bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 35: Diacylglycerol kinase zeta-mediated regulation of mTOR and SREBP-1 offers new opportunities for cancer management
Authors: Torres-Ayuso PEDRO , Jones David , Tello-Lafoz Maria , Avila-Flores Antonia , Merida Isabel
Published: August 2015

Diacylglycerol (DAG) and phosphatidic acid (PA) are two lipids that lie at the core of several metabolic and signaling pathways, which allows integrated control of cell growth and nutrient sensing with cell metabolism. Diacylglycerol kinases (DGK) are a family of enzymes that balance the levels of both lipids. Some DGK isoforms, mainly DGKα, have been recently suggested as potential targets for cancer treatment (Dominguez et al, 2013; Torres-Ayuso et al, 2014). Previous observations from us indicate that the DGKζ isoform contributes the most to DGK activity in cancer cells, and DGKζ-derived PA is required for mTOR activation, a master regulator of cell growth and metabolism (Avila-Flores et al, 2005).

To address the potential of DGKζ as new therapeutic target for cancer treatment, we used several breast and colon cancer cell lines, and analyzed the effect of its pharmacological inhibition or siRNA-mediated knock down in the proliferation, survival and metabolism of these cells.

We found that DGKζ protein levels where higher in malignant cells compared to their non-transformed counterparts, and its depletion abrogated cancer cell growth in colony assays. DGKζ was prominent over other PA sources to activate mTOR, and its activity was induced by rapamycin. We also investigated whether DGKζ-derived PA functions were coordinated with DAG metabolism and signaling. We observed that DGKζ-dependent DAG consumption was crucial for cell homeostasis via control of the SREBP-1 transcription factor, which was negatively regulated by DAG. Our data provide new evidence of the crosstalk between mTOR and lipid metabolism, and suggest that DGKζ represents an effective target for cancer control.

References

Avila-Flores A, Santos T, Rincon E, Merida I (2005) Modulation of the mammalian target of rapamycin pathway by diacylglycerol kinase-produced phosphatidic acid. The Journal of biological chemistry 280: 10091-10099

Dominguez CL, Floyd DH, Xiao A, Mullins GR, Kefas BA, Xin W, Yacur MN, Abounader R, Lee JK, Wilson GM, Harris TE, Purow BW (2013) Diacylglycerol kinase alpha is a critical signaling node and novel therapeutic target in glioblastoma and other cancers. Cancer discovery 3: 782-797

Torres-Ayuso P, Daza-Martin M, Martin-Perez J, Avila-Flores A, Merida I (2014) Diacylglycerol kinase alpha promotes 3D cancer cell growth and limits drug sensitivity through functional interaction with Src. Oncotarget 5: 9710-9726

Citation Format: PEDRO Torres-Ayuso, David Jones, Maria Tello-Lafoz, Antonia Avila-Flores, Isabel Merida. Diacylglycerol kinase zeta-mediated regulation of mTOR and SREBP-1 offers new opportunities for cancer management. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 35. doi:10.1158/1538-7445.AM2015-35
Abstract 3522: BET and BRAF inhibitors act synergistically against BRAF mutant melanoma
Authors: Paoluzzi Luca , Hanniford Douglas , Sokolova Elena , Osman Iman , Darvishian Farbod , Wang Jinhua , Bradner James E. , Hernando Eva
Published: August 2015

Purpose: Despite major advances in the treatment of metastatic melanoma, treatment failure is still inevitable in most cases. Manipulation of key epigenetic regulators, including inhibition of Bromodomain and extra-terminal domain (BET) family members impairs cell proliferation in vitro and tumor growth in vivo in different cancers, including melanoma. BET inhibition is thought to impair downstream transcriptional activity of the MAPK (MAPK) pathway and other key oncogenic signaling pathways, suggesting complementarity with MAPK inhibition. Here, we investigated the effect of combining the BET inhibitor JQ1 with the BRAF inhibitor Vemurafenib in in vitro and in vivo models of BRAF-mutant melanoma. We additionally screened potential synergism of JQ-1 with 27 additional compounds including inhibitors of the proteasome, histone deacetylases (HDACi) and MEK.

Methods: We performed cytotoxicity and apoptosis assays, and a xenograft mouse model to determine the in vitro and in vivo efficacy of JQ1 in combination with Vemurafenib against BRAF-mutant melanoma cell lines. Further, to investigate the molecular mechanisms underlying the effects of this combined treatment, we conducted antibody arrays of in vitro drug-treated cell lines and RNA sequencing of drug-treated xenograft tumors. Synergism between JQ1 and other targeted compounds was examined by in vitro cytotoxicity assays.

Results: The combination of JQ1 and Vemurafenib acted synergistically in BRAF-mutant cell lines. Additional combinations that were found synergistic included JQ-1 plus proteasome inhibitors (carfilzomib and marizomib, HDAC inhibitors (romidepsin and panobinostat), or anti-Bcl2 small molecules (Obatoclax and TW-37). The combination of JQ-1 and Vemurafenib resulted in marked apoptosis in vitro, with up-regulation of several pro-apoptotic proteins involved in both the intrinsic and extrinsic pathways of apoptosis. In vivo, this combination treatment suppressed tumor growth and significantly improved survival compared to either drug alone. RNA sequencing of tumor tissues revealed almost four thousand genes that were uniquely modulated by the combination. Furthermore, several anti-apoptotic genes such as MCL-1, BCL-XL and BCL-2 were significantly down-regulated.

Conclusions: Collectively, our data provide a rationale for combinations of BET inhibitors with additional compounds including BRAF inhibitors, as a novel strategy for the treatment of melanoma.

