The discovery of an HIV-1 cure remains a medical challenge because the virus rebounds quickly after the cessation of combination antiretroviral drug therapy (cART). Here, we investigate the potential of an engineered tandem bi-specific broadly neutralizing antibody (bs-bnAb) as an innovative product for HIV-1 prophylactic and therapeutic interventions. We discovered that by preserving two scFv binding domains of each parental bnAb, a single-gene-encoded tandem bs-bnAb, namely BiIA-SG, displayed significantly improved breadth and potency. BiIA-SG neutralized all 124 HIV-1 pseudotyped viruses tested, including global subtypes/recombinant forms, transmitted/founder viruses, and variants less or not susceptible to parental and many bnAbs, with an average IC50 value of 0.073 µ/ml (range < 0.001 to 1.03 µg/ml). In humanized mice, an injection of BiIA-SG conferred sterile protection when administered prior to challenges with diverse live HIV-1 stains. Moreover, while BiIA-SG delayed viral rebound in a short-term therapeutic setting when combined with cART, a single injection of AAV-transferred BiIA-SG gene resulted dose-dependently in prolonged in vivo expression of BiIA-SG, which was associated with complete viremia control and subsequent elimination of infected cells in humanized mice. These results warrant the clinical development of BiIA-SG as a promising bs-bnAb-based biomedical intervention for prevention and treatment of HIV-1 infections.
Xilin Wu, Jia Guo, Mengyue Niu, Minghui An, Li Liu, Hui Wang, Xia Jin, Qi Zhang, Ka Shing Lam, Tongjin Wu, Hua Wang, Qian Wang, Yanhua Du, Jingjing Li, Lin Cheng, Hang Ying Tang, Hong Shang, Linqi Zhang, Paul Zhou, Zhiwei Chen
The immune system is tightly controlled by regulatory processes that allow for the elimination of invading pathogens, while limiting immunopathological damage to the host. In the present study, we found that conditional deletion of the cell surface receptor Toso on B cells unexpectedly resulted in impaired proinflammatory T cell responses, which led to impaired immune protection in an acute viral infection model, while, in a chronic inflammatory context, was associated with reduced immunopathological tissue damage. Toso exhibited its B cell-inherent immunoregulatory function by negatively controlling the pool of IL-10-competent B1 and B2 B cells, which were characterized by a high degree of self-reactivity and were shown to mediate immunosuppressive activity on inflammatory T cell responses in vivo. Our results indicate that Toso is involved in the differentiation/maintenance of regulatory B cells by fine-tuning B cell receptor (BCR)-activation thresholds. Furthermore, we showed that during influenza A-induced pulmonary inflammation the application of Toso-specific antibodies selectively induced IL-10-competent B cells at the site of inflammation and resulted in decreased proinflammatory cytokine production by lung T cells. These findings suggest that Toso may serve as a novel therapeutic target to dampen pathogenic T cell responses via the modulation of IL-10-competent regulatory B cells.
Jinbo Yu, Vu Huy Hoang Duong, Katrin Westphal, Andreas Westphal, Abdulhadi Suwandi, Guntram A. Grassl, Korbinian Brand, Andrew C. Chan, Niko Föger, Kyeong-Hee Lee
Genetic forms of vitamin D–dependent rickets (VDDRs) are due to mutations impairing activation of vitamin D or decreasing vitamin D receptor responsiveness. Here we describe two unrelated patients with early-onset rickets, reduced serum levels of the vitamin D metabolites 25-hydroxyvitamin D and 1,25-dihydroxyvitamin D, and deficient responsiveness to parent and activated forms of vitamin D. Neither patient had a mutation in any genes known to cause VDDR, however, using whole exome sequence analysis we identified a recurrent de novo missense mutation c.902T>C (p.I301T) in CYP3A4 in both subjects that alters the conformation of substrate-recognition-site 4 (SRS-4). In vitro, the mutant CYP3A4 oxidized 1,25-dihydroxyvitamin D with 10-fold greater activity than wild-type CYP3A4 and 2-fold greater activity than CYP24A1, the principal inactivator of vitamin D metabolites. As CYP3A4 mutations have not previously been linked to rickets, these findings provide new insight into vitamin D metabolism, and demonstrate that accelerated inactivation of vitamin D metabolites represents a previously undescribed mechanism for vitamin D deficiency.
