bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3071: Exploring the molecular mechanism underlying SETD2-PI3Kβ synthetic-lethal interaction in renal cell carcinoma (RCC)
Authors: Terzo Esteban , Chiang Jeanne , Rathmell W. Kimryn
Published: July 2017

Epigenetic regulators are emerging as major drivers of renal cell carcinoma (RCC). SETD2, a tumor suppressor commonly mutated in RCC, is the sole enzyme responsible for tri-methylating histone H3 on lysine 36 (H3K36me3) of actively transcribed genes. We recently discovered that SETD2 also trimethylates the non-histone target α-tubulin on lysine 40 (α-TubK40Me3) of mitotic microtubules. We have demonstrated that mutations altering SETD2’s ability to trimethylate H3K36 and α-TubK40 are associated with genomic instability in cancer progression. We hypothesized that defective SETD2 methylating activity gives rise to deleterious cell signaling cascades that promote the tumorigenic features of SETD2 loss. Preliminary data show that RCC-derived 786-0 cells that are deleted for SETD2 (SETD2-KO) or harbor pathogenic mutations are more sensitive to treatment with the PI3Kβ-specific inhibitor TGX-221 than their wild type counterparts. Phosphatidylinositol 3kinases (PI3Ks) are a family of lipid kinases that coordinate signals from growth factors, cytokines and other environmental cues, translating them into intracellular signals controlling diverse signaling pathways. Several members of the PI3K/AKT signaling pathway, including the PI3K α and β isoforms, are frequently mutated in a variety of cancers, making this pathway a prime drug target for anti-cancer therapy. These pathways control numerous biological processes, such as cell proliferation, growth, and motility. Examination of structurally variant SETD2 mutants suggests that the loss of SETD2 methylating activity mediates this interaction, and future studies will examine the functional consequences of inhibiting this pathway. In summary, these data suggest a synthetic-lethal type of interaction between SETD2 loss (and specifically loss of methylating activity) and an aberrant PI3K/AKT pathway.

Citation Format: Esteban Terzo, Jeanne Chiang, W. Kimryn Rathmell. Exploring the molecular mechanism underlying SETD2-PI3Kβ synthetic-lethal interaction in renal cell carcinoma (RCC) [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 3071. doi:10.1158/1538-7445.AM2017-3071
Abstract 308: The PP2A-B56 phosphatase opposes cyclin E autocatalytic degradation via site-specific dephosphorylation
Authors: Davis Ryan J. , Swanger Jherek , Clurman Bruce E.
Published: July 2017

Cyclin E, in conjunction with its catalytic partner cyclin-dependent kinase 2 (CDK2), regulates cell cycle progression as cells exit quiescence and enter S-phase. Multiple mechanisms control cyclin E-CDK2 activity during the cell cycle, including phosphorylation-dependent cyclin E ubiquitylation by the SCFFbw7 ubiquitin ligase. Serine 384 (S384) is the critical cyclin E phosphorylation site that stimulates Fbw7 binding and subsequent cyclin E degradation. Because S384 is autophosphorylated by CDK2, cyclin E therefore instigates its own degradation in an autocatalytic manner. This presents a paradox as to how cyclin E-CDK2 is able to phosphorylate its numerous substrates prior to cyclin E autophosphorylation-catalyzed degradation. Here we find that the PP2A-B56 phosphatase specifically dephosphorylates cyclin E at S384, thereby uncoupling cyclin E degradation from cyclin E-CDK2 activity. Furthermore, the rate of S384 dephosphorylation is high in interphase and low in mitosis, allowing PP2A-B56 to oppose autocatalytic cyclin E degradation and maintain cyclin E-CDK2 activity at the G1/S transition.

Citation Format: Ryan J. Davis, Jherek Swanger, Bruce E. Clurman. The PP2A-B56 phosphatase opposes cyclin E autocatalytic degradation via site-specific dephosphorylation [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 308. doi:10.1158/1538-7445.AM2017-308
Abstract 3115: Characterization of CDCA1 protein as a novel prognostic biomarker and therapeutic target for oral cancer
Authors: Phung Thang Manh , Takano Atsushi , Yoshitake Yoshihiro , Shinohara Masanori , Murakami Yoshinori , Daigo Yataro
Published: July 2017

Oral cavity carcinoma (OCC) is one of the most common causes of cancer-related death worldwide. Current therapies for OCC still show poor clinical outcome with five-year survival rate of less than 63%. Therefore, next generation biomarkers and therapeutic strategies for OCC are eagerly awaited. We selected genes that were specifically overexpressed in the majority of OCC using our gene expression profile database, and identified cell division cycle associated 1 (CDCA1) that appeared to encode nuclear protein as a candidate. Immunohistochemical analysis using tissue microarray covering 99 OCC tissues detected that CDCA1 protein was expressed in 67 cases of 99 OCC tissues (68%), but not in normal oral epithelia. In addition, high levels of CDCA1 protein expression was significantly associated with poor prognosis for OCC patients (P = 0.0244 by log-rank test). Furthermore, knockdown of CDCA1 expression by siRNAs significantly inhibited the growth of tumor cells. Moreover, flow cytometric analysis, multiple apoptosis assays, and live cell imaging experiment revealed that CDCA1 could be important for the growth and survival of OCC cells, probably through the regulation of mitotic prophase and/or apoptosis. Our findings suggest that CDCA1 is likely to be a novel prognostic biomarker and a therapeutic target for OCC patients.

