Tag Archives: epigenetics

neutropenia and garlic

All cancer chemotherapy is unpleasant, but the chemo we use for breast cancer is particularly nasty. Megan is 48 and was admitted this week with a fever and diarrhoea, a week after her third course of CEF – cyclophosphamide, epirubicin and 5-flurouracil.  Her breast cancer was diagnosed a few months ago – she noticed a lump in her left breast when washing herself in the bath. Megan could tell from the face of her GP when she was examined that it was going to be bad.

Then, shortly after, there was the ultrasound, mammogram and trucut needle biopsy at the breast clinic which hurt more than she was expecting – Megan was really scared by this time.

mammogram of breast showing a cancer bottom right
mammogram of breast showing a cancer bottom right

She remembers vividly the meeting with the breast surgeon a week later, but only as a jumble of “surgery”, “partial mastectomy”, “total mastectomy”, “node clearance”, “chemotherapy”, “radiotherapy”. Her husband Jack was with her, but it didn’t make much sense to him either. They went home with a pile of leaflets and phone numbers.

The surgery went fine, everyone was really nice and caring. Then a meeting with the oncologist. She advised chemotherapy then radiotherapy. That seemed much more simple. But chemotherapy was awful. Megan felt sick and hopeless. All she could do was sit in bed and do crosswords. Just as she was starting to feel better the next round of chemo was due.

epirubicin is an anthracycline antibiotic chemotherapy agent which works by damaging DNA
epirubicin is an anthracycline antibiotic chemotherapy agent which works by damaging DNA

When I met her she had the look of a usually bright and cheerful person who was wondering if she could take any more. I told her that her white cell count was low, and we needed to give her intravenous antibiotics until her fever resolved and white cell count recovered.

How does cancer chemotherapy work? Why does it kill cancer cells in preference to normal cells in our body?

The normal answer I get from the young doctors and students is that chemotherapy targets rapidly-dividing cells. It really is not quite as simple as that. Breast cancer cells divide much less rapidly than bone marrow or gut epithelial cells, but chemotherapy is designed to kill all the cancer cells and leave our bone marrow and intestinal lining intact. To answer the question properly, we need to understand what makes cancer cells malignant.

We have a much better understanding of the molecular basis of cancer than we did thirty years ago. Cancer cells usually have a number of genetic abnormalities which result in the cells no longer behaving in a useful and regulated way. Cells, whether they are from breast, colon, lung or any other tissue need to behave in a regulated and responsible way. They need to co-operate with the cells around them, working together to make sure the organ works properly. Like in normal society, where people have to abide by rules which make sure everything works properly. If someone misbehaves badly, the police force and courts act to stop them doing the bad thing. Similarly, if a cell is damaged and behaves abnormally, the body has ways of making sure that is either removed or repaired. Our immune system is constantly removing abnormal cells, and all cells have a mechanism to detect and repair DNA damage. One of the main mechanisms to identify DNA damage is the p53 system. The p53 gene is turned on when DNA damage is detected. This causes p53 protein to be made and cause the cell to stop dividing until the DNA damage is repaired. If the DNA cannot be repaired the cell undergoes programmed cell death. Harsh, you may think, but necessary to prevent the cell dividing and making more abnormal cells – “here is a pistol – I think you need to go to the library and consider your options”.

When cells have damaged DNA which cannot be repaired they do the decent thing
When normal (non-malignant) cells have damaged DNA which cannot be repaired they do the decent thing

The problem comes when the law enforcers are compromised. Corruption in the police force or judges is disastrous for society and for our bodies. Most common cancers have damage to the p53 gene, meaning that cells will divide even when DNA is damaged. Understanding this helps us understand why traditional chemotherapy targets cancer cells.

Cyclophosphamide is a nitrogen mustard alkylating agent which damages DNA by permanently crosslinking the two DNA strands. Nitrogen mustard has been molecule of the month: find out more about its chemisty here:

http://www.bris.ac.uk/Depts/Chemistry/MOTM/mustard/mustard.htm

In normal cells this damage is detected and prompts an activation of the p53 gene causing production of p53 protein –  with consequent shutdown of cell division until the damage is repaired (or, if the damage cannot be repaired the cell commits sucicide). In cancer cells which cannot make p53, the cells will continue to divide and render them more susceptible to further DNA damage. This is because when cells are dividing the DNA is unravelled and more exposed to the damaging chemotherapy agent.

