bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 3722: Extracellular arginine starvation imposes DNA replication fork stall and permits DNA damage tolerance
Authors: Wang Yi-Chang , Kelso Andrew A. , Chen Yi-Hsuan , Hsieh Chi-An , Chen Wei-Kai , Stark Jeremy M. , Kung Hsing-Jien , Ann David K. 
Published: September 2020

Abstract
Regional nutrients deprivation, which is frequently encountered in many solid tumors due to poor vascularization, induces metabolic rewiring and genetic dysregulation to support the demands of proliferating cancer cells. Numerous chemotherapies exert their S-phase dependent cytotoxicity by producing deleterious effects on DNA to kill proliferating tumor cells. How regional nutrients deprivation affects the response of cancer cells to chemotherapy, especially DNA damaging genotoxicity, and what is the key nutrient dominates these effects is an active area of interest across tumor biology. Of all the amino acids we examined, only arginine is regional deprived in tumor core and indispensable for S-phase progression in multiple breast cancer cells and non-tumorigenic breast epithelial cells. We found that arginine deprivation stalls DNA replication in an mTORC1 inactivation, insufficient metabolites, or the integrated stress response pathway independent manner. We also found that low levels of newly synthesized Histone H4 marks (H4K5ac and H4K12ac), which were reported to be associated with adverse outcomes in various cancers including basal-like and HER2-positive breast cancer, are positively correlated with arginine bioavailability and could serve as surrogate markers of arginine sufficiency. Mechanistically, extracellular arginine is required for the translation of DNA-synthesis-dependent histone H4, but not H3, to promote DNA replication. Knockdown of arginyl-tRNA-transferase, like arginine deprivation, recapitulates the loss of histone H4 lysine 5 and 12 acetylation marks and compromises replication fork speed. Notably, one round of arginine deprivation and addition did not cause an increase in γH2AX, a hallmark of nuclear DNA double-stand breaks. However, multiple rounds of arginine fluctuation promotes the emergence of ATR-Chk1 checkpoint independent-nuclear p53-binding protein 1 bodies in the subsequent G1-phase of daughter cells and induces DNA damage tolerance and resistance to genotoxic agents. Thus, intratumoral variation in the arginine microenvironment synchronizes DNA replication with therapeutic response.
Citation Format: Yi-Chang Wang, Andrew A. Kelso, Yi-Hsuan Chen, Chi-An Hsieh, Wei-Kai Chen, Jeremy M. Stark, Hsing-Jien Kung, David K. Ann. Extracellular arginine starvation imposes DNA replication fork stall and permits DNA damage tolerance [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3722.
Abstract 3737: Mutant isocitrate dehydrogenase driven metabolic reprogramming results in therapeutic vulnerability to lactate dehydrogenase inhibition
Authors: Bi Aiwei , Xu Jun , Jin Nan , Lan Xiaojing , Tang Shuai , Shou Matthew , Liu Jia , Ding Jian , Geng Meiyu , Lin Shuhai , Huang Min 
Published: September 2020

Abstract
Mutant isocitrate dehydrogenase (mIDH), with a gain-of-function in the production of an oncometabolite (R)-2-hydroxyglutarate (2-HG), is considered as a promising target for cancers carrying IDH mutations. IDH1 mutations, typically involving an amino acid substitution in the active site of the enzyme in codon 132, have been identified in a broad spectrum of solid tumors. The massive production of 2-HG remodels the metabolic network in cancer cells and results in epigenetic alterations facilitating malignant phenotypes. Of note, while the pharmacological inhibition of mIDH1 effectively leads to the remarkable reduction in the intracellular 2-HG level, it seems not associated with the sufficient suppression of solid tumor growth. We hence asked whether it is possible to identify extra metabolic vulnerability in IDH1 mutant tumors. To this end, we performed 13C isotope tracing analysis of glucose and glutamine, two major nutrient supply for 2-HG production, in solid tumor cell lines including HT1080 cells with spontaneous IDH1 mutation and IDH1 mutant glioma cell lines expressing mIDH1. [U-13C6]-glucose tracking enabled us to discover that IDH1 mutated tumors exhibit increased glycolysis and decreased tricarboxylic acid (TCA) metabolism, which can be reversed by mIDH1 inhibition. Meanwhile, cell mitochondrial stress test indicated decreased oxygen consumption rate (OCR) compared with the wildtype counterpart. These data suggest that 2-HG production paralyzes TCA metabolism and subsequent oxidative phosphorylation but enhances aerobic glycolysis, thus reprogramming tumor cells prone to Warburg effect. In line with this hypothesis, lactate dehydrogenase (LDH) inhibition that shut down lactate production increased 2-HG levels in IDH1 mutant cells. Further, LDH inhibition obtained the improved response in IDH1 mutant tumors while combined inhibition of mIDH1 abolished this effect. Together, we discovered that mutant IDH1 impaired TCA cycle by production of millimolar level 2HG which may lead to more dependency on glycolysis.