Citation Format: Thang Manh Phung, Atsushi Takano, Yoshihiro Yoshitake, Masanori Shinohara, Yoshinori Murakami, Yataro Daigo. Characterization of CDCA1 protein as a novel prognostic biomarker and therapeutic target for oral 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 3115. doi:10.1158/1538-7445.AM2017-3115
Abstract 3118: Histological subtypes correlate with distinct genetic abnormalities in defining therapeutic targets in medulloblastoma
Authors: Gelnick Samuel , Amin Anubhav G. , Gordon Molly , Medeiros Raphael Salles Scortegagna de , Zanon Nelci , Murali Raj , Jhanwar-Uniyal Meena
Published: July 2017

Medulloblastoma (MB) is the most common primary pediatric malignant brain tumor. Genetic classification and distinct histologic subtypes defines MB into 4 groups: classic (WNT), sonic hedgehog (Shh), group 3 and group 4. In addition, recent studies have provided evidence that p53 abnormalities is seen in various sub-groups of MB and that p53-mutated SHH MBs commonly harbor genetic anomalies including MYCN and GLI2 amplifications, which confers drug resistance. Here, we test the hypothesis that specific immunohistological markers, their correlation with the amplification of the oncogene MYC, and abnormalities in tumor suppressor gene p53, may define their metastatic potential.

Materials and Methods: Immunohistological analysis of MB tumors (n=41) was evaluated for the expression of glioma transcription factor 1 (GLI-1), Grb2-associated binding protein 1 (GAB-1), natriuretic peptide receptor (NPR), voltage-gated potassium channel (KV1) and mutant p53. FISH analysis was performed to determine MYC amplification or iso- P53. p53-mutant MB cell line was used to investigate the signaling pathway leading to proliferation, migration, and drug resistance using HDAC (LBH-589) and PI3K/mTOR (BKM-120/rapamycin) inhibitors.

Results: we showed that: 1) GAB-1 was highly expressed in the Shh group (82%) and KV1 expression was evenly distributed in all subtypes; 2) No obvious correlation with expression of GLI-1, GAB-1, NPR, or KV1 with metastasis was seen; 3) Analysis of loss of p53 and overexpression of MYC varied in each subtype; 4) Combined Treatment with LBH-589 and BKM-120 reduced cell proliferation, migration and S-phase entry, however, MB cells were resistant to BKM -120 treatments, while LBH-589 caused massive apoptosis; 5) Tumor formation was suppressed by BKM-120 given with mTOR inhibitors.

Conclusion: the result of this study suggests that expression of GLI-1, GAB-1, NPR, KV1 and p53 was important in defining the subgroups of MB, although their metastatic potential remains to be understood. MB cells shows reduced cell proliferation, cell cycle and migration, when treated with HDAC inhibitor LBH-589, however these cells display complete drug resistance to PI3K inhibitors.

Citation Format: Samuel Gelnick, Anubhav G. Amin, Molly Gordon, Raphael Salles Scortegagna de Medeiros, Nelci Zanon, Raj Murali, Meena Jhanwar-Uniyal. Histological subtypes correlate with distinct genetic abnormalities in defining therapeutic targets in medulloblastoma [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 3118. doi:10.1158/1538-7445.AM2017-3118
Abstract 3120: Application of a lectin as a drug carrier for glycan-targeting cancer therapy
Authors: Oda Tatsuya , Shimomura Osamu , Tateno Hiroaki , Hirabayashi Jun , Noguchi Masayuki , Chiba Shigeru , Asashima Makoto , Ohkohchi Nobuhiro
Published: July 2017

Various cancers such as pancreatic ductal adenocarcinoma (PDAC) remain intractable despite recent advances in tumour-targeting antibody drugs. Although the outermost coating of every cell type is predominantly composed of cell-specific glycans, antibody strategies have hit walls of technical difficulties in intentional glycan targeting, and medico-economic burden. Lectins, proteins with glycan-binding potential, could be used as an alternative scaffold; however, the prejudice that all lectins mediate harmful haemagglutination limited its use only in experimental and diagnostic applications. Here, we show the successful in vivo application of a lectin as a drug carrier for cancer therapy. We have employed rBC2LC-N lectin since this has already shown a specific affinity between PDAC cell-surface glycans. When a lectin-drug conjugate (LecD) was developed by fusion with a bacterial exotoxin, the 50% inhibitory concentration (1.04 pg/ml=0.0195 pM) was 1000 times lower than that of conventional immunotoxins (on the order of ng/ml). In addition, we have revealed that rBC2LC-N lectin was safely administrable to mice without haemagglutination. Administration of this LecD in a PDAC mouse model of subcutaneous nodules and peritoneal dissemination, generated prominent therapeutic effects. The utilization of lectins as an in vivo drug-carrier targeting cancer glycan shows realistic potential when paired with advanced lectin engineering technologies, and their applications could be expanded by coupling with small molecule and/or nanoparticle drugs.

