New method for cost-effective genome-wide DNA methylation analysis

Cytosine, one of the four DNA bases, can be chemically modified by the addition of a molecule known as a methyl group to form 5-methylcytosine. This “epigenetic” modification has long been known to regulate gene expression and plays a critical role in processes like embryonic development. Its levels and distribution are also distinct in different tissues and are significantly altered in cancers. Analysing methylation patterns of DNA shed into blood and other bodily fluids by tumours can thus reveal both the presence and the location of a cancerous growth.

In 2019, Dr Chunxiao Song (Ludwig Institute for Cancer Research, Oxford Branch, Nuffield Department of Medicine) and his team developed TET-assisted pyridine borane sequencing (TAPS) for mapping DNA methylation. The technology was spun out in 2020 to establish the biotechnology company Base Genomics, which was acquired for $410 million by Exact Sciences in October 2020. Compared to the previous gold standard for sequencing DNA methylation, TAPS is far more cost-effective and sensitive, and generates cleaner data to allow for additional genetic analysis.

Yet despite its advantages, TAPS still relies on whole-genome sequencing, which remains an expensive approach for detecting DNA methylation since just ~4% of all cytosines in the genome are methylated. Chunxiao and his team have now developed a new method that cuts costs further by sequencing only those regions of the genome that contain methylated cytosines.

Building on the TAPS method, postdocs Dr Jingfei Cheng and Dr Paulina Siejka-Zielińska made use of molecular scissors called endonucleases that recognise and cut specific DNA sites. During TAPS, methylated cytosines are chemically converted to an altered base called dihydrouracil (DHU). The researchers found an endonuclease called USER enzyme that specifically cuts at DHU. Because of the enzyme specificity, they knew that all the DNA fragments produced had methylation sites at the beginnings and ends. By then size-selecting the DNA to exclude the larger, uncut DNA, only the smaller, cut DNA fragments with methylation sites are sequenced, making this approach more cost-effective for studying DNA methylation at base-pair resolution.

The team has named the new technique endonuclease enrichment TAPS (eeTAPS), and details on the method can be found in their publication in Nucleic Acids Research.

Using Herpesvirus to fight cancer

The Seymour lab at the Department of Oncology, University of Oxford, has published a new paper investigating the use of oncolytic herpes virus-1 as a vector to augment immunotherapy in cancer

Oxford success at the early detection sandpit on pancreatic cancer

Pancreatic cancer is a devastating disease with low survival rates that have hardly improved in the last 40 years. These cancers are very challenging to treat, in part due to their frequently late diagnosis when the cancer is already advanced.

To address this need for earlier detection, Cancer Research UK, Pancreatic Cancer UK and the Engineering and Physical Sciences Research Council convened a 3-day virtual workshop in November 2020. Multidisciplinary teams worked together to generate innovative research ideas for detecting pancreatic cancer earlier. At the end of the workshop, the teams pitched their ideas to receive seed funding for feasibility testing from a Cancer Research UK Early Detection Innovation Award. Two successful teams involved Oxford researchers.

Team ReTHOMS: Real-time high-sensitivity optrode metabolic sensor for pancreatic cyst fluids

Team ‘ReTHOMS’ includes Oxford’s Professor Eric O’Neill (Department of Oncology) who is working with Dr Paolo Bertoncello (Swansea University), Dr David Chang (University of Glasgow) and Dr George Gordon (University of Nottingham). The team aims to develop a new sensor device to detect malignant transformation in people with pancreatic cysts, a condition that puts them at higher risk of pancreatic cancer.

Pancreatic cysts are fluid-filled sacs on or in the pancreas that are mostly benign. However, 2-3% are precancerous and can develop into pancreatic cancer. Cysts are often identified incidentally and are then monitored for malignant transformation using either clinical imaging or analysis of the cyst fluid for cancer biomarkers such as mucins. Despite this surveillance regime, early cancers are still being missed since these methods have limited sensitivity and specificity.

To improve the early detection of malignant cyst transformation, the team aims to develop real-time and highly sensitive detection of an expanded range of cancer biomarkers. In addition to mucin, raised cellular levels of the chemical hydrogen peroxide are associated with cancer. So-called optrode technology will be used to detect hydrogen peroxide and mucin in cyst fluid. Optrodes are optical sensor devices that detect light emitted as a result of an electrochemical reaction with the biomarkers being analysed.

During this short project, the team will build the optrode device for measuring hydrogen peroxide and mucin, and undertake technical and biological validation. The longer-term aim of this research is to detect pancreatic cancer earlier by screening pancreatic cyst fluid at the point-of-care and determining further action based on the risk of cancer.