Citation Format: Luca Paoluzzi, Douglas Hanniford, Elena Sokolova, Iman Osman, Farbod Darvishian, Jinhua Wang, James E. Bradner, Eva Hernando. BET and BRAF inhibitors act synergistically against BRAF mutant melanoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3522. doi:10.1158/1538-7445.AM2015-3522
Abstract 3552: Metabolite profiling reveals the glutathione biosynthetic pathway as a therapeutic target in triple negative breast cancers
Authors: Beatty Alexander
Published: August 2015

Identifying metabolic pathway alterations that are critical to support cancer growth is a key hurdle for developing therapeutic strategies that exploit these pathways. We have applied metabolomic and pharmacological approaches to identify targetable pathways in triple negative breast cancer (TNBC). TNBC is an aggressive and genomically heterogeneous subset of breast cancer that is resistant to existing targeted therapies. To identify dysregulated metabolic pathways in TNBC, we conducted mass spectrometry-based metabolomics of TNBC and control cells. The relative levels of 155 intracellular metabolites distinguished TNBC from non-transformed breast epithelia, and revealed two metabolic subtypes within TNBC that, unexpectedly, correlate with markers of basal-like versus non-basal-like status. Distinguishing metabolites included amino acids, lipids and the cellular redox buffer glutathione. Levels of glutathione were generally lower in TNBC cell lines compared to controls, and markedly lower in the metabolic subtype containing non-basal-like TNBC. Further, these cell lines showed enhanced sensitivity to inhibition of glutathione biosynthesis, showing a dependence on glutathione production for survival. These findings demonstrate the potential of targeting the glutathione biosynthetic pathway as a therapeutic strategy in TNBC, and suggest that existing clinical biomarkers may provide a means for stratifying TNBC tumors to identify likely responders to anti-glutathione therapy.

Citation Format: Alexander Beatty. Metabolite profiling reveals the glutathione biosynthetic pathway as a therapeutic target in triple negative breast cancers. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3552. doi:10.1158/1538-7445.AM2015-3552
Abstract 3594: P-glycoprotein attenuates Src activation and DNA repair activity via increased C-terminal Src kinase-binding protein, a negative regulator of Src, in multidrug-resistant cells
Authors: Lin Li-Fang , Wu Ming-Hsi , Su Tsann-Long , Lee Te-Chang
Published: August 2015

Multidrug resistance (MDR) remains a significant obstacle to the success of cancer chemotherapy. MDR is often associated with increased expression of ATP-binding cassette transporter family members, which extrude anticancer drugs out of cells. The MDR1 gene product, P-glycoprotein (P-gp) is one of the most well-known ABC transporters and expels a broad range of anticancer drugs, such as vinblastine, vincristine, doxorubicin and paclitaxel. Overexpression of P-gp in tumor tissues is thus a prognostic indicator associated with poor response to chemotherapy and poor clinical outcome. Our previous study has shown that P-gp overexpression attenuates the repair activity of DNA interstrand crosslinks (ICLs) and hence contributes the susceptibility of MDR cells to DNA ICL agents [1]. In the present study, we explored the molecular mechanisms underlying which P-gp interferes with Src-activated DNA repair activity. We first confirmed that ectopic expression of P-gp attenuates DNA ICL repair activity and becomes more susceptible to DNA ICL agents than vector control cells. Our present results showed that the C-terminal Src kinase-binding protein (Cbp), a negative regulator of Src, is significantly enhanced in MDR cells. Cbp silencing by siRNA technique apparently enhances cisplatin-induced Src activation and DNA repair activity, and hence increases the resistance of MDR cells to ICL agents. By aid of immuneprecipitation and immunofluorescence staining techniques, we demonstrated that P-gp not only interacts with Cbp and Src but also enhances the formation of inhibitory Csk-Cbp complex that reduces phosphorylation of Src activation residue Tyr416 and increases phosphorylation of Src negative regulatory Tyr527 residue. Taken together, the current results indicate that P-gp overexpression in MDR cells attenuates DNA repair activity by mediating Cbp dependent mechanism to interfere with Src activation. These results implicate that increasing the expression of Cbp is potentially a therapeutic strategy to increase the susceptibility of cancer cells to chemotherapeutic agents.

1. Lee PC, Lee HJ, Kakadiya R, Sanjiv K, Su TL, Lee TC: Multidrug-resistant cells overexpressing P-glycoprotein are susceptible to DNA crosslinking agents due to attenuated Src/nuclear EGFR cascade-activated DNA repair activity. Oncogene 2013, 32:1144-1154.

Citation Format: Li-Fang Lin, Ming-Hsi Wu, Tsann-Long Su, Te-Chang Lee. P-glycoprotein attenuates Src activation and DNA repair activity via increased C-terminal Src kinase-binding protein, a negative regulator of Src, in multidrug-resistant cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3594. doi:10.1158/1538-7445.AM2015-3594
Abstract 3607: Loss-of-function screens using haploid KBM7 and HAP1 cells to identify mechanisms of anti-cancer drug resistance
Authors: Gerhards Nora M. , Guyader Charlotte , Blomen Vincent A. , Küçükosmanoğlu Aslı , van Tellingen Olaf , Vis Daniel J. , Wessels Lodewyk F. , Brummelkamp Thijn R. , Borst Piet , Rottenberg Sven
Published: August 2015

The lack of biomarkers to predict anti-cancer therapy response remains a major obstacle in the treatment of various cancers. This handicap not only affects novel targeted therapies. Also for classical chemotherapeutic drugs such as DNA cross-linking agents, topoisomerase inhibitors or microtubule-targeting compounds, which are all frequently used in the clinic, we do not understand well why some patients have a major benefit of the therapy whereas others do not. A decreased intracellular drug accumulation has been proposed as one potential mechanism, but the clinical relevance of known uptake and efflux mechanisms is still controversial for various chemotherapeutic drugs. To identify novel factors that may be relevant for the intracellular concentration of drugs, we carried out loss-of-function screens using haploid KBM7 and HAP1 cells. In particular, we used the PARP inhibitor olaparib, the topoisomerase I inhibitor topotecan, platinum drugs, and the microtubule-targeting compounds docetaxel and vinorelbine to select resistant clones. We will present results from these screens and their validation, including independent cell lines and TCGA data sets.

