EXTH-76. DEVELOPING THE NOVEL COMBINATION THERAPY OPTIONS FOR CANCER THERAPY USING TUMOR TREATING FIELDS TOGETHER WITH THE CHEMO AGENTS TARGETING THE REPLICATION STRESS PATHWAY

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi180-vi181
Author(s):  
Narasimha Kumar Karanam ◽  
Michael Story
Keyword(s):  
Dna Repair ◽  
Cancer Therapy ◽  
Clinical Benefit ◽  
Critical Role ◽  
Replication Stress ◽  
Cell Killing ◽  
Dna Breaks ◽  
Repair Capacity ◽  
Tumor Treating Fields ◽  
Chemotherapy Agents

Abstract Tumor Treating Fields (TTFields) as a component of cancer therapy has been shown to provide significant clinical benefit. The disruption of mitosis was identified as the first mechanism of action, however, a novel role for TTFields outside of mitosis where TTFields downregulates the Fanconi’s Anemia genes and proteins results in replication stress and reduced DNA repair capacity. Given these results we hypothesized that TTFields would increase the sensitivity of tumor cells to chemotherapy agents that increase replication stress. An analysis of agents that target replication stress in different ways was initiated. PARP1: Targeting PARP1 protects DNA breaks by recruiting DNA repair and checkpoint proteins to the sites of damage. Using the PARP1 inhibitor olaparib followed by radiation resulted in synergistic cell killing compared to radiation or olaparib alone or in combination. Etoposide: Etoposide forms a ternary complex with topoisomerase II and prevents re-ligation of DNA strands to elicit DNA strand breaks and replication stress. When combined with TTFields cell killing increased synergistically vs. etoposide alone. AZD6738: ATR is an essential kinase regulator of the replication checkpoint that plays a critical role in safeguarding genome integrity from replication stress. The advantage of combining TTFields with the ATR inhibitor-AZD6738 was tested for cell killing and the combination vs. AZD6738 was found to be synergistic. Irinotecan: Irinotecan traps topoisomerase I- DNA in a ternary cleavage complex and inhibits the initial cleavage reaction and re-ligation steps resulting in increased replication stress. Irinotecan and TTFields was tested and found to be highly effective at tumor cell killing. Collectively, our results suggest that it is likely of considerable clinical benefit to combine TTFields with chemotherapy agents that cause replication stress. This notion may explain the results of a number of clinical trials and suggests that there may be novel TTFields/replication stress-inducing chemotherapy agent combinations worth exploring.

scholarly journals EXTH-24. EXPOSURE TO TUMOR TREATING FIELDS INHIBITS DNA REPAIR, INDUCES REPLICATION STRESS AND RENDERS TUMOR CELLS SENSITIVE TO AGENTS THAT IMPINGE UPON THESE PATHWAYS

2018 ◽  
Vol 20 (suppl_6) ◽  
pp. vi90-vi90
Author(s):  
Michael Story ◽  
Narasimha Karanam ◽  
Lianghao Ding ◽  
Brock Sishc ◽  
Debabrata Saha
Keyword(s):  
Dna Repair ◽  
Tumor Cells ◽  
Replication Stress ◽  
Tumor Treating Fields

scholarly journals Glutathione depletion sensitizes cisplatin- and temozolomide-resistant glioma cells in vitro and in vivo

2014 ◽  
Vol 5 (10) ◽  
pp. e1505-e1505 ◽  
Author(s):  
C R R Rocha ◽  
C C M Garcia ◽  
D B Vieira ◽  
A Quinet ◽  
L C de Andrade-Lima ◽  
...  
Keyword(s):  
Dna Repair ◽  
Cisplatin Resistance ◽  
Critical Role ◽  
Combined Treatment ◽  
Glioma Cells ◽  
Glutathione Depletion ◽  
Apoptotic Pathway ◽  
Repair Capacity ◽  
Intracellular Glutathione

