bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1731: Analysis of prostate cancer circulating tumor cells at the mRNA and protein level using a microfluidic graphene oxide chip
Authors: Kozminsky Molly , Day Kathleen C. , Hiles Guadalupe Lorenzatti , Fouladdel Shamileh , Azizi Ebrahim , Day Mark L. , Morgan Todd , Nagrath Sunitha
Published: July 2017

While the relative five-year survival rate for men with localized prostate cancer is nearly 100%, it plummets to 28% in the case of distant metastases. This underscores the importance of investigating those cells able to spread and form these metastases: circulating tumor cells (CTCs). Though rare and surrounded by other blood cells, CTCs can be isolated using the sensitive nanomaterial-based microfluidic technology the graphene oxide (GO) Chip. Whole blood samples from 54 prostate cancer patients were obtained with consent and processed on parallel chips for immunofluorescence and RNA analysis. All of the samples were analyzed for CTC enumeration, where captured cells were stained for cytokeratin 7/8 (CK), CD45, and DAPI. CTCs were identified as DAPI+/CK+/CD45- cells. CTCs were detected in all of the samples. Interestingly, 25 samples showed CTC clusters ranging from 2-10 CTCs per cluster. To explore the potential role of EGFR in metastatic progression and CTC survival, a subset of samples was also assayed for EGFR expression with 9/10 samples showing EGFR+ CTCs. RNA extracted from a parallel device was reverse transcribed to cDNA. The cDNA was analyzed via qPCR for expression levels of 96 genes of interest, including housekeeping genes; epithelial and mesenchymal genes; oncogenes and tumor suppressor genes; prostate specific genes; extracellular matrix and inflammatory genes; and others. The results show our ability to examine RNA from CTCs to examine the role of relevant pathways in the cells in transit. With analysis capabilities at both the RNA and protein levels, the GO Chip is an example of a clinically relevant microfluidic technology.

Citation Format: Molly Kozminsky, Kathleen C. Day, Guadalupe Lorenzatti Hiles, Shamileh Fouladdel, Ebrahim Azizi, Mark L. Day, Todd Morgan, Sunitha Nagrath. Analysis of prostate cancer circulating tumor cells at the mRNA and protein level using a microfluidic graphene oxide chip [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 1731. doi:10.1158/1538-7445.AM2017-1731
Abstract 1739: Analysis of EML4-ALK fusion transcripts in plasma and platelets to monitor response to crizotinib in EML4-ALK positive non-small cell lung cancer patients (NSCLC)
Authors: Aguado Cristina , Teixido Cristina , Gimenez-Capitan Ana , Gil Maria de los Llanos , Rodriguez Sonia , Viteri Santiago , Karachaliou Niki , Aldeguer Erika , Peg Vicente , Alonso Lidia , Molina-Vila Miguel Angel , Rosell Rafael
Published: July 2017

Background: Rearrangements in anaplastic lymphoma kinase (ALK) gene can be detected in 5-7% of EGFR and KRAS wild-type advanced NSCLC patients (p). Fluorescent in situ hybridization (FISH) and immunohistochemistry (IHC) are currently used for screening but are unable to identify the specific fusion partner and are unpractical to monitor clinical responses due to difficulty of obtaining rebiopsies. The RT-PCR technique has the potential to overcome this pitfall and allow patient monitorization in blood.

Methods: A total of 405 formalin-fixed paraffin-embedded (FFPE) samples from advanced NSCLC were analyzed by ALK IHC (Ventana D5F3) and FISH (Vysis). Positive patients were confirmed by RT-PCR and submitted to Sanger in order to identify the variant. In a subset of 36 patients with EML4-ALK-rearranged tumors who were treated with crizotinib, fusion transcripts were analyzed by RT-PCR in mRNA purified from plasma and platelets and correlated with clinical response.

Results: ALK IHC was analyzed in 405 NSCLC patients and 37 tested positive (9.1%) whereas 25 (7.7%) were identify as translocated by FISH (n=323). ALK fusion transcripts were analyzed by RT-PCR and a new fusion variant of ALK was identified. A total of 36 p benefited from crizotinib treatment, including the p with the new variant. Monitoring of EML4-ALK fusion transcripts in the plasma ad platelets of 35 ALK positive patients revealed a good correlation with clinical outcome to crizotinib treatment, with the fusion transcripts becoming undetectable in p with good clinical responses.

Conclusions: Analysis of ALK fusion transcripts in mRNA purified from plasma and platelets can have a value in patients with no biopsy available and to monitor the course of the disease.

Citation Format: Cristina Aguado, Cristina Teixido, Ana Gimenez-Capitan, Maria de los Llanos Gil, Sonia Rodriguez, Santiago Viteri, Niki Karachaliou, Erika Aldeguer, Vicente Peg, Lidia Alonso, Miguel Angel Molina-Vila, Rafael Rosell. Analysis of EML4-ALK fusion transcripts in plasma and platelets to monitor response to crizotinib in EML4-ALK positive non-small cell lung cancer patients (NSCLC) [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 1739. doi:10.1158/1538-7445.AM2017-1739
Abstract 176: Early development of GMC1, a novel molecule targeting FKBP52 for the treatment of hormone-refractory prostate cancer
Authors: Xie Huan , Ekpenyong Oscar
Published: July 2017

Purpose: GMC1 directly inhibits FKBP52, effectively blocking androgen receptor dependent gene expression and androgen-stimulated proliferation. This make it an attractive option for the treatment of hormone-dependent and hormone-independent prostate cancer. This study investigated an analytical method for GMC1 quantification, pre-formulation characteristics of GMC1, and developed intravenous formulations for the evaluation of GMC1 in animal models.

