bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A18: High-content 3D image analysis of patient-derived organoids
Authors: Spiller Erin , Lau Roy , Choung Sarah , Mumenthaler Shannon M.
Published: March 2017

Organoids are an emerging model system that more closely recapitulates the in vivo environment compared to traditional monolayer systems. Using patient-derived samples grown in a 3D matrix, we hope to advance personalized medicine by providing more patient-specific treatment plans. We have begun building a patient-derived organoid library from primary colon cancer tumors and metastases. Once cells are isolated and seeded into a Basement Membrane Extract (BME) matrix we analyze features on a high content imaging platform, the Operetta (Perkin Elmer). Using the Operetta we are able to obtain quantitative phenotypic data that includes the number, size, and morphology of organoids. In addition, we can capture the dynamics of organoid formation, growth, and death by tracking the same organoids over time using live-cell imaging.

More specifically, for each organoid, we capture z-stack images at various heights, and using the Harmony software, we perform quantitative analysis on the maximum projection of the complied images. Using a building block approach, we created a workflow that first relied on identifying regions of interest (ROI), which correspond to organoids. In each ROI we then obtained information such as geometric center, length and width measurements, and texture features. We used this information to determine inter-patient heterogeneity, as well as differences across samples isolated from the primary location (i.e. colon) versus metastatic sites (i.e. liver). After culturing patient-derived organoids for several weeks in laboratory conditions, we perturbed various aspects of the tumor microenvironment (e.g. drug and oxygen levels) and subsequently tracked changes in organoid number, size, and morphology over time. This workflow represents a unique method for image-based quantitative phenotypic analysis of organoids under varying environmental conditions.

Citation Format: Erin Spiller, Roy Lau, Sarah Choung, Shannon M. Mumenthaler. High-content 3D image analysis of patient-derived organoids. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr A18.
Abstract IA10: Renal cancer tumorgrafts: A validated model to evaluate novel targeted therapies
Authors: Pavia-Jimenez Andrea , Chen Wenfang , Hill Haley , Christie Alana , Yousuf Qurratulain , Williams Noelle , Xie Xian-Jin , Kapur Payal , Brugarolas James
Published: March 2017

Renal cell carcinoma (RCC) is diagnosed in over 50,000 Americans annually and in the metastatic setting remains largely incurable. The development of new drugs is hampered by the lack of suitable preclinical animal models reproducing the characteristics of human tumors. To date, we have implanted tumors from over 700 kidney cancer patients orthotopically in mice. Our models include common tumor types, such as clear cell and papillary, but also rare tumor types such as translocation carcinomas and fumarate hydratase (FH)-deficient renal tumors from HLRCC patients. We have shown that tumor fragments implanted orthotopically into NOD/SCID mice result in tumors that reproduce: (i) histology, (ii) gene expression, (iii) DNA copy number alterations and (iv) mutations. Tumorgraft (TG)-bearing mice also reproduce paraneoplastic syndromes (paraneoplastic hypercalcemia). Furthermore, TGs recapitulate the treatment responsiveness of RCC in patients. We performed pharmacokinetic studies to determine appropriate administration regimens resulting in comparable exposures. TGs responded to drugs effective against RCC in patients (sunitinib and sirolimus, which accounts for 70% of the circulating drug after temsirolimus administration) and were resistant to ineffective drugs (erlotinib). TGs provide a superb platform to evaluate promising novel therapeutics. Data will be presented on the application of our TG platform to the evaluation of a first-in-class inhibitor of arguably the most important driver of RCC, the HIF-2 transcription factor. TGs were used to assess activity, validate a putative pharmacodynamic biomarker, identify biomarkers of sensitivity, and to assess the development of resistance. By leveraging a large and diverse TG platform, preclinical drug testing may be rigorously conducted, the most promising agents may be identified, and future clinical development may be effectively informed.

Citation Format: Andrea Pavia-Jimenez, Wenfang Chen, Haley Hill, Alana Christie, Qurratulain Yousuf, Noelle Williams, Xian-Jin Xie, Payal Kapur, James Brugarolas. Renal cancer tumorgrafts: A validated model to evaluate novel targeted therapies. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr IA10.
Abstract IA14: PDX models in head and neck cancers: Present status and future application for immune-oriented biology and therapy development
Authors: Jimeno Antonio
Published: March 2017

Targeted therapy development in head and neck squamous cell carcinoma (HNSCC) is challenging given the rarity of activating mutations. Additionally, HNSCC incidence is dramatically increasing related to human papillomavirus (HPV). We sought to develop an in vivo model derived from patients reflecting the evolving HNSCC epidemiologic landscape, and use it to identify new therapies. Primary and relapsed tumors from over 100 HNSCC patients, both HPV+ and HPV-, have been implanted on mice, giving rise to over 50 strains. Resulting xenografts were characterized by detecting key mutations, measuring protein expression by IHC and gene expression/pathway analysis by mRNA-sequencing. The genetic makeup of these tumors was consistent with the frequencies of TP53, PI3KCA, NOTCH1, and NOTCH2 mutations found in landmark genomic characterization of large HNSCC patient cohorts. Tumors maintained their original morphology, genetic profiles, and drug susceptibilities through serial passaging. Cancer cells deceive the host to create an advantageous microenvironment and evade immune surveillance. A limitation to studying human cancer is the limited availability of in vivo models with appropriate human stroma and immunity. To address this unmet need, we generated an allogeneic humanized mouse (aHM) model where HNSCC were implanted on NOD/SCID/IL2rg-/- (NSG) mice reconstituted with human hematopoietic stem cells (HSCs) and other precursors from an unrelated donor. In aHM, bone marrow-derived human immune and mesenchymal cells homed into the growing tumor and interacted with human cancer cells as they would in a patient, partially reversing the gene expression drift that occurs after tumor passage on immune compromised mice. Further work is aimed at improving upon this platform. In summary, we have developed a HNSCC model covering its clinical spectrum whose major genetic alterations and susceptibility to anticancer agents represent contemporary HNSCC. Then, we have generated an aHM model to enable immune biology and therapy exploration in HNSCC. These models will enable prospectively testing therapeutic-oriented hypotheses leading to enhanced precision medicine, especially in the field of immune therapy.

