Poly(ADP-ribosyl)polymerase Inhibitor Research Articles

Oxidized mC modulates synthetic lethality to PARP inhibitors for the treatment of leukemia

TET2 haploinsufficiency is a driving event in myeloid cancers and is associated with a worse prognosis in patients with acute myeloid leukemia (AML). Enhancing residual TET2 activity using vitamin C increases oxidized 5-methylcytosine (mC) formation and promotes active DNA demethylation via base excision repair (BER), which slows leukemia progression. We utilize genetic and compound library screening approaches to identify rational combination treatment strategies to improve use of vitamin C as an adjuvant therapy for AML. In addition to increasing the efficacy of several US Food and Drug Administration (FDA)-approved drugs, vitamin C treatment with poly-ADP-ribosyl polymerase inhibitors (PARPis) elicits a strong synergistic effect to block AML self-renewal in murine and human AML models. Vitamin-C-mediated TET activation combined with PARPis causes enrichment of chromatin-bound PARP1 at oxidized mCs and γH2AX accumulation during mid-S phase, leading to cell cycle stalling and differentiation. Given that most AML subtypes maintain residual TET2 expression, vitamin C could elicit broad efficacy as a PARPi therapeutic adjuvant.

Synergetic effect of poly (ADP-ribosyl) polymerase (PARP) inhibitor and cisplatin on ovarian cancer

Purpose: To investigate the effect of the combination of poly (ADP-ribosyl) polymerase (PARP) inhibitor, talazoparib (BMN673), and cisplatin on the proliferation and apoptosis of ovarian cancer cells in vitro, and on xenograft tumors of ovarian cancer cells in vivo.
 Methods: Cell viability was determined by CCK-8 assay, while cell proliferation and cell cycle were assessed using colony formation assay and flow cytometry, respectively. Cell apoptosis was evaluated by flow cytometry and TUNEL assays. Western blot assay was used to measure the expression levels of proliferation- and apoptosis-related proteins.
 Results: The PARP inhibitor, BMN673, produced a dose-dependent synergistic effect with cisplatin (p < 0.05). Compared with when cisplatin or BMN673 was used alone, the combination of cisplatin and BMN673 significantly inhibited the growth of transplanted tumors (p < 0.05). Immunohistochemical analysis showed that cisplatin and BMN673 treatment increased the number of cells positive for TUNEL, but reduced the population of cells positive for Ki67.
 Conclusion: Thus, BMN673 and cisplatin are synergistic against ovarian cancer cells, and therefore, should be subjected to further investigations, including clinical trials, to determine the potentials of the combination for the management of ovarian carcinoma.
 Keywords: PARP inhibitor; Ovarian cancer; Xenografted tumor; Cisplatin

PARP Inhibitor Olaparib Causes No Potentiation of the Bleomycin Effect in VERO Cells, Even in the Presence of Pooled ATM, DNA-PK, and LigIV Inhibitors.

Poly(ADP-ribosyl)polymerase (PARP) synthesizes poly(ADP-ribose) (PAR), which is anchored to proteins. PAR facilitates multiprotein complexes’ assembly. Nuclear PAR affects chromatin’s structure and functions, including transcriptional regulation. In response to stress, particularly genotoxic stress, PARP activation facilitates DNA damage repair. The PARP inhibitor Olaparib (OLA) displays synthetic lethality with mutated homologous recombination proteins (BRCA-1/2), base excision repair proteins (XRCC1, Polβ), and canonical nonhomologous end joining (LigIV). However, the limits of synthetic lethality are not clear. On one hand, it is unknown whether any limiting factor of homologous recombination can be a synthetic PARP lethality partner. On the other hand, some BRCA-mutated patients are not responsive to OLA for still unknown reasons. In an effort to help delineate the boundaries of synthetic lethality, we have induced DNA damage in VERO cells with the radiomimetic chemotherapeutic agent bleomycin (BLEO). A VERO subpopulation was resistant to BLEO, BLEO + OLA, and BLEO + OLA + ATM inhibitor KU55933 + DNA-PK inhibitor KU-0060648 + LigIV inhibitor SCR7 pyrazine. Regarding the mechanism(s) behind the resistance and lack of synthetic lethality, some hypotheses have been discarded and alternative hypotheses are suggested.

Neoadjuvant Treatment for Triple Negative Breast Cancer: Recent Progresses and Challenges.

