Section 7: Platinum Anti-Cancer Drugs Flashcards
What is cis-platis
(NH3)2-Pt-(Cl)2 First Synthesis-1825 Clical use approved 1980 Very successful drug for testicular and ovarian cancer (>95% survival rate) Head, neck, bladder, cervical cancers Lymphoma and melonoma
What are the loose rules for platinum drug anticancer activity
for PL2X2
1) cis leaving groups
2) electrically neutral complex with Pt(II) or (IV)
3) Leaving groups should be moderately strongly boune eg. anionic. Tightly bound=low activity highly labile=toxic
4) Non leaving group is crucial: amine group with at least one N-H
How is the anti cancer activity usually tested?
-in vitro on mouse leukaemia cell lines
-testing on animals
-several stages of chemical trials
1 in 10,000 screened drugs make in to approved clinical use
What is the active species?
The monoaqua complex
195Pt and 15N nmr studies show this
The positive charge of this complex means it is attracted to the negative surface of the DNA.
What happens time wise once the drug is administered
2-3h delay in sensitization after administration is due to the slow formation of the monoaqua complex
What happens when the drug is injected into the body?
the plasma of the blood has a relatively high concentration of Cl- there is little hydration of the complex
however the complex diffuses (passive diffusion? mechanism not known
tumours have immature cell membranes- permeated by molecules more easily
in the cytoplasm chlorine concentration much less and hydrataion starts occuring and the drug can become active
ligand exchange- energy required- csfe
PICTURE
What are the biological targets of cis-platin
Many targets
-Pt-DNA binding focused on
Because:
-Diseases where DNA processes are deficient are hypersensitive to CP
-correlations have been shown between Pt-DNA adducts in peripheral blood cells and disease response in cis-platin patients.
- treatment of HeLa cells with a high dose of radiolabelled CP shows:
1 Pt per 10^4/10^5 proteins
1Pt per 10-1000 RNA
1Pt per 1 DNA
showing it has the highest affinity for DNA
How does Pt bind to mononucleotides
Picture from slide
Both theoretical and experimental studies show Guanine N7 (the free N in the smaller ring) to be the most electron rich centre
G (N7)> C(N7) and A(N1)
Picture
The Pt-G(N7) bond is very strong and requires a strong nucleophile to break the bond
H bond between O of G and NH of Pt stabilises adduct
adduct can be easily detected in NMR studies increase in delta (7.8 to 8.8) and pt satellites
very distictive peak v useful as well as X-ray crystalloraphy and H NMR NOE
What is the structure of DNA?
5 member ring sugars (deoxyribose) joined by phosphate groups attatched to the sugar are nucleic bases A,C,G,T that strongly h bond with another base on another string of DNA
They form a double helix
bases complementary
w/ minor and major grooves and nezymes fit in these
PICTURE
Pt binding to oligonucleotides
oligonucleotides are short duplexes(2-15) nucleotides long used for model studies
studies found that cis platin almost exclusivelt binds at G(N7)-p-G(N7) to produce a cis guanine-guanine product
found using x-ray crystallography and 1H,1H NOE NMR
G-T-G platinum can still bind
or on different strands G connect
Why does the Pt not bind to the backbone of DNA
Oxygen too hard for Pt, but will still attract it
NOESY spectrum etc
bloop
what is a typical experiment to study where CP binds to actual DNA?
Usually involves
1) in vitro incubation of DNA and cis-platin
2) extraction of DNA from cells
3) digestion of the DNA using enzymes
4) separation (HPLC)
5) characterisation (1H NMR w/ much more analysable “chunks” of DNA
How does PC bind to DNA?
60-65% G,G 1,2-intrastrand 20-25% G,A 1,2-intrastrand 5-10% G,G 1,3 intrastrand or interstrand 2-5% monoadduct (time dependant 1% DNA-protein binding this can lead to side effects and is only observed in whole cell studies
inorganic way of representing G,G intrastrand etc
cis-[Pt(NH3)2(GpG)]
or ApG
(GMP)=guanine monophosphate
1,3 intra and inter =GMP2
what is the conclusion from the DNA studies
1,2-intrastrand crosslinks are important to anti-cancer activity
because
1) they are the major adducts formed when Pt reacts with DNA
2) clinically inactive compounds fail to form these links eg. trans-platin
What is the mechanism of reaction of CP w DNA
PICTURE
Aquation of the CP is the RDS. Reaction of the Pt complex w DNA strand is fast
H-bonding from NH3 and OH2 ligands are possibly important in orientating the Pt complex - speeding up the reaction
Example of the damage to DNA on CP binding
PICTURE
- 2 G groups in Head to head configuration
- Dehedral angle between rings of approx 80 degrees (disruption of base stacking- 0 normally parallel)
- 17 membered ring quite stable
- H bonding from NH3 to 5’-phosphate group
Generalised damage to bigger oligonucleotides on CP binding
Different adducts distort the DNA in diff ways
but the main effects of intrastrand cross-links are:
-Bending towards major groove (which is stabilised by h bonding??)
-Unwinding of duplex (around 20%)
-Minor grove widened
Distortion of watson-crick base pairing
-Dublex destabilisation
lower m.p. and calorimetric experniments
DNA on CP binding effect
The general damage reasons blocks replication and inhibits transcription. Replication stops at site corresponding to one nucleotide before the first Pt-G residue and at positions opposite the two Pt-G residues.
There is also evidence suggesting CP induces cells to undergo apoptesis (programmed cell death)
DNA repair: what is it?
Dna is continually checked for errors in its sequence by various proteins. the proteins activate enzymes to eradicate the errors. Studies show that cells deficient in DNA repair are much more sensitive to CP
How do Pt 1,2 intrastrand cross-links inhibit repair?
As seen in the picture:
Usually DNA can be repaired, the Pt complex would be cut out and the section would be replaced
However, it is hypothesised that this does not work as a High Mobility Group (HMG0 protein) binds to the 1,2 intrastrand complex, shielding the repair and leading to apoptosis (cell death)
Binding occurs by HMG inserting a phenyl group protruding from it’s backbone into the notch created when cis-platin complexes to DNA. pi stacking involved too.
The bend in the DNA increases to 60to90 degrees
Biotransformation of CP
Platinum is able to bind to S atoms (strong soft interaction).
Many biomoleculaes contain sulphur centres which are potential coordination sites for platinum