Citation Format: Antonio Jimeno. PDX models in head and neck cancers: Present status and future application for immune-oriented biology and therapy development. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr IA14.
Abstract IA19: Phase II human clinical trials in mice-modeling inter-patient response heterogeneity in PDXs
Authors: Gao Hui
Published: March 2017

The current strategy of profiling candidate therapeutics using a limited set of cancer models during pre-clinical development rarely enables accurate predictions of clinical efficacy, or the identification of factors underlying heterogeneous patient responses that inform clinical patient selection strategies. Novartis established ~1000 patient-derived tumor xenograft models (PDXs), where mutation and copy number based genomic landscape analyses revealed a relatively broad conservation of driver mutations found in human cancers. Using this panel of PDXs, we performed population-based in vivo compound screens using a 1x1x1 experimental design (PDX clinical trial or PCT). We demonstrate both the reproducibility and translatability of this approach by identifying associations between genotype and drug response, and clinically established mechanisms of resistance. We therefore propose a new experimental paradigm that can potentially revolutionize preclinical evaluation of novel treatment modalities and improve our ability to predict clinical trial response.

Citation Format: Hui Gao. Phase II human clinical trials in mice-modeling inter-patient response heterogeneity in PDXs. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr IA19.
Abstract IA27: Small cell lung cancer circulating tumor cell-derived explants: A new preclinical platform for examining drug response
Authors: Frese Kristopher K. , Morrow Christopher , Miller Crispin , Blackhall Fiona , Dive Caroline
Published: March 2017

Small cell lung cancer (SCLC) is a highly aggressive disease with a 5 year survival of 5% that has remained relatively unchanged for the past several decades. Despite a robust response the first line chemotherapy, 95% of patients relapse within a few months. Due in part to the paucity of SCLC tissue samples available for preclinical studies, the molecular mechanisms unpinning disease relapse and acquired drug resistance are poorly understood. Non-invasive collection of liquid biopsies have proven useful in for tumor diagnosis and monitoring, however they have traditionally provided limited utility in mechanistically interrogating drug sensitivity. We have recently demonstrated that SCLC circulating tumor cells (CTCs) are tumorigenic in mice and the resulting patient CTC-derived explants (CDX) mirror patients' disease. We have developed a library CDX where CTCs were enriched either at baseline or upon progression from chemosensitive or chemorefractory SCLC patients. Strikingly, these CDX faithfully recapitulate the histological, genomic, and chemoresistance aspects of patient tumors, suggesting they may be a suitable platform for identifying new drug targets, testing novel drugs, and exploring drug resistance mechanisms. Studies underway are aimed at providing mRNA and genomic DNA analysis from the same CTC or CDX sample to establish direct linkage between mutational changes and emergent mRNA profiles to gain insights into mechanisms underpinning treatment responses.

Citation Format: Kristopher K. Frese, Christopher Morrow, Crispin Miller, Fiona Blackhall, Caroline Dive. Small cell lung cancer circulating tumor cell-derived explants: A new preclinical platform for examining drug response. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr IA27.
Abstract A09: A bioinformatics workflow to identify neoantigens using next-generation sequencing
Authors: Zhang Shile , So Alex , Kaplan Shannon , Kruglyak Kristina
Published: March 2017

Immuno-oncology is an emerging field of research that represents tremendous promise for cancer patients. Accumulating evidence suggests that tumor-specific neoantigens can persistently stimulate cytotoxic T lymphocytes, which may eradicate cancer cells in a favorable immune microenvironment. Next-generation sequencing (NGS) is a key technology enabling advances in this field. Here we present a complete bioinformatics workflow using tumor/normal whole-exome sequencing (WES) and tumor-only whole-transcriptome sequencing (WTS) to predict neoantigens presented by cancer cells. We evaluate existing WES probe designs for MHC class I genes and show that accurate four-digit HLA typing can be achieved using standard WES solutions, thus allowing for a simplified experimental workflow by removing the need for targeted HLA assays. Final neoantigen candidates are prioritized based on in silico peptide-HLA binding predictions, variant allele frequencies for clonal somatic mutations, tumor RNA expression, and self-similarities between mutant peptide to wildtype peptide. This workflow will be available through BaseSpace® Sequence Hub, which can be accessed via an intuitive web-based interface or Linux-based command line tool, providing any researcher with rapid analysis of NGS data and neoantigen predictions.