Continued division of cancer cells will also make them more susceptible to 5-fluorouracil. This drug inhibits the enzyme thymidylate synthase, preventing the production of normal thymine, one of the DNA bases. The cells are trying to divide but cannot make thymine and suffer a fate known as “thymineless death”.  Cells which have normal p53 will have shut down and stopped dividing, and will not suffer in this way.

Epirubicin is a member of a group of drugs called anthracycline antibiotics, derived from steptomyces bacteria. These substances also work by binding to DNA and interfering with cell division, again targeting the breast cancer cells which are still dividing, despite having their DNA damaged, because of defective p53.

This combination of drugs work well to kill the breast cancer cells, but the shutdown of normal bone marrow production and intestinal epithelial cell division still causes a major problem, as evidenced by Megan and her neutropenic sepsis and diarrhoea. Without white cells and gut epithelial cell production she cannot not defend herself against normal germs.

Patients undergoing chemotherapy are given a whole pile of leaflets including advice about neutropenic sepsis
Patients undergoing chemotherapy are given a whole pile of leaflets including advice about neutropenic sepsis

Damage to the p53 gene is not the only abnormality that causes breast and other cancers. Most breast cancer cells have a large number of genetic abnormalities, many of which we know are important in making the cells cancerous, but we do not know how. There is a really good video which explains what we know about the genetic abnormalities in breast cancer and other cancers by Prof Sir Mike Stratton FRS –

Also, it is being recognised that epigenetic abnormalities are common in cancer cells, with differences in DNA methylation between normal and cancer cells. As I mentioned in a previous post, epigenetic changes are likely to be important in ageing, as well as cancer. I think there maybe an important link between these.

What causes the DNA damage in breast cells?  That is a big problem to understand. It is easy to imagine how lung epithelial cells are exposed to cigarette smoke which contains carcinogens, and how colon epithelial cells may be damaged by nasty substances in our diet. But breast cells – why do they get damaged? The main risk factors for breast cancer are:

1 early menarche (onset of menstrual periods at an early age

2 late first pregnancy

3 fewer pregnancies

4 obesity

5 alcohol

How do these factors damage DNA and cause cancer? I do not know. Answers please.

Now on to the food link – garlic.

garlic has its characteristic odour because it contains the molecule allicin, which helps protect it from bacteria and insects which want to eat it
garlic has its characteristic odour because it contains the molecule allicin, which helps protect it from bacteria and insects which want to eat it

Cyclophosphamide is a nitrogen mustard. It is called that because mustard gas was thought to smell like mustard. In fact it smells more like garlic.

poster from first world war
poster from first world war – wikimedia commons

Garlic contains a substance known as allicin, which has a similar molecular shape to nitrogen mustard.

allicin and nitrogen mustard have a similar molecular shape - which is maybe why they have a similar smell
allicin and nitrogen mustard have a similar molecular shape – which is maybe why they have a similar smell

It has been suggested that garlic has all manner of wonderful properties to keep us healthy, including lowering cholesterol, preventing heart attacks and protecting us against infection (and vampires). The hard evidence is not there. But as Neils Bohr, the eminent physicist, said when a visiting journalist remarked “ why Professor Bohr, do you have a horseshoe above your front door – surely you cannot believe that it will bring you good luck?” replied:

“Of course I don’t believe in it, but I understand it brings you luck, whether you believe in it or not.”

That’s more or less what I think about garlic.

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old age and malbec

Dorothy was born in 1911. She is 102. When someone over 100 is admitted to our ward there is always a lot of interest – but it is happening more and more. Dorothy, or Dot, as she is known by her friends, has been living on her own in sheltered housing. She has her own ground floor flat, but eats meals with the other elderly residents and has help with washing and dressing in the morning, and there is always a manager on hand if there are problems. Slide1She can make herself simple meals and cups of tea, and her 78 year-old son, David, usually visits every Sunday.  I saw her in our ward last week. She had fallen over on Sunday night and bashed her head on the edge of a table, resulting in a huge bruise over her right eye. She has atrial fibrillation and is on warfarin – the ambulance crew brought her in to hospital as she was a bit confused and they were worried she might have a blood clot on her brain.

The CT head scan on the left of a healthy 96 year old woman, the one on the right is of a 23 year old woman. The dark portion in the middle of the old brain is fluid, not brain. The younger person's head is completely full of brain.
The CT head scan on the left is that of a healthy 96 year old woman, the one on the right is that of a 23 year old woman. The dark portion in the middle of the old brain is fluid, not brain. The younger person’s head is completely full of brain.

