Inhibiting stages in the DNA damage response pathway

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Overview

It has long been known that many cancers have defects in the way in which cells monitor and repair damaged DNA, collectively termed the DNA damage response, or DDR.1 These deficiencies in DDR pathways render the cells more susceptible to DNA damage, and traditional cancer treatments such as radiotherapy and DNA-damaging chemotherapy are based on this premise.1 However, such treatments are often accompanied by significant collateral damage and unwanted side effects.1 Therefore, developing treatments which target specific DDR deficiencies to preferentially kill cancer cells, while minimising the impact on normal cells, has potential for more selective, better tolerated therapies to improve survival in multiple cancers.1

Since the 1990s, scientists have been working to research how errors in DDR pathways can trigger cancer formation, in order to develop treatments targeting the DDR deficiencies associated with cancers. AstraZeneca has had a focus in this area for several years, demonstrated by the PARP inhibitor, olaparib, which was approved for use in ovarian cancer patients in 2014. We are also exploring a number of other targets in DDR.

DDR is one of AstraZeneca’s four key platforms in oncology, in addition to immuno-oncology, antibody drug conjugates and tumour drivers and resistance. We are committed to continuing to investigate potential DDR targets, in order to harness the potential of this science to benefit patients.

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Hijacking the DNA damage response pathway to treat cancer

An introduction to DNA damage response

DNA damage response (DDR) is a term describing the network of cellular pathways that minimise the daily impact of DNA damage.1 Currently, many cancers are known to have defects in DDR pathways,1 which makes them dependent on and therefore, highly sensitive to inhibition of the remaining DDR pathways.1 Targeting DDR deficiencies to preferentially kill cancer cells, while minimising the impact on normal cells, has potential for more selective, better tolerated therapies to improve survival in multiple cancers.1

“People would be really surprised to find out just how much DNA damage occurs every day in a person’s body,” says Dr Mark O’Connor, Chief Scientist in AstraZeneca’s Oncology IMED. “I mean, there are tens of thousands of damage events happening daily to the DNA in our cells, either naturally through cellular processes or because of external or environmental factors, like ultraviolet radiation or from smoking.” Dr O’Connor has devoted his career to understanding how breaks in DNA are dealt with, and more importantly how their presence in cancer cells can be leveraged to kill a tumour. Today, he is one of the senior principal scientists leading AstraZeneca’s development of a potentially new class of anti-cancer drugs— the DNA damage response (DDR) inhibitors.

To appreciate the emergence of this new class of compounds we have to go back to the late 1990’s and a privately-owned UK start-up company called KuDOS which was founded by Professor Stephen Jackson from Cambridge University. Dr O’Connor joined KuDOS not long after studying for a PhD in molecular genetics at Bristol University, UK. At KuDOS, together with Professor Jackson, as well as Dr Graeme Smith (now both at AstraZeneca) and Dr Alan Lau, he researched the cellular network of pathways that minimise the daily impact of DNA damage and studied how errors in these pathways can occur, triggering cancer formation.

Our cells have developed different ways to combat this damage through a network of cellular pathways known as the DNA damage response (DDR).1 This system is made up of at least 450 proteins responsible for monitoring the integrity of our genome,1 initiating repair where possible or instructing our cells to stop growing (or even die when necessary) in order to minimise the impact of the damage.1,2

The time period in which cells pause to carry out internal quality control checks are called cell cycle checkpoints,2 and they form an important part of the DDR for two main reasons. Firstly, they cannot be easily bypassed if the cell’s DNA is damaged. Secondly, the best repair pathway available for the specific type of DNA damage often varies depending on the cell-cycle stage.1 As such, DNA repair and cell-cycle checkpoint regulators are inherently interlinked in the DDR process.1

