NEW YORK (GenomeWeb) – The University Medical Center Utrecht is offering a range of next-generation sequencing-based tests, including noninvasive prenatal testing on Life Technologies' SOLiD 5500xl Wildfire system, exome sequencing for intellectual disability on the Illumina HiSeq, and targeted gene panels on the SOLiD 5500xl. In addition, it is working with the department of immunology and hematology to develop HLA typing assays on the MiSeq and is working with the department of pathology to run oncology panels on the Ion Torrent PGM.
Hans Kristian Ploos van Amstel, who heads UMC Utrecht's Genome Diagnostics Laboratory, told Clinical Sequencing News that the laboratory has ISO 15189 certification, similar to CLIA certification in the US, and that the laboratory launched its NIPT test for trisomies 21, 18, and 13 in April for women at higher risk of fetal aneuploidy.
For NIPT, the lab is using Life Tech's SOLiD 5500xl platform with the Wildfire sample prep upgrade and has two such instruments dedicated for that test, van Amstel said. Life Tech launched its Wildfire sample prep technology for the SOLiD 5500 platform in 2012. Wildfire uses isothermal amplification, rather than emulsion PCR, in the sample prep workflow. In addition, sample prep is done directly on the flow cell by essentially coating both sides of the flow cell with DNA, which is then isothermally amplified. These features enable two- to five-fold higher throughput per lane and reduced sample prep time and cost, compared to the 5500 system without Wildfire.
According to Edwin Cuppen, who heads the research lab in the department of human genetics at UMC Utrecht out of which many of the NGS-based diagnostics are initially developed, the Wildfire instrument is well suited for NIPT applications since the elimination of emPCR reduces hands-on time and the increased throughput helps reduce cost while still generating millions of reads per sample.
For short tag-based counting approaches, it is the "cheapest platform," Cuppen told CSN.
In the first week after the test launched, the lab received clinical samples from 34 patients, van Amstel said, adding that turnaround time is around two weeks.
The lab runs the test using a whole-genome shotgun sequencing approach with read lengths of 35 bases. The short reads keep sequencing times down to around two days, Cuppen said, and enable a raw output of around 500 million reads per lane. For NIPT and other counting applications, the number of mappable reads is more important than read length, he added.
The Dutch government recently approved next-gen sequencing-based NIPT in high-risk women as part of a study dubbed TRIDENT, for Trial by Dutch Laboratories for Evaluation of Noninvasive Prenatal Testing. UMC Utrecht and others are part of the NIPT consortium that will evaluate NIPT in high-risk women and, among other things, will aim to determine a price at which such testing will be reimbursed if it is ultimately found to be clinically useful and cost-effective.
Van Amstel said that in the UMC Utrecht laboratory, reagent costs are around €200 ($276) per test with the total raw costs, including reagents, equipment, and personnel, around €600 ($828).
In the Netherlands, all postnatal NGS tests are reimbursed at a flat rate of €800 ($1,100), he added. Aside from NIPT, van Amstel said the laboratory currently offers targeted sequencing panels. Those are run on the SOLiD platform without the Wildfire configuration. It has three systems devoted for these diagnostic panels.
Van Amstel said that Wildfire is convenient for NIPT applications because it enables the sequencing libraries to be loaded directly onto the flow cell, but that it is not ideal for targeted panels because the read quality is lower. Read quality is not as critical for NIPT, which bases aneuploidy calls on the proportion of reads that correspond to the chromosome of interest, but quality is crucial when looking for point mutations assessed by targeted panels.
Physicians have 84 different conditions for which they can order testing. Those tests are grouped together into nine enrichment panels from which "we select for sub-panels of genes," explained van Amstel. The nine enrichment panels comprise genes related to distinct subtypes of cardiomyopathies, congenital heart defects, epilepsy, kidney disease, primary immune deficiencies, red cell membrane disorders, thrombopenia, developmental dental anomalies, and neurological disorders. Only the genes related to the specific disorder for which testing is ordered are analyzed.
Finally, UMC Utrecht is also performing diagnostic exome sequencing on the Illumina HiSeq for children with intellectual disability or other likely recessive disorders where the causative gene is unknown.
Because clinical exome sequencing is still so new, van Amstel said that a reimbursement price is still "under discussion," and authorities are working out whether the current €800 price point for other genetic tests is suitable for exome sequencing.
UMC Utrecht began offering diagnostic exomes in February and has so far tested around 20 patients and found the likely causative mutation in about one-third of those patients, van Amstel said. He noted that the main challenge with exome sequencing has been in interpretation. It's "not a plug and play," he said.
Within a research setting, van Amstel said that the laboratory is testing the MiSeq on smaller gene panels and for HLA typing, and it's also testing the PGM on cancer panels in collaboration with other groups at UMC Utrecht.
Going forward, he said that while the diagnostic lab is open to bringing in new technology, the panels that currently run on the SOLiD are performing well and the lab would only consider switching to a different system if significant improvements in turnaround time, quality, or cost could be demonstrated. "There's a limit to what you can invest in," he said, and validating new platforms for a diagnostic setting takes time.