Poly ADP-ribose Polymerase Research Articles

Combined Effects of Carbon Ion Radiation and PARP Inhibitor on Non-Small Cell Lung Carcinoma Cells: Insights into DNA Repair Pathways and Cell Death Mechanisms

The utilization of high linear energy transfer (LET) carbon ion (12C-ion) in radiotherapy has witnessed a notable rise in managing highly metastatic, recurrent, and chemo/radio-resistant human cancers. Non-small cell lung cancer (NSCLC) presents a formidable challenge due to its chemo-resistance and aggressive nature, resulting in poor prognosis and survival rates. In a previous study, we demonstrated that the combination of 12C-ion with the poly (ADP-ribose) polymerase (PARP) inhibitor (PARPi) olaparib significantly mitigated metastasis in A549 cells. Here, we delve into the underlying rationale behind the combined action of olaparib with 12C-ion, focusing on DNA repair pathways and cell death mechanisms in asynchronous NSCLC A549 cells following single and combined treatments. Evaluation included analysis of colony-forming ability, DNA damage assessed by γH2AX foci, expression profiling of key proteins involved in Homologous Recombination (HR) and Non-Homologous End Joining (NHEJ) repair pathways, caspase-3 activation, apoptotic body formation, and autophagic cell death. Our findings reveal that both PARPi olaparib and rucaparib sensitize A549 cells to 12C-ion exposure, with olaparib exhibiting superior sensitization. Moreover, 12C-ion exposure alone significantly downregulates both HR and NHEJ repair pathways by reducing the expression of MRE11-RAD50-RAD51 and Ku70-Ku80 protein complexes at 24h post-treatment. Notably, the combination of olaparib pre-treatment with 12C-ion markedly inhibits both HR and NHEJ pathways, culminating in DNA damage-induced apoptotic and autophagic cell death. Thus we are the first to demonstrate that olaparib sensitizes NSCLC cells to carbon ion by interfering with HR and NHEJ pathway. These insights underscore the promising therapeutic potential of combining PARP inhibition with carbon ion exposure for effective NSCLC management.

Abstract C046: PARP inhibitors as immune modulators in metastatic ovarian cancer treatment

Abstract Epithelial ovarian carcinoma (EOC) is one of the top five causes of cancer-related death in women. Most patients with EOC achieve initial remission after primary or interval cytoreductive surgery combined with platinum-taxane chemotherapy. However, mutations in BRCA1 or BRCA2 genes, which lead to homologous recombination (HR) defects, play a crucial role in platinum sensitivity and justify the use of poly (ADP-ribose) polymerase (PARP) inhibitors as maintenance therapy, commonly linked to extended progression-free survival (PFS). Besides their direct cytotoxic and cytostatic effects, PARP inhibitors (PARPi) have shown significant immunostimulatory properties by disrupting DNA repair in cancer cells, and opening possibilities for synergy with immune checkpoint inhibitors (ICIs). In this study, we investigate the immunomodulatory effects of PARPi using multiparametric flow cytometry, multiplexed immunolabeling, single-cell transcriptomics, and functional assays in an experimental BR5Brca1-/- syngeneic mouse model and human EOC tumor samples. We examine the molecular and cellular mechanisms that can be exploited to develop more effective combination therapies. PARPi may increase the mutational burden in EOCs due to unresolved DNA damage and the release of damage-associated molecular patterns (DAMPs), thereby increasing T-cell infiltration. In addition, PARPi appear to promote potent type I interferon (IFN) secretion through the activation of cGAMP synthase and the stimulator of the interferon genes (STING) pathway. In combination with ICIs, PARPi showed a beneficial effect on the balance between adaptive anti-tumor immunity and innate myeloid components, leading to an improved cytotoxic T-cell response, as observed in mouse models and HGSOC tumor samples. These observations emphasize the role of strategically designed combinations of PARPi and immunotherapeutic agents, which could be the key to overcoming immunosuppression in the EOC microenvironment, thereby improving clinical outcomes. Citation Format: Sarka Vosahlikova, Peter Holicek, Irena Moserova, Michal Hensler, Romana Mikyskova, Lenka Kasikova, Josef Pasuvka, Jana Drozenova, Katerina Mojzisova, Marek Kovar, Iain McNiesh, Michael Halaska, Lukas Rob, Sandra Orsulic, Milan Reinis, Lorenzo Galluzzi, Radek Spisek, Jitka Palich Fucikova. PARP inhibitors as immune modulators in metastatic ovarian cancer treatment [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr C046.

