@misc{SteinbrechtKiebistKoenigetal., author = {Steinbrecht, Susanne and Kiebist, Jan and K{\"o}nig, Rosalie and Thiessen, Markus and Schmidtke, Kai-Uwe and Kammerer, Sarah and K{\"u}pper, Jan-Heiner and Scheibner, Katrin}, title = {Synthesis of cyclophosphamide metabolites by a peroxygenase from Marasmius rotula for toxicological studies on human cancer cells}, series = {AMB Express}, volume = {10}, journal = {AMB Express}, issn = {2191-0855}, doi = {10.1186/s13568-020-01064-w}, pages = {13}, abstract = {Cyclophosphamide (CPA) represents a widely used anti-cancer prodrug that is converted by liver cytochrome P450 (CYP) enzymes into the primary metabolite 4-hydroxycyclophosphamide (4-OH-CPA), followed by non-enzymatic generation of the bioactive metabolites phosphoramide mustard and acrolein. The use of human drug metabolites as authentic standards to evaluate their toxicity is essential for drug development. However, the chemical synthesis of 4-OH-CPA is complex and leads to only low yields and undesired side products. In past years, fungal unspecific peroxygenases (UPOs) have raised to powerful biocatalysts. They can exert the identical selective oxyfunctionalization of organic compounds and drugs as known for CYP enzymes with hydrogen peroxide being used as sole cosubstrate. Herein, we report the efficient enzymatic hydroxylation of CPA using the unspecific peroxygenase from Marasmius rotula (MroUPO) in a simple reaction design. Depending on the conditions used the primary liver metabolite 4-OH-CPA, its tautomer aldophosphamide (APA) and the overoxidized product 4-ketocyclophosphamide (4-keto-CPA) could be obtained. Using a kinetically controlled approach 4-OH-CPA was isolated with a yield of 32\% (purity > 97.6\%). Two human cancer cell lines (HepG2 and MCF-7) were treated with purified 4-OH-CPA produced by MroUPO (4-OH-CPAUPO). 4-OH-CPAUPO-induced cytotoxicity as measured by a luminescent cell viability assay and its genotoxicity as measured by γH2AX foci formation was not significantly different to the commercially available standard. The high yield of 4-OH-CPAUPO and its biological activity demonstrate that UPOs can be efficiently used to produce CYP-specific drug metabolites for pharmacological assessment.}, language = {en} } @misc{VagiannisNovotnaSkarkaetal., author = {Vagiannis, Dimitrios and Novotna, Eva and Skarka, Adam and Kammerer, Sarah and K{\"u}pper, Jan-Heiner and Chen, Si and Guo, Lei and Staud, Frantisek and Hofman, Jakub}, title = {Ensartinib (X-396) Effectively Modulates Pharmacokinetic Resistance Mediated by ABCB1 and ABCG2 Drug Efflux Transporters and CYP3A4 Biotransformation Enzyme}, series = {Cancers}, volume = {12}, journal = {Cancers}, number = {4}, issn = {2072-6694}, doi = {10.3390/cancers12040813}, pages = {27}, abstract = {Ensartinib (X-396) is a promising tyrosine kinase inhibitor currently undergoing advanced clinical evaluation for the treatment of non-small cell lung cancer. In this work, we investigate possible interactions of this promising drug candidate with ATP-binding cassette (ABC) drug efflux transporters and cytochrome P450 biotransformation enzymes (CYPs), which play major roles in multidrug resistance (MDR) and pharmacokinetic drug-drug interactions (DDIs). Accumulation studies showed that ensartinib is a potent inhibitor of ABCB1 and ABCG2 transporters. Additionally, incubation experiments with recombinant CYPs showed that ensartinib significantly inhibits CYP3A4 and CYP2C9. Subsequent molecular docking studies confirmed these findings. Drug combination experiments demonstrated that ensartinib synergistically potentiates the antiproliferative effects of daunorubicin, mitoxantrone, and docetaxel in ABCB1, ABCG2, and CYP3A4-overexpressing cellular models, respectively. Advantageously, ensartinib's antitumor efficiency was not compromised by the presence of MDR-associated ABC transporters, although it acted as a substrate of ABCB1 in Madin-Darby Canine Kidney II (MDCKII) monolayer transport assays. Finally, we demonstrated that ensartinib had no significant effect on the mRNA-level expression of examined transporters and enzymes in physiological and lung tumor cellular models. In conclusion, ensartinib may perpetrate clinically relevant pharmacokinetic DDIs and modulate ABCB1-, ABCG2-, and CYP3A4-mediated MDR. The in vitro findings presented here will provide a valuable foundation for future in vivo investigations.