Immunology & cell biology of bacterial infections

Description

Our research within this program focuses on key aspects of host pathogen interactions, particularly in mycobacterial infections, and on anti-microbial resistance. Several clusters of scientists work together to unravel fundamental mechanisms of host defence against the responsible pathogens, using advanced immunomonitoring and other advanced approaches to identify and refine biomarker signatures of infection, disease risk, disease activity and protective immunity, as well as the design of novel therapies that target drug-resistant bacterial pathogens. This work provides the translational basis for the development of diagnostics, novel host-directed therapies, immune modulatory and antibacterial molecules strategies as well as novel vaccines.

Our clinical research activities focus particularly on 1) host defence against infectious diseases in the immunocompromised host (genetic and acquired immunodeficiencies, due to HIV, treatment with biologicals or other immunosuppressants); 2) on clinical vaccination studies (tuberculosis, leprosy, malaria, yellow fever, rabies, Ebola, and vaccination in immunosuppressed individuals); 3) on antibiotic stewardship in empirical treatment strategies; 4) on experimental human infection models, and 5) on tropical and travel medicine.

The cutting-edge approaches developed by the Department’s basic scientific research are used in cross sectional and prospective (endemic) population field studies to unravel the immunological and molecular basis of susceptibility to the mentioned poverty-related diseases of major public health significance. In multiple, long-term international collaborations with excellent research institutes in countries where these diseases are endemic, the emergence of non-communicable diseases and their interactions with infectious diseases is also being studied, with a focus on the emerging epidemic of type 2 diabetes.

All research activities are embedded in the interdepartmental Center of Infectious Diseases (CID) and in the LUMC Profiling Area “Immunity, Infection and Tolerance” (IIT).

Aim and focus

Our program encompasses the following interrelated research themes:

The theme immunodeficiencies is a clinically important research theme, and focuses on the identification and treatment of genetic immunodeficiencies, using immunological and molecular-genetic diagnostic technologies, as well as research into acquired immunodeficiencies, in patients eligible for treatment with immunosuppressants or monoclonal antibodies (“biologicals”). These efforts are directly related to the TRF “immunodeficiency” of the Department. The department has long term expertise in the screening for latent Mycobacterium tuberculosis infection, an important risk factor in the immunocompromised, particularly in those with iatrogenic immunosuppression. In addition, our vaccination and travel clinic provides optimized vaccination and counselling schemes to such patients with suppressed immunity. Complementary research at the vaccination clinic includes the study of the efficacy of (modified) active immunisation schedules to induce protective immunity in the immunosuppressed traveller. The vaccination clinic also explores the efficacy and safety of alternative routes of vaccine administration.

Tuberculosis research is a major theme in the research program. Tuberculosis (TB) is a tremendous global public health problem: over 10 million people develop active TB and 1.8 million die from TB every year (WHO report 2016).  Around one fourth of the world’s population is latently infected with Mtb, of which 5-10% will go on to develop active TB during their lifetime. The past two centuries over a billion people have died from TB, more than from any other infectious disease.  There is no effective vaccine – BCG affords only partial protection and has limited impact on TB transmission, urging for better vaccines.  Current TB diagnostic tools are insufficient and lack sensitivity and specificity for differential diagnosis of TB, and adequate tools to predict treatment outcome at an early stage are lacking.  TB drug resistance is a rapidly increasing problem, calling urgently for novel treatments.  There are no current “Correlate of Risk” tests that can predict who of those infeed with Mtb will proceed to developing active TB disease. Current efforts will fail to achieve the WHO ambition to eliminate this disease before 2050. Our TB research responds to these challenges, and is clinical, translational and fundamental in nature and including humanized animal models of TB (mice, zebrafish). Research efforts aim to develop better vaccines and diagnostic, prognostic and therapy monitoring, “biomarker signatures”. Our vaccine program has led to a number of first-in-man clinical studies with newly developed, molecularly defined synthetic TB vaccines. Systems biology, chemical genetics, transcriptomics, metabolomics, lipidomics and immunomics approaches are used to identify the key cellular signalling pathways in host defence to intracellular pathogens in general, and to M. tuberculosis, Salmonellae, and MRSA in particular. Identified key host regulatory networks and molecules will be used as targets for host-directed therapy to develop alternatives for antibiotic treatment to combat the rise in drug resistant pathogens. Correlates of protection and prospective correlates of risk of developing TB are being identified. Novel immunological subsets (both innate and adaptive) have also been identified and provide novel insights which are instrumental for development of novel vaccines and treatment strategies. Using technologies developed in the TB research program, recent work also enabled identification of biomarkers of Ebola-vaccine induced human responses. Moreover, there is increasing interest in the areas of co-infections (helminth infections; HIV) and co-morbidities such as type-2 diabetes, a major newly identified risk factor for TB. Extensive international collaborative networks have been established, which include leading partners from EU, US and TB endemic areas/countries (South Africa, Gambia, Ethiopia, Indonesia, China, Peru, Brazil, S Korea, etc.). Multiplex, POC diagnostic tests are being developed to allow diagnosis of active TB in field settings as well as to monitor treatment responses and efficacy by “personalised treatment monitoring”.

