Cationic poly(amidoamine) promotes cytosolic delivery of bovine RNase A in melanoma cells, while maintaining its cellular toxicity

Abstract

Ribonucleases are known to cleave ribonucleic acids, inducing cell death. RNase A, a member of the ribonuclease family, generally displayed poor in vitro activity. This has been attributed to factors such as low intracellular delivery. Poly(amidoamine)s have been used to promote the translocation of non-permeant proteins to the cytosol. Our objective was to demonstrate that poly(amidoamine)s could potentially promote the delivery of RNase A to selected cell line. Interactions of three cationic poly(amidoamine)s (P1, P2 and ISA1) with wild-type bovine RNase A were investigated using gel retardation assays, DLS and microcalorimetry. Although the polymers and the protein are essentially cationic at physiological pH, complexation between the PAAs and RNase A was observed. The high sensitivity differential scanning calorimetry (HSDSC) thermograms demonstrated that the thermal stability of the protein was reduced when complexed with ISA1 (T_max decreased by 6.5 °C) but was not affected by P1 and P2. All the polymers displayed low cytotoxicity towards non-cancerous cells (IC₅₀ > 3.5 mg mL⁻¹). While RNase A alone was not toxic to mouse melanoma cells (B16F1), P1 was able to promote cytosolic delivery of biologically active RNase A, increasing cell death (IC₅₀ = 0.09 mg mL⁻¹).