Oncology
Abstract
NK cells, lymphocytes of the innate immune system, are important for defense against infectious pathogens and cancer. Classically, the CD56dim NK cell subset is thought to mediate antitumor responses, whereas the CD56bright subset is involved in immunomodulation. Here, we challenge this paradigm by demonstrating that brief priming with IL-15 markedly enhanced the antitumor response of CD56bright NK cells. Priming improved multiple CD56bright cell functions: degranulation, cytotoxicity, and cytokine production. Primed CD56bright cells from leukemia patients demonstrated enhanced responses to autologous blasts in vitro, and primed CD56bright cells controlled leukemia cells in vivo in a murine xenograft model. Primed CD56bright cells from multiple myeloma (MM) patients displayed superior responses to autologous myeloma targets, and furthermore, CD56bright NK cells from MM patients primed with the IL-15 receptor agonist ALT-803 in vivo displayed enhanced ex vivo functional responses to MM targets. Effector mechanisms contributing to IL-15–based priming included improved cytotoxic protein expression, target cell conjugation, and LFA-1–, CD2-, and NKG2D-dependent activation of NK cells. Finally, IL-15 robustly stimulated the PI3K/Akt/mTOR and MEK/ERK pathways in CD56bright compared with CD56dim NK cells, and blockade of these pathways attenuated antitumor responses. These findings identify CD56bright NK cells as potent antitumor effectors that warrant further investigation as a cancer immunotherapy.
Authors
Julia A. Wagner, Maximillian Rosario, Rizwan Romee, Melissa M. Berrien-Elliott, Stephanie E. Schneider, Jeffrey W. Leong, Ryan P. Sullivan, Brea A. Jewell, Michelle Becker-Hapak, Timothy Schappe, Sara Abdel-Latif, Aaron R. Ireland, Devika Jaishankar, Justin A. King, Ravi Vij, Dennis Clement, Jodie Goodridge, Karl-Johan Malmberg, Hing C. Wong, Todd A. Fehniger
Abstract
Tumors adapt to an unfavorable microenvironment by controlling the balance between cell proliferation and cell motility, but the regulators of this process are largely unknown. Here, we show that an alternatively spliced isoform of syntaphilin (SNPH), a cytoskeletal regulator of mitochondrial movements in neurons, is directed to mitochondria of tumor cells. Mitochondrial SNPH buffers oxidative stress and maintains complex II–dependent bioenergetics, sustaining local tumor growth while restricting mitochondrial redistribution to the cortical cytoskeleton and tumor cell motility. Conversely, introduction of stress stimuli to the microenvironment, including hypoxia, acutely lowered SNPH levels, resulting in bioenergetics defects and increased superoxide production. In turn, this suppressed tumor cell proliferation but increased tumor cell invasion via greater mitochondrial trafficking to the cortical cytoskeleton. Loss of SNPH or expression of an SNPH mutant lacking the mitochondrial localization sequence resulted in increased metastatic dissemination in xenograft or syngeneic tumor models in vivo. Accordingly, tumor cells that acquired the ability to metastasize in vivo constitutively downregulated SNPH and exhibited higher oxidative stress, reduced cell proliferation, and increased cell motility. Therefore, SNPH is a stress-regulated mitochondrial switch of the cell proliferation-motility balance in cancer, and its pathway may represent a therapeutic target.
Authors
M. Cecilia Caino, Jae Ho Seo, Yuan Wang, Dayana B. Rivadeneira, Dmitry I. Gabrilovich, Eui Tae Kim, Ashani T. Weeraratna, Lucia R. Languino, Dario C. Altieri
Abstract
The gene that encodes de novo DNA methyltransferase 3A (DNMT3A) is frequently mutated in acute myeloid leukemia genomes. Point mutations at position R882 have been shown to cause a dominant negative loss of DNMT3A methylation activity, but 15% of DNMT3A mutations are predicted to produce truncated proteins that could either have dominant negative activities or cause loss of function and haploinsufficiency. Here, we demonstrate that 3 of these mutants produce truncated, inactive proteins that do not dimerize with WT DNMT3A, strongly supporting the haploinsufficiency hypothesis. We therefore evaluated hematopoiesis in mice heterozygous for a constitutive null Dnmt3a mutation. With no other manipulations, Dnmt3a+/– mice developed myeloid skewing over time, and their hematopoietic stem/progenitor cells exhibited a long-term competitive transplantation advantage. Dnmt3a+/– mice also spontaneously developed transplantable myeloid malignancies after a long latent period, and 3 of 12 tumors tested had cooperating mutations in the Ras/MAPK pathway. The residual Dnmt3a allele was neither mutated nor downregulated in these tumors. The bone marrow cells of Dnmt3a+/– mice had a subtle but statistically significant DNA hypomethylation phenotype that was not associated with gene dysregulation. These data demonstrate that haploinsufficiency for Dnmt3a alters hematopoiesis and predisposes mice (and probably humans) to myeloid malignancies by a mechanism that is not yet clear.