We anticipate to establish a ‘TB Research Center of Excellence (TBRC)’ to enhance collaboration, visibility and permit rapid and efficient implementation of novel technologies in the design of TB therapies, vaccines and diagnostic tools. TBRC will harness, integrate and synergize the uniquely available knowledge and cutting-edge technologies at LUMC, Univ. Leiden (LACDR, LIC, IBL, CARES), RIVM and KNCV to create a highly innovative, trans-disciplinary TB research community.  TBRC aims to discover and develop innovative vaccines, biomarkers, diagnostics and therapies for mycobacterial infectious diseases.  TBRC’s integration of fundamental, preclinical, translational, clinical and public health research is unique in the world, and its model is translatable to other infectious diseases.  TBRC is linked to national programmes including NCOH and NADP.

Leprosy research has been a major research theme at the LUMC since the 1970s. This resulted in the identification of the first HLA-disease associations, the first human helper and regulatory T-cell clones, followed by the identification of M.leprae unique antigens that activated those cells.  Besides the M.leprae unique antigens, we currently  apply the identification of host immune, metabolic and transcriptomic biomarker profiles for early diagnosis of infection and disease to develop diagnostic tests.  Moreover, these biomarkers are used to investigate nerve damaging mechanisms and prospective correlates of risk for leprosy reactions (“flares”). Collaborations with endemic field sites within the three main continents reporting leprosy are firmly established (Brazil, Bangladesh, Ethiopia, Nepal, India) exemplified by a large scale BCG vaccination field trial in Bangladesh. In collaboration with the LUMC Dept. Molecular Cell Biology, user-friendly, field-applicable, first generation lateral flow tests have been developed based on multiple discriminating host biomarkers and the mentioned M. leprae unique antigens. Since leprosy is considered a model disease to study human immuneregulation and inflammation, test development is now also being extended to other chronic diseases (rheumatoid arthritis and Crohn’s disease) for which POC tests allowing treatment monitoring may also be beneficial.  Recent new projects include additional focus on mechanisms of mycobacterial transmission using a One health approach (human, animal, environment). Our Department is part of the Leprosy Expertise Center (housed at EMC), functions as the national reference centre for routine serological diagnosis of leprosy, and provides this service also to European partners. In addition, the group has established a large biobank of leprosy patients’ samples including sera, cells and urine.

Research on malaria vaccines, in collaboration with the Department of Parasitology (LUMC) and Radboud University Medical Center, focuses on challenging volunteers with live Plasmodium falciparum parasites to follow the response to immunisation under chemoprophylaxis (“vaccination”) and infection. Detailed immunological and transcriptomic “biomarker signatures” are being determined to measure vaccine responses using platform technologies (dcRT-MLPA) developed for TB and other diseases with the purpose of advanced immunomonitoring. In order to focus our research portfolio more on bacterial infectious diseases, this research line is now fully embedded within the Dept of Parasitology (LUMC).

