gms | German Medical Science

Objective: Organotypic slice cultures have been used instead of dissociated cell cultures to create an environment where the cytoarchitecture of the original tissue is preserved. We established an assay for drug efficacy testing with explants of surgical specimens of gliomas and meningeomas. Intratumoral drug delivery of the mTor inhibitors Everolimus and Temsirolimus is facilitated by immersed silica gel. The influence of both mTOR inhibitors on composition and proliferation of the cultivated cell complex is studied after immunohistochemical staining.

Method: Surgical specimens were received immediately after extirpation. Serial 400µm slices were made and transferred to sterile, porous (8µm) membranes (Cell Culture Inserts, BD Falcon) in 12well culture plates and incubated overnight in 1ml DMEM supplemented with 10% FCS in an incubator at 37°C in a humidified atmosphere. Silica gel beads were implanted into the tumor slice after incubation with the dissolved drugs. After 24h of incubation, the slices were fixed in 4% PFA, dehydrated, frozen in liquid nitrogen and then 10µm frozen sections were made. The treated samples were stained for CD31, GFAP, EMA, Iba-1, Ki-67, and cleaved Caspase 3, as well as PAS and HE, in order to analyze for the presence of blood vessels, glial cells, proliferative activity, apoptosis and necrosis.

Results: The cultivated tumor explant slices showed an increase in diameter during a 48h cultivation period. We detected Ki-67 expression, but no apoptosis or necrosis. After mTOR-inhibitor treatment, we demonstrated an immigration of microglia into the (mTOR inhibitor) treated region for all analyzed tumor entities. A decrease in the number of proliferative cells, and an increase of apoptotic cells and necrotic areas was observed.

Conclusions: We were able to cultivate tumor explant slices over a period of 48h. By incubation of tumor slices with mTOR-inhibitors, we achieved a repression of tumor cell growth and a formation of necrosis. Cultivation of tumor explant slices is an organotypic surrounding makes it possible to determine the efficacy of any chemotherapeutic agent on heterogenous tumor cells with a reasonable degree of cellular differentiation and organization resembling that of the native tissue.