TABLE 2

Cell-based counter assays

Inhibition by R406 was tested in cell-based counter assays measuring a variety of Syk-independent signaling pathways. Each assay was optimized independently and run with the appropriate controls.

Cell Type Stimulation Receptor Type Readout EC50 S.D.
μM
CHMC Anti-IgE ITAM (FcϵRI) Degranulation 0.056 0.02
Ramos B-cell line IL-4 Cytokine (IL-4R) CD23 up-regulation 0.192 0.01
Primary human T cells IL-2 Cytokine (IL-2R) Cell proliferation 0.190 0.07
Primary human T cells αCD3/CD28 ITAM (TCR/CD28) IL-2 production 0.448 0.09
A549 epithelial line TNFα Cytokine (TNFR) CD54 up-regulation 4.8 1.82
A549 epithelial line IL-1 Cytokine (IL-1R) CD54 up-regulation 7.87 3.76
A549 epithelial line IFN-γ Cytokine (IFNR) CD54 up-regulation 2.14 0.78
Jurkat T cell line PMA PKC CD69 up-regulation >20
Primary human neutrophils PMA PKC Oxidative burst >10
CHMC Ionomycin Calcium channels Degranulation >10
Monocytic line, THP-1 LPS TLR-4 TNFα 2.10 1.2
Primary human macrophage LPS TLR-4 TNFα 1.15 0.1
Huh7 hepatocyte line Serum General cell growth Cell proliferation 15.1 7.48
A549 epithelial line Serum General cell growth Cell proliferation 2.9 0.57
H1299 lung cancer line Serum General cell growth Cell proliferation 6.3 3.5
HeLa Insulin Insulin receptor P-AKT (ICW) 1.19 0.28
HeLa EGF EGF receptor P-EGFR (ICW) 17.46 11.57
  • ICW, In-cell Western; EGF, epidermal growth factor; EGFR, EGF receptor; PKC, protein kinase C.