bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5042: Defining an optimal single time point sampling strategy representative of overall capecitabine pharmacokinetics
Authors: Welch Stephen , Teft Wendy , Lenehan John , Tirona Rommel , Lumsden Karen , Winquist Eric , Kim Richard B.
Published: July 2017

Background: Capecitabine is an oral chemotherapy pro-drug used to treat advanced colorectal cancer. Patients may experience hand-foot syndrome and diarrhea, among other side effects, that affect quality of life and may necessitate dose modification or discontinuation. There is significant regional variation in capecitabine tolerability, related to a myriad of factors including pharmacogenomics, dietary and cultural differences. Capecitabine dose modification, when necessary, is empirical based on toxicity suggesting a personalized dosing approach might better optimize therapy. Objective: In phase I of a personalized dosing approach, our objective was to define an optimal time point for blood sampling that best represented overall exposure of capecitabine and its metabolites.

Methods: A single-arm prospective pharmacokinetic cohort study of patients with advanced or metastatic colorectal cancer prescribed capecitabine monotherapy was done. Blood samples were collected pre-dose and at timed intervals between 0 and 8 hours post-dose. Plasma concentration of capecitabine and its major metabolites, 5'-deoxy-5-fluorocytidine (5'-DFCR) and 5'-deoxy-5-fluorouracil (5'-DFUR), were measured by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS).

Results: 26 patients were enrolled; 65% were male and 42.3% had metastatic disease. Mean capecitabine dose was 2854 ± 944 mg. Hand-foot symptoms (60%), fatigue (53%) and diarrhea (30%) were the most common adverse drug reactions. Dose normalized mean (SD) AUC0-8h for capecitabine, 5’-DFCR and 5’-DFUR were 6.74 (3.0), 4.19 (1.5) and 6.33 (2.8) ng/ml*h, respectively. Spearman correlation between dose normalized concentrations and AUC at each blood draw was performed. The best estimated time points for capecitabine, 5’-DFCR and 5’-DFUR were 1.5, 2 and 2 hours with r2 values of 0.6 (p <0.01), 0.64 (p <0.001) and 0.51 (p <0.01), respectively. There was a significant correlation seen between capecitabine AUC and need for subsequent dose reduction (p<0.05).

Conclusions: Blood samples obtained between 1.5 and 2 hours post-dose provide the best estimate of capecitabine exposure. Further pharmacokinetic analysis in this cohort is ongoing. This blood draw strategy will be used in a larger trial intended to develop a personalized capecitabine dosing algorithm.

Citation Format: Stephen Welch, Wendy Teft, John Lenehan, Rommel Tirona, Karen Lumsden, Eric Winquist, Richard B. Kim. Defining an optimal single time point sampling strategy representative of overall capecitabine pharmacokinetics [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 5042. doi:10.1158/1538-7445.AM2017-5042
Abstract 5081: Disruption of DNA methyltransferase (DNMT) 1 confers resistance to DNMT inhibitors in human colorectal cancer cells
Authors: Laranjeira Angelo B. , Nguyen Dat , Huang Erich , Doroshow James H. , Yang Sherry X.
Published: July 2017

DNA methyltransferase (DNMT) 1 is responsible for the maintenance and propagation of DNA methylation pattern, and plays an important role in cell survival and proliferation. Thus, targeting DNMT represents a promising approach for cancer treatment. However, the role of DNMT1 in treatment efficacy with DNMT inhibitors remains controversial. The aim of this study was to investigate potential alterations of DNMT inhibitor-induced cytotoxicity and cellular survival after disruption/knockout of DNMT1 in cancer cells. The human colorectal cancer cell line HCT116 and its isogenic DNMT1 knockout pair were treated with increasing concentrations of 4’-thio-5-aza-2’-deoxycytidine (aza-TdCyd), a novel DNMT inhibitor, and 5-aza-2’-deoxycytidine (aza-dCyd). Cytotoxic effects and cell survival were evaluated by MTT and clonogenic assays. Apoptosis was measured by annexin-V/propidium iodide test, and expression of DNMT1, DNMT3A and DNMT3B was detected by western blot. After 96h of drug exposure, IC50s of aza-TdCyd and aza-dCyd were 0.031µM and 0.28 µM, respectively, in HCT116 cells. In contrast, DNMT1 knockout cells were remarkably resistant to both drugs with IC50s all higher than 10 µM. The treatments resulted in a significant increase in apoptosis in the parental line versus its knockout counterpart. As for the cell survival, IC50s were ~0.12 µM for aza-TdCyd and ~4.62 µM for aza-dCyd in HCT116 cells, compared to ~43.77 µM and ~62.09 µM in DNMT1 knockout cells. As such, disruption of DNMT1 caused over 300-fold and 10-fold increase in resistance to aza-TdCyd and aza-dCyd. Moreover, we observed a G2/M arrest produced by aza-TdCyd versus aza-dCyd in the parental cells, and no apparent alteration in cell cycle phase distribution in DNMT1 knockout cells. Notably, aza-TdCyd was more potent than aza-dCyd in depleting DNMT3A and/or DNMT1 in both cell lines after 48h of treatment. Our findings demonstrate that disruption of DNMT1 leads to a significantly increased resistance to aza-TdCyd and aza-dCyd, suggesting that DNMT1 is critical to the antitumor activity of DNMT inhibitors. Aza-TdCyd is much more potent than aza-dCyd for both inhibition of DNMTs and growth of cancer cells.

