Lecture 12 - New targets in Chemo Flashcards
Fundamental principles of the outcomes of DNA damaging therapies
- If you can recover the damage by a repair mechanism from the drug then you have failed
- If you can inhibit the repair mechanisms you can get a better result
Example of a drug & target where there is direct inhibition of a repair enzyme
Enzyme targeted: MGMT
Drug: O6-Benzylguanine, patrin
Examples of targets of inhibitors of DNA control systems
- PARP-1
- Checkpoint kinase
- DNA dependent protein kinases
Example of PARP-1 inhibitors
- Olaparib
- rucaparib
- veliparib
- niraparib
Where do alkylating agents preferentially alkylate in DNA?
N7 position
What is another site which DNA alkylators target in DNA, other than the most common one?
O6-alkyl guanine
What removes O6-alkyl guanine in DNA?
MGMT enzyme
How can resistance to O6-alkylators be overcome and drugs for this?
- By MGMT depletion
- Thus making the DNA more susceptible to damage from the O6 lesions
- O6-Benzylguanine and Partin in clinical trials
What is the issue with depleting MGMT?
- Can cause myelosuppression
- lower doses of alkylating agents would be required
What is PAR?
The third nucleic acid in mammalian cell
- Poly(ADP-ribose)
Features of PAR
- Polyanionic polymer
- linear or branched
- built from ADP-ribose units derived from NAD+
- Usually build onto Glu side chains in target proteins
What substrate do PARPs use?
NAD+
What is PARP-1?
- Poly(ADP-ribose)polymerase-1
- Found in nuclei
- essential for initiating DNA repair
What is the reaction of PARP-1?
- PARP-1 uses NAD+
- Forms intermediate, then lone pair of electron on Oxygen forms a double bond
- Break the carbon bond
- Oxygen quenched by reaction with an alcohol
- Forms polymer chain
What is the structure of PARP-1?
N terminal –> C terminal
- DNA binding domain with 2 ZNF
- Auto modification domain containing GLUTAMATE with Nucleuar localisation signal and caspase-3 cleavage site
- NAD+ binding domain
Where does polymerisation occur in PARP-1?
at NAD+ binding domain
What role does PARP-1 play?
Repairing damaged DNA
Steps in repair of DNA by PARP-1
- SSB
- PARP-1 recruited to damage, forms PAR chain on histone near damage
- Histone removed from DNA
- Repair enzymes bought to damage
- PARP-1 adds PAR chains to itself, removal of PARP-1
- PARG snips PAR chains off PARP-1 and histone
- return back to repaired DNA with histone and PARP-1 back in place
- Inhibition of PARP-1 = no base repair
What is the cancer therapeutic application of inhibition of PARP-1?
- Radiotherapy, cytotoxic electrophilic drugs and inhibitors of topisomerase II cause DNA SSB
- If you can repair via PARP-1 then cells survive
- Inhibit PARP-1 to inhibit repair and potentiate these therapies
What is the other therapeutic applications of inhibition of PARP-1?
- Reduction of organ damage following reperfusion injury
Regarding drug design of inhibitors of PARP, what important obstacle was discovered which prompted new drug design of PARP-1 inhibitors
Full analogues of NAD+ such as TAD do not inhibit PARP activity as they are inacitve
When designing a drug to inhibit PARP-1, what features of simple analogues DONT make a difference when wanting to increase potency of the inhibitor?
- Polarity of 3-subsituent
- Hydrogen bonding
- Size
When designing a drug to inhibit PARP-1, what features of simple analogues are favoured to increase potency of the inhibitor?
- electron neutral or electron donating group in the ring (pushes the electron density towards)
- Side chain on 3rd position of ring (not 2nd or 4th)
- Amide group > sulphur group as side chain
- Control of steric clashes/conformation by building another ring/benzene ring to lock amide in place
- A bulkier subsitent
When designing a drug to inhibit PARP-1, what features of simple analogues are disfavoured to increase potency of the inhibitor?