Method: An LC/MS/MS method for the quantification of GMC1 in solution, plasma and urine was developed, validated and applied to the determination of the stability, log P, plasma protein binding and solubility of GMC1 in various solvents. Liposomal formulations and co-solvent systems with various ratios of high capacity vehicles were formulated and the optimal formulation applied, at 2 mg/kg single IV bolus dose, to the pharmacokinetic study of GMC1 in a rat model.

Result: The intra- and inter-day accuracy (%RE) and precision (%CV) of the LC/MS/MS method ranged from 1.6 – 11.7 % and 1.4 – 8.8 %, respectively. GMC1 is stable in solid and solution state, moderately lipophilic (log P = 1.38 ± 0.05), poorly water soluble (0.4 ± 0.01 mg/mL), and highly plasma protein bound (>71%). The optimal formulation consisting of PEG 300 and Labrasol ® (1:1, v/v) allowed us to achieve a GMC1 concentration of 10 mg/mL, and tolerated an aqueous environment. GMC1 has a tri-exponential disposition with a Cmax of 7.6 ± 1.97 mg/L, clearance of 0.53 L/kg/hr, α-distribution, β-phase and terminal elimination half-lives of 0.1 ± 0.04 hr, 1.2 ± 0.34 hr, and 19.7 ± 5.09 hr respectively.

Conclusion: The LC/MS/MS method, formulations and pharmacokinetic study can be applied to the pre-clinical and clinical development of GMC1.

Grant Support: This work was performed under funding from NIH/NIGMS grant (5SC3GM102018) and NIH/NIMHD/RCMI grant (5G12MD007605).

Citation Format: Huan Xie, Oscar Ekpenyong. Early development of GMC1, a novel molecule targeting FKBP52 for the treatment of hormone-refractory prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 176. doi:10.1158/1538-7445.AM2017-176
Abstract 1765: Molecular and immunological characterization of non-small cell lung cancer harboring cMET alterations
Authors: Saigi Maria , Pereira Carolina , Pros Eva , Brambilla Elisabeth , Nadal Ernest , Sánchez-Céspedes Montse
Published: July 2017

Purpose: To explore the correlation between the expression of immunological biomarkers in a subset of NSCLC tumors presenting cMET activation.

Methods: We retrospectively reviewed 200 tumor samples from a cohort of patients diagnosed with NSCLC in 3 different hospitals, from Spain and France. cMET activation was defined by the presence of one or more of the following criteria: 1- cMET mutations inducing exon 14 skipping (METex14), 2- cMET amplification, assessed by FISH 3- cMET overexpression, assessed by immunohistochemistry (IHC). METex14 mutations were determined by genomic sequencing and RT-PCR. FISH was interpreted according to Capuzzo scoring system, considering 5 or more gene copies as the mean value for positive amplification. Concurrent genetic alterations in EGFR and ERBB2 tyrosine kinase receptors were also determined. PD-L1 expression and density of tumoral infiltrating CD8+ lymphocytes were assessed by IHC. Statistical correlation analysis was performed by Chi-square and Fisher’s exact test.

Results: METex14, cMET amplification and cMET overexpression were detected in 2%, 7.8% and 31% of tumors, respectively. Histological subtypes included: 70% adenocarcinoma, 20% squamous cell carcinoma, 8% large cell, 2% pulmonary sarcomatoid. Curiously, one of the METex14 adenocarcinoma was concurrent with ERBB2 amplification. Those tumors harbouring METex14 or cMET amplification were correlated with positivity for PD-L1 expression (p = 0.02). However, this association was not observed in cMET overexpressed tumors.

Conclusions: We observed a direct correlation between PD-L1 positivity and METex14 or cMET amplification. Functional analysis and a large cohort of tumors are required to better characterize the potential role of cMET pathway in the regulation of PD-L1 expression.

Citation Format: Maria Saigi, Carolina Pereira, Eva Pros, Elisabeth Brambilla, Ernest Nadal, Montse Sánchez-Céspedes. Molecular and immunological characterization of non-small cell lung cancer harboring cMET alterations [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 1765. doi:10.1158/1538-7445.AM2017-1765
Abstract 1768: A practical tool for probing protein glycosylation in clinical samples: Application to characterizing subtypes of MUC5AC glycosylation in distinct subsets of pancreatic cancer patients
Authors: Hsueh Peter Y.
Published: July 2017

