@article{SeethalerHertleinHopkeetal.2022, author = {Seethaler, Marius and Hertlein, Tobias and Hopke, Elisa and K{\"o}hling, Paul and Ohlsen, Knut and Lalk, Michael and Hilgeroth, Andreas}, title = {Novel effective fluorinated benzothiophene-indole hybrid antibacterials against S. aureus and MRSA strains}, series = {Pharmaceuticals}, volume = {15}, journal = {Pharmaceuticals}, number = {9}, issn = {1424-8247}, doi = {10.3390/ph15091138}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-288253}, year = {2022}, abstract = {Increasing antibacterial drug resistance threatens global health, unfortunately, however, efforts to find novel antibacterial agents have been scaled back by the pharmaceutical industry due to concerns about a poor return on investment. Nevertheless, there is an urgent need to find novel antibacterial compounds to combat antibacterial drug resistance. The synthesis of novel drugs from natural sources is mostly cost-intensive due to those drugs' complicated structures. Therefore, it is necessary to find novel antibacterials by simple synthesis to become more attractive for industrial production. We succeeded in the discovery of four antibacterial compound (sub)classes accessible in a simple one-pot reaction based on fluorinated benzothiophene-indole hybrids. They have been evaluated against various S. aureus and MRSA strains. Structure- and substituent-dependent activities have been found within the (sub)classes and promising lead compounds have been identified. In addition, bacterial pyruvate kinase was found to be the molecular target of the active compounds. In conclusion, simple one-pot synthesis of benzothiophene-indoles represents a promising strategy for the search of novel antimicrobial compounds.}, language = {en} } @article{UmstaetterWernerZerlinetal.2022, author = {Umst{\"a}tter, Florian and Werner, Julia and Zerlin, Leah and M{\"u}hlberg, Eric and Kleist, Christian and Klika, Karel D. and Hertlein, Tobias and Beijer, Barbro and Domhan, Cornelius and Zimmermann, Stefan and Ohlsen, Knut and Haberkorn, Uwe and Mier, Walter and Uhl, Philipp}, title = {Impact of linker modification and PEGylation of vancomycin conjugates on structure-activity relationships and pharmacokinetics}, series = {Pharmaceuticals}, volume = {15}, journal = {Pharmaceuticals}, number = {2}, issn = {1424-8247}, doi = {10.3390/ph15020159}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-255197}, year = {2022}, abstract = {As multidrug-resistant bacteria represent a concerning burden, experts insist on the need for a dramatic rethinking on antibiotic use and development in order to avoid a post-antibiotic era. New and rapidly developable strategies for antimicrobial substances, in particular substances highly potent against multidrug-resistant bacteria, are urgently required. Some of the treatment options currently available for multidrug-resistant bacteria are considerably limited by side effects and unfavorable pharmacokinetics. The glycopeptide vancomycin is considered an antibiotic of last resort. Its use is challenged by bacterial strains exhibiting various types of resistance. Therefore, in this study, highly active polycationic peptide-vancomycin conjugates with varying linker characteristics or the addition of PEG moieties were synthesized to optimize pharmacokinetics while retaining or even increasing antimicrobial activity in comparison to vancomycin. The antimicrobial activity of the novel conjugates was determined by microdilution assays on susceptible and vancomycin-resistant bacterial strains. VAN1 and VAN2, the most promising linker-modified derivatives, were further characterized in vivo with molecular imaging and biodistribution studies in rodents, showing that the linker moiety influences both antimicrobial activity and pharmacokinetics. Encouragingly, VAN2 was able to undercut the resistance breakpoint in microdilution assays on vanB and vanC vancomycin-resistant enterococci. Out of all PEGylated derivatives, VAN:PEG1 and VAN:PEG3 were able to overcome vanC resistance. Biodistribution studies of the novel derivatives revealed significant changes in pharmacokinetics when compared with vancomycin. In conclusion, linker modification of vancomycin-polycationic peptide conjugates represents a promising strategy for the modulation of pharmacokinetic behavior while providing potent antimicrobial activity.