bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3075: Nuclear export inhibitor selinexor (KPT-330) demonstrates anti-tumor efficacy alone and in combination with chemotherapy in multiple breast cancer models
Authors: Paez Arango Natalia , Evans Kurt , Zhao Ming , Yuca Erkan , Scott Stephen , Kim Charissa , Naing Aung , Meric-Bernstam Funda
Published: July 2016

Background: The nuclear exporter XPO1 (Exportin1 or CRM1), mediates the transport of over 200 proteins, including several tumor suppressors. For this reason, XPO1 is being pursued as a promising target for cancer therapy options. Selinexor (KPT-330), a selective inhibitor of nuclear export, is an oral agent that has been shown to inhibit XPO1 and is currently in phase 2 trials for hematologic and solid tumors. We sought to determine the antitumor effect of selinexor in breast cancer cells in vitro and in vivo

Methods: We studied the effects of selinexor in vitro using cell proliferation assays; the half maximal inhibitory concentration (IC50) was calculated using isobologram curves after 3 days of treatment. We also tested the effects in combination with chemotherapy and calculated the combination index by the method of Chou and Talalay. In vivo efficacy was tested in triple negative breast cancer (TNBC) patient derived xenografts (PDXs) with varying levels of paclitaxel sensitivity, as single agent and in combination therapy. T/C ratio was calculated using the formula: [(median tumor volume of treated group)/(median tumor volume of control group) x 100]

Results: Selinexor treatment inhibited growth of 12 breast cancer cell lines representing different subtypes (TNBC as well as ER+, HER+) with a median IC50 of 375nM (range 3-750nM). Selinexor significantly reduced tumor growth in vivo in 4 of 5 different TNBC PDX models (with varying status of TP53 and PIK3CA, and gene expression subtypes), with a median T/C score of 48% (range 34-59%) In Multiple TNBC cell lines, selinexor was synergistic with paclitaxel, carboplatin, eribulin and doxorubicin in vitro. In vivo, selinexor in combination with paclitaxel or eribulin had greater antitumor efficacy than either agent alone.

Conclusions: Collectively these findings strongly suggest that selinexor is a promising therapeutic option for breast cancer.

Citation Format: Natalia Paez Arango, Kurt Evans, Ming Zhao, Erkan Yuca, Stephen Scott, Charissa Kim, Aung Naing, Funda Meric-Bernstam. Nuclear export inhibitor selinexor (KPT-330) demonstrates anti-tumor efficacy alone and in combination with chemotherapy in multiple breast cancer models. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3075.
Abstract 3084: The effects of NT-1044, a novel AMPK activator, on endometrial cancer cell proliferation, apoptosis, cell stress, and tumor growth
Authors: Roque Dario R. , Wysham Weiya Z. , Zhou Chunxiao , Batchelor Ken W. , Brewster Wendy R. , Bae-Jump Victoria L.
Published: July 2016

Objectives: Obesity and diabetes are associated with increased risk and worse outcomes in endometrial cancer (EC). Anti-diabetic biguanide drugs such as metformin may have anti-tumorigenic effects by behaving as AMPK activators and mTOR inhibitors. Metformin requires organic cation transporters (OCTs) for entry into cells, and NT-1044 (NovaTarg Therapeutics) is an AMPK activator designed to have greater affinity for two of these transporters, OCT1 and 3. We sought to compare the effects of NT-1044 on cell proliferation in human EC cell lines and on tumor growth in an endometrioid EC mouse model.

Methods: Cell proliferation was assessed in two EC cell lines, ECC-1 and Ishikawa, by MTT assay after exposure to NT-1044. Apoptosis was analyzed by Annexin V-FITC assay. Cell cycle progression was evaluated by flow cytometry. Reactive oxygen species (ROS) were measured using a DCFH-DA assay. Western immunoblotting was performed to evaluate the effects of NT-1044 on the downstream targets of the AMPK/mTOR pathway and proteins related to cell cycle and cellular stress. For in vivo studies, we utilized the LKB1f/f/p53f/f mouse model of endometrioid endometrial cancer. Mice were treated with placebo or NT-1044 (200 mg/kg/day, oral gavage) following tumor onset for 4 weeks.

Results: NT-1044 significantly inhibited proliferation in a dose-dependent manner in both EC cell lines after 72 hours of exposure (IC50 218 uM for ECC-1; 87 uM for Ishikawa). Treatment with NT-1044 resulted in G1 cell cycle arrest, induced apoptosis and increased ROS production in both cell lines. NT-1044 increased phosphorylation of AMPK and decreased phosphorylation of S6, a key downstream target of the mTOR pathway. Expression of the cell cycle proteins CDK4, CDK6 and cyclin D1 decreased in a dose dependent-fashion while cellular stress protein expression was induced in both cell lines. As compared to placebo, NT-1044 inhibited endometrial tumor weight in the LKB1f/f/p53f/f mice by 56% (p = 0.031).

Conclusions: NT-1044 suppressed EC cell growth through G1 cell cycle arrest, induction of apoptosis, cellular stress and inhibition of the AMPK/mTOR pathway. In addition, NT-1044 inhibited EC tumor growth in vivo. More work is needed to determine if this novel biguanide will be beneficial in the treatment of women with EC, a disease strongly impacted by obesity and diabetes.

