Proteolytically cleavable chimeric polypeptides and methods of use thereof
Inventors
Lim, Wendell A. • Roybal, Kole T. • Williams, Jasper Z.
Assignees
University of California San Diego UCSD
Publication Number
US-12358988-B2
Publication Date
2025-07-15
Expiration Date
2037-08-22
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Abstract
The instant disclosure provides chimeric polypeptides which modulate various cellular processes following a cleavage event induced upon binding of a specific binding member of the polypeptide with its binding partner. Methods of using chimeric polypeptides to modulate cellular functions, including e.g., induction of gene expression, are also provided. Nucleic acids encoding the subject chimeric polypeptides and associated expression cassettes and vectors as well as cells that contain such nucleic acids and/or expression cassettes and vectors are provided. Also provided, are methods of treating a subject using the described components and methods as well as kits for practicing the subject methods.
Core Innovation
The invention provides chimeric polypeptides that modulate cellular processes following a cleavage event triggered upon binding of a specific binding member of the polypeptide with its binding partner. The chimeric polypeptides include an extracellular domain with a specific binding member, a proteolytically cleavable Notch receptor polypeptide with one or more proteolytic cleavage sites, and an intracellular domain comprising a transcriptional activator or repressor. Binding of the specific binding member to a peptide-major histocompatibility complex (peptide-MHC) induces cleavage of the Notch receptor polypeptide, releasing the intracellular domain to modulate cellular functions such as gene expression.
This invention addresses limitations of conventional inducible expression systems, which affect cell behavior globally or require user input for spatial and temporal control. The disclosed chimeric polypeptides enable autonomous cellular reprogramming to detect specific signals, especially intracellular antigens presented in the context of MHC, and transduce these inputs into desired activity outputs. The constructed nucleic acids encoding these chimeric polypeptides and associated expression cassettes, vectors, and genetically modified cells facilitate targeted modulation of cellular activities, including therapeutic applications such as cancer treatment.
Claims Coverage
The patent includes three independent claims covering methods of treatment using host cells expressing specific chimeric polypeptides that activate transcription of therapeutic components upon binding to peptide-MHC or cancer antigens.
Proteolytically cleavable chimeric polypeptide with specific binding to peptide-MHC
A chimeric polypeptide comprising an extracellular domain with a specific binding member that binds a first peptide-MHC comprising a first intracellular cancer antigen; a proteolytically cleavable Notch receptor domain with Lin12 repeats, heterodimerization domain, transmembrane domain, and proteolytic cleavage sites; and an intracellular domain containing a transcriptional activator, whereby binding to the first peptide-MHC induces cleavage and releases the intracellular domain.
Nucleic acid encoding inducible CAR or TCR driven by chimeric polypeptide intracellular domain
A nucleic acid encoding a chimeric antigen receptor (CAR) or engineered T cell receptor (TCR) operably linked to a transcriptional control element responsive to the released intracellular domain, such that binding to the first peptide-MHC induces cleavage of the Notch receptor, intracellular domain release, and expression of the CAR or TCR which specifically binds a second peptide-MHC comprising a second intracellular cancer antigen present in the cancer expressing the first and second antigens.
Method of treatment using host cells expressing chimeric polypeptides targeting cancer via intracellular and surface antigens
Methods of administering to a cancer patient a host cell expressing a chimeric polypeptide with an extracellular domain binding a first cancer antigen on the cancer cell surface, a proteolytically cleavable Notch receptor polypeptide, and an intracellular transcriptional activator; together with a nucleic acid encoding a chimeric bispecific binding member or an anti-Fc CAR or an innate-immune response inducer operably linked to a transcriptional control element responsive to the intracellular domain; wherein binding to the first antigen induces cleavage and expression of the therapeutic polypeptide that targets a second cancer antigen also expressed by the cancer.
The independent claims disclose methods of treatment using host cells expressing chimeric polypeptides possessing a specific binding domain for peptide-MHC or surface cancer antigen, coupled with a proteolytically cleavable Notch receptor and transcriptional activator domain, inducing expression of therapeutic agents such as CARs, TCRs, chimeric bispecific binding members, anti-Fc CARs or innate immune response inducers, for selective targeting of cancers expressing defined intracellular or surface antigens.
Stated Advantages
Allows modulation of cellular processes specifically triggered by binding to peptide-MHC complexes, enabling precise control of gene expression and cellular functions.
Enables autonomous and antigen-specific control of immune cell activity, improving specificity and avoiding off-target effects common in global inducible systems.
Facilitates targeting of intracellular cancer antigens presented by MHC, expanding the range of targetable tumor-associated antigens.
Supports multiplexed antigen recognition and logic-gated responses, allowing engineered cells to respond only to defined antigen combinations, improving therapy safety and selectivity.
Permits spatially restricted expression of therapeutic polypeptides, including CARs, TCRs, antibodies, bispecific binding members, and immune modulators, reducing systemic toxicity and enhancing therapeutic efficacy.
Documented Applications
Treatment of cancers expressing defined intracellular cancer antigens using host cells engineered with chimeric polypeptides targeting peptide-MHC complexes to induce therapeutic receptor expression.
Cancer immunotherapy employing dual-antigen gated circuits where chimeric polypeptides induce expression of CARs or TCRs against second cancer antigens, allowing selective T cell activation and tumor killing.
Localization and controlled expression of bispecific T cell engagers (BiTEs) in T cells using synNotch circuits activated by tumor specific antigen binding.
Induction of local innate immune responses via targeted expression of innate-immune response inducers such as flagellin in engineered immune cells under synNotch control.
Suppression of immune responses in autoimmune disease contexts using engineered cells expressing immune suppression factors (e.g., IL-10, PD-L1) controlled by antigen-specific chimeric polypeptides.
Treatment of heterogeneous tumors using engineered immune cells where priming antigen recognition induces expression of therapeutic molecules targeting additional tumor antigens.
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