Research on antimicrobial peptides aims to develop alternatives for antibiotic treatment in the face of the emerging antimicrobial resistance and tolerance. Naturally occurring peptides are selected as lead structures which are further optimized for application and efficacy. Several human 3-D models, such as skin, airway epithelium and bladder epithelium and animal models are used to study and select optimal variants peptides. Peptides are being developed for topical applications, such as treatment of MRSA carriage, wound infections, biomaterial-associated infections and urinary tract infections with highly resistant microbes. The next stage in the antimicrobial peptide research is the development of innovative formulations that promotes the controlled release of the peptide at the site of infection. A first study with synthetic antimicrobial peptide in patients with chronic suppurative otitis media has been successfully completed (Peek et al, 2017). Further proof of concept studies in man are being planned for 2018.  Based on our experience and contacts a center of expertise on antimicrobial peptides (Leiden Antimicrobial Peptide Platform (LAPP) has been initiated. This research line is embedded in NCOH, CARES and NADP initiatives.

Recently, our Dept. has been involved in researching infections due to Clostridium difficile, a debilitating pathogen especially among the elderly with chronic co-morbidities.  Studies have been initiated in collaboration with the Dept.  of Medical Microbiology and the Dutch Donor Faeces Bank to enable faecal transplantations in patients with relapsing CDI or in patients with recurrent urinary tract infections with multi-resistant Gram-negative bacteria. The latter fits very well with our TRF profile of “complicated urinary tract infections”.

Research on Antimicrobial stewardship / Choices in empiric antimicrobial treatment strategies

A set of projects are performed on the topic of selecting optimal empiric antibacterial- and antifungal therapy strategies.  The projects directly relate to- and are guided by current clinical questions that arise with the emergence of AMR and antifungal resistance. Empiric and prophylactic treatments in particular are affected in their efficacy by an increasing incidence of AMR, and thus relevant to large groups of patients. Hence it is also a societal challenge.
With regard to antibiotic resistance, determinants that allow early de-escalation of antimicrobial therapy in cases with suspected sepsis are being identified. Furthermore, a standardized evaluation method for the need of escalating antimicrobial treatment policy on an institutional scale is developed. In addition, a separate project has been set up to explore and study the ethics of escalating empirical sepsis therapy on institutional and national levels, and to identify socio-behavioural factors that influence antibiotic prescription patterns. Another example is the study of optimal treatment of prosthetic joint infections - the research includes the study of the early effect of specific antibiotics (in vivo and in vitro) and antimicrobial peptides (in vitro) on biofilm, which is crucial for the treatment of prosthetic joint infection.      
Increasing resistance is also a problem when selecting optimal treatment for fungal infections. After performing several complex epidemiological studies, a mathematical model is being constructed to estimate the clinical impact of antifungal resistance and to support treatment decisions. This, and previous research projects by the department on the topic of personalizing antifungal treatment according to immunological markers, instigated the formation of the Leiden Fungal Infections Study Group (LFI-SG). By collaboration of the LFI-SG in the Dutch-Belgium Mycosis Study Group, access to participation in-, and roll out of future high quality research on this topic is pursued.                      
Connections with more basic research are developed with the research lines named above of antimicrobial peptides (alternative treatments, anticipated clinical trial in 2018) and e.g. Leprosy (biomarker profiles in suspected bacteraemia, in a submitted ZonMW grant application). Development of this clinical research theme benefits from the Department’s positions held in regional and national committees that govern policy making on antimicrobial treatment (e.g. SWAB, the national working party on antibiotic policy)..

Position in international context

The Department has extensive multinational and multilateral collaborative research- as well as operational- programs with laboratories and field sites worldwide.

