Advanced Genetic Sequencing Uncovers New Variants in Vascular Anomalies

The Children’s Hospital of Philadelphia (CHOP) researchers have utilized advanced “deep sequencing” to uncover new genetic variants related to vascular anomalies in patients’ genomes. Over 60% of patients exhibited improvements in their condition as a result of tailored therapies informed by these new discoveries, paving the way for future non-invasive diagnostic methods for these life-threatening conditions. The study revealed genetic variants at significantly low levels, resulting in the initiation of new treatments for many patients.

In this meticulous study, CHOP researchers applied “deep sequencing” to the genomes of patients with serious vascular anomalies, using tissue samples and cell-free DNA. This approach identified numerous disease-related genetic variants missed by conventional genetic sequencing techniques. As a result, over 60% of patients witnessed improvements after targeted therapies based on these newly discovered variants. The findings were published in the journal Nature Medicine.

Vascular anomalies represent a range of conditions affecting veins, arteries, and the lymphatic system. These can be categorized as vascular tumors or vascular malformations, both benign or malignant. Some vascular anomalies resolve naturally, while others can lead to visible deformities, severely impair vital organ functions, or cause intense pain. Certain vascular anomalies can even be life-threatening.

A previous study by CHOP researchers, also published in Nature Medicine, was the first to discover a genetic variant causing a vascular anomaly in the lymphatic system. This led to the successful repurposing of an existing drug to treat a patient, greatly improving their breathing capacity and significantly reducing leg swelling, symptoms of their condition.

CHOP researchers suspected that other patients with vascular anomalies might also possess disease-driving mutations that could benefit from targeted therapies. However, limitations in accessing affected tissue samples or inadequate genomic sequencing data meant these disease-causing gene variants might go undetected with conventional genetic testing.

Senior study author Hakon Hakonarson, MD, PhD, said, “While some patients have inherited variants detectable in a blood sample, about 90% of patients with vascular anomalies possess acquired somatic mutations, usually present in very low frequencies and only in certain cell or tissue types.” He added, “In many instances, the disease-causing variant in the mutated gene of interest is present in frequencies of less than 1%, making them difficult to detect with conventional sequencing methods.”

To address this challenge, the research team analyzed DNA from CD31+ cells or cell-free DNA obtained from lymphatic fluid or plasma from 356 patients, including 104 with primary complex lymphatic anomalies. The DNA underwent deep sequencing, revealing several new somatic variants. This technique achieved a variant allele frequency of 0.15%, enabling the detection of variants with a frequency as low as 0.15% in a specific sample.

Identifying these variants enabled the researchers to offer a molecular diagnosis in 41% of patients with primary complex lymphatic anomalies and 72% of patients with vascular malformations. As a result, 69 patients initiated or planned to start new medical treatment, and 63% of patients experienced significant symptom improvement.

Study co-leader Sarah Sheppard, MD, PhD, highlighted the significance of their research, saying, “Our study comprehensively demonstrated the bedside to bench and back approach – from the molecular studies that found the low allele frequency variants to the functional studies in organoids and zebrafish that ultimately benefited the patients by directing medical therapy.”

Another study co-leader, Dong Li, PhD, said, “Our findings pave the way for future applications of cfDNA technology to be an innovative, non-invasive molecular diagnostic for all patients with vascular anomalies. We believe the time is right to transform the understanding of these complex diseases and identify and test new therapies for these life-threatening and life-altering conditions.”

This study was made possible by the support of a Children’s Hospital of Philadelphia Frontier Program Grant, K-Readiness Grant and Endowed Chair in Genomic Research; the National Center for Advancing Translational Sciences of the National Institutes of Health under grant 5R21TR00333; a research grant from the Lymphatic Malformation Institute; and the Eunice Kennedy Shriver National Institute of Child Health and Human Development under grant ZIA-HD009003-01.