- Electron withdrawing groups which pull electron denisity away from the ring (DONT WANT A NITROGEN)
- Side chain on 2,4 position of ring
- Steric clashes
Examples of PARP-1 inhibitors
- Olaparib
- Rucaparib
- Veliparib
- Niraparib
Features of Olaparib
- Inhibits PARP 1,2,3
- Ovarian, fallopian tube and primary peritoneal cancer
Features of Rucaparib
- Inhibits PARP-1, PARP-2, tankyrase-1, tankyrase-2
- For ovarian, fallopian tube, peritoneal cancer AND treatment of patients with BRCA mutation (germline and/or somatic) taking platinum-based chemotherapy
- in combination with temozolomide
Features of Veliparib
- Inhibits PARP-1, PARP-2, tankyrases
- Discontinued in trials
Features of Niraparib
- Inhibits PARP-1, PARP-2, PARP-4, tankyrases
- ovarian, fallopian and peritoneal cancer after response to first line platinum therapy
Overview of achieving synthetic lethality (BRCA-1/2)
- cancer cells have damage repair mechanism
- Damaged base gives a SSB, this can be repaired by BER with PARP-1 playing a role and the cell would survive
- DSB leads to HR and cell survives
- A persistent break would lead to cell death
- Patients can be heterozygous for mutant BRCA1/2, but the tumour could be homozygous
- means tumour might be unable to perform HR on DSB and if you inhibit PARP-1 then you might end up with synthetic lethality, you kill the cells
- PARP-1 as a single agent
What are the typical features of a tumour?
- Poor vascular structure
- Disorganised network
- leakiness
- high internal pressure
What regions are present in tumours?
Oxic region
Hypoxic region
Necrotic region
Function blood vessel
Sensitivity to chemo and radiotherapy of the Hypoxic and Oxic region of tumours
Oxic - sensitive
Hypoxic - insensitive
How can hypoxia be a therapeutic opportunity?
- Radiotherapy requires O2 –> hypoxic tissue less sensitive to radiotherapy
- radiosensitsing drugs & PARP-1 inhibitors –> increase sensitivity
- Can use hypoxia selective drugs –> behave as prodrugs in oxic tissue –> only display toxicity in hypoxic regions
Example of a radiosensitising drug
Etanidazole
How do Oxygen-mimetic radiosensitizers work?
- work in hypoxic cells
- replace oxygen in the chemical reactions that lead to DNA damage
How does Etanidazole work?
- radio sensitising drug
- Reduces glutathione concentration
- inhibits glutathione S-transferase
- tissues become more sensitive to ionising radiation
How do hypoxia selective drugs (prodrugs) work?
- target the hypoxic cells of tumours –> which cause resistance to conventional therapies
- inactive in normal tissues that are well oxygenated, but becomes active at the low oxygen levels
Examples of Hypoxia selective prodrugs
- Mitomycin C
- Nitromidazole
In a hypoxia associated tumour microenvironment, what is the correlation of resistance and oxygen levels?
- Drug resistance increases in a tumour as the oxygen level decreases
Therapeutic approaches targeting telomerase/telomeres
- Inhibition of telomerase activity –-> binding to G-quadruplex
- Inhibition of tankyrase-1 –-> preventing telomerase from binding to telomere
Why is MGMT not considered a proper enzyme?
- not regenerated
- 1:1 stoichiometric
What is the rationale for combination therapy of Temozolomide with pseudo substrates for MGMT?
- TMZ resistance mediated by MGMT protein
- Tumours lacking MGMT activity –> more sensitive to TMZ
Why would we want to target PARP-1 with inhibitors in the context of Reduction of organ damage following ischaemia/reperfusion injury?
- Blood supply to an organ interrupted –> cells hypoxic
- Reperfusion –> rapid resupply of O2 to the hypoxic cells
- O2 –> oxidising diradical –> damages DNA
- PARP-1 over-activated
- Cells depleted of NAD+
- Cells die and organ failure
Methods to overcome hypoxia
- reduce cellular oxygen consumption
- Increase tumour oxygenation
