Toxin-Based Therapeutic Approaches

Shapira, Assaf; Benhar, Itai

In another study, which was performed by Law et al [434] , HIV protease-activated toxins were designed on the basis of the type III maize ribosome-inactivating protein, which in its native form is synthesized as inactive precursor (pro-RIP) and is activated by proteolytic removal of a 25 amino-acid inhibitory internal peptide in-planta [218, 219] . In this work, the first and the last 10 residues of the internal inactivation region were replaced with two HIV-PR recognition sequences, and an 11 amino-acids transduction peptide derived from the HIV-1 Tat protein was fused to the N-termini of the modified RIPs for promoting cell entry [519] . These modifications resulted in the generation of re-engineered pro-RIPs which underwent an efficient cleavage in vitro by recombinant HIV-PR or in vivo (in HIV infected cells) by the viral encoded protease. Upon treatment of infected cells, the N-glycosidase and anti-viral activities of the modified cleavable RIPs were found to be higher than those of an uncleavable-nonactivated pro-RIP and resembled those of an activated mutant in which the inhibitory region was genetically removed. The cytotoxic activity of the cleavable RIPs against infected cells was not reported, but cytotoxicity toward uninfected cells was found to be lower than that of the activated mutant. Thus, the fact that the cleavable toxins behave more like a constitutively active form toward infected cells and in a similar way to a non-activated form toward uninfected cells, may suggest that they are specifically activated by the HIV protease and demonstrates the possibility of using this cleavable internal inactivation region as a switch to activate the toxin inside infected cells [434] .