bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2158: Pharmacologic, pharmacodynamic action of MELK kinase inhibitor OTS167 in cancer cells
Authors: Chung Suyoun , Kijima Kyoko , Harada Yosuke , Takamatsu Naofumi , Miyamoto Takashi , Matsuo Yo , Nakamura Yusuke
Published: July 2016

Cancer is the second most common cause of death in US in 2015 and has been the leading cause of death in Japan since 1981. Despite of improved therapeutic strategies in cancer, only limited treatment options can still be available to a subset of patients. Many efforts have been made to develop molecular targeted drugs, but the success rate did not reach to patients’ expectation. Unlike the conventional chemotherapy, the use of maximum tolerated dose (MTD) and the measurement of bulk tumor volume are not always appropriate for the clinical evaluation of molecular targeted drugs. To optimize and personalize the dosing for targeted agents, detailed molecular pathway and biomarkers that are affected by the drug should be elucidated. OTS167 was developed as a novel and potent MELK kinase inhibitor that we previously reported. The expression of MELK is elevated in various human cancers both in solid and hematological tumor and MELK is known to be associated with cancer progression and poor prognosis. Because MELK is indicated its critical roles in cancer stem cell proliferation as well, targeting MELK is an attractive and promising therapeutic strategy for cancer patient.

Here, we report the molecular mechanism of action of OTS167 in preclinical model. OTS167 suppressed MELK activity and promoted MELK protein degradation by inhibition of autophosphorylation. OTS167-treated cells showed drastic morphological transformation with the induction of p53 and p21 expression. We also investigated the expression of stem cell markers to elucidate whether OTS167 suppresses cancer stem-like properties through inhibition of MELK. Furthermore, we evaluated antitumor activity of OTS167 using human xenograft model and molecular changes in tumor tissue. The expression of MELK and downstream molecules were decreased in OTS167-treated xenograft tumor tissues by IHC. The change of expression and pharmacological effect were well correlated. Our data provide the evidence for the concept that OTS167 suppresses tumor growth through the inhibition of MELK signaling pathway and suggest the possibility of biomarkers for the assessment of clinical efficacy.

Citation Format: Suyoun Chung, Kyoko Kijima, Yosuke Harada, Naofumi Takamatsu, Takashi Miyamoto, Yo Matsuo, Yusuke Nakamura. Pharmacologic, pharmacodynamic action of MELK kinase inhibitor OTS167 in cancer 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 2158.
Abstract 2169: High content screening in MCF7 and MCF10A cells show differential responses depending on oxygen levels and mechanistic readout for viability
Authors: Yan Michelle , O’Grady Michael , April Anderson , Clarke Scott , Low Quentin , DeMarco Carolyn , Calderon Veronica , Kihn Kathy , Montoya Leticia , Finnessy Carmen , Wickett Marcy
Published: July 2016

Oxygen levels in typical cell culture conditions do not accurately reflect the oxygen levels cells are exposed to within the body. Furthermore, oxygen levels can vary within the tumor microenvironment. These variances can affect how cells respond to a variety of drugs and small molecules often used during cancer treatment. Previous studies have shown altered drug sensitivities in MCF7 cells depending on the oxygen levels the cells are grown in. To further understand how oxygen levels affect drug sensitivity, the response of tumorigenic MCF7 cells were compared to non-malignant MCF10A cells, cultured under low and high oxygen.

The goal of this study was to examine the differences in the sensitivity of MCF7 and MCF10A cells to drugs from the Tocriscreen Total compound library when cultured under low and high oxygen levels and for different drug exposure time periods. Cells were grown using standard cell culture conditions (19% oxygen) or low oxygen (5%). Using high-content imaging and high-throughput analysis methods, viability was assessed using two different mechanistic readouts; cellular metabolic activity and membrane permeability. Post-screening analysis was performed to confirm positive hits by performing dose-responses and determining IC50 concentrations using the same reagents as in the screen, and reagents to assess oxidative stress and cellular proliferation.

Results showed that viability differed depending on mechanistic readout and platform method used to determine hits as well as duration of exposure to compound. In addition, the response of MCF10A cells was not always identical to that of the MCF7 cells. Post-screening analysis of “hits” indicated different potencies of compounds tested depending on oxygen level and cell type. These data suggest that some drugs may affect MCF7 and MCF10A cells differently depending on environmental oxygen levels and mechanistic readout used to determine cellular health.

Citation Format: Michelle Yan, Michael O’Grady, Anderson April, Scott Clarke, Quentin Low, Carolyn DeMarco, Veronica Calderon, Kathy Kihn, Leticia Montoya, Carmen Finnessy, Marcy Wickett. High content screening in MCF7 and MCF10A cells show differential responses depending on oxygen levels and mechanistic readout for viability. [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 2169.
Abstract 2180: Drug-like inhibitors of SGK1: Discovery and optimization of low molecular weight fragment leads
Authors: Zapf James , Meyer Todd , Wade Warren , Lingardo Laura , Batova Ayse , Alton Gordon , Pallai Peter
Published: July 2016