Abstract Malignant glioma is a severe type of brain tumor with a poor prognosis and few options for therapy. The main chemotherapy protocol for this type of tumor is based on temozolomide (TMZ), albeit with limited success. Cisplatin is widely used to treat several types of tumor and, in association with TMZ, is also used to treat recurrent glioma. However, several mechanisms of cellular resistance to cisplatin restrict therapy efficiency. In that sense, enhanced DNA repair, high glutathione levels and functional p53 have a critical role on cisplatin resistance. In this work, we explored several mechanisms of cisplatin resistance in human glioma. We showed that cellular survival was independent of the p53 status of those cells. In addition, in a host-cell reactivation assay using cisplatin-treated plasmid, we did not detect any difference in DNA repair capacity. We demonstrated that cisplatin-treated U138MG cells suffered fewer DNA double-strand breaks and DNA platination. Interestingly, the resistant cells carried higher levels of intracellular glutathione. Thus, preincubation with the glutathione inhibitor buthionine sulfoximine (BSO) induced massive cell death, whereas N-acetyl cysteine, a precursor of glutathione synthesis, improved the resistance to cisplatin treatment. In addition, BSO sensitized glioma cells to TMZ alone or in combination with cisplatin. Furthermore, using an in vivo model the combination of BSO, cisplatin and TMZ activated the caspase 3–7 apoptotic pathway. Remarkably, the combined treatment did not lead to severe side effects, while causing a huge impact on tumor progression. In fact, we noted a remarkable threefold increase in survival rate compared with other treatment regimens. Thus, the intracellular glutathione concentration is a potential molecular marker for cisplatin resistance in glioma, and the use of glutathione inhibitors, such as BSO, in association with cisplatin and TMZ seems a promising approach for the therapy of such devastating tumors.

scholarly journals Tumor treating fields cause replication stress and interfere with DNA replication fork maintenance: Implications for cancer therapy

2020 ◽  
Vol 217 ◽  
pp. 33-46 ◽  
Author(s):  
Narasimha Kumar Karanam ◽  
Lianghao Ding ◽  
Asaithamby Aroumougame ◽  
Michael D. Story
Keyword(s):  
Dna Replication ◽  
Cancer Therapy ◽  
Replication Fork ◽  
Replication Stress ◽  
Tumor Treating Fields

Randomized phase II study of 2nd-line FOLFIRI versus modified FOLFIRI with PARP inhibitor ABT-888 (veliparib) (NSC-737664) in metastatic pancreatic cancer (mPC): SWOG S1513.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. TPS4147-TPS4147
Author(s):  
E. Gabriela Chiorean ◽  
Shannon McDonough ◽  
Philip Agop Philip ◽  
Elizabeth M. Swisher ◽  
Michael J. Pishvaian ◽  
...  
Keyword(s):  
Dna Repair ◽  
Phase Ii ◽  
Phase Ii Study ◽  
Parp Inhibitor ◽  
New Drugs ◽  
Parp Inhibition ◽  
Dna Breaks ◽  
Antitumor Effects ◽  
Repair Capacity ◽  
Randomized Phase Ii

