Over the past few years, Stephen Kingsmore, MB ChB BAO, D.Sc., has pioneered the use of clinical next-generation sequencing (NGS) for the diagnosis of babies admitted into the newborn intensive care unit (NICU). That expertise was honed at Children’s Mercy Hospital in Kansas City, Missouri before Kingsmore moved in 2015 to the Rady Children’s Hospital in San Diego, California.

In a talk at the American Society of Human Genetics annual conference, Dr. Kingsmore presented his vision for enhancing and expediting the pipeline, which he believes will become widely adopted at children’s hospitals around the world. Kingsmore already holds the Guinness World Record for the fastest genetic diagnosis—26 hours—which was set using Illumina’s HiSeq 2500 sequencing instrument (in rapid run mode).

About 5 percent of all newborns are admitted to the NICU. The ensuing diagnostic odyssey can either be short or long. “Some babies will die. Others will be discharged without a diagnosis,” said Dr. Kinsgmore. At the center of Dr. Kingsmore’s vision is Illumina’s latest sequencer, the NovaSeq 6000. The goal is “to obtain a diagnosis as rapidly as possible” which would enable “scalable, high-throughput analysis to be used at various hospitals around the world.”

In brief, the Rady pipeline uses natural language processing to extract information from the patient’s medical record. This is converted into a standard vocabulary that enables pattern recognition, which is then compared against some 8,000 known genetic diseases, which are listed in rank order.

Whole-genome sequencing (WGS) is performed on the newborns and their parents (if possible). For the most urgent cases, the newborn’s genome is currently sequenced on the HiSeq 2500, which typically takes about 30 hours. The sequence data are fed into a rapid analysis pipeline using Edico Genome’s DRAGEN processing platform and Fabric Genomics’ Opal clinical variant platform for interpretation. “We can obtain a provisional diagnosis in two days if [the newborn is] sick enough to warrant that,” Dr. Kingsmore said.

Fuel the Vision

The Rady team’s experience with the first 42 cases in this paradigm “fuel the vision”, said Dr. Kingsmore. Eighteen (40 percent) of the 42 newborns received a diagnosis after genome analysis, of which 13 obtained a change in disease management. Overall, 11 (24 percent) of NICU newborns sequenced received a change in outcome.

“These are remarkable findings that are going to change the way medicine is practiced in the NICU,” said Dr. Kingsmore.

“This [approach] is still not well advertised, well known, or well implemented in children’s hospitals around the world,” he added. “We believe this will become a paradigm for treating children around the world.”

Dr. Kingsmore described a recent successful outcome in a case known as Infant 6041. A two-day-old baby girl named Sebastiana experienced seizures beginning just 16 hours after delivery. Standard drugs were ineffective or required such high doses that the baby was lethargic and couldn’t feed.

Within 68 hours, Kingsmore’s team had reached a diagnosis of Ohtahara syndrome, having discovered a de novo missense mutation in a gene encoding a potassium channel, KCNQ2. A change in treatment resulted in control over the seizures, and Sebastiana was discharged 18 days after admission. A similar case at the hospital had been admitted a year earlier, but without the full genome analysis, it took doctors two months to reach the same diagnosis, and the patient in that case has major brain damage. Moreover, the reduced time to Sebastiana’s diagnosis resulted in cost savings of about $180,000.

Enter the NovaSeq

While the current Rady pipeline is among the most efficient in the world, there is room for improvement. “The HiSeq 2500 does a beautiful job,” Dr. Kingsmore said, “but it only runs one sample at a time and it is rather costly.”

Rady has just validated the NovaSeq instrument, placing it into clinical (CLIA/CAP) production earlier this month. “Almost everything is different about the platform,” Dr. Kingsmore said. Initial results on 52 whole genomes sequenced showed excellent coverage of the entire genome (comparable to the HiSeq), including known Mendelian disease genes. Dr. Kingsmore said he favors the 2×100-nucleotide (compared to 2×150) approach, which he says saves a lot of time.

Dr. Kinsgmore believes in 40x coverage for newborn screening. “That’s our clinical specification,” he said, adding that the NovaSeq offers “an exquisitively sensitive and specific test.” Whereas the HiSeq 2500 produced a single human genome (at 40x coverage) over about 26 hours, requiring two flow cells, the NovaSeq can generate six genomes (also 40x) in a 24-hour run on a single flow-cell, for a third of the cost of the HiSeq.

Dr. Kingsmore’s groundbreaking work was recently profiled in TIME magazine in a story by Alice Park. He had hoped to announce an even shorter turnaround time, using a new technology that would prepare the DNA library directly from a blood sample without having to extract the DNA as an intermediate step. But that technique wasn’t ready in time.

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