bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1566: Establishment of analytical performance features of two immunohistochemistry assays for PD-L1 expression in lung cancer
Authors: Zhang Lei , Begot Claire , McGee Robert , Anderson Steven M.
Published: August 2018

Therapies that target PD-1/PD-L1 and overcome checkpoint inhibition have shown great promise in the treatment of a variety of cancers, with PD-L1 expression demonstrated as a biomarker of therapy response in a variety of tumor types. There are multiple IHC assays that are used to measure PD-L1 expression in formalin-fixed, paraffin-embedded tissues (FFPE) with significant variability in primary antibody, assay platform, scoring criteria, and assay cut-offs. This situation creates potential confusion in applications of PD-L1 immunohistochemistry (IHC) assays for the appropriate intended use. In this study we evaluated two assays (Dako pharmDx PD-L1 28-8 assay and Ventana PD-L1 SP263) in 45 non-small cell lung cancer samples encompassing predominately squamous cell carcinomas (35) and adenocarcinomas (10). PD-L1 protein expression, measured by the percentage of tumor cell showing positivity for PD-L1 staining, has been compared between two assays. PD-L1 results were interpreted by using various assay cutoffs. Correlation of PD-L1 expression measured by positivity in percentage of tumor cell staining with each of the two assays achieved a significant concordance (r2 = 0.91). Similar trends in percentage of positive of tumor cells were demonstrated across a broad dynamic range of staining. Cutoffs that have been established in clinical studies and referenced in the respective assay package inserts (>1% positivity percentage of tumor cells for 28-8 assay and >25% positivity percentage of tumor cells for the SP263 assay) were used as the reference, for interpreting the test results as positive or negative. When these established cutoffs were applied, a low-to-medium concordance between two assays has been demonstrated (r2 = 0.58). When the same cutoff (1% or 25%) was applied, the correlation has been improved markedly (r2 = 0.73 or 0.95 respectively). In this sample set the 25% cutoff produced the more consistent data when comparing the two methods. Changes in the classification of PD-L1 status was also observed for both Dako pharmDx PD-L1 28-8 assay (29%, 13/45) and Ventana PD-L1 SP263 (22%, 10/45) assays when the established cutoff was replaced. The findings of this study are consistent with others such as those from Blueprint phase I study. This comparison study showed that the two assays (Dako pharmDx PD-L1 28-8 assay and Ventana PD-L1 SP263) were analytically comparable. Further investigation is warranted to explore both the analytical and clinical performance of the various PD-L1 assays across multiple assay cut-points.

Citation Format: Lei Zhang, Claire Begot, Robert McGee, Steven M. Anderson. Establishment of analytical performance features of two immunohistochemistry assays for PD-L1 expression in lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1566.
Abstract 1568: Advancing convergence and innovation in cancer research: National Cancer Institute Center for Strategic Scientific Initiatives (CSSI)
Authors: Elder Sarah G. , Lee Jerry S. , Berny-Lang Michelle A. , Hanlon Sean E.
Published: August 2018

Since 2003, the National Cancer Institute's Center for Strategic Scientific Initiatives (CSSI) has worked to develop the resources and infrastructures investigators need to surmount roadblocks in cancer research. To this end, CSSI manages programs that promote technology development and cross-disciplinary collaboration and provide support for investigators in nascent and challenging research fields. This support includes funding opportunities, shared reagent and database resources, and assistance in the development of standards and protocols. CSSI also provides a network of partners in industry and government that can help NCI-funded researchers advance their technologies toward commercialization and translation. This presentation will highlight technologies including single-cell isolation and analysis techniques that have been supported through various CSSI mechanisms from proof of concept to translation into the clinic.

Citation Format: Sarah G. Elder, Jerry S. Lee, Michelle A. Berny-Lang, Sean E. Hanlon. Advancing convergence and innovation in cancer research: National Cancer Institute Center for Strategic Scientific Initiatives (CSSI) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1568.
Abstract 1570: Loss of mismatch repair protein MSH6 sensitizes diffuse-type gastric cancer cells to DNA-PKcs inhibitor KU60648
Authors: Krishnan Vaidehi , Lau Wen Min , Teng Eileen , Tan Jin Wei , Sankar Haresh , Lim Debbie Xiu En , Pang Crystal Kok Yan , Teh Ming , So Jimmy Bok Yan , Tan Patrick , Ito Yoshiaki , Chan Shing Leng
Published: August 2018