Abstract C034: Investigating residual disease in its spatial context in BRCA1 p53-deficient mammary tumors

Abstract Although various effective anti-cancer treatments have become available over the last decades, resistance to all available therapies remains the major cause of death of cancer patients with disseminated tumors. Striking examples are patients with triple-negative breast cancer (TNBC), which are frequently defective in the repair of DNA double strand breaks, e.g. due to loss of BRCA1 function. Because of this defect, the patients initially respond very well to DNA damage-inducing chemotherapy. Unfortunately, disseminated tumors are usually not eradicated and resistant tumor cells are eventually selected from residual primary or metastatic tumor sites. It is therefore crucial to understand the molecular mechanisms underlying the drug tolerance of the residual tumor cells. To study residual disease, we used the K14cre;Brca1 F/F ;p53 F/F (KB1P) mouse model for hereditary breast cancer, which provides the unique opportunity to explore and target those mechanisms in a fully immunocompetent model. The mammary tumors that spontaneously develop highly resemble their human counterparts, both morphologically and in their therapy response. For example, tumors shrink in response to poly(ADP-ribose) polymerase inhibition, platinum-based treatment or the commonly used doxorubicin, docetaxel and cyclophosphamide (TAC) combination therapy. But despite repeated drug sensitivity, the KB1P mammary tumors are not eradicated, not even by a frequent dosing schedule. By combining single-cell RNA sequencing, spatial transcriptomics and imaging mass cytometry, we dissected the intratumoral heterogeneity and alterations in the tumor microenvironment of residual disease. We identified specific subpopulations of tumor cells that have a survival benefit after treatment, and these occurred together with an altered microenvironment characterized by a highly reactive stroma infiltrated with both anti-inflammatory and pro-tumoral immune cells. Interestingly, those structural and transcriptional changes are reversed in the relapsed tumors, highlighting the plasticity of drug tolerance. To investigate the mechanisms of the altered tumor-stroma interactions that are relevant for drug tolerance, we have designed a custom-made CRISPR/Cas9 library based on differential gene expression of the residual tumor cells. This library enables us to functionally test relevant mechanisms of drug tolerance in vivo and we expect that these analyses will provide useful insights into the tumor-stroma interactions that contribute to residual disease. Taken together, our detailed analyses demonstrate the substantial remodeling of tumor cells in their microenvironment upon treatment. To develop new therapeutic approaches to eradicate drug-tolerant tumor cells and thereby circumvent tumor relapse, it is essential to better understand the heterogeneous cellular composition of residual tumors in its spatial context. With this project, we hope to provide comprehensive data to develop better therapeutic strategies that target the tumor-stroma interaction and eradicate residual tumors. Citation Format: Morgane Decollogny, Demeter Túrós, Astrid Chanfon, Myriam Siffert, Ismar Klebic, Sven Rottenberg. Investigating residual disease in its spatial context in BRCA1 p53-deficient mammary tumors [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr C034.

Germline BRCA1/2 status and chemotherapy response score in high-grade serous ovarian cancer.

High-grade serous ovarian cancer (HGSOC) can be treated with platinum-based neoadjuvant chemotherapy (NACT) and delayed primary surgery (DPS). Histopathological response to NACT can be assessed using Böhm's chemotherapy response score (CRS). We investigated whether germline BRCA1/2 (gBRCA1/2) genotype associated with omental CRS phenotype. A retrospective study of patients with newly diagnosed FIGO stage IIIC/IV HGSOC prescribed NACT and tested for gBRCA1/2 pathogenic variants (PVs) between September 2017 and December 2022 at The Christie Hospital. The Cox proportional hazards model evaluated the association between survival and key clinical factors. The chi-square test assessed the association between CRS3 (no/minimal residual tumour) and gBRCA1/2 status. Of 586 eligible patients, 393 underwent DPS and had a CRSreported. Independent prognostic factors by multivariable analysis were gBRCA1/2 status (PV versus wild type [WT]), CRS (3 versus 1 + 2), surgical outcome (complete versus optimal/suboptimal) and first-line poly (ADP-ribose) polymerase-1/2 inhibitor maintenance therapy (yes versus no) (all P < 0.05). There was a non-significant trend for tumours with a gBRCA2 PV having CRS3 versus WT (odds ratio [OR] = 2.13, 95% confidence intervals [CI] 0.95-4.91; P = 0.0647). By contrast, tumours with a gBRCA1 PV were significantly less likely to have CRS3 than WT (OR = 0.35, 95%CI 0.14-0.91; P = 0.0291). Germline BRCA1/2 genotype was not clearly associated with superior omental CRS. Further research is required to understand how HGSOC biology defines CRS.