Triple negative (ER, PR and HER2/neu negative) breast cancers (TNBC) constitute 15-24% of breast cancers, but account for a disproportionate share of mortality. TNBC tumors are more aggressive, lack a targeted therapy, and are prone to relapse and metastasis after cytotoxic drug treatments, the only drug therapy currently available. Serum and glucocorticoid-regulated kinase 1 (SGK1) promotes the growth, invasiveness and chemo-resistance of cancer cells and is overexpressed in 48% of breast cancers, yet is not detected in normal breast tissue. In particular, SGK1 is overexpressed in TNBC cells and knockdown of SGK1 blocks the proliferation and invasiveness of TNBC cells. Thus, SGK1 is an attractive targeted therapy for TNBC. In the SGK1 inhibitors that have been reported previously, poor physicochemical properties have prevented these inhibitors from being active in vivo and advancing to the clinic. To discover drug-like inhibitors of SGK1, we used our technology platform (Leap-To-Lead™) and a fragment-based screening approach to find SGK1 inhibitor scaffolds. Low molecular weight inhibitors like ours are more readily optimized for potency while retaining drug-like properties. From this screen, we identified 11 scaffolds as novel inhibitors of SGK1. Four scaffold series had clear structure activity relationships (SAR). To illustrate the potential of these scaffolds, we carried out a limited synthetic expansion around one scaffold and improved potency more than 600-fold (SGK1 IC50 = 1 μM; LE = 0.4). Several compounds dose-dependently inhibited the proliferation of a TNBC cell line (MDA-MB-231) and inhibited the phosphorylation of the biomarker protein N-Myc downstream regulated 1 (NDRG-1). Unlike previously reported SGK1 inhibitors, our fragment lead compounds show excellent cellular penetration in Caco-2 assays.

In summary, a fragment-based screening approach was used to identify four novel scaffolds with clear SAR trends. A single scaffold series was further optimized and shows anti-proliferative activity and biomarker modulation in a TNBC cell line. These SGK1 inhibitor series represent excellent starting points for further SAR studies and development of a novel preclinical candidate.

Citation Format: James Zapf, Todd Meyer, Warren Wade, Laura Lingardo, Ayse Batova, Gordon Alton, Peter Pallai. Drug-like inhibitors of SGK1: Discovery and optimization of low molecular weight fragment leads. [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 2180.
Abstract 2190: Isolated cannabidiol from Cannabis sativa plant extracts inhibit growth and induce apoptosis in cervical cancer cells
Authors: Motadi Lesetja Raymond , lukhelo. sindiswa
Published: July 2016

Cervical cancer remains a global health related issue among females of Sub-Saharan Africa, with over half a million new cases reported each year. Different therapeutic regimens have been suggested in various regions of Africa, however, over a quarter of a million women die of cervical cancer, annually. Therefore, it is important to search for new drugs through effective screening of medicinal plant extracts to identify lead anti-cervical cancer drugs. The aim of this study was to evaluate for the anti-growth effects of Cannabis sativa extracts and its isolate, cannabidiol on cervical cancer cell lines HeLa, SiHa, and ME-180. To determine for the presence of important constituents and evaluate for the anti-growth effects, phytochemical screening, MTT assay, cell growth analysis, flow cytometry, morphology analysis, Western blot, caspase 3/7 assay, and ATP measurement assay were conducted were conducted. Results obtained indicate that both plant extracts induced cell death at an IC50 of 50 - 100μg/ml and the Inhibition of cell growth was cell line dependent. Flow cytometry confirmed that, with or without cell cycle arrest, the type of induced cell death was apoptosis. Cannabis sativa extracts led to the up-regulation of apoptosis proteins (p53, Bax, caspase-3, and caspase-9) and the down regulation of anti-apoptosis proteins (Bcl-2 and RBBP6), signalling the execution of apoptosis. Apoptosis induction was further confirmed by morphological changes, an increase in Caspase 3/7 and a decrease in the ATP levels. In conclusion, this data implies Cannabis sativa crude extracts has the potential to inhibit growth and induce apoptosis in cervical cancer cell lines, which may be due to the presence of cannabidiol.

Citation Format: Lesetja Raymond Motadi, sindiswa lukhelo. Isolated cannabidiol from Cannabis sativa plant extracts inhibit growth and induce apoptosis in cervical cancer 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 2190.
Abstract 2191: Imaging the kinetics of targeted nanoparticles to pancreatic cancer
Authors: Moreno Jonathan , Fei Qin qin , Lambros Maria P.
Published: July 2016

Purpose: The purpose of this work is to image the kinetics of targeted lipid nanoparticles to an SCL transporter of pancreatic cancer using a near infrared (NIR) fluorescent dye DyLight® 747 and a mouse orthotopic model of fluorescent pancreatic cancer.

Methods: Initially and before the in vivo experiments, the uptake of nanoparticles by human pancreatic cancer cells was evaluated in vitro using fluorescent microscopy and flow cytometry. A quantity of 1.5 x105 PANC-1, MIA PaCa, or BxPC-3 cells/ml was placed in each well of a six-well plate and incubated for 5 hours with different formulations of fluorescent lipid targeted and non-targeted nanoparticles encapsulation daunorubicin which has natural fluorescence. After incubation, the growth media were removed and the cells were washed with PBS. The uptake of the fluorescent lipid nanoparticles by the cancer cells was evaluated an EVOS® microscope. All the in vitro experiments were done in triplicate. For the in vivo targeting studies, nude mice (Balb/c Ola Hsd-Fox1 nu) were implanted orthotopically (in the tail of the pancreas) with 3 x106 PANC-1 cells expressing GFP. The injection volume for the implantation was 50 microliters and contained 3 x106 PANC-1-GFP cells mixed with serum free media and HC Matrigel (1:1). One month after the orthotopic implantation, the implanted pancreatic tumor was visualized in live, anesthetized animals using UVP Scientia (UVP, Upland, CA). The animals were separated into three groups (three animals in each group) and injected via the tail vein as follows: Group A, was injected with only the fluorescent dye, Dylight 747; group B was injected with fluorescent non-targeted nanoparticles; and group C was injected with fluorescent targeted nanoparticles.