Citation Format: Tatsuya Oda, Osamu Shimomura, Hiroaki Tateno, Jun Hirabayashi, Masayuki Noguchi, Shigeru Chiba, Makoto Asashima, Nobuhiro Ohkohchi. Application of a lectin as a drug carrier for glycan-targeting 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 3120. doi:10.1158/1538-7445.AM2017-3120
Abstract 3128: TRPM7 regulates glioma cell proliferation and migration through different function domains
Authors: Liu Mingli
Published: July 2017

Background: Transient Receptor Potential Melastatin-related 7 (TRPM7) is a non-selective cation channel fused with a functional kinase domain. TRPM7 channel expression is aberrant in numerous cancers. We have previously reported that suppression of TRPM7 inhibits the proliferation, migration, and invasion of malignant human glioma cells using A172 glioma cell line and brain tissue of human glioma [CNS Neuroscience & Therapeutics 2015,21(3):252-61]. We further found that TRPM7 channels regulate glioma stem cell growth/proliferation through STAT3 and Notch signaling pathways [Cellular Signaling, 2014; 26: 2773-2781]. Since TRPM7 is a bifunctional protein with channel and α-kinase domain, we next investigated whether the changes in glioma cancer cell proliferation and migration might be caused through channel domain-mediated and/or kinase domain mediated TRPM7 activation.

Methods: 1) A172 cells expressing wild-type human TRPM7 (wtTRPM7) or constructs in which the α-kinase domain was deleted (Δkinase) or rendered inactive with a point mutation in the ATP binding site of the α-kinase domain (K1648R) were studied. 2) The effects of TRPM7 on glioma cell proliferation and invasion were determined using MTT assay and transwell invasion assay by TRPM7 expression using wtTRPM7, Δkinase, and K1648R, respectively.

Results: 1) Treatment of A172 cells with Δkinase and K1648R significantly increased TRPM7 expression demonstrated by Western blot assay. 2) The cell invasion was significantly reduced when treated with Δkinase and K1648R which indicate that TRPM7 kinase activity is required for changes in cell migration and invasion. 3) The cell proliferation was not significantly changed when treated with Δkinase and K1648R which indicate that TRPM7 channel rather than kinase activity is required in cell growth of A172 cells.

Conclusion: TRPM7 regulates glioma cell proliferation and migration/invasion through different function domains. The channel activity of TRPM7 is required for the cell growth while the kinase domain is required for the cell migration/invasion. The finding is important for designing and developing drugs targeting TRPM7 in human malignancies.

Note: This abstract was not presented at the meeting.

Citation Format: Mingli Liu. TRPM7 regulates glioma cell proliferation and migration through different function domains [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 3128. doi:10.1158/1538-7445.AM2017-3128
Abstract 3141: Combined cetuximab and pemetrexed therapy enhances cytotoxicity against crizotinib-resistant non-small cell lung cancer cells by downregulating thymidylate synthase
Authors: Kim Hwang-Phill , Han Sae-Won , Song Sang-Hyun , Kim Tae-You
Published: July 2017

Although non-small cell lung cancer (NSCLC) cells with anaplastic lymphoma kinase (ALK)-rearranged initially show a dramatic response to crizotinib, these cells eventually develop resistance to crizotinib. This resistance is caused by secondary mutations and copy number gain in the ALK gene. However, other resistance mechanisms through activation of the bypass tracts have yet to be clearly elucidated. To investigate the mechanisms of acquired resistance to ALK fusion-directed treatment in NSCLC, we generated crizotinib-resistant NSCLC cell lines in vitro through chronic exposure of an ALK fusion NSCLC line (SNU2292) to crizotinib. Interestingly, the resultant SNU2292-CR cells maintained activation of epidermal growth factor receptor (EGFR) and expressed increased levels of transforming growth factor alpha (TGF-α), an EGFR ligand. Additionally, we found that thymidylate synthase (TS) was overexpressed in SNU2292-CR cells compared with the parental cells. We showed reduction of TS enhanced synergistic inhibition of cell proliferation when cetuximab, an EGFR inhibitor, was combined with pemetrexed, a TS inhibitor. As a result, combined therapy was found to exhibit a synergistic growth inhibitory effect against crizotinib-resistant cells by downregulating TS expression. Taken together, these results support a potential role of activation of the bypass tracts in acquired resistance to ALK-directed treatment in ALK-rearranged NSCLC, and provide insights into strategies for preventing and/or overcoming this resistance in patients

Citation Format: Hwang-Phill Kim, Sae-Won Han, Sang-Hyun Song, Tae-You Kim. Combined cetuximab and pemetrexed therapy enhances cytotoxicity against crizotinib-resistant non-small cell lung cancer cells by downregulating thymidylate synthase [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 3141. doi:10.1158/1538-7445.AM2017-3141
Abstract 3145: Mechanisms of resistance to FGFR-targeted therapy in bladder cancer
Authors: Pettitt Geoffrey A. , McPherson Helen R. , Hurst Carolyn D. , Burns Julie E. , Alder Olivia A. , Dunning Matthew C. , Knowles Margaret A.
Published: July 2017

Fibroblast growth factor receptor 3 (FGFR3) overexpression, point mutations or gene fusions are found in ~80% of non muscle-invasive and ~15% of muscle-invasive bladder cancer. FGFR inhibitors have entered clinical trials in advanced bladder cancer however, as with other targeted therapies, intrinsic and acquired resistance are expected to limit treatment efficacy. We have used an in vitro model to explore possible mechanisms of resistance.

