bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2784: Simultaneous checkpoint-checkpoint or checkpoint-costimulatory receptor targeting with bispecific antibodies promotes enhanced human T cell activation
Authors: Hedvat Michael , Bonzon Christine , Bernett Matthew J. , Moore Gregory L. , Avery Kendra , Rashid Rumana , Nisthal Alex , Schubert Suzanne , Varma Rajat , Lee Sung-Hyung , Bogaert Liz , Leung Irene W.L. , Chu Seung , Muchhal Umesh , Desjarlais John
Published: August 2018

Combination checkpoint blockade promotes productive anti-tumor clinical responses often associated with an increase in immune-related adverse events. We developed optimized bispecific antibody candidates that simultaneously engage PD1 and CTLA4 (XmAb20717), CTLA4 and LAG3 (XmAb22841), and PD1 and ICOS (XmAb23104), that preferentially bind to cells co-expressing the targeted receptors. Because tumor infiltrating lymphocytes (TILs) typically express multiple immune checkpoints and costimulatory receptors, we hypothesized that these bispecific antibodies will enable selective targeting of tumor-reactive TILs, leading to safer and more cost-effective combination checkpoint blockade. The PD1 x CTLA4 bispecific antibody XmAb20717 and the PD1 x ICOS bispecific antibody XmAb23104 promoted superior in vitro T cell activation when compared to an anti-PD1 bivalent antibody. The CTLA4 x LAG3 bispecific antibody XmAb22841 combines productively with an anti-PD1 bivalent antibody achieving additive T cell activation through triple checkpoint blockade. Treatment with bispecific antibodies caused superior engraftment and activation of human T cells in NSG mice compared to an anti-PD1 bivalent antibody. Bispecific antibodies also potentiate allogeneic anti-tumor activity against human cancer cells in vivo. Each bispecific antibody promoted a unique RNA gene signature suggesting distinct mechanisms of action. These bispecific antibodies demonstrate compelling immune modulatory activity suggesting clinical development is warranted for the treatment of human malignancies.

Citation Format: Michael Hedvat, Christine Bonzon, Matthew J. Bernett, Gregory L. Moore, Kendra Avery, Rumana Rashid, Alex Nisthal, Suzanne Schubert, Rajat Varma, Sung-Hyung Lee, Liz Bogaert, Irene W.L. Leung, Seung Chu, Umesh Muchhal, John Desjarlais. Simultaneous checkpoint-checkpoint or checkpoint-costimulatory receptor targeting with bispecific antibodies promotes enhanced human T cell activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2784.
Abstract 2875: Calycosin is a double-edged sword in the treatment of pancreatic cancer
Authors: Zhang Zhu , Ko Joshua KS
Published: August 2018

Calycosin is known for its suppression of breast cancer cell growth. We are the first group to report the effects of calycosin in pancreatic cancer development. Growth inhibition of the human pancreatic cell line MIA PaCa-2 was evaluated by MTT assay. Cell cycle distribution was determined by flow cytometry. Cell migratory activity was investigated by a wound healing assay. Subcutaneous xenografting of MIA PaCa-2 cells was conducted in Balb/C nude mice, while orthotopic inoculation of the mouse Pan01 pancreatic cancer cells was performed in C57BL/6 mice. Our results have shown that calycosin exhibited modest growth inhibition of MIA PaCa-2 cells, together with induction of S phase arrest and restoration of p53. Besides, promotion of both caspase-dependent apoptosis and autophagy implicate that the anticancer mechanism of calycosin strongly rely on programmed cell death. Alternatively, calycosin also inhibited the migration of MIA PaCa-2 cells in line with downregulation of epithelial–mesenchymal transition (EMT) and pro-metastatic biomarkers. The above in vitro benefits of calycosin was confirmed by a significant reduction of tumor mass in the MIA PaCa-2 xenograft. Nevertheless, when we look closely on the in vivo anticancer effect of calycosin in the Pan01 orthotopic mice model, although both attenuation of tumor growth and proapoptotic action were confirmed by histological findings, the tumor tissue expression of Snail, vimentin and CD31 was all elevated, indicating possible promotion of EMT and pro-angiogenic potential of the drug. These controversial findings suggest that calycosin could be an effective anticancer agent by promoting programmed cell death and inhibition of cell proliferation of pancreatic cancer cells, while it may also facilitate metastatic development of tumor, particular at the advanced stages. It would be essential to further dissect the conditions and factors that differentiate the dual actions of calycosin in pancreatic cancer therapy.

Citation Format: Zhu Zhang, Joshua KS Ko. Calycosin is a double-edged sword in the treatment of pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2875.
Abstract 3012: Urban neighborhood and residential factors associated with breast cancer in African American women: A systematic review
Authors: Smith Brandi P. , Madak-Erdogan Zeynep
Published: August 2018

Residential characteristics in urban neighborhoods impacts health and might be an important factor contributing to health disparities, especially in the African American population. The purpose of this systematic review is to understand the relationship between urban neighborhood and residential factors and breast cancer incidence and prognosis in African American women. Using PubMed and Web of Science, the existing literature was reviewed. Observational, cross-sectional, cohort, and prospective studies until February 2017 were examined. Studies including populations of African American women, setting in “urban” areas, and a measure of a neighborhood or residential factor were reviewed. Four parameters related to neighborhood or residential factors were extracted including: neighborhood socioeconomic status (nSES), residential segregation, spatial access to mammography, and residential pollution. Our analysis showed that African American women living in low nSES have greater odds of late stage diagnosis and mortality. Furthermore, African American women living in segregated areas (higher percentage of Blacks) have higher odds of late stage diagnosis and mortality compared to White and Hispanic women living in less segregated areas (lower percentage of Blacks). Late stage diagnosis was also shown to be significantly higher in areas with poor mammography access and areas with higher Black residential segregation. Lastly, residential pollution did not affect breast cancer risk in African American women. Overall, this systematic review provides a qualitative synthesis of major neighborhood and residential factors on breast cancer outcomes in African American women.

