Coordinating gene expression using RNA destabilizing elements

Inventors

Wang, Benjamin • Zeiner, Gusti • McNally, Krista

Assignees

Chimera Bioengineering Inc

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

The invention provides an isolated primary T-cell that includes a receptor that is a chimeric antigen receptor or a T-cell receptor, together with heterologous nucleic acids encoding transgenes. Each transgene is operably linked to an RNA degradation element, and at least a first RNA degradation element and a second RNA degradation element are AU rich elements. The heterologous nucleic acids are transcribed to make transcripts encoding the transgenes operably linked to the corresponding AU rich element RNA degradation elements.

The isolated primary T-cell includes a mechanism in which a glycolytic enzyme with RNA degradation element binding activity binds the AU rich element RNA degradation element and regulates expression of the transgene. In the two-transgene configuration, a first glycolytic enzyme with RNA degradation element binding activity binds the first AU rich element RNA degradation element to regulate expression of the first transgene, and a second glycolytic enzyme with RNA degradation element binding activity binds the second AU rich element RNA degradation element to regulate expression of the second transgene. Receptor activation increases the amount of polypeptide made from the first transgene and the second transgene by reducing the amount of the glycolytic enzymes available to bind the RNA degradation elements.

The document further describes that RNA degradation elements can be used to tune transgene expression in time and in relation to metabolic and redox state, including temporal regulation dependent on receptor activation. It also describes an optional ligand-controlled RNA control device, where RNA degradation element behavior is linked to ligand-mediated control, thereby providing coordinated control of transgene expression. The system is presented as enabling regulation of multiple payloads in a multitransgene manner using separate RNA degradation element-linked transcripts and separate RNA degradation element-binding glycolytic enzyme regulation.

Claims Coverage

The provided claims coverage centers on a dual-transgene isolated primary T-cell in which AU rich element RNA degradation elements are used as regulatory RNA features, and receptor activation increases transgene polypeptide output by diminishing availability of glycolytic enzymes that bind the RNA degradation elements. The excerpt includes one independent claim and dependent claims that narrow payload options and receptor binding targets.

Overall, the claim coverage centers on a dual-transgene isolated primary T-cell in which AU rich element RNA degradation elements are used as regulatory RNA features, and receptor activation increases transgene polypeptide output by diminishing availability of glycolytic enzymes that bind the RNA degradation elements. Dependent claims specify IL-12 and other immune modulators, and define CAR binding targets and cancer cell contexts.

Stated Advantages

Documented Applications

No documented applications found