Medicines Design Flashcards
All but Andy Thompsons
How do Vinca Alkaloids work?
Eg, Vincristine/Vinblastine/Vinorelbine
They bind strongly to tubulin dimers, causing a conformational change and preventing them form binding to the microtubule, and so preventing it from growing
They then condense into paracrystalline aggregates
Vinblastine binds to the (+) microtubule, preventing tubulin dimers from binding also
Using various assays, what was found to be favourable in a PARP-1 inhibitor, and what was found to not matter?
Not Matter –> Polarity of the 3-substituent, H-bonding and size
Favoured –> Electron donating groups, only a bottom substituent on the benzene, presence of an amide and a bulky substituent in the right bottom corner
What is hormone therapy?
Slowing or stopping the production of hormones like progesterone or oestrogen to prevent the growth of hormone-sensitive tumours
So you cant take HRT with a positive tumour as it would ber counteracting the action of the hormone therapy!
Why can inhibiting PARP-1 be useful following ischaic/reperfusion injuries?
Reperfusion after hypoxia will flood the cells with oxygen, which in itself is a diradical and can damage DNA
This causes overactivation of PARP-1, but due to the lack of NAD+ (due to the earlier hypoxia) there is not enough to power the cell and be the substrate for PARP-1…..meaning the enzyme functions but the cell dies due to a lack of energy! This can lead to organ faliure
So inhibiting PARP-1 will ensure that more NAD+ is readily avalaible to the cell
What are the 3 ways that hormone therapy can be used?
Adjuvant (post) treatment –> Prevent reoccureance
In advanced/metastatic breast cancer
As a neoadjuvant (pre) treatment –> To shrink the tumour
How does the Scintillation Counter work?
Beta particles strike the scintillant molecule, which then gives off photons, which can be detected by photomultiplier tubes, which we can use to quantify the amount of radiation that is present
What is positron?
A positively charged electron
When it combines with an electron it produces 2 gamma waves at 180 degree angles from each other
How does Autoradiography work?
Tritium is used to unionise silver, using beta radiation
This will mean that where ionization has occured will show silver spots
How do Bicalutamide and Enzalutamide work?
These are androgen receptors blockers
Bicalutamide –> Binds to the AR, causing internalisation of the complex (but no conformational change), but it cannot get into the nucleus. Has some agonist activity, especially at mutant AR
Enzalutamide –> The same as bicalutamide but has no agonist activity
Name 3 inhibitors of CYP17A1
Why are these good inhibitors?
Ketoconazole –> Non-specific and an antifungal
Abiraterone –> Selectivie inhibitor of CYP17A1
Abiraterone Acetate –> Pro-drug of abiraterone
These are good as they all have a nitrogen with a lone pair of electrons, allowing them to bind to metals (such as the haem needed to bind CYP17A1)
How does PARP-1 help repair DNA?
When DNA damage is detected, Poly(ADPr) binds to PARP-1 and Histone, removing the histone
The removal of the histone allows DNA repair proteins (blue) to be activated
Poly(ADP-Ribose)glycohydrolase (PARG) then removes the Poly(ADPr)s from PARP-1 and Histone, allowing them to return to normal
Why do we want to inhibit PARP-1 when treating cancer?
As it repairs ssDNA!
When giving chemotherapy we want to damage the DNA and cause the cell to die, but PARP-1 will prevent this from occuring
So giving a PARP-1 inhibitor will potentiate chemotherapy when given in combination
Explain the self-potentiation of Gemcitabine
UTP is broken down to dCTP, which inhibits dCK….so low levels of dCTP will activate dCK
dCK inhibits the conversion of Gemcitabine to F2dCMP, which we dont want!
F2dCMP inhibits the conversion of UTP to dCTP, which means that dCK is activated, and so more Gemcitabine can be converted to F2dCMP
How do Colchicine like drugs work?
Eg, Colchicine/Combretastatin A-4
They bind to colchcine binding sites on the beta subunit of tubulin dimers
If they bind to the free tubulin dimers then it prevents the microtubules from growing
If they bind to the microtubules, then it prevents them from breaking down
How does PARP-1 bind using zinc fingers?
And where would the inhibitor Olaparib bind?
The Zn2+ binds to the thiols of 3 Cys residues, and a His imidazole binds to the DNA-binding domain
Olaparib binds to the NAD+ binding domain (at the C-terminus)
How does gamma imaging and positron emmision tomography work?