Numerous ongoing international collaborations exist with amongst others: Hazel Dockrell (London School of Hygiene and Tropical Medicine, UK), Peter Andersen (Statens Serum Institute, Denmark), Stefan Kaufmann, (Max Planck Institute for Infection Biology, Germany), Gary Schoolnik (Stanford University School of Medicine, USA), Gerhard Walzl (Stellenbosch University , South Africa), Thomas Scriba (University of Cape Town, South Africa), Helen McShane (University of Oxford, UK), Delia Goletti (INMI, Italy), Jayne Sutherland (MRC, The Gambia) many other TBVI researchers (www.tbvi.eu), Robert Modlin (David Geffen School of Medicine, Univ. of California, Los Angeles),  Stewart Cole (Ecole Polytechnique Féderale Lausanne and Pasteur Institute Paris), Bob Hancock (Univ. British Columbia), David Pritchard (Univ. Nottingham), Jeff Tomberlin (Texas A&M Univ.), Karl Lohner (Univ. Graz), Karin Thevissen (Univ. Leuven), Galit Alter, Sarah Fortune (Harvard Medical School, Boston).

Ongoing national collaborations exist with amongst others: Robert Sauerwein, Reinout van Crevel and Mihai Netea (Radboud UMC, Nijmegen), Herman Spaink and Annemarie Meijer (Institute of Biology, Leiden), Jacques Neefjes and Huib Ovaa (Chemical Immunology, LUMC), Hermen Overkleeft and Mario van der Stelt (Leiden Institute Chemistry, Leiden), Gilles van Wezel and Ariane Briegel (Institute of Biology, Leiden), Bas Zaat (Amsterdam Medical Center), Willem van Wamel (Erasmus Medical Center), Jan Hendrik Richardus (Erasmus Medical Center), Colette van Hees (Lepra Expertise Centrum; Erasmus Medical Center), Esther Middelkoop (VUMC),  Gijs van der Marel (LIC) and Joke Bouwstra and Wim Jiskoot (LACDR) Robert Rissmann, Koos Burggraaf (CHDR), Remko van Leeuwen (Madam TRX).

The Department’s laboratories have acquired many national and international projects during the past 15 years, including from The Bill & Melinda Gates Foundation, NIH, NWO- and KNAW-program grants, industry-research grants, STW/TTW grants, Netherland’s Leprosy Relief Foundation (currently Leprosy Research Initiative) project grants, TIP-Pharma, TSK Health Impulse, Novartis Foundation, Turing Foundation, EU- and IMI2 and a series of EDCTP grants. We have recently been awarded a prestigious LEAD fellow in the first round of applications.

Content / highlights / achievements

1. Zak DE et al. and other members of the ACS and GC6-74 cohort study teams. Prospective blood RNA signatures of tuberculosis disease risk. Lancet. 2016 Jun 4;387(10035):2312-22 – IF 47·831
2. Joosten SA et al. Patients with tuberculosis have a dysfunctional circulating B-cell compartment, which normalizes following successful treatment.  PLoS Pathogens 2016 Jun 15;12(6):e1005687 – IF 6.608
3. Coppola M et al. New Genome-Wide Algorithm Identifies Novel In-Vivo Expressed Mycobacterium tuberculosis Antigens Inducing Human T-Cell Responses with Classical and Unconventional Cytokine Profiles. Scientific Reports, 2016 Nov 28;6:37793 – IF 4.259
4. van Meijgaarden KE et al. Human CD8+ T-cells recognizing peptides from Mycobacterium tuberculosis (Mtb) presented by HLA-E have an unorthodox Th2-like, multifunctional, Mtb inhibitory phenotype and represent a novel human T-cell subset. PLoS Pathogens 2015 Mar 24;11(3):e1004671. – IF 6.608
5. Ottenhoff THM et al. Genome-wide Expression Profiling Identifies Novel Interferon Response Pathways In Active Mycobacterium tuberculosis Infection. PLoS One. 2012;7(9):e45839 – IF 2.806
6. Kaforou M et al. on behalf of the ILULU consortium. Detection of tuberculosis in HIV-infected and -uninfected African adults using whole blood RNA expression signatures: a case-control study. PLoS Med 2013 10(10): e1001538 – IF 11.862 
7. de Breij A et al. The antimicrobial peptide SAAP-148 Combats Drug-Resistant Bacteria and Biofilms. Sci Transl Med, accepted for publication. 2017 – IF 15,84
8. van Hooij A et al. Field-friendly serological tests for determination of M. leprae-specific antibodies. Scientific Reports 2017 7(1):8868 – IF 4.259
9. van Hooij, A et al. Quantitative lateral flow strip assays as user-friendly tools to detect biomarker profiles for leprosy.Scientific Reports 2016 6, 34260 – IF 4.259
10. Tran TM et al. Transcriptomic evidence for modulation of host inflammatory responses during febrile Plasmodium falciparum malaria. Sci Rep. 2016 Aug 10;6:31291 – IF 4.259

List of the unit’s five most important societal publications and/or other societal outputs in the past six years (minimum of 5, maximum of 10 publications/outputs)

1. Arend SM et al. Antigenic equivalence of human T-cell responses to Mycobacterium tuberculosis-specific RD1-encoded protein antigens ESAT-6 and culture filtrate protein 10 and to mixtures of synthetic peptides. Infect Immun 2000 Jun;68(6):3314-21. Basic study that was the start of the development of the IGRA test for LTBI
2. Zak DE et al. and other members of the ACS and GC6-74 cohort study teams. Prospective blood RNA signatures of tuberculosis disease risk. Lancet. 2016 Jun 4;387(10035):2312-22. First biomarker signature identifying individuals at risk of devolping TB
3. Roset Bahmanyar, E et al. Leprosy diagnostic test development as a prerequisite towards elimination: requirements from the user’s perspective. PLoS Negl Trop Dis 2016; 10, e0004331. => TPP for a diagnostic test for leprosy for application in PEP treatment in health programs
4. Corstjens, PLAM et al. Field-friendly test for monitoring multiple immune response markers during onset and treatment of exacerbated immunity in leprosy. Clin.Vaccine Immunol 2016; 23 (6):515-519. Basis for POC diagnostic test for diagnosis of flares
5. Visser LG1, Roukens AH. Modelling a way out of yellow fever. Lancet. 2016 Dec 10;388(10062):2847-2848
6. Comment on “17DD yellow fever vaccine: a double blind, randomized clinical trial of immunogenicity and safety on a dose-response study”. [Hum Vaccin Immunother. 2013], “Intradermally administered yellow fever vaccine at reduced dose induces a protective immune response: a randomized controlled non-inferiority trial”. [PLoS One. 2008] and Fractional dosing of yellow fever vaccine to extend supply: a modelling study. [Lancet. 2016]Our study (2008) was one of the two pivotal studies on which the WHO policy of fractional dose yellow fever vaccination was based to allow massive preventive vaccination of over 6 million population to curb the yellow fever outbreak in Angola and DRC (2016).

Future themes

Our future vision is to focus the clinical research of the Department increasingly on our TRFs, notably the prevention and treatment of recurrent and chronic infections, in particular those in the immunocompromised host, or because of infections with multi-resistant microorganisms refractory to standard treatments. We want to strengthen our TRF in these areas by continued innovative research efforts aimed towards developing novel vaccination strategies and anti-microbial treatments and diagnostic tests. This builds on top of our primary central research theme, notably to gain insight in key aspects of the immunology and cell biology of chronic and   recurrent bacterial and parasitic infectious diseases through advanced immunological monitoring to innovate the development of vaccines, diagnostic tests and treatment.

Our research efforts in the area of vaccination will continue to cover and integrate both fundamental aspects of vaccination, such as antigen discovery, development and testing of candidate vaccines in preclinical animal models, with translational efforts including discovery of immune markers for evaluation of immunogenicity and vaccine efficacy, all the way to the execution of innovative clinical trials in humans. Current studies focus mostly on tuberculosis, leprosy, Ebola and malaria. The aim is to develop molecularly and genetically characterized novel vaccines that are superior to, and safer than current vaccines (if available at all) in pre- and post-infection prophylaxis for major chronic and recurrent human infectious diseases. Our Department has a longstanding strong international position in these fields, particularly in poverty related and Neglected Diseases.

A second, more recent ambition is the development of innovative therapies and better clinical strategies to combat infectious diseases, specifically those that are caused by drug resistant organisms and infections refractory to treatments for other reasons. This is particularly relevant because of the increasing burden of antimicrobial resistance that makes even more common infections sometimes difficult to treat.

