Organotypic brain explant culture as a drug evaluation system for malignant brain tumors.

Noriaki Minami,Takashi Sasayama,Koji Maruyama,Shunsuke Shibao,Oltea Sampetrean,Yoshimi Arima,Fumiharu Ohka,Hideyuki Saya,Yutaka Kondo,Eiji Kohmura,Masatoshi Kusuhara,Yusuke Maeda

doi:10.1002/cam4.1174

Abstract

Therapeutic options for malignant brain tumors are limited, with new drugs being continuously evaluated. Organotypic brain slice culture has been adopted for neuroscience studies as a system that preserves brain architecture, cellular function, and the vascular network. However, the suitability of brain explants for anticancer drug evaluation has been unclear. We here adopted a mouse model of malignant glioma based on expression of H‐RasV12 in Ink4a/Arf −/− neural stem/progenitor cells to establish tumor‐bearing brain explants from adult mice. We treated the slices with cisplatin, temozolomide, paclitaxel, or tranilast and investigated the minimal assays required to assess drug effects. Serial fluorescence‐based tumor imaging was sufficient for evaluation of cisplatin, a drug with a pronounced cytotoxic action, whereas immunostaining of cleaved caspase 3 (a marker of apoptosis) and of Ki67 (a marker of cell proliferation) was necessary for the assessment of temozolomide action and immunostaining for phosphorylated histone H3 (a marker of mitosis) allowed visualization of paclitaxel‐specific effects. Staining for cleaved caspase 3 was also informative in the assessment of drug toxicity for normal brain tissue. Incubation of explants with fluorescently labeled antibodies to CD31 allowed real‐time imaging of the microvascular network and complemented time‐lapse imaging of tumor cell invasion into surrounding tissue. Our results suggest that a combination of fluorescence imaging and immunohistological staining allows a unified assessment of the effects of various classes of drug on the survival, proliferation, and invasion of glioma cells, and that organotypic brain slice culture is therefore a useful tool for evaluation of antiglioma drugs.

Highlights

Knowledge of the genetic and molecular underpinnings of malignant glioma has increased greatly in recent years [1,2,3]
To take full advantage of the microenvironment provided by brain explants, we chose a syngeneic murine glioma model based on transduction of Ink4a/Arf–null neural stem/progenitor cells with the oncogene H-RasV12
We have here asked whether they are suitable for a systematic evaluation of the antitumor effects of various types of drug on murine glioma cells

Summary

Introduction

Knowledge of the genetic and molecular underpinnings of malignant glioma has increased greatly in recent years [1,2,3]. The development of new and effective drugs for this condition has not kept up with such advances [4]. A lack of assays that balance feasibility, sufficient throughput, and reconstruction of the architecture and environment of the normal brain is one technical reason for the slow pace of drug discovery. Cell-based assays [5] remain the gold standard for high-. Throughput screening, whereas animal models are important for validation of promising lead compounds and evaluation of safety and dosing schedules.

Methods

Results

Conclusion