bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2808: Simultaneous inhibition of SHP2 phosphatase at two allosteric sites
Authors: Fodor Michelle , Price Edmund , Wang Ping , Lu Hengyu , Argintaru Andreea , Chen Zhouliang , Glick Meir , Hao Huai-Xiang , Kato Mitsunori , Koenig Robert , LaRochelle Jonathan R. , Liu Gang , McNeill Eric , Majumdar Dyuti , Nishiguchi Gisele , Perez Lawrence , Paris Greg , Quinn Christopher , Ramsey Timothy , Sendzik Martin , Shultz Michael , Williams Sarah , Stams Travis , Blacklow Stephen C. , LaMarche Matthew J. , Acker Michael G.
Published: August 2018

SHP2 is a cytoplasmic non-receptor tyrosine phosphatase involved in the propagation of extracellular signaling through receptor tyrosine kinases. Aberrant SHP2 activity has been identified as a driver in multiple cancers and SHP2 has also been implicated in the PD-1/PD-L1-mediated exhaustion of effector T-cells, leading to immune system evasion of tumors. Recently, we reported the identification of SHP099, an allosteric inhibitor of SHP2 with in in vivo efficacy against multiple RTK-driven tumor xenograft models. Here we report the use of alternate screening paradigms to identify a novel allosteric inhibitor which binds to a previously uncharacterized pocket on SHP2. Like SHP099, the second allosteric inhibitor stabilizes a closed conformation of SHP2, which blocks access to the phosphatase active site. Structure based drug design led to improvements in potency, and combination studies in biochemical, biophysical and cellular assays confirm dual occupation of SHP099 and the second allosteric molecule, resulting in improved potency. This work highlights a rare opportunity for dual occupation of inhibitors for a single target and provides additional tools for the exploration of SHP2 biology.

Citation Format: Michelle Fodor, Edmund Price, Ping Wang, Hengyu Lu, Andreea Argintaru, Zhouliang Chen, Meir Glick, Huai-Xiang Hao, Mitsunori Kato, Robert Koenig, Jonathan R. LaRochelle, Gang Liu, Eric McNeill, Dyuti Majumdar, Gisele Nishiguchi, Lawrence Perez, Greg Paris, Christopher Quinn, Timothy Ramsey, Martin Sendzik, Michael Shultz, Sarah Williams, Travis Stams, Stephen C. Blacklow, Matthew J. LaMarche, Michael G. Acker. Simultaneous inhibition of SHP2 phosphatase at two allosteric sites [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2808.
Abstract 282: Inferring transcriptional regulatory programs in gynecological cancers
Authors: Osmanbeyoglu Hatice U. , Jelinic Petar , Douglas Douglas , Leslie Christina
Published: August 2018

Cancer cells acquire genetic and epigenetic alterations that often lead to dysregulation of oncogenic signal transduction pathways, which in turn alter downstream transcriptional programs. Transcription factors (TFs) are the main link between signaling pathways and the transcriptional regulatory programs. The Cancer Genome Atlas (TCGA) has studied several of the most common and aggressive gynecologic tumors including high-grade serous ovarian carcinomas (HGSOC), uterine carcinosarcoma (UCS), and the serous-like subset of endometrial cancer (UCEC), together with basal breast cancer, which shares many genomic features with serous ovarian tumors. TFs impacts on gene regulation have not been well characterized in gynecological and basal breast cancers. The majority of these tumors lack accurate predictors of response and resistance and share an unmet need for adequate treatment of recurrent disease. We developed a multitask learning framework for integrating regulatory sequence from ATAC-mapped promoters and enhancers from cell line models with RNA-seq data from patient tumors in order to infer transcription factor (TF) regulatory activities and explore similarities and differences between uterine, ovarian, and basal breast tumors. We showed that our multitask learning framework enables us to selectively share the information across tumors and strongly improves the accuracy of gene expression prediction models for gynecological and basal breast tumors. Our analysis identified histologic type specific and common TF regulators of gene expression as well as predicted distinct dysregulated transcriptional regulators downstream of somatic alterations in these different cancers. Moreover, many of the identified TF regulators were significantly associated with survival outcome within the histological subtype. Computationally dissecting the role of TFs in these cancers may ultimately lead to new therapeutics tailored to groups of subtypes or individuals.

Citation Format: Hatice U. Osmanbeyoglu, Petar Jelinic, Douglas Douglas, Christina Leslie. Inferring transcriptional regulatory programs in gynecological cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 282.
Abstract 2825: PIN1-SCFFBW7 proteolytic signaling regulates the Fanconi anemia pathway
Authors: Wang Jingming , Chan Bryan , Tong Michael , Seeliger Markus , Kim Hyungjin
Published: August 2018