Jeffrey D. Roizen, Dong Li, Lauren O'Lear, Muhammad K. Javaid, Nicholas J. Shaw, Peter R. Ebeling, Hanh H. Nguyen, Christine P. Rodda, Kenneth E. Thummel, Tom D Thacher, Hakon Hakonarson, Michael A. Levine
HLA-B*57 control of HIV involves enhanced CD8+ T cell responses against infected cells, but extensive heterogeneity exists in level of HIV control among B*57+ individuals. Using whole genome sequencing of untreated B*57+ HIV-1 infected controllers and non-controllers, we identified a single variant (rs643347A/G) encoding an isoleucine to valine substitution at position 47 (I47V) of the inhibitory killer cell immunoglobulin-like receptor, KIR3DL1, as the only significant modifier of B*57 protection. The association replicated in an independent cohort and across multiple outcomes. The modifying effect of I47V was confined to B*57:01, and was not observed for the closely related B*57:03. Positions 2, 47, and 54 track one another nearly perfectly, and two KIR3DL1 allotypes differing only at these three positions showed significant differences in binding B*57:01 tetramers, where the protective allotype showed lower binding. Thus, variation in an immune natural killer cell receptor that binds B*57:01 modifies its protection. These data speak to exquisite specificity of KIR-HLA interactions in human health and disease.
Maureen P. Martin, Vivek Naranbhai, Patrick R. Shea, Ying Qi, Veron Ramsuran, Nicolas Vince, Xiaojiang Gao, Rasmi Thomas, Zabrina L. Brumme, Jonathan M. Carlson, Steven M. Wolinsky, James J. Goedert, Bruce D. Walker, Florencia P. Segal, Steven G. Deeks, David W. Haas, Stephen A. Migueles, Mark Connors, Nelson Michael, Jacques Fellay, Emma Gostick, Sian Llewellyn-Lacey, David A. Price, Bernard A. Lafont, Phillip Pymm, Philippa M. Saunders, Jacqueline Widjaja, Shu Cheng Wong, Julian P. Vivian, Jamie Rossjohn, Andrew G. Brooks, Mary Carrington
Synthetic lethality is an efficient mechanism-based approach to selectively target DNA repair defects. ERCC1 deficiency is frequently found in non-small cell lung cancers, making this DNA repair protein an attractive target for exploiting synthetic lethal approaches in this disease. Using unbiased proteomic and metabolic high-throughput profiling on a unique in-house generated isogenic model of ERCC1 deficiency, we found marked metabolic rewiring of ERCC1-deficient populations, including decreased levels of the metabolite NAD+ and reduced expression of the rate-limiting NAD+ biosynthetic enzyme nicotinamide phosphoribosyltransferase (NAMPT). We further evidenced reduced NAMPT expression in NSCLC samples with low levels of ERCC1. These metabolic alterations were a primary effect of ERCC1 deficiency, and caused selective exquisite sensitivity to small molecule NAMPT inhibitors, both in vitro — ERCC1-deficient cells being approximately 1000 times more sensitive — and in vivo. Using transmission electronic microscopy and functional metabolic studies, we found that ERCC1-deficient cells harbor mitochondrial defects. We propose a model where NAD+ acts as a regulator of ERCC1-deficient NSCLC fitness. These findings open therapeutic opportunities that exploit a yet undescribed nuclear — mitochondrial synthetic lethal relationship in cancer cells, and highlight the potential for targeting DNA repair/metabolic crosstalks for cancer therapy.
