Journal: Cancers

Article Title: Drug-Induced Resistance and Phenotypic Switch in Triple-Negative Breast Cancer Can Be Controlled via Resolution and Targeting of Individualized Signaling Signatures

doi: 10.3390/cancers13195009

Figure Lengend Snippet: TNBC tissues are represented by different patient-specific signaling signatures, majority of which do not include EGFR. ( A ) Fold changes in expression levels of EGFR and pEGFR in TNBC and non-TNBC tumors are shown. Y1068 and Y1173 are EGFR phosphorylation sites; ( B ) Examples for patient-specific sets of active unbalanced processes are shown. Each sample harbors a set of 1–3 active unbalanced processes (PaSSS), represented schematically by a barcode. In each barcode active unbalanced processes are represented by black or gray squares, inactive white. Negative/positive amplitude denotes how the patients are correlated with respect to a particular process. Suggested PaSSS-based therapies appear below each barcode; ( C ) Heterogeneity index of 3 subgroups, represented by a ratio between the number of distinct PaSSSs and the number of samples in each subset, is shown for the TNBC subset of tissues, the entire set (3467 samples from 11 cancer types) and the subset of non-TNBC samples. (Abbreviations: TNBC—Triple Negative Breast Cancer, PaSSS—Patient-specific signaling signature, EGFR—Epidermal Growth Factor Receptor, VEGFR2—Vascular Endothelial Growth Factor Receptor 2, Her2—Human Epidermal growth factor Receptor 2, Src—Proto-oncogene tyrosine-protein kinase Src).

Article Snippet: The following conjugated antibodies were used: anti-p-EGFR (Y1068) (R&D Systems, Minneapolis, MN, USA, cat. no. IC3570G), anti-p-ERK2 (Thr202/Tyr204) (BioLegend, San Diego, CA, USA, cat. No. 675503), anti-p-S6 (Ser235/236) (BioLegend, cat. no. 608605), and anti-GAPDH (Santa Cruz Biotechnology, Dallas, Texas, USA, cat. no. sc-47724AF594).

Techniques: Expressing, Phospho-proteomics

Journal: British journal of pharmacology

Article Title: Veklury® (remdesivir) formulations inhibit initial membrane-coupled events of SARS-CoV-2 infection due to their sulfobutylether-β-cyclodextrin content.

doi: 10.1111/bph.16063

Figure Lengend Snippet: FIGURE 2 Methyl-β cyclodextrin (MβCD)-induced reductions in ACE2 binding of Wuhan-Hu-1 strain of SARS-CoV-2 (WT) and Delta SARS- CoV-2 spike receptor-binding domains (RBDs) negatively correlate with the applied RBD concentrations. HEK/ACE2 + TMPRSS2 cells were treated with different concentrations of MβCD ranging between 0.25 and 10 mM. Fluorescence intensities of cells were measured using flow cytometry for 5 min continuously starting immediately after the addition of GFP-conjugated WT (a) or Delta (b) SARS-CoV-2 spike RBDs applied at various concentrations between 0.1 and 5 μgml1, or 0.05 and 2.5 μgml1, respectively. Moving averages of time-correlated fluorescence intensities of at least 50,000 cells of normal morphology were determined with a window size of 10 s and subsequently normalized to the average of intensities obtained in the last time window of untreated control samples. Mean ± SD values were calculated from data of six independent samples per treatment condition and plotted as a function of time (starting 20 s after RBD addition). The extents of inhibition of WT (a) and Delta (b) RBD binding at the end of the measurement time in response to various MβCD concentrations were calculated and plotted as mean ± SD as a function of the applied RBD concentration in the right panels. Asterisks indicate significant differences of samples treated with the lowest versus highest RBD concentrations at each applied CD concentration (*P < 0.05; two-way ANOVA followed by Tukey's HSD test). See details of statistical analysis in Table S2.

Article Snippet: When using Omicron spike trimer conjugated with Alexa Fluor 488 (R&D Systems; AFG11060), experiments were repeated after SBECD, Veklury® P or Veklury® S treatments in HEK/ACE2 + TMPRSS2 and Calu-3 cells using the trimer at 0.5 and 2.5 μg ml 1, respectively.

Techniques: Binding Assay, Fluorescence, Flow Cytometry, Control, Inhibition, Concentration Assay

Journal: British journal of pharmacology

Article Title: Veklury® (remdesivir) formulations inhibit initial membrane-coupled events of SARS-CoV-2 infection due to their sulfobutylether-β-cyclodextrin content.

doi: 10.1111/bph.16063

Figure Lengend Snippet: FIGURE 6 Veklury® (remdesivir) formulations effectively decrease ACE2 binding of Omicron SARS-CoV-2 spike receptor-binding domain (RBD) and the cellular uptake of Omicron SARS-CoV-2 spike trimer due to their SBECD content. (a) HEK/ACE2 + TMPRSS2 (left panel) or Calu-3 cells (right panel) were treated with SBECD, Veklury® P (powder) or Veklury® S (solution) at CD concentrations of 1, 5 and 10 mM. Then, HEK/ ACE2 + TMPRSS2 cells were incubated in the presence of 0.2-μgml1 Omicron SARS-CoV-2 spike RBD–GFP, whereas Calu-3 cells were labelled with 1-μgml1 Omicron RBD–GFP for 4 min. The emitted fluorescence intensities of individual cells were subsequently measured using flow cytometry, and the mean intensity was calculated from data of at least 10,000 cells of normal morphology per sample. The average values of six independent measurements (±SD) were calculated, normalized to the mean value determined in untreated control samples and plotted as a function of the applied concentrations of CD. Asterisks indicate significant differences of samples treated with the highest applied concentrations compared to the control samples (*P < 0.05; two-way ANOVA followed by Tukey's HSD test). See details of statistical analysis in Table S6. (b) HEK/ACE2 + TMPRSS2 (left panel) or Calu-3 cells (right panel) treated with SBECD, Veklury® P or Veklury® S at CD concentrations of 1 and 5 mM for 1 h were subsequently incubated for 4 h in the presence of Omicron SARS-CoV-2 spike trimers conjugated with Alexa Fluor 488. The average fluorescence intensity values emitted by Alexa Fluor 488-labelled trimers were calculated using data of intracellular pixels identified using the membrane marker N-[3-(40-dihexylamino-3-hydroxy-flavonyl-6-oxy)-propyl]N,N-dimethyl-N-(3-sulfopropyl)-ammonium inner salt (F66) as described in Figure 5 for individual cells, which were subsequently normalized to the median value determined in untreated control samples. The number of cells obtained from five independent experiments and involved in the analysis was from left to right: 382, 348, 373, 396, 388, 402 and 393 for Omicron trimer uptake in HEK/ACE2 + TMPRSS2 and 308, 250, 256, 223, 222, 232 and 296 for Omicron trimer uptake in Calu-3. Data points indicated in the figure were obtained from individual cells and plotted along with median values with quartiles. Grey shaded areas show ranges between 0.5 and 1. Asterisks indicate significant differences compared to the control samples (*P < 0.05; one-way ANOVA followed by Tukey's HSD test). See details of statistical analysis in Table S6.

Article Snippet: When using Omicron spike trimer conjugated with Alexa Fluor 488 (R&D Systems; AFG11060), experiments were repeated after SBECD, Veklury® P or Veklury® S treatments in HEK/ACE2 + TMPRSS2 and Calu-3 cells using the trimer at 0.5 and 2.5 μg ml 1, respectively.

Techniques: Binding Assay, Incubation, Fluorescence, Flow Cytometry, Control, Membrane, Marker