How cell-free chromatin particles could be the missing link to the long-awaited treatment of sepsis

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Septicaemia is now responsible for nearly 20% of the global death toll due to life-threatening inflammation. As we begin to truly grapple with the scale of the problem, researchers and clinicians have struggled to develop treatments of a condition with unknown pathological mechanisms. However, Professor Indraneel Mittra has led research into cell-free chromatin particles, that had been hitherto neglected, and has linked them to many diseases due to the ability to induce DNA damage and inflammation, including sepsis. With this paradigm-shifting new approach, Professor Mittra and his colleagues have looked at new treatments for sepsis, which could help treat other diseases, including COVID-19.

If there is one concept that people have recently become all too familiar with, it’s the concept of a public health crisis. Even discounting our current COVID-19 pandemic which has brought the world to a halt, our history is littered with many pandemics, epidemics, and healthcare crises. You may think of pandemics of the past, such as the AIDS pandemic or the BSE crisis. Or you might think of current conditions which induce fears from doctors and experts around the world, like Type 2 diabetes. But you likely aren’t aware of the crisis surrounding septicaemia.

This perhaps shouldn’t come as such a surprise. Indeed, the World Health Organization has only published their first global report of septicaemia (or sepsis) this September, one which admits to “an urgent need for better data”. Even our healthcare systems will admit they need to improve their awareness of sepsis. The WHO’s pioneering report found that worldwide, there were nearly 49 million cases and 11 million deaths in 2017, accounting for nearly 20% of the global death toll. That’s more deaths than from cancer, every single type of cancer, combined. Like many diseases, sepsis doesn’t afflict its potential patients equally. The WHO report discovered a startling 85% of cases were recorded in newborn children, pregnant women, and low-income people, underscoring its unequal impact.

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Attempting to Understand Septicaemia

Septicaemia isn’t a disease or infection. Instead, it is a condition which arises as a consequence of a microbial infection. Sepsis describes the tragic aftermath following pathogenic microbes entering the blood, as the infection induces a severe immune reaction towards the invading infection. The reaction becomes a potentially deadly overreaction within 24 hours, as the body’s immune response ends up destroying its own cells, thus harming its own organs.

The ‘overreaction’ is defined by multiple simultaneous processes, overwhelming the body with tissue damage, organ damage, and the paralysis of the immune system. The body enters a hyper-inflammatory state from a cytokine storm, where chemical signals overwhelm and paralyse the body’s immune system. This induces apoptosis, the cell’s self-destruct mechanism programmed into itself, triggered upon severe damage to the cell, or from its activation by the immune system. The apoptosis of cells in the spleen, thymus, kidney, and liver leads to multiple organ failure, internal bleeding due to an inability to clot the blood, and haemorrhaging.

This begins a deadly positive feedback loop, where the host cell now undergoing apoptosis inevitably releases cell-free chromatin particles.

However, the physiological mechanisms that underpin sepsis have been elusive to researchers. This is likely why clinical trials for sepsis treatments have been universal failures. Scientists believed that sepsis directly resulted from an immune response to certain microbial infection, but this was more of an educated assumption than a definitive model. Consequently, the call for research into treatments is no longer merely looking for new compounds, but a wholesale paradigm shift for a new approach towards developing a sepsis treatment. No current treatment exists, as every drug candidate falls at the hurdle of clinical trials. One group that has answered this call is Professor Indraneel Mittra’s research group at the Tata Memorial Centre, Mumbai, who have found that cell-free chromatin particles could provide the paradigm shift for a new – and successful – way to fight sepsis.

Apoptosis is the process of programmed cell death.

Cell-free chromatin

Eagled-eyed readers will be right to think of chromosomes. As the name suggests, cell-free chromatin is chromosomal material which has escaped the confines of its cell. But rather than being a freed criminal or prisoner, cell-free chromatin molecules are closer to the fragments of the blown up prison. They are ejected from cells that have undergone apoptosis – the cell’s hardwired process for its own destruction, triggered by the cell upon detecting severe damage to itself.