Citation Format: Brandi P. Smith, Zeynep Madak-Erdogan. Urban neighborhood and residential factors associated with breast cancer in African American women: A systematic review [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3012.
Abstract 3055: A model of CSC converted from iPSC in the conditioned medium of HCC paving the way to establish HCC CSC
Authors: Afify Said M , Calle Anna Sanchez , Kumon Kazuki , Nawara Hend M , Khairani Apriliana C , Mahmud Hafizah , Oo Aung Ko Ko , Juan Du , Zahara Maram H , Seno Akimasa , Kasai Tomonari , Iwasaki Yoshiaki , Seno Masaharu
Published: August 2018

Hepatocellular carcinoma (HCC) represents the major histological subtype, accounting for 70%–85% of cases of primary liver cancer. Liver CSCs sustain their self-renewal remains largely unknown. The self-renewal and pluripotency of liver CSC are maintained by several signaling cascades. However, the mechanism of regulating these signaling is unknown.

In this study, we tried to convert mouse iPSCs into CSCs with the conditioned medium (CM) from human HCC cell line Huh7 cells without any genetic manipulation aiming at establishing liver CSCs.

First of all, CM was collected from confluent culture of Huh7 cells. Then, mouse iPSCs cells without MEF feeder cells were cultured in the presence of 50% CM for 4 weeks. The medium was changed every day with fresh medium containing 50% of CM. Mouse iPSCs cultured in the complete medium with LIF were used as a control.The survived cells (5x105 cells) were suspended in HBSS and injected into the liver of BALB/c nude mice. After 25 days malignant tumor was formed in the liver while benign teratoma was formed by the injection of iPSCs. Tumors were then excised and partly fixed in 10% neutral formalin buffer solution for HE staining and immunohistochemical analysis. The rest of tumors were subjected to rt-qPCR anaylsis and primary culture. Immunohistochemical analysis with GFP antibody showed that malignant tumor sustained GFP expression while teratoma from miPSCs did not. Immunocytochemistry of the primary cells from malignant tumor showed high expression of both glypican-3 (GPC3) and cytokeratin19 (CK19) when compared to that in miPSCs. Expression of stem cell markers (Nanog, Oct3/4, Sox2, Klf4) and CSC markers (CD44, EpCAM) were also confirmed by rt-qPCR in both the CSCs converted from iPSCs (miPS-Huh7cm cells) and the primary culture cells. These results indicate that the primary cells from the malignant tumor are rich in CSCs with high expression of GPC3 and CK19, which paves the way to establish a model of HCC CSC. This model should be very important and useful to assess the significant molecular mechanisms necessary to maintain HCC CSC, which will help develop effective therapy of liver cancer.

Citation Format: Said M Afify, Anna Sanchez Calle, Kazuki Kumon, Hend M Nawara, Apriliana C Khairani, Hafizah Mahmud, Aung Ko Ko Oo, Du Juan, Maram H Zahara, Akimasa Seno, Tomonari Kasai, Yoshiaki Iwasaki, Masaharu Seno. A model of CSC converted from iPSC in the conditioned medium of HCC paving the way to establish HCC CSC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3055.
Abstract 3278: Performance assessment of de novo assembly-based structural variation detection in the human genome
Authors: Xiao Chunlin ,   SEQC2 Working Group #3
Published: August 2018

Structural variations (SVs) contribute to genetic diversity of human populations, affect biologic functions, and cause various human disorders. However, accurately identifying SVs with correct sizes and locations in the human genome remains challenging due to the complexity of the human genome, limitations of sequencing technologies, and drawbacks of analysis methods. The advancement of next-generation sequencing technologies has dramatically decreased the sequencing cost, while substantially increased the lengths of the sequencing reads. Thus, using de novo assembly-based approaches for discovering a full spectrum of SVs in human genome becomes appealing. While various assembly methods have been developed and proposed for general use by the community, the relative efficiency and predictive accuracy of SVs calling based on these assembly methods have not been fully evaluated. In this study, we applied several popular de novo assembly tools to the sequencing read data that were generated using multiple sequencing technologies with technical replicates for NA12878/HG001, a well-studied individual from NIST-led Genome-in-a-Bottle (GIAB) project: a HapMap Caucasian trio and a Chinese Quartet from FDA-led Sequencing Quality Control Phase II (SEQC2) project. Assemblies and SVs callsets were generated for each of the eight samples, and repeatability in the SVs of the technical replicates and reproducibility across sequencing sites were evaluated. The assembly-based SVs callsets have been compared with alignment-based SVs callsets. These results allow better understanding of the impacts of de novo assembly methods on SVs calling, thus providing a better insight to precision medicine.

