In 1999, D. discoideum was chosen by the NIH (USA), as a non-mammalian model for biomedical researches (http://www.nih.gov/science/models/).

As related in our first paper [8], D. discoideum cells are highly resistant to the DNA vital stain Hoechst 33342 (HO342). These cells are also resistant to many structurally unrelated xenobiotics, such as the carcinogenic hydrocarbon benzo(a)pyrene, the antitumoral antibiotic daunorubicin, the mitochondrial vital stain Rhodamine 123 and a carbocyanine dye, JC-1. All these xenobiotics are known substrates of the P-gp, a 170-kDa protein, mediating classical multidrug resistance.

We first demonstrated, by Western blot analysis with the anti-P-gp monoclonal antibody (mAb) JSB-1, the presence of a constitutive P-gp in D. discoideum cells [10]. This was confirmed by flow cytometry analysis using two other anti-P-gp mAbs, C219 and MRK-16. However, despite the presence of P-gp, the classical P-gp-mediated multidrug resistance appeared non-functional in D. discoideum cells.

Searching for a new resistance mechanism to explain the high endogeneous resistance of D. discoideum cells, we observed that, when assays of vitally staining the cells with the DNA-targeted dye, HO342 were performed, a fluorescent material was released in the culture medium. The spectral characteristics of the fluorescence allowed us to identify HO342 as the emitting compound [8,9]. After concentration, the vesicular nature of this fluorescent extracellular material was revealed by electron microscopy and phospholipid identification. Nucleic acids were also found associated with these vesicles [8]. The new extracellular organelles were further characterized by cryo-electron microscopy and protein analysis [11]. Thus was evidenced in D. discoideum cells, an unknown multidrug resistance mechanism [8], sustained by a detoxification process, by means of which the cells get rid of different structurally unrelated xenobiotics. Moreover, these Dictyostelium extracellular vesicles were able to transfer their drug content to human cell lines, such as the erythroleukemic K562 cells [9] and tumoral HeLa cells [11]. This “Trojan-horse” capacity of Dictyostelium extracellular vesicles was accredited for therapeutic non-viral drug delivery¹, whereas their immunogenicity was checked by a first in vivo study, showing a specific antibody response, but no pyrogenic response nor any inflammation, as measured by five pertinent cytokines (study performed by Genosafe, Evry, France) [11].

Although being a very primitive eukaryotic ancestor of mammalian cells [12], Dictyostelium was recently recognized as a model for human disease [13]. This model turned valuable for the fundamental study of many unrelated human pathologies, such as pathogen-host interactions [14,15], pathobiology of cell motility [16], analyses of lissencephaly-related proteins [17], mechanisms involved in sensitivity to cisplatin and other anticancer drugs [18], signaling in bipolar disorder [19], lysosomal and trafficking diseases [20] and mitochondrial disease [21].

Taking into account both the above consideration and the amazing continuity in many biological processes from primitive to higher organisms, we suggest a new hypothesis about the mechanism of multidrug resistance in mammalian cells. The newly observed vesicle-mediated detoxification mechanism in D. discoideum cells [8,9,11,22] might also be involved in mammalian cells, and explain, at least partially, the failure of antitumoral chemotherapy.

Already in 2003, Shedden et al. mentioned that, when treated with doxorubicin or other antitumoral compounds, many different tumor cells release vesicles, inholding the antitumoral compound [23]. Some recent clinical observations fit with our suggestion and underline the importance of membrane vesicles in tumorigenesis [24]. Very recently, Goler-Baron and Assaraf observed extracellular vesicles mediating multidrug resistance between two attached MCF-7/MR cells [25]. Fig. 1 depicts schematically the suggested new “dynamic” multidrug resistance (MDR), mediated via cell-derived vesicles, originating from the endocytic intracellular traffic. The classical multidrug resistance (MDR), mediated via known transporters, like P-glycoprotein, multidrug resistance proteins (MRP) (and others), corresponding to a “static” mechanism preventing many therapeutic drugs to enter the cells, is shown for comparison.

Two lines of experiments are proposed to check the hypothesis that cell-derived vesicles are also drug transporters in human cells.

First, our observation that Dictyostelium cells are resistant to the vital staining of their nuclei by HO342 [8,9] could be linked with the fact that the detection by flow cytometry of the few hematopoietic stem cells in a cell population has also long been based on their high resistance to the vital staining of their nuclei by HO342 [26,27]. Therefore, it would be worth searching whether a stem cell population, grown in the presence of HO342 should depict the same vesicle-mediated detoxification mechanism, as the one, which we evidenced in D. discoideum cells. Very recently, tumors where suggested to harbour stem cells [4] and it would, therefore, be interesting to evidence such a multidrug resistance mechanism in stem cells.

On the other hand, with regard to non-solid tumors, the human erythroleukemia K562s cell line, normally sensitive to chemotherapy, can become a resistant K562r cell line by an appropriate long term drug treatment [28]. Up to now, this resistance was thought to be mainly mediated by the P-glycoprotein. K562 cells are also known to constitutively expel “exosome-like” vesicles in their extracellular medium [29]. We wonder whether these vesicles could also be involved in a multidrug resistance mechanism, like Dictyostelium vesicles, which turned out to be both constitutive without drug, and involved in drug detoxification [8]. These two cell lines, K562s and K562r, would, indeed, be a good choice to check the general occurrence of a multidrug “vesicle-mediated” resistance mechanism. Vesicles could be concentrated from both extracellular growth media, following incubation of the two cell lines with a fluorescent P-gp substrate, like HO342, and P-gp expression should be compared in the vesicles originating from K562s and K562r cells, respectively. If P-gp might be transported by vesicles, originating from K562r cells, it might, possibly, also be transferred to K562s cells by means of these vesicles. This could sign a way of propagating multidrug resistance both in vitro and in vivo, as already observed in the vesicle-mediated propagation of a few tumors [24].

D. discoideum has already shown its interest as a model for human disease [13–21]. It could as well be thought as a model for unraveling the mechanisms of multidrug resistance, still severely impairing the antitumoral chemotherapeutic treatments.

Based on our previous results, which evidenced a new vesicle-mediated multidrug resistance in D. discoideum cells [8,9,11], the observed detoxification mechanism is suggested to be a quite general eukaryotic process, also implied in antitumoral multidrug resistance. Two lines of experiments with human cells are proposed to check this hypothesis.

Moreover, the newly recognised biological importance of cell-derived vesicles in intercellular communication² [30–32], as well as their importance in tumorigenesis [24,33] could also be advantageously further questioned by using Dictyostelium as a model.

The author declare that he has no conflicts of interest concerning this article.