Diffuse-type gastric cancer (DGC) is a histologic subtype characterized by poor cellular cohesiveness, increased metastasis and poor prognosis. As preclinical models of DGC that reliably predict clinical activity of novel compounds are lacking, we generated patient-derived xenograft (PDX) models using malignant ascites from DGC patients that closely recapitulated the primary cancer. To uncover novel therapeutic targets, we applied the concept of synthetic lethality and asked if DGCs require DNA repair proteins for survival. Major DNA repair kinases ATM, ATR and DNA-PKcs in established DGC cell lines and ex vivo cell lines from PDXs (GAGA3, GAGA6 and GAS24) were inhibited using small-molecule inhibitors KU55933, VE-821 and KU60648, respectively. Interestingly, most diffuse-type gastric cells exhibited elevated lethality when DNA-PKcs, a central kinase that regulates the non-homologous end joining pathway (NHEJ), was inhibited by KU60648 as compared to the nontransformed gastric epithelial line, HFE-145. We confirmed that genetic depletion of DNA-PKcs by siRNA resulted in cell death in two PDX lines GAS24 and GAGA6 cells but not in HFE-145 cells, suggesting the dependence of these cells on DNA-PKcs for survival. Two mechanisms appear to sensitize DGC cells to DNA-PKcs inhibition; firstly, impaired homologous recombination repair in GAS24 and GAGA6 as shown by compromised ability to resolve radiation-induced γH2AX foci, and secondly, transcriptional downregulation of the mismatch repair protein, MSH6, in GAGA3 and GAGA6. Knockdown of MSH6 in GAS24 cells enhanced their sensitivity to KU60648, suggesting a functional link between MSH6 and DNA-PK function. Lastly, to develop MSH6 as a biomarker for predicting DNA-PKcs sensitivity, we analyzed MSH6 levels by immunohistochemical staining in formalin-fixed, paraffin-embedded (FFPE) tissue sample sections derived from PDXs. In these preliminary studies, MSH6 expression was lower with high heterogeneity in GAGA3 and GAGA6 compared to GAS24. In this study, we show for the first time the downregulation of DNA mismatch protein MSH6 in sporadic diffuse gastric cancer, and its potential as a therapeutic predictor for DNA-PKcs inhibition treatment.

Citation Format: Vaidehi Krishnan, Wen Min Lau, Eileen Teng, Jin Wei Tan, Haresh Sankar, Debbie Xiu En Lim, Crystal Kok Yan Pang, Ming Teh, Jimmy Bok Yan So, Patrick Tan, Yoshiaki Ito, Shing Leng Chan. Loss of mismatch repair protein MSH6 sensitizes diffuse-type gastric cancer cells to DNA-PKcs inhibitor KU60648 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1570.
Abstract 1572: The value of p16 or HPV as predictive marker of anti-EGFR treatment in recurrent/metastatic HNSCC
Authors: Lim Joo Han , Hong Soon-Sun , Kim Jin-Chul , Lee Moon Hee , Cho Jinhyun
Published: August 2018

Introduction: The anti-epidermal growth factor receptor (EGFR) agent was approved and has been a mainstay of treatment of recurrent/metastatic head and neck squamous cell cancers (HNSCC). However, it is unclear whether human papilloma virus (HPV) modulates the response to targeting the EGFR. Some subset analysis of the randomized phase III study is confusing and recent The Cancer Genome Atlas data showed that EGFR gene is rarely abnormal in HPV-positive HNSCC tumors while it is frequently altered in HPV-negative HNSCC tumors. The purpose of our meta-analysis is to evaluate the effects of anti-EGFR agents regarding HPV status.

Materials and Methods: We conducted a pooled analysis to evaluate the association of p16 expression status with outcomes in patients with recurrent/metastatic HNSCC receiving palliative anti-EGFR treatment with or without chemotherapy. A literature search of PubMed, EMBASE, and meeting abstracts of the American Society of Clinical Oncology and European Society of Medical Oncology through December 2017 was conducted.

Results: A total of seven trials met the selection criteria. The total number of patients was 700 (554 with the HPV-negative group and 146 with the HPV-positive group). Anti-EGFR treatment was associated with higher response rate in p16-negative patients compared with in p16-positive patients (OR 2.18; 95% CI, 1.18-3.99, p=0.01).

Conclusion: A meta-analysis of trials with anti-EGFR agents in p16-negative RMHNSCC patients versus in p16-positive recurrent/metastatic HNSCC patients yielded that p16 status could be a predictive biomarker for anti-EGFR treatment in recurrent/metastatic HNSCC. Prospective study is warranted to determine the value of p16 or HPV as a predictive marker of anti-EGFR treatment in recurrent/metastatic HNSCC.

Citation Format: Joo Han Lim, Soon-Sun Hong, Jin-Chul Kim, Moon Hee Lee, Jinhyun Cho. The value of p16 or HPV as predictive marker of anti-EGFR treatment in recurrent/metastatic HNSCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1572.
Abstract 1573: A clinical biomarker assay to quantitate MIP-1a in human serum
Authors: Wong Li Chin , Wong Oitak , Laterza Omar F. , Zhao Xuemei
Published: August 2018

Macrophage inflammatory protein 1-alpha (MIP-1alpha, MIP-1α, CCL3) is a cytokine that is involved in the acute inflammatory state in recruitment and activation of polymorphonuclear leukocytes through binding to the receptors CCR1, CCR4 and CCR5. MIP-1α is expressed by macrophages, dendritic cells and lymphocytes. Crucial in immune response against infection and inflammation, MIP-1α induces the synthesis and release of IL-6, IL-10 and TNFα. As a proinflammatory chemokine, MIP-1α is primarily associated with cell adhesion and migration of monocytes to inflammatory tissues. In addition, MIP-1 α inhibits the proliferation of hematopoietic stem cells in vitro and in vivo. Expression levels of MIP-1 α have provided important information in the progression of diseases such as multiple myeloma, lung cancer, multiple sclerosis, HIV infection, allergic asthma and sepsis.

