The accrual of myeloid-derived suppressor cells (MDSCs) represents a major obstacle to effective immunotherapy in cancer patients, but the mechanisms underlying this process in the human setting remain elusive. Here, we describe a set of microRNAs (miR-146a, miR-155, miR-125b, miR-100, let-7e, miR-125a, miR-146b, miR-99b) that are associated with MDSCs and with resistance to treatment with immune checkpoint inhibitors in melanoma patients. The miRs were identified by transcriptional analyses as being responsible for the conversion of monocytes into MDSCs (CD14+HLA-DRneg cells) mediated by melanoma extracellular vesicles (EVs) and were shown to recreate MDSC features upon transfection. In melanoma patients, these miRs are increased in circulating CD14+ monocytes, plasma and tumor samples, where they correlate with the myeloid cell infiltrate. In plasma, their baseline level clusters with the clinical efficacy of CTLA-4 or PD-1 blockade. Hence, MDSC-related miRs represent an indicator of MDSC activity in cancer patients and a potential blood marker of a poor immunotherapy outcome.
Veronica Huber, Viviana Vallacchi, Viktor Fleming, Xiaoying Hu, Agata Cova, Matteo Dugo, Eriomina Shahaj, Roberta Sulsenti, Elisabetta Vergani, Paola Filipazzi, Angela De Laurentiis, Luca Lalli, Lorenza Di Guardo, Roberto Patuzzo, Barbara Vergani, Elena Casiraghi, Mara Cossa, Ambra Gualeni, Valentina Bollati, Flavio Arienti, Filippo De Braud, Luigi Mariani, Antonello Villa, Peter Altevogt, Viktor Umansky, Monica Rodolfo, Licia Rivoltini
Concordant activation of MYC and BCL-2 oncoproteins in double-hit lymphoma (DHL) results in aggressive disease that is refractory to treatment. By integrating activity-based proteomic profiling and drug screens, polo-like kinase-1 (PLK1) was identified as an essential regulator of the MYC-dependent kinome in DHL. Notably, PLK1 was expressed at high levels in DHL, correlated with MYC expression and connoted poor outcome. Further, PLK1 signaling augmented MYC protein stability and, in turn, MYC directly induced PLK1 transcription, establishing a feed-forward MYC-PLK1 circuit in DHL. Finally, inhibition of PLK1 triggered degradation of MYC and of the anti-apoptotic protein MCL1, and PLK1 inhibitors showed synergy with BCL-2 antagonists in blocking DHL cell growth, survival and tumorigenicity, supporting clinical targeting of PLK1 in DHL.
Yuan Ren, Chengfeng Bi, Xiaohong Zhao, Tint Lwin, Cheng Wang, Ji Yuan, Ariosto S. Silva, Bijal D. Shah, Bin Fang, Tao Li, John M. Koomen, Huijuan Jiang, Julio C. Chavez, Lan Pham, Praneeth R. Sudalagunta, Lixin Wan, Xuefeng Wang, William S. Dalton, Lynn C. Moscinski, Kenneth H. Shain, Julie Vose, John L. Cleveland, Eduardo M. Sotomayor, Kai Fu, Jianguo Tao
MASTL, a Ser/Thr kinase that inhibits PP2A-B55 complexes during mitosis, is mutated in autosomal dominant thrombocytopenia. However, the connections between the cell cycle machinery and this human disease remain unexplored. We report here that, whereas Mastl ablation in megakaryocytes prevented proper maturation of these cells, mice carrying the thrombocytopenia-associated mutation developed thrombocytopenia as a consequence of aberrant activation and survival of platelets. Activation of mutant platelets was characterized by hyper-stabilized pseudopods mimicking the effect of PP2A inhibition and actin polymerization defects. These aberrations were accompanied by abnormal hyper-phosphorylation of multiple components of the actin cytoskeleton and were rescued both in vitro and in vivo by inhibiting upstream kinases such as PKA, PKC, or AMPK. These data reveal an unexpected role of Mastl in actin cytoskeleton dynamics in postmitotic cells, and suggest that the thrombocytopenia-associated mutation in MASTL is a pathogenic dominant mutation that mimics decreased PP2A activity resulting in altered phosphorylation of cytoskeletal regulatory pathways.
