BACKGROUND. VRC01, a potent, broadly-neutralizing monoclonal antibody, inhibits simian-HIV infection in animal models. HVTN 104 assessed VRC01 safety and pharmacokinetics in humans. We extend the clinical evaluation to determine intravenous-infused VRC01 distribution and protective function at mucosal sites of HIV-1 entry. METHODS. Healthy, HIV-1-uninfected men (n=7) and women (n=5) receiving VRC01 every two months provided mucosal and serum samples once, 4-13 days post-infusion. Eleven male and 8 female HIV-seronegative volunteers provided untreated control samples. VRC01 levels were measured in serum, secretions and tissue, and HIV-1 inhibition was determined in tissue explants. RESULTS. Median VRC01 levels were quantifiable in serum (96.2 µg/ml or 1.3 pg/ng protein), rectal tissue (0.11 pg/ng protein), rectal secretions (0.13 pg/ng protein), vaginal tissue (0.1 pg/ng protein) and cervical secretions (0.44 pg/ng protein) from all recipients. VRC01/IgG ratios in male serum correlated with those in paired rectal tissue (r=0.893, P=0.012) and rectal secretions (r=0.9643, P=0.003). Ex vivo HIV-1Bal26 challenge infected 4/21 rectal explants from VRC01-infused versus 20/22 from controls (P=0.005); HIV-1 Du422.1 infected 20/21 rectal explants of VRC01 recipients and 12/12 from controls (P=0.639). HIV-1Bal26 infected 0/14 vaginal explants of VRC01 recipients compared to 23/28 control explants (P=0.003). CONCLUSION. Intravenous VRC01 distributes into the female genital and male rectal mucosa and retains anti-HIV-1 functionality, inhibiting a highly neutralization-sensitive but not a highly-resistant HIV-1 strain in mucosal tissue. These findings lend insight into VRC01 mucosal infiltration and provide perspective to in vivo protective efficacy. FUNDING. National Institute of Allergy and Infectious Diseases and Bill & Melinda Gates Foundation.
Rena D. Astronomo, Maria P. Lemos, Sandeep R. Narpala, Julie Czartoski, Lamar Ballweber Fleming, Kelly E. Seaton, Madhu Prabhakaran, Yunda Huang, Yiwen Lu, Katharine Westerberg, Lily Zhang, Mary K. Gross, John Hural, Hong-Van Tieu, Lindsey R. Baden, Scott Hammer, Ian Frank, Christina Ochsenbauer, Nicole Grunenberg, Julie E. Ledgerwood, Kenneth Mayer, Georgia Tomaras, Adrian B. McDermott, M. Juliana McElrath
The 12q13-q14 chromosomal region is recurrently amplified in 25% of fusion-positive (FP) rhabdomyosarcoma (RMS) cases and is associated with a poor prognosis. To identify amplified oncogenes in FP RMS, we compared the size, gene composition and expression of 12q13-q14 amplicons in FP RMS with other cancer categories (glioblastoma multiforme, lung adenocarcinoma and liposarcoma) in which 12q13-q14 amplification frequently occurs. We uncovered a 0.2 Mb region that is commonly amplified across these cancers and includes CDK4 and six other genes that are overexpressed in amplicon-positive samples. Additionally, we identified a 0.5 Mb segment that is only recurrently amplified in FP RMS and includes four genes that are overexpressed in amplicon-positive RMS. Among these genes, only SHMT2 was overexpressed at the protein level in an amplicon-positive RMS cell line. SHMT2 knockdown in amplicon- positive RMS cells suppressed growth, transformation and tumorigenesis, whereas overexpression in amplicon-negative RMS cells promoted these phenotypes. High SHMT2 expression reduced sensitivity of FP RMS cells to SHIN1, a direct SHMT2 inhibitor, but sensitized cells to pemetrexed, an inhibitor of the folate cycle. In conclusion, our study demonstrated that SHMT2 contributes to tumorigenesis in FP RMS and that SHMT2 amplification predicts differential response to drugs targeting this metabolic pathway.
Thanh H. Nguyen, Prasantha L. Vemu, Gregory E. Hoy, Salah Boudjadi, Bishwanath Chatterjee, Jack F. Shern, Javed Khan, Wenyue Sun, Frederic G. Barr
Multisystem inflammatory syndrome in children (MIS-C) occurs during, or recently following SARS-CoV-2 infection and is characterized by persistent fever, inflammation and severe illness requiring hospitalization. The majority of MIS-C cases also present with gastrointestinal (GI) symptoms, including abdominal pain, vomiting and diarrhea. In a recent issue of the JCI, Yonker and Gilboa et al., identify zonulin as a biomarker of GI permeability in children with MIS-C, and present the results of an intriguing proof-of-concept study which suggests that zonulin may represent a potential therapeutic target for MIS-C treatment and prevention. Together, these findings suggest that intestinal mucosal dysfunction and epithelial barrier breakdown may represent a biological mechanism underlying the development of MIS-C in SARS-CoV-2-infected children.