}, language = {en} } @article{HungDreherDiessneretal.2022, author = {Hung, Sophia and Dreher, Liane and Diessner, Joachim and Schwarz, Stefan and Ohlsen, Knut and Hertlein, Tobias}, title = {MRSA infection in the thigh muscle leads to systemic disease, strong inflammation, and loss of human monocytes in humanized mice}, series = {Frontiers in Immunology}, volume = {13}, journal = {Frontiers in Immunology}, issn = {1664-3224}, doi = {10.3389/fimmu.2022.892053}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-278050}, year = {2022}, abstract = {MRSA (Methicillin-resistant Staphylococcus aureus) is the second-leading cause of deaths by antibiotic-resistant bacteria globally, with more than 100,000 attributable deaths annually. Despite the high urgency to develop a vaccine to control this pathogen, all clinical trials with pre-clinically effective candidates failed so far. The recent development of "humanized" mice might help to edge the pre-clinical evaluation closer to the clinical situation and thus close this gap. We infected humanized NSG mice (huNSG: (NOD)-scid IL2R\(_γ\)\(^{null}\) mice engrafted with human CD34+ hematopoietic stem cells) locally with S. aureus USA300 LAC* lux into the thigh muscle in order to investigate the human immune response to acute and chronic infection. These mice proved not only to be more susceptible to MRSA infection than wild-type or "murinized" mice, but displayed furthermore inferior survival and signs of systemic infection in an otherwise localized infection model. The rate of humanization correlated directly with the severity of disease and survival of the mice. Human and murine cytokine levels in blood and at the primary site of infection were strongly elevated in huNSG mice compared to all control groups. And importantly, differences in human and murine immune cell lineages surfaced during the infection, with human monocyte and B cell numbers in blood and bone marrow being significantly reduced at the later time point of infection. Murine monocytes in contrast behaved conversely by increasing cell numbers. This study demonstrates significant differences in the in vivo behavior of human and murine cells towards S. aureus infection, which might help to sharpen the translational potential of pre-clinical models for future therapeutic approaches.}, language = {en} } @article{GehrmannHertleinHopkeetal.2021, author = {Gehrmann, Robin and Hertlein, Tobias and Hopke, Elisa and Ohlsen, Knut and Lalk, Michael and Hilgeroth, Andreas}, title = {Novel small-molecule hybrid-antibacterial agents against S. aureus and MRSA strains}, series = {Molecules}, volume = {27}, journal = {Molecules}, number = {1}, issn = {1420-3049}, doi = {10.3390/molecules27010061}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-252371}, year = {2021}, abstract = {Ongoing resistance developments against antibiotics that also affect last-resort antibiotics require novel antibacterial compounds. Strategies to discover such novel structures have been dimerization or hybridization of known antibacterial agents. We found novel antibacterial agents by dimerization of indols and hybridization with carbazoles. They were obtained in a simple one-pot reaction as bisindole tetrahydrocarbazoles. Further oxidation led to bisindole carbazoles with varied substitutions of both the indole and the carbazole scaffold. Both the tetrahydrocarbazoles and the carbazoles have been evaluated in various S. aureus strains, including MRSA strains. Those 5-cyano substituted derivatives showed best activities as determined by MIC values. The tetrahydrocarbazoles partly exceed the activity of the carbazole compounds and thus the activity of the used standard antibiotics. Thus, promising lead compounds could be identified for further studies.}, language = {en} } @article{MuehlbergUmstaetterDomhanetal.2020, author = {M{\"u}hlberg, Eric and Umst{\"a}tter, Florian and Domhan, Cornelius and Hertlein, Tobias and Ohlsen, Knut and Krause, Andreas and Kleist, Christian and Beijer, Barbro and Zimmermann, Stefan and Haberkorn, Uwe and Mier, Walter and Uhl, Philipp}, title = {Vancomycin-lipopeptide conjugates with high antimicrobial activity on vancomycin-resistant enterococci}, series = {Pharmaceuticals}, volume = {13}, journal = {Pharmaceuticals}, number = {6}, issn = {1424-8247}, doi = {10.3390/ph13060110}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-205879}, year = {2020}, abstract = {Multidrug-resistant bacteria represent one of the most important health care problems worldwide. While there are numerous drugs available for standard therapy, there are only a few compounds capable of serving as a last resort for severe infections. Therefore, approaches to control multidrug-resistant bacteria must be implemented. Here, a strategy of reactivating the established glycopeptide antibiotic vancomycin by structural modification with polycationic peptides and subsequent fatty acid conjugation to overcome the resistance of multidrug-resistant bacteria was followed. This study especially focuses on the structure-activity relationship, depending on the modification site and fatty acid chain length. The synthesized conjugates showed high antimicrobial potential on vancomycin-resistant enterococci. We were able to demonstrate that the antimicrobial activity of the vancomycin-lipopeptide conjugates depends on the chain length of the attached fatty acid. All conjugates showed good cytocompatibility in vitro and in vivo. Radiolabeling enabled the in vivo determination of pharmacokinetics in Wistar rats by molecular imaging and biodistribution studies. An improved biodistribution profile in comparison to unmodified vancomycin was observed. While vancomycin is rapidly excreted by the kidneys, the most potent conjugate shows a hepatobiliary excretion profile. In conclusion, these results demonstrate the potential of the structural modification of already established antibiotics to provide highly active compounds for tackling multidrug-resistant bacteria.