BMN 673, a Novel and Highly Potent PARP1/2 Inhibitor for the Treatment of Human Cancers with DNA Repair Deficiency

<div>Abstract<p><b>Purpose:</b> PARP1/2 inhibitors are a class of anticancer agents that target tumor-specific defects in DNA repair. Here, we describe BMN 673, a novel, highly potent PARP1/2 inhibitor with favorable metabolic stability, oral bioavailability, and pharmacokinetic properties.</p><p><b>Experimental Design:</b> Potency and selectivity of BMN 673 was determined by biochemical assays. Anticancer activity either as a single-agent or in combination with other antitumor agents was evaluated both <i>in vitro</i> and in xenograft cancer models.</p><p><b>Results:</b> BMN 673 is a potent PARP1/2 inhibitor (PARP1 IC<sub>50</sub> = 0.57 nmol/L), but it does not inhibit other enzymes that we have tested. BMN 673 exhibits selective antitumor cytotoxicity and elicits DNA repair biomarkers at much lower concentrations than earlier generation PARP1/2 inhibitors (such as olaparib, rucaparib, and veliparib). <i>In vitro</i>, BMN 673 selectively targeted tumor cells with <i>BRCA1</i>, <i>BRCA2</i>, or <i>PTEN</i> gene defects with 20- to more than 200-fold greater potency than existing PARP1/2 inhibitors. BMN 673 is readily orally bioavailable, with more than 40% absolute oral bioavailability in rats when dosed in carboxylmethyl cellulose. Oral administration of BMN 673 elicited remarkable antitumor activity <i>in vivo</i>; xenografted tumors that carry defects in DNA repair due to <i>BRCA</i> mutations or PTEN deficiency were profoundly sensitive to oral BMN 673 treatment at well-tolerated doses in mice. Synergistic or additive antitumor effects were also found when BMN 673 was combined with temozolomide, SN38, or platinum drugs.</p><p><b>Conclusion:</b> BMN 673 is currently in early-phase clinical development and represents a promising PARP1/2 inhibitor with potentially advantageous features in its drug class. <i>Clin Cancer Res; 19(18); 5003–15. ©2013 AACR</i>.</p></div>

<div>Abstract<p><b>Purpose:</b> PARP1/2 inhibitors are a class of anticancer agents that target tumor-specific defects in DNA repair. Here, we describe BMN 673, a novel, highly potent PARP1/2 inhibitor with favorable metabolic stability, oral bioavailability, and pharmacokinetic properties.</p><p><b>Experimental Design:</b> Potency and selectivity of BMN 673 was determined by biochemical assays. Anticancer activity either as a single-agent or in combination with other antitumor agents was evaluated both <i>in vitro</i> and in xenograft cancer models.</p><p><b>Results:</b> BMN 673 is a potent PARP1/2 inhibitor (PARP1 IC<sub>50</sub> = 0.57 nmol/L), but it does not inhibit other enzymes that we have tested. BMN 673 exhibits selective antitumor cytotoxicity and elicits DNA repair biomarkers at much lower concentrations than earlier generation PARP1/2 inhibitors (such as olaparib, rucaparib, and veliparib). <i>In vitro</i>, BMN 673 selectively targeted tumor cells with <i>BRCA1</i>, <i>BRCA2</i>, or <i>PTEN</i> gene defects with 20- to more than 200-fold greater potency than existing PARP1/2 inhibitors. BMN 673 is readily orally bioavailable, with more than 40% absolute oral bioavailability in rats when dosed in carboxylmethyl cellulose. Oral administration of BMN 673 elicited remarkable antitumor activity <i>in vivo</i>; xenografted tumors that carry defects in DNA repair due to <i>BRCA</i> mutations or PTEN deficiency were profoundly sensitive to oral BMN 673 treatment at well-tolerated doses in mice. Synergistic or additive antitumor effects were also found when BMN 673 was combined with temozolomide, SN38, or platinum drugs.</p><p><b>Conclusion:</b> BMN 673 is currently in early-phase clinical development and represents a promising PARP1/2 inhibitor with potentially advantageous features in its drug class. <i>Clin Cancer Res; 19(18); 5003–15. ©2013 AACR</i>.</p></div>