Glycans are critical to protein biology and are useful as disease biomarkers. Many studies of glycans rely on clinical specimens, but the low amount of sample available for some specimens limits the experimental options. We have a method to obtain information about protein glycosylation using a minimal amount of protein, and we applied it to the study of biomarkers in the blood plasma of pancreatic cancer patients. The method works by treating proteins that were captured by antibody microarrays (2.2 x 2.2 mm) with exoglycosidases to successively expose underlying features, and then probing the native or exposed features using a panel of lectins or glycan-binding antibodies. We use an algorithm to interpret the data and provide predictions about the glycan motifs that are present in the sample. We applied the method to the analysis of the glycosylation of a biomarker, MUC5AC, found in the plasma of patients with pancreatic cancer. An O-linked glycan on MUC5AC that is elevated in the plasma of about 75% of patient had mostly 2,3 linked sialic acid on type-1 N-acetyl-lactosamine (LacNAc), whereas other glycans on MUC5AC had a 2,6 linkage of sialic acid on type-2 LacNAc. Another glycan, fucosylated and sialylated/sulfated type-2 LacNAc, which is elevated in distinct subset of patients, was on N-linked glycans, representing an unusual finding of N-linked glycans on mucins associated with cancer. Further developments are enabling the testing for consistent structural features associating with subtypes of cancer, which could improve our ability to detect and target the subtypes. The on-chip glycan modification and probing (on-chip GMAP) method provides a platform for analyzing protein glycosylation in clinical specimens and could facilitate the characterization and discovery of glycan biomarkers.

Citation Format: Peter Y. Hsueh. A practical tool for probing protein glycosylation in clinical samples: Application to characterizing subtypes of MUC5AC glycosylation in distinct subsets of pancreatic 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 1768. doi:10.1158/1538-7445.AM2017-1768
Abstract 1792: Immune system pathway activation for prediction of the response to neoadjuvant chemotherapy in triple-negative breast cancer
Authors: Sawada Takeshi , Hilhorst Riet , Rangarajan Savithri , Yoshida Masayuki , Tamura Kenji , Beuningen Rinie van , Ruijtenbeek Rob , Tsuda Hitoshi , Koizumi Fumiaki
Published: July 2017

Background: Triple-negative breast cancer patients (TNBCs) who receive neoadjuvant chemotherapy (NAC) generally show a response to NAC, but about 5% experience progressive disease (PD). Although TNBCs that respond to NAC have been well-studied, reports about TNBCs with PD during NAC are absent. We aimed to compare kinase activity profiles of TNBCs of both groups to identify a biological predictor of NAC non-responders.

Methods: Tyrosine kinase activity profiles of lysates of fresh frozen cancer tissues from NAC non-responders and responders, the latter represented by TNBCs who did not receive NAC, were determined with a peptide microarray system (Cancer Res. 2009; 69(14):5987-95). Tumor infiltrating lymphocytes (TILs) were counted in these tissues.

Results: Among 740 TNBCs, 20 NAC non-responders were identified. For the 7 samples from PD confirmed patients and 10 TNBCs not receiving NAC, no correlation was observed between NAC response and age, menopausal status, tumor size or axillary lymph node status. Kinase activity measurements showed large differences in overall tyrosine kinase activity between responders and non-responders. 35 peptides had significantly (p < 0.05) higher phosphorylation in the responder group. Kinases that phosphorylate these peptides (upstream kinases) were identified from literature. Among others, the immune related kinases Zap70, Syk and JAK2 were identified as upstream kinases. Pathway analysis yielded evidence for increased activity in the responder group in immune-related pathways and in pathways involved in development and signal transduction. Responders had a significantly higher (p = 0.0053) TILs count.

Conclusion: Kinases related to the immune system are less activated in NAC non-responders. The low TILs count in the non-responders confirmed that activation of the immune system in non-responders is lower than in NAC responders. These data merit a further investigation of the use of kinase activity profiling either alone or in combination with TIL scoring for the prediction of the response to NAC in TNBC with a larger patient cohort.

Citation Format: Takeshi Sawada, Riet Hilhorst, Savithri Rangarajan, Masayuki Yoshida, Kenji Tamura, Rinie van Beuningen, Rob Ruijtenbeek, Hitoshi Tsuda, Fumiaki Koizumi. Immune system pathway activation for prediction of the response to neoadjuvant chemotherapy in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1792. doi:10.1158/1538-7445.AM2017-1792
Abstract 1799: CRM1 expression in pancreatic adenocarcinoma correlates with survivin expression and the proliferative activity
Authors: Saulino David M. , Younes Pamela S. , Bailey Jennifer M. , Younes Mamoun
Published: July 2017

Introduction: CRM1 is a nuclear export chaperone that mediates the export of proteins essential to growth regulation and tumor suppression. Its overexpression in tumors was found to be associated with poor prognosis. Selective inhibitors of nuclear export are in phase I and II clinical trials for several tumor types. The expression of CRM1 in human pancreatic adenocarcinoma (PAC) and its relation to survivin expression and tumor proliferative activity is largely unknown.

Experimental procedures: Sections of tissue microarray containing 77 formalin fixed and paraffin embedded PAC were stained by immunohistochemistry (IHC) for CRM1, survivin, and Cyclin A. Expression levels of CRM1 and survivin in tumor cells was determined using a quantitative digital image analysis solution (OTMIAS). The tumor proliferative activity was determined by measuring the S-phase fraction (SPF) in sections stained for Cyclin A, also using OTMIAS.

Summary: Sixty-six of the 77 (86%) PAC showed at least some positive staining for CRM1, and 11 (14%) were completely negative. The mean CRM1 expression levels ranged from 0.3 to 53 units and the median from 0.3 to 45 units. There was significant positive correlation between the mean and median expression levels of CRM1 in tumor cells with the mean and median levels of survivin (p<0.001). Moreover, there was positive correlation between the mean and median CRM1 levels in tumor cells the SPF (p=0.005).