Gamma Imaging –> Tc is put into the brain and then detected
- Due to its short life it is made on site from Molybdenum
PET –> 18F is used as a glucose mimic, which is taken up into the body and the gamma radiation thay is produced is detected
What is the main problem with inhibiting PARP-1?
They all function by blocking its substrate (NAD+)….but NAD+ is used everywhere! So there are so many side effects across the body
How can we block ovarian function?
Remove the ovaries (oophorectomy) or by using radiation….this is permanent
Use of GnRH/LH-RH agonists such as Goserelin (Zoladex) to supprese ovarian function
Why are microtubules a good cancer target?
As they are resposnible for maintaining structure of the cell and seperating chromosomes during mitosis (cell division)….so interfering with them will inhibit mitosis and so kill the cell
How does 6-MP and 6-TG work?
They are mimics of Hypoxanthine and Guanine respectively, but with a sulfur instead of a carbonyl (as part of the amide group)
This means that the same pathway can occur, but not fully correctly
Why are SSRIs problematic for people taking tamoxifen?
SSRIs inhibit CYP2D6, which is a vital enzyme for breaking down tamoxifen into more active metabolites
What are the 3 regions of PARP-1?
DNA Binding Domain –> Contains 2 zinc fingers that bind to the DNA tightly
Auto-modification centre –> Where it binds to glutamic acid
NAD+ Binding Domain
Explain alpha, beta and gamma decay
Alpha –> Slow moving helium nucleus (very efficient ionisation)
Beta –> High speed electron (also releases gamma radiation)
Gamma –> A photon (inefficient ionisation)
What are antimetabolite drugs?
And what are the 4 classes of drugs?
Drugs that kill cancer cells by inhibiting critical enzymes that are involved in the biosynthesis of DNA
Folate Antagonists –> Methotrexate
Pyrimidine Antagonists –> 5-FU
Pure Antagnoists –> 6-Mercaptopurine
Sugar Modified Nucleosides –> Cytarabine
What is triple negative breast cancer?
And what drug cant be used in this case?
When the tumour is ER -ve, PR -ve and HER2 -ve
Cannot take Herceptin (Trastuzumab) as no HER2 receptors
How do we block oestrogen production?
Using drugs called aromatase inhibitors to block the enzyme aromastase which produces oestrogen in the ovaries
Can only be used in post-menopausal women as pre-menopausal women create too much aromastase to inhibit
Examples –> Anastrozole (temproary), Letrozole (temporary), and Exemestane (permanent)
How does 5-FU inhibit TS?
It’s metabolised to dUMP, but with a F present
The F cannot be removed (unlike the H that is normally present) due to its large electronegative charge (so cant be F+) and so the pathway stops dead, so dTMP and DHF cant be made
What was the major problem with Iniparib?
It was a non-specific thiol-reacting compound, ejecting Zn2+ from PARP-1
So it reacted with anything that had zinc fingers
Why can Pemetrexed and Raltitrexed inhibit TS?
They are analogues of 5,10-CH2-tetrahydrofolate but without the CH2, meaning it can bind to the tetrahydrofolate binding site (as a competitive inhibitor), and inhibit TS
How do Taxols work?
Eg, Paclitaxel/Docetaxel (Taxotere)
Bind to taxol binding sites inside of the microtubules, preventing the breakdown of the microtubules by stabilising it
This means that there is less tubulin dimers present, so less microtubules can be built
What is a gatekeeper mutation?
And what is the one in EGFR?
A mutation that is caused by a selection pressure (usually by a drug) that causes the drug to not bind anymore
In EGFR the mutation T790M occurs, where the Met resiude is much bigger, causing a steric clash that means Gefitnib cant bind
However ATP can still bind to the kinase after the T790M, so cell proliferation can still carry on
In what situations can we image the inactive conformation of EGFR via crystallisation?
In the presence of the inhibitor Lapatinib
In the presence of the V948R mutation
How do Sugar-Modified Nucleotides work?
They inhibit DNA polymerases by being analogues
How does testosterone cause cell proliferation?
It binds to the androgen receptor, which causes it to have a conformational change
It then translocates to the nucleus, where it signals for proliferation
How does a Geiger-Muller Detector work?
And which region is radiation detectable?
Beta ionization creates ion pairs, which split and move towards the anode/cathode, which creates something that’s detectable
We use the Geiger Region as it is the only place where we cannot discriminate between the type of radiation –> The avalanche cascade occurs consistently in this region
Do analogues of NAD+ inhibit PARP-1?
