Deploying esters of the Krebs cycle metabolites to eliminate cancer stem cells in acute myeloid leukaemia
Primary supervisor: Kamil R Kranc, Queen Mary University of London
Secondary supervisor: Andrew Finch, Queen Mary University of London, Richard Dillon, King’s College London, Christopher Schofield, University of Oxford
AML is an aggressive clonal disorder of haematopoietic stem cells (HSCs) and progenitors, in which they acquire driver mutations, resulting in the generation of treatment-resistant leukaemic stem cells (LSCs). Current conventional therapies fail to eliminate LSCs, which propagate leukaemia and its relapses. As a consequence, many AML patients suffer from severe disease relapses with only 30% of adults and 60% of children surviving beyond 5 years. It is therefore critical to identify new treatments that eradicate LSCs, while sparing normal haematopoiesis. In order to discover new potent anti-leukaemic compounds, we screened a small library of metabolic compound esters against multiple paediatric and adult AML cells. We found that esters of the Krebs cycle metabolites potently induced apoptosis in AML cells, but do not compromise normal HSC/progenitor cell survival. Notably, they were most potent in AML cells (including LSCs) strongly expressing carboxylesterases, which convert these esters to active compounds. These data highlighted esters of the Krebs cycle metabolites as attractive therapeutic agents to eliminate AML cells, while sparing normal haematopoiesis.
We aim to address the hypothesis that esters of the Krebs cycle metabolites represent potent therapeutics for selective LSC eradication.
This preclinical study will address the following specific questions:
1. Can we eradicate LSCs in AML by esters of the Krebs cycle metabolites?
We will employ clinically-relevant AML mouse models and patient-derived samples combined with state-of-the-art stem cell assays in vivo. Given that mouse MLL-AF9+ AML cells (both paediatric and adult) express high levels of carboxylesterases and strongly respond to esters of the Krebs cycle metabolites, we will utilise MLL-AF9 models to determine the impact of these esters on AML progression and LSC elimination. In parallel, we will test the impact of the esters on survival on human primary LSCs, focusing on those expressing high levels of carboxylesterases. Finally, given our unpublished data that BCL-2 inhibitor Venetoclax currently used in AML treatment potentiates the anti-leukaemic effect of the esters, we will investigate the impact of this combination. Together, we will reveal esters of the Krebs cycle metabolites as potent therapeutics in AML in their own right and in combination with other anti-leukaemia agents.
2. What is the therapeutic window for employing these novel therapies?
In order for our novel therapies to be broadly applicable, they must eliminate LSCs without perturbing normal haematopoiesis. Here we will determine the impact of esters of the Krebs cycle metabolites administered singly and in combination with Venetoclax on HSC/progenitor cell compartments at different levels of the haematopoietic hierarchy and multilineage haematopoiesis. This aim will reveal the therapeutic window for our new therapies, thus informing a design of a clinical trial in AML.
3. What are the molecular mechanisms by which esters of the Krebs cycle metabolites eliminate LSCs?
We found that the esters competitively inhibit 2-oxoglutarate-dependent oxygenases. We now intend to inhibit these targets singly and in combination using novel specific inhibitors and CRISPR/Cas9 approaches in primary mouse AML cells to dissect mechanisms of how these esters impacts on AML. In unbiased approaches, we will employ transcriptomics, proteomics and metabolomic approaches on ester-treated LSCs, combined with functional validation, to uncover other unknown mechanisms of anti-leukaemic activity of esters of the Krebs cycle metabolites.
Together, we will uncover therapeutic potential of esters of the Krebs cycle metabolites or their combined application with other therapeutics to selectively eliminate LSCs in paediatric and adult AML, and reveal molecular mechanisms of their action.
- Hewitson, K. S., Liénard, B. M., McDonough, M. A., Clifton, I. J., Butler, D., Soares, A. S., Oldham, N. J., McNeill, L. A., Schofield, C. J. Structural and mechanistic studies on the inhibition of the hypoxia-inducible transcription factor hydroxylases by tricarboxylic acid cycle intermediates. J Biol Chem. 2;282(5):3293-301, DOI: 10.1074/jbc.M608337200 (2006).
- Edwards-Hicks, J., Su, H., Mangolini, M., Yoneten, K. K., Wills, J., Blanco, G., Young, C., Cho, K., Barker, H., Muir, M., Guerrieri, A., Li, X,-F., White, R., Manasterski, P., Mandrou, E.,Wills, K., Chen, J., Abraham, E., Sateri, K., Qian, B.-Z., Bankhead, P., Arends, M., Gammoh, N., Kriegsheim, A., Patti, G., Sims, A., Acosta, J.C., Brunton, V., Kranc, K.R., Christophorou, M., Pearce, E., Ringshausen, I., & Finch, A. MYC sensitises cells to apoptosis by driving energetic demand. Nature Communications 13: 4674, DOI: 10.1038/s41467-022-32368-z (2022).
- 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).