Conclusions: CRM1 is expressed in a significant proportion of PAC, and increased CRM1 expression correlates with increased survivin expression and increased proliferative activity, suggesting that selective inhibitors of nuclear export may be effective against PAC.

Note: This abstract was not presented at the meeting.

Citation Format: David M. Saulino, Pamela S. Younes, Jennifer M. Bailey, Mamoun Younes. CRM1 expression in pancreatic adenocarcinoma correlates with survivin expression and the proliferative activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1799. doi:10.1158/1538-7445.AM2017-1799
Abstract 181: Discovery of novel targeted therapeutics for metastatic breast cancer
Authors: Valle Cristina Del , Hernández Eliud , Vlaar Cornelis P. , Cubano Luis A. , Dharmawardhane Suranganie , Castillo-Pichardo Linette
Published: July 2017

Metastatic disease is the primary cause of breast cancer mortality, due to the lack of effective therapy. The Rho GTPase Rac is integral for the promotion of cancer cell migration/invasion, proliferation, and survival. Since metastatic breast cancers often overexpress or exhibit high Rac activity, inhibition of Rac is a viable strategy against metastatic cancer. Recently, we characterized EHop-016, a small molecule that inhibits Rac activity of metastatic breast cancer cells with an IC50 of 1 µM. EHop-016 is 10-100 times more active than previously available Rac inhibitors, and is the first compound shown to inhibit the activation of Rac by the oncogenic GEF Vav. EHop-016 inhibits the activity of the Rac downstream effector p21 activated kinase (PAK), lamellipodia extension, and cell migration of metastatic breast cancer cells. We also reported that EHop-016 at ≥ 25 mg/kg Body Weight (BW) significantly reduced tumor growth, metastasis, and angiogenesis in a mouse model. However, our recent pharmacokinetic study of EHop-016 in a mouse model demonstrated that the bioavailability of Ehop-016 needs to be improved for further pharmacological development. Therefore our hypothesis is that improvement of the EHop-016 structure will provide probes with increased potency against Rac and, therefore, increased bioavailability. Herein we have tested several Ehop-016 derivatives for their effects on breast cancer cell viability and Rac activation. Using MTT assays we found that the Ehop-016 derivatives, HV-107 and HV-118, significantly inhibit the viability of metastatic breast cancer cell lines MDA-MB-231 and MDA-MB-435. The effects of HV-107 and HV-118 on the inhibition of Rac activation were tested by ELISA-based Rac activity assays and pulldown assays. Results show that at 250nM, HV-107 inhibits Rac activation by 55%, whereas HV-118 has a similar effect at 100nM in MDA-MB-231 and MDA-MB-435 cells. Taken together, our findings suggest HV-107 and HV-118 as promising Ehop-016 derivatives with potential as anti-metastatic agents, which should be further characterized.

This study was supported by awards from the Susan Komen for the Cure, NIH/NIMHHD U54MD008149, and the Puerto Rico Science and Technology Trust to SD; NIH/NCRR R25GM061838 to UPR MSC; NIH/NIMHHD RCMI 8G12MD007583RCMI, Title V PPOHA 031M10505 and Title V Cooperative P031S130068 from U.S. Department of Education to UCC; and PRINBRE (NIH/NIGMS P20GM103475-13) Pilot Project to LCP.

Citation Format: Cristina Del Valle, Eliud Hernández, Cornelis P. Vlaar, Luis A. Cubano, Suranganie Dharmawardhane, Linette Castillo-Pichardo. Discovery of novel targeted therapeutics for metastatic 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 181. doi:10.1158/1538-7445.AM2017-181
Abstract 1816: Endogenous ADAMTS-13 regulates angiogenesis in cultured human endothelial cells
Authors: Tang Huiyuan , Lee Manfai , Kim Eun Ho , Bishop Daniel , Rodgers George M.
Published: July 2017

ADAMTS-13, a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13, is a zinc-containing metalloprotease that cleaves von Willebrand factor (vWf). Previous publications by our laboratory have shown that ADAMTS-13 may also be involved in angiogenesis; for example; addition of exogenous ADAMTS-13 enhances endothelial cell (Lee et al. Microvascular Res 2012; 84:109) and glioblastoma cell (Lee et al. Blood 2013; 122:#2306) migration and proliferation. For this study, we report the successful transient knockdown of endogenous ADAMTS-13 in human umbilical vein endothelial cells (HUVEC) via siRNA. 30pmols of ADAMTS-13 siRNA in a six-well plate inhibited HUVEC ADAMTS-13 expression levels by 90% after 24hr incubation, whereas control siRNA did not affect endogenous ADAMTS-13 levels. The knockdown effect was decreased over time: 72hr after the initial knockdown, ADAMTS-13 expression was reduced by only 50%. The effects of reduced endogenous ADAMTS-13 on HUVEC angiogenesis functions were studied. Transfection of HUVEC with 10pmols of ADAMTS-13 siRNA in a 24-well plate resulted in a 21% and 22% decrease in proliferation after 24hr and 48hr incubation, respectively. The effects of ADAMTS-13 knockdown on migration of HUVEC across a scratch wound were also evaluated. 24hr after transfection with control siRNA, there was robust cell migration across the scratch wound. This dramatic migration did not occur with ADAMTS-13 knockdown cells. Decreased expression of endogenous ADAMTS-13 also affected angiogenesis as measured by endothelial cell tube formation using a Matrigel matrix method. The tube lengths, sizes and junction numbers of the ADAMTS-13 knockdown cells were all significantly lower compared to control cells by about 40%. Activity of the AKT pathway, one of the angiogenesis downstream signaling pathways was down-regulated by ADAMTS-13 siRNA while ERK, a component of the MAP kinase pathway was not affected upon knockdown of ADAMTS-13. These data indicate that in cultured endothelial cells, one role of endogenous ADAMTS-13 is regulation of angiogenesis, mediated through the AKT signaling pathway. Overall, our data suggest an additional model of endogenous ADAMTS-13 functionality, beyond that of cleaving von Willebrand factor.

