Coordinating gene expression using RNA destabilizing elements
Inventors
Wang, Benjamin • Zeiner, Gusti • McNally, Krista
Assignees
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Publication Number
US-10675305-B2
Publication Date
2020-06-09
Expiration Date
Abstract
Control Devices are disclosed including RNA destabilizing elements (RDE), and RNA control devices, combined with transgenes, including Chimeric Antigen Receptors (CARs) in eukaryotic cells. RDEs can be combined with RNA control devices to make RDEs that include ligand mediated control. These smart RDEs and other RDEs can be used to optimize expression of transgenes, e.g., CARs, in the eukaryotic cells so that, for example, effector function is optimized. CARs and transgene payloads can also be engineered into eukaryotic cells so that the transgene payload is expressed and delivered at desired times from the eukaryotic cell.
Core Innovation
A primary T-cell comprising a chimeric antigen receptor is obtained together with a heterologous nucleic acid encoding a transgene. The transgene is operably linked to a RNA degradation element (RDE), where the RDE is AU101 (Interferon gamma or IFNg) or AU14 (IL6), and the RDE tagged transgene transcript is regulated by an RDE binding protein that binds to the RDE and regulates expression of the transgene.
The primary T-cell is exposed to a ligand for the chimeric antigen receptor. Binding of the ligand activates the primary T-cell, and the primary T-cell proliferates, and activation changes binding of the RDE by the RDE binding protein.
Transgene expression is increased after activation relative to the level of transgene expression before activation. The framework focuses on controlling the expression of an RDE-linked transgene in CAR T-cells so that ligand-driven activation yields increased expression of the transgene.
Claims Coverage
1 independent claim is identified, centered on CAR T-cell activation coupled to an RDE binding protein regulated transgene transcript. The claim covers ligand-activated CAR signaling that changes RDE binding and increases transgene expression after activation versus before activation.
Rde-linked transgene control in a car t-cell using AU101 or AU14
A primary T-cell comprising a chimeric antigen receptor is used with a heterologous nucleic acid encoding a transgene operably linked to a RNA degradation element (RDE), where the RDE is AU101 (IFNg) or AU14 (IL6), and an RDE binding protein binds to the RDE to regulate expression of the transgene.
Ligand-activated CAR changes RDE binding and increases transgene expression
The primary T-cell is exposed to a ligand for the chimeric antigen receptor, wherein ligand binding activates the primary T-cell and proliferation occurs, and the activation changes binding of the RDE by the RDE binding protein; the transgene is expressed with increased levels of expression after activation compared to before activation.
The independent claim covers controlling transgene expression in CAR T-cells by coupling CAR ligand activation to RDE binding protein regulation of an AU101- or AU14-linked transgene transcript, producing increased transgene expression after activation.
Stated Advantages
Increased levels of transgene expression after activation compared to before activation.
Documented Applications
Not explicitly described in patent.
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