Citation Format: Dario R. Roque, Weiya Z. Wysham, Chunxiao Zhou, Ken W. Batchelor, Wendy R. Brewster, Victoria L. Bae-Jump. The effects of NT-1044, a novel AMPK activator, on endometrial cancer cell proliferation, apoptosis, cell stress, and tumor growth. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3084.
Abstract 3085: The BET family bromodomain inhibitor ABBV-075 targets multiple pathogenesis factors in multiple myeloma and exhibits robust in vivo efficacies as a single agent and in combination with bortezomib
Authors: Uziel Tamar , Lin Xiaoyu , Faivre Emily , Sarthy Aparna , Albert Daniel H. , Li Leiming , Wilcox Denise , Huang Xiaoli , Magoc Terry , Lam Lloyd , Elmore Steven W. , McDaniel Keith , Kati Warren , Shen Yu
Published: July 2016

Small molecule inhibitors of the bromodomain and extraterminal domain (BET) proteins represent a potential new class of cancer therapeutic agents. ABBV-075 is a potent and selective BET family bromodomain inhibitor that was recently advanced to Phase 1 studies. Here we report the in vitro and in vivo characterization of ABBV-075 in preclinical models of multiple myeloma. ABBV-075 exhibited significant anti-proliferative activities and triggered robust apoptosis across many multiple myeloma cell lines, including cell lines that were engineered to become resistant to bortezomib. As reported for other BET inhibitors, ABBV-075 potently inhibited super-enhancer regulated potential cancer drivers such as Myc, IRF8 etc. In addition to directly impacting multiple myeloma cell growth and survival, ABBV-075 also diminished stroma cell derived IL-6 secretion and prevented HUVEC cell proliferation in vitro. Given the importance of angiogenesis and the survival factor IL-6 for multiple myeloma pathogenesis, ABBV-075 could potentially produce significant therapeutic benefits in myeloma by simultaneously targeting multiple critical pathogenesis mechanisms for this disease. ABBV-075 caused significant tumor growth delay in the OPM2 flank tumor model as a monotherapy. Combining ABBV-075 with Bortezomib provided further therapeutic benefit compared with the treatment of either of the two agents individually. Taken together, our results suggest that ABBV-075 could be a promising option to explore for the treatment of multiple myeloma.

Citation Format: Tamar Uziel, Xiaoyu Lin, Emily Faivre, Aparna Sarthy, Daniel H. Albert, Leiming Li, Denise Wilcox, Xiaoli Huang, Terry Magoc, Lloyd Lam, Steven W. Elmore, Keith McDaniel, Warren Kati, Yu Shen. The BET family bromodomain inhibitor ABBV-075 targets multiple pathogenesis factors in multiple myeloma and exhibits robust in vivo efficacies as a single agent and in combination with bortezomib. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3085.
Abstract 3108: PD-L1 expressing circulating tumor cells (CTCs) in patients with head and neck squamous cell carcinoma (HNSCC)
Authors: Strati Areti , Koutsodontis George , Angelidis Ilias , Sasaki Clarence , Avgeris Margaritis , Psyrri Amanda , Lianidou Evi S.
Published: July 2016

Background: Predictive biomarkers for response to anti-PD1 therapy are lacking. Because therapy with checkpoint inhibitors is cost intensive, noninvasive tools for prediction of responders are of major interest.

Methods: The “Liquid Biopsy’’in head and neck squamous cell carcinoma (HNSCC) project involved the isolation of Circulating Tumor Cells (CTC) from patients with HNSCC at baseline, at different time points during treatment and at relapse. Herein, we assessed gene expression of PD-L1 on CTCs in a prospective fashion in a cohort of locally advanced inoperable HNSCC patients treated with curative intent (n = 61), in a second cohort of recurrent/metastatic HNSCC patients (n = 18) and in 20 healthy individuals, used as normal controls. For the quantification of PD-L1mRNA in CTCs, we developed a highly sensitive, specific, and robust RT-qPCR assay that was firstly analytically validated prior to its application in HNSCC patients.

Results: In patients with locally advanced disease, 54/61(88.5%) samples were evaluable for CTCs at baseline. Twenty four samples were obtained after induction chemotherapy (IC) and 34(55.7%) at the end of concurrent chemo-radiation. At baseline 22/54(40.7%) pts were found to be positive for PD-L1 overexpression, at the post-IC samples 12/24(50%) patients were positive for PD-L1overexpressionand at the end of treatment, 11/34 (32.4%) patients were positive for PD-L1 overexpression. Patients with PD-L1 positive CTCs at the end of treatment had shorter progression-free survival (PFS) (p = 0.011) and overall survival (OS) (p = 0.004). Multivariate analysis showed that PD-L1 overexpression in patients at the end of treatment was independent prognostic factor of PFS (HR = 2422.4; p = 0.014) and OS (HR = 32.23; p = 0.014). Five R/M patients were found to be positive for PD-L1 out of the 18 (27.8%) at baseline.

Conclusions: We demonstrate for the first time that detection of PD-L1+ CTCs at the end of treatment in patients with locally advanced disease is associated with shorter PFS and OS. Serial PD-L1 expression assessment has potential to select and monitor pts for PD-1 checkpoint inhibitors. These data support testing of PD-1 inhibitors in the adjuvant setting in patients with locally advanced HNSCC in whom PD-L1 positive CTCs are detected at the end of treatment.

Citation Format: Areti Strati, George Koutsodontis, Ilias Angelidis, Clarence Sasaki, Margaritis Avgeris, Amanda Psyrri, Evi S. Lianidou. PD-L1 expressing circulating tumor cells (CTCs) in patients with head and neck squamous cell carcinoma (HNSCC). [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3108.
Abstract 3132: Genome-wide scan for genetic markers modifying survival of pancreatic cancer patients
Authors: Tang Hongwei
Published: July 2016

Background: Genetic factors that are associated with pancreatic cancer survival have previously been examined in Genome-Wide Association Studies (GWAS), but the results are inconsistent due to limited sample size.