Citation Format: Shile Zhang, Alex So, Shannon Kaplan, Kristina Kruglyak. A bioinformatics workflow to identify neoantigens using next-generation sequencing. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A09.
Abstract A37: CBP501 induces tumor immunogenic cell death and CD8 T cell infiltration into tumor in combination with platinum, thereby increasing the efficacy of immune checkpoint inhibitors against tumors in mice
Authors: Sakakibara Keiichi , Sato Takuji , Kufe Donald W. , VonHoff Daniel D. , Kawabe Takumi
Published: March 2017

Introduction: CBP501, a calmodulin-binding peptide, is an anti-cancer drug candidate that has completed two Phase II clinical trials for patients with malignant pleural mesothelioma and non-small cell lung carcinoma (NSCLC). CBP501 enhances the uptake of platinum agents by cancer cells specifically and induces immunogenic cell death within the clinically achievable dose levels of both CBP501 and cisplatin (CDDP) (EORTC 2015 abstract#C106). Here we treated tumor bearing mice with the combination of CBP501, platinum agents (CDDP and carboplatin), and immune checkpoint inhibitors and analyzed the tumor growth and behaviors of immune cells.

Methods: Immuno-competent BALB/c mice were subcutaneously inoculated with CT26WT (5 x105 cells/mouse). A week later, mice were apportioned into 8 groups (6 mice/group) and treated with 2 dosing cycles for 3 anti-cancer agents, alone or in different combinations [CDDP: 4 mg/kg x 1/week, CBP501: 6.0 mg/kg x 1/week, anti-mPD1 mAb (RMP1-14) or anti-PD-L1 mAb (10F.9G2): 200 ug/mice x 2/week].

Results: In BALB/c mice bearing CT26WT tumor, CDDP- or immune checkpoint inhibitor-treated mice showed a minimal anti-tumor effect compared to the vehicle-treated mice. CDDP/immune checkpoint inhibitor-treated mice showed an additive anti-tumor effect. CBP501 significantly enhanced the anti-tumor effect of CDDP/immune checkpoint inhibitor combination. Flow cytometry analysis of single suspension cells from the tumor tissue showed that treatment of CDDP/CBP501 or CDDP/CBP501/anti-mPD1 mAb but not CDDP or anti-mPD1 mAb increased the percentage of tumor-infiltrating CD8+ T cell.

Conclusion: An increase of tumor-infiltrating CD8+ T cells in CDDP/CBP501-treated mice supported the conclusion that CDDP/CBP501 treatment induced immunogenic cell death not only in vitro but also in vivo. It also supported the notion that CBP501 would not increase cytotoxicity of platinum agents to non-tumor cells. The fact that CBP501 enhanced anti-tumor activity of immune checkpoint inhibitors in combination with platinum agents (CDDP and carboplatin) would support further clinical development of CBP501.

Citation Format: Keiichi Sakakibara, Takuji Sato, Donald W. Kufe, Daniel D. VonHoff, Takumi Kawabe. CBP501 induces tumor immunogenic cell death and CD8 T cell infiltration into tumor in combination with platinum, thereby increasing the efficacy of immune checkpoint inhibitors against tumors in mice. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A37.
Abstract B32: Loss of MK2 in tumor stroma promotes aggressive tumor development in an advanced stage lung cancer model
Authors: Sriram Ganapathy , Suarez-Lopez Lucia , Yaffe Michael B.
Published: March 2017

MAPKAP Kinase-2 (MK2) is a serine/threonine protein kinase that is activated by p38MAPK in response to a variety of stresses including DNA damage, osmotic shock, thermal injury, and inflammation/innate immune signaling downstream of TLR4 activation. MK2 controls the synthesis of many pro-inflammatory cytokines, modulates the actin cytoskeleton in stressed cells, and is responsible for maintaining both the G1/S and G2/M cell cycle checkpoints in p53-defective tumor cells after genotoxic damage. We recently created a “Cre-reversible" MK2 knock-out mouse model that allows the function of MK2 within either the tumor cells or within the tumor microenvironment to be studied independently. We found that MK2-deficient tumor cells in an MK2 wild-type stroma were dramatically sensitized to the anti-tumor effects of cisplatinum and doxorubicin (Morandell et al., Cell Reports, 2013). Here, we report that loss of MK2 in the tumor stroma with preservation of MK2 in tumor cells results in an aggressive tumor-promoting phenotype in an immuno-competent transplant K-RasG12D/p53-/- model of NSCLC. A similar tumor aggressive phenotype was observed in Tie2-Cre-driven endothelial and bone marrow-specific MK2-knockout mice compared to wild-type controls. Tumor-bearing mice with MK2-null stroma display increased total levels of MIP-1α, G-CSF, GM-CSF and IL-6 in the lung. Myeloperoxidase immunohistochemistry reveals enhanced tumor infiltration by neutrophils in these stromal MK2-deficient animals, consistent with the cytokine profile. In contrast with loss of MK2 in the stroma, direct activation of the p38MAPK/MK2 pathway in the tumor microenvironment by weekly intra-tracheal LPS exposure dramatically reduces tumor formation in an autochthonous K-RasG12D/p53-/- murine NSCLC model. Collectively, these results suggest that MK2 signaling in the endothelium and/or immune cells suppresses NSCLC tumor progression and modulates the inflammatory milieu in the tumor microenvironment.