When a cell needs to repair breaks in its double helix of DNA, possible response pathways include non-homologous end joining (NHEJ), homologous recombination repair (HRR), mismatch repair (MMR), nucleotide excision repair (NER) and base excision repair (BER). DNA polymerases are also critical players in DNA repair processes.1 Each response is specific to the type of DNA damage it repairs.1 For instance, MMR responds to cell replication errors, while NER is responsible primarily for dealing with damage due to ultraviolet light.1 Double-strand breaks (DSBs) in DNA can induce both HRR and NHEJ.1 HRR requires the presence of undamaged, replicated DNA to fix the damage, while NHEJ repair isn’t wholly accurate and can lead to DNA changes.1

We now know that most common cancers have defects in DDR pathways and this is an early event in their evolution.1 These defects are shared among all daughter cells3 and cause the cancer to rely on remaining DDR pathways, resulting in an increase in genetic errors.1

“We’ve evolved multiple different pathways to try and deal with all the damage that the DNA in our cells experiences; these are DDR pathways. In a normal scenario, you can have more than one DDR pathway that deals optimally with a given type of damage plus others that can fill in but aren’t quite as good. What happens early on in cancers is that the cells tend to lose one or more DDR pathway and become highly dependent on a remaining, compensating pathway.” This state, described scientifically as “synthetic lethality,”1 provided the therapeutic opportunity that the KuDOS scientists were looking for.

Their efforts centred on the BER pathway which is responsible for repairing breaks in single strands of DNA (ssDNA).1 The team was investigating PARP (poly ADP-ribose polymerase), an essential protein in the BER pathway.1 Inhibiting PARP forces single strand breaks to form double strand DNA breaks which are repaired via the alternative HRR pathway.1 Cancer cells with BRCA mutations have a faulty HRR pathway,1 and when PARP proteins are also blocked, the cell has no reliable alternative routes available to repair damage to its DNA; double strand breaks, if left unrepaired, are lethal to the cell and the cancer cell dies.1 This is the essence of synthetic lethality.1 By contrast, healthy cells still have fully functioning DDR pathways; if one is blocked, in the case of PARP inhibition, others can step in and still enable DNA repair.1 Leveraging this opportunity to kill cancer cells selectively and leave normal cells unharmed,1 KuDOS discovered the compound called olaparib which inhibits the action of PARP.

The story is taken up by Dr Susan Galbraith, Head of the Oncology Innovative Medicines Group at AstraZeneca. “The reason AstraZeneca has a DNA damage response portfolio today is because back in 2005 we acquired KuDOS in order to gain access to their PARP inhibitor. We were fortunate enough to have several of their leading scientists join us, such as Mark O’Connor, Graeme Smith, Alan Lau and Steve Durrant. They’re all involved in the DDR programmes that we’ve been building at AstraZeneca for many years. Now we’re starting to see that PARP inhibition can work in multiple cancer indications beyond ovarian and we’re becoming more confident that inhibition of DNA damage response pathways is going to be an important mechanism to treat cancer.”

Inhibiting PARP forces cells to repair DNA damage via the alternative DDR pathway, HR.1 However, many cancer cells have mutations – often found in BRCA1, BRCA2, or ATM genes – which result in HR pathway deficiencies.1 As a result, when PARP proteins are also blocked by PARP inhibitors, the cancer cell has no back up pathways left to repair damage to its DNA, and subsequently dies.1 However—and this point is key to the potential value of DDR compounds as anti-cancer agents—normal cells have no DDR deficit and are still able to repair the build-up of DNA damage caused by PARP inhibition, meaning they are unaffected by PARP inhibitors.1 As a result, PARP inhibitors are relatively nontoxic to normal cells but strikingly toxic tumour cells with a defective HR pathway, providing a highly targeted treatment option.1


Meet the team

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A broad portfolio of DDR inhibitors

With a growing team of scientists focused on DDR, AstraZeneca began to explore more targets in DDR. One compound is aimed at the target ataxia telangiectasia and rad3 related (ATR) kinase which activates DNA damage checkpoints in the cell cycle when damage is present and slows the progression of DNA replication, preventing premature entry in to cell division.4 Inhibition of ATR disrupts repair leading to tumour cell death.4

Following the principle of synthetic lethality, ATR inhibitors show potential in cancers that have lost function in the repair pathway involving ataxia telangiectasia mutated (ATM) kinase (non-small cell lung cancer, breast, prostate and gastric cancers).4 ATM inhibitors cause double strand breaks to accumulate, leading to cell death.4

“We have both ATM and ATR inhibitors in Phase 1 trials and other programmes that are coming through preclinical research,” says Dr Galbraith. “While there are other companies developing PARP inhibitors there’s no company that has anything close to the depth and breadth of the portfolio that we have.”