Method: We performed a two-stage study on pancreatic cancer survival. The discovery phase took advantage of the existing GWAS data of 868 patients with European descendent from a case-control study conducted at MD Anderson. The validation study on ten top hits was conducted in another 820 white patients with Taqman. The missing genotypes were imputed using IMPUTE2 based on 1000 Genomes Project. Cox regression model was fitted to calculate allele-specific hazard ratios (HR) and 95% confidence intervals (95% CI) with adjustment for demographics, tumor stage, resection, and chemotherapy.

Results: After imputation, 7,738,399 SNPs with a Quality Score >0.03, minor allele frequency >0.01, and Hardy-Weinberg equilibrium were obtained for analysis. Three loci in complete LD at chromosome 2 were associated with overall survival (OS) at genome-wide significance (P = 1 ×): rs113988120 in PAIP2B (poly(A) binding protein interacting protein 2B), rs112493246 and rs138529893 in DYSF (dysferlin). The variant allele was associated with 3.02 fold higher risk of dying (95% CI: 2.07-4.40). SNP rs113988120 and nine others were examined in the validation study and rs113988120 remained nominally significant (P = 2.85 ×; HR: 1.45, 95% CI: 1.04-2.03). In the combined datasets of 1688 patients, this SNP was significantly associated with OS (P = 1.46 × and HR = 1.80 (95% CI: 1.42-2.30).

Conclusion: These observations support our hypothesis that genetic variations to a certain degree influence patient survival in pancreatic cancer.

Key words: Pancreatic cancer, GWAS, Survival

Citation Format: Hongwei Tang. Genome-wide scan for genetic markers modifying survival of pancreatic cancer patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3132.
Abstract 3168: Your targeted population might not be what you predict: Changes in tumor genetic landscapes post standard of care
Authors: Greenawalt Danielle , Dulak Austin , Ramakrishna Manasa , Lai Zhongwu , Johnson Justin , Dai Hongyue , Mitchell Melissa , Barrett Carl , Fawell Stephen , Dry Jonathan
Published: July 2016

Our current understanding of the mutation spectrum of relapse/refractory patients is limited. Public sequencing efforts have focused by design, on primary tumors, while re-biopsy of patients who are refractory to first line therapy or who relapse on treatment is not standard protocol. We have performed whole exome sequencing on post standard of care biopsies from 3 patient cohorts, 47 DLBCL patients post-relapse following R-CHOP, 49 triple negative breast (TNBC) following taxol alone or in combination and 41 lung adenocarcinoma (LUAD) following platinum alone or in combination. Comparative analysis of the mutation spectrum of these cohorts was performed against available clinically matched primary tumor cohorts. Comparison of the mutational signatures, somatic variant frequencies and copy number variation between matched pre- and post-treatment samples, and treated-primary tumor cohorts were performed. Significant differences were identified in the frequency of oncogenic drivers including PIK3CA in TNBC and KRAS in LUAD. We also observed mutational signature differences between cohorts that could impact treatment regimens post standard of care. Collectively these data further highlight the value of re-assessing the tumor genetic landscape at the time of treatment rather than relying on primary tumor samples, which may not be representative of the cohort to be treated.

Citation Format: Danielle Greenawalt, Austin Dulak, Manasa Ramakrishna, Zhongwu Lai, Justin Johnson, Hongyue Dai, Melissa Mitchell, Carl Barrett, Stephen Fawell, Jonathan Dry. Your targeted population might not be what you predict: Changes in tumor genetic landscapes post standard of care. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3168.
Abstract 3190: Genomic profiling of salivary gland tumors identifies novel and targetable alterations
Authors: Majewska Hanna , Müller Judith , Lakis Sotirios , Czapiewski Piotr , Gorczyński Adam , Iliszko Mariola , Skalova Alena , Dziadziuszko Rafal , Jassem Jacek , Biernat Wojciech , Menon Roopika , Heuckmann Johannes M. , Heukamp Lukas C.
Published: July 2016

Background

Salivary gland neoplasms represent a heterogeneous group of benign and malignant tumors that comprise 6% of all head and neck cancers. Due to their low incidence, these tumors are poorly understood and remain a diagnostic and therapeutic challenge.

Methods

Out of 182 analyzed salivary gland tumors with various histologies, eight were positive for ALK immunohistochemistry. The cut-off was 15% positive cells in either membranous, nuclear or cytoplasmic compartments. The ALK positive tumors were first subjected to FISH analysis. Subsequently, these tumors were analyzed using hybrid capture based next generation sequencing to confirm possible ALK gene fusions or copy number alterations, and to detect additional genomic alterations of clinical relevance.

Results

An in-depth genomic analysis of the samples resulted in the detection of inactivating mutations in BRAF (p.D594N) and TP53 (p.C238S), as well as amplifications of ERBB2 and ALK. Strikingly, a novel MYO18A (Exon1-40)-ALK (exon 20-29) gene fusion was detected in a patient with adenocarcinoma not otherwise specified. This tumor was FISH positive and 100% of cells showed strong membranous staining for ALK. MYO18A (Exon1-40)-ALK (exon 20-29) gene fusion resulted in the retention of the kinase domain of ALK and the coiled-coil domain of MYO18A. Similarly to other ALK fusions, we hypothesize that the coiled-coil domain of MYO18A mediates the dimerization and activation of MYO18A-ALK, thereby resulting in an overexpression of constantly activated ALK.

