March 15, 2018  |  Human genetics research

Study: SMRT sequencing can improve the safety of gene therapy protocols

A new publication in the journal of the American Society of Gene & Cell Therapy demonstrates the novel use of SMRT Sequencing for improving the safety and quality of a type of gene therapy delivered through a viral vector. The work established new quality control standards and revealed previously undetected risks for gene therapy delivery.

Adeno-Associated Virus Genome Population Sequencing Achieves Full Vector Genome Resolution and Reveals Human-Vector Chimeras,” published in Molecular Therapy — Methods & Clinical Development, comes from lead author Phillip Tai, senior author Guangping Gao, and collaborators from the University of Massachusetts Medical School and other institutions. The team aimed to characterize the adeno-associated virus (AAV) vectors that have gained traction recently as delivery vehicles for gene therapy. Previously, there had been no reliable, comprehensive method for assessing the integrity of those AAV vectors prior to introducing them to patients.

The team used PacBio long-read sequencing to analyze a population of AAV vectors, providing the first high-resolution view of the DNA sequences they would deliver to a patient. They found more problems than expected: some vectors contained less than half of the DNA sequence they should have, while genetic errors called chimeras were discovered in other vectors. In a patient, these vectors would have reduced the likelihood of successful gene therapy outcomes.

With this approach, known as AAV-genome population sequencing or AAV-GPseq, “we can comprehensively profile packaged genomes as a single intact molecule and directly assess vector integrity without extensive preparation,” the scientists report. This detailed view allowed the team to spot reverse-packaged genomes, chimeras with inverted terminal repeat-containing sequences, and truncated genomes. “These discoveries redefine quality control standards for viral vector preparations and highlight the degree of foreign products in [recombinant AAV]-based therapeutic vectors,” the team writes.

The full AAV-GPseq method is available and “can be easily adapted for research-grade and clinical vector manufacturing QC pipelines,” the scientists add.

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