Background: Pediatric and adult high-grade glioma (HGG) frequently harbor PDGFRA alterations. We hypothesized that co-treatment with everolimus may improve the efficacy of dasatinib in PDGFRα-driven glioma through combinatorial synergism and increased tumor accumulation of dasatinib. Methods: Dose response, synergism studies, P-gp inhibition and pharmacokinetic studies were performed on in vitro and in vivo human and mouse models of HGG. Six patients with recurrent PDGFRα-driven glioma were treated with dasatinib and everolimus. Results: Dasatinib effectively inhibited the proliferation of mouse and human primary HGG cells with a variety of PDGFRA alterations. Dasatinib exhibited synergy with everolimus in the treatment of HGG cells at low nanomolar concentrations of both agents, with reduction in mTOR signaling that persists after dasatinib treatment alone. Prolonged exposure to everolimus significantly improved the CNS retention of dasatinib and extended survival of PPK tumor bearing mice. Pediatric patients (n=6) with glioma tolerated this combination without significant adverse events. Recurrent patients (n=4) demonstrated median overall survival of 8.5 months. Conclusion: Efficacy of dasatinib treatment of PDGFRα-driven HGG is improved with everolimus and suggests a promising route for improving targeted therapy for this patient population. Trial Registration: ClinicalTrials.gov NCT03352427 Funding: The authors thank the patients and their families for participation in this study. CKis supported by NIH/NINDS K08-NS099427-01, the University of Michigan Chad Carr Pediatric Brain Tumor Center, the Chad Tough Foundation, Hyundai Hope on Wheels, Catching up With Jack, Prayers from Maria Foundation, U CAN-CER VIVE FOUNDATION, Morgan Behen Golf Classic, and the DIPG Collaborative. The PEDS-MIONCOSEQ study was supported by grant 1UM1HG006508 from the National Institutes of Health Clinical Sequencing Exploratory Research Award (PI: Arul Chinnaiyan).
Zachary Miklja, Viveka Nand Yadav, Rodrigo T. Cartaxo, Ruby Siada, Chase C. Thomas, Jessica R. Cummings, Brendan Mullan, Stefanie Stallard, Alyssa Paul, Amy K. Bruzek, Kyle Wierzbicki, Tao Yang, Taylor Garcia, Ian Wolfe, Marcia Leonard, Patricia L. Robertson, Hugh J.L. Garton, Daniel R. Wahl, Hemant A. Parmar, Jann N. Sarkaria, Cassie Kline, Sabine Mueller, Theodore Nicolaides, Chana Glasser, Sarah E. S. Leary, Sriram Venneti, Chandan Kumar-Sinha, Arul M. Chinnaiyan, Rajen Mody, Manjunath P. Pai, Timothy N. Phoenix, Bernard L. Marini, Carl Koschmann
Diabetic patients develop endothelial dysfunction shortly after diabetes onset that progresses to vascular disease underlying the majority of diabetes associated comorbidities. Increased lipid peroxidation, mitochondrial calcium overload and mitochondrial dysfunction are characteristics of dysfunctional endothelial cells in diabetic patients. We here identified that targeting the lipid peroxidation product 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE) induced activation of the intracellularly located cation channel transient receptor potential vanilloid 1 (TRPV1) in endothelial cells is a means to causally control early stage vascular disease in type I diabetic mice. Mice with an inducible, endothelial specific 12/15 lipoxygenase (12/15Lo) knock out were similarly protected from type I diabetes induced endothelial dysfunction and impaired vascular regeneration following arterial injury as TRPV1 knock out mice. Both 12(S)-HETE in concentrations found in diabetic patients and TRPV1 agonists triggered mitochondrial calcium influx and mitochondrial dysfunction in endothelial cells and 12(S)-HETE effects were absent in endothelial cells from TRPV1 knock out mice. As a therapeutic consequence, we found that a peptide targeting 12(S)-HETE induced TRPV1 interaction at the TRPV1 TRP box ameliorated diabetes-induced endothelial dysfunction and augmented vascular regeneration in diabetic mice. Our findings suggest that pharmacological targeting of increased endothelial lipid peroxidation can attenuate diabetes induced comorbidities related to vascular disease.
