Novel therapeutic strategies, including immunotherapeutics, targeting glioblastoma (GBM) often fail in the clinic, at least partly because available preclinical models in which hypotheses are being tested, do not recapitulate the human disease. To address this challenge, we took advantage of our previously developed spontaneous Qk/trp53/Pten (QPP) triple-knockout model of human GBM, and compared its immune microenvironment components with those of patient-derived tumors in effort to determine whether this model might provide an opportunity for gaining insights into tumor physiopathology as well as for preclinical evaluation of therapeutic agents. Immune profiling analyses and single-cell sequencing of implanted and spontaneous tumors from QPP mice as well as from GBM patients revealed intratumoral immune components that were predominantly myeloid cells (e.g. monocytes, macrophages, and microglia) with minor populations of T, B, and NK cells. When comparing spontaneous and implanted mouse samples, we found that there were more neutrophils, T and NK cells in the implanted model. Neutrophils, T and NK cells were increased in abundance in samples derived from human high-grade glioma (HGG) compared to those derived from low grade glioma (LGG). Overall, our data demonstrate that our implanted and spontaneous QPP models recapitulate the immunosuppressive myeloid dominant nature of the tumor microenvironment of human gliomas. Our model provides a suitable tool for investigating the complex immune compartment of gliomas and it may contribute to a better understanding of the resistance of human glioblastoma to currently available immunotherapeutics.
Daniel B. Zamler, Takashi Shingu, Laura M. Kahn, Kristin Huntoon, Cynthia Kassab, Martina Ott, Katarzyna Tomczak, Jintan Liu, Yating Li, Ivy Lai, Rocio Zorilla-Veloz, Cassian Yee, Kunal Rai, Betty Y.S. Kim, Stephanie S. Watowich, Amy B. Heimberger, Giulio F. Draetta, Jian Hu
Apolipoprotein C-III (apoC-III) is a critical regulator of triglyceride metabolism and correlates positively with hypertriglyceridemia and cardiovascular disease (CVD). ApoC-III also induces sterile inflammation via inflammasome activation, another CVD risk factor. It remains unclear if therapeutic apoC-III lowering reduces CVD risk, nor is it understood if the CVD correlation depends on the lipid-lowering or anti-inflammatory properties. Therefore, we determined the impact of interventional apoC-III lowering on atherogenesis via apoC-III antisense oligonucleotide (ASO) administration in two hypertriglyceridemic mouse models where the intervention lowers plasma triglycerides (Apoe-/-Ndst1f/fAlb-Cre+, Ldlr-/-Ndst1f/fAlb-Cre+) and in a third lipid-refractory model where the ASO cannot lower plasma triglycerides (Ldlr-/-Lrp1f/fAlb-Cre+). A high-cholesterol Western diet ApoC-III ASO treatment did not alter atherosclerotic lesion size but did significantly attenuate advanced and unstable plaque development in the two triglyceride responsive mouse models. In contrast, no lesion size or composition improvement was observed with apoC-III ASO in the lipid-refractory Ldlr-/-Lrp1f/fAlb-Cre+ mice. To circumvent confounding effects of continuous high cholesterol feeding, we tested the impact of interventional apoC-III lowering when switching to a cholesterol-poor diet after 12-weeks of Western diet. In this diet-switch regimen, ApoC-III ASO treatment significantly reduced plasma triglycerides, atherosclerotic lesion progression, and necrotic core area and increased fibrous cap thickness in Ldlr-/-Ndst1f/fAlb-Cre+ mice. Again, ApoC-III ASO treatment did not alter triglyceride levels, lesion development and lesion composition in Ldlr-/-Lrp1f/fAlb-Cre+ mice after the diet-switch. Thus, therapeutic apoC-III targeting increased fibrous cap thickness, and reduced necrotic core area and lesion size after diet intervention when triglyceride-lowering is achieved in murine models. Our findings suggest that interventional apoC-III lowering might be an effective strategy to reduce atherosclerosis lesion size and improve plaque stability.
