ADP-ribose Polymerase Research Articles

Saponins Derived from the Korean Endemic Plant Heloniopsis koreana Inhibit Diffuse-type Gastric Cancer Cells.

Diffuse-type gastric cancer (GC) is a type of stomach cancer that occurs in small clusters of cells that are widely spread. It does not typically manifest with symptoms until the advanced stages and often goes undetected in routine imaging tests. In addition, there is no specific targeted therapy for diffuse-type GC and it has a high mortality risk. Hence, it is worthwhile to discover molecules against this cancer. In this study, the extract of Heloniopsis koreana, which is endemic to Korea, exhibited cytotoxicity against two diffuse-type GC cell lines, MKN1 and SNU668. This led to the isolation of 10 compounds, including a new cinnamic acid glycoside. Of the compounds, saponin Th (4) and SNF 11 (5) showed potent activities with IC50 values of 3.66 and 3.85 μM, respectively, in MKN1 cells, and 1.8 and 1.98 μM, respectively, in SNU668 cells. These compounds prevented cancer cell division, invasion, and colony formation in both cell lines. In addition, these compounds induced cancer cell death through conventional cell death pathways, showing an increase in ADP-ribose polymerase, caspase 3, and BAX and a decrease in BCL2.

Regulation of radiation-induced liver damage by modulation of SIRT-1 activity: In vivo rat model.

Silent information regulator 1 (SIRT-1), a nicotinamide adenine dinucleotide-dependent deacetylase, was found to regulate cell apoptosis, inflammation, and oxidative stress response in living organisms. Therefore, the role of SIRT-1 in regulating forkhead box O/poly ADP-ribose polymerase-1 (FOXO-1/PARP-1) signaling could provide the necessary validation for developing new pharmacological targets for the promotion or inhibition of SIRT-1 activity toward radiation sensitivity. In the present study, the SIRT-1 signaling pathway is being investigated to study the possible modulatory effect of resveratrol (RSV, SIRT-1 activator) versus nicotinamide (NAM, SIRT-1 inhibitor) in case of liver damage induced by whole-body gamma irradiation. Rats were exposed to 6 Gy gamma radiation after being pretreated with either RSV (10 mg/kg/day) or NAM (100 mg/kg/day) for 5 days, and subsequent examining hepatic morphological changes and apoptotic markers were assessed. The expression of SIRT-1, FOXO-1, and cleaved PARP-1 in the liver was analyzed. RSV improved radiation-induced apoptosis, mitochondrial dysfunction, and inflammation signified by low expression of caspase-3, lactate dehydrogenase, complex-I activity, myeloperoxidase, and total nitric oxide content. RSV increased the expression of SIRT-1, whereas cleaved PARP-1 and FOXO-1 were suppressed. These protective effects were suppressed by inhibition of SIRT-1 activity using NAM. These findings suggest that RSV can attenuate radiation-induced hepatic injury by reducing apoptosis and inflammation via SIRT-1 activity modulation.

Tumor elimination by clustered microRNAs miR-306 and miR-79 via noncanonical activation of JNK signaling.

JNK signaling plays a critical role in both tumor promotion and tumor suppression. Here, we identified clustered microRNAs (miRNAs) miR-306 and miR-79 as novel tumor-suppressor miRNAs that specifically eliminate JNK-activated tumors in <i>Drosophila</i>. While showing only a slight effect on normal tissue growth, miR-306 and miR-79 strongly suppressed growth of multiple tumor models, including malignant tumors caused by Ras activation and cell polarity defects. Mechanistically, these miRNAs commonly target the mRNA of an E3 ubiquitin ligase ring finger protein 146 (RNF146). We found that RNF146 promotes degradation of tankyrase (Tnks), an ADP-ribose polymerase that promotes JNK activation in a noncanonical manner. Thus, downregulation of RNF146 by miR-306 and miR-79 leads to hyper-enhancement of JNK activation. Our data show that, while JNK activity is essential for tumor growth, elevation of miR-306 or miR-79 overactivate JNK signaling to the lethal level via noncanonical JNK pathway and thus eliminate tumors, providing a new miRNA-based strategy against cancer.

Poly (ADP-ribose) polymerase inhibitors (PARPi) for advanced malignancies with multiple DNA-repair genetic aberrations

ABSTRACT Introduction Poly (ADP-ribose) polymerase inhibitors (PARPi) have been approved for the treatment of advanced tumors with defects in genes involved in homologous recombination repair (HRR), including cancers of the prostate, pancreas, breast, and ovary. In these advanced tumors, PARPi afford ‘synthetic lethality’ by blocking the PARP-associated repair pathway in cancer cells with HRR genetic mutations, resulting in chromosome instability and cellular apoptosis. According to the synthetic lethality theory, patients with a greater burden of genetic alterations, in proportion (relative quantity) or category, would have more satisfactory outcomes after PARPi administration. However, this issue remains obscure based on the existing sporadic evidence. Areas covered We summarize the therapeutic effects of PARPi in advanced tumors with multiple HRR genetic mutations, and attempted to compare these results with those obtained for cancers with a single mutation. Expert opinion Limited evidence has provided a possibly encouraging response to PARPi among patients carrying multiple HRR genetic mutations compared with those with a single mutation (although the treatment effect was negative in some patients). Further research is needed to understand the role of PARPi in tumor cells with multiple HRR genetic mutations.