Citation Format: Nora M. Gerhards, Charlotte Guyader, Vincent A. Blomen, Aslı Küçükosmanoğlu, Olaf van Tellingen, Daniel J. Vis, Lodewyk F. Wessels, Thijn R. Brummelkamp, Piet Borst, Sven Rottenberg. Loss-of-function screens using haploid KBM7 and HAP1 cells to identify mechanisms of anti-cancer drug resistance. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3607. doi:10.1158/1538-7445.AM2015-3607
Abstract 3609: In vitro comparison of conventional hyperthermia and electro-hyperthemia
Authors: Chi Kwan-Hwa , Wang Yu-Shan Samuel , Huang Cheng-Chung , Chang Chao-Chun , Chi Mau-Shin , Yang Kai-Lin , Tsang Yuk-Wah , Andocs Gabor
Published: August 2015

Purpose:

Deep heating hypertermia methods are a promising cancer treatment modalities used to enhance conventional treatment. This could be achieved through radiofrequency field en-bloc heating by Thermotron RF-8 and selective tumor heating by Oncotherm-Labehy, respectively. This is the first study to evaluate the different cytotoxic effect and action mechanism on cancer cells after mild hyperthermia (MHT) by Thermotron RF-8 or Oncothermia-Labehy.

Experimental Design:

Hepatoma HepG2 cells were used for MHT treatment (42°C for 30minute). Either by Oncotherm-Labehy or Thermotron RF-8 or water bath treatment were compared. Apoptosis evaluated 24h after MTH as measured by Annexin-V assay and cell cycle analysis. Mitochondria-targeted damage as measured by ROS production with or without Mito-TEMPO treatment and membrane-targeted damage as measured by caspase-8 expression, adhesion molecule expression, heat shock protein (HSP) release and calcium channel disturbance were evaluated and compared.

Results:

MHT resulted in similar cell cycle arrest on G2/M. Oncothermia treatment resulted in higher apoptosis rate (38.87±6.58) than RF-8 (25.13±2.26), water bath (18.77±0.35) and control (14.93±0.76). The expression of Caspase-8 is higher by oncothermia treatment, while caspase-3 and 9 levels were similar between 3 types of treatment. HSPs were upregulated by MHT, however a marked HSP release outside cell and E-cadherin overexpression on the membrane were only observed after oncothermia treatment.

Conclusion:

In this report, we found that different hyperthermia devices may induce different mechanisms on their cytotoxic effects and cell responses to heat. Oncothermia may selectively target on cell membrane before targeting on mitochondria, while RF-8 has no membrane targeting effect. These results may provide rational basis for future clinical application of hyperthermia treatment.

Citation Format: Kwan-Hwa Chi, Yu-Shan Samuel Wang, Cheng-Chung Huang, Chao-Chun Chang, Mau-Shin Chi, Kai-Lin Yang, Yuk-Wah Tsang, Gabor Andocs. In vitro comparison of conventional hyperthermia and electro-hyperthemia. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3609. doi:10.1158/1538-7445.AM2015-3609
Abstract 3610: DMH4, A VEGFR2 inhibitor effectively suppresses proliferation and migration of lung cancer cells
Authors: Li Hao , Bae Chay , Gazy Nicky , Hao Jijun , Zhong Li
Published: August 2015

Background

Currently there is a major unmet medical need for effective therapy of the non-small cell lung cancer (NSCLC). It is well known that vascular endothelial growth factor (VEGF) signaling plays a critical role in regulating tumor angiogenesis, and bevacizumab is the only approved antiangiogenic agent for the treatment of NSCLC to date. Due to individual differences in drug response, developing novel angiogenic inhibitors is essential for NSCLC treatment. We have recently developed DMH4, a small molecule which specifically inhibits VEGF signaling by targeting VEGF receptor 2 (VEGFR2) (Hao, et al 2010). Here, we show that DMH4 can effectively suppress proliferation and migration of NSCLC cells.

Method

Briefly, NSCLC cell line H460, A549 and a lung fibroblast cell line MRC5 was treated by DMH4 at concentrations of 0.25 ug/ml, 0.5 ug/ml,1 ug/ml, 2 ug/ml, 4 ug/ml, 8 ug/ml and 16 ug/ml. MTT assay was taken out for testing cell viability at 24 hr, 48 hr and 72 hr. Meanwhile, FITC Annexin V/PI double staining was taken out to distinguish apoptosis and necrosis. Scratch assay was taken out to evaluate cell migration. Cell lysate was collected at the same time. Phospho-VEGF Receptor 2 level and Phospho-Akt level were tested through Western Blot. 43 apoptosis related protein levels were measured using human apoptosis antibody array assay.