Mandy Otto, Clarissa Bucher, Wantao Liu, Melanie Müller, Tobias Schmidt, Marina Kardell, Marvin Noel Driessen, Jan Rossaint, Eric R. Gross, Nana-Maria Wagner
The correlation of HIV-specific Antibody-Dependent Cellular Cytotoxicity (ADCC) responses with protection from, and delayed progression of HIV-1 infection provides a rationale to leverage ADCC-mediating antibodies for treatment purposes. We evaluated ADCC mediated by different combinations of two to six neutralizing and non-neutralizing anti-HIV-1-Envelope (Env) monoclonal antibodies (mAbs), using concentrations ≤ 1 µg/mL, to identify combinations effective at targeting latent reservoir HIV-1 viruses (LRVs) from ten individuals. We found that within 2 hours, combinations of three mAbs mediated >30% killing of HIV-infected primary CD4+ T cells in presence of autologous NK cells, with the combination of A32 (C1C2), DH511.2K3 (MPER), and PGT121 (V3) mAbs being the most effective. Increasing the incubation of target and effector cells in presence of mAb combinations from 2 to 24 hours resulted in increased specific killing of infected cells, even with neutralization-resistant viruses. The same combination eliminated reactivated HIV-1 latently-infected cells in an ex vivo qualitative viral outgrowth (QVOA) assay. Therefore, administration of a combination of three mAbs should be considered when planning in vivo studies seeking to eliminate persistently HIV-1 infected cells.
Marina Tuyishime, Carolina Garrido, Shalini Jha, Matthew Moeser, Dieter Mielke, Celia LaBranche, David Montefiori, Barton F. Haynes, Sarah B. Joseph, David M. Margolis, Guido Ferrari
NF-kB transcription factors, driven by the IRAK-IKK cascade, confer treatment resistance in pancreatic ductal adenocarcinoma (PDAC), a cancer characterized by near universal KRAS mutation. Through reverse-phase protein array and RNAseq we discovered IRAK4 also contributes substantially to MAPK activation in KRAS-mutant PDAC. IRAK4 ablation completely blocked RAS-induced transformation of human and murine cells. Mechanistically, expression of mutant KRAS stimulated an inflammatory, autocrine IL-1b signaling loop that activated IRAK4 and MAPK pathway. Downstream of IRAK4, we uncovered TPL2/MAP3K8 as the essential kinase that propels both MAPK and NF-kB cascades. Inhibition of TPL2 blocked both MAPK and NF-kB signaling, and suppressed KRAS-mutant cell growth. To counter chemotherapy-induced genotoxic stress, PDAC cells upregulated TLR9, which activated pro-survival IRAK4-TPL2 signaling. Accordingly, TPL2 inhibitor synergized with chemotherapy to curb PDAC growth in vivo. Finally, from TCGA we characterized two MAP3K8 point mutations that hyperactivate MAPK and NF-kB cascades by impeding TPL2 protein degradation. Cancer cell lines naturally harboring these MAP3K8 mutations are strikingly sensitive to TPL2 inhibition, underscoring the need to identify these potentially targetable mutations in patients. Overall, our study establishes TPL2 as a promising therapeutic target in RAS- and MAP3K8-mutant cancers and strongly prompts development of TPL2 inhibitors for pre-clinical and clinical studies.
Paarth B. Dodhiawala, Namrata Khurana, Daoxiang Zhang, Yi Cheng, Lin Li, Qing Wei, Kuljeet Seehra, Hongmei Jiang, Patrick M. Grierson, Andrea Wang-Gillam, Kian-Huat Lim
Despite effective antiretroviral therapy, HIV-1-nfected cells continue to produce viral antigens and induce chronic immune exhaustion. We propose to identify HIV-1-suppressing agents which can inhibit HIV-1 reactivation and reduce HIV-1-induced immune activation. Using a novel dual reporter system and a high-throughput drug screen, we identified FDA-approved drugs which can suppress HIV-1 reactivation in both cell line models and CD4+ T cells from virally suppressed, HIV-1-infected individuals. We identified 11 cellular pathways required for HIV-1 reactivation as druggable targets. Using differential expression analysis, gene set enrichment analysis and exon-intron landscape analysis, we examined the impact of drug treatment on the cellular environment at a genome-wide level. We identified a new function of a JAK inhibitor filgotinib which suppresses HIV-1 splicing. First, filgotinib preferentially suppresses spliced HIV-1 RNA transcription. Second, filgotinib suppresses HIV-1-driven aberrant cancer-related gene expression at the integration site. Third, we found that filgotinib suppresses HIV-1 transcription by inhibiting T cell activation and by modulating RNA splicing. Finally, we found that filgotinib treatment reduces the proliferation of HIV-1-infected cells. Overall, the combination of a drug screen and transcriptome analysis provides systemic understanding of cellular targets required for HIV-1 reactivation and drug candidates that may reduce HIV-1-related immune activation.
