Identifying Regulators for EAG1 Channels with a Novel Electrophysiology and Tryptophan Fluorescence Based Screen

Ether-à-go-go (EAG) channels are expressed throughout the central nervous system and are also crucial regulators of cell cycle and tumor progression. The large intracellular amino- and carboxy- terminal domains of EAG1 each share similarity with known ligand binding motifs in other proteins, yet EAG1 channels have no known regulatory ligands.Here we screened a library of small biologically relevant molecules against EAG1 channels with a novel two-pronged screen to identify channel regulators. In one arm of the screen we used electrophysiology to assess the functional effects of the library compounds on full-length EAG1 channels. In an orthogonal arm, we used tryptophan fluorescence to screen for binding of the library compounds to the isolated C-terminal region.Several compounds from the flavonoid, indole and benzofuran chemical families emerged as binding partners and/or regulators of EAG1 channels. The two-prong screen can aid ligand and drug discovery for ligand-binding domains of other ion channels

Targeting and killing of glioblastoma with activated T cells armed with bispecific antibodies

Abstract Background Since most glioblastomas express both wild-type EGFR and EGFRvIII as well as HER2/neu, they are excellent targets for activated T cells (ATC) armed with bispecific antibodies (BiAbs) that target EGFR and HER2. Methods ATC were generated from PBMC activated for 14 days with anti-CD3 monoclonal antibody in the presence of interleukin-2 and armed with chemically heteroconjugated anti-CD3×anti-HER2/neu (HER2Bi) and/or anti-CD3×anti-EGFR (EGFRBi). HER2Bi- and/or EGFRBi-armed ATC were examined for in vitro cytotoxicity using MTT and 51Cr-release assays against malignant glioma lines (U87MG, U118MG, and U251MG) and primary glioblastoma lines. Results EGFRBi-armed ATC killed up to 85% of U87, U118, and U251 targets at effector:target ratios (E:T) ranging from 1:1 to 25:1. Engagement of tumor by EGFRBi-armed ATC induced Th1 and Th2 cytokine secretion by armed ATC. HER2Bi-armed ATC exhibited comparable cytotoxicity against U118 and U251, but did not kill HER2-negative U87 cells. HER2Bi- or EGFRBi-armed ATC exhibited 50—80% cytotoxicity against four primary glioblastoma lines as well as a temozolomide (TMZ)-resistant variant of U251. Both CD133– and CD133+ subpopulations were killed by armed ATC. Targeting both HER2Bi and EGFRBi simultaneously showed enhanced efficacy than arming with a single BiAb. Armed ATC maintained effectiveness after irradiation and in the presence of TMZ at a therapeutic concentration and were capable of killing multiple targets. Conclusion High-grade gliomas are suitable for specific targeting by armed ATC. These data, together with additional animal studies, may provide the preclinical support for the use of armed ATC as a valuable addition to current treatment regimens

The Helicobacter pylori Genome Project : insights into H. pylori population structure from analysis of a worldwide collection of complete genomes

Helicobacter pylori, a dominant member of the gastric microbiota, shares co-evolutionary history with humans. This has led to the development of genetically distinct H. pylori subpopulations associated with the geographic origin of the host and with differential gastric disease risk. Here, we provide insights into H. pylori population structure as a part of the Helicobacter pylori Genome Project (HpGP), a multi-disciplinary initiative aimed at elucidating H. pylori pathogenesis and identifying new therapeutic targets. We collected 1011 well-characterized clinical strains from 50 countries and generated high-quality genome sequences. We analysed core genome diversity and population structure of the HpGP dataset and 255 worldwide reference genomes to outline the ancestral contribution to Eurasian, African, and American populations. We found evidence of substantial contribution of population hpNorthAsia and subpopulation hspUral in Northern European H. pylori. The genomes of H. pylori isolated from northern and southern Indigenous Americans differed in that bacteria isolated in northern Indigenous communities were more similar to North Asian H. pylori while the southern had higher relatedness to hpEastAsia. Notably, we also found a highly clonal yet geographically dispersed North American subpopulation, which is negative for the cag pathogenicity island, and present in 7% of sequenced US genomes. We expect the HpGP dataset and the corresponding strains to become a major asset for H. pylori genomics