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Theodore S. Lawrence - One of the best experts on this subject based on the ideXlab platform.

  • Targeting the DNA Damage Response for Radiosensitization
    Cancer Drug Discovery and Development, 2020
    Co-Authors: Matthew T. Mcmillan, Theodore S. Lawrence, Meredith A Morgan
    Abstract:

    Radiation is a DNA-damaging agent that exerts its lethal anti-tumoral effects predominately by inducing DNA double-strand breaks. Tumor cells can evade radiation-induced cell death through upregulating elements of the DNA damage response (DDR) that promote DNA repair and cell cycle checkpoint activation. These critical mediators include PARP1, WEE1, DNA-PK, ATM, ATR, and CHK1. Agents that inhibit these components of the DDR can act as radiosensitizers that enhance tumor cell killing. In this chapter, we discuss actionable DDR targets and their role in Radiosensitization. This includes detailing their normal physiological roles, mechanisms for Radiosensitization, immunomodulatory properties, clinical trials, and mechanistically based rational treatment combinations.

  • parp1 trapping and dna replication stress enhance Radiosensitization with combined wee1 and parp inhibitors
    Molecular Cancer Research, 2017
    Co-Authors: Leslie A Parsels, David Karnak, Joshua D Parsels, Qiang Zhang, Jonathan Velezpadilla, Zachery R Reichert, Daniel R Wahl, Jonathan Maybaum, Mark J Oconnor, Theodore S. Lawrence
    Abstract:

    KRAS mutations in non–small cell lung cancer (NSCLC) cause increased levels of DNA damage and replication stress, suggesting that inhibition of the DNA damage response (DDR) is a promising strategy for Radiosensitization of NSCLC. This study investigates the ability of a WEE1 inhibitor (AZD1775) and a PARP inhibitor (olaparib) to radiosensitize KRAS-mutant NSCLC cells and tumors. In addition to inhibiting the DDR, these small-molecule inhibitors of WEE1 and PARP induce DNA replication stress via nucleotide exhaustion and PARP trapping, respectively. As monotherapy, AZD1775 or olaparib alone modestly radiosensitized a panel of KRAS-mutant NSCLC lines. The combination of agents, however, significantly increased Radiosensitization. Furthermore, AZD1775-mediated Radiosensitization was rescued by nucleotide repletion, suggesting a mechanism involving AZD1775-mediated replication stress. In contrast, Radiosensitization by the combination of AZD1775 and olaparib was not rescued by nucleosides. Whereas both veliparib, a PARP inhibitor that does not efficiently trap PARP1 to chromatin, and PARP1 depletion radiosensitized NSCLC cells as effectively as olaparib, which does efficiently trap PARP, only olaparib potentiated AZD1775-mediated Radiosensitization. Taken together, these mechanistic data demonstrate that although nucleotide depletion is sufficient for Radiosensitization by WEE1 inhibition alone, and inhibition of PARP catalytic activity is sufficient for Radiosensitization by olaparib alone, PARP1 trapping is required for enhanced Radiosensitization by the combination of WEE1 and PARP inhibitors. Implications: This study highlights DNA replication stress caused by nucleotide depletion and PARP1 trapping as an important mechanism of Radiosensitization in KRAS-mutant tumors and supports further development of DNA replication as a therapeutic target. Mol Cancer Res; 16(2); 222–32. ©2017 AACR.

  • abstract 519 combined inhibition of wee1 and parp1 radiosensitizes kras mutant non small cell lung cancers via inhibition of dsb repair
    Cancer Research, 2016
    Co-Authors: Leslie A Parsels, Theodore S. Lawrence, David Karnak, Joshua D Parsels, Zachery R Reichert, Jonathan Maybaum, Meredith A Morgan
    Abstract:

    Mutant KRAS is found in approximately 30% of non-small cell lung cancers (NSCLC) and is associated with poor prognosis. Despite the use of radiation (RT) therapy for the treatment of locally advanced disease, local recurrence remains an issue. The observation that mutations in KRAS lead to both replication stress and DNA double-strand breaks (DSBs) suggests these cancers may have a greater reliance on DNA damage response pathways and therefore may be preferentially radiosensitized by therapies targeting DNA repair. In this study, we investigated the combined inhibition of WEE1 and PARP1 using the small molecule inhibitors, AZD1775 and olaparib, respectively as a radiosensitizing strategy in KRAS mutant NSCLC. We began by comparing Radiosensitization by AZD1775+olaparib, in KRAS isogenic H1703 lung cancer cells and found that KRAS mutant cells were preferentially radiosensitized (ER, enhancement ratio 1.8) compared to KRAS wild type cells (ER 1.4). Combined WEE1 and PARP1 inhibition also radiosensitized KRAS mutant Calu-6 and NCI-H23 lung cancer cells (ER 1.9 and 1.5, respectively). These findings were further confirmed in vivo: Calu-6 tumor xenografts were significantly radiosensitized by AZD1775+olaparib, as evidenced by an 11 day delay in tumor volume doubling time relative to RT treatment. Given that WEE1 and PARP1 function to prevent and manage replication stress, respectively, we hypothesized that Radiosensitization by AZD1775+olaparib results from persistent replication stress. While replication stress did contribute to AZD1775-mediated Radiosensitization in Calu-6 cells, as evidenced by pan-nuclear γH2AX staining, and the ability of exogenous nucleosides to protect cells from Radiosensitization by AZD1775 alone, nucleosides had little effect on Radiosensitization by AZD1775+olaparib. These results suggest that replication stress is not required for Radiosensitization by AZD1775+olaparib. As WEE1 and PARP1 both promote repair of radiation-induced DNA damage, we hypothesized that Radiosensitization by AZD1775+olaparib results from persistent, unrepaired DSBs. Assessment of the kinetics of DSB repair by γH2AX flow cytometry demonstrated that while total γH2AX levels in cells treated with RT alone had returned to control levels within 24 h, AZD1775+olaparib treatment significantly delayed the resolution of γH2AX following RT, with 44.8% or 46.2% of Calu-6 or KRAS mutant H1703 cells, respectively remaining γH2AX-positive 24 h post-RT. This delay corresponded with the inhibition of radiation-induced RAD51 foci by AZD1775, indicating inhibition of homologous recombination repair. Taken together these data demonstrate the efficacy of combined inhibition of WEE1 and PARP1 with radiation in KRAS mutant lung cancer. Furthermore, these results suggest that although replication stress occurs in response to AZD1775 and olaparib, persistent DSBs are the cause of Radiosensitization. Citation Format: Leslie A. Parsels, David Karnak, Joshua D. Parsels, Zachery Reichert, Jonathan Maybaum, Theodore S. Lawrence, Meredith Ann Morgan. Combined inhibition of WEE1 and PARP1 radiosensitizes KRAS mutant non-small cell lung cancers via inhibition of DSB repair. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 519.

