Targeting Cell Cycle Checkpoint Defects Flashcards
Name 3 features of Cell cycle
1) Directional - Distinct Cyclin-CDK complexes act during distinct phases of the cell cycle ensuring 1-way directionality 2) Robust 3) Highly accurate
Basic mechanism regulating Cyclin-CDK (Cell-Cycle Checkpoint concept)
Signal can either be activatory or inhibitory which results in a STOP signal.
This STOP signal then regulates Cyclin-CDK function
Name the features of the G1 (R) Checkpoint
- Growth Factor/Nutrients
- Cell-Cell Contact
- Morphogens
- Size
- DNA Integrity
What does RB checkpoint sense for?
- Cell cycle checkpoint regulator
- tumor suppressor protein
- Deleterious mutation (Inactivation results in cancer)
Types of cancer implicated in:
Eye cancer
Sarcoma
Lung cancer
breast cancer
glioblastoma
urinary tract cancer
What part of the cell cycle does RB control
RB, product of tumor suppressor gene, controls cell cycle progression from G1 to S phase.
How is RB regulated on a basic level?
Cyclin E-CDK2 controlled by the RB checkpoint
Transcriptional target of E2F
- E2F required to drive transcription of Cyclin E-CDK2 BUT this is -vely regulated by RB protein, preventing passage into the next cell cycle
- RB is under control of CDK4/6 (Cyclin D1-D3) which inactivate RB by phosphorylation
Name ways in which Cyclin D can be controlled by Oncogenic signalling
EGFR - Colon, Lung, Glioblastoma
RAS, RAF - 30% of all cancers
TcF/LeF, ß-Cat, Wnts - Breast, Colorectal, Melanoma, Prostate, Lung, Ovarian, Oesophageal
NF-KB
Cytokine R
How do signalling pathways affect Cyclin-D levels
Cyclin D levels can be controlled by proto-oncogenic signalling
Consequence = Singal independant activation of CDK4/6
Explain the mutual exclusivity concept with regards to Cyclin D
1) Signalling pathways which converge onto Cyclin D-CDK4/6 can have a pro-oncogenic function (FOS-JUN, AP1, EGFR, RAS/RAF, TcF-LeF-ß Catenin-Wnts, NF-KB)
2) CDK4/6 can also be directly mutated
Idea of mutual exlusivity is that if an RB mutation exists (GOF/LOF) then there is usually no GOF/LOF of other pathways in the cell (CDK4/6)
RB functionality is so critical to tumor function - if RB loss then the the tumor does not need other GOF/LOF of G1 checkpoint components
Name the G1 checkpoint components
1) Cyclin D - CDK4/6
2) Retinoblastoma Tumor Suppressor (RB1)
3) CNDN2/INK4: Inhibitors of CDK4/6
4) CDKN1/CIP-KIP: Inhibitors of CDK
- 2,3,4 control the cell cycle transit by preventing CDK2 activation
RB loss or INK4a loss - what is worse?
- INK4A loss and RB loss both occur in the context of the G1 checkpoint
- However, if you lose one particular side of the inhibitor (e.g: loss of INK4A) then you are still checkpoint sensitive for the signals of the other side
- INK4a loss is not that dramatic
- If you lose RB then you don’t need the kinase in the upper left OR upper right – this is because the only purpose for this kinase is to inactivate the RB
- RB loss is much more dramatic than INK4A loss or CIP/KIP loss
-
- Cancer with RB loss are v aggressive
- Cancers with INK4A loss are less aggressive as they will just result in a an increase Cyclin D1 conc, which then obviously affects the RB protein
- However, RB loss is much worse
Explain the rationale behind INK4a loss and RB loss
Loss of G1 checkpoint is a key mechanism affected in cancer
CDK4/6 complexes are highly druggable targets - they are sufficiently different from other enzymes that we can specifically target this as an enzyme
E.g: Mouse models of CDK4/6 treatment are resistant to ErbB2 Breast cancer
Example of CDK4/6 inhibitors: Ribociclib, Palbociclib
Explain the efficacy curve of Palbociclib and its implication for resistance.
- Palbociclib is a CDK4/6 inhibitor which demonstrates a much longer PFS than Placebo (Palbociclib + Fulvesterant vs Placebo + Fulvestrant)
- However, its outcome survival is just the same
- This is due to primary and secondary resistance emergence:
i) if RB1 mutation exists then CDK4/6i will have no impact as the RB1 mutation will just continue to drive the cancer
ii) Thus the RB1 mutation is selected for and evolves in patients with CDK4/6i
Explanation of CT results:
i) RB1 mutations occurred at the end of treatment in CDK4/6i and Estrogen inhibitor patients
ii) No mutations occurred in Estrogen Ri alone
iii) No difference in freq of mutations between CDK4/6i, Estrogen Ri combination or single Estrogen Ri
What are stromal cues
- TK network involving MET/FAK which permits cancer cell proliferation independant of CDK4/6 catalysis
- They undermine the CDK4/6i repsonse
- Stromal cues confer CDK4/6 independant proliferation capability and CDK4/6i resistance
- Cause degredation of CIP/KIP
- HGF/MET/FAK signalling upregulates CDK2 and degrades CIP/KIP proteins
G2 DNA Integrity Checkpoint Activation
- DNA damage (ds breakages, ss unreplication) is recognised by ATM/ATR kinase
- ATM/ATR has 2 different arms: P53, CHK1/2 (Checkpoint Kinase 1/2)
- CHK1/2 activation: Can phosphorylate P53 resulting in the Downstream activation of CDKN1A/p21CIP proteins (CDK Inhibitors)
- CHK1/2 activation = phosphorylation and degredation of CDC25 phosphatase (CDC25 phosphatase usually v important for CDK activation as it clips the inhibitory phosphate on CDK)
- CDKN1A and p21CIP is inhibiting kinases which have already been activated.
Explain the idea of mutual exclusivity in the G2 DNA integrity checkpoint:
- Checkpoint machinery can be widely mutated as in G1 cancer
- P53 is highly mutated >50%
- CHK1 mutation is not as freq
- Reason for P53>CHK1 mutations freq: P53 mutation probably gains more than the CHK1 mutation as it saves you from the apoptosis
What are the consequences of P53 loss (basic concept of synthetic lethality)
- Losing P53 does not mean that we lose the entire checkpoint
- But there is now critical dependancy on the CHK1/2 kinases
- introduces the idea of synthetic lethality: can we design inhibitors to prevent repair and thus indice apoptosis of the cell?
Explain the idea of Normal tissue in P53 normal cancer (idea behind synthetic lethality)
Explain the idea of synthetic lethality in P53 defective cancer
Rationale:
- idea that if there is a P53 defective cancer, then this checkpoint solely relies on CHK1/2
- If you apply a small molecule inhibitor for CHK1/2 then this means that there is no active checkpoint for the cancerous cell, you cause mitotic catastrophe of the cancer cell - thus, apoptosis of the cancer cell ONLY
- Termed: Synthetic Lethality
- CT: Tumor growth massively inhibited as a result of CHK1/2 inhibitor application.
- EG CHK1/2 inhibitor: SAR-020106
