Figure 4.
IL10RA overexpression modulates sensitivity to ALK inhibition. (A) Fold change in expression levels of IL10RA for each of the 3 sgRNAs targeting IL10RA vs nontargeting (NT) control sgRNA in the indicated ALCL cell lines. Data are means ± standard deviation (SD) (n = 3). (B) Viability of SUP-M2 cells based on normalized CellTiter-Blue fluorescence reads on exposure to increasing concentrations of crizotinib, alectinib, brigatinib, or lorlatinib for 48 hours when expressing 1 of 3 of the indicated sgRNAs inducing overexpression of IL10RA in the presence or absence of 10 ng/mL IL-10. Data are means ± SD (n = 3). (C) Proliferation of ALCL cell lines expressing sgRNAs inducing overexpression of IL10RA over 4 days. Data are means ± SD (n = 3). (D) Modulation of apoptosis upon expression of sgRNAs inducing overexpression of IL10RA in the indicated ALCL cell lines. The percentage of apoptotic cells was determined by APC-annexin V and propidium iodide (PI) staining of ALCL cells treated with 125 nM crizotinib (SU-DHL-1), 312.5 nM crizotinib (SUP-M2) or 1250 nM crizotinib (DEL) for 48 hours (left panel). Data are means ± SD of technical replicates; experiments were performed independently 3 times. Representative flow cytometry plots of APC-annexin V/PI staining intensities corresponding to IL10RA sgRNA promoting survival vs NT control sgRNA in SUP-M2 cells (right panel). (E) Modulation of apoptotic response upon expression of sgRNAs inducing overexpression of IL10RA in the indicated ALCL cell lines. The percentage of apoptotic cells was determined by annexin V and PI staining of ALCL cells treated with 125 nM crizotinib (SU-DHL-1), 312.5 nM crizotinib (SUP-M2) or 1250 nM crizotinib (DEL) in the presence of 10 ng/mL IL-10 for 48 hours. Data are means ± SD of technical replicates; experiments were performed independently 3 times. (F) Results from the mini CRISPR-Cas9–based knockout screen. Read counts of 6 sgRNAs targeting IL10/IL10RA/IL10RB in the SUP-M2–derived TS cell line before and after a 14-day incubation with DMSO or 80 nM crizotinib. Data are means ± SD with read counts for individual sgRNAs (n = 6) plotted as dots. *P < .05. **P < .01, ***P < .001, Welch 2-sample t test (B,D-E); unpaired Welch-corrected t test (F). ns, not significant.

IL10RA overexpression modulates sensitivity to ALK inhibition. (A) Fold change in expression levels of IL10RA for each of the 3 sgRNAs targeting IL10RA vs nontargeting (NT) control sgRNA in the indicated ALCL cell lines. Data are means ± standard deviation (SD) (n = 3). (B) Viability of SUP-M2 cells based on normalized CellTiter-Blue fluorescence reads on exposure to increasing concentrations of crizotinib, alectinib, brigatinib, or lorlatinib for 48 hours when expressing 1 of 3 of the indicated sgRNAs inducing overexpression of IL10RA in the presence or absence of 10 ng/mL IL-10. Data are means ± SD (n = 3). (C) Proliferation of ALCL cell lines expressing sgRNAs inducing overexpression of IL10RA over 4 days. Data are means ± SD (n = 3). (D) Modulation of apoptosis upon expression of sgRNAs inducing overexpression of IL10RA in the indicated ALCL cell lines. The percentage of apoptotic cells was determined by APC-annexin V and propidium iodide (PI) staining of ALCL cells treated with 125 nM crizotinib (SU-DHL-1), 312.5 nM crizotinib (SUP-M2) or 1250 nM crizotinib (DEL) for 48 hours (left panel). Data are means ± SD of technical replicates; experiments were performed independently 3 times. Representative flow cytometry plots of APC-annexin V/PI staining intensities corresponding to IL10RA sgRNA promoting survival vs NT control sgRNA in SUP-M2 cells (right panel). (E) Modulation of apoptotic response upon expression of sgRNAs inducing overexpression of IL10RA in the indicated ALCL cell lines. The percentage of apoptotic cells was determined by annexin V and PI staining of ALCL cells treated with 125 nM crizotinib (SU-DHL-1), 312.5 nM crizotinib (SUP-M2) or 1250 nM crizotinib (DEL) in the presence of 10 ng/mL IL-10 for 48 hours. Data are means ± SD of technical replicates; experiments were performed independently 3 times. (F) Results from the mini CRISPR-Cas9–based knockout screen. Read counts of 6 sgRNAs targeting IL10/IL10RA/IL10RB in the SUP-M2–derived TS cell line before and after a 14-day incubation with DMSO or 80 nM crizotinib. Data are means ± SD with read counts for individual sgRNAs (n = 6) plotted as dots. *P < .05. **P < .01, ***P < .001, Welch 2-sample t test (B,D-E); unpaired Welch-corrected t test (F). ns, not significant.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal