Ru(III)-based complexes are generally considered attracting antitumor drugs for their remarkable anticancer and/or antimetastatic properties combined with general low toxicity in vivo, as well-demonstrated for two iconic Ru(III) compounds, known as KP1019 and NAMI-A. Intrigued by the excellent pharmacological activity of several Ru(III)-based drugs, some years ago, we started a study on a structurally-related NAMI-A compound, named AziRu, inserting it into nucleolipidic structures. These nucleolipidic-based Ru(III) complexes were then further co-formulated with biocompatible lipids, thus forming stable liposomes featured by a significant antiproliferative activity and selectivity for cancer cells both in vitro and in vivo. With the aim of enriching the arsenal of the available Ru(III) compounds, we recently designed a set of novel Ru(III) complexes characterized by lipophilic appendages decorating the structure of AziRu. Thus, the pyridine ligand of the ruthenium ion was conjugated with selected lipids (fatty acids or a cholesterol derivative) providing different lipid-functionalized Ru(III) complexes obtained with simple and high yielding synthetic schemes. Their molecular and self-assembling properties in proper buffer solutions mimicking the extracellular environment were investigated using several biophysical techniques. In vitro biological experiments, performed on a selected panel of cancer and healthy cell lines, proved that the lipophilic compounds carrying a palmitic and a stearic acid-based tail, named PalmiPyRu and StePyRu respectively, exhibited an interesting antiproliferative activity on specific breast cancer phenotypes without showing toxicity on normal cells, used as control. These lipophilic Ru(III) complexes were thus selected for more in-depth investigations aimed at evaluating their ability to interact with potential biomacromolecular targets, such as the bovine serum albumin (BSA), as a model serum protein, and a harpin duplex and two unimolecular G-quadruplex structures, as DNA model systems. These studies, performed by spectroscopic techniques in comparison with AziRu, evidenced the unique ability of PalmiPyRu to interact with both BSA and the selected G-quadruplex structures, one from the human telomere and the other from the c-myc oncogene promoter. In addition, PalmiPyRu exhibited a 20-fold enhanced cell uptake compared to AziRu in two breast cancer cell lines of different histological origin. Overall these results indicated PalmiPyRu as the best candidate in the designed series of lipid-conjugated Ru(III) complexes with marked improved cytotoxicity and cell uptake ability than AziRu. Ongoing experiments are devoted to better understand the in vitro distribution of PalmiPyRu after its administration in cell and identify the specific cell death pathway that this candidate drug is able to activate.