Combined inhibition of BADSer99 phosphorylation and PARP ablates models of recurrent ovarian carcinoma

BackgroundPoly (ADP-ribose) polymerase inhibitors (PARPis) have been approved for the treatment of recurrent epithelial ovarian cancer (EOC), regardless of BRCA status or homologous recombination repair deficiency. However, the low response of platinum-resistant EOC, the emergence of resistance in BRCA-deficient cancer, and therapy-associated toxicities in patients limit the clinical utility of PARPis in recurrent EOC.MethodsThe association of phosphorylated (p) BADS99 with clinicopathological parameters and survival outcomes in an EOC cohort was assessed by immunohistochemistry. The therapeutic synergy, and mechanisms thereof, between a pBADS99 inhibitor and PARPis in EOC was determined in vitro and in vivo using cell line and patient-derived models.ResultsA positive correlation between pBADS99 in EOC with higher disease stage and poorer survival is observed. Increased pBADS99 in EOC cells is significantly associated with BRCA-deficiency and decreased Cisplatin or Olaparib sensitivity. Pharmacological inhibition of pBADS99 synergizes with PARPis to enhance PARPi IC50 and decreases survival, foci formation, and growth in ex vivo culture of EOC cells and patient-derived organoids (PDOs). Combined inhibition of pBADS99 and PARP in EOC cells or PDOs enhances DNA damage but impairs PARPi stimulated DNA repair with a consequent increase in apoptosis. Inhibition of BADS99 phosphorylation synergizes with Olaparib to suppress the xenograft growth of platinum-sensitive and resistant EOC. Combined pBADS99-PARP inhibition produces a complete response in a PDX derived from a patient with metastatic and chemoresistant EOC.ConclusionsA rational and efficacious combination strategy involving combined inhibition of pBADS99 and PARP for the treatment of recurrent EOC is presented.

SAT319 - The crucial role of PARP [poly (ADP-ribose polymerase] on the post-ischemic liver injury and inflammation

SAT319 - The crucial role of PARP [poly (ADP-ribose polymerase] on the post-ischemic liver injury and inflammation

DNA Repair Enzymes Promising in Cancer Therapy; Poly (ADP-Ribose) Polymerase Inhibitors

DNA Repair Enzymes Promising in Cancer Therapy; Poly (ADP-Ribose) Polymerase Inhibitors

Abstract 5296: Poly(ADP-ribose) polymerase-1 expression is a good survival marker in pancreatic cancers, unlike other solid cancers

Abstract Purpose: Poly(ADP-ribose) Polymerases-1 (PARP-1) is well known as poor survival factor in various solid cancers and its inhibitors have shown promising results in clinical trials. However, in pancreatic ductal adenocarcinoma (PDAC), there is only one study with good prognostic result. We assessed the prognostic significance of PARP-1 in PDAC Methods: From 1998 to 2020, 194 patients with radical surgery for PDAC were enrolled in Yeouido St. Mary’s hospital (n = 84) and Uijeongbu St. Mary’s hospital (n=110). After making tumor microarray, PARP-1 immunohistochemistry was conducted and we analyzed H score with image analyzer (QuantCenter, 3DHistech, Budapest, Hungary). Results: High expression of PARP1 (OS: p = 0.004, CSS: p = 0.007, DFS: p &amp;lt; 0.001) was good survival factor in univariate analysis. Old age (OS: p = 0.018, CSS: p = 0.041, DFS: p = 0.050 ), high stage (OS: p = 0.004, CSS: p = 0.009, DFS: p = 0.817), lymphatic invasion (OS: p &amp;lt; 0.001, CSS: p &amp;lt; 0.001, DFS: p = 0.001), venous invasion (OS: p &amp;lt; 0.001, CSS: p &amp;lt; 0.001, DFS: p = 0.109), perineural invasion (OS: p &amp;lt; 0.001, CSS: p &amp;lt; 0.001, DFS: p &amp;lt; 0.001) and high grade tumor differentiation (OS: p = 0.005, CSS: p = 0.003, DFS: p = 0.001 ) was poor survival factor in log rank test. In multivariate analysis, PAPR1 was independent good survival factor (OS: p = 0.002, CSS: p = 0.005, DFS: p &amp;lt; 0.001). Conclusion: Our study revealed that PARP1 expression was good survival factor in PDAC unlike other solid cancers. In our observation, studies with larger datasets or on the mechanism of PARP1 in PDAC are necessary in order to use PARP inhibitor in PDAC. Citation Format: Kwangil Yim, Nishant Thakur, Mohammad Rizwan Alam, Jamshid Abdul-Ghafar, Yosep Chong, Kyung Jin Seo, Ok Ran Shin. Poly(ADP-ribose) polymerase-1 expression is a good survival marker in pancreatic cancers, unlike other solid cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5296.