In addition to DDR defects, cancer cells can also lose the ability to pause and check DNA at a checkpoint in the cell cycle.5 For example, the protein kinase WEE1 stops the cell cycle after DNA damage occurs, allowing tumour cells time to repair the damage.5 WEE1 is overexpressed in many breast,5 lung,6 and colon cancers,5 suggesting that the tumour relies heavily on this mechanism to survive. AstraZeneca is developing an inhibitor of WEE1 which forces the cell cycle to continue without checking for damage, instead carrying any errors through to cell division, leading to tumour cell death.5 “Just like DNA damage repair deficiencies, we know that when there’s a check point in the cell cycle that’s already missing, if you then knock out another then you drive the cell to accumulate more and more unsustainable DNA damage,” says Dr Galbraith.

Another target is Aurora B kinase which assists in DNA chromosome alignment during cell division.7 Its inhibition causes either unequal splitting of the chromosomes between the daughter cells or failure of the cell to divide, leading to cell death. Aurora B kinase is known to be over-expressed in liver, colon, breast, renal, lung and thyroid cancers.7

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Enhanced specificity

The selectivity of DDR inhibitors for cancer cells over normal cells makes them potentially better tolerated therapies for patients,1 allowing for longer term dosing schedules in contrast with the limited cycles of chemotherapy that patients can sustain.

Dr O’Connor explains the rationale for this improved tolerability: “One of the basic concepts when we first started to think about targeting the DNA damage response was to find a much more specific way of killing cancer cells, than say, chemotherapy. Chemotherapies essentially are working on the same principles that is you’re trying to induce so much DNA damage that the cancer cell can’t survive it; the problem is that it’s not a very precise approach and you also damage normal tissue, which means that patients experience significant side effects. We hope that by identifying specific DDR defects present in tumours but not in normal cells that we can have a much greater effect on the cancer than normal tissue, and consequently we can reduce side effects of treatment.”


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Next generation sequencing

Identifying which patients have which DDR defect is key explains Dr Darren Hodgson, Senior Principal Scientist in the oncology iMed. “It’s extremely exciting to be working in the DDR area at present because we’re understanding the fundamental science of what has gone wrong in the development of those tumours, and then we’re trying to turn that against them by using the right pharmaceutical intervention. Of course, in order to know what has gone wrong within those tumours we need to be able to sequence their DNA accurately and reliably so that we can match the right treatment to the right patient.”

Advances in understanding the biology of DDR pathways have coincided perfectly with progress in gene sequencing technology, which has delivered next generation sequencing, or NGS. This technology allows whole genomes to be sequenced in days rather than years which has made sequencing sufficiently cost effective to become a real tool for the lab. The plan is for tumours to be rapidly sequenced to reveal their molecular profile, showing which of the DDR pathway or cell cycle defects they contain, allowing them to be matched to inhibitors of their alternate pathway.

“It’s incredibly exciting because for years and years oncology trials have not been able to say ‘this is how my drug works and this is what’s gone wrong with the tumour,’ and now we are able to put these two things together and not only understand at the basic level but translate that into a test that will identify the right patient for the right drug,” says Dr Hodgson. “DNA damage response research within AstraZeneca is a great example of what science can do; not only are we leading the basic science and the understanding of how these agents work, but we’re also leading in how we match the biology of the agents with the biology of the tumours and it’s next generation sequencing that is enabling us to do that.”


