Note for: PARP and PARG inhibitors in cancer treatment

Note for: PARP and PARG inhibitors in cancer treatment

Doi: 10.1101/gad.334516.119

-        Chemotherapy and radiotherapy are nonselectively means of killing cancer cells.

-        PARP is the successful drug targeting DNA damage response

-        It was first approved for treating cancer with BRCA mutated ovarian and breast cancer – using synthetic lethality approach

-        PAPR inhibitor destabilize replication fork

o   Trap PARP to DNA

o   Cell death through replication stress-induced mitotic catastrophe

-        Inhibition of PARG – exacerbate replication deficiencies of cancer cells

-        Highlight 4 PARP inhibitors used in cancer therapy

o   Olaparib

o   Rucaparib

o   Niraparib

o   Talazoparib

-        Cancer cells produce many ROS, thus, having higher oxidative stress over than normal cells

-        Inflammatory cells – macrophages/neutrophil -- > release ROS

-        PARPi exploits the defective in HR but PARGi exploits the defective in replication machinery

-        PARP

o   Catalyzing the addition of poly(ADP-ribose) from NAD+ to protein and release nicotinamide and protein-mono/poly-ADP-ribose as products

o   PARP1

§  major cellular PAR

§  activated through DNA-binding (binding to DNA causes the conformational change which the loop is opened for NAD+)

§  Step by step procedures

·        N-terminal bind to DNA through Zn finger domain

·        Induce conformational change – unfolding the helical domain (HD)

·        NAD+ incorporate into the catalytic pocket

·        Catalysis occur through three key AA

§  Inhibitor

·        1963 – enzyme was discovered

·        1971 – nicotinamide itself

·        1980 – 3-aminobenzamide (3AB)

·        Key pharmacophores – nicotinamide/benzamide – compete with NAD

·        Interaction involved;

o   Hydrogen bonds with Gly, Ser, Glu

o   Hydrophobic interaction with two Tyr in NAD binding pocket

o   Catalytic AA are conserved among PARP but the AAs nearby are not, thus, drug design should take this information into consideration

·        New generation of PARP1 inhibitors

o   Phthalazinone and tetrahyydropyridophthalazinone

§  Scaffold for

·        Olaparib

·        Talazoparib

o   Benzimidazole and indazole carboxamide

§  Scaffold for

·        Veliparib

·        Niraparib

·        Most selective to PARP1/2

o   Veliparib>niraparib (>100 fold selectivity compared with W/O drug)

§  Based on the formation of hydrogen bond interaction with regulatory subdomain residue (D766, PARP1)

§  D766 conserved in PARP1/2 not other PARP

o   Olaparib and talazoparib

§  15- to 20-  fold selectivity

o   Rucaparib least selective

o   Off-target report for

§  Rucaparib

·        H6PD (hexose-6-phosphate)

§  Niraparib

·        DCK (deoxycytidine)

-        Catalytic inhibitory effects of clinically relevant PARP inhibitors olaparib, rucaparib, niraparib, and talazoparib are comparable

-        Thiers potency in trapping PARP-DNA complexes are different

-        Thus, the trapping was proposed to relying on allosteric changes in PARP1 DNA-binding domain upon inhibitor binding

-        Highest trapping efficiency

o   Talazoparib>>niraparib>olaparib=rucaparib>>veliparib

o   Velaparib is one of the weakest PAPR1/ inhibitor

-         

      PARG

-        Hydrolyzes ribose-ribose bonds within PAR

-        Has macro domain that binds ADP-ribose moiety and PARG-specific loop

Cellular mechanisms of PARP and PARG inhibitors (focus PARP1 only)

Functions in DNA repair and replication fork protection

-        DNA repair, folk protection, and transcription regulation

-        PARP1 relates in many DNA repair pathways

o   SSB

o   NER

o   Alt-NHEJ

o   HR

-        PARP1

o   Under replication stress (stalled replication fork – can be induced by Misincorporation of ribonucleotides, Unusual DNA structures, Conflicts between replication and transcription, Insufficiency of essential replication factors, Common fragile sites, Overexpression or constitutive activation of oncogenes, Chromatin inaccessibility)