  • Targeting mcl-1 for Radiosensitization of pancreatic cancers.
    Translational oncology, 2015
    Co-Authors: Dongping Wei, Theodore S. Lawrence, Leslie A Parsels, Qiang Zhang, Jason S. Schreiber, Fardokht A. Abulwerdi, Tasneem Kausar, Yi Sun, Zaneta Nikolovska-coleska, Meredith A Morgan
    Abstract:

    In order to identify targets whose inhibition may enhance the efficacy of chemoradiation in pancreatic cancer, we previously conducted an RNAi library screen of 8,800 genes. We identified Mcl-1 (myeloid cell leukemia-1), an anti-apoptotic member of the Bcl-2 family, as a target for sensitizing pancreatic cancer cells to chemoradiation. In the present study we investigated Mcl-1 inhibition by either genetic or pharmacological approaches as a radiosensitizing strategy in pancreatic cancer cells. Mcl-1 depletion by siRNA produced significant Radiosensitization in BxPC-3 and Panc-1 cells in association with Caspase-3 activation and PARP cleavage, but only minimal Radiosensitization in MiaPaCa-2 cells. We next tested the ability of the recently identified, selective, small molecule inhibitor of Mcl-1, UMI77, to radiosensitize in pancreatic cancer cells. UMI77 caused dissociation of Mcl-1 from the pro-apoptotic protein Bak and produced significant Radiosensitization in BxPC-3 and Panc-1 cells, but minimal Radiosensitization in MiaPaCa-2 cells. Radiosensitization by UMI77 was associated with Caspase-3 activation and PARP cleavage. Importantly, UMI77 did not radiosensitize normal small intestinal cells. In contrast, ABT-737, an established inhibitor of Bcl-2, Bcl-XL, and Bcl-w, failed to radiosensitize pancreatic cancer cells suggesting the unique importance of Mcl-1 relative to other Bcl-2 family members to radiation survival in pancreatic cancer cells. Taken together, these results validate Mcl-1 as a target for Radiosensitization of pancreatic cancer cells and demonstrate the ability of small molecules which bind the canonical BH3 groove of Mcl-1, causing displacement of Mcl-1 from Bak, to selectively radiosensitize pancreatic cancer cells.

  • combined inhibition of wee1 and parp1 2 for Radiosensitization in pancreatic cancer
    Clinical Cancer Research, 2014
    Co-Authors: David Karnak, Leslie A Parsels, Jonathan Maybaum, Tasneem Kausar, Mary A. Davis, Carl G Engelke, Jordan R Robertson, Katherine Marsh, Lili Zhao, Theodore S. Lawrence
    Abstract:

    Purpose: While the addition of radiation to chemotherapy improves survival in patients with locally advanced pancreatic cancer, more effective therapies are urgently needed. Thus, we investigated the radiosensitizing efficacy of the novel drug combination of Wee1 and PARP1/2 inhibitors (AZD1775 and olaparib, respectively) in pancreatic cancer. Experimental Design: Radiosensitization of AsPC-1 or MiaPaCa-2 human pancreatic cancer cells was assessed by clonogenic survival and tumor growth assays. Mechanistically, the effects of AZD1775, olaparib, and radiation on cell cycle, DNA damage (γH2AX), and homologous recombination repair (HRR) were determined. Results: Treatment of AsPC-1 and MiaPaCa-2 cells with either AZD1775 or olaparib caused modest Radiosensitization, whereas treatment with the combination significantly increased Radiosensitization. Radiosensitization by the combination of AZD1775 and olaparib was associated with G 2 checkpoint abrogation and persistent DNA damage. In addition, AZD1775 inhibited HRR activity and prevented radiation-induced Rad51 focus formation. Finally, in vivo , in MiaPaCa-2–derived xenografts, olaparib did not radiosensitize, whereas AZD1775 produced moderate, yet significant, Radiosensitization ( P P Conclusions: Taken together, these results demonstrate the efficacy of combined inhibition of Wee1 and PARP inhibitors for radiosensitizing pancreatic cancers and support the model that Wee1 inhibition sensitizes cells to PARP inhibitor–mediated Radiosensitization through inhibition of HRR and abrogation of the G 2 checkpoint, ultimately resulting in unrepaired, lethal DNA damage and Radiosensitization. Clin Cancer Res; 20(19); 5085–96. ©2014 AACR .

Meilan Liu - One of the best experts on this subject based on the ideXlab platform.

