Bruton tyrosine kinase (BTK) is present in a wide variety of cells and may thus have important non-B cell functions. Here we explored the function of this kinase in macrophages with studies of its regulation of the NLRP3 inflammasome. We found that bone marrow-derived macrophages (BMDMs) from BTK-deficient mice or monocytes from X-linked agammaglobulinemia patients exhibit increased NLRP3 inflammasome activity; this was also the case with BMDMs exposed to low doses of BTK inhibitor such as ibrutinib and monocytes from chronic lymphocytic leukemia patients being treated with ibrutinib. In mechanistic studies, we found that BTK binds to NLRP3 during the priming phase of inflammasome activation and in doing so inhibits LPS/nigericin-induced assembly of the NLRP3 inflammasome during the activation phase of inflammasome activation. This inhibitory effect was caused by BTK inhibition of PP2A-mediated dephosphorylation of Ser5 in the pyrin domain of NLRP3. Finally, we showed that BTK-deficient mice are subject to severe experimental colitis and such colitis is normalized by administration of anti-IL-β or an inhibitor of IL-1β signaling, anakinra. Together, these studies strongly suggest that BTK functions as a physiologic inhibitor of NLRP3 inflammasome activation; they thereby explain the fact that XLA patients are prone to develop Crohn’s disease.
Liming Mao, Atsushi Kitani, Eitaro Hiejima, Kim Montgomery-Recht, Wenchang Zhou, Ivan Fuss, Adrian Wiestner, Warren Strober
Cancer–related anemia is present in over 60% of newly diagnosed cancer patients and is associated with substantial morbidity and high medical costs. Drugs that enhance erythropoiesis are urgently required to decrease transfusion rates and improve quality of life. Clinical studies have observed an unexpected improvement in hemoglobin and red blood cell (RBC) transfusion-independence in AML patients treated with the isocitrate dehydrogenase 2 (IDH2) mutant-specific inhibitor, enasidenib, leading to improved quality of life without a reduction in AML disease burden. Here, we demonstrate that enasidenib enhanced human erythroid differentiation of hematopoietic progenitors. The phenomenon was not observed with other IDH1/2 inhibitors and occurred in IDH2-deficient CRIPSR-engineered progenitors independently of D-2-hydroxyglutarate. The effect of enasidenib on hematopoietic progenitors was mediated by protoporphyrin accumulation, driving heme production and erythroid differentiation in committed CD71+ progenitors rather than hematopoietic stem cells. Our results position enasidenib as a promising therapeutic agent for improvement of anemia and provide the basis for a clinical trial using enasidenib to decrease transfusion dependence in a wide array of clinical contexts.
Ritika Dutta, Tian Yi Zhang, Thomas Köhnke, Daniel Thomas, Miles Linde, Eric Gars, Melissa Stafford, Satinder Kaur, Yusuke Nakauchi, Raymond Yin, Armon Azizi, Anupama Narla, Ravindra Majeti
BACKGROUND. Residual C-peptide is detected in many people for years following the diagnosis of type 1 diabetes; however, the physiologic significance of low levels of detectable C-peptide is not known. METHODS. We studied sixty-three adults with type 1 diabetes classified by peak mixed-meal tolerance test (MMTT) C-peptide as negative (<0.007; n =15), low (0.017–0.200; n =16), intermediate (>0.200–0.400; n =15), or high (>0.400 pmol/mL; n =17). We compared the groups’ glycemia from continuous glucose monitoring (CGM), β-cell secretory responses from a glucose-potentiated arginine (GPA) test, insulin sensitivity from a hyperinsulinemia euglycemic (EU) clamp, and glucose counterregulatory responses from a subsequent hypoglycemic (HYPO) clamp. RESULTS. Low and intermediate MMTT C-peptide groups did not exhibit β-cell secretory responses to hyperglycemia, whereas the high C-peptide group showed increases in both C-peptide and proinsulin (P ≤0.01). All groups with detectable MMTT C-peptide demonstrated acute C-peptide and proinsulin responses to arginine that were positively correlated with peak MMTT C-peptide (P <0.0001 for both analytes). During the EU-HYPO clamp, C-peptide levels were proportionately suppressed in the low, intermediate, and high C-peptide compared to the negative group (P ≤0.0001), whereas glucagon increased from EU to HYPO only in the high C-peptide group compared to negative (P =0.01). CGM demonstrated lower mean glucose and more time-in-range for the high C-peptide group. CONCLUSION. These results indicate that in adults with type 1 diabetes, β-cell responsiveness to hyperglycemia and α-cell responsiveness to hypoglycemia are only observed at high levels of residual C-peptide that likely contribute to glycemic control.