Abstract CT170: A phase 1/2 study of olaparib in combination with ramucirumab in metastatic gastric and gastroesophageal junction adenocarcinoma (GEJ)

Abstract Background: Ramucirumab is a monoclonal antibody that binds to the vascular endothelial growth factor receptor 2, increasing hypoxia in the tumor microenvironment. In advanced gastric cancer, single agent ramucirumab is well tolerated and improves overall survival (OS) compared to placebo although it is largely cytostatic with a response rate ≤3%. Preclinical in vitro and in vivo experiments reveal that hypoxia induces homologous recombination deficiency (HRD) in tumors, which may be exploited by the addition of inhibitors of poly (ADP-ribose) polymerase (PARPi). We thus designed NCI10066 for metastatic gastric cancer with the hypothesis that ramucirumab would sensitize tumors to the PARPi olaparib. Methods: NCI 10066 (NCT03008278) was a phase 1/2, multicenter, open-label clinical trial sponsored by the Cancer Therapy Evaluation Program. Eligible patients had histologically confirmed metastatic gastric or GEJ adenocarcinoma with measurable disease refractory to ≥ 1 prior therapy that did not contain ramucirumab. Patients were treated with escalating doses of olaparib with ramucirumab 8 mg/kg every 2 weeks in a 3+3 design. Dose escalation evaluated two dose levels: olaparib 200 mg twice daily with ramucirumab 8 mg/kg every 2 weeks (dose level 1) and olaparib 300 mg twice daily with ramucirumab 8 mg/kg every 2 weeks (dose level 2). The primary objective of dose escalation was to assess safety and measure dose limiting toxicities (DLT) to determine the recommended phase 2 dose (RP2D). The primary objective in phase 2 was to evaluate clinical activity at the RP2D in a 40-patient expansion cohort. Results: From 2/8/18 - 9/17/21, 51 patients were enrolled at 10 centers throughout the Experimental Therapeutics Clinical Trials Network and 28/51 (55%) patients had ≥2 prior lines of therapy. In dose escalation 3 patients were treated at dose level 1 and 8 at dose level 2. No DLTs were observed at dose level 1 and at dose level 2 there was 1 DLT (fatigue) in 6 evaluable patients. The RP2D was determined to be olaparib 300 mg twice daily and ramucirumab 8 mg/kg every 2 weeks. Forty patients were treated in dose expansion at the RP2D. Of 46 patients evaluable for a response there were 5/46 (11%) partial responses (PR) and the disease control rate (PR/SD) was 29/46 (63%) at 6 weeks. The median progression free survival (PFS) was 2.8 months (95% CI 2.3 - 4.2) and the median OS was 7.3 months (95% CI 5.6 - 13.0). Grade ≥ 3 treatment related adverse events were seen in 13 (25%) of patients and there were no treatment related deaths. Conclusions: The combination of olaparib and ramucirumab is well tolerated with a RP2D of olaparib 300 mg twice daily and ramucirumab 8 mg/kg every 2 weeks. The ORR, PFS, and OS exceed the historical control with ramucirumab monotherapy, suggesting that the combination is clinically active. Biomarkers are critical to defining patient subsets more likely to benefit from the combination and HRD biomarker analysis is ongoing. Citation Format: Michael Cecchini, Joseph Chao, Yu Shyr, James Cleary, Nataliya Uboha, May Cho, Anthony Shields, Shubham Pant, Laura Goff, Kristin Spencer, Edward Kim, Chih-Yuan Hsu, Stacey Stein, Jaykumar Thumar, Jeremy Kortmansky, John Kunstman, Patricia LoRusso, Percy Ivy, Jill Lacy. A phase 1/2 study of olaparib in combination with ramucirumab in metastatic gastric and gastroesophageal junction adenocarcinoma (GEJ) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr CT170.