Authors
Christopher B. Cole, David A. Russler-Germain, Shamika Ketkar, Angela M. Verdoni, Amanda M. Smith, Celia V. Bangert, Nichole M. Helton, Mindy Guo, Jeffery M. Klco, Shelly O’Laughlin, Catrina Fronick, Robert Fulton, Gue Su Chang, Allegra A. Petti, Christopher A. Miller, Timothy J. Ley
Abstract
Blood vessels in the tumor periphery have high pericyte coverage and are resistant to vascular disrupting agents (VDAs). VDA treatment resistance leads to a viable peripheral tumor rim that contributes to treatment failure and disease recurrence. Here, we provide evidence to support a hypothesis that shifting the target of VDAs from tumor vessel endothelial cells to pericytes disrupts tumor peripheral vessels and the viable rim, circumventing VDA treatment resistance. Through chemical engineering, we developed Z-GP-DAVLBH (from the tubulin-binding VDA desacetylvinblastine monohydrazide [DAVLBH]) as a prodrug that can be selectively activated by fibroblast activation protein α (FAPα) in tumor pericytes. Z-GP-DAVLBH selectively destroys the cytoskeleton of FAPα-expressing tumor pericytes, disrupting blood vessels both within the core and around the periphery of tumors. As a result, Z-GP-DAVLBH treatment eradicated the otherwise VDA-resistant tumor rim and led to complete regression of tumors in multiple lines of xenografts without producing the drug-related toxicity that is associated with similar doses of DAVLBH. This study demonstrates that targeting tumor pericytes with an FAPα-activated VDA prodrug represents a potential vascular disruption strategy in overcoming tumor resistance to VDA treatments.
Authors
Minfeng Chen, Xueping Lei, Changzheng Shi, Maohua Huang, Xiaobo Li, Baojian Wu, Zhengqiu Li, Weili Han, Bin Du, Jianyang Hu, Qiulin Nie, Weiqian Mai, Nan Ma, Nanhui Xu, Xinyi Zhang, Chunlin Fan, Aihua Hong, Minghan Xia, Liangping Luo, Ande Ma, Hongsheng Li, Qiang Yu, Heru Chen, Dongmei Zhang, Wencai Ye
Abstract
Claudin-low breast cancer is an aggressive subtype that confers poor prognosis and is found largely within the clinical triple-negative group of breast cancer patients. Here, we have shown that intrinsic and immune cell gene signatures distinguish the claudin-low subtype clinically as well as in mouse models of other breast cancer subtypes. Despite adaptive immune cell infiltration in claudin-low tumors, treatment with immune checkpoint inhibitory antibodies against cytotoxic T lymphocyte–associated protein 4 (CTLA-4) and programmed death receptor 1 (PD-1) were ineffective in controlling tumor growth. CD4+FoxP3+ Tregs represented a large proportion of the tumor-infiltrating lymphocytes (TILs) in claudin-low tumors, and Tregs isolated from tumor-bearing mice were able to suppress effector T cell responses. Tregs in the tumor microenvironment highly expressed PD-1 and were recruited partly through tumor generation of the chemokine CXCL12. Antitumor efficacy required stringent Treg depletion combined with checkpoint inhibition; delays in tumor growth were not observed using therapies that modestly diminished the number of Tregs in the tumor microenvironment. This study provides evidence that the recruitment of Tregs to the tumor microenvironment inhibits an effective antitumor immune response and highlights early Treg recruitment as a possible mechanism for the lack of response to immune checkpoint blockade antibodies in specific subtypes of cancer that are heavily infiltrated with adaptive immune cells.