Triple negative breast cancer (TNBC) is an aggressive breast cancer with historically poor outcomes, primarily due to the lack of effective targeted therapies. The tumor molecular heterogeneity of TNBC has been well recognized, yet molecular subtype driven therapy remains lacking. While neoadjuvant anthracycline and taxane-based chemotherapy remains the standard of care for early stage TNBC, the optimal chemotherapy regimen is debatable. The addition of carboplatin to anthracycline, cyclophosphamide, and taxane (ACT) regimen is associated with improved complete pathologic response (pCR). Immune checkpoint inhibitor (ICI) combinations significantly increase pCR in TNBC. Increased tumor infiltrating lymphocyte (TILs) or the presence of DNA repair deficiency (DRD) mutation is associated with increased pCR. Other targets, such as poly-ADP-ribosyl polymerase inhibitors (PARPi) and Phosphatidylinositol-3-kinase/Protein Kinase B/mammalian target of rapamycin (PI3K-AKT-mTOR) pathway inhibitors, are being evaluated in the neoadjuvant setting. This review examines recent progress in neoadjuvant therapy of TNBC, including platinum, ICI, PARPi, phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) pathway targeted therapies, and novel tumor microenvironment (TME) targeted therapy, in addition to biomarkers for the prediction of pCR.

Advances in the use of PARP inhibitors for BRCA1/2-associated breast cancer: talazoparib.

Poly-ADP-ribosyl polymerase (PARP) enzymes PARP-1 and PARP-2 recognize DNA damage and set off a cascade of cellular mechanisms required for multiple types of DNA damage repair. PARP inhibitors are small molecule mimetics of nicotinamide which bind to PARP's catalytic domain to inhibit poly-ADP-ribosylation (PARylation) of target proteins, including PARP-1 itself. PARP inhibitors olaparib, veliparib, talazoparib, niraparib and rucaparib have predominantly been studied in women with breast or ovarian cancers associated with deleterious germline mutations in BRCA1 and BRCA2 (gBRCA1/2+). The BRCA1 and BRCA2 proteins are involved in DNA repair by homologous recombination. This review will focus on talazoparib, a PARP inhibitor approved by the US FDA for the treatment of metastatic gBRCA1/2+ breast cancers in October 2018.

Poly-ADP-ribosyl-polymerase inhibitor resistance mechanisms and their therapeutic implications.

Poly-ADP-ribosyl-polymerase (PARP) inhibitors are an increasingly-utilized therapy in women with high-grade serous ovarian carcinoma, but tumor resistance to PARP inhibitor monotherapy is inevitable. PARP inhibitors have been most studied in patients with breast and ovarian cancers associated with deleterious germline BRCA1 or BRCA2 mutations, though their role has expanded to include use as maintenance therapy in women with platinum-sensitive high-grade serous ovarian cancer due to the high propensity of such cancers to have defects in DNA repair by homologous recombination. As mechanisms of PARP inhibitor resistance are elucidated, rationale combination strategies can be devised to extend therapeutic benefits and to abrogate resistance. Mechanisms of resistance include restoration of homologous recombination repair proficiency, loss of cancer cell reliance on PARP, and increased intracellular signaling through cell growth pathways.

Abstract 3784: Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells

Abstract The field of ovarian cancer therapeutics has made a substantial leap forward with the discovery that ovarian cancers with a BRCA mutation, can be exploited with “synthetic lethality.” In patients who have a BRCA germline mutation, this phenomenon is achieved by blocking PARP (poly-ADP-ribosyl-polymerase), which has a role in compensating for the missing or low levels of the BRCA protein. PARP activity is essential for DNA repair in patients with BRCA mutations as their cells attempt to maintain a normal genotype; however, PARP is not enough to compensate for the BRCA mutation, and these patients eventually develop ovarian cancer. Out of the five major mechanisms of DNA repair, PARP compensates for missing BRCA1 and/or BRCA2 proteins required for homologous repair, base excision repair and nuclear excision repair. If PARP is inhibited in BRCA-mutant patients, DNA mutations accumulate and are not repaired because PARP is not available to ribosylate the areas of cut or damaged DNA, which would signal that repair is needed. The BRCA-mutant genotype, in which there is a germline-inactivating mutation in one allele and loss of heterozygosity or somatic mutation in the other allele, only occurs in 5-10% of the cancer population. We wondered, “is there a way to apply the concept of “synthetic lethality” to the 90-95% of the ovarian cancer patients who do not have BRCA mutations?” We tried to answer that query by testing different gene-expression, modulating agents. We found that low-concentration hydroxyurea can sometimes potentiate the cytotoxicity of alkylating agents, suggesting a mechanism that involves DNA repair. We found synergistic apoptosis in 4 (ovcar-8, TOV21G, PA-1 and UACC1598) ovarian cancer cell lines treated with hydroxyurea and the PARP inhibitor, Olaparib, and in cells treated with curcumin with Olaparib. When the Ovcar-8 cancer cell line was treated with a single agent, the percentage of apoptosis was ≤ 17%, and when treated with synergistic agents together, relative apoptosis increased to ≤ 45%. In these cell lines, both curcumin and hydroxyurea significantly decreased BRCA1 and BRCA2 mRNA and protein levels. We also found that curcumin triggered BRCA1,2 promoter methylation, which led to cessation of gene expression. Whereas, hydroxyurea, not only caused BRCA1,2 promoter methylation, but also elicited an increase in miRNA hsa-miR-17-5p and hsa-miR-182-5p, causing BRCA1 mRNA degradation. This finding suggests greater than 5-10% of wild-type, BRCA ovarian cancer patients may benefit from PARP inhibitors and DNA cross-linking agents, such as mitomycin C. if they can be induced into “BRCAness,” Citation Format: Yuehua Mao, Richard D Dinnen, Robert L. Fine. Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3784. doi:10.1158/1538-7445.AM2014-3784