Team EDPAN: Earlier detection of pancreatic cancer through personalised assessment of risk combined with non-invasive infrared spectroscopy

Oxford’s Dr Pui San Tan (Nuffield Department of Primary Care Health Sciences) will work as part of team EDPAN with Dr Pilar Acedo Nunez (University College London), Dr Aida Santaolalla (King’s College London), Dr Paul Brennan (University of Edinburgh), Dr Lucy Oldfield (University of Liverpool), Dr Andrew Kunzmann (Queen’s University Belfast) and Dr Mohammad Golbabaee (University of Bath). This team aims to identify individuals at higher risk of pancreatic cancer for further diagnostic screening.

One of the reasons that pancreatic cancer is often diagnosed late is that symptoms are non-specific and cannot discriminate those that require investigation for pancreatic cancer. Team EDPAN will develop an approach for personalised risk stratification to identify individuals at higher risk that would benefit from more in-depth screening for pancreatic cancer.

During the project, the team will make use of existing cohorts for pancreatic cancer (ADEPTS (UCL) and PanDIA (Liverpool)) and larger cohorts for epidemiology research (UK Biobank, AMORIS). They will combine clinical and demographic information with analysis of serum and urine samples using a technique called infrared spectroscopy. They will also evaluate changes in immune components. These approaches aim to identify individuals at high-risk of pancreatic cancer and investigate whether addition of infrared spectroscopy data and immune analysis improves the accuracy of the risk prediction model.

Further funding secured to hunt out cancer using innovative radiotherapy techniques

Professor Bart Cornelissen and Dr Tiffany Chan, from the Department of Oncology, have received an additional £408,338 award from the charity Prostate Cancer Research (PCR) to continue their innovative work to help a new type of radiotherapy, designed to hunt out cancer even after it has spread, to benefit even more men with prostate cancer. Prostate cancer is now the most commonly diagnosed cancer in the UK and their work could lead to more personalised treatment for those with prostate cancer.

Bart and Tiffany are working with a type of radionuclide therapy called 177Lu-PSMA. PSMA seeks out a protein found almost exclusively on prostate cancer cells, and by linking it to radioactive Lutetium (Lu), it can guide the radiotherapy directly inside tumour cells. ‘An advantage of 177Lu-PSMA is that we don’t need to know where all the cancer cells are before treatment, unlike in external beam radiotherapy’ explains Tiffany. ‘In theory, even if we just have a single cell that has split away from the main tumour, if it expresses PSMA, we should still be able to target it.’

Some Lu-PSMA treatments are already used in the UK, but on a private basis only and at the moment they are primarily used for pain relief. The Oxford researchers aim to combine Lu-PSMA with other therapies, and their initial results, from testing nearly 2,000 drugs, have led to the discovery of a group of drugs that may be able to help 177Lu-PSMA hunt out prostate cancer better and make it more effective for more patients. This discovery, which led to further funding from PCR for them to continue this exciting work, comes at an important time for radionuclide therapies. ‘There’s a very large Phase 3 clinical trial with Lu-PSMA and that seems to suggest that you actually get benefits in overall survival from this treatment’ explains Bart. ‘Rather than just being pain relief, we can now start to think of these as cures as well.’

“Speaking as a patient whose prostate cancer has been previously treated with radiotherapy and is likely to be so again in the future, I find this work to be a very welcome addition to the treatments available for the disease,’ said prostate cancer patient David Matheson. ‘It is heartening to see such progress with this treatment, and I look forward to it becoming more widespread in the future.’

During lockdown, the closure of their lab meant they had to find alternative ways to reach their goal. They found an innovative solution – reversing their original plans and developing new and efficient ways to analyse results in lockdown first, and then conducting the experiments when they could return to the lab. Tiffany developed a network analysis tool to enable them to predict which combinations might work. ‘I’m a Londoner, so I like to think of it like a tube map, where have all of our different tube stops, connected by different tube lines, with some lines being more efficient than others. The idea behind mapping the system in this way is that we can hopefully find the best line, or in this case, the best biological pathway, that is most likely to lead to synergism with Lu-PSMA,’ she said. PCR initially awarded £100,000 to Bart and Tiffany in 2019 to test up to 1,000 drugs in combination with 177Lu-PSMA. Despite the challenges brought on by the Covid-19 pandemic, the team surpassed their target and managed to test an incredible number of drugs.