Abstract 804: Trapped PARP1 cytotoxicity is modulated by the ubiquitin-dependentsegregase p97

Abstract Poly-(ADP-ribose) polymerase inhibitors (PARPi) elicit anti-tumor activity in homologous recombination defective cancers by promoting cytotoxic, chromatin-bound, “trapped” PARP1. Here we have elucidated a pathway that process trapped PARP1. By exploiting wild-type or trapping-resistant PARP1 transgenes combined with either a rapid immunoprecipitation mass-spectrometry of endogenous proteins (RIME)-based approach or PARP1 Apex2-proximity labelling linked to mass-spectrometry, we generated proteomic profiles of trapped and non-trapped PARP1 complexes. These experiments demonstrated an increase in the SUMO1/2 modifications upon trapping, as well as an interaction with the ubiquitin-regulated p97 ATPase (aka VCP). Subsequent experiments demonstrated that upon trapping, PARP1 is SUMOylated by the SUMO-ligase PIAS4 and subsequently ubiquitinated by the SUMO-targeted E3-ubiquitin ligase, RNF4, events that promote p97 recruitment and p97 ATPase-mediated removal of trapped-PARP1 from chromatin. Consistently, small molecule p97 complex inhibitors, including a metabolite of the clinically-used drug disulfiram, CuET that acts as a p97 sequestration agent, prolong PARP1 trapping and thus enhance PARPi-induced cytotoxicity in homologous recombination defective tumor cells and patient-derived tumor organoids. Taken together, these results suggest that p97 ATPase plays a key role in the processing of trapped PARP1 from chromatin and the response of homologous recombination defective tumor cells to PARPi. Citation Format: Dragomir B. Krastev, Chris Lord. Trapped PARP1 cytotoxicity is modulated by the ubiquitin-dependentsegregase p97 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 804.

Abstract 1288: STING agonism enhances anti-tumor immune responses and therapeutic efficacy of PARP inhibition in BRCA-associated breast cancer

Abstract Poly (ADP-ribose) polymerase (PARP) inhibitors exert their efficacy by inducing synthetic lethal effects, as well as cGAS/STING-mediated immune responses in BRCA- and other homologous recombination repair-deficient cancer cells. Here, we investigated whether the immunologic and therapeutic effects of PARP inhibition in BRCA-deficient breast cancer models could be augmented by synthetic cyclic dinucleotide agonists of STING. In in vitro experiments, combined olaparib-mediated PARP inhibition and ADU-S100-mediated STING agonism induced a greater degree of STING pathway activation and proinflammatory cytokine production compared to monotherapies in BRCA1-deficient human MDA-MB-436 cells and mouse triple-negative breast cancer cell lines derived from a genetically engineered model of BRCA-deficient breast cancer. In vivo experiments were conducted in mice bearing completely established tumors derived from the genetically engineered mouse model expanded syngeneically. In this model, the combination also induced an improved immune response compared to either monotherapy alone, evidenced by a greater degree of cytotoxic T cell recruitment and activation, and enhanced dendritic cell activation and antigen presentation. Nanostring mRNA analysis indicated that combinatorial effects were the result of augmented interferon signaling and antigen processing, as well as heightened leukocyte and dendritic cell functions. The combination markedly improved anti-tumor efficacy in vivo compared to monotherapy treatment, with evidence of complete tumor clearance and prolongation of survival. Finally, in mice treated with the combination, tumors could not be re-established upon rechallenge, indicating the induction of immunologic memory. These results support the development of combined PARP inhibition and STING agonism in BRCA-associated breast cancer. Citation Format: Heta Jadhav, Constantia Pantelidou, Aditi Kothari, Renyan Liu, Gerburg Wulf, Jennifer L. Guerriero, Geoffrey I. Shapiro. STING agonism enhances anti-tumor immune responses and therapeutic efficacy of PARP inhibition in BRCA-associated breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1288.

Abstract 1051: Combination of senaparib with temozolomide for the treatment of cancer

Abstract Poly (ADP-ribose) polymerase (PARP) is a critical enzyme for DNA repair and PARP inhibitors have been found to be efficacious against cancers with defects in DNA repair pathways such as with BRCA mutations. In addition, in vitro and in vivo studies have found that for PARP inhibitors with good trapping activity, combination with temozolomide (TMZ) produced the best synergetic effects among different types of chemotherapies. Several clinical combination studies of PARP inhibitors with TMZ have been reported, such as the combination of olaparib with TMZ in small cell lung cancer (SCLC). In this presentation, we will report in vitro and in vivo studies of PARP inhibitor senaparib (IMP4297) in combination with TMZ. Synergetic effects were observed when senaparib was combined with TMZ in vitro against human SCLC NCI-H209 cells. In vivo studies of high-dose senaparib in combination with low-dose TMZ once a day continuously in human SCLC NCI-H209 xenograft model was found to be highly synergetic and well tolerated. A clinical study of senaparib in combination with TMZ in patients with advanced solid tumors and SCLC is ongoing (ClinicalTrials.gov Identifier: NCT04434482). Citation Format: Sui Xiong Cai, Ning Ma, Xiaozhu Wang, Jie Ma, Mingchuan Guo, Ruiyu Zhou, Lan Liu, Chih-Yi Hsieh, Ye Edward Tian. Combination of senaparib with temozolomide for the treatment of cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1051.