Prostate cancer is the most diagnosed cancer among men in the United States. African American men are diagnosed with and succumb to prostate cancer at higher rates than other demographic groups. However, the biological drivers contributing to poorer outcomes in African American men compared to European American men with prostate cancer are still poorly defined. One understudied area is DNA repair defects, which drive genomic instability in cancers. Defects in homologous recombination are often at the forefront of repair defects examined and exploited for therapeutic intervention. However, defects in other DNA repair pathways may contribute to poor disease outcomes and chemoresistance. We demonstrated that defects in the base excision repair (BER) pathway contribute to chemoresistance and alter growth in breast cancer. Others have also shown chemoresistant effects from dysregulated BER protein expression in gastric, colorectal, and ovarian cancers. Examination of the TCGA database show dysregulation of key BER protein like X-ray cross complementing protein 1 (XRCC1) and DNA polymerase β (POLβ) are common across several cancers. Therefore, we decided to explore DNA damage and repair defects in prostate cancer to explore if altered DNA repair contributes to poorer outcomes in African American men. To measure DNA repair defects in formalin-fixed paraffin-embedded (FFPE) tissues, we employed a novel assay, Repair Assisted Damage Detection (RADD), which measures DNA lesions within the tissues. RADD fluorescently tags DNA lesions for quantification and is compatible with immunohistochemistry detection of DNA repair or other proteins of interest. Using the tissue microarrays, we used RADD to discover that prostate tumors from African American patients have more uracil and pyrimidine lesions, elevated uracil DNA glycosylase (UNG) levels, and reduced XRCC1 levels than European American tumors, which indicate defects in the BER pathway. In addition, these men had higher uracil monophosphate (UMP) and lower expression of folate cycle metabolites, suggesting that metabolic rewiring may also contribute to the dysregulation of BER. Defects in the BER pathway may offer new therapeutic opportunities for African American prostate cancer patients. These defects may be targeted by therapeutics which pressure the BER pathways. This work demonstrates that functional detection of DNA repair defects through RADD offers new insight into race-specific DNA repair defects offering new molecular targets or therapeutic strategies to reduce racial disparities in prostate cancer. Citation Format: Kaveri Goel, Kimiko Krieger, Manoj Sonavane, Arun Sreekumar, Natalie R. Gassman. Race-associated base excision repair defects alter the DNA damage landscape in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6114.

3rd International Genome Dynamics in Neuroscience Conference: “DNA repair and neurological disease”

DNA repair mechanisms and their biologic implications in mammalian cells: Report of a NATO advanced research workshop, Fontevraud, France, Oct. 2–7, 1988

The FASEB JournalVolume 32, Issue 12 p. 6355-6355 EditorialFree Access DNA repair and cancer: the semicentennial First published: 05 November 2018 https://doi.org/10.1096/fj.181201ufmCitations: 1AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat “Where the rivers change direction, across the great divide” The 1950s saw the establishment of “DNA constancy,” in which measurements of the DNA of somatic cells in a given organism revealed very similar contents, and such measurements on germ cells revealed half as much. The revelation of DNA constancy played a tremendous heuristic role in the growing acceptance of DNA as the gene. In addition, and this took longer to be absorbed, the fact that the somatic cells of an organism have the same DNA content gave rise to the variable gene theory of differentiation and development, in which it is not the selective deletion or retention of genes but rather the varying patterns of their expression from a constant keyboard. But even prior to this period, DNA mutation had been discovered, and so DNA constancy, although hugely important, was nonetheless something of a misnomer: DNA was constant only if not changed. The induction of damage without repair (repair being unanticipated) was why Hermann J. Muller's X-irradiation experiments resulted in phenotypic change and was why the experiments of Beadle and Tatum in Neurospora were successful (leading to the principle of “one gene, one enzyme”), and why the acridine dye-induced frameshift mutations, uncorrected, allowed Sydney Brenner and Francis Crick and colleagues to deduce that the genetic code was a triplet nucleotide language. The notion that DNA damage can sometimes be repaired seems obvious in hindsight, but like all concepts in biologic science, it had to earn its wings. It was first seen in bacteria and later in mammalian cells. Discoveries in DNA repair were recognized by the 2015 Nobel Prize in Chemistry. Although not viewed as a parallel epistemological corridor at the time of the aforementioned mutagenesis-based discoveries, the question of cancer-causing genes subsequently arose. This concept was launched by Peyton Rous for a chicken sarcoma and then steadily rose, culminating in the discovery of viral genes that had been incorporated into the human genome. The notion that endogenous genes could undergo mutation into oncogenes was the key conceptual advance. But another essential concept had already been well established—a link between deficiencies in DNA repair and cancer. This discovery preceded the oncogene era, and this year is its 50th anniversary. A connection between UV sensitivity and skin cancer had long been known, but in 1967, a young research fellow at the University of California, San Francisco, James Cleaver (Fig. 1), had the idea that the reason patients with the UV-sensitive disease xeroderma pigmentosum (XP) had an elevated incidence of skin cancer was perhaps because they could not repair the DNA lesions. This was a bold conceptual leap, viz.,that a defect in DNA repair could be carcinogenic. In short order, Cleaver demonstrated that cells from patients with XP were indeed defective in DNA repair (1, 2). This milestone paper generated further discoveries that solidified the connection between DNA repair deficiencies and many human cancers. Deficiencies in DNA repair are now known to lie at the pathogenesis of many human diseases beyond cancer. At the 50th anniversary of his seminal paper, let us take this opportunity to salute James Cleaver for his keen nose and all that it has meant for the modern era of the field of DNA repair and cancer. Figure 1Open in figure viewer James Cleaver. Epilogue: The quoted line is by the American songwriter Kate Wolf (1942–1986), who penned it as a metaphor for changing one's world view, just as ideas about genes and cancer eventually also changed, not across the western orogenic mountain range of the North American continent, but from a century of oppositional views. REFERENCES 1Cleaver, J. E. (1968) Defective repair replication of DNA in xeroderma pigmentosum. Nature 218, 652– 656 2Wood, R. D. (2018) Fifty years since DNA repair was linked to cancer. Nature 557, 648– 649 Citing Literature Volume32, Issue12September 2018Pages 6355-6355 FiguresReferencesRelatedInformation