Result

The data from MTT assay showed that DMH4 treatment decreased cell viability in a dose-depend manner. At the concentrations of 1 ug/ml, DMH4 effectively decreased viabilities of both H460 (82.1%, P<0.01) and A549 (78.5%, P<0.01) NSCLC cells, but had no effect on the normal lung MRC5 cells (103.6%). In addition, our study demonstrated that DMH4 promoted apoptosis in NSCLC cells, but did not show significantly effects in normal lung cells. Our further study indicated that DMH4 also suppressed NSCLC migration. Through Western Blot, phospho-VEGFR2 and phosphor-Akt levels were both detected lower in H460 and A549 than in MRC5.

Conclusion

As it can effectively suppress cell proliferation and migration in NSCLC cells but not in normal lung cells, DMH4 may be a good drug lead as a new VEGFR2 inhibitor for NSCLC treatment in the future.

Note: This abstract was not presented at the meeting.

Citation Format: Hao Li, Chay Bae, Nicky Gazy, Jijun Hao, Li Zhong. DMH4, A VEGFR2 inhibitor effectively suppresses proliferation and migration of lung cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3610. doi:10.1158/1538-7445.AM2015-3610
Abstract 3621: Deubiquitination of AXIN1 by USP9X promotes apoptosis in melanoma in response to BRAF or MEK inhibition
Authors: Potu Harish , Peterson Luke F. , Kandarpa Malathi , Fearon Eric , Talpaz Moshe , Donato Nicholas J.
Published: August 2015

The Wnt/β-catenin pathway is important in development, transformation and self-renewal of various tissues and tumors. In contrast, Wnt/β-catenin signaling appears to play a paradoxical role in melanoma where its activation is anti-oncogenic and a marker of good prognosis. Previous studies have shown that activation of the NRAS/BRAF/MEK pathway suppresses Wnt/β-catenin signaling and reduces apoptotic sensitivity to BRAF or MEK inhibitors in melanoma. MEK pathway activation stabilized AXIN1, a Wnt/β-catenin inhibitor, in some melanoma cell lines through a proteasome-dependent process and co-treatment with Wnt3A and BRAF/MEK kinase inhibitors resulted in a reduction of AXIN1 and full activation of apoptosis. Further, AXIN1 knockdown sensitized BRAF/MEK inhibitor insensitive cells to kinase inhibition, providing a prominent role for AXIN1 in determining melanoma cell apoptotic responsiveness and exposing a potential mechanism for improving kinase inhibitor clinical activity. While AXIN1 protein levels appear to be regulated by ubiquitination, the enzymes involved in control of its ubiquitination or deubiquitination are not fully defined. Here we show that ubiquitin-specific peptidase 9, X-linked (USP9X), a deubiquitinating enzyme whose expression is elevated in melanoma, binds and deubiquitinates AXIN1. USP9X knockdown resulted in increased ubiquitination and proteasome-mediated depletion of AXIN1 (but not AXIN2), resulting in a reduction in AXIN1 cellular half-life. USP9X knockdown elevated β-catenin response genes (>2-fold) in melanoma cells treated with Wnt3A. USP9X depletion also increased apoptotic responsiveness to Wnt3A or BRAF/MEK inhibitor and overcame BRAF inhibitor resistance in A375R cells. In contrast, USP34 which was previously shown to regulate AXIN1 (and AXIN2) deubiquitination, did not alter AXIN1 levels or apoptotic responsiveness in melanoma. A previously identified small molecule USP9X inhibitor (EOAI3402143 or G9) replicated the effects of USP9X knockdown on AXIN1 depletion in vitro and in vivo, suppressed melanoma tumor growth in animals and fully blocked in vivo tumor growth when combined with BRAF kinase inhibitor. Thus, we identify USP9X as a potential therapeutic target in melanoma and established G9 as a lead compound for the development of AXIN1 depleting agents that may add to the targeted therapies already used in the treatment of melanoma.

Note: This abstract was not presented at the meeting.

Citation Format: Harish Potu, Luke F. Peterson, Malathi Kandarpa, Eric Fearon, Moshe Talpaz, Nicholas J. Donato. Deubiquitination of AXIN1 by USP9X promotes apoptosis in melanoma in response to BRAF or MEK inhibition. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3621. doi:10.1158/1538-7445.AM2015-3621
Abstract 3626: Expression of BARD1 splice variant impairs homologous recombination and can predict PARP1 inhibitor therapy in colorectal cancer
Authors: Ozden Ozkan , Bishehsari Faraz , Bauer Jessica , Baik Seung Hyua , Jung Barbara
Published: August 2015

Background & Aims: BRCA1 associated RING domain protein 1 (BARD1) stabilizes BRCA1 protein by forming a heterodimeric RING-RING complex. Its alterations impact function of BRCA1, disabling HR (homologous recombination) repair leading to sensitivity to PARP-1 inhibition in cancer. In colorectal cancer (CRC), we observed expression of BARD1 splice variants (SVs) and loss of full-length is associated with poor prognosis despite wild type expression of BRCA1. Here, we examined the roles of BARD1 SV and its therapeutic impact on PARP-1 inhibition, oxaliplatin and irinotecan sensitivity in CRC cell lines.

Methods: A panel of CRC cell lines was treated with PARP-1 inhibitor, ABT888, and HR proficiency assessed via RAD51 foci formation. BARD1 splice variant (SV B1/4) transcription was determined via real-time PCR, and protein expression assessed via immunoblotting. Tumorigenesis of CRC cells expressing SV B1/4 and wild type BARD1 was measured by epithelial-mesenchymal transitions (EMT), colony formation abilities, and migration assays.

