Antonia Franceschi

Self-potentiation of ligand-toxin conjugates containing ricin A chain fused with viral structures.

A chimeric protein was obtained by fusing together the ricin toxin A chain (RTA) gene and a DNA fragment encoding the N terminus of protein G of the vesicular stomatitis virus. Chimeric RTA (cRTA) retained full enzymic activity in a cell-free assay, but was 10-fold less toxic against human leukemic cells than either native RTA (nRTA) or unmodified recombinant RTA (rRTA). However, conjugates made with cRTA and human transferrin (Tfn) showed 10-20-fold greater cell killing efficacy than Tfn-nRTA or Tfn-rRTA conjugates despite equivalent binding of the three conjugates to target tumor cells. As a consequence, by fusion of the KFT25 peptide to the RTA sequence, the specificity factor (i.e. the ratio between nonspecific and specific cytotoxicity) of Tfn-cRTA was increased 90-240 times with respect to those of Tfn-nRTA and Tfn-rRTA. cRTA interacted with phospholipid vesicles with 15-fold faster kinetics than nRTA at acidic pH. Taken together, our results suggest that the ability of vesicular stomatitis virus protein G to interact with cell membranes can be transferred to RTA to facilitate its translocation to the cell cytosol. Our strategy may serve as a general approach for potentiating the cytotoxic efficacy of antitumor immunotoxins.

Enhancement of ricin toxin a chain immunotoxin activity: Synthesis, ionophoretic ability, and in vitro activity of monensin derivatives

Site-selective toxin delivery was achieved by coupling monoclonal antibody to the A chain subunit of ricin (RTA-IT). The cell-killing potency of RTA-IT can be drastically increased in vitro by using ionophores such as monensin. To reduce the intrinsic toxicity of monensin and to enhance its in vitro and in vivo activity, we synthesized 7 derivatives characterized by different lipophilicities. These derivatives were also analyzed for ionophoretic activity on intact cells, toxicity, and RTA-IT-enhancing activity. Two different RTA-IT were assayed on a human leukemia cell line. A correlation between lipophilicity, ionophoretic activity, and RTA-IT enhancement was observed. The compounds with the highest polar charge showed low intrinsic toxicity, revealed moderate ionophoretic activity, and were able to enhance RTA-IT only at high concentrations, whereas more lipophilic compounds (with a C28 tail or a phenyl group) showed significant ionophoretic activity and good enhancing properties.

Hepcidin and DNA promoter methylation in hepatocellular carcinoma

The liver hormone hepcidin regulates iron homoeostasis that is often altered in hepatocellular carcinoma (HCC). Epigenetic phenomena control gene expression through a dynamic fashion; therefore, considering the plasticity of both iron homoeostasis and epigenetic mechanisms and their role in liver carcinogenesis, we investigated whether hepcidin gene (HAMP) expression is modulated by DNA methylation, thus affecting iron status in human HCC.