bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Amplification of Oncolytic Vaccinia Virus Widespread Tumor Cell Killing by Sunitinib through Multiple Mechanisms
Authors: Kim Minah , Nitschké Maximilian , Sennino Barbara , Murer Patrizia , Schriver Brian J. , Bell Alexander , Subramanian Aishwarya , McDonald Corry E. , Wang Jiahu , Cha Howard , Bourgeois-Daigneault Marie-Claude , Kirn David H. , Bell John C. , De Silva Naomi , Breitbach Caroline J. , McDonald Donald M.
Published: March 2023

Abstract
Oncolytic viruses pose many questions in their use in cancer therapy. In this study, we assessed the potential of mpJX-594 (mouse-prototype JX-594), a replication-competent vaccinia virus administered by intravenous injection, to target the tumor vasculature, produce immune activation and tumor cell killing more widespread than the infection, and suppress invasion and metastasis. These actions were examined in RIP-Tag2 transgenic mice with pancreatic neuroendocrine tumors that developed spontaneously and progressed as in humans. mpJX-594 initially infected tumor vascular endothelial cells, leading to vascular pruning and prolonged leakage in tumors but not in normal organs; parallel effects were observed in U87 gliomas. Viral infection spread to tumor cells, where tumor cell killing was much more widespread than the infection. Widespread tumor cell killing at 5 days was prevented by depletion of CD8+ T lymphocytes and did not require GM-CSF, as mpJX-594 variants that expressed human, mouse, or no GM-CSF produced equivalent amounts of killing. The antivascular, antitumor, and antimetastatic effects of mpJX-594 were amplified by concurrent or sequential administration of sunitinib, a multitargeted receptor tyrosine kinase inhibitor. These effects were not mimicked by selective inhibition of VEGFR2 despite equivalent vascular pruning, but were accompanied by suppression of regulatory T cells and greater influx of activated CD8+ T cells. Together, our results showed that mpJX-594 targets tumor blood vessels, spreads secondarily to tumor cells, and produces widespread CD8+ T-cell–dependent tumor cell killing in primary tumors and metastases, and that these effects can be amplified by coadministration of sunitinib.

Significance: These findings reveal multiple unrecognized features of the antitumor properties of oncolytic vaccinia viruses, all of which can be amplified by the multitargeted kinase inhibitor sunitinib. Cancer Res; 78(4); 922–37. ©2017 AACR.
Data from miR-139-5p Modulates Radiotherapy Resistance in Breast Cancer by Repressing Multiple Gene Networks of DNA Repair and ROS Defense
Authors: Pajic Marina , Froio Danielle , Daly Sheridan , Doculara Louise , Millar Ewan , Graham Peter H. , Drury Alison , Steinmann Angela , de Bock Charles E. , Boulghourjian Alice , Zaratzian Anaiis , Carroll Susan , Toohey Joanne , O’Toole Sandra A. , Harris Adrian L , Buffa Francesca M. , Gee Harriet E. , Hollway Georgina E. , Molloy Timothy J.
Published: March 2023

Abstract
Radiotherapy is essential to the treatment of most solid tumors and acquired or innate resistance to this therapeutic modality is a major clinical problem. Here we show that miR-139-5p is a potent modulator of radiotherapy response in breast cancer via its regulation of genes involved in multiple DNA repair and reactive oxygen species defense pathways. Treatment of breast cancer cells with a miR-139-5p mimic strongly synergized with radiation both in vitro and in vivo, resulting in significantly increased oxidative stress, accumulation of unrepaired DNA damage, and induction of apoptosis. Several miR-139-5p target genes were also strongly predictive of outcome in radiotherapy-treated patients across multiple independent breast cancer cohorts. These prognostically relevant miR-139-5p target genes were used as companion biomarkers to identify radioresistant breast cancer xenografts highly amenable to sensitization by cotreatment with a miR-139-5p mimetic.

Significance: The microRNA described in this study offers a potentially useful predictive biomarker of radiosensitivity in solid tumors and a generally applicable druggable target for tumor radiosensitization. Cancer Res; 78(2); 501–15. ©2017 AACR.
Data from Fc-Mediated Anomalous Biodistribution of Therapeutic Antibodies in Immunodeficient Mouse Models
Authors: Sharma Sai Kiran , Chow Andrew , Monette Sebastien , Vivier Delphine , Pourat Jacob , Edwards Kimberly J. , Dilling Thomas R. , Abdel-Atti Dalya , Zeglis Brian M. , Poirier John T. , Lewis Jason S.
Published: March 2023

Abstract
A critical benchmark in the development of antibody-based therapeutics is demonstration of efficacy in preclinical mouse models of human disease, many of which rely on immunodeficient mice. However, relatively little is known about how the biology of various immunodeficient strains impacts the in vivo fate of these drugs. Here we used immunoPET radiotracers prepared from humanized, chimeric, and murine mAbs against four therapeutic oncologic targets to interrogate their biodistribution in four different strains of immunodeficient mice bearing lung, prostate, and ovarian cancer xenografts. The immunodeficiency status of the mouse host as well as both the biological origin and glycosylation of the antibody contributed significantly to the anomalous biodistribution of therapeutic monoclonal antibodies in an Fc receptor-dependent manner. These findings may have important implications for the preclinical evaluation of Fc-containing therapeutics and highlight a clear need for biodistribution studies in the early stages of antibody drug development.