Conclusion

Using hybrid capture based next generation sequencing, we identified in salivary gland tumors numerous genomic alterations in therapeutically relevant genes and a novel gene fusion (MYO18A-ALK). Patients harboring this fusion may potentially benefit from treatment with ALK inhibitors.

Citation Format: Hanna Majewska, Judith Müller, Sotirios Lakis, Piotr Czapiewski, Adam Gorczyński, Mariola Iliszko, Alena Skalova, Rafal Dziadziuszko, Jacek Jassem, Wojciech Biernat, Roopika Menon, Johannes M. Heuckmann, Lukas C. Heukamp. Genomic profiling of salivary gland tumors identifies novel and targetable alterations. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3190.
Abstract 321: S49076, a novel kinase inhibitor of MET, AXL and FGFR, demonstrates marked in vitro and in vivo efficacy in hepatocellular carcinoma
Authors: Tijeras-Raballand Annemilai , Pasteur-Rousseau Adrien , Martinet Matthieu , Bonnin Philippe , Pocard Marc , Banquet Sébastien , Cattan Valérie , Raymond Eric , de Gramont Armand
Published: July 2016

Introduction: Hepatocellular carcinoma (HCC) is the second cause of cancer-related deaths worldwide. While sorafenib may improve overall survival in advanced HCC, new therapies are needed. Both preclinical and clinical data support a role of MET and AXL receptor tyrosine kinases in HCC progression. We thus evaluated S49076, a novel MET/AXL/FGFR inhibitor in HCC cell lines in vitro and in a transgenic mouse model of HCC.

Materials and Methods: MET and AXL expressions were assessed in a panel of HCC cell lines by western blot. Downstream signaling pathways were investigated in the presence of HGF, GAS6 and S49076. Cell invasion was evaluated on matrigel assay. Transgenic mice developing stage-defined HCC were treated with S49076 (50mg/kg). Tumor response was evaluated measuring liver volume by echo-doppler and the number of tumor nodules at intermediary (12weeks) and final sacrifice (16 weeks).

Results: Both MET and AXL are expressed in the majority of HCC lines screened. Notably, a high level of AXL was observed in the SK-HEP1 cells which were therefore selected for further investigation. MET and AXL pathways were strongly stimulated by their respective ligands, HGF and GAS6. Combination of HGF and GAS6 led to marked activation of ERK. S49076 inhibited MET and AXL pathway activation and HGF- and GAS6- induced invasion at nanomolar concentrations. In vivo anti-tumor efficacy of S49076 was demonstrated in a transgenic MET-expressing mouse model of HCC.

Conclusion: S49076 displayed strong MET and AXL pathway inhibition and anti-invasive properties in SK-HEP1 HCC cells. Moreover, S49076 demonstrated antitumor activity in a mouse model of HCC. Together, these results would support development of S49076 as an innovative treatment in HCC patients.

Citation Format: Annemilai Tijeras-Raballand, Adrien Pasteur-Rousseau, Matthieu Martinet, Philippe Bonnin, Marc Pocard, Sébastien Banquet, Valérie Cattan, Eric Raymond, Armand de Gramont. S49076, a novel kinase inhibitor of MET, AXL and FGFR, demonstrates marked in vitro and in vivo efficacy in hepatocellular carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 321.
Abstract 3215: GITRL-Fc can significantly reduce tumor growth by stimulating innate and adaptive immunity
Authors: Jie Hyun-Bae , Srivastava Minu K. , Mayes Erin , Yun Rui , Axelrod Fumiko , Monteon Jorge , Gurney Austin , Park. Angie In-Kyung
Published: July 2016

Although the interaction between GITR (Glucocorticoid-Induced Tumor Necrosis Factor Receptor) and GITRL (its ligand) is important for the development of immune responses, the cellular mechanisms underlying anti-tumor immunity including both innate and adaptive immunity is not fully understood. We generated a novel single-gene linkerless GITRL trimer fused to an immunoglobulin Fc domain (GITRL-Fc), and investigated its effect on controlling tumor growth and immune responses in preclinical tumor models. Treatment with GITRL-Fc significantly reduced tumor growth in several preclinical tumor models by inducing Th1 biased anti-tumor immunity and reducing Treg-mediated immune suppression. Immune cell depletion studies showed that anti-tumor immunity induced by GITRL-Fc depended on CD8+ T cells as well as NK cells. Furthermore, the combination of GITRL-Fc with PD-1 blockade significantly reduced the tumor growth in the Renca murine kidney adenocarcinoma tumor model. Taken together, these results suggest that GITRL-Fc can improve cancer treatment as a single agent or in combination therapy by enhancing innate and adaptive cellular immunity.

