RNA sequencing has emerged as a powerful supplement to DNA sequencing for Mendelian disease diagnosis, but clinical translation of diagnostic RNA-seq has not been widely achieved. Researchers at Baylor College of Medicine's Medical Genetics and Multiomics Laboratory published the clinical validation of the first RNA sequencing test for diagnostic whole-transcriptome analysis for genetic disorders. The findings, published in the American Journal of Human Genetics, establish a comprehensive transcriptome sequencing pipeline for clinical use, expanding the role of RNA sequencing beyond targeted analysis.
"This validation represents the first attempt to unleash the full transcriptome potential of RNA sequencing for clinical diagnostics," said corresponding author Dr. Pengfei Liu, associate professor of molecular and human genetics and director of the Medical Genetics and Multiomics Laboratory (MGML) at Baylor. "While RNA sequencing has been used in clinical testing, it has been restricted to targeted gene analysis. In fact, when we applied for accreditation from the CAP (College of American Pathologists), we worked with them to create the first whole-transcriptome sequencing activity as a reportable assay."
The RNA-seq test uses samples from fibroblasts or blood and analyzes for outliers in gene expression and splicing patterns. Researchers validated the test using 130 samples, including 40 samples with a positive molecular diagnosis and 90 negative samples from apparently healthy people for control. They developed benchmarks for expression and splicing using RNA-seq data from a lymphoblastoid sample in the publicly available Genome in a Bottle Consortium. The team established reference ranges for each gene based on control data.
To assess clinical performance, the team evaluated the test using samples with previously identified diagnostic findings from the Undiagnosed Diseases Network (UDN). The study confirmed the assay's ability to detect diagnostic RNA findings through both transcriptome-driven and DNA-driven analysis modes, demonstrating its potential to enhance molecular diagnosis.
This test is the first launched by the MGML and now is being offered for clinical RNA sequencing in the UDN.
Other authors who contributed to this work include Sen Zhao, Kristina Macakova, Jefferson C. Sinson, Hongzheng Dai, Jill Rosenfeld, Gladys E. Zapata, Shenglan Li, Patricia Ward, Christiana Wang, Chunjing Qu, Becky Maywald, Brendan Lee and Christine Eng. They are affiliated with one or more of the following institutions: Baylor College of Medicine, Baylor Genetics and the University of Texas MD Anderson Cancer Center School of Health Professions.
This research was supported by the National Institutes of Health Common Fund (U01HG007709 and U01HG007942) and the National Human Genome Research Institute (R35HG011311).