Genome instability caused by defective DNA repair mechanisms is a hallmark of cancer and drives tumorigenesis. The Fanconi anemia (FA) DNA repair pathway contributes to the integrity of the genome by resolving DNA interstrand cross-links (ICLs) encountered during DNA replication. Deregulation of the FA pathway is associated with cancer predisposition and affects therapeutic outcomes against DNA-damaging cytotoxic chemotherapy. FANCD2 monoubiquitination by a multi-subunit ubiquitin E3 ligase, the FA core complex, is an essential gateway that connects the DNA damage response to enzymatic steps of DNA ICL repair. We have previously identified FAAP20 as a critical component of the FA pathway; and demonstrated that the phosphorylation of the Cdc4 phospho-degron (CPD) motif in FAAP20 by GSK3β is required for its proteasomal degradation mediated by the E3 ligase, SCFFBW7 complex. However, the upstream signaling that governs the FAAP20 phosphorylation status and its detailed mode of action for FAAP20 degradation remain elusive. Here, we show that PIN1, a phosphorylation-specific prolyl isomerase, is a key regulator of the integrity of the FA core complex and, therefore, FA pathway activity as a whole. By catalyzing a proline cis-trans isomerization of the phosphorylated backbone and thereby converting the substrate into a conformation that is favorable or refractory to downstream proteolytic signaling, PIN1 modulates the rate of protein turnover. We demonstrate that PIN1 interacts and catalyzes a phosphorylated Ser48-Pro49 motif of FAAP20 and by inducing a conformational change of FAAP20, PIN1 enhances the affinity of FAAP20 with PP2A (Protein phosphatase 2), the phosphatase of the CPD motif in FAAP20 and thereby prevents its degradation from the GSK3β-FBW7 mediated proteolytic signaling. Collectively, our studies uncover PIN1-dependent isomerization as a new regulatory mechanism for DNA ICL repair. Given that PIN1 overexpression is prevalent in diverse human cancers, identifying a way to regulate PIN1 activity in the FA pathway may help develop a PIN1 inhibitor as a chemosensitizer for cytotoxic chemotherapy to increase its therapeutic efficacy.

Citation Format: Jingming Wang, Bryan Chan, Michael Tong, Markus Seeliger, Hyungjin Kim. PIN1-SCFFBW7 proteolytic signaling regulates the Fanconi anemia pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2825.
Abstract 2831: Collateral lethality: A new target for personalized oncology
Authors: Lin Yu-Hsi , Hammoudi Naima , Satani Nikunj , Ackroyd Jeffrey , Khadka Sunada , Georgiou Dimitra , Marszalek Joe , Sun Yuting , Protopopova Marina , Francesco Maria E. Di , Czako Barbara , Wang Alan Y. , DePinho Ronald A. , Muller Florian L.
Published: August 2018

Large-scale genomic profiling efforts, such as The Cancer Genome Atlas (TCGA) have painted an unprecedentedly detailed picture of the genetic alterations that underlie carcinogenesis, yet the key challenge remains as to how to turn this information into actionable therapies. Genomic deletions are a frequent event in diverse cancers, which inactivate a limited number of tumor suppressor genes (“driver”-events) but frequently include many chromosomal neighbors as “passengers”, some of which play critical but redundant roles in normal cellular housekeeping. The overall hypothesis we propose to test is that collateral deletions of such “passenger” genes can be utilized as novel targets of synthetic lethality, an idea which we term “collateral lethality." The large number of passenger deleted genes, playing diverse functions in cell homeostasis, offers a rich repertoire of pharmacologically targetable vulnerabilities presenting novel opportunities for the development of personalized anti-neoplastic therapies. We have provided in vitro proof-of-principal of a collaterally deleted glycolytic gene Enolase 1 (ENO1) at the 1p36 tumor suppressor locus in glioblastoma (GBM), that leads to dramatic sensitization to inhibition of the redundant paralogue, ENO2. The next step is to take this concept to the clinic. Our overall goal is to generate a clinical candidate Enolase inhibitor for tumors with ENO1-deletions, like GBM for which there is no other treatment option and the therapeutic benefit could be quite substantial.

Citation Format: Yu-Hsi Lin, Naima Hammoudi, Nikunj Satani, Jeffrey Ackroyd, Sunada Khadka, Dimitra Georgiou, Joe Marszalek, Yuting Sun, Marina Protopopova, Maria E. Di Francesco, Barbara Czako, Alan Y. Wang, Ronald A. DePinho, Florian L. Muller. Collateral lethality: A new target for personalized oncology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2831.
Abstract 2837: Single-cell heterogeneity with transcriptional memory confers resistance to cancer therapy
Authors: Shaffer Sydney , Emert Benjamin , Raj Arjun
Published: August 2018

Targeted therapies for cancer are a promising class of drugs that inhibit the specific molecular alterations that underlie the uncontrolled proliferation seen in cancer. The primary shortcoming of targeted therapy is disease relapse, which is driven by a subpopulation of cells that are resistant to these drugs. This phenomenon is generally thought to be genetic in origin; however, our recent work on melanoma shows that nongenetic cellular plasticity may provide a mechanism of resistance to these therapies. Furthermore, we showed that through the addition of the drug itself, cells transition from this transient plasticity into a new, stably resistant cell state via cellular reprogramming, suggesting that the time an individual cell exists in a state is important for producing the divergent resistance phenotype. However, there are currently no methods available to quantify the timescale of these fluctuations for the whole transcriptome. Thus, broadly generalizing this concept of timescales for cellular plasticity, we developed a novel method for genome-wide quantification of the timescales of gene expression memory based on a modern version of the ingenious Luria-Delbrück fluctuation analysis. Specifically, we isolated individual cells, allowed them to expand, and then performed RNA sequencing on each of the samples. For genes with uniform expression, all the samples should have similar RNA sequencing counts; however, for genes that occasionally turn on in a subset of cells, the RNA sequencing counts across all samples should show high variance. In melanoma, this method revealed the gene expression state of rare cells resistant to targeted therapy. In a completely new model, triple-negative breast cancer, this method revealed a novel rare subpopulation of cells exhibiting resistance to chemotherapy. More generally, this method has the potential to reveal other new phenotypes associated with rare cell biology, including metastasis and the early stages of stem cell differentiation.