Mehdi Touat, Tony Sourisseau, Nicolas Dorvault, Roman M. Chabanon, Marlène Garrido, Daphné Morel, Dragomir B. Krastev, Ludovic Bigot, Julien Adam, Jessica Frankum, Sylvère Durand, Clement Pontoizeau, Sylvie Souquère, Mei-Shiue Kuo, Sylvie Sauvaigo, Faraz Mardakheh, Alain Sarasin, Ken A. Olaussen, Luc Friboulet, Frédéric Bouillaud, Gérard Pierron, Alan Ashworth, Anne Lombès, Christopher J. Lord, Jean-Charles Soria, Sophie Postel-Vinay
Major histocompatibility (MHC) class II molecules are strongly associated with many autoimmune disorders. In type 1 diabetes, the DQ8 molecule is common, confers significant disease risk and is involved in disease pathogenesis. We hypothesized blocking DQ8 antigen presentation would provide therapeutic benefit by preventing recognition of self-peptides by pathogenic T cells. We used the crystal structure of DQ8 to select drug-like small molecules predicted to bind structural pockets in the MHC antigen-binding cleft. A limited number of the predicted compounds inhibited DQ8 antigen presentation in vitro with one compound preventing insulin autoantibody production and delaying diabetes onset in an animal model of spontaneous autoimmune diabetes. An existing drug of similar structure, methyldopa, specifically blocked DQ8 in recent-onset patients with type 1 diabetes along with reducing inflammatory T cell responses toward insulin, highlighting the relevance of blocking disease-specific MHC class II antigen presentation to treat autoimmunity.
David A. Ostrov, Aimon Alkanani, Kristen A. McDaniel, Stephanie Case, Erin E. Baschal, Laura Pyle, Samuel Ellis, Bernadette Pöllinger, Katherine J. Seidl, Viral N. Shah, Satish K. Garg, Mark A. Atkinson, Peter A. Gottlieb, Aaron W. Michels
Immune evasion and the suppression of anti-tumor responses during cancer progression are considered hallmarks of cancer and are typically attributed to tumor-derived factors. Although the molecular basis for the crosstalk between tumor and immune cells is an area of active investigation, whether host-specific germline variants can dictate immunosuppressive mechanisms has remained a challenge to address. A commonly occurring germline mutation (c.1162G>A/rs351855 G/A) in the FGFR4 (CD334) gene enhances STAT3 signaling and is associated with poor prognosis and accelerated progression of multiple cancer types. Here, using rs351855 single nucleotide polymorphism (SNP) knock-in transgenic mice and Fgfr4 knockout mice, we reveal the genotype-specific gain of immunological function of suppressing the CD8/CD4+FOXP3+CD25+ve regulatory T cell ratio in vivo. Furthermore, using knock-in transgenic mouse models for lung and breast cancers, we establish the host-specific tumor-extrinsic functions of STAT3-enhancing germline variants in impeding the tumor infiltration of CD8 T cells. Thus, STAT3-enhancing germline receptor variants contribute to immune evasion through their pleiotropic functions in immune cells.
Daniel Kogan, Alexander Grabner, Christopher Yanucil, Christian Faul, Vijay Kumar Ulaganathan
In these studies we evaluated the contribution of the NLRP3 inflammasome to Crohn’s disease (CD) in a kindred containing individuals having a missense mutation in CARD8, a protein known to inhibit this inflammasome. Whole exome sequencing and PCR studies identified that the affected individuals had a V44I mutation in a single allele of the T60 isoform of CARD8. The serum levels of IL-1β in the affected individuals were increased compared with that in healthy controls and their peripheral monocytes produced increased amounts of IL-1β when stimulated by NLRP3 activators. Immunoblot studies probing the basis of these findings showed that mutated T60 CARD8 fails to down-regulate the NLRP3 inflammasome because it does not bind to NLRP3 and inhibit its oligomerization. In addition, these studies showed that mutated T60 CARD8 exerts a dominant negative effect by its capacity to bind to and form oligomers with unmutated T60 or T48 CARD8 that impede their binding to NLRP3. Finally, inflammasome activation studies revealed that intact but not mutated CARD8 prevents NLRP3 deubiquitination and serine dephosphorylation. CD due to a CARD8 mutation was not effectively treated by anti-TNF-α, but did respond to IL-1β inhibitors. Thus, patients with anti-TNF-α-resistant CD may respond to this treatment option.
