Ian Cowell – Newcastle University Biosciences Institute
Newcastle University E-mail ian.cowell@ncl.ac.uk
Research Interests – DNA topoisomerases, chromatin dynamics and genome stability.
The genetic material of eukaryotic cells is packaged into chromosomes. These highly organized but dynamic structures support the stable maintenance and faithful replication of the genome, while at the same time permitting transcription, the execution of genetic programs and repair of damaged DNA. How these processes occur and how they are coordinated are fundamental questions in modern biology, with great significance for conditions such as cancer and ageing. DNA topoisomerases carry out DNA topological transactions, including altering DNA supercoiling and decatenation of interlinked DNA duplexes. These enzymes play an essential role in chromosome structure and organisation, DNA replication and transcription and furthermore, are the target for a class of widely use and effective anticancer agents.
Ongoing and Previous Research:
Reducing myelotoxicity of DNA topoisomerase poisons.
While cancer survival rates continue to improve, it remains a challenge to develop therapies that cause fewer adverse effects, such as bone marrow toxicity. In the acute setting, this toxicity includes chemotherapy-induced neutropenia, which is associated with substantial morbidity and mortality and is a common cause of dose reduction and delay. As a late effect, some of the most effective anti-cancer drugs, including DNA topoisomerase II (TOP2) poisons, are associated with therapy-induced acute myeloid leukaemia that can arise years after treatment of the original cancer (see below). We are carrying out research to determine whether myeloperoxidase inhibitors have the potential to protect bone marrow cells from short-term and long-term toxicities of anti-cancer drugs. These toxicities result largely from DNA damage induced in bone marrow cells. Recently, using a cell line model system, we have shown that the enzyme myeloperoxidase (MPO) amplifies the DNA damage inflicted by TOP2 poison anti-cancer drugs (Atwal et al 2017). Since MPO is expressed exclusively in neutrophils and their bone marrow progenitors, we hypothesize that pharmacological inhibition of MPO will protect bone marrow during chemotherapy without affecting the efficacy of the cancer treatment. To this end we are currently assessing the capacity of MPO inhibitors to protect bone marrow progenitor cells from TOP2 poisons using ex-vivo toxicity assays
Processing and Repair of TOP2 poison mediated DNA damage.
TOP2 enzymes alter DNA topology via a transient enzyme-bridged DNA double-strand break. In this conformation subunits of the dimeric TOP2 enzyme remain covalently attached to each end of the DSB via a 5’-phosphotyrosyl linkage. A second DNA segment then passes through the enzyme-bridged DNA gate, and the break is then re-ligated. TOP2 poisons used in anti-cancer therapies inhibit the religation step of this reaction cycle, resulting in the persistence of covalently linked TOP2-DNA complexes. In sufficient quantities, these complexes lead to cell death. The lesions can be converted to DNA DSBs via cellular processing pathways and the DSBs are repaired predominantly via NHEJ. The conversion of TOP2-DNA covalent complexes to “frank” DSBs appears to proceed by several alternative nucleolytic or proteolytic pathways [Lee 2012, Lee 2016], but the major pathway involves proteasomal degradation of the DNA-coupled TOP2 protein, followed by end polishing by TDP2. As part of this research we recently discovered that the deubiquitinating enzyme (DUB) inhibitor PR-619 is a potent TOP2 poison that leads to TOP2A and TOP2B-DNA complexes accumulating in nucleoli (Cowell et al 2019).
Mechanisms of chromosomal translocation in leukaemia.
Therapy-related leukaemia is a rare but unfortunate side effect of otherwise successful treatment for primary cancers using DNA damage-inducing anti-cancer drugs [Cowell & Austin 2012]. Leukaemia cells from these cases display chromosome aberrations, including characteristic recurrent chromosome translocations, and these genetic lesions are thought to be causative or at least early contributory events in the generation of these leukaemias. Precisely how recurrent translocations occur in therapy-related and de novo leukaemia is unknown, but the mechanism presumably involves chromosome breakage and incorrect rejoining. A major part of our work has been supported by Bloodwise (formerly known as Leukaemia and Lymphoma Research ) programme grant funding, to investigate how anti-TOP2 drugs used in primary cancer therapy can contribute to therapy related leukaemia [see Cowell and Austin for review] with the aim of identifying strategies that can reduce the rate of this devastating side. We have shown that the TOP2 isoform TOP2B is responsible for the genotoxic properties of etoposide using a cell line model [Cowell et al 2012], and gathered evidence for a transcription-related mechanism for generating chromosome translocations [Cowell and Austin 2012]. Work is ongoing to further understand DNA damage induced in translocation-prone genes by TOP2 poisons such as etoposide.
Changes in chromatin and nuclear organization associated with DNA damage and genome stability.
The histone variant H2A.X is rapidly phosphorylated after cellular exposure to DNA-damaging agents such as ionizing radiation or the topoisomerase poison etoposide, resulting in chromatin domains enriched for phosphorylated H2A.X (gammaH2AX) around sites of double-strand DNA breaks. I previously showed that heterochromatin is a barrier to H2AX phosphorylation [Cowell et al 2007], and work subsequently published in Molecular Cell and Nature has confirmed this, showing that removal of the heterochromatin protein HP1 is a necessary part of the DNA damage response. Histone deacetylase inhibitors (HDACIs) cause histone hyperacetylation and alter the properties of heterochromatin relieving the block to H2AX phosphorylation. DNA topoisomerase II poisons such as etoposide are mainstream cytotoxic anticancer drugs, and recently it has been shown that their cytotoxic effects can be potentiated by HDACIs. The interaction of HDACIs and other drugs such as TOP2 poisons is an area of interest and we have shown that HDACIs cause a redistribution of TOP2B from heterochromatin to euchromatin in mouse cells and a change in the genomic distribution of topoisomerase-mediated DNA damage [Cowell et al 2011].
Targeting DNA-repair proteins as a means to improve the efficacy of anti-cancer treatments.
Previously I contributed to work validating the PIKK kinases DNA-PKcs and ATM as drug targets for anti-cancer therapies [Cowell et al 2005, Zhao et al 2006, Willmore et al 2008] with Drs Durkacz and Wilmore in the NICR at Newcastle on an LLR-funded project to determine the role of DNA damage-inducible kinases in the cellular responses to nucleoside analogues used in leukaemia therapy.
Histone lysine methylation and the function of chromodomain proteins.
Work I started at Newcastle and then continued at the Babraham and the Roslin Institutes focused on histone modifications and the function of chromodomain proteins [Cowell & Austin 1997; Jones et al 2000; Wang et al 2000; Kourmouli et al 2000; Cowell et al 2002]. Chromodomain proteins, particularly the heterochromatin protein HP1 bind to, and form the “readout” for modifications including trimethylation of histone H3 at lysine 9. I was amongst the first to report the evolutionarily widespread occurrence of chromatin domains enriched for histone H3 trimethylated at lysine 9, and the association of this epigenetic modification with heterochromatin and a transcriptionally silent state [Cowell et al 2002].