The effective delivery of nucleic acid therapeutics into tumor cells is of great importance in terms of cancer gene therapy. Successful application of siRNAs is restricted by the requirement to overcome several biological barriers, i.e. cell membrane penetration, endosomal escape, enzymatic degradation in cytosol, etc. Cationic polymers are promising delivery systems due to their biodegradability and effective binding with siRNA. A key characteristic of the particle trafficking into the cell is a noticeable change in pH during this process. This change can be used as a natural stimulus for pH-sensitive polymer particles. Such polymers are able to buffer the environment of a late endosome that leads to an abnormal flow of ions and water followed by endosomal swelling and disrupt and quick escape of the carriers into a cytoplasm. In present work, we synthesized a series of amphiphilic positively charged biocompatible pH-sensitive polypeptides forming polyplexes with nucleic acids. Four amino acids were used to compose the delivery systems, namely l-lysine, l-glutamic acid, l-phenylalanine and l-histidine. The number and the ratio of amino acids were varied in order to establish the optimal composition. The characteristics of polypeptides were studied by ¹H NMR spectroscopy, high performance liquid chromatography and static light scattering. All synthesized polypeptides tended to a self-assembly in aqueous media and formed nanoparticles with hydrodynamic diameters ranging from 140 to 360 nm. The formation of polyplexes with short double-stranded oligonucleotides as a physicochemical siRNA model as well as the release of cargo in buffer media mimicking lysosome (pH∼5) and cytosol or bloodstream (pH 7.4) were studied. Cellular uptake of the particles was efficient and their cytotoxicity was negligible. Successful RNAi-mediated down-regulation of GFP gene expression was revealed in MDA-MB-231/GFP breast cancer cells. The amphiphilic polypeptides obtained can be considered as promising non-viral candidates for siRNA delivery.