Hyperthermic intraperitoneal chemotherapy for recurrent ovarian cancer (CHIPOR): a randomised, open-label, phase 3 trial.

Hyperthermic intraperitoneal chemotherapy (HIPEC) at interval cytoreductive surgery for ovarian cancer improves overall survival but its role in recurrent disease is uncertain. We aimed to compare outcomes in patients treated with or without HIPEC during surgery for recurrent ovarian cancer. The multicentre, open-label, randomised, phase 3 CHIPOR trial was conducted at 31 sites in France, Belgium, Spain, and Canada, and enrolled patients with first relapse of epithelial ovarian cancer at least 6 months after completing platinum-based chemotherapy. Eligible patients were aged 18 years or older with WHO performance status of less than 2. After six cycles of platinum-based chemotherapy (and optional bevacizumab), patients amenable to complete cytoreductive surgery were randomly assigned centrally in a 1:1 ratio, using a web-based system and a minimisation procedure, during surgery to receive HIPEC (cisplatin 75 mg/m2 in 2 L/m2 of serum at 41±1°C for 60 min) or not, stratified by centre, completeness of cytoreduction score, platinum-free interval, and latterly, planned poly(ADP-ribose) polymerase inhibitor use. The primary endpoint was overall survival, analysed on an intention-to-treat basis in all randomly assigned patients. This ongoing trial is registered with ClinicalTrials.gov, NCT01376752. Between May 11, 2011, and May 14, 2021, 415 female patients were randomly assigned (207 HIPEC, 208 no HIPEC). At the primary analysis (median follow-up 6·2 years, IQR 4·1-8·1), 268 (65%) patients had died (126 [61%] of 207 in the HIPEC group; 142 [68%] of 208 in the no-HIPEC group). Overall survival was significantly improved with HIPEC (stratified hazard ratio 0·73, 95% CI 0·56-0·96; p=0·024). Median overall survival was 54·3 months (95% CI 41·9-61·7) with HIPEC versus 45·8 months (38·9-54·2) without. Grade 3 or worse adverse events within 60 days after surgery occurred in 102 (49%) of 207 patients receiving HIPEC versus 56 (27%) of 208 receiving no HIPEC, the most common being anaemia (47 [23%] vs 30 [14%]), hepatotoxicity (23 [11%] vs 18 [9%]), electrolyte disturbance (28 [14%] vs two [1%]), and renal failure (20 [10%] vs three [1%]). There were three deaths within 60 days of surgery, all in the no-HIPEC group. Adding HIPEC to cytoreductive surgery after response to platinum-based chemotherapy at first epithelial ovarian cancer recurrence significantly improved overall survival. When treating patients with late first relapse of high-grade serous or high-grade endometrioid ovarian cancer amenable to complete cytoreductive surgery at specialist centres, platinum-based HIPEC should be considered to extend overall survival. French National Cancer Institute and French League Against Cancer.

Investigating the Structure of the Components of the PolyADP-Ribosylation System in Fusarium Fungi and Evaluating the Expression Dynamics of Its Key Genes

Poly(ADP-ribose) polymerase (PARP) is the key enzyme in polyADP-ribosylation, one of the main post-translational modifications. This enzyme is abundant in eukaryotic organisms. However, information on the PARP structure and its functions in members of the Fungi kingdom is very limited. In this study, we performed a bioinformatic search for homologs of PARP and its antagonist, PARG, in the genomes of four Fusarium strains using their whole-genome sequences annotated and deposited in databases. The F. graminearum PH-1, F. proliferatum ET-1, and F. oxysporum Fo47 strains were shown to possess a single homolog of both PARP and PARG. In addition, the F. oxysporum f. sp. lycopersici strain 4287 contained four additional proteins comprising PARP catalytic domains whose structure was different from that of the remaining identified homologs. Partial nucleotide sequences encoding the catalytic domains of the PARP and PARG homologs were determined in 11 strains of 9 Fusarium species deposited in all-Russian collections, and the phylogenetic properties of the analyzed genes were evaluated. In the toxigenic F. graminearum strain, we demonstrated up-regulation of the gene encoding the PARP homolog upon culturing under conditions stimulating the production of the DON mycotoxin, as well as up-regulation of the gene encoding PARG at later stages of growth. These findings indirectly indicate involvement of the polyADP-ribosylation system in the regulation of the genes responsible for DON biosynthesis.

Metabolically inducing defects in DNA repair sensitizes BRCA-wild-type cancer cells to replication stress.