Michael R. Rickels, Carmella Evans-Molina, Henry T. Bahnson, Alyssa Ylescupidez, Kristen J. Nadeau, Wei Hao, Mark A. Clements, Jennifer L. Sherr, Richard E. Pratley, Tamara S. Hannon, Viral N. Shah, Kellee M. Miller, Carla J. Greenbaum
Hematopoietic stem cell (HSC) attrition is considered the key event underlying progressive bone marrow failure (BMF) in Fanconi anemia (FA), the most frequent inherited BMF disorder in humans. However, despite major advances, how the cellular, biochemical and molecular alterations reported in FA lead to HSC exhaustion remains poorly understood. Here, we demonstrated in human and mouse cells that loss-of-function of FANCA or FANCC, products of two genes affecting more than 80% of FA patients worldwide, is associated with constitutive expression of the transcription factor Microphthalmia (MiTF) through the cooperative, unscheduled activation of several stress signaling pathways, including the SMAD2/3, p38MAPK, NF-kB and AKT cascades. We validated the unrestrained Mitf expression downstream of p38 in Fanca-/- mice, which display hallmarks of hematopoietic stress, including loss of HSC quiescence, DNA damage accumulation in HSCs and reduced HSC repopulation capacity. Importantly, we demonstrated that shRNA-mediated downregulation of Mitf expression or inhibition of p38 signaling rescued HSC quiescence and prevented DNA damage accumulation. Our data support the hypothesis that HSC attrition in FA is the consequence of defects in the DNA damage response combined with chronic activation of otherwise transiently activated signaling pathways, which jointly prevent the recovery of HSC quiescence.
Alessia Oppezzo, Julie Bourseguin, Emilie Renaud, Patrycja Pawlikowska, Filippo Rosselli
Oncogenic KRAS is a major driver in lung adenocarcinoma (LUAD) that has yet to be therapeutically conquered. Here we report that the SLC7A11/glutathione axis displays metabolic synthetic lethality with oncogenic KRAS. Through metabolomics approaches, we found that mutationally activated KRAS strikingly increased the intracellular cystine level and glutathione biosynthesis. SLC7A11, a cystine/glutamate antiporter conferring specificity for cystine uptake, was overexpressed in patients with KRAS-mutant LUAD and showed positive association with tumor progression. Furthermore, SLC7A11 inhibition either by genetic depletion or pharmacological inhibition by sulfasalazine resulted in selective killing across a panel of KRAS-mutant cancer cells in vitro and tumor growth inhibition in vivo, suggesting the functionality and specificity of SLC7A11 as a therapeutic target. Importantly, we further identified a potent SLC7A11 inhibitor, HG106 that markedly decreased cystine uptake and intracellular glutathione biosynthesis. Furthermore, HG106 exhibited selective cytotoxicity towards KRAS-mutant cells by increasing oxidative stress- and endoplasmic reticulum stress-mediated cell apoptosis. Of note, treatment of KRAS-mutant LUAD with HG106 in several lung cancer preclinical mouse models led to marked tumor suppression and prolonged mouse survival. Overall, our findings reveal that KRAS-mutant LUAD cells are vulnerable to SLC7A11 inhibition, providing promising therapeutic approaches to the treatment of this currently incurable disease.
Kewen Hu, Kun Li, Jing Lv, Jie Feng, Jing Chen, Haigang Wu, Feixiong Cheng, Wenhao Jiang, Jieqiong Wang, Haixiang Pei, Paul J. Chiao, Zhenyu Cai, Yihua Chen, Mingyao Liu, Xiufeng Pang
Staphylococcus aureus remains a leading cause of human infection. These infections frequently recur when the skin is a primary site of infection, especially in infants and children. In contrast, invasive staphylococcal disease is less commonly associated with reinfection, suggesting that tissue-specific mechanisms govern the development of immunity. Knowledge of how S. aureus manipulates protective immunity has been hampered by a lack of antigen-specific models to interrogate the T cell response. Utilizing a chicken egg ovalbumin (OVA)-expressing S. aureus strain to analyze OVA-specific T cell responses, we demonstrated that primary skin infection is associated with impaired development of T cell memory. Conversely, invasive infection induced antigen-specific memory and protected against reinfection. This defect in adaptive immunity following skin infection was associated with a loss of dendritic cells, attributable to S. aureus α-toxin (Hla) expression. Genetic and immunization-based approaches to protect against Hla during skin infection restored the T cell response. Within the human population, exposure to α-toxin through skin infection may modulate the establishment of T cell-mediated immunity, adversely impacting long-term protection. These studies prompt consideration that vaccination targeting S. aureus may be most effective if delivered prior to initial contact with the organism.