Citation Format: Hyun-Bae Jie, Minu K. Srivastava, Erin Mayes, Rui Yun, Fumiko Axelrod, Jorge Monteon, Austin Gurney, Angie In-Kyung Park. GITRL-Fc can significantly reduce tumor growth by stimulating innate and adaptive immunity. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3215.
Abstract 3240: Prostate cancer exosomes and their effects on macrophage engulfment and polarization
Authors: Weagel Evita G. , Meng Wei , Robison Richard A. , O’Neill. Kim L.
Published: July 2016

The purpose of this study is to explore the effects of prostate cancer-derived exosomes on macrophage engulfment and polarization. Exosomes are small vesicles secreted by virtually all cells, which contain nucleic acids, lipids, proteins, and carbohydrates. Exosomes can be abundantly found in cancer patient secretions (serum, urine, etc.) and contain a high amount of RNA molecules, some of them being miRNA, and parts of the DNA of the cell of origin. One of the functions of exosomes is to mediate cellular communication. This occurs as they are secreted from one cell and then distribute their contents into other cells. Some exosomes specifically target immune cells. The miRNA the exosomes carry can silence genes involved in the activation of these immune cells, leading to immune suppression and tumor growth. Macrophages are often found surrounding the tumor microenvironment, as they are recruited by tumor signals to promote angiogenesis and metastasis. These macrophages, which are often called tumor associated macrophages (TAMs), are exposed to various signals derived from the tumor microenvironment, including exosomes. Previous data generated in our lab suggests that the co-culture of macrophages with PC3 and DU145 cell lines results in a decrease in macrophage phagocytosis and an M2-like polarization state. For this study, we exposed U937 cells to phorbol 12-myristate 13-acetate (PMA) for 24 hours to allow for differentiation. Then, we co-cultured these macrophages with isolated exosomes from spent media from both PC3 and DU145 cell lines at 1, 2, 3, 4, 5, 6, 12, and 24 hours. We then allowed the macrophages to phagocytose fluorescent microbeads for 1 hour. Flow cytometry analysis of these macrophages revealed that the total bead phagocytosis starts decreasing at 3 hours of exosome co-culture compared to control. Interestingly, total bead phagocytosis resumes to normal at 12 and 24 hours of exosome co-culture. Gene expression analysis of these macrophages suggests an M2 phenotype from hours 2-24. These results suggest that prostate cancer-derived exosomes can target macrophages and affect their phagocytosis. Further research is required to elucidate the specific genes these exosomes target.

Citation Format: Evita G. Weagel, Wei Meng, Richard A. Robison, Kim L. O’Neill. Prostate cancer exosomes and their effects on macrophage engulfment and polarization. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3240.
Abstract 3246: Phytotherapy boosts innate immunity to maximize cancer survival
Authors: Lau Benjamin H.
Published: July 2016

We studied surgery, radiation, and chemotherapy in animals and found all these modalities suppressed cancer growth but they also impaired the immune functions of the animal host. We also studied phytotherapy (use of plant chemicals) and found it to have no side effects and with more lasting benefits. Using in vitro cell culture system with both malignant and non-malignant cells, and immune cells (macrophages and natural killer cells), we compared crude extract of Pacific yew and the chemotherapeutic drug Paclitaxel derived from Pacific yew. The crude extract or the drug was incubated separately with: human malignant ovarian cells, nonmalignant ovarian cells, macrophages, and natural killer cells. Cell viability was determined by MTT (methylthiazol tetrazolium) assay. Results show the drug damaged all these four types of cells while the crude plant extract only damaged the cancer cells showing its selective toxicity for cancer cells. Study with other phytochemicals also shows selective toxicity for cancer cells while enhancing the activities of immune cells. Phytotherapy is thus a non-toxic chemotherapy and immunotherapy. In our clinical observation, 160 men and women diagnosed with five types of cancer (72 with breast cancer, 28 with prostate cancer, 18 with colon cancer, 14 with lung cancer, 28 with ovarian cancer) chose to use phytotherapy (a plant-based diet) plus lifestyle modification. The five year survival of these patients were 90%, 89%, 83%, 86%, and 82%, respectively. Compared with five year survival data from National Cancer Institute (www.seer.cancer.gov) for these five types of cancer: 82%, 50%, 62%, 15%, and 45%, respectively, the phytotherapy group exhibits greater cancer survival. Since numerous studies have now revealed that cancer is attributable to our lifestyle habits – the way we eat, and the way we live, our data suggest that regardless of treatment choices, a change in diet and lifestyle is likely to improve cancer survival.

Citation Format: Benjamin H. Lau. Phytotherapy boosts innate immunity to maximize cancer survival. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3246.
Abstract 3255: Inhibition of intracrine VEGF signaling prevents colorectal cancer cell migration and invasion
Authors: Bhattacharya Rajat , Xia Ling , Fan Fan , Wang Rui , Boulbes Delphine , Ye Xiang-Cang , Ellis Lee
Published: July 2016

Introduction: Vascular endothelial growth factor (VEGF) is a key regulator of angiogenesis and vascular function. The mechanisms of VEGF signaling in angiogenesis have been extensively studied. However, its effects on tumor cell function remain to be elucidated. Our studies on depletion of VEGF by somatic knockout of the VEGF gene or by siRNA in human colorectal cancer (CRC) cell lines have demonstrated that loss of VEGF expression led to significantly decreased cell growth, increased apoptosis, and enhanced chemo-sensitivity of CRC cells through an intracrine signaling mechanism. These intracrine effects were mediated through VEGF by regulation of the activity of multiple receptor tyrosine kinases (e.g. EGFR, cMET) and downstream AKT signaling. Similar effects were observed by depletion of VEGF receptor 1 (VEGFR1), supporting the role of a novel intracellular VEGF-VEGFR1 complex in CRC cell survival. Since VEGF signaling has been shown to effect endothelial and tumor cell migration, we attempted to determine if inhibition of intracrine VEGF signaling affected CRC cell motility.

Methods: Migration and invasion of CRC cells, with and without VEGF depletion, were assayed using Boyden chambers and a serum gradient. Changes in cell morphology and epithelial to mesenchymal transition (EMT) markers and markers of cell motility were assessed by microscopy, immunostaining, and western blot analyses.

