RESUMO
Targeted tumor therapy can provide the basis for the inhibition of tumor growth. However, a number of toxin-based therapeutics lack efficacy because of insufficient endosomal escape after being internalized by endocytosis. To address this problem, the potential of glycosylated triterpenoids, such as SO1861, as endosomal escape enhancers (EEE) for superparamagnetic iron oxide nanoparticle (SPION)-based toxin therapy was investigated. Herein, two different SPION-based particle systems were synthesized, each selectively functionalized with either the targeted toxin, dianthin-epidermal growth factor (DiaEGF), or the EEE, SO1861. After applying both particle systems in vitro, an almost 2000-fold enhancement in tumor cell cytotoxicity compared to the monotherapy with SPION-DiaEGF and a 6.7-fold gain in specificity was observed. Thus, the required dose of the formulation was appreciably reduced, and the therapeutic window widened.
RESUMO
Protein-based targeted toxins play an increasingly important role in targeted tumor therapies. In spite of their high intrinsic toxicity, their efficacy in animal models is low. A major reason for this is the limited entry of the toxin into the cytosol of the target cell, which is required to mediate the fatal effect. Target receptor bound and internalized toxins are mostly either recycled back to the cell surface or lysosomally degraded. This might explain why no antibody-targeted protein toxin has been approved for tumor therapeutic applications by the authorities to date although more than 500 targeted toxins have been developed within the last decades. To overcome the problem of insufficient endosomal escape, a number of strategies that make use of diverse chemicals, cell-penetrating or fusogenic peptides, and light-induced techniques were designed to weaken the membrane integrity of endosomes. This review focuses on glycosylated triterpenoids as endosomal escape enhancers and throws light on their structure, the mechanism of action, and on their efficacy in cell culture and animal models. Obstacles, challenges, opportunities, and future prospects are discussed.
RESUMO
The therapeutic relevance of immunotoxins is based on the conjugation of monoclonal antibodies to toxins. In cancer therapies, the conjugated antibodies not only direct the binding of immunotoxins to cancer-specific receptors and mediate the elimination of tumor cells through the innate immune system, but also increase target cytotoxicity by the intrinsic toxin activity. In the present study, the therapeutic antibodies Cetuximab (anti-EGFR, Erbitux(®)), Panitumumab (anti-EGFR, Vectibix(®)) and Trastuzumab (anti-HER2, Herceptin(®)) were chemically conjugated to the toxin dianthin. In the first instance, recombinant dianthin was characterized by mass spectrometry and its stability was analyzed by circular dichroism. Dianthin showed increased cytotoxicity on MCF-7 cells when tested in combination with a glycosylated triterpenoid (SO1861) in a real-time impedance-based cytotoxicity assay. In data obtained by live cell imaging, SO1861 specifically mediated the endo/lysosomal escape of dianthin without disrupting the plasma membrane. The purity of immunotoxins was confirmed by SDS-PAGE and Western blot. Their cytotoxicity was evaluated in the presence of SO1861 and dianthin-Cetuximab presented a GI50 (50% growth inhibition) of 5.3pM, dianthin-Panitumumab of 1.5pM, and dianthin-Trastuzumab of 23pM. Finally, the specificity of these immunotoxins was validated in a fluorescence-based real-time assay, where their binding to target cells was prevented by preincubation with an excess of label-free unconjugated antibody. Based on these data, we propose the use of dianthin and SO1861 as a new platform technology to enhance the efficacy of therapeutic antibodies.