Authors
Nicholas A. Taylor, Sarah C. Vick, Michael D. Iglesia, W. June Brickey, Bentley R. Midkiff, Karen P. McKinnon, Shannon Reisdorf, Carey K. Anders, Lisa A. Carey, Joel S. Parker, Charles M. Perou, Benjamin G. Vincent, Jonathan S. Serody
Abstract
The histone H3K36 methyltransferase SETD2 is frequently mutated or deleted in a variety of human tumors. Nevertheless, the role of SETD2 loss in oncogenesis remains largely undefined. Here, we found that SETD2 counteracts Wnt signaling and its inactivation promotes intestinal tumorigenesis in mouse models of colorectal cancer (CRC). SETD2 was not required for intestinal homeostasis under steady state; however, upon irradiation, genetic inactivation of Setd2 in mouse intestinal epithelium facilitated the self-renewal of intestinal stem/progenitor cells as well as tissue regeneration. Furthermore, depletion of SETD2 enhanced the susceptibility to tumorigenesis in the context of dysregulated Wnt signaling. Mechanistic characterizations indicated that SETD2 downregulation affects the alternative splicing of a subset of genes implicated in tumorigenesis. Importantly, we uncovered that SETD2 ablation reduces intron retention of dishevelled segment polarity protein 2 (DVL2) pre-mRNA, which would otherwise be degraded by nonsense-mediated decay, thereby augmenting Wnt signaling. The signaling cascades mediated by SETD2 were further substantiated by a CRC patient cohort analysis. Together, our studies highlight SETD2 as an integral regulator of Wnt signaling through epigenetic regulation of RNA processing during tissue regeneration and tumorigenesis.
Authors
Huairui Yuan, Ni Li, Da Fu, Jiale Ren, Jingyi Hui, Junjie Peng, Yongfeng Liu, Tong Qiu, Min Jiang, Qiang Pan, Ying Han, Xiaoming Wang, Qintong Li, Jun Qin
Abstract
The most frequent chromosomal structural loss in hepatocellular carcinoma (HCC) is of the short arm of chromosome 8 (8p). Genes on the remaining homologous chromosome, however, are not recurrently mutated, and the identity of key 8p tumor-suppressor genes (TSG) is unknown. In this work, analysis of minimal commonly deleted 8p segments to identify candidate TSG implicated GATA4, a master transcription factor driver of hepatocyte epithelial lineage fate. In a murine model, liver-conditional deletion of 1 Gata4 allele to model the haploinsufficiency seen in HCC produced enlarged livers with a gene expression profile of persistent precursor proliferation and failed hepatocyte epithelial differentiation. HCC mimicked this gene expression profile, even in cases that were morphologically classified as well differentiated. HCC with intact chromosome 8p also featured GATA4 loss of function via GATA4 germline mutations that abrogated GATA4 interactions with a coactivator, MED12, or by inactivating mutations directly in GATA4 coactivators, including ARID1A. GATA4 reintroduction into GATA4-haploinsufficient HCC cells or ARID1A reintroduction into ARID1A-mutant/GATA4-intact HCC cells activated hundreds of hepatocyte genes and quenched the proliferative precursor program. Thus, disruption of GATA4-mediated transactivation in HCC suppresses hepatocyte epithelial differentiation to sustain replicative precursor phenotype.
Authors
Francis O. Enane, Wai Ho Shuen, Xiaorong Gu, Ebrahem Quteba, Bartlomiej Przychodzen, Hideki Makishima, Juraj Bodo, Joanna Ng, Chit Lai Chee, Rebecca Ba, Lip Seng Koh, Janice Lim, Rachael Cheong, Marissa Teo, Zhenbo Hu, Kwok Peng Ng, Jaroslaw Maciejewski, Tomas Radivoyevitch, Alexander Chung, London Lucien Ooi, Yu Meng Tan, Peng Chung Cheow, Pierce Chow, Chung Yip Chan, Kiat Hon Lim, Lisa Yerian, Eric Hsi, Han Chong Toh, Yogen Saunthararajah
Abstract
Accumulating evidence suggests that glioma stem cells (GSCs) are important therapeutic targets in glioblastoma (GBM). In this study, we identified NIMA-related kinase 2 (NEK2) as a functional binding protein of enhancer of zeste homolog 2 (EZH2) that plays a critical role in the posttranslational regulation of EZH2 protein in GSCs. NEK2 was among the most differentially expressed kinase-encoding genes in GSC-containing cultures (glioma spheres), and it was required for in vitro clonogenicity, in vivo tumor propagation, and radioresistance. Mechanistically, the formation of a protein complex comprising NEK2 and EZH2 in glioma spheres phosphorylated and then protected EZH2 from ubiquitination-dependent protein degradation in a NEK2 kinase activity–dependent manner. Clinically, NEK2 expression in patients with glioma was closely associated with EZH2 expression and correlated with a poor prognosis. NEK2 expression was also substantially elevated in recurrent tumors after therapeutic failure compared with primary untreated tumors in matched GBM patients. We designed a NEK2 kinase inhibitor, compound 3a (CMP3a), which efficiently attenuated GBM growth in a mouse model and exhibited a synergistic effect with radiotherapy. These data demonstrate a key role for NEK2 in maintaining GSCs in GBM by stabilizing the EZH2 protein and introduce the small-molecule inhibitor CMP3a as a potential therapeutic agent for GBM.