Citation Format: Sydney Shaffer, Benjamin Emert, Arjun Raj. Single-cell heterogeneity with transcriptional memory confers resistance to cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2837.
Abstract 2846: Extracellular ATP induces different types of drug resistances in cancer cells through ATP internalization-mediated intracellular ATP level increase
Authors: Chen Xiaozhuo , Wang Xuan , Zhang Haiyun , Qian Yanrong , Li Yunsheng , Cao Yanyang , Shriwas Pratik
Published: August 2018

Cancer cells are able to uptake extracellular ATP (eATP) via macropinocytosis to elevate intracellular ATP (iATP) levels, enhancing their survival from drug treatment (1). However, the involved drug resistance mechanisms are unknown. Here we investigated the roles of eATP as either an energy or a phosphorylating molecule in general drug resistance mediated by ATP internalization and iATP elevation. We report that eATP increased iATP levels and promoted drug resistance to various tyrosine kinase inhibitors (TKIs) and chemo-drugs in human cancer cell lines of five cancer types, including non-small cell lung cancer, breast cancer, colon, liver cancer, and pancreatic cancer (2). In A549 lung cancer cells, the resistance was downregulated by macropinocytosis inhibition or siRNA knockdown of PAK1, an essential macropinocytosis enzyme, strongly suggesting that the resistance is mediated at least in part by micropinocytosis (and therefore ATP internalization) (2). The elevated iATP upregulated the efflux activity of ABC transporters in A549 and SK-Hep-1 cells, as well as phosphorylation of PDGFRα and proteins in the PDGFR-mediated Akt-mTOR and Raf-MEK signaling pathways in A549 cells, increasing cell cycle progression and decreasing drug-induced apoptosis. Similar phosphorylation upregulations were found in A549 tumors. The resistance was not significantly affected by purinergic receptor-mediated signaling. These results demonstrate, for the first time, that eATP induces different types of drug resistance by eATP internalization and iATP elevation, implicating the ATP-rich tumor microenvironment in cancer drug resistance, expanding our understanding of the roles of eATP in the Warburg effect and offering new anticancer drug resistance targets.

References

1.Qian, Y. et al. Mol Cancer Res 14(11); 1087-1096 (2016)

2.Wang, X. et al. Oncotarget 8, 87860-87877 (2017).

Citation Format: Xiaozhuo Chen, Xuan Wang, Haiyun Zhang, Yanrong Qian, Yunsheng Li, Yanyang Cao, Pratik Shriwas. Extracellular ATP induces different types of drug resistances in cancer cells through ATP internalization-mediated intracellular ATP level increase [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2846.
Abstract 2847: Investigation of the combinatorial treatment strategy for BRAFmutant anaplastic thyroid cancer based on real time monitoring technology
Authors: Byeon Hyung Kwon , Ku Minhee , Choi Ji Hye , Kee Hyunjung , Kim Ji-Hoon , Koh Yoon Woo , Yang Jaemoon
Published: August 2018

Purpose Previously, we have identified that c-Met-mediated reactivation of PI3K/AKT pathway contributes to acquired resistance to BRAF inhibitor in BRAF mutant (BRAFm) anaplastic thyroid cancer (ATC). So dual inhibition of BRAF and c-Met led to sustained treatment response. However, previous RTK array result in 8505C cells following PLX4032 treatment revealed increased expression of Src together with c-Met. Therefore further in-depth investigation is required to elucidate the optimal combinatorial strategy to maximize the treatment effects in BRAFm ATC. Using non-invasive, real-time monitoring in vitro and in vivo technologies, the best treatment strategy was investigated with consideration of not only the specific treatment regimens but also the mode of combinatorial treatment.

Methods Real-time cell analysis (RTCA), a cell-based label-free technology using electronic impedance sensors was adopted as a strategy for real-time screening of cellular response to drugs. Live wound healing assays with supporting live cell microscopy and confocal microscopy were conducted for in vitro analyses and both ectopic xenograft and metastatic mouse models were utilized for in vivo analyses. Tumor characteristics were evaluated using 9.4T MR diffusion weighted imagings.

Results Dual inhibition of BRAF and c-Met was not fully effective in overcoming BRAF inhibitor resistance in BRAFm ATC. Src-mediated signal has a greater contribution than c-Met-mediated signaling in conferring BRAF inhibitor resistance. Inhibition of Src exerted a more potent effect than c-Met inhibition. Simultaneous drug treatment was more effective than sequential therapy. Combinatorial treatment of BRAF inhibitor (PLX4032) and Src inhibitor (dasatinib) presented the maximal therapeutic effect against BRAFm ATC.

Conclusions Real-time/non-invasive monitoring assays sensitively provides more accurate information regarding drug response.