Citation Format: Chunlin Xiao, SEQC2 Working Group #3. Performance assessment of de novo assembly-based structural variation detection in the human genome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3278.
Abstract 3432: Evolving genomes of breast cancer aromatase inhibitor resistance models
Authors: Hoppe Reiner , Kandabarau Siarhei , Fan Ping , Jordan V Craig , Brauch Hiltrud B.
Published: August 2018

Acquired endocrine resistance is a severe barrier in the anti-hormonal treatment of breast cancer and is responsible for its failure in about half of the treated patients. In vitro models of long-term estrogen deprived breast cancer cells serve as laboratory tools to explore new therapeutic strategies. The models MCF-7:5C and MCF-7:2A mimic clinical AI resistance but regress with E2 due to the reconfiguration of survival signaling known as E2-inducible apoptosis (5C rapid, 2A delayed reaction). Alterations of stress related pathways including the accumulation of endoplasmic reticulum stress, oxidative stress, and inflammatory stress that occur prior to E2-induced apoptosis (Ariazi et al. 2011; Fan et al. 2013, 2015; Sweeney et al. 2014) and specific miRNA expression profiles (Hoppe et al Oncotarget 2016) have been linked with these resistance and E2-inducible apoptosis phenotypes. Little is known on their underlying genetic variability. Here we compare the mutational profiles of 5C and 2A clones with those of the estradiol stimulated growth sensitive MCF-7:WS8 and standard MCF-7 (ATCC) cells. Whole genome libraries were sequenced on the Illumina HiSeq platform (2x150bp reads). Alignments were manually inspected (bcftools mpileup) to avoid false negative variant calling in poorly (<20x) covered regions. Loci with at least 10x coverage and absence of any variant reads were classified as wildtype. The coding genome positions defined as mutant or wildtype in all four cell types served as a basis to select candidate mutations for the verification of MCF-7 variants via comparison to the COSMIC database (http://cancer.sanger.ac.uk/cosmic) and identification of novel mutations by variant calling (GATK HaplotypeCaller). Putative protein effects of variants were predicted by the SnpEff tool. We verified 421 point mutations in MCF-7 (COSMIC) of which 203 mutations were lost in our models, with additional specific losses of 4, 4, and 18 mutations in WS8, 2A and 5C, respectively. In 5C cells new mutations affected 261 genes with 31 high impact mutations based on splice site alterations in 7 genes (i.e. CERS2, SEMA3A, TDRD7, ATP5H) and gain of stop codons in 24 genes (i.e. CACNA1S, NOTCH4, SUN1, JPH3, SERPINB2). Similarly, in 2A cells there were 12 high impact mutations among 122 affected genes (e.g. ARHGEF5, ARHGAP35, ARAP2). We will report on the integrative genomic analysis including larger structural variants (short indels and CNVs) towards the evolution of these models for a comprehensive understanding of the biology of E2-inducible apoptosis.

Citation Format: Reiner Hoppe, Siarhei Kandabarau, Ping Fan, V Craig Jordan, Hiltrud B. Brauch. Evolving genomes of breast cancer aromatase inhibitor resistance models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3432.
Abstract 3544: Ras GTPases are modified by SUMOylation
Authors: Choi Byeong Hyeok , Chen Changyan , Philips Mark , Dai Wei
Published: August 2018

RAS proteins are GTPases that participate in multiple signal cascades, regulating crucial cellular processes including cell survival, proliferation, differentiation, and autophagy. Mutations or deregulated activities of RAS are frequently the driving force for oncogenic transformation and tumorigenesis. Given the important roles of the small ubiquitin-related modifier (SUMO) pathway in controlling the stability, activity, or subcellular localization of key cellular regulators, we investigated here whether RAS proteins are post-translationally modified (i.e., SUMOylated) by the SUMO pathway. We observed that all three RAS protein isoforms (HRAS, KRAS, and NRAS) were modified by the SUMO3 protein. SUMOylation of KRAS protein, either endogenous or ectopically expressed, was observed in multiple cell lines. The SUMO3 modification of KRAS proteins could be removed by SUMO1/sentrin-specific peptidase 1 (SENP1) and SENP2, but not by SENP6, indicating that RAS SUMOylation is a reversible process. A conserved residue in RAS, Lys-42, was a site that mediates SUMOylation. Results from biochemical and molecular studies indicated that the SUMO-E3 ligase PIASγ specifically interacts with RAS and promotes its SUMOylation. Moreover, SUMOylation of RAS appeared to be associated with its activation. In summary, our study reveals a new post-translational modification for RAS proteins. Since we found that HRAS, KRAS, and NRAS can all be SUMOylated, we propose that SUMOylation might represent a mechanism by which RAS activities are controlled.

Citation Format: Byeong Hyeok Choi, Changyan Chen, Mark Philips, Wei Dai. Ras GTPases are modified by SUMOylation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3544.
Abstract 3560: Targeting the MIF receptor CD74 in bladder cancer tumorigenesis
Authors: Penticuff Justin , Shanmugam Sambantham , Warrick Joshua , Chase Brian , Abbott Erika , Woolbright Benjamin L. , Taylor John A.
Published: August 2018

Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that can stimulate cellular proliferation in multiple tumors. Our previous data indicates genetic deletion of MIF results in T1 stage arrest in the N-Butyl-N-(4-hydroxybutyl) nitrosamine (BBN) induced bladder cancer model. Similarly, pharmacological inhibition of MIF results in lower stage tumors and reduced bladder weights when given therapeutically. CD74 is a human leukocyte antigen class II trafficking and processing protein that also functions as an extracellular receptor for MIF. Additionally, CD74 complexes with CD44 to act as the cognate MIF receptor complex, and upregulation of CD74 has been noted in high grade bladder cancer. We hypothesized CD74 would be upregulated in malignant bladder cancer cell lines, and knockout of CD74 would reduce bladder cancer tumorigenesis in the murine BBN model. The objectives of this study were to determine the effect of CD74 knockout on development and progression of urothelial cell carcinoma; as well as evaluate expression of the putative MIF signaling axis in BCa cell lines. BCa cell lines HTB-9, HTB-5, RT-4, T24 and HT-1376 and benign urothelial UROtsa cells were analyzed for MIF, CD74, and CD44 expression via quantitative PCR or western blot to assess expression of the MIF signaling axis. CD74 and CD44 expression by q-PCR were dramatically elevated in BCa cell lines vs. benign cell lines. Protein levels of CD74 and CD44 were elevated in multiple cell lines as well. MIF expression levels did not change between cell lines. To ascertain differences in BCa tumorigeneis, WT C57Bl/6J mice or CD74-/- mice were given 0.05% BBN in the drinking water for 22 weeks. Bladders were weighed and evaluated for pathologic staging by histology. After exposure to BBN, bladders from CD74-/- mice had a 33% reduction in weight vs. WT bladders (p=.062). Pathological analysis revealed lower stage tumors in bladders of CD74-/- mice (33% incidence; 6.7% pTa, 0% pTis, 6.7% pT1, 20% pT2, 6.7% pT3) as compared to bladders of WT mice (81.3% incidence; 0% pTa, 6.3% pTis, 25% pT1, 12.5% pT2, 37.5% pT3), which was a statistically significant reduction in tumor stage. Lower tumor stage correlated significantly with the CD74-/- genotype (p=0.02, Fisher's Exact Test). These data suggest CD74 likely mediates a portion of the effects of MIF in BCa, raising the possibility of both CD74 dependent, and independent MIF signaling pathways.

Citation Format: Justin Penticuff, Sambantham Shanmugam, Joshua Warrick, Brian Chase, Erika Abbott, Benjamin L. Woolbright, John A. Taylor. Targeting the MIF receptor CD74 in bladder cancer tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3560.
Abstract 3720: Augmented loading efficiency of doxorubicin into glioma-derived exosomes by an integrated microfluidic device
Authors: Thakur Abhimanyu , Zou Heng , Yang Mengsu , Lee Youngjin
Published: August 2018

Glioma is a lethal brain cancer in the central nervous system (CNS). The therapeutic usage of potent anticancer drugs for glioma is restricted by their poor penetration across the blood-brain barrier (BBB). Current reports have revealed that cell-derived exosomes can cross the BBB, suggesting their prospective use as drug delivery vehicles for targeting glioma as well as neurological disorders. However, conventional isolation of intact, specific, and optimal exosomes for drug delivery from various donor cells is laborious and time-consuming. Particularly, increasing loading efficiency of non-hydrophobic anticancer drugs, such as doxorubicin, into exosomes has been challenged. Therefore, a single integrated microfluidic device has been developed, based on the principles of immunocapture and density-gradient-free centrifugation, using soft lithography for efficient exosome isolation and drug loading consecutively. The isolated glioma-exosomes were characterized by transmission electron microscopy (TEM), immunogold-EM, and nano tracking analyzer. The loading efficiency of doxorubicin into glioma-exosomes was evaluated by UV-visible spectrophotometer as well high performance liquid chromatography (HPLC). Furthermore, the doxorubicin loaded glioma-exosomes were treated to autologous and heterologous glioma cells, followed by evaluation with confocal microscopy. The present study has shown that our integrated microfluidic device isolates pure exosomes from glioma cells, and it dramatically increases loading efficiency of doxorubicin into the exosomes, supporting its usefulness to study the exosome-mediated drug delivery for glioma therapy.

Citation Format: Abhimanyu Thakur, Heng Zou, Mengsu Yang, Youngjin Lee. Augmented loading efficiency of doxorubicin into glioma-derived exosomes by an integrated microfluidic device [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3720.
Abstract 3731: Deciphering the transcriptional role of estrogen receptor alpha in ovarian cancer
Authors: Lee Irene , Brown Myles
Published: August 2018

