Background: High risk neuroblastoma (HRNB) patients have a poor response to chemotherapy, with 20% unable to achieve complete response. Genomic studies identified 6 targeted agents whose pathways are active in HRNB. The low frequency of established driver mutations, along with the variety of molecular mechanisms for cancer pathway dysregulation, suggest development of gene expression-based predictive biomarkers in addition to sequence-based biomarkers. Our platform for developing gene expression-based biomarkers includes primary NB cell lines, high throughput quantification of cell survival, exome sequencing, and genome-wide expression profiling. We apply this platform to six commonly used targeted therapies.
Methods: Cell survival was quantified for each of thirty primary neuroblastoma cell lines in the presence of each of six drugs (bortezomib, crizotinib, dasatinib, lapatinib, sorafenib, and vorinostat) using high throughput cell survival measurements. Titration curves were based on vehicle and sixteen concentrations of each drug (replicated within and between plates). Univariate summaries of the response of cell line/drug were obtained using the four parameter logistic model as well as the area under the curve. Exome sequences were obtained using Illumina HiSeq technology. Genome-wide expression profiles were obtained using Affymetrix GeneChip (U133 Plus 2.0). Exploratory multivariate analysis included hierarchical clustering, principal component analysis, and multidimensional scaling. Inference was based on parametric univariate and non-parametric multivariate tests.
Results: For three of the drugs (bortezomib, dasatinib, and vorinostat) variation in response between cell lines was statistically significant (p < 0.05; F ≥3) and assay results are reproducible in the context of the differences between cell lines (the other three drugs the estimated standard deviation of log10 EC50 ≤ 0.2). Genome-wide expression profiles were associated with variation in drug response between cell lines (example: high PDGFRB expression indicates sensitivity to dasatinib, p < 0.001).
Conclusion: High throughput cell survival measurements provided for identification of predictive biomarkers of Neuroblastoma drug sensitivity. These biomarkers are currently being studied in the NMTRC PEDS-PLAN (Pediatric Precision Laboratory Advanced Neuroblastoma Therapy) clinical trial.