The Transmission and Toxicity of Phpma Copolymer-Bound Doxorubicin Containing Exosomes Derived from Adherent Two-Dimensional Human Breast Adenocarcinoma Cell Line and Three-Dimensional Spheroids

Exosomes are endosomally-derived vesicles. Their composition is significantly affected by physiological state of the donor cells and can reflect changes in the cell microenvironment as well as of the whole body. Exosomes can manipulate local and systemic environment to influence cancer progression and dissemination and also modulate immune response. Moreover, exosomes have been investigated as possible transmission nanocarriers for therapeutics. In this study, exosome loading was achieved by incubation of free doxorubicin (DOX) and DOX bound to N -(2-hydroxypropyl) methacrylamide (HPMA)-based copolymers (pHPMA) with an adherent human breast adenocarcinoma cell line and its derived spheroids. Both free and p(HPMA)-bound DOX were successfully loaded into exosomes with different loading efficiencies. Spheroids secreted significantly more exosomes than adherent cells. These exosomes differed in abundance of their exosomal markers. The adherent cell line showed a decreased viability after treatment with free DOX or pHPMA bound-DOX-loaded exosomes, confirming the successful transmission of both, free cancerostatics and polymer bound cancerostatics by exosomes. To our knowledge, this is the first proof of pHPMA-drug conjugates secretion by extracellular vesicles, providing a new perspective on the transmission of drug/polymeric drugs in tumor tissue via exosomes. Interestingly, results obtained within the manuscript contribute to the explanation of high therapeutic activity of pH-sensitive polymer-DOX conjugates, which we suppose, is based on the exosome-based cell-to-cell delivery within the tumor tissue after the passive accumulation of the polymer-drug system.