Mike Threadgill Flashcards
Antimetabolites
Kill (cancer) cells by inhibiting critical cellular processes, especially those involved in growth
Generally inhibit critical enzymes involved in DNA biosynthesis, because DNA biosynthesis is essential for tumour cell proliferation
Main groups of antimetabolite drugs
There are 4:
- Folate “antagonists” e.g. methotrexate, non-classical lipophilic antifolates, pemetrexed, raltitrexed
- Pyrimidine “antagonists” e.g. 5-fluorouracil, fluorodeoxyuridine, azacytidine
- Purine “antagonists” eg. 6-mercaptopurine, thioguanine, tiazofurin
- Sugar-modified nucleosides e.g. cytarabine, fludarabine, gemcitabine
Why “antagonists”?
The drugs are more like inhibitors than classical antagonists
Folate cycle enzymes
Dihydrofolate –> tetrahydrofolate = DHFR (dihydrofolate reductase)
Tetrahydrofolate –> 5,10-CH2-tetrahydrofolate = SHMT (serine hydroxymethyltransferase)
5,10-CH2-tetrahydrofolate –> dihydrofolate = TS (thymidylate synthetase)
Methotrexate
Analogue of dihydrofolate
Binds to DHFR at folate binding site
Very potent competitive inhibitor (Ki = 5 pM for human DHFR)
Too polar for passive diffusion into cells - needs to be taken up via RFC (reduced folate carrier) and must be polyglutamylated for intracellular retention
Widely used against many cancer types, often in combination with leucovorin (folinic acid) to rescue normal cells and reduce toxic effects of methotrexate
Problems associated with methotrexate
Cancer cells can develop several mechanisms of resistance to methotrexate e.g.
Mutations in DHFR that modify the folate binding site (so methotrexate can’t bind, but folate may still be able to bind)
Mutations in RFC therefore reducing the uptake of methotrexate into cancer cells
“Multi-drug resistance” phenotype, where the cancer cell actively pumps the drug out
Non-classical lipophilic antifolates
Also inhibit DHFR
Can enter cells by passive diffusion (RFC not required, circumvents potential resistance)
No glutamate side chain so not polyglutamylated, which means administration via continuous infusion is required due to lack of intracellular retention
Pyrimethamine
Mainly used as antibacterial
Piritrexim
Potent, active in several tumour types
Nolatrexed
Can inhibit DHFR and TS due to pyridine ring
(Non-competitive inhibitor of TS)
Use in liver carcinomas
Methylbenzoprim
Very potent experimental lipophilic inhibitor of DHFR
Ki ~ 10 pM
TS inhibitors
e.g. pemetrexed, raltitrexed
Analogues of the endogenous TS substrate
Competitive inhibitors of TS, bind at the same site as 5,10-CH2-THF
Require RFC for uptake
TS is over expressed in tumour cells so inhibition of TS is an important strategy in the development of chemotherapeutic drugs
TS mechanism
Draw
Reductive methylation, 5,10-CH2-THF serves as both methyl donor and reducing agent
Effect of inhibition of TS
Blocks DNA synthesis, eventually leading to cell death (“thymineless death”)
This reaction is the sole de novo source of thymidine, which is necessary for DNA replication and repair
Raltitrexed
Direct and specific inhibitor of TS
Water-soluble, non-nephrotoxic
Transported into cells by RFC and extensively polyglutamylated by folylpolyglutamate synthase (FPGS)
Polyglutamylation increases raltitrexed inhibitory activity by > 100-fold and is retained within cells for a prolonged period of time
Effective against metastatic colorectal cancer, but usage is now generally restricted to patients intolerant to FUra
Pemetrexed
New-generation antifolate - TS is its primary target by also inhibits other enzymes involved in pyrimidine synthesis e.g. DHFR
Can be taken up by RFC and PCFT (proton-coupled folate transporter), meaning its activity is preserved even if RFC is lost/impaired
Extensive polyglutamylation so persists in cells for a prolonged period of time
Inhibitory effect against TS is insensitive to dUMP accumulation because dUMP and pemetrexed bind to TS at different sites
Pemetrexed + cisplatin = first-line treatment for NSCLC
And shows activity in a number of other tumours
5-fluorouracil metabolism
5-Fu —> FdURD —> FdUMP
Mechanism of TS inhibition by 5-Fu
Fluorine substituent fails to dissociate from the pyrimidine ring, meaning elimination cannot occur to give loss of -S-TS
Therefore TS remains bound to FdUMP and is inhibited and unable to catalyse other processes
5-Fu also has other mechanisms of action e.g. metabolised to FdUTP and FUTP which can be misincorporated into DNA and RNA, respectively
Very insoluble
Major use in colorectal cancer
Azacytidine
Weak TS inhibitor (binds at substrate binding site but doesn’t inhibit mechanism)
Phosphorylated to form azacytidine triphosphate, that is then incorporated into RNA as a cytidine mimetic
However, azacytidine is unstable and decomposes, causing damage to RNA
Also inhibits DNA methyltransferases which has epigenetic effects and affects regulation
Limitations to clinical use of 5-Fu
Drug resistance
Inefficiently converted into FdUMP - some is catabolised to toxic metabolites
Dihydrofolate
Precursor to 5,10-CH2-THF
Required for pyrimidine biosynthesis
Mechanism of resistance to 5-Fu
5-Fu acutely induces TS expression due to inhibition of a negative feedback mechanism whereby ligand-free TS binds to and inhibits the translation of TS mRNA
When TS is bound by FdUMP, it is no longer able to bind to its mRNA and suppress its own translation, resulting in increased TS expression
This acute increase in TS levels would facilitate recovery of enzyme activity
HPRT
Hypoxanthine phosphoribosyltransferase
6-MP –> Thio-IMP
6-TG –> Thio-GMP
(I = inosine)
Metabolism of Thio-GMP
2 phosphate groups added to give Thio-GTP which is incorporated into RNA (TG-RNA)
Can also lose OH in 2 position on ribose to give Thio-dGTP which is incorporated into DNA (TG-DNA)
