Chapter 11: Fighting the Enemy Within 

Alexander Fleming wasn’t simply some fortunate man who struck lucky. --[기호] it is very unlikely that Fleming was the first scientist whose bacterial cultures had become infected with mould.  

Alexander Fleming wasn’t simply some fortunate man who struck lucky. His insight wasn’t a fluke. --à it is very unlikely that Fleming was the first scientist whose bacterial cultures had become infected with mould.→→his achievement came in recognizing that something unusual had happened, and appreciating its significance. Knowledge and training had prepared Fleming’s mind to make the most of the chance event. He saw what probably many others had seen before him but he thought what nobody else had thought.  

Epigenetic modifications control cell fate →→ its’s these processes by which liver cells, for example, stay as live cells and don’t turn into ther cell types.  Cancer represents a breaksown in normal control of cell fate, because liver cells stop being liver cells and become cancer cells, suggesting that epigenetic regulation has become abnormal in cancer. →→ we should therefore aim to de3velop drugs that influence this epigenetic mis-regulation.

The accidental epigeneticist

Peter Jones →→ a young South African scientist working with 5 -azacytidine.

5-azaxytidine it’s very similar in chemical structure to base C(cytidine)from DNA and RNA. →→ it was assumed that 5-azacytidine get added into DNA and RNA chains. Once there, it somehow disrupted normal copying of DNA and transcription or activity of RNA. →→ if 5-azacytidine was interfering with one or both of these processes, it would probably hamper the growth and divisions of the cancer cells. →→ one morning, after the cells had been cultured with 5-azaxytindine for several weeks, the researchers found that here was a strange lumpy bit in one of the culture flask. To the naked eye, this initially looked like a mold infection. Instead of discarding the flask. Peter Jones looked at the lump more closely and discovered it wasn’t stray bit of mold at all. It was a big mass of cells, which had fused to form giant cells containing lots of nuclei. These were little muscle fiber. →→→ cell lines treated with 5 azacytidine could also form other cell types, fat cells, chondrocytes. →→ 5-azacytidine causes a reversion to a more pluripotent state. →→in interview for university of southern California, Robert Stellwagen asked “have you thought of DBA methylation? →→ Peter Jones admitted he not only hadn’t thought of it he hadn’t even heard of it. →→ after that, Peter began to focus on DNA methylation and in a very short time showed that this was indeed key to the effects of 5-azacytidine. 5-azacytidine inhibited DNA methylation.  

The methylation Cu-de-sac

DNMT1 binds at region where the DNA contains 5-azacytidine instead of the moral cytidine, the DNMT1 becomes stuck there. This marooned enzyme then sent to a different part of the cell and is broken down.→→→ because of this the total levels of DNMT1 enzyme in the cell fall. →→The combination of this decrease in the amount of DNMT1, and the fact that 5-azacytidine can’t be methylated, means that the amount of DNA methylation in the cell keeps dropping. We’ll come back in a little while to why this drop in DNA methylation has an anti-cancer effect.  [kp1] 

Another happy accident

Charlotte Friend DMSO(dimethyl sulfoxide) had an odd effect on the cancer cells from a mouse model of leukemia. →→  Ronald Breslow Paul Marks and Richard Rifkind create a new set of chemicals, using the structure of DMSO →→→ after many iterations, learning from each new and more complicated set of structures,  created SAHA(suberoylanilide hydroxamic acid)→→ was really effective at stopping growth and /or causing cell death in cancer cell lines.

Trichostatin A  or TSA→→→ known to be able to stop cells proliferating. →→ when cells were treated with TSA, the levels of histone acetylation went up.  This is because TSA was inhibiting the enzymes that remove acetyl groups from these chromatin protein.

Fig 11.2 compare the structure for TSA and ASHA

TSA and SAHA was also an HDAC inhibitor.  SAHA prevents HAAC enzymes from removing acetyl groups from histone proteins, and as a result, the histones carry lots of acetyl groups.

Beyond coincidence  

5-azacytidine and SAHA both decrease cancer cell proliferation, and both inhibit the activity of epigenetic enzymes.

Epigeneticists believe that is far from being a coincidence. →→ DNA methyltransferase enzymes add a methyl group to the cytidine base. Higher concentrations of this base are found in the long CG-rich stretches of DNA known as CpG island is heavily methylated, the gene controlled by that promoter is switched off. In other words. DNA methylation is a repressive modification. DNMT activity increased DNA methylation and therefore represses gene expression. By inhibiting these enzymes with 5-azacytidine, we can drive gene expression up.