Significance: Fc/FcγR-mediated immunobiology of the experimental host is a key determinant to preclinical in vivo tumor targeting and efficacy of therapeutic antibodies. Cancer Res; 78(7); 1820–32. ©2018 AACR.
Data from Silencing the Snail-Dependent RNA Splice Regulator ESRP1 Drives Malignant Transformation of Human Pulmonary Epithelial Cells
Authors: Walser Tonya C. , Jing Zhe , Tran Linh M. , Lin Ying Q. , Yakobian Natalie , Wang Gerald , Krysan Kostyantyn , Zhu Li X. , Sharma Sherven , Lee Mi-Heon , Belperio John A. , Ooi Aik T. , Gomperts Brigitte N. , Shay Jerry W. , Larsen Jill E. , Minna John D. , Hong Long-sheng , Fishbein Michael C. , Dubinett Steven M.
Published: March 2023

Abstract
Epithelial-to-mesenchymal transition (EMT) is organized in cancer cells by a set of key transcription factors, but the significance of this process is still debated, including in non–small cell lung cancer (NSCLC). Here, we report increased expression of the EMT-inducing transcription factor Snail in premalignant pulmonary lesions, relative to histologically normal pulmonary epithelium. In immortalized human pulmonary epithelial cells and isogenic derivatives, we documented Snail-dependent anchorage-independent growth in vitro and primary tumor growth and metastatic behavior in vivo. Snail-mediated transformation relied upon silencing of the tumor-suppressive RNA splicing regulatory protein ESRP1. In clinical specimens of NSCLC, ESRP1 loss was documented in Snail-expressing premalignant pulmonary lesions. Mechanistic investigations showed that Snail drives malignant progression in an ALDH+CD44+CD24− pulmonary stem cell subset in which ESRP1 and stemness-repressing microRNAs are inhibited. Collectively, our results show how ESRP1 loss is a critical event in lung carcinogenesis, and they identify new candidate directions for targeted therapy of NSCLC.

Significance: This study defines a Snail-ESRP1 cancer axis that is crucial for human lung carcinogenesis, with implications for new intervention strategies and translational opportunities. Cancer Res; 78(8); 1986–99. ©2018 AACR.
Data from CD163 Is Required for Protumoral Activation of Macrophages in Human and Murine Sarcoma
Authors: Shiraishi Daisuke , Fujiwara Yukio , Horlad Hasita , Saito Yoichi , Iriki Toyohisa , Tsuboki Junko , Cheng Pan , Nakagata Naomi , Mizuta Hiroshi , Bekki Hirofumi , Nakashima Yasuharu , Oda Yoshinao , Takeya Motohiro , Komohara Yoshihiro
Published: March 2023

Abstract
Recent findings have shown the significance of CD163-positive macrophages in tumor progression, yet there have been few studies on the function of CD163 in macrophages. Here, we uncover the role of CD163 in macrophage activation using CD163-deficient mice and human samples. We detected CD163 in 62 undifferentiated pleomorphic sarcoma samples, in which a high percentage of CD163-positive macrophages was associated with decreased overall survival and higher histologic grade. We observed macrophage-induced tumor cell proliferation in cocultures of human monocyte-derived macrophages and leiomyosarcoma (TYLMS-1) and myxofibrosarcoma (NMFH-1) cell lines, which was abrogated by silencing of CD163. Tumor development of sarcoma (MCA205 and LM8) cells in CD163-deficient mice was significantly abrogated in comparison with wild-type (WT) mice. Coculture with WT peritoneal macrophages significantly increased proliferation of MCA205 cells but decreased in the presence of CD163-deficient macrophages. Production of IL6 and CXCL2 in CD163-deficient macrophages was suppressed in comparison with WT macrophages, and overexpression of CD163 in CD163-deficient macrophages induced production of IL6 and CXCL2. Silencing of IL6 but not CXCL2 abrogated macrophage-induced proliferation of MCA205 cells. Taken together, our results show that CD163 is involved in protumoral activation of macrophages and subsequent development and progression of tumors in mice and humans.

Significance: Macrophage CD163-mediated induction of IL6 promotes tumor development and progression in murine and human malignant tumors. Cancer Res; 78(12); 3255–66. ©2018 AACR.
Data from LSR Antibody Therapy Inhibits Ovarian Epithelial Tumor Growth by Inhibiting Lipid Uptake
Authors: Hiramatsu Kosuke , Serada Satoshi , Enomoto Takayuki , Takahashi Yusuke , Nakagawa Satoshi , Nojima Satoshi , Morimoto Akiko , Matsuzaki Shinya , Yokoyama Takuhei , Takahashi Tsuyoshi , Fujimoto Minoru , Takemori Hiroshi , Ueda Yutaka , Yoshino Kiyoshi , Morii Eiichi , Kimura Tadashi , Naka Tetsuji
Published: March 2023

Abstract
Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy, but it still lacks effective treatment options. In this study, we utilized proteomic technology to identify lipolysis-stimulated lipoprotein receptor (LSR) as a new tumor antigen of EOC. Immunohistochemical analysis of EOC tissues in conjunction with survival analysis of EOC patients showed that high expression of LSR is associated with poor prognosis. High LSR expression also occurred in tumor metastases including to the lymph node and omentum. To evaluate the possible benefits of blocking this antigen in EOC, we raised a new monoclonal antibody (mAb) to human LSR (hLSR). In mouse xenograft models of hLSR+ EOC (cell lines or patient-derived tumors), we found that administration of anti-hLSR mAb inhibited tumor growth in a manner independent of both antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity. Mechanistic investigations showed that hLSR expression increased incorporation of very-low-density lipoprotein (VLDL) into EOC cells and that anti-hLSR mAb inhibited lipid uptake in vitro and in vivo. Moreover, VLDL promoted cell proliferation in hLSR-positive EOC cells in vitro, and this effect was inhibited by anti-hLSR mAb. While the anti-hLSR mAb studied cross reacted with the mouse antigen, we observed no adverse effects on normal organs and lipid metabolism in murine hosts. Our findings suggest that hLSR plays a key functional role in EOC development and that this antigen can be therapeutically targeted by specific mAb to improve EOC treatment.