Citation Format: Huiyuan Tang, Manfai Lee, Eun Ho Kim, Daniel Bishop, George M. Rodgers. Endogenous ADAMTS-13 regulates angiogenesis in cultured human endothelial 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 1816. doi:10.1158/1538-7445.AM2017-1816
Abstract 1817: Resistin induces angiogenesis and lymphangiogenesis in human chondrosarcoma
Authors: Chi Meng-Ju , Lin Chih-Yang , Tang Chih-Hsin
Published: July 2017

Chondrosarcoma is a common kind of bone cancers, and it may develop distant metastasis, followed by a significant decline in overall survival. However, there are still no specific therapeutic methods for it today. For tumors to metastasis, angiogenesis and lymphangiogenesis are both important in the early processes. Therefore, inhibiting the development of angiogenesis and lymphangiogenesis could be a method to decline tumor metastasis. Resistin was discovered as an adipocyte-secreting adipokine, which may play a critical role in modulating cancer pathogenesis. In our lab, we previously found that resistin appears to increase MMP-2 expression and then promotes metastasis in human chondrosarcoma cells. Nevertheless, the role of resistin in angiogenesis and lymphangiogenesis of human chondrosarcoma is still unknown. To examine angiogenetic and lymphangiogenetic effects of resistin, we used human endothelial progenitor cells (EPCs) and lymphatic endothelial cells (LECs) to mimic capillary and lymphatic vessels formation. The results indicated that resistin-treated chondrosarcoma cell lines promoted EPCs VEGF-A-dependent as well as LECs VEGF-C-dependent tube formation and cell migration. Then we confirmed that treating cells with resistin increased VEGF-A and VEGF-C expression in human chondrosarcoma cell lines. Moreover, we found resistin-induced VEGF-A and VEGF-C expressions are mediated by PI3K/AKT signaling and by the activation of c-Src separately. In addition, resistin decreased the expression of miR-16-5p via PI3K/AKT pathway, and so of miR-186 via c-Src. We also demonstrated that miR-186 directly targeted on VEGF-C 3’ untranslated region, and regulated the VEGF-C production. Besides, we found the expressions of resistin, VEGF-A and VEGF-C was higher in human chondrosarcoma biopsy tissues than those in normal cartilage. Taken together, resistin not only promotes human chondrosarcoma angiogenesis through the activation of PI3K/AKT signaling pathway and down-regulating miR-16-5p expression, but also promotes human chondrosarcoma lymphangiogenesis through the activation of c-Src and down-regulating miR-186. Consequently, resistin may represent a potential novel molecular therapeutic target for human chondrosarcoma therapeutic treatment.

Citation Format: Meng-Ju Chi, Chih-Yang Lin, Chih-Hsin Tang. Resistin induces angiogenesis and lymphangiogenesis in human chondrosarcoma [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 1817. doi:10.1158/1538-7445.AM2017-1817
Abstract 1832: Bioluminescent pharmacokinetics of luciferin in preclinical brain metastases of breast cancer models
Authors: Shah Neal , Adkins Chris E. , Mohammad Afroz S. , Lockman Paul R.
Published: July 2017

Background: Approximately 20% of breast cancer patients with disseminated disease will develop brain metastases. Preclinical models of brain metastases of breast cancer rely on ex-vivo histology to evaluate drug efficacy. However, bioluminescence imaging allows more precise quantification of tumor burden and progression through counting photons per second without requiring animal sacrifice. This methodology has not readily been applied to hematogenously derived brain metastases models given the heterogeneity of tumor burden and growth. Herein we demonstrate repeatable methodology to quantify brain metastases of breast cancer progression which is verified with histology.

Methods: Approximately 1.75 x 105 JIMT-1 (n=3) and 231-BrLuc (n=4) brain seeking subclones were injected intracardially to produce metastatic brain lesions. Mice were given 150 mg/kg of luciferin IP and luminescence was captured every 2-minutes for 60 minutes starting 24 hours after cell implantation. Radiance (photons per second per square centimeter per steridian) was plotted to observe peak luminescence. Imaging was repeated twice weekly to evaluate progression until euthanasia.

