Zonal necrosis prevented by transduction of the artificial anti-death FNK protein

Abstract

Protection of cells from necrosis would be important for many medical applications. Here, we show protein transduction domain (PTD)-FNK therapeutics based on protein transduction to prevent necrosis and acute hepatic injury with zonal death induced by carbon tetrachloride (CCl 4 ). PTD-FNK is a fusion protein comprising the HIV/Tat PTD and FNK, a gain-offunction mutant of anti-apoptotic Bcl-x L . PTD-FNK protected hepatoma HepG2 from necrotic death induced by CCl 4 , and additionally, increased the apoptotic population among cells treated with CCl 4 . A concomitant treatment with a pancaspase inhibitor Z-VAD-FMK (N-benzyloxycarbonyl-Val-AlaAsp-fluoromethylketone), which alone could not prevent the necrosis, protected these cells from the apoptosis. When preinjected intraperitoneally, PTD-FNK markedly reduced zonal liver necrosis caused by CCl 4 . Moreover, injection of PTD-FNK accompanied by Z-VAD-FMK suppressed necrotic injury even after CCl 4 administration. These results suggest that PTD-FNK has great potential for clinical applications to prevent cell death, whether from apoptosis or necrosis, and organ failure.

Transduction of anti-cell death protein FNK protects isolated rat hearts from myocardial infarction induced by ischemia/reperfusion

“…PTD-FNK affected the movement of calcium ions, leading to the protection of primary neurons against glutamate toxicity, directly or indirectly (Asoh et al, 2002). We also demonstrated that PTD-FNK retains the mitochondrial membrane potential and the intracellular ATP level to protect hepatoma cells from necrosis induced by carbon tetrachloride (Asoh et al, 2005). In the ischemia/reperfusion injury model using Langendorff's heart, it is likely that PTD-FNK contributes to calcium homeostasis and/or mitochondrial activity to inhibit myocardial cell death, regardless of the form of cell death.…”

“…PTD-FNK was shown to suppress hepatic necrosis caused by carbon tetrachloride in vitro and in vivo as well as apoptosis (Asoh et al, 2005). Importantly, PTD-FNK is effective for post-treatment (Asoh et al, 2005). Moreover, PTD-FNK protected neurons from delayed neuronal cell death caused by ischemic brain injury, when intraperitoneally injected into animal models (Asoh et al, 2002).…”

“…Ozaki et al (2004) also reported that the PTD-FNK protein protects chondrocytes in cartilage slices from cell death induced by anti-Fas antibody and nitrogen oxide. PTD-FNK was shown to suppress hepatic necrosis caused by carbon tetrachloride in vitro and in vivo as well as apoptosis (Asoh et al, 2005). Importantly, PTD-FNK is effective for post-treatment (Asoh et al, 2005).…”

Topical application of the antiapoptotic TAT-FNK protein prevents aminoglycoside-induced ototoxicity

“…7,8 We first constructed a powerful artificial antiapoptotic protein, FNK (originally designated Bcl-xFNK by Asoh et al 9 ), which has three amino-acid substitutions, Tyr-22 to Phe(F), Gln-26 to Asn(N) and Arg-165 to Lys(K), to strengthen the cytoprotective activity of Bcl-x L . We then demonstrated that fusion of FNK with TAT enabled FNK to penetrate highly negatively charged chondrocytes 9,10 and the blood --brain barrier, 11 and that TAT-FNK showed an antiapoptotic effect in a model of brain and hepatic ischemia. 11,12 When injected intraperitoneally into guinea pigs in vivo, we observed that TAT-FNK was distributed widely in the cochlea and that it reduced the expression of cleaved poly-(ADP-ribose)-polymerase (PARP), auditory brainstem response (ABR) threshold shifts, and HC loss induced by a combination of ethacrynic acid (EA) and kanamycin sulfate (KM), in vivo.…”

Mitochondrial drug delivery and mitochondrial disease therapy – An approach to liposome-based delivery targeted to mitochondria

“…Asoh and co-workers showed that the PTD-fused FNK protein, constructed from Bcl-X L by the site-directed mutagenesis of three amino acids improves cytoprotective activity Yamada, Y., et al 10 (Asoh et al, 2000), protected a certain type of cells from both necrotic and apoptotic cell death (Asoh et al, 2002, Asoh et al, 2005 (Fig. 1 C).…”