Results: Depletion of VEGF by siRNAs in multiple CRC cell lines led to a very strong inhibition of migration and invasion of CRC cells. Such inhibition was not observed when CRC cells were treated with bevacizumab to inhibit paracrine or autocrine signaling. Further examination revealed no significant changes in cell morphology. Other than Twist, there were no significant differences between specific markers of EMT indicating that differences in migration and invasion were not due to classic alterations in EMT. However, significant changes in activation of mediators of cell motility were observed. VEGF depleted CRC cells demonstrated significantly lower levels of phosphorylated focal adhesion kinase (p-FAK) compared to cells with normal VEGF expression. Analyses of upstream regulators indicated strong reduction in either p-cMET or p-SRC depending on cell type.

Conclusions: Inhibition of intracrine VEGF signaling strongly inhibits CRC cell migration and invasion by regulating proteins involved in cell motility.

Citation Format: Rajat Bhattacharya, Ling Xia, Fan Fan, Rui Wang, Delphine Boulbes, Xiang-Cang Ye, Lee Ellis. Inhibition of intracrine VEGF signaling prevents colorectal cancer cell migration and invasion. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3255.
Abstract 3276: The tissue and cellular destination of therapeutic IgGs in glioblastoma
Authors: Muller-Greven Gaelle , Carlin Cathleen , Toms Steven , Ahluwalia Manmeet , Bredel Markus , Lathia Justin , Rich Jeremy , Hamerlik Petra , Gladson Candece L.
Published: July 2016

Most patients with recurrent glioblastoma (GBM) are treated with bevacizumab, a humanized monoclonal antibody (mAb) that binds VEGF-A and inhibits its binding to VEGFR. Approximately 30% of GBM patients are non-responsive to bevacizumab and the underlying mechanism for the lack of response is not known. It has been assumed that bevacizumab solely targets circulating VEGF-A in blood. We hypothesized that bevacizumab and human IgGs in general gain access to the perivascular niche that contains cancer stem cells (CSCs) in GBM. We found that bevacizumab gains access to the perivascular tumor area through leaky blood vessels and was internalized by tumor cells in an orthotopic xenograft mouse model of GBM. In vitro, CSCs (CD133+) from GBM rapidly internalized either bevacizumab or human IgG into membrane protrusions that contained actin and internalization was significantly inhibited by a macropinocytosis inhibitor (EIPA), suggesting CSCs internalize bevacizumab or human IgG via macropinocytosis. Furthermore, bevacizumab or human IgG was largely detected in the Rab4+ “fast” recycling compartment at 5 min, and both were largely detected in the LAMP1+ compartment (late endosome/lysosome) at 3 hr in the CSCs. CSCs (CD133+) from GBM do not express the neonatal Fc receptor, the canonical pathway for recycling of IgG. Administration of bevacizumab to an orthotopic xenograft mouse model of established GBM showed that bevacizumab was partially co-localized with Rab4+ or with LAMP1+ in perivascular tumor cells, consistent with our in vitro findings. Taken together, our data show that in GBM, humanized IgG, including bevacizumab, gains access to the perivascular tumor space and is then macropinocytosed by CSCs and trafficked to a recycling compartment or to the late endosome/lysosome. These data suggest that alterations in endocytosis or recycling in the CSCs could impact the fate of therapeutic IgGs like bevacizumab and ultimately influence a patients’ response to GBM therapy.

Citation Format: Gaelle Muller-Greven, Cathleen Carlin, Steven Toms, Manmeet Ahluwalia, Markus Bredel, Justin Lathia, Jeremy Rich, Petra Hamerlik, Candece L. Gladson. The tissue and cellular destination of therapeutic IgGs in glioblastoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3276.
Abstract 3283: APRIL/BCMA activation promotes human multiple myeloma progression and further induces immunosuppressive bone marrow microenvironment
Authors: Tai Yu-Tzu , Acharya Chirag , An Gang , Moschetta Michele , Zhong Mike Y. , Feng Xiaoyan , Cea Michele , Cagnetta Antonia , Wen Kenneth , van Eenennaam Hans , van Elsas Andrea , Munshi Nikhil , Anderson Kenneth
Published: July 2016

Elevated B cell maturation antigen (BCMA) and its ligand a proliferation-inducing ligand (APRIL) may directly advance human multiple myeloma (MM) malignancy in vivo. Here, we first show that BCMA downregulation potently diminishes viability and colony formation of MM cells while BCMA overexpression augments MM cell growth and survival via induction of AKT, MAPK, and NFκB signaling cascades and molecules essential for proliferation and anti-apoptosis. Importantly, BCMA itself forces accelerated tumor growth of xenografted MM cells harboring p53 mutation in mice. BCMA-overexpressing tumors exhibit significantly enhanced CD31/microvessel density and VEGF than paired control tumors. Concurrently, these tumors express increased factors crucial for osteoclast activation, adhesion, and angiogenesis/metastasis, as well as immune inhibition including PD-L1, TGFβ and IL-10 which further triggers 38 IL-10 signaling molecules. In parallel, these identified target genes are induced by paracrine APRIL binding to BCMA in MM cells and they are potently blocked by an antagonistic anti-APRIL monoclonal antibody hAPRIL01A (01A). 01A is cytotoxic against MM cells protected by abnormal bone marrow (BM) myeloid cells, i.e., osteoclasts, macrophages, and plasmacytoid dendritic cells. It further decreases APRIL-induced adhesion and migration of MM cells via blockade of canonical and non-canonical NFκB pathways. It prevents in vivo MM cell growth within implanted human bone chips in SCID mice. 01A-inhibited MM cell viability induced by osteoclasts is further enhanced by lenalidomide and bortezomib. Taken together, these results further characterize new molecular mechanisms of APRIL/BCMA-induced in vivo MM progression and immunosuppressive BM microenvironment, strongly supporting targeting this prominent pathway in MM.