TPS4147 Background: PC is characterized by multiple DNA repair defects, including in BRCA1/ 2, and other homologous recombination (HR) genes such as FANC, ATM, ATR (Waddell N, Nature 2015). Folinic acid/5-fluorouracil/ irinotecan (FOLFIRI) is a 2nd line therapy option in mPC, but overall survival (OS) averages only 6 mos (Yoo C, Br J Cancer 2009). It is known that PARP facilitates repair from topoisomerase 1-associated DNA damage, and that preclinically PARP inhibitors (PARPi) increase DNA breaks from camptothecins, resulting in synergistic antitumor effects (Smith LM, Clin Cancer Res 2005, Davidson D, Invest New Drugs 2013). PARPi are active in mPC harboring BRCA1/2 mutations. Given the preclinical synergism between ABT-888 with irinotecan, and the safety and preliminary efficacy noted in a phase I trial (Berlin J, J Clin Oncol 2014; abstr 2574), we designed a randomized phase II study of mFOLFIRI /ABT-888 vs FOLFIRI alone for 2nd line mPC patients (pts). Blood and tumor samples are collected at baseline to retrospectively analyze biomarkers related to DNA repair capacity, including the HRD assay and BROCA-HR, a targeted multi-gene sequencing to detect alterations within the Fanconi Anemia-BRCA (HR), non-homologous end joining (NHEJ), and DNA mismatch repair pathways, and correlate with efficacy. Methods: Phase II study in 143 pts randomized (1:1) to mFOLFIRI/ABT-888 or FOLFIRI. For optimal PARP inhibition, ABT-888 is dosed Days (D) 1-7 and mFOLFIRI (no 5-FU bolus) D3-5 in 14D-cycles. In the control arm, FOLFIRI is dosed D1-3 in 14D-cycles. Primary endpoint: compare OS between treatment arms; secondary endpoints: safety, progression-free survival, response rates; translational: correlate germline/somatic BRCA1/2 mutations, and other DNA repair biomarkers with efficacy in each arm. Standard eligibility criteria apply. Assuming that the addition of ABT-888 will increase OS from 6 to 9 mos, 128 eligible pts (143 pts total) are required, based on a one-sided type 1 error of 10%, and 80% power. Kaplan-Meier methodology will be used to estimate median OS for each treatment arm. This study is open to accrual (NCT02890355). Clinical trial information: NCT02890355.

scholarly journals Polymorphisms of Homologous RecombinationRAD51,RAD51B,XRCC2, andXRCC3Genes and the Risk of Prostate Cancer

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Maria Nowacka-Zawisza ◽  
Ewelina Wiśnik ◽  
Andrzej Wasilewski ◽  
Milena Skowrońska ◽  
Ewa Forma ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Dna Repair ◽  
Cancer Risk ◽  
Cancer Susceptibility ◽  
Critical Role ◽  
Prostate Cancer Risk ◽  
Nucleotide Polymorphisms ◽  
Repair Capacity ◽  
Dna Repair Capacity ◽  
Risk Of Cancer

Genetic polymorphisms in DNA repair genes may induce individual variations in DNA repair capacity, which may in turn contribute to the risk of cancer developing. Homologous recombination repair (HRR) plays a critical role in maintaining chromosomal integrity and protecting against carcinogenic factors. The aim of the present study was to evaluate the relationship between prostate cancer risk and the presence of single nucleotide polymorphisms (SNPs) in the genes involved in HRR, that is,RAD51(rs1801320 and rs1801321),RAD51B(rs10483813 and rs3784099),XRCC2(rs3218536), andXRCC3(rs861539). Polymorphisms were analyzed by PCR-RFLP and Real-Time PCR in 101 patients with prostate adenocarcinoma and 216 age- and sex-matched controls. A significant relationship was detected between theRAD51gene rs1801320 polymorphism and increased prostate cancer risk. Our results indicate that theRAD51gene rs1801320 polymorphism may contribute to prostate cancer susceptibility in Poland.

scholarly journals Inhibition of DNA2 nuclease as a therapeutic strategy targeting replication stress in cancer cells

2017 ◽  
Author(s):  
Sandeep Kumar ◽  
Xiangdong Peng ◽  
James M. Daley ◽  
Lin Yang ◽  
Jianfeng Shen ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Therapeutic Potential ◽  
Synthetic Lethality ◽  
Replication Stress ◽  
Nuclease Activity ◽  
Dna Breaks ◽  
Repair Capacity ◽  
Genetic Ablation ◽  
Associated Lesions ◽  
Dna End Resection