  • abstract 6271 hormone receptor inhibition as a strategy for Radiosensitization of breast cancer
    Cancer Research, 2020
    Co-Authors: Anna R Michmerhuizen, Meilan Liu, Amanda Zhang, Rachel Schwartz, Andrea M Pesch, Benjamin C Chandler, Cassandra L Ritter, Kari Wilderromans, Daniel E Spratt, Daniel R Wahl
    Abstract:

    Purpose: Expression of the androgen receptor (AR) has been identified as a driver of tumor growth in triple negative breast cancers (TNBC), and previous work has nominated AR as a target for Radiosensitization. In addition, 70-95% of all estrogen receptor (ER) positive (ER+) breast cancers also have coexpression of AR, suggesting extended utility of AR inhibition in the Radiosensitization of these AR+, ER+ tumors. Here we assessed the efficacy of AR inhibition in ER+, AR+ breast cancers to better understand the role of AR signaling across breast cancer models. Further, we also investigated the effect of ER inhibition on Radiosensitization of ER+ breast cancer models. Methods: IC50 values were determined for MDV3100 (enzalutamide), ARN-509 (apalutamide), and ODM-201 (darolutamide) in TNBC cell lines (AR+ TNBC: MDA-MB-453, ACC-422, and SUM-185PE, and AR- TNBC: MDA-MB-231) and ER+ breast cancer cell lines (AR+, ER+: ZR-75-1, BT-474, CAMA-1, and AR-, ER+: MCF-7). IC50 values for tamoxifen were determined for ER+ breast cancer cell lines (MCF-7, T47D, ZR-75-1), and ER- (SUM-159) cells. Clonogenic survival assays were performed to assess Radiosensitization with ER or AR inhibition with tamoxifen or second generation anti-androgens, respectively, in TNBC and ER+ breast cancer models. Results: AR inhibition with enzalutamide, apalutamide, and darolutamide showed limited single agent growth inhibition efficacy in AR+ TNBC and AR+, ER+ breast cancer cell lines (IC50 > 10 μM). AR inhibition with enzalutamide did not induce radiosensitivity in vitro. In AR+, ER+ CAMA-1 cells, AR blockade with enzalutamide had a radioprotective effect with enhancement ratios (enhR) of 0.76-0.83. No Radiosensitization was observed in BT-474 (enhR: 0.92-1.01) or ZR-75-1 cells (enhR: 0.94-1.00). Radiosensitization was also assessed with anti-androgens apalutamide and darolutamide in AR+ breast cancer models. Inhibition of ER with tamoxifen, however, induced Radiosensitization in MCF-7 (enhR: 1.14-1.50) and T47D (enhR: 1.33-1.60) cells. No Radiosensitization was observed with tamoxifen in ER- SUM-159 cells. Conclusion: Although AR is a mediator of radioresistance in AR+ TNBC, AR inhibition does not provide comparable Radiosensitization in AR+, ER+ models and may actually confer a radioprotective effect. In contrast, our results demonstrate ER inhibition is an effective radiosensitizing strategy in ER+ breast cancers, independent of AR status. This work highlights the complexities of androgen and estrogen receptor signaling in AR+, ER+ breast tumors and underscores the necessity for understanding context dependent effects when translating into patients with AR+ breast cancer. Citation Format: Anna R. Michmerhuizen, Amanda Zhang, Rachel Schwartz, Andrea M. Pesch, Benjamin C. Chandler, Cassandra L. Ritter, Meilan Liu, Kari Wilder-Romans, Daniel E. Spratt, Daniel R. Wahl, Shyam Nyati, Lori J. Pierce, Corey Speers. Hormone receptor inhibition as a strategy for Radiosensitization of breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6271.

  • abstract 2926 nomination and characterization of ttk for Radiosensitization in basal like breast cancers
    Cancer Research, 2019
    Co-Authors: Benjamin C Chandler, Meilan Liu, Anna R Michmerhuizen, Andrea M Pesch, Kari Wilderromans, Shyam Nyati, Leah Moubadder, Cassie Ritter, Meleah Cameron, Eric Olsen
    Abstract:

    Background: Increased rates of locoregional recurrence have been observed in basal-like breast cancer despite the use of radiation therapy (RT), therefore approaches that result in Radiosensitization of basal-like breast cancer are critically needed. Studies detailing the poor response of basal-like tumors to adjuvant RT underscore the biologic differences and as of yet undefined oncogenic drivers of these particular types of breast cancer. Methods: 4 independent datasets were used to correlate gene expression with local recurrence (LR) after 3 years. Kaplan-Meier analysis was used to validate the impact of TTK expression on LR. The TCGA and institutional breast cancer dataset were used to determine TTK expression in BC subtypes. TTK RNA and protein levels were measured using qPCR and western blot at baseline and correlated to intrinsic radiosensitivity. Clonogenic survival assays were used to determine the Radiosensitization of cell lines after TTK inhibition (TTKi). Mice models were used to assess TTKi in combination with RT in vivo. DNA damage was quantified using γH2AX staining. HR and NHEJ efficiency assays were performed using HR/NHEJ specific reporter systems. HR competency was also assessed using RAD51 foci formation assays. Results: Ten genes were found to significantly correlate with early LR (≤3 years) across 4 distinct datasets (N=896 pts) (OR of recurrence > 2, p-value Conclusion: These data support TTK inhibition as a rationale radiosensitizing strategy for clinical development in basal-like breast cancer patients and that the mechanism of Radiosensitization is, at least in part, through impaired HR repair. Citation Format: Benjamin C. Chandler, Leah Moubadder, Cassie Ritter, Kari Wilder-Romans, Meleah Cameron, Meilan Liu, Shyam Nyati, Andrea Pesch, Anna Michmerhuizen, Eric Olsen, Yashar Niknafs, Arul Chinnaiyan, Corey Speers. Nomination and characterization of TTK for Radiosensitization in basal-like breast cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2926.