Results: Caco-2 CRC cells were significantly more sensitive to PARP-1 inhibition when compared to the other CRC cell lines. This sensitivity was not associated with loss of BRCA1 or full-length BARD1 expression but rather with expression of BARD1 SV 1/4. Caco-2 showed significantly impaired HR response following PARP-1 inhibition with ABT888 and characterized by decreased RAD51 foci formation. Exogenous expression of the BARD1 SV B1/4 in SW480 cells exhibits a more transformed phenotype, decreased RAD51 foci formation, reduced BRCA1 E3 ubiquitin ligase activity, and increased sensitivity to PARP-1 inhibitor. Addition of irinotecan to ABT888 pre-treated cells created higher specificity and sensitivity with respect to killing of SV B1/4 expressing SW480 cells. Further immunofluorescence and confocal microscopy studies are underway to investigate the effect of BARD1 SV B1/4 on BRCA1 homologous recombination DNA repair function.

Conclusions: Expression of BARD1 SV might be a potential biomarker for therapeutic sensitivity to PARP-1 inhibitors in treatment of CRC.

Citation Format: Ozkan Ozden, Faraz Bishehsari, Jessica Bauer, Seung Hyua Baik, Barbara Jung. Expression of BARD1 splice variant impairs homologous recombination and can predict PARP1 inhibitor therapy in colorectal cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3626. doi:10.1158/1538-7445.AM2015-3626
Abstract 3630: Inhibition of PARP by hyperthermia or pharmacological inhibitors synergize with cisplatin and doxorubicin but not with 5-FU or paclitaxel
Authors: Schaaf Lea , Ulmer Christoph , Steurer Wolfgang , Mürdter Thomas E. , Aulitzky Walter E. , van der Kuip Heiko
Published: August 2015

Short-term, high-dose chemotherapy together with hyperthermia (HIPEC) is widely used for treatment of peritoneal carcinomatosis. However, it is fairly known which agents currently used for HIPEC actually act synergistically with elevated temperature. Furthermore, the molecular mechanisms behind the potentiating effects are incompletely understood. To mirror the clinical HIPEC setting we cultivated 2 ovarian and 2 colon cancer cell lines under hypoxic conditions and performed a 1-hour treatment at atmospheric oxygen levels with Cisplatin, Doxorubicin, 5FU, or Paclitaxel at either 37°C or 42°C. Cells were then evaluated for long-term survival, DNA strand break repair, poly(ADP-Ribosy)lation, and γH2AX and P-53BP1 foci formation. Importantly, we found that Cisplatin as well as Doxorubicin were potentiated by hyperthermia as revealed by a significantly reduced colony forming ability whereas high temperature had no effects together with 5FU or Paclitaxel on long-term survival. The synergistic effects on survival were due to compromised DNA strand break repair capacity and followed by a significantly enhanced number of double strand breaks. Interestingly, this was paralleled by a diminished poly(ADP-Ribosy)lation indicating a reduced activity of PARP. The role of this enzyme for the hyperthermia effects was confirmed by using clinically relevant PARP inhibitors. Combination of PARP inhibitors was synergistic with the same subset of compounds as observed with hyperthermia. Short-term treatment with PARP inhibitors mimicked the effects of elevated temperature both on survival and DNA repair capacity. Hyperthermia and PARP inhibitors also led to a comparable increase of γH2AX and P-53BP1 foci in cells treated with Doxorubicin or Cisplatin.

In conclusion our data indicate that the cytotoxic effects of only a subset of currently used chemotherapeutics can be potentiated by hyperthermia. The molecular mechanisms behind these synergistic effects are related to PARP functions in DNA repair.

Citation Format: Lea Schaaf, Christoph Ulmer, Wolfgang Steurer, Thomas E. Mürdter, Walter E. Aulitzky, Heiko van der Kuip. Inhibition of PARP by hyperthermia or pharmacological inhibitors synergize with cisplatin and doxorubicin but not with 5-FU or paclitaxel. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3630. doi:10.1158/1538-7445.AM2015-3630
Abstract 3634: A gain-of-function mutation in tyrosine phosphatase SHP-2 enhances breast carcinoma malignant phenotypes
Authors: Hu Zhongqian , Fang Haoshu , Wang Siying
Published: August 2015

Breast cancer is the most common malignant tumor in women worldwide, and it is the second leading cause of cancer-related mortality in the US women. Evidences suggested that overexpression of Src homologous protein phosphotyrosyl phosphatase 2 (SHP2) has been observed in most of the breast cancer cell lines and in breast cancer tissues, and accompanied with lymph nodes metastasis. Inhibition of SHP2 in the breast cancer cell lines abolishes the growth and decrease survival of tumor cells, which leads to the differentiation of malignant cells into normal breast epithelial phenotype. These observations suggest that SHP2 promotes the tumor development though increase tumor formation and metastasis. However, the pathophysiologic role of SHP2 (overexpression or gain-of-function mutation) and its relevance to tumorigenesis are still largely unknow. In this study, we focused on the in vivo and in vitro effects of the gain-of-function mutant SHP-2 on phenotypes of breast cancer. Mutant PTPN11 cDNA (D61G) was transduced into MDA-MB231 and MCF-7 cells. The effects of D61G mutant on breast cancer tumorigenesis and malignant behaviours, such as cell adhesion, proliferation, migration and invasion were tested. The potential underlying molecular mechanisms, i.e., the activation of the Gab1-Ras-Erk axis were also examined. In vitro studies found that, in SHP2 D61G mutant groups, the tumour adhesion, proliferation, migration and invasion were increased significantly. Consistently, in vivo studies also found that the incidence of tumour and the tumour weights of the SHP-2 D61G-MB231 group were increased significantly (P<0.001), which is also associated with tumour metastasis. Mechanistically, the PTPN11 D61G mutation resulted in the activation of Ras-ErK pathway. And interestingly, the interaction between Gab1 and mutant SHP2 was also significantly increased. The mutant SHP-2 significantly promoted tumour migration and invasion at least partially through the activation of Gab1-Ras-Erk axis. In conclusion, our findings clearly establish a role for the SHP-2 GOF mutant in breast tumour growth and provide novel insight into the mechanisms of SHP-2 GOF mutant-mediated tumor promotion. This finding could have direct implications for breast cancer therapy.