Significance: These findings offer preclinical evidence of the therapeutic efficacy of a novel targeted antibody therapy against deadly epithelial ovarian cancers. Cancer Res; 78(2); 516–27. ©2017 AACR.
Data from CCR5+ Myeloid-Derived Suppressor Cells Are Enriched and Activated in Melanoma Lesions
Authors: Blattner Carolin , Fleming Viktor , Weber Rebekka , Himmelhan Bianca , Altevogt Peter , Gebhardt Christoffer , Schulze Torsten J. , Razon Hila , Hawila Elias , Wildbaum Gizi , Utikal Jochen , Karin Nathan , Umansky Viktor
Published: March 2023

Abstract
Accumulation of myeloid-derived suppressor cells (MDSC) in melanoma microenvironment is supported by chemokine receptor/chemokine signaling. Although different chemokines were suggested to be involved in this process, the role of CCR5 and its ligands is not established. Using a Ret transgenic mouse melanoma model, we found an accumulation of CCR5+ MDSCs in melanoma lesions associated with both increased concentrations of CCR5 ligands and tumor progression. Tumor-infiltrating CCR5+ MDSCs displayed higher immunosuppressive activity than their CCR5− counterparts. Upregulation of CCR5 expression on CD11b+Gr1+ myeloid cells was induced in vitro by CCR5 ligands and other inflammatory factors. In melanoma patients, CCR5+ MDSCs were enriched at the tumor site and correlated with enhanced production of CCR5 ligands. Moreover, they exhibited a stronger immunosuppressive pattern compared with CCR5− MDSCs. Blocking CCR5/CCR5 ligand interactions increased survival of tumor-bearing mice and was associated with reduced migration and immunosuppressive potential of MDSCs in tumor lesions. Our findings define a critical role for CCR5 in recruitment and activation of MDSCs, suggesting a novel strategy for melanoma treatment.

Significance: These findings validate the importance of the CCR5/CCR5 ligand axis not only for MDSC recruitment but also for further activation of their immunosuppressive functions in the tumor microenvironment, with potentially broad therapeutic implications, given existing clinically available inhibitors of this axis. Cancer Res; 78(1); 157–67. ©2017 AACR.
Data from Improved Evaluation of Antivascular Cancer Therapy Using Constrained Tracer-Kinetic Modeling for Multiagent Dynamic Contrast-Enhanced MRI
Authors: Hectors Stefanie J. , Jacobs Igor , Lok Jasper , Peters Johannes , Bussink Johan , Hoeben Freek J. , Keizer Henk M. , Janssen Henk M. , Nicolay Klaas , Schabel Matthias C. , Strijkers Gustav J.
Published: March 2023

Abstract
Dynamic contrast–enhanced MRI (DCE-MRI) is a promising technique for assessing the response of tumor vasculature to antivascular therapies. Multiagent DCE-MRI employs a combination of low and high molecular weight contrast agents, which potentially improves the accuracy of estimation of tumor hemodynamic and vascular permeability parameters. In this study, we used multiagent DCE-MRI to assess changes in tumor hemodynamics and vascular permeability after vascular-disrupting therapy. Multiagent DCE-MRI (sequential injection of G5 dendrimer, G2 dendrimer, and Gd-DOTA) was performed in tumor-bearing mice before, 2 and 24 hours after treatment with vascular disrupting agent DMXAA or placebo. Constrained DCE-MRI gamma capillary transit time modeling was used to estimate flow F, blood volume fraction vb, mean capillary transit time tc, bolus arrival time td, extracellular extravascular fraction ve, vascular heterogeneity index α−1 (all identical between agents) and extraction fraction E (reflective of permeability), and transfer constant Ktrans (both agent-specific) in perfused pixels. F, vb, and α−1 decreased at both time points after DMXAA, whereas tc increased. E (G2 and G5) showed an initial increase, after which, both parameters restored. Ktrans (G2 and Gd-DOTA) decreased at both time points after treatment. In the control, placebo-treated animals, only F, tc, and Ktrans Gd-DOTA showed significant changes. Histologic perfused tumor fraction was significantly lower in DMXAA-treated versus control animals. Our results show how multiagent tracer-kinetic modeling can accurately determine the effects of vascular-disrupting therapy by separating simultaneous changes in tumor hemodynamics and vascular permeability.

Significance: These findings describe a new approach to measure separately the effects of antivascular therapy on tumor hemodynamics and vascular permeability, which could help more rapidly and accurately assess the efficacy of experimental therapy of this class. Cancer Res; 78(6); 1561–70. ©2018 AACR.
Data from Remodeling the Tumor Microenvironment Sensitizes Breast Tumors to Anti-Programmed Death-Ligand 1 Immunotherapy
Authors: Clift Renee , Souratha Jennifer , Garrovillo Sheryl A. , Zimmerman Susan , Blouw Barbara
Published: March 2023

Abstract
Immunotherapies targeting immune checkpoint inhibitors have changed the landscape of cancer treatment, however, many patients are resistant or refractory to immunotherapy. The sensitivity of tumor cells to immunotherapy may be influenced by hyaluronan (HA) accumulation in the tumor microenvironment (TME). Enzymatic degradation of HA by pegvorhyaluronidase alfa (PEGPH20; PVHA) remodels the TME. This leads to reduced tumor interstitial pressure and decompressed tumor blood vessels, which are both associated with increased exposure of tumor cells to chemotherapy drugs. Here, we demonstrate PVHA increased the uptake of anti-programmed death-ligand 1 (PD-L1) antibody in HA-accumulating animal models of breast cancer. The increased levels of anti-PD-L1 antibody were associated with increased accumulation of T cells and natural killer cells and decreased myeloid-derived suppressor cells. PD-L1 blockade significantly inhibited tumor growth when combined with PVHA, but not alone. Our results suggest that PVHA can sensitize HA-accumulating tumors to anti-PD-L1 immunotherapy.
Significance:
These findings show removal of hyaluronan in the tumor microenvironment improves immune cells and checkpoint inhibitors access to tumors.
Data from Gaussian Mixture Models for Probabilistic Classification of Breast Cancer
Authors: Prabakaran Indira , Wu Zhengdong , Lee Changgun , Tong Brian , Steeman Samantha , Koo Gabriel , Zhang Paul J. , Guvakova Marina A.
Published: March 2023

