RESEARCH COLLABORATION - BIOMARKERS IN THE FIGHT AGAINST INFECTIOUS DISEASES
Biomarkers in the Development of Diagnostic Assays
Projects D1 - D7 of the research consortium mainly focus on the development of biomarker based diagnostic (D) detection systems.
Within this overall focus on diagnostics, projects D1 – D4 address the topics of verification, differentiation and targeted treatment of bacterial and fungal infections.
Project D1 (Prof. Dr. Dr. Gessner, University of Regensburg / Hyglos GmbH) looks into bacterial permeability increasing protein (BPI) as a novel biomarker for tests allowing a fast distinction between bacterial and viral infections. This distinction prior to complex serological, molecular and cell-culture-based tests, would, especially in cases of major, life-threatening infections, result in improved therapeutic approaches to the benefit of patients. The sensitive and reliable detection of BPI, which is released acutely from neutrophil granulocytes after the contact with bacteria, could serve as a new biomarker for the detection of systemic bacterial infections. Due to its selectivity against bacterial pathogens, BPI could replace or supplement other biomarkers such as CRP.
Projects D2 and D3 investigate the possibilities of a serological distinction of related pathogens to allow for a faster and more precise therapy as is currently possible. Project D2 (Prof. Dr. Haas and Dr. Fischer, LMU Munich / NovaTec Immundiagnostica GmbH) targets the development of a detection system which utilizes the differences in the so-called plasticity region of different Helicobacter pylori strains to improve the current diagnostics in regard to the likelihood of the emergence of a duodenal ulcer. The work focuses on the validation and verification of the duodenal ulcer promoting gene (dupA) encoded by the PZ region, as well as singular components of the dupA-associated secretion system (T4SS), which have been linked to the onset of a duodenal ulcer, as a biomarker for the development of an ELISA or Western-blot-based serological detection process.
The focus of project D3 (Prof. Dr. Ebel, LMU Munich / Mikrogen GmbH) is the establishment of immunohistological and serological processes by which non-aspergillus fungi and candida can be detected and differentiated regarding clinically relevant species with different resistance profiles against echinocandines, azoles and polyenes, (e.g. C. albicans, C. glabrata and C. krusei). This is to be achieved by producing monoclonal antibodies against glycan structures through the immunization with supernatants of fungal cultures, by testing and analyzing their specific reaction with non-aspergillus fungi and selected candida and translating this into a diagnostically relevant detection process. Advances in the differential diagnosis will enable a more targeted treatment of fungal infections and reduce the high mortality rate associated with invasive mycoses for immunosuppressed patients (e.g. following chemotherapy or in connection with an organ transplantation).
Next to the differentiation between pathogens, resistance testing is a major diagnostic parameter important for therapy success. This is especially important because of the drastic development of resistances, e.g. with gram-positive (MRSA, VRE) and gram-negative bacteria (ESBL enterobacteria, Pseudomonas aeruginosa, Acinetobacter baumannii) as well as the differential susceptibility of fungi against antimycotics. Current procedures for resistance profiling and therapy control are tied to the growth of the tested organism, giving results often only after a long incubation period.
Therefore, within project D4 (Prof. Dr. Schubert, LMU Munich / Bruker Daltonik GmbH) the principally fast MALDI-TOF MS technology will be used to (i) identify antibiotic resistances of bacteria in vitro , (ii) enable a faster in vitro testing of antimycotics against Candida and fungi and (iii) display bacterial resistances in vivo during therapy (therapeutic biomarkers) for the first time.
Projects D5 – D7 look into the detection and assessment of virus infections through antigen-specific T cells as a new biomarker for therapy monitoring (e.g. after transplantations), the design of innovative vaccine components as well as the idea of a cell-based therapy process.
Common basis for projects D5 and D6 is a process developed by their industry partner Lophius Biosciences GmbH which inserts recombinantly manufactured, soluble antigens also into the MHC class I pathway to stimulate virus-specific CD8+- cells for in vitro immunomonitoring. The effectiveness of the process is in part determined by the intrinsic characteristics of the treated protein, which so far are not fully understood.
The objective of project D5 (Prof. Dr. Wagner and Dr. Asbach, University of Regensburg / Lophius Biosciences GmbH) is the analysis of genetically modified variants of a model antigen in regard to a standardized ex vivo activation of selected phenotypic and functional biomarkers of the innate and adaptive immune systems. The HCMV IE1 antigen serves as a model antigen, which in the near future could be a useful addition to an already available and validated test for the immune monitoring of transplant patients, allowing for a more differentiated diagnosis. In addition it will be examined whether the in vitro stimulation of HCMV-antigen-specific T cells , which is so far only focussed on diagnostic applications, can be extended towards expansion of pathogen-specific T cells for cell-based therapy approaches as well as a basis for vaccination concepts.
Project D6 (Prof. Dr. Protzer and Dr. Bauer, TUM Munich / Lophius Biosciences GmbH) will then focus in detail on the pharmacodynamic T cell monitoring of antiviral immune responses as a prognostic biomarker following a kidney transplantation. In the course of the proposed project, antiviral T cell responses will be verified in the context of a planned multicentric study as a measurement for the severity of the immune suppression in relation to already established and validated immunoassay and correlated with clinical data such as (i) the reactivation of HCMV or BKV infections as well as (ii) rejection episodes. Through the definition of the specific immune response before and during immune suppression it will be examined if T cell monitoring of a virus-specific immune response can be used as a prognostic marker and for therapy monitoring, to measure the level of immune suppression and to be able to individually adjust the therapy accordingly. Besides the number of antigen-specific T cells, the binding strength of the T cell receptor (TCR) to its antigen (HLA-presented peptides) mainly determines the quality and possible protectivity of an immune response.
Prof. Dirk Busch and his colleagues recently could establish a new measuring system based on a reversible MHC streptamer technology devised in their team by which an important parameter of TCR avidity, the MHC / peptide dissociation-rate, can be measured exactly on the surface of living antigen-specific T cells. Based on this technology a test system should be developed during the course of project D7 (Prof. Dr. Busch and Dr. Nauerth, TUM Munich / Juno Therapeutics GmbH) which will be optimized for clinical diagnostic use and which will be (project D5 and D6) tested and validated in existing clinical cohorts on HCMV specific CD8+ T cells. Like project D5, project D7 can make an important contribution to the design of T-cell-based therapies.