Tiffany R. Hensley-McBain, Jennifer A. Manuzak
Broadly reactive antibodies targeting the influenza A virus hemagglutinin (HA) head domain are thought to be rare and to require extensive somatic mutations or unusual structural features to achieve breadth against divergent HA subtypes. Here we describe common genetic and structural features of protective human antibodies from several individuals recognizing the trimer interface (TI) of the influenza A HA head, a recently identified site of vulnerability. We examined the sequence of TI-reactive antibodies, determined crystal structures for TI antibody-antigen complexes, and analyzed the contact residues of the antibodies on HA to discover common genetic and structural features of TI antibodies. Our data reveal that many TI antibodies are encoded by a light chain variable gene segment incorporating a shared somatic mutation. In addition, these antibodies have a shared acidic residue in the heavy chain despite originating from diverse heavy chain variable gene segments. These studies show that the TI region of influenza A HA is a major antigenic site with conserved structural features that are recognized by a common human B cell public clonotype. The canonical nature of this antibody-antigen interaction suggests that the TI epitope might serve as an important new target for structure-based vaccine design.
Seth J. Zost, Jinhui Dong, Iuliia M. Gilchuk, Pavlo Gilchuk, Natalie J. Thornburg, Sandhya Bangaru, Nurgun Kose, Jessica A. Finn, Robin Bombardi, Cinque Soto, Elaine C. Chen, Rachel S. Nargi, Rachel E. Sutton, Ryan P. Irving, Naveenchandra Suryadevara, Jonna B. Westover, Robert H. Carnahan, Hannah L. Turner, Sheng Li, Andrew B. Ward, James E. Crowe Jr.
To the Editor: Povysil G. et al. report that “rare loss-of-function (LOF) variants in type I interferon (IFN) immunity genes are not associated with severe COVID-19” (1). We disagree with the authors’ interpretation of our data and their own (2), for six reasons: 1) Only predicted LOF (pLOF) variants are relevant for comparison between the two studies, because, unlike us, these authors did not test variants experimentally. The relevant proportion in our data is therefore not 24/659=3.5%, but 9/659= 1.36%, whereas theirs is 1/713=0.14%. 2) Our definitions of ‘severe/critical’ patients are different: we defined critical disease as severity grades 6-10 of the WHO scale (3), whereas they restricted their recruitment to grades 7-10 (i.e., excluding patients on high-flow oxygen, considered in our study). Their cohort of ‘mild’ cases may therefore include ‘severe’ COVID-19 cases (grade 6), such as perhaps their ‘mild’ TLR3 pLOF carrier. 3) Their ‘controls’ are subjects from the general population, without depletion of COVID-19 genetic risk factors, whereas we used pauci-/asymptomatic infected subjects (grades 1-3) as ‘controls’. Consequently their power computation in Figure 1 is based on an incorrect hypothesis about the odds ratio, which would be expected to be lower when using general population controls (as they did), than when using pauci- and asymptomatic infected individuals (as we did). 4) The ethnic origin of the patients differs between the two studies: 58% of our 659 patients (and 8 of our 9 pLOF carriers) were European, versus only 10% of their 713 patients with severe disease (and their pLOF carrier is East Asian). 5) Age is a key factor neglected in their comparison: our sample was much younger (mean age: 51.8 years) than theirs (mean: 65.9 years), and seven of our nine pLOF carriers were < 60 years old. We performed a comparison stratified by age (<60/≥60 years), and no significant difference in pLOF proportion was found between the two studies, even ignoring the only patient carrying a pLOF they found (of unknown age): 7/458 in our sample vs. 0/192 in their sample (p=0.11, Fisher’s exact test) for patients <60 years old, and 2/201 vs. 0/521 (p=0.07) for patients ≥60 years old. 6) Finally, and crucially, the authors did not exclude patients with autoantibodies against type I IFN, which account for at least 10% of critical cases and are much more frequent in patients > 60 years of age, particularly men (4).