Citation Format: Aiwei Bi, Jun Xu, Nan Jin, Xiaojing Lan, Shuai Tang, Matthew Shou, Jia Liu, Jian Ding, Meiyu Geng, Shuhai Lin, Min Huang. Mutant isocitrate dehydrogenase driven metabolic reprogramming results in therapeutic vulnerability to lactate dehydrogenase inhibition [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3737.
Abstract 375: In vivo metabolic autofluorescence imaging of macrophage heterogeneity across normal and cancerous tissue
Authors: Heaster Tiffany , Esbona Karla , Sondel Paul M. , Skala Melissa C. 
Published: September 2020

Abstract
Tissue microenvironmental conditions affect macrophage metabolism and function. However, standard methods to measure macrophage function require tissue dissociation (e.g. flow cytometry) or tissue fixation (e.g. immunohistochemistry), which remove the cells from their native microenvironment and prevent in vivo monitoring. Autofluorescence imaging enables non-invasive, dynamic imaging of cell metabolism in live, intact samples by measuring fluorescence from metabolic co-enzymes, NAD(P)H and FAD, naturally present in all cells.
Here, we test autofluorescence imaging to assess macrophage function in vivo within normal skin and tumor microenvironments. Transgenic mice harboring a floxed mCherry fusion protein controlled by the Csf1r promoter were crossed with Lyz2-Cre mice to generate a model of mCherry-expressing macrophages. Multiphoton wavelength mixing was used to simultaneously image mCherry, NAD(P)H, and FAD fluorescence in spectrally distinct channels. NAD(P)H and FAD fluorescence intensities and lifetimes were quantified for tissue-resident macrophages in the mouse ear dermis and tumor-associated macrophages in Panc02 pancreatic tumors engrafted on the flank.
Spatial distributions of macrophages were visualized at each tissue site. Dermal macrophages were predominantly localized around hair follicles in the ear, while tumor-associated macrophages were randomly distributed throughout the tumor. Co-localization of mCherry and FAD signal indicated high FAD intensities within macrophages across both tissues. Population density modeling of autofluorescence variables on a single cell level in each tissue site indicate metabolic heterogeneity of macrophages within normal and cancerous tissues. Immunofluorescence of standard macrophage markers (CD86/CD206) further characterize diversity in macrophage phenotypes across both tissue microenvironments. These studies demonstrate metabolic autofluorescence imaging as a novel non-invasive tool to assess tissue-specific macrophage metabolism at the single cell level in vivo. Ultimately, this imaging approach could provide insight on the role of macrophage plasticity in tumor growth and treatment response in vivo.
Citation Format: Tiffany Heaster, Karla Esbona, Paul M. Sondel, Melissa C. Skala. In vivo metabolic autofluorescence imaging of macrophage heterogeneity across normal and cancerous tissue [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 375.
Abstract 3754: PPP6C regulation of ERK signaling in melanoma
Authors: Cho Eunice , Lou Hua Jane , Turk Benjamin E. 