  • abstract 5839 androgen receptor ar a novel target for Radiosensitization in triple negative breast cancers tnbc
    Cancer Research, 2017
    Co-Authors: Benjamin C Chandler, Daniel R Wahl, Corey Speers, Shuang G Zhao, Meilan Liu, Kari Wilderromans, Daniel E Spratt, Eric Olsen, Shyam Nyati, Daniel F Hayes
    Abstract:

    Purpose: Increased rates of local recurrence (LR) have been observed in TNBC despite chemotherapy and radiation (RT). Thus, approaches that result in Radiosensitization in TNBC are critically needed. We characterized the RT response of 21 breast cancer cell (BCC) lines using clonogenic survival assays and paired this with high-throughput drug screen data, identifying AR as a top target for Radiosensitization. We demonstrate that AR inhibition confers Radiosensitization in vitro and in vivo, identified a biomarker of response, and characterize the mechanism of AR-mediated Radiosensitization in TNBC. Materials/Methods: Clonogenic survival assays determined the intrinsic RT sensitivity of 21 BCC lines. IC50 values were determined for 130 clinical compounds and correlation coefficients were calculated using IC50 values and SF-2Gy. Gene and protein expression was measured using RNA Seq and RPPA arrays, respectively, in tumor samples (n=2,061) and BCC lines (n=51). AR function was assessed using gene knockdown (KD) or functional inhibition with anti-androgen drugs. We measured in vivo tumor growth with varying control and treatment groups (16-20 tumors/group). Kaplan-Meier analysis was performed to estimate local control. A Cox proportional hazards model and MVA were used to determine variables associated with LRF survival. Results: Our unbiased drug radiosensitizer screen nominated bicalutamide as an effective drug in treating RT-resistant BCC lines (R2= 0.46, p-value 2000 human breast tumor samples and 51 BCC lines and found heterogeneity in AR expression with strongly correlated expression of protein and RNA levels in TNBC (R2=0.89, p-value Conclusion: Our results implicate AR as a mediator of radioresistance in breast cancer and support the rationale for developing clinical strategies, including clinical trials, to inhibit AR as a novel radiosensitizing target in TNBC. Citation Format: Benjamin C. Chandler, Corey W. Speers, Shuang G. Zhao, Meilan Liu, Kari Wilder-Romans, Eric Olsen, Shyam Nyati, Daniel Spratt, Daniel Wahl, Daniel Hayes, Felix Y. Feng, Lori J. Pierce. Androgen receptor (AR): A novel target for Radiosensitization in triple-negative breast cancers (TNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5839. doi:10.1158/1538-7445.AM2017-5839

  • abstract p6 13 05 androgen receptor ar a novel target and mechanism for Radiosensitization and treatment in triple negative breast cancers tnbc
    Cancer Research, 2016
    Co-Authors: Corey Speers, Shuang G Zhao, Meilan Liu, James M Rae, Daniel F Hayes, Felix Y Feng, Lori J Pierce
    Abstract:

    Background: Increased rates of locoregional recurrence have been observed in TNBC despite chemotherapy and radiation (RT). Thus, approaches that result in radiosensitizaton in TNBC are critically needed. We characterized the RT response of 21 breast cancer cell (BCC) lines using clonogenic survival assays. We paired this with high-throughput drug screen data to identify AR as a top target for Radiosensitization and assess AR inhibition as a Radiosensitization strategy for TNBC. Methods: Clonogenic survival assays were used to determine the intrinsic RT sensitivity of 21 BCC lines. IC50 values were determined for 130 clinically available compounds and correlation coefficients were calculated using IC50 values and SF-2Gy. Gene expression was measured using RNA Seq and protein expression was measured using RPPA arrays in human tumor samples (n=2,061) and BCC lines (n=51). AR function was assessed using siRNA knockdown or functional inhibition with MDV3100 (enzalutamide). We measured in vivo tumor growth with varying control and treatment groups (16-20 tumors/group). Kaplan-Meier analysis was performed to estimate local control and survival. A Cox proportional hazards model was used to identify factors of survival, and MVA was used to determine variables associated with LRF survival. Results: Our radiosensitizer screen nominated bicalutamide as a most effective drug in treating RT-resistant BCC lines (R2= 0.46, p-value 2000 human breast tumor samples and found significant heterogeneity in AR expression with enrichment of expression at the protein and RNA level in TNBC. This same heterogeneity was also identified in human BCC lines. There was a strong correlation between AR RNA expression and protein expression (R2= 0.72, p Conclusion: Our results implicate AR as a mediator of radioresistance in breast cancer and support the rationale for developing clinical strategies, including clinical trials, to inhibit AR as a novel radiosensitizing target in TNBC. Citation Format: Speers C, Zhao SG, Liu M, Rae JM, Hayes DF, Feng FY, Pierce LJ. Androgen receptor (AR): A novel target and mechanism for Radiosensitization and treatment in triple-negative breast cancers (TNBC). [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-13-05.

Donna S. Shewach - One of the best experts on this subject based on the ideXlab platform.

  • Late DNA Damage Mediated by Homologous Recombination Repair Results in Radiosensitization with Gemcitabine.
    Radiation research, 2016
    Co-Authors: Sheryl A. Flanagan, Jeffrey J. Ackroyd, Brendan J. Knapp, Aaron Kramer, Donna S. Shewach
    Abstract:

    Gemcitabine (dFdCyd) shows broad antitumor activity in solid tumors in chemotherapeutic regimens or when combined with ionizing radiation (Radiosensitization). While it is known that mismatches in DNA are necessary for dFdCyd Radiosensitization, the critical event resulting in Radiosensitization has not been identified. Here we hypothesized that late DNA damage (≥24 h after drug washout/irradiation) is a causal event in Radiosensitization by dFdCyd, and that homologous recombination repair (HRR) is required for this late DNA damage. Using γ-H2AX as a measurement of DNA damage in MCF-7 breast cancer cells, we demonstrate that 10 or 80 nM dFdCyd alone produced significantly more late DNA damage compared to that observed within 4 h after treatment. The combination of dFdCyd treatment followed by irradiation did not produce a consistent increase in DNA damage in the first 4 h after treatment, however, there was a synergistic increase 24–48 h later relative to treatment with dFdCyd or radiation alone. RNAi sup...