Brandon Lee, Reuben Olaniyi, Jakub Kwiecinski, Juliane Bubeck Wardenburg
Deficits in social interaction (SI) are a core symptom of Autism Spectrum Disorders (ASD), however treatments for social deficits are notably lacking. Elucidating brain circuits and neuromodulatory signaling systems that regulate sociability could facilitate a deeper understanding of ASD pathophysiology and reveal novel treatments for ASD. Here we found that in vivo optogenetic activation of the basolateral amygdala-nucleus accumbens (BLA-NAc) glutamatergic circuit reduced SI and increased social avoidance in mice. Furthermore, we found that 2-arachidonoylglycerol (2-AG) endocannabinoid (eCB) signaling reduced BLA-NAc glutamatergic activity, and that pharmacological 2-AG augmentation via administration of JZL184 blocked SI deficits associated with in vivo BLA-NAc stimulation. Additionally, optogenetic inhibition of the BLA-NAc circuit significantly increased SI in the Shank3B-/-, an ASD model with substantial SI impairment, without affecting SI in wild-type mice. Finally, we demonstrated that JZL184 delivered systemically or directly to the NAc also normalized SI deficits in Shank3B-/-mice, while ex vivo JZL184 application corrected aberrant NAc excitatory and inhibitory neurotransmission and reduced BLA-NAc-elicited feedforward inhibition of NAc neurons in Shank3B-/- mice. These data reveal circuit-level and neuromodulatory mechanisms regulating social function relevant to ASD and suggest 2-AG augmentation could reduce social deficits via modulation of excitatory and inhibitory neurotransmission in the NAc.
Oakleigh M. Folkes, Rita Báldi, Veronika Kondev, David J. Marcus, Nolan D. Hartley, Brandon D. Turner, Jade K. Ayers, Jordan J. Baechle, Maya P. Misra, Megan Altemus, Carrie A. Grueter, Brad A. Grueter, Sachin Patel
BACKGROUND. Undifferentiated systemic autoinflammatory diseases (USAID) present diagnostic and therapeutic challenges. Chronic interferon (IFN) signaling and cytokine dysregulation may identify diseases with available targeted treatments. METHODS. Sixty-six consecutively-referred USAID patients underwent standardized evaluation of Type-I IFN-response-gene-signature (IRG-S); cytokine profiling, and genetic evaluation by next-generation sequencing. RESULTS. Thirty-six USAID patients (55%) had elevated IRG-S. Neutrophilic panniculitis (40% vs 0%), basal ganglia calcifications (46% vs 0%), interstitial lung disease (47% vs 5%), and myositis (60% vs 10%) were more prevalent in patients with elevated IRG-S. Moderate IRG-S elevation and highly-elevated serum IL-18 distinguished eight patients with pulmonary alveolar proteinosis (PAP) and recurrent macrophage activation syndrome (MAS). Among patients with panniculitis and progressive cytopenias, two patients were compound heterozygous for novel LRBA mutations, four patients harbored novel splice variants in IKBKG/NEMO, and six patients had de novo frameshift mutations in SAMD9L. Of additional 12 patients with elevated IRG-S and CANDLE-, SAVI- or Aicardi-Goutières-Syndrome (AGS)-like phenotypes, five patients carried mutations in either SAMHD1, TREX1, PSMB8 or PSMG2. Two patients had anti-MDA5 autoantibody-positive juvenile dermatomyositis, and seven could not be classified. Patients with LRBA, IKBKG/NEMO and SAMD9L mutations showed a pattern of IRG elevation that suggests prominent NF-κB activation different from the canonical interferonopathies CANDLE, SAVI and AGS. CONCLUSIONS. In patients with elevated IRG-S, we identified characteristic clinical features and 3 additional autoinflammatory diseases: IL-18-mediated PAP and recurrent MAS (IL-18PAP-MAS), NEMO∆5-associated autoinflammatory syndrome (NEMO-NDAS), and SAMD9L-associated autoinflammatory disease (SAMD9L-SAAD). The IRG-S expands the diagnostic armamentarium in evaluating USAIDs and points to different pathways regulating IRG expression.