Citation Format: Angelo B. Laranjeira, Dat Nguyen, Erich Huang, James H. Doroshow, Sherry X. Yang. Disruption of DNA methyltransferase (DNMT) 1 confers resistance to DNMT inhibitors in human colorectal cancer cells [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 5081. doi:10.1158/1538-7445.AM2017-5081
Abstract 5088: Tumor penetrating RNA delivery for therapeutic benefit of pancreatic cancer
Authors: Hao Liangliang , Lo Justin , Bhatia Sangeeta
Published: July 2017

Pancreatic cancer remains a deadly disease with a median survival of less than one year, in part because we lack effective therapeutic strategies. Mutations of the oncogene KRAS and the tumor suppressor genes TP53, INK4A, and DPC4 play important roles in the pathogenesis of pancreatic ductal adenocarcinoma (PDAC); however, these driver events have proven to be challenging targets for drug development. RNA interference through small interfering RNA (siRNA) delivery has the potential to have an impact on silencing ‘undruggable’ targets; however, siRNA delivery in tumors has been challenging in general and worsened in PDAC by the stromal barriers to drug delivery. Herein, we integrated biomedical engineering, materials science, and pre-clinical investigation to establish a novel class of tumor-penetrating nanoparticles (TPNs) to enhance delivery of siRNAs in PDAC for therapeutic benefit. Specifically, we adapted self-assembled nanoparticles comprised of siRNA complexed with tandem tumor-penetrating and membrane-translocating peptides, which enabled the specific delivery of siRNA deep into the tumor parenchyma. Specifically, to establish efficacious siRNA delivery vehicles, we characterized and selected tandem peptides that incorporate novel tumor-penetrating peptide domains known to target PDAC in mice. In parallel, we optimized the particle stability through incorporation of hydrophilic molecules, focusing on achieving desirable pharmacokinetic properties which will reduce off-target delivery and increase potency of a given siRNA dose. The resulting TPNs carry siRNA into the cytosol of monolayer cultures of tumor cells, and also exhibit penetration in pancreas organoids derived from primary mouse and human tumors. As a proof-of-concept, we used TPN in vivo to knockdown KRAS in a murine PDAC model, using subcutaneously-implanted, autochthonous KRASG12D/+p53-/- tumors. Treatment of tumor-bearing mice with systemically administrated KRAS-specific TPNs suppressed the growth of established tumors. With the delivery platform established, the ongoing goals of this project are to identify siRNA targets that synergize with the suppression of oncogenic KRAS, and to validate synthetic lethal targets by interfering with signature PDAC driver events uncovered by functional genomic efforts. Together, these studies will provide a foundation for translational studies involving new targets and delivery technology in pancreatic cancer.

Citation Format: Liangliang Hao, Justin Lo, Sangeeta Bhatia. Tumor penetrating RNA delivery for therapeutic benefit of pancreatic cancer [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 5088. doi:10.1158/1538-7445.AM2017-5088
Abstract 5093: In vitro, in vivo and molecular effects of stromal selective targeting by uPAR retargeted oncoytic measles virus on breast cancer progression
Authors: Jing Yuqi , Chavez Valery , Acquavella Nicolas , El-Ashry Doraya , Ban Yuguang , Chen Xi , Merchan Jaime
Published: July 2017

The tumor microenvironment is a relevant target for novel biological therapies. While it has been demonstrated that tumor-stromal cell interactions are important in the sensitivity of the cancer cells to oncolytic virues, few studies have investigated the direct effects of an oncolytic virus on the stroma, and its implications on the virus antitumor efficacy.

MV-m-uPA and MV-h-uPA are fully retargeted, species specific, oncolytic measles viruses directed against murine or human urokinase receptor (uPAR). The effects of stromal selective targeting by uPAR retargeted MVs were investigated. In vitro infection, gene expression and cytotoxicity by MV-h-uPA and MV-m-uPA were demonstrated in human and murine cancer cells and cancer associated fibroblasts in a species specific manner. In murine fibroblast (3T3)/human breast cancer (T47D) 3-D co-cultures, selective fibroblast targeting by MV-m-uPA inhibited breast cancer cell growth.  These effects were validated in vivo, in a human breast cancer xenograft (MDA-MB-231), where intravenous administration of the murine specific MV-m-uPA led to significant tumor growth delay and improved survival compared to controls. Experiments comparing the effects of tumor (MV-h-uPA) vs. stromal (MV-m-uPA) vs. combined treatment showed that tumor and stromal targeting was associated with improved tumor control. Correlative studies demonstrated in vivo tumor targeting, increased apoptosis, and MV-m-uPA induced differential regulation of both stromal (murine) genes and cancer (human) genes associated with inflammation, angiogenesis and survival, among others, indicating viral induced modulation of tumor-stromal interactions.

Our results demonstrate for the first time the feasibility of stromal selective targeting by an oncolytic MV, viral induced modulation of the tumor microenvironment, and subsequent tumor growth delay. These findings further validate the critical role of stromal uPAR in cancer progression and the potential of oncolytic viruses as anti-stromal agents.

Citation Format: Yuqi Jing, Valery Chavez, Nicolas Acquavella, Doraya El-Ashry, Yuguang Ban, Xi Chen, Jaime Merchan. In vitro, in vivo and molecular effects of stromal selective targeting by uPAR retargeted oncoytic measles virus on breast cancer progression [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 5093. doi:10.1158/1538-7445.AM2017-5093
Abstract 5100: AB61, a new potent nucleoside cytostatic: Molecular mechanisms of action and preclinical activity
Authors: Dzubak Petr , Hajduch Marian , Perlikova Pavla , Rylová Gabriela , Naus Petr , Elbert Tomas , Tloustova Eva , Bourderioux Aurelie , Slavetinska Lenka , Motyka Kamil , Dolezal Dalibor , Znojek Pawel , Nova Alice , Harvanova Monika , Siler Michal , Hlavac Jan , Hocek Michal
Published: July 2017

