New technique for analysing aggressive brain tumour tissue

Immunocytochemical image of
cultured tumour cells: Katrina Kan

by Anne Crawford

A Department of Neuroscience PhD student has helped develop a new, cost-effective and reliable method for studying glioblastoma, a devastating brain cancer.

Ms Katrina Kan from Dr Mastura Monif’s laboratory, was first author on a paper about a new protocol for using primary human glioblastoma tissue, published last month in Methods and Protocols.

Glioblastoma is the most aggressive and most common brain tumour, accounting for more than 60% of all brain cancer in adults. Current treatments – mainly chemotherapy, radiotherapy and surgery – are of limited efficacy; patients typically survive for only 14 to 15 months after diagnosis. Research providing deeper understanding of glioblastoma pathology and the mechanisms of tumour pathogenesis is vital.

Ms Kan said studies into glioblastoma traditionally used cell lines and animal models. Cell lines, which use culture grown from a single glioblastoma cell isolated from a patient a number of years ago, had several disadvantages.

“They often contain only one type of cell from a homogenous cell population – that doesn’t represent what actually happens in a physiological sense,” Ms Kan said. “They don’t reflect the heterogeneity of tumours in vivo.”

The new methodology developed in the Monif laboratory surgically resects, prepares and cultures fresh human glioblastoma tissue.

Primary glioblastoma resembled most closely the microenvironment of the original tumour mass and the range of other cells present including infiltrating immune cells such as lymphocytes, neutrophils and microglia, Ms Kan said. “So it’s able to reflect what the immune system does on the tumour and therefore more closely resemble true disease, compared to cell lines.”

According to the method, a sample of the tumour is resected at the time of surgery and immediately sent to the laboratory. It is immersed in an enzyme solution that breaks down the various tissues and leaves the glioblastoma cells, making it easier to create a single cell suspension, Ms Kan explained. Supplementary material is added and the cells sealed in plates then left in an incubator for about seven days at which time it was about 80% confluent (the percentage of the culture dish covered by cells).

“From then we can add different treatments to assess their effect on the tumour,” Ms Kan said.

“It’s a reliable and cost-effective way to culture glioblastoma cells which are usually quite hard to culture,” she said. “It also represents the tumour microenvironment better than cell lines and because we’re creating cultures from different patients, it accounts for patient heterogeneity and allows us to see if potential drugs work on different patients.”

Moreover, it is already proving its effectiveness. The method, previously trialled in the Monif lab, was used to discover a compound that could reduce tumour growth by almost 40% – a finding in a paper that has recently been submitted for publication, Ms Kan said.

Ms Kan, and colleagues, made that discovery in her Honours year in 2018. “I’m hoping to take this further in my PhD studies,” she said.

Originally studying immunology and oncology, Ms Kan embarked on research solely because she wanted to get into medicine before being inspired by Dr Monif to pursue both. “I didn’t really want to do research and never thought I’d be doing neuroscience until I met Mastura, who was really approachable.

“The translational side of this project really intrigued me. That’s why I picked it and since then I’ve continued with the same supervisors,” she said.

Ms Kan said Dr Monif, her main supervisor, and co-supervisors Professor Terence O’Brien, Professor Kate Drummond AM (Director of Neurosurgery, Royal Melbourne Hospital) and Professor David Williams Department of Physiology, University of Melbourne), as well as researcher Associate Professor Martin Hunn (Director of Neurosurgery, The Alfred) all played an important part in the study.

She now wants to specialise in neurology and continue researching. “Hopefully I’ll be studying brain tumours and can translate my research to apply to my patients,” she said.

Kan LK, Drummond KJ, Hunn M, Williams DA, O'Brien TJ, Monif M. A simple and reliable protocol for the preparation and culturing of fresh surgically resected human glioblastoma tissue. Methods Protoc. 2020 Jan 22;3(1). pii: E11. doi: 10.3390/mps3010011.