Citation Format: Hyung Kwon Byeon, Minhee Ku, Ji Hye Choi, Hyunjung Kee, Ji-Hoon Kim, Yoon Woo Koh, Jaemoon Yang. Investigation of the combinatorial treatment strategy for BRAFmutant anaplastic thyroid cancer based on real time monitoring technology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2847.
Abstract 2860: The chemosensitizing and oncolytic effects of Dihydrotanshinone in glioblastoma
Authors: Kumar Varun , Leonardi Donna
Published: August 2018

Temozolomide is the primary chemotherapeutic agent used to treat glioblastoma. However, many tumors are initially resistant to or develop resistance to temozolomide, mainly due to high levels of O6-methylguanine DNA transferase (MGMT) which repairs DNA damage traditionally caused by temozolomide. Dihydrotanshinone (DHT) is extracted from Salvia miltiorrhiza, a Chinese medicinal plant, and has been shown to have antiproliferative effects on various cancer cell lines. DHT has also been to shown to induce apoptosis via induction endoplasmic reticulum stress, that can reportedly sensitize cells to temozolomide. MTS cellular proliferation assays or trypan blue viability assays were used to determine the effects of DHT/temozolomide combinatorial treatment. Enzyme-linked immunosorbent assay (ELISA) was used to determine effects on MGMT, P-glycoprotein, AKT, p65, and IlkB levels after singular and combinatorial treatment. DNA microarray was used to gauge gene expression and miRNA levels. Solute clearance of the blood-brain barrier was analyzed using a co-culture model system and confocal microscopy. DHT had a selectively cytotoxic synergistic oncolytic effect in a MGMT-deficient cell line and a sensitizing effect in a MGMT-expressing cell line. Cytotoxicity due to DHT was shown to be reactive oxygen species-dependent, while the combinatorial effect of DHT and temozolomide synergistically reduced MGMT and P-glycoprotein expression and protein levels. In addition, the NFkB complex was sensitive to combinatorial treatment, while AKT activity and gene expression was downregulated in response to treatment. DHT was shown to augment temozolomide efficacy, indicating that, since DHT can penetrate the blood–brain barrier, temozolomide in combination with DHT may represent a promising therapeutic option for glioblastoma.

Citation Format: Varun Kumar, Donna Leonardi. The chemosensitizing and oncolytic effects of Dihydrotanshinone in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2860.
Abstract 2862: Inhibition of DNA repair pathways in breast cancer is a potential mechanism of action of IT-139
Authors: Conde Isabel , Fallah Yassi , Demas Diane , Bakewell Suzanne , Lynce Filipa , Shajahan-Haq Ayesha N.
Published: August 2018

IT-139 is a novel small-molecule drug that demonstrated a manageable safety profile at the maximum tolerated dose and modest antitumor activity as a monotherapy in a phase I clinical trial. A high affinity for albumin and unique chemistry allow increased uptake and activation of IT-139 within tumor cells, respectively. To date, IT-139 has been reported to inhibit the endoplasmic reticulum stress-sensing protein GRP78. However, different cancer cells show disparate responses to IT-139 and the precise mechanism of action in high-response versus low-response cancer cells remains unclear. In vitro studies have indicated that IT-139 induces cytostatic rather than cytotoxic effects as a monotherapy. In this study, to understand IT-139 mediated signaling mechanism in breast cancer cells, we used estrogen receptor-positive (ER+) MCF7 breast cancer cells to obtain gene-metabolite integrated models following IT-139 treatment (10 or 100 μM for 72 h). Our data showed that IT-139 significantly reduced expression of genes involved in the DNA repair pathway, including BRCA1 and RAD51, following treatment with IT-139 in MCF7 cells. Validation of our findings in other breast cancer cell lines including MDA-MB-175 (ER+), MDA-MB-231 and MDA-MB-468 (both ER-) cells also showed increased phosphorylation of H2AX (Ser139) following IT-139 treatment, suggesting lack of proper DNA surveillance and subsequent accumulation of DNA double-stranded breaks (DSB). Consequently, we investigated the effects of combination treatment approaches with IT-139 and other clinical anticancer therapies that interfere with nucleotide synthesis or DNA damage repair pathways, particularly in ER- breast cancers. Our data showed additive effects of IT-139 in combination with capecitabine (5-FU prodrug) or olaparib (PARP inhibitor) and a synergistic effect with carboplatin in ER- breast cancer cell lines. Further analysis of IT-139 mediated signal transduction in breast cancer cells will reveal whether it can be combined with aforementioned chemotherapies to circumvent resistance.

Citation Format: Isabel Conde, Yassi Fallah, Diane Demas, Suzanne Bakewell, Filipa Lynce, Ayesha N. Shajahan-Haq. Inhibition of DNA repair pathways in breast cancer is a potential mechanism of action of IT-139 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2862.
Abstract 2863: Dissecting the structural basis for inhibitors of RNA-binding proteins
Authors: Lan Lan , Xing Minli , Wu Xiaoqing , Gao Philip , Douglas Justin T. , Zhan Yu , Gardashova Gulhumay , Liu Jiajun , Hanzlik Robert P. , Aubé Jeffrey , Neufeld Kristi L. , Oakley Berl R. , Guzman Roberto N. De , Xu Liang
Published: August 2018