Citation Format: Li Chin Wong, Oitak Wong, Omar F. Laterza, Xuemei Zhao. A clinical biomarker assay to quantitate MIP-1a in human serum [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1573.
Abstract 1576: Clonal diversity revealed by morphoproteomic and copy number profiles of single prostate cancer cells at diagnosis
Authors: Malihi Paymaneh D. , Morikado Michael , Welter Lisa , Carlsson Anders , Velasco Carmen Ruiz , Kolatkar Anand , Rodriguez-Lee Mariam , Hicks James , Kuhn Peter , Liu Sandy T. , Miller Eric T. , Cadaneanu Radu M. , Knudsen Beatrice S. , Lewis Michael S. , Garraway Isla P.
Published: August 2018

Tumor heterogeneity is prevalent in both treatment-naïve and end-stage metastatic castration-resistant prostate cancer (PCa), and may contribute to the broad range of clinical presentation, treatment response, and disease progression. To characterize molecular heterogeneity associated with de novo metastatic PCa, multiplatform single cell profiling was performed using High Definition Single Cell Analysis (HD-SCA). HD-SCA enabled morphoproteomic and morphogenomic profiling of single cells from touch preparations of tissue cores (prostate and bone marrow biopsies) as well as liquid samples (peripheral blood and bone marrow aspirate). Morphology, nuclear features, copy number alterations, and protein expression were analyzed. Tumor cells isolated from prostate tissue touch preparation (PTTP) and bone marrow touch preparation (BMTP) as well as metastatic tumor cells (MTCs) isolated from bone marrow aspirate were characterized by morphology and cytokeratin expression. Although peripheral blood was examined, circulating tumor cells were not definitively observed. Targeted proteomics of PTTP, BMTP, and MTCs revealed cell lineage and luminal prostate epithelial differentiation associated with PCa, including co-expression of EpCAM, PSA, and PSMA. Androgen receptor expression was highest in MTCs. Hallmark PCa copy number alterations, including PTEN and ETV6 deletions and NCOA2 amplification, were observed in cells within the primary tumor and bone marrow biopsy samples. Genomic landscape of MTCs revealed to be a mix of both primary and bone metastatic tissue. This multiplatform analysis of single cells reveals several clonal origins of metastatic PCa in a newly diagnosed, untreated patient with polymetastatic disease. This case demonstrates that real-time molecular profiling of cells collected through prostate and bone marrow biopsies is feasible and has the potential to elucidate the origin and evolution of metastatic tumor cells. Altogether, biological and genomic data obtained through longitudinal biopsies can be used to reveal the properties of PCa and can impact clinical management.

Citation Format: Paymaneh D. Malihi, Michael Morikado, Lisa Welter, Anders Carlsson, Carmen Ruiz Velasco, Anand Kolatkar, Mariam Rodriguez-Lee, James Hicks, Peter Kuhn, Sandy T. Liu, Eric T. Miller, Radu M. Cadaneanu, Beatrice S. Knudsen, Michael S. Lewis, Isla P. Garraway. Clonal diversity revealed by morphoproteomic and copy number profiles of single prostate cancer cells at diagnosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1576.
Abstract 1579: IMI's CANCER-ID: Implications of liquid biopsy technology standardization for its clinical use
Authors: Pantel Klaus , Terstappen Leon W. , Baggiani Barbara , Krahn Thomas , Panabières Catherine , Lianidou Evi , Naume Bjørn , Farace Françoise , Schlange Thomas
Published: August 2018

Liquid biopsies offer the opportunity to improve the diagnosis and timely adaption of therapy options to the progression and heterogeneity of cancer. The CANCER-ID (www.cancer-id.eu) consortium was established in early 2015 as part of the Innovative Medicines Initiative (IMI) and has currently 37 partners from 14 countries, including the USA and Singapore. The goal of the CANCER-ID program is to foster implementation of liquid biopsy technologies in multicenter clinical studies and the potential regulatory approval of devices or protocols in the field. Within the project best practice protocols for using technologies aiming at the enrichment, isolation and analysis of Circulating Tumor Cells (CTCs), circulating free tumor DNA (ctDNA) and microRNAs (miRNAs) are being developed and benchmarking data for the performance of different technologies are collected. To this end defined standards for the different biomarker classes are crucial and their introduction and design is being supported by CANCER-ID. Another integral part of our work is the development of integrated workflows from patient sample collection, pre-analytical sample handling, sample and bioinformatics analyses down to the actionable information guiding patient selection and personalized treatment.

CANCER-ID partners are currently planning clinical studies to assess the clinical utility of analyzing PD-L1 expression on CTCs in their core indications Non-Small Cell Lung Cancer (NSCLC) and metastatic breast cancer. While the benefit of immune checkpoint inhibition has been impressively demonstrated in NSCLC, most advanced breast cancer patients do not benefit from targeting PD-1 with the exception of the triple-negative disease type. The liquid biopsy workflows established by the CANCER-ID consortium will be used to correlate CTC PD-L1 expression in patients treated with PD-1 blocking antibodies with clinical response and in parallel perform mutational load analysis in blood. The aim will be to attempt to develop a liquid biopsy-based rationale for patient selection guiding effective immune checkpoint therapies. The study plan and current status of this multicenter effort will be presented.

This work is supported by IMI JU & EFPIA (grand no. 115749, CANCER-ID). Samples from patients and healthy volunteers, respectively, were collected under signed informed consent.