Citation Format: Ganapathy Sriram, Lucia Suarez-Lopez, Michael B. Yaffe. Loss of MK2 in tumor stroma promotes aggressive tumor development in an advanced stage lung cancer model. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr B32.
Abstract B45: Short-term EGFR blockade enhances immune-mediated cytotoxicity of EGFR mutant lung cancer cells: Rationale for combination therapies
Authors: Dominguez Charli , Tsang Kwong-Yok , Palena Claudia
Published: March 2017

The epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor erlotinib has been approved for years as a first-line therapy for patients harboring EGFR sensitizing mutations. With the promising implementation of immunotherapeutic strategies for the treatment of lung cancer, there is a growing interest in developing combinatorial therapies that could utilize immune approaches in the context of conventional or targeted therapies. Tumor cells are known to evade immune attack by multiple strategies, including undergoing phenotypic plasticity via a process designated as the epithelial-mesenchymal transition (EMT). As signaling through EGFR is a major inducer of EMT in epithelial cells, we have investigated the effect of EGFR inhibition with erlotinib on tumor phenotype and susceptibility to immune attack. Utilizing a panel of lung carcinoma cell lines with mutated EGFR, we have evaluated the ability of erlotinib to modulate tumor phenotype and to modulate cytotoxic responses to effector NK cells and T cells specific for brachyury and MUC-1. The effect of erlotinib treatment was also evaluated with single cell clonal populations of lung cancer cells isolated based on their expression of epithelial and mesenchymal markers. Our data shows that short-term exposure of tumor cells to low dose erlotinib leads to a remarkable enhancement of tumor lysis mediated by innate NK cells and antigen-specific T cells. This effect positively correlated with the ability of short-term EGFR blockade to modulate tumor phenotype towards a more epithelial one, as well as altered apoptotic signaling through enhanced caspase cleavage. The effect, however, was lost when erlotinib was utilized for long periods of time in vitro or in vivo, which resulted in gain of mesenchymal features and decreased (rather than increased) tumor lysis in response to immune effector mechanisms. Additionally, with the utilization of single cell clones, it was evident that there was a selection of mesenchymal populations during erlotinib treatment, which could explain the acquired resistance to immune attack seen at longer time points. Our data provides rationale for potential combinations of erlotinib and immunotherapies for the treatment of lung carcinomas in the early setting, prior to the establishment of tumor relapse with long-term EGFR inhibition.

Citation Format: Charli Dominguez, Kwong-Yok Tsang, Claudia Palena. Short-term EGFR blockade enhances immune-mediated cytotoxicity of EGFR mutant lung cancer cells: Rationale for combination therapies. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr B45.
Abstract B47: Stabilization of programmed death ligand-1 by epidermal growth factor enhances cancer cell immune escape
Authors: Lim Seung-Oe , Li Chia-Wei , Hung Mien-Chie
Published: March 2017

Extracellular interaction between programmed death ligand-1 (PD-L1) and programmed cell death protein-1 (PD-1) leads to tumor-associated immune escape. Despite efforts to limit the interaction between cancer cells and immune cells, the molecular mechanism by which cancer cells initiate immune escape is less understood. Here, we show that the immunosuppression activity of PD-L1 is stringently modulated by the spatial competition between ubiquitination and N-glycosylation. We identified glycogen synthase kinase 3β (GSK3β) as a novel protein that interacts with PD-L1 and can induce phosphorylation-dependent proteasome degradation by β-TrCP. In depth analysis of PD-L1 N192, N200, and N219 glycosylation suggests glycosylation antagonizes GSK3β binding. In this regard, only non-glycosylated PD-L1 forms a complex with GSK3β and β-TrCP. We also demonstrate that epidermal growth factor (EGF) stabilizes PD-L1 via GSK3β inactivation in basal-like breast cancer (BLBC). Inhibition of EGF signaling by gefitinib destabilizes PD-L1, enhances antitumor T cell immunity, and sensitizes BLBC cells to PD-1 blockade. Together, our results link ubiquitination and glycosylation pathways with stringent regulation of PD-L1, which could open new therapeutic strategies to enhance cancer immune therapy efficacy.

Citation Format: Seung-Oe Lim, Chia-Wei Li, Mien-Chie Hung. Stabilization of programmed death ligand-1 by epidermal growth factor enhances cancer cell immune escape. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr B47.
Abstract B49: Sustained IFNalpha treatment inhibits metastasis in 4T1 mammary carcinoma and correlates with decreasing myeloid-derived suppressor cells
Authors: Kochel Christina , Pande Kalyan , Ueda Roanna , Gorman Dan , Laface Drake
Published: March 2017