7-(2-Thienyl)-7-deazaadenosine (AB61) showed nanomolar cytotoxic activities against various cancer cell lines but only mild (micromolar) activities against normal fibroblasts. The selectivity of AB61 was found to be due to inefficient phosphorylation of AB61 in normal fibroblasts. The phosphorylation of AB61 in the leukemic CCRF-CEM cell line proceeds well and it was shown that AB61 is incorporated into both DNA and RNA, preferentially as a ribonucleotide. It was further confirmed that a triphosphate of AB61 is a substrate for both RNA and DNA polymerases in enzymatic assays. Gene expression analysis suggests that AB61 affects DNA damage pathways and protein translation/folding machinery. Indeed, the formation of large 53BP1 foci was observed in nuclei of AB61-treated U2OS-GFP-53BP1 cells indicating DNA damage. Random incorporation of AB61 into RNA blocked its translation in an in vitro assay and reduction of reporter protein expression was also observed in mice after 4-hour treatment with AB61. AB61 also significantly reduced tumor volume in mice bearing SK- OV-3, BT-549, HT-29 and MDA-MB231 xenografts. The results indicate that AB61 is a promising compound with the unique mechanism of action and deserves further development as an anticancer agent. This work was supported by the Ministry of Education of the Czech Republic (LO1304).

Citation Format: Petr Dzubak, Marian Hajduch, Pavla Perlikova, Gabriela Rylová, Petr Naus, Tomas Elbert, Eva Tloustova, Aurelie Bourderioux, Lenka Slavetinska, Kamil Motyka, Dalibor Dolezal, Pawel Znojek, Alice Nova, Monika Harvanova, Michal Siler, Jan Hlavac, Michal Hocek. AB61, a new potent nucleoside cytostatic: Molecular mechanisms of action and preclinical activity [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 5100. doi:10.1158/1538-7445.AM2017-5100
Abstract 5106: In vitro and in vivo evaluation of WAC-224, a novel quinolone class of topoisomerase II inhibitor for cancer therapy
Authors: Ueshima Taichi , Yamaguchi Tomonori , Itoh Kenji , Kashimoto Naoki , Hirano Tatsuya , Shimabara Rumiko , Kawakubo Yohei , Sato Masayuki , Yamashita Junpei , Yazaki Akira , Tamura Koichi
Published: July 2017

Introduction: DNA topoisomerases (Topo) are classical but still attractive targets for drug therapy in multiple types of cancers. Topo inhibitors, such as Etoposide and Daunorubicin, have been effectively used; however, their clinical use is often limited by drug resistance in cancer cell population. Therefore, the development of a novel chemical class of Topo inhibitors has been desired to overcome drug resistance. Recently, Vosaroxin (QINPREZO) is identified as a first-in-class anti-cancer quinolone derivative targeting Topo II, which is under development for relapsed or refractory acute myeloid leukemia (AML). In this study, we evaluated the in vitro and in vivo activities of WAC-224, a novel quinolone derivative for Topo II inhibition, for various cancer cell lines including multi-drug resistant cells and in mouse xenograft models, respectively.

Materials and Methods: In vitro anti-proliferative activities against over 20 cell lines were determined using WST cell proliferation reagent. Apoptosis was measured by Annexin-V staining. In vivo anti-tumor activity was determined in immunodeficient mice bearing multi-drug resistant human uterine sarcoma MES-SA/DX5.

Results: WAC-224 accomplished extensive anti-proliferative activities for cancer cell lines including multi-drug resistant ones with the EC50 range of 0.001 to 1 microM. Especially, in MV4-11 AML cell line, WAC-224 showed a potent and superior anti-proliferative activity compared with Doxorubicin (EC50: 1.4 nM for WAC-224 v.s. 2.5 nM for Doxorubicin). Molecular and cellular mechanisms of WAC-224 were defined as inhibition of human Topo II, induction of G2/M phase cell cycle arrest and apoptosis. Additionally, significant antitumor effects of WAC-224 were confirmed in mouse xenograft models. Furthermore, WAC-224 showed no toxic effect on small intestinal crypts in mice.

Conclusion: This study demonstrated that WAC-224 has strong in vitro activities against broad cancer cell types along with the potent in vivo efficacy and well tolerability, without gastrointestinal toxicity. These results indicate that WAC-224 is promised to provide a new therapeutic option for various cancers including multi-drug resistant and hematological ones.

Citation Format: Taichi Ueshima, Tomonori Yamaguchi, Kenji Itoh, Naoki Kashimoto, Tatsuya Hirano, Rumiko Shimabara, Yohei Kawakubo, Masayuki Sato, Junpei Yamashita, Akira Yazaki, Koichi Tamura. In vitro and in vivo evaluation of WAC-224, a novel quinolone class of topoisomerase II inhibitor for cancer therapy [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 5106. doi:10.1158/1538-7445.AM2017-5106
Abstract 5111: Norcantharidin impairs tumor growth in vivo and inhibits stemness of triple-negative breast cancer cells
Authors: Berardi Damian E. , Cicuttin Guido , Taruselli Maria A. , Cirigliano Stefano M. , Joffé Elisa D. Bal de Kier , Urtreger Alejandro J. , Todaro Laura B.
Published: July 2017

Triple-negative breast cancer (TNBC) is characterized by an abundance of treatment-resistant cancer stem cells (CSC). The absence of a molecular target, coupled with its highly aggressiveness, leads to the lack of an effective therapy for TNBC. Norcantharidin (NCTD) is a synthetic demethylated small-molecule analog of the naturally occurring cantharidin isolated from blister beetles (Mylabris phalerata Pall). Unlike the conventional chemotherapeutics, NCTD toxicity is higher to cancer cells than normal ones, making this small molecule promising for cancer treatment.

