Dissecting the molecular determinants of clinical PARP1 inhibitor selectivity for tankyrase1

Poly ADP ribosyltransferases play a critical role in DNA repair and cell death, and PARP1 is a particularly important therapeutic target for the treatment of breast cancer due to its synthetic lethal relationship with BRCA1/2. Numerous PARP1 inhibitors have been developed, and their efficacy in cancer treatment is attributed to both the inhibition of enzymatic activity and their ability to trap PARP1 on to the damaged DNA, which is cytotoxic. Of the clinical PARP inhibitors, talazoparib is the most effective at trapping PARP1 on damaged DNA. Biochemically, talazoparib is also suspected to be a potent inhibitor of PARP5a/b (tankyrase1/2), which is an important regulator of Wnt/β-catenin pathway. Here we show using competition experiments in cell lysate that, at a clinically relevant concentration, talazoparib can potentially bind and engage tankyrase1. Using surface plasmon resonance, we measured the dissociation constants of talazoparib, olaparib, niraparib and veliparib for their interaction with PARP1 and tankyrase1. The results show that talazoparib has strong affinity for PARP1 as well as uniquely strong affinity for tankyrase1. Finally, we used crystallography and hydrogen deuterium exchange mass spectroscopy to dissect the molecular mechanism of differential selectivity of these PARP1 inhibitors. From these data, we conclude that subtle differences between the ligand binding sites of PARP1 and tankyrase1, differences in the electrostatic nature of the ligands, protein dynamics, and ligand conformational energetics contribute to the different pharmacology of these PARP1 inhibitors. These results will help in the design of drugs to treat Wnt-β-catenin pathway-related cancers, such as colorectal cancers.

Download Full-text

The Naturally Occurring Mutants of DDB Are Impaired in Stimulating Nuclear Import of the p125 Subunit and E2F1-Activated Transcription

Molecular and Cellular Biology ◽

10.1128/mcb.19.7.4935 ◽

1999 ◽

Vol 19 (7) ◽

pp. 4935-4943 ◽

Cited By ~ 72

Author(s):

Pavel Shiyanov ◽

Steven A. Hayes ◽

Manjula Donepudi ◽

Anne F. Nichols ◽

Stuart Linn ◽

...

Keyword(s):

Dna Binding ◽

Critical Role ◽

Binding Activity ◽

Single Amino Acid ◽

Nuclear Accumulation ◽

Stable Complex ◽

Deficiency Disease ◽

Naturally Occurring ◽

Repair Deficiency ◽

Damaged Dna

ABSTRACT The human UV-damaged-DNA binding protein DDB has been linked to the repair deficiency disease xeroderma pigmentosum group E (XP-E), because a subset of XP-E patients lack the damaged-DNA binding function of DDB. Moreover, the microinjection of purified DDB complements the repair deficiency in XP-E cells lacking DDB. Two naturally occurring XP-E mutations of DDB, 82TO and 2RO, have been characterized. They have single amino acid substitutions (K244E and R273H) within the WD motif of the p48 subunit of DDB, and the mutated proteins lack the damaged-DNA binding activity. In this report, we describe a new function of the p48 subunit of DDB, which reveals additional defects in the function of the XP-E mutants. We show that when the subunits of DDB were expressed individually, p48 localized in the nucleus and p125 localized in the cytoplasm. The coexpression of p125 with p48 resulted in an increased accumulation of p125 in the nucleus, indicating that p48 plays a critical role in the nuclear localization of p125. The mutant forms of p48, 2RO and 82TO, are deficient in stimulating the nuclear accumulation of the p125 subunit of DDB. In addition, the mutant 2RO fails to form a stable complex with the p125 subunit of DDB. Our previous studies indicated that DDB can associate with the transcription factor E2F1 and can function as a transcriptional partner of E2F1. Here we show that the two mutants, while they associate with E2F1 as efficiently as wild-type p48, are severely impaired in stimulating E2F1-activated transcription. This is consistent with our observation that both subunits of DDB are required to stimulate E2F1-activated transcription. The results provide insights into the functions of the subunits of DDB and suggest a possible link between the role of DDB in E2F1-activated transcription and the repair deficiency disease XP-E.

