الفهرس 
Introduction
Back ground about neuroblastomaOnly 14 pages are availabe for public view
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Abstract
Neuroblastoma is one of the most common solid cancers among children. Prognosis of advanced neuroblastoma is still poor despite the recent advances in chemo/radiotherapies.
In view of improving the clinical outcome of advanced neuroblastoma, it is important to identify the key molecules responsible for the pathogenesis of neuroblastoma and to develop effective drugs that target these molecules.
Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase, initially identified through the analysis of a specific translocation associated with a rare subtype of non-Hodgkin’s lymphoma.
Recently it was demonstrated that ALK is frequently mutated in sporadic cases with advanced neuroblastoma. Recently, it was demonstrated that ALK is frequently mutated in sporadic cases with advanced neuroblastoma (Ogawa S, et al. 2010).
The purpose of our study was to examine ALK expression level in human NBL tissue samples and tumor cell lines.
Patients
343 human NBL tumor tissue specimens were collected retrospectively in a cohort study in Chiba Cancer Center Research Institute, Chiba University, Japan. Patients were diagnosed clinically as well as pathologically as primary neuroblastoma.
By analyzing these 343 human neuroblastoma tissue specimens for ALK mutation (Sequence), the results were that 16 (4.6%) cases had ALK gene mutation and 327 (95.4%) cases were without ALK mutation.
Then we randomly selected from the mutation free cases (327 cases) 79 cases for determination of ALK gene expression by (quantitative real-time PCR) to know whether there is a relationship between the expression level of ALK gene without any mutations and the prognosis of neuroblastoma patients.
Expression levels of ALK mRNA were examined by the quantitative real-time RT-PCR, analyzed by statistical studies, and our results indicated that, the expression level of ALK gene is significantly associated with the previously known prognostic factors and the findings in survival analysis indicated that all the previously reported prognostic factors carry a better outcome in neuroblastoma patients which agree with the previous publications.
Univariate analysis demonstrated that Shimadas’ histology, patient age, tumor stage, MYCN amplification status, TrkA 1 expression and DNA index were of prognostic importance in neuroblastoma patients, supporting the results of log-rank test.
On the other hand, the multivariate analysis (logistic regression test) showed that ALK expression was not significantly associated with survival after controlling Shimadas΄ histology, MYCN amplification, age, tumor stage ,TrkA expression, tumor origin and DNA ploidy , suggesting that ALK expression was not an independent prognostic factor from the other factors and its effect on the prognosis of neuroblastoma depend on the other previously reported factors(age, pathology, stage, TRKA expression level, MYCN amplification status, DNA ploidy and tumor origin).
The other part of the study which include analysis of 27 human neuroblastoma cell lines for ALK gene mutations and expression level, analysis of these 27 human neuroblastoma cell lines for ALK mutation showed that 9 (33.33%) cell lines had ALK mutation and 18 (66.66%) cell lines were without ALK mutation(wild type).
ALK expression was quantified in the same 27 human neuroblastoma tumor cell lines used for the mutation search by quantitative real-time RT-PCR and we found that 3 (11.1 %) cell lines had relatively high ALK expression these 3 cell lines were among the MYCN amplified cells and 2 of them had ALK gene mutations, this suggest that their might be a relationship between ALK expression, MYCN gene amplification and ALK gene mutations.
By lowering the level endogenous ALK by using siRNA against ALK gene in human neuroblastoma cell lines, we found that down regulation of ALK expression levels leads to decreased cellular proliferation of theses tumor cells by the MTT assay either in all cell lines.
ALK over expression in the 3 cell lines (PC12, SK-N-AS and NBL-S.) showed that ALK-transfected PC12 cells showed neurite outgrowth without NGF treatment This observation outlines that; there could be a relationship between ALK gene over expression and the neurite cell differentiation as established previously (Iwahara T, et al, 1997).
All our findings suggested that full-length ALK protein may not be enzymatically active (and hence may not be involved in oncogenic transformation) in the cell lines also this support the view that its presence may be a physiological rather than an oncogenic phenomenon.
Conclusion:
The present study was performed to examine the level of ALK mRNA gene expression in primary neuroblastoma patient tumor tissue samples and to assess the relation between ALK gene expression and other previously reported prognostic factors of neuroblastoma.
Collectively our present findings suggest that, high expression of wild type ALK is associated with good prognosis of NBL and that, as the mutation and amplification of ALK are reported previously to play an important role in NBL, also the expression level of wild type ALK gene might also have some function in cell growth as well as cell differentiation in neuroblastoma.
These observations open new opportunities for the development of novel therapeutic strategies based upon ALK targeting and the possibility to develop alternative therapeutic strategies based on ALK molecular targeting for advanced/metastatic NBLs is an attractive opportunity.
Recommendations
• Neuroblastoma patients above one year should be followed up carefully as they considered of bad prognosis.
• Family history and family planning should be done for any case of neuroblastoma to exclude familial neuroblastoma.
• Chromosomal study for neuroblastoma patients is recommended.
• Mutation study for ALK gene in neuroblastoma patients is recommended to detect patients with ALK gene mutations who considered of bad prognosis.
• The development of novel therapeutic strategies based upon ALK targeting and the possibility to develop alternative therapeutic strategies based on ALK molecular targeting.