title: Towards a Safer, More Randomized Lentiviral Vector Integration Profile Exploring Artificial LEDGF Chimeras.
abstract: The capacity to integrate transgenes into the host cell genome makes retroviral vectors an interesting tool for gene therapy. Although stable insertion resulted in successful correction of several monogenic disorders, it also accounts for insertional mutagenesis, a major setback in otherwise successful clinical gene therapy trials due to leukemia development in a subset of treated patients. Despite improvements in vector design, their use is still not risk-free. Lentiviral vector (LV) integration is directed into active transcription units by LEDGF/p75, a host-cell protein co-opted by the viral integrase. We engineered LEDGF/p75-based hybrid tethers in an effort to elicit a more random integration pattern to increase biosafety, and potentially reduce proto-oncogene activation. We therefore truncated LEDGF/p75 by deleting the N-terminal chromatin-reading PWWP-domain, and replaced this domain with alternative pan-chromatin binding peptides. Expression of these LEDGF-hybrids in LEDGF-depleted cells efficiently rescued LV transduction and resulted in LV integrations that distributed more randomly throughout the host-cell genome. In addition, when considering safe harbor criteria, LV integration sites for these LEDGF-hybrids distributed more safely compared to LEDGF/p75-mediated integration in wild-type cells. This approach should be broadly applicable to introduce therapeutic or suicide genes for cell therapy, such as patient-specific iPS cells.
authors: Laboratory for Molecular Virology and Drug discovery, KU Leuven, Belgium.
authors: Lenard S
authors: LS
authors: Vranckx
authors: Laboratory for Molecular Virology and Drug discovery, KU Leuven, Belgium.
authors: Jonas
authors: J
authors: Demeulemeester
authors: Laboratory for Molecular Virology and Drug discovery, KU Leuven, Belgium.
authors: Zeger
authors: Z
authors: Debyser
authors: Laboratory for Molecular Virology and Drug discovery, KU Leuven, Belgium.|Laboratory for Viral Vector Technology and Gene Therapy, KU Leuven, Belgium.|Leuven Viral Vector Core, KU Leuven, Belgium.
authors: Rik
authors: http://orcid.org/0000-0003-0191-3904
authors: R
authors: Gijsbers
mesh_terms: D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000378:metabolism; D018014:Gene Transfer Techniques / Q000009:adverse effects; D005822:Genetic Vectors / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics*; D016086:Lentivirus / Q000235:genetics* / Q000502:physiology*; D015202:Protein Engineering; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics*; D014161:Transduction, Genetic; D016662:Virus Integration
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins; C119380:lens epithelium-derived growth factor