Journal: Leukemia

Article Title: AML/T cell interactomics uncover correlates of patient outcomes and the key role of ICAM1 in T cell killing of AML

doi: 10.1038/s41375-024-02255-1

Figure Lengend Snippet: A Elimination efficiency of U937 cell line after CRISPR/Cas9-mediated knock-out (KO) of selected genes by CD4 IL10 cells; each dot represents a CD4 IL10 donor ( n = 3). WT = wild type; U937 WT: positive control, K562 WT: negative control. Representative FACS plots from one donor against U937- ITGB2 -KO and U937- ICAM1 -KO are shown on the right; numbers indicate the percentage in the AML gate. B Degranulation of CD8 + T cells, expressed as % CD8 + CD107a + cells within live singlet CD3 + T cells, against U937 WT and U937- ICAM1 -KO cells; each circle represents a CD8 + T cell donor ( n = 7). Representative FACS plots from one donor are shown on the right; numbers indicate percent degranulating CD8 + T cells. C Elimination efficiency of U937 WT and U937- ICAM1 -KO cells by CD8 + T cells ( n = 10). Wilcoxon test, p < 0.05. Representative FACS plots from one donor are shown on the right; numbers indicate the percentage in the AML gate. D Elimination efficiency of U937 cell line 24 h after treatment with LFA-1 inhibitors at indicated concentrations; each line represents a CD4 IL10 donor ( n = 3); circles are color-coded by the inhibitor as indicated. Dunn’s post hoc test results are shown, following Friedman ANOVA ( p < 0.0001). E CD8 + T cells were co-cultured for 24 h with indicated target cells; activation was measured as a percent of CD69 + CD25 +/ − cells within the live CD8 + T cell gate. F Elimination efficiency of indicated target cell lines and primary AML cells by CD8 + T cells after 3 days of co-culture in the absence or presence of LFA-1 inhibitor BMS587101 (10 μM). G Spearman correlation of percent activation of CD8 + T cells and the elimination efficiency of target cells. H Timeline of mice injections; n = 10 mice per group. I Graphs showing total flux (log normalized) obtained from imaging data on days 8 and 12. 1-way ANOVA with Bonferroni multiple comparison test; p = n.s. at day 8, p = 0.0075 at day 12.

Article Snippet: When indicated, LFA-1 inhibitors (BIRT377, BMS688571, A286982 from Tocris Bioscience, UK; BMS587101 from MedChemExpress, NJ, USA) were added at 0.1, 1, and 10 μM, and EE was measured 24 h after co-culture.

Techniques: CRISPR, Knock-Out, Positive Control, Negative Control, Cell Culture, Activation Assay, Co-Culture Assay, Imaging, Comparison

Journal: British Journal of Pharmacology

Article Title: Anti‐αLβ2 antibodies reveal novel endocytotic cross‐modulatory functionality

doi: 10.1111/bph.14996

Figure Lengend Snippet: Effect of mAbs and compounds on integrin αLβ2, α4β1, and α4β7 surface expression. (a) Surface expression of αLβ2, α4β1, and α4β7 on CD2+ T cells after 24 hr of treatment with anti‐αL (efalizumab, R7.1, and TS1/22), anti‐α4 (natalizumab), anti‐α4β7 (vedolizumab) mAbs (all 10 μg·ml−1), small‐molecule inhibitors targeting αL (LFA878 and BIRT377), αLβ2 (XVA143 and lifitegrast), and α4β1/α4β7 (firategrast and RO0505376; all 10 μM) and respective controls. Each bar represents the mean value ± SEM of five independent experiments using blood samples from different donors. Statistical significance was determined by using one‐way ANOVA, *P < .05, versus control. (b) Flow cytometry dot plots of α4β1 CD2+ T cells. Representative flow cytometry dot plots of CD2+ T cells treated for 24 hr with efalizumab, R7.1, natalizumab and respective isotype controls are shown. Dots in the upper right quadrant indicate α4β1 positive CD2+ T cells. The samples were acquired using a Cytoflex cytometer and analysed using FlowJo software

Article Snippet: BIRT‐377 (Tocris Cat# 4776, PubChem CID: 9803375).

Techniques: Expressing, Control, Flow Cytometry, Cytometry, Software

Journal: British Journal of Pharmacology

Article Title: Anti‐αLβ2 antibodies reveal novel endocytotic cross‐modulatory functionality

doi: 10.1111/bph.14996

Figure Lengend Snippet: Effect of αLβ2 inhibitors on the conformational state of αLβ2, α4β1, and CD69 expression. Exposure of (a) m24 epitope, (b) MEM148 epitope, and (c) HUTS‐21 on CD2+ T cells after 40 min of treatment with anti‐αL (efalizumab, R7.1, and TS1/22), anti‐α4 (natalizumab), anti‐α4β7 (vedolizumab) mAbs (all 10 μg·ml−1), small‐molecule inhibitors targeting αL (LFA878 and BIRT377), αLβ2 (XVA143 and lifitegrast) and α4 (firategrast and RO0505376; all 10 μM), and respective controls. (d) Surface expression of CD69 on CD2+ T cells after 24 hr of treatment with efalizumab (10 μg·ml−1), natalizumab (10 μg·ml−1), and respective isotype controls (hIgG1 and hIgG4). Anti‐CD3 (OKT3) was used as positive control for the induction of CD69 surface expression. Each bar represents the mean value ± SEM of five independent experiments using blood samples from different donors. Statistical significance was determined by using one‐way ANOVA, *P < .05, versus control

Article Snippet: BIRT‐377 (Tocris Cat# 4776, PubChem CID: 9803375).

Techniques: Expressing, Positive Control, Control