AbstractReplication stress is a characteristic feature of cancer cells, which is resulted from sustained proliferative signaling induced by activation of oncogenes or loss of tumor suppressors. In cancer cells, oncogene-induced replication stress manifests as replication-associated lesions, predominantly double-strand DNA breaks (DSBs). An essential mechanism utilized by cells to repair replication-associated DSBs is homologous recombination (HR). In order to overcome replication stress and survive, cancer cells often require enhanced HR repair capacity. Therefore, the key link between HR repair and cellular tolerance to replication-associated DSBs provides us with a mechanistic rationale for exploiting synthetic lethality between HR repair inhibition and replication stress. Our studies showed that DNA2 nuclease is an evolutionarily conserved essential component of HR repair machinery. Here we demonstrate that DNA2 is indeed overexpressed in pancreatic cancers, one of the deadliest and more aggressive forms of human cancers, where mutations in the KRAS are present in 90%-95% of cases. In addition, depletion of DNA2 significantly reduces pancreatic cancer cell survival and xenograft tumor growth, suggesting the therapeutic potential of DNA2 inhibition. Finally, we develop a robust high-throughput biochemistry assay to screen for inhibitors of the DNA2 nuclease activity. The top inhibitors were shown to be efficacious against both yeast Dna2 and human DNA2. Treatment of cancer cells with DNA2 inhibitors recapitulates phenotypes observed upon DNA2 depletion, including decreased DNA end resection and attenuation of HR repair. Similar to genetic ablation of DNA2, chemical inhibition of DNA2 selectively attenuates the growth of various cancer cells with oncogene-induced replication stress. Taken together, our findings open a new avenue to develop a new class of anti-cancer drugs by targeting druggable nuclease DNA2. We 4, 16. In propose DNA2 inhibition as new strategy in cancer therapy by targeting replication stress, a molecular property of cancer cells that is acquired as a result of oncogene activation instead of targeting undruggable oncoprotein itself such as KRAS.

The Recql5 Helicase Is a Novel Downstream Target of Jak2V617F and Maintains Genome Stability in Response to Replication Stress

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 822-822
Author(s):  
Edwin Chen ◽  
Jong-Sook Ahn ◽  
Lawrence J Breyfogle ◽  
Anthony R Green ◽  
Benjamin L. Ebert ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Dna Repair ◽  
Myeloproliferative Neoplasms ◽  
Critical Role ◽  
Replication Stress ◽  
Therapeutic Window ◽  
Replication Forks ◽  
Erythroid Progenitor ◽  
Stalled Replication Forks