Metabolic reprogramming from oxidative respiration to glycolysis is generally considered to be advantageous for tumor initiation and progression. However, we found that breast cancer cells forced to perform glycolysis acquired a vulnerability to PARP inhibitors. Small-molecule inhibition of mitochondrial respiration-using glyceollin I, metformin, or phenformin-induced overproduction of the oncometabolite lactate, which acidified the extracellular milieu and repressed the expression of homologous recombination (HR)-associated DNA repair genes. These serial events created so-called "BRCAness," in which cells exhibit an HR deficiency phenotype despite lacking germline mutations in HR genes such as BRCA1 and BRCA2, and, thus, sensitized the cancer cells to clinically available poly(ADP-ribose) polymerase inhibitors. The increase in lactate repressed HR-associated gene expression by decreasing histone acetylation. These effects were selective to breast cancer cells; normal epithelial cells retained HR proficiency and cell viability. These mechanistic insights into the BRCAness-prone properties of breast cancer cells support the therapeutic utility and cancer cell-specific potential of mitochondria-targeting drugs.

DNAR-12. STEAROYL-COA DESATURASE REGULATES DNA DAMAGE REPAIR IN GBM BY MODULATING PARP1 ACTIVITY

Abstract Glioblastoma (GBM) is the most prevalent malignant brain cancer in adults, notorious for its aggressiveness and resistance to therapy. GBM patients commonly receive radiation therapy combined with temozolomide (TMZ), which engages tumor cells to promote lethal DNA damage and subsequent cytotoxicity. Active DNA repair mechanisms, particularly in a subset population of GBM stem-like cells (GSCs), drive therapeutic resistance, thus largely contributing to the bleak prognosis of GBM. Poly (ADP-ribose) polymerases (PARPs) are essential in various cellular processes, including DNA repair. We have previously reported that fatty acid desaturation mediated by Stearoyl-CoA desaturase 1 (SCD1) plays a cytoprotective role in GSCs. Inhibiting SCD prevents tumor initiation and inhibits DNA damage repair, hence sensitizing to radiation and TMZ. Yet, the mechanistic role of SCD in DNA damage regulation remained unknown. Here, we found that the expression of both human isoforms of SCD, SCD1, and SCD5, directly correlates with PARP protein levels and subcellular localization. Downregulating SCD1/5 in GSCs resulted in significantly lower levels of PARP1, coupled with increased DNA damage, as well as reduced DNA damage repair and chromosome assembly. Conversely, cells overexpressing SCD1/5 exhibited higher PARP1 expression and lower levels of basal DNA damage compared to control GSCs. Notably, SCD1/5 overexpression also led to increased resistance to TMZ. We show that SCD1/5 regulates PARP activity by controlling PARP1 subcellular localization. In addition to decreased total levels of PARP following SCD1/5 downregulation, we also found increased PARP1 self-PARylation. Using confocal microscopy, we show that when SCD1/5 is repressed, PARylated PARP translocates from the nucleus to the cytosol, rendering it unable to bind DNA and recruit DNA-repairing machinery. In conclusion, our study provides a direct link between fatty acid metabolism and PARP1-mediated DNA damage repair.

Olaparib-induced myelodysplasia – A case report

Olaparib is an antineoplastic agent that is a poly ADP ribose polymerase (PARP) inhibitor and is Food and Drug Administration-approved for the treatment of ovarian, breast, pancreatic, and prostate cancer. PARP inhibitors can cause DNA damage and epigenetic changes, leading to hematological transformations and increasing the risk of myelodysplasia. We present a case of a 58-year-old female with malignant breast cancer initiated on Olaparib after failing multiple treatment lines. The patient had persistent declining blood cell counts, and a bone marrow biopsy revealed hypercellular bone marrow with predominant myeloid cells with increasing number of blasts and promyelocytes. Olaparib was stopped. The patient elected for supportive care and hospice.

CTNI-50. CLINICAL EFFICACY AND PK/PD RESPONSE IN A PHASE 0/2 STUDY OF NIRAPARIB PLUS RADIOTHERAPY FOR NEWLY-DIAGNOSED, MGMT-UNMETHYLATED GLIOBLASTOMA