Citation Format: Yu-Tzu Tai, Chirag Acharya, Gang An, Michele Moschetta, Mike Y. Zhong, Xiaoyan Feng, Michele Cea, Antonia Cagnetta, Kenneth Wen, Hans van Eenennaam, Andrea van Elsas, Nikhil Munshi, Kenneth Anderson. APRIL/BCMA activation promotes human multiple myeloma progression and further induces immunosuppressive bone marrow microenvironment. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3283.
Abstract 3285: HTLV-1 viral oncogene Hbz induces leukemia with osteolytic bone involvement in mice
Authors: Esser Alison , Rauch Dan , Xiang Jingyu , Harding John , Kohart Nicole , Green Patrick , Niewiesk Stefan , Rosol Thomas , Ratner Lee , Weilbaecher Katherine
Published: July 2016

Adult T-cell leukemia/lymphoma (ATL) develops in a subset of patients infected with the HTLV-1 virus. Most ATL patients become refractory to chemotherapy and have a median survival time of 6 months. Although uncommon in hematologic malignancies, 80% of ATL patients develop osteolytic lesions and hypercalcemia of malignancy. Bone resident and metastatic tumors release paracrine factors that modulate the bone microenvironment to facilitate disease progression and decrease survival. HTLV-1 encodes 2 viral oncogenes, Tax and Hbz. Tax is critical to ATL development and regulates tumor growth and proliferation in part through trans-activation of NFκB and CREB. We have previously shown Tax expression driven by the Granzymbe B promoter is sufficient for the development of leukemia/lymphoma with osteolytic lesions and hypercalcemia. We and others have shown that Tax alters the expression of paracrine factors that modulate the bone microenvironment through effects on bone forming osteoblasts (OB) and bone resorping osteoclasts (OC). Tax is expressed in early lymphocyte transformation with low expression in advanced ATL. HBZ is expressed early in lymphocyte transformation and throughout ATL progression. We hypothesize that in ATL cells, HTLV-1 viral oncogenes Tax and Hbz cooperate to modulate bone metabolism in a paracrine manner to enhance ATL tumor growth and progression. Mice with Granzyme B driven Hbz expression (T and NK cells) develop leukemia/lymphoproliferative disease in lymph nodes correlating with increased spleen weight. We found that lymphoproliferative disease is also present in the bone marrow. Hbz mice have decreased trabecular bone at 18 months by microCT and radiographic analysis. These data suggest Hbz can alter bone metabolism. Future studies will define the effects of Hbz on bone formation, OB and OC specific effects and tumor progression. Understanding HTLV-1 oncogene modulation of the bone microenvironment will uncover critical pathways in tumor/bone cross talk enabling the development of novel targeted therapies for ATL patients.

Citation Format: Alison Esser, Dan Rauch, Jingyu Xiang, John Harding, Nicole Kohart, Patrick Green, Stefan Niewiesk, Thomas Rosol, Lee Ratner, Katherine Weilbaecher. HTLV-1 viral oncogene Hbz induces leukemia with osteolytic bone involvement in mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3285.
Abstract 3295: Microbiome profiling and immunological characterization on gastrointestinal tract tumors
Authors: Zhang Chao , Cleveland Kyle , Schnoll-Sussman Felice , McClure Bridget , Bigg Michelle , Schultz Nikolaus , Shah Manish A. , Betel Doron
Published: July 2016

Motivation: Conservative estimates are that around 20% of all malignancies may be attributed to microbiota. Especially in the gastrointestinal (GI) tract, infection can cause chronic inflammation and further influence immune response to create a proinflammatory microenvironment for cancer development. This can account in part for the reason that the immunotherapy in colon cancer is less efficacious than in non-GI tract cancers. We proposed a systematic computational study to profile the microbiome from next generation sequencing of gastric biopsy samples, and evaluate the relationships among microbiome composition, host immune response and genomic characterization.

Method: Patients undergoing upper endoscopy without chronic inflammatory disease or chronic NSAID use were eligible for participation, including cancer/non-cancer, Helicobacter pylori positive and negative patients. In total, 62 endoscopic biopsies from 3 locations in the gastric antrum/body were freshly frozen for whole genome sequencing and transcriptome analysis. In addition, TCGA whole genome sequencing (WGS) data from 37 gastric cancer, 102 colon cancer, and 46 rectal cancer samples were analyzed for microbiome composition.

Results: H. pylori was detected in all positive infection gastric biopsy samples by our pipeline, and the results were successfully validated by qPCR. Comparing to other traditional bacterial identification methods, our pipeline shows a strong capacity for profiling microbiome from low bacterial content biopsy samples. We observed a high positive correlation between the H. pylori content and total non-H. pylori bacteria content. And we also identified significantly differentially expressed immune-related genes between active H. pylori infection and negative samples. Finally, we analyzed collected TCGA WGS data, and identified the dominant species from the different cancer studies, such as Bacteroides fragilis in rectum adenocarcinoma, and Fusobacterium nucleatum in colon adenocarcinoma. The correlation among microbiome composition, the expression of immune-related genes and cancer genomic characterization were also demonstrated in this study.