Luckily the CT scan of her head showed no evidence of serious bleeding, although her brain was considerably shrunken compared to a young person. She was quite deaf, but clearly had most of her marbles. She recounted how her father had fought, and was wounded, in the Battle of the Somme in the first world war, and her husband Rodney, who died in 1980 had fought in the second. She worked in a munitions factory during the war and had three children – all boys. It’s strange calling elderly people children, but offspring or any other word does not seem quite right. Only the youngest was still alive. She could remember the names of all 5 grandchildren and 7 great-grandchildren, but did struggle with the name of the Prime Minister and what year it was. Her brain was old as well as her skin and her ears, and all the rest of her.

How much longer will Dot live? There was an entertaining paper in the Christmas BMJ in 2011. http://www.bmj.com/content/343/bmj.d7679

It has been shown that for any cohort of elderly people, the main determinant of longevity is how old you are – as we get older the more likely we are to die sooner. The second most important factor is gender – women live longer than men. The third, somewhat surprisingly, is how fast you can walk. This is probably because being able to walk fast requires a lot of things to be working well – good lungs, good cardiac function, good muscles, good joints, good brain function, good eyesight, good peripheral nerve function etc. The paper suggests another alternative explanation – that if you walk slowly the grim reaper will catch up with you.

One explanation of why elderly people die soon if they cannot walk fast. Illustration by Belle Mellor - http://bellemellor.com (with permission - thanks Belle)
One explanation of why elderly people die soon if they cannot walk fast. Illustration by Belle Mellor – http://bellemellor.com
(with permission – thanks Belle)

Why do people get old? This is a difficult question, and I cannot give you a definite answer, but will give you my thoughts.  Ageing (aging in the US) seems inevitable.  We can guess how old someone is by their skin, which becomes wrinkled – their hair becomes thin and grey, and their posture becomes bent. The cells in our skin, bones, brain and just about everywhere else continually become older, but some of our cells do not age in the same way. Think about a sixty year old man who becomes a new father.  His sperm have contributed to make a baby that is not old, but brand new. The cells in his testes which have made the sperm are old, but they must have in them the information about how to make a cell which is not old. This, to me, very much suggests that old cells have not lost the information in the form of DNA sequence that they had when they were new. The creation of Dolly the sheep confirms this. Dolly was made from the nucleus of  her “mother’s” mammary gland cell (she was called Dolly after Dolly Parton, who has very impressive mammary glands). In a test-tube this nucleus was inserted into an egg cell of another sheep (also her mother?) and processed to de-differentiate it.

Dolly Parton - from Wikimedia commons
Dolly Parton – from Wikimedia commons

This means the nucleus of the mammary gland cell was changed so that instead of only being able to make more, old, mammary gland cells, it could make all the new and different cells of baby Dolly the sheep. So, I’ll ask the question again, why do cells get old? It’s not that they have lost the information about how to be new cells, but nature has determined that after a certain period of time they change into old cells. What is happening in the cells to make them old? – clearly they are not so damaged that the DNA sequence is irreparably altered. There must be a clock inside cells that tells them how old they are – this is suggested by the classic studies of Leonard Hayflick, who showed that cells from human babies only divide about 50 times, and then stop.

cells must have some sort of clock inside them - it may well be the amount of methylation of DNA in the nucleus
cells must have some sort of clock inside them – it may well be the amount of methylation of DNA in the nucleus

Cells from older people divide far fewer times. It was thought for many years that the reason cells could only divide a certain number of times was because each time division took place, the teleomeres at the end of each chromosome become shorter. When they become too short, the cell can no longer divide. Teleomeres have been likened to like the plastic bits at the end of shoelaces – they stop the lace from fraying and are necessary in chromosomes to keep the DNA strands from coming apart.