Published: September 2020

Abstract
Melanoma, the deadliest form of skin cancer, is characterized by aberrant hyperactivation of the ERK mitogen-activated protein kinase signaling pathway. Inhibitors of BRAF and MEK1/2, both members of this pathway, have shown clinical efficacy and are used in combination to treat BRAFV600E melanoma, the most common melanoma genomic subtype. However, not all BRAFV600E melanomas respond to these inhibitors, and those that do respond eventually acquire resistance. To better understand mechanisms of drug susceptibility in BRAFV600E melanoma, we performed a loss-of-function screen to identify kinases and phosphatases that modulate sensitivity to two clinical MEK1/2 inhibitors. In this screen, we identified PPP6C, the catalytic subunit of protein phosphatase 6 (PP6), as a factor promoting sensitivity to MEK1/2 inhibition. PPP6C is a potential tumor suppressor mutated in 7-12% of melanomas, and has roles in cell cycle regulation and DNA damage repair. shRNA-mediated knockdown and CRISPR/Cas9-mediated knockout of PPP6C in BRAF mutant melanoma cells both lead to hyperactivation of MEK1/2 and ERK1/2, resulting in decreased sensitivity to pathway inhibitors. This finding was confirmed in a panel of BRAFV600E melanoma cell lines. Through in vitro biochemical and cell-based studies, we have established that PP6 directly dephosphorylates MEK1/2. As with complete loss of PPP6C, cells expressing most recurrent cancer-associated PPP6C mutants similarly have hyperactive ERK signaling. Our studies support a novel negative regulatory role for PPP6C in ERK signaling that contributes to its activity as a tumor suppressor and promotes pathway inhibitor sensitivity in BRAFV600E melanoma.
Citation Format: Eunice Cho, Hua Jane Lou, Benjamin E. Turk. PPP6C regulation of ERK signaling in melanoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3754.
Abstract 3758: Impact of ANXA7 I1 expression on PDGFRA and MET endosomal trafficking in glioblastoma
Authors: White Zachary B. , Hurt Centdrika Dates , Rajbhandari Rajani , Nair Sindhu , Nozell Susan , Bredel Markus 
Published: September 2020

Abstract
Purpose/Objective(s): Glioblastomas (GBM) are highly malignant tumors that arise from astrocytes. GBMs rely on signaling through receptor tyrosine kinases (RTK) to ensure tumorigenicity. EGFR, PDGFRA and MET are distinct RTKs; each is capable of binding a ligand, which promotes receptor dimerization and kinase activation. Annexin A7 (ANXA7), a protein involved in membrane binding and vesicle trafficking, is spliced into 2 isoforms - isoforms 1 (I1) and isoform 2 (I2). A normal adult brain will express both isoforms, however, in GBM, I1 expression is prevented. We determined that I1, but not I2, is tumor suppressive, in part, because it promotes the endocytic degradation of EGFR. Because I1 is typically not expressed in GBM, EGFR signaling persists unabated and tumors thrive. Recently, we extended these observations and determined that I1 also impacts additional RTKs, including PDGFRA and MET. By understanding how ANXA7 I1 impacts PDGFRA and MET signaling, we hope to target multiple tumorigenic RTKs and improve therapy for patients with GBM.
Materials/Methods: To understand how and when I1 downregulates PDGFRα and MET, U251-malignant glioma (MG) cells, which only express I2 (P), were modified to express I1. Cells were serum starved overnight, and then stimulated with PDGFAA and HGF(20 ng/ml), which activate the aforementioned receptors respectively. As a positive control, cells were stimulated with EGF to activate EGFR. Total protein was then collected and analyzed by western blot analyses. Next, we sought to determine whether I1 was competent to down-regulate several RTKs coincidentally. To do this, P and I1 cells were serum starved, stimulated with EGF, PDGFAA or both, and total protein levels were evaluated.
Results: The levels of activated EGFR, PDGFRα and MET are dramatically reduced in those cells that express I1 as compared to cells expressing I2 (P). Upon activation with both PDGFAA and EGF, in cells expressing I1 (I1), levels of PDGFRα and EGFR were markedly reduced 30 minutes post-stimulation as compared to cells expressing I2.