Qian Zhang, Aurélie Cobat, Paul Bastard, Luigi D. Notarangelo, Helen C. Su, Laurent Abel, Jean-Laurent Casanova
The authors reply: We appreciate the interest of Dr. Zhang and colleagues in our manuscript. The main difference between our publication and that of Zhang et al. (1), was that we assessed all rare predicted loss-of-function variants (pLOFs) meeting the same criteria in cases and controls, which is a well-established paradigm in the field (2). On the other hand, Zhang et al. included specific variants which were experimentally confirmed only in cases, but not controls, precluding a valid case-control comparison. We matched patients as closely as possible to the previous study, and the inclusion of more severe cases (WHO grades 7-10) should only strengthen the signal against population controls. The use of population controls is standard in such settings and has minimal impact on power, because only a small proportion of individuals exposed to SARS-Cov-2 develop severe disease (3). Additionally, for the pLOF model we report adequate power even for an odds ratio of 5.5, which is considerably lower than the one reported by Zhang et al. We tested the same dominant model as Zhang et al., even though LOF variants in these genes have only been reported to cause disease under recessive inheritance (4). We have serious concerns about confounding by ancestry in the analysis by Zhang et al. in which the pLOF carriers were mostly European, but functionally validated missense variants were found in various nationalities from Asia, Europe, Latin America, and the Middle East. Because the rates of pLOFs vary considerably across populations, adjusting for only 3 principal components of ancestry in rare-variant association tests of multi-ethnic cohorts does not provide adequate control for population structure. While we noted that age differences may contribute to the discrepancies between the two studies, Zhang et al. do not discuss the role of age in the interpretation of their results stating: “Inborn errors of TLR3- and IRF7-dependent type I IFN immunity at eight loci were found in as many as 23 patients (3.5%) of various ages (17 to 77 years) and ancestries (various nationalities from Asia, Europe, Latin America, and the Middle East) and in patients of both sexes.” We also note that the patients with autoantibodies were not excluded from the primary analysis by Zhang et al., but this was done only in the post-hoc analysis. Most importantly, our negative findings are in full agreement with the recently published independent study of 586,157 individuals, including 20,952 cases of COVID-19 (4,928 hospitalized and 1,304 requiring ventilation or resulting in death) (5). There were no significant associations with any of the 13 candidate genes examined either individually or in aggregate, or when comparisons included all hospitalized cases or only the most severe cases. Indeed, none of the associations displayed even marginal significance. Therefore, consistent with our study, these findings do not support substantial contributions of inborn errors in type I IFN immunity to COVID-19 severity. These negative results underscore the importance of proper study design, selection of appropriate genetic models, adequate control for genetic ancestry, and adherence to unbiased methods for genetic discovery rather than focusing only on a candidate biological pathway.
Gundula Povysil, Guillaume Butler-Laporte, Ali G. Gharavi, J. Brent Richards, David B. Goldstein, Krzysztof Kiryluk
Patients with neuropathic pain often experience comorbid psychiatric disorders. Cellular plasticity in the anterior cingulate cortex (ACC) is assumed as a critical interface for pain perception and emotion. However, substantial efforts thus far are focused on intracellular mechanisms of plasticity rather than extracellular alterations that might trigger and facilitate intracellular changes. Laminin is a key element of extracellular matrix (ECM) consisting of one α-, β- and γ-chain and implicated in several pathophysiological processes. Here we showed that Laminin β1 (LAMB1) in ACC is significantly downregulated upon peripheral neuropathy. Knocking down ACC LAMB1 exacerbated pain sensitivity and induced anxiety and depression. Mechanistic analysis revealed that loss of LAMB1 causes actin dysregulation via interaction with integrin beta1 and subsequent Src-dependent RhoA/LIMK/cofilin pathway, leading to increased presynaptic transmitter release probability and abnormal postsynaptic spine remodeling, which in turn orchestrates structural and functional plasticity of pyramidal neurons and eventually results in pain hypersensitivity and anxiodepression. This study shed new light on the functional capability of ECM, LAMB1 in modulating pain plasticity and revealed a mechanism that conveys extracellular alterations to intracellular plasticity. Moreover, we identified cingulate LAMB1/integrin β1 as a promising therapeutic strategy for treatment of neuropathic pain and associated anxiodepression.