The urothelial cancer cell line RT112 expresses an FGFR3-TACC3 fusion protein and is sensitive to FGFR inhibition. Isogenic resistant cell lines, termed R1, R2 and R3, were derived by long-term culture of RT112 in the presence of the FGFR inhibitor PD173074. Compared to parental RT112, R1 and R2 show reduced proliferation and have a more mesenchymal morphology, decreased expression of FGFR3 and increased expression of N-cadherin. R3 has a faster growth rate, more epithelial morphology and lower N-cadherin expression than R1 and R2.

The changes in morphology and gene expression between parental and resistant derivatives R1 and R2 were reversed when the resistant cells were cultured without PD173074 for 4 passages. Despite this, the cells retained their resistance when re-exposed to PD173074.

Exome sequencing, RNA microarray analysis and phospho-receptor tyrosine kinase array data on the parental cells and resistant derivatives and will be presented. Our data suggests that diverse mechanisms of resistance occur following prolonged FGFR inhibition.

Citation Format: Geoffrey A. Pettitt, Helen R. McPherson, Carolyn D. Hurst, Julie E. Burns, Olivia A. Alder, Matthew C. Dunning, Margaret A. Knowles. Mechanisms of resistance to FGFR-targeted therapy in bladder 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 3145. doi:10.1158/1538-7445.AM2017-3145
Abstract 3169: SerpinB2 enhances invadopodia-like structure protrusions and is down-regulated in acquired gefitinib-resistant non-small cell lung cancer cells
Authors: Bae Song Yi , Kim Donghwa , Park Hyen Joo , Byun Woong Sub , Hong Ji-Young , Lee Hye-Jung , Lee Sang Kook
Published: July 2017

Non-small cell lung cancer (NSCLC) is a major type of lung cancer which accounts for approximately 80-85% of all lung cancers. The targeted therapies have significantly improved the survival of advanced NSCLC patients, but the failure of targeted therapy due to the resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), such as gefitinib, is now considered a major problem. SerpinB2 is a component of the urokinase plasminogen activator (uPA) system and has been recognized as a biomarker for the progression and metastasis of lung cancer. We found that SerpinB2 is down-regulated in gefitinib-resistant (H292-Gef) cells compared to gefitinib-sensitive (H292) cells. The low SerpinB2 levels in H292-Gef cells were also associated with an enhancement in invasiveness and increase in the length of invadopodia-like structures in the cells. The effect on invasiveness and gefitinib sensitivity was confirmed by knockdown and overexpression of SerpinB2. In addition, an antitumor agent yuanhuadine (YD) was used to test the possibility to overcome the resistance through the up-regulation of SerpinB2. YD effectively elevated SerpinB2 levels and suppressed invasive properties in H292-Gef cells. Collectively, these findings demonstrate the prospective role of SerpinB2 as a novel biomarker for acquired gefitinib resistance and a potential target for NSCLC treatment.

Acknowledgement

This study was funded by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2015R1D1A1A02062012).

Citation Format: Song Yi Bae, Donghwa Kim, Hyen Joo Park, Woong Sub Byun, Ji-Young Hong, Hye-Jung Lee, Sang Kook Lee. SerpinB2 enhances invadopodia-like structure protrusions and is down-regulated in acquired gefitinib-resistant non-small cell lung 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 3169. doi:10.1158/1538-7445.AM2017-3169
Abstract 3173: AKT1low quiescent cancer cells persist after neoadjuvant chemotherapy in triple-negative breast cancer patients
Authors: Kabraji Sheheryar , Sole Xavier , Huang Ying , Bango Clyde , Bowden Michaela , Bardia Aditya , Sgroi Dennis , Loda Massimo , Ramaswamy Sridhar
Published: July 2017

The mechanisms that allow triple negative breast cancer tumors to survive neoadjuvant chemotherapy are incompletely understood. Evidence suggests that proliferative heterogeneity may contribute to primary chemotherapy resistance in patients with triple negative breast cancer. AKT1low quiescent cancer cells (QCCs) are a quiescent, epigenetically plastic, and chemotherapy resistant subpopulation initially identified in experimental cancer models. Here, we identify QCCs in primary and metastatic human breast tumors using automated, quantitative, immunofluorescence microscopy coupled with computational and statistical analysis. We show that QCCs exist as non-random and heterogeneously distributed clusters within primary tumors. In addition, these QCC clusters persist after treatment with multi-agent, multi-cycle, neoadjuvant chemotherapy in both residual primary tumors as well as nodal and distant metastases in patients with triple negative breast cancer. Together, these data qualify QCCs as a non-genetic mechanism of chemotherapy resistance in triple negative breast cancer patients that warrants further study.

