They have the world record for fastest sequencing-based diagnosis in the world, at 19.5 hours, but Rady Children’s Institute for Genomic Medicine (RCIGM) isn’t stopping there. “We aim to use sequencing to diagnose 10,000 children with rare diseases by sometime in 2022,” said Stephen Kingsmore, M.D., president and CEO of the Institute. As of spring 2019, they had sequenced and interpreted the genomes of more than 750 children and were able to make a diagnosis in about 25% of those cases.
Sequencing for rare disease diagnosis is one of the fastest growing fields in genomic medicine. The cost of sequencing has dipped significantly, while the number of known variants that cause such diseases has steadily risen. As a result, more and more hospitals are moving toward earlier, and more thorough, sequencing for infants and children with undiagnosed diseases.
Hurdles still abound. More than half of undiagnosed patients will not get answers, even after sequencing, mainly because all the possible mutations that cause disease have not yet been uncovered. Approximately 7,000 rare diseases have been described and more than 250 new ones are found each year.
GeneDx, for example, is one of the world’s most active DNA testing laboratories, providing services in more than 55 countries including the U.S. The company, which is a subsidiary of BioReference Laboratories, recently announced it has completed 100,000 exome sequences. “Over the past few years, GeneDx has helped discover and publish on more than 62 disease-causing genes,” said Managing Director Ben Solomon, M.D.
The company’s database now contains more than 150,000 exomes. Additional clinical information, including imaging tests and family history, is combed through by in-house GeneDx curators who enter terms in the database to help document phenotypes. “This continually expanded dataset, along with our proprietary state-of-the-art variant analysis software, lets us provide highly accurate variant interpretation results for making definitive diagnoses possible, even in very complex cases,” Solomon said.
Since this is a newer, but rapidly expanding field, there also aren’t enough physicians who know how to request these genomic tests, let alone how to interpret or act upon their results. “When we get a diagnosis, we have to ask where the patient goes next,” Kingsmore explained. “Many physicians still have not been trained in how to follow up on a genetic diagnosis.”
The most challenging holdup may not be in the clinic (See “Bringing Payers Along” page 15). “Hands down, reimbursement is the biggest problem,” said Catherine Brownstein, Ph.D., scientific director of the Manton Center for Orphan Disease Research and research associate in the Division of Genetics and Genomics at Boston Children’s Hospital. “I have confidence that many more clinicians will adopt NGS for rare diseases once reimbursement is settled.”
But these challenges are not enough to slow pioneers in the field. Experts in the clinic and industry are working to achieve a bold vision where “it’s not just a single test that you do or don’t do, it’s a part of an integrated health system where the process is seamless and children with rare diseases anywhere are getting the right treatment, quickly,” Kingsmore said. That process should not just be quicker, it also should be more intuitive so that clinicians can easily understand how to interpret each patient’s sequencing results.
Full Speed Ahead
Rady Children’s is well-recognized as the leader in this field. Its team has worked with several leading technology and data-sciences providers to create a rapid Whole Genome Sequencing (rWGS) process, which was detailed in Science Translational Medicine in April 2019. This pipeline “required minimal user intervention, increasing usability and shortening time to diagnosis,” the team reported. A provisional diagnosis was reached in a median time of less than 24 hours.
The pipeline’s current core components include Illumina’s Nextera DNA Flex Library preparation, the NovaSeq 6000 for sequencing, and DRAGEN (Dynamic Read Analysis for GENomics) for secondary analysis. It also includes Clinithink’s clinical natural language processing platform CliX ENRICH to extract phenotype information from a patient’s electronic medical record. Diploid’s MOON is used to automate genome interpretation using AI and filters and ranks likely pathogenic variants, taking just a few minutes to reveal a causal mutation out of the 4.5 million variants usually found in a whole genome. Alexion’s “rare disease and data science expertise enables the translation of clinical information into a computable format for guided variant interpretation,” according to a Rady press release. And Fabric Genomics’ AI-based clinical decision support software (Fabric Enterprise) confirms the output of the automated pipeline.
Thanks to this optimized platform, Rady is able to reach a diagnosis for a patient using sequencing data in 40 hours or less, whenever speed is of the essence. “There are times when you must get the results to the NICU as soon as possible to try and save the child’s life,” Kingsmore said.
Rady also offers this service to others. It currently has collaborations with 23 hospitals to provide them sequencing services for rare diseases. The Institute’s goal is to have 40 such partnerships before the end of next year. It is also teaching others how to incorporate WGS into clinical practice. Through its collaboration with Vermont Oxford Network (VON), Rady is part of a learning network that helps NICU teams improve their competency in genomic medicine. VON is a nonprofit voluntary collaboration of more than 1,200 hospitals working to improve neonatal care around the globe. “We’ve optimized the steps for everything from what type of test to order through delivering results,” said Kingsmore. “The algorithm just spits out the next step you need to take to properly manage the patient.”
Congenica, similarly, offers a complete solution for sequence interpretation. The company’s Congenica decision support platform leverages its proprietary datasets, knowledgebase, and expertise for variant detection and interpretation. “Once you move to an exome or whole genome you get so many thousand more putative variants; it’s crucial to filter them efficiently,” said Nick Lench, Congenica’s chief scientific officer and cofounder. Congenica essentially put a wrapper around a mix of top public and private databases and algorithms for sequence analysis. “We also bring proprietary information and the ability to handle data at scale,” Lench added. One of the company’s clients is Genomics England and its 100,000 Genomes Project, which has reached its first milestones and is now aiming to sequence five million genomes in five years.