Hepatitis B virus (HBV) is implicated in liver cancer. The aim of this study was to find out whether HBV or its components [HBV surface antigen (HBsAg), HBV core protein (HBc), and HBV X protein (HBx)] could interfere with the host DNA damage response and repair pathway. The full HBV genome or individual HBV open-reading frame (ORF) was introduced into HepG2 cells to examine the effect on host genomic stability, DNA repair efficacy in response to double-strand DNA damage, and DNA damage-induced cell death. Responses to apoptosis induction in the HBV ORF-transfected HepG2 cells were also compared with those in HBV-positive and HBV-negative human hepatocellular carcinoma (HCC) cells. In the absence of HBV replication, accumulation of HBsAg in liver cells without other HBV proteins enhanced DNA repair protein and tumor suppressor promyelocytic leukemia (PML) degradation, which resulted in resistance to apoptosis induction and deficient double-strand DNA repair. However, HBsAg-positive cells exhibited increased cell death with exposure to the poly(ADP-ribose) polymerase inhibitor that blocks single-strand DNA repair. These results indicate that suppression of PML by HBsAg disrupts cellular mechanisms that respond to double-strand DNA damage for DNA repair or apoptosis induction, which may facilitate hepatocarcinogenesis and open up a synthetic lethality strategy for HBsAg-positive HCC treatment.

There is growing evidence that many solid tumors have defective DNA repair pathways and cell cycle checkpoints. This has spurred development of therapeutics to specifically target cancer cells with defective DNA repair, but few have been successful outright due to problems such as general toxicity. One novel anti-cancer agent of interest to us is 3E10, a unique cell-penetrating, anti-DNA autoantibody. This molecule is non-toxic to normal cells on its own but has deleterious effects on cells with DNA repair deficiencies. Specifically, 3E10 has been shown to be synthetically lethal in BRCA2-deficient human cancer cells, to be synergistic with radiation and other commonly used DNA damaging therapies, and to reduce the efficiency of Rad51 mediated strand exchange and thus homologous recombination (HR). The goal of this project is to better understand how 3E10 can be used as a new paradigm for cancer treatment. New evidence suggests that 3E10 inhibits Exo1 exonuclease activity. Additionally, preliminary data shows that 3E10 physically interacts with Rad51, ultimately inhibiting Rad51 nuclear localization and foci formation. Additionally, a point mutation in 3E10 that confers increased DNA binding mediates increased cell death in BRCA2-deficient human cancer cells. Furthermore, studies investigating the potential therapeutic effect of 3E10 in patient derived primary melanoma cell lines show that 3E10 is synthetically lethal with PTEN deficiency. This data suggests that 3E10 inhibits HR by binding to DNA at double strand breaks and occupying sites normally processed by core repair machinery, as well as by physically interacting with Rad51. Overall, 3E10 holds great promise as a novel therapeutic agent for various cancers, including PTEN deficient melanomas. Reference: Hansen, J. et al. Targeting cancer with a lupus autoantibody. Science Translational Medicine 4, (2012) Citation Format: Audrey L. Turchick, Peter M. Glazer. Targeting DNA repair deficient cancers with the cell-penetrating autoantibody 3E10. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2748.

