@misc{RezaniaKammererLietal., author = {Rezania, Simin and Kammerer, Sarah and Li, Chouyang and Steinecker-Frohnwieser, Bibiane and Gorischek, Astrid and Devaney, Trevor Thomas Joseph and Verheyen, Sarah and Passegger, Christina and Ghaffari Tabrizi-Wizsy, Nassim and Hackl, Hubert and Platzer, Dieter and Zarnani, A.H. and Malle, Ernst and Jahn, Stephan Wenzel and Bauernhofer, Thomas and Schreibmayer, Wolfgang}, title = {Overexpression of KCNJ3 gene splice variants affects vital parameters of the malignant breast cancer cell line MCF-7 in an opposing manner}, series = {BMC Cancer}, volume = {16}, journal = {BMC Cancer}, issn = {1471-2407}, doi = {10.1186/s12885-016-2664-8}, abstract = {Background Overexpression the KCNJ3, a gene that encodes subunit 1 of G-protein activated inwardly rectifying K+ channel (GIRK1) in the primary tumor has been found to be associated with reduced survival times and increased lymph node metastasis in breast cancer patients. Methods In order to survey possible tumorigenic properties of GIRK1 overexpression, a range of malignant mammary epithelial cells, based on the MCF-7 cell line that permanently overexpress different splice variants of the KCNJ3 gene (GIRK1a, GIRK1c, GIRK1d and as a control, eYFP) were produced. Subsequently, selected cardinal neoplasia associated cellular parameters were assessed and compared. ResultsAdhesion to fibronectin coated surface as well as cell proliferation remained unaffected. Other vital parameters intimately linked to malignancy, i.e. wound healing, chemoinvasion, cellular velocities / motilities and angiogenesis were massively affected by GIRK1 overexpression. Overexpression of different GIRK1 splice variants exerted differential actions. While GIRK1a and GIRK1c overexpression reinforced the affected parameters towards malignancy, overexpression of GIRK1d resulted in the opposite. Single channel recording using the patch clamp technique revealed functional GIRK channels in the plasma membrane of MCF-7 cells albeit at very low frequency. DiscussionWe conclude that GIRK1d acts as a dominant negative constituent of functional GIRK complexes present in the plasma membrane of MCF-7 cells, while overexpression of GIRK1a and GIRK1c augmented their activity. The core component responsible for the cancerogenic action of GIRK1 is apparently presented by a segment comprising aminoacids 235-402, that is present exclusively in GIRK1a and GIRK1c, but not GIRK1d (positions according to GIRK1a primary structure). Conclusions The current study provides insight into the cellular and molecular consequences of KCNJ3 overexpression in breast cancer cells and the mechanism upon clinical outcome in patients suffering from breast cancer.}, language = {en} } @misc{KammererJahnWinteretal., author = {Kammerer, Sarah and Jahn, Stephan Wenzel and Winter, Elke and Eidenhammer, Sylvia and Rezania, Simin and Regitnig, Peter and Pichler, Martin and Schreibmayer, Wolfgang and Bauernhofer, Thomas}, title = {Critical evaluation of KCNJ3 gene product detection in human breast cancer: MRNA in situ hybridisation is superior to immunohistochemistry}, series = {Journal of Clinical Pathology}, volume = {69}, journal = {Journal of Clinical Pathology}, number = {12}, issn = {1472-4146}, doi = {10.1136/jclinpath-2016-203798}, pages = {1116 -- 1121}, abstract = {Increased expression levels of KCNJ3 have been correlated with lymph node metastases and poor prognosis in patients with breast cancer, suggesting a prognostic role of KCNJ3. We aimed to establish protocols for the detection of KCNJ3 in formalin-fixed, paraffin-embedded (FFPE) breast cancer tissue. Several antibodies were tested for sensitivity and specificity by western blot, followed by optimisation of the immunohistochemistry (IHC) procedure and establishment of KCNJ3 mRNA in situ hybridisation (ISH). Methods were validated by processing 15 FFPE breast cancer samples for which microarray data were available. Spearman's rank correlation analysis resulted in borderline significant correlation for IHC versus ISH (rS: 0.625; p<0.05) and IHC versus microarray (rS: 0.668; p<0.01), but in significant correlation for ISH versus microarray (rS: 0.861; p<0.001). The ISH method was superior to IHC, regarding robustness, sensitivity and specificity and will aid to further study expression levels of KCNJ3 in both malignant and physiological conditions.