It may well be that short teleomeres contribute to the cellular clock, but it seems they are not the only answer.

teleomeres are on the end of chromosomes like the bits of plastic on the end of shoelaces - to stop them fraying
teleomeres are on the end of chromosomes like the bits of plastic on the end of shoelaces – to stop them fraying

More recent research suggests that epigenetic changes in DNA may be responsible. Epigenetics is a very fashionable science. Once a cell is differentiated in the growing embryo – say into a skin cell – its DNA becomes progressively altered. In particular methyl groups (CH3) become attached to a cytosine nucleotide when a cytosine and guanine are next to each other in the nucleic acid chain (CpG). This methyl group attached to the cytosine is copied when the cell divides. Recent research suggests that the age of cells is closely related to the number of methyl groups attached to CpG sequences in DNA.

http://genomebiology.com/2013/14/10/r115

What to me is really interesting, is that when sperm cells meet egg cells and their nuclei fuse to make the start of a new baby, all the methyl groups are stripped from the CpG sequences in DNA.  This looks like a definite candidate for the cellular clock.

But why do we need to age? What if cells stayed “new”? What if we could stop the cellular clock? If we could inject a substance into Dot which reversed the epigenetic changes would she suddenly get lovely wrinkle-free baby-like skin and a fully functioning brain, ears and eyes? I don’t know, but I suspect not – answers please.

Why did evolution invent a clock in our cells to stop them dividing as we get older? One explanation for this is that cells become damaged with time, and dividing may not be a good thing for damaged cells as it is more likely that they will become malignant. We now know that most malignant cells develop a sequence of gene defects over time, which when added together cause the cell to become cancerous. Maybe the clock, which limits the number of divisions, is a way of preventing further DNA damage and malignancy. Certainly it seems that reactive oxygen compounds that can cause damage to DNA also cause CpG methylation – in a way causing premature ageing of cells.

I used to think that the reason our cells got older was so that we would die and no longer be competition for our children, who, through evolution, had become more fit than us.  It seems this is not the explanation because there is very good evidence that early humans did not die from age-related causes, but from infections and trauma.  The average life-span in prehistoric times was much lower than in modern times, most people dying before 30 and nearly all before 40 years of age.  This means that it is not possible for evolution to select for, or against, those people who lived longer, because there were so few of them. Instead it seems that evolution only cares about keeping us alive until we have produced and reared babies – after that it has no interest or influence one way or the other. If there was a new gene that made a prehistoric human live longer, it would be extremely unlikely that the owner would survive longer and there would be no positive or negative pressure to keep it. This does not mean that our genes cannot influence how long we live. An important factor in determining how long we live is how long our parents lived. Another is where we live and our lifestyle.

Roger Corder, scientist and author of The Red Wine Diet
Roger Corder, scientist and author of The Red Wine Diet

There is a wonderful book by an erstwhile colleague of mine, Roger Corder, called The Red Wine Diet, which suggests that eating foods with certain polyphenols can make us live longer.

polyphenols are large molecules made up of lots of phenols - they are useful antioxidants which trap oxygen radicals
polyphenols such as tannins are large molecules made up of lots of phenols – they are useful antioxidants which trap oxygen radicals

Polyphenols are found in plants and are often brightly coloured. Tannins are polyphenols, and cause a characteristic “furry” taste in the mouth because they react with the protein molecules on the surface of our tongue. These molecules are powerful antioxidants. What does that mean, and why do plants make them? In the case of a grape, one of its main concerns is DNA damage by ultraviolet light. Although short wavelength UV light can directly damage DNA, it seems that less powerful, longer wavelength rays can generate very reactive oxygen radicals such as singlet oxygen and hydroxyl, which then combine with, and damage DNA. Polyphenols can rapidly “mop up” or detoxify these oxygen radicals and protect DNA.  If you live in the mountains of SW France, Sardinia, Crete or Sicily, the plants, and especially grapes, are subjected to large amounts of UV irradiation and produce lots of polyphenols. The people who live eat and drink lots of these polyphenols live to an unexpected old age. It might be a coincidence, but, for me, the inconvenience, expense and sheer tedium of drinking three glasses of red wine a day seems worth a try. But don’t overdo it and do remember Kevin (vodka and sweetbreads) below. The food link this week is a drink – Malbec.

Argentinian Malbec is full of tannins
Argentinian Malbec is full of tannins

This is a grape variety, which, according to Roger Corder, contains very high levels of procyanidins – tannins which he thinks helps protect us from heart disease and cancer, and makes us live longer.  Antioxidants prevent the oxidation of LDL cholesterol (see previous post on chest pain and horsemeat lasagne) and reduce DNA damage that can make normal cells malignant and age more quickly. It is grown in the mountains of SW France and is the main variety in Cahors wine – a particularly dark and astringent glassful. Argentinian Malbec is more available in the UK and is also grown high up in the mountains where it gets plenty of ultraviolet light.