Suppression of human T cell proliferation by the caspase inhibitors, z-VAD-FMK and z-IETD-FMK is independent of their caspase inhibition properties

C.P Lawrence,S.C Chow

doi:10.1016/j.taap.2012.09.002

C.P Lawrence, S.C Chow

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https://doi.org/10.1016/j.taap.2012.09.002

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The caspase inhibitors, benzyloxycarbony (Cbz)-l-Val-Ala-Asp (OMe)-fluoromethylketone (z-VAD-FMK) and benzyloxycarbonyl (Cbz)-Ile-Glu (OMe)-Thr-Asp (OMe)-FMK (z-IETD-FMK) at non-toxic doses were found to be immunosuppressive and inhibit human T cell proliferation induced by mitogens and IL-2 in vitro. Both caspase inhibitors were shown to block NF-κB in activated primary T cells, but have little inhibitory effect on the secretion of IL-2 and IFN-γ during T cell activation. However, the expression of IL-2 receptor α-chain (CD25) in activated T cells was inhibited by both z-VAD-FMK and z-IETD-FMK, whereas the expression of the early activated T cell marker, CD69 was unaffected. During primary T cell activation via the antigen receptor, both caspase-8 and caspase-3 were activated and processed to their respective subunits, but neither caspase inhibitors had any effect on the processing of these two caspases. In sharp contrast both caspase inhibitors readily blocked apoptosis and the activation of caspases during FasL-induced apoptosis in activated primary T cells and Jurkat T cells. Collectively, the results demonstrate that both z-VAD-FMK and z-IETD-FMK are immunosuppressive in vitro and inhibit T cell proliferation without blocking the processing of caspase-8 and caspase-3.

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The important role of caspases, caspase-8 in T cell activation and proliferation is firmly established (Chun et al, 2002)
In the present study we examined the immunosuppressive properties of the peptidyl-FMK caspase inhibitors, z-VAD-FMK and z-IETD-FMK, and determined whether their inhibition of mitogeninduced T cell proliferation is due to the blocking of caspase processing during T cell activation
About 9% of control activated T cells took up propidium iodide (PI) after activation, whereas in the presence of 100 μM of z-VAD-FMK and z-IETD-FMK cell death increased to 18% and 23%, respectively (Fig. 1C)

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Introduction

The important role of caspases, caspase-8 in T cell activation and proliferation is firmly established (Chun et al, 2002). Besides blocking mitogen-induced T cell proliferation (Chun et al, 2002; Falk et al, 2004) these caspase inhibitors were shown to reduce the expression of the α-subunit of the IL-2 receptor, CD25 and inhibit IL-2 secretion in activated T cells (Falk et al, 2004; Kennedy et al, 1999). All peptidyl-FMK caspase inhibitors contain a peptide sequence based on the target cleavage sequence of the substrate and act as competitive inhibitors by mimicking the substrate. These enzymes recognise a sequence of four amino acids in the substrates, designated P4-P3-P2-P1 and cleave substrates after an Asp residue at P1 (Yuan et al, 1993).

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