Ovarian cancer is the leading cause of death among gynecologic malignancies. Despite years of research, the 5-year survival rate for ovarian cancer patients has remained poor and the standard of care is largely the same as it was decades ago. Epidemiological evidence and previous studies have suggested a role for steroid hormones in the pathogenesis of ovarian cancer, however, the role of steroid hormones in ovarian cancer remains understudied. We hypothesized that Estrogen Receptor α (ERα) is transcriptionally active and drives a transcriptional program that is sufficient to promote ovarian cancer cell proliferation and survival in a subset of ER+ ovarian cancers. Preliminary data generated in our laboratory has suggested that estradiol (E2) treatment increases cell proliferation in PEO1 cells, an ERα+ ovarian cancer cell line. This increase in cell proliferation can be inhibited by co-administration with the selective estrogen receptor modulator, Tamoxifen, and the selective estrogen receptor degrader, Fulvestrant. To further interrogate the mechanism of action of ERα in ovarian cancer, we performed RNA-seq on PEO1 cells treated with Vehicle, 10 nM E2, 100 nM Fulvestrant, and E2+Fulvestrant for 24 hours. We identified 659 significantly differentially expressed genes following E2 treatment. Fulvestrant inhibited the majority of E2-induced differentially expressed genes, confirming that these genes are dependent upon ERα. Gene Set Enrichment Analysis (GSEA) indicated that the Hallmark early and late estrogen responses are enriched in the E2 dataset, confirming that classical ER activity is intact in these cells. Furthermore, we identified G2/M checkpoint as positively enriched in our dataset, indicating that proliferation genes are upregulated by ERα in these cells. Additionally, we found that apoptosis was negatively enriched in our dataset, indicating that apoptosis pathway genes are suppressed by E2 treatment. Future investigation in this project will center on exploring the mechanisms of ERα transcriptional activity through examining the ERα-dependent cistrome, regulation of target genes, and its interactions with other cofactors. These data will give us insight into how ER is regulated and what pathways and processes ER is driving. Additionally, we will use CRISPR/Cas9 screening to uncover the key genes downstream of ER that are executing the estrogen-dependent effects on proliferation and survival. We believe that these studies will provide additional drug targets that may suggest a combinatorial therapeutic approach in conjunction with endocrine therapies for the treatment of ovarian cancer.

Citation Format: Irene Lee, Myles Brown. Deciphering the transcriptional role of estrogen receptor alpha in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3731.
Abstract 3783: Mitochondrial metabolism in tumor-associated macrophages: The role of MIF and tumor-derived lactate
Authors: Noe Jordan , Rendon Beatriz , Kim Eun Jung , Mitchell Robert
Published: August 2018

Purpose: Within the tumor microenvironment, tumor-associated macrophages (TAMs) polarized to an "M2" phenotype promote angiogenesis, metastasis, and the suppression of anti-tumor immune responses. Previously, we have found that macrophage migration inhibitory factor (MIF) promotes lactate-enhanced M2-polarization, however the underlying mechanisms were not elucidated. As cancer cells that exhibit the Warburg Effect secrete copious amounts of lactate into the tumor microenvironment, highly glycolytic cancers may promote tumor progression by producing lactate to support an immunosuppressive tumor microenvironment. The goal of this study is to investigate the mechanism by which MIF and tumor-derived lactate support pro-tumorigenic M2-TAMs.

Methods: Using primary bone marrow-derived macrophages (BMDMs), we explored the mechanisms of MIF and lactate-enhanced M2 polarization using metabolic inhibitors and gene expression assays. To investigate the role of MIF in mitochondrial metabolism, we conducted extracellular flux analysis to determine mitochondrial OCR and ECAR in macrophages polarized to an M0 and M2 phenotype.

Results: Blocking mitochondrial pyruvate uptake reduces both M2 polarization and HIF1α stabilization. Blocking mitochondrial metabolism and respiration inhibits M2 polarization. MIF deficiency decreases mitochondrial OCR, NRF2 stability, and M2 polarization. Increasing CSN5 activity "rescues" the loss of M2 polarization with MIF deficiency.

Conclusions: Tumor-derived lactate per se does not promote M2 polarization but instead requires subsequent metabolism to pyruvate. Additionally, we have identified that mitochondrial pyruvate metabolism is a previously undetermined requirement for M2 macrophages. Further, we have found a potential link between MIF and metabolism, where MIF likely regulates CSN5 activity and NRF2 stability to increase mitochondrial respiration. As inhibiting mitochondrial respiration blocks M2 polarization, MIF may promote lactate-enhanced M2 polarization by enhancing mitochondrial metabolism. Subsequent studies will be conducted to further investigate the metabolic pathways and mechanisms required for the mitochondrial metabolism of tumor-derived lactate in promoting M2-TAM polarization.

Citation Format: Jordan Noe, Beatriz Rendon, Eun Jung Kim, Robert Mitchell. Mitochondrial metabolism in tumor-associated macrophages: The role of MIF and tumor-derived lactate [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3783.
Abstract 3797: Chromosome instability as a prognostic factor& an immunotherapeutic target in acute myeloid leukemia
Authors: Jin Ning , Burkard Mark
Published: August 2018

Chromosome instability (CIN) is an increased rate of chromosome mis-segregation due to errors in mitosis. CIN generates genetic and karyotypic diversity and is common in hematological malignancies. Low to moderate levels of CIN are well tolerated, and moderate levels can promote cancer proliferation by generating genetic diversity for tumor evolution. However, very high levels of CIN are lethal and enhancing CIN can be therapeutic. Therefore, CIN status could be prognostic for clinical outcomes as well as a therapeutic target. Despite its promise, little is known about CIN in AML.

We first determined if CIN is correlated with clinical outcome in patients with acute myeloid leukemia (AML). We quantified the chromosome mis-segregation frequency (lagging chromosomes and chromatin bridges) in dividing AML cells on bone marrow core biopsies. Our study reveals a wide distribution of CIN in AML core biopsy specimens, ranging from 20% to 67%. High CIN correlates with a better overall survival in patients with AML.