Abstract
In the era of omics-driven research, it remains a common dilemma to stratify individual patients based on the molecular characteristics of their tumors. To improve molecular stratification of patients with breast cancer, we developed the Gaussian mixture model (GMM)–based classifier. This probabilistic classifier was built on mRNA expression data from more than 300 clinical samples of breast cancer and healthy tissue and was validated on datasets of ESR1, PGR, and ERBB2, which encode standard clinical markers and therapeutic targets. To demonstrate how a GMM approach could be exploited for multiclass classification using data from a candidate marker, we analyzed the insulin-like growth factor I receptor (IGF1R), a promising target, but a marker of uncertain importance in breast cancer. The GMM defined subclasses with downregulated (40%), unchanged (39%), upregulated (19%), and overexpressed (2%) IGF1R levels; inter- and intrapatient analyses of IGF1R transcript and protein levels supported these predictions. Overexpressed IGF1R was observed in a small percentage of tumors. Samples with unchanged and upregulated IGF1R were differentiated tumors, and downregulation of IGF1R correlated with poorly differentiated, high-risk hormone receptor–negative and HER2-positive tumors. A similar correlation was found in the independent cohort of carcinoma in situ, suggesting that loss or low expression of IGF1R is a marker of aggressiveness in subsets of preinvasive and invasive breast cancer. These results demonstrate the importance of probabilistic modeling that delves deeper into molecular data and aims to improve diagnostic classification, prognostic assessment, and treatment selection.
Significance:
A GMM classifier demonstrates potential use for clinical validation of markers and determination of target populations, particularly when availability of specimens for marker development is low.
Data from Stathmin Is Required for Normal Mouse Mammary Gland Development and Δ16HER2-Driven Tumorigenesis
Authors: Segatto Ilenia , Zompit Mara De Marco , Citron Francesca , D'Andrea Sara , Vinciguerra Gian Luca Rampioni , Perin Tiziana , Berton Stefania , Mungo Giorgia , Schiappacassi Monica , Marchini Cristina , Amici Augusto , Vecchione Andrea , Baldassarre Gustavo , Belletti Barbara
Published: March 2023

Abstract
Postnatal development of the mammary gland relies on the maintenance of oriented cell division and apicobasal polarity, both of which are often deregulated in cancer. The microtubule (MT) network contributes to control these processes; however, very little is known about the impact of altered MT dynamics in the development of a complex organ and on the role played by MT-interacting proteins such as stathmin. In this study, we report that female stathmin knock-out (STM KO) mice are unable to nurse their litters due to frank impairment of mammary gland development. In mouse mammary epithelial cells, loss of stathmin compromised the trafficking of polarized proteins and the achievement of proper apicobasal polarity. In particular, prolactin receptor internalization and localization was altered in STM KO mammary epithelial cells, leading to decreased protein stability and downmodulation of the Prl/PrlR/STAT5 signaling pathway. Absence of stathmin induced alterations in mitotic spindle orientation, accumulation of mitotic defects, and apoptosis, overall contributing to tissue disorganization and further decreasing the expansion of the mammary epithelial compartment. Loss of stathmin in MMTV-Δ16HER2 transgenic mice decreased the incidence and increased the latency of these very aggressive mammary carcinomas. Collectively, these data identify the essential mammary protein stathmin as protumorigenic and suggest it may serve as a potential therapeutic target in breast cancer.
Significance:
Stathmin expression is critical to maintain oriented cell division and apicobasal polarity in normal mammary glands and to establish a protumorigenic program that eventually sustains HER2-positive breast cancer formation in mice.
Data from Asporin Restricts Mesenchymal Stromal Cell Differentiation, Alters the Tumor Microenvironment, and Drives Metastatic Progression
Authors: Hughes Robert M. , Simons Brian W. , Khan Hamda , Miller Rebecca , Kugler Valentina , Torquato Samantha , Theodros Debebe , Haffner Michael C. , Lotan Tamara , Huang Jessie , Davicioni Elai , An Steven S. , Riddle Ryan C. , Thorek Daniel L.J. , Garraway Isla P. , Fertig Elana J. , Isaacs John T. , Brennen W. Nathaniel , Park Ben H. , Hurley Paula J.
Published: March 2023

Abstract
Tumor progression to metastasis is not cancer cell autonomous, but rather involves the interplay of multiple cell types within the tumor microenvironment. Here we identify asporin (ASPN) as a novel, secreted mesenchymal stromal cell (MSC) factor in the tumor microenvironment that regulates metastatic development. MSCs expressed high levels of ASPN, which decreased following lineage differentiation. ASPN loss impaired MSC self-renewal and promoted terminal cell differentiation. Mechanistically, secreted ASPN bound to BMP-4 and restricted BMP-4–induced MSC differentiation prior to lineage commitment. ASPN expression was distinctly conserved between MSC and cancer-associated fibroblasts (CAF). ASPN expression in the tumor microenvironment broadly impacted multiple cell types. Prostate tumor allografts in ASPN-null mice had a reduced number of tumor-associated MSCs, fewer cancer stem cells, decreased tumor vasculature, and an increased percentage of infiltrating CD8+ T cells. ASPN-null mice also demonstrated a significant reduction in lung metastases compared with wild-type mice. These data establish a role for ASPN as a critical MSC factor that extensively affects the tumor microenvironment and induces metastatic progression.
Significance:
These findings show that asporin regulates key properties of mesenchymal stromal cells, including self-renewal and multipotency, and asporin expression by reactive stromal cells alters the tumor microenvironment and promotes metastatic progression.
Data from Nanoparticle Encapsulation of Synergistic Immune Agonists Enables Systemic Codelivery to Tumor Sites and IFNβ-Driven Antitumor Immunity
Authors: Atukorale Prabhani U. , Raghunathan Shruti P. , Raguveer Vanitha , Moon Taylor J. , Zheng Carolyn , Bielecki Peter A. , Wiese Michelle L. , Goldberg Amy L. , Covarrubias Gil , Hoimes Christopher J. , Karathanasis Efstathios
Published: March 2023