Abstract BACKGROUND Poly (ADP-ribose) polymerase (PARP) mediates DNA damage response; niraparib is a PARP1/2-selective inhibitor. This Phase 0/2 study evaluates tumor pharmacokinetics (PK), pharmacodynamics (PD), and clinical efficacy of niraparib plus fractionated radiotherapy in newly diagnosed, O6-methylguanine methyltransferase (MGMT)-unmethylated glioblastoma (GBM). METHODS Newly-diagnosed GBM patients received 4 days of niraparib (300/200 mg QD) prior to planned resection 3-5 (cohort 1) or 8-10 hours (cohort 2) following the last dose. Tumor tissue (gadolinium-enhancing and non-enhancing regions), cerebrospinal fluid (CSF), and plasma were collected. Total and unbound drug concentrations were measured using validated LC-MS/MS methods. PARP inhibition was assessed by quantification of PAR induction after ex vivo irradiation vs non-irradiated control. A PK response was defined as unbound [niraparib] &amp;gt; 5-fold biochemical IC50 (19 nM) in non-enhancing tumor. Patients with MGMT-unmethylated tumors and a PK response were eligible for Phase 2 dosing of niraparib plus radiotherapy followed by niraparib monotherapy. RESULTS All Phase 0 patients (n=46) met the PK threshold. The mean unbound concentrations of niraparib in non-enhancing tumor regions were 335.1 nM (cohort 1; n=43) and 331.9 nM (cohort 2; n=3). Ex vivo PAR suppression was observed in 73% of the patients (24/33). 71.4% patients with unmethylated tumors were enrolled into Phase 2. Five patients in Phase 2 experienced Grade 4 thrombocytopenia related to niraparib and all adverse events resolved without sequelae. At a mean clinical follow-up of 13.4 months, median overall survival was 20.3 months. CONCLUSIONS Niraparib achieves pharmacologically relevant concentrations in non-enhancing, newly-diagnosed GBM tissue. Accompanying PD effects were observed in tumor tissue, no new safety signals were observed, and clinical outcomes were promising. A global Phase 3, open-label, randomized 2-arm study comparing niraparib versus temozolomide in adult patients with newly diagnosed, MGMT-unmethylated glioblastoma is underway.

CTIM-32. PHASE I/II STUDY OF STEREOTACTIC RADIOSURGERY WITH CONCURRENT OLAPARIB FOLLOWED BY ADJUVANT DURVALUMAB AND PHYSICIAN’S CHOICE SYSTEMIC THERAPY IN SUBJECTS WITH BREAST CANCER BRAIN METASTASES (SOLARA)

Abstract BACKGROUND Despite progress in the treatment of brain metastasis for HER2+ breast cancer, outcomes for patients with HER2-negative breast cancer brain metastases remain poor. Preclinical studies show inhibitors of poly(ADP-ribose) polymerase (PARP) are effective in combination with radiation therapy as a DNA damage response inhibitor. Triple-negative breast cancer (TNBC) has higher rates of homologous recombination deficiency compared to other subtypes, and together with HER2-negative, BRCA-mutated breast cancer would be particularly sensitive to PARP inhibition. PARP inhibition has also demonstrated promising efficacy combined with immunotherapy in patients with germline BRCA-mutant and metastatic TNBC in clinical trials. In addition, immunotherapy with stereotactic radiosurgery (SRS) is associated with favorable intracranial control and survival in patients with brain metastases. We hypothesize that this biologically-driven combination will enhance local control of SRS-treated brain metastases through synergy with PARP inhibition, while controlling micrometastatic disease via potentiation of the immune response. METHODS We are conducting a multi-institution, Phase I/II trial of SRS plus olaparib, followed by durvalumab (with physician’s choice systemic therapy), for patients with TNBC (any BRCA status) or HER2-negative with BRCA-mutated (germline or somatic) breast cancer brain metastases [NCT04711824]. A total of 41 patients are planned for enrollment at 8 sites. The primary objectives are to evaluate safety and tolerability (Phase I) and determine intracranial disease control at 6 months (Phase II). Secondary objectives include assessing intracranial and global progression-free survival, overall survival, and intracranial and extracranial response rate. Exploratory objectives will assess potential biomarkers of treatment response, including changes in circulating tumor cells and DNA in blood and cerebrospinal fluid, germline and tumor mutations in DNA repair pathway genes, and PD-L1 expression, as well as quality of life and patient-reported outcomes. As of May 2024, phase I has been completed without dose-limiting toxicity, and phase II is currently enrolling. Funding/drug provision from AstraZeneca.

DDDR-15. UTILIZING DEEP DOCKING AND ARTIFICIAL INTELLIGENCE FOR THE DISCOVERY OF NOVEL PARP1-SELECTIVE INHIBITORS FOR USE AGAINST BRAIN TUMORS

