Therapeutic targeting of HIF prolyl hydroxylases in acute myeloid leukaemia

Lead Research Organisation: Queen Mary, University of London
Department Name: Barts Cancer Institute

Abstract

Studies on the blood system have in the past pioneered stem cell transplantation and delivered new therapies for cancer. Normal blood stem cells reside in the bone marrow and are responsible for life-long production of red and white blood cells. In blood cancers such as acute myeloid leukaemia (AML), normal blood stem cells are damaged and turn into cancer stem cells (CSCs) that are unable to make normal blood cells. Instead, they generate and fuel the leukaemia bulk which has devastating consequences to many tissues and organs. A major problem in cancer treatments is that the currently available therapies shrink the leukaemic bulk but they fail to eliminate CSCs, which cause the disease to return in a more aggressive form. Therefore, it is of immense importance to design new therapies that efficiently target CSCs and permanently eradicate them.

In order to develop curative therapies that eliminate CSCs, it is essential to understand how these rogue cells are made and how they manage to survive currently available leukaemia treatments. The central aim of our laboratory is to understand the biology of CSCs and harness this knowledge to pharmacologically target the key biological processes in CSCs in order to therapeutically eliminate them.

Recent fundamental discoveries have revealed that the bone marrow, where CSCs are generated, has remarkably low levels of oxygen (i.e. is hypoxic). Cells typically respond to hypoxia by producing molecules called 'hypoxia-inducible factors' (Hifs) that help them to adapt to low oxygen levels. We have teamed up with key international experts in hypoxia biology, Prof. Sir Peter Ratcliffe and Prof. Chris Schofield at Oxford, to understand the role of Hifs in leukaemia. We found that normal blood stem cells do not require Hifs to generate all blood cells (Guitart et al, Blood, 2013 & Vukovic et al, Blood, 2016) but, importantly, under pathological conditions, Hifs strongly protect blood stem cells from becoming CSCs (Vukovic et al, J. Exp. Med, 2015). Based on this knowledge and promising pilot studies, we now intend to conduct a pre-clinical investigation testing the hypothesis that Hifs can be used as powerful weapons to combat CSCs. Using different state-of-the-art experimental strategies in mice harbouring murine and human leukaemia we will investigate whether high levels of Hifs have the ability to kill CSCs and rescue the mice from the disease. Importantly, one of these strategies will involve testing new drugs known to pharmacologically induce high levels of Hifs and thus, if successful in mice, can be rapidly translated to the clinic. This work will provide a framework for clinical trials aiming for curative therapies in AML. We will work closely with our clinical collaborators (including Prof. Tim Somervaillle in Manchester and Dr Christoph Lutz in Heidelberg, Germany) to translate our findings for patient benefit as quickly as possible.

We are in a unique position to perform this research. We have all the necessary reagents and track record and expertise in haematology and stem cell biology. Our close collaborators in Oxford are internationally recognised experts in hypoxia and cancer, and have significant interest in therapeutically manipulating Hifs in human diseases. This project will greatly benefit from such cross-discipline collaboration. Finally, MRC Centre for Regenerative Medicine has a long-term track record in pioneering discoveries in the stem cell field and this institute is an ideal place to successfully perform this study.

Technical Summary

Acute myeloid leukaemia (AML) is a clonal disorder of haematopoietic stem cells (HSCs) and progenitor cells, which acquire mutations and form treatment-resistant leukaemic stem cells (LSCs) that propagate the disease. Since current therapies often fail to fully eradicate LSCs, it is essential to identify new therapeutic targets for LSC elimination. Considering that normal and malignant haematopoiesis occur under hypoxic conditions of the bone marrow, we investigated the role of hypoxia-inducible factor-1alpha (Hif-1alpha) and Hif-2alpha, the main mediators of cellular responses to hypoxia, in these processes. We found that while HSCs do not require Hif-1alpha and Hif-2alpha to self-renew and sustain haematopoiesis (Guitart et al., Blood, 2013 and Vukovic et al., Blood, 2016), Hif-1alpha and Hif-2alpha synergise to suppress development and maintenance of LSCs in several mouse models of AML (Vukovic et al., J. Exp. Med., 2015). We now intend to investigate the therapeutic significance of a constitutive activation of Hif-1/Hif-2 in AML. Firstly, we will delete or suppress Phd2 (a Hif prolyl hydroxylase whose activity normally results in Hif-1/2alpha degradation) in different mouse models of AML and test the impact of this manipulation on the development and maintenance of LSCs. Secondly, we will establish the therapeutic potential of pharmacological Phd2 inhibition in mouse AML models and immunocompromised mice xenografted with different subtypes of human AML. Finally, we will identify and functionally validate the mechanisms through which Phd2 inactivation compromises LSC functions. This project will validate Phd2 as a therapeutic target in AML and reveal further Hif-dependent targets for LSC eradication. We will next be in a position to harness this knowledge to design improved treatment regimens in AML and test them in clinical trials.