Professor Mittra’s past research has included work on understanding the fundamental science behind cell-free chromatin. They are invasive fragments of DNA tightly coupled with histone proteins, entering healthy cells and inserting themselves into the genome of the host cells. This integration is a catastrophic event for the cell leading to disruption and re-writing of its genome that sets off alarm bells in the cell. This catastrophic event often leads to apoptosis or death of the cell with release of cell-free chromatin particles. The latter triggers an inflammatory response in the surrounding cells. Thus, begins a deadly positive feedback loop, or a vicious cycle, leading to more cell death and inflammation in the surrounding bystander cells.

Cell-free chromatin has already been linked to cardiovascular disease, diabetes, Alzheimer’s disease, and multiple types of cancers. And Professor Mittra now believes it to be the root cause of sepsis. Cell-free chromatin break DNA, inducing a systematic inflammatory response, and apoptosis of cells in vital organs and tissues, including the spleen and thymus. This new mechanism for sepsis could provide the paradigm shift for new, successful, treatments.

When small levels of copper were combined with resveratrol, there was a significant improvement in outcomes.

A Paradigm shift in septicaemia?

Professor Mittra and his colleagues put this theory to the scientific test using mouse models to experiment with three different chromatin deactivating compounds, all potential treatments for sepsis. The mice were injected with lipopolysaccharides (LPS), a large chain of sugars and lipids which are found on the outer membranes of certain bacteria. Its association with pathogenic bacteria, as well as its constant shedding from bacteria, makes it a common inflammatory molecule that sounds the alarm of the immune system. By every parameter that could measure and describe sepsis, there were significant improvements: from preventing the inflammatory response, tissue cell death and paralysis of the immune system, to less coagulopathy (dangerous levels of bleeding due to an inability to form clots at wounds) and a lower mortality rate.

It was resveratrol that proved to be the most interesting compound. Resveratrol is a polyphenol – organic compounds found in plants which include tannins and flavonoids found in wine and plant-based foods. Resveratrol specifically are found in the skins of red grapes. It has long attracted medical interest as an anti-oxidant, though not as the body of Western research on the clinical efficacy for resveratrol were wanting, as it failed to show any clinically significant results. However, when small levels of copper were combined with resveratrol, there was a significant improvement in outcomes. The resveratrol-copper combination was found to be the most effective treatment for sepsis, with a 40% greater survival rate than the control mice that received no treatment. This is because of the free radicals provided by the copper, living up to its reputation as an excellent conductor.

Their tendency towards destructive relationships with DNA molecules suddenly becomes a useful ally of convenience, albeit a temporary alliance.

A radical solution

Free radicals are ion molecules with a free electron which makes them highly reactive as they search the body for another electron to pair with. They’re naturally occurring, with free radicals being by-products of the body’s metabolic processes, as well as being found in alcohol, tobacco, fried foods and air pollutants. As those foods and drinks suggests, this normally spells trouble for your body as the electron pairing reaction can damage DNA and cells. But their tendency towards destructive relationships with DNA molecules suddenly becomes a useful ally of convenience, albeit a temporary alliance. The free radicals that resveratrol-copper provides will bind to cell-free chromatin, just as they bind to cellular DNA. The cell-free chromatin molecules are destroyed through this binding, and the symptoms disappear.

With the success of resveratrol-copper in the laboratory, Professor Mittra and his team are now conducting human clinical trials in sepsis patients. There are still multiple hurdles to overcome. Alongside the general uncertainty that plagues medical treatments, Mittra notes that the mouse model experiments administered the treatments 4 hours before the sepsis-triggering LPS. In real life, any treatment would be provided at a time after symptoms were detected, sometimes hours after initial onset.

Anti-oxidants: natural polyphenols. Anastasiya Litvinenka/Shutterstock.com

Nonetheless, the potential of these new treatments is evident. Not only could this be the antidote for the alarming level of deaths in cases of sepsis, but it could also be used to treat a multitude of diseases and conditions. Sepsis is a source of toxicity from chemotherapy and radiotherapy, as cell death causes cell-free chromatin release, which is also a hallmark of COVID-19. A major symptom of COVID-19 is cytokine storm from inflammation, and resveratrol-copper’s remarkable ability to curtail the incredibly aggressive cytokine storm could make it the key to treating this devastating symptom of COVID-19.