Citation Format: Klaus Pantel, Leon W. Terstappen, Barbara Baggiani, Thomas Krahn, Catherine Panabières, Evi Lianidou, Bjørn Naume, Françoise Farace, Thomas Schlange. IMI's CANCER-ID: Implications of liquid biopsy technology standardization for its clinical use [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1579.
Abstract 1583: Clinical significance of PIK3CA, AKT1, and ESR1 mutation in plasma cell-free DNA from estrogen receptor-positive breast cancer patients
Authors: Takeshita Takashi , Tomiguchi Mai , Sueta Aiko , Yamamoto-Ibusuki Mutsuko , Yamamoto Yutaka , Iwase Hirotaka
Published: August 2018

Background: In endocrine therapy (ET) resistance mechanisms, PI3K / AKT pathway abnormality and ESR1 mutation are drawing attention. In recent large-scale clinical trials, it was revealed that identification of these genetic abnormalities in cell-free DNA (cfDNA) was useful for rapid assessment and monitoring of the therapeutic effect in ET-resistant breast cancer (BC). However, the frequency of them per treatment line and its clinical significance have not been verified. Here we studied the clinical significance of PIK3CA, AKT1, and ESR1 mutation according to the treatment line in estrogen receptor (ER)-positive BC.

METHODS: From 2003 to 2017, a total of 251 plasma specimens were collected from 128 patients with ER positive BC treated at our hospital. The breakdown were 133 plasma samples from 73 primary BC (PBC) patients and 118 plasma samples from 68 metastatic BC (MBC) patients. CfDNA was extracted from 500 μL of plasma. The hotspot of PIK3CA, AKT1, and ESR1 mutation in plasma cfDNA was verified using multiplex digital PCR method.

RESULTS: In the PBC patient group, PIK3CA mutation was recognized at 15.1%, AKT1 mutation at 1.4%, and ESR1 mutation at 2.7%. The presence or absence of PIK3CA mutation did not affect clinical outcome. In the MBC patient group, the frequency of PIK3CA mutation increased from 16% to 32% and that of ESR1 mutation increased from 23% to 41.9% as the treatment line advanced. Furthermore, we examined the time to treatment failure (TTF) by dividing into early treatment line and late treatment line. In the early treatment group, patients with PIK3CA mutation had significantly shorter TTF (P = 0.035). However, the presence or absence of ESR1 mutation did not affect TTF. On the other hand, in the late treatment group, patients with ESR1 mutation had significantly shorter TTF (P = 0.048). However, the presence or absence of PIK3CA mutation did not affect TTF. Since AKT1 mutation was rare in both PBC patients and MBC patients, its clinical significance was unknown.

CONCLUSION: We showed clinical significance of verification of PIK3CA, AKT1, and ESR1 mutation in cfDNA according to treatment line in ER positive BC patients.

Citation Format: Takashi Takeshita, Mai Tomiguchi, Aiko Sueta, Mutsuko Yamamoto-Ibusuki, Yutaka Yamamoto, Hirotaka Iwase. Clinical significance of PIK3CA, AKT1, and ESR1 mutation in plasma cell-free DNA from estrogen receptor-positive breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1583.
Abstract 1587: Detection of PD-L1 and PD-1 positive circulating tumor cells (CTCs) in non-small cell lung cancer (NSCLC) patients treated with nivolumab
Authors: Kallergi Galatea , Koutsopoulos Anastasios , Lagoudaki Eleni , Agouraki Despoina , Vetsika Eleni-Kiriaki , Voumvouraki Anastasia , Martin Stuart S. , Koinis Filippos , Stournaras Christos , Georgoulias Vassilis , Kotsakis Athanasios
Published: August 2018

Introduction: Circulating tumor cells (CTCs) are considered as a “liquid biopsy” that allows the assessment of tumor changes over time. Tumor cells may escape from the immune system through the activation of PD-1/PD-L1 axis. Targeting these molecules with monoclonal antibodies has shown encouraging results at many types of cancers, including NSCLC. In the current study we investigated the expression of PD-1/PD-L1 molecules on the CTCs isolated from NSCLC patients treated with Nivolumab.

Methods: CTCs were isolated based on their size using the ISET platform from 27 patients before treatment, after 1 cycle and 3 cycles. CTCs were detected with Giemsa staining and immunofluorescence (IF) experiments, using either pancytokeratin (A45-B/B3) (CK7)/PD-1/CD45 or (A45-B/B3)(CK7)/PD-L1/CD45 combination of antibodies and analysis with the ARIOL system. Spiking experiments using the NSCLC cell lines: H460, H1299, HCC827 and SKMES in normal blood were used to evaluate the detection method.

Results: Giemsa evaluation in Nivolumab-treated patients at baseline (25 evaluable samples), after the 1st (9 evaluable samples) and the 3rd (8 evaluable patients) cycle of treatment showed that CTCs could be detected in 48% (12/25), 33.3% (3/9) and 50% (4/8) of patients, respectively. IF could also reveal the presence of CK-positive cells in 44.4% (12/27), 22% (2/9) and 75% (6/8) patients, respectively. PD-1 (+) CTCs were detected in 33.3% (4/12) of patients at baseline, in, 0% after the 1 and 16.7% (1/6) of patients after the 3rd cycle. The same percentages were identified for PD-L1 expression in the same cohort of patients. The expression of PD-1 at baseline was associated with poorer OS (p=0.022) and PFS (p=0.011), while the expression of PD-L1 was associated with shorter PFS (p=0.011). Multivariate analysis revealed that the presence of CK-positive cells is an independent prognostic factor for OS (p=0.028)

Conclusion: Nivolumab reduced the number of PD-1- and PD-L1-expressing CTCs in advanced NSCLC patients. Furthermore the expression of both markers at baseline is associated with the clinical outcome.