Liming Mao, Atsushi Kitani, Morgan Similuk, Andrew J. Oler, Lindsey Albenberg, Judith Kelsen, Atiye Aktay, Martha Quezado, Michael Yao, Kim Montgomery-Recht, Ivan J. Fuss, Warren Strober
A modifier variant can abrogate risk of a monogenic disorder. DFNM1 is a locus on chromosome 1 encoding a dominant suppressor of human DFNB26 recessive, profound deafness. Here, we report that DFNB26 is associated with a substitution (p.Gly116Glu) in the pleckstrin-homology-domain of GAB1, an essential scaffold in the MET/HGF pathway. A dominant substitution (p.Arg544Gln) of METTL13, encoding a predicted methyltransferase, is the DFNM1 suppressor of GAB1-associated deafness. In zebrafish, human METTL13 mRNA harboring the modifier allele rescues the GAB1 associated morphant phenotype. In mouse, GAB1 and METTL13 co-localize in auditory sensory neurons, and METTL13 co-immunoprecipitates with GAB1 and SPRY2, indicating at least a tripartite complex. Expression of MET-signaling genes in human lymphoblastoid cells of individuals homozygous for p.Gly116Glu GAB1 revealed dysregulation of HGF, MET, SHP2, and SPRY2, all of which have reported variants associated with deafness. However, SPRY2 was not dysregulated in normal-hearing humans homozygous for both the GAB1 DFNB26 deafness variant and the dominant METTL13 deafness suppressor, indicating a plausible mechanism of suppression. Identification of METTL13-based modification of MET-signaling provides potential therapeutic strategy for a wide range of associated hearing disorders. Furthermore, MET-signaling is essential for diverse functions in many tissues including the inner ear. Therefore, identification of the modifier of MET-signaling is likely to have broad clinical implications.
Rizwan Yousaf, Zubair M. Ahmed, Arnaud P.J. Giese, Robert J. Morell, Ayala Lagziel, Alain Dabdoub, Edward R. Wilcox, Sheikh Riazuddin, Thomas B. Friedman, Saima Riazuddin
Transient vanilloid potential 1 (TRPV1) agonists are emerging as highly efficacious non-opioid analgesics in preclinical studies. These drugs selectively lesion TRPV1+ primary sensory afferents, which are responsible for the transmission of many noxious stimulus modalities. Resiniferatoxin (RTX) is a very potent and selective TRPV1 agonist and is a promising candidate for treating many types of pain. Recent work establishing intrathecal application of RTX for the treatment of pain resulting from advanced cancer has demonstrated profound analgesia in client-owned dogs with osteosarcoma. The present study uses transcriptomics and histochemistry to examine the molecular mechanism of RTX action in rats, in clinical canine subjects, and in one human subject with advanced cancer treated for pain using intrathecal RTX. In all three species we observe a strong analgesic action, yet this was accompanied by limited transcriptional alterations at the level of the DRG. Functional and neuroanatomical studies demonstrated that intrathecal RTX largely spares susceptible neuronal perikarya, which remain active peripherally, but unable to transmit signals to the spinal cord. The results demonstrate that central chemo-axotomy of the TRPV1+ afferents underlies RTX analgesia and refine the neurobiology underlying effective clinical use of TRPV1 agonists for pain control.
Matthew R. Sapio, John K. Neubert, Danielle M. LaPaglia, Dragan Maric, Jason M. Keller, Stephen J. Raithel, Eric L. Rohrs, Ethan M. Anderson, John A. Butman, Robert M. Caudle, Dorothy C. Brown, John D. Heiss, Andrew J. Mannes, Michael J. Iadarola
Insulin resistance and type 2 diabetes are associated with low levels of high-density lipoprotein-cholesterol (HDL-C). The insulin-repressible FoxO transcription factors are potential mediators of insulin’s effect on HDL-C. FoxOs mediate a substantial portion of insulin-regulated transcription, and poor FoxO repression is thought to contribute to the excessive glucose production in diabetes. In this work, we show that mice with liver-specific triple FoxO knockout (L-FoxO1,3,4), which are known to have reduced hepatic glucose production, also have increased HDL-C. This was associated with decreased expression of HDL-C clearance factors, scavenger receptor class B type I (SR-BI) and hepatic lipase, and defective selective uptake of HDL-cholesteryl ester by the liver. The phenotype could be rescued by re-expression of SR-BI. These findings demonstrate that hepatic FoxOs are required for cholesterol homeostasis and HDL-mediated reverse cholesterol transport to the liver.