Replication protein A (RPA) is the primary ssDNA-binding protein in eukaryotes. RPA is essential for DNA replication, repair, and recombination. Mutation of a conserved leucine residue to proline in the high-affinity DNA binding site of RPA (residue L221 in human RPA) has been shown to have defects in DNA repair and a high rate of chromosomal rearrangements in yeast. The homologous mutation in mice was found to be lethal when homozygous and to cause high rates of cancer when heterozygous. To understand the molecular defect causing these phenotypes, we created the homologous mutation in the human RPA1 gene (L221P) and analyzed its properties in cells and in vitro. RPA1(L221P) does not support cell cycle progression when it is the only form of RPA1 in HeLa cells. This phenotype is caused by defects in DNA replication and repair. No phenotype is observed when cells contain both wild-type and L221P forms of RPA1, indicating that L221P is not dominant. Recombinant L221P polypeptide forms a stable complex with the other subunits of RPA, indicating that the mutation does not destabilize the protein; however, the resulting complex has dramatically reduced ssDNA binding activity and cannot support SV40 DNA replication in vitro. These findings indicate that in mammals, the L221P mutation causes a defect in ssDNA binding and a nonfunctional protein complex. This suggests that haploinsufficiency of RPA causes an increase in the levels of DNA damage and in the incidence of cancer.

The Involvement of DNA-Damage and -Repair Defects in Neurological Dysfunction

Fanconi Anemia (FA) is an autosomal recessive disorder characterized by diverse clinical manifestations such as aplastic anemia, cancer predisposition, and developmental defects including hypogonadism, microcephaly, organ dysfunction, infertility, hyperpigmentation, microphthalmia, and skeletal defects. In addition to the well-described defects in DNA repair, mitochondrial dysfunction due to defects in mitochondrial autophagy (mitophagy) is also associated with FA, although its contribution to FA phenotypes is unknown. This study focused on the FANCC gene, which, alongside other FA genes, is integral to DNA repair and mitochondrial quality control. In the present study, we created a FANCC mutant mouse model, based on a human mutation (FANCC c.67delG) that is defective in DNA repair but proficient in mitophagy. We found that the FANCC c.67delG mutant mouse model recapitulates some phenotypes observed in FA patients, such as cellular hypersensitivity to DNA cross-linking agents and hematopoietic defects. In contrast, FA phenotypes such as microphthalmia, hypogonadism, and infertility, present in FANCC-deficient mice, were absent in the FANCC c.67delG mice, suggesting that the N-terminal 55 amino acids of FANCC are dispensable for these developmental processes. Furthermore, the FANCC c.67delG mutation preserved mitophagy, and unlike the FANCC null mutation, did not lead to the accumulation of damaged mitochondria in cells or tissues. This study highlights the multifaceted nature of the FANCC protein, with distinct domains responsible for DNA repair and mitophagy. Our results suggest that developmental defects in FA may not solely stem from DNA repair deficiencies but could also involve other functions, such as mitochondrial quality control.

