Introduction: Gliobastoma is one of the most aggressive forms of brain cancer. Each year 18,000 people are diagnosed with glioblastomas with an average survival of 10 to 12 months. MSC are an attractive candidate for cell based therapy since they can be easily isolated, manipulated and have no immune rejection. MSC transduced with TRAIL are capable of migrating towards the tumor cells and kill the tumor cells due to bystander effect[1]. However for cell based therapy migration, survival and retention are the major factors. MSC in spheoridal form has shown to have increased expression of CXCR4 which is critical for migration towards tumor cells[2]. The spheroidal form also might help the cells to survive for longer and prevent it from getting washed out due to the extracellular matrix surrounding the cells and larger size respectively. Hence MSC in spheroidal created using a microfluidic platform was used for cell based therapy to treat glioma.
Materials and Methods: Microfluidic device to generate double emulsion was prepared using soft lithography techniques. Silicon mold was patterned using SU-8 3025 to be of 200 um in height. To prepare the cell spheroids 10 million cels/ml was used as the inner phase. The oil phase used was HFE-7500 supplemented with Pico-Surf 1 and Pluronic F-127 was used as the outer phase. IBIDI cell culture insert was used to compare MSC spheroids and MSC migration towards U87 on either side of a 500 um gap. MSC was then transduced with sTRAIL followed by the formation of the spheroids. The condition media collected from the MSC-sTRAIL was used to evaluate the caspase 3/7 activation and toxicity on three different glioblastoma cells.
Results and Discussion: The MSC self assembled to form spheroids within the double emulsion in 150 mins. Live/dead staining demonstrated high viability of the cells in the spheroid (Fig 1).The size of the spheroid was controlled using the cell density of the inner fluid.
Semi-quantitative RT-PCR demonstrated enhanced expression of CXCR4 in the MSC spheroids. This translated to higher migration of the MSC spheroids compared to MSC as seen by increased directionality, velocity, euclidian distance and displacement of the center of mass.
Condition media collected from MSC-sTRAIL single and spheroids were collected and showed higher caspase 3/7 activation. The condition media also effectively killed the cancer cells. Co-Culture of MSC-sTRAIL Spheroids and U87 led to high killing of the cancer cells.
Conclusions: MSC spheroids were prepared using a microfluidic device in as little as 2.5 hrs. These MSC spheroids showed higher expression of CXCR4, enhanced migration toward tumor cells, and effectively killed the cancer cells by activating caspase 3/7 pathway. Hence MSCs in spheroidal form are an excellent candidate for treating glioblastomas.
Schlumberger Faculty for the future fellowship
References:
[1] Sasportas, L. S. et al. Assessment of therapeutic efficacy and fate of engineered human mesenchymal stem cells for cancer therapy. Proc. Natl. Acad. Sci. U.S.A. 106, 4822–4827 (2009).
[2] Bartosh, T. J. et al. Aggregation of human mesenchymal stromal cells (MSCs) into 3D spheroids enhances their antiinflammatory properties. Proc. Natl. Acad. Sci. U.S.A. 107, 13724–13729 (2010).