Extracellular vesicles of Janthinobacterium lividum as violacein carriers in melanoma cell treatment

Violacein is a natural indole-derived purple pigment of microbial origin that has attracted attention for its remarkable biological properties. Due to its poor solubility in aqueous media, most studies of this pigment use extracts of the compound obtained with common solvents. Violacein is also transported in bacterial extracellular vesicles (EVs) and transferred via this type of carrier remains stable in an aqueous environment. This paper is the first to present an in-depth study of Janthinobacterium lividum EVs as violacein carriers. J. lividum EVs were studied for their contribution to violacein translocation, size, morphology and protein composition. The production of violacein encapsulated in EVs was more efficient than the intracellular production of this compound. The average size of the violacein-containing EVs was 124.07 ± 3.74 nm. Liquid chromatography-tandem mass spectrometry analysis (LC–MS/MS) revealed 932 proteins common to three independent EVs isolations. The high proportion of proteins with intracellular localisation, which are involved in many fundamental cellular processes, suggests that J. lividum EVs could be generated in a cell lysis model, additionally stimulated by violacein production. Using human keratinocytes and melanoma cell lines, it was confirmed that J. lividum EVs are able to react with and deliver their cargo to mammalian cells. The EVs-delivered violacein was shown to retain its activity against melanoma cells, and the dose and timing of treatment can be selected to target only cancer cells. The characterisation of J. lividum EVs, described in the following paper, represents a milestone for their future potential anticancer application.

• This report focuses on the investigation of Janthinobacterium lividum EVs as a new delivery vehicle for violacein, a compound with a previously demonstrated broad spectrum of activity.

• EVs were characterised for size, morphology and protein composition.

• Studies on human keratinocytes and a melanoma cell model confirmed that the activity of violacein applied in the encapsulated form of EVs is similar to that of its organic solvent extract, but their production is much more environmentally friendly.