Begoña Hurtado, Marianna Trakala, Pilar Ximénez-Embún, Aicha El Bakkali, David Partida, Belén Sanz-Castillo, Mónica Álvarez-Fernández, María Maroto, Ruth Sánchez-Martínez, Lola Martínez, Javier Muñoz, Pablo García de Frutos, Marcos Malumbres
Acute Myeloid Leukemia and Myelodysplastic Syndromes are associated with disease-initiating stem cells that are not eliminated by conventional therapies. Transcriptomic analysis of stem and progenitor populations in MDS and AML demonstrated overexpression of STAT3 that was validated in an independent cohort. STAT3 overexpression was predictive of a shorter survival and worse clinical features in a large MDS cohort. High STAT3 expression signature in MDS CD34+ cells was similar to known pre-leukemic gene signatures. Functionally, STAT3 inhibition by a clinical, antisense oligonucleotide, AZD9150, led to reduced viability and increased apoptosis in leukemic cell lines. AZD9150 was rapidly incorporated by primary MDS/AML stem and progenitor cells and led to increased hematopoietic differentiation. STAT3 knockdown also impaired leukemic growth in vivo and led to decreased expression of MCL1 and other oncogenic genes in malignant cells. These studies demonstrate that STAT3 is an adverse prognostic factor in MDS/AML and provide a pre-clinical rationale for studies using AZD9150 in these diseases.
Aditi Shastri, Gaurav Choudhary, Margarida Teixeira, Shanisha Gordon-Mitchell, Nandini Ramachandra, Lumie Bernard, Sanchari Bhattacharyya, Robert Lopez, Kith Pradhan, Orsolya Giricz, Goutham Ravipati, Li-Fan Wong, Sally Cole, Tushar D. Bhagat, Jonathan Feld, Yosman Dhar, Matthias Bartenstein, Victor J. Thiruthuvanathan, Amittha Wickrema, B. Hilda Ye, David A. Frank, Andrea Pellagatti, Jacqueline Boultwood, Tianyuan Zhou, Youngsoo Kim, A. Robert MacLeod, Pearlie K. Epling-Burnette, Minwei Ye, Patricia McCoon, Richard Woessner, Ulrich Steidl, Britta Will, Amit K. Verma
Obesity is a major risk factor for developing nonalcoholic fatty-liver disease (NAFLD). NAFLD is the most common form of chronic liver disease and closely associated with insulin resistance, ultimately leading to cirrhosis and hepatocellular carcinoma. However, knowledge of the intracellular regulators of obesity-linked fatty-liver disease remains incomplete. Here we showed that hepatic Rho-kinase 1 (ROCK1) drives obesity-induced steatosis in mice through stimulation of de novo lipogenesis. Mice lacking ROCK1 in the liver were resistant to diet-induced obesity due to increased energy expenditure and thermogenic gene expression. Constitutive expression of hepatic ROCK1 was sufficient to promote adiposity, insulin resistance, and hepatic lipid accumulation in mice fed a high-fat diet. Correspondingly, liver-specific ROCK1 deletion prevented the development of severe hepatic steatosis and reduced hyperglycemia in obese diabetic (ob/ob) mice. Of pathophysiologic significance, hepatic ROCK1 was markedly up-regulated in humans with fatty-liver disease and correlated with risk factors clustering around NAFLD and insulin resistance. Mechanistically, we found that hepatic ROCK1 suppresses AMPK activity and a ROCK1-AMPK pathway is necessary to mediate cannabinoid-induced lipogenesis in the liver. Furthermore, treatment with metformin, the most widely used anti-diabetes drug, reduced hepatic lipid accumulation by inactivating ROCK1, resulting in activation of AMPK downstream signaling. Taken together, our findings establish a ROCK1-AMPK signaling axis that regulates de novo lipogenesis, providing a unique target for treating obesity-related metabolic disorders such as NAFLD.