The aims of this work were: A) To study the effect of NCTD on 4T1 cell line proliferation in vitro. B) To analyze the effect of NCTD on 4T1 derived CSC on self-renewal and clonogenic capacity. C) To evaluate the effect of NCTD on 4T1 tumor growth in vivo.

We employed the well-known 4T1 triple-negative breast cancer cell model, which presents a huge proportion of CSC.

We observed that NCTD treatment during 96 h significantly reduced 4T1 cell proliferation in vitro. In addition, the IC50 value of NCTD was 27.35 ± 2.83 μM.

Related to CSC, NCTD pre-treatment for 96 h impaired CSC self-renewal (Number of secondary mammospheres: Control: 276±39; NCTD: 163±18; p≤0.05) as well as the clonogenic capacity (Number of colonies: Control: 359±38; NCTD: 122±11; p≤0.05). By q-PCR, we observed that NCTD treatment for 48 h significantly induced an increase of Gli-1 and Smooth in CSC, keys member of Sonic Hedgehog pathway.

Finally, we performed an in vivo assay, where 4T1 cells were orthotopically inoculated on mammary gland of BALB/c mice, and NCTD was i.p. inoculated twice a week (5mg/kg). We observed that NCTD treatment significantly reduced tumor growth in vivo.

Our data suggest that NCTD treatment reduces tumor growth both in vitro and in vivo, possibly through the direct effect on CSC self-renewal and clonogenic capacity, by modulating Sonic Hedgehog pathway.

Citation Format: Damian E. Berardi, Guido Cicuttin, Maria A. Taruselli, Stefano M. Cirigliano, Elisa D. Bal de Kier Joffé, Alejandro J. Urtreger, Laura B. Todaro. Norcantharidin impairs tumor growth in vivo and inhibits stemness of triple-negative breast cancer cells [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 5111. doi:10.1158/1538-7445.AM2017-5111
Abstract 5119: Evaluation of Minnelide as potential targeted therapy for triple negative breast cancer
Authors: Singh Mahendra K. , Shah Soham , Sharma Nikita Satish , Giri Bhuwan , Banerjee Sulagna , Saluja Ashok
Published: July 2017

Recent advances in diagnostics and better understanding of molecular mechanism underlying breast cancer has let to the better therapeutic options and disease outcome for majority of breast cancer patients. However, ~10 - 20% of all breast cancers often referred to as “triple negative” as they lack expression of the estrogen (ER), progesterone (PR), and human epidermal growth factor 2 (HER2) receptors convey a poor prognosis due in part to a lack of targeted therapies. The aim of the current study is to evaluate whether triptolide and its water soluble analog Minnelide is effective against triple negative breast cancer cells. We have previously shown that triptolide/Minnelide not only reduces tumor growth in various cancer models but it also regulates epithelial -mesenchymal transition (EMT), an important mechanism underlying metastasis. In our preliminary findings using three triple negative breast cancer (TNBC) cell lines, MDA-MB-231, MDA-MB-468, and MDA-MB-157, we demonstrate that triptolide not only inhibits the proliferation of TNBC cells but also regulates the protein levels of EMT markers including β-Catenin and Vimentin. In order to elucidate the mechanism underlying triptolide mediated inhibition of cellular proliferation and regulation of EMT markers in TNBC cells, we identified Src kinase and Aurora kinase A as two new targets for triptolide action in TNBC cells. By targeting Src and Aurora kinase, triptolide disrupts the integrity of focal adhesion structures and reduces cell spreading via regulating FAK activity. Our preliminary findings regarding potential use of triptolide/Minnelide in TNBC based on in vitro experiments are promising. However, considering the complex pathophysiology of breast cancer and other biological factors playing role in a disease setting, in-vivo experiments to test the efficacy of Minnelide in relevant mouse models for mammary cancers are currently underway.

Citation Format: Mahendra K. Singh, Soham Shah, Nikita Satish Sharma, Bhuwan Giri, Sulagna Banerjee, Ashok Saluja. Evaluation of Minnelide as potential targeted therapy for triple negative breast cancer [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 5119. doi:10.1158/1538-7445.AM2017-5119
Abstract 5128: Induction of immunogenic cell death and tumor regression in murine animal models by a novel cytolytic compound, LTX-401
Authors: Mauseth Brynjar , Eike Liv-Marie , Shi Ji-Hua , Camilio Ketil , Zhou Heng , Sauvat Allan , Gomes-da-Silva Lígia C , Durand Sylvèr , Forveille Sabrina , Iribarren Kristina , Yamazaki Takahiro , Souquere Sylvie , Bezu Lucillia , Müller Kevin , Leduc Marion , Liu Peng , Zhao Liwei , Marabelle Aurélien , Zitvogel Laurence , Kepp Oliver , Kroemer Guido , Rekdal Øystein , Line Pål-Dag , Sveinbjørnsson Baldur
Published: July 2017

LTX-401 is a de novo designed cytolytic compound that shares many chemical features with anticancer peptides, such as amphipathicity, hydrophobicity and overall net charge. In vitro cytotoxicity studies revealed that LTX-401 was highly active against a panel of malignant cells including murine and human cancer cell lines while displaying selectivity towards human red blood cells. Furthermore, LTX-401 was found to induce immunogenic cell death as demonstrated by the release of Damage-Associated Molecular Pattern molecules, or ‘danger signals’, such as High-Mobility Group Box-1 protein, ATP and cytochrome c. Flow cytometric and confocal microscopy studies also demonstrated reduced signal from lysosomal dye Lysotracker-DND-26, hence indicating loss of lysosomal integrity upon induction of cell death. The latter was supported using transmission electron microscopy, showing distinct morphological characteristics of necrosis. Moreover, at low concentrations, LTX-401 selectively enriched in the Golgi apparatus and initiated a lethal signaling event that in part could be inhibited by Brefeldin A. Complete tumor regression has been obtained in several rodent models, including the B16 mouse melanoma model and the JM1 rat hepatocellular carcinoma model by intratumoral injection with LTX-401. Additionally, previously cured animals showed protection against rechallenge with live tumor cells, indicating the induction of tumor-specific immune memory. The increased survival benefit of LTX-401-treated animals provides a rationale for further evaluation of the compound as an immunotherapeutic agent against solid malignancies.