Citation Format: Chao Zhang, Kyle Cleveland, Felice Schnoll-Sussman, Bridget McClure, Michelle Bigg, Nikolaus Schultz, Manish A. Shah, Doron Betel. Microbiome profiling and immunological characterization on gastrointestinal tract tumors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3295.
Abstract 33: NSCLC cells internalize ATP in vitro and in vivo using multiple endocytotic pathways
Authors: Qian Yanrong , Wang Xuan , Li Yunsheng , Cao Yanyang , Chen Xiaozhuo
Published: July 2016

Upregulated glycolysis in cancer, the Warburg effect, has been studied for almost one century. However, the biological reasons for the upregulation are still debated and the effect is far from fully understood. One of the major controversies related to the effect is the role of ATP synthesis. Intratumoral ATP concentrations are found to be 10⁁3-10⁁4 times higher than those in normal tissues of the same cell origin. However, whether and how cancer cells use the abundant extracellular ATP was unknown until we recently reported that cancer cells internalize ATP by macropinocytosis (1). Extracellular ATP was found to increase intracellular ATP concentration and promote cell proliferation and drug resistance in cancer cells (1). Here we report that, using a nonhydrolyzable fluorescent ATP (NHF-ATP) as an ATP surrogate, high and low molecular weight dextrans as endocytosis tracers and fluorescence microscopy detection, cultured human NSCLC A549 and H1299 cells as well as xenografted A549 tumors were found to internalize ATP at concentrations within the reported intratumoral ATP concentrations. Endocytosis inhibitor studies show that in addition to macropinocytosis, clathrin- and caveolae-mediated endocytoses contribute to the ATP internalization, which led to a ∼30% increase in intracellular ATP level after 45 min of ATP incubation. This increase cannot be accounted for by purinergic receptor signaling or increased intracellular ATP synthesis rates. Internalization of NHF-ATP and dextran exhibits time-dependent profiles parallel to the extracellular ATP-induced intracellular ATP level elevation. All these demonstrate the observed ATP increase is a result of internalization of extracellular ATP and NHF-ATP is a powerful tool for studying ATP internalization. These findings significantly deepen our understanding of the Warburg effect by showing how cancer cells in tumors take up intratumoral ATP and perform previously unrecognized biological functions. It strongly suggests ATP-sharing among cancer and stromal cells and identifies multiple novel anticancer targets (2).

1. Qian Y, Wang X, et al., Cancer Lett. 351(2014): 242-5.

2. Chen X, Qian Y, Wu S. Free. Radic. Biol. Med. 79(2015): 253-263.

Citation Format: Yanrong Qian, Xuan Wang, Yunsheng Li, Yanyang Cao, Xiaozhuo Chen. NSCLC cells internalize ATP in vitro and in vivo using multiple endocytotic pathways. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 33.
Abstract 3312: Transcriptional regulation of chemokine receptor 4 (CXCR4) by nuclear respiratory factor 1 (NRF1) controls estrogen-induced malignant transformation of breast epithelial cells to breast cancer stem cells
Authors: Das Jayanta Kumar , Roy Deodutta
Published: July 2016

Chemokine receptor 4 (CXCR4) is involved in the maintenance normal stem cells and metastasis of cancer stem cells. NRF1 is one of the major transcription factor that controls transcription of CXCR4. The functional regulation of transcription of NRF1 target genes has not been explored in breast cancer. In this study, we have investigated whether transcriptional regulation of CXCR4 by NRF1 regulates estrogen-induced malignant transformation of breast epithelial cells to breast cancer stem cells (CSCs). We have previously shown that NRF1 may be involved in 17 β-estradiol (E2) induced malignant transformation of breast epithelial cells, however, the mechanism of transcriptional regulation of the NRF1 target genes remains unknown. In this study we showed that NRF1 and E2 jointly contributed in reprogramming of breast epithelial cells to CSCs. Levels of breast cancer stem cell markers (CD44+CD24+ALDH1+CD133+) were significantly increased by E2 treatment in NRF1 overexpressing cells compared to cells transfected with vector receiving E2. E2-induced increases of spheroid formation, cell survival and growth of cancer stem cells were modulated by functional gain or loss of NRF1. For morphological details we used a holographic microscope (HoloMonitor M4), we observed that E2 generated breast cancer stem cells with CD44+CD24+ALDH1+CD133+ markers produced large live spheroids in NRF1 overexpressing cells compared to cells transfected with vector receiving E2. Overexpression of NRF1 promoted the transition of E2-treated breast epithelial MCF-10A cells to mesenchymal stem cell-like phenotype. The ChIP qPCR, RT-qPCR, Western blotting and immunofluorescence microscopic assays showed that NRF1 mediated transcriptional changes of its target genes CXCR4, BNIP3, and DJ-1 correlated with malignant phenotypic changes. The suppression of NRF-1 activity induced growth arrest and apoptosis, and reduced stemness, growth and tumorigenic property of cancer stem cells. In summary, our findings for the first time showed that transcriptional regulation of CXCR4 by NRF1 may contribute in the induction of a pre-malignant phenotype by estrogen presumably by promoting generation of breast cancer stem cells. These data suggest NRF1 as an emerging potential target for therapeutic intervention against breast cancer.

Citation Format: Jayanta Kumar Das, Deodutta Roy. Transcriptional regulation of chemokine receptor 4 (CXCR4) by nuclear respiratory factor 1 (NRF1) controls estrogen-induced malignant transformation of breast epithelial cells to breast cancer stem cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3312.