The purpose of the proposed study is to develop a method based on fast atom bombardment mass spectrometry and tandem mass spectrometry (FABMS/MS) for measuring cell membrane damage in the central nervous system (CNS). This damage is the result of whole brain radiation (WB RTX) and intrathecal methotrexate (IT MTX) used in the treatment of children with acute lymphoblastic leukemia (ALL) and medulloblastoma. The proposed FABMS/MS method will be used in combination with normal phase high performance liquid chromatography (HPLC).
The specific aims of the proposed study are: 1. The identification of the phospholipid classes and species in cerebrospinal fluid (CSF) collected from children with ALL and medulloblastoma before and during CNS treatment. Samples will be fractionated using normal phase HPLC and analyzed off line using FABMS and FABMS/MS techniques. 2. Identification of phospholipid markers of CNS membrane damage following WB RTX and IT MTX treatment by FABMS/MS in combination with HPLC. 3. Quantitation of phospholipid markers of CNS membrane damage by fast atom bombardment mass spectrometry (FABMS) and HPLC. 4. Explore the possibility of using FABMS/MS alone to identify and quantify the cell membrane damage by analysis of total CSF lipid extracts. A repeated measures comparative three group design will be used. Subjects will be grouped by type of CNS treatment: (Group I:IT MTX only; Group II:IT MTX, ARA-C and hydrocortisone; Group III: WB RTX). Total CSF lipid extracts from children before, during and after WB RTX and IT MTX will be analyzed to completely characterize the phospholipid classes and molecular species present, and to identify. markers of CNS cell membrane damage. Quantitation of the markers will be done using calibration curves (HPLC) and internal standards (FABMS). Changes over time will be calculated by repeated measures analysis of variance. Direct analysis of total CSF lipid extracts will be done using (+) and FABMS/MS techniques. The results of this research will provide important information on the mechanism of membrane damage during WB RTX and IT MTX. Identification of phospholipid markers of membrane damage will provide a method of fundamental clinical importance for monitoring the CNS toxicity of therapy in diverse patient populations to minimize the long-term adverse effects.