What is clear that cell-free chromatin particles could be the elusive target needed to treat not just septicaemia, but many other diseases – even the one currently that has paralysed our world.


What inspired your initial interest in cell-free chromatins?

My interest in cell-free chromatin (cfCh) was the result of a serendipitous finding over 20 years ago. In 1996 two Nature papers described the detection of tumour-derived DNA in circulation. My immediate question was: can circulating DNA be involved in cancer metastasis? I tested this hypothesis on cancer cell lines for a whole year, to no avail. I then ran with the new hypothesis that it might be fragmented chromatin particles, rather than DNA, that caused cells to transform. Low and behold, this turned out to be the case and laid the foundation of what would go on to become a decade-long pursuit to understand the role of circulating cfCh in human physiology. But horizontal transfer of chromatin particles has much wider implications than just cancer mutation. Indeed, my work has gone on to show the expanded role of cfCh in a variety of phenomenon including cancer metastasis, sepsis and toxicity from chemotherapy. I am now pursuing the role of cfCh in Darwinian evolution, which I hope will help to explain how tumours evolve under clonal and selective pressure of continuous genomic damage induced by cfCh released from dying cells within the tumour.

 

References

  • Mittra, I., Pal, K., Pancholi, P., Tidke, P., Siddiqui, S., Rane, B., D’souza, J., Shaikh, A., Parab, S., Shinde, S., Jadhav, V., Shende, S., Raghuram, G. (2020). Cell-free chromatin particles released from dying host cells are global instigators of endotoxin sepsis in mice. PLoS One [online], 15 (I3), e0229017.
DOI
10.26904/RF-134-118121

Research Objectives

Professor Mittra’s research interests are wide ranging and encompass clinical, laboratory and public health research in cancer.

Funding

This study was supported by the Department of Atomic Energy, Government of India, through its grant CTCTMC to Tata Memorial Centre awarded to Professor Mittra.

Bio

Professor Mittra obtained his medical degree from University of Delhi, and did his Post Graduate Training in Surgery in the UK. He became a fellow of the Royal College of Surgeons of England. He obtained his Ph.D. degree from University of London and did his post-doctoral work with Dr Renato Dulbecco, Nobel Laureate, at the Imperial Cancer Research Fund Laboratories in London. On returning to India in 1982, Professor Mittra joined the Tata Memorial Hospital, Mumbai as a Consultant Surgeon in the Department of Surgical Oncology. Professor Mittra set up the first dedicated Breast Unit in India and became the Chief of Breast Cancer Service at TMH. Simultaneously, he held the position of Head of the Division of Laboratory Medicine that he set up at TMH. Professor Mittra has published extensively in the fields of clinical, basic as well as public health research in cancer in major international journals. He has the distinction of having published his first ever scientific contribution as a single author paper in Nature in 1974, and happens to be the first Indian scientist to have published in the journal Cell. Professor Mittra was conferred the Roll of Honours by the International Union against Cancer in 1994. He has been visiting professor to several international institutions and held the Prestigious Pearce Gould Professorship at University College London in 1998. Professor Mittra is/has been on the editorial/advisory board of British Medical Journal, Lancet Oncology, Nature Clinical Practice Oncology, British Journal of Cancer, European Journal of Cancer, International Journal of Surgery, The Breast, Journal of Biosciences and Current Science. Professor Mittra is a fellow of the Indian Academy of Sciences and Indian National Science Academy.

Professor Mittra

Contact
Professor Indraneel (Neel) Mittra
MBBS, PhD. (London), FRCS (England), FASc, FNA
Dr Ernest Borges Chair in Translational Research and Professor Emeritus, Department of Surgical Oncology
Tata Memorial Centre
Advanced Centre for Treatment, Research and Education in Cancer (ACTREC)
Kharghar, Navi Mumbai – 410 210.
India

E: neelmittra01@gmail.com
W: https://orcid.org/0000-0002-5768-3821

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