Potential Therapeutic Effects of Exosomesloaded with Paclitaxel in Glioblastoma cell line

Background: Glioblastoma multiforme (GBM) is the deadliesttype of brain cancer with a poor prognosis and a survival rateof about ۸ to ۱۵ months after initial diagnosis. It is estimatedthat more than ۲۱۰,۰۰۰ individuals succumb to GBM annually.Currently, surgery (as the first-line treatment), radiotherapy,and temozolomide chemotherapy are the common therapeuticapproaches for GBM therapy. However, with regard to theheterogenous infiltrative nature of GBM, these treatments havenot been successful enough. Paclitaxel (PTX) is approximately۱۴۰۰-fold more potent than temozolomide. However, it is notused in the clinic due to its inadequate penetration across theblood–brain barrier. The main obstacle in the chemotherapy ofGBM is how to deliver therapeutic cargo into cancer cells. Exosomesare nanoscale extracellular vesicles ranging from ۳۰ to۱۲۰nm, and play pivotal roles in cell communications. Duringthe last decade, exosomes have shown promising achievementsto be applied as a potential delivery vehicle.Here, we purified the naïve exosomes from HEK-۲۹۳T cellsusing ۲۰% Polyethylene glycol (PEG). Then, an appropriateamount of PTX was loaded into the exosomes through the extracellularsonication method. Finally, the inhibitory effect ofthe engineered exosomes was evaluated on the tumorigenesisof U۸۷ cells.Materials and Methods: HEK۲۹۳T cells were cultured inexosome-depleted media under optimal growth condition for۷۲ hours (h). Then, the supernatant was harvested and the exosomeswere precipitated using ۲۰% PEG ۸۰۰۰ according tothe protocol. The purified exosomes were characterized basedon size, using Dynamic light scattering (DLS), and morphology,using field emission scanning electron microscopy (FE-SEM).The mixture was sonicated using an ultrasonic probe with ۲۰%amplitude for ۶ cycles. To recover the exosomal membrane aftersonication, the obtained solution was incubated at ۳۷ °C for۱ h. Finally, the amount of loaded PTX into the exosomes wasdetermined using High-performance liquid chromatography.The cytotoxic effects of the engineered exosomes (PTX-loadedexosomes; PTX-exo) on U۸۷ cells were evaluated using theMTT assay at ۲۴ and ۴۸ hours after treatment.Results: Exosomes collected from the supernatant of HEK-۲۹۳T cells were characterized by size (۱۱۲ nm in diameter)using DLS and morphology (spherical shape and size of ۳۰-۱۰۰ nm) using FE-SEM. By using HPLC it was estimated thatthe amount of PTX loaded into exosomes with sonication was۰.۹۱%. The percent of cell proliferation was significantly decreasedin U۸۷ cells that were treated with PTX-exo (۲۸.۲%),while it was ۹۷% for the cells which were treated with naïveexosomes. According to this, PTX-exo decreased cell proliferationby ۳.۴-fold.Conclusion: Exosomes, as natural nanoscale extravesicularparticles, can overcome the limitations of other drug-deliverysystems. However, some obstacles in exosomal formulationssuch as the choice of exosome extraction protocols and drugloading procedures should be addressed. Our findings elucidatedthat the exosomes purified from the HEK-۲۹۳T cell linecould be a promising vehicle to deliver PTX into GBM cells.Furthermore, our results confirmed the efficiency of PTX in inhibitingthe proliferation of U۸۷ cells. Further studies are neededto investigate the in vivo efficiency of PTX-exo

کلیدواژه ها:

Exosome ، paclitaxel ، glioblastoma ، drug loading