Application of thermally responsive polypeptides directed against c-Myc transcriptional function for cancer therapy

Abstract

Elastin-like polypeptides are biopolymers composed of the pentapeptide repeat Val-Pro-Gly-Xaa-Gly. Elastin-like polypeptides are soluble in aqueous solution below their transition temperature, but they hydrophobically collapse and aggregate when the temperature is raised above the transition temperature. Previous studies have suggested that the aggregation of these polypeptides in response to externally applied hyperthermia may be exploited in the use of elastin-like polypeptide for thermally targeted drug delivery. This work shows the application of elastin-like polypeptide as a delivery vehicle for a short peptide that can inhibit the transcriptional function of a specific oncogene. The coding sequence for elastin-like polypeptide was modified by the addition of the membrane translocating sequence penetratin and a peptide derived from helix 1 of the helix-loop-helix region of c-Myc (H1-S6A,F8A), known to inhibit c-Myc transcriptional function. The designed polypeptide (Pen-ELP-H1) was then expressed and purified from Escherichia coli. Cellular uptake of Pen-ELP-H1 is enhanced by both the penetratin sequence and by the hyperthermia-induced phase transition as shown by flow cytometry studies. Using immunofluorescence and reverse transcription-PCR, we show that Pen-ELP-H1 is able to disrupt the nuclear localization of c-Myc and inhibit transcriptional activation by c-Myc. Cell proliferation studies showed that Pen-ELP-H1 inhibits growth of MCF-7 cells. Furthermore, the use of hyperthermia increased the antiproliferative effect of a thermally responsive Pen-ELP-H1 f2-fold compared with a nonthermally responsive control polypeptide. These studies show that genetically engineered elastin-like polypeptide carriers may provide a new way to thermally target specific oncogene inhibitors to solid tumors. [Mol Cancer Ther 2005;4(7):1076 -85]

Peptide-Based Polymer Therapeutics

“…Chilkoti et al have reviewed multiple alternatives for these systems [106,107]. ELPs have been successfully fused to a peptide (H1) inhibitor of an oncogene [108] together with penetratin (Pen) in order to allow membrane translocation demonstrating that Pen-ELP-H1 inhibits cell growth and more importantly, that hyperthermia significantly enhances treatment effectiveness. The well known chemotherapeutic agent Dox has been also conjugated to ELPs [109][110][111] alone or in combination with cell penetrating peptides [112], positioning the ELP carriers as promising candidates for thermally responsive polymer-drug conjugates in cancer treatment.…”

Peptide-based biopolymers in biomedicine and biotechnology

“…The CPP AntP showed the greatest in vitro increase in uptake in the unimer phase. An AntP-ELP fusion protein was also conjugated to the H1 peptide that is used to disrupt transcriptional activation of cells by inhibiting the c-Myc oncogene [141]. AntP-ELP-H1 caused a 2-fold decrease in cell proliferation when administered in hyperthermic versus normothermic conditions.…”

Development and characterization of a fusion protein between thermally responsive elastin‐like polypeptide and interleukin‐1 receptor antagonist: Sustained release of a local antiinflammatory therapeutic

“…Biologic activity was also retained by ELP fused to Penetratin, which underwent effective membrane translocation, as well as by ELP fused to H1, a c-myc oncogene inhibitor, which antagonized the transcriptional activation and retarded the growth of the breast carcinoma cell line MCF-7 (25). In addition, surfaces coated with an ELP fused to the RGD or fibronectin CS5 cell-binding sequence retained the ability to promote endothelial cell adhesion and spreading in vitro (26).…”