No abstract

Hepatocyte growth factor (HGF) and macrophage-stimulating protein (MSP) have an intrinsic dual nature: they are trophic cytokines preventing apoptosis on one side and scatter factors promoting invasion on the other. For therapeutic use, their anti-apoptotic activity must be separated from their pro-invasive activity. To this end, we engineered chimeric factors containing selected functional domains of HGF and/or MSP in different combinations, and tested their biological activity. Here we present a chimeric cytokine derived from the alpha-chains of HGF and MSP, named Metron factor 1 for its ability to concomitantly activate the HGF receptor (Met) and the MSP receptor (Ron). We provide evidence that Metron factor 1 prevents apoptosis and stimulates cell proliferation at nanomolar concentrations, but is devoid of any pro-invasive activity. In an in vivo murine model of drug-induced nephrotoxicity, intravenous injection of recombinant Metron factor 1 prevented renal damage and preserved tubular integrity.

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Ras proteins are small GTPases playing a pivotal role in cell proliferation and di erentiation. Their activation state depends on the competing action of GTPase Activating Proteins (GAP) and Guanine nucleotide Exchange Factors (GEF). A tryptophan residue (Trp1056 in CDC25 Mm -GEF), conserved in all rasspeci®c GEFs identi®ed so far has been previously shown to be essential for GEF activity. Its substitution with glutamic acid results in a catalytically inactive mutant, which is able to e ciently displace wild-type GEF from p21 ras and to originate a stable ras/GEF binary complex due to the reduced a nity of the nucleotide-free ras/ GEF complex for the incoming nucleotide. We show here that this`ras-sequestering property' can be utilized to attenuate ras signal transduction pathways in mouse ®broblasts transformed by oncogenic ras. In fact overexpression of the dominant negative GEF W1056E in stable transfected cells strongly reduces intracellular ras´GTP levels in k-ras transformed ®broblasts. Accordingly, the transfected ®broblasts revert to wild-type phenotype on the basis of morphology, cell cycle and anchorage independent growth. The reversion of the transformed phenotype is accompanied by DNA endoreduplication. The possible use of dominant negative ras-speci®c GEFs as a tool to down-regulate tumor growth is discussed.

No abstract