In order to model pharmacokinetics and pharmacodynamics of pegylated IFNalpha, we utilized hydrodynamic delivery technology using minicircle DNA vectors encoding universal IFNalpha as a rapid and cost-effective method to obtain sustained levels of serum IFNalpha2alpha1. This approach is particularly useful for delivering secreted molecules, such as cytokines, as transfected hepatocytes secrete the transgene product in a paracrine manner providing a serum exposure profile resembling that reported in PEG-intron-treated patients. We found that sustained exposure of IFNalpha2alpha1 induced inhibition of metastasis. Primary 4T1 tumor growth was partially inhibited, while significant decreases in pulmonary metastatic burden were consistently observed. Further evidence that the observed inhibition of metastasis was independent of potential effects on the primary tumor was tested by resecting the primary tumor prior to treatment, which did not alter the outcome. Importantly, sustained IFNalpha2alpha1 treatment correlated with a significant increase in the survival of tumor-bearing animals. IFNalpha effects are plieotropic resulting in potential activation of numerous immune cell types associated with both innate and adaptive immunity. However, the mechanisms associated with IFNalpha-induced inhibition of lung metastasis were largely engendered by innate immune cells since treatment in BALB/c mice deficient for T and B cells did not affect the capacity of IFNalpha to inhibit pulmonary metastasis when compared to wild-type animals. The importance of NK cells has recently been reported as an important innate component for IFNalpha induced reduction of metastatic burden. However, little is known about IFNalpha-regulation of other innate cell types that may contribute to the inhibition of tumor metastasis. Here we report IFNalpha2alpha1 treatment induced marked decreases in 4T1-induced expansion and activation of immature CD11b+ Gr-1+ myeloid cells (aka: MDSC), which have been reported to contribute to metastatic potential. These results correlated with similar decreases in metastatic burden in mice following chlodronate-liposome depletion of CD11b+ Gr-1+ myeloid cells.

Citation Format: Christina Kochel, Kalyan Pande, Roanna Ueda, Dan Gorman, Drake Laface. Sustained IFNalpha treatment inhibits metastasis in 4T1 mammary carcinoma and correlates with decreasing myeloid-derived suppressor cells. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr B49.
Abstract B63: Therapeutic potential of GPR64 specific CAR T cells in pediatric sarcomas
Authors: Storz Isabel , Schirmer David , Wisskirchen Karin , Feederle Regina , Schmidt Oxana , Abken Hinrich , Protzer Ulrike , Burdach Stefan , Richter Günther HS
Published: March 2017

Ewing Sarcoma (ES) is the second most common bone malignancy in children and young adolescents with a high potential of dissemination into lung and bones. Patients with localized disease receiving current treatment, have an approximate long-term survival of >65%. Patients with disseminated disease into the bone have an approximate long-term survival rate of only 10%, compelling the search for new therapeutic treatment modalities like engineered T cell therapy. Here the therapeutic potential of chimeric antigen receptor (CAR) transgenic T cells directed against G-protein coupled receptor 64 (GPR64), an orphan receptor with normal expression restricted to human epididymis and significant overexpression in ES, was examined. Therefore, two different monoclonal antibodies (mAb) directed against the extracellular region of GPR64 were generated and characterized. Subsequently, retroviral constructs containing second generation CARs together with the scFv fragments of the respective mAbs were designed. Primary lymphocytes were transduced and tested in vitro via flow cytometry, ELISpot and xCelligence assay. Antibodies specifically stained ES cells as determined by flow cytometry. The signal intensity was reduced after RNAi mediated down-regulation of GPR64 in ES cell lines confirming specificity of mAbs. Following sequence determination of those mAbs two different CAR constructs were designed. Retroviruses containing such CARs transduced primary lymphocytes with good efficiency. The CAR transgenic T cells could be enriched for CD8+CAR+ cells via microbead isolation and showed strong proliferative capacities in vitro. Furthermore, target structures were specifically recognized as determined by ELISpot and xCelligence assays. CAR transgenic T cells targeting GPR64 show a promising approach to transfer the success of CARs in hematological malignancies to solid tumors. The cells generated in this study show strong specificity towards GPR64 and are able to control tumor cell growth in vitro. Since GPR64 expression is not restricted to ES but also up-regulated in a number of carcinomas derived from prostate, kidney or lung, GPR64-specific CARs may also be a future treatment option for other tumor entities.

Citation Format: Isabel Storz, David Schirmer, Karin Wisskirchen, Regina Feederle, Oxana Schmidt, Hinrich Abken, Ulrike Protzer, Stefan Burdach, Günther HS Richter. Therapeutic potential of GPR64 specific CAR T cells in pediatric sarcomas. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr B63.
Abstract IA20: PET imaging of the immune system using new nucleoside analog probes
Authors: Radu Caius G.
Published: March 2017

Similar to other major metabolic networks, nucleotide metabolism consists of redundant and convergent biosynthetic pathways. Thus, nucleotides can be produced both de novo, from glucose and amino acids, and by salvage pathways, via scavenging and re-utilization of preformed nucleosides and nucleobases. The differential utilization of de novo and salvage nucleotide biosynthetic pathways in normal and malignant is poorly understood. We are studying this process in lymphocytes and cancer cells using newly developed mass spectrometry and positron emission tomography (PET) approaches. Deoxycytidine kinase (dCK), a rate-limiting enzyme in the cytosolic deoxyribonucleoside salvage pathway, is significantly upregulated following T cell activation and therefore represents a promising PET imaging target. PET probes for dCK have been previously developed by our group and shown to be effective in mice but their specificity and sensitivity in humans remained suboptimal. Recently we have identified [18F]CFA (2-chloro-2′-deoxy-2′-[18F]fluoro-9-β-D-arabinofuranosyl-adenine) as a suitable third generation nucleoside analog probe for clinical dCK PET imaging. First-in-human [18F]CFA PET/CT studies showed probe accumulation in tissues with high dCK expression: e.g., hematopoietic bone marrow and secondary lymphoid organs. The potential use of [18F]CFA PET for imaging systemic immune activation induced by various cancer immunotherapies will be discussed.