§  Preserving or replication forks

-        PARP1-- > SSB

o   Interacting with scaffold XRCC1 to SSB

-        PARP1 -- > response to replication stress

o   Interact with replication machinery

o   Active during S-phase

o   Slow down replication forks to promote fork reversal by antagonizing REQ1 helicase

Role of PARP1

PARP1 -- > transcription regulation

-        Stimulating activity of transcription factor E2F1 – regulate the expression of replication and HR genes

PARP1 -- > HR

-        Promote rapid recruitment of MRE11, EXO1, BRCA1+2 to DNA damage sites

-        Counteract NHEJ – preventing the binding of KU to DNA end

Actually, other members of the PARP family -- > implicated in DSB repair and replication fork stability -- > PARP2, PARP3, PARP10 and PARP14

 

Genotoxic agents used routinely in cancer therapy -- > induce high level of DNA damage -- > causing cancer cell to undergo cell-death because its high proliferative rate (more and more damage acculation)

Synergistic effects are additionally achieved in patients with genetic defect in DNA repair pathways ex.

-        Pt drugs – improve response rate in BRCA mutated TNBC

-        Genotoxic agents has side effects like myelosuppression

PARP inhibitor is the successful example of targeted therapy

-        Using synthetic lethality concept – targeting the compensatory pw which cancer relies on

-        Compensatory -- > HR defect/replicative stress defect (genes cause replicative stress)

Not only BRCA1/2 defective – other genes on DNA damage response (ATM, PRKDC, ATR, RPA1, DSS1, NBN, RAD51, RAD54, CHEK1, CHEK2, FANC, ERCC1, POLB, FEN1, and CDK12) – shown synthetic lethality in combination with PARP inhibitors

Amplifying genomic instability with PARP and PARG inhibitors (focus on PARP1 only)

-        Depletion of PARP1 (under replication stress situation)

o   Increase replication fork speed

o   Impairs replication fork reversal

o   Cause untimely fork restart

o   Consequence -- >

§  Accumulation of DNA damage in S-phase

§  S-phase stalling

§  G2-delay

§  DSB arise

-        PARP inhibitor cytotoxicity – correlate with strength of PARP-DNA entrapment rather than a reduction on PARP1 catalytic activity

-        CSF (common fragile sites) – genome regions which sensitive to impaired fork progression

-        CSF – late replicating and remain underreplicated at G2/M transition

-        PARP inhibitor efficient in

o   HR-defective cells

o   HR-proficient but defect in oxidative stress system/replication stress

o   PARP-inhibitor efficacy correlate wit basal levels of replication stress in cancer cells

-        PARP1 also modulates the activity of different transcription regulators implicated in cancer or inflammation.

Determinants of PARP inhibitor sensitivity in cancer cells

-        PARP1 inhibitor is very effective with cell which defective in BRCA1/2 or cell with phenocopies BRCA1/2 (BRCAness)

-        Biomarkers used to identify the patients

o   HR deficiency (mutation in DNA repair genes)

o   DNA repair expression levels

o   Replication stress markers

o   Transcriptome profiles (to check the PARP inhibitor response)

-        Mutational signature is used for preliminary screening for PARP1- receiver

o   Defective in BRCA1/2 caused special feature in genetic defects (due to the repair through NHEJ as backup pathway)

o   This cannot be used in case of HR restoration

-        Genomic signature for HR deficiency

o   LOH

o   Telomeric allelic imbalances (TAI)

o   Large scale state transition (LSTs)

-        HRDetect and HRD scores (at DNA level) -- > used to predict PAPR inhibitor sensitivity in clinical setting

-        Gene expression profile of BRCA mutated ovarian cancers could be also used as the predictor of PARP sensitivity

