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Deep Whole-Genome Sequencing Diagnoses Early Infantile Epileptic Encephalopathy

NEW YORK (GenomeWeb) – Using deep sequencing and computational tools, researchers were able to tease out the genetic root of early infantile epileptic encephalopathy (EIEE) in a dozen patients.

Early infantile epileptic encephalopathy affects a small number of children, but can lead to developmental delays, intellectual impairment, and even early death. While variants in more than 50 genes have been linked to the condition, University of Utah researchers said gene panel- and whole-exome sequencing-based diagnostics have a yield of about 60 percent.

The researchers combined deep whole-genome sequencing and a new reference-free de novo variant calling algorithm they developed and applied it to a cohort of 14 patients for whom previous testing led to no diagnosis. They uncovered causal mutations in 12 of these patients. As they wrote in npj Genomic Medicine today, the researchers said their findings suggest whole-genome analysis could soon be a standard approach for diagnosing EIEE.

"These families have been drifting through expensive prolonged testing with little hope of finding an answer," first author Betsy Ostrander from University of Utah School of Medicine said in a statement. "We can now identify the genetic cause of EIEE and select medications best suited to each patient to decrease the frequency of seizures earlier and hopefully prevent developmental delays."

Ostrander and her colleagues used the Illumina HiSeq X platform to sequence the 14 individuals they recruited into their study, as well both parents of each child, to an average 65X coverage. The researchers said they sought deep coverage to boost their accuracy in uncovered de novo variants.

They took two approaches to tease out variants within the trios. One relied on a combination of the GATK pipeline to find SNVs and indels with the LUMPY structural variant detection tool and the SVTyper tool that generates SV genotypes. They also used RUFUS, an alignment-free algorithm they developed to compare trio data to tease out de novo mutations. RUFUS, they reported, was able to identify all types of de novo variation in one step and with higher specificity.

The researchers prioritized the variants they found based on whether they were missense, frameshift, or nonsense variants that fell in known EIEE genes, candidate EIEE genes, or other regions using the GEMINI framework.

For nine patients, the researchers uncovered single, de novo variants in which they had high confidence in pathogenicity. For seven, they found de novo missense mutations affecting ion channel genes like SCNA1, SCN2A, and KCNQ2 that have known links to EIEE. In the two others, they found missense mutations affecting the EEF1A2 and PIGA genes.

In two additional patients, the researchers uncovered de novo missense of putative loss-of-function mutations in the protein-coding region of genes that hadn't before been associated with EIEE: DEAF1 and CAMK2G.

Since these mutations all fell within exonic regions, the researchers noted that exome sequencing rather than whole-genome sequencing might have been able to pick them up. However, they added that three of those patients had previously been evaluated using commercial, exome-based gene panels and hadn't received a diagnosis.

In one case, the researchers uncovered a de novo, inverted, balanced translocation affecting the X chromosome and chromosome 2. This, they said, likely alters X-inactivation and transcription patterns, possibly affecting the regulation of MECP2, and would only have been picked up through whole-genome sequencing.

The researchers argued that whole-genome sequencing offers a number of advantages for diagnosing monogenic diseases like EIEE. One such advantage, they said is cost. Even though the cost of sequencing remains high, they noted the participants in their study had undergone, in some cases, years of diagnostics tests — each had undergone a minimum of 24 diagnostic tests, costing an average $30,866. Whole-genome sequencing, they concluded, could save both time and money.