Recently, Congenica and BGI Genomics inked a deal that combines BGI’s exome sequencing service with Congenica’s interpretation and clinical expertise. Lench said Congenica has more than 700 registered users from 40 to 50 organizations, mostly in the U.K., the U.S., and China. BGI operates in more than 100 regions and works with more than 3,000 medical institutions and more than 300 hospitals. Through this collaboration, the two companies anticipate “We will be able to offer partners across the world access to a trusted, premium clinical research whole-exome service that supports diagnostic confidence at extremely accessible pricing,” said Ning Li, vice president of BGI Group in a statement.
Attention has also been drawn to the lack of diversity in most of the genomic data we have. The majority of sequencing has taken place in the U.S. and Europe, which means there are potentially many variants that are specific, or just more common, in certain populations that are not yet known.
With regard to Singapore, for example, researchers have found an unexpectedly high carrier frequency for certain treatable inherited disorders. That research was carried out by Roger Foo, M.D., colleagues, and collaborators. Foo is senior consultant and professor at National University Heart Centre and National University of Singapore.
In a study of 831 Singaporeans, they found that citrin deficiency and Wilson Disease had rate of 1 in 41 and 1 in 103 respectively. That study was published in Genetics in Medicine last year. Singapore is rapidly increasing the amount of sequencing it performs and Foo says his group is keeping a database of the variants they find. It shares these through publications and Matchmaker Exchange, a project started in 2013 that links projects and research efforts around the world to aggregate genomic data for the purpose of better understanding and diagnosing rare diseases.
India recently announced it will launch its first human genome cataloging project—Genome India—in fall of 2019. The project’s first phase aims to sequence 10,000 Indian nationals. One of the institutions participating in this effort is the newly inaugurated Next Generation Sequencing facility at the CSIR-Centre for Cellular and Molecular Biology (CCMB) in Hyderabad. That facility now boasts top flight equipment from manufacturers such as Illumina, Thermo Fisher, Oxford Nanopore, and 10x Genomics. The CCMB can sequence up to 30 human genomes or 384 diagnostic samples in a day. “It used to take about seven to ten days to do one sample,” noted K Thangaraj, chief scientist at CCMB.
While the facility will generate diagnoses, it will also help India catch up in gathering genomic data of its population. The genetic causes of many conditions affecting Indians are unknown, but it is known the country has a substantial rate of genetic disease. “Many Indians have been marrying within a specific community for a long time,” Thangaraj explained. “While there are several thousand such communities, it is likely that there is a concentration of variants among certain groups.”
Other countries are already well advanced in gathering genomes from its residents, including England, Qatar, and Finland.
Centogene may have an advantage in leveraging that data. Genomic-data and interpretation company Centogene believes it may have an advantage in leveraging broad, population-based genomic data. The company focuses on rare disease, and has clients around the world, which help it build a knowledgebase that is increasing in diversity.
“Everything we have learned about genetics tells us that ethnic background plays a role,” said Oved Amitay, chief business officer at Centogene. “We receive samples from about 110 different companies around the globe,” he added. “Meanwhile, most other groups like ours are focused on Caucasian populations.” He pointed out that transthyretin (TTR) amyloidosis, for example, progresses more slowly in Portuguese natives than in people of different heritage. Likewise, BRCA1 conveys a different level of risk in people from Canada, versus those from Japan.
“No one yet knows why these geographic variations in phenotype happen,” said Amitay. Centogene believes it has an advantage because “Sixty percent of the variations in our database will not be found in other databases,” he added. Most recently, the company signed a Memorandum of Understanding with the Republic of Tajikistan, to “stimulate and facilitate the well being of patients with genetic diseases,” and more, according to a company release.
Tackling More Complex Diseases
Scientists are now also pushing to get more use of sequencing from complex disorders, such as mental illness and autism. One challenge, Brownstein pointed out, is that some genetic variants produce multiple phenotypes.
Duplications or deletions in 16p13.11, for example, most often lead to developmental delay, intellectual deficiency, or autism spectrum disorder. However, they can also lead to a range of other symptoms, including heart defects. As a result, a patient who is diagnosed with one of these variations should be screened for heart disease, whether or not they have obvious symptoms. “We need to understand what is causing that variability,” said Brownstein. CNVs may, for example, be acting together. “To sort that out, we need more samples and more collaborations,” Brownstein added.
One thing that is clear is that the shift toward WGS has many potential benefits. Boston Children’s Hospital clinicians, for example, see many patients who have been referred, sometimes from other countries. And they may have undergone many types of testing already. “A lot of patients come to us, and they have had a panel done, or maybe an exome, but not WGS yet,” she explained. “Or maybe the patient was sequenced and nothing was found, but the parents haven’t been sequenced.” More thorough sequencing should help scientists find more variants that count and thereby help more patients.
There are also certain variants that are harder to detect than others, such as repeat expansions and structural variants, but we need more data about all of these to get a truly complete picture of the genome.
“People want the most comprehensive, cost-effective, and accurate clinical diagnosic test with the shortest turn-around-time,” said Christine Stanley, chief director of clinical genomics at Variantyx. But because of reimbursement challenges, for most genetic testing “You have one shot at getting the answer,” she explained.
Variantyx aims to solve that dilemma by offering comprehensive whole-genome clinical interpretation software that analyzes single nucleotide variants (SNVs), structural variants (SVs), short tandem repeats (STRs), and mitochondrial variants. “PCR-free WGS in conjunction with Variantyx Genomic Intelligence software eliminates the need for multiple tests by providing results on all types of genetic variants in one report,” Stanley said. The company published on their technology’s ability to detect and analyze structural variants in the July 2019 issue of BMC Genomics. The platform “identifies structural variants down to single base pair resolution,” they reported.