Citation Format: Brynjar Mauseth, Liv-Marie Eike, Ji-Hua Shi, Ketil Camilio, Heng Zhou, Allan Sauvat, Lígia C Gomes-da-Silva, Sylvèr Durand, Sabrina Forveille, Kristina Iribarren, Takahiro Yamazaki, Sylvie Souquere, Lucillia Bezu, Kevin Müller, Marion Leduc, Peng Liu, Liwei Zhao, Aurélien Marabelle, Laurence Zitvogel, Oliver Kepp, Guido Kroemer, Øystein Rekdal, Pål-Dag Line, Baldur Sveinbjørnsson. Induction of immunogenic cell death and tumor regression in murine animal models by a novel cytolytic compound, LTX-401 [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 5128. doi:10.1158/1538-7445.AM2017-5128
Abstract 5161: Inhibition of TBK1/IKBKE impacts tumor growth of KRAS-mutant colorectal cancers
Authors: Stinson Susanna , He Jianhua , Jia Zhiheng , Walker Katie S. , Cui Zhi-Hua , Kaplan Joshua A. , Venkataramani Chandrasekar , Wang Yujin , Guevara Fatima , Nelson Timothy , Sicinska Ewa , Ng Kimmie , Fuchs Charles S. , Dornan David
Published: July 2017

Mutations in RAS occur in an estimated one third of all human cancers.  While direct targeting of RAS remains challenging, targeting downstream RAS effector molecules has been shown to impact the oncogenic function of RAS and elicit anti-tumor activity.  TANK Binding Kinase 1 (TBK1) and I-Kappa B Kinase Epsilon (IKBKE) serine/threonine kinases are non-canonical IKK family members. TBK1 is directly downstream of the RAS effector Ral Guanine Nucleotide Dissociation Stimulator (RALGDS) and may be required for RAS to elicit its full oncogenic potential. 

To explore this possibility, we identified a potent and selective TBK1/IKBKE inhibitor to determine whether inhibition of TBK1 impacts the growth of RAS-mutant cancer models. KRAS-mutant cancer cell lines showed preferential sensitivity to TBK1/IKBKE inhibition over KRAS wild-type cell lines.  The TBK1/IKBKE inhibitor was then evaluated in vivo with two KRAS-mutant colorectal cancer patient-derived xenograft (PDX) models, one of which harbors co-occurring PIK3CA and KRAS mutations. The TBK1/IKBKE inhibitor caused moderate tumor growth inhibition in comparison to vehicle in both PDX models.  Since RAS exerts its oncogenic effects via additional effectors, such as RAF, and that inhibition of MEK1/2 resulted in compensatory activation of TBK1 in a KRAS-mutant cancer cell line, we tested if inhibition of TBK1/IKBKE in combination with the MEK inhibitor, trametinib, would result in superior efficacy in PDX models.  The combination of a TBK1/IKBKE inhibitor and trametinib resulted in greater than 90% tumor growth inhibition within both of these KRAS mutant PDX models compared to either single agent alone.  We also tested TBK1/IKBKE inhibition in a KRAS wild-type colorectal PDX model containing a BRAF mutation. In this model, TBK1/IKBKE did not inhibit tumor growth and, although there was significant tumor growth inhibition with trametinib alone, there was no further enhancement of efficacy with the addition of a TBK1/IKBKE inhibitor.

Together, these data suggest that a TBK1/IKBKE inhibitor in combination with a MEK inhibitor may impact KRAS-mutant cancers.

Citation Format: Susanna Stinson, Jianhua He, Zhiheng Jia, Katie S. Walker, Zhi-Hua Cui, Joshua A. Kaplan, Chandrasekar Venkataramani, Yujin Wang, Fatima Guevara, Timothy Nelson, Ewa Sicinska, Kimmie Ng, Charles S. Fuchs, David Dornan. Inhibition of TBK1/IKBKE impacts tumor growth of KRAS-mutant colorectal cancers [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 5161. doi:10.1158/1538-7445.AM2017-5161
Abstract 5213: Aberrant tumor metabolism enables GR takeover in enzalutamide-resistant prostate cancer
Authors: Li Jianneng
Published: July 2017