RNA-binding proteins (RBPs) are key regulators of cellular functions, especially in post-transcriptional regulations. Dysregulation of RBPs is implicated in many diseases including cancer. One of the RBPs that is overexpressed in a variety of human cancer is Musashi-2 (MSI2). Elevated MSI2 expression is associated with ectopic oncogenic pathways, including but not limited to NUMB/Notch, PTEN/mTOR, TGF-β/SMAD3, making MSI2 a promising therapeutic target for cancer. In the case of NUMB, MSI2 binds to and negatively regulates translation of NUMB, a negative regulator of Notch signaling. Protein structure is critical for drug discovery and structure-based rational design. However, so far there is no structure available for MSI2 protein. Recently, using nuclear magnetic resonance (NMR) we solved the first solution structure of MSI2 RNA-recognition Motif 1 (MSI2-RRM1) that forms the major long narrow RNA-binding pocket. Using a fluorescence polarization (FP) chemical libraries screening, we identified several groups of hit compounds that disrupt the binding of MSI2-NUMB RNA potently. These compounds induced apoptosis, inhibited cancer cell proliferation, invasion and metastasis in multiple tumor models tested in vitro and in vivo. They also interfered with cancer stem cell functions with reduced tumorsphere formation. Several lead compounds showed promising efficacy in animal tumor models of human breast, prostate and colon cancer. To investigate the specificity of the compounds towards other RBPs, the compounds were tested in biophysical binding assays such as FP, time resolved Fluorescence Resonance Energy Transfer assay (TR-FRET), NMR spectroscopy and cell-based assays. We compared the affinity of these MSI2 inhibitors towards other RBPs such as MSI1 and HuR. Using docking and molecular dynamics simulation, we analyzed the structure-activity relationship of these inhibitors towards different RBP targets. This work adds significant information to the structure of MSI2-RRM1 and the structural basis for designing more potent and specific inhibitors of RBPs.

Citation Format: Lan Lan, Minli Xing, Xiaoqing Wu, Philip Gao, Justin T. Douglas, Yu Zhan, Gulhumay Gardashova, Jiajun Liu, Robert P. Hanzlik, Jeffrey Aubé, Kristi L. Neufeld, Berl R. Oakley, Roberto N. De Guzman, Liang Xu. Dissecting the structural basis for inhibitors of RNA-binding proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2863.
Abstract 2873: Targeting RAGE for prostate cancer treatment
Authors: Ernzen Kyle J. , Dasari Subramanyam , Munirathinam Gnanasekar
Published: August 2018

Prostate cancer (PCa) is the second most common cancer among men and is the third leading cause of cancer-related death in the United States. In 2017 alone, an estimated 161,360 men will be diagnosed with PCa and approximately 26,730 deaths will result from this disease in the United States. The most common methods for PCa treatment include surgery, radiotherapy, and hormone therapy. These treatment options unfortunately have serious drawbacks such as the possibility of organ damage, adverse side effects, and eventual PCa resistance to treatment. Due to the severe threat that PCa poses across the male population, it is vital that more effective and safe treatment options are explored. Previous research has indicated that the receptor for advanced glycation end products (RAGE) plays a vital role in the survival of prostate cancer cells, indicating that RAGE could be a potential therapeutic target. In the present study, we focused on targeting RAGE using two different antagonists, RAGE aptamer and azeliragon as potential alternative therapies for PCa. Confocal microscopy assays initially confirmed that RAGE aptamer expression strongly binds to VCaP cells while azeliragon treatment decreased the expression of RAGE in these cells. MTT cell viability assays indicated that RAGE aptamer and azeliragon decrease PCa cell proliferation in a dose-dependent manner for VCaP and DU-145 cell lines respectively. Cell cycle analysis suggested that RAGE aptamer and azeliragon-treated DU-145 cells enforced cellular G0/G1 phase arrest in a dose-dependent manner. Annexin V FITC and ROS immunofluorescence assays showed that RAGE aptamer and azeliragon induce apoptosis potentially by inducing oxidative stress in PCa cells. Apoptosis activation in RAGE aptamer and azeliragon-treated VCaP cells was confirmed by the detection of PARP-1 cleavage through western blotting. Taken together, our findings suggest that RAGE aptamer and azeliragon may be a promising alternative form of therapy for PCa patients.

Citation Format: Kyle J. Ernzen, Subramanyam Dasari, Gnanasekar Munirathinam. Targeting RAGE for prostate cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2873.
Abstract 2874: Benzaldehyde inhibits the multiple signals in cancer by suppressing the binding activity of overexpressed 14-3-3ζ and represses the pRb/E2F transcriptional pathway
Authors: Saitoh Jun , Saya Hideyuki
Published: August 2018