}, language = {en} } @misc{LauRangarajanKruegerGengeetal., author = {Lau, S. and Rangarajan, R. and Kr{\"u}ger-Genge, Anne and Braune, Steffen and K{\"u}pper, Jan-Heiner and Lendlein, Andreas and Jung, Friedrich}, title = {Age-related morphology and function of human arterial endothelial cells}, series = {Clinical Hemorheology and Microcirculation}, volume = {74}, journal = {Clinical Hemorheology and Microcirculation}, number = {1}, issn = {1875-8622}, doi = {10.3233/CH-199238}, pages = {93 -- 107}, abstract = {Endothelialization of cardiovascular implants is regarded as a promising strategy for long-term compatibility. While umbilical vein endothelial cells are typically applied in research, human arterial endothelial cells (HAEC) from elderly donors would be the obvious source for autologous cellularization strategies. In our approach, HAEC from 16 donors of varying age (16-63 years) were divided into two groups (<30 years and >30 years) and analyzed regarding morphology, viability, proliferation, function and senescence status. No age-related differences were found regarding morphology, viability, density, prostacyclin and nitrite secretion or collagen and laminin production. However, the metabolic activity was slightly decreased (p = 0.0374) and the membrane integrity marginally impaired (p = 0.0404) in cells from older donors. Two out of three senescence assays detected more senescence markers in cells from older donors. According to the assays applied here, HAEC from young and elderly donors up to the age of 63 years could be judged equally suitable for autologous cellularization strategies. However, this finding should be regarded with caution due to the extremely large variability between individual donors. Further studies comprising a larger sample size are necessary to investigate this issue more thoroughly.}, language = {en} } @misc{KruegerGengeSteinbrechtJungetal., author = {Kr{\"u}ger-Genge, Anne and Steinbrecht, Susanne and Jung, Conrad H. G. and Westphal, Sophia and Kl{\"o}pzig, Stefanie and Waldeck, Peter and K{\"u}pper, Jan-Heiner and Storsberg, J. and Jung, Friedrich}, title = {Arthrospira platensis accelerates the formation of an endothelial cell monolayer and protects against endothelial cell detachment after bacterial contamination}, series = {Clinical Hemorheology and Microcirculation}, volume = {78}, journal = {Clinical Hemorheology and Microcirculation}, number = {2}, issn = {1386-0291}, doi = {10.3233/CH-201096}, pages = {151 -- 161}, abstract = {Within the last years a comprehensive number of scientific studies demonstrated beneficial effect of Arthropira platensis (AP) as dietary supplement due to a high content of proteins, minerals and vitamins. Positive effects like promoting the immune system, reducing inflammation and an anti-oxidant capacity are reported. In this study, the effect of an aqueous AP extract on primary human venous endothelial cells (HUVEC) was investigated. In addition, the effect of AP on HUVEC treated with a bacterial toxin (lipopolysaccharide, LPA), inducing an activation of HUVEC and cellular detachment, was analyzed. Depending on the concentration of AP extract a significantly accelerated formation of an endothelial cell monolayer was observed. Furthermore, the detachment of HUVEC after LPA addition was dramatically reduced by AP. In conclusion, the data are promising and indicatory for an application of Arthrospira platensis in the clinical field.}, language = {en} } @misc{SchulzKruegerGengeLendleinetal., author = {Schulz, Christian and Kr{\"u}ger-Genge, Anne and Lendlein, Andreas and K{\"u}pper, Jan-Heiner and Jung, Friedrich}, title = {Potential Effects of Nonadherent on Adherent Human Umbilical Venous Endothelial Cells in Cell Culture}, series = {International Journal of Molecular Science}, volume = {22}, journal = {International Journal of Molecular Science}, number = {3}, doi = {10.3390/ijms22031493}, pages = {15}, abstract = {The adherence and shear-resistance of human umbilical venous endothelial cells (HUVEC) on polymers is determined in vitro in order to qualify cardiovascular implant materials. In these tests, variable fractions of HUVEC do not adhere to the material but remain suspended in the culture medium. Nonadherent HUVEC usually stop growing, rapidly lose their viability and can release mediators able to influence the growth and function of the adherent HUVEC. The aim of this study was the investigation of the time dependent behaviour of HUVEC under controlled nonadherent conditions, in order to gain insights into potential influences of these cells on their surrounding environment in particular adherent HUVEC in the context of in vitro biofunctionality assessment of cardiovascular implant materials. Data from adherent or nonadherent HUVEC growing