Proteomic Identification of Glutamine Synthetase as a Differential Marker for Oligodendrogliomas and Astrocytomas

A review of:

Proteomic Identification of Glutamine Synthetase as a Differential Marker for Oligodendrogliomas and Astrocytomas

Zhengping Zhuang et al. (2011)

Mohamed A. Mohamed

Genomic Proteomic Engineering

Dr. Tianfu Wu

Spring 2014

Part one: Paper Review

To provide an appropriate prognosis and treatment of brain cancers, it is not sufficient to only know the size and morphology of tumors. Treating the tumor in the hopes of preventing its resurgence requires awareness of its cellular composition. What type of cell a tumor is primarily made of plays heavily on the severity of the cancer and the appropriate treatment. It is, thus, important to develop markers to determine the cellular makeup of various tumors.

Zhunag and his colleagues set out to do just that [1]. They have identified glutamine synthetase (GS), an enzyme involved in the metabolism of nitrogen, to differentiate between two types of brain cancers, oligodendrogliomas (ODs) and astrocytomas (ACs). GS catalyzes the condensation of glutamate and ammonia to form glutamine. The reaction is ATP driven and results in the attachment of an amine group, from ammonia, which contains the nitrogen to metabolize, to glutamate to form glutamine. This is an essential reaction predominantly occurring in the brain, kidney, and liver. In the brain, this translates into detoxification of ammonia, as well as recycling and termination of glutamate, a neurotransmitter [2]. GS was initially considered to be exclusively produced by astrocytes, but starting in the late 1980s it has been shown to also be produced by oligodendrocytes [3].

According to the World Health Organization (WHO), grade two and three ODs are more sensitive to chemotherapy than ACs of the same grade. As a result, patients stricken with ODs live twice as long after the onset of the cancer. Currently established markers used to differentiate between the two cancers, such as glial fibrillary acidic protein (GFAP), an intermediate filament found in astrocytes but not ODs, are inadequate. Zhuang claims that since the two cancers consist of cells which derive from different progenitor cells, a more unique protein marker may be found for more specific immunohistopathological cancer typing. To pursue their goal, they used 2D-PAGE, mass spectrometry, and bioinformatics to identify proteins which are uniquely expressed between astrocytes and oligodendrocytes.

Zhuang obtained tumor samples of grade two, three, and four ACs, as well as grade two and three ODs to use for protein expression profiling. To validate the new markers, they obtained samples of oligoastrocytomas (OACs), which contain both cell types, as well as normal mouse cortex for a negative control. After selectively choosing microdissections of clearly tumorous tissue, they dissolved the specimens in an extraction buffer, then mixed and centrifuged to collect the supernatant. The supernatant was then combined with a rehydration buffer overnight in an immobilized pH gradient (IPG). For the first dimension of separation, isoelectric focusing was performed by subjecting the IPG strips to high voltages. For the second dimension, a 4-7 pH gradient gel was used under a constant voltage. Protein spots were detected with a silver staining kit.