Bart and Tiffany are hopeful that Lu-PSMA could become more widespread in the clinic but believe more research needs to be done. ‘Lu-PSMA is the new kid on the block, it’s a very new technology. I think there’s still a lack of understanding about how Lu-PSMA itself works, and there’s a lot of biology we can learn to improve its efficacy’ says Tiffany. ‘So that’s what we’re trying to achieve, particularly with our network analysis approach to map out the biology behind it.’

‘Bart and Tiffany’s project is already showing promising results on the route to improving radiotherapy for men with prostate cancer. We look forward to continuing to support their project on a larger and long-term basis and hope it will mean that more people can benefit from enhanced radiotherapy, without the side effects’

– Dr Naomi Elster, Head of Research and Communications, PCR

‘There are Lu-PSMA treatments that are already given in the UK but on a private basis’ Bart explains. ‘Whether that will hit the NHS depends on approval by NICE but given the fantastically positive data out there, the upcoming results of the VISION Phase 3 clinical trials that are very positive, and given the improvement in actual survival of patients, I think there is good hope there that that will be approved.’

– Professor Bart Cornelissen

“Perhaps, what is most exciting is that, by targeting PSMA, this therapy delivers the radiotherapy directly to the sites of the cancer, wherever they are located. It is heartening to see such progress with this treatment, and I look forward to it becoming more widespread in the future.”

– David Matheson, prostate cancer patient

For more information, please visit: www.pcr.org.uk. Full story on the Department of Oncology website.

Artwork donated to Oxford ward for patients undergoing cancer clinical trials

A generous donation by artist Lida Sherafatmand has seen three prints installed into the Early Phase Clinical Trials Unit at the Churchill Hospital in Oxford.

T-cell landscape mapping identifies new targets for pancreatic cancer immunotherapy

Pancreatic cancer has one of the worst prognoses of any cancer, with pancreatic ductal adenocarcinoma (PDAC) patients having an average survival rate of 7%.

T-cells (the lymphocytes that play a wide range of roles in shaping the body’s immune response to cancer) are known to be less active in pancreas tumours. So far, checkpoint therapy trials, a type of immune-therapy that targets T-cells and have curative properties on other cancer types, have had minimal effect on pancreatic cancer with a response rate of only 5-10%. Furthermore there has been no lasting impact on a patient’s survival chance and current approved checkpoint therapies are focused on only targeting only two T-cell checkpoints, known as PD-1 and CTLA4.

In order to better understand checkpoint treatment has had a minimal impact on pancreatic cancer, and how to improve their efficacy, there is a need to understand the specific sub-populations of T-cells that are involved in pancreatic cancer. Even though we know T-cells exist in the microenvironment of pancreatic cancer, not much is known about why they are less active. There is also a need to identify new checkpoint therapy targets, beyond the two currently used, so that new, more impactful drugs may be developed.

A new study from researchers in the PancrImmune network at the University of Oxford, has characterised the immune landscape, and specifically the different T-cells, in pancreatic cancer patients, in the hope of understanding the features to aid drug development and novel therapeutics for this disease. This is the first comprehensive characterisations of T-cells in primary human pancreatic ductal adenocarcinoma.

The team looked at 32,000 T-cells from 8 cancer patients, to see if there were any unique T-cell subtypes in the tumour microenvironment. First, this data confirmed that the microenvironment of pancreatic cancer is extremely suppressive and could be a major driver of poor prognosis. Secondly, they have also identified important genetic components of these T-cell subtypes that may be driving this immunosuppression, which could be potential targets for future immunotherapy drugs.

Specifically, their observations showed an activated regulatory T-cell population, which was characterized by a highly immunosuppressive state with high TIGIT, ICOS and CD39 gene expression. The exhausted CD8 T-cells had lower PD1 levels but high levels of TIGIT and tim3. As well as the presence of a significant senescent T-cell population – this is when cells have gone down an irreversible cell cycle arrest and are no longer responsive to antigen stimuli.

This means that new potential checkpoint immunotherapy avenues in TIGIT, ICOS, CD39 and Tim3, that target these populations, may have more potential to improving the prognosis of pancreatic cancer.

The next step in this research is to take the newly-identified immunotherapy target and begin clinical experiments using targeted immunotherapy drugs. This may eventually lead to clinical trials to test these drugs in patients.

Mapping the immune landscape in PDAC patients is a huge step in pancreatic cancer research, as PDAC tumours represent 50% of all pancreatic cancer diagnoses. So new targets identified in this study has the potential to generate novel drugs that could benefit a large range of patients.

