Poor intercellular transport and absence of enhanced antiproliferative activity after non-viral gene transfer of VP22-P53 or P53-VP22 fusions into p53 null cell lines in vitro or in vivo.

BACKGROUND: The herpes simplex virus type 1 (HSV-1) VP22 protein has the property to mediate intercellular trafficking of heterologous proteins fused to its C- or N-terminus. We have previously shown improved delivery and enhanced therapeutic effect in vitro and in vivo with a P27-VP22 fusion protein. In this report, we were interested in studying the spread and biological activity of VP22 fused to the P53 tumor suppressor. METHODS: Expression of the VP22-P53 and P53-VP22 fusion proteins was shown by Western blot and intercellular spreading was monitored by immunofluorescence on transiently transfected cells. In vitro antiproliferative activity of wild-type (wt) P53 and P53-VP22 was assessed by proliferation assays and transactivating ability was studied by a reporter gene test and a gel-shift assay. Antitumor activity was also tested in vivo by intratumoral injections of naked DNA in a model of subcutaneous tumors implanted in nude mice. RESULTS: Our results show that the C-terminal fusion or the N-terminal P53-VP22 fusion proteins are not able to spread as efficiently as VP22. Moreover, we demonstrate that VP22-P53 does not possess any transactivating ability. P53-VP22 has an antiproliferative activity, but this activity is not superior to the one of P53 alone, in vitro or in vivo. CONCLUSIONS: Our study indicates that a gene transfer strategy using VP22 cannot be considered as a universal system to improve the delivery of any protein.

VP22-mediated and light-activated delivery of an anti-c-raf1 antisense oligonucleotide improves its activity after intratumoral injection in nude mice.

VP22, a protein of the herpes simplex virus tegument, can form complexes with fluorescein-labeled oligonucleotides. These particles, termed "Vectosomes," are efficiently taken up by cells and remain stable in the cell cytoplasm without any particular activity. Interestingly, these Vectosomes can be disrupted by light, which releases the antisense activity. Here we show that anti-c-raf1 Vectosomes are efficiently activated by light in vivo after injection into subcutaneous A549 (non-small-cell lung cancer) tumors implanted in nude mice. Moreover, two injections per week of anti-c-raf1 Vectosomes followed by illumination result in a stronger inhibition of tumor growth than injections of the antisense alone or of the different control Vectosomes. This effect correlates with a strong inhibition of the c-Raf1 protein expression. As a consequence of c-Raf1 loss, apoptosis was also detected in these tumors. Vectosomes thus represent a new powerful tool to improve the delivery of oligonucleotides in vitro and in vivo.