Recent data suggest that chromosome mis-segregation promotes senescence and initiates immune clearance. Our hypothesis is that chromosome instability is immunogenic due to DNA damage, proteotoxic/oxidative stress with a high mutagenic potential, which can be recognized by immune system. To test this, we used an inhibitor of Monopolar spindle 1 (Mps1) to enhance CIN and tested if that altered immunogenicity. Mps1 is a key component of the mitotic checkpoint and is also involved with chromosome alignment. Inhibition of Mps1 by AZ3146 results in severe chromosome mis-segregation, micronuclei formation, and apoptosis in AML cell lines. In addition, AZ3146 treatment causes activation of γH2AX due to DNA damage, and calreticulin exposure from endoplasmic reticulum stress. This generates danger signals to activate innate immune system (natural killer cells and dendritic cells) and induces immunogenic cell death (ICD) through priming of cytotoxic T cells. This effect is mainly seen in synchronized mitotic cells, suggesting AZ3146 acts specifically on mitosis. Additionally, we observed the overexpression of programed death ligand-1 (PD-L1) after the treatment of AZ3146 in AML cells. As evidence showed, PD-L1 participates in the immune evasion and protects AML blasts from cytotoxic T cells in mouse models of leukemia.

Taken together, our observations in Mps1 inhibitor treatment indicate the immunogenicity of CIN in AML. Additionally, these data support combination treatment of mitotic checkpoint inhibitor with immunotherapy to improve therapeutic efficacy in AML patients.

Citation Format: Ning Jin, Mark Burkard. Chromosome instability as a prognostic factor& an immunotherapeutic target in acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3797.
Abstract 3831: Development of near-infrared photoimmunotherapy targeting GD2-positive neuroblastoma
Authors: Nouso Hiroshi , Tazawa Hiroshi , Tanimoto Terutaka , Tani Morimichi , Oyama Takanori , Sato Hiroaki , Noma Kazuhiro , Kagawa Shunsuke , Kobayashi Hisataka , Noda Takuo , Fujiwara Toshiyoshi
Published: August 2018

Background: Neuroblastoma (NB) is a primary malignant tumor of the peripheral sympathetic nervous system. Despite the recent advances in the treatment of NB, patients with high-risk NB still have poor prognosis. Immunotherapy using anti-GD2 monoclonal antibody (mAb), which binds to disialoganglioside GD2 on the surface of NB cells, has recently become a standard treatment for high-risk NB. However, the therapeutic efficacy of anti-GD2 mAb is insufficient in many clinical trials. Therefore, the enhancement of therapeutic potential of anti-GD2 mAb is needed to improve the clinical outcome of patients with high-risk NB. Near-infrared photoimmunotherapy (NIR-PIT) is a promising antitumor strategy to enhance the therapeutic potential of immunotherapy using an antibody-photoabsorber conjugate (APC). In this study, we investigated the therapeutic efficacy of anti-GD2-based NIR-PIT using anti-GD2 mAb conjugated to the photoabsorber IR700 (anti-GD2-IR700) in human NB cells.

Methods: We used 3 human NB cell lines, including IMR-32, CHP-134 and SK-N-SH. The expression of GD2 on the cell surface of NB cells was analyzed by flow cytometric analysis. Anti-GD2-IR700 was synthesized by incubating anti-GD2 mAb and IR700. The mixture was purified with a Sephadex G50 column. The antitumor effect of anti-GD2-IR700 was evaluated using XTT assay. NB cells were incubated with anti-GD2-IR700, anti-GD2 mAb or PBS and NIR light was irradiated at 2.0J/cm2.

Results: IMR-32 and CHP-134 cells, but not SK-N-SH cells, expressed GD2 protein on the cell surface. In GD2-positive IMR-32 and CHP-134 cells, administration of anti-GD2-IR700 significantly suppressed the cell viability compared to anti-GD2 mAb or PBS when combined with NIR light irradiation. In contrast, in GD2-negative SK-N-SH cells, anti-GD2-IR700 showed no significant inhibition of cell viability in combination with NIR light irradiation.

Conclusion: These results suggest that NIR-PIT using an anti-GD2-IR700 is a promising antitumor strategy to promote the therapeutic efficacy of anti-GD2 immunotherapy for high-risk NB.

Citation Format: Hiroshi Nouso, Hiroshi Tazawa, Terutaka Tanimoto, Morimichi Tani, Takanori Oyama, Hiroaki Sato, Kazuhiro Noma, Shunsuke Kagawa, Hisataka Kobayashi, Takuo Noda, Toshiyoshi Fujiwara. Development of near-infrared photoimmunotherapy targeting GD2-positive neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3831.
Abstract 3881: Formulating a cell membrane lipid-extracted nanoliposome (CLENS) drug platform for the treatment of prostate cancer
Authors: Omaer Abubker , Alqarni Ali , Randulfe Carlos , Alharbi Hanan , Campbell Robert
Published: August 2018

Background: Liposome-encapsulated doxorubicin has proven to be an effective drug delivery method to cancer cells. Lipid extracts derived from human prostate cancer cells along with conventional components were employed to develop cell membrane lipid-extracted nanoliposomes (CLENs) for prostate cancer treatment. In an effort to balance efficiency of drug loading with cellular targeting efficiency we report on physicochemical characteristics (i.e., particle size, zeta potential, drug loading efficiency) and cellular uptake studies.

Methods: Five preparations of CLENS were prepared consisting of variable concentrations of DOPC and CRL-1740 lipid extract material varying as a function of optimized fixed ratios of conventional components (Cholesterol and DSPE-PEG5000). The controls used to investigate the effect of the different components on drug incorporation efficiency were preparations of DOPC (100%) and CRL-1740 (100%). Doxorubicin hydrochloride (5 mol%) was employed during the preparation of all formulations. Samples were readily prepared by thin film evaporation. For cellular uptake studies CRL-1740 cells were seeded at 1 x 104 mL in 48-well plate and incubated for 24 hours prior to use. The cells were next exposed to various amounts of rhodamine labeled CLENS for 24 hrs. Cellular uptake was determined using fluorescence intensity values determined by a fluorescence microplate reader. Fluorescence microscopy supplement cellular uptake studies.