Inherited defects in DNA repair and susceptibility to DNA-damaging agents

DNA Lesion Measurement in Human Tissues.- Measurement of DNA Adducts by Immunoassays.- Detection of Human DNA Adducts by 32P-Postlabeling.- Immunologic Methods for the Detection of Carcinogen Adducts in Humans.- Lesion Measurement in Non-Radioactive DNA by Quantitative Gel Electrophoresis.- Fluorescence Detection of Lesions in DNA.- Damage and Repair in Skin.- Solar Radiation Induced Skin Cancer and DNA Photoproducts in Humans.- Defective DNA Repair in Humans: Clinical and Molecular Studies of Xeroderma Pigmentosum.- 5-MOP Induced Protection Against Epidermal DNA Damage by Ultraviolet Radiation in Human Skin.- Variability in DNA Repair in Human Skin.- Effects of Chemicals on Photobiologic Reactions of Skin.- Repair of 8-MOP Photoadducts in Human Lymphocytes.- Human Skin: Biochemistry and Molecular Biology.- DNA Damage and Repair in Human Skin: Pathways and Questions.- Unscheduled DNA Synthesis in Human Skin.- DNA Repair in Mammalian Tissues and Cells.- Damage Specific Mammalian Endonucleases.- Model Systems for Human Skin.- Characteristics of DNA Excision Repair in Nondividing Xeroderma Pigmentosum Cells, Complementation Group C.- Prospects for Epithelial Gene Therapy.- The Significance of DNA Damage and Repair Mechanisms in Health Risk Assessment.- DNA Damage and Repair in Human Blood Cells.- Measurements of Genomic and Gene-Specific DNA Repair of Alkylation Damage in Cultured Human T-Lymphocytes.- Peripheral Blood Leukocytes as a Surrogate Marker for Cisplatin Drug Resistance: Studies of Adduct Levels and the Repair Gene ERCCl.- Factors Which Affect DNA Repair in Human Lymphocytes.- Methyl Transferase Activity in Secondary Leukemia.- The Single Cell Gel Assay: A Sensitive Technique for Evaluating Intracellular Differences in DNA Damage and Repair.- Detection and Analysis of Human Mutations.- Cytogenetic Investigations of DNA Damage in Aging: A Twin Study.- Studies of DNA Alterations in In Vivo Somatic Cell Mutations in Humans.- Mechanisms of Induction of Specific Chromosomal Alterations.- Unfolding Perspectives on the Genetic Effects of Human Exposures to Radiation.- Effects of Caloric Restriction on the Maintenance of Genetic Fidelity.- Internal Organs.- Oncogenes and Tumor Suppressor Genes Involved in Human Lung Carcinogenesis.- Activation of Carcinogens by Human Liver Cytochromes P-450.- Repair of 06-Methylguanine Damage in Normal Human Tissues.- Repair of 04-Alkylthymine Damage in Human Cells.- Alkylation Repair in Human Tissues.- Participants.

Background Aberrant or impaired repair of double-strand DNA breaks is a common feature of de novo acute myeloid leukemia and myelodysplastic syndromes. Since poly (ADP-ribose) polymerase (PARP) inhibitors have been recently shown to selectively target cells with defects in double-strand DNA repair, the aim of this study was to explore the possibility of exploiting defects in DNA repair in leukemic cells using PARP inhibitors. Leukemic cell lines were exposed to various PARP inhibitors alone and in combination with non-cytotoxic concentrations of DNA methyltransferase inhibitor, 5' aza-2'-deoxycytidine and/or the histone deacetylase inhibitor, MS275, to test for potentiation of apoptosis with these agents. PARP inhibitors, KU-0058948 and PJ34, induced cell cycle arrest and apoptosis of primary myeloid leukemic cells and myeloid leukemic cell lines in vitro. Immunofluorescence analysis also revealed that PARP inhibitor sensitivity in these leukemic cells was due to a defect in homologous recombination DNA repair. Addition of 5' aza-2'-deoxycytidine failed to increase the cytotoxicity of PARP inhibitors. In contrast, MS275 potentiated the cytotoxic effect of KU-0058948 and PJ34 in all PARP inhibitor-sensitive leukemic cells. Immunofluorescence analysis supported the idea that histone deacetylase inhibitors potentiate cytotoxicity by inhibiting DNA repair processes. Conclusions On the basis of the data presented here, we suggest that PARP inhibitors can potentially exploit defects in double-strand DNA break repair in leukemic cells, paving the way for testing the therapeutic potential of these agents in myelodysplastic syndromes and acute myeloid leukemia.

Human cells have numerous repair mechanisms to counteract various insults incurred on the DNA. Any mutation in these repair mechanisms can lead to accumulation of DNA errors and carcinogenesis. This review aims to discuss the therapeutic options in the two most common DNA repair deficient cancer syndromes, namely Lynch syndrome (hereditary non-polyposis colorectal cancer) and breast cancer susceptibility gene (BRCA) associated ovarian and breast cancer. Deficiency in DNA repair mechanisms renders these tumors with increased sensitivity to platinum agents. There has been increasing amount of information on the utility of the defects in DNA repair as targets for cancer therapy in these syndromes. Novel therapies like poly (ADP-ribose) polymerase (PARP) inhibitors are one of such example where the induction of double stranded breaks in DNA leads to tumoricidal effect in patients with homologous DNA repair deficiency. Interestingly, patients with DNA repair deficiencies tend to have a more favorable prognosis than sporadic malignancies. In microsatellite high colorectal cancer patients, this has been attributed to increased recruitment of CD8+ T lymphocytes in tumor microenvironment. However, these tumors are able to limit the host immune response by activation of immune checkpoints that seem like attractive targets of therapy in the future.