Drugs That Act On DNA Topoisomerases

Question 1

Describe DNA topoisomerases

Answer 1

Present in all bacterial and eukaryotic cells
Change number of twists in DNA
Untangle DNA molecules
Decatenate

Question 2

What is the unction of topoisomerases

Answer 2

To unwind DNA that has become overwound ahead of the replication fork in order to allow for replication

Question 3

What are the 2 classes of topoisomerase enzymes

Answer 3

Type 1 = cuts 1 strand
Type 2 = both strands cut

Question 4

What is the function of topoisomerase I

Answer 4

DNA replication and txn
Topo I part of txn complexes

K/o in mice = lethal

Question 5

What is the function of topoisomerase II

Answer 5

A: chromsome segregation, resolves intertwined chromosomes at mitosis
B: neuronal/development and txn initiation

Chromatin condensations and decondensation

Question 6

How does topoisomerase II work

Answer 6

Is covalently bound to DNA and breaks both strand
Change in conformation allows 2nd DNA strand to pass through
DNA relegates and topo II leaves

Question 7

Name 2 topoisomerase II poisons

Answer 7

Etoposide
Teniposide

Question 8

Describe the mechanism of action of Etoposide

Answer 8

Forms a cleavable complex w topo II and DNA
Blocks reaction and stops topo II from opening and closing
Doesn’t allow 2nd strand through

Question 9

Why is etoposide a poison rather than an inhibitor

Answer 9

Blocks the reaction instead of interacting directly and inhibiting it

Question 10

How can you detect cleavable complexes formed in cells

Answer 10

Alkaline elation -> TARDIS (trapped in agarose DNA immunostaining)

Question 11

Describe TARDIS

Answer 11

Liv cells embedded in layer of agarose gel
Cells listed to remove proteins leaving only DBNA
Fluorescent antibodies added that bind to topo protein

Question 12

Why is TARDIS useful

Answer 12

Allows to look at the effect of drug on topo
Can check if the addition of a second drug will affect the interaction between drugs and topo

Question 13

Describe anthracyclines

Answer 13

Flat 4 ring structure that can intercalated and distort double helix
Hydroxy radical formed = damage DNA and cell membranes

Question 14

Name to anthracyclines and their use

Answer 14

Doxorubicin: solid tumours e.g breast, bladder etc
Daunorubicin: leukaemia

Question 15

Name a topisomerase I poison

Answer 15

Camptothecin

Question 16

Describe camptothecin

Answer 16

Poorly water soluble
Incredibly potent but too toxic
-> need to change drug structure to change toxicity profile

Question 17

Name the 2 derivatives of camptothecin

Answer 17

Topotecan
Irinotecan

Overcome the toxicities

Question 18

Describe the metabolism of irinotecan (CPT-11)

Answer 18

Converted to CES1 and CES2 to SN-38
SN-38 primarily metabolised to inactive glucuronide by UGTs
Produces inactive form excreted in bile

Question 19

When would someone require a lower does of irniotecan

Answer 19

When have SNPs in CES1 and CES2 that cuase low amounts of glucuronides and high conc on SN38

Question 20

When does cytotoxicity occur with topoisomerase poisons

Answer 20

Inhibition of enzyme activity and formation of stabilised cleavable complexes

Question 21

What can happen once DNA damage occurs after topoisomerase poisions

Answer 21

Delay of cell cycle progression and repair/ removal of damage
-> cell survives or mitotic catastrophe or apoptosis triggered by damage

Question 22

Why are there severe toxic side effects with topo poisons

Answer 22

Not very specific

Question 23

Why do cytotoxic drugs preferentially attack tumours

Answer 23

Cells more sensitive during DNA replication
Many dividing cells in tumours

Question 24

What resistance mechanisms do topo poisons have

Answer 24

Less access to target
Altered target or less target
Altered response to DNA damage

Question 25

Describe how a cell can reduce access of topo poisons to target

Answer 25

Over expression of export pump PGP that exports a wide range of large, hydrophobic molecules

Question 26

Describe how a cell can alter the target of topo poisons to increase resistance

Answer 26

Reduce levels of topo 1 or topo 2

Question 27

How can tumour cells increase repair or tolerance of damage by topo

Answer 27

Overexpress enzymes that remove covalently bound topo
TPD1/2

Question 28

Describe TPD1

Answer 28

Hydrolyses the bind that links Top1 to a DNA strand break

Question 29

Describe TPD2

Answer 29

Causes dependent NHEJ which protects against Top2 induced DNA breaks

Question 30

What are current developments to improve Topo-targeted therapies

Answer 30

Dual inhibitors
Novel topo poisons
Novel delivery of topo poisons