Bastian Ramms, Sohan Patel, Xiaoli Sun, Ariane R. Pessentheiner, G. Michelle Ducasa, Adam E. Mullick, Richard G. Lee, Rosanne M. Crooke, Sotirios Tsimikas, Joseph L. Witztum, Philip L.S.M. Gordts
Vaccine-elicited SARS-CoV-2 antibody responses are an established correlate of protection against viral infection in humans and non-human primates. However, it is less clear that vaccine-induced immunity is able to limit infection-elicited inflammation in the lower respiratory tract. To assess this, we collected bronchoalveolar lavage fluid samples post-SARS-CoV-2 strain USA-WA1/2020 challenge from rhesus macaques vaccinated with mRNA-1273 in a dose-reduction study. Single-cell transcriptomic profiling revealed a broad cellular landscape 48 hours post-challenge with distinct inflammatory signatures that correlated with viral RNA burden in the lower respiratory tract. These inflammatory signatures included phagocyte-restricted expression of chemokines such as CXCL10 (IP10) and CCL3 (MIP-1A) and the broad expression of interferon-induced genes such as MX1, ISG15, and IFIT1. Induction of these inflammatory profiles was suppressed by prior mRNA-1273 vaccination in a dose-dependent manner, and negatively correlated with pre-challenge serum and lung antibody titers against SARS-CoV-2 spike. These observations were replicated and validated in a second independent macaque challenge study using the B.1.351/beta-variant of SARS-CoV-2. These data support a model wherein vaccine-elicited antibody responses restrict viral replication following SARS-CoV-2 exposure, including limiting viral dissemination to the lower respiratory tract and infection-mediated inflammation and pathogenesis.
Adam T. Waickman, Kaitlin Victor, Krista Newell, Tao Li, Heather Friberg, Kathryn E. Foulds, Mario Roederer, Diane L. Bolton, Jeffrey R. Currier, Robert Seder
Most patients with neovascular age-related macular degeneration (nvAMD), the leading cause of severe vision loss in elderly Americans, respond inadequately to current therapies targeting a single angiogenic mediator, vascular endothelial growth factor (VEGF). Here we report that aqueous levels of a second vasoactive mediator, angiopoietin-like 4 (ANGPTL4), can help predict the response of nvAMD patients to anti-VEGF therapies. ANGPTL4 expression was higher in patients who required monthly treatment with anti-VEGF therapies compared to patients who could be effectively treated with less frequent injections. We further demonstrate that ANGPTL4 acts synergistically with VEGF to promote the growth and leakage of choroidal neovascular (CNV) lesions in mice. Targeting ANGPTL4 expression was as effective as targeting VEGF expression for treating CNV in mice, while simultaneously targeting both was more effective than targeting either factor alone. To help translate these findings to patients, we used a soluble receptor that binds to both VEGF and ANGPTL4 and effectively inhibited the development of CNV lesions in mice. Our findings provide an assay that can help predict the response of nvAMD patients to anti-VEGF monotherapy and suggest that therapies targeting both ANGPTL4 and VEGF will be a more effective approach for the treatment of this blinding disease.
Yu Qin, Aumreetam Dinabandhu, Xuan Cao, Jaron C. Sanchez, Kathleen Jee, Murilo Rodrigues, Chuanyu Guo, Jing Zhang, Jordan Vancel, Deepak Menon, Noore-Sabah Khan, Tao Ma, Stephany Y. Tzeng, Yassine J. Daoud, Jordan J. Green, Gregg L. Semenza, Silvia Montaner, Akrit Sodhi
Women of African ancestry suffer higher rates of breast cancer mortality compared to all other groups in the United States. Though the precise reasons for these disparities remain unclear, many recent studies have implicated a role for differences in tumor biology. Using an epitope-validated antibody against the endoplasmic reticulum-associated degradation (ERAD) E3 ubiquitin ligase, gp78, we show that elevated levels of gp78 in patient breast cancer cells predict poor survival. Moreover, high levels of gp78 are associated with poor outcomes in both ER-positive and ER-negative tumors, and breast cancers expressing elevated amounts of gp78 protein are enriched in gene expression pathways that influence cell cycle, metabolism, receptor-mediated signaling, and cell stress response pathways. In multivariate analysis adjusted for subtype and grade, gp78 protein is an independent predictor of poor outcomes in women of African ancestry. Furthermore, gene expression signatures, derived from patients stratified by gp78 protein expression, are strong predictors of recurrence and pathological complete response in retrospective clinical trial data and share many common features with gene sets previously identified to be overrepresented in breast cancers based on race. These findings implicate a prominent role for gp78 in tumor progression and offer new insights into our understanding of racial differences in breast cancer outcomes.