Benzaldehyde (BA) is the simplest aromatic aldehyde composed of only benzene ring and aldehyde. It is constituents of many fruits and flavor essence. Anti-cancer effect of BA was first reported in 1980, and multi institutional clinical trials by using derivatives of BA were performed in 1980's in Japan. However, clinical study was aborted due to the policy and strategic change in the company. Only the safety of the compound was confirmed. Given that we had multiple effective clinical cases including pancreatic cancers and chemo-resistant non Hodgkin lymphoma, we attempted to elucidate the mechanism of anti-cancer effect of BA. We have previously reported that BA inhibits cancer-related multiple pathways such as PI3K/AKT/mTOR, STAT3, NFκB and ERK, leading to cell death and growth suppression in BA sensitive pancreatic cancer cell line BxPC3, but not in normal cells. We found that such mechanism is based on the BA-mediated inhibition of 14-3-3ζ binding to its client proteins. To further analyze the BA-mediated regulation of 14-3-3ζ, we generated HEK293 cells overexpressing 14-3-3ζ and confirmed that BA suppressed mTOR pathway in the 14-3-3ζ expressing HEK293 cells but not in parental HEK293 cells. Furthermore, we found that depletion of 14-3-3ζ suppresses mTOR signal in BxPC3 and BA-induced reduction of phosphorylation of mTOR signal proteins. Moreover, we performed microarray analysis on three pancreatic cancer cell lines (BxPC-3, AsPC1, PANC1) and found that stress response genes were upregulated in BxPC-3 cells by BA treatment. E2F2 and E2F targeted genes and histone cluster genes in 6p22 were suppressed in not only BxPC-3 but also BA low-sensitive PANC1. We confirmed that BA treatment suppresses the expression of E2F2 and E2F target genes. Of note, protein levels of pRb and cyclin D were also suppressed by BA treatment. Those data suggest that BA inhibits overexpressed 14-3-3ζ mediated multiple signaling pathway and consequently represses pRB/E2F and E2F target genes, resulting in inhibition of cell cycle progression, DNA replication and DNA repair. We propose that those mechanisms can explain why this safe compound is effective for refractory cancers.

Citation Format: Jun Saitoh, Hideyuki Saya. Benzaldehyde inhibits the multiple signals in cancer by suppressing the binding activity of overexpressed 14-3-3ζ and represses the pRb/E2F transcriptional pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2874.
Abstract 2881: RET signaling in pancreatic cancer: A novel target by metformin to suppress cancer progression
Authors: Chang Huailong , Xiao Zhiyong , Li Tao , Zhang Lanjing , Lin Yong , Carpizo Darren , Tan Xianglin
Published: August 2018

Metformin has recently gained attention as an anti-cancer drug and/or a chemoprevention agent because of its roles in inhibiting mTOR, lowering hyperinsulinemia, modulating inflammatory responses, and selectively killing cancer stem cells. However, the key underlying molecular mechanisms for the inhibitory effects of metformin on pancreatic cancer progression remain largely unknown. RET (REarranged during Transfection), a single-pass transmembrane receptor tyrosine kinase and its ligand, glial cell-derived neurotrophic factor (GDNF), were strongly expressed in pancreatic cancer and correlated to invasion and worse survival after surgical resection. In this study, we investigated the roles of RET in the inhibitory effects of metformin on pancreatic cancer cell growth and migration. Real-time PCR and Western blot were used to determine mRNA and protein levels of molecular markers, respectively. siRNAs were adopted to specifically knockdown target genes. Boyden chamber assay was applied to assess cell migration in vitro. We observed that metformin treatment significantly reduced the mRNA and protein expression of RET in a dose-dependent manner in pancreatic cancer cells PANC-1 and MIA PaCa-2. The inhibitory effects of metformin on RET expression were not diminished when AMPK was knockdown by AMPK siRNA or the specific inhibitor, Compound C, suggesting that metformin may suppress RET in an AMPK-independent manner. Metformin treatment or RET knockdown by RET siRNA significantly decreased the phosphorylation of NF-κB and p70S6K, while modestly reduced the phosphorylation of the ERK, AKT, or STAT3. Furthermore, metformin treatment or RET knockdown significantly inhibited GDNF-induced cell migration. These data indicate that targeting RET with metformin may be an attractive and novel strategy for the prevention and treatment of pancreatic cancer progression and metastasis. Further in vitro and in vivo studies are warranted to investigate how metformin modulates RET signaling to inhibit the progression and metastasis of pancreatic cancer.

Citation Format: Huailong Chang, Zhiyong Xiao, Tao Li, Lanjing Zhang, Yong Lin, Darren Carpizo, Xianglin Tan. RET signaling in pancreatic cancer: A novel target by metformin to suppress cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2881.
Abstract 2882: Elucidation of the mechanism of action of the multi-kinase inhibitor midostaurin in NSCLC
Authors: Ctortecka Claudia , Kuenzi Brent , Fang Bin , Sumi Natalia , Izumi Victoria , Kinose Fumi , Haura Eric , Koomen John , Rix Uwe
Published: August 2018

Background: Non-small cell lung cancer (NSCLC) constitutes a strong unmet medical need in spite of recent successes with targeted drugs and immunotherapy in some subsets. We had observed the recently FDA-approved drug midostaurin to exhibit potent anticancer activity in a subset of NSCLC cell lines that was unrelated to its known targets FLT3 or PKC. We here explored a new repurposing opportunity and investigated the mechanism of action (MoA) of midostaurin in NSCLC utilizing a multilayered functional proteomics approach.

Methods: Chemical proteomics identified kinase targets and global and tyrosine phosphoproteomics generated a network view of intracellular signaling upon midostaurin treatment. Functional validation was performed using RNA interference and selective small-molecule probes. Cell viability assays were carried out using CellTiterGlo and colony formation assays. Induction of apoptosis was determined by immunoblotting and fluorescent microscopy. Flow cytometry assessed cell cycle stages. Signaling changes were determined by immunoblotting.