Citation Format: Sheheryar Kabraji, Xavier Sole, Ying Huang, Clyde Bango, Michaela Bowden, Aditya Bardia, Dennis Sgroi, Massimo Loda, Sridhar Ramaswamy. AKT1low quiescent cancer cells persist after neoadjuvant chemotherapy in triple-negative breast cancer patients [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 3173. doi:10.1158/1538-7445.AM2017-3173
Abstract 3176: Inhibition of HER2/β-catenin signaling by penfluridol overcomes resistance to paclitaxel in breast cancer
Authors: Gupta Nehal , Gupta Parul , Srivastava Sanjay
Published: July 2017

Paclitaxel is a first line treatment option for patients with metastatic breast cancer. However, inherited or acquired resistance is a limiting factor for therapy with paclitaxel. The mechanism of paclitaxel resistance remains obscured and hinders the development of therapeutic strategies. HER2 is an oncogene overexpressed in about 30% of breast cancer patients and plays role in drug resistance leading to poor prognosis. To identify more clinically relevant mechanism of paclitaxel resistance, we developed resistance to paclitaxel in MCF-7 and 4T1 breast cancer cell lines. The continuous exposure to paclitaxel for several months resulted in >1000 fold resistance in MCF-7 cells and >100 fold resistance in 4T1 cells. Western blot analysis showed enhanced expression of HER2, β-catenin and downstream molecules such as TCF/LEF, c-Myc, Cyclin D in these resistant cells. We have recently demonstrated that penfluridol, an anti-psychotic drug, suppresses the growth of triple negative metastatic breast cancer cells (Ranjan and Srivastava, Cancer Res 2016; 76(4): 877-890), giving us the rationale to evaluate whether penfluridol inhibits HER2 and β-catenin signaling. Our current results showed that penfluridol treatment not only suppressed HER2 but also inhibited β-catenin expression. We also observed down regulation of LEF-1/TCF, Cyclin D1 and c-Myc expression with penfluridol treatment in paclitaxel sensitive as well as resistant cells resulting in reduced survival of cells. Our results further showed that penfluridol treatment synergistically enhanced the growth suppressive effects of paclitaxel in MCF-7 and 4T1 paclitaxel resistant cells. Treatment of paclitaxel resistant 4T1 cells with 1.5μM of penfluridol in combination with 50nM of paclitaxel resulted in 65% of cell growth suppression whereas either treatment alone was not cytotoxic at all. We also observed an enhanced down regulation of proteins involved in paclitaxel resistance such as HER2, β-catenin, c-Myc and Cyclin D1 as well as increase in apoptotic markers such as Cl-PARP and Cl-Caspase3 when paclitaxel treatment was combined with penfluridol in resistant cells. Taken together, our results provided a novel insight into the mechanism of resistance to paclitaxel and also opened new avenues for application of penfluridol in cancer therapeutics. Further detailed mechanistic and in vivo studies are in progress. (Supported in part by RO1 grant CA129038, awarded by National Cancer Institute, NIH).

Citation Format: Nehal Gupta, Parul Gupta, Sanjay Srivastava. Inhibition of HER2/β-catenin signaling by penfluridol overcomes resistance to paclitaxel in 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 3176. doi:10.1158/1538-7445.AM2017-3176
Abstract 3177: KITENIN leads to resistance against temozolomide through enhancement of cancer stemness factors in mouse glioma model
Authors: Lee Kyung-Hwa , Ahn Eun-Jung , Jung Shin , Lee Jae-Hyuk , Kim Kyung-Keun , Moon Kyung-Sub
Published: July 2017

Background: Recently, a new research spotlighted the role of KITENIN (KAI1 COOH-terminal interacting tetraspanin) on glioma invasiveness and progression, associated with the up-regulation of EMT (epithelial-mesenchymal transition) and cancer stemness markers. In this study, furthermore, it is investigated whether KITENIN leads to resistance against TMZ through enhancement of cancer stemness factors using mouse glioma model.

Materials and Methods: TMZ-resistant T98G cell line (T98G/TR) was developed. EMT and cancer stemness markers and KITENIN expression were assessed by Western blotting and qRT-PCR. With KITENIN modulated U251 (knockdown) and GL261 (overexpression) cells, cell viability and apoptotic cell death factors after TMZ treatment were evaluated. Biological role of KITENIN on TMZ-resistance was investigated using magnetic resonance imaging (MRI) and immunofluorescence analysis for tumor sections of orthotopic glioma model. Using human glioma samples and primary cells, the mechanistic link between KITENIN and cancer stemness factors was confirmed.

Results: T98G/TR cells showed increased expression of the KITENIN and EMT and cancer stemness factors. Also, In vitro assays revealed that KITENIN knockdown inhibited cell viability against TMZ treatment, whereas KITENIN overexpression promoted their viability, via apoptotic cell death pathway. In orthotopic glioma models, mice implanted with KITENIN-overexpressed cells showed resistance to TMZ on MRI and histopathological examination. The expression of KITENIN/ALDH1 or KITENIN/CD44 were co-localized and significantly higher in tumor sections of mouse transplanted with KITENIN-overexpressed cells than in tumor sections of mouse transplanted with control cells. Genetic down-regulation of KITENIN for KITENIN-overexpressed cell line and primary glioma cells led decreased expression of ALDH1 and CD44. In human glioma samples, high expression of KITENIN was closely related with high expression of ALDH1.

Conclusion: KITENIN affects TMZ-resistance in malignant gliomas through induction of ALDH1 and CD44. Therefore, it could be suggested that KITENIN is therapeutic target to overcome TMZ-resistance of malignant gliomas.