Abstract 5408: Targeting ATR and AKT pathways promotes cell death in PARP inhibitor-resistant ovarian cancer cells by increasing DNA damage and lethal replication stress

Abstract Poly (ADP-ribose) polymerase inhibitors (PARPis) have changed the treatment paradigm in the management of high-grade serous ovarian cancer (HGSOC). However, most HGSOC ultimately develops resistance to PARPi, leading to unmet medical needs for better therapeutic strategies. ATR-mediated G2/M cell cycle checkpoints are important for cell survival and DNA repair, especially in HGSOC due to its dysfunctional p53-mediated G1/S checkpoint, making them an attractive therapeutic target. However, only modest monotherapy activity of ATR inhibitors (ATRis) has been reported in patients with HGSOC, highlighting the need for combination treatment. We conducted a high-throughput drug combination screen of ATRi (ceralasertib) with 1,912 drugs in two PARPi-resistant BRCA2-mutant PEO1 (PEO1/OlaR and PEO1/OlaJR) HGSOC cell lines. In this screen, 234 drugs showed synergistic cytotoxicity with ATRi, including 31 (13.2%) cell cycle checkpoint inhibitors, 27 (11.5%) chemotherapeutic drugs, and 13 (5.6%) PI3K/AKT pathway inhibitors. We prioritized the PI3K/AKT pathway inhibitors for ATRi combination because the PI3K/AKT pathway is upregulated in &amp;gt;70% of HGSOC and associated with an aggressive phenotype. Moreover, PI3K/AKT signaling is found to be activated in HGSOC cells with PARPi resistance. We separately validated that an ATRi (ceralasertib &amp;lt; 1 μM) in combination with an AKT inhibitor (AKTi; capivasertib &amp;lt; 10 μM) yielded additive/synergistic cytotoxic effects (combination index &amp;lt; 1) in various HGSOC cell lines, including acquired and de novo PARPi-resistant (PEO1/OlaR, PEO1/OlaJR, and PEO4) and platinum-resistant HGSOC cell lines (OVCAR3), by XTT and colony formation assays. With clinically attainable concentrations of ATRi (1 μM) and AKTi (10 μM), the combination treatment increased cell apoptosis compared to ATRi (increased 2.1 to 95.2-fold; P &amp;lt; 0.001) or AKTi alone (increased 3.3 to 92.2-fold; P &amp;lt; 0.001) in both PARPi-sensitive and -resistant HGSOC cells. ATRi+AKTi treatment also showed greater DNA damage as evidenced by increased percentage of cells with ≥ 5 γH2AX foci relative to ATRi (increased 1.1 to 7.6-fold; not significant in PEO1 and P &amp;lt; 0.001 in PARPi-resistant cells) or AKTi (increased 2.3 to 6.2-fold; P &amp;lt; 0.001) in both PARPi-sensitive and -resistant cells. Furthermore, ATRi+AKTi treatment augmented replication stress as measured by increased phospho-RPA+/γH2AX+ populations compared with ATRi (increased 1.7 to 4.7-fold; P &amp;lt; 0.001) or AKTi (increased 3.7 to 12.0-fold; P &amp;lt; 0.001). Overall, our results suggest that dual inhibition of ATR and AKT pathways results in greater cell death by increasing DNA damage and replication stress in HGSOC cells with PARPi resistance. Citation Format: Tzu-Ting Huang, Chih-Yuan Chiang, Nitasha Gupta, Kelli Wilson, Ken Chih-Chien Cheng, Jung-Min Lee. Targeting ATR and AKT pathways promotes cell death in PARP inhibitor-resistant ovarian cancer cells by increasing DNA damage and lethal replication stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5408.

Abstract 3494: Characterizing the relationship between genotype and phenotype in DNA damage response-altered pancreatic cancer models