This study was supported by a grant from the National Natural Science Foundation of China (codes: 30873046, 30973424, and 81072663) and by the Fundamental Research Funds for the Central Universities (code:3290004402).

Note: This abstract was not presented at the meeting.

Citation Format: Zhongqian Hu, Haoshu Fang, Siying Wang. A gain-of-function mutation in tyrosine phosphatase SHP-2 enhances breast carcinoma malignant phenotypes. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3634. doi:10.1158/1538-7445.AM2015-3634
Abstract 3647: Targeting cancer-induced anemia with hepcidin lowering ALK2 inhibitors
Authors: Peterson Peter W.
Published: August 2015

Anemia of cancer (AOC) is a cytokine driven condition that frequently occurs in patients due to chronic inflammation associated with cancer development. It differs from chemotherapy-induced anemia because the anemia is a result of the disease itself and not a side effect of a treatment regimen, although the both causes can work in concert. AOC is caused by high levels of the liver peptide hormone, hepcidin, which sequesters iron in epithelial cells and macrophages, thereby limiting bioavailable iron required for erythropoiesis. Reducing elevated hepcidin levels in cancer patients can reverse the hypoferremia and anemia seen in AOC. Hepcidin levels are controlled by inflammatory cytokines through pathways dependent on bone morphogenetic proteins (BMP). BMPs up-regulate hepcidin by activating the SMAD signaling pathway through the activin-like kinase receptor 2 (ALK2). Therefore, ALK2 has emerged as an attractive therapeutic target to modulate hepcidin levels in cancer patients and treat their AOC. We have developed novel small molecule ALK2 inhibitors which have shown promising results in tissue culture models as well as a preclinical cancer model of AOC. The two most promising compounds downregulate hepcidin expression in BMP-induced cell culture studies at concentrations of 100 nM or lower. Importantly, this activity was observed at concentrations that exhibited little cytotoxicity suggesting the compounds have a clean selectivity profile. The two compounds have been tested in a TC-1 syngeneic mouse model of AOC where anemia is induced after tumor formation, in a manner similar to human cancer caused anemia. Treatment with the lead candidates reversed the induction of hepcidin expression and also decreased the symptoms of anemia as measured by serum iron and red blood cell levels. From these data, we have nominated a candidate to advance into IND-enabling studies that has favorable drug-like properties. We anticipate a clinical development strategy that focuses on AOC.

Citation Format: Peter W. Peterson. Targeting cancer-induced anemia with hepcidin lowering ALK2 inhibitors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3647. doi:10.1158/1538-7445.AM2015-3647
Abstract 3649: Broad profiling reveals opportunities for selective inhibition of disease-associated mutant kinases
Authors: Duong-Ly Krisna C. , Devarajan Karthik , Liang Shuguang , Horiuchi Kurumi , Wang Yuren , Ma Haiching , Peterson Jeffrey R.
Published: August 2015

Small molecule kinase inhibitors are promising therapeutic agents in a number of diseases, most notably cancer. However, mutations in kinases, both intrinsic and acquired, can drastically alter inhibitor sensitivity. To identify inhibitors of disease-associated mutant kinases, we conducted an unbiased functional screen of 182 small molecule kinase inhibitors against 76 mutated recombinant kinases arising from 21 cognate wild-type kinases. The results revealed novel lead compounds that were exquisitely selective for mutant kinases, including several that exhibited preferred inhibition of mutant kinases over their cognate wild-type kinases. This study provides a resource for the development of novel small molecule inhibitors against disease-associated mutant kinases and illustrates the potential of unbiased large-scale profiling as an approach to compound-centric kinase inhibitor discovery.

Citation Format: Krisna C. Duong-Ly, Karthik Devarajan, Shuguang Liang, Kurumi Horiuchi, Yuren Wang, Haiching Ma, Jeffrey R. Peterson. Broad profiling reveals opportunities for selective inhibition of disease-associated mutant kinases. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3649. doi:10.1158/1538-7445.AM2015-3649
Abstract 3671: Loading and molecular labeling of cell-specific exosomes by HDL-like AuNPs
Authors: Angeloni Nicholas L. , McMahon Kaylin M. , Thaxton C. Shad
Published: August 2015

