Multiplexing Phenotype and Function for More Biologically Relevant Insights - 19

More Biologically Relevant Insights | APPLICATION NOTE

uncover an experimental artifact-general cytotoxicity rather than specific CMC
inhibition-that could have led to misinterpretation of the results in the absence
of the effector cell data. For this assay we selected a fixed 2:1 ratio of NK-92 cells to
Jurkat cells, which results in baseline values of ~75% of Jurkat cells staining positive
for caspase 3/7 activation, and ~60% of Jurkat cells staining positive for membrane
permeability (non-viability). The effect of pathway inhibitor compounds on CMC

Figure 5. Pathway-specific inhibition of CMC. Four different compounds show varying
degrees of CMC inhibition on Jurkat target cells (blue curves) as assessed by cell membrane
permeability (A, C, E, G) and caspase 3/7 activation (B, D, F, H). For each data point n = 3, with

19

| January, 2019

can be seen in Figure 5, with increasing inhibitor concentration generally resulting
in less cell membrane permeability (Figure 5A, C, E, and G) and caspase 3/7 activation (Figure 5B, D, F, and H) in Jurkat cells. There are two intriguing exceptions.
Figure 5A and B show the effects of sunitinib on both Jurkat target cells and NK-92
effector cells. While this tyrosine kinase inhibitor appears to have no effect on CMC
based on the stable levels of membrane permeability and caspase 3/7 activation

the data fit using a four parameter logistic nonlinear regression model. Notably, sunitinib
and U73122 also have an effect on the NK-92 effector cells (red curves), which would not
have been detected using a traditional Cr51 release assay.



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Multiplexing Phenotype and Function for More Biologically Relevant Insights

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