title: INSPIIRED: Quantification and Visualization Tools for Analyzing Integration Site Distributions.
abstract: Analysis of sites of newly integrated DNA in cellular genomes is important to several fields, but methods for analyzing and visualizing these datasets are still under development. Here, we describe tools for data analysis and visualization that take as input integration site data from our INSPIIRED pipeline. Paired-end sequencing allows inference of the numbers of transduced cells as well as the distributions of integration sites in target genomes. We present interactive heatmaps that allow comparison of distributions of integration sites to genomic features and that support numerous user-defined statistical tests. To summarize integration site data from human gene therapy samples, we developed a reproducible report format that catalogs sample population structure, longitudinal dynamics, and integration frequency near cancer-associated genes. We also introduce a novel summary statistic, the UC50 (unique cell progenitors contributing the most expanded 50% of progeny cell clones), which provides a single number summarizing possible clonal expansion. Using these tools, we characterize ongoing longitudinal characterization of a patient from the first trial to treat severe combined immunodeficiency-X1 (SCID-X1), showing successful reconstitution for 15 years accompanied by persistence of a cell clone with an integration site near the cancer-associated gene CCND2. Software is available at https://github.com/BushmanLab/INSPIIRED.
authors.affiliation: Department of Family Medicine and Public Health, UC San Diego, La Jolla, CA 92093, USA.
authors.forename: Charles C
authors.initials: CC
authors.lastname: Berry
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Nobles
authors.affiliation: Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, 75015 Paris, France; INSERM 24, Laboratory of Human Lymphohematopoiesis, 75015 Paris, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Six
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Yinghua
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Nirav
authors.initials: N
authors.lastname: Malani
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Sherman
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Anatoly
authors.initials: A
authors.lastname: Dryga
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: John K
authors.initials: JK
authors.lastname: Everett
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Frances
authors.initials: F
authors.lastname: Male
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Aubrey
authors.initials: A
authors.lastname: Bailey
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Kyle
authors.initials: K
authors.lastname: Bittinger
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Mary J
authors.initials: MJ
authors.lastname: Drake
authors.affiliation: Biotherapy Department, Necker Children's Hospital, Assistance Publique-Hôpitaux de Paris, 75014 Paris, France; Biotherapy Clinical Investigation Center, Groupe Hospitalier Universitaire Ouest, Assistance Publique-Hôpitaux de Paris, INSERM, 75014 Paris, France.
authors.forename: Laure
authors.initials: L
authors.lastname: Caccavelli
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Bates
authors.affiliation: Biotherapy Department, Necker Children's Hospital, Assistance Publique-Hôpitaux de Paris, 75014 Paris, France; Biotherapy Clinical Investigation Center, Groupe Hospitalier Universitaire Ouest, Assistance Publique-Hôpitaux de Paris, INSERM, 75014 Paris, France.
authors.forename: Salima
authors.initials: S
authors.lastname: Hacein-Bey-Abina
authors.affiliation: Biotherapy Department, Necker Children's Hospital, Assistance Publique-Hôpitaux de Paris, 75014 Paris, France; Biotherapy Clinical Investigation Center, Groupe Hospitalier Universitaire Ouest, Assistance Publique-Hôpitaux de Paris, INSERM, 75014 Paris, France.
authors.forename: Marina
authors.initials: M
authors.lastname: Cavazzana
authors.affiliation: Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104-6076, USA.
authors.forename: Frederic D
authors.initials: FD
authors.lastname: Bushman
keywords: SCID-X1; data visualization; gammaretrovirus; gene therapy; insertional mutagenesis; lentivirus; mutagenesis; recombination; retrovirus; vector; vector driving