on polystyrene-based cell adhesive tissue culture plates (TCP) or nonadhesive low attachment plates (LAP) allow to calculate the number of mediators released into the culture medium either from adherent or nonadherent cells. Thus, the source of the inflammatory mediators can be identified. For nonadherent HUVEC, a time-dependent aggregation without further proliferation was observed. The rate of apoptotic/dead HUVEC progressively increased over 90\% within two days. Concomitant with distinct blebbing and loss of membrane integrity over time, augmented releases of prostacyclin (PGI2, up to 2.91 ± 0.62 fg/cell) and platelet-derived growth factor BB (PDGF-BB, up to 1.46 ± 0.42 fg/cell) were detected. The study revealed that nonadherent, dying HUVEC released mediators, which can influence the surrounding microenvironment and thereby the results of in vitro biofunctionality assessment of cardiovascular implant materials. Neglecting nonadherent HUVEC bears the risk for under- or overestimation of the materials endothelialization potential, which could lead to the loss of relevant candidates or to uncertainty with regard to their suitability for cardiac applications. One approach to minimize the influence from nonadherent endothelial cells could be their removal shortly after observing initial cell adhesion. However, this would require an individual adaptation of the study design, depending on the properties of the biomaterial used.}, language = {en} } @misc{GeburekRutzGaoetal., author = {Geburek, Ina and Rutz, Lukas and Gao, Lan and K{\"u}pper, Jan-Heiner and These, Anja and Schrenk, Dieter}, title = {The metabolic pattern of hepatotoxic pyrrolizidine alkaloids in liver cells}, series = {Chemical Research in Toxicology}, volume = {34}, journal = {Chemical Research in Toxicology}, number = {4}, doi = {10.1021/acs.chemrestox.0c00507}, pages = {1101 -- 1113}, abstract = {Contamination with 1,2-unsaturated pyrrolizidine alkaloids (PAs) is a serious problem for certain phytomedicines, foods, and animal feeds. Several of these PAs are genotoxic and carcinogenic, primarily in the liver, upon cytochrome P450 (CYP)-catalyzed activation into reactive (pyrrolic and pyrrole-like) metabolites. Here we investigated the metabolism of selected PAs (echimidine, europine, lasiocarpine, lycopsamine, retrorsine, and senecionine) in rat hepatocytes in primary culture and in human CYP3A4-transfected HepG2 cells. The open-chained diesters echimidine and lasiocarpine and the cyclic diester senecionine were extensively metabolized in rat hepatocytes into a broad spectrum of products released into the medium. A large portion of unidentified, possibly irreversibly bound, products remained in the cells while detectable amounts of reactive and other metabolites were found in the incubation media. In HepG2-CYP3A4 cells, lasiocarpine was more extensively metabolized than echimidine and senecionine which also gave rise to the release of pyrrolic metabolites. In human cells, no pyrrolic metabolites were detected in retrorsine or lycopsamine incubations, while no such metabolites were detected from europine in both cell types. Other types of metabolic changes comprised modifications such as side chain demethylation or oxygenation reactions like the formation of N-oxides. The latter, considered as a detoxification step, was a major pathway with cyclic diesters, was less distinctive for echimidine and lycopsamine and almost negligible for lasiocarpine and europine. Our data are in agreement with previously published cyto- and genotoxicity findings and suggests that the metabolic pattern may contribute substantially to the specific toxic potency of a certain congener. In addition, marked differences were found for certain congeners between rat hepatocytes and transfected human HepG2 cells, whereby a high level of bioactivation was found for lasiocarpine, whereas a very low level of bioactivation was observed for monoesters, in particular in human cells.