Abstract Alterations in DNA damage response (DDR) proteins are prevalent across cancers and present unique opportunities for the therapeutic induction of synthetic lethality (i.e., two gene deficiencies in the same pathway synergize to kill tumor cells). In pancreatic adenocarcinoma, patients harboring alterations in DDR genes, such as in homology-directed repair (HDR), comprise the 10-20% of patients that may benefit from the recent FDA approval of Olaparib, a poly (ADP-ribose) polymerase inhibitor that disrupts non-homologous end joining (NHEJ). Despite being the best-in-class targeted therapy for DDR-altered pancreatic cancers, a subset of patients fails to respond to Olaparib and those that initially respond eventually go on to develop resistance. The mechanisms underlying resistance are poorly elucidated but suggest that some patients harboring DDR-altered tumors are HDR-proficient or revert to HDR-proficiency during therapy, bypassing the second gene deficiency requisite for synthetic lethal induction with targeted therapy (e.g., Olaparib). Utilizing patient-derived models of cancer, we are actively investigating DDR-altered, pancreatic tumor cell lines and exploring the relationship between DDR-alterations and HDR-deficiency with three goals: [1] To evaluate the prevalence of HDR-proficiency in BRCA-altered tumors, [2] to understand the extent to which patient-derived models of cancer are concordant and predictive of clinical outcomes, and [3] to identify novel therapeutic strategies for resistant patients. So far, we have observed HDR-proficiency by functional assays (e.g., RAD51 foci formation) in DDR-altered patient-derived cell lines that corresponds with poor response to Olaparib in vitro, and resistance to platinum therapy clinically. Whereas DDR-altered HDR-deficient lines had greater sensitivity to Olaparib and platinum therapy. These early data suggest that measures of HDR by functional assay may be more predictive of clinical therapeutic responses than the use of genetic signatures alone. Citation Format: James R. Carroll, Jason M. Link, Jonathan R. Brody. Characterizing the relationship between genotype and phenotype in DNA damage response-altered pancreatic cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3494.

Abstract 1792: FGFR3 mediated PARP1 tyrosine 158 phosphorylation promotes PARP inhibitor resistance

Abstract Poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi), which induce DNA damage by inhibiting PARP1 enzymatic activity and trapping PARP on the damaged DNA, are used to eliminate BRCA1/2-mutated (BRCAm) cancer. However, clinical observations suggest that BRCAm tumors develop PARPi resistance. Current strategies to overcome PARPi resistance include impeding multiple DNA repair pathways to induce excessive DNA damage. Here, we propose a novel strategy targeting oncogenic receptor tyrosine kinases to enhance PARP trapping. By developing triple-negative breast cancer (TNBC) cells with acquired talazoparib resistance, we observed a high prevalence of activated fibroblast growth factor receptor 3 (FGFR3) kinase in these cells through kinase antibody array analysis. Mass spectrometry analysis and in vitro kinase assay suggested that FGFR3 phosphorylated PARP1 at tyrosine residues 158 and 176. Biochemistry studies suggested that only PARP1 tyrosine 158 phosphorylation contributes to PARPi resistance in the cells we developed. We then developed a monoclonal antibody against tyrosine 158 phosphorylated PARP1, and found that high-level PARP1 tyrosine 158 phosphorylation positively correlated with PARPi resistance in breast cancer patient-derived xenograft models. We further demonstrated that the combination of FGFR inhibitor and PARPi delayed DNA repair with prolonged PARP trapping. Moreover, synergy between PARPi and FGFR inhibition was observed in multiple TNBC cell lines with PARPi resistance in vitro. The combination of PARPi and FGFR inhibitor also showed synergism in vivo, and treatment with the combination of PARPi and FGFR inhibitor was tolerated in mouse models. These findings reveal that PARP1 tyrosine 158 phosphorylation facilitates resolving of the PARPi-induced PARP-trapping, and that the tyrosine 158 phosphorylated PARP1 may be an effective biomarker to indicate FGFR3 mediated PARPi resistance. Citation Format: MeiKuang Chen, Yuan Gao, Weiya Xia, Yu-Han Wang, Jennifer K. Litton, Yu-Yi Chu, Funda Meric-Bernstam, Helen Piwnica-Worms, Banu Arun, Jordi Rodon Ahnert, Yongkun Wei, Wei-Chao Chang, Hung-Ling Wang, Coya Tapia, Constance T. Albarracin, Shao-Chun Wang, Ying-Nai Wang, Gabriel N. Hortobagyi, Chunru Lin, Liuqing Yang, Dihua Yu, Mien-Chie Hung. FGFR3 mediated PARP1 tyrosine 158 phosphorylation promotes PARP inhibitor resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1792.

Abstract 5399: Synergistic combination nanomedicine of doxorubicin and oligo hyaluronic acid inhibits DNA damage repair and overcomes drug resistance in metastatic triple-negative breast cancer