Planned Impact

Strategic importance:

Understanding leukaemic/cancer stem cell biology is an immense strategic imperative as it has broad ramifications for several fields including oncology, haematology, and drug discovery. Eradication of leukaemic/cancer stem cells is essential to achieve curative therapies in blood malignancies and other cancers. Our project is therefore of immense strategic importance and addresses key areas of unmet clinical needs. It has a strong potential to rapidly translate to patient, family and societal benefits and fits well with MRC's mission and strategic goals.

Who benefits:

AML patients: We aim to achieve efficient eradication of AML leukaemic stem cells thus providing curative treatments. Patient benefit will depend on the efficient translation of our work to clinical need. We have strong links with pharma and clinical haematologists around the UK, and we will liaise with them to achieve this as rapidly as possible.

Patients with other blood malignancies: Once we provide a proof of concept in AML treatments, we will test whether similar therapies can be applied to other blood malignancies (e.g. chronic myeloid leukaemia, myeloma, lymphomas).

Cancer patients: In the longer term, we will collaborate with our colleagues focusing on different cancers to test whether our treatment strategies can be employed in other cancers.

Industry: Our results will inform drug development within the pharmaceutical industry aiming to achieve leukaemic/cancer stem cell eradiation. We will boost the development of spinouts and provide opportunities for commercialisation. This will generate new business in the UK contributing to its economic competitiveness.

Patients' families: Blood malignancies are devastating conditions that also immensely affect the wellbeing of patients' relatives. Curative treatments will undoubtedly have a major positive impact on these beneficiaries and their function in society.

The NHS budget: Curative leukaemia treatments that target leukaemic stem cells (and potentially cancer stem cells in other tumours) will reduce the necessity to treat disease relapses, minimising the cost of treatments.

Clinicians: Therapeutic targeting of leukaemic stem cells and cancer stem cells is currently ineffective. Improvements in this area will be welcomed by haematologists and oncologists. They will benefit from the development of novel and more effective therapies aimed at targeting LSCs in AML, making the treatment of AML easier, more efficient and more cost-effective.

Research community & collaborators: Given the multidisciplinary nature of this project a broad academic community will benefit. This includes leukaemia biologists, clinical haematologists, cancer stem cell researchers, drug discovery researchers, and hypoxia biologists.

This project will not only uncover the impact of HIF prolyl hydroxylases on leukaemic/cancer stem cell functions but it will also reveal novel fundamental aspects and leukaemia biology. Our work will set the stage for further investigations into Phds and importantly also other protein hydroxylases in blood malignancies. The vulnerability of leukaemic stem cells upon PHD inhibition may prove a common feature of other cancer stem cells in many solid tumours. Through this project, we may provide a broader cancer community with important clues to eradicate cancer stem cells.

Lay audience: Through public lectures, press releases, comments for national media and interviews we often engage with the public. We explain science to school pupils, fundraisers, teachers, and cancer patients. We liaise with the University Press Office and give interviews to national and international press.

Biomedical charities: Our work may inform strategic priorities of cancer charities and we will regularly update them about our findings.

Young scientists: This project will provide an excellent training and create opportunities for career progression.
 
