LM 12.2: Targeted Therapy; Epigenetics

Question 1

what is epigenetic?

Answer 1

any process that alters gene activity without changing the DNA sequence and leads to modifications that can be transmitted to daughter cells

involves DNA methylation and chromatin modification such as nucleosome positioning, histone acetylation and methylation

Question 2

what is DNA methylation?

Answer 2

involves the addition of a methyl group to cytosines contained within CpG residues

DNA methyl transferases (DNMts) catalyze the reaction using S- adenosl-L-methionine as a co-factor

Question 3

on what residue does DNA methylation happen?

Answer 3

occurs on cytosines at the 5 carbon position of the pyrimidine ring

Question 4

how does DNA methylation play a role in cancer?

Answer 4

In cancer, hyper-methylation of promoters regulating tumor suppressor genes silences their expression while hypo-methylation of promoters regulating oncogenes activates their expression

cancer is characterized by an overall decrease in DNA methylation genome-wide

Question 5

what are the normal functions of DNA methylation?

Answer 5

1. transcriptional gene silencing
2. chromatin compaction
3. genome stability
4. suppression of homologous recombination between repeats
5. genome defense
6. x-chromosome inactivation in females
7. imprinting

Question 6

what is CIMP?

Answer 6

CGI methylation phenotype

indicates an abundance of cancer-specific hyper-methylated promoter CpG islands/shores was first found in colon cancer and later demonstrated in other cancers

CIMP phenotype is a cancer specific increase in DNA methylation at CpG island and shores that is characteristic of many cancers

Question 7

how can you reverse hyper-methylation on a TSG CpG island and island shores?

Answer 7

DNA hypomethylating drugs can reverse hyper-methylation!

DNA methylation inhibitors are nucleoside analogues that exert their de-methylating activity through the establishment of an irreversible covalent bond with DNMTs after their incorporation into DNA

Question 8

what are DNA methylation inhibitors?

Answer 8

nucleoside analogues that exert their de-methylating activity through the establishment of an irreversible covalent bond with DNMTs after their incorporation into DNA

nucleoside analogs such as 5-azacytidine and decitabine bind to DNA methyltransferases and inhibit their activity

these drugs can be used to reverse the hyper-methylation that occurs on CpG island and island shores that encompass the promoters of tumor suppressor genes

Question 9

which drugs are DNA methylation inhibitors?

Answer 9

5-azacytidine

decitabine

Question 10

what’s the structure of a chromosome?

Answer 10

1. DNA is wrapped around a core of histone proteins to form nucleosomes. The nucleosome is the basic unit of chromatin.
2. Each nucleosome contains two copies of 4 different histone proteins with DNA wrapped around the core, 1.65 times. The diagram on the right shows how the amino (N) terminal ends of the histone proteins project out as tails.
3. There is additional folding of nucleosomes to form higher order chromatin structure.

Question 11

what is the basic unit of chromatin?

Answer 11

nucleosome

Question 12

how does chromatin regulate gene expression?

Answer 12

nucleosomes over regulatory regions can block access to RNA Polymerase II (PolII), thus silencing gene expression.

ATP dependent chromatin remodeling enzymes like HATs unwind DNA from histone proteins and thus alter the positioning of nucleosomes

this allows PolII, transcription factors and other regulatory proteins access to promoters, enhancers, and other regulatory regions to activate gene expression

Question 13

what part of the histone is modified to allow for gene regulation?

Answer 13

the amino terminal ends of histone proteins are commons sites for covalent modifications that include addition of acetyl and methyl groups

these reversible covalent modifications can form a code which determines if a gene is expressed or silent

Question 14

what are writers, readers and erasers?

Answer 14

enzymes that add covalent groups to histones are referred to as “writers”

enzymes that remove covalent groups are “erasers”

proteins that recognize specific modifications are referred to as “readers” of the code

writers, erasers, and readers can be targeted with drugs that inhibit their activity in cancer

Question 15

what does histone acetylation do?

Answer 15

acetylation on various amino acid residues within histone H3 and histone H4 enhances gene expression

it neutralizes the positive charge on histone proteins thus loosening DNA-histone contacts

Question 16

what are HATs?

Answer 16

Histone acetylation is catalyzed by histone acetyl transferases (HATS) which are referred to as “writers”

Question 17

what are bromodomains?

Answer 17

an acetyl group being added to a histone also attracts other regulatory proteins with bromodomains

bromodomain (BRD) containing proteins are called “Readers” because they recognize the acetylation in order to facilitate downstream events that lead to gene activation

Question 18

what are the two groups of HDACs?

Answer 18

1. classical HDACs

2. Sirtuins

Question 19

what are classical HDACs?

Answer 19

zinc dependent

HDAC 1-8

Question 20

what are sirtuins?

Answer 20

HDACs that are NAD+ dependent

Sirt 1-7

Question 21

what do HDAC inhibitors do?

Answer 21

HDAC inhibitors block HDAC activity and can restore expression of silenced tumor suppressor genes by promoting activation of transcription

depending on the cancer type, treatment with HDAC inhibitors can curb cancer cell proliferation, induce apoptosis, or promote cellular differentiation

however, a limitation of these drugs is that they often target multiple HDACs and have unwanted side effects

Question 22

what drug is an HDAC inhibitor?

Answer 22

vorinostat

it was the first HDAC inhibitor to be approved for treatment of cutaneous T cell lymphoma

Question 23

what are histone methyl transferases?

Answer 23

HMT = writers
HDM = erasers

enzymes that add methyl groups to lysines or arginines on histone proteins using S-adenosyl-L-methionine

histone demethylases remove methyl groups

Question 24

what does histone methylation do?

Answer 24

histone methylation can activate or repress gene expression depending on the amino acid being modified

enzymes that add or remove methyl groups are specific for particular amino acids on histones H3 or H4

Question 25

what are the readers of histone methylation?

Answer 25

readers of histone methylation are proteins with chromodomains, tudor domains, PHD, or MBT which are all methyl-binding domains that recognize specific methylations and can activate or silence gene expression