Zhen-Zhen Li, Wen-Juan Han, Zhi-Chuan Sun, Yun Chen, Jun-Yi Sun, Guo-Hong Cai, Wan-Neng Liu, Tao-Zhi Wang, Yang-Dan Xie, Hong-Hui Mao, Fei Wang, Sui-Bin Ma, Fu-Dong Wang, Rou-Gang Xie, Sheng-Xi Wu, Ceng Luo
Clear Cell Sarcoma (CCS) is a deadly malignancy affecting adolescents and young adults. It is characterized by reciprocal translocations resulting in the expression of the chimeric EWSR1-ATF1 or EWSR1-CREB1 fusion proteins, driving sarcomagenesis. Besides these characteristics, CCS has remained genomically uncharacterized. Copy number analysis of human CCSs showed frequent amplifications of the MITF locus and chromosomes 7 and 8. Few alterations were shared with Ewing sarcoma or desmoplastic small round cell tumors, other EWSR1-rearranged tumors. Exome sequencing in mouse tumors generated by expressing EWSR1-ATF1 from the Rosa26 locus demonstrated no other repeated pathogenic variants. Additionally, we generated a new CCS mouse by Cre-loxP-induced chromosomal translocation between Ewsr1 and Atf1, resulting in copy number loss of chromosome 6 and chromosome 15 instability, including amplification of a portion syntenic with human chromosome 8, surrounding Myc. Additional experiments in the Rosa26 conditional model demonstrated that Mitf or Myc can contribute to sarcomagenesis. Copy number observations in human tumors and genetic experiments in mice render, for the first time, a functional landscape of the CCS genome. These data advance efforts to understand the biology of CCS with innovative models, in which we can eventually validate preclinical therapies, necessary to move toward longer and better survival of the young victims of this disease.
Emanuele Panza, Benjamin B. Ozenberger, Krystal M. Straessler, Jared J. Barrott, Li Li, Yanliang Wang, Mingchao Xie, Anne Boulet, Simon W. A. Titen, Clinton C. Mason, Alexander J. Lazar, Li Ding, Mario R. Capecchi, Kevin B. Jones
Inter-individual immune variability is driven predominantly by environmental factors including exposure to chronic infectious agents such as cytomegalovirus (CMV). We investigated the effects of rhesus CMV (RhCMV) on composition and function of the immune system in young macaques. Within months of infection, RhCMV was associated with impressive changes in antigen presenting cells, T cells, and NK cells — and marked expansion of innate-memory CD8+ T cells. These cells express high levels of NKG2A/C and the IL-2- and IL-15-receptor beta chain, CD122. IL-15 was sufficient to drive differentiation of the cells in vitro and in vivo. Expanded NKG2A/C+CD122+CD8+ T cells in RhCMV-infected macaques, but not their NKG2-negative counterparts, were endowed with cytotoxicity against class I-deficient K562 targets and prompt IFN-ɣ production in response to stimulation with IL-12 and IL-18. Because RhCMV clone 68-1 forms the viral backbone of RhCMV-vectored SIV vaccines, we also investigated immune changes following administration of RhCMV 68-1-vectored SIV vaccines. These vaccines led to impressive expansion of NKG2A/C+CD8+ T cells with capacity to inhibit SIV replication ex vivo. Thus, CMV infection and CMV-vectored vaccination drive expansion of functional innate-like CD8 cells via host IL-15 production, suggesting that innate-memory expansion could be achieved by other vaccine platforms expressing IL-15.
Gema Méndez-Lagares, Ning Chin, W.L. William Chang, Jaewon Lee, Míriam Rosás-Umbert, Hung T. Kieu, David Merriam, Wenze Lu, Sungjin Kim, Lourdes Adamson, Christian Brander, Paul A. Luciw, Peter A. Barry, Dennis J. Hartigan-O’Connor
Transplant recipients were excluded from the initial clinical trials determining safety and efficacy of the landmark COVID-19 vaccines. Further, there is increasing evidence that immunosuppressed transplant recipients have a blunted antibody response to COVID-19 vaccination. In a concerning report by Sattler, et al. in this issue of the JCI, kidney transplant recipients not only lacked a humoral response following two doses of Pfizer BNT162b2, but also displayed substantial impairment of the cellular response to SARS-CoV-2 antigens. This commentary addresses potential strategies for transplant providers to evaluate and augment vaccine immunogenicity given the likelihood that COVID-19 will remain a world-wide threat to the health of transplant recipients.
Peter G. Stock, Timothy J. Henrich, Dorry L. Segev, William A. Werbel
Glioblastoma, the most aggressive brain cancer, recurs because glioblastoma stem cells (GSCs) are resistant to all standard therapies. We showed that GSCs, but not normal astrocytes, are sensitive to lysis by healthy allogeneic natural killer (NK) cells in vitro. Mass cytometry and single cell RNA sequencing of primary tumor samples revealed that glioblastoma-infiltrating NK cells acquired an altered phenotype associated with impaired lytic function relative to matched peripheral blood NK cells from glioblastoma patients or healthy donors. We attributed this immune evasion tactic to direct cell-cell contact between GSCs and NK cells via integrin-mediated TGF-β activation. Treatment of GSC-engrafted mice with allogeneic NK cells in combination with inhibitors of integrin or TGF-β signaling, or with TGFBR2 gene-edited allogeneic NK cells prevented GSC-induced NK cell dysfunction and tumor growth. These findings revealed an important mechanism of NK cell immune evasion by GSCs and implicated the integrin-TGF-β axis as a potentially useful therapeutic target in glioblastoma.
