ORIGINAL ARTICLE Detection of cathepsin S cysteine protease in human brain tumour microdialysates in vivo T. FLANNERY 1,2,3,4 , R. S. MCCONNELL 1 , S. MCQUAID 2 , G. MCGREGOR 2 , M. MIRAKHUR 2 , L. MARTIN 3 , C. SCOTT 3 , R. BURDEN 3 , B. WALKER 3 , C. MCGOOHAN 3 & P. G. JOHNSTON 4 Departments of 1 Neurosurgery & 2 Pathology, Royal Victoria Hospital, Belfast, and 3 School of Pharmacy and 4 Centre for Cancer Research & Cell Biology, Queen’s University Belfast, Northern Ireland Abstract Microdialysis enables the chemistry of extracellular fluid in body tissues to be measured. Extracellular proteases such as the cysteine protease, cathepsin S (CatS), are thought to facilitate astrocytoma invasion. Microdialysates obtained from human brain tumours in vivo were subjected to cathepsin S activity and ELISA assays. Cathepsin S ELISA expression was detected in five out of 10 tumour microdialysates, while activity was detected in five out of 11 tumour microdialysates. Cathepsin S expression was also detected in microdialysate from the normal brain control although no activity was found in the same sample. While some refinements to the technique are necessary, the authors demonstrate the feasibility and safety of microdialysis in human astrocytomas in vivo. Characterisation of the extracellular environment of brain tumours in vivo using microdialysis may be a useful tool to identify the protease profile of brain tumours. Key words: Cathepsin S, microdialysis, astrocytomas. Introduction Microdialysis enables the chemistry of extracellular fluid in body tissues to be measured. 1 It has been applied in patients with head injury, 2 subarachnoid haemorrhage 3 and epilepsy. 4 The principle involves implanting a catheter lined with a dialysis membrane into the area of interest. 5 This catheter is perfused with physiological fluid at ultra-low flow rates by using a precision pump. Low molecular-weight substances diffuse across the membrane into the solution, which is then collected for analysis. Astrocytomas are primary brain tumours that diffusely infiltrate normal brain precluding complete surgical excision. Astrocytoma invasion is thought to be facilitated by the release of proteases, which can degrade the surrounding extracellular matrix (ECM). This process may be facilitated by microglia present in tumour stroma. 6 Proteases shown to be involved in astrocytoma invasion include metallo- proteases, serine proteases and cysteine proteases. Cathepsin S (CatS) is one of the lysosomal cysteine proteases. It is synthesised as an inactive precursor (36 kDa), which is activated in the acidic environ- ment of lysosomes by proteolytic cleavage of its propeptide. 7 Activated CatS is a 24 kDa single chain polypeptide capable of degrading a range of ECM macromolecules. 8 Unlike other cysteine proteases, such as cathepsins B and L, CatS is markedly more stable at extracellular pH indicating a possible role in extracellular proteolysis. 9 Immunohistochemical analysis of astrocytomas demonstrates that CatS is expressed in astrocytoma tumour cells and macrophages/microglial cells, but is absent from other normal glial and endothelial cells. 10 Cathepsin S expression and activity in tumour specimens is highest in WHO grade IV astrocytomas compared with astrocytoma grades I, II and III. 10,11 In vitro studies also demonstrate that CatS is expressed in astrocytoma cells with highest levels of secreted activity found in glioblastoma cultures. Inhibition of CatS activity by a synthetic compound (LHVS) reduces in vitro invasion in a glioblastoma cell line (U251MG) indicating a possi- ble role for CatS in astrocytoma invasion. 10 This potential role in invasion may contribute to the shorter survival associated with glioblastoma tu- mours expressing high levels of CatS. 11 The expres- sion and secretion of CatS in macrophages/microglial cells in vitro has also been demonstrated. 8 However, Correspondence: Mr Robert S. McConnell, Department of Neurosurgery, Royal Victoria Hospital, Grosvenor Road, Belfast BT12 6BA, Northern Ireland. Tel: 028 90240503. Fax: 028 90237733. E-mail: Roy.McConnell@royalhospitals.n-i.nhs.uk Received for publication 15 September 2006. Accepted 30 January 2007. British Journal of Neurosurgery, April 2007; 21(2): 204 – 209 ISSN 0268-8697 print/ISSN 1360-046X online ª The Neurosurgical Foundation DOI: 10.1080/02688690701248190 Br J Neurosurg Downloaded from informahealthcare.com by Queens University on 06/25/13 For personal use only.