Abstract Brain tumors, including primary brain tumors and central nervous system (CNS) metastases, remain among the most challenging malignancies to treat, with therapeutic options often limited by the inability of drugs to penetrate the blood-brain barrier. Poly(ADP-ribose) polymerase (PARP) is a key enzyme in DNA repair, and inhibition of PARP1 specifically drives synthetic lethality in BRCA-mutated disease. While they’ve achieved commercial success, first generation PARP inhibitors are limited in their utility as they cannot readily pass through the blood-brain barrier and have adverse side effects, likely driven by the collateral inhibition of PARP2. Development of a PARP1-selective, CNS penetrant inhibitor could reduce toxicity, while providing a new therapeutic option for brain tumors. Traditional drug discovery methods are time-consuming and costly, necessitating innovative approaches. Here, we describe the application of Deep Docking, an advanced artificial intelligence (AI) approach, to discover a novel, PARP1-selective inhibitor for use against brain tumors. Deep Docking utilizes deep learning to accelerate the prediction of the binding affinity of a large number of compounds to target proteins, streamlining the virtual screening process and allowing for rapid docking of billions of compounds against the PARP1 protein. Additionally, state-of-the-art generative algorithms and deep learning techniques for predicting drug-like properties such as metabolism, permeability, and safety profiles, can be combined to rapidly perform hit-to-lead optimization. We will present preliminary Deep Docking screening results from billions of compounds, identifying several with predicted high binding affinities for PARP-1. Validating in vitro and in vivo data, including PARP1 selectivity, metabolic stability, pharmacokinetic profile, CNS penetration and safety profile, will be described. Application of the Deep Docking AI platform is being used to significantly accelerate the discovery of a selective PARP-1 inhibitor for brain tumors. This approach will not only improve the efficiency of drug discovery but also enhance the specificity and efficacy of potential therapeutics.

A NETWORK PHARMACOLOGY-BASED DRUG REPURPOSING STUDY OF LEVETIRACETAM UNCOVERS ITS INTERACTION WITH MULTI-DRUG TARGETS IN PARKINSON'S DISEASE

Objective: The current study utilized network pharmacology to examine how Levetiracetam interacts with specific drug targets associated with Parkinson's Disease (PD) treatment. Methods: We used information from Kyoto Encyclopedia of Genes and Genome (KEGG) studies and Protein-Protein Interaction (PPI) pathway analysis to create a network that depicts the relationships between Levetiracetam and PD targets. Further investigation involved PPI analysis, molecular docking, and Molecular Dynamics (MD) simulation studies, ultimately pinpointing five protein targets. Their participation in pathways such as Ribonucleic acid Polymerase II-specific Deoxyribonucleic acid binding Transcription Factor Binding (Gene Ontology [GO]:0061629), Axon (GO: 0030424), and Excitatory Postsynaptic Potential was emphasized by GO and KEGG pathway enrichment. Additionally, Dopamine Receptor D2 (DRD2), Solute Carrier Family 6 Member 3 (SLC6A3), Glycogen Synthase Kinase 3 Beta (GSK3B), Poly (ADP-ribose) Polymerase 1 (PARP1) and Myeloperoxidase (MPO) were identified as protein targets through PPI and molecular docking analysis. Results: The results of molecular docking showed that protein targets, SLC6A3, have highest binding affinity with Levetiracetam. The MD Simulation result of Levetiracetam-SLC6A3 docked complex represented the complex to be quite stable with few conformational changes in the SLC6A3 structure. DRD2, SLC6A3, GSK3B, PARP1, MPO were recognized as the likely protein targets of Levetiracetam for treating PD. SLC6A3 was considered as a target of Levetiracetam in PD. Conclusion: Our study revealed the mechanism of Levetiracetam in the treatment of PD and can contribute to more effective treatment for the same. By identifying key protein targets, this research lays the groundwork for future studies that could further explore Levetiracetam’s efficacy.

The radiosensitivity of niraparib tosylate to esophageal squamous cell carcinoma is stronger under hypoxia and higher-pressure conditions

Background: Radiotherapy is an effective method for esophageal squamous cell carcinoma (ESCC) treatment show reduced effectiveness in long-term due to reduced sensitivity of cancer cells to radiotherapy. Niraparib tosylate, a poly ADP-ribose polymerase (PARP) inhibitor may enhance the radiosensitivity of these cells. Hence, the present study was aimed to study the effects of niraparib tosylate on ESCC proliferation, apoptosis and cell cycle along with radiosensitivity enhancement efficiency. Materials and methods: ESCC cell lines, KYSE-30 were subjected to niraparib tosylate or irradiation treatments or combination of both. CCK8 and colony formation assays were performed to monitor cell growth status. Cell cycle and apoptosis level were investigated through flow cytometry. The expressions of targeted genes were detected at protein levels by western blot. Besides, xenografts were successfully established in immunocompromised mice. Student’s t-test was used to compare data and P &lt; 0.05 was considered statistically significant. Results: Treatment with niraparib tosylate showed G2/M cycle arrest and apoptosis in KYSE-30 cells and the rate was observed to be more significant with combination therapy. These effects were further strengthened when cells were cultured under hypoxia and high atmospheric pressure. An enhanced susceptibility to radiotherapy was observed with niraparib tosylate combination. The homologous recombination related regulator, Rad51, showed remarkable upregulation in cancer cells exposed to combination of drug-radiation treatment. The tumor burden was also relieved in xenograft mouse models with combination of drug-radiation treatment, indicating the potential of niraparib tosylate use clinical cancer therapy. Conclusion: From the results, a novel function of niraparib tosylate can be established, impairing DNA damage repair efficiency to increase the susceptibility of ESCC to radiotherapy, resulting in cell apoptosis and G2/M cycle arrest in vitro and in vivo.

