Figure 3
Figure 3. CT-011 and lenalidomide enhance NK-cell trafficking toward MM cell line media and MM patient serum. (A) To test whether or not CT-011 affects NK-cell trafficking, Transwell migration assays were conducted whereby NK cells were cultured in RPMI 1640 media with 10% FBS and 150 IU/mL IL-2 with isotype control or CT-011 for 72 hours. Data shown summarize 3 independent experiments. All differences within treatment conditions are statistically significant. Across treatment conditions, CT-011 enhanced migration beyond control into U266 media (*P < .05) and MM patient serum (**P < .05). (B) By flow cytometry, we systematically evaluated for changes in the surface expression of trafficking antigens known to be expressed by human NK cells. Only CXCR4 showed a statistically significant increase in expression in response to 72 hours in 150 IU/mL IL-2 with CT-011, P < .01 (representative data from one patient shown, n = 5). (C) Transwell migration assays were conducted with NK cells pretreated for 72 hours in 150 IU/mL IL-2 with control or CT-011 into normal media or media enriched with SDF-1α. CT-011 enhanced NK-cell trafficking into SDF-1α–enriched media (, data collated from 3 independent experiments), suggesting a functional relevance to the increase in CXCR4 expression observed on NK cells in response to these agents. CT-011 enhanced migration over control condition (*P < .05). However, when NK cells were pretreated with the CXCR4 inhibitor AMD-3100 (10 μg/mL) for 90 minutes before assay, trafficking was virtually abolished in all conditions (, data from 2 independent experiments, pairwise comparisons within treatments; P < .05).

CT-011 and lenalidomide enhance NK-cell trafficking toward MM cell line media and MM patient serum. (A) To test whether or not CT-011 affects NK-cell trafficking, Transwell migration assays were conducted whereby NK cells were cultured in RPMI 1640 media with 10% FBS and 150 IU/mL IL-2 with isotype control or CT-011 for 72 hours. Data shown summarize 3 independent experiments. All differences within treatment conditions are statistically significant. Across treatment conditions, CT-011 enhanced migration beyond control into U266 media (*P < .05) and MM patient serum (**P < .05). (B) By flow cytometry, we systematically evaluated for changes in the surface expression of trafficking antigens known to be expressed by human NK cells. Only CXCR4 showed a statistically significant increase in expression in response to 72 hours in 150 IU/mL IL-2 with CT-011, P < .01 (representative data from one patient shown, n = 5). (C) Transwell migration assays were conducted with NK cells pretreated for 72 hours in 150 IU/mL IL-2 with control or CT-011 into normal media or media enriched with SDF-1α. CT-011 enhanced NK-cell trafficking into SDF-1α–enriched media (, data collated from 3 independent experiments), suggesting a functional relevance to the increase in CXCR4 expression observed on NK cells in response to these agents. CT-011 enhanced migration over control condition (*P < .05). However, when NK cells were pretreated with the CXCR4 inhibitor AMD-3100 (10 μg/mL) for 90 minutes before assay, trafficking was virtually abolished in all conditions (, data from 2 independent experiments, pairwise comparisons within treatments; P < .05).

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