Adriana A. de Jesus, Yanfeng Hou, Stephen Brooks, Louise Malle, Angelique Biancotto, Yan Huang, Katherine R. Calvo, Bernadette Marrero, Susan Moir, Andrew J. Oler, Zuoming Deng, Gina A. Montealegre Sanchez, Amina Ahmed, Eric Allenspach, Bita Arabshahi, Edward Behrens, Susanne Benseler, Liliana Bezrodnik, Sharon Bout-Tabaku, AnneMarie C. Brescia, Diane Brown, Jon M. Burnham, María Soledad Caldirola, Ruy Carrasco, Alice Y. Chan, Rolando Cimaz, Paul Dancey, Jason Dare, Marietta DeGuzman, Victoria Dimitriades, Ian Ferguson, Polly Ferguson, Laura Finn, Marco Gattorno, Alexei A. Grom, Eric P. Hanson, Philip J. Hashkes, Christian M. Hedrich, Ronit Herzog, Gerd Horneff, Rita Jerath, Elizabeth Kessler, Hanna Kim, Daniel J. Kingsbury, Ronald M. Laxer, Pui Y. Lee, Min Ae Lee-Kirsch, Laura Lewandowski, Suzanne Li, Vibke Lilleby, Vafa Mammadova, Lakshmi N. Moorthy, Gulnara Nasrullayeva, Kathleen M. O’Neil, Karen Onel, Seza Ozen, Nancy Pan, Pascal Pillet, Daniela G.P. Piotto, Marilynn G. Punaro, Andreas Reiff, Adam Reinhardt, Lisa G. Rider, Rafael Rivas-Chacon, Tova Ronis, Angela Rösen-Wolff, Johannes Roth, Natasha Mckerran Ruth, Marite Rygg, Heinrike Schmeling, Grant Schulert, Christiaan Scott, Gisela Seminario, Andrew Shulman, Vidya Sivaraman, Mary Beth Son, Yuriy Stepanovskyy, Elizabeth Stringer, Sara Taber, Maria Teresa Terreri, Cynthia Tifft, Troy Torgerson, Laura Tosi, Annet Van Royen-Kerkhof, Theresa Wampler Muskardin, Scott W. Canna, Raphaela Goldbach-Mansky
Few therapies are currently available for patients with KRAS-driven cancers, highlighting the need to identify new molecular targets that modulate central downstream effector pathways. Here we found the miRNA cluster mir181ab1 as a key modulator of KRAS-driven oncogenesis. Ablation of Mir181ab1 in genetically-engineered mouse models of Kras-driven lung and pancreatic cancer was deleterious to tumor initiation and progression. Expression of both resident miRNAs in the Mir181ab1 cluster, miR181a1 and miR181b1, was necessary to rescue the Mir181ab1-loss phenotype underscoring their non-redundant role. In human cancer cells, depletion of miR181ab1 impaired proliferation and 3D growth, whereas overexpression provided a proliferative advantage. Lastly, we unveiled miR181ab1-regulated genes responsible for this phenotype. These studies identified what we believe to be a previously unknown role for miR181ab1 as a potential therapeutic target in two highly aggressive and difficult to treat KRAS-mutated cancers.
Karmele Valencia, Oihane Erice, Kaja Kostyrko, Simone Hausmann, Elizabeth Guruceaga, Anuradha Thathireddy, Natasha M. Flores, Leanne C. Sayles, Alex G. Lee, Rita Fragoso, Tian-Qiang Sun, Adrian Vallejo, Marta Roman, Rodrigo Entrialgo-Cadierno, Itziar Migueliz, Nerea Razquin, Puri Fortes, Fernando Lecanda, Jun Lu, Mariano Ponz-Sarvise, Chang-Zheng Chen, Pawel K. Mazur, E. Alejandro Sweet-Cordero, Silvestre Vicent
Genomics of primary prostate cancer differs from that of metastatic castration-resistant prostate cancer (mCRPC). We studied genomic aberrations in primary prostate cancer biopsies from patients who developed mCRPC, also studying matching, same patient, diagnostic and mCRPC biopsies following treatment. We profiled 470 treatment-naïve, prostate cancer diagnostic biopsies and for 61 cases, mCRPC biopsies using targeted and low-pass whole genome sequencing (n = 52). Descriptive statistics were used to summarize mutation and copy number profile. Prevalence was compared using Fisher's exact test. Survival correlations were studied using log-rank test. TP53 (27%) and PTEN (12%) and DDR gene defects (BRCA2 7%; CDK12 5%; ATM 4%) were commonly detected. TP53, BRCA2, and CDK12 mutations were significantly commoner than described in the TCGA cohort. Patients with RB1 loss in the primary tumour had a worse prognosis. Among 61 men with matched hormone-naïve and mCRPC biopsies, differences were identified in AR, TP53, RB1, and PI3K/AKT mutational status between same-patient samples. In conclusion, the genomics of diagnostic prostatic biopsies acquired from men who develop mCRPC differs to that of the primary prostatic cancers. RB1/TP53/AR aberrations are enriched in later stages, but the prevalence of DDR defects in diagnostic samples is similar to mCRPC.