Results: Both the flow cytometry and fluorescence microscopy data show a significantly higher uptake of the targeted lipid nanoparticles compared to the non-targeted ones. The imaging of the fluorescent lipid nanoparticles in live mice shows that both targeted and nontargeted nanoparticles reach the pancreatic tumor. The nontargeted nanoparticles reside less than 48 hours on the tumor, whereas the targeted nanoparticles can reside for 96 hours on the tumor. The non-encapsulated fluorescent dye did not reach the tumor and was excreted in the urine.

Conclusion: The results show that Dylight® 747 can be used to image the kinetics of a drug delivery system in a live animal. Furthermore while both lipid nanoparticles reach the tumor, the targeted lipid nanoparticles reside significantly longer on the tumor and this may translate to a better therapeutic outcome.

Citation Format: Jonathan Moreno, Qin qin Fei, Maria P. Lambros. Imaging the kinetics of targeted nanoparticles to pancreatic cancer. [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 2191.
Abstract 2199: TLR3-induced immune response using PLGA nanoparticles for dendritic cell-based cancer immunotherapy
Authors: HAN. HEEDONG
Published: July 2016

Dendritic cells (DCs) are potent professional antigen presenting cells (APCs) capable of initiating a primary immune response and activating T cells, and they play a pivotal role in host immune responses to cancer. Prior to antigen presentation, efficient antigen and adjuvant uptake by DCs is necessary to induce their maturation and cytokine generation. Nanoparticles (NPs) are capable of intracellular delivery of both antigen and adjuvant to DCs. Here, we developed an advanced poly(lactic-co-glycolic acid) (PLGA)-NP encapsulating both ovalbumin (OVA) as model antigen and poly I:C as adjuvant to increase intracellular delivery, and promote DC maturation. These NPs were taken up by mouse primary DCs and their uptake greatly facilitated MHC class I antigen presentation in vitro. Moreover, vaccination with PLGA-NP treated DCs led to the generation of OVA-specific CD8+ T cells, and the resulting antitumor efficacy was significantly increased in EG.7 tumor-bearing mice compared to control (p < 0.01). Taken together, we demonstrate that this PLGA-NP platform may be an effective method for delivering tumor-specific antigens or adjuvant to DCs.

Citation Format: HEEDONG HAN. TLR3-induced immune response using PLGA nanoparticles for dendritic cell-based cancer immunotherapy. [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 2199.
Abstract 2215: Pembrolizumab and ipilimumab reduce human RKO colon carcinoma tumor growth in HLA matched CD34-NSG humanized mice
Authors: Lasaro Marcio , Davis Jason M. , Ellison Mark , Graf Martin R. , Mahajan Vivek , Synnott Aidan , Mullin Robert J. , Miliani De Marval. Paula L.
Published: July 2016

Over the past decade there has been an increasing demand for preclinical models useful for evaluating the efficacy of checkpoint inhibition-based cancer immunotherapies. The recently developed humanized mouse models composed of various types of human immune cells has the potential to recapitulate the human anti-tumor response modulated by checkpoint inhibitors. Tumor cells may develop multiple mechanisms to evade immune surveillance, including PD-L1 upregulation which suppresses T-cell effector responses towards the tumor. In the present study, we evaluated the efficacy of the checkpoint inhibitors pembrolizumab (anti-PD-1) and ipilimumab (anti-CTLA-4) in the human RKO colon carcinoma xenograft model in CD34-NSG humanized mice. These mice were engrafted with human hematopoietic stem cells (HSC) that matched the HLA haplotype of the RKO cells (HLA-A*01). Flow cytometry analysis confirmed high level of expression of PD-L1 in RKO colon carcinoma cells. The results from this study revealed that pembrolizumab and ipilimumab monotherapies significantly inhibited tumor growth. Surprisingly, combination therapy did not provide additive or synergistic effects and resulted in the same level of efficacy as the individual regimens. In order to understand the impact of these therapies on distinct immune cell populations, we analyzed the distribution of CD4 and CD8 T lymphocytes, Treg cells, NK cells and B cells in peripheral blood, spleen and tumor samples from tumor bearing humanized mice, via flow cytometry. We found that these checkpoint inhibitors can enhance effector functions of CD4+ and CD8+ T cells associated with increased expression of IFN-Γ. A parallel study with these checkpoint inhibitors carried out with humanized mice engrafted with HSC of the same haplotype as the RKO cells but from a different donor resulted in identical anti-tumor efficacy. In summary, these studies validate the use of humanized mouse models to test the tumor response to immune-checkpoint based therapies as it shows significant anti-tumor growth as a result of pembrolizumab and ipilimumab monotherapies associated with activation of T lymphocytes.