Citation Format: Caius G. Radu. PET imaging of the immune system using new nucleoside analog probes. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr IA20.
Abstract IA24: T cell recognition and tumor resistance in human cancer
Authors: Schumacher Ton
Published: March 2017

The T cell-based immune system has evolved to recognize and destroy cells that differ from self on the basis of the antigens they express. Many human tumors contain large numbers of mutations, of which many hundreds can be present within expressed genes. As the resulting altered protein sequences are foreign to the immune system, immune recognition of such ‘neo-antigens’ has been proposed as a major factor in the activity of clinically used immunotherapeutics such as anti-CTLA-4 and anti-PD-1.

In support of this hypothesis, using in-house developed technologies we have demonstrated that both CD4 and CD8 T cell recognition of the consequences of DNA damage is a common feature in human melanoma and can be boosted by therapy. Furthermore, to extend these data to other tumor types, we have benchmarked the foreign antigenic space in a variety of human cancers against a series of human pathogens. Collectively, these data, and data from other groups, establish tumor foreignness as one of the critical determinants in effective cancer immunotherapy. However, the sensitivity of tumor cells to this T cell attack forms an obvious additional requirement. To address this issue, we are currently determining mechanisms of immune escape upon T cell targeting of human tumors. Results from this analysis are expected to contribute to the further definition of biomarkers that predict the outcome of the interaction between human tumors and the T cell-based immune system.

Citation Format: Ton Schumacher. T cell recognition and tumor resistance in human cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr IA24.
Roles of Ileal ASBT and OSTα-OSTβ in Regulating Bile Acid Signaling
Authors: Dawson Paul A.
Published: March 2017

Background: In addition to their classical role as detergents, bile acids function as signaling molecules to regulate gastrointestinal physiology, carbohydrate and lipid metabolism, and energy expenditure. The pharmacodynamic potential of bile acids is dependent in part on the tight pharmacokinetic control of their concentration and metabolism, properties governed by their hepatic synthesis, enterohepatic cycling, and biotransformation via host and gut microbiota-catalyzed pathways. Key Messages: By altering the normal cycling and compartmentalization of bile acids, changes in hepatobiliary or intestinal transport can affect signaling and lead to the retention of cytotoxic hydrophobic bile acids and cell injury. This review discusses advances in our understanding of the intestinal transporters that maintain the enterohepatic cycling of bile acids, signaling via bile acid-activated nuclear and G protein receptors, and mechanisms of bile acid-induced cell injury. Conclusions: Dysregulated expression of the Asbt and Ostα-Ostβ alters bile acid signaling via the gut-liver farnesoid X receptor-fibroblast growth factor 15/19 axis and may contribute to other bile acid-regulated metabolic and cell injury pathways.
Hair Cortisol Concentrations Are Associated with Hair Growth Rate
Authors: Xiang Lianbin , Sunesara Imran , Rehm Kristina E. , Marshall Jr. Gailen D.
Published: March 2017

There is a growing interest in hair cortisol concentrations as a valuable biomarker for the assessment of metabolic diseases and chronic psychological stress. Fifty-three volunteers were recruited, and hair segments proximal to the scalp were collected from each individual. A cost-effective ball mill was used for the preparation of hair samples, and ELISA was performed to analyze cortisol concentrations. Results indicate that the frequency of hair washing affects the hair cortisol concentration. The group that washed their hair every day had significantly lower cortisol concentrations than the group that washed it less often. However, no significant differences were detected between cosmetic-treated and nontreated hair samples. The study also shows that hair cortisol concentrations in the first 3 cm of hair segments proximal to the scalp corresponded to average hair growth rate based on 1 cm/month. Thus, hair cortisol concentrations of segments 3 cm proximal to the scalp may represent cumulative stress exposure over the previous 3 months. These findings will allow more widespread research to validate the utility of hair cortisol as a potential biomarker to assess chronic stress.
The Effect of apoM Polymorphism Associated with HDL Metabolism on Obese Korean Adults
Authors: Lee Myoungsook , Kim Jung-Im , Choi Seojin , Jang Yangsoo , Sorn Sungbin Richard
Published: March 2017