Joaquin Mateo, George Seed, Claudia Bertan, Pasquale Rescigno, David Dolling, Ines Figueiredo, Susana Miranda, Daniel Nava Rodrigues, Bora Gurel, Matthew Clarke, Mark Atkin, Rob Chandler, Carlo Messina, Semini Sumanasuriya, Diletta Bianchini, Maialen Barrero, Antonella Petremolo, Zafeiris Zafeiriou, Mariane Sousa Fontes, Raquel Perez-Lopez, Nina Tunariu, Ben A. Fulton, Robert Jones, Ursula B. McGovern, Christy Ralph, Mohini Varughese, Omi Parikh, Suneil Jain, Tony Elliott, Shahneen Sandhu, Nuria Porta, Emma Hall, Wei Yuan, Suzanne Carreira, Johann S. de Bono
Recent findings have shown that inhibitors targeting BET (bromodomain and extraterminal domain) proteins, such as the small molecule JQ1, are potent growth inhibitors of many cancers and hold promise for cancer therapy. However, some reports also have revealed that JQ1 can activate additional oncogenic pathways and may affect EMT (epithelial mesenchymal transition). Therefore, it is important to address the potential unexpected effect of JQ1 treatment, such as cell invasion and metastasis. Here, we showed that in prostate cancer, JQ1 inhibited cancer cell growth but promoted invasion and metastasis in a BET protein independent manner. Multiple invasion pathways including EMT, BMP (bone morphogenetic protein) signaling, chemokine signaling and focal adhesion pathway were activated by JQ1 to promote invasion. Notably, JQ1 induced upregulation of invasion genes through inhibition of FOXA1, an invasion suppressor in prostate cancer. JQ1 directly interacted with FOXA1, inactivated FOXA1 binding to its interacting repressors, TLE3, HDAC7, and NFIC, thus blocking FOXA1 repressive function and activating the invasion genes. Our finding indicates that JQ1 has an unexpected effect of promoting invasion in prostate cancer. Thus, the ill effect of JQ1 or its derived therapeutic agents cannot be ignored during cancer treatment, especially in FOXA1 related cancers.
Leiming Wang, Mafei Xu, Chung-Yang Kao, Sophia Y. Tsai, Ming-Jer Tsai
The incidence of human papillomavirus (HPV)+ head and neck squamous cell carcinoma (HNSCC) has surpassed that of cervical cancer and is projected to increase rapidly until 2060. The co-evolution of HPV with transforming epithelial cells leads to the shutdown of host immune detection. Targeting proximal viral nucleic acid-sensing machinery is an evolutionarily conserved strategy among viruses to enable immune evasion. However, E7 from the dominant HPV subtype-16 in HNSCC shares low homology with HPV18 E7, which was shown to inhibit the STING-DNA-sensing pathway. The mechanisms by which HPV16 suppresses STING remain unknown. Recently, we characterized the role of the STING-type-I interferon (IFN-I) pathway in maintaining immunogenicity of HNSCC in mouse models. Here we extended those findings into clinical domain utilizing tissue microarrays and machine-learning-enhanced profiling of STING signatures with immune subsets. We additionally showed that HPV16 E7 employs distinct mechanisms than HPV18 E7 to antagonize the STING pathway. We identified NLRX1 as a critical intermediary partner to facilitate HPV16 E7-potentiated STING turnover. The depletion of NLRX1 resulted in significantly improved IFN-I-dependent T-cell infiltration profiles and tumor control. Overall, we discovered a unique HPV16 viral strategy to thwart host innate immune detection that can be further exploited to restore cancer immunogenicity.
