Aims: Advanced melanoma is characterized by poor outcome. Despite the number of treatments having been increased over the last decade, current pharmacological strategies are only partially effective. Therefore, the improvement of the current systemic therapy is worthy of investigation. Methods: a nanotechnology-based poly-chemotherapy was tested at preclinical level. Temozolomide, rapamycin, and bevacizumab were co-loaded as injectable nanoemulsions for total parenteral nutrition (Intralipid^®), due to suitable devices, and preliminarily tested in vitro on human and mouse cell models and in vivo on the B16-F10 melanoma mouse model. Results: Drug combination was efficiently loaded in the liquid lipid matrix of Intralipid^®, including bevacizumab monoclonal antibody, leading to a fast internalization in tumour cells. An increased cytotoxicity towards melanoma cells, as well as an improved inhibition of tumour relapse, migration, and angiogenesis were demonstrated in cell models for the Intralipid^®-loaded drug combinations. In preliminary in vivo studies, the proposed approach was able to reduce tumour growth significantly, compared to controls. A relevant efficacy towards tumour angiogenesis and mitotic index was determined and immune response was involved. Conclusions: In these preliminary studies, Intralipid^® proved to be a safe and versatile poly-chemotherapy delivery system for advanced melanoma treatment, by acting on multiple mechanisms.

Despite recent progressions in cancer genomic and immunotherapies, advanced melanoma still represents a life threat, pushing to optimise new targeted nanotechnology approaches for specific drug delivery to the tumour. To this aim, owing to their biocompatibility and favourable technological features, injectable lipid nanoemulsions were functionalised with proteins owing to two alternative approaches: transferrin was chemically grafted for active targeting, while cancer cell membrane fragments wrapping was used for homotypic targeting. In both cases, protein functionalisation was successfully achieved. Targeting efficiency was preliminarily evaluated using flow cytometry internalisation studies in two-dimensional cellular models, after fluorescence labelling of formulations with 6-coumarin. The uptake of cell-membrane-fragment-wrapped nanoemulsions was higher compared to uncoated nanoemulsions. Instead, the effect of transferrin grafting was less evident in serum-enriched medium, since such ligand probably undergoes competition with the endogenous protein. Moreover, a more pronounced internalisation was achieved when a pegylated heterodimer was employed for conjugation (p < 0.05).

Poor prognosis in high-grade gliomas is mainly due to fatal relapse after surgical resection in the absence of efficient chemotherapy, which is severely hampered by the blood–brain barrier. However, the leaky blood–brain–tumour barrier forms upon tumour growth and vascularization, allowing targeted nanocarrier-mediated drug delivery. The homotypic targeting ability of cell-membrane fragments obtained from cancer cells means that these fragments can be exploited to this aim. In this experimental work, injectable nanoemulsions, which have a long history of safe clinic usage, have been wrapped in glioma-cell membrane fragments via co-extrusion to give targeted, homogeneously sized, sterile formulations. These systems were then loaded with three different chemotherapeutics, in the form of hydrophobic ion pairs that can be released into the target site thanks to interactions with physiological components. The numerous assays performed in two-dimensional (2D) and three-dimensional (3D) cell models demonstrate that the proposed approach is a versatile drug-delivery platform with chemo-tactic properties towards glioma cells, with adhesive interactions between the target cell and the cell membrane fragments most likely being responsible for the effect. This approach’s promising translational perspectives towards personalized nanomedicine mean that further in vivo studies are foreseen for the future.