Background: Apolipoprotein M (apoM) is a recently identified apolipoprotein associated with high-density lipoprotein (HDL) in coronary artery disease (CAD), but the association between apoM polymorphism and obesity has not been reported. Aim: To investigate the association between apoM polymorphism and obesity prevalence in 584 Korean adults. Methods: A total of 584 individuals aged between 30 and 80 years were recruited from Yonsei Medical Center in Seoul, Korea, and divided into obese (OB; body mass index, BMI ≥25) and nonobese (non-OB; BMI <25) groups. Anthropometric variables, lipid profiles, insulin-resistant profiles, reverse cholesterol transport (RCT) enzymes, HDL subfraction, and apoM polymorphism were determined. Results: In OB with T-855C polymorphism, TT genotype carriers significantly showed 6.2% higher diastolic blood pressure (DBP), 1.3% lower amount of HDL2b subfraction, and 19.7% higher lecithin-cholesterol acyltransferase (LCAT) mass than TC+CC carriers. OB subjects with the T allele of T-778C polymorphism significantly demonstrated 43% higher plasma insulin, 17.7% higher total cholesterol, 26.7% higher triglyceride, 40.7% higher leptin, 1.6% lower HDL2b, and 12.6% higher LCAT mass than those with the C allele. These results were reversed in non-OB with T-778C polymorphism regarding HDL subfractions and RCT enzymes. Conclusion: apoM T-855C and T-778C polymorphisms were found to be associated with obesity by regulating HDL metabolism, and the T alleles of apoM T-778C were shown to be more strongly correlated.
Effects of Dietary Supplementation with Sea Buckthorn (Hippophae rhamnoides L.) Seed Oil on an Experimental Model of Hypertensive Retinopathy in Wistar Rats
Authors: Bouras Konstantinos , Kopsidas Konstantinos , Bariotakis Michael , Kitsiou Paraskevi , Kapodistria Katerina , Agrogiannis Giorgos , Vergados Ioannis , Theodossiadis Panagiotis , Perrea Despoina
Published: March 2017

Background/Aims: Sea buckthorn (Hippophae rhamnoides L.) oil is a rich source of phytosterols, flavonoids, unsaturated fatty acids, and carotenoids, known for their antioxidant and neuroprotective activity. In this study, we investigated the neuroprotective and antioxidant effect of sea buckthorn oil on rat retina in hypertensive retinopathy. Methods: Twenty-eight male 6-month-old Wistar rats were separated into 3 groups: (1) controls, (2) unilateral nephrectomized rats receiving drinking water with 1% NaCl, (3) unilateral nephrectomized rats receiving 0.5 mL sea buckthorn oil and drinking water with 1% NaCl. Systemic pressures were being measured with the tail-cuff method. The antiapoptotic effect of sea buckthorn was determined by measuring glial fibrillary acidic protein (GFAP), cleaved caspase-3, and glutamine synthetase levels with immunohistochemistry and Western blot. Results: Nephrectomy and salt intake caused increases in both systolic and diastolic pressures. Both types of analysis showed that group 2 had statistically significant increases in the expression of GFAP and cleaved caspase-3, while group 3 showed no significant differences compared with the control group. The expression of glutamine synthetase showed no significant differences between the 3 groups. Conclusions: Our findings suggest that sea buckthorn could notably protect the retina from damage induced by hypertensive retinopathy.
Clinical Factors Predicting and Stratifying the Risk of Lymph Node Invasion in Localized Prostate Cancer
Authors: Porcaro Antonio Benito , de Luyk Nicolò , Corsi Paolo , Sebben Marco , Tafuri Alessandro , Mattevi Daniele , Processali Tania , Cerruto Maria A. , Brunelli Matteo , Zecchini Antoniolli Stefano , Siracusano Salvatore , Artibani Walter
Published: March 2017

Purpose: The study aimed to investigate clinical factors associating with occult lymph node micrometastases (pN1 disease) in a contemporary cohort of organ-confined prostate cancer (PCA) patients staged as cN0. Materials and Methods: The study evaluated 184 consecutive patients. Associations of clinical factors with pN1 disease were assessed by multivariate logistic regression analysis. Results: Lymph node invasion was detected in 33 cases (17.9%). Independent factors associating with pN1 status were prostate specific antigen (PSA; OR 1.054; p = 0.004), percentage of positive biopsy cores (PPC; OR 1.030; p = 0.013), and biopsy Gleason pattern (bGP) >4 + 3 (OR 3.666; p = 0.004). A clinical model predicting the risk of pN1 disease identified 4 prognostic groups of pN1 disease. Conclusions: In a contemporary cohort of PCA patients, lymph node invasion was detected in 17.9% of cases. An independent clinical disease showed that the risk of lymph node invasion was directly proportional to PPC and more stratification of the risk of pN1 disease was operated by PSA and BGP. The model allowed the stratification of the patient population in 4 groups and showed that the risk of lymph node invasion progressively increased as the risk group ranked from 1 to 4.
Cashew Nut Positioning during Stone Tool Use by Wild Bearded Capuchin Monkeys (Sapajus libidinosus)
Authors: Falótico Tiago , Luncz Lydia V. , Svensson Magdalena S. , Haslam Michael
Published: March 2017

Wild capuchin monkeys (Sapajus libidinosus) at Serra da Capivara National Park, Brazil, regularly use stone tools to break open cashew nuts (Anacardium spp.). Here we examine 2 approaches used by the capuchins to position the kidney-shaped cashew nuts on an anvil before striking with a stone tool. Lateral positioning involves placing the nut on its flatter, more stable side, therefore requiring less attention from the monkey during placement. However, the less stable and never previously described arched position, in which the nut is balanced with its curved side uppermost, requires less force to crack the outer shell. We observed cashew nut cracking in a field experimental setting. Only 6 of 20 adults, of both sexes, were observed to deliberately place cashew nuts in an arched position, which may indicate that the technique requires time and experience to learn. We also found that use of the arched position with dry nuts, but not fresh, required, in 63% of the time, an initial processing to remove one of the cashew nut lobes, creating a more stable base for the arch. This relatively rare behaviour appears to have a complex ontogeny, but further studies are required to establish the extent to which social learning is involved.
Association between Basal Total Testosterone Levels and Tumor Upgrading in Low and Intermediate Risk Prostate Cancer
Authors: Porcaro Antonio Benito , De Luyk Nicolò , Corsi Paolo , Sebben Marco , Tafuri Alessandro , Processali Tania , Cerasuolo Mattia , Mattevi Daniele , Cerruto Maria A. , Brunelli Matteo , Siracusano Salvatore , Artibani Walter
Published: March 2017