Abstract
Effective cancer immunotherapy depends on the robust activation of tumor-specific antigen-presenting cells (APC). Immune agonists encapsulated within nanoparticles (NP) can be delivered to tumor sites to generate powerful antitumor immune responses with minimal off-target dissemination. Systemic delivery enables widespread access to the microvasculature and draining to the APC-rich perivasculature. We developed an immuno-nanoparticle (immuno-NP) coloaded with cyclic diguanylate monophosphate, an agonist of the stimulator of interferon genes pathway, and monophosphoryl lipid A, and a Toll-like receptor 4 agonist, which synergize to produce high levels of type I IFNβ. Using a murine model of metastatic triple-negative breast cancer, systemic delivery of these immuno-NPs resulted in significant therapeutic outcomes due to extensive upregulation of APCs and natural killer cells in the blood and tumor compared with control treatments. These results indicate that NPs can facilitate systemic delivery of multiple immune-potentiating cargoes for effective APC-driven local and systemic antitumor immunity.
Significance:
Systemic administration of an immuno-nanoparticle in a murine breast tumor model drives a robust tumor site–specific APC response by delivering two synergistic immune-potentiating molecules, highlighting the potential of nanoparticles for immunotherapy.
Data from A Microfluidic Chip Enables Isolation of Exosomes and Establishment of Their Protein Profiles and Associated Signaling Pathways in Ovarian Cancer
Authors: Dorayappan Kalpana Deepa Priya , Gardner Miranda L. , Hisey Colin L. , Zingarelli Roman A. , Smith Brentley Q. , Lightfoot Michelle D.S. , Gogna Rajan , Flannery Meghan M. , Hays John , Hansford Derek J. , Freitas Michael A. , Yu Lianbo , Cohn David E. , Selvendiran Karuppaiyah
Published: March 2023

Abstract
Because of limits on specificity and purity to allow for in-depth protein profiling, a standardized method for exosome isolation has yet to be established. In this study, we describe a novel, in-house microfluidic-based device to isolate exosomes from culture media and patient samples. This technology overcomes contamination issues because sample separation is based on the expression of highly specific surface markers CD63 and EpCAM. Mass spectrometry revealed over 25 exosome proteins that are differentially expressed in high-grade serous ovarian cancer (HGSOC) cell lines compared with normal cells—ovarian surface epithelia cells and fallopian tube secretory epithelial cells (FTSEC). Top exosome proteins were identified on the basis of their fold change and statistical significance between groups. Ingenuity pathway analysis identified STAT3 and HGF as top regulator proteins. We further validated exosome proteins of interest (pSTAT3, HGF, and IL6) in HGSOC samples of origin-based cell lines (OVCAR-8, FTSEC) and in early-stage HGSOC patient serum exosome samples using LC/MS-MS and proximity extension assay. Our microfluidic device will allow us to make new discoveries for exosome-based biomarkers for the early detection of HGSOC and will contribute to the development of new targeted therapies based on signaling pathways that are unique to HGSOC, both of which could improve the outcome for women with HGSOC.
Significance:
A unique platform utilizing a microfluidic device enables the discovery of new exosome-based biomarkers in ovarian cancer.
Data from CXCR7 Reactivates ERK Signaling to Promote Resistance to EGFR Kinase Inhibitors in NSCLC
Authors: Becker Jeffrey H. , Gao Yandi , Soucheray Margaret , Pulido Ines , Kikuchi Eiki , Rodríguez María L. , Gandhi Rutu , Lafuente-Sanchis Aranzazu , Aupí Miguel , Alcácer Fernández-Coronado Javier , Martín-Martorell Paloma , Cremades Antonio , Galbis-Caravajal José M. , Alcácer Javier , Christensen Camilla L. , Simms Patricia , Hess Ashley , Asahina Hajime , Kahle Michael P. , Al-Shahrour Fatima , Borgia Jeffrey A. , Lahoz Agustín , Insa Amelia , Juan Oscar , Jänne Pasi A. , Wong Kwok-Kin , Carretero Julian , Shimamura Takeshi
Published: March 2023

Abstract
Although EGFR mutant–selective tyrosine kinase inhibitors (TKI) are clinically effective, acquired resistance can occur by reactivating ERK. We show using in vitro models of acquired EGFR TKI resistance with a mesenchymal phenotype that CXCR7, an atypical G protein-coupled receptor, activates the MAPK–ERK pathway via β-arrestin. Depletion of CXCR7 inhibited the MAPK pathway, significantly attenuated EGFR TKI resistance, and resulted in mesenchymal-to-epithelial transition. CXCR7 overexpression was essential in reactivation of ERK1/2 for the generation of EGFR TKI–resistant persister cells. Many patients with non–small cell lung cancer (NSCLC) harboring an EGFR kinase domain mutation, who progressed on EGFR inhibitors, demonstrated increased CXCR7 expression. These data suggest that CXCR7 inhibition could considerably delay and prevent the emergence of acquired EGFR TKI resistance in EGFR-mutant NSCLC.
Significance:
Increased expression of the chemokine receptor CXCR7 constitutes a mechanism of resistance to EGFR TKI in patients with non–small cell lung cancer through reactivation of ERK signaling.
Data from Multiparametric MRI and Coregistered Histology Identify Tumor Habitats in Breast Cancer Mouse Models
Authors: Jardim-Perassi Bruna V. , Huang Suning , Dominguez-Viqueira William , Poleszczuk Jan , Budzevich Mikalai M. , Abdalah Mahmoud A. , Pillai Smitha R. , Ruiz Epifanio , Bui Marilyn M. , Zuccari Debora A.P.C. , Gillies Robert J. , Martinez Gary V.
Published: March 2023