}, language = {en} } @misc{KammererSokolowskiHackletal., author = {Kammerer, Sarah and Sokolowski, Armin Andreas and Hackl, Hubert and Platzer, Dieter and Jahn, Stephan Wenzel and El-Heliebi, Amin and Schwarzenbacher, Daniela and Stiegelbauer, Verena and Pichler, Martin and Rezania, Simin and Fiegl, Heidelinde and Peintinger, Florentia and Regitnig, Peter and H{\"o}fler, Gerald and Schreibmayer, Wolfgang and Bauernhofer, Thomas}, title = {KCNJ3 is a new independent prognostic marker for estrogen receptor positive breast cancer patients}, series = {OncoTarget : open access impact journal}, volume = {7}, journal = {OncoTarget : open access impact journal}, number = {51}, doi = {10.18632/oncotarget.13224}, pages = {84705 -- 84717}, abstract = {Numerous studies showed abnormal expression of ion channels in different cancer types. Amongst these, the potassium channel gene KCNJ3 (encoding for GIRK1 proteins) has been reported to be upregulated in tumors of patients with breast cancer and to correlate with positive lymph node status. We aimed to study KCNJ3 levels in different breast cancer subtypes using gene expression data from the TCGA, to validate our findings using RNA in situ hybridization in a validation cohort (GEO ID GSE17705), and to study the prognostic value of KCNJ3using survival analysis. In a total of > 1000 breast cancer patients of two independent data sets we showed a) that KCNJ3 expression is upregulated in tumor tissue compared to corresponding normal tissue (p < 0.001), b) that KCNJ3 expression is associated with estrogen receptor (ER) positive tumors (p < 0.001), but that KCNJ3 expression is variable within this group, and c) that ER positive patients with high KCNJ3 levels have worse overall (p < 0.05) and disease free survival probabilities (p < 0.01), whereby KCNJ3 is an independent prognostic factor (p <0.05). In conclusion, our data suggest that patients with ER positive breast cancer might be stratified into high risk and low risk groups based on the KCNJ3 levels in the tumor.}, language = {en} } @misc{VerbruggeAlAssarafetal., author = {Verbrugge, Sue Ellen and Al, Marjon and Assaraf, Yehuda G. and Kammerer, Sarah and Chandrupatla, Durga and Honeywell, Richard and Musters, Rene and Giovannetti, Elisa and O`Toole, Tom and Scheffer, George L. and Krige, David and Gruijl, Tanja de and Niessen, Hans W.M. and Lems, Willem F. and Kramer, Pieternella A. and Scheper, Rik J. and Cloos, Jacqueline and Ossenkoppele, Gert and Peters, Godefridus J. and Jansen, Gerrit}, title = {Multifactorial resistance to aminopeptidase inhibitor prodrug CHR2863 in myeloid leukemia cells: Down-regulation of carboxylesterase 1, drug sequestration in lipid droplets and pro-survival activation ERK/Akt/mTOR}, series = {OncoTarget : open access impact journal}, volume = {7}, journal = {OncoTarget : open access impact journal}, number = {5}, issn = {1949-2553}, doi = {10.18632/oncotarget.6169}, pages = {5240 -- 5257}, abstract = {Aminopeptidase inhibitors are receiving attention as combination chemotherapeutic agents for the treatment of refractory acute myeloid leukemia. However, the factors determining therapeutic efficacy remain elusive. Here we identified the molecular basis of acquired resistance to CHR2863, an orally available hydrophobic aminopeptidase inhibitor prodrug with an esterase-sensitive motif, in myeloid leukemia cells. CHR2863 enters cells by diffusion and is retained therein upon esterase activity-mediated conversion to its hydrophilic active metabolite drug CHR6768, thereby exerting amino acid depletion. Carboxylesterases (CES) serve as candidate prodrug activating enzymes given CES1 expression in acute myeloid leukemia specimens. We established two novel myeloid leukemia sublines U937/CHR2863(200) and U937/CHR2863(5uM), with low (14-fold) and high level (270-fold) CHR2863 resistance. The latter drug resistant cells displayed: (i) complete loss of CES1-mediated drug activation associated with down-regulation of CES1 mRNA and protein, (ii) marked retention/sequestration of the prodrug, (iii) a substantial increase in intracellular lipid droplets, and (iv) a dominant activation of the pro-survival Akt/mTOR pathway. Remarkably, the latter feature coincided with a gain of sensitivity to the mTOR inhibitor rapamycin. These finding delineate the molecular basis of CHR2863 resistance and offer a novel modality to overcome this drug resistance in myeloid leukemia cells.