  • Mismatched nucleotides as the lesions responsible for Radiosensitization with gemcitabine: a new paradigm for antimetabolite radiosensitizers
    Molecular cancer therapeutics, 2007
    Co-Authors: Sheryl A. Flanagan, Blaine W. Robinson, Christina M. Krokosky, Donna S. Shewach
    Abstract:

    Radiation sensitization by 2′,2′-difluoro-2′-deoxycytidine (dFdCyd) has correlated with dATP depletion [dFdCDP-mediated inhibition of ribonucleotide reductase (RR)] and S-phase accumulation. We hypothesized that Radiosensitization by dFdCyd is due to nucleotide misincorporations in the presence of deoxynucleotide triphosphate pool imbalances, which, if not repaired, augments cell death following irradiation. The ability of dFdCyd to produce misincorporations was measured as pSP189 plasmid mutations in hMLH1-deficient [mismatch repair (MMR) deficient] and hMLH1-expressing (MMR proficient) HCT116 cells. Only MMR-deficient cells showed a significant increase in nucleotide misincorporations (2- to 3-fold increase; P ≤ 0.01) after radiosensitizing concentrations of dFdCyd ± 5 Gy radiation, which persisted for at least 96 h. dFdCyd (10 nmol/L) did not radiosensitize MMR-proficient HCT116 or A549 cells, but following small interfering RNA–mediated suppression of hMLH1, this concentration produced excellent Radiosensitization (radiation enhancement ratios = 1.6 ± 0.1 and 1.5 ± 0.1, respectively; P < 0.05) and a 2.5-fold increase in mutation frequency in A549 cells. Cytosine arabinoside (1-β-d-arabinofuranosylcytosine), which can be incorporated into DNA but does not inhibit RR, failed to radiosensitize MMR-deficient cells or increase mutation frequency in the MMR-deficient and MMR-proficient cells. However, the RR inhibitor hydroxyurea radiosensitized MMR-deficient cells and increased nucleotide misincorporations (≥5-fold increase; P < 0.05), thus further implicating the inhibition of RR as the mechanism underlying Radiosensitization by dFdCyd. These data showed that the presence and persistence of mismatched nucleotides is integral to Radiosensitization by dFdCyd and suggest a role for hMLH1 deficiency in eliciting the radiosensitizing effect. [Mol Cancer Ther 2007;6(6):1858–68]

  • Mismatch repair status plays a role in Radiosensitization by gemcitabine
    Cancer Research, 2006
    Co-Authors: Sheryl A. Flanagan, Blaine W. Robinson, Donna S. Shewach
    Abstract:

    Proc Amer Assoc Cancer Res, Volume 47, 2006 4641 The chemotherapeutic drug gemcitabine (2’,2’-difluroro-2’-deoxycytidine;dFdCyd) is a potent radiosensitizer in tumor cells and in vivo . Incorporation of dFdCTP into DNA correlates with cytotoxicity and inhibition of ribonucleotide reductase (RR) by dFdCDP results in the depletion of dATP which is strongly correlated to Radiosensitization. We hypothesized that Radiosensitization by dFdCyd is the result of incorrect nucleotide incorporation into DNA, produced by DNA replication in the presence of imbalanced dNTP pools, which, if not repaired, augments cell death following irradiation. This hypothesis was supported by the finding that mismatch repair (MMR)-deficient HCT116 cells were better radiosensitized by dFdCyd compared to isogenic MMR-proficient cell lines. The plasmid shuttle vector pSP189 was used to examine the ability of dFdCyd to produce nucleotide misincorporations by measuring plasmid mutation rates in the MMR-deficient and MMR-proficient cell lines. Only the MMR-deficient cells showed significant nucleotide misincorporations after incubation with dFdCyd, with a ≥ 2-fold increase in mutation rate (p ≤ 0.01). Furthermore, the increased mutation frequency occurred only at dFdCyd concentrations that produced Radiosensitization. Treatment with dFdCyd for 24 hr + 5 Gy produced a ≥ 2-fold increase in the plasmid mutation rates. However, 72 hr after treatment this increase persisted only in the group that received both dFdCyd and 5 Gy (p < 0.05). While MMR-proficient HCT116 cells and A549 cells cannot be radiosensitized with dFdCyd at concentrations ≤ IC50 or ≤ IC10, respectively, these concentrations produced excellent Radiosensitization following siRNA directed knockdown of hMLH1 protein. Furthermore, plasmid mutation rates observed in A549MLH1- cells with 10 nM dFdCyd (IC10) were 2.5-fold greater than observed in the A549MLH1+ control cells. To further evaluate the relative roles of RR inhibition by dFdCDP vs. dFdCTP incorporation into DNA as the cause of the increase in mutation frequency with dFdCyd, we treated cells with the RR inhibitor hydroxyurea (HU) or the nucleoside analog cytosine arabinoside (araC). When HCT116 cell lines were treated with the IC50 for HU, only the MMR-deficient cell lines were radiosensitized and produced increased nucleotide misincorporations. Treatment with araC, which can be incorporated into DNA but does not inhibit RR, failed to produce Radiosensitization in MMR-deficient cell lines and did not increase plasmid mutation frequency. However, the combination of HU and araC produced a 2-fold increase in mutation frequency (p< 0.05). These results support earlier correlative studies that suggest the action of dFdCDP on RR as the mechanism underlying Radiosensitization by dFdCyd. Finally, these data provide evidence that the mismatched nucleotide lesion is integral to Radiosensitization by dFdCyd and support a role for MMR status in eliciting the radiosensitizing effect.

  • Radiosensitization by gemcitabine.
    Oncology, 1999
    Co-Authors: Theodore S. Lawrence, Avraham Eisbruch, Cornelius J. Mcginn, Marc T. Fields, Donna S. Shewach
    Abstract:

    Gemcitabine is a potent radiosensitizer in both laboratory studies and in the clinic. Initial laboratory studies showed that gemcitabine radiosensitizes a wide variety of rodent and human tumor cells in culture. Maximum

  • Gemcitabine-mediated Radiosensitization.
    Seminars in Oncology, 1997
    Co-Authors: Theodore S. Lawrence, Avraham Eisbruch, Donna S. Shewach
    Abstract:

    : Gemcitabine is a potent radiosensitizer of human tumor cells. This review summarizes our preclinical and early clinical studies designed to elucidate the mechanism of action of gemcitabine and phase I trials conducted to determine the optimal dose and schedule. Gemcitabine was found to radiosensitize a wide variety of human tumor cells in culture, particularly cells derived from cancers of the pancreas, breast, and head and neck. Radiosensitization occurs under conditions in which cells demonstrate concurrent redistribution into S phase and deoxyadenosine triphosphate pool depletion. These conditions can be produced by either a long (24-hour) exposure to a low concentration of gemcitabine (10 nmol/L) or by a brief (2-hour) treatment with higher but clinically relevant concentrations (100 nmol/L to 3 micromol/L). Under the latter conditions, sensitization can be detected 4 hours after treatment and last for up to 2 days. These preclinical data were useful in the design of a gemcitabine dose escalation trial in combination with standard radiation for patients with unresectable head and neck cancer. Although this trial is not yet complete, the starting dose of gemcitabine, which is far below the maximum tolerated dose for the drug when used alone, significantly potentiates the toxicity of radiation treatment. We conclude that gemcitabine is a promising radiation sensitizer that needs to be developed cautiously if excessive normal tissue toxicity is to be avoided.