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Power in combination

With a breadth of inhibitors available covering major DDR pathways, AstraZeneca is trialling them in combination with each other and with other anti-cancer agents with the intent of leaving no escape route for cancer cells.1 For example, combining DDR inhibitors with each other could simultaneously increase tumour DNA damage in the S-phase of the cell cycle and prevent its repair at checkpoints.1 There is also scope for combining DDR inhibitors with other, new targeted therapies and with treatments such as chemo and radiotherapy.1

Dr Nina Mojas, Global Medicines Leader for olaparib is encouraged by the science behind combinability. “We believe there are significant synergies between DDR agents and the new immuno-oncology therapies. DDR agents can serve as primers for immunotherapy meaning that treatment with DDR agents will cause tumours to act a little bit like centres of inflammation. When inflammation happens in the body our immune system tends to attack it and get rid of it. So tumours somehow transform into that centre of inflammation that then our own natural system will try to eradicate. We have several ongoing studies exploring this combination.”

AstraZeneca DDR combinations under investigation include:

  • DDR-DDR agent combinations providing broader and more effective response than DDR-based monotherapy, as shown by preclinical data on the combination of olaparib with WEE1 inhibitor (AZD1775). Olaparib is also being tested with the ATM inhibitor (AZD0156) and ATR inhibitor (AZD6738)
  • The MEDIOLA study is testing the combination of olaparib with durvalumab alongside traditional chemotherapy and radiotherapy, which can serve to sensitise cells to DDR targets.8 AZD1775 and AZD6738 are also each being tested in combination with durvalumab.
  • Combinations with targeted agents affecting other hallmarks of cancer, such as the aberrant blood vessels associated with tumour growth.9 Olaparib is in Phase II trials with the VEGF inhibitor, cediranib.

AstraZeneca’s DDR portfolio will be on display at ASCO 2016 where 17 abstracts on DDR have been accepted for presentation. Data on overall survival of women with ovarian cancer treated with olaparib (Study 19) has been selected as a “Best of ASCO” abstract (Abstract # 5501). The combination potential of DDR with immuno-oncology therapies or with a VEGF inhibitor will also be described (Abstract # 3015).

These days, Dr O’Connor admits just a little surprise at how well the idea of exploiting defects in DNA damage response is panning out as a potentially new class of anti-cancer drugs. “I think what’s been really exciting for the team and me, and perhaps a little unexpected if I think back to the start, is that we’ve now truly arrived at a position where we have a world-leading position in terms of DDR agents. It’s humbling, but very exciting, to have been one of those at the forefront of developing completely new ways in which we can target cancers.”


References

1 O’Connor M, 'Targeting The DNA Damage Response In Cancer' (2015) 60 Molecular Cell

2 'Eukaryotes, Cell Cycle | Learn Science At Scitable' (Nature.com, 2016) <http://www.nature.com/scitable/topicpage/eukaryotes-and-cell-cycle-14046014> accessed 23 May 2016

3 'Cell Division, Cancer | Learn Science At Scitable' (Nature.com, 2016) <http://www.nature.com/scitable/topicpage/cell-division-and-cancer-14046590> accessed 17 May 2016

4 Weber A and Ryan A, 'ATM And ATR As Therapeutic Targets In Cancer' (2015) 149 Pharmacology & Therapeutics

5 Vriend L and others, 'WEE1 Inhibition And Genomic Instability In Cancer' (2013) 1836 Biochimica et Biophysica Acta (BBA) - Reviews on Cancer

6 De Witt Hamer P and others, 'WEE1 Kinase Targeting Combined With DNA-Damaging Cancer Therapy Catalyzes Mitotic Catastrophe' (2011) 17 Clinical Cancer Research

7 Gavriilidis P, Giakoustidis A and Giakoustidis D, 'Aurora Kinases And Potential Medical Applications Of Aurora Kinase Inhibitors: A Review' (2015) 7 J Clin Med Res

8 Phase 1 And 2 Study Of MEDI4736 In Combination With Olaparib Or Cediranib For Advanced Solid Tumors And Recurrent Ovarian Cancer - Full Text View - Clinicaltrials.Gov' (Clinicaltrials.gov, 2016) <https://clinicaltrials.gov/ct2/show/NCT02484404> accessed 24 May 2016

9 Hanahan D and Weinberg R, 'Hallmarks Of Cancer: The Next Generation' (2011) 144 Cell