Histone protein are also found at the promoters of genes. Histone acetylation is the most straightforward in terms of its effects on gene expression. If the histones upstream of a gene are heavily acetylated, the gene is likely to be highly expressed. If the histones are lacking acetylation, the gene is likely to be switched off.  Histone deacetylation is a repressive change. Histone deacetylases (HDACs) remove the acetyl groups from histone protein and will therefore repress gene expression. By inhibiting the enzymes with SAHA, we can drive gene expression up.

Cancer biology 101

Cancer is the result of abnormal and uncontrolled proliferation of cells. Normally, the cells of our body divide and proliferate at exactly the right rate. This is controlled by a complex balancing act between networks of genes in our cells. Certain genes promote cell proliferation. These are sometimes referred to as proto-oncogenes. They were represented by a plus sign in the see-saw diagram fig 10.5

Other genes hold the cell back, preventing too much proliferation. These genes are called tumor suppressors. Represented by a negative sign on the same diagram.

Cancer cells divide too often, they are also abnormal and can start to invade other tissue. →→ the more of these moles or polys you have, the greater the likelihood that one of them will go next step, and develop an abnormality that will take it further along the path towards full-brown cancer.  →→→ cancer is a multi-step process. →→→ cells accumulate defects as they move increasingly close to becoming cancerous.  These defects must be transmitted from other cell to daughter cell,

Two copies of each tumor suppressor gene. →→ cell becomes increasingly cancerous, both copies of key tumor suppressor genes usually get switched off(inactivated)→ these is called somatic mutations to distinguish them from genetic mutations, the one that are transmitted from parent to child.

If the DNA at the promoter becomes excessively methylated or the histones are covered in repressive modifications, the tumor suppressor will be switched off. The gene has been inactivated without changing the underlying blueprint.

The epigenetic frontier in cancer.

Clear cell renal carcinoma→→→ inactivation of a specific tumor suppressor gene called VHL→→ analyze the CpG island in front of the VHL gene. the DNA of the island was hypermethylated.

BRCA1 tumor suppressor gene in breast cancer.  Which no mutation in BRCA1 → BRCA1 CpG island was hypermethylated.

The histones associated with t tumor suppressor gene called ARH1 had low levels of acetylation in breast cancer. →→ a similar relationship exists for the PER1 tumor suppressor in a form of lung cancer called non-small cell. →→ in both cases, there was a relationship between the levels of histone acetylation and the expression of the tumor suppressor – the lower the levels of acetylation, the lower the expression of the gene. →→→ because these genes are both tumor suppressors, their decreased expression would mean that the cell would find it harder to put the brakes on proliferation.

Tumor suppressor genes are often silence by epigenetic mechanisms. →→→ creates a new way of treating cancer. Tumor suppressor genes back on in cancer cells, there is a fighting chance of reining in the crazy proliferation rate of those cells.

Gene therapy when tumor suppressor were inactivated by mutations of deletions. Gene therapy →→ very difficult to get the genes delivered into the right cells. Relatively rare cases where the gene therapy itself has led to cancer, expensive and difficult. →→ that’s why there is so much excitement about the development of epigenetic drugs to treat cancer.

Two drug that inhibit the DNMT1 enzyme are 5 azacytidine and 2-aza-5 deoxycytidine. →→ two HDAC inhibitors have also been licensed. →→→ SAHA and romidepsin

Stop looking for miracles

DNMT1 inhibitors and HDAC inhibitors seem to rein in the cancer cell growth, retarding and repressing it. They control rather than cure.

No easy wins

Fig11.3 →→ repressive histone modifications attract DNA methyltransferases, which deposit DNA methylation near those histones. →→ this methylation in turn attract more repressive histone modifying enzymes, creating a self perpetuating cycle 못 lead to an increasingly hostile region for gene expression.

Repressive histone modifications can act as the bait to attract DNA methylation to the promoter of a tumor suppressor  gene. A key example of this is an epigenetic enzyme we met in the previous chapter, called EZH2. →→ The EZH2 protein adds methyl groups  to the lysine amino acid at position 27 on histone H3. This amino acid is known as H3K27.

An uneasy truce

Most evolutionary pressures are connected with the drive to leave behind as many offspring as possible. For a human to reach reproductive age, it’s incredibly important that early development occur as efficiently as possible. After all, you can’’t reproduce if you never make it past the embryo stage. Once we’ve reached reproductive age and had the opportunity to reproduce, there is little to be gained in evolutionary terms in us staying alive for several decades afterwards.

Regulatory systems that allow cells to be flexible are inevitable also systems that allow cells to go wrong. →→ in evolutionary terms, it’s the price we have to pay for our Goldilocks scenario. →→ our epigenetic pathways make sure some of our cells aren’t completely pluripotent or completely differentiated. Instead, they are just right, hovering somewhere near the top of Waddington’s epigenetic landscape, but ready to roll down at any time.

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 [kp1]