Download Full-text

Abstract A15: Downregulation of c-myc is synthetic lethal with PARP inhibitors in high MYC cancers independent of BRCA status

10.1158/1538-8514.synthleth-a15 ◽

2017 ◽

Author(s):

Jason PW Carey ◽

Smruthi Vjayaraghavan ◽

Kelly Hunt ◽

Khandan Keyomarsi

Keyword(s):

Parp Inhibitors ◽

Synthetic Lethal

Download Full-text

Therapeutic relevance of homologous recombination repair (HRR) pathway variants in metastatic colorectal cancer (mCRC).

Journal of Clinical Oncology ◽

10.1200/jco.2018.36.4_suppl.677 ◽

2018 ◽

Vol 36 (4_suppl) ◽

pp. 677-677

Author(s):

Ravi Kishore Narra ◽

Arun K. Singavi ◽

Paul S. Ritch ◽

James P. Thomas ◽

Abdel Alqwasmi ◽

...

Keyword(s):

Critical Role ◽

Molecular Data ◽

Parp Inhibitors ◽

Dna Breaks ◽

Characterization Methods ◽

Variants Of Unknown Significance ◽

Platinum Based Chemotherapy ◽

Unknown Significance ◽

Formalin Fixed Paraffin Embedded ◽

Chemotherapy Patients

677 Background: Intact HRR pathway genes play a critical role in repairing double-stranded DNA breaks. Genomic alterations (GA) in several HRR pathway genes have been well characterized, demonstrating prognostic and predictive significance. However, there are many variants of unknown significance (VUS) in the HRR pathway genes that need better characterization. Methods: Patients (pts) with mCRC that harbored VUSs in HRR pathway genes (HRR- VUS; BRCA1/2, ATM, ATR, RAD50, RAD51, PALB2, CHEK1, CHEK2) and treated with front line platinum therapy were identified by review of our institutional molecular data base (MDB) and EMR. Time to 2nd line therapy (TT2L) and Overall Survival (OS) were calculated. DNA was extracted from formalin fixed paraffin embedded clinical specimens and Next Generation Sequencing (NGS) was performed on hybrid-capture, adaptor ligation based libraries to a mean coverage depth of > 600 unique reads utilizing the Foundation Medicine NGS platform. Results: Among the 873 pts in our institutional MDB, 96 (11%) had mCRC, 20 (21%) harbored HRR pathway GAs, 15 (16%) were categorized as VUSs. HRR-VUS pts had a median age of 57 at diagnosis, 5 (33%) were male, 3 (20%) were right-sided. Distribution of HRR-VUSs in this cohort is summarized in table 1. All pts with HRR-VUSs were micro-satellite stable (MSS), median TMB was 4.5 (range 0 – 8.1). Median TT2L and OS in mCRC pts with HRR-VUSs were 8.0 and 43 months (m) respectively. Two pts with HRR-VUSs had TT2L > 24 m and OS > 40 m; one of them had a BRCA1P1464A mutation, while the other had a BRCA2 amplification. Conclusions: BRCA2, ATM and BRCA1 were the HRR genes that harbored VUSs most frequently in pts with mCRC. Majority with HRR-VUSs had left sided primaries, were MSS and had low TMB. BRCA1P1464A mutation in the ATM binding domain needs further characterization on account of prolonged TT2L with platinum based chemotherapy. Patients with HRR-VUSs that are predictive of benefit with platinum based chemotherapies could be considered for maintenance therapy with PARP inhibitors [Table: see text]

Download Full-text

Poly(ADP-Ribose) polymerase (PARP) inhibitors: Exploiting a synthetic lethal strategy in the clinic

CA A Cancer Journal for Clinicians ◽

10.3322/caac.20095 ◽

2011 ◽

Vol 61 (1) ◽

pp. 31-49 ◽

Cited By ~ 126

Author(s):

Timothy A. Yap ◽

Shahneen K. Sandhu ◽

Craig P. Carden ◽

Johann S. de Bono

Keyword(s):

Parp Inhibitors ◽

Synthetic Lethal

Download Full-text

NR1D1 regulation by Ran GTPase via miR4472 identifies an essential vulnerability linked to aneuploidy in ovarian cancer

Oncogene ◽

10.1038/s41388-021-02082-z ◽

2021 ◽

Author(s):

Zied Boudhraa ◽

Kossay Zaoui ◽

Hubert Fleury ◽

Maxime Cahuzac ◽

Sophie Gilbert ◽

...