Authors
Jia Wang, Peng Cheng, Marat S. Pavlyukov, Hai Yu, Zhuo Zhang, Sung-Hak Kim, Mutsuko Minata, Ahmed Mohyeldin, Wanfu Xie, Dongquan Chen, Violaine Goidts, Brendan Frett, Wenhao Hu, Hongyu Li, Yong Jae Shin, Yeri Lee, Do-Hyun Nam, Harley I. Kornblum, Maode Wang, Ichiro Nakano
Abstract
The tumor microenvironment plays an important role in tumor growth and metastasis. However, the mechanism by which tumor cells regulate the cell and non-cell constituents of surrounding stroma remains incompletely understood. Promyelocytic leukemia (PML) is a pleiotropic tumor suppressor, but its role in tumor microenvironment regulation is poorly characterized. PML is frequently downregulated in many cancer types, including lung cancer. Here, we identify a PML ubiquitination pathway that is mediated by WD repeat 4–containing cullin-RING ubiquitin ligase 4 (CRL4WDR4). Clinically, this PML degradation pathway is hyperactivated in lung cancer and correlates with poor prognosis. The WDR4/PML axis induces a set of cell-surface or secreted factors, including CD73, urokinase-type plasminogen activator receptor (uPAR), and serum amyloid A2 (SAA2), which elicit paracrine effects to stimulate migration, invasion, and metastasis in multiple lung cancer models. In xenograft and genetically engineered mouse models, the WDR4/PML axis elevates intratumoral Tregs and M2-like macrophages and reduces CD8+ T cells to promote lung tumor growth. These immunosuppressive effects were all reversed by CD73 blockade. Our study identifies WDR4 as an oncoprotein that negatively regulates PML via ubiquitination to promote lung cancer progression by fostering an immunosuppressive and prometastatic tumor microenvironment, suggesting the potential of immune-modulatory approaches for treating lung cancer with aberrant PML degradation.
Authors
Ya-Ting Wang, Jocelyn Chen, Chou-Wei Chang, Jayu Jen, Tzu-Yu Huang, Chun-Ming Chen, Roger Shen, Suh-Yuen Liang, I-Cheng Cheng, Shuenn-Chen Yang, Wu-Wei Lai, Kuang-Hung Cheng, Tao-Shih Hsieh, Ming-Zong Lai, Hung-Chi Cheng, Yi-Ching Wang, Ruey-Hwa Chen
Abstract
Current anti-VEGF therapies for colorectal cancer (CRC) provide limited survival benefit, as tumors rapidly develop resistance to these agents. Here, we have uncovered an immunosuppressive role for nonclassical Ly6Clo monocytes that mediates resistance to anti-VEGFR2 treatment. We found that the chemokine CX3CL1 was upregulated in both human and murine tumors following VEGF signaling blockade, resulting in recruitment of CX3CR1+Ly6Clo monocytes into the tumor. We also found that treatment with VEGFA reduced expression of CX3CL1 in endothelial cells in vitro. Intravital microscopy revealed that CX3CR1 is critical for Ly6Clo monocyte transmigration across the endothelium in murine CRC tumors. Moreover, Ly6Clo monocytes recruit Ly6G+ neutrophils via CXCL5 and produce IL-10, which inhibits adaptive immunity. Preventing Ly6Clo monocyte or Ly6G+ neutrophil infiltration into tumors enhanced inhibition of tumor growth with anti-VEGFR2 therapy. Furthermore, a gene therapy using a nanoparticle formulated with an siRNA against CX3CL1 reduced Ly6Clo monocyte recruitment and improved outcome of anti-VEGFR2 therapy in mouse CRCs. Our study unveils an immunosuppressive function of Ly6Clo monocytes that, to our knowledge, has yet to be reported in any context. We also reveal molecular mechanisms underlying antiangiogenic treatment resistance, suggesting potential immunomodulatory strategies to enhance the long-term clinical outcome of anti-VEGF therapies.
Authors
Keehoon Jung, Takahiro Heishi, Omar F. Khan, Piotr S. Kowalski, Joao Incio, Nuh N. Rahbari, Euiheon Chung, Jeffrey W. Clark, Christopher G. Willett, Andrew D. Luster, Seok Hyun Yun, Robert Langer, Daniel G. Anderson, Timothy P. Padera, Rakesh K. Jain, Dai Fukumura
Copyright © 2017 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)