Abstract
It is well-recognized that solid tumors are genomically, anatomically, and physiologically heterogeneous. In general, more heterogeneous tumors have poorer outcomes, likely due to the increased probability of harboring therapy-resistant cells and regions. It is hypothesized that the genomic and physiologic heterogeneity are related, because physiologically distinct regions will exert variable selection pressures leading to the outgrowth of clones with variable genomic/proteomic profiles. To investigate this, methods must be in place to interrogate and define, at the microscopic scale, the cytotypes that exist within physiologically distinct subregions (“habitats”) that are present at mesoscopic scales. MRI provides a noninvasive approach to interrogate physiologically distinct local environments, due to the biophysical principles that govern MRI signal generation. Here, we interrogate different physiologic parameters, such as perfusion, cell density, and edema, using multiparametric MRI (mpMRI). Signals from six different acquisition schema were combined voxel-by-voxel into four clusters identified using a Gaussian mixture model. These were compared with histologic and IHC characterizations of sections that were coregistered using MRI-guided 3D printed tumor molds. Specifically, we identified a specific set of MRI parameters to classify viable-normoxic, viable-hypoxic, nonviable-hypoxic, and nonviable-normoxic tissue types within orthotopic 4T1 and MDA-MB-231 breast tumors. This is the first coregistered study to show that mpMRI can be used to define physiologically distinct tumor habitats within breast tumor models.
Significance:
This study demonstrates that noninvasive imaging metrics can be used to distinguish subregions within heterogeneous tumors with histopathologic correlation.
Data from Deletion of Calcineurin Promotes a Protumorigenic Fibroblast Phenotype
Authors: Lieberman Allyson , Barrett Richard , Kim Jaewon , Zhang Kathy L. , Avery Diana , Monslow James , Kim Hyunsoo , Kim Bang-Jin , Puré Ellen , Ryeom Sandra
Published: March 2023

Abstract
Fibroblast activation is a crucial step in tumor growth and metastatic progression. Activated fibroblasts remodel the extracellular matrix (ECM) in primary tumor and metastatic microenvironments, exerting both pro- and antitumorigenic effects. However, the intrinsic mechanisms that regulate the activation of fibroblasts are not well-defined. The signaling axis comprising the calcium-activated Ser/Thr phosphatase calcineurin (CN), and its downstream target nuclear factor of activated T cells, has been implicated in endothelial (EC) and immune cell activation, but its role in fibroblasts is not known. Here, we demonstrate that deletion of CN in fibroblasts in vitro altered fibroblast morphology and function consistent with an activated phenotype relative to wild-type fibroblasts. CN-null fibroblasts had a greater migratory capacity, increased collagen secretion and remodeling, and promoted more robust EC activation in vitro. ECM generated by CN-null fibroblasts contained more collagen with greater alignment of fibrillar collagen compared with wild-type fibroblast-derived matrix. These differences in matrix composition and organization imposed distinct changes in morphology and cytoskeletal architecture of both fibroblasts and tumor cells. Consistent with this in vitro phenotype, mice with stromal CN deletion had a greater incidence and larger lung metastases. Our data suggest that CN signaling contributes to the maintenance of fibroblast homeostasis and that loss of CN is sufficient to promote fibroblast activation.
Significance:
Calcineurin signaling is a key pathway underlying fibroblast homeostasis that could be targeted to potentially prevent fibroblast activation in distant metastatic sites.
Data from A Multigene Assay Determines Risk of Recurrence in Patients with Triple-Negative Breast Cancer
Authors: Stewart Rachel L. , Updike Katherine L. , Factor Rachel E. , Henry N. Lynn , Boucher Kenneth M. , Bernard Philip S. , Varley Katherine E.
Published: March 2023

Abstract
Approximately 40% of patients with stage I–III triple-negative breast cancer (TNBC) recur after standard treatment, whereas the remaining 60% experience long-term disease-free survival (DFS). There are currently no clinical tests to assess the risk of recurrence in TNBC patients. We previously determined that TNBC patients with MHC class II (MHCII) pathway expression in their tumors experienced significantly longer DFS. To translate this discovery into a clinical test, we developed an MHCII Immune Activation assay, which measures expression of 36 genes using NanoString technology. Preanalytical testing confirmed that the assay is accurate and reproducible in formalin-fixed paraffin-embedded (FFPE) tumor specimens. The assay measurements were concordant with RNA-seq, MHCII protein expression, and tumor-infiltrating lymphocyte counts. In a training set of 44 primary TNBC tumors, the MHCII Immune Activation Score was significantly associated with longer DFS (HR = 0.17; P = 0.015). In an independent validation cohort of 56 primary FFPE TNBC tumors, the Immune Activation Score was significantly associated with longer DFS (HR = 0.19; P = 0.011) independent of clinical stage. An Immune Activation Score threshold for identifying patients with very low risk of relapse in the training set provided 100% specificity in the validation cohort. The assay format enables adoption as a standardized clinical prognostic test for identifying TNBC patients with a low risk of recurrence. Correlative data support future studies to determine if the assay can identify patients in whom chemotherapy can be safely deescalated and patients likely to respond to immunotherapy.
Significance:
The MHCII Immune Activation assay identifies TNBC patients with a low risk of recurrence, addressing a critical need for prognostic biomarker tests that enable precision medicine for TNBC patients.
Data from Deletion of the Miz-1 POZ Domain Increases Efficacy of Cytarabine Treatment in T- and B-ALL/Lymphoma Mouse Models
Authors: Ross Julie , Rashkovan Marissa , Fraszczak Jennifer , Joly-Beauparlant Charles , Vadnais Charles , Winkler René , Droit Arnaud , Kosan Christian , Möröy Tarik
Published: March 2023