Citation Format: Marcio Lasaro, Jason M. Davis, Mark Ellison, Martin R. Graf, Vivek Mahajan, Aidan Synnott, Robert J. Mullin, Paula L. Miliani De Marval. Pembrolizumab and ipilimumab reduce human RKO colon carcinoma tumor growth in HLA matched CD34-NSG humanized 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 2215.
Abstract 2226: Anti-human PD-1 antibody BGB-A317 exhibits potent immune cell activation
Authors: Zhang Tong , Song Jing , Li Yucheng , Ma Jie , Shi Yingdi , Wu Xiaoran , Xu Lanlan , Liu Qi , Cao Yanping , Wang Yali , Peng Hao , Hou Hongjia , Zhao Xu , Song Xiaomin , Wang Lai , Wei Min , Luo Lusong , Li Kang
Published: July 2016

Background: PD-1 is a “check point” inhibitory receptor, which is primarily expressed in activated and/or exhausted T cells. Engagement of PD-1 receptor by its ligand PD-L1 or PD-L2 (expressed on APCs and some tumor cells), leads to negative regulatory signaling in T-cells, promoting functional exhaustion of tumor-infiltrating T-lymphocytes. BGB-A317 is a novel humanized IgG4 anti-PD-1 antibody under clinical development. The immunomodulatory activity of BGB-A317 was evaluated in in vitro assays and reported here.

Materials and methods: BGB-A317 was generated through hybridoma fusion, humanized by CDR grafting and structural simulation. The Fc and hinge regions of BGB-A317 were engineered to remove Fc gamma receptor (FcγR) binding and to stabilize the hinge region. The binding affinity and specificity were studied by ELISA, FACS and SPR (Biacore). The immunomodulatory functions of BGB-A317 were evaluated using both T-cell lines and primary immune cells.

Results: BGB-A317 binds to the extracellular domain of human PD-1 with high affinity (KD = 0.15nM) and specificity. In competition assays, BGB-A317 efficiently blocks the interactions between PD-1 and its ligands. In vitro cell-based assays, BGB-A317 significantly enhances cytokine (IL-2 and IFN-γ) production using T-cell line or primary T cells cocultured with signal-sending cells in a dose-dependent manner. BGB-A317 also potently increases NK cell-mediated IFN-γ production and cytotoxicity against PD-L1+ tumor cells. As an important functional attribute, BGB-A317 exhibits no binding to any of FcγRs, including FcγRI, FcγRIIA, FcγRIIB and FcγRIIIA. Therefore, no antibody-dependent cell-mediated (ADCC) or complement-dependent cytotoxicity (CDC) has been observed in activated T cells treated with high concentration of BGB-A317 up to 100 μg/ml. BGB-A317 is stable as bivalent antibody unlike the wild type human IgG4 going through dynamic conformational change, forming bispecific antibody by Fab-arm exchange.

Conclusions: BGB-A317 demonstrated potent immune cell activation in in vitro assays, supporting its clinical development for the treatment of human cancers.

Keywords: Anti-PD-1, immunotherapy, functional assays

Citation Format: Tong Zhang, Jing Song, Yucheng Li, Jie Ma, Yingdi Shi, Xiaoran Wu, Lanlan Xu, Qi Liu, Yanping Cao, Yali Wang, Hao Peng, Hongjia Hou, Xu Zhao, Xiaomin Song, Lai Wang, Min Wei, Lusong Luo, Kang Li. Anti-human PD-1 antibody BGB-A317 exhibits potent immune cell activation. [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 2226.
Abstract 223: Comparison of human-specific versus cross-reactive Complex I inhibitor for in vivo tumor efficacy
Authors: Christian Sven , Algire Carolyn , Schwede Wolfgang , Mowat Jeffrey S. , Ehrmann Alexander , Menz Stephan , Bauser Marcus , Haegebarth Andrea
Published: July 2016

Mitochondria are both key regulators of energy supply and apoptotic cell death. The mitochondrial electron transport chain (ETC) consists of four enzyme complexes that transfer electrons from NADH to oxygen. During electron transfer, the ETC (Complex I to IV) pumps protons into the inter-membrane space, generating a gradient across the inner mitochondrial membrane that is used by Complex V to drive ATP synthesis. Recent publications have shown that tumor cells harboring specific mutations (LKB1, mIDH and others) are more sensitive to Complex I inhibition, compared to cells that do not have these mutations. We have identified an optimized human/mouse cross-reactive Complex I inhibitor that allows profiling of Complex I inhibitors in pharmacological models. We have pursued different approaches based on the literature, an unbiased screen and in-house results generated with the human-specific Complex I inhibitor BAY 872243 to identify sensitive in vivo tumor models. However, using the cross-reactive Complex I inhibitor we were unable to identify sensitive models apart from weakly sensitive LKB1-deficient tumors (A549, G361) when animals were treated at maximum tolerated dose (MTD). In addition, all approaches for combination therapy failed to improve efficacy in vivo. Direct comparison of human-specific Complex I inhibitor BAY 87-2243 and cross-reactive inhibitor BAY179 in a sensitive LKB1-deficient melanoma model, G361, demonstrated that inhibition of Complex I specifically in the tumor is a valid approach as it results in tumor growth inhibition of ∼50%. However, cross-reactive compounds do not reach exposures at MTD to generate comparable effects.