Citation Format: Galatea Kallergi, Anastasios Koutsopoulos, Eleni Lagoudaki, Despoina Agouraki, Eleni-Kiriaki Vetsika, Anastasia Voumvouraki, Stuart S. Martin, Filippos Koinis, Christos Stournaras, Vassilis Georgoulias, Athanasios Kotsakis. Detection of PD-L1 and PD-1 positive circulating tumor cells (CTCs) in non-small cell lung cancer (NSCLC) patients treated with nivolumab [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1587.
Abstract 16: A wide-spectrum analysis of angiogenesis-related factors in ovarian cancer reveals a signature potentially for prognosis and therapeutic targeting
Authors: Kuo Chih-Lin
Published: August 2018

Ovarian cancer is the first leading cause of death from woman gynecological malignancies worldwide. It is typically diagnosed at a late stage with a poor 5-year survival rate less than 30%. In the progress of ovarian cancer, angiogenesis-related factors not only induce neovascularization for supporting tumor growth but also accelerate ascites production, which helps the peritoneal metastasis and is associated with poor disease prognosis. Therefore, angiogenesis-related factors are thought to be particularly crucial in the persistence of ovarian cancer and have become key targets in clinical research. Although the expression of some angiogenesis-related factors in ovarian cancer has been sporadically reported, analysis of the profile of angiogenesis-related factors in a whole spectrum manner is needed to evaluate their cruciality and relationship. In our study, totally 104 angiogenesis-related factors were used to analyze their transcriptional changes along with ovarian cancer progression in the data of 601 patients obtained from TCGA datasets (AgilentG4502A_07_2 and AgilentG4502A_07_3). 33 candidates with significantly differential expression (p < 0.05) between normal and any stage were initially revealed. To narrow down the candidates, these factors were ranked by the significance of expressional changes between normal and all various stages and 13 factors with the most significant difference (p < 0.0001) were selected and their fold changes were further shown in a heat-map manner; these included MMP9, VEGFA, CXCL10, TIMP2, AGRP, MMP8, IGFBP-1, ENG, TEK, FGF7, CSF3, ANGPTL5, ANGPTL7. Moreover, Ingenuity Pathway Analysis (IPA) and DAVID database analysis were applied to analyze their functional categories in angiogenesis. In conclusion, we developed a signature of angiogenesis-related factors for patients with ovarian cancer. With appropriate validation, it can be applied as a prognostic index or used to select certain candidates for developing targeted therapies.

Citation Format: Chih-Lin Kuo. A wide-spectrum analysis of angiogenesis-related factors in ovarian cancer reveals a signature potentially for prognosis and therapeutic targeting [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 16.
Abstract 1602: Urine cell-free DNA as early biomarker in immunotherapy response in NSCLC
Authors: Wekken Anthonie J. van der , Elst Arja ter , Miedema Anneke , Hiltermann Thijo J. , Goen Harry J. , Schuuring Ed
Published: August 2018

Introduction: Immunotherapy shows a durable tumor response in approximately 20% of advanced NSCLC patients. Imaging by CT is used to evaluate tumor response. PD-L1 immunohistochemistry is generally used to predict survival advantages, however with low predictive value. Therefore, it is important that alternative predictive biomarkers are investigated. Minimal invasive techniques may be a solution. The role of circulating cell-free DNA (cfDNA) was found to be an early biomarker for tumor response in KRAS positive Non-Small Cell Lung Cancer (NSCLC) patients treated with immunotherapy (N=6). In this study we will determine the role of urine cell-free DNA (ucfDNA) in an extended cohort. The non-invasive method of urine collection makes it an excellent source for mutation detection and follow-up.

Methods: In an ongoing study KRAS positive patients treated with immunotherapy, are being tested for cfDNA at baseline (before treatment, 1, 2, 4, 6 and thereafter every 12 weeks until disease progression) with a KRAS G12-13 or Q61H digital droplet PCR (ddPCR) screening assay (BioRad). In a pilot study, urine from these patients is collected in a tube with 5 ml of STRECK reagent at baseline and at 6 weeks.

Results: In 16/27 patients with KRAS positive advanced NSCLC, the mutation was detected in plasma at baseline and subsequent evaluation time points. Urine cfDNA as a predictive marker seems promising and results will be reported and presented at the AACR meeting.

Conclusion: Among patients with advanced NSCLC treated with immunotherapy plasma and urine cfDNA are collected and will be presented as non-invasive early biomarker for tumor response evaluation.

Citation Format: Anthonie J. van der Wekken, Arja ter Elst, Anneke Miedema, Thijo J. Hiltermann, Harry J. Goen, Ed Schuuring. Urine cell-free DNA as early biomarker in immunotherapy response in NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1602.
Abstract 1604: Highly sensitive digital detection of circulating DNA cancer mutations using synthetic genome standards
Authors: Wood-Bouwens Christina M. , St.Onge Robert P. , Ji Hanlee P.
Published: August 2018

With the emergence of precision cancer medicine, there is an increasing need for new longitudinal diagnostic tests to evaluate patients at the initial diagnosis, during treatment, and for routine monitoring. As a general solution, we developed a single-color digital PCR assay technology that detects and quantifies circulating DNA cancer mutations collected from the plasma of cancer patients. The assay has a sensitivity of 0.1% mutation allelic fraction and can be designed for nearly any cancer mutation. In addition, this assay demonstrates design flexibility, low cost, and robust performance, making it a powerful approach for longitudinal monitoring of cancer patients during treatment.