Hila Shaim, Mayra Shanley, Rafet Basar, May Daher, Joy Gumin, Daniel B. Zamler, Nadima Uprety, Fang Wang, Yuefan Huang, Konrad Gabrusiewicz, Qi Miao, Jinzhuang Dou, Abdullah Alsuliman, Lucila N. Kerbauy, Sunil Acharya, Vakul Mohanty, Mayela Mendt, Sufang Li, JunJun Lu, Jun Wei, Natalie W. Fowlkes, Elif Gokdemir, Emily Ensley, Mecit Kaplan, Cynthia Kassab, Li Li, Gonca Ozcan, Pinaki P. Banerjee, Yifei Shen, April L. Gilbert, Corry M. Jones, Mustafa Bdiwi, Ana K. Nunez-Cortes, Enli Liu, Jun Yu, Nobuhiko Imahashi, Luis Muniz-Feliciano, Ye Li, Jian Hu, Giulio Draetta, David Marin, Dihua Yu, Stephan Mielke, Matthias Eyrich, Richard E. Champlin, Ken Chen, Frederick F. Lang, Elizabeth J. Shpall, Amy B. Heimberger, Katayoun Rezvani
Pyridoxine-dependent epilepsy (PDE-ALDH7A1), also known as antiquitin deficiency, is an inborn error of lysine metabolism that presents with refractory epilepsy in newborns. Bi-allelic ALDH7A1 variants lead to deficiency of α-aminoadipic semialdehyde dehydrogenase, resulting in accumulation of piperideine-6-carboxylate (P6C), and secondary deficiency of the important co-factor pyridoxal-5’-phosphate (PLP, active vitamin B6) through its complexation with P6C. Vitamin B6 supplementation resolves epilepsy in patients, but despite this treatment, intellectual disability may occur. Early diagnosis and treatment, preferably based on newborn screening, potentially optimize long-term clinical outcome. However, the currently known diagnostic PDE-ALDH7A1 biomarkers are incompatible with newborn screening procedures. Combining of the innovative analytical methods untargeted metabolomics and infrared ion spectroscopy, we were able to discover a novel biomarker for PDE-ALDH7A1,2S,6S- and 2S,6R-oxopropylpiperidine-2-carboxylic acid (2-OPP), and confirmed 6-oxopiperidine-2-carboxylic acid (6-oxoPIP)as biomarker. We demonstrated the applicability of 2-OPP as a PDE-ALDH7A1 biomarker in newborn screening. Additionally, we showed that 2-OPP accumulates in brain tissue of patients and aldh7a1 knock-out mice, and induced epilepsy-like behavior in a zebrafish model system. We speculate that 2-OPP may contribute to ongoing neurotoxicity, also in treated PDE-ALDH7A1 patients. As 2-OPP formation appears to increase upon ketosis, we emphasize the importance of avoiding catabolism in PDE-ALDH7A1 patients.
Udo F.G. Engelke, Rianne E. van Outersterp, Jona Merx, Fred A.M.G. van Geenen, Arno van Rooij, Giel Berden, Marleen C.D.G. Huigen, Leo A.J. Kluijtmans, Tessa M.A. Peters, Hilal H. Al-Shekaili, Blair R. Leavitt, Erik de Vrieze, Sanne Broekman, Erwin van Wijk, Laura A. Tseng, Purva Kulkarni, Floris P.J.T. Rutjes, Jasmin Mecinovic, Eduard A. Struys, Laura A. Jansen, Sidney M. Gospe, Jr., Saadet Mercimek-Andrews, Keith Hyland, Michel A.A.P. Willemsen, Levinus A. Bok, Clara D.M. Van Karnebeek, Ron A. Wevers, Thomas J. Boltje, Jos Oomens, Jonathan Martens, Karlien L.M. Coene
Disordered lysosomal/autophagy pathways initiate and drive pancreatitis, but the underlying mechanisms and links to disease pathology are poorly understood. Here, we show that mannose-6-phosphate (M6P) pathway of hydrolase delivery to lysosomes critically regulates pancreatic acinar cell cholesterol metabolism. Ablation of the Gnptab gene coding for a key enzyme in M6P pathway disrupted acinar cell cholesterol turnover, causing accumulation of non-esterified cholesterol in lysosomes/autolysosomes, its’ depletion in the plasma membrane, and upregulation of cholesterol synthesis and uptake. We found similar dysregulation of acinar cell cholesterol, and a decrease in GNPTAB levels, in both WT experimental pancreatitis and human disease. The mechanisms mediating pancreatic cholesterol dyshomeostasis in Gnptab-/- and experimental models involve disordered endolysosomal system, resulting in impaired cholesterol transport through lysosomes and blockage of autophagic flux. By contrast, in Gnptab-/- liver the endolysosomal system and cholesterol homeostasis were largely unaffected. Gnptab-/- mice developed spontaneous pancreatitis. Normalization of cholesterol metabolism by pharmacologic means alleviated responses of experimental pancreatitis, particularly trypsinogen activation, the disease hallmark. The results reveal the essential role of M6P pathway in maintaining exocrine pancreas homeostasis and function, and implicate cholesterol disordering in the pathogenesis of pancreatitis.