Citation Format: Kyung-Hwa Lee, Eun-Jung Ahn, Shin Jung, Jae-Hyuk Lee, Kyung-Keun Kim, Kyung-Sub Moon. KITENIN leads to resistance against temozolomide through enhancement of cancer stemness factors in mouse glioma model [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 3177. doi:10.1158/1538-7445.AM2017-3177
Abstract 318: Identification of FERM domain-containing protein 5 (FRMD5) as a novel target of β-catenin/TCF7L2 complex
Authors: Furukawa Yoichi , Zhu Chi , Ohsugi Tomoyuki , Terakado Yumi , Noguchi Rei , Ikenoue Tsuneo , Yamaguchi Kiyoshi
Published: July 2017

Deregulated Wnt signaling is one of the earliest steps in colorectal tumorigenesis. The impairment results most frequently from APC mutations, which leads to the accumulation of β-catenin and subsequent activation of TCF7L2. Although previous studies have identified a number of target genes of the activated β-catenin/TCF7L2 transcriptional complex such as cMYC and CCND1, the comprehensive effect of the activation on tumorigenesis remains to be elucidated. To understand the precise molecular mechanisms underlying colorectal cancer, we searched for novel genes regulated by the complex in colorectal tumors. We performed expression profile analysis of HCT116 and SW480 colon cancer cells treated with β-catenin siRNAs, and combined these data with public microarray data of LS174 cells treated with a dominant-negative form of TCF7L2. As a result, we identified a total of 134 genes that were regulated by both β-catenin and TCF7L2. Subsequent ChIP-sequence with TCF7L2 antibody selected 11 genes from the 134 genes as candidates of direct target genes. In this study, we focused on FERM domain-containing protein 5 (FRMD5) among the 11 candidates. Quantitative PCR confirmed that its expression was reduced by β-catenin siRNA in HCT116 and SW480 cells. ChIP-quantitative PCR analysis with ant-TCF7L2 antibody also corroborated the interaction of TCF7L2 with a region (hg19; chr15:44,449,680-44,450,487) within intron 1 of FRMD5. Reporter assay with plasmids containing this region revealed that the reporter activity was down-regulated by the knockdown of β-catenin. These data suggested that FRMD5 is a direct target of β-catenin/TCF7L2 complex, and that the region is involved in the transcriptional activation through an interaction with the complex. Consistently, its expression was elevated in colorectal tumors compared to normal colonic mucosa in public microarray data. To uncover the role of FRMD5 in colorectal carcinogenesis, we analyzed expression profile of HCT116 cells treated with FRMD5 siRNA, and combined the data with profile with β-catenin siRNA. Gene set analysis with 53 commonly up-regulated and 36 commonly down-regulated genes by both FRMD5 and β-catenin disclosed that the gene sets of DNA replication, cell cycle, and extracellular matrix (ECM) were altered by FRMD5. These data may be useful for the future studies of colorectal carcinogenesis.

Citation Format: Yoichi Furukawa, Chi Zhu, Tomoyuki Ohsugi, Yumi Terakado, Rei Noguchi, Tsuneo Ikenoue, Kiyoshi Yamaguchi. Identification of FERM domain-containing protein 5 (FRMD5) as a novel target of β-catenin/TCF7L2 complex [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 318. doi:10.1158/1538-7445.AM2017-318
Abstract 3182: Cytoskeletal modulation results in increased tumor survival and drug resistance through attenuation of p53 dependent apoptosis
Authors: Wang Victoria E. , Doench John , Root David , Bernards Rene , Settleman Jeffrey , McCormick Frank
Published: July 2017

One of the major challenges to eradicating cancer involves the evolution of drug resistant clones and persistence of residual disease that escapes our current limit of detection but may proliferate upon cessation of therapy. These microscopic foci of residual diseases often exhibit stem-cell like properties and are intrinsically more resistant to drug therapy. A better understanding of the mechanisms underlying the innate drug resistance of these cellular populations may lead to improved treatment strategies, resulting in more durable remissions and ultimately improved patient survival.

We utilize functional genomic platforms to identify mechanisms responsible for the persistence of drug tolerant cells. A genome wide shRNA enrichment screen was performed using the c-Met addicted cell line GTL-16 to identify genes whose knockdown conferred DTP survival. Inhibition of the RhoA-ROCK-myosin pathway promotes drug resistance in a variety of tumor models, including those treated with either targeted therapy or conventional chemotherapy. Furthermore, pharmacological validation using multiple small molecule inhibitors of ROCK1 phenocopied both myosin heavy chain (MYH9) and light chain (MYL12) shRNA knockdown. More recently, utilizing a combination of gene expression and biochemical approaches, we have identified attenuation of p53 induced apoptosis to be a key event in mediating survival of these drug tolerant persisters. Reactivation of p53 using nutlin results in increased cell death.

These findings demonstrate that modulation of cytoskeleton is an important, but underappreciated, mechanism of drug resistance across many tumor types. These downstream effectors may serve as novel therapeutic targets for intervention and also biomarkers to stratify patients and their response to treatment.

Citation Format: Victoria E. Wang, John Doench, David Root, Rene Bernards, Jeffrey Settleman, Frank McCormick. Cytoskeletal modulation results in increased tumor survival and drug resistance through attenuation of p53 dependent apoptosis [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 3182. doi:10.1158/1538-7445.AM2017-3182
Abstract 3186: Taxane resistance in prostate cancer: A role for miRNA 181a
Authors: Armstrong Cameron M. , Liu Chengfei , Lou Wei , Gao Allen C.
Published: July 2017

Introduction: Docetaxel (DTX) is one of the primary drugs used for treating castration resistant prostate cancer (CRPC). Unfortunately, over time patients invariably develop resistance to DTX therapy and their disease will continue to progress. The mechanisms by which resistance develops are still incompletely understood. This study seeks to determine the involvement of miRNAs, specifically miR-181a, in DTX resistance in CRPC.