The research spans from basic research which includes the discovery of anti-microbial peptides with broad activity against major human pathogens; innovative host directed therapies in which host molecules are modulated by chemical genetic strategies to optimize host defense to control infection; to testing and optimizing new antibiotic regimens and anti-microbial peptides in human clinical efficacy trials. Efforts are directed particularly but not exclusively to tuberculosis, salmonellosis, MRSA, urinary tract infections caused by multi-resistant bacteria and the use of specific antibodies against amongst others C. difficile. The Department aims to strengthen and expand its position in fundamental and translational immunology and immunogenetics, including clinical implementation of discoveries and application of new biomarkers, also to more generally occurring infectious diseases such as pyelonephritis and pneumonia in the compromised host through immunosuppression or advanced age. Main areas in which clinical output is pursued are novel treatments, vaccination and diagnostic strategies, point-of-care testing and immune modulation.

A key goal of the Dept. will also be to facilitate the establishment of TBRC which we consider to be a unique asset to the Dept. and the Center of Infectious Diseases, with a huge national and international visibility and profile.
Finally, we want to further integrate research, education and patient care within the Center of Infectious Diseases.

Cohesion within LUMC

The research groups in the Dept. play key roles in the LUMC “Research Profile” of “Immunity, Infection and Tolerance " and in the Center for Infectious Diseases together with the Department of Medical Microbiology and Department of Parasitology. It is well embedded within the overall LUMC profiling areas. The Department collaborates with numerous partners within LUMC (e.g. Departments of Chemical Immunology, Molecular Cell Biology, Pediatrics, Parasitology, Dermatology, Immunohematology and Blood Transfusion, Pharmacy, Pulmonary Diseases, Molecular Epidemiology, the Center for Proteomics and Metabolomics, Endocrinology/Internal Medicine, Clinical Oncology etc.) as well as Leiden University (LIC, CARES, IBL). It has a longstanding high international reputation and is involved in numerous collaborative networks, as outlined above in section 1. Below we provide several relevant examples. 

Infection,Immunity and Tolerance
Collaboration with other research programs within the biomedical research profile “Infection, Immunity and Tolerance” is actively pursued by 1) plans to establish a TBRC (see above), which will integrate activities of many Depts. inside and outside of LUMC around TB research and clinical application; 2) studying biomarkers of disease progression found in leprosy (flares) for their predictive role in patients with early arthritis (rheumatology, program code 02.07.01) ) and Crohn’s disease (MDL); 3) initiating collaborative research on the microbiome of gut, bladder, nose and skin to open ways for innovative treatment strategies to resolve recurrent infections with multi-resistant pathogens such as through controlled recolonization (medical microbiology, program code 4.06.02).

Our department is strongly committed to support the department of Parasitology in realizing a center for controlled human infections, which would give the LUMC a head start in evaluating new vaccines and in exploratory research on host-pathogen interactions under extremely well controlled conditions.

Collaboration with the Neefjes and Ovaa, Overkleeft labs (LUMC and UL) in screening of FDA approved compound libraries for their ability to control drug resistant intracellular infections, as well as synthesizing and screening libraries of new compounds. Many innovative leads have been identified (NWO-TOP and TTW projects)
Together with the department of Medical Microbiology (Prof dr E. Kuijper) we participate in the interdisciplinary research network CARES (coordinated by Gillis van Wezel, Leiden University). CARES is the center of excellence in the development of novel antimicrobial agents and strategies and is engaged as partner in NCOH, NADP and international collaborative research projects. Collaborative research between LUMC, CARES and CHDR offers unique opportunities for the development and evaluation of new antimicrobial compounds to this aim.

Cancer pathogenesis and Therapy: In a joint collaboration with the department of Epidemiology and Hematology we are studying the risk profile for invasive aspergillosis, validating stopping rules, modeling the consequences of increasing azole resistance and investigating the role of T-cell immunity in fungal infections (supervision Dr. Mark de Boer, Dr. H. Jolink, R. van de Peppel) (Hematology, program code 4.01.03).