Xiaobo Luo, Christopher R. Donnelly, Wang Gong, Blake R. Heath, Yuning Hao, Lorenza A. Donnelly, Toktam Moghbeli, Yee Sun Tan, Xin Lin, Emily Bellile, Benjamin A. Kansy, Thomas E. Carey, J. Chad Brenner, Lei Cheng, Peter J. Polverini, Meredith A. Morgan, Haitao Wen, Mark E. Prince, Robert L. Ferris, Yuying Xie, Simon Young, Gregory T. Wolf, Qianming Chen, Yu L. Lei
The protein-protein interaction between menin and Mixed Lineage Leukemia 1 (MLL1) plays a critical role in acute leukemias with translocations of the MLL1 gene or with mutations in the Nucleophosmin 1 (NPM1) gene. As a step toward clinical translation of menin-MLL1 inhibitors, we report development of MI-3454, a highly potent and orally bioavailable inhibitor of the menin-MLL1 interaction. MI-3454 profoundly inhibited proliferation and induced differentiation in acute leukemia cells and primary patient samples with MLL1 translocations or NPM1 mutations. When applied as a single agent, MI-3454 induced complete remission or regression of leukemia in mouse models of MLL1-rearranged or NPM1-mutated leukemia, including patient-derived xenograft models, through downregulation of key genes involved in leukemogenesis. We also identified MEIS1 as a potential pharmacodynamic biomarker of treatment response with MI-3454 in leukemia, and demonstrated that this compound is well tolerated and did not impair normal hematopoiesis in mice. Overall, this study demonstrates for the first time profound activity of the menin-MLL1 inhibitor as a single agent in clinically relevant PDX models of leukemia. These data provide a strong rationale for clinical translation of MI-3454 or its analogs for leukemia patients with MLL1-rearrangements or NPM1 mutations
Szymon Klossowski, Hongzhi Miao, Katarzyna Kempinska, Tao Wu, Trupta Purohit, EunGi Kim, Brian M. Linhares, Dong Chen, Gloria Jih, Eric Perkey, Huang Huang, Miao He, Bo Wen, Yi Wang, Ke Yu, Stanley Chun-Wei Lee, Gwenn Danet-Desnoyers, Winifred Trotman, Malathi Kandarpa, Anitria Cotton, Omar Abdel-Wahab, Hongwei Lei, Yali Dou, Monica Guzman, Luke Peterson, Tanja A. Gruber, Sarah M. Choi, Duxin Sun, Pingda Ren, Lian-Sheng Li, Yi Liu, Francis J. Burrows, Ivan Maillard, Tomasz Cierpicki, Jolanta Grembecka
Acute graft-versus-host disease (GVHD) can affect the central nervous system (CNS). The role of microglia in CNS-GVHD remains undefined. In agreement with microglia activation, we found that profound morphological changes, MHC-II- and CD80-upregulation occurred upon GVHD induction. RNA-sequencing-based analysis of purified microglial obtained from mice with CNS-GVHD revealed TNF upregulation. Selective TNF gene deletion in microglia of Cx3cr1creER:Tnffl/-mice reduced MHC-II-expression, decreased CNS T-cell infiltrates and VCAM-1+ endothelial cells. GVHD increased microglia TGF-β-activated kinase-1 (TAK1) activation and NF-κB/p38-MAPK-signaling. Selective Tak1-deletion in microglia using Cx3cr1creER:Tak1fl/fl-mice resulted in reduced TNF-production, microglial MHC-II, and improved neurocognitive-activity. Pharmacological TAK1-inhibition reduced TNF-production and MHC-II-expression by microglia, Th1 and Th17 T-cell infiltrates, VCAM-1+ endothelial cells and improved neurocognitive activity, without blocking graft-versus-leukemia effects. Consistent with these findings in mice, we observed increased activation and TNF-production of microglia in the CNS of GVHD-patients. In summary, we prove a role for microglia in CNS-GVHD, identify the TAK1/TNF/MHC-II axis as mediator of CNS-GVHD and provide a novel TAK1 inhibitor-based approach against GVHD-induced neurotoxicity.
Nimitha R. Mathew, Janaki M. Vinnakota, Petya Apostolova, Daniel Erny, Shaima’a Hamarsheh, Geoffroy Andrieux, Jung-Seok Kim, Kathrin Hanke, Tobias Goldmann, Louise Chappell-Maor, Nadia El-Khawanky, Gabriele Ihorst, Dominik Schmidt, Justus Duyster, Jürgen Finke, Thomas Blank, Melanie Boerries, Bruce R. Blazar, Steffen Jung, Marco Prinz, Robert Zeiser
Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex disease with no known cause or mechanism. There is an increasing appreciation for the role of immune and metabolic dysfunction in the disease. ME/CFS has historically presented in outbreaks, often has a flu-like onset, and results in inflammatory symptoms. Patients suffer from severe fatigue and post-exertional malaise. There is little known about the metabolism of specific immune cells in ME/CFS patients. To investigate immune metabolism in ME/CFS, we isolated CD4+ and CD8+ T cells from 53 ME/CFS patients and 45 healthy controls. We analyzed glycolysis and mitochondrial respiration in resting and activated T cells, along with markers related to cellular metabolism, and plasma cytokines. We found that ME/CFS CD8+ T cells have reduced mitochondrial membrane potential compared to healthy controls. Both CD4+ and CD8+ T cells from ME/CFS patients had reduced glycolysis at rest, while CD8+ T cells also had reduced glycolysis following activation. ME/CFS patients had significant correlations between measures of T cell metabolism and plasma cytokine abundance that differed from healthy control subjects. Our data indicate that patients have impaired T cell metabolism consistent with ongoing immune alterations in ME/CFS that may illuminate the mechanism behind this disease.