It is well known that castration resistant prostate cancer (CRPC) is still driven by the androgen axis. Therefore, the advent of new AR-targeting drugs, such as Enzalutamide (ENZ), has been beneficial for many patients with CRPC. ENZ is a potent FDA-approved AR antagonist which prolongs survival for patients with CRPC, even after docetaxel chemotherapy. Unfortunately, although ENZ has proven effective clinically, treatment resistance eventually arises in nearly all patients with CRPC. Emerging data suggest that the glucocorticoid receptor (GR) is upregulated in this context, stimulating expression of AR-target genes that permit continued growth despite AR blockade. Hence, we hypothesize that similar to metabolic mechanisms that elicit dihydrotestosterone (DHT) synthesis, which in turn stimulate AR in CRPC, a role for GR in ENZ resistance would be accompanied by a tumor metabolic switch that provides sustained tissue cortisol concentrations which enable GR activation. Here, we found that long-term ENZ treatment sustains tumor cortisol concentrations in models of prostate cancer and patient tissues by using [3H]-cortisol and high performance liquid chromatography. Further investigations with immunoblot, co-immunoprecipitation as well as in vitro ubiquitination assays in ENZ resistant prostate cancer models demonstrated that autocrine motility factor receptor (AMFR), an ubiquitin E3-ligase, and one of its adaptors, Erlin-2, mediate loss of 11β-hydroxysteroid dehydrogenase-2 (11β-HSD2), which sustains tumor cortisol concentrations, resulting in the continuous stimulation of GR and ENZ resistance. Remarkably, reinstatement of 11β-HSD2 expression, or AMFR loss, reverses ENZ resistance in mouse xenograft tumors.

Together, these findings reveal a surprising metabolic mechanism of ENZ resistance that may be targeted with a strategy that circumvents a requirement for systemic GR ablation.

Citation Format: Jianneng Li. Aberrant tumor metabolism enables GR takeover in enzalutamide-resistant prostate cancer [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 5213. doi:10.1158/1538-7445.AM2017-5213
Abstract 524: Characterizing the in vitro and in vivo effects of the PIM kinase inhibitor HS140 in triple-negative human breast cancer
Authors: Cobb Michael , Hunter Lucas , Carlson David , Darr David , Haystead Timothy , Baines Antonio T.
Published: July 2017

Triple-negative breast cancer (TNBC) lacks expression of the estrogen receptor (ER), the progesterone receptor (PR) and the ERBB2 (also known as HER2) receptor, making it the worse subtype of all breast cancers. Currently, there are no molecular targeted therapies for this cancer and chemotherapy is only successful in a limited number of patients. Recent studies suggest that members of the oncogenic PIM kinase family, especially PIM-1, play a significant role in the growth of TNBC. However, there is very limited information on the role other PIM family members have in the growth and development of TNBCs. As an attempt to address this concern, we treated a panel of TNBC cell lines with HS140, a PIM-2 kinase inhibitor developed in the laboratory. Using cytotoxicity assays, we were able to demonstrate a decrease in anchorage-dependent growth of cell lines at different concentrations of HS140. Also, a triple negative breast cancer GEMM (C3TAg) was used for an efficacy study. Mice (FVB/N background) have C(3)SV40 T-antigen resulting in inactivation of p53 and Rb. Nine mice received HS140 treatment (80mg/kg BIW IP) and 14 left untreated for control (non-treatment [NT]). All mice were monitored for weight loss and timespan to tumor development 5 times weekly. Tumor volumes at 21 days were significantly (Mann-Whitney, p=0.0002) reduced in the treated cohort with a mean of 109mm3 (range of 0-5000) compared to untreated 1393mm3 (0-239). We observed no toxicities with body mass stable at the treatment dose of 80mg/kg. Additionally, three mice from untreated and three mice from treated cohorts were chosen for blood sampling pre- and post-treatment via a submandibular bleed; no significant differences were noted in WBC, RBC, PLT, or HGB between untreated and treated cohorts. Overall, these results suggest that other members of the PIM kinase family, including PIM-2, have an important function in the growth and development of TNBC and may serve as a potential molecular target for future therapeutics.

Citation Format: Michael Cobb, Lucas Hunter, David Carlson, David Darr, Timothy Haystead, Antonio T. Baines. Characterizing the in vitro and in vivo effects of the PIM kinase inhibitor HS140 in triple-negative human breast cancer [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 524. doi:10.1158/1538-7445.AM2017-524
Abstract 5261: Tocopherols inhibit the estrogen-stimulated expansion of cancer stem cells via down-regulation of OCT4 and NFκB
Authors: Bak Min Ji
Published: July 2017

Cancer stem cells have been suggested to play a role in tumor initiation, maintenance, resistance, and metastasis. Breast cancer stem cells are defined as CD44+/CD24- cell populations that have the properties of tumor-initiating ability, self-renewal and differentiation. Although the importance of estrogen in breast cancer is well established, the effects of estrogen on breast cancer stem cells are not fully understood. Vitamin E, which is consisted of different forms of tocopherols and tocotrienols, has anti-tumor effects in various cancers, including breast cancer. In the present study, we examined the effects of α-, γ-, and δ-tocopherol against estrogen-stimulated expansion of cancer stem cells. To determine the effects of tocopherols on breast cancer stem cells, the MCF-7 tumorsphere cell culture system, which enriches for mammary progenitor cells and putative breast cancer stem cells, was utilized. Treatment of MCF-7 tumorsphere with estrogen for four days resulted in an increase in CD44+/CD24- stem cell populations in tumorspheres as well as the number and size of tumorspheres. Tocopherols at 1 and 10 μM inhibited formation of estrogen-induced stem cell enriched tumorspheres, indicating that tocopherols reversed estrogen-induced expansion of breast cancer stem cells. Tocopherols decreased the expression of estrogen-related genes such as TFF/pS2, cathepsin D, progesterone receptor and SerpinA1 in estrogen-stimulated tumorspheres. In addition, the mRNA levels of stem cell markers, including OCT4, NFκB, SOX-2 and CD44, were upregulated by estrogen, which were downregulated by the treatment with tocopherols. Our results suggest that tocopherols repress the expansion of breast cancer stem cell population by regulating OCT4 and NFκB signaling. Tocopherols could be effective natural agents for the prevention and treatment of estrogen-induced breast cancer. (This work was supported in part by the National Institutes of Health grant R01 AT007036.)