Abstract The JAK2V617F mutation is present in a majority of patients with chronic myeloproliferative neoplasms (MPNs). Mutant JAK2 induces hyperactivation of multiple downstream signaling processes with the net effect of conferring cells with a pro-survival advantage. In particular, JAK2V617F-expressing cells tolerate increased DNA damage and higher levels of intracellular reactive oxygen species. However, the mechanisms by which increased genotoxic tolerance is mediated remain unclear. Previously, we performed gene expression analysis on autologous wildtype and JAK2V617F-heterozygous erythroblasts from 36 MPN patients, and observed increased expression of the RECQL5 helicase in JAK2-mutant erythroblasts. Increased Recql5 transcript and protein levels were also validated in Hoxb8-immortalized, GMP-like cell lines derived from wildtype and Jak2V617F knock-in mice (WT-B8 and VF-B8 cells, respectively). Recql5 up-regulation was dependent on the Pi3k-Akt pathway, and was independent of Stat1/5 and Mapk/Erk activity. As the Recql family of helicases plays a critical role in replication fork stability, we tested whether Recql5 could modulate sensitivity of JAK2-expressing cells to agents which promote replication stress, such as hydroxyurea (HU) and aphidicolin (APH). Strikingly, VF-B8 cells transduced with two different Recql5 shRNAs were more susceptible to HU- and APH-induced apoptosis when compared to Recql5-deficient WT-B8 cells. Replication stress-induced cytotoxicity was accompanied by increased gamma-H2Ax-marked double stranded breaks (DSBs) and activation of DNA repair pathways. Importantly, re-introduction of an shRNA-resistant Recql5 cDNA successfully rescued Recql5-deficient VF-B8 cells from HU- and APH-cytotoxicity. Molecularly, we show that Recql5 plays two roles to protect against DSB formation and cell death in Jak2-mutant cells. First, we visualized replication tracts on individual DNA fibers by chromosome combing and observed that Recql5-deficient VF-B8 cells treated with HU exhibit increased numbers of stalled replication forks. Moreover, Recql5 deficiency also led to an inability to restart forks stalled by HU treatment. This indicates that the absence of Recql5 leads to replication forks which are unstably arrested upon HU treatment, leading to fork collapse and the generation of DSBs. Second, we quantified the rate of single-stranded annealing (SSA) repair following Recql5 knockdown. Consistent with previous reports, we observed increased rates of SSA repair in VF-B8 cells compared to WT-B8 cells. However, this difference in the rate of SSA repair is abrogated upon Recql5 knockdown, suggesting that Recql5 functions as a mediator for the SSA DNA repair pathway. Cumulatively, these findings suggest that Recql5 up-regulation in Jak2V617F-expressing cells plays a role in protecting against DNA damage-induced cell death through (1) stabilization of stalled replication forks thus preventing their collapse, and (2) promoting rapid (albeit error prone) SSA DNA repair to ameliorate genomic instability. Finally, we tested whether modulation of RECQL5 could also increase sensitivity of JAK2V617F-positive cells from primary MPN patients to HU. Following depletion with RECQL5, c-kit-enriched peripheral blood mononuclear cells from 2 essential thrombocythemia and 3 myelofibrosis patients were grown in semi-solid medium supplemented with HU for 14 days. Strikingly, we observed more specific eradication of JAK2V617F-positive erythroid progenitor colonies compared to autologous wildtype colonies from all patients examined. In contrast, no specific killing of JAK2V617F-positive erythroblasts was seen following transduction of control hairpins. This suggests that RECQL5 knockdown may potentially open a therapeutic window by sensitizing Jak2V617F-expressing cells to HU and other agents that induce replication stress. Disclosures No relevant conflicts of interest to declare.

scholarly journals Hydrogen Peroxide-Induced DNA Damage and Repair through the Differentiation of Human Adipose-Derived Mesenchymal Stem Cells

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Mahara Valverde ◽  
Jonathan Lozano-Salgado ◽  
Paola Fortini ◽  
Maria Alexandra Rodriguez-Sastre ◽  
Emilio Rojas ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Stem Cells ◽  
Dna Damage ◽  
Dna Repair ◽  
Mesenchymal Stem Cells ◽  
Adipocyte Differentiation ◽  
Dna Lesions ◽  
Dna Breaks ◽  
Repair Capacity ◽  
Dna Repair Capacity

Human adipose-derived mesenchymal stem cells (hADMSCs) are recognized as a potential tool in cell tissue therapy because of their capacity to proliferate and differentiate in vitro. Several studies have addressed their use in regenerative medicine; however, little is known regarding their response to DNA damage and in particular to the reactive oxygen species (ROS) that are present in the microenvironment of implantation. In this study, we used the ROS-inducing agent hydrogen peroxide to explore the responses of (1) hADMSCs and (2) derived terminally differentiated adipocytes to oxidatively generated DNA damage. Using single cell gel electrophoresis, a dose-related increase was found for both DNA breaks and oxidative lesions (formamidopyrimidine DNA glycosylase-sensitive sites) upon exposure of hADMSCs to hydrogen peroxide. DNA repair capacity of hADMSCs was affected in cells exposed to 150 and 200 μM of hydrogen peroxide. An increase in the basal levels of DNA breaks and oxidative DNA lesions was observed through adipocyte differentiation. In addition, hydrogen peroxide-induced DNA damage increased through adipocyte differentiation; DNA repair capacity also decreased. This study is the first follow-up report on DNA repair capacity during adipogenic differentiation. Remarkably, in terminally differentiated adipocytes, DNA breakage repair is abolished while the repair of DNA oxidative lesions remains efficient.

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