Alexandra H. Mandarano, Jessica Maya, Ludovic Giloteaux, Daniel L. Peterson, Marco Maynard, C. Gunnar Gottschalk, Maureen R. Hanson
Cantu Syndrome (CS) is a complex disorder caused by gain-of-function (GoF) mutations in ABCC9 and KCNJ8, which encode the SUR2 and Kir6.1 subunits, respectively, of vascular smooth muscle (VSM) KATP channels. CS includes dilated vasculature, marked cardiac hypertrophy, and other cardiovascular abnormalities. There is currently no targeted therapy, and it is unknown whether cardiovascular features can be reversed once manifest. Using combined transgenic and pharmacological approaches in a knock-in mouse model of CS, we have shown that reversal of vascular and cardiac phenotypes can be achieved (1) by genetic downregulation of KATP channel activity specifically in VSM, and (2) by chronic administration of the clinically-used KATP channel inhibitor, glibenclamide. These findings demonstrate (i) that VSM KATP channel GoF underlies CS cardiac enlargement, (ii) reversibility of CS-associated abnormalities and (iii) evidence of in vivo efficacy of glibenclamide as a therapeutic agent in CS.
Conor McClenaghan, Yan Huang, Zihan Yan, Theresa Harter, Carmen M. Halabi, Rod Chalk, Attila Kovacs, Gijs van Haaften, Maria S. Remedi, Colin G. Nichols
BACKGROUND. Cerebral malaria (CM) accounts for nearly 400,000 deaths annually in African children. Current dogma suggests that CM results from infected RBC (iRBC) sequestration in the brain microvasculature and resulting sequelae. Therapies targeting these events have been unsuccessful; findings in experimental models suggest that CD8+ T cells drive disease pathogenesis. However, these data have largely been ignored because corroborating evidence in humans is lacking. This work fills a critical gap in our understanding of CM pathogenesis that is impeding development of therapeutics. METHODS. Using multiplex immunohistochemistry, we characterized cerebrovascular immune cells in brain sections from 34 children who died from CM or other causes. Children were grouped by clinical diagnosis (CM+ or –), iRBC sequestration (Seqhi, lo, or 0) and HIV status (HIV+ or –). RESULTS. We identified effector CD3+CD8+ T cells engaged on the cerebrovasculature in 69% of CM+ HIV– children. The number of intravascular CD3+CD8+ T cells was influenced by CM status (CM+ vs –, P = 0.004) and sequestration level (Seqhi > lo, P = 0.010). HIV co-infection significantly increased T cell numbers and shifted cells from an intravascular (P = 0.004) to perivascular (P < 0.0001) distribution. CONCLUSION. Within the studied cohort, CM is associated with cerebrovascular engagement of CD3+CD8+ T cells, which is exacerbated by HIV coinfection. Thus, CD3+CD8+ T cells are highly promising targets for CM adjunctive therapy, opening new avenues for the treatment of this deadly disease. FUNDING. This research was supported by the Intramural Research Program of the National Institutes of Health.
Brittany A. Riggle, Monica Manglani, Dragan Maric, Kory R. Johnson, Myoung-Hwa Lee, Osorio Lopes Abath Neto, Terrie E. Taylor, Karl B. Seydel, Avindra Nath, Louis H. Miller, Dorian B. McGavern, Susan K. Pierce
Background: DICER1 is the only miRNA biogenesis component associated with an inherited tumor syndrome, featuring multinodular goiter (MNG) and rare pediatric-onset lesions. Other susceptibility genes for familial forms of MNG likely exist. Methods: Whole exome sequencing of a kindred with early-onset MNG and schwannomatosis was followed by investigation of germline pathogenic variants that fully segregated with the disease. Genome wide analyses were performed on 13 tissue samples from familial and non-familial DGCR8-E518K positive tumors, including MNG, schwannomas, papillary thyroid cancers (PTC) and Wilms Tumors. MiRNA profiles of four tissue types were compared, and sequencing of miRNA, pre-miRNA and mRNA was performed in a subset of 9 schwannomas, four of which harbor DGCR8-E518K. Results: We identified c.1552G>A;p.E518K in DGCR8, a microprocessor located in 22q, in the kindred. The variant identified is a somatic hotspot in Wilms Tumors and has been identified in two PTCs. Copy number loss of chromosome 22q, leading to loss of heterozygosity at the DGCR8 locus, was found in all 13 samples harboring c.1552G>A;p.E518K. miRNA profiling of PTC, MNG, schwannomas and Wilms Tumors revealed a common profile among E518K hemizygous tumors. In vitro cleavage demonstrated improper processing of pre-miRNA by DGCR8-E518K. MicroRNA and RNA profiling show that this variant disrupts precursor microRNA production, impacting populations of canonical microRNAs and mirtrons. Conclusions: We identified DGCR8 as the cause of an unreported autosomal dominant mendelian tumor susceptibility syndrome: familial multinodular goiter with schwannomatosis. Funded by CIHR, Compute Canada, Alex’s Lemonade Stand Foundation, and the Mia Neri Foundation for Childhood Cancer.