About the study

This study was co-authored by researchers in the PancrImmune Network, including  Dr Shivan Sivakumar (Dept of Oncology), Dr Enas Abu-Shah (Kennedy Institute of Rheumatology, Prof Mark Middleton (Dept of Oncology), Dr Rachael Bashford-Rogers (Wellcome Trust Centre for Human Genomics),  Prof Michael Dustin (Kennedy Institute of Rheumatology), Mr Michael Silva (University Hospitals NHS foundations Trust) and Mr Zahir Soonawalla (University Hospitals NHS foundations Trust).

New clinical prediction tools for myeloma

Myeloma is a cancer of the bone marrow that caused 117,077 deaths worldwide in 2020 (International Agency for Research on Cancer). Earlier diagnosis improves the rate of survival but unfortunately, delays in myeloma diagnosis are common and result in poorer patient outcomes.

One of the reasons for the diagnostic delay is that myeloma symptoms are non-specific and relatively common in people without cancer. For example, back pain is associated with myeloma yet there are many other non-myeloma causes of this symptom. Additional measures are therefore needed to highlight the possibility of myeloma in patients where GPs do not originally suspect this disease.

GPs frequently order simple laboratory tests, such as the full blood count, to investigate patients presenting with non-specific symptoms. Previous work by Dr Constantinos Koshiaris, Dr Jason Oke, Dr Brian Nicholson and colleagues from Oxford’s Nuffield Department of Primary Care Health Sciences and the University of Exeter identified certain abnormalities in blood test results that indicate a higher risk of myeloma, such as low haemoglobin which can be observed up to 2 years before a myeloma diagnosis.

In this paper published recently in the British Journal of General Practice, the Oxford researchers have developed new clinical prediction models for myeloma that incorporate both symptoms and blood test results. Using the Clinical Practice Research Datalink (GOLD version), a primary care database containing electronic health records for more than 11 million patients in the UK, the team identified the most common symptoms and full blood count results recorded for patients with myeloma. The most predictive of these were included in the models they developed and the new tools were validated against a set of test data. Decisions made using their prediction models resulted in fewer false positives and more true positives when compared to single tests or symptoms alone.

By identifying patients at highest risk of myeloma in primary care, these new prediction rules have the potential to reduce diagnostic delays by a substantial amount. Further research is now needed to understand more about the feasibility and implementation of this tool in the primary care setting and the impact it will have on the diagnostic pathway and patient outcomes.

Oxford at the American Association for Cancer Research Annual Meeting

The American Association for Cancer Research (AACR) Annual Meeting is being held digitally this year from the 10-15 April and 17-21 May. Oxford cancer researchers will be involved in live-streamed presentations, panel discussions and poster sessions.

The AACR Annual Meeting program covers the latest discoveries across the spectrum of cancer research—from population science and prevention; to cancer biology, translational, and clinical studies; to survivorship and advocacy—and highlights the work of the best minds in research and medicine from institutions all over the world.

Register to attend this event here or check out what Oxford’s researchers will be talking about below.

 

Drug Development & Therapies

Immunology

Genetics

Data Science

Early Detection Award for research into the clinical application of single cell genomics

Myelodysplastic syndromes (MDS) are a group of blood cancers in which the bone marrow fails to make normal levels of blood cells. MDS can be broadly classified into two major groups: high-risk MDS, in which patients progress to acute myeloid leukaemia with a very poor survival rate; and low-risk MDS, in which the disease is less aggressive but patients still suffer from a huge burden of symptoms, often the result of anaemia.

There are a number of exciting new targeted treatment options for low-risk MDS. However, these do not work in all patients and, particularly given the high economic cost of newer treatments, current biomarkers are not sufficiently predictive of treatment response. There is a need to more precisely categorise MDS to predict the disease trajectory and the response to therapy so that the most effective treatment can be given to each patient.

Large investments in sequencing technology in clinical laboratory services are enabling precision medicine in certain cancers and revolutionising patient care. Dr Onima Chowdhury, MRC Clinical Academic Research Fellow and Consultant Haematologist (MRC Weatherall Institute of Molecular Medicine and Oxford University Hospitals) is working with Professor Adam Mead, Dr Supat Thongjuea and Dr Lynn Quek at the MRC WIMM to explore the use of single-cell genomics in the clinical diagnosis and management of MDS. Funded by a Cancer Research UK Early Detection and Diagnosis Primer Award, the team will seek to develop a simple, clinically applicable processing and analysis pipeline, as well as identifying biomarkers that correlate and can perhaps supersede current diagnostic modalities.

Long-term, the team hope that this approach will be able to improve outcomes of patients through improved diagnosis, risk prediction and targeted treatment in MDS and other haematological malignancies.