Results: Viability screening confirmed anticancer activity in several midostaurin-sensitive NSCLC cell lines at clinically relevant concentrations independent of its canonical targets. Using chemical proteomics, we identified multiple midostaurin kinase targets. Network-based integration of the chemical proteomics and quantitative tyrosine and global phosphoproteomics data using STRING for protein-protein interactions and NetworKIN for substrate-kinase predictions revealed several nodes to be relevant for the MoA of midostaurin. Subsequent functional validation showed that simultaneous inhibition of TBK1, PDK1 and AURKA was required to mimic midostaurin's cellular effects in these cells. Immunoblotting for downstream targets of these kinases revealed PLK1 as a critical signaling effector. Rational combination of midostaurin with the more potent PLK1 inhibitor BI2536 caused strong synergy.

Conclusion: Midostaurin showed activity in a subset of NSCLC cells. Combination of complementary functional proteomics approaches and subsequent network-based integration revealed novel insight into the MoA of midostaurin and NSCLC vulnerabilities. We utilized this mechanistic knowledge for the rational combination of midostaurin with the clinical PLK1 inhibitor BI2536, which is highly synergistic and has strong potential for clinical translation.

Citation Format: Claudia Ctortecka, Brent Kuenzi, Bin Fang, Natalia Sumi, Victoria Izumi, Fumi Kinose, Eric Haura, John Koomen, Uwe Rix. Elucidation of the mechanism of action of the multi-kinase inhibitor midostaurin in NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2882.
Abstract 2887: Evaluating the effectiveness of MAPK, AKT, and mTOR inhibitors in reducing proliferation in cellular models of papillary and follicular thyroid cancer
Authors: LeBoeuf Brianna , Anderson Braxton , Young Margaret , Franco Aime , MacDonald Laura
Published: August 2018

Thyroid cancer is the most common endocrine malignancy, and incidence has been steadily increasing, suggesting occurrence will exceed that of colon cancer by 2030. Papillary and follicular thyroid cancer subtypes are most common, which are activated by BRAF and HRAS mutations, respectively. Interestingly, papillary and follicular thyroid cancers are associated with different pathologies and metastases. Currently, both papillary and follicular thyroid cancers are treated by surgical removal of the thyroid, followed by radioactive iodine treatment to eliminate any remaining tumor cells. Unfortunately, for individuals with progressive thyroid cancer, these treatment options are not effective, highlighting a need for increased investigation into mechanisms of drug sensitivity. In this study, we sought to evaluate mechanisms of differential drug sensitivity through RNA-seq analysis of cell lines derived from mouse models of papillary and follicular thyroid cancer. Additionally, we evaluated the effects of MAPK, AKT, and mTOR inhibitors on murine papillary and follicular thyroid cancer cell proliferation. To accurately assess drug efficacy, we calculated GR50 concentrations for each inhibitor in six papillary and follicular thyroid cancer cell lines. We discovered that papillary and follicular thyroid cancer cell lines responded differently to MAPK and AKT inhibitors, suggesting that treatment approaches should be tailored to subtype despite having mutations in the same signaling pathway.

Citation Format: Brianna LeBoeuf, Braxton Anderson, Margaret Young, Aime Franco, Laura MacDonald. Evaluating the effectiveness of MAPK, AKT, and mTOR inhibitors in reducing proliferation in cellular models of papillary and follicular thyroid cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2887.
Abstract 2891: Molecular profiling of hormone-resistant prostate cancer cells with TRPV6 oncochannel knockout/knockdown
Authors: Lutes Tyler , Davey Michelle , Rice Christopher , MacCormack Tyson , Stewart John M. , Dugourd Dominique
Published: August 2018

TRPV6 calcium channel is a recognized oncochannel over-expressed in a number of epithelial cancers (e.g. breast, ovarian, prostate). TRPV6, a member of one of seven subfamilies of TRP channels, plays an important role in Ca2+ absorption from the intestinal lumen. TRPV6 over-expression is associated with a poor prognosis particularly with breast and prostate cancers. Although some TRPV6 mechanistic work is available and indicates that NFAT is involved, there is a need to better understand pathways involved in its mechanism of action (MOA). These data could allow the identification of clinically relevant biomarkers and identify potential for synergy of TRPV6-targeting therapies with other anti-cancer treatments. To determine which key pathways TRPV6 influences in prostate cancer, two TRPV6 knockouts (KO) were produced in a castration-resistant prostate cancer cell line (PC-3) using the CRISPR-Cas9 approach with two different gRNAs. In addition, knockdowns (KD) were produced by transfecting PC-3 cells with two TRPV6 siRNA targeting different regions of TRPV6 mRNA. The mRNA expression of 184 genes was analyzed using an RT-qPCR TaqMan array. The panel consisted of genes involved in cancer calcium signalling, genes that have been associated with the MOA of TRPV6 and genes directly or indirectly involved in NFAT signalling. Gene profiling results were generally consistent between the TRPV6 KD and KO cells. Results showed a dramatic reduction in expression of TRPV6 mRNA in the KO/KD cells (14 to 166-fold). TRPV6 KO/KD affected multiple genes involved in cancer cell proliferation (e.g. ESR1, CDH1), metastasis (e.g. SNAI1, MUC1/16), resistance to apoptosis (e.g. Bcl-2-type proteins, Fos) and angiogenesis (e.g. VEGFA/B, FLT4), as well as genes involved in the M2 immune tumor microenvironment (e.g. IL6 and VEGFA). The TRPV6 KO/KD data confirm the basic MOA of TRPV6, activating NFAT by increasing cytosolic calcium and provide information on the downstream pathways involved (e.g. NF-kB, estrogen and MAPK). The data allow for the identification of clinical efficacy biomarkers for TRPV6 inhibitors (e.g. a panel of CXCL12, FLT4, MUC16 and IL-6). Furthermore, results indicate TRPV6 inhibitors have the potential to modify the immune composition of the tumor microenvironment from a tumor promoting to a tumor control response, and thus may help trigger an anti-cancer immune defence. This study demonstrates a central role of TRPV6 and calcium signalling in cancer development.