Conclusion: Our results suggest I1 may be a master regulator of RTK endocytosis and degradation. Further investigations of the impact of I1 on endocytic trafficking patterns using immunofluorescence and confocal microscopy are underway. Collectively, understanding how I1 functions and targets multiple tumorigenic RTKs will provide the foundation for future selective therapies that restore I1 expression in GBM and reduce tumorigenicity.
Citation Format: Zachary B. White, Centdrika Dates Hurt, Rajani Rajbhandari, Sindhu Nair, Susan Nozell, Markus Bredel. Impact of ANXA7 I1 expression on PDGFRA and MET endosomal trafficking in glioblastoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3758.
Abstract 376: Intravital microscopy reveals HER2-dependent and -independent effects of myeloid cells on the single-cell biodistribution of trastuzumab
Authors: Li Ran , Weissleder Ralph , Miller Miles A. 
Published: September 2020

Abstract
Advents in single-cell profiling have revealed that tumor tissues are highly heterogenous, and this cell-to-cell heterogeneity can greatly affect therapeutic efficacy. This is especially true for monoclonal antibody (mAb) therapies that exhibit variability in both delivery and action within solid tumors. Moreover, recent studies have highlighted the importance of tumor microenvironment in determining mAb responses. Yet, it has been traditionally difficult to analyze the kinetics of mAb biodistribution and delivery at a single-cell resolution, as well as how mAb interacts with tumor-associated stromal cells in vivo. To address these issues, we developed a high-resolution intravital microscopy (IVM) pipeline to study single-cell pharmacokinetics and pharmacodynamics of mAb in a tumor consists of cancer cells with variable target expression. We applied this imaging strategy to track the in vivo binding of trastuzumab (Tzm), a HER2-targeting mAb, to fluorescently-labeled HER2 in the heterogenous tumor microenvironment. We observed efficient binding of Tzm to HER2+ cancer cells, and the amount of Tzm on cancer cells correlated with HER2 expression level. We also discovered a large accumulation of Tzm in tumor-associated macrophages (TAMs). Time-lapse IVM imaging further revealed an increase in Tzm uptake in TAMs over the period of 72 hrs, with concomitant decrease in Tzm level on the HER2+ cancer cells. Further controlled experiments demonstrated that the baseline ability of TAMs to take up Tzm was not dependent on HER2 expression in the surrounding cancer cells, yet HER2 over-expression can increase the levels of Tzm in TAMs. All in all, these results revealed crucial roles that tumor-associated stromal cells play in mAb delivery and action. They also demonstrated the utility of IVM in dissecting mAb behaviors in heterogenous tumor microenvironment, which can significantly impact our understanding of mAb treatment efficacy.
Citation Format: Ran Li, Ralph Weissleder, Miles A. Miller. Intravital microscopy reveals HER2-dependent and -independent effects of myeloid cells on the single-cell biodistribution of trastuzumab [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 376.