Olga A. Mareninova, Eszter T. Vegh, Natalia Shalbueva, Carli J.M. Wightman, Dustin L. Dillon, Sudarshan Malla, Yan Xie, Toshimasa Takahashi, Zoltan Rakonczay Jr, Samuel W. French, Herbert Y. Gaisano, Frederick Sanford Gorelick, Stephen J. Pandol, Steven J. Bensinger, Nicholas O. Davidson, David W. Dawson, Ilya Gukovsky, Anna S. Gukovskaya
BACKGROUND. Matrix metalloproteinases (MMPs) are implicated as key regulators of tissue destruction in tuberculosis (TB) and may be a target for host-directed therapy. Here, we conducted a Phase 2 randomized, double-blind, placebo-controlled trial investigating doxycycline, a licensed broad spectrum MMP inhibitor, in pulmonary TB patients. METHODS. Thirty pulmonary TB patients were enrolled within 7 days of initiating anti-TB treatment and randomly assigned to receive either doxycycline 100 mg or placebo twice a day for 14 days in addition to standard care. RESULTS. There were significant changes in the host transcriptome, and suppression of systemic and respiratory markers of tissue destruction with the doxycycline intervention. Whole blood RNA-sequencing demonstrated that doxycycline accelerated restoration of dysregulated gene expression patterns in TB towards normality, with more rapid down-regulation of type I and II interferon and innate immune response genes and concurrent up-regulation of B-cell modules relative to placebo. The effects persisted for 6 weeks after doxycycline was discontinued, concurrent with suppression of plasma MMP-1. In respiratory samples, doxycycline reduced MMP-1, -8, -9, -12 and -13 concentrations, suppressed type I collagen and elastin destruction, and reduced pulmonary cavity volume despite unchanged sputum Mycobacterium tuberculosis loads between the study arms. Two weeks of adjunctive doxycycline with standard anti-TB treatment was well-tolerated, with no serious adverse events related to doxycycline. CONCLUSION. These data demonstrate that adjunctive doxycycline with standard anti-TB treatment suppresses pathological MMPs in pulmonary tuberculosis patients, and suggest that larger studies on adjunctive doxycycline to limit immunopathology in TB are merited.
Qing Hao Miow, Andres F. Vallejo, Yu Wang, Jia Mei Hong, Chen Bai, Felicia S.W. Teo, Alvin Dingyuan Wang, Hong Rong Loh, Tuan Zea Tan, Ying Ding, Hoi Wah She, Suay Hong Gan, Nicholas I. Paton, Josephine Lum, Alicia Tay, Cynthia B.E. Chee, Paul A. Tambyah, Marta E. Polak, Yee Tang Wang, Amit Singhal, Paul Elkington, Jon S. Friedland, Catherine W.M. Ong
Tuberculosis (TB) is a persistent global pandemic and standard treatment has not changed for thirty years. Mycobacterium tuberculosis (Mtb) has undergone prolonged co-evolution with humans, and patients can control Mtb even after extensive infection, demonstrating the fine balance between protective and pathological host responses within infected granulomas. We hypothesised that whole transcriptome analysis of human TB granulomas isolated by laser capture microdissection could identify therapeutic targets, and that comparison with a non-infectious granulomatous disease, sarcoidosis, would identify disease-specific pathological mechanisms. Bioinformatic analysis of RNAseq data identified numerous shared pathways between TB and sarcoidosis lymph nodes, and also specific clusters demonstrating TB results from a dysregulated inflammatory immune response. To translate these insights, we compared three primary human cell culture models at the whole transcriptome level, and demonstrated that the 3D collagen granuloma model most closely reflected human TB disease. We investigated shared signaling pathways with human disease and identified twelve intracellular enzymes as potential therapeutic targets. Sphingosine kinase 1 inhibition controlled Mtb growth, concurrently reducing intracellular pH in infected monocytes and suppressing inflammatory mediator secretion. Immunohistochemical staining confirmed that sphingosine kinase 1 is expressed in human lung TB granulomas, and therefore represents a host therapeutic target to improve TB outcomes.