Methods: Total RNA from parental C4-2B prostate cancer cells and DTX resistant C4-2B cells (C4-2B TaxR) was submitted for small RNA deep sequencing. Data was analyzed to ascertain which miRNAs expressions were most altered in C4-2B TaxR cells compared to parental cells. Having identified an increase in miR-181a in resistant cells, its expression was modulated in C4-2B and C4-2B TaxR cells by transfecting them with miR-181a mimics or antisense, respectively. Following transfection, cell number was determined after 48 h with or without DTX. Cross resistance to cabazitaxel induced by miR-181a was also determined. Western blots were used to determine ABCB1 protein expression and rhodamine assays used to assess activity. Phospho-p53 expression was assessed by western blot and apoptosis was measured by ELISA in C4-2B TaxR cells with inhibited miR-181a expression with or without DTX.

Results: miR-181a is significantly upregulated in C4-2B TaxR cells compared to parental C4-2B cells as analyzed by small RNA sequencing. Overexpression of miR-181a in C4-2B cells confers DTX and cabazitaxel resistance. Knockdown of miR-181a in C4-2B TaxR cells re-sensitizes them to treatment with both DTX and cabazitaxel. miR-181a knockdown alone induced apoptosis in C4-2B TaxR cells which is further enhanced by DTX. We next assessed if miR-181a altered expression or activity of ABCB1, which is overexpressed/active in C4-2B TaxR cells and promotes resistance to DTX by pumping the drug out of cells. We found that miR-181a does not impact ABCB1 expression or activity. Since we previously demonstrated that phospho-p53 can modulate DTX sensitivity, we determined if miR-181a can alter p53 expression in C4-2B TaxR cells. Knockdown of miR-181a in C4-2B TaxR cells induced phospho-p53 expression, suggesting that miR-181a induced resistance to DTX is mediated by inhibition of phospho-p53 expression.

Conclusions: Overexpression of miR-181a in C4-2B TaxR cells contributes to their resistance to DTX and inhibition of miR-181a expression can restore treatment response. This is due, in part, to modulation of p53 phosphorylation and induction of apoptosis.

Note: This abstract was not presented at the meeting.

Citation Format: Cameron M. Armstrong, Chengfei Liu, Wei Lou, Allen C. Gao. Taxane resistance in prostate cancer: A role for miRNA 181a [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 3186. doi:10.1158/1538-7445.AM2017-3186
Abstract 3196: Genetic and pharmacologic approaches to overcome epithelial to mesenchymal mediated chemoresistance in breast cancer
Authors: Crowley Michael J. , Altorki Nasser , Mittal Vivek , Gao Dingcheng
Published: July 2017

Development of resistance to conventional chemotherapy remains a major barrier to effective treatment of breast cancer. We and others have recently demonstrated in breast and pancreatic cancer models that epithelial-mesenchymal transition (EMT) may not be a critical mediator of cancer metastasis, however, it contributes to cancer drug resistance (Fischer et al. Nature 2015; Zeng et al Nature 2015). Importantly, blocking EMT through miR-200 family abrogated chemoresistance, indicating potentials for clinical translation. Using our novel EMT lineage tracing system, we have performed both genetic (CRISPR/Cas9 mediated genome-wide targeted mutagenesis) and pharmacological (high-throughput small molecule libraries) screens, to identify potential candidates to overcome EMT-mediated chemoresistance in breast cancer metastasis. The identified molecules provide not only novel mechanistic insights but also attractive anti-metastatic strategies for breast cancer treatment and the design of future clinical trials.

Citation Format: Michael J. Crowley, Nasser Altorki, Vivek Mittal, Dingcheng Gao. Genetic and pharmacologic approaches to overcome epithelial to mesenchymal mediated chemoresistance in 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 3196. doi:10.1158/1538-7445.AM2017-3196
Abstract 3197: Splicing factors and the role of Navitoclax in drug-resistant ovarian cancer
Authors: Coley Helen M. , Lattanzio Laura , Garrone Ornella , Merlano Marco , Vivenza Daniela , Nigro Cristiana Lo , Syed Nelofer , Thompson Alistair , Rao Bhavya , Crook Tim
Published: July 2017