DNA damage-induced EMT controlled by the PARP-dependent chromatin remodeler ALC1 promotes DNA repair efficiency through RAD51 in tumor cells.

Epithelial-to-mesenchymal transition (EMT) allows cancer cells to metastasize while acquiring resistance to apoptosis and chemotherapeutic agents with significant implications for patients' prognosis and survival. Despite its clinical relevance, the mechanisms initiating EMT during cancer progression remain poorly understood. We demonstrate that DNA damage triggers EMT and that activation of poly (ADP-ribose) polymerase (PARP) and the PARP-dependent chromatin remodeler ALC1 (CHD1L) was required for this response. Our results suggest that this activation directly facilitates access to the chromatin of EMT transcriptional factors (TFs) which then initiate cell reprogramming. We also show that EMT-TFs bind to the RAD51 promoter to stimulate its expression and to promote DNA repair by homologous recombination. Importantly, a clinically relevant PARP inhibitor reversed or prevented EMT in response to DNA damage while resensitizing tumor cells to other genotoxic agents. Overall, our observations shed light on the intricate relationship between EMT, DNA damage response, and PARP inhibitors, providing potential insights for in cancer therapeutics.

PARP1 condensates differentially partition DNA repair proteins and enhance DNA ligation.

Poly(ADP-ribose) polymerase 1 (PARP1) is one of the first responders to DNA damage and plays crucial roles in recruiting DNA repair proteins through its activity - poly(ADP-ribosyl)ation (PARylation). The enrichment of DNA repair proteins at sites of DNA damage has been described as the formation of a biomolecular condensate. However, it remains unclear how exactly PARP1 and PARylation contribute to the formation and organization of DNA repair condensates. Using recombinant human single-strand repair proteins in vitro, we find that PARP1 readily forms viscous biomolecular condensates in a DNA-dependent manner and that this depends on its three zinc finger (ZnF) domains. PARylation enhances PARP1 condensation in a PAR chain length-dependent manner and increases the internal dynamics of PARP1 condensates. DNA and single-strand break repair proteins XRCC1, LigIII, Polβ, and FUS partition in PARP1 condensates, although in different patterns. While Polβ and FUS are both homogeneously mixed within PARP1 condensates, FUS enrichment is greatly enhanced upon PARylation whereas Polβ partitioning is not. XRCC1 and LigIII display an inhomogeneous organization within PARP1 condensates; their enrichment in these multiphase condensates is enhanced by PARylation. Functionally, PARP1 condensates concentrate short DNA fragments, which correlates with PARP1 clusters compacting long DNA and bridging DNA ends. Furthermore, the presence of PARP1 condensates significantly promotes DNA ligation upon PARylation. These findings provide insight into how PARP1 condensation and PARylation regulate the assembly and biochemical activities of DNA repair factors, which may inform on how PARPs function in DNA repair foci and other PAR-driven condensates in cells.

Screening and design of PARP12 inhibitors from traditional Chinese medicine small molecules using computational modeling and simulation

The poly (ADP-ribose) polymerase (PARP) family of enzymes plays a pivotal role in orchestrating a multitude of cellular processes, including DNA repair mechanisms, transcriptional regulation, and modulation of immune responses. Within this family, PARP12 emerges as a noteworthy candidate for targeted cancer therapeutics. Consequently, this investigation endeavors to screen and design potential PARP12 inhibitors derived from traditional Chinese medicinal compounds by employing sophisticated molecular modeling and computational medicinal chemistry approaches. The compound RBN2397 is utilized as a benchmark, and the binding efficacies of the newly identified small molecules are assessed against a spectrum of criteria, encompassing molecular interactions, binding free energy, and extensive post-simulation analyses. The outcomes demonstrated that the identified small molecules, specifically tcm8650 and its derivative XC-1, possess remarkable binding affinities and exhibit reduced binding free energies compared to RBN2397. The molecular docking and interaction profiles of these compounds were also comprehensively scrutinized. Moreover, ADMET profiling meticulously evaluated the pharmacokinetic profiles and physicochemical characteristics of these promising molecules and their projected human physiological impact. These computational studies indicated their potential therapeutic applicability and predicted acceptable safety profile, advocating their further exploration as viable candidates in cancer treatment.