Abstract
Acute lymphoblastic leukemia (ALL) is an aggressive blood cancer that mainly affects children. Relapse rates are high and toxic chemotherapies that block DNA replication and induce DNA damage lead to health problems later in life, underlining the need for improved therapies. MYC is a transcription factor that is hyperactive in a large proportion of cancers including leukemia but is difficult to target in therapy. We show that ablation of the function of the BTB/POZ domain factor Zbtb17 (Miz-1), an important cofactor of c-Myc, significantly delayed T- and B-ALL/lymphoma in mice and interfered with the oncogenic transcriptional activity of c-Myc. Leukemic cells that still emerged in this system activated DNA replication pathways that could be targeted by current chemotherapeutic drugs such as cytarabine. Acute ablation of the Miz-1 POZ domain enhanced the effect of cytarabine treatment. The combined treatment was effective in both Eμ-Myc and Notch ICN-driven leukemia models and prolonged survival of tumor-bearing animals by accelerating apoptosis of leukemic cells. These observations suggest that targeting MIZ-1 could render current ALL chemotherapies more effective, with a better outcome for patients.
Significance:
Ablation of the POZ domain of Miz-1 perturbs its interaction with c-MYC and delays the generation of T- and B-cell leukemias and lymphomas.
Data from The HPV16 Genome Is Stable in Women Who Progress to In Situ or Invasive Cervical Cancer: A Prospective Population-Based Study
Authors: Arroyo-Mühr Laila-Sara , Lagheden Camilla , Hultin Emilie , Eklund Carina , Adami Hans-Olov , Dillner Joakim , Sundström Karin
Published: March 2023

Abstract
The human papillomavirus (HPV) rate of evolution is essential for cancer-preventive strategies targeting HPV. We analyzed variability over time in a prospective, population-based nested case–control study of in situ (CIS) and invasive squamous cervical cancer (SCC). Among 757,690 women who participated in cervical screening in Sweden during 1969 to 2002, we identified 94 women who had HPV16 persistence in two serial cervical screening samples (median 24 months apart, range 0.5–178 months) and later were diagnosed with CIS (n = 59), SCC (n = 32), or remained healthy (n = 3). Whole-HPV16-genome sequencing and comparison of sequences in the serial samples revealed that all women had the same HPV16 lineage, particularly lineage A, in both serial smears. Fifty-six percent of women had an identical 7,906 base pair HPV16 sequence in both samples, and no woman had more than 15 nucleotide substitutions. The median substitution rate was 0 substitutions/site/year (95% confidence interval, 0–0.00008), with no variation between quartiles of follow-up. We concluded that in most women with HPV16 persistence preceding disease, the nucleotide substitution rate was not measurable within up to 15-years follow-up. This slow rate of evolution has important implications for both HPV-based screening and HPV vaccination.
Significance:
These findings show there is no genomic variation over time in HPV16 infections progressing to cervical cancer, which could influence risk stratification of women when screening for cervical cancer and inform HPV vaccination strategies.
Data from SIX2 Mediates Late-Stage Metastasis via Direct Regulation of SOX2 and Induction of a Cancer Stem Cell Program
Authors: Oliphant Michael U.J. , Vincent Melanie Y. , Galbraith Matthew D. , Pandey Ahwan , Zaberezhnyy Vadym , Rudra Pratyaydipta , Johnson Katherine R. , Costello James C. , Ghosh Debashis , DeGregori James , Espinosa Joaquin M. , Ford Heide L.
Published: March 2023

Abstract
The capacity for tumor cells to metastasize efficiently is directly linked to their ability to colonize secondary sites. Here we identify Six2, a developmental transcription factor, as a critical regulator of a breast cancer stem cell program that enables metastatic colonization. In several triple-negative breast cancer (TNBC) models, Six2 enhanced the expression of genes associated with embryonic stem cell programs. Six2 directly bound the Sox2 Srr2 enhancer, promoting Sox2 expression and downstream expression of Nanog, which are both key pluripotency factors. Regulation of Sox2 by Six2 enhanced cancer stem cell properties and increased metastatic colonization. Six2 and Sox2 expression correlated highly in breast cancers including TNBC, where a Six2 expression signature was predictive of metastatic burden and poor clinical outcome. Our findings demonstrate that a SIX2/SOX2 axis is required for efficient metastatic colonization, underscoring a key role for stemness factors in outgrowth at secondary sites.
Significance:
These findings provide novel mechanistic insight into stemness and the metastatic outgrowth of triple-negative breast cancer cells.
Data from Parallel Signaling through IRE1α and PERK Regulates Pancreatic Neuroendocrine Tumor Growth and Survival
Authors: Moore Paul C. , Qi Jenny Y. , Thamsen Maike , Ghosh Rajarshi , Peng Justin , Gliedt Micah J. , Meza-Acevedo Rosa , Warren Rachel E. , Hiniker Annie , Kim Grace E. , Maly Dustin J. , Backes Bradley J. , Papa Feroz R. , Oakes Scott A.
Published: March 2023