Samuel X. Lee, Markus Heine, Christian Schlein, Rajasekhar Ramakrishnan, Jing Liu, Gabriella Belnavis, Ido Haimi, Alexander W. Fischer, Henry Ginsberg, Joerg Heeren, Franz Rinninger, Rebecca A. Haeusler
Ribosomal proteins (RP) regulate specific gene expression by selectively translating subsets of mRNAs. Indeed, in Diamond–Blackfan anaemia and 5q- syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anaemia. Little is known about the molecular mechanisms of selective mRNA translation and involvement of ribosomal-associated factors in this process. Ribonuclease inhibitor (RNH1) is an ubiquitously expressed protein that binds to and inhibits pancreatic-type ribonucleases. Here we report that RNH1 binds to ribosomes and regulates erythropoiesis by controlling translation of the erythroid transcription factor GATA1. Rnh1-deficient mice die between embryonic days E8.5 to E10 due to impaired production of mature erythroid cells from progenitor cells. In Rnh1-deficient embryos, mRNA levels of Gata1 are normal, but GATA1 protein levels are decreased. At the molecular level, we found that RNH1 binds to the 40S subunit of ribosomes and facilitates polysome formation on Gata1 mRNA to confer transcript-specific translation. Further, RNH1 knock down in human CD34+ progenitor cells decreased erythroid differentiation without affecting myelopoiesis. Our results reveal an unsuspected role for RNH1 in the control of GATA1 mRNA translation and erythropoiesis.
Vijaykumar Chennupati, Diogo F.T. Veiga, Kendle M. Maslowski, Nicola Andina, Aubry Tardivel, Eric Chi-Wang Yu, Martina Stilinovic, Cedric Simillion, Michel A. Duchosal, Manfredo Quadroni, Irene Roberts, Vijay G. Sankaran, H. Robson MacDonald, Nicolas Fasel, Anne Angelillo-Scherrer, Pascal Schneider, Trang Hoang, Ramanjaneyulu Allam
Tuberous sclerosis complex (TSC) is a dominantly inherited disease, caused by hyperactivation of the mTORC1 pathway and characterized by the development of hamartomas and benign tumors, also in the brain. Among the neurological manifestations associated with TSC, the tumor progression of static subependymal nodules (SENs) into subependymal giant cell astrocytomas (SEGAs) is one of the major causes of morbidity and shortened life expectancy. To date, mouse modeling has failed in reproducing these two lesions. Here we report that simultaneous hyperactivation of mTORC1 and Akt pathways by codeletion of Tsc1 and Pten, selectively in postnatal neural stem cells (pNSCs), is required for the formation of bona fide SENs and SEGAs. Notably, both lesions closely recapitulate the pathognomonic morphological and molecular features of the corresponding human abnormalities. The establishment of long-term expanding pNSC lines from mouse SENs and SEGAs made possible the identification of mTORC2 as one of the mediators conferring tumorigenic potential to SEGA pNSCs. Of note, in spite of concurrent Akt hyperactivation in mouse brain lesions, single mTOR inhibition by rapamycin was sufficient to strongly impair mouse SEGA growth. This study provides the first evidence that, concomitant with mTORC1 hyperactivation, sustained activation of Akt and mTORC2 in pNSCs is a mandatory step for the induction of SENs and SEGAs and, at the same time, makes available an unprecedented NSC-based in vivo/in vitro model to be exploited for identifying actionable targets in TSC.