-        Cytological signature of HR deficiency – number of RAD51 foci (reduced in Rad51 foci – markers for PARP sensitivity)

Replication stress markers

-        Loss of TP53+RB1+amplification of MYC -- > replication stress -- > sensitize small cell lung cancers

-        Schlafen – marker for the replication stress

Clinically recommended does and severity of side effects correlate with PARP inhibitor trapping potency

Olaparib – approve 2017 – maintenance therapy in Pt-sensitive high-grade ovarian cancer patients irrespective of BRCA status

Rucaparib – approve 2016 – advanced ovarian cancer with germline and somatic mutation of BRCA1/2

Niraparib and rucaparib – approve 2017, 2018 – maintenance treatment of recurrent, epithelial ovarian fallopian tube, primary peritoneal cancer irrespective of BRCA status

Talazoparib – approve 2018 – BRCA mutated and HER2-negative breast cancer

No PARG inhibitor reached clinical trials – low metabolic stability

Resistant mechanism (anything that causes PARP dissociation easily -- > resistant to PARP)

-        Restoration of HR pathway

o   Secondary mutation caused the restoration of HR

o   Changing in gene expression levels in HR

-        Inactivation of different NHEJ-promoting factors that inhibit DNA end resection can restore HR in BRCA1-deficienct cells

-        Stabilization of replication forks in BRCA1/2-deficienct cells

-        PARP1 mutation – R591C -- > WGR domain -- > critical for interdomain communication between WGR and DNA-binding domain -- > it dissociates very quick

-        Drug efflux

o   Overexpression of ATP-binding cassette (ABC) -- > drug transporters -- > associate with drug resistant

o   Olaparib and rucaparib

§  Substrate of ATP-dependent drug efflux P-glycoprotein (P-gp) pump (MDR1)

§  Long term treat -- > increase in MDR1 -- > intracellular PARP inhibitor is reduced

PARP inhibitor combination therapy

-        Combination of PARP inhibitors with cell cycle check-point inhibitors -- > promising strategy -- > fight against PARP inhibitor resistance caused by replication fork stabilization

-        Inhibition of S/G2 checkpoint kinases -- > ATR and CHK1 -- > unscheduled replication origin firing, exhausts nuclear RPA pools (excess ssDNA), depletes dNTP -- > underreplicated DNA + unrepaired DNA damage entering mitotic -- > cell death by mitotic catastrophe

-        Inhibition of BRD4 (global transcription regulator) – synthetic lethal with PARP inhibitors

o   BRD4 inhibition -- > induces HR deficiency -- > lowering the expression of CTIP, BRCA1 and RAD51

-        Histone deacetylase (HDAC – epigenetic modifiers) – synergize with PARP inhibitors

-        Agents that pharmacologically induce HR deficiency like androgen receptor inhibitors, PI3K-AKT-mTOR inhibitors, and antiangiogenic agents

-        Combination with immune checkpoint inhibitors -- > DNA damage caused by PARP inhibitor -- > activate cGAS-STING pathway -- > trigger expression of immune checkpoint in cancer cells (PD1) -- > administration of anti-PD-L1 -- > helping T-cell to proliferate and release cytokine -- > kill cancer cells

Conclusion

-        Cancer cell characteristic

o   High level of replication stress

§  Destability of replication folk

o   Harbor germline or somatic mutation in DNA damage response genes

§  Mutation in HR – prevent restoration of fork stability

-        PARP and PARG inhibitors

o   Exploit these underlying mechanisms

§  Destabilizing replication folks

§  Inducing DNA damage

-        Synthetic lethality for PARP

o   Besides BRCA1/2 extent to HR deficiency genes

o   Replication stress

-        Future research on PARP and PARG

o   Dynamic between these two proteins in regulating replication folk stability

o   How replication stress and genomic instability -- > induce mitotic defect and cell death through replication and mitotic catastrophe

o   Looking for the marker to indicate PARP sensitivity