Michaela T. Reichmann, Liku B. Tezera, Andres F. Vallejo, Milica Vukmirovic, Rui Xiao, James Reynolds, Sanjay Jogai, Susan Wilson, Ben Marshall, Mark G. Jones, Alasdair Leslie, Jeanine M. D'Armiento, Naftali Kaminski, Marta E. Polak, Paul Elkington
After extensive exposure to Mycobacterium tuberculosis (Mtb), most individuals acquire latent Mtb infection (LTBI) defined by a positive tuberculin skin test (TST) or interferon-γ release assay (IGRA). To identify mechanisms of resistance to Mtb infection, we compared transcriptional profiles from highly-exposed contacts who resist TST/IGRA conversion (resisters, RSTRs) and controls with LTBI using RNAseq. Gene sets related to carbon metabolism and free fatty acid (FFA) transcriptional responses enriched across two independent cohorts suggesting RSTR and LTBI monocytes have distinct activation states. We compared intracellular Mtb replication in macrophages treated with FFAs and found that palmitic acid (PA), but not oleic acid (OA), enhanced Mtb intracellular growth. This PA activity correlated with its inhibition of pro-inflammatory cytokines in Mtb-infected cells. Mtb growth restriction in PA-treated macrophages was restored by activation of AMP kinase (AMPK), a central host metabolic regulator known to be inhibited by PA. Finally, we genotyped AMPK variants and found seven SNPs in PRKAG2, which encodes the AMPKγ subunit, that strongly associated with RSTR status. Taken together, RSTR and LTBI phenotypes are distinguished by FFA transcriptional programs and by genetic variation in a central metabolic regulator, which suggests immunometabolic pathways regulate TST/IGRA conversion.
Jason D. Simmons, Phu T. Van, Catherine M. Stein, Violet Chihota, Thobani Ntshiqa, Pholo Maenetje, Glenna J. Peterson, Anthony Reynolds, Penelope Benchek, Kavindhran Velen, Katherine L. Fielding, Alison D. Grant, Andrew D. Graustein, Felicia K. Nguyen, Chetan Seshadri, Raphael Gottardo, Harriet Mayanja-Kizza, Robert S. Wallis, Gavin Churchyard, W. Henry Boom, Thomas R. Hawn
As a result of impressive increases in our knowledge of rodent and human immunology, the pathophysiological mechanisms underlying graft-versus-host disease (GVHD) have dramatically improved in the past 15 years. Despite improved knowledge, translation to clinical care has not proceeded rapidly, with results from experimental models being inconsistent in their ability to predict the clinical utility of new therapeutic agents. In parallel, new tools in immunology have allowed in depth analyses of the human system and have been recently been applied in the field of clinical GVHD. Notwithstanding these advances, there is a relative paucity of mechanistic insights into human translational research, and this remains an area of high unmet need. Here we review selected recent advances both in preclinical experimental transplantation and translational human studies, including new insights into human immunology, the microbiome, and regenerative medicine. We focus on the fact that both approaches can interactively improve our understanding of both acute and chronic GVHD biology and open the door to improved therapeutics and successes.
Gérard Socié, Leslie S. Kean, Robert Zeiser, Bruce R. Blazar
Novel mRNA-based vaccines have been proven powerful tools to combat the global pandemic caused by SARS-CoV2 with BNT162b2 (trade name: Comirnaty) efficiently protecting individuals from COVID-19 across a broad age range. Still, it remains largely unknown how renal insufficiency and immunosuppressive medication affect development of vaccine induced immunity. We therefore comprehensively analyzed humoral and cellular responses in kidney transplant recipients after the standard second vaccination dose. As opposed to all healthy vaccinees and the majority of hemodialysis patients, only 4/39 and 1/39 transplanted individuals showed IgA and IgG seroconversion at day 8 ± 1 after booster immunization with minor changes until day 23 ± 5, respectively. Although most transplanted patients mounted spike-specific T helper cell responses, frequencies were significantly reduced compared to controls and dialysis patients, accompanied by a broad impairment in effector cytokine production, memory differentiation and activation-related signatures. Spike-specific CD8+ T cell responses were less abundant than their CD4+ counterparts in healthy controls and hemodialysis patients and almost undetectable in transplant patients. Promotion of anti-HLA antibodies or acute rejection was not detected after vaccination. In summary, our data strongly suggest revised vaccination approaches in immunosuppressed patients, including individual immune monitoring for protection of this vulnerable group at risk to develop severe COVID-19.