Results: The DOPC drug-loaded liposome preparation consistently achieved the lowest percent of drug incorporation. Likewise, 100% CRL-1740 CLENs demonstrated a significant degree of formulation instability, resulting in a relatively low percent of drug incorporation compared to preparations that consisted of lipid extract and conventional components. The finding were supported by values determined for zeta potential and particle size. Cellular uptake studies also favored the preparations that consisted of lipid extract material and conventional components compared to each alone. The results collectively support the importance of balancing lipid extract material derived from prostate cancer target cells with cholesterol and DSPE-PEG5000 for formulation development and cellular uptake.

Conclusion: We formulated seven preparations of CLENS as part of the drug platform. The most promising candidates were determined following completion of the study. Future studies should evaluate the cytotoxicity and safety profile of the most promising formulations vivo.

Citation Format: Abubker Omaer, Ali Alqarni, Carlos Randulfe, Hanan Alharbi, Robert Campbell. Formulating a cell membrane lipid-extracted nanoliposome (CLENS) drug platform for the treatment of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3881.
Abstract 3889: RPL28 promoter polymorphism rs4806668 is associated with reduced survival in FOLFIRI-treated metastatic colorectal cancer patients
Authors: Labriet Adrien , Lévesque Éric , Mattia Elena De , Cecchin Erika , Jonker Derek , Couture Félix , Simonyan David , Buonadonna Angela , D'Andrea Mario , Villeneuve Lyne , Toffoli Giuseppe , Guillemette Chantal
Published: August 2018

Fluorouracil (5-FU), folinic acid, and irinotecan (FOLFIRI) is a standard treatment of metastatic colorectal cancer (mCRC). Our study aimed to investigate genetic variability in candidate genes in relation to patients' outcomes using two independent cohorts of 417 FOLFIRI-treated mCRC cases recruited in Canada and Italy. We used a haplotype-tagging polymorphism (htSNP) approach to maximize the coverage of genetic variability of the selected genes and genotyping was performed using time-of-flight mass spectrometry iPLEX Sequenom Technology. Associations between polymorphisms and clinical outcomes were tested using Cox proportional hazards model adjusted for covariates. Of the genes tested, RPL28 encodes a ribosomal protein and its silencing was shown to enhance sensitivity to 5-FU and irinotecan in vitro.1 RPL28 rs4806668G>T was associated with a shorter progression-free survival (PFS) in the Canadian (hazard ratio (HR) 3.23, P = 0.013), Italian (HR 3.28, P = 0.021) and combined (HR 3.36, P < 0.001) cohorts. This marker was also associated with a reduced overall survival (OS) in the Canadian (HR 3.09, P = 0.032), Italian (HR 3.05, P = 0.030) and combined (HR 3.07, P = 0.002) cohorts. These carriers of the rs4806668 TT genotype associated with reduced survival represent less than 5% of the population of European ancestry and its frequency varies greatly among ethnic groups. This htSNP, located in the promoter region of RPL28, is in strong linkage disequilibrium with six other polymorphisms and may affect RPL28 gene expression. In support, the rs4806668T allele was associated with increased RPL28 expression in colon tissues of healthy individuals of the GTEx cohort, compared to rs4806668G carriers. Functional investigations are required to elucidate the underlying molecular mechanism. These results suggest a role for the ribosomal RPL28 protein in cancer cell response to FOLFIRI treatment.

This work is supported by the Canadian Institutes of Health Research.

1Allen et al. Mol Cancer Ther. 2012 Jan;11(1):119-31

Citation Format: Adrien Labriet, Éric Lévesque, Elena De Mattia, Erika Cecchin, Derek Jonker, Félix Couture, David Simonyan, Angela Buonadonna, Mario D'Andrea, Lyne Villeneuve, Giuseppe Toffoli, Chantal Guillemette. RPL28 promoter polymorphism rs4806668 is associated with reduced survival in FOLFIRI-treated metastatic colorectal cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3889.
Abstract 4190: Synergistic anticancer effect of sulforaphane and cisplatin on sirtuins-mediated epithelial-to-mesenchymal signaling in triple negative breast cancer
Authors: SINHA SONAM , Shrivastava Neeta
Published: August 2018

Purpose:

Triple negative breast cancer (TNBC) overexpresses sirtuins (SIRTs), a class III HDACs, which induces epithelial-to-mesenchymal (EMT) transition and facilitates Cisplatin (CIS) resistance. We hypothesized that Sulforaphane (SFN), an HDAC inhibitor, when combined with CIS, could potentiate CIS efficacy against TNBC metastasis and overcome its resistance.

Material and Methods:

The effect of SFN and CIS combination on cellular proliferation and apoptosis of two TNBC cell lines, MDA-MB-231 and MDA-MB-468 were assessed by MTT assay and Annexin and PI method followed by FACS analysis respectively. The anti-metastatic capabilities of the compounds were evaluated using wound healing assay, invasion assay, chemotaxis assay, colony formation and tumorsphere assay. The regulation of key proteins involved in sirtuin-mediated EMT signaling and metastasis was determined by zymography and western blot analysis.