Exosomes are 30-100 nm diameter lipid vesicles produced by most cells. Exosomes carry diverse cargo, such as RNA and proteins. Cancer cells produce an abundant amount of exosomes, which are released from cells of the primary tumor into the tumor microenvironment and the circulation. Data show that exosomes can promote cancer progression and metastasis by delivering, for instance, pro-metastatic factors to pre-metastatic sites. It is not currently possible to specifically separate tumor-derived exosomes from complex matrices. This ability would provide a better understanding of their contribution to tumorigenesis and cancer progression. Furthermore, the targeting properties of exosomes make them appealing vehicles for encapsulating and delivering therapeutic cargo. However, it is challenging to isolate and then load exosomes with specific drugs. Uniquely, our data reveal that synthetic high-density lipoprotein nanoparticles (HDL NPs) can be tailored to achieve cell-specific exosome labeling and loading of therapeutic cargo. HDL NPs are synthesized using a 5 nm diameter spherical gold nanoparticle. The gold NP serves as a template to control size and shape. Apolipoprotein A-I and phospholipids are conjugated to the gold surface, yielding a synthetic HDL NP similar in size, shape, surface chemistry, and function to some natural HDLs. Our data demonstrate that, upon addition of HDL NPs to cultured CWR 22Rv1 prostate cancer cells, they are packaged into exosomes that are secreted from these cells. Furthermore, HDL NPs made using phospholipids functionalized with either biotin or fluorophore molecules leads to the incorporation of the phospholipids into the exosomes. Thus, the specific cell-derived exosomes are labeled with gold, a molecular fluorophore label, and/or biotin. In each case, the specific exosomes are easy to isolate using methods such as fluorescent isolation or by the robust streptavidin-biotin interaction. Isolated exosomes retain the same size and molecular markers as exosomes from untreated cells. Overall, this unique method may provide control over exosomes for a myriad of diagnostic and therapeutic uses.

Citation Format: Nicholas L. Angeloni, Kaylin M. McMahon, C. Shad Thaxton. Loading and molecular labeling of cell-specific exosomes by HDL-like AuNPs. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3671. doi:10.1158/1538-7445.AM2015-3671
Abstract 3678: Imaging AZD1152HQPA Accurin™ nanoparticle accumulation in preclinical tumors
Authors: Goodwin Richard , Swales John , Nilsson Anna , Andren Per , Strittmatter Nicola , Takats Zoltan , Howes Colin , Taylor Paula , Ashton Susan , Jewsbury Philip , Barry Simon T.
Published: August 2015

AZD1152HQPA Accurin is a passively targeted nanoparticle containing the active metabolite of the Aurora B kinase prodrug AZD1152. In preclinical studies, AZD1152HQPA Accurin displays reduced bone marrow toxicity associated with AZD1152 together with increases in anti-tumour activity. Nanoparticles are proposed to enable delivery and retention of drug to the tumour thereby increasing efficacy and modifying therapeutic index. While it is possible to demonstrate increased duration of modulation of pharmacodynamic biomarkers and drug exposure by nanoparticle delivery, directly demonstrating that nanoparticles accumulate in the tumour is challenging. To investigate the intra tumoural distribution of AZD1152HQPA Accurin nanoparticles we have employed mass spectrometry techniques to image samples derived from human colorectal cancer SW620 studies in rats. We have separately identified the nanoparticle itself, drug encapsulated in nanoparticle, and drug released from nanoparticle; the latter by detecting a metabolite that is only generated from released drug. First MALDI (matrix assisted laser desorption ionization) which offers 20 μm spatial resolution, was used to detect AZD1152-hQPA (released and encapsulated). Second LESA (liquid extraction surface analysis), which offers 1000 μm spatial resolution, was used to detect drug and metabolite, enabling co-distribution to be demonstrated. Finally DESI (desorption electrospray ionization) which offers 100 μm spatial resolution was used to detect drug, metabolite and nanoparticle constituent showing the co-distribution in more detail. Using this combination of approaches it is possible to demonstrate the presence of nanoparticle-delivered AZD1152HQPA metabolite in the tumour at extended timepoints; co-located with nanoparticle and unchanged (encapsulated) drug. At these time points no drug remains if delivered as a simple IV infusion. Collectively this imaging mass spectrometry analysis demonstrates AZD1152HQPA Accurin accumulation in pre-clinical tumours and confirms that the Accurin accesses the tumour and achieves sustained drug release within the tumour.

Citation Format: Richard Goodwin, John Swales, Anna Nilsson, Per Andren, Nicola Strittmatter, Zoltan Takats, Colin Howes, Paula Taylor, Susan Ashton, Philip Jewsbury, Simon T. Barry. Imaging AZD1152HQPA Accurin™ nanoparticle accumulation in preclinical tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3678. doi:10.1158/1538-7445.AM2015-3678
Abstract 37: Role and regulation of CYP24A1 in endometrial cancer
Authors: Bokhari Amber A. , Lee Laura R. , Dewayne Raboteau , Hamilton Chad A. , Maxwell George L. , Rodriguez Gustavo C. , Syed Viqar
Published: August 2015

The cytochrome P450 enzyme, 24-hydroxylase, encoded by CYP24A1 is critical for the catabolism of 1,25(OH)2D3 (calcitriol). The unbalanced high levels of CYP24A1 seem to be a determinant of calcitriol resistance in tumors. We have previously shown that progesterone enhances calcitriol antitumor activity by upregulating vitamin D receptor expression and promoting apoptosis in endometrial cancer cells. In the present study, we evaluated CYP24A1 protein expression in normal and endometrial tumor tissues, assessed the effect of progesterone and calcitriol on CYP24A1 and its spliced variant expression in endometrial cancer cell lines and correlated this with tumor cell growth. Expression of CYP24A1 was assessed in tissue microarrays by immunohistochemistry. A grade-dependent increase of CYP24A1 expression was found in endometrial carcinomas. Endometrial cancer cells expressed high levels of CYP24A1 compared to immortalized endometrial epithelial cells. Furthermore, CYP24A1 is induced in cells in response to calcitriol. Regulation of CYP24A1 by progesterone, progestin derivatives, calcitriol and their combination was examined by RT-PCR and Western blotting following 8, 24, 72 and 120 h exposure of cells to hormones. In all cancer cell lines, progesterone, medroxyprogesterone acetate and norgestrol attenuated

calcitriol-induced CYP24A1, spliced variant CYP24SV transcripts and protein expression at 72 and 120 h of treatment. Furthermore, knockdown of CYP24A1 gene expression by siRNA sensitized endometrial cancer cells to the growth suppressive effect of calcitriol. The data suggest that CYP24A1 overexpression limits calcitriol anti-proliferative signaling in cancer cells, and provide evidence that progestins may be beneficial in preserving calcitriol action in endometrial cancer.

