Abstract:
:Extraocular dissemination is the main cause of death in patients with retinoblastoma (RB) in developing countries, and there are few molecular markers that are useful for the evaluation of minimal disseminated disease. The GD2 ganglioside is known to be expressed by RB cells that metastasize in bone marrow, and the activity of the enzyme responsible for its synthesis, GD2 synthase, can be detected in neuroblastoma, which shares many phenotypic features with RB. The purpose of the present study was to optimize the detection of GD2 synthase expression by reverse transcription-polymerase chain reaction (RT-PCR) followed by nested-PCR in human RB cell lines and patient samples. The optimization strategy was carried out using the RB cell lines Y79 and WERI-Rb1 and specific primers designed for the human sequence of GD2 synthase mRNA. We detected GD2 synthase expression with at least 200 and 40 pg of total RNA extracted from cultured RB cells using a first round of RT-PCR amplification or a second round of nested-PCR, respectively. We also confirmed the expression of GD2 synthase by RT-PCR and immunohistochemical detection of the ganglioside in human RB tumors xenotransplanted in nude mice. Using tumor bank specimens from eight RB patients, we were able to demonstrate the presence of GD2 synthase mRNA in blood and cerebrospinal fluid samples in cases of extraocular dissemination of the tumor. The sequence was not detected in samples derived from children with low-risk disease or healthy adult volunteers. Hence, GD2 synthase mRNA detection through an optimized nested RT-PCR assay is a promising tool for the assessment of minimal disseminated disease in enucleated patients.
journal_name
Mol Med Repjournal_title
Molecular medicine reportsauthors
Laurent VE,Otero LL,Vazquez V,Camarero S,Gabri MR,Labraga M,De Dávila MT,Chantada GL,Alonso DFdoi
10.3892/mmr_00000248subject
Has Abstractpub_date
2010-03-01 00:00:00pages
253-9issue
2eissn
1791-2997issn
1791-3004journal_volume
3pub_type
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