}, language = {en} } @article{SeethalerHertleinWeckleinetal.2019, author = {Seethaler, Marius and Hertlein, Tobias and Wecklein, Bj{\"o}rn and Ymeraj, Alba and Ohlsen, Knut and Lalk, Michael and Hilgeroth, Andreas}, title = {Novel small-molecule antibacterials against Gram-positive pathogens of Staphylococcus and Enterococcus species}, series = {Antibiotics}, volume = {8}, journal = {Antibiotics}, number = {4}, issn = {2079-6382}, doi = {10.3390/antibiotics8040210}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-193130}, year = {2019}, abstract = {Defeat of the antibiotic resistance of pathogenic bacteria is one great challenge today and for the future. In the last century many classes of effective antibacterials have been developed, so that upcoming resistances could be met with novel drugs of various compound classes. Meanwhile, there is a certain lack of research of the pharmaceutical companies, and thus there are missing developments of novel antibiotics. Gram-positive bacteria are the most important cause of clinical infections. The number of novel antibacterials in clinical trials is strongly restricted. There is an urgent need to find novel antibacterials. We used synthetic chemistry to build completely novel hybrid molecules of substituted indoles and benzothiophene. In a simple one-pot reaction, two novel types of thienocarbazoles were yielded. Both indole substituted compound classes have been evaluated as completely novel antibacterials against the Staphylococcus and Enterococcus species. The evaluated partly promising activities depend on the indole substituent type. First lead compounds have been evaluated within in vivo studies. They confirmed the in vitro results for the new classes of small-molecule antibacterials.}, language = {en} } @article{StelznerBoynyHertleinetal.2021, author = {Stelzner, Kathrin and Boyny, Aziza and Hertlein, Tobias and Sroka, Aneta and Moldovan, Adriana and Paprotka, Kerstin and Kessie, David and Mehling, Helene and Potempa, Jan and Ohlsen, Knut and Fraunholz, Martin J. and Rudel, Thomas}, title = {Intracellular Staphylococcus aureus employs the cysteine protease staphopain A to induce host cell death in epithelial cells}, series = {PLoS Pathogens}, volume = {17}, journal = {PLoS Pathogens}, number = {9}, doi = {10.1371/journal.ppat.1009874}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-263908}, year = {2021}, abstract = {Staphylococcus aureus is a major human pathogen, which can invade and survive in non-professional and professional phagocytes. Uptake by host cells is thought to contribute to pathogenicity and persistence of the bacterium. Upon internalization by epithelial cells, cytotoxic S. aureus strains can escape from the phagosome, replicate in the cytosol and induce host cell death. Here, we identified a staphylococcal cysteine protease to induce cell death after translocation of intracellular S. aureus into the host cell cytoplasm. We demonstrated that loss of staphopain A function leads to delayed onset of host cell death and prolonged intracellular replication of S. aureus in epithelial cells. Overexpression of staphopain A in a non-cytotoxic strain facilitated intracellular killing of the host cell even in the absence of detectable intracellular replication. Moreover, staphopain A contributed to efficient colonization of the lung in a mouse pneumonia model. In phagocytic cells, where intracellular S. aureus is exclusively localized in the phagosome, staphopain A did not contribute to cytotoxicity. Our study suggests that staphopain A is utilized by S. aureus to exit the epithelial host cell and thus contributes to tissue destruction and dissemination of infection. Author summary Staphylococcus aureus is an antibiotic-resistant pathogen that emerges in hospital and community settings and can cause a variety of diseases ranging from skin abscesses to lung inflammation and blood poisoning. The bacterium can asymptomatically colonize the upper respiratory tract and skin of humans and take advantage of opportune conditions, like immunodeficiency or breached barriers, to cause infection. Although S. aureus was not regarded as intracellular bacterium, it can be internalized by human cells and subsequently exit the host cells by induction of cell death, which is considered to cause tissue destruction and spread of infection. The bacterial virulence factors and underlying molecular mechanisms involved in the intracellular lifestyle of S. aureus remain largely unknown. We identified a bacterial cysteine protease to contribute to host cell death of epithelial cells mediated by intracellular S. aureus. Staphopain A induced killing of the host cell after translocation of the pathogen into the cell cytosol, while bacterial proliferation was not required. Further, the protease enhanced survival of the pathogen during lung infection. These findings reveal a novel, intracellular role for the bacterial protease staphopain A.