Sandeep K. Singhal, Jung S. Byun, Tingfen Yan, Ryan Yancey, Ambar Caban, Sara Gil Hernandez, Sediqua Bufford, Stephen M. Hewitt, Joy Winfield, Jaya Sarin Pradhan, Vesco Mustkov, Jasmine A. McDonald, Eliseo J. Pérez-Stable, Anna Maria Napoles, Nasreen Vohra, Adriana De Siervi, Clayton Yates, Melissa B. Davis, Mei Yang, Yien Che Tsai, Allan M. Weissman, Kevin Gardner
LAMA2-deficiency, resulting from a defective or absent laminin α2-subunit, is a common cause of congenital muscular dystrophy. It is characterized by muscle weakness from myofiber degeneration and neuropathy from Schwann cell amyelination. Previously it was shown that transgenic muscle-specific expression of αLNNd, a laminin-γ1-binding linker protein that ena-bles polymerization in defective laminins, selectively ameliorates the muscle abnormality in mouse disease models. Here, adeno-associated virus (AAV) was used to deliver linker mini-genes to dystrophic dy2J/dy2J mice for expression of either αLNNd in muscle, or αLNNdΔG2’, a shortened linker, in muscle, nerve and other tissues. Linker and laminin-α2 levels were higher in αLNNdΔG2’-treated mice. Both αLNNd- and αLNNdΔG2’-treated mice exhibited increased fore-limb grip strength. Further, αLNNdΔG2’-treated mice achieved hindlimb and all-limb grip strength levels approaching those of wild-type mice as well as ablation of hindlimb paresis and contractures. Improvement of muscle histology was evident in the muscle-specific αLNNd-ex-pressing mice but more extensive in the αLNNdΔG2'-expressing mice, along with restoration of sciatic nerve axonal envelopment and myelination. The results reveal that an αLN-linker mini-gene, driven by a ubiquitous promoter is superior to muscle-specific delivery through higher ex-pression that extends to peripheral nerve. These studies support a novel approach of somatic gene therapy.
Karen K. McKee, Peter D. Yurchenco
Liver diseases have become a major comorbidity health concern in people living with HIV-1 (PLWH) under combination antiretroviral therapy (cART). To investigate if HIV-1 infection and cART interact to lead to liver diseases, humanized mice reconstituted with progenitor cells from human fetal livers were infected with HIV-1 and treated with cART. We report here that chronic HIV-1 infection with cART induced hepatitis and liver fibrosis in humanized mice, associated with accumulation of M2-like macrophages (M2LM), elevated TGFβ and interferon signaling in the liver. Interestingly, IFN-I and TGFβ cooperatively activated human hepatic stellate cells (HepSC) in vitro. Mechanistically, IFN-I enhanced TGFβ-induced SMAD2/3 activation in HepSC. Finally, blockade of IFN-I signaling reversed HIV/cART-induced liver diseases in humanized mice. Consistent with the findings in humanized mice with HIV-1 and cART, we detected elevated markers of liver injury, M2-like macrophages, and of interferon signaling in blood specimens from PLWH over healthy individuals. These findings identify the IFN-I/M2LM/HepSC axis in HIV/cART-induced liver diseases and suggest that inhibiting IFN-I signaling or M2LM may provide a novel therapeutic strategy for treating HIV/cART-associated liver diseases in PLWH under ART.
James Ahodantin, Kouki Nio, Masaya Funaki, Xuguang Zhai, Eleanor Wilson, Shyamasundaran Kottilil, Liang Cheng, Guangming Li, Lishan Su
Disruption of the neurovascular unit (NVU) underlies the pathophysiology of various CNS diseases.(1-3) One strategy to repair NVU dysfunction would use stem/progenitor cells to provide trophic support to the NVU’s functionally coupled and interdependent vasculature and surrounding CNS parenchyma.(4) A subset of endothelial progenitor cells, endothelial colony forming cells (ECFCs) with high expression of the CD44 hyaluronan receptor (CD44hi), provides such neurovasculotrophic support via a paracrine mechanism.(5) Here, we report that bioactive extracellular vesicles from CD44hi ECFCs (EVshi) are paracrine mediators, recapitulating the effects of intact cell therapy in murine models of ischemic/neurodegenerative retinopathy; vesicles from ECFCs with low expression levels of CD44 (EVslo) were ineffective. Small RNA sequencing comparing the microRNA (miR) cargo from EVshi and EVslo identified candidate miRs that contribute to these effects. EVshi may be used to repair NVU dysfunction through multiple mechanisms to stabilize hypoxic vasculature, promote vascular growth, and support neural cells.