Paola Zordan, Manuela Cominelli, Federica Cascino, Elisa Tratta, Pietro L. Poliani, Rossella Galli
BACKGROUND. Drugs and vaccines that can interrupt the transmission of Plasmodium falciparum will be important for malaria control and elimination. However, models for early clinical evaluation of candidate transmission-blocking interventions are currently unavailable. Here we describe a new model for evaluating malaria transmission from humans to Anopheles mosquitoes using controlled human malaria infection (CHMI). METHODS. Seventeen healthy malaria-naïve volunteers underwent CHMI by intravenous inoculation of P. falciparum-infected erythrocytes to initiate blood-stage infection. Seven to eight days after inoculation participants received piperaquine (480 mg) to attenuate asexual parasite replication while allowing gametocytes to develop and mature. Primary endpoints were development of gametocytemia, the transmissibility of gametocytes from humans to mosquitoes, and the safety and tolerability of the CHMI transmission model. To investigate in-vivo gametocytocidal drug activity in this model, participants were either given an experimental antimalarial, artefenomel (500 mg), a known gametocytocidal drug, primaquine (15 mg), or remained untreated during the period of gametocyte carriage. RESULTS. Male and female gametocytes were detected in all participants, and transmission to mosquitoes was achieved from 8/11 (73%) participants evaluated. Compared to untreated controls (n = 7), primaquine (15 mg, n = 5) significantly reduced gametocyte burden (P = 0.01), while artefenomel (500 mg, n = 4) had no effect. Adverse events (AEs) were mostly mild or moderate. Three AEs were assessed as severe — fatigue, elevated alanine aminotransferase, and elevated aspartate aminotransferase — and were attributed to malaria infection. Transaminase elevations were transient, asymptomatic, and resolved without intervention. CONCLUSION. We report the safe and reproducible induction of P. falciparum gametocytes in healthy malaria-naïve volunteers at densities infectious to mosquitoes, thereby demonstrating the potential for evaluating transmission-blocking interventions in this model. TRIAL REGISTRATION. ClinicalTrials.gov NCT02431637 and NCT02431650 FUNDING. Bill & Melinda Gates Foundation
Katharine A. Collins, Claire Y.T. Wang, Matthew Adams, Hayley Mitchell, Melanie Rampton, Suzanne Elliott, Isaie J. Reuling, Teun Bousema, Robert Sauerwein, Stephan Chalon, Jörg J. Möhrle, James S. McCarthy
BACKGROUND. Amongst non-diabetic individuals, mild glucose decrements alter brain activity in regions linked to reward, motivation and executive control. Whether these effects differ in T1DM patients with and without hypoglycemia awareness remains unclear. METHODS. 42 individuals (13 healthy control subjects (HC), 16 T1DM individuals with hypoglycemia awareness (T1DM-Aware) and 13 T1DM individuals with hypoglycemia unawareness (T1DM-Unaware)) underwent BOLD fMRI brain imaging during a 2-step hyperinsulinemic euglycemic (90 mg/dl)-hypoglycemic (60 mg/dl) clamp for assessment of neural responses to mild hypoglycemia. RESULTS. Mild hypoglycemia in HC altered activity in the caudate, insula, prefrontal cortex, and angular gyrus, whereas T1DM-Aware subjects showed no caudate and insula changes, but showed altered activation patterns in the prefrontal cortex and angular gyrus. Most strikingly, in direct contrast to HC and T1DM-Aware subjects, T1DM-Unaware subjects failed to show any hypoglycemia-induced changes in brain activity. These findings were also associated with blunted hormonal counterregulatory responses and hypoglycemia symptoms scores during mild hypoglycemia. CONCLUSION. In T1DM, and in particular T1DM-Unaware patients, there is a progressive blunting of brain responses in cortico-striatal and fronto-parietal neurocircuits in response to mild-moderate hypoglycemia. These findings have implications for understanding why individuals with impaired hypoglycemia awareness fail to respond appropriately to falling blood glucose levels. FUNDING. This study was supported in part by grants from the NIH R01DK020495 and P30 DK045735 (Sherwin), K23DK109284 (Hwang), K08AA023545 (Seo), the Yale Center for Clinical Investigation supported by the Clinical Translational Science Award (UL1 RR024139).