Description Collaboration with Prof Christopher Schofield 
Organisation University of Oxford
Country United Kingdom 
Sector Academic/University 
PI Contribution Investigating the role of 2-oxoglutarate-dependent oxygenase in cancer stem cell development and maintenance. Characterisation of mutant mice and small molecules in partnership with the Oxford collaborators.
Collaborator Contribution Provision of small molecule inhibitors and exchange of unpublished knowledge. Advise on drug discovery.
Impact not yet
Start Year 2013
 
Description Collaboration with Professor Bertie Gottgens 
Organisation University of Cambridge
Department School of Clinical Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution Generation and provision of novel state-of-the-art mouse models to uncover haematopoietic trajectories upon infection and ageing.
Collaborator Contribution Internationally-recognised expertise in single cell genomics
Impact Not yet.
Start Year 2019
 
Description Collaboration with Professor Donal O'Carroll 
Organisation University of Edinburgh
Country United Kingdom 
Sector Academic/University 
PI Contribution Prof. Donal O'Carroll is a co-investigator on my CRUK Programme grant
Collaborator Contribution Prof. Donal O'Carroll is a co-investigator on my CRUK Programme grant
Impact This collaboration is at the intersection of RNA metabolism, stem cell and cancer biology and has immense therapeutic potential. Outputs: Paris, J., Morgan, M., Campos, J., Spencer, G.J., Shmakova, A., Ivanova, I., Mapperley, C., Lawson, H., Wotherspoon, D.A., Sepulveda, C., Vukovic, M., Allen, L., Sarapuu, A., Tavosanis, A., Guitart, A.V., Villacreces, A., Much, C., Choe, J., Azar, A., van de Lagemaat, L.N, Vernimmen, D., Nehme, A., Mazurier, F., Somervaille, T.C.P., Gregory, R.I., O'Carroll, D. & Kranc, K.R. Targeting the RNA m6A reader YTHDF2 selectively compromises cancer stem cells in acute myeloid leukemia. Cell Stem Cell 25(1), 127-148 (2019). Guitart, A.V., Panagopoulou, T.I., Villacreces, A., Vukovic, M., Sepulveda, C., Allen, L., Carter, R.N., van de Lagemaat, L. N., Morgan, M., Giles, P., Sas, Z., Vila Gonzalez, M., Lawson, H., Paris, J., Edwards-Hicks, J., Schaak, K., Subramani, C., Gezer, D., Armesilla-Diaz, A., Wills, J.C., Easterbrook, A., Coman, D., So, C.W.E., O'Carroll, D., Vernimmen, D., Rodrigues, N.P., Pollard, P.J., Morton, N.M., Finch. A. & Kranc, K.R. Fumarate hydratase (Fh1) is a critical metabolic regulator of haematopoietic stem cell functions. J. Exp. Med. 214(3), 719-735 (2017). Vukovic, M., Guitart, A., Sepulveda, C., Villacreces, A., O'Duibhir, E., Panagopoulou, T., Ivens, A., Menendez-Gonzalez, J., Iglesias, J.M., Allen, L., Glykofrydis, F., Subramani, C., Armesilla-Diaz, A., Post, A., Schaak, K., Gezer, D., So, C.W.E., Holyoake, T., Wood, A., O'Carroll, D., Ratcliffe, P. & Kranc, K.R. Hif-1a and Hif-2a synergise to suppress AML development but are dispensable for disease maintenance. J. Exp. Med. 212, 2223-2234 (2015). Patent application: Haematopoietic stem cell treatment International Patent Application No PCT/GB2019/050918 co-inventors: Kamil R Kranc and Donal O'Carroll
Start Year 2015
 