}, language = {en} } @article{UmstaetterDomhanHertleinetal.2020, author = {Umst{\"a}tter, Florian and Domhan, Cornelius and Hertlein, Tobias and Ohlsen, Knut and M{\"u}hlberg, Eric and Kleist, Christian and Zimmermann, Stefan and Beijer, Barbro and Klika, Karel D. and Haberkorn, Uwe and Mier, Walter and Uhl, Philipp}, title = {Vancomycin Resistance Is Overcome by Conjugation of Polycationic Peptides}, series = {Angewandte Chemie International Edition}, volume = {59}, journal = {Angewandte Chemie International Edition}, number = {23}, doi = {10.1002/anie.202002727}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-215550}, pages = {8823 -- 8827}, year = {2020}, abstract = {Multidrug-resistant bacteria represent one of the biggest challenges facing modern medicine. The increasing prevalence of glycopeptide resistance compromises the efficacy of vancomycin, for a long time considered as the last resort for the treatment of resistant bacteria. To reestablish its activity, polycationic peptides were conjugated to vancomycin. By site-specific conjugation, derivatives that bear the peptide moiety at four different sites of the antibiotic were synthesized. The most potent compounds exhibited an approximately 1000-fold increased antimicrobial activity and were able to overcome the most important types of vancomycin resistance. Additional blocking experiments using d-Ala-d-Ala revealed a mode of action beyond inhibition of cell-wall formation. The antimicrobial potential of the lead candidate FU002 for bacterial infection treatments could be demonstrated in an in vivo study. Molecular imaging and biodistribution studies revealed that conjugation engenders superior pharmacokinetics.}, language = {en} } @article{HanzelmannJooFranzWachteletal.2016, author = {Hanzelmann, Dennis and Joo, Hwang-Soo and Franz-Wachtel, Mirita and Hertlein, Tobias and Stevanovic, Stefan and Macek, Boris and Wolz, Christiane and G{\"o}tz, Friedrich and Otto, Michael and Kretschmer, Dorothee and Peschel, Andreas}, title = {Toll-like receptor 2 activation depends on lipopeptide shedding by bacterial surfactants}, series = {Nature Communications}, volume = {7}, journal = {Nature Communications}, doi = {10.1038/ncomms12304}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-165975}, pages = {12304}, year = {2016}, abstract = {Sepsis caused by Gram-positive bacterial pathogens is a major fatal disease but its molecular basis remains elusive. Toll-like receptor 2 (TLR2) has been implicated in the orchestration of inflammation and sepsis but its role appears to vary for different pathogen species and clones. Accordingly, Staphylococcus aureus clinical isolates differ substantially in their capacity to activate TLR2. Here we show that strong TLR2 stimulation depends on high-level production of phenol-soluble modulin (PSM) peptides in response to the global virulence activator Agr. PSMs are required for mobilizing lipoproteins, the TLR2 agonists, from the staphylococcal cytoplasmic membrane. Notably, the course of sepsis caused by PSM-deficient S. aureus is similar in wild-type and TLR2-deficient mice, but TLR2 is required for protection of mice against PSM-producing S. aureus. Thus, a crucial role of TLR2 depends on agonist release by bacterial surfactants. Modulation of this process may lead to new therapeutic strategies against Gram-positive infections.}, language = {en} } @article{AshrafYasrebiHertleinetal.2017, author = {Ashraf, Kerolos and Yasrebi, Kaveh and Hertlein, Tobias and Ohlsen, Knut and Lalk, Michael and Hilgeroth, Andreas}, title = {Novel effective small-molecule antibacterials against \(Enterococcus\) strains}, series = {Molecules}, volume = {22}, journal = {Molecules}, number = {12}, doi = {10.3390/molecules22122193}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-172628}, year = {2017}, abstract = {\(Enterococcus\) species cause increasing numbers of infections in hospitals. They contribute to the increasing mortality rates, mostly in patients with comorbidities, who suffer from severe diseases. \(Enterococcus\) resistances against most antibiotics have been described, including novel antibiotics. Therefore, there is an ongoing demand for novel types of antibiotics that may overcome bacterial resistances. We discovered a novel class of antibiotics resulting from a simple one-pot reaction of indole and \(o\)-phthaldialdehyde. Differently substituted indolyl benzocarbazoles were yielded. Both the indole substitution and the positioning at the molecular scaffold influence the antibacterial activity towards the various strains of \(Enterococcus\) species with the highest relevance to nosocomial infections. Structure-activity relationships are discussed, and the first lead compounds were identified as also being effective in the case of a vancomycin resistance.}, language = {en} }