Yang-Hui Jimmy Yeh, Katharine M. Jenike, Rachela M. Calvi, Jennifer Chiarella, Rebecca Hoh, Steven G. Deeks, Ya-Chi Ho
Mechanisms driving tumor progression from less aggressive subtypes to more aggressive states represent key targets for therapy. We identified a subset of Luminal A primary breast tumors to give rise to HER2-enriched (HER2E) subtype metastases, but remain clinically HER2 negative (cHER2-). By testing the unique genetic and transcriptomic features of these cases, we developed the hypothesis FGFR4 likely participates in this subtype switching. To evaluate this, we developed two FGFR4 genomic signatures using a PDX model treated with a FGFR4 inhibitor, which inhibited PDX growth in vivo. Bulk tumor gene expression analysis and single cell RNAseq demonstrated that the inhibition of FGFR4 signaling caused molecular switching. In the METABRIC breast cancer cohort,FGFR4-induced and FGFR4-repressed signatures each predicted overall survival. Additionally, FGFR4-induced signature was also an independent prognostic factor beyond subtype and stage. Supervised analysis of 77 primary tumors with paired metastasis revealed that the FGFR4-induced signature was significantly higher in luminal/ER+ tumor metastases compared with their primaries. Finally, multivariate analysis demonstrated that the FGFR4-induced signature also predicted site-specific metastasis for lung, liver and brain, but not for bone or lymph nodes. These data identify a link between FGFR4-regulated genes and metastasis, suggesting treatment options for FGFR4-positive patients, whose high expression is not caused by mutation or amplification.
Susana Garcia-Recio, Aatish Thennavan, Michael P. East, Joel S. Parker, Juan M. Cejalvo, Joseph P. Garay, Daniel P. Hollern, Xiaping He, Kevin R. Mott, Patricia Galván, Cheng Fan, Sara R. Selitsky, Alisha R. Coffey, David Marron, Fara Brasó-Maristany, Octavio Burgues, Joan Albanell, Federico Rojo, Ana Lluch, Eduardo Martinez de Dueñas, Jeffrey M. Rosen, Gary L. Johnson, Lisa A. Carey, Aleix Prat, Charles M. Perou
In mammalian heart, left ventricle (LV) rapidly becomes more dominant in size and function over right ventricle (RV) after birth. The molecular regulators responsible for this chamber specific differential growth are largely unknown. We found the cardiomyocytes in neonatal mouse RV had lower proliferation, more apoptosis and smaller sizes comparing to the LV. Such chamber specific growth pattern was associated with a selective activation of p38 MAPK activity in the RV and simultaneous inactivation in the LV. Cardiomyocyte-specific deletion of both mapk14 and mapk11 genes in mice results in loss of p38 MAP kinase expression and activity in the neonatal heart. Inactivation of p38 activity led to marked increase in myocytes proliferation and hypertrophy but diminished myocyte apoptosis, specifically in the RV. Consequently, the p38 inactivated hearts showed RV specific enlargement postnatally, progressing to pulmonary hypertension and right heart failure at adult stage. Chamber-specific p38 activity was associated with differential expression of dual-specific phosphatases (DUSPs) in neonatal hearts, including Dusp26. Unbiased transcriptome analysis revealed IRE1/XBP mediated gene regulation contributed to p38 MAPK dependent regulation of neonatal myocyte proliferation and binucleation. These findings establish an obligatory role of DUSP-p38-IRE1 signaling in myocytes for chamber specific growth in postnatal heart.