Citation Format: Min Ji Bak. Tocopherols inhibit the estrogen-stimulated expansion of cancer stem cells via down-regulation of OCT4 and NFκB [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 5261. doi:10.1158/1538-7445.AM2017-5261
Abstract 5270: Disease subtype independent biomarkers of breast cancer prevention by withaferin a
Authors: Hahm Eun-Ryeong , Samanta Suman K. , Sehrawat Anuradha , Kim Su-Hyeong , Pore Subrata K. , Singh Krishna B. , Christner Susan M. , Shuai Yongli , Beumer Jan H. , Roy Ruchi , Davidson Nancy E. , Singh Shivendra V.
Published: July 2017

Breast cancer is a rather complex and heterogeneous disease broadly grouped into four major subtypes, including luminal-type, basal-like, HER2 amplified, and normal-like, and each with a distinct molecular signature. A non-toxic chemopreventive intervention efficacious against different subtypes of breast cancer is still a clinically unmet need. The present study not only demonstrates chemoprevention of breast cancer in rats by the Ayurvedic medicine phytochemical withaferin A (WA) but also identifies its mechanistic biomarkers common to different subtypes of this disease. Chemopreventive efficacy of WA (4 and 8 mg per kg body weight) was determined using a rat model of breast cancer induced by N-methyl-N-nitrosourea (MNU). The mechanisms underlying breast cancer chemoprevention by WA were elucidated by western blotting, biochemical assays, immunohistochemistry, and cytokine profiling using plasma and tumors from the MNU-rat and/or mouse mammary tumor virus-neu (MMTV-neu) models. Inhibitory effect of WA on exit from mitosis and leptin-induced oncogenic signaling was determined using MCF-7 and MDA-MB-231 cells. Incidence, multiplicity, and burden of MNU-induced breast cancer in rats were decreased by WA administration. For example, the tumor weight in the 8 mg per kg group was lower by 67% compared with controls (P = 0.004). Mitotic arrest and apoptosis induction were common determinants of breast cancer chemoprevention by WA in the MNU-rat and MMTV-neu models. Cytokine profiling showed suppression of plasma leptin levels by WA in rats. WA inhibited leptin-induced oncogenic signaling in cultured MCF-7 and MDA-MB-231 cell lines. WA is a promising phytochemical with the ability to inhibit at least two different subtypes of breast cancer, including neu-driven estrogen receptor negative (ER-) breast cancer in MMTV-neu model and MNU-induced ER+ breast cancer in rats. This study was supported by the grant RO1 CA142604-07 awarded by the National Cancer Institute.

Citation Format: Eun-Ryeong Hahm, Suman K. Samanta, Anuradha Sehrawat, Su-Hyeong Kim, Subrata K. Pore, Krishna B. Singh, Susan M. Christner, Yongli Shuai, Jan H. Beumer, Ruchi Roy, Nancy E. Davidson, Shivendra V. Singh. Disease subtype independent biomarkers of breast cancer prevention by withaferin a [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 5270. doi:10.1158/1538-7445.AM2017-5270
Abstract 5323: Obesity and ovarian cancer survival revisited
Authors: Bandera Elisa V. , Lee Valerie , Qin Bo , Rodriguez Lorna , Powell Bethan , Kushi Lawrence H.
Published: July 2017

Research on the impact of obesity on ovarian cancer survival is inconsistent but previous studies did not consider the possible impact of ascites, bowel obstruction, or cachexia, which commonly occur in late-stage disease. We evaluated this association in a cohort study of primary invasive epithelial ovarian cancers diagnosed from 2000-2013 in Kaiser Permanente Northern California (KPNC) (n=1,157). Deaths were identified through December 2014, with median follow-up of 37 months. Proportional hazards regression was used to estimate overall and ovarian cancer-specific mortality, accounting for prognostic variables including age at diagnosis, race, stage, grade, histology, comorbidities, treatment, post-treatment CA125 levels, ascites, and bowel obstruction. There was no evidence of an association between BMI and overall or ovarian cancer-specific survival. However, we found strong effect modification by stage (Pinteraction<.01). Compared to normal pre-diagnosis BMI (18.5-24.9 kg/m2), for women who were obese before diagnosis (BMI≥35 kg/m2) risk of ovarian cancer-specific mortality was increased among those diagnosed at stages I/II (Hazard ratio [HR]: 3.40; 95% confidence interval [CI]: 1.16-9.99), but reduced among those diagnosed with stage IV disease (HR: 0.58; 95%CI: 0.35-0.96). Associations persisted after excluding those diagnosed with cachexia (n=91) and further adjusting for ascites and bowel obstruction, with no evidence of effect modification by these factors. Associations of obesity with ovarian cancer survival differ by stage, with decreased survival among those with localized disease and increased survival among those with late-stage disease. Stage-specific effects of obesity on survival suggest a tailored approach to improve prognosis may be appropriate.

Citation Format: Elisa V. Bandera, Valerie Lee, Bo Qin, Lorna Rodriguez, Bethan Powell, Lawrence H. Kushi. Obesity and ovarian cancer survival revisited [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 5323. doi:10.1158/1538-7445.AM2017-5323
Abstract 5340: Retrovirus insertion induces syncytin mediated exosomes delivery in prostate cancer cells
Authors: Uygur Berna , Chernomordik Leonid
Published: July 2017