Citation Format: Sven Christian, Carolyn Algire, Wolfgang Schwede, Jeffrey S. Mowat, Alexander Ehrmann, Stephan Menz, Marcus Bauser, Andrea Haegebarth. Comparison of human-specific versus cross-reactive Complex I inhibitor for in vivo tumor efficacy. [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 223.
Abstract 2230: Evaluation of PD-L1 expression in ovarian carcinosarcoma by immunohistochemistry and RNAscope in-situ hybridization
Authors: Liang Li , Chen Jun-Song , Jiang Yi , Niu Na , Zhang Connie , Park Emily , Ma Xiao-Jun , Liu Jinsong
Published: July 2016

Ovarian carcinosarcoma, also called malignant Müllerian mixed tumor, is a rare, but very aggressive biphasic tumor with both malignant epithelial and stromal components. PD-L1 is one of the ligands of immune inhibitory receptor PD-1. PD-L1/PD-1 pathway suppresses T cell immune response, increases programmed cell death of activated T cells and plays an important role in tumor immune escape. Anti-PD-L1 immunotherapy has shown promising results in certain malignancies, such as melanoma and lung cancer. Therefore, we performed PD-L1 immunohistochemical staining on 30 specimens of ovarian carcinosarcoma collected from patients that underwent surgeries at our institution between 2002 and 2015. In addition, we performed RNAscope in-situ hybridization to examine PD-L1 mRNA expression in 16 specimens that were collected within past 5 years (2010-2015). Our result demonstrated focally membranous expression of PD-L1 protein in 23 of 30 cases (77%). PD-L1 mRNA expression correlated with membranous, but not cytoplasmic expression of PD-L1 protein. In conclusion, PD-L1 protein and mRNA are frequently expressed in ovarian carcinosarcoma, suggesting anti-PD-L1 immunotherapy may be helpful in these patients.

Citation Format: Li Liang, Jun-Song Chen, Yi Jiang, Na Niu, Connie Zhang, Emily Park, Xiao-Jun Ma, Jinsong Liu. Evaluation of PD-L1 expression in ovarian carcinosarcoma by immunohistochemistry and RNAscope in-situ hybridization. [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 2230.
Abstract 2231: IO125 is a potent inducer of tumor immunogenicity
Authors: Mylavarapu Sanghamitra , Kaushik Shelly , Yadav Yashpal , Roy Monideepa , Sengupta Aniruddha , Sengupta Shiladitya
Published: July 2016

Combinatorial treatments with chemotherapeutics and immunotherapies are currently being investigated in several cancers. Chemotherapeutics can increase the immunogenicity of tumors besides modulating the immune system. Here we describe the development of a novel platinum-based therapeutic, IO-125, which assembles into stable nanoscale supramolecular structures. IO125 demonstrates enhanced activity compared with standard Pt drug across cancer lines. IO-125 preferentially targets tumors in vivo, and demonstrated improved efficacy in triple negative breast cancer (4T1), Lewis lung carcinoma (LLC) and melanoma (B16/F10) tumor models. Treatment with IO-125 elevates expression of immune-markers (including IGKC, PD-L1, PD-1, CD8A), which are associated with good prognosis. In addition, combinatorial treatment with IO-125 and antibody against immune checkpoint inhibitor(s) resulted in superior antitumor efficacy compared to treatment with antibody alone. This study shows that supramolecular therapeutics, especially IO125, can emerge as a novel approach to focally modulate the tumor immune contexture.

Citation Format: Sanghamitra Mylavarapu, Shelly Kaushik, Yashpal Yadav, Monideepa Roy, Aniruddha Sengupta, Shiladitya Sengupta. IO125 is a potent inducer of tumor immunogenicity. [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 2231.
Abstract 2238: Understanding immune phenotypes and their spatial relationships to breast adenocarcinoma in FFPE tissues
Authors: Zheng Yi , Thuse Pallavi , Liu Linying , Stack Edward C. , Campisano Michael , Johnson Kent , Unfricht Darryn , Narayanan Nara , Hoyt Clifford , Rajopadhye Milind
Published: July 2016

The relationship between elements of the immune system and breast tumors in situ requires an approach that leverages multiplexed immunohistochemistry (mIHC) with multispectral imaging to facilitate precise image analyses. To achieve this, we developed a novel 7-color mIHC assay, based on tyramide signal amplification, that allowed us to reliably interrogate CD3, CD4, CD8, FoxP3, CD68, and cytokeratin, in formalin-fixed, paraffin-embedded (FFPE) samples of human breast cancer. Imaging was performed using the multispectral Mantra system and inForm image analysis software. Using this specific mIHC panel and the cell segmenting and phenotyping tools in inForm, we were able to reliably identify cytotoxic T cells (CD3+ CD8+), helper T cells (CD3+ CD4+), regulatory T cells (CD3+ CD4+ FoxP3+), tumor associated macrophages (CD68+) and breast tumor cells (CK+). With cell phenotypes within the tumor microenvironment determined based on specific colocalized staining combinations, we then employed spatial point pattern analyses to examine spatial relationships between specific phenotypes. With this analysis, we were able to describe distances between cytotoxic T cells and regulatory T cells, cytotoxic T cells and tumor associated macrophages, as well as cytotoxic T cells and breast tumor cells. With this combined mIHC, multispectral imaging and advanced image analysis, we demonstrate a novel method that allows for unique tumor microenvironment assessments within in breast cancer. Through the preservation of tumor architecture available in archival FFPE tissues, these methods can advance our understanding of unique tumor microenvironment interactions, and could provide the ability to stratify responses to immunotherapies.

