PARP inhibitors Flashcards
Explain the rationale behind original cisplatiin molecules in E.coli growth
Filamentous growth due to the action of platinum compounds formed by reaction of the electrodes with ammonium in the culture medium. Filamentous growth - this is what happens when bacteria fail to replicate properly
Explain Connor’s hypothesis
Cisplatin is a molecule wit bi-amine groups, and bi-leaving groups. - Bi-amine groups: Biologically stable, affect the physical properties, confer non-cross resistance with other analogues - Bi-Leaving groups: 2 parts: STABLE (low potency and toxicity); REACTIVE: (higher potency and toxicity). The leaving groups can dissociate from the main molecule which allows nucleophilic reaction with DNA, thus causing DNA cross-links (DNA cross-linking agent)
Describe Carboplatin structure and functionality
- Stable – cis leaving groups
- Good toxicity profile in patients
- At least as active as cisplatin in ovarian cancer
- But: Cross-resistant with cisplatin – no improvement in the spectrum
- BUT: Patients also develop v low platelet counts as a result of treatment with this drug
- Its tetrahedral carbon structure adds spacer regions to the molecule, this allows interaction with water molecules - increased solubility..
Rationale behind the development of carboplatin dosing formula?
Calvert Formula:
- Carboplatin is renally excreted and in some patients this caused huge thrombocytopenia and leukopenia.
- Calvert formula (Total dose=AUC(desired)×(GFR+25)^2) allowed clinicians to work out how much to give to patients to avoid side effects.
- The area under the curve you wanted would take into account for the lack of leukocytes (leucopenia) and lack of blood platelets (thromobocytopenia) – allowed to take account for these factors which subsequently made the drug v widely available for a lot of patients.
Clinical uses of platinum based drugs?
•Cisplatin:
–Lung cancer (+ gemcitabine / pemetrexed)
–Germ Cell Tumours
–Bladder, Head and Neck, (Ovarian)
•Carboplatin:
–Ovarian
–Lung cancer
–Her2 amplified breast cancer breast cancer
•Oxaliplatin:
–Colon cancer
–Pancreatic cancer
Describe Methotrexate and Folate metabolism and its role in cancer
- Patients are given methotrexate as a cancer therapy.
- Methotrexate depletes FH4 (essential for purine and thymidine synthesis)
- Inhibition of DHFR (dihydro folate reductase enzyme) prevents FH4 being made which is crucial to Thymidine AND Purine synthesis.
- Thus, inhibition of DNA synthesis
- BUT: Essential to give Folinic Acid (anti-dote to methotrexate) to ameliorate the toxic effects of methotrexate
- Methotrexate in Child Brain Cancer:
- The first drug to induce remissions
- Median survival increased from a few weeks to a few months
- Survival increased to several years with the addition of other drugs (especially vincristine and 6-mercaptopurine)
- Relapse and death still ocurred due to meningeal leukaemia
- The blood-brain barrier forms a pharmacokinetic sanctuary
- High dose methotrexate + folinic acid rescue allows the brain to be treated
- this allows for high doses of methotrexate to be given and orevent the formation of pharmacokinetic sanctuaries
Development of more specific anti-proliferative folate metabolism drugs?
- PERMATREXED
- Idea of blocking Thymidine synthase directly. More specific anti-proliferative anti-folate effect.
- This still allows the production of purines (DNA, RNA)
- profile of permetrexed:
- Active in a range of solid tumours (lung, breast, colon, mesothelioma, pancreas, cervix, head and neck, gastric, and others)
- Dose-limiting myelosuppression
- Other toxicities: Rash, mucosal, transaminases, asthenia
Sporadic toxicities of Antifolates
•Raltitrexed – reported drug-related deaths
–16/699 (2.2%) in three Phase III Trials†
•Alimta (Permetrexed)
–4% in early Phase II trials without vitamin supplementation§
•Not possible to predict these toxicities on the basis of plasma or red-cell folate levels
- WHY?