Positive controls are a critical component for highly sensitive detection of circulating DNA cancer mutations. However, there are minimal resources for providing standard curves. Commercially available cancer cell lines cover a very limited number of cancer mutations. Amplicons or oligonucleotides pose a high risk of contamination due to the sheer number of molecules generated, an important consideration when working with rare molecular species such as cell-free DNA.

To address this challenge, we developed a novel technology that enables a single copy of synthetic human genome sequence to be introduced into a non-mammalian eukaryotic genome. The system highly-scalable and capable of producing thousands of unique clones which can be archived for future use, accessed individually, or combined to create customizable target pools on-demand. These custom designed synthetic positive controls are therefore readily renewable, require no amplification to obtain, and most importantly allow precise control over the number of molar equivalents of each target in a reaction. Using these synthetic controls, we simulate low mutation-allele fractions to validate the specificity and sensitivity of our assay.

As a proof of concept, we have designed approximately 400 cancer mutation controls in driver genes such as KRAS and EGFR, and developed a design process to generate multiplex digital PCR assays targeting these driver genes. We will validate the digital PCR assays with standard bulk PCR and will validate the template by comparing to results generated using cancer cell line derived DNA. In the era of circulating DNA diagnostics, these genome engineered synthetic controls meet a critical need for reliable and customizable positive controls.

Citation Format: Christina M. Wood-Bouwens, Robert P. St.Onge, Hanlee P. Ji. Highly sensitive digital detection of circulating DNA cancer mutations using synthetic genome standards [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1604.
Abstract 162: UBE2T: A molecular regulator for cancer stemness and drug resistance in hepatocellular carcinoma
Authors: Ho Pui Yu , Lee Kin Wah
Published: August 2018

Hepatocellular carcinoma (HCC) is one of the common cancers worldwide. Poor prognosis of HCC patients is attributed to high frequency of tumor relapse and chemoresistant nature of this disease. Increasing evidence showed the critical role of cancer stem cells (CSCs) on tumor relapse and therapeutic resistance. Since normal stem cells and CSCs share high similarity, we would like to identify the molecules/pathways crucial in liver CSCs by determining what molecules involved in normal liver stem cells during liver regeneration. Comparison of expression profiles between early regenerating liver and intact liver revealed the upregulation of DNA Damage Response pathways in the self-renewing liver, in which Ubiquitin Conjugating Enzyme E2 T (UBE2T) was the most significantly upregulated. This, together with the publicly available dataset (GSE5975) shows upregulation of UBE2T in EpCAM-enriched liver CSC populations, suggest the potential role of UBE2T on regulation of cancer stemness. By qPCR analysis UBE2T mRNA level was overexpressed in 91% of the clinical HCC specimens (n=56) and associated with aggressive tumor behavior and poorer patients' survival. The overexpression of UBE2T was further confirmed by western blot and immunohistochemical analysis. By lentiviral based knock-down approach, we demonstrated the role of UBE2T in regulation of liver CSC properties, including self-renewal, tumorigenicity, drug resistance and expression of liver CSC markers. In orthotopic HCC model, UBE2T was found to play pivotal role in lung metastasis in vivo. Mechanistically, UBE2T is found to interact directly with E3 ligase Mule and regulate its protein expression via ubiquitination. Since Mule is found to directly degrade β-catenin protein, we are examining whether UBE2T mediates liver T-IC function through direct regulation of Mule-mediated β-catenin degradation. In conclusion, we have uncovered the novel role of UBE2T signaling cascade in regulation liver CSCs, which provides attractive therapeutic target for potential treatment of HCC.

Citation Format: Pui Yu Ho, Kin Wah Lee. UBE2T: A molecular regulator for cancer stemness and drug resistance in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 162.
Abstract 163: Glioblastoma, cancer stem cells, and reactive species balances: A case for GTP cyclohydrolase 1
Authors: Tran Anh N. , Walker Kiera , Harrison David G. , Chen Wei , Mobley James , Hocevar Lauren , Hackney James R. , Sedaka Randee , Pollock Jennifer , Goldberg Matthew S. , Hambardzumyan Dolores , Cooper Sara J. , Gillespie G Yancey , Hjelmeland Anita B.
Published: August 2018

Glioblastoma (GBM), or grade IV astrocytoma, is a deadly disease due in part to the high degree of intratumoral heterogeneity that contributes to treatment failures. Previous studies have shown the importance of reactive species balances, partially controlled by the coupling of nitric oxide synthases (NOS) with their cofactor, in maintenance of glioma stem cell (GSC) phenotype as well as survival of cancer cells in general. In this study, we investigated the roles of GTP cyclohydrolase 1 (GCH1), which is the first and rate-limiting enzyme of the pathway producing of NOS cofactor producing pathway, in GBM stem cell phenotypes via redox balances. We found that GCH1 RNA and protein expression were increased in GSCs in comparison to non-GSCs, but that GCH1 was not exclusive to the GSC fraction. Indeed, GCH1 was elevated in GBM in comparison to normal brain. Overexpression of GCH1 in GBM cells increased cell growth in vitro and neurosphere-forming capability and decreased survival in an intracranial GBM mouse model. In contrast, GCH1 knockdown with short hairpin RNA in GBM cells led to growth inhibition in vitro as well as increased survival in animal models. GCH1 increased CD44 expression and was upregulated in the detrimental mesenchymal GBM subtype in which CD44 served as a marker. Mechanistically, we found that the expression of GCH1 increased BH4 production, as well as augmented multiple antioxidant pathways, including the expression of PARK7, was critical for controlling reactive species balance, including suppressing reactive oxygen species production. In silico analyses demonstrated that higher GCH1 levels in glioma patients correlate with higher glioma grade, recurrence and worse survival. Together, our data suggest that upregulation of GCH1 in GSCs promotes tumor maintenance and is a key regulator of reactive oxygen species in GBM, and GCH1 pathway is a potential target for therapy.