Citation Format: Yi Zheng, Pallavi Thuse, Linying Liu, Edward C. Stack, Michael Campisano, Kent Johnson, Darryn Unfricht, Nara Narayanan, Clifford Hoyt, Milind Rajopadhye. Understanding immune phenotypes and their spatial relationships to breast adenocarcinoma in FFPE tissues. [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 2238.
Abstract 224: The role of the microphthalmia associated transcription factor in autophagy regulation in melanoma
Authors: Moller Katrin , Sigurbjornsdottir Sara , Dilixiati Remina , Brynjolfsdottir Solveig H. , Larue Lionel , Thorsson Vesteinn , Steingrimsson Eirikur , Ogmundsdottir Margret H.
Published: July 2016

Autophagy is considered a therapeutic target in various cancer types, including melanoma. The transcription factors TFEB and TFE3 have been shown to play a key role in autophagy regulation in various cell types by affecting gene expression required for this process. A close relative to these factors is the transcription factor MITF; often termed the master regulator of melanocyte development and function. MITF activity has also been shown to be important for various stages of melanoma formation and progression.

Analysis of The Cancer Genome Atlas expression data from melanoma tumors reveals that MITF expression correlates with lysosomal and autophagy gene expression in metastatic tumors. Furthermore, analysis of expression data from 23 melanoma cell lines shows that MITF expression correlates with lysosomal and autophagy gene expression, and the analysis of available MITF ChIP sequencing data shows that MITF binds the promoters of lysosomal and autophagy genes. This binding was experimentally verified by ChIP experiments in 501Mel melanoma cells. MITF over-expression and knock down experiments show effects of MITF on lysosomal and autophagy gene expression, as well as an effect on the autophagy activity.

Autophagy is high in melanoma and has been linked to tolerance against cancer treatment. We have found that MITF regulates autophagy genes and activity in melanoma cell lines. In light of both MITF and autophagy being considered a therapeutic target in melanoma, this relationship needs to be taken into consideration.

Citation Format: Katrin Moller, Sara Sigurbjornsdottir, Remina Dilixiati, Solveig H. Brynjolfsdottir, Lionel Larue, Vesteinn Thorsson, Eirikur Steingrimsson, Margret H. Ogmundsdottir. The role of the microphthalmia associated transcription factor in autophagy regulation in melanoma. [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 224.
Abstract 2245: FDG PET quantification for prediction of early recurrence in patients with stage I and II non small cell lung cancer
Authors: Burger Irene A. , Arvanitakis Michael , Steiger Seraina , Weder Walter , Hillinger Sven
Published: July 2016

Background: Although surgical resection remains the optimal treatment for early-stage NSCLC, up to 50% of patients with stage I and II relapse and die within 5 years after curative resection. Therefore prognostic markers are important as these patients might benefit from adjuvant therapy. The goal of this study was to evaluate established PET quantification metrics including: maximal standard uptake volume (SUVmax), metabolic tumor volume (MTV) and total lesion glycolysis (TLG) as prognostic markers for early recurrence and overall survival in resected early stage lung cancer.

Methods: Between January 2003 and December 2010 182 surgically resected patients with stage I-II NSCLC who underwent 18 F FDG PET/CT less than one month prior to surgery have been evaluated. All patients had at least 5 years of follow-up. Cox proportional hazard model was used to determine the association between variables and survival respectively time to recurrence. For the multivariate analysis the following variables have been included: tumor size on CT, age tumor stage, histology, SUVmax, TLG (for TLG42% (threshold at 42% SUVmax) and TLG2.5 (cut-off at SUV 2.5) and MTV42% and MTV2.5).

Results: 133 patients were included, 71 with adeno carcinoma, 62 with squamous cell carcinoma. TLG2.5 and MTV2.5 values have been a significant prognostic factor for recurrence (P<0.0001). Patients with a MTV2.5 above 42 cm3 had a mean recurrence time of 0.8±0.9 years, while patients with MTV2.5 ≤ 42 cm3 recurred within 2.8±1.3 years. TLG2.5 and MTV2.5 PET volume metrics have not been predictable for overall survival in adenocarcinoma patients, recurrence or overall survival in squamous cell cancer. SUVmax, TLG42% and MTV42 were not predictable for early recurrence or survival for both histologies.

Conclusions: TLG2.5 and MTV2.5 may be useful prognostic variables in stage I-II NSCLC depending on the tumor type. Using a cut-off at 42 cm3 for early stage adenocarcinoma patients a high risk of recurrence within one year might be identified and adjuvant therapy following surgical resection could improve outcome for those patients.

Citation Format: Irene A. Burger, Michael Arvanitakis, Seraina Steiger, Walter Weder, Sven Hillinger. FDG PET quantification for prediction of early recurrence in patients with stage I and II non small cell lung cancer. [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 2245.
Abstract 2247: A multicenter clinical trial of lung cancer circulating tumor cell assay with the largest sample size (1210 cases) in China
Authors: Lou Jiatao , Zhou Caicun , Wu Jing , Qiao Lihua , Liang Xiaohui , Wang Xiaoqian , Chen Xiaoxia , Li Xuefei , Zhao Chao
Published: July 2016

Background: As a novel biomarker of primary tumors, circulating tumor cells (CTCs) are well known to play an important role in cancer diagnosis, recurrence and metastasis, and disease monitoring. This study investigated the application of CTCs in lung cancer therapy by quantitative determination of folate receptor-positive CTCs.

