction, which reduces transcription at oncogenic loci, such as c-myc, Bcl-2 and cdk4/6 [1].  Although a heterogeneous disease, most AML retains wild type p53 [2]. However, p53 is often rendered functionally deficient by over-expression of MDM2 [3]. Accordingly, we hypothesized that dual inhibition of MDM2 and BET would be synthetic lethal to p53 wild type AML. In extensive studies, we have confirmed this hypothesis both in vitro and in vivo.

Hypothesis: We hypothesize that BET inhibitors potentiate activation of p53 to promote cell cycle arrest and apoptosis. We will: 1) investigate the mechanism by which BET inhibitors potentiate activation of p53 by nutlin; 2) Investigate the mechanism of p53-dependent cell cycle arrest and cell killing.

Objectives:

1. Investigate mechanism by which BET inhibitors potentiate activation of p53 by nutlin. We hypothesize that BET inhibitors potentiate activation of p53 by promoting binding of p53 to specific target gene regulatory sides and/or enhancing the activity of chromatin-bound p53. We further hypothesize that this depends on the ability of BET inhibitors to control p53 post-translational modification and/or interaction with associated proteins. We will dissect these hypotheses to elucidate the mechanism of p53 activation by BET inhibitors.
2. Investigate mechanism of cell cycle arrest and cell killing. We hypothesize that marked G1 cell cycle arrest and apoptosis induced by the drug combination depends, at least in part, on expression of synergistically activated pro-apoptotic and cell cycle arrest p53 target genes, such as PUMA and CDKN1A (p21CIP1) respectively.

Completion of these studies will justify the testing of nutlin and BET inhibitors in a human clinical trial to be led from Glasgow.

References:

1. Gallipoli P, Giotopoulos G, Huntly BJ: Epigenetic regulators as promising therapeutic targets in acute myeloid leukemia. Ther Adv Hematol 2015, 6:103-119.
2. Hou HA, Chou WC, Kuo YY, Liu CY, Lin LI, Tseng MH, Chiang YC, Liu MC, Liu CW, Tang JL, et al: TP53 mutations in de novo acute myeloid leukemia patients: longitudinal follow-ups show the mutation is stable during disease evolution. Blood Cancer J 2015, 5:e331.
3. Kojima K, Konopleva M, Samudio IJ, Shikami M, Cabreira-Hansen M, McQueen T, Ruvolo V, Tsao T, Zeng Z, Vassilev LT, Andreeff M: MDM2 antagonists induce p53-dependent apoptosis in AML: implications for leukemia therapy. Blood 2005, 106:3150-3159.

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Overview

We are part of a national centre of excellence in the fight against cancer carrying out a programme of world-class science directed at understanding the molecular changes that cause cancer. We are working to translate scientific discoveries into new drugs or diagnostic and prognostic tools that benefit cancer patients, taking new therapies through preclinical and clinical trials.

The School of Cancer Sciences is a major component of the Cancer Research UK West of Scotland Cancer Centre. There are currently 51 research groups housed in magnificent new research buildings at the Beatson Institute for Cancer Research, the Paul O’Gorman Leukaemia Research Centre, the CRUK clinical trials unit (CTU) and the Wolfson Wohl Cancer Research Centre. Our facilities house a number of state-of-the-art technologies that underpin our key research themes.

Individual research projects are tailored around the expertise of principal investigators within our Schools. Basic and clinical projects are also available for study.

A variety of approaches are used, including molecular biology, biochemistry, bioinformatics, genetics, cancer modelling and cell biology (including advanced in vitro and in vivo imaging), immunology and polyomics (genomics, transcriptomics, proteomics and metabolomics).

Specific areas of interest include:

• cancer biology and cell signalling
• epigenetics
• cancer stem cell biology
• cancer imaging
• chemoresistance in cancer
• cancer and ageing
• regulation of cancer cell death processes
• genetics, genomics and systems medicine 
• immunotherapy for cancer
• cancer clinical trials

Study options

PhD

• Duration: 3/4 years full-time; 5 years part-time

Individual research projects are tailored around the expertise of principal investigators.

Integrated PhD programmes (5 years)

Our Integrated PhD allows you to combine masters level teaching with your chosen research direction in a 1+3+1 format. 

International students with MSc and PhD scholarships/funding do not have to apply for 2 visas or exit and re-enter the country between programmes. International and UK/EU students may apply.

Year 1

Taught masters level modules are taken alongside students on our masters programmes. Our research-led teaching supports you to fine tune your research ideas and discuss these with potential PhD supervisors. You will gain a valuable introduction to academic topics, research methods, laboratory skills and the critical evaluation of research data. Your grades must meet our requirements in order to gain entry on to a PhD research programme. If not, you will receive the masters degree only.

Years 2, 3 and 4

PhD programme with research/lab work, completing an examinable piece of independent research in year 4.

Year 5

Thesis write up.

MSc (Research)

• Duration: 1 year full-time; 2 years part-time