Hu Huang, Seung-Hwan Lee, Inês Sousa-Lima, Sang Soo Kim, Won Min Hwang, Yossi Dagon, Won-Mo Yang, Sungman Cho, Min-Cheol Kang, Ji A Seo, Munehiko Shibata, Hyunsoo Cho, Getachew Debas Belew, Jinhyuk Bhin, Bhavna N. Desai, Min Jeong Ryu, Minho Shong, Peixin Li, Hua Meng, Byung-Hong Chung, Daehee Hwang, Min Seon Kim, Kyong Soo Park, Paula Macedo, Morris White, John Jones, Young-Bum Kim
Sugar- and lipid-derived aldehydes are reactive carbonyl species (RCS) frequently used as surrogate markers of oxidative stress in obesity. A pathogenic role for RCS in metabolic diseases of obesity remains controversial, however, due in part to their highly diffuse and broad reactivity, and to lack of specific RCS-scavenging therapies. Naturally occurring histidine dipeptides (e.g., anserine and carnosine) possess RCS reactivity, but their therapeutic potential in humans is limited by serum carnosinases. Here we present the rational design, characterization and pharmacological evaluation of ‘carnosinol’ (i.e. (2S)-2-(3-amino propanoylamino)-3-(1H-imidazol-5-yl)propanol) a derivative of carnosine with high oral bioavailability that is resistant to carnosinases. Carnosinol displayed a suitable ADMET profile and was determined to have the greatest potency and selectivity toward α,β-unsaturated aldehydes (e.g. 4-hydroxynonenal, HNE, acrolein) among all others so far reported. In rodent models of diet-induced obesity and metabolic syndrome, carnosinol dose-dependently attenuated HNE-adduct formation in liver and skeletal muscle while simultaneously mitigating inflammation, dyslipidemia, insulin resistance, and steatohepatitis. These improvements in metabolic parameters with carnosinol were not due to changes in energy expenditure, physical activity, adiposity or body weight. Collectively, our findings illustrate a pathogenic role for RCS in obesity-related metabolic disorders, and provide validation for a promising new class of carbonyl-scavenging therapeutic compounds rationally derived from carnosine.
Ethan J. Anderson, Giulio Vistoli, Lalage A. Katunga, Katsuhiko Funai, Luca Regazzoni, T. Blake Monroe, Ettore Gilardoni, Luca Cannizzaro, Mara Colzani, Danilo De Maddis, Giuseppe Rossoni, Renato Canevotti, Stefania Gagliardi, Marina Carini, Giancarlo Aldini
Heart failure (HF) remains a major source of morbidity and mortality in the U.S. The multifunctional Ca2+/calmodulin-dependent kinase II (CaMKII) has emerged as a critical regulator of cardiac hypertrophy and failure, although the mechanisms remain unclear. Previous studies have established that the cytoskeletal protein βIV-spectrin coordinates local CaMKII signaling. Here we sought to determine the role of a spectrin/CaMKII complex in maladaptive remodeling in HF. Chronic pressure overload (6 weeks transaortic constriction, TAC) induced a decrease in cardiac function in WT mice but not in animals expressing truncated βIV-spectrin lacking spectrin/CaMKII interaction (qv3J). Underlying observed differences in function was an unexpected differential regulation of STAT3-related genes in qv3J TAC hearts. In vitro experiments demonstrate that βIV-spectrin serves as a target for CaMKII phosphorylation, which regulates its stability. Cardiac-specific βIV-spectrin knockout (βIV-cKO) mice show STAT3 dysregulation, fibrosis and decreased cardiac function at baseline similar to WT TAC. STAT3 inhibition restored normal cardiac structure and function in βIV-cKO and WT TAC hearts. Our studies identify a novel spectrin-based complex essential for regulation of the cardiac response to chronic pressure overload. We anticipate that strategies targeting the new spectrin-based “statosome” will be effective at suppressing maladaptive remodeling in response to chronic stress.