Methods: This study enrolled 1210 subjects (including 560 lung cancer patients, 350 patients with benign lung diseases, 150 healthy subjects and 150 non-lung cancer malignant tumor patients) and quantified the tiny amounts of CTCs in peripheral blood by negative enrichment using immunomagnetic beads in combination with folate receptor-directed polymerase chain reaction (PCR).

Results: Following ROC curve analyses of CTC levels in the patients with benign lung diseases or benign lung diseases and the healthy subjects, the cut-off of CTCs was 8.70 CTC Units/3mL with a specificity of 88.2% (441/500) and a sensitivity of 79.6% (446/560) as determined by the method mentioned above. The ROC area under

curve of more than 0.8 (AUC = 0.8797, P<0.0001) suggested the excellence of ligand-directed PCR to diagnose lung cancer. For stage II-IV lung cancer patients, the sensitivities were 70.8%(34/48), 79.6% (125/157), and 90.2% (185/205) respectively, and the sensitivity in stage I patients can reach up to 68.0% (102/150) as well. CTC levels appeared to have greater accuracy to diagnose lung cancer (AUC: 0.8833, P<0.0001) than the combination of the other four lung cancer-specific serum tumor markers (CEA, NSE, CYFRA21-1 and SCC)(AUC: 0.8557, P<0.0001). CTCs were related to tumor stage and tumor size other than pathological type. By quantitative determination of folate receptor-positive cells in 150 different types of cancers, the CTC level in lung cancer was found to be higher than those in gastric, breast, colorectal, liver, and esophageal cancers.

Conclusions: LT-PCR achieves quantitative determination of folate receptor-positive CTCs to diagnose lung cancer with high sensitivity and specificity, and significantly greater diagnosis accuracy than the model of combined lung cancer-specific serum tumor markers.

Citation Format: Jiatao Lou, Caicun Zhou, Jing Wu, Lihua Qiao, Xiaohui Liang, Xiaoqian Wang, Xiaoxia Chen, Xuefei Li, Chao Zhao. A multicenter clinical trial of lung cancer circulating tumor cell assay with the largest sample size (1210 cases) in China. [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 2247.
Abstract 226: PI3K pathway inhibition induces a different metabolic response in NSCLC cells harboring WT and G12C mutant KRAS
Authors: Caiola Elisa , Brunelli Laura , Marabese Mirko , Pastorelli Roberta , Lupi Monica , Broggini Massimo
Published: July 2016

Lung cancer is the most common cancer worldwide and, because of its high fatality, it also represents the most common cause of cancer related deaths. Non-small-cell lung cancers (NSCLCs) constitute about 85% of lung cancers and the 5-year survival rate is currently below 20%. Only a limited number of NSCLC patients, carrying alterations in specific genes, can benefit from small molecules therapies, highlighting the need for new treatment strategies to improve the outcome of the remaining patients.

KRAS is among the most frequently mutated oncogene in NSCLC and 90% of the mutations affect codon 12, with the G12C substitution being the most frequent. KRAS mutations in NSCLC are supposed to be associated to a poor prognosis and poor response to chemotherapy. Despite KRAS being one of the earliest known oncogenic drivers in NSCLC, it is considered practically undruggable. Mutations in KRAS gene lead to the activation of PI3K/akt/mTOR pathway, whose effective inhibition remains a challenging clinical target.

Since PI3K/akt/mTOR pathway and KRAS oncogene mutations have both a role in cancer cell metabolism, we investigated whether the activity of PI3K/akt/mTOR inhibitors (BEZ235 and BKM120) in cells harboring different KRAS status is related to their effect at metabolic level.A targeted metabolomics approach was applied to a robust and well characterized isogenic system to profile the metabolic adaptations occurring in KRAS-G12C and KRAS-wild type (WT) cells in response to treatment with BEZ235 and BKM120. KRAS-G12C and KRAS-WT harboring cells displayed similar sensitivities to BEZ235 and BKM120 treatments. Metabolomics analysis indicated that the different KRAS isoforms induced a distinct metabolic response to PI3K inhibitor treatments. In particular, impairment of glutamine and serine metabolism, in KRAS-G12C and KRAS-WT respectively, was observed after pharmacological blockade of the PI3K signaling. In addition, PI3K inhibitors caused autophagy in KRAS-WT isoform, but not in KRAS-G12C, where a striking decreased in ammonia production was found as probable consequence of glutamine metabolism impairment. At doses relevant for autophagy, differences in apoptosis and cell cycle phase distribution were not detected in KRAS-WT and KRAS-G12C clones.

These preliminary results would set the basis for more effective therapeutic combinations possibly exploiting the different metabolic response to PI3K/akt/mTOR inhibitors in NSCLC cells harboring a different KRAS status.