No!
What type of DNA breaks can PARP-1 fix?
And which ones can BRCA1/2 fix?
PARP-1 –> Single stranded only
BRCA1/2 –> Double stranded only
How does Azacytidine work?
A weak inhibitor of TS and an inhibitor of DNA methyltransferases
A mimic of cytidine
It is phosphorylated then incorporated into RNA, causing it to become unstable and degrade
Why is Gefetinib more responsive in those with the L858R mutation when compared to the wild-type?
As it binds more tighty to the mutant version
This is because it preferentially binds to the active conformation (which the mutation pushes the protein towards)
How does Tiazofurin work?
And is it approved for treatment?
Forms TAD, which is a mimic of NAD+, and so can bind to its binding sites
It therefore prevents the conversion of IMP –> XMP, by inhibiting IMP dehydrogenase, and so prevents the formation of guanosine monophosphate (GMP)
NO –> It is an expermental drug
How does testosterone bind to the AR naturally? (in terms of chemistry)
2 ends are hydrophillic and so can H-bond via its carbonyl (Arg752) and alcohol (Thr877) group
The central core is hydrophobic and so can bind via hydrophobic interactions to the pocket (Phe764/Met787/Leu704)
How can we block oestrogens effects?
By using SERMs (Selective Oestrogen Receptor Modulators) like tamoxifen or toremifene
These work only of HR+ breast cancers
Tamoxifen has partial agonist activity in other tissues than breast, whereas Fulvestrant has none, and so is purely antioestrogen
Why do Nolatrexed, Piritrexim, Methylbenzoprim and Pyrimethamine not need RFCs to be uptaken into cells to function?
As they are more lipophillic, so can get in via passive diffusion
Why is Thymidylate Synthase (TS) vital in DNA biosynthesis?
As it converts dUMP to dTMP, which is tyrosine containing which is vital in DNA
dUMP is used in RNA, whereas dTMP is for DNA
L-Serine is also vital as it contains the CH2 that is needed to form dTMP
What causes prostatic cancer to become androgen independent?
Mutation of the Androgen Receptor gene
What is the importance of Haem in the production of testosterone?
Fe acts as an oxidising agent, so when in close proximity to progesterone (or other precursers) it can allow oxidation (using the CYP17A1 enzmye) to the next step
The enzyme also binds in part via Arg
How does Bicalutamide bind chemically to the AR?
And how does this differ from testosterone?
Contains a Fluorophenyl group that binds to a seperate hydrophobic side pocket which stabilises the drug more, meaning it is a better competitor inhibitor
Binds through Asn705 instead of Thr877
What enzyme is vital in all stages of testosterone biosynthesis?
CYP17A1
So blocking this will prevent testosterone production, which can be beneficial in androgen sensitive tumours
What is PAR and PARP-1?
PAR –> Poly(ADP-Ribose), which is the third nucleic acid after DNA and RNA
Usually builds onto glutamic acid side chains, being either branched or linear
PARP-1 –> PAR Polymerase-1, an enzmye in the nuclei that has the substrate of NAD+ that is used to fix DNA damage
What is the most common intracellular kinase domain mutation in EGFR?
And what effect does this have on the proteins conformation?
L858R
This makes the inactive protein less stable
The aC helix will move in, and the activation loop isnt created (these are present in the inactive conformation which stabilises it)
The Leu is hydrophillic and so binds to the solvent, but when mutated to Arg….it wont bind, and so pushes the confrmation towards active…which will cause cell growth
How does Methotrexate work?
And what are the resistance mechansims?
Binds to DHFR (by being a analogue of dihydrofolate), preventing the formation of tetrahydrofolate
Needs to be taken up via Reduced Folate Carriers (RFCs) and polyglutamated in the cell to be retained
Resistance –> Mutations to DHFR, Efflux of the drug and mutations to RFCs causing less uptake
What are the following components of this cell during early metaphase?
How does NAD+ (and therefore most inhibitors) bind to PARP-1?
Ser904/Gly863 –> H-bond to the amide
Tyr907 –> Binds via pie-interactions
What PARP inhibitors are approved for BRCA resistant ovarian cancer?
Olaparib
Rucaparib
What PARP inhibitors can bind via ionic interactions between glutamic acid?
Rucaparib
Veliparib
What PARP inhibitor can bind via ionic intercations between Aspartic Acid?
Niraparib