Activation of retroviruses in normal cells can induce tumors and a wide array of cancers contains retrovirus-like particles. The potential role of these endogenous retroviruses in the development and progression of tumors has been discussed in many studies. However the effects of retrovirus insertion into cancer cells have not yet been fully examined. In this study, we explore genetic material transfer either between retrovirus-transduced prostate cancer PC3 cells or between PC3 cells and their host environment through extracellular vesicles (EV) and virus-like particles. For these purposes, we transduced PC3 cells with either GFP-encoding retrovirus or with m-cherry-encoding lentivirus. Co-incubation of GFP-expressing cells with m-cherry-expressing cells for 72h yielded yellow PC3 cells. Yellow cells were also observed in the experiments where lentiviral m-cherry expressing PC3 cells were cultured with tissue culture medium from retrovirus GFP expressing PC3 cells. These results indicated that horizontal genetic information transfer involves neither fusion nor contact between the cells. Since incubation of the retrovirus GFP-expressing PC3 cells with the medium from the lentivirus m-cherry expressing PC3 cells, did not yield yellow cells the genetic material transfer requires retrovirus insertion in PC3 cells. Using Western blot and Q-PCR, we found the exosomes from the medium generated by retrovirus GFP expressing PC3 cells to carry human endogenous retroviral proteins syncytin 1 and 2 and contain RNAs encoding these proteins. Exosomes from the lentivirus m-cherry expressing PC3 cells had lower protein expression of Syncytin 1&2 than exosomes from retrovirus GFP expressing PC3 cells. Application of Syncytin 1 or 2 antibodies to the medium generated by or co-cultured retrovirus GFP expressing PC3 and lentivirus m-cherry expressing PC3 cells suppressed horizontal transfer of the genetic information from retrovirus GFP expressing PC3 cells to other cells. The specific mechanisms by which retroviral transduction and syncytins control the generation of a GFP-encoding pseudotyped retrovirus or EV and the delivery of their content, as well as the role of the virus/EV-mediated horizontal genetic material transfer between prostate cancer cells and between prostate cancer and their microenvironment remain to be clarified.

Citation Format: Berna Uygur, Leonid Chernomordik. Retrovirus insertion induces syncytin mediated exosomes delivery in prostate cancer cells [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 5340. doi:10.1158/1538-7445.AM2017-5340
Abstract 5347: Molecular barcoding improves confidence in variant calling with low input FFPE tissue samples
Authors: Arezi Bahram , Hsue Bilan , Chen Keith , Rogers Brenda , Lecocq Christian , Ashutosh Ashutosh , Ghosh Jayati , Amparo Gilbert , Ramaiah Madhuvanthi , Pabon Carlos , Hogrefe Holly
Published: July 2017

FFPE tissue archiving is the most widely used method for clinical sample preservation, and provides a valuable source of diverse genetic information for cancer biomarker discovery. NGS library preparation is often challenging, however, as FFPE-derived DNAs exhibit varying degrees of fragmentation and chemical modification depending on how tissues were handled and processed. Moreover, the presence of low allele frequency variants because of tumor heterogeneity, sample degradation, and high rate of formalin-induced mutations make SNV calling a challenging task. To improve variant calling and consistency across FFPE samples of varying quality, we developed a complete workflow solution that features DNA pre-qualification, molecular barcodes for uniquely tagging each DNA fragment, pre-capture indexing for tracking up to 192 samples, accelerated hybrid capture, and one-tube library prep, to enable the preparation of complex enriched libraries in less than 8 hours from 10-200ng of FFPE-derived DNA. We incorporate molecular barcodes at the ligation step to enable de-duplication and generation of consensus reads using SureCall software, which significantly improves confidence of variant calling (99% of calls >150 variant call quality threshold) at low allelic frequencies.

Using ClearSeq Comprehensive Cancer Panel which targets 151 genes frequently mutated in solid and hematological cancers, we routinely obtain high quality libraries (with 1000X sequencing depth and after deduplication) with >99% of the targeted region covered at 20X for high quality gDNA (FF tissue), and greater than 95% and 90% covered at 20X for medium and low quality FFPE samples, respectively. Using multiplex reference samples, we demonstrate detection of low frequency variants (at 1-5%) including SNVs, small indels, and RET and ROS1 translocations. As we will further show, FFPE integrity scores determined by qPCR and TapeStation (ddCq and DIN) enable researchers to choose the appropriate quantification method, PCR cycling, and sequencing depth for optimal sequencing coverage.

Citation Format: Bahram Arezi, Bilan Hsue, Keith Chen, Brenda Rogers, Christian Lecocq, Ashutosh Ashutosh, Jayati Ghosh, Gilbert Amparo, Madhuvanthi Ramaiah, Carlos Pabon, Holly Hogrefe. Molecular barcoding improves confidence in variant calling with low input FFPE tissue samples [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 5347. doi:10.1158/1538-7445.AM2017-5347
Abstract 5367: Ngago: no evidence of targeted endonuclease activity
Authors: Topka Sabine , Kazzaz Sara , Offit Kenneth , Joseph Vijai
Published: July 2017

The development of a versatile and easy to use genome editing tool based on the bacterial CRISPR/Cas9 system has led to a vast increase in applications of genome editing in laboratories around the world. As an extremely powerful and versatile tool it has facilitated important discoveries in both basic and translational research as well as in a potentially wide range of therapeutic applications. Natronobacterium gregoryi Argonaute (NgAgo) protein was described as a favorable alternative to Cas9 due to its simplicity and higher endonuclease specificity. Thus far, there are no formal reports documenting the success of NgAgo in other laboratory settings and cellular systems. Utilizing the identical cellular background and protocol that was made publicly available by the authors of the initial study, we were unable to reproduce the endonuclease function of NgAgo. These results do not support the role of NgAgo in gene editing experiments.

Note: This abstract was not presented at the meeting.

Citation Format: Sabine Topka, Sara Kazzaz, Kenneth Offit, Vijai Joseph. Ngago: no evidence of targeted endonuclease activity [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 5367. doi:10.1158/1538-7445.AM2017-5367