Ultra-High Density SNParray in Neuroblastoma Molecular Diagnostics

Abstract

Neuroblastoma serves as a paradigm for applying tumor genomic data for determining patient prognosis and thus for treatment allocation. MYCN status, i.e., amplified vs. non-amplified, was one of the very first biomarkers in oncology to discriminate aggressive from less aggressive or even favorable clinical courses of neuroblastoma. However, MYCN amplification is by far not the only genetic change associated with unfavorable clinical courses. So called “segmental chromosomal aberrations,” (SCAs) i.e., gains or losses of chromosomal fragments, can also indicate tumor aggressiveness. The clinical use of these genomic aberrations has, however, been hampered for many years by methodical and interpretational problems. Only after reaching worldwide consensus on markers, methodology, and data interpretation, information on SCAs has recently been implemented in clinical studies. Now, a number of collaborative studies within COG, GPOH, and SIOPEN use genomic information to stratify therapy for patients with localized and metastatic disease. Recently, new types of DNA based aberrations influencing the clinical behavior of neuroblastomas have been described. Deletions or mutations of genes like ATRX and a phenomenon referred to as “chromothripsis” are all assumed to correlate with an unfavorable clinical behavior. However, these genomic aberrations need to be scrutinized in larger studies applying the most appropriate techniques. Single nucleotide polymorphism arrays have proven successful in deciphering genomic aberrations of cancer cells; these techniques, however, are usually not applied in the daily routine. Here, we present an ultra-high density (UHD) SNParray technique which is, because of its high specificity and sensitivity and the combined copy number and allele information, highly appropriate for the genomic diagnosis of neuroblastoma and other malignancies.

Genomic Copy Number Profiling Using Circulating Free Tumor DNA Highlights Heterogeneity in Neuroblastoma

“…In neuroblastoma, it has been shown that the overall tumor genomic profile determined by copy number analysis of tumor material is of prognostic impact, and genomic copy number profiling using material from tumor cells is considered as a reference for obtaining a tumor genomic profile at the time of diagnosis (11,13,14). We have now undertaken a study of 70 plasma samples obtained at diagnosis to determine the feasibility of using cfDNA isolated from plasma for the study of an overall genomic profile.…”

“…Current treatment strategies take into account the genomic copy number profile both in patients with low-risk neuroblastoma, as well as in certain patients with high-risk neuroblastoma, aged <18 months (11,12). A genomic copy number profile is routinely assessed on diagnostic tumor tissue, most frequently using pangenomic techniques, such as MLPA, array comparative genomic hybridization (aCGH), or SNPa (11,13,14). However, in some instances, biopsies from a primary or metastatic tumor site are not available, for instance, in severely ill young infants with disseminated disease, whereas in older children biopsies might harbor only a low percentage of tumor cells, thus impacting on molecular genetic analysis.…”

Impact of Disseminated Neuroblastoma Cells on the Identification of the Relapse-Seeding Clone

“…23,30). CytoScan HD has a genome-wide coverage with more than 2.6 million copy number and SNP markers.…”

“…Because it includes 750,000 SNP probes, it is possible to detect regions with copy-neutral LOH and low level of mosaicisms. Subclonal frequencies were calculated by evaluating the smooth signal level and the allele difference track pattern and by considering the tumor cell content of the samples as described before (23,30,31).…”

Rawcopy: Improved copy number analysis with Affymetrix arrays

“…Genome-wide copy number analysis is commonly performed in hypothesis-generating genomics research, including many recent large-scale cancer studies3. It is also growing rapidly in clinical diagnostics as a high-resolution alternative or complement to in situ chromosome analysis45.…”