Resveratrol Inhibits Nucleosome Binding and Catalytic Activity of PARP1

The natural polyphenol resveratrol is a biologically active compound that interacts with DNA and affects the activity of some nuclear enzymes. Its effect on the interaction between nucleosomes and poly(ADP-ribose) polymerase-1 (PARP1) and on the catalytic activity of PARP1 was studied using Western blotting, spectrophotometry, electrophoretic mobility shift assay, and single particle Förster resonance energy transfer microscopy. Resveratrol inhibited PARP1 activity at micro- and sub-micromolar concentrations, but the inhibitory effect decreased at higher concentrations due to the aggregation of the polyphenol. The inhibition of PARP1 by resveratrol was accompanied by its binding to the enzyme catalytic center and a subsequent decrease in PARP1 affinity to nucleosomal DNA. Concurrent binding of talazoparib to the substrate binding pocket of PARP1, which occurs in the presence of resveratrol, restores the interaction of PARP1 with nucleosomes, suggesting that the binding sites of resveratrol and talazoparib overlap. The data suggest that resveratrol can be classified as a natural inhibitor of PARP1.

The combination of venetoclax and quercetin exerts a cytotoxic effect on acute myeloid leukemia

Venetoclax is a BH3 mimetic that was recently approved for the treatment of acute myeloid leukemia (AML) treatment. However, the effect of venetoclax on AML remains limited, and a novel strategy is required. Here, we demonstrate for the first time that the cytotoxic effect of venetoclax drastically increased when by combined with the naturally occurring flavonoid quercetin. Combined treatment with venetoclax and quercetin caused most of AML KG-1 cells to exhibit a condensed morphology. Cell cycle analysis revealed that the combination strongly induced cell death. Caspase inhibitor blocked this cell death, and the combination induced poly (ADP-ribose) polymerase (PARP) cleavage, indicating that apoptosis was the primary mechanism. These effects were also observed in another AML cell line Kasumi-1 but not in chronic myeloid leukemia (CML) K562 cells. Public data analysis demonstrated that B-cell/CLL lymphoma 2 (Bcl-2) expression is increased in AML cells compared to other malignant tumors, and the survival and the growth of AML cell line depends on Bcl-2. We found that quercetin increased Bcl-2-associated X protein (Bax) expression in KG-1. Our study provides a novel function for quercetin and presents a promising strategy for AML treatment using venetoclax.

PARP7i Clinical Candidate RBN-2397 Exerts Antiviral Activity by Modulating Interferon-β Associated Innate Immune Response in Macrophages.

Polyadenosine diphosphate-ribose polymerase 7 (PARP7) acts as a suppressor of the type I interferon (IFN) signaling pathway via suppressing TANK-binding protein 1 (TBK1). Research study indicates that inhibition of PARP7 could potentially regulate tumor immunity. However, the effect of PARP7 inhibition on innate antiviral immunity in macrophages as well as the underlying mechanism have not been demonstrated else well. We report herein that PARP7 inhibitor clinical candidate RBN-2397 could augment type I interferon (IFN-I) production in macrophages by elevating retinoic acid-inducible gene I (RIG-I) and stimulator of interferon genes (STING) signaling pathways. Treatment with RBN-2397 leads to increased pattern recognition ligands-induced interferon-β production in primary bone marrow-derived macrophages (BMDM) and RAW264.7 cells. Additionally, RBN-2397 suppresses viral replication efficiency in macrophages infected by vesicular stomatitis virus (VSV) and amplifies the expression of interferon-stimulated chemokine genes (ISGs). Mechanistically, RBN-2397 promotes TBK1 phosphorylation, consequently leading to the amplified activation of RIG-I and STING signaling pathways. Furthermore, RBN-2397 enhances the phosphorylation of signal transducer and activator of transcription 1 (STAT1) and STAT2 induced by IFN-α/β and the expression of chemokine genes in macrophages in response to IFN stimulation. In vivo experiments demonstrated that RBN-2397 enhances innate antiviral immunity in mice infected with VSV, resulting in increased serum IFN-β levels, reduced viral loads, and alleviated pulmonary inflammatory responses of the VSV-infected mice. In conclusion, our findings highlight the potential of RBN-2397 as a promising antiviral therapeutic agent for enhancing the IFN-relative antiviral immune defense in host.