Abstract 3761: HSK26784: An oral PROTAC-BTK degrader for multiple B lymphocyte derived malignancies
Authors: Qian Meilin , Ye Fei , Zhang Chen , Wang Jianming , Zhang Yuanli , Cui Yangyang , Li Linli , Gou Xiaoli , Ni Jia 
Published: September 2020

Abstract
PROteolysis TArgeting Chimeras (PROTACs) have emerged as a new and promising modality utilizing an event-driven MOA, whereby protein levels could be modulated by PROTAC-induced degradation by hijacking the ubiquitin-proteasome system. Starting from 2001, many target proteins have been studied, such as BET and kinase families as well as nuclear receptors. Bruton's tyrosine kinase (BTK) is an essential component of BCR pathway, which plays an important role in development, activation, proliferation, and survival of B lymphocytes. Ibrutinib is an irreversible inhibitor of BTK, and shows promising activity against multiple B lymphocyte derived malignancies. However, there remains a big concern about the development of Ibrutinib induced resistance by BTK C481S mutant. In our study, HSK26784 was found as a novel orally available BTK-PROTAC molecule, which could selectively degrade the BTK proteins. HSK26784 selectively degraded BTK proteins in Mino cell lines, with a half degradation concentration (DC50) of 22.9 nM. HSK26784 also effectively induced BTK degradation in mouse spleen, with a DC50 of 3.8 mg/kg. In addition, HSK26784 significantly inhibit the expression of CD69 on human PBMC cells after ex vivo BCR stimulation. HSK26784 could inhibit cell proliferation in many malignant B cell lines. In Ibrutinib sensitive cell lines, the inhibitory activity of HSK26784 is similar to that of Ibrutinib. While in the Ibrutinib non-sensitive cell lines, HSK26784 was more potent than Ibrutinib. Importantly, in the xenograft models, HSK26784 could also inhibit the tumor growth in a dose dependent manner. And HSK26784 achieved much better therapeutic effects than Ibrutinib. HSK26784 could inhibit the kinase activity of BTK wt and BTK C481S mutant, with IC50 of 11 nM and 14 nM respectively. It indicated that the efficacy of HSK26784 in BTK wt and BTK C481S mutant was comparable. HSK26784 showed good orally bioavailability in different species. The oral bio availabilities in mice, rats and dogs were 15.4 %, 11.8 % and 12.8 % respectively. In conclusion, HSK26784 is a promising BTK PROTAC degrader with better efficacy than Ibrutinib in multiple B lymphocytes.
Citation Format: Meilin Qian, Fei Ye, Chen Zhang, Jianming Wang, Yuanli Zhang, Yangyang Cui, Linli Li, Xiaoli Gou, Jia Ni. HSK26784: An oral PROTAC-BTK degrader for multiple B lymphocyte derived malignancies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3761.
Abstract 3772: Comprehensive characterization of the phosphoproteome of gastric cancer from endoscopic biopsy specimens
Authors: ADACHI Jun , Abe Yuichi , Hirano Hidekazu , Shoji Hirokazu , Gunji Daigo , Honda Kazufumi , Boku Narikazu , Tomonaga Takeshi 
Published: September 2020

Abstract
Cancer phosphoproteomics can provide insights regarding kinases that can be targeted for therapeutic applications. Monitoring the phosphoproteomics in cancer is expected to play a key role in optimizing treatments with kinase inhibitors. Clinical phosphoproteomics in surgical tissues and patient-derived models has been studied intensively. However, the reported data may not accurately reflect the phosphosignaling status in patients due to the effect of ischemia occurring during surgery or changes in the characteristics of cancer cells when establishing the models. In contrast, endoscopic biopsies have an advantage for clinical phosphoproteomics because they can be rapidly cryo-preserved. We aimed to develop a highly sensitive method for phosphoproteomics in endoscopic biopsies of gastric cancer.Three tumor/normal biopsies from gastric cancer tissue were obtained by endoscopy, and subjected to our optimized phosphoproteomics. Phosphopeptides were enriched with an immobilized metal affinity chromatography, and labeled with Tandem Mass Tag reagent. Quantified phosphosites were compared between the pairs of tumor/normal biopsies within same patient. Cancer-specific activated pathways and kinases were identified by pathway enrichment analysis and kinase-substrate enrichment analysis.Our protocol enabled the identification of more than 10,000 class-1 phosphosites from endoscopic biopsies. A comparison between samples from cancer tissue and normal mucosa demonstrated differences in the phosphosignaling, including biomarkers of response to DNA damage. Finally, cancer-specific activation of DNA damage response signaling was validated by additional phosphoproteomics of other patients and western blotting of gastric cancer/normal cells. In summary, our pioneering approach will facilitate more accurate clinical phosphoproteomics in endoscopic biopsies, which can be applied to monitor the activities of therapeutic kinases and, ultimately, can be a useful tool to precision medicine.
Citation Format: Jun ADACHI, Yuichi Abe, Hidekazu Hirano, Hirokazu Shoji, Daigo Gunji, Kazufumi Honda, Narikazu Boku, Takeshi Tomonaga. Comprehensive characterization of the phosphoproteome of gastric cancer from endoscopic biopsy specimens [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3772.