Abstract Background: Triple-negative breast cancer (TNBC) is a highly aggressive breast cancer subtype. Despite the early response to chemotherapy, high incidence of recurrence leads to short overall survival and poor prognosis. Native hyaluronic acid (HA) interacts with CD44 and receptors for hyaluronan mediated motility (RHAMM) overexpressed in TNBC mediating chemo-resistance and metastasis. In contrast, oligomeric HA (oHA) can disrupt HA-CD44/RHAMM interactions and attenuate oncogenic and pro-metastatic pathways, such as the mitogen-activated protein kinase/extracellular-signal-regulated kinase (MAPK/ERK) pathway, reducing downstream DNA repair and drug resistance markers. Our previous work demonstrated that oHA can exert synergistic anti-tumor and anti-metastatic effects when combined with doxorubicin (DOX) and co-loaded in an integrin-targeted iRGD-modified nanoparticle system (iRGD-DOX-oHA-PLNs). This study aims to investigate the inhibitory effect of iRGD-DOX-oHA-PLNs on both DNA single-strand break (SSB) and double-strand break (DSB) repair proteins and drug efflux pumps responsible for multidrug resistance to enhance DOX efficacy in breast cancer gene 1 (BRCA1) mutant and non-mutant TNBC. Methods: The cytotoxicity of DOX, oHA and their combinations in free solution or in nanoparticles was evaluated by clonogenic assay in human MDA-MB-231-luc-D3H2LN and MDA-MB-436 (BRCA1 mutant) TNBC cells. The in vitro expression of a DNA DSB marker, DNA repair markers, and drug efflux pump P-glycoprotein (P-gp) were measured by Western blot. The in vivo expression level of BRCA1 and Rad51 in an orthotopic TNBC mouse model was determined by immunohistochemical staining. Results: The combination of DOX-oHA showed synergism against both BRCA1 mutant and non-mutant TNBC cells. The iRGD-DOX-oHA-PLNs induced great increases in DNA DSBs demonstrated by the highest γH2AX level compared to other treatment groups. These nanoparticles also showed inhibitory effects on the expression of both DNA SSB repair protein (poly (ADP-ribose) polymerase) and DNA DSB repair proteins (Rad50 and Rad51), contributing to the enhanced efficacy of chemotherapy. The immunohistochemical staining of tumor tissues indicated lower levels of BRCA1 and Rad51 after iRGD-DOX-oHA-PLN treatment than the formulation without oHA, attributable to the effect of intracellularly delivered oHA on limiting the MAPK signaling. Additionally, iRGD-DOX-oHA-PLNs reduced the expression of the drug efflux pump P-gp as compared to DOX treatment groups without oHA. Conclusion: The co-delivery of oHA and DOX in the iRGD-DOX-oHA-PLNs efficiently blocked DNA damage repair and down-regulated the drug efflux pump P-gp, thus improving the efficacy of DOX. Collectively, this nanoparticle system could be a promising option for metastatic TNBC treatment. Citation Format: Pei Zhi, Ibrahim Alradwan, Tian Zhang, HoYin Lip, Abdulmutalib Zetrini, Chunsheng He, Jeffery Henderson, Andrew Michael Rauth, Xiao Yu Wu. Synergistic combination nanomedicine of doxorubicin and oligo hyaluronic acid inhibits DNA damage repair and overcomes drug resistance in metastatic triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5399.

Real-world effectiveness of first-line maintenance olaparib in women with BRCA-mutated advanced ovarian cancer: U.S. retrospective cohort study.

5518 Background: Following cytoreductive surgery and first-line (1L) platinum based chemotherapy, 70% of patients (pts) with newly diagnosed advanced ovarian cancer (AOC) experience relapse within 3 years. 1L maintenance treatment (tx) with olaparib monotherapy, a poly (ADP-ribose) polymerase inhibitor, extended progression free survival in pts with BRCA mutated (BRCAm) AOC in the SOLO-1 (NCT01844986) randomized controlled trial (RCT). We aimed to describe real world characteristics, tx patterns, and clinical outcomes of pts with newly diagnosed, BRCAm AOC. Methods: Data were collected from 457 pts via electronic case report forms in the US. Pt eligibility criteria were positive BRCAm test result between Jan-June 2019 (6 months post 1L olaparib FDA approval), newly diagnosed AOC (no maintenance tx prior to index), aged ≥18 years, no RCT enrollment, no active neoplasia, no prior tx for metastatic/stage IV cancer. Pt follow up was until July 2021. Kaplan-Meier (KM) estimation was used for time to event analysis using progression proxies: Time to subsequent therapy and discontinuation (or death for both) from initiation of 1L maintenance olaparib monotherapy. Results: Pt mean age (±SD) at index was 62 (±10) years, 64% of pts were white and 18% African American; further clinical characteristics are in the table. 95% of pts received ≥1 pharmacological tx. At 1L, 36% of pts (156/433) received maintenance tx after chemotherapy, with 64% of pts (277/433) receiving routine surveillance (no maintenance tx). Olaparib was the most common maintenance tx (67%, 105/156), with 67% of pts (70/105) prescribed olaparib monotherapy. At 24 months, the KM model estimated 91% of pts (95% confidence interval: 82, 96) would not progress to their subsequent therapy, with a median discontinuation of olaparib monotherapy of 25.0 (interquartile range [IQR]: 21.0, not estimable) months, over a median follow-up of 22.0 (IQR: 20.6, 24.0) months. Conclusions: This real world study complements SOLO-1 RCT data by demonstrating prolonged benefit of 1L olaparib in newly diagnosed pts with BRCAm AOC in terms of lower likelihood of progression (tx switch/death). To assess real world tx effectiveness globally, data collection across further eight countries is due to be completed in 2022. [Table: see text]

Lurbinectedin in patients with pretreated BRCA1/2-associated metastatic breast cancer: Results from a phase II basket study.