Results: Following intracardiac injection, both brain seeking metastatic models produced a maximum luminescence signal between 14-20 minutes after luciferin injection. For all subsequent imaging, 5 minute imaging between 15-20 minutes after luciferin injection was used. Longitudinally, both tumor cell lines produce bioluminescence 24 hours after cell injection, which is used to ensure tumor implantation and randomization. The BLI signal decreases to undetectable limits in both models between 3-7 days after cell implantation. Re-emergence of signal occurs on day 14 for the JIMT-1 line and day 21 for 231-Br line. JIMT-1 bioluminescence on day 14 begins at ~105 photons/sec/cm2/sr and then over time signal increases with an ~slope of 5.6x105 until reaching a maximum of 107 units on day 28. 231-Br bioluminescence on day 21 begins at ~104 photons/sec/cm2/sr and then over time signal increases with an ~ slope of 1.1x106 until reaching a maximum of ~107 on day 53.

Conclusion: To accurately measure bioluminescence as a surrogate for tumor burden in hematogenously implanted metastases, the optimal circulation time is approximately 15 minutes with a 5 minute imaging period. Bioluminescence of the 231Br line initially starts at a lower magnitude and later in time than the JIMT-1 line, but slopes of both intracardiac models are similar and result in similar bioluminescent curves.

Citation Format: Neal Shah, Chris E. Adkins, Afroz S. Mohammad, Paul R. Lockman. Bioluminescent pharmacokinetics of luciferin in preclinical brain metastases of breast cancer models [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 1832. doi:10.1158/1538-7445.AM2017-1832
Abstract 1834: Constitutive Nedd9 null genotype promotes lung cancer aggressiveness
Authors: Deneka Alexander , Kopp Meghan , Nikonova Anna S. , Gaponova Anna , Nagele Alexandra , Hensley Harvey , Golemis Erica
Published: July 2017

Non-small cell lung cancer (NSCLC) has a low survival rate, with metastasis contributing to the vast majority of deaths. The NEDD9 (HEF1/Cas-L) protein has been reported to be elevated in expression and to promote metastasis in a large subset of lung cancers and in other malignancies. NEDD9 functions as a scaffold for multiple critical effectors in integrin/FAK/SRC and receptor tyrosine kinase signaling cascades, as well as for the mitotic kinase Aurora-A, with overexpression thought to sustain signaling by these pro-oncogenic proteins. Previous studies demonstrated depletion of NEDD9 by RNAi reduced the growth and invasion of established lung cell lines and tumors, based on cell culture and xenograft assays. We have now investigated the consequences of a null genotype for Nedd9 from the earliest stages of tumor formation, crossing Nedd9 null mice to a 129S/Sv-Krastm3Tyj/Trp53tm1Brn (KP) model in which Kras mutation is induced specifically in lung tissue by inhalation of adenovirus bearing the Cre gene. Unexpectedly, based on in vivo imaging, the Nedd9 null genotype significantly accelerated tumor growth in KP mice. Pathological examination of tissues indicated Nedd9 null genotype also was associated with higher invasive capacity in vivo, including direct invasion to the heart, as well as elevated proliferation rate, decreased apoptotic activity, and changes in the expression of proteins such as vimentin, associated with mesenchymal status. Although Nedd9 function has been implicated in immune cell activity, we found minimal differences in tumor infiltration of myeloid cells or macrophages based on Nedd9 genotype. Rather, use of Reverse Phase Protein Array (RPPA) analysis to characterize signaling in isolated tumors suggested tumor intrinsic changes in glycolytic and proliferative signaling pathways, which were subsequently validated using direct functional assays. These results for the first time imply that NSCLC tumor progression beyond early stages in the absence of Nedd9 requires reprogramming of intrinsic tumor signaling to compensate for the absence of this protein, emphasizing activities distinct from those associated with elevated Nedd9 in late stage tumors.

Citation Format: Alexander Deneka, Meghan Kopp, Anna S. Nikonova, Anna Gaponova, Alexandra Nagele, Harvey Hensley, Erica Golemis. Constitutive Nedd9 null genotype promotes lung cancer aggressiveness [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 1834. doi:10.1158/1538-7445.AM2017-1834
Abstract 1846: Immunocompetent mouse allograft models for development of therapies to target breast cancer metastasis therapies to target breast cancer metastasis
Authors: Yang Yu-an , Yang Howard , Hu Ying , Watson Peter , Liu Huaitian , Geiger Thomas R. , Anver Miriam R. , Haines Diana , Martin Philip , Lee Maxwell P. , Hunter Kent W. , Wakefield Lalage M.
Published: July 2017

Effective drug development to combat metastatic disease in breast cancer would be aided by the availability of well-characterized preclinical animal models that (a) metastasize with high efficiency, (b) metastasize in a reasonable time-frame, (c) have an intact immune system, and (d) capture some of the heterogeneity of the human disease. To address these issues, we have assembled a panel of twelve mouse mammary cancer cell lines that can metastasize efficiently on implantation into syngeneic immunocompetent hosts. Genomic characterization shows that more than half of the 30 most commonly mutated genes in human breast cancer are represented within the panel. Transcriptomically, most of the models fall into the luminal A or B intrinsic molecular subtypes, despite the predominance of an aggressive, poorly-differentiated or spindled histopathology in all models. Patterns of immune cell infiltration, proliferation rates, apoptosis and angiogenesis differed significantly among models. Inherent within-model variability of the metastatic phenotype mandates large cohort sizes for intervention studies but may also capture some relevant non-genetic sources of variability. The varied molecular and phenotypic characteristics of this expanded panel of models should aid in model selection for development of anti-metastatic therapies in vivo, and serve as a useful platform for predictive biomarker identification.

