Prognostic Impact of Gene Expression–Based Classification for Neuroblastoma

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Purpose: Genomic alterations of the anaplastic lymphoma kinase (ALK) gene have been postulated to contribute to neuroblastoma pathogenesis. This study aimed to determine the interrelation of ALK mutations, ALK expression levels, and clinical phenotype in primary neuroblastoma.Experimental Design: The genomic ALK status and global gene expression patterns were examined in 263 primary neuroblastomas. Allele-specific ALK expression was determined by cDNA cloning and sequencing. Associations of genomic ALK alterations and ALK expression levels with clinical phenotypes and transcriptomic profiles were compared.Results: Nonsynonymous point mutations of ALK were detected in 21 of 263 neuroblastomas (8%). Tumors with ALK mutations exhibited about 2-fold elevated median ALK mRNA levels in comparison with tumors with wild-type (WT) ALK. Unexpectedly, the WT allele was preferentially expressed in 12 of 21 mutated tumors. Whereas survival of patients with ALK mutated tumors was significantly worse as compared with the entire cohort of WT ALK patients, it was similarly poor in patients with WT ALK tumors in which ALK expression was as high as in ALK mutated neuroblastomas. Global gene expression patterns of tumors with ALK mutations or with high-level WT ALK expression were highly similar, and suggested that ALK may be involved in cellular proliferation in primary neuroblastoma.Conclusions: Primary neuroblastomas with mutated ALK exhibit high ALK expression levels and strongly resemble neuroblastomas with elevated WT ALK expression levels in both their clinical and molecular phenotypes. These data suggest that high levels of mutated and WT ALK mediate similar molecular functions that may contribute to a malignant phenotype in primary neuroblastoma. Clin Cancer Res; 17(15); 5082-92. Ó2011 AACR.

Section: Discussionmentioning

confidence: 99%

“…To examine the influence of genomic ALK alterations on ALK transcript levels, relative ALK mRNA expression levels were determined in all 263 tumors by using microarrays (21,26), and validated in 81 samples by qRT-PCR (Supplementary Fig. S2).…”

Section: Genomic Alk Alterations Are Associated With Elevated Alk Expmentioning

confidence: 99%

High ALK Receptor Tyrosine Kinase Expression Supersedes ALK Mutation as a Determining Factor of an Unfavorable Phenotype in Primary Neuroblastoma

Schulte

Bachmann

Brockmeyer

et al. 2011

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“…In our study, we corroborate this result and furthermore show by analysis of a large neuroblastoma cohort representing the whole spectrum of the disease that PLK1 expression is an independent marker of poor outcome in this malignancy. However, in light of the high prognostic accuracy of current risk-stratification strategies and complex DNA-or RNA-based prognostic classifiers (21,22,(37)(38)(39), it remains questionable, whether PLK1 expression will be utilized for risk estimation of neuroblastoma patients in the future. Western blot analysis revealed that PLK1 transcript levels correlate well with PLK1 protein levels ( Fig.…”

Section: Discussionmentioning

confidence: 99%

Polo-Like Kinase 1 is a Therapeutic Target in High-Risk Neuroblastoma

Ackermann

Goeser

Schulte

et al. 2011

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Purpose: High-risk neuroblastoma remains a therapeutic challenge for pediatric oncologists. The Pololike kinase 1 (PLK1) is highly expressed in many human cancers and is a target of the novel small-molecule inhibitor BI 2536, which has shown promising anticancer activity in adult malignancies. Here, we investigated the effect of BI 2536 on neuroblastoma cells in vitro and in vivo to explore PLK1 as a potential target in high-risk neuroblastoma therapy.Experimental Design: PLK1 transcript levels were analyzed by microarrays in 476 primary neuroblastoma specimens, and correlation with prognostic markers and patient outcome was examined. To explore the effect of PLK1 inhibition on neuroblastoma cells, 7 cell lines were treated with BI 2536 and changes in growth properties were determined. Furthermore, nude mice with IMR-32 and SK-N-AS xenografts were treated with BI 2536.Results: PLK1 is highly expressed in unfavorable neuroblastoma and in neuroblastoma cell lines. Expression of PLK1 is associated with unfavorable prognostic markers such as stage 4, age >18 months, MYCN amplification, unfavorable gene expression-based classification, and adverse patient outcome (P < 0.001 each). On treatment with nanomolar doses of BI 2536, all neuroblastoma cell lines analyzed showed significantly reduced proliferation, cell cycle arrest, and cell death. Moreover, BI 2536 abrogated growth of neuroblastoma xenografts in nude mice.Conclusions: Elevated PLK1 expression is significantly associated with high-risk neuroblastoma and unfavorable patient outcome. Inhibition of PLK1 using BI 2536 exhibits strong antitumor activity on human neuroblastoma cells in vitro and in vivo, opening encouraging new perspectives for the treatment of high-risk neuroblastoma.

“…To validate how well the cell line-based ALK downstream transcriptional profile recapitulates ALK oncogenic activity, we evaluated ALK signature scores in 2 independent gene expression datasets consisting of respectively 252 and 283 neuroblastoma tumors (19,24,25), as well as data of 2 in vivo neuroblastoma model systems (20). In human primary tumors with ALK amplification or ALK mutations, ALK signature scores were significantly higher than ALK wild-type tumors ( Fig.…”

Section: Aldh6a1mentioning

confidence: 99%

Upregulation of MAPK Negative Feedback Regulators and RET in Mutant ALK Neuroblastoma: Implications for Targeted Treatment

Lambertz

Kumps

Claeys

et al. 2015

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Purpose: Activating ALK mutations are present in almost 10% of primary neuroblastomas and mark patients for treatment with small-molecule ALK inhibitors in clinical trials. However, recent studies have shown that multiple mechanisms drive resistance to these molecular therapies. We anticipated that detailed mapping of the oncogenic ALK-driven signaling in neuroblastoma can aid to identify potential fragile nodes as additional targets for combination therapies.Experimental Design: To achieve this goal, transcriptome profiling was performed in neuroblastoma cell lines with the ALK F1174L or ALK R1275Q hotspot mutations, ALK amplification, or wild-type ALK following pharmacologic inhibition of ALK using four different compounds. Next, we performed cross-species genomic analyses to identify commonly transcriptionally perturbed genes in MYCN/ALK F1174L double transgenic versus MYCN transgenic mouse tumors as compared with the mutant ALKdriven transcriptome in human neuroblastomas.Results: A 77-gene ALK signature was established and successfully validated in primary neuroblastoma samples, in a neuroblastoma cell line with ALK F1174L and ALK R1275Q regulable overexpression constructs and in other ALKomas. In addition to the previously established PI3K/AKT/mTOR, MAPK/ERK, and MYC/ MYCN signaling branches, we identified that mutant ALK drives a strong upregulation of MAPK negative feedback regulators and upregulates RET and RET-driven sympathetic neuronal markers of the cholinergic lineage.Conclusions: We provide important novel insights into the transcriptional consequences and the complexity of mutant ALK signaling in this aggressive pediatric tumor. The negative feedback loop of MAPK pathway inhibitors may affect novel ALK inhibition therapies, whereas mutant ALK induced RET signaling can offer novel opportunities for testing ALK-RET oriented molecular combination therapies.