Benjamin C Chandler - One of the best experts on this subject based on the ideXlab platform.

  • abstract 6271 hormone receptor inhibition as a strategy for Radiosensitization of breast cancer
    Cancer Research, 2020
    Co-Authors: Anna R Michmerhuizen, Meilan Liu, Amanda Zhang, Rachel Schwartz, Andrea M Pesch, Benjamin C Chandler, Cassandra L Ritter, Kari Wilderromans, Daniel E Spratt, Daniel R Wahl
    Abstract:

    Purpose: Expression of the androgen receptor (AR) has been identified as a driver of tumor growth in triple negative breast cancers (TNBC), and previous work has nominated AR as a target for Radiosensitization. In addition, 70-95% of all estrogen receptor (ER) positive (ER+) breast cancers also have coexpression of AR, suggesting extended utility of AR inhibition in the Radiosensitization of these AR+, ER+ tumors. Here we assessed the efficacy of AR inhibition in ER+, AR+ breast cancers to better understand the role of AR signaling across breast cancer models. Further, we also investigated the effect of ER inhibition on Radiosensitization of ER+ breast cancer models. Methods: IC50 values were determined for MDV3100 (enzalutamide), ARN-509 (apalutamide), and ODM-201 (darolutamide) in TNBC cell lines (AR+ TNBC: MDA-MB-453, ACC-422, and SUM-185PE, and AR- TNBC: MDA-MB-231) and ER+ breast cancer cell lines (AR+, ER+: ZR-75-1, BT-474, CAMA-1, and AR-, ER+: MCF-7). IC50 values for tamoxifen were determined for ER+ breast cancer cell lines (MCF-7, T47D, ZR-75-1), and ER- (SUM-159) cells. Clonogenic survival assays were performed to assess Radiosensitization with ER or AR inhibition with tamoxifen or second generation anti-androgens, respectively, in TNBC and ER+ breast cancer models. Results: AR inhibition with enzalutamide, apalutamide, and darolutamide showed limited single agent growth inhibition efficacy in AR+ TNBC and AR+, ER+ breast cancer cell lines (IC50 > 10 μM). AR inhibition with enzalutamide did not induce radiosensitivity in vitro. In AR+, ER+ CAMA-1 cells, AR blockade with enzalutamide had a radioprotective effect with enhancement ratios (enhR) of 0.76-0.83. No Radiosensitization was observed in BT-474 (enhR: 0.92-1.01) or ZR-75-1 cells (enhR: 0.94-1.00). Radiosensitization was also assessed with anti-androgens apalutamide and darolutamide in AR+ breast cancer models. Inhibition of ER with tamoxifen, however, induced Radiosensitization in MCF-7 (enhR: 1.14-1.50) and T47D (enhR: 1.33-1.60) cells. No Radiosensitization was observed with tamoxifen in ER- SUM-159 cells. Conclusion: Although AR is a mediator of radioresistance in AR+ TNBC, AR inhibition does not provide comparable Radiosensitization in AR+, ER+ models and may actually confer a radioprotective effect. In contrast, our results demonstrate ER inhibition is an effective radiosensitizing strategy in ER+ breast cancers, independent of AR status. This work highlights the complexities of androgen and estrogen receptor signaling in AR+, ER+ breast tumors and underscores the necessity for understanding context dependent effects when translating into patients with AR+ breast cancer. Citation Format: Anna R. Michmerhuizen, Amanda Zhang, Rachel Schwartz, Andrea M. Pesch, Benjamin C. Chandler, Cassandra L. Ritter, Meilan Liu, Kari Wilder-Romans, Daniel E. Spratt, Daniel R. Wahl, Shyam Nyati, Lori J. Pierce, Corey Speers. Hormone receptor inhibition as a strategy for Radiosensitization of breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6271.

  • abstract 2926 nomination and characterization of ttk for Radiosensitization in basal like breast cancers
    Cancer Research, 2019
    Co-Authors: Benjamin C Chandler, Meilan Liu, Anna R Michmerhuizen, Andrea M Pesch, Kari Wilderromans, Shyam Nyati, Leah Moubadder, Cassie Ritter, Meleah Cameron, Eric Olsen
    Abstract:

    Background: Increased rates of locoregional recurrence have been observed in basal-like breast cancer despite the use of radiation therapy (RT), therefore approaches that result in Radiosensitization of basal-like breast cancer are critically needed. Studies detailing the poor response of basal-like tumors to adjuvant RT underscore the biologic differences and as of yet undefined oncogenic drivers of these particular types of breast cancer. Methods: 4 independent datasets were used to correlate gene expression with local recurrence (LR) after 3 years. Kaplan-Meier analysis was used to validate the impact of TTK expression on LR. The TCGA and institutional breast cancer dataset were used to determine TTK expression in BC subtypes. TTK RNA and protein levels were measured using qPCR and western blot at baseline and correlated to intrinsic radiosensitivity. Clonogenic survival assays were used to determine the Radiosensitization of cell lines after TTK inhibition (TTKi). Mice models were used to assess TTKi in combination with RT in vivo. DNA damage was quantified using γH2AX staining. HR and NHEJ efficiency assays were performed using HR/NHEJ specific reporter systems. HR competency was also assessed using RAD51 foci formation assays. Results: Ten genes were found to significantly correlate with early LR (≤3 years) across 4 distinct datasets (N=896 pts) (OR of recurrence > 2, p-value Conclusion: These data support TTK inhibition as a rationale radiosensitizing strategy for clinical development in basal-like breast cancer patients and that the mechanism of Radiosensitization is, at least in part, through impaired HR repair. Citation Format: Benjamin C. Chandler, Leah Moubadder, Cassie Ritter, Kari Wilder-Romans, Meleah Cameron, Meilan Liu, Shyam Nyati, Andrea Pesch, Anna Michmerhuizen, Eric Olsen, Yashar Niknafs, Arul Chinnaiyan, Corey Speers. Nomination and characterization of TTK for Radiosensitization in basal-like breast cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2926.

