A novel MVMp-based vector system specifically designed to reduce the risk of replication-competent virus generation by homologous recombination.

Recent work highlights the potential usefulness of MVM-based vectors as selective vehicles for cancer gene therapy (Dupont et al, Gene Therapy, 2000; 7: 790-796). To implement this strategy, however, it is necessary to develop optimized methods for producing high-titer, helper-free parvovirus stocks. Recombinants of MVMp (rMVMp) are currently generated by transiently co-transfecting permissive cell lines with a plasmid carrying the vector genome and a helper plasmid expressing the capsid genes (replaced with a foreign gene in the vector genome).

Xath2, a bHLH gene expressed during a late transition stage of neurogenesis in the forebrain of Xenopus embryos.

We have identified a Xenopus bHLH gene, Xath2, which is the homologue of the murine MATH-2/NEX-1 gene, using a functional expression screening approach. Overexpression of this gene in neurula embryos induces the expression of the N-tubulin neuronal marker but does not stimulate the expression of the X-ngnr-1 and NeuroD proneural genes. Expression of Xath2 begins in stage 32 embryos and is restricted to the dorsal telencephalon.

Cloning and sequencing of defective particles derived from the autonomous parvovirus minute virus of mice for the construction of vectors with minimal cis-acting sequences.

The production of wild-type-free stocks of recombinant parvovirus minute virus of mice [MVM(p)] is difficult due to the presence of homologous sequences in vector and helper genomes that cannot easily be eliminated from the overlapping coding sequences. We have therefore cloned and sequenced spontaneously occurring defective particles of MVM(p) with very small genomes to identify the minimal cis-acting sequences required for DNA amplification and virus production. One of them has lost all capsid-coding sequences but is still able to replicate in permissive cells when nonstructural proteins are provided in trans by a helper plasmid.

XSIP1, a Xenopus zinc finger/homeodomain encoding gene highly expressed during early neural development.

We have isolated a Xenopus homologue of the zinc finger/homeodomain-containing transcriptional repressor Smad-interacting protein-1 (SIP1) from mouse. XSIP1 is activated at the early gastrula stage and transcription occurs throughout embryogenesis. At the beginning of gastrulation, XSIP1 is strongly expressed in prospective neurectoderm.

Tumor-selective gene transduction and cell killing with an oncotropic autonomous parvovirus-based vector.

A recombinant MVMp of the fibrotropic strain of minute virus of mice (MVMp) expressing the chloramphenicol acetyltransferase reporter gene was used to infect a series of biologically relevant cultured cells, normal or tumor-derived, including normal melanocytes versus melanoma cells, normal mammary epithelial cells versus breast adenocarcinoma cells, and normal neurons or astrocytes versus glioma cells. As a reference cell system we used normal human fibroblasts versus the SV40-transformed fibroblast cell line NB324K. After infection, we observed good expression of the reporter gene in the different tumor cell types, but only poor expression if any in the corresponding normal cells.