Citation Format: Tyler Lutes, Michelle Davey, Christopher Rice, Tyson MacCormack, John M. Stewart, Dominique Dugourd. Molecular profiling of hormone-resistant prostate cancer cells with TRPV6 oncochannel knockout/knockdown [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2891.
Abstract 2897: Therapeutic efficacy of antibodies targeting domain 1 of HSPG2 in breast cancer
Authors: Khanna Vidhi , Kalscheuer Stephen , Panyam Jayanth , Yang Da , Li Sihan
Published: August 2018

Significant advancements in chemotherapy have improved the survival rates of patients presenting with local or regional breast cancer to as high as 99% and 85%, respectively. On the contrary, patients with metastatic breast cancer have a dismal 5-year survival rate of 26%. Thus, there is an urgent need for research strategies directed towards treatment of metastasis. Our lab used a phage display-based cell panning procedure to develop two fully humanized antibodies (Clone 6 and AM6) that are able to bind specifically to breast cancer metastatic cells. Target deconvolution revealed HSPG2/Perlecan Domain 1 as the cell surface antigen bound by the antibodies. Immunohistochemistry studies revealed high HSPG2 expression across various tumor subtypes including melanoma, bladder cancer, glioblastoma and ovarian cancer. There was a significant correlation between high HSPG2 and with poor survival in bladder and ovarian cancers. Interestingly, we observed significant tumor growth inhibition in the triple-negative MDA-MB-231 breast cancer xenograft model. This efficacy was significantly reduced when the tumor growth inhibition studies were repeated in NSG mice, suggesting NK cell-mediated antibody dependent cellular cytotoxicity (ADCC) as the potential mechanism of action. The data presented here point to the relevance of HSPG2 as a novel target for not only breast cancer but also other malignancies. We also show the potential of Clone 6 and AM6 as therapeutic and targeting agents. Further studies are required to understand the significance of HSPG2 overexpression and its correlation with tumor progression as well as to confirm the mechanism of action with Clone 6 and AM6 antibodies.

Citation Format: Vidhi Khanna, Stephen Kalscheuer, Jayanth Panyam, Da Yang, Sihan Li. Therapeutic efficacy of antibodies targeting domain 1 of HSPG2 in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2897.
Abstract 2898: Inhibition of HDAC6 as a novel targeted therapy in breast cancer
Authors: Zeleke Tizita Z. , Putcha Preeti , Yu Jiyang , Califano Andrea , Silva Jose M.
Published: August 2018

The identification and use of biomarkers is critical to guided clinical care. Cancer treatment has made use of many biomarkers to determine which therapeutic avenues are adequate for each patient. For example, the use of Trastuzumab is intimately linked to the presence of HER2/NEU amplification in breast cancers. Previous work from our group established the importance of HDAC6 in maintaining the homeostasis of inflammatory breast cancer cells (IBC). We also reconstructed the HDAC6 regulon using gene expression profile signatures from over 900 primary breast cancers to obtain a biomarker that guide the use of HDAC6 inhibitors towards the right patient group. This method identified a regulon of 162 transcripts as a set of transcriptional targets whose expression is affected by HDAC6 master regulator activity. Finally, the expression of all transcripts that are co-regulated by HDAC6 in the breast cancer samples were integrated into a single score termed as the HDAC6 score. Through this work, we established that HDAC6 score is associated with cancer cell response to HDAC6 inhibitor ACY1215.

HDAC6, one of eighteen deactylases, is distinct from all other HDACs as it is located mainly in the cytosol and has non-chromatin substrates such as α-tubulin and HSP90. HDAC6 inhibitors have gained a lot of momentum in the clinic for hematological malignancies and our computational analysis has also revealed that these tumors also exhibit a high HDAC6 score. Therefore, by identifying the HDAC6 score for tumors, we can widen the scope of using HDAC6 inhibitors to a wide range of cancers.

Our increasing understanding of tumor biology has led to the use of combinatorial therapies to fully benefit from cancer therapy. This project will encompass the use of combinatorial therapy containing HDAC6 inhibitors and paclitaxel to demonstrate an increased potential in treating breast cancer. These studies will closely mirror the clinical trial that we are undertaking with our collaborators from Acetylon where metastatic breast cancer patients are being treated using HDAC6 inhibitor ACY1215 in combination with nab-paclitaxel (NCT02632071). This project aims to provide the frame work to utilize for the first time the HDAC6 score as a marker for identifying tumors that will be most susceptible to HDAC6 inhibition in the clinic.

Citation Format: Tizita Z. Zeleke, Preeti Putcha, Jiyang Yu, Andrea Califano, Jose M. Silva. Inhibition of HDAC6 as a novel targeted therapy in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2898.