Purpose: The study aimed to evaluate associations of basal levels of total testosterone (TT) with tumor upgrading to high risk disease in low-intermediate risk prostate cancer (PCA). Materials and Methods: We retrospectively evaluated the records of 135 patients undergoing radical prostatectomy. Evaluated factors included age, body mass index, prostate specific antigen (PSA), TT, prostate volume, PSA density (PSAD), proportion of biopsy positive cores (P+), clinical tumor stage, and biopsy grading system (1 or 2). Factors associating with tumor upgrading were investigated by the multivariate logistic regression analysis. Results: Tumor upgrading rate to high risk disease was 8.9%. TT, PSA, and PSAD were associated with tumor upgrading. On multivariate analysis, independent factors predicting tumor upgrading were PSA (OR 1.324; p = 0.001) and TT (OR 1.005; p = 0.015). Basal TT was dichotomized up to the third quartile (TT > q3) vs. TT ≤ q3 (426.0 ng/dL). The assessed tumor upgrading risk model showed that TT dichotomized to third quartile (TT > q3 vs. TT ≤ q3) stratified the risk of tumor upgrading (OR 6.577; p = 0.010) along increasing levels of PSA (OR 1.3; p < 0.0001). Conclusions: Low and intermediate risk PCA patients show a not negligible risk of tumor upgrading to high risk disease. In this particular subset of patients, basal levels of TT stratify the risk of tumor upgrading.
Childhood Estimates of Glomerular Filtration Rate Based on Creatinine and Cystatin C: Importance of Body Composition
Authors: Miliku Kozeta , Bakker Hanneke , Dorresteijn Eiske M. , Cransberg Karlien , Franco Oscar H. , Felix Janine F. , Jaddoe Vincent W.V.
Published: March 2017

Background: Creatinine and cystatin C concentrations are commonly used to estimate glomerular filtration rate (eGFR) in clinical practice and epidemiological studies. To estimate the influence of different body composition measures on eGFR from creatinine and cystatin C blood concentrations, we compared the associations of different anthropometric and body composition measures with eGFR derived from creatinine (eGFRcreat) and cystatin C (eGFRcystC) blood concentrations. Methods: In a population-based cohort study among 4,305 children aged 6.0 years (95% range 5.7-8.0), we measured weight and height and calculated body mass index (BMI) and body surface area (BSA), and lean and fat mass using dual-energy X-ray absorptiometry. At the same age, we measured creatinine and cystatin C blood concentrations and estimated the GFR. Results: Correlation between eGFR based on creatinine and cystatin C concentrations was r = 0.40 (p value <0.01). Higher BMI was associated with lower eGFRcystC but not with eGFRcreat. Higher BSA was associated with higher eGFRcreat and lower eGFRcystC (p value <0.05). Lean and fat mass percentages were associated with eGFRcreat but not with eGFRcystC. Conclusion: Our findings suggest that both eGFRcreat and eGFRcystC are influenced by BMI and BSA. eGFRcreat is more strongly influenced by body composition than eGFRcystC.
LAPTM4B Predicts Axillary Lymph Node Metastasis in Breast Cancer and Promotes Breast Cancer Cell Aggressiveness in Vitro
Authors: Xiao Min , Yang Shanshan , Meng FanLing , Qin Yu , Yang Yue , Jia Shusheng , Cai XiuMing , Li Changli , Huang Yuanxi , Ning Xiaoming
Published: March 2017

Purpose: Lysosome-associated protein transmembrane-4 beta (LAPTM4B) is associated with the prognosis of several human malignancies. In this study, the role of LAPTM4B in the metastatic potential of breast cancer (BC) and its underlying molecular mechanisms were investigated. Methods: The relationship between LAPTM4B expression and axillary lymph node metastasis was determined in 291 BC specimens by immunohistochemistry. The expression of LAPTM4B in paired BC cells was overexpressed and inhibited to analyse the role of LAPTM4B in the aggressiveness of BC. Cell proliferation, migration and invasion were assessed in vitro. Metastasis-related protein levels were detected through Western blot. Results: Immunohistochemical staining demonstrated that high expression level of LAPTM4B was independently associated with axillary lymph node metastasis (odds ratio=2.428; 95%CI=1.333- 4.425; P=0.004). The LAPTM4B inhibition in MCF-7 cells inhibited cell proliferation, migration, invasion, and resulted in simultaneous downregulation of phosphorylated N-cadherin, vimentin, and upregulation of E-cadherin. By contrast, the LAPTM4B overexpression promoted cell proliferation, migration, invasion, and led to simultaneous upregulation of N-cadherin, vimentin, and downregulation of E-cadherin in T47D cells. Conclusions: High expression level of LAPTM4B predicts tumor metastatic potential in patients with BC. Our results provide the first evidence of the role of LAPTM4B as an Epithelial-mesenchymal transition (EMT) inducer that promotes aggressiveness in BC cells.