Abstract
Master regulators of the unfolded protein response (UPR), IRE1α and PERK, promote adaptation or apoptosis depending on the level of endoplasmic reticulum (ER) stress. Although the UPR is activated in many cancers, its effects on tumor growth remain unclear. Derived from endocrine cells, pancreatic neuroendocrine tumors (PanNET) universally hypersecrete one or more peptide hormones, likely sensitizing these cells to high ER protein-folding stress. To assess whether targeting the UPR is a viable therapeutic strategy, we analyzed human PanNET samples and found evidence of elevated ER stress and UPR activation. Genetic and pharmacologic modulation of IRE1α and PERK in cultured cells, xenograft, and spontaneous genetic (RIP-Tag2) mouse models of PanNETs revealed that UPR signaling was optimized for adaptation and that inhibiting either IRE1α or PERK led to hyperactivation and apoptotic signaling through the reciprocal arm, thereby halting tumor growth and survival. These results provide a strong rationale for therapeutically targeting the UPR in PanNETs and other cancers with elevated ER stress.
Significance:
The UPR is upregulated in pancreatic neuroendocrine tumors and its inhibition significantly reduces tumor growth in preclinical models, providing strong rationale for targeting the UPR in these cancers.
Data from Curatopes Melanoma: A Database of Predicted T-cell Epitopes from Overly Expressed Proteins in Metastatic Cutaneous Melanoma
Authors: Lischer Christopher , Eberhardt Martin , Jaitly Tanushree , Schinzel Cornelia , Schaft Niels , Dörrie Jan , Schuler Gerold , Vera Julio
Published: March 2023

Abstract
Therapeutic anticancer vaccination has been adapted as an immunotherapy in several solid tumors. However, the selection of promising candidates from the total quantity of possible epitopes poses a challenge to clinicians and bioinformaticians alike, and very few epitopes have been tested in experimental or clinical settings to validate their efficacy. Here, we present a comprehensive database of predicted nonmutated peptide epitopes derived from genes that are overly expressed in a group of 32 melanoma biopsies compared with healthy tissues and that were filtered against expression in a curated list of survival-critical tissues. We hypothesize that these "self-tolerant" epitopes have two desirable properties: they do not depend on mutations, being immediately applicable to a large patient collective, and they potentially cause fewer autoimmune reactions. To support epitope selection, we provide an aggregated score of expected therapeutic efficiency as a shortlist mechanism. The database has applications in facilitating epitope selection and trial design and is freely accessible at https://www.curatopes.com.
Significance:
A database is presented that predicts and scores antitumor T-cell epitopes, with a focus on tolerability and avoidance of severe autoimmunity, offering a supplementary epitope set for further investigation in immunotherapy.
Data from Comprehensive Benchmarking and Integration of Tumor Microenvironment Cell Estimation Methods
Authors: Jiménez-Sánchez Alejandro , Cast Oliver , Miller Martin L.
Published: March 2023

Abstract
Various computational approaches have been developed for estimating the relative abundance of different cell types in the tumor microenvironment (TME) using bulk tumor RNA data. However, a comprehensive comparison across diverse datasets that objectively evaluates the performance of these approaches has not been conducted. Here, we benchmarked seven widely used tools and gene sets and introduced ConsensusTME, a method that integrates gene sets from all the other methods for relative TME cell estimation of 18 cell types. We collected a comprehensive benchmark dataset consisting of pan-cancer data (DNA-derived purity, leukocyte methylation, and hematoxylin and eosin–derived lymphocyte counts) and cell-specific benchmark datasets (peripheral blood cells and tumor tissues). Although none of the methods outperformed others in every benchmark, ConsensusTME ranked top three in all cancer-related benchmarks and was the best performing tool overall. We provide a Web resource to interactively explore the benchmark results and an objective evaluation to help researchers select the most robust and accurate method to further investigate the role of the TME in cancer (www.consensusTME.org).
Significance:
This work shows an independent and comprehensive benchmarking of recently developed and widely used tumor microenvironment cell estimation methods based on bulk expression data and integrates the tools into a consensus approach.
Data from Macrophage-Derived IL1β and TNFα Regulate Arginine Metabolism in Neuroblastoma
Authors: Fultang Livingstone , Gamble Laura D. , Gneo Luciana , Berry Andrea M. , Egan Sharon A. , De Bie Fenna , Yogev Orli , Eden Georgina L. , Booth Sarah , Brownhill Samantha , Vardon Ashley , McConville Carmel M. , Cheng Paul N. , Norris Murray D. , Etchevers Heather C. , Murray Jayne , Ziegler David S. , Chesler Louis , Schmidt Ronny , Burchill Susan A. , Haber Michelle , De Santo Carmela , Mussai Francis
Published: March 2023

Abstract
Neuroblastoma is the most common childhood solid tumor, yet the prognosis for high-risk disease remains poor. We demonstrate here that arginase 2 (ARG2) drives neuroblastoma cell proliferation via regulation of arginine metabolism. Targeting arginine metabolism, either by blocking cationic amino acid transporter 1 (CAT-1)–dependent arginine uptake in vitro or therapeutic depletion of arginine by pegylated recombinant arginase BCT-100, significantly delayed tumor development and prolonged murine survival. Tumor cells polarized infiltrating monocytes to an M1-macrophage phenotype, which released IL1β and TNFα in a RAC-alpha serine/threonine-protein kinase (AKT)–dependent manner. IL1β and TNFα established a feedback loop to upregulate ARG2 expression via p38 and extracellular regulated kinases 1/2 (ERK1/2) signaling in neuroblastoma and neural crest–derived cells. Proteomic analysis revealed that enrichment of IL1β and TNFα in stage IV human tumor microenvironments was associated with a worse prognosis. These data thus describe an immune-metabolic regulatory loop between tumor cells and infiltrating myeloid cells regulating ARG2, which can be clinically exploited.
Significance:
These findings illustrate that cross-talk between myeloid cells and tumor cells creates a metabolic regulatory loop that promotes neuroblastoma progression.