- Methionine synthase can take a methyl group from FH4 to convert homocysteine compounds into methionine
- Methionine then used in cellular methylation reactions and recycled back to homocysteine (shuttle reaction)
- If you don’t have enough folate then the MS does not work properly and the levels of homocysteine in the blood go up. This means that there would be greater toxicity of the drug in patients with high levels of homocysteine (low folate)
- But you can ID these patients and give additional vit b12 which means that the drug (pemotrexate) is now well tolerated
Major mechanisms of DNA repair
Catalytic activity of PARP
- PARP uses NAD as a substrate and makes Poly-ADP Ribose chains attached to the histone proteins and themselves
Mechanism of PARP in BER
- PARP is activated by breaks in the DNA strands
- Makes polymers which are then incorporated into the histone structures which maintain the structure of the DNA
- ADP ribose polymer is –vely charged and so is the DNA, the histone proteins are forced away from the DNA backbone; this allows specific enzymes to repair the ss break
- If you inhibit PARP, you inhibit the repair of DNA breaks; concept of SYNTHETIC LETHALITY
Is PARP a truly cancer specific target?
- Whenever we talk about PARP, we talk about PARP-1 (there are 17 isoforms)
- Present in high activity in most tissues
- Activated by DNA strand breaks and involved with single-strand break repair
–“Housekeeping” function
- Utilises NAD as a substrate to form APD-ribose polymers on histone proteins and itself (automodification)
- Involved in numerous other processes
–Epigenetic regulation of chromatin structure and gene expression
–Interacts with transcription factors and co-factors (NF)-kB, PAX6, AP-2, b-Myb, TEF1
–Interacts with kinetochore proteins
Structure of PARPi
- All contain an active amide group which is the active part of the drug
Applications of PARPi
•Treatment of BRCA1 or BRCA2 related cancers or tumours with a homologous recombination (HR) repair deficient phenotype.
- BRCA1/2 related cancers are extremely sensitive to PARPi. TI is ~250-1000; most cancer drugs are 1:1.
•To potentiate chemotherapy, especially
–Monomethylating agents (eg temozolomide)
–Topo-isomerase I inhibitors (eg toptecan / irinotecan)
•Radiopotentiation
BRCA1/2 Cancer predisposition genes are v sensitive to PARPi. Why?
- Mutation carriers (BRCA 1/2 (+/-), (-/-)) are predisposed to breast, ovarian, prostate, pancreatic and other cancers
- BRCA1 and BRCA2 are involved in HR repair – error-free repair of double strand breaks.
- Carriers have one allele carrying a mutant, non-functioning gene. Damage to the functioning copy results in error-prone DNA repair and is oncogenic. Patients with this dysfuncitonality will have to repair DNA by NHEJ (Protein Kinase (Ku proteins)). This is v error prone.
Propose the mechanism for PARPi synthetic lethality
- IN PATIENTS with defective HR
- Normal ssDNA break and functional PARP then repair just occurs normally. NHEJ is the mechanism for repair (error-prone, oncogenic)
- PARP inhibitor treatment induces phosphorylation of DNA-dependent protein kinase substrates and stimulates error-prone nonhomologous end joining (NHEJ) selectively in HR-deficient cells
- If there is a PARP inhibitor, it means that PARP repair cannot occur. When the DNA damage gets to the replication fork, then it cannot be fixed. Usually error-free HR would just take care of this. But, BRCA patients have dysfunctional HR and so therefore cannot repair themselves. The cell has a crisis moment and will die.
- loss of PARP1 activity is thought to result in accumulation of DNA single-strand breaks (SSBs), which are subsequently converted to DNA double-strand breaks (DSBs) by the cellular replication and/or transcription machinery. These DSBs, which are repaired by HR in BRCA-positive cells, are presumed to accumulate in BRCA1- or BRCA2-deficient cells, leading to subsequent cell death.
- PARP inhibition preferentially enhances error-prone NHEJ activity in HR-deficient cells, these errors are then killed by the cell when the damage gets to the replication fork
- This idea is termed SYNTHETIC LETHALITY
Proposed mechanism of PARPi and its Therapeutic potential
- Endogenously formed SSB are normally repaired by PARP-dependent BER.
- If PARP is inhibited SSB persist.
- SSB form DSB at replication, which are repaired by HR.
- If HR is defective the breaks are not repaired and the cell dies.
- This is the first exploitation of synthetic lethality in cancer therapy.
- Tumour selective.
Discuss the PARPi drugs
- a class of drugs where the indication is determined by the the presence of a deficiency in HR repair, rather than by the tissue of origin of the cancer.