Sathya D. Unudurthi, Drew M. Nassal, Amara Greer-Short, Nehal J. Patel, Taylor Howard, Xianyao Xu, Birce Onal, Tony Satroplus, Deborah Y. Hong, Cemantha M. Lane, Alyssa Dalic, Sara N. Koenig, Adam C. Lehnig, Lisa A. Baer, Hassan Musa, Kristin I. Stanford, Sakima A. Smith, Peter J. Mohler, Thomas J. Hund
The subthalamic nucleus (STN) is an effective therapeutic target for deep brain stimulation (DBS) for Parkinson’s disease (PD) and histamine level is elevated in the basal ganglia in PD patients. However, the endogenous histaminergic modulation on STN neuronal activities and the neuronal mechanism underlying STN-DBS are unknown. Here we report that STN neuronal firing patterns are more crucial than firing rates for motor control. Histamine excited STN neurons, but paradoxically ameliorated parkinsonian motor deficits, which we attributed to regularizing firing patterns of STN neurons via HCN2 channel coupled to H2 receptor. Intriguingly, DBS increased histamine release in the STN and regularized STN neuronal firing patterns under parkinsonian conditions. HCN2 contributed to the DBS-induced regularization of neuronal firing patterns, suppression of excessive beta oscillations, and alleviation of motor deficits in PD. The results reveal an indispensable role for regularizing STN neuronal firing patterns in amelioration of parkinsonian motor dysfunction and a functional compensation for histamine in parkinsonian basal ganglia circuitry. The findings provide insights into mechanisms of STN-DBS as well as potential therapeutic targets and STN-DBS strategies for PD.
Qian-Xing Zhuang, Guang-Ying Li, Bin Li, Chang-Zheng Zhang, Xiao-Yang Zhang, Kang Xi, Hong-Zhao Li, Jian-Jun Wang, Jing-Ning Zhu
Experimental autoimmune encephalomyelitis (EAE) is an inflammatory demyelinating disease of the central nervous system (CNS), induced by the adoptive transfer of myelin-reactive CD4+ T cells into naïve syngeneic mice. It is widely used as a rodent model of multiple sclerosis (MS). EAE lesion development is initiated when transferred CD4+ T cells access the CNS and are reactivated by local antigen presenting cells (APC) bearing endogenous myelin peptide/ MHC Class II complexes. The identity of the CNS resident, lesion-initiating APC is widely debated. Here we demonstrate that classical dendritic cells (cDC) normally reside in the meninges, brain, and spinal cord in the steady state. These cells are unique among candidate CNS APC in their ability to stimulate naïve, as well as effector, myelin-specific T cells to proliferate and produce pro-inflammatory cytokines directly ex vivo. cDC expanded in the meninges and CNS parenchyma in association with disease progression. Selective depletion of cDC led to a decrease in the number of myelin-primed donor T cells in the CNS and reduced the incidence of clinical EAE by half. Based on our findings, we propose that cDC, and the factors that regulate them, be further investigated as potential therapeutic targets in MS.
David A. Giles, Patrick C. Duncker, Nicole M. Wilkinson, Jesse M. Washnock-Schmid, Benjamin M. Segal
ASXL1 is frequently mutated in myeloid malignancies and is known to co-occur with other gene mutations. However, the molecular mechanisms underlying the leukemogenesis associated with ASXL1 and cooperating mutations remain to be elucidated. Here we report that Asxl1 loss cooperated with haploinsufficiency of Nf1, a negative regulator of the RAS signaling pathway, to accelerate the development of myeloid leukemia in mice. Loss of Asxl1 and Nf1 in hematopoietic stem and progenitor cells resulted in a gain-of-function transcriptional activation of multiple pathways critical for leukemogenesis, such as MYC, NRAS, and BRD4. The hyperactive MYC and BRD4 transcription programs were correlated with elevated H3K4 tri-methylation at the promoter regions of genes involving these pathways. Furthermore, pharmacological inhibition of both MAPK pathway and BET bromodomain prevented leukemia initiation and inhibited disease progression in Asxl1Δ/Δ;Nf1Δ/Δ mice. Concomitant mutations of ASXL1 and RAS pathway genes were associated with aggressive progression of myeloid malignancies in patients. This study sheds light on the understanding of the cooperative effect between epigenetic alterations and signaling pathways in accelerating the progression of myeloid malignancies and provides a rational therapeutic strategy for the treatment of myeloid malignancies with ASXL1 and RAS pathway gene mutations.
Peng Zhang, Fuhong He, Jie Bai, Shohei Yamamoto, Shi Chen, Lin Zhang, Mengyao Sheng, Lei Zhang, Ying Guo, Na Man, Hui Yang, Suyun Wang, Tao Cheng, Stephen D. Nimer, Yuan Zhou, Mingjiang Xu, Qian-Fei Wang, Feng-Chun Yang
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