Description Collaboration with Sir Peter Ratcliffe 
Organisation University of Oxford
Country United Kingdom 
Sector Academic/University 
PI Contribution We characterised mutant mice and small molecules in partnership with the Oxford collaborators.
Collaborator Contribution Expertise in hypoxia signalling. Sharing unpublished mutant mice and other relevant and unique reagents.
Impact Yamamoto, A., Hester, J., Macklin, P.S., Kawai, K., Uchiyama, M., Biggs, D., Bishop, T., Bull, K.R., Cheng, X., Cawthorne, E., Crockford, T.L., Davies, B., Dow, L.E., Goldin, R., Kranc, K.R., Kudo, H., Lawson, H., McAuliffe, J., Milward, K., Scudamore, S.L., Soilleux, E.J., Issa, F., Ratcliffe, P.J., Pugh, C.W. Systemic silencing of HIF hydroxylase PHD2 causes reversible immunoregulatory dysfunction. J. Clin. Invest. 130(9), 3640-3656 (2019). Vukovic, M., Guitart, A., Sepulveda, C., Villacreces, A., O'Duibhir, E., Panagopoulou, T., Ivens, A., Menendez-Gonzalez, J., Iglesias, J.M., Allen, L., Glykofrydis, F., Subramani, C., Armesilla-Diaz, A., Post, A., Schaak, K., Gezer, D., So, C.W.E., Holyoake, T., Wood, A., O'Carroll, D., Ratcliffe, P. & Kranc, K.R. Hif-1a and Hif-2a synergise to suppress AML development but are dispensable for disease maintenance. J. Exp. Med. 212, 2223-2234 (2015) (Commentary in an Insights article by Prof. Mick Bhattia in J. Exp. Med.) Vukovic, M., Sepulveda, C., Subramani, C., Guitart, A., Mohr, J., Allen, L., Panagopoulou, T., Paris, J., Lawson, H., Villacreces, A., Armesilla-Diaz, A., Gezer, D., Holyoake, T.L., Ratcliffe, P. & Kranc, K.R. Adult haematopoietic stem cells lacking Hif-1a self-renew normally. Blood 127(23), 2841-6 (2016) (Commentary in Insight Blood Commentaries by Dr Stephen M. Sykes in Blood) Guitart, A.V., Subramani, C., Armesilla-Diaz, A., Smith G, Sepulveda, C., Gezer, D., Vukovic, M., Dunn, K., Pollard, P., Holyoake, T.L., Enver, T., Ratcliffe, P.J. & Kranc, K.R. Hif-2alpha is not essential for cell-autonomous hematopoietic stem cell maintenance. Blood 122(10), 1741-5 (2013).
Start Year 2013
 
Description Bloodwise visits 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Patients, carers and/or patient groups
Results and Impact Lectures on stem cell transplantation and cancer stem cells and AML to leukaemia patients, fundraisers and Bloodwise supporters
Year(s) Of Engagement Activity 2014,2015,2018,2019
 
Description Cancer Research UK Parliamentary Reception 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Policymakers/politicians
Results and Impact Public outreach event for senior CRUK researchers, MSPs, ministers, fundraisers, and cancer patients at the Scottish Parliament. This involved focused group conversations with these groups about my cancer research.
Year(s) Of Engagement Activity 2016,2018
 
Description Expert opinion 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Media (as a channel to the public)
Results and Impact Expert opinion on Woll et al, Cancer Cell, 2014 in The Independent (UK national newspaper)
http://www.independent.co.uk/life-style/health-and-families/health-news/first-evidence-of-cancer-stem-cells-brings-hope-for-possible-future-treatment-9380201.html
Year(s) Of Engagement Activity 2014
URL https://www.independent.co.uk/life-style/health-and-families/health-news/first-evidence-of-cancer-st...
 
Description Motion in Scottish Parliament 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Policymakers/politicians
Results and Impact Motion in Scottish Parliament congratulating the laboratory on 'Major Milestone in Tackling Leukaemia'
Year(s) Of Engagement Activity 2016
URL https://www.parliament.scot/parliamentarybusiness/BusinessBulletin/95602.aspx
 
Description Our research in the news 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Media (as a channel to the public)
Results and Impact Articles in UK national newspapers (the Scotsman, Herald, Metro, Scottish Daily Express and Daily Mail) commenting on Vukovic et al, J. Exp. Med., 2015
Year(s) Of Engagement Activity 2015
URL https://www.scotsman.com/regions/scientists-hail-find-major-milestone-tackling-blood-cancer-1487662
 
Description Public outreach in the media 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Media (as a channel to the public)
Results and Impact Articles in UK national newspapers (Herald, Evening Standard, Daily Express and Daily Mail) commenting on Paris et al, Cell Stem Cell, 2019 (Targeting the RNA m6A Reader YTHDF2 Selectively Compromises Cancer Stem Cells in Acute Myeloid Leukemia.)

https://www.standard.co.uk/news/health/scientists-discover-achilles-heel-in-deadly-blood-cancer-that-could-boost-treatment-a4126516.html
https://www.heraldscotland.com/news/17598846.research-sparks-fresh-hope-of-leukaemia-cure/
Year(s) Of Engagement Activity 2019
URL https://www.standard.co.uk/news/health/scientists-discover-achilles-heel-in-deadly-blood-cancer-that...