}, language = {en} } @misc{LiRezaniaKammereretal., author = {Li, Chouyang and Rezania, Simin and Kammerer, Sarah and Sokolowski, Armin Andreas and Devaney, Trevor Thomas Joseph and Gorischek, Astrid and Jahn, Stephan Wenzel and Hackl, Hubert and Groschner, Klaus and Windpassinger, Christian and Malle, Ernst and Bauernhofer, Thomas and Schreibmayer, Wolfgang}, title = {Piezo1 forms mechanosensitive ion channels in the human MCF-7 breast cancer cell line}, series = {Scientific Reports}, volume = {5}, journal = {Scientific Reports}, issn = {2045-2322}, doi = {10.1038/srep08364}, pages = {9}, abstract = {Mechanical interaction between cells - specifically distortion of tensional homeostasis-emerged as an important aspect of breast cancer genesis and progression. We investigated the biophysical characteristics of mechanosensitive ion channels (MSCs) in the malignant MCF-7 breast cancer cell line. MSCs turned out to be the most abundant ion channel species and could be activated by negative pressure at the outer side of the cell membrane in a saturable manner. Assessing single channel conductance (GΛ) for different monovalent cations revealed an increase in the succession: Li⁺ < Na⁺ < K⁺ ≈Rb⁺ ≈ Cs⁺. Divalent cations permeated also with the order: Ca²⁺ < Ba²⁺. Comparison of biophysical properties enabled us to identify MSCs in MCF-7 as ion channels formed by the Piezo1 protein. Using patch clamp technique no functional MSCs were observed in the benign MCF-10A mammary epithelial cell line. Blocking of MSCs by GsMTx-4 resulted in decreased motility of MCF-7, but not of MCF-10A cells, underscoring a possible role of Piezo1 in invasion and metastatic propagation. The role of Piezo1 in biology and progression of breast cancer is further substantiated by markedly reduced overall survival in patients with increased Piezo1 mRNA levels in the primary tumor.}, language = {en} } @misc{KammererKuepper, author = {Kammerer, Sarah and K{\"u}pper, Jan-Heiner}, title = {Human Hepatocyte Systems for in vitro Toxicology Analysis.}, series = {Journal of Cellular Biotechnology}, volume = {3}, journal = {Journal of Cellular Biotechnology}, number = {2}, issn = {2352-3697}, doi = {10.3233/JCB-179012}, pages = {85 -- 93}, language = {en} } @misc{KammererKuepper, author = {Kammerer, Sarah and K{\"u}pper, Jan-Heiner}, title = {Optimized protocol for induction of cytochrome P450 enzymes 1A2 and 3A4 in human primary-like hepatocyte cell strain HepaFH3 to study in vitro toxicology}, series = {Clinical Hemorheology and Microcirculation}, volume = {70}, journal = {Clinical Hemorheology and Microcirculation}, number = {4}, issn = {1386-0291}, doi = {10.3233/CH-189321}, pages = {563 -- 571}, language = {en} } @misc{SteinbrechtKoenigSchmidtkeetal., author = {Steinbrecht, Susanne and K{\"o}nig, Rosalie and Schmidtke, Kai-Uwe and Herzog, Natalie and Scheibner, Katrin and Kr{\"u}ger-Genge, Anne and Jung, Friedrich and Kammerer, Sarah and K{\"u}pper, Jan-Heiner}, title = {Metabolic activity testing can underestimate acute drug cytotoxicity as revealed by HepG2 cell clones overexpressing cytochrome P450 2C19 and 3A4}, series = {Toxicology}, volume = {412}, journal = {Toxicology}, issn = {0300-483X}, doi = {10.1016/j.tox.2018.11.008}, pages = {37 -- 47}, language = {en} } @misc{NowakKammererKuepper, author = {Nowak, Elisabeth and Kammerer, Sarah and K{\"u}pper, Jan-Heiner}, title = {ATP-based cell viability assay is superior to trypan blue exclusion and XTT assay in measuring cytotoxicity of anticancer drugs Taxol and Imatinib, and proteasome inhibitor MG-132 on human hepatoma cell line HepG2.}, series = {Clinical Hemorheology and Microcirculation}, volume = {69}, journal = {Clinical Hemorheology and Microcirculation}, number = {1-2}, issn = {1386-0291}, doi = {10.3233/CH-189120}, pages = {327 -- 336}, language = {en} } @misc{HofmanSorfVagiannisetal., author = {Hofman, Jakub and Sorf, Ales and Vagiannis, Dimitrios and Sucha, Simona and Novotna, Eva and Kammerer, Sarah and K{\"u}pper, Jan-Heiner and Ceckova, Martina and Staud, Frantisek}, title = {Interactions of Alectinib with Human ATP-Binding Cassette Drug Efflux Transporters and Cytochrome P450 Biotransformation Enzymes: Effect on Pharmacokinetic Multidrug Resistance}, series = {Drug Metabolism and Disposition}, volume = {47}, journal = {Drug Metabolism and Disposition}, number = {7}, issn = {1521-009X}, doi = {10.1124/dmd.119.086975}, pages = {699 -- 709}, language = {en} }