Keyword(s):

Ovarian Cancer ◽

Dna Repair ◽

Critical Role ◽

Inverse Correlation ◽

Dna Damages ◽

Ran Gtpase ◽

Synthetic Lethal ◽

Damage Repair ◽

Repair Pathways

AbstractWhile aneuploidy is a main enabling characteristic of cancers, it also creates specific vulnerabilities. Here we demonstrate that Ran inhibition targets epithelial ovarian cancer (EOC) survival through its characteristic aneuploidy. We show that induction of aneuploidy in rare diploid EOC cell lines or normal cells renders them highly dependent on Ran. We also establish an inverse correlation between Ran and the tumor suppressor NR1D1 and reveal the critical role of Ran/NR1D1 axis in aneuploidy-associated endogenous DNA damage repair. Mechanistically, we show that Ran, through the maturation of miR4472, destabilizes the mRNA of NR1D1 impacting several DNA repair pathways. We showed that NR1D1 interacts with both PARP1 and BRCA1 leading to the inhibition of DNA repair. Concordantly, loss of Ran was associated with NR1D1 induction, accumulation of DNA damages, and lethality of aneuploid EOC cells. Our findings suggest a synthetic lethal strategy targeting aneuploid cells based on their dependency to Ran.

Download Full-text

Identifying chemogenetic interactions from CRISPR knockout screens with drugZ

10.1101/232736 ◽

2017 ◽

Cited By ~ 2

Author(s):

Medina Colic ◽

Gang Wang ◽

Michal Zimmermann ◽

Keith Mascall ◽

Megan McLaughlin ◽

...

Keyword(s):

Gene Mutations ◽

Drug Efficacy ◽

Chemical Compounds ◽

Parp Inhibitors ◽

Resistance Mechanisms ◽

Synthetic Lethal ◽

Drug Mechanism ◽

Damage Repair ◽

Synthetic Lethal Interactions ◽

Additional Cell

AbstractChemogenetic profiling enables the identification of gene mutations that enhance or suppress the activity of chemical compounds. This knowledge provides insights into drug mechanism-of-action, genetic vulnerabilities, and resistance mechanisms, all of which may help stratify patient populations and improve drug efficacy. CRISPR-based screening enables sensitive detection of drug-gene interactions directly in human cells, but until recently has largely been used to screen only for resistance mechanisms. We present drugZ, an algorithm for identifying both synergistic and suppressor chemogenetic interactions from CRISPR screens. DrugZ identifies synthetic lethal interactions between PARP inhibitors and both known and novel members of the DNA damage repair pathway. Additionally, drugZ confirms KEAP1 loss as a resistance factor for ERK inhibitors in oncogenic KRAS backgrounds and identifies additional cell-specific mechanisms of drug resistance. The software is available at github.com/hart-lab/drugz.

Download Full-text

Poly (ADP-Ribose) Polymerases (PARPs) and PARP Inhibitor-Targeted Therapeutics

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520618666181109164645 ◽

2019 ◽

Vol 19 (2) ◽

pp. 206-212 ◽

Cited By ~ 1

Author(s):

Nan Li ◽

Yifan Wang ◽

Weiye Deng ◽

Steven H. Lin

Keyword(s):

Small Molecule ◽

Anticancer Agents ◽

Current Knowledge ◽

Parp Inhibitor ◽

Parp Inhibitors ◽

Parp Inhibition ◽

Post Translational Modification ◽

Unique Type ◽

Synthetic Lethal ◽

Therapeutic Tools

Background:Poly-ADP-ribosylation, that is, adding ADP-ribose moieties to a protein, is a unique type of protein post-translational modification that regulates various cellular processes such as DNA repair, mitosis, transcription, and cell growth. Small-molecule inhibitors of poly-ADP-ribose polymerase 1 (PARP1) have been developed as anticancer agents because inhibition of PARP enzymes may be a synthetic lethal strategy for cancers with or BRCA2 mutations. However, there are still questions surrounding PARP inhibitors.Methods/Results:Data were collected from Pubmed, Medline, through searching of these keywords: “PARP”, “BRCA”, “Synthetic lethal” and “Tankyrase inhibitors”. We describe the current knowledge of PARP inhibition and its effects on DNA damage; mechanisms of resistance to PARP inhibitors; the evolution of PARP inhibitors; and the potential use of PARP5a/b (tankyrases) inhibitors in cancer treatment.Conclusion:PARP inhibitors are already showing promise as therapeutic tools, especially in the management of BRCA-mutated breast and ovarian cancers but also in tumors with dysfunctional BRCA genes. Small-molecule tankyrase inhibitors are important for increasing our understanding of tankyrase biology.

Download Full-text