Citation Format: Elisa Caiola, Laura Brunelli, Mirko Marabese, Roberta Pastorelli, Monica Lupi, Massimo Broggini. PI3K pathway inhibition induces a different metabolic response in NSCLC cells harboring WT and G12C mutant KRAS. [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 226.
Abstract 2287: Final results of the pharmacodynamic (PD) data of PQR309-001, a first-in-human trial of a combined PI3K/mTOR inhibitor in advanced solid tumors
Authors: Wicki Andreas , Prêtre Vincent , Ritschard Reto , Brown Nicholas , Bize Vincent , Fabbro Thomas , Cmiljanovic Natasa , Dimitrijevic Sasa , Schmitz Deborah , Stumm Michael , Kristeleit Rebecca
Published: July 2016

Background: PQR309 is a novel, oral, balanced pan-class 1 PI3K, mTORC1 and mTORC2 inhibitor. PQR309-001 was a first-in-man dose escalation study evaluating PQR309 in patients with advanced solid tumours.

The primary objective of the study was to establish the maximum tolerated dose (MTD) of PQR309, its safety and tolerability. Secondary objectives included characterization of PQR309 pharmacokinetics (PK) and pharmacodynamics (PD).

Methods: We investigated PQR309 treatment-induced PD, including changes of 16 PI3K-mTOR and MAPK associated phospho-proteins, and 84 mRNAs in serial tumor biopsies obtained from patients enrolled in study PQR309-001. 13 patients were eligible for PD analysis after histologic evaluation of fresh-frozen tissue. A solid-phase assay with phospho-specific antibodies was used for the investigation of PI3K/mTOR and MAPK signalling. RNA analysis was performed with an RTK/PI3K-specific RNA array.

Results: Under treatment with PQR309, phospho-Akt (p_T308 and p_S473), phospho-mTOR and phospho-S6 were significantly downregulated compared to baseline (p<0.05, Wilcoxon signed-rank test). Moreover, patients with radiographic tumor shrinkage (n = 7) had significantly stronger phospho-Akt (p_T308), phospho-mTOR and phospho-S6 reduction than patients whose tumor was growing (n = 6) (p<0.05, Mann-Whitney test). Importantly, ERK1/2 phosphorylation, as the readout of MAPK activity, was decreased overall with no statistically significant difference between shrinking and growing tumors. The analysis of mRNAs showed a trend towards upregulation of PDGFRA (3.2 times, not statistically significant) after 21 days of therapy with PQR309. Intriguingly, no consistent changes of other PI3K-related RNAs was observed.

Conclusion: PD assessments in tumour tissue samples from patients treated with PQR309 demonstrated a reduction of phosphorylation of direct targets for PQR309 (PI3K and mTOR) as well as downstream effectors of mTOR. Furthermore, the data indicate that downregulation of phospho-Akt (p_T308) and mTORC1 activity may be predictive of tumor shrinkage while upregulation of PDGFRA may represent a mechanism of resistance against PI3K/mTOR inhibition.

Citation Format: Andreas Wicki, Vincent Prêtre, Reto Ritschard, Nicholas Brown, Vincent Bize, Thomas Fabbro, Natasa Cmiljanovic, Sasa Dimitrijevic, Deborah Schmitz, Michael Stumm, Rebecca Kristeleit. Final results of the pharmacodynamic (PD) data of PQR309-001, a first-in-human trial of a combined PI3K/mTOR inhibitor in advanced solid 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 2287.
Abstract 2288: Targeting mTOR pathway in rare and recalcitrant tumors
Authors: Rojas Veronica , Ganesan Shridar , Hirshfield Kim M. , Dipaola Robert S. , Rodriguez-Rodriguez Lorna
Published: July 2016

INTRODUCTION: Genomic alterations affecting the PI3K/AKT/mTOR pathway are commonly seen among various cancer types [Kandoth 2013]. The objective of this study is to determine the clinical utility of using comprehensive genomic profiling (CGP) in the course of clinical care to identify genomic alterations affecting the PI3K/AKT/mTOR pathway in patients with either rare or refractory cancers.

METHODS: CGP was performed by Foundation Medicine, Inc. Genomic alterations were reviewed by members of an institutional molecular tumor board (MTB). Consensus recommendations on genomically targeted, clinically relevant FDA-approved, on- and off-label therapies and clinical trials were sent to the treating physician for their consideration as treatment alternatives. Ninety-seven tumors with alterations in the PI3K/AKT/mTOR pathway were analyzed.

RESULTS: Thirty-three patients with cancers that were either rare or refractory to standard therapy received drug targeting the PI3K/AKT/mTOR pathway. Age range was 18-78 years old. Thirteen patients were non-Hispanic White, three non-Hispanic Black, two Hispanic and one Asian. Tumors tested included 6 gynecologic, 5 renal, 2 sarcomas and one each of pancreatic, melanoma, T cell lymphoma, bladder, adrenal, and skin-not melanoma. Treatment outcomes were available for 19/33 patients (one patient died soon after presentation at the MTB and could not be treated). Seven (37%) patients derived clinical benefit from a drug targeting the PI3K/AKT/mTOR pathway; the duration of clinical benefit ranged from 90-245 days. The clinical benefit was not tumor type specific and included two endometrial tumors, 2 renal tumors, one sarcoma, one adrenal paraganglioma and one micropapillary urothelial carcinoma. Eight patients have not been started on PI3K/AKT/mTOR targeted therapy yet.

CONCLUSION: These data suggest that tumors with alterations in the PI3K/AKT/mTOR pathway may respond to targeted therapy regardless of tumor of origin.

Citation Format: Veronica Rojas, Shridar Ganesan, Kim M. Hirshfield, Robert S. Dipaola, Lorna Rodriguez-Rodriguez. Targeting mTOR pathway in rare and recalcitrant 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 2288.