}, language = {en} } @misc{JungWaldeckPetricketal., author = {Jung, Conrad H. G. and Waldeck, Peter and Petrick, Ingolf and Braune, Steffen and K{\"u}pper, Jan-Heiner and Jung, Friedrich}, title = {Bioreactor for the cultivation of Arthrospira platensis under controlled conditions}, series = {Journal of Cellular Biotechnology}, volume = {7}, journal = {Journal of Cellular Biotechnology}, number = {1}, doi = {10.3233/JCB-210032}, pages = {35 -- 40}, abstract = {A vertical flat-type bioreactor consisting of transparent polyethylene (food safe) was constructed, which is characterized by a flexible design and allows the adjustment of a light path of 1 cm - 8.5 cm and a working volume of 1.5 l - 4 l. To characterize the performance of the bioreactor, cultivation experiments were performed with the cyanobacterium Arthrospira platensis (AP). The growth was assessed continuously by monitoring optical density and intermittently by measuring the dry weight of the AP biomass. An on-line measurement technique for estimating biomass production rate in a photosynthetic microalgae culture was developed. The oxygen produced by AP in the culture medium was flushed out sparging using a mixture of air and CO2 (1\%). Factors which might influence the AP growth were monitored: pH, temperature, oxygen concentration and the filling level were corrected automatically to compensate evaporation losses. As an example, the huge influence of the light intensity on the AP growth was tested. The increase of the photon flux density of 15 to 1200μmol/(l · d) led to a 22-fold increase of the productivity and a 3.1-fold shorter doubling time. Using an online measurement technique - together with the control of the growth process via a wireless local area network (WLAN) router and virtual private network - allows monitoring the growth of Arthrospira platensis remotely.}, language = {en} } @misc{RahmanMoolaKuepper, author = {Rahman, Ateeq and Moola, Nyambe and K{\"u}pper, Jan-Heiner}, title = {Namibian algae species: A review of their distribution, medicinal uses and chemical constituents}, series = {Journal of Cellular Biotechnology}, volume = {6}, journal = {Journal of Cellular Biotechnology}, number = {2}, doi = {10.3233/JCB-209010}, pages = {139 -- 159}, abstract = {The use of indigenous or remote popular knowledge to identify new drugs against diseases or infections is a well-known approach in medicine. The inhabitants of coastal regions in Namibia and other African countries are known to prepare algae extracts for the treatment of disorders and ailments such as wounds, fever and stomach aches, as well as for the prevention of arrhythmia, cancer, and many other diseases. Algae survive in a competitive environment and, therefore, developed defense strategies that have resulted in a significant level of chemical structural diversity in various metabolic pathways. The exploration of these organisms for pharmaceutical, nutritional and medical purposes has provided important chemical candidates for the discovery of new agents against neglected tropical diseases and stimulated the use of sophisticated physical techniques. This current review provides a broad picture on the taxonomy, various medical and nutritional uses of algae, which thus should be of relevance for the African continent and underdeveloped countries in the Global South.}, language = {en} } @misc{BrauneKuepperJung, author = {Braune, Steffen and K{\"u}pper, Jan-Heiner and Jung, Friedrich}, title = {Effect of Prostanoids on Human Platelet Function: An Overview}, series = {International Journal of Molecular Science}, volume = {21}, journal = {International Journal of Molecular Science}, number = {23}, doi = {10.3390/ijms21239020}, pages = {20}, abstract = {Prostanoids are bioactive lipid mediators and take part in many physiological and pathophysiological processes in practically every organ, tissue and cell, including the vascular, renal, gastrointestinal and reproductive systems. In this review, we focus on their influence on platelets, which are key elements in thrombosis and hemostasis. The function of platelets is influenced by mediators in the blood and the vascular wall. Activated platelets aggregate and release bioactive substances, thereby activating further neighbored platelets, which finally can lead to the formation of thrombi. Prostanoids regulate the function of blood platelets by both activating or inhibiting and so are involved in hemostasis. Each prostanoid has a unique activity profile and, thus, a specific profile of action. This article reviews the effects of the following prostanoids: prostaglandin-D2 (PGD2), prostaglandin-E1, -E2 and E3 (PGE1, PGE2, PGE3), prostaglandin F2α (PGF2α), prostacyclin (PGI2) and thromboxane-A2 (TXA2) on platelet activation and aggregation via their respective receptors.}, language = {en} } @misc{BrauneKruegerGengeKammereretal., author = {Braune, Steffen and Kr{\"u}ger-Genge, Anne and Kammerer, Sarah and Jung, Friedrich and K{\"u}pper, Jan-Heiner}, title = {Phycocyanin from Arthrospira platensis as Potential Anti-Cancer Drug: Review of In Vitro and In Vivo Studies}, series = {Life}, volume = {11}, journal = {Life}, number = {2}, doi = {10.3390/life11020091}, pages = {91}, language = {en} }