Janice Jin Hwang, Lisa Parikh, Cheryl Lacadie, Dongju Seo, Wai Lam, Muhammad Hamza, Christian Schmidt, Feng Dai, Anne-Sophie Sejling, Renata Belfort-DeAguiar, R. Todd Constable, Rajita Sinha, Robert Sherwin
Myc activation is a primary oncogenic event in many human cancers; however, these transcription factors are difficult to inhibit pharmacologically, suggesting that Myc-dependent downstream effectors may be more tractable therapeutic targets. Here we show that Myc overexpression induces endoplasmic reticulum (ER) stress and engages the IRE1α-XBP1 pathway through multiple molecular mechanisms in a variety of c-Myc- and N-Myc-dependent cancers. In particular, Myc-overexpressing cells require IRE1α-XBP1 signaling for sustained growth and survival in vitro and in vivo, dependent on elevated stearoyl-CoA-desaturase 1 (SCD1) activity. Pharmacological and genetic XBP1 inhibition induces Myc-dependent apoptosis, which is alleviated by exogenous unsaturated fatty acids. Of note, SCD1 inhibition phenocopies IRE1α RNase activity suppression in vivo. Furthermore, IRE1α inhibition enhances the cytotoxic effects of standard chemotherapy drugs used to treat c-Myc-overexpressing Burkitt’s lymphoma, suggesting that inhibiting the IRE1α-XBP1 pathway is a useful general strategy for treatment of Myc-driven cancers.
Hong Xie, Chih-Hang Anthony Tang, Jun H. Song, Anthony Mancuso, Juan R. Del Valle, Jin Cao, Yan Xiang, Chi V. Dang, Roy Lan, Danielle J. Sanchez, Brian Keith, Chih-Chi Andrew Hu, M. Celeste Simon
Lupus nephritis (LN) often results in progressive renal dysfunction. The inactive Rhomboid 2 (iRhom2) is a newly identified key regulator of A disintegrin and metalloprotease 17 (ADAM17), whose substrates, such as TNF-α and heparin-binding EGF (HB-EGF), have been implicated in the pathogenesis of chronic kidney disease. Here we demonstrate that deficiency of iRhom2 protects the lupus-prone Fcgr2b–/– mice from developing severe kidney damage without altering anti-double stranded (ds) DNA Ab production, by simultaneously blocking the HB-EGF/EGFR and the TNF-α signaling in the kidney tissues. Unbiased transcriptome profiling of kidneys and kidney macrophages revealed that TNF-α and HB-EGF/EGFR signaling pathways are highly upregulated in Fcgr2b–/– mice; alterations that were diminished in the absence of iRhom2. Pharmacological blockade of either TNF-α or EGFR signaling protected Fcgr2b–/– mice from severe renal damage. Finally, kidneys from LN patients showed increased iRhom2 and HB-EGF expression, with interstitial HB-EGF expression significantly associated with chronicity indices. Our data suggest that activation of iRhom2/ADAM17-dependent TNF-α and EGFR signaling plays a crucial role in mediating irreversible kidney damage in LN, thereby uncovering a novel target for selective and simultaneous dual inhibition of two major pathological pathways in the effector arm of the disease.
Xiaoping Qing, Yurii Chinenov, Patricia Redecha, Michael Madaio, Joris J.T.H. Roelofs, Gregory Farber, Priya D. Issuree, Laura Donlin, David R. McIlwain, Tak W. Mak, Carl P. Blobel, Jane E. Salmon
T cells specific for neoantigens encoded by mutated genes in cancers are increasingly recognized as mediators of tumor destruction after immune checkpoint inhibitor therapy or adoptive cell transfer. Unfortunately, most neoantigens result from random mutations and are patient specific, and some cancers contain few mutations to serve as potential antigens. We describe a patient with stage IV acral melanoma who obtained a complete response following adoptive transfer of tumor infiltrating lymphocytes (TIL). Tumor exome sequencing surprisingly revealed less than 30 somatic mutations, including oncogenic BRAF V600E. Analysis of the specificity of TIL identified rare CD4 T cells specific for BRAFV600E and diverse CD8 T cells reactive to non-mutated self-antigens. These specificities increased in blood after TIL transfer and persisted long term suggesting they contributed to the effective antitumor immune response. Gene transfer of the BRAFV600E-specific T cell receptor (TCR) conferred recognition of class II MHC positive cells expressing the BRAF mutation. Therapy with TCR engineered BRAFV600E-specific CD4+ T cells may have direct antitumor effects and augment CD8+ T cell responses to self and/or mutated tumor antigens in patients with BRAF mutated cancers.