Alternative RNA splicing allows processing of a pre-mRNA into various mature mRNAs. In this way multiple protein products can be derived from a single gene. In cancer it is known that specific splice variants may facilitate malignant transformation and therapeutic resistance. Our current studies focus on the problem of drug resistance in ovarian cancer and we have approached this in a variety of ways. Following sequencing analysis of a panel of ovarian cancer cell lines we identified the RNA splicing factors TCERG and SRSF2 as being reduced in drug resistant ovarian cancer cell lines, compared with their drug sensitive counterpart. TCERG is important in regulating the splicing of variant forms of the BClx gene: BClxL (long form) and BClxs (short form). There are clear implications for drug resistance whereby low TCERG results in reduced BClx splicing to yield the pro-apoptotic BClxs form. In the SKOV-3 human ovarian cancer cell line panel both paclitaxel resistant SKOV-3TaxR and SKOV-3CR carboplatin resistant cells showed reduced levels of TCERG mRNA using qPCR analysis. In addition, qPCR analysis for BClxl and BClxs levels indicated increased ratio of long:short form with drug resistance: SKOV-3 parent line ratio 1.0; SKOV-3TaxR 1.8; SKOV-3CR 1.65. Western immunoblotting showed relatively decreased levels of Bax protein in drug resistant versus drug sensitive cell line in line with a reduced apoptotic reduced. We then explored the use of Navitoclax (ABT0263), the small-molecule Bcl-2 family protein inhibitor for Bcl-xL, Bcl-2 and Bcl-w in our cell line panel. Navitoclax in combination with paclitaxel for 48h gave rise to significantly increased apoptotic response in drug-resistant versus drug sensitive ovarian cancer cell line counterparts, using the Annexin V assay with PI, indicative of re-sensitisation to paclitaxel. Synergistic effects were also seen for the parental cell line but the effects were more marked for the resistant cell lines. Our future work will also involve the use of clinical biopsies of ovarian cancer cases.

Citation Format: Helen M. Coley, Laura Lattanzio, Ornella Garrone, Marco Merlano, Daniela Vivenza, Cristiana Lo Nigro, Nelofer Syed, Alistair Thompson, Bhavya Rao, Tim Crook. Splicing factors and the role of Navitoclax in drug-resistant ovarian 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 3197. doi:10.1158/1538-7445.AM2017-3197
Abstract 320: Potential of targeting of MYC-driven cancers via CIP2A without detrimental physiological effects
Authors: Qiao Xi
Published: July 2017

Transcription factor MYC is a driver of many human cancers due to its widespread effects on gene expression. An understanding of the mechanisms determining MYC’s transcriptional and proliferation-promoting activities in vivo could facilitate approaches for MYC targeting. However, post-translational mechanisms that control MYC function in vivo are poorly understood. Here, we demonstrate that MYC phosphorylation at serine 62 enhances MYC accumulation on Lamin A/C-associated nuclear structures and that the protein phosphatase 2A (PP2A) inhibitor protein CIP2A is required for this process. CIP2A is also critical for serum-induced MYC phosphorylation and for MYC-elicited proliferation induction in vitro. Complementary transgenic approaches and an intestinal regeneration model further demonstrated the in vivo importance of CIP2A and serine 62 phosphorylation for MYC activity upon DNA damage-induced proliferation. However, targeting of CIP2A did not influence the normal function of intestinal crypt cells or general well-being of the mice. Meanwhile, in breast cancer cells, CIP2A depletion was shown to inhibit the MYC serine 62 phosphorylation, MYC-mediated gene expression, and anchorage-independent growth. Furthermore, CIP2A supports MDA-MB-231 xenograft growth in nude mice. Therefore, results of these studies collectively suggest for a novel opportunity to target MYC’s function in MYC- driven cancers via CIP2A, and without detrimental physiological effects.

Citation Format: Xi Qiao. Potential of targeting of MYC-driven cancers via CIP2A without detrimental physiological effects [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 320. doi:10.1158/1538-7445.AM2017-320
Abstract 3206: A potent and selective CSF-1R inhibitor, DCR-0064, inhibits colony stimulating factor 1 signaling in vitro and in vivo
Authors: Liao Chu-Bin , Peng Shao-Zheng , Ho Chen-Hsuan , Lee Chao-Pin , Chang Jia-Ming , Huang Hung-Jyun , Ciou Sian-Yi , Pan Yu-Chih , Chen Yu-Kai
Published: July 2017

Tumor-associated macrophages (TAMs) are major components of leukocytic infiltrate of tumors. It has been reported that high TAMs density is associated with poor clinical prognosis. Within the tumor microenvironment, TAMs rely on signaling through CSF-1/CSF-1 Receptor kinase axis to promote tumor growth, angiogenesis, and metastasis. Therefore, decrease of TAMs density in tumors through inhibition of CSF-1R kinase is considered a potential target for drug development. In this study, we report that a novel small molecule DCR-0064 inhibits CSF-1R kinase with IC50 value below 10 nM and possesses specificity for CSF-1R over other tyrosine or serine/threonine kinases. DCR-0064 selectively inhibits CSF-1 induced growth of M-NFS-60 myeloid cells and effectively suppresses intracellular CSF-1R activation with IC50 value less than 100 nM. The potency and selectivity of DCR-0064 is superior to the current lead CSF-1R inhibitor, PLX3397, which is in phase III clinical development for treating tenosynovial giant cell tumor. The selectively growth inhibitory activity of DCR-0064 is consistent with inhibition of CSF-1R signaling. Oral administration of DCR-0064 reduces TAMs density in vivo in mice. DCR-0064 is being developed as a candidate for preclinical development.

Citation Format: Chu-Bin Liao, Shao-Zheng Peng, Chen-Hsuan Ho, Chao-Pin Lee, Jia-Ming Chang, Hung-Jyun Huang, Sian-Yi Ciou, Yu-Chih Pan, Yu-Kai Chen. A potent and selective CSF-1R inhibitor, DCR-0064, inhibits colony stimulating factor 1 signaling in vitro and in vivo [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 3206. doi:10.1158/1538-7445.AM2017-3206