  • abstract 5839 androgen receptor ar a novel target for Radiosensitization in triple negative breast cancers tnbc
    Cancer Research, 2017
    Co-Authors: Benjamin C Chandler, Daniel R Wahl, Corey Speers, Shuang G Zhao, Meilan Liu, Kari Wilderromans, Daniel E Spratt, Eric Olsen, Shyam Nyati, Daniel F Hayes
    Abstract:

    Purpose: Increased rates of local recurrence (LR) have been observed in TNBC despite chemotherapy and radiation (RT). Thus, approaches that result in Radiosensitization in TNBC are critically needed. We characterized the RT response of 21 breast cancer cell (BCC) lines using clonogenic survival assays and paired this with high-throughput drug screen data, identifying AR as a top target for Radiosensitization. We demonstrate that AR inhibition confers Radiosensitization in vitro and in vivo, identified a biomarker of response, and characterize the mechanism of AR-mediated Radiosensitization in TNBC. Materials/Methods: Clonogenic survival assays determined the intrinsic RT sensitivity of 21 BCC lines. IC50 values were determined for 130 clinical compounds and correlation coefficients were calculated using IC50 values and SF-2Gy. Gene and protein expression was measured using RNA Seq and RPPA arrays, respectively, in tumor samples (n=2,061) and BCC lines (n=51). AR function was assessed using gene knockdown (KD) or functional inhibition with anti-androgen drugs. We measured in vivo tumor growth with varying control and treatment groups (16-20 tumors/group). Kaplan-Meier analysis was performed to estimate local control. A Cox proportional hazards model and MVA were used to determine variables associated with LRF survival. Results: Our unbiased drug radiosensitizer screen nominated bicalutamide as an effective drug in treating RT-resistant BCC lines (R2= 0.46, p-value 2000 human breast tumor samples and 51 BCC lines and found heterogeneity in AR expression with strongly correlated expression of protein and RNA levels in TNBC (R2=0.89, p-value Conclusion: Our results implicate AR as a mediator of radioresistance in breast cancer and support the rationale for developing clinical strategies, including clinical trials, to inhibit AR as a novel radiosensitizing target in TNBC. Citation Format: Benjamin C. Chandler, Corey W. Speers, Shuang G. Zhao, Meilan Liu, Kari Wilder-Romans, Eric Olsen, Shyam Nyati, Daniel Spratt, Daniel Wahl, Daniel Hayes, Felix Y. Feng, Lori J. Pierce. Androgen receptor (AR): A novel target for Radiosensitization in triple-negative breast cancers (TNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5839. doi:10.1158/1538-7445.AM2017-5839

Daniel R Wahl - One of the best experts on this subject based on the ideXlab platform.

  • abstract 6271 hormone receptor inhibition as a strategy for Radiosensitization of breast cancer
    Cancer Research, 2020
    Co-Authors: Anna R Michmerhuizen, Meilan Liu, Amanda Zhang, Rachel Schwartz, Andrea M Pesch, Benjamin C Chandler, Cassandra L Ritter, Kari Wilderromans, Daniel E Spratt, Daniel R Wahl
    Abstract:

    Purpose: Expression of the androgen receptor (AR) has been identified as a driver of tumor growth in triple negative breast cancers (TNBC), and previous work has nominated AR as a target for Radiosensitization. In addition, 70-95% of all estrogen receptor (ER) positive (ER+) breast cancers also have coexpression of AR, suggesting extended utility of AR inhibition in the Radiosensitization of these AR+, ER+ tumors. Here we assessed the efficacy of AR inhibition in ER+, AR+ breast cancers to better understand the role of AR signaling across breast cancer models. Further, we also investigated the effect of ER inhibition on Radiosensitization of ER+ breast cancer models. Methods: IC50 values were determined for MDV3100 (enzalutamide), ARN-509 (apalutamide), and ODM-201 (darolutamide) in TNBC cell lines (AR+ TNBC: MDA-MB-453, ACC-422, and SUM-185PE, and AR- TNBC: MDA-MB-231) and ER+ breast cancer cell lines (AR+, ER+: ZR-75-1, BT-474, CAMA-1, and AR-, ER+: MCF-7). IC50 values for tamoxifen were determined for ER+ breast cancer cell lines (MCF-7, T47D, ZR-75-1), and ER- (SUM-159) cells. Clonogenic survival assays were performed to assess Radiosensitization with ER or AR inhibition with tamoxifen or second generation anti-androgens, respectively, in TNBC and ER+ breast cancer models. Results: AR inhibition with enzalutamide, apalutamide, and darolutamide showed limited single agent growth inhibition efficacy in AR+ TNBC and AR+, ER+ breast cancer cell lines (IC50 > 10 μM). AR inhibition with enzalutamide did not induce radiosensitivity in vitro. In AR+, ER+ CAMA-1 cells, AR blockade with enzalutamide had a radioprotective effect with enhancement ratios (enhR) of 0.76-0.83. No Radiosensitization was observed in BT-474 (enhR: 0.92-1.01) or ZR-75-1 cells (enhR: 0.94-1.00). Radiosensitization was also assessed with anti-androgens apalutamide and darolutamide in AR+ breast cancer models. Inhibition of ER with tamoxifen, however, induced Radiosensitization in MCF-7 (enhR: 1.14-1.50) and T47D (enhR: 1.33-1.60) cells. No Radiosensitization was observed with tamoxifen in ER- SUM-159 cells. Conclusion: Although AR is a mediator of radioresistance in AR+ TNBC, AR inhibition does not provide comparable Radiosensitization in AR+, ER+ models and may actually confer a radioprotective effect. In contrast, our results demonstrate ER inhibition is an effective radiosensitizing strategy in ER+ breast cancers, independent of AR status. This work highlights the complexities of androgen and estrogen receptor signaling in AR+, ER+ breast tumors and underscores the necessity for understanding context dependent effects when translating into patients with AR+ breast cancer. Citation Format: Anna R. Michmerhuizen, Amanda Zhang, Rachel Schwartz, Andrea M. Pesch, Benjamin C. Chandler, Cassandra L. Ritter, Meilan Liu, Kari Wilder-Romans, Daniel E. Spratt, Daniel R. Wahl, Shyam Nyati, Lori J. Pierce, Corey Speers. Hormone receptor inhibition as a strategy for Radiosensitization of breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6271.