Tomohiro Yokota, Jin LI, Jijun Huang, Zhaojun Xiong, Qing Zhang, Tracey W. Chan, Yichen Ding, Christoph D. Rau, Kevin Sung, Shuxun Ren, Rajan P. Kulkarni, Tzung Hsiai, Xinshu Xiao, Marlin Touma, Susumu Minamisawa, Yibin Wang
While the advent of combination antiretroviral therapy (ART) has significantly improved survival, tuberculosis (TB) remains the leading cause of death in the HIV-infected population. We employed Mtb/Simian Immunodeficiency Virus (SIV) co-infected macaques to model Mtb/HIV co-infection and study the impact of ART on TB reactivation due to HIV-infection. While ART significantly reduced viral loads and increased CD4+ T cell counts in whole blood and BAL samples, it did not reduce the relative risk of SIV- induced TB reactivation in ART treated macaques in the early phase of treatment. CD4+ T cells were poorly restored specifically in the lung interstitium, despite their significant restoration in the alveolar compartment of the lung as well as in the periphery. IDO1 induction on myeloid cells in the iBALT likely contributed to dysregulated T cell homing and impaired lung immunity. Thus, while ART is indispensable in controlling viral replication, CD4+ T cells restoration and preventing opportunistic infection, it appears inadequate in reversing clinical signs of TB reactivation during the relatively short duration of ART and follow-up during this study. This warrants modeling concurrent treatment of TB and HIV to potentially reduce the risk of reactivation of TB due to HIV. The current and future studies like this have the potential to inform treatment strategies in patients with Mtb/HIV co-infection.
Shashank R. Ganatra, Allison N. Bucsan, Xavier Alvarez, Shyamesh Kumar, Ayan Chatterjee, Melanie Quezada, Abigail I. Fish, Dhiraj K. Singh, Bindu Singh, Riti Sharan, Tae-Hyung Lee, Uma Shanmugasundaram, Vijayakumar Velu, Shabaana A. Khader, Smriti Mehra, Jyothi Rengarajan, Deepak Kaushal
The amyloid hypothesis posits that the amyloid-beta (Aβ) protein precedes and requires microtubule-associated protein tau in a sort of trigger-bullet mechanism leading to Alzheimer’s disease (AD) pathology. This sequence of events has become dogmatic in the AD field and is used to explain clinical trial failures due to a late start of the intervention when Aβ already activated tau. Here, using a multidisplinary approach combining molecular biological, biochemical, histopathological, electrophysiological and behavioral methods we demonstrated that tau suppression did not protect against Aβ-induced damage of long-term synaptic plasticity and memory, as well as amyloid deposition. Tau suppression could even unravel a defect in basal synaptic transmission in a mouse model of amyloid deposition. Similarly, tau suppression did not protect against exogenous oligomeric tau induced impairment of long-term synaptic plasticity and memory. The protective effect of tau suppression was, in turn, confined to short-term plasticity and memory. Taken together, our data suggest that therapies downstream of Aβ and tau together are more suitable to combat AD than therapies against one or the other alone.
Daniela Puzzo, Elentina K. Argyrousi, Agnieszka Staniszewski, Hong Zhang, Elisa Calcagno, Elisa Zuccarello, Erica Acquarone, Mauro Fà, Domenica Donatella Li Puma, Claudio Grassi, Luciano D'Adamio, Nicholas M. Kanaan, Paul E. Fraser, Ottavio Arancio
The biology of harlequin ichthyosis (HI), a devastating skin disorder, caused by loss of function mutations in the gene ABCA12, is poorly understood and to date no satisfactory treatment has been developed. We sought to investigate pathomechanisms of HI which could lead to the identification of new treatments to improve patients’ quality of life. In this study, RNA-Seq and functional assays were performed to define the effects of loss of ABCA12, using HI patient skin samples and an engineered CRISPR-Cas9 ABCA12 KO cell line. The HI living skin equivalent (3D model) recapitulated the HI skin phenotype. The cytokines IL-36α and IL-36γ were upregulated in HI skin whereas the innate immune inhibitor, IL-37, was strongly downregulated. We also identified STAT1 and its downstream target inducible nitric oxide synthase (NOS2) to be upregulated in the in vitro HI 3D model and HI patient skin samples. Inhibition of NOS2 using the inhibitor, 1400W, or the JAK inhibitor, tofacitinib, dramatically improved the in vitro HI phenotype by restoring the lipid barrier in the HI 3D model. Our study has identified dysregulated pathways in HI skin that are feasible therapeutic targets.
Florence Enjalbert, Priya Dewan, Matthew P. Caley, Eleri M. Jones, Mary A. Morse, David P. Kelsell, Anton J. Enright, Edel A. O'Toole
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