Z-IETD-FMK (Z-IE(OMe)TD(OMe)-FMK) is a cell-permeable, selective inhibitor of caspase 8.

Z-IETD-FMK, which inhibits the cleavage of caspase-8 and partially inhibits the cleavage of caspase-3 and PARP, prevents the execution of apoptosis in retinal cells exposed to different apoptotic stimuli. [1] Z-IETD-FMK (50 μM) reduces ceramide-induced cardiomyocyte death and significantly inhibits the activation of caspase 3. [2] Inhibition of caspase-8 by Z-IETD-FMK affects the generation of activated/memory T cells and T cell cytokine production, and decreases NF-kappaB responses to TCR:CD3 engagement in T cell cultures. [3]

In vivo, inhibition of caspase-8 by Z-IETD-FMK reduces memory/activated CD4 and CD8 T cells, and increases susceptibility to T. cruzi infection. [3] Z-IETD-FMK promotes neuronal survival and regeneration of injured retinal ganglion cells after CNS injuries. [4]

Proteasome assay: Exponentially growing cells on a 96-well clustered plate are treated with different concentrations of drugs or left untreated (control) for 6 hours. Proteasomes extracted with 0.5% NP40 buffer are mixed with equal amounts of samples in 100 μL total volume, and then incubated with 25 μmol/L of fluorogenic substrates (LRR- specific for trypsin-like activity, LLE-specific for caspase-like activity, and SUVY-specific for chymotrypsin-like activity) in black-bottom 96-well plates at 37°C. Fluorescence is monitored every 5 minutes at the wavelength of 360 nm (excitation) and 480 nm (emission).