Citation Format: Yu-an Yang, Howard Yang, Ying Hu, Peter Watson, Huaitian Liu, Thomas R. Geiger, Miriam R. Anver, Diana Haines, Philip Martin, Maxwell P. Lee, Kent W. Hunter, Lalage M. Wakefield. Immunocompetent mouse allograft models for development of therapies to target breast cancer metastasis therapies to target breast cancer metastasis [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 1846. doi:10.1158/1538-7445.AM2017-1846
Abstract 1848: Autocrine IGF1 signaling mediates the survival of pancreatic cancer cells following the ablation of oncogenes
Authors: Rajbhandari Nirakar
Published: July 2017

Pancreatic cancer is an extremely lethal disease, which is mostly associated with the gain-of-function mutation in KRAS gene. Due to a very high frequency of activating mutations within the KRAS gene, mutant KRAS is considered a rational therapeutic target in pancreatic cancer. In line with the existing idea that oncogenic KRAS is required for the maintenance of pancreatic cancer, we show a complete macroscopic regression of both primary and orthotopically transplanted tumors in our ligand-regulatable (Doxycycline) mouse model of KRASG12D-driven pancreatic cancer. This finding indicate that the expression, and provides a strong rationale for the development of KRAS targeted therapy for the treatment of pancreatic cancer.

However, in our current study, despite the complete macroscopic regression of PDACs upon downregulation of oncogenic KRAS, few cancer cells always survived and remained dormant for a protracted duration. These residual cancer cells were responsible for the rapid recurrence upon KRAS reactivation. Using a genome-wide gene expression (RNA-Seq) analysis of in vivo-derived bulk and residual cancer cells followed by biochemical analysis, we identified a significant increase in autocrine IGF-1/AKT signaling in residual cancer cells. We also identified IGF1 signaling as a common survival mechanism of residual cancer cells in c-MYC-driven pancreatic tumor model. Pharmacological inhibition of IGF-1R signaling significantly delayed the tumor recurrence in both the tumor models, suggesting the crucial role of IGF-1R signaling in the survival of cancer cells in the absence of oncogenic drivers. Our findings from animal models were also validated in human pancreatic cancer cell lines harboring KRAS mutations, by demonstrating a compensatory increase in IGF1 signaling in response to a conditional knockdown of KRAS in vitro and in vivo. Collectively, all our findings from mouse model and human cancer cell studies strongly indicate that residual cancer cells can survive in the absence of tumor driving oncogenes by upregulation of autocrine IGF1 signaling loop. Thus, targeting oncogenic drivers along with IGF-1R signaling might be an effective strategy for prevention of tumor recurrence in pancreatic cancer.

Citation Format: Nirakar Rajbhandari. Autocrine IGF1 signaling mediates the survival of pancreatic cancer cells following the ablation of oncogenes [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 1848. doi:10.1158/1538-7445.AM2017-1848
Abstract 1850: Development of a NOTCH1 loss of function and PIK3CA mutant transgenic model of HNSCC
Authors: Devenport Samantha N. , Michmerhuizen Nicole L. , Leonard Elizabeth , Jewell Brittany , Swanson Jacob , Wong Sunny , Carey Thomas E. , Brenner J Chad
Published: July 2017

Head and neck squamous cell carcinoma (HSNCC) is a debilitating disease with a poor 5-year survival rate. The use of in vivo models to study tumorigenesis and to identify potential targeted therapies, especially those that work in combination with immune checkpoint inhibitors, is essential in improving overall patient care. However, the overall number of HNSCC transgenic models with recurrent genetic lesions identified by the HNSCC TCGA project has been limited. For example, TCGA data has shown both Notch1 loss of function mutations and PIK3CA activating mutations as some of the most common in HNSCC, but bi-genic models of these lesions do not yet exist. Here, we have designed a model which utilizes K14-CreERT to drive a bi-genic model with Notch1flox and PIK3CAH1047R mutations in epithelial compartments. Characterization of this line both in the context of spontaneous and carcinogen induced tumor formation is a tool which can be utilized to better understand common mutations in HNSCC. Immunohistochemistry confirms the overexpression of PIK3CA in tamoxifen induced mice. Furthermore, early characterization of this model with chronic treatment of the smoking analogue 4-Nitroquinoline N-oxide (4NQO) in drinking water shows tumor formation on the tongue and surrounding head and neck regions in as little as 12-16 weeks. Hematoxylin and Eosin staining confirmed the squamous tumor formation, while Ki67 staining showed enhanced proliferation of the transformed cells. In the future, this model can be used to study tumor formation over time and in response to targeted inhibitors and common therapies that are being advanced as companion diagnostic strategies for either NOTCH1 deficient or PIK3CA aberrant HNSCC tumors. Further detailed characterization of this model will allow for a deeper understanding of the mechanisms which drive HNSCC and will translate to improved patient care.