Barbara Rivera, Javad Nadaf, Somayyeh Fahiminiya, Maria Apellaniz-Ruiz, Avi Saskin, Anne-Sophie Chong, Sahil Sharma, Rabea Wagener, Timothée Revil, Vincenzo Condello, Zineb Harra, Nancy Hamel, Nelly Sabbaghian, Karl Muchantef, Christian Thomas, Leanne de Kock, Marie-Noëlle Hébert-Blouin, Angelia V. Bassenden, Hannah Rabenstein, Ozgur Mete, Ralf Paschke, Marc P. Pusztaszeri, Werner Paulus, Albert Berghuis, Jiannis Ragoussis, Yuri E. Nikiforov, Reiner Siebert, Steffen Albrecht, Robert Turcotte, Martin Hasselblatt, Marc R. Fabian, William D. Foulkes
Background. An increase in intrahepatic triglyceride (IHTG) is the hallmark feature of nonalcoholic fatty liver disease (NAFLD) and is decreased by weight loss. Hepatic de novo lipogenesis (DNL) contributes to steatosis in people with NAFLD. The physiological factors that stimulate hepatic DNL and the effect of weight loss on hepatic DNL are not clear.Methods. Hepatic DNL, 24-h integrated plasma insulin and glucose concentrations, and both liver and whole-body insulin sensitivity were determined in people who were lean (n = 14), obese with normal IHTG content (Obese, n = 26) and obese with NAFLD (Obese-NAFLD, n = 27). Hepatic DNL was assessed by using the deuterated water method corrected for the potential confounding contribution of adipose tissue DNL. Liver and whole-body insulin sensitivity were assessed by using the hyperinsulinemic-euglycemic clamp procedure in conjunction with glucose tracer infusion. Six subjects in the Obese-NAFLD group were also evaluated before and after 10% diet-induced weight loss.Results. The contribution of hepatic DNL to IHTG-palmitate was 11%, 19% and 38% in the Lean, Obese and Obese-NAFLD groups, respectively. Hepatic DNL was inversely correlated with hepatic and whole-body insulin sensitivity, but directly correlated with 24-h plasma glucose and insulin concentrations. Weight loss decreased IHTG content, in conjunction with a decrease in hepatic DNL and 24-h plasma glucose and insulin concentrations. Conclusions. These data suggest hepatic DNL is an important regulator of IHTG content, and that increases in circulating glucose and insulin stimulate hepatic DNL in people with NAFLD. Weight loss decreases IHTG content, at least in part, by decreasing hepatic DNL.
Gordon I. Smith, Mahalakshmi Shankaran, Mihoko Yoshino, George G. Schweitzer, Maria Chondronikola, Joseph W. Beals, Adewole L. Okunade, Bruce W. Patterson, Edna Nyangau, Tyler Field, Claude B. Sirlin, Saswata Talukdar, Marc K. Hellerstein, Samuel Klein
Epidermal growth factor receptor (EGFR) and MEK inhibitors (EGFR/MEKi) are beneficial for the treatment of solid cancers but are frequently associated with severe therapy-limiting acneiform skin toxicities. The underlying molecular mechanisms are poorly understood. Using gene expression profiling we identified IL-36γ and IL-8 as candidate drivers of EGFR/MEKi skin toxicity. We provide molecular and translational evidence that EGFR/MEKi in concert with the skin commensal bacterium Cutibacterium acnes act synergistically to induce IL-36γ in keratinocytes and subsequently IL-8, leading to cutaneous neutrophilia. IL-36γ expression was the combined result of C. acnes-induced NF-κB activation and EGFR/MEKi-mediated expression of the transcription factor Krüppel-like factor 4 (KLF4), due to the presence of both NF-κB- and KLF4-binding sites in the human IL-36γ gene promoter. EGFR/MEKi increased KLF4 expression by blockade of the EGFR-MEK-ERK pathway. These results provide an insight into understanding the pathological mechanism of the acneiform skin toxicities induced by EGFR/MEKi and identify IL-36γ and the transcription factor KLF4 as potential therapeutic targets.
Takashi K. Satoh, Mark Mellett, Barbara Meier-Schiesser, Gabriele Fenini, Atsushi Otsuka, Hans-Dietmar Beer, Tamara Rordorf, Julia-Tatjana Maul, Jürg Hafner, Alexander A. Navarini, Emmanuel Contassot, Lars E. French