Genomics & Bioinformatics
Gene editing is a revolutionary technology for curing genetic disorders. It is accomplished by using designer nucleases (CRISPR-Cas, TALEN), base editors, prime editors, designer nickases, and other tools that enable targeted gene knockout or correction. Despite the general high specificity of these genome editors, unintended alterations in the genome (off-target effects) have been observed. If such off-target activity occurs within a gene or regulatory region of the genome, it can lead to unexpected side effects, potentially resulting in tumorigenesis. Consequently, the activity and specificity of each genome editor must be thoroughly validated before any clinical application.
Our lab has developed several methods to detect off-target effects of genome editors, incl. CAST-Seq, a highly sensitive assay that identifies chromosomal rearrangements and nominates off-targets sites (patents US11319580B2/EP3856928B1). CAST-Seq can be employed in in vivo or ex vivo edited primary cells. Our established bioinformatics pipeline filters the CAST-Seq results and reliably nominates off-targets.
Additionally, the nominated off-targets can be validated by a combination of next generation sequencing (NGS)-based amplicon sequencing (short-read Illumina) and/or rhAmpSeq™ assays.
If you are interested in our CAST-Seq technology or want to request our off-target service as a fee-for-service, please contact Sandra Ammann (sandra.ammann@uniklinik-freiburg.de).
Our publications using CAST-Seq:
| Year | target(s) | Nuclease | Journal | Cells and Methode | DOI |
| 2024 | HBB, CCR5, GAPDH + DNA-PKcs inhibitor AZD7648 | spCas9 | Nature Biotech. | human HSCs, RPE
| 10.1038/s41587-024-02488-6 |
| 2024 | NCF1 | spCas9, Cas12a | Communications Biology | HSC CAST-Seq | 10.1038/s42003-024-06959-z |
| 2024 | CD40LG + DNA-PKcs inhibitor AZD7648 | TALEN, spCas9 | Nature Biotech. | human HSCs CAST-Seq, | 10.1016/j.omtm.2024.101297 |
| 2024 | mAlb | Cas9, AAV-Hiti | Cell Rep Med | in vivo CAST-Seq | 10.1016/j.xcrm.2024.101619 |
| 2024 | COL7A1, COL17A, LAMA3 | SpCas9, D10ASaCas9 | Mol Ther | CAST | 10.1016/j.ymthe.2024.03.006 |
| 2024 | ELANE | Cas9HiFi, Cas9D10A | Mol Ther | D-CAST | 10.1016/j.ymthe.2024.03.037 |
| 2023 | CCR5 (TALEN) | TALEN | Front Genome Ed. | T-CAST | 10.3389/fgeed.2023.1130736 |
| 2023 | CD40L, CSF2 | Cas9 | Cells | double KO, CAST-Seq | 10.3390/cells12212581 |
| 2023 | mUnc13d | spCas9 | JACI | in vivo, mouse HSCs, CAST-Seq | 10.1016/j.jaci.2023.08.003 |
| 2023 | CCR5 CRISPR/Cas9-CtIP-dnRNF168 | spCas9, Cas9-CtIP-dnRNF168 | NAR | CAST-Seq | 10.1093/nar/gkad255 |
| 2023 | CD3Z | spCas9 | Blood | T and NK cells CAST-Seq | 10.1182/blood.2023020973 |
| 2023 | CD123 | Cas9 | J Exp Med | human HSCs CAST-Seq | 10.1084/jem.20231235 |
| 2023 | HBB (p53i) | Cas9 | Nat Commun | human HSCs CAST-Seq | 10.1038/s41467-023-43413-w |
| 2023 | globin | Cas9 | Mol Ther Nucleic Acids | human HSC, CAST-Seq | 10.1016/j.omtn.2023.04.024 |
| 2023 | mHao1 | D10ASaCas9 | EMBO Mol Med | in vivo, D-CAST | 10.1038/s44321-023-00008-8 |
| 2021 | rhCD33 | Cas9 | Mol Ther | NHP, CAST-Seq | 10.1016/j.ymthe.2021.06.016 |
| 2021 | CCR5, HBB, FANCF, VEGFA, RAG1 | Cas9, Cas9HIFI, TALEN | Cell Stem Cell | HSCs T-CAST | 10.1016/j.stem.2021.02.002 |
Dr. Sandra Ammann
Project Manager
Research & Development
Medical Center Freiburg
Institute for Transfusion Medicine and Gene Therapy
at Center for Translational Cell Research
Breisacher Straße 115
79106 Freiburg
+49 761 270 77748
sandra.ammann@uniklinik-freiburg.de