Citation Format: Samantha N. Devenport, Nicole L. Michmerhuizen, Elizabeth Leonard, Brittany Jewell, Jacob Swanson, Sunny Wong, Thomas E. Carey, J Chad Brenner. Development of a NOTCH1 loss of function and PIK3CA mutant transgenic model of HNSCC [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 1850. doi:10.1158/1538-7445.AM2017-1850
Abstract 1865: Enhancing the in vivo detection of cancer by manipulating magnetic fields applied to tumor targeting superparamagnetic iron oxide nanoparticles
Authors: Karaulanov Todor , Vreeland Erika C. , Gomez Andrew , Minser Kayla E. , Weldon Caroline L. , Anderson William H. , Nettles Christopher , Paciotti Giulio
Published: July 2017

Superparamagnetic Relaxometry (SPMR) is combination technology for the early detection of cancer. Conceptually, PEGylated superparamagnetic iron oxide (Fe3O4) nanoparticles (NPs) coupled with a tumor targeting monoclonal antibody are systemically administered and target solid tumors by both passive (EPR) and active (receptor targeting) mechanisms. Once bound to the target cells, the NPs are magnetized by a brief, low field magnetic pulse to create macroscopic magnetization. Once the field is removed, the particle’s magnetization decays and is measurable by superconducting quantum interference device (SQUID) detectors. The pattern of decay, known as Néel relaxation, exhibits a difference in latency specific to bound nanoparticles and differs from the Brownian decay exhibited by free/unbound particles. As a proof of concept, we developed PEGylated NPs that are covalently bound to an anti-Her-2 monoclonal antibody. In vitro, the nanoconstruct exhibits specific binding to the Her-2 overexpressing BT-474 breast cancer cells, with little to no binding to Her-2 negative MCF-7cells. Similar patterns of selective targeting were observed in vivo.

To improve the specific detection of SPMR signals in tumors we developed a novel system that selectively enhances the magnetic signal at tumor sites and reduces the contribution of similar signals at off target sites such as the liver and spleen. For these studies, we created phantoms of varying strength to mimic tumor and non-specific signals. We observed that by creating non-homogenous excitation magnetic field patterns, we reduced the contribution of the non-specific signals by an order of magnitude, effectively increasing the signal to noise ratio of the tumor signal. These results are clinically relevant and support the use of SPMR in the detection of small tumors in cancer patients.

Citation Format: Todor Karaulanov, Erika C. Vreeland, Andrew Gomez, Kayla E. Minser, Caroline L. Weldon, William H. Anderson, Christopher Nettles, Giulio Paciotti. Enhancing the in vivo detection of cancer by manipulating magnetic fields applied to tumor targeting superparamagnetic iron oxide nanoparticles [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 1865. doi:10.1158/1538-7445.AM2017-1865
Abstract 1869: Improved image-guided surgical resection of glioblastoma with [18F]-fluoroethyltyrosine Cerenkov luminescence imaging
Authors: Lewis David Y. , Mair Richard , Wright Alan J. , Allinson Kieren , Lyons Scott K. , Booth Tom , Bielik Robert , Soloviev Dmitry , Brindle Kevin M.
Published: July 2017

The ability of a surgeon to completely resect a tumor is directly related to outcome in high- and low-grade gliomas. However current intraoperative navigational tools are useful only in a subset of glioma patients and do not detect the full extent of disease.

We modelled orthotopic brain tumors by stereotactically implanting human U87, rat F98 and C6 glioblastoma cells into the forebrains of rats. We show here that a new hybrid PET-Cerenkov luminescence imaging approach using O-(2-[18F]fluoroethyl)-L-tyrosine (FET) can accurately delineate tumor margins pre- and intra-operatively. We demonstrate consistency in the successful localisation and quantification of tumor burden using PET and Cerenkov luminescence imaging. The Cerenkov signal in individual tumors was directly proportional to the signal detected in corresponding FET PET scans (y = 1.06x - 0.01; R2 = 0.98; p < 0.0001) and subsequent autoradiography indicated equivalence between modalities. Cerenkov luminescence was better able to discriminate tumor from healthy brain tissue than the current ‘gold standard’ for intraoperative mapping, 5-ALA (5-aminolevulinic acid), indicated by a greater area under the ROC curve in human xenograft (0.968 ± 0.003 vs. 0.893 ± 0.019; p = 0.003) and syngeneic rat glioma models (0.970 ± 0.010 vs. 0.774 ± 0.046; p = 0.006). The quantitative accuracy of Cerenkov luminescence enabled us to determine a threshold of 2.1 x 102 p-1 sec-1 cm-2 sr-1 MBq-1 that separated U87 human glioblastoma and normal brain tissue, more precisely guiding tumor excision than 5-ALA. At the optimal threshold the specificity for detecting syngeneic rat and human xenograft gliomas was better with FET Cerenkov than 5-ALA (91.1 ± 2.7 % vs. 67.2 ± 5.2 %; p = 0.007 and 91.2 ± 1.0 % vs. 82.9 ± 1.8 %; p = 0.005 respectively). We confirmed and extended these findings at higher spatial resolution on cryosections using FET autoradiography and 5-ALA confocal microscopy, with FET demonstrating higher specificity and sensitivity for tumor detection than 5-ALA.

FET PET-Cerenkov luminescence imaging has the potential for guiding resections in a much broader range of glioma patients than current approaches. Previous clinical experience with FET offers a facile route for clinical translation of this technology.

Citation Format: David Y. Lewis, Richard Mair, Alan J. Wright, Kieren Allinson, Scott K. Lyons, Tom Booth, Robert Bielik, Dmitry Soloviev, Kevin M. Brindle. Improved image-guided surgical resection of glioblastoma with [18F]-fluoroethyltyrosine Cerenkov luminescence imaging [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 1869. doi:10.1158/1538-7445.AM2017-1869