Citation Format: Amber A. Bokhari, Laura R. Lee, Raboteau Dewayne, Chad A. Hamilton, George L. Maxwell, Gustavo C. Rodriguez, Viqar Syed. Role and regulation of CYP24A1 in endometrial cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 37. doi:10.1158/1538-7445.AM2015-37
Abstract 3702: Cumulative incidence rates of breast cancer for Filipinos and Chamorros in Guam and the United States
Authors: Badowski Grazyna , Leon Guerrero Rachael , Simsiman Brayan , Novotny Rachel , Wilkens Lynne
Published: August 2015

Background

Breast cancer is the leading cause of cancer-related death among women on Guam. However incidence rates vary between Chamorros and Filipinos, the two predominant ethnic groups on Guam.

Objective

Our objective was to compare age-specific cumulative incidence rates among ethnic groups in Guam and US. Cumulative incidence rates are relatively stable and thus provide a good estimate of cumulative risk of breast cancer.

Methods

This study included all breast cancer cases (n = 576) reported to the Guam Cancer Registry from 2000 to 2009 and all breast cancer cases (n = 678,637) reported to the Surveillance, Epidemiology,and End Results (SEER) 18-Registry Database. Cumulative incidence rates were calculated for Filipinos and Chamorros for two time periods: 2000-2004 and 2005-2009. For comparison, cumulative incidence rates were also calculated for all women in Guam and the US, and for Filipino only in the US.

Results

Remarkable differences were seen in cumulative incidence rates by age and ethnicity. During the 2000-2004 time period, the cumulative incidence rates by age 40 in Filipinos on Guam were the highest (0.7%) and almost threefold higher than the rate in Chamorros (0.24%). But after age 40, the rates for Chamorros increased rapidly. The lifetime cumulative incidence rates (age 0-75) for Chamorros was highest in Guam (11.4%) and close to the US rate (12%). Interestingly, there was a marked difference in lifetime cumulative incidence rates between Filipino women in Guam and Filipinos in the US (7% and 10.9% respectively). Similar results were found for the 2005-2009 time period.

Conclusions

The observed disparity among ethnic groups suggests ethnic differences in exposure or response to certain risk factors. Further study is needed to examine possible differences in risk behaviors between Filipinos and Chamorros in Guam, as well as between Filipinos in Guam and Filipinos in the US.

Citation Format: Grazyna Badowski, Rachael Leon Guerrero, Brayan Simsiman, Rachel Novotny, Lynne Wilkens. Cumulative incidence rates of breast cancer for Filipinos and Chamorros in Guam and the United States. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3702. doi:10.1158/1538-7445.AM2015-3702
Abstract 3728: Rapid deployment of statewide tobacco cessation program for cancer patients
Authors: Petersen Laura , Baca Hilary , Brown Karen , Lindsay Jamie , Mott Sarah , Severson Jane , Schulze Anna , Warren Graham
Published: August 2015

Purpose

Tobacco use is not only associated with the development of cancer, but after a patient develops cancer, its continued use is associated with poorer cancer treatment outcomes. Despite this fact, most oncologists do not regularly provide tobacco cessation support. The purpose of this study was to implement an expedited process of patient identification and tobacco cessation for cancer patients.

Methods

The Michigan Oncology Quality Consortium (MOQC) collaborated with the Michigan Department of Community Health, Blue Cross and Blue Shield of Michigan, and the Michigan Cancer Consortium to enhance tobacco cessation support for cancer patients. A statewide tobacco cessation demonstration project using a lean engineering approach was developed to provide structured tobacco assessments, referral to state quitline resources, and provide tailored cessation support to cancer patients consisting of behavioral counseling and free pharmacotherapy where appropriate. These steps were rapidly deployed in over 30 locations including those in private oncology practices and cancer centers in the first year. Data were analyzed to evaluate participation, retention, and quit rates for cancer patients enrolled in the program.

Results

Previous reports have described high rates of patient contact and participation in the cessation program. In the 690 patients with at least 12 months since program enrollment, 200 (29%) participated in only 1 call, 313 (45%) participated in 1-2 follow-up calls, and 177 (26%) participated in 3 or more follow-up calls. Follow-up 30 day quit rate data was obtained in 198, 138, and 199 patients at 3, 6, and 12 months respectively. Self-reported quit rates remained largely consistent with 29.8% at 3 months, 27.5% at 6 months, and 30.2% at 12 months.

Conclusions

The MOQC demonstration project represents a model partnership between payors, clinicians, and statewide quitlines that maintained consistent and moderate quit rates in cancer patients. Large-scale deployment of this model program may provide a clinically efficient and effective mechanism to enhance widespread tobacco cessation efforts for cancer patients.

Citation Format: Laura Petersen, Hilary Baca, Karen Brown, Jamie Lindsay, Sarah Mott, Jane Severson, Anna Schulze, Graham Warren. Rapid deployment of statewide tobacco cessation program for cancer patients. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3728. doi:10.1158/1538-7445.AM2015-3728