Kyle V. Marra, Edith Aguilar, Wei Guoqin, Ayumi Usui-Ouchi, Yochiro Ideguchi, Susumu Sakimoto, Martin Friedlander
Long non-coding RNA (lncRNA) plays a crucial role in the pathogenesis of various diseases, including colorectal cancer (CRC). The gene mutations of Adenomatous polyposis coli (APC) were found in most colorectal cancer patients. They are functioned as an important inducer of tumorigenesis. Based on our microarray results, we identified a specific upregulated lncRNA in colorectal cancer (SURC). Further analysis showed that high SURC expression correlated with poorer disease-free survival and overall survival in patients with colorectal cancer. Besides, we found that mutated APC genes can promote the transcription of SURC by reducing the degradation of β-catenin protein in colorectal cancer. Functional assays revealed that knockdown of SURC impaired CRC cell proliferation, colony formation, cell cycle and tumor growth. Additionally, SURC can promote CCND2 expression by inhibiting the expression of miR-185-5p in CRC cells. In conclusion, we demonstrate that SURC is a specific upregulated lncRNA in CRC and the SURC/miR-185-5p/CCND2 axis may be targetable for CRC diagnosis and therapy.
Junshu Li, Yanhong Ji, Na Chen, Huiling Wang, Chao Fang, Xiaonan Yin, Zhiyuan Jiang, Zhexu Dong, Dan Zhu, Jiamei Fu, Wencheng Zhou, Ruiyi Jiang, Ling He, Zhang Hantao, Gang Shi, Lin Cheng, Xiaolan Su, Lei Dai, Hongxin Deng
The role of immune responses to previously seen endemic coronavirus epitopes in severe acute respiratory coronavirus 2 (SARS-CoV-2) infection and disease progression has not yet been determined. Here, we show that a key characteristic of fatal coronavirus disease (COVID-19) outcomes is that the immune response to the SARS-CoV-2 spike protein is enriched for antibodies directed against epitopes shared with endemic beta-coronaviruses, and has a lower proportion of antibodies targeting the more protective variable regions of the spike. The magnitude of antibody responses to the SARS-CoV-2 full-length spike protein, its domains and subunits, and the SARS-CoV-2 nucleocapsid also correlated strongly with responses to the endemic beta-coronavirus spike proteins in individuals admitted to intensive care units (ICU) with fatal COVID-19 outcomes, but not in individuals with non-fatal outcomes. This correlation was found to be due to the antibody response directed at the S2 subunit of the SARS-CoV-2 spike protein, which has the highest degree of conservation between the beta-coronavirus spike proteins. Intriguingly, antibody responses to the less cross-reactive SARS-CoV-2 nucleocapsid were not significantly different in individuals who were admitted to ICU with fatal and non-fatal outcomes, suggesting an antibody profile in individuals with fatal outcomes consistent with an original antigenic sin type-response.
Anna L. McNaughton, Robert S. Paton, Matthew Edmans, Jonathan C.W. Youngs, Judith Wellens, Prabhjeet Phalora, Alex Fyfe, Sandra Belij-Rammerstorfer, Jai S. Bolton, Jonathan Ball, George W. Carnell, Wanwisa Dejnirattisai, Christina Dold, David W. Eyre, Philip Hopkins, Alison Howarth, Kreepa Kooblall, Hannah Klim, Susannah Leaver, Lian N. Lee, César López-Camacho, Sheila F. Lumley, Derek C. Macallan, Alexander J. Mentzer, Nicholas M. Provine, Jeremy Ratcliff, Jose L. Slon-Campos, Donal T. Skelly, Lucas B. Stolle, Piyada Supasa, Nigel Temperton, Chris Walker, Beibei Wang, Duncan Wyncoll, Peter Simmonds, Teresa Lambe, John K. Ballie, Malcolm G. Semple, Peter J.M. Openshaw, Uri Obolski, Marc Turner, Miles Carroll, Juthathip Mongkolsapaya, Gavin Screaton, Stephen H. Kennedy, Lisa M. Jarvis, Eleanor Barnes, Susanna Dunachie, José Lourenço, Philippa C. Matthews, Tihana Bicanic, Paul Klenerman, Sunetra Gupta, Craig P. Thompson
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