Citation Format: Anh N. Tran, Kiera Walker, David G. Harrison, Wei Chen, James Mobley, Lauren Hocevar, James R. Hackney, Randee Sedaka, Jennifer Pollock, Matthew S. Goldberg, Dolores Hambardzumyan, Sara J. Cooper, G Yancey Gillespie, Anita B. Hjelmeland. Glioblastoma, cancer stem cells, and reactive species balances: A case for GTP cyclohydrolase 1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 163.
Abstract 1638: Is short interval colonoscopy at the discretion of the gastroenterologist
Authors: Brim Hassan , Tushir Abhimanyu , Manchegowda Shashidhar , Farsi Negin , Lee Edward L. , Adeyinka Laiyemo O. , Nouraie Seyed M. , Ashktorab Hassan
Published: August 2018

BACKGROUND: Gastroenterologists do not always follow guidelines for performing colonoscopies. Short interval colonoscopies are recommended at the discretion of gastroenterologists depending on various factors. We here evaluated such factors in an African American patient population.

AIM: To assess factors contributing to short interval (within 3 months) colonoscopies.

METHODS: Data was retrieved retrospectively from Howard University Hospital database (Jan. 2010 to Dec. 2016). African American patients who had repeated colonoscopies within a period of 3 months were selected for this study. We analyzed patients' demographics, family history, polyp characteristics, and reasons for repeat colonoscopy along with latest follow-up findings.

RESULTS: There were 36 patients who underwent two colonoscopies within 3 months, 13 females and 23 males with mean age of 61 years. Out of these, 7 patients (19.4%) had poor bowel preparation. For the remaining 29 patients, the reason for second colonoscopy were partial removal and suspicion of partial removal (61.1%), difficult access (16.7%), and follow-up for Crohn's disease (2.8%). Follow-up of these 36 patients after their last colonoscopy revealed that 8 patients (22.2%) had colon cancer/resection, 4 other patients (11.1%) had cancer of other organs. Of the remaining, 6 (16.6%) had future colon polyp resection, 3 (8.3%) had normal screening colonoscopy and 4 (11.1%) visited the hospital for reasons unrelated to gastroenterology while 11 patients (30.6%) had no follow up.

CONCLUSION: Even though there are no recommendations to repeat colonoscopy within 3 months, endoscopists perform such colonoscopies at their own discretion based on factors like high risk individuals, poor bowel preparation, multiple lesions, type of lesions and resection clearance. Of interest, a significant number of patients in our study had colon cancer/resection on eventual follow-ups.

Citation Format: Hassan Brim, Abhimanyu Tushir, Shashidhar Manchegowda, Negin Farsi, Edward L. Lee, Laiyemo O. Adeyinka, Seyed M. Nouraie, Hassan Ashktorab. Is short interval colonoscopy at the discretion of the gastroenterologist [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1638.
Abstract 1649: Impact of linker and conjugation site on tubulysin M ADC stability and in vivo activity
Authors: Burke Patrick J. , Hamilton Joseph Z. , Pires Thomas A. , Leiske Christopher I. , Cochran Julia H. , Setter Jocelyn R. , Emmerton Kim K. , Waight Andrew B. , Senter Peter D. , Lyon Robert P. , Jeffrey Scott C.
Published: August 2018

The tubulysins are a potent class of microtubule-disrupting agents consisting of natural products and designed analogues that have become a compelling cytotoxic payload for drug-targeting applications. This is due in large part to their high cytotoxic potency on cancer cells, including those that express transporters conferring the multidrug resistance (MDR+) phenotype. All potent tubulysin natural products contain an acetoxy group at the C11-position in the central tubuvaline residue. Deacetylation results in >100-fold loss of cellular and biochemical potency, underscoring the importance of this structural element. Recently, we developed quaternary ammonium linker systems to conjugate tertiary amine-containing payloads, and applied them to the tubulysins. In this work, protease- and β-glucuronidase-cleavable quaternary ammonium linkers of tubulysin M were evaluated as ADC payloads. In vitro, both dipeptide and glucuronide linker systems provided ADCs with comparable biologic properties. However, in vivo, linker chemistry and conjugation site were important parameters impacting acetate group stability. Higher degrees of stability translated to greater activity in xenograft models. ADCs were prepared with both dipeptide and glucuronide linkers as heterogeneous ADCs loaded at an average of 4-drugs/Ab (DAR 4), and as homogeneous ADCs loaded at 2-drugs/Ab (DAR 2) at the engineered S239C sites. Increased in vivo acetate stability was observed for glucuronide-based conjugates relative to the dipeptide analogue. Further stabilization was achieved by conjugating at the S239C sites (DAR 2) relative to native antibody cysteines (DAR 4). Tubulysin M acetate stabilization by both linker and conjugation site selection resulted in increased ADC potency in preclinical models and, combined, serves as an enabling strategy for antibody-mediated tubulysin M drug delivery.