  • parp1 trapping and dna replication stress enhance Radiosensitization with combined wee1 and parp inhibitors
    Molecular Cancer Research, 2017
    Co-Authors: Leslie A Parsels, David Karnak, Joshua D Parsels, Qiang Zhang, Jonathan Velezpadilla, Zachery R Reichert, Daniel R Wahl, Jonathan Maybaum, Mark J Oconnor, Theodore S. Lawrence
    Abstract:

    KRAS mutations in non–small cell lung cancer (NSCLC) cause increased levels of DNA damage and replication stress, suggesting that inhibition of the DNA damage response (DDR) is a promising strategy for Radiosensitization of NSCLC. This study investigates the ability of a WEE1 inhibitor (AZD1775) and a PARP inhibitor (olaparib) to radiosensitize KRAS-mutant NSCLC cells and tumors. In addition to inhibiting the DDR, these small-molecule inhibitors of WEE1 and PARP induce DNA replication stress via nucleotide exhaustion and PARP trapping, respectively. As monotherapy, AZD1775 or olaparib alone modestly radiosensitized a panel of KRAS-mutant NSCLC lines. The combination of agents, however, significantly increased Radiosensitization. Furthermore, AZD1775-mediated Radiosensitization was rescued by nucleotide repletion, suggesting a mechanism involving AZD1775-mediated replication stress. In contrast, Radiosensitization by the combination of AZD1775 and olaparib was not rescued by nucleosides. Whereas both veliparib, a PARP inhibitor that does not efficiently trap PARP1 to chromatin, and PARP1 depletion radiosensitized NSCLC cells as effectively as olaparib, which does efficiently trap PARP, only olaparib potentiated AZD1775-mediated Radiosensitization. Taken together, these mechanistic data demonstrate that although nucleotide depletion is sufficient for Radiosensitization by WEE1 inhibition alone, and inhibition of PARP catalytic activity is sufficient for Radiosensitization by olaparib alone, PARP1 trapping is required for enhanced Radiosensitization by the combination of WEE1 and PARP inhibitors. Implications: This study highlights DNA replication stress caused by nucleotide depletion and PARP1 trapping as an important mechanism of Radiosensitization in KRAS-mutant tumors and supports further development of DNA replication as a therapeutic target. Mol Cancer Res; 16(2); 222–32. ©2017 AACR.

  • abstract 5839 androgen receptor ar a novel target for Radiosensitization in triple negative breast cancers tnbc
    Cancer Research, 2017
    Co-Authors: Benjamin C Chandler, Daniel R Wahl, Corey Speers, Shuang G Zhao, Meilan Liu, Kari Wilderromans, Daniel E Spratt, Eric Olsen, Shyam Nyati, Daniel F Hayes
    Abstract:

    Purpose: Increased rates of local recurrence (LR) have been observed in TNBC despite chemotherapy and radiation (RT). Thus, approaches that result in Radiosensitization in TNBC are critically needed. We characterized the RT response of 21 breast cancer cell (BCC) lines using clonogenic survival assays and paired this with high-throughput drug screen data, identifying AR as a top target for Radiosensitization. We demonstrate that AR inhibition confers Radiosensitization in vitro and in vivo, identified a biomarker of response, and characterize the mechanism of AR-mediated Radiosensitization in TNBC. Materials/Methods: Clonogenic survival assays determined the intrinsic RT sensitivity of 21 BCC lines. IC50 values were determined for 130 clinical compounds and correlation coefficients were calculated using IC50 values and SF-2Gy. Gene and protein expression was measured using RNA Seq and RPPA arrays, respectively, in tumor samples (n=2,061) and BCC lines (n=51). AR function was assessed using gene knockdown (KD) or functional inhibition with anti-androgen drugs. We measured in vivo tumor growth with varying control and treatment groups (16-20 tumors/group). Kaplan-Meier analysis was performed to estimate local control. A Cox proportional hazards model and MVA were used to determine variables associated with LRF survival. Results: Our unbiased drug radiosensitizer screen nominated bicalutamide as an effective drug in treating RT-resistant BCC lines (R2= 0.46, p-value 2000 human breast tumor samples and 51 BCC lines and found heterogeneity in AR expression with strongly correlated expression of protein and RNA levels in TNBC (R2=0.89, p-value Conclusion: Our results implicate AR as a mediator of radioresistance in breast cancer and support the rationale for developing clinical strategies, including clinical trials, to inhibit AR as a novel radiosensitizing target in TNBC. Citation Format: Benjamin C. Chandler, Corey W. Speers, Shuang G. Zhao, Meilan Liu, Kari Wilder-Romans, Eric Olsen, Shyam Nyati, Daniel Spratt, Daniel Wahl, Daniel Hayes, Felix Y. Feng, Lori J. Pierce. Androgen receptor (AR): A novel target for Radiosensitization in triple-negative breast cancers (TNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5839. doi:10.1158/1538-7445.AM2017-5839