Reversal of neurobehavioral and neurochemical alterations in STZ-induced diabetic rats by FeTMPyP, a peroxynitrite decomposition catalyst and 1,5-Isoquinolinediol a poly(ADP-ribose) polymerase inhibitor

Objective:In this study, we have evaluated the involvement of nitrosative stress and poly-ADP ribosyl polymerase (PARP) in diabetes induced neurobehavioral and neurochemical changes using pharmacological agents peroxynitrite decomposition catalyst (FeTMPyP) and a PARP inhibitor (1,5-Isoquinolinediol) in diabetic brains.Methods:The extent of neurobehavioral changes was assessed by functional observation battery, motor coordination activity (rota rod performance) and passive avoidance test. Neurochemical changes were assessed by measuring nicotinamide adenine dinucleotide (NAD), malondialdehyde, acetylcholinesterase, neurotransmitters (GABA and glutamate) levels in the hippocampus. GABA and glutamate were measured by high-performance liquid chromatography with electrochemical detection method.Results:Two weeks’ treatment with FeTMPyP (3 mg/kg, i.p.) and 1,5-Isoquinolinediol (3 mg/kg, i.p.) improved the cognitive deficits in diabetic rats as observed in passive avoidance test. Both the agents inhibited lipid peroxidation and improves the acetylcholinesterase level in the hippocampus. 1,5-Isoquinolinediol treatment also improves the NAD, neurotransmitter level in the hippocampus.Discussion:These results suggest that peroxynitrite decomposition catalyst and PARP inhibitor have beneficial effects in neurobehavioral alterations induced by diabetes. Improvement in neurobehavioral alteration may be attributed to reversal of neurotransmitter homeostasis deficits.

Correlation of increased PARP14 and CCL26 expression in biopsies from children with eosinophilic esophagitis

Correlation of increased PARP14 and CCL26 expression in biopsies from children with eosinophilic esophagitis

BRCA1- and BRCA2-related mutations: therapeutic implications in ovarian cancer

Ovarian cancer is the deadliest among gynecologic cancers. Hereditary cancer related to BRCA1/2 gene mutations account for ∼10%–12% of ovarian cancers. The BRCA1/2 proteins are important in homologous recombination (HR) repair of DNA. Patients with BRCA1/2 mutations have been reported to have improved chemosensitivity to platinum agents, longer disease-free intervals, and longer survivals than nonhereditary counterparts. Recent interest in poly(ADP-ribosyl) polymerase (PARP) proteins which are key components of base excision repair, has led to the development of PARP inhibitors; tumors arising in BRCA1/2 mutation carriers and/or with HR deficiency (HRD) are particularly sensitive to the action of these drugs. As 60%–80% of all advanced ovarian cancers are high-grade serous type, exhibiting HRD in at least 50% (referred as BRCAness) future antitumor strategies may depend on identifying these defects through molecular testing. Once HRD becomes amenable to routine testing, a larger group of ovarian cancer patients than are currently considered for PARP inhibitor trials, may benefit from such targeted therapy.