Results:

The results revealed that SFN in combination with CIS synergistically inhibited TNBC cells through inhibition of cellular proliferation, induction of apoptosis, in-silico and in-vitro modulation of SIRTs (1-7) expression. Anti-metastatic capabilities of the combinatorial treatment regimen were associated with inhibition of migratory, invasive, mammosphere forming potential and downregulation of matrix metallopeptidase (MMP-9 and MMP-2) activity of TNBC cells. When TNBC cells were treated with SFN, the expression of epithelial marker, E-cadherin was up-regulated, while the expression of mesenchymal markers, ZEB1, SNAIL, TWIST and associated SIRTs were down-regulated. In contrast, when the cells were treated with CIS, the expressions of mesenchymal markers were up-regulated. Interestingly, the combinatorial treatment of SFN and CIS synergistically inhibited EMT transition in TNBC cells. In-vivo studies are warranted to confirm our results.

Conclusion:

Collectively our findings demonstrated a SIRTs-mediated EMT-suppression leading to synergistic antitumor effect by combination of SFN and CIS in TNBC cells. This combination therapy may serve as a novel therapeutic strategy for TNBC management.

Citation Format: SONAM SINHA, Neeta Shrivastava. Synergistic anticancer effect of sulforaphane and cisplatin on sirtuins-mediated epithelial-to-mesenchymal signaling in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4190.
Abstract 4193: Biochanin A in pancreatic cancer
Authors: Desai Vilas , Tadinada Satya Murthy , Lai James C K , Bhushan Alok
Published: August 2018

Pancreatic cancer, with its dismally low survival rate (<5%), is one of the most aggressive forms of cancer and is the fourth leading cause of cancer-related deaths in the U.S. Gemcitabine, the first-line drug approved by the FDA, has been marginally effective as it improves survival by only six months. Epidemiologic studies have implicated a role for dietary isoflavones in lowering the incidence of different cancers in the Asian population. Isoflavones such as genistein, biochanin A and daidzein have been tested for their anticancer potential in different cancers; however, the clastogenic effect associated with genistein limits its therapeutic potential. Primarily used in management of hyperlipidemia and cardiovascular diseases, statins have been documented to exert antiproliferative effects. We hypothesized that a combination treatment of biochanin A and atorvastatin exerts enhanced anticancer effects on pancreatic cancer cells viz. AsPC1, PANC-1 and MIAPaCa-2. Our studies showed a decrease in cell viability on combination drug treatment with differential effects on mitogenic and cell signaling pathways in the cell lines investigated. In PANC-1 cells, biochanin A and atorvastatin combination treatment decreased invasiveness, dampened the expression of activated STAT3, and critical mediators of cell cycle progression. We also observed the involvement of enhanced proapoptotic effect of the combination treatment in AsPC-1 cell line and potential cell cycle arrest phenomenon in PANC-1 cells. Cell metabolism studies using seahorse XFp analyzer showed an increase in mitochondrial respiration and a decrease in glycolytic activity on combination treatment. Further, the XF-Plasma Membrane Permeabilizer (XF-PMP) assay used to evaluate the mitochondrial ETC complex activity showed a complete impairment of complex I and II (prominently), with MIAPaCa-2 being more susceptible to the combination treatment than AsPC1 and PANC-1. Together, the combination treatment-induced effects on pancreatic cancer cell lines may have potential implications in developing novel and improved therapies for the treatment of pancreatic cancer.

Citation Format: Vilas Desai, Satya Murthy Tadinada, James C K Lai, Alok Bhushan. Biochanin A in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4193.
Abstract 4200: New potential therapeutic applications of WX-UK1, as a specific and potent inhibitor of human trypsin-like proteases
Authors: Oldenburg Emil , Schar Christine R. , Lange Eva , Plasse Terry F. , Abramson Danielle T. , Fathi Reza , Towler Eric M. , Levitt Mark , Jensen Jan K.
Published: August 2018

WX-UK1 (the active metabolite of upamostat) was originally developed as an inhibitor of the serine protease urokinase (uPA) with a Ki ~1 uM. To identify more sensitive targets, we performed a bioinformatic analysis of the ~200 human trypsin-like serine proteases, many of which play crucial roles in homeostasis and disease. Among these we selected a subset for biochemical analysis based on an inspection of modelled 3D structures of WX-UK1:protease complexes and sequence alignment of binding site residues. Samples of the selected proteases were prepared and characterized for their binding to WX-UK1; enzymatically with respect to inhibition constant (Ki) and by surface plasmon resonance with respect to dissociation constant (kd). We now report that WX-UK1 is a potent and specific inhibitor of five human serine proteases (trypsin-3, trypsin-2, trypsin-1 and matriptase-1 and trypsin-6), with Ki's down to the low nanomolar range, 19 nM for trypsin-3. Several of these serine proteases are known to be associated with cancer progression and metastasis. As a compound with an established clinical safety profile, targeted use of upamostat in oncology and extension to non-oncology indications may be assessed.

Citation Format: Emil Oldenburg, Christine R. Schar, Eva Lange, Terry F. Plasse, Danielle T. Abramson, Reza Fathi, Eric M. Towler, Mark Levitt, Jan K. Jensen. New potential therapeutic applications of WX-UK1, as a specific and potent inhibitor of human trypsin-like proteases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4200.