Joshua R. Veatch, Sylvia M. Lee, Matthew Fitzgibbon, I-Ting Chow, Brenda Jesernig, Thomas Schmitt, Ying Ying Kong, Julia Kargl, A. McGarry Houghton, John A. Thompson, Martin McIntosh, William W. Kwok, Stanley R. Riddell
Aberrant activation of MAPK signaling leads to activation of oncogenic transcriptomes. How MAPK signaling is coupled with transcriptional response in cancer is not fully understood. In gastrointestinal stromal tumor and melanoma, both with oncogenic MAPK activation, we find that ETV1 and other Pea3-ETS transcription factors are critical nuclear effectors of MAPK signaling that are regulated through protein stability. Expression of stabilized Pea3-ETS factors can partially rescue the MAPK transcriptome and cell viability after MAPK inhibition. To identify players involved in this process, we performed a pooled genome-wide RNAi screen using a novel fluorescence-based ETV1 protein stability sensor, and identified COP1, DET1, DDB1, UBE3C, PSMD4, and COP9 signalosome members. COP1 and DET1 loss led to decoupling between MAPK signaling and downstream transcriptional response, where MAPK inhibition failed to destabilize Pea3 factors and fully inhibit the MAPK transcriptome, thus resulting in decreased sensitivity to MAPK pathway inhibitors. We identified multiple COP1 and DET1 mutations in human tumors that were defective in degradation of Pea3-ETS factors. Two melanoma patients had de novo DET1 mutations arising after vemurafenib treatment. These observations indicate that MAPK signaling-dependent regulation of Pea3-ETS protein stability is a key signaling node in oncogenesis and therapeutic resistance to MAPK pathway inhibition.
Yuanyuan Xie, Zhen Cao, Elissa W.P. Wong, Youxin Guan, Wenfu Ma, Jenny Q. Zhang, Edward G. Walczak, Devan Murphy, Leili Ran, Inna Sirota, Shangqian Wang, Shipra Shukla, Dong Gao, Simon R.V. Knott, Kenneth Chang, Justin Leu, John Wongvipat, Cristina R. Antonescu, Gregory Hannon, Ping Chi, Yu Chen
Disordered coagulation contributes to death in sepsis and lacks effective treatments. Existing markers of disseminated intravascular coagulation (DIC) reflect its sequelae rather than its causes, delaying diagnosis and treatment. Here we show that disruption of the endothelial Tie2 axis is a sentinel event in septic DIC. Proteomics in septic DIC patients revealed a network involving inflammation and coagulation with the Tie2 antagonist, Angiopoietin-2 (Angpt-2), occupying a central node. Angpt-2 was strongly associated with traditional DIC markers including platelet counts, yet more accurately predicted mortality in two large independent cohorts (combined N = 1077). In endotoxemic mice, reduced Tie2 signaling preceded signs of overt DIC. During this early phase, intravital imaging of microvascular injury revealed excessive fibrin accumulation, a pattern remarkably mimicked by Tie2 deficiency even without inflammation. Conversely, Tie2 activation normalized pro-thrombotic responses by inhibiting endothelial tissue factor and phosphatidylserine exposure. Critically, Tie2 activation had no adverse effects on bleeding. These results mechanistically implicate Tie2 signaling as a central regulator of microvascular thrombus formation in septic DIC and indicate that circulating markers of the Tie2 axis could facilitate earlier diagnosis. Finally, interventions targeting Tie2 may normalize coagulation in inflammatory states while averting the bleeding risks of current DIC therapies.
Sarah J. Higgins, Karen De Ceunynck, John Kellum, Xiuying Chen, Xuesong Gu, Sharjeel A. Chaudhry, Sol Schulman, Towia A. Libermann, Shulin Lu, Nathan I. Shapiro, David C. Christiani, Robert Flaumenhaft, Samir M. Parikh