Abstract B30: Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells

Abstract The field of ovarian cancer therapeutics has recently made a substantial leap forward with the discovery that ovarian cancers with a BRCA mutation can be exploited with “synthetic lethality”. This phenomenon takes advantage of the role of PARP (poly-ADP ribosyl-polymerase), which compensates for the missing or low BRCA protein in these patients with a BRCA germline mutation. PARP activity is essential for these patients to maintain a normal genotype, albeit not completely normal since they developed ovarian cancer mainly because of their BRCA mutation. Thus, in the 5 major mechanisms of DNA repair PARP compensates for missing BRCA1 or BRCA2 in homologous repair, base excision repair and nuclear excision repair. In these patients, if PARP is inhibited, then massive DNA mutations accumulate and are not repaired because PARP ribosylates the areas of cut or damaged DNA to mark the areas needed for repair to occur. The concept of inhibiting PARP in these BRCA mutated tumors is an elegant idea. The caveat here is the BRCA mutant genotype where there is a germline, inactivating mutation in one allele and LOH or somatic mutation of the other allele only occurs in 5-10% of the cancer population. Thus, could there be a way to apply this concept of synthetic lethality to the other 90-95% of the cancer population without BRCA mutations in ovarian cancer patients? We tried to answer that question by testing different agent which could modulate gene expression, and found that curcumin and hydroxyurea, at low concentrations, could sometimes increase the cytotoxicity of alkylating agents which suggests a mechanism involving DNA repair. Four (ovcar-8, TOV21G, PA-1 and UACC1598) out of nine pretreated ovarian cancer cell lines with hydroxyurea or curcumin has synergistically increase apoptosis induced by PARP inhibitor - Olaparid. In the tested Ovcar-8 cancer cell line, Curcumin (5 μM) alone induced 10% apoptosis, hydroxyurea (100 μM) alone induced 17% apoptosis, Olaparid (2.5 μM) alone induced 7% cell death. Curcumin and Olaparid together induce 45% cell death (165% increase), hydroxurea and Olaparid together induced 40% (67% increase). Both curcumin and hydroxyurea decrease BRCA1 and BRCA2 mRNA and protein level in the tested sensitive cell lines. Further study, we found that curcumin prompt epigenetically BRCA1,2 promoter methylation and lead to gene expression shutting down whereas hydroxyurea induced miRNA hsa-miR-17-5p and hsa-miR-182-5p and lead BRCA1 mRNA degradation. This finding suggest If wt BRCA patients with ovarian cancer could be induced into BRCAness, then perhaps a larger percentage than 5-10% could benefit from PARP inhibitors and DNA crosslinkers like mitomycin C. Citation Format: Yuehua Mao, Richard D Dinnen, Robert L Fine. Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: From Concept to Clinic; Sep 18-21, 2013; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2013;19(19 Suppl):Abstract nr B30.

Inhibition of matrix metalloproteinase-2 by PARP inhibitors

Matrix metalloproteinase-2 (MMP-2), a ubiquitously expressed zinc-dependent endopeptidase, and poly(ADP-ribosyl) polymerase (PARP), a nuclear enzyme regulating DNA repair, are activated by nitroxidative stress associated with various pathologies. As MMP-2 plays a detrimental role in heart injuries resulting from enhanced nitroxidative stress, where PARP and MMP inhibitors are beneficial, we hypothesized that PARP inhibitors may affect MMP-2 activity. Using substrate degradation assays to determine MMP-2 activity we found that four PARP inhibitors (3-AB, PJ-34, 5-AIQ, and EB-47) inhibited 64 kDa MMP-2 in a concentration-dependent manner. The IC 50 values of PJ-34 and 5-AIQ were in the high micromolar range and comparable to those of known MMP-2 inhibitors doxycycline, minocycline or o-phenanthroline, whereas those for 3-AB and EB-47 were in the millimolar range. Co-incubation of PARP inhibitors with doxycycline showed an additive inhibition of MMP-2 that was significant for 3-AB alone. These data demonstrate that the protective effects of some PARP inhibitors may include inhibition of MMP-2 activity.

Different Basal NAD Levels Determine Opposite Effects of Poly(ADP-Ribosyl)Polymerase Inhibitors on H 2O 2-Induced Apoptosis

We have recently described that poly(ADP-ribosyl)-polymerase (PARP) inhibitors rescue U937 cells from apoptosis induced by 1 m M H 2O 2 oxidative stress; PARP activation leads to a reversible drop in NAD level, which could be blocked by PARP inhibitors (Nosseri et al., 1994, Exp. Cell Res. 212, 367-373). A phenotypic variant of U937 is characterized by a lower basal NAD level (low NAD, LN U937, as opposed to the original high NAD, HN U937). In LN cells treatment with 1 m M H 2O 2, although activating PARP, does not lower NAD concentration; puzzlingly, PARP inhibitors increase (instead of decreasing, as occurs in HN cells) tile extent of stress-induced apoptosis, leading to a reduced cell survival. NAD concentration could he increased in LN cells by adding nicotinamide (5- and 25-fold increase) to the culture medium. These cells (LN +) behaved as HN U937: oxidative stress induced a NAD drop, the extent of which is dependent on the cells' basal NAD level; moreover, PARP inhibitors could rescue LN + cells from peroxide-induced apoptosis. H 2O 2-induced apoptosis is not triggered by NAD depletion, but instead it takes place only when NAD levels have been preserved or have recovered: on HN U937, peroxide doses (5 and 10 m M) which lead to necrosis induce an irreversible NAD drop, whereas apoptosis occurs only at lower doses, where NAD depletion is reversible; on LN cells NAD levels do not drop even upon 10 m M H 2O 2 treatment, and these cells die only by apoptosis; moreover, in HN cells apoptosis is not detectable until 8 h posttreatment, when NAD levels recover, whereas in LN cells, where NAD is always present, apoptosis begins to take place as early as 3 h after stress.