lecture 10 - antimetabolites

Question 1

what kind of enzymes do antimetabolites inhibit?

Answer 1

critical enzymes involved in DNA biosynthesis

Question 2

What are the four main groups of antimetabolite drugs?

Answer 2

Folate antagonists (e.g. methotrexate, lipophilic antifolates)
Pyrimidine antagonists (e.g. 5-fluorouracil)
Purine antagonists (e.g. 6-mercaptopurine)
Sugar-modified nucleosides (e.g. cytarabine, fludarabine and gemcitabine)

Question 3

What is folic acid required for and what are its derivatives?

Answer 3

Folic acid is required for the synthesis of DNA and RNA, as well as amino acid metabolism
Dihydrofolate (one additional hydrogen) and tetrahydrofolate (two additional hydrogens) - reduced forms

Question 4

What are the steps of the folate cycle?

Answer 4

• There are three enzyme-mediated reactions
• 1) Tetrahydrofolate is converted by serine hydroxymethyltransferase (SHMT) into 5,10-CH2-tetrahydrofolate
• It does this by taking a carbon unit from a serine (which is converted into glycine in the process), and attaching the carbon unit to tetrahydrofolate
• Now formed a new ring in the tetrahydrofolate system, and distorted the shape
• 2) dUMP comes along and thymidylate synthetase (TS) converts dUMP into dTMP – it does this by taking a carbon unit off 5,10-CH2-tetrahydrofolate and adding it to the aromatic group (the uracil), so the uracil is converted to thymine (dTMP)
• At the same time, the 5,10-CH2 loses the carbon unit and is converted into dihydrofolate
• 3) Now need to regenerate tetrahydrofolate. This occurs by dihydrofolate reductase (DHFR) and it also involves NADPH (reducing agent) – it reduces the carbon bond across the carbon and nitrogen imine, and dihydrofolate is formed again

Question 5

How does methotrexate work and what are some issues with it?

Answer 5

• Analogue of dihydrofolate – binds to DHFR at folate-binding site
• Very potent competitive inhibitor of DHFR (Ki = 5 pM (human DHFR))
• Too polar for passive diffusion into cells – taken up through reduced folate carrier (RFC)
• Must be polyglutamylated to be retained in cells
• Often used in high-dose regimen, with leucovorin (folate) rescue of normal cells
• Widely used drug against many cancer types

Question 6

What are some mechanisms of resistance to methotrexate?

Answer 6

• Mutations to DHFR enzyme, modifying folate binding site
• Multi-drug resistance phenotype, causing active efflux of drug
• Mutations to RFC, reducing uptake

Question 7

What are some examples of lipophilic antifolates?

Answer 7

Methylbenzoprim, piritrexim and nolatrexed

Question 8

What are some examples of inhibitors of thymidylate synthetase?

Answer 8

Pemetrexed and raltitrexed are competitive inhibitors

Question 9

What is the mechanism of thymidylate synthetase?

Answer 9

• get an attack by the sulphur molecule in a 1,4 fashion
• The sulphur from the enzyme attacks in this fashion – break this bond and form a new one
  Can exist in two forms – opened the 5-membered ring and presenting an imine
• Intermediate is able to unwind and attacks at the carbon unit – this attacks and creates a ternary complex (3 parts)
• The enzyme itself, and the dUMP inside of it, and the CH2-THF
• Lose a hydrogen (proton)
• Now turned dTMP into dUMP – this will then get converted into tetrahydrofolate
• Important: ternary complex breaking apart is trigged by the loss of a proton (H+)

Question 10

How does azacytidine work?

Answer 10

It is a weak inhibitor of TS - binds competitively
it is phosphorylated to form azacytidine triphosphate, and then incorporated into RNA
It mimics the C in RNA, but is unstable and decomposes, causing damage to RNA
It can inhibit DNA methyltransferases

Question 11

How do purine analogues work as antimetabolites?

Answer 11

Idk :(
look it up later

Question 12

How gemcitabine activated?

Answer 12

deoxycitidine kinase is able to add a phosphate - then UMP adds another phosphate
NDP then finally adds the last phosphate, making it triphosphate