Arne Sattler, Eva Schrezenmeier, Ulrike A. Weber, Alexander Potekhin, Friederike Bachmann, Henriette Straub-Hohenbleicher, Klemens Budde, Elena Storz, Vanessa Proß, Yasmin Bergmann, Linda M.L. Thole, Caroline Tizian, Oliver Hölsken, Andreas Diefenbach, Hubert Schrezenmeier, Bernd Jahrsdörfer, Tomasz Zemojtel, Katharina Jechow, Christian Conrad, Sören Lukassen, Diana Stauch, Nils Lachmann, Mira Choi, Fabian Halleck, Katja Kotsch
Inhibitors of mPges-1 are in the early phase of clinical development. Deletion of mPges-1 in mice confers analgesia, restrains atherogenesis and fails to accelerate thrombogenesis, while suppressing PGE2, but increasing biosynthesis of PGI2. In Ldlr-/- mice, this last effect represents the dominant mechanism by which mPges-1 deletion restrains thrombogenesis, while suppression of PGE2 accounts for its anti-atherogenic effect. However, the impact of mPges-1 depletion on BP in this setting remains unknown. Here, mPges-1 depletion significantly increased the BP response to salt loading in male Ldlr-/- mice, whereas, despite the direct vasodilator properties of PGI2, Ipr deletion suppressed it. Furthermore, combined deletion of the Ipr abrogated the exaggerated BP response in male mPges-1-/- mice. Interestingly, these unexpected BP phenotypes were not observed in female mice fed a high salt diet. This is attributable to the protective effect of estrogen in Ldlr-/- mice and in Ipr-/- /Ldlr-/- mice. Thus, estrogen compensates for a deficiency in PGI2 to maintain BP homeostasis in response to high salt in hyperlipidemic female mice. In males, by contrast, augmented formation of ANP plays a similar compensatory role, restraining hypertension and oxidant stress in the setting of Ipr depletion. Hyperlipidemic males on a high salt diet might be at risk of a hypertensive response to mPGES-1 inhibitors.
Soon Y. Tang, Hu Meng, Seán T. Anderson, Dimitra Sarantopoulou, Soumita Ghosh, Nicholas F. Lahens, Katherine N. Theken, Emanuela Ricciotti, Elizabeth J. Hennessy, Vincent Tu, Kyle Bittinger, Aalim M. Weljie, Gregory R. Grant, Garret A. FitzGerald
Androgen receptor (AR)-positive prostate cancers (PCa) and estrogen receptor (ER)-positive luminal breast cancers (BCa) are generally less responsive to immunotherapy compared to certain tumor types such as melanoma. However, the underlying mechanisms are not fully elucidated. Here we found that FOXA1 overexpression inversely correlated with interferon (IFN) signature and antigen presentation gene expression in PCa and BCa patients. FOXA1 bound STAT2 DNA binding domain and suppressed STAT2 DNA binding activity, IFN signaling gene expression and cancer immune response independently of the transactivation activity of FOXA1 and its mutations detected in prostate and breast cancers. Increased FOXA1 expression promoted cancer immuno- and chemotherapy resistance in mice and PCa and BCa patients. These findings were also validated in bladder cancer expressing high level FOXA1. FOXA1 overexpression could be a prognostic factor to predict therapy resistance and a viable target to sensitize luminal prostate, breast and bladder cancer to immuno- and chemotherapy.
Yundong He, Liguo Wang, Ting Wei, Yu-Tian Xiao, Haoyue Sheng, Hengchuan Su, Daniel P. Hollern, Xiaoling Zhang, Jian Ma, Simeng Wen, Hongyan Xie, Yuqian Yan, Yunqian Pan, Xiaonan Hou, Xiaojia Tang, Vera J. Suman, Jodi M. Carter, Richard Weinshilboum, Liewei Wang, Krishna R. Kalari, Saravut J. Weroha, Alan H. Bryce, Judy C. Boughey, Haidong Dong, Charles M. Perou, Dingwei Ye, Matthew P. Goetz, Shancheng Ren, Haojie Huang