Citation Format: Patrick J. Burke, Joseph Z. Hamilton, Thomas A. Pires, Christopher I. Leiske, Julia H. Cochran, Jocelyn R. Setter, Kim K. Emmerton, Andrew B. Waight, Peter D. Senter, Robert P. Lyon, Scott C. Jeffrey. Impact of linker and conjugation site on tubulysin M ADC stability and in vivo activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1649.
Abstract 1651: In vitro and in vivo profile of the preclinical candidate and MPS1 kinase inhibitor CCT289346
Authors: Woodward Hannah , Innocenti Paolo , Cheung Kwai-Ming J. , Naud Sébastien , Faisal Amir , Mak Grace W. , Hayes Angela , O'Fee Lisa , Saville Harry , Brandon Alexis De Haven , Roberts Jennie , Box Gary , Valenti Melanie , Heneley Alan T. , Walsh Katie , Burke Rosemary , Eccles Suzanne A. , Raynaud Florence I. , Montfort Rob L. van , Blagg Julian , Linardopoulos Spiros , Hoelder Swen
Published: August 2018

The mitotic kinase MPS1 (also known as TTK) is one of the main components of the spindle assembly checkpoint. MPS1 is required for chromosome alignment and kinetochore-microtubule error correction. Cancer cells are dependent on MPS1 to cope with chromosomal instability resulting from aberrant numbers of chromosomes. Moreover, MPS1 has been found to be deregulated in a large number of tumor types. MPS1 kinase inhibitors induce cancer cells to prematurely exit mitosis with incorrectly attached and unaligned chromosomes, causing severe chromosome mis-segregation, aneuploidy and cell death. These data stimulated us to pursue MPS1 as a cancer target. Extensive work by us and other groups has shown that MPS1 inhibitors are effective against a variety of cancers, particularly when used in combination with other drugs, such as paclitaxel. Here we disclose CCT289346, an MPS1 inhibitor currently completing late stage preclinical development. We describe the final stages of chemical optimisation and the data driven selection and nomination of CCT289346 as our preclinical candidate. We report key in vitro and in vivo preclinical data such as kinase profiling, PK in mouse, rat and dog, PK/PD relationship and efficacy in different in vivo models.

Citation Format: Hannah Woodward, Paolo Innocenti, Kwai-Ming J. Cheung, Sébastien Naud, Amir Faisal, Grace W. Mak, Angela Hayes, Lisa O'Fee, Harry Saville, Alexis De Haven Brandon, Jennie Roberts, Gary Box, Melanie Valenti, Alan T. Heneley, Katie Walsh, Rosemary Burke, Suzanne A. Eccles, Florence I. Raynaud, Rob L. van Montfort, Julian Blagg, Spiros Linardopoulos, Swen Hoelder. In vitro and in vivo profile of the preclinical candidate and MPS1 kinase inhibitor CCT289346 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1651.
Abstract 1654: Chemical and structure-guided optimization of BAX trigger site activators for cancer therapy
Authors: Reyna Denis E. , Kopp Felix , Gavathiotis Evripidis
Published: August 2018

Cancer cells evade mitochondrial apoptosis most commonly by over-expressing anti-apoptotic BCL-2 proteins that suppress the activation of pro-apoptotic proteins. Pro-apoptotic BAX is a critical BCL-2 family effector protein that upon death stimuli is activated and induces mitochondrial outer membrane permeabilization that leads to cell death. The vast majority of cancer cells express wild-type BAX, therefore, direct BAX activation represents a potential therapeutic strategy in cancer. We previously presented the discovery of BTSA1 as a selective BAX activator with anti-leukemic activity and provided proof-of-concept for BAX as a druggable target in acute myeloid leukemia (AML) (Reyna et al, Cancer Cell, 2017). Here, we investigated chemical optimization of BTSA1 core scaffold to rationally optimize the potency and efficacy of the BAX trigger site activators in AML models. We designed compounds using a structural model of BTSA1 bound to the BAX trigger site that also fits with the previous established pharmacophore model. Synthesized compounds were evaluated for binding potency using a fluorescence polarization binding assay that measures compound competition of the fluorescein-label stapled BIM BH3 peptide from the BAX trigger site. Compounds with improved binding activity were evaluated for their potency in biochemical assays of BAX activation using isolated liposomal and mitochondrial membrane assays. Improved binding to BAX of BTSA1 derivatives correlated with better activation capacity. Moreover, compounds were evaluated for their efficacy in induction of BAX-mediated cell death in AML cell lines including Venetoclax-resistant cells. We have identified compounds with similar or superior cellular efficacy compared to BTSA1. Compounds were evaluated for ADME and in vivo pharmacokinetic profiles and these are compared to efficacy and pharmacodynamics data using in vivo AML xenograft models. Our work highlights our strategy for chemical and structure-guided optimization of BTSA1 demonstrating the potential for improved therapeutic induction of apoptosis in AML by BAX Trigger Site Activators.

Citation Format: Denis E. Reyna, Felix Kopp, Evripidis Gavathiotis. Chemical and structure-guided optimization of BAX trigger site activators for cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1654.