1092 Background: Lurbinectedin (L) is a selective inhibitor of oncogenic transcription that leads to cell apoptosis and shows antitumor activity against homologous recombination repair-deficient cell lines. A previous phase II study (Cruz et al. JCO 2018;36:3134-3143) demonstrated antitumor activity in patients (pts) with pretreated metastatic breast cancer (median of 1 prior advanced chemotherapy line) and BRCA1/2-mutated tumors with L 3.5 mg/m2 or 7.0 mg flat dose (equivalent to 4.0 mg/m2) every three weeks [q3wk]). This report focuses on the outcomes in the BRCA1/2-associated breast cancer cohort of a phase II Basket multitumor trial. Methods: This phase II study evaluated L 3.2 mg/m2 1-hour intravenous (i.v.) infusion q3wk in a cohort of 21 female pts with pretreated BRCA1/2-associated breast cancer. The primary efficacy endpoint was ORR according to RECIST v1.1. Secondary endpoints included duration of response (DoR), progression-free survival (PFS), OS and safety. Results: Median age was 45 years (range, 29-73 years). Hormone receptor (HR)+ disease was observed in 76.2% of pts, triple negative disease in 19.0% and HER2+ in 9.5%. BRCA1 and BRCA2 were reported in 47.6% and 52.4% of pts, respectively. Median number of prior lines of chemotherapy for advanced disease was 2 (range, 0-3 lines). Prior poly(ADP-ribose) polymerase inhibitors and platinum compounds had been administered to 23.8% and 47.6% of pts, respectively. Confirmed partial response (PR) was observed in six pts (ORR = 28.6%; 95% CI, 11.3-52.2%). Lurbinectedin was active in both BRCA mutations: four PRs in 11 pts (36.4%) in BRCA2 and two PRs in 10 pts (20.0%) in BRCA1. Median DoR was 8.6 months, median PFS was 4.1 months and median OS was 16.1 months. Stable disease (SD) was observed in ten pts (47.6%), including three pts with unconfirmed response in a subsequent tumor assessment (ORR unconfirmed = 42.9% [95%CI, 21.8-66.0]). Clinical benefit rate (PR + SD≥4 months) was 76.2% (95% CI, 52.8-91.8%). The most common grade 3/4 toxicity was neutropenia (42.9%; grade 4, 23.8%; with no febrile neutropenia). Conclusions: This phase II study met its primary endpoint and confirmed the activity of L in pretreated BRCA1/2-associated breast cancer pts. L 3.2 mg/m2 1-hour i.v. infusion q3wk showed an acceptable, predictable and manageable safety profile. Considering the exploratory aim of this trial as well as previous results in other phase II study, further development of L in this indication is warranted. Clinical trial information: NCT02454972.

Homologous Recombination Deficiency in Pancreatic Cancer: Poly (ADP-ribose) Polymerase Inhibition, Checkpoint Inhibition, or a Combination of Both?

Homologous Recombination Deficiency in Pancreatic Cancer: Poly (ADP-ribose) Polymerase Inhibition, Checkpoint Inhibition, or a Combination of Both?

Modeling the Structure of Human tRNA-Guanine Transglycosylase in Complex with 7-Methylguanine and Revealing the Factors that Determine the Enzyme Interaction with Inhibitors

tRNA-guanine transglycosylase, an enzyme catalyzing replacement of guanine with queuine in human tRNA and participating in the translation mechanism, is involved in the development of cancer. However, information on the small-molecule inhibitors that can suppress activity of this enzyme is very limited. Molecular dynamics simulations were used to determine the amino acid residues that provide efficient binding of inhibitors in the active site of tRNA-guanine transglycosylase. It was demonstrated using 7-methylguanine molecule as a probe that the ability of the inhibitor to adopt a charged state in the environment of hydrogen bond acceptors Asp105 and Asp159 plays a key role in complex formation. Formation of the hydrogen bonds and hydrophobic contacts with Gln202, Gly229, Phe109, and Met259 residues are also important. It has been predicted that introduction of the substituents would have a different effect on the ability to inhibit tRNA-guanine transglycosylase, as well as the DNA repair protein poly(ADP-ribose) polymerase 1, which can contribute to the development of more efficient and selective compounds.