Engineered host cells and methods of use thereof
Inventors
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MiNK TherapeuticsMiNK Therapeutics is a clinical-stage biotechnology company focused on developing allogeneic invariant natural killer T (iNKT) cell therapies for cancer, pulmonary, and immune-mediated diseases. The company advances a platform of off-the-shelf, native and engineered iNKT cell programs capable of targeting tumors and modulating immune responses, with demonstrated clinical evidence in oncology, acute respiratory distress syndrome (ARDS), and pulmonary fibrosis. Its manufacturing platform enables large-scale GMP-compliant production and clinical delivery of iNKT cell therapies without lymphodepletion or HLA matching. MiNK collaborates with leading academic and industry partners to advance novel cell-based immunotherapies.
MiNK Therapeutics is a clinical-stage biotechnology company focused on developing allogeneic invariant natural killer T (iNKT) cell therapies for cancer, pulmonary, and immune-mediated diseases. The company advances a platform of off-the-shelf, native and engineered iNKT cell programs capable of targeting tumors and modulating immune responses, with demonstrated clinical evidence in oncology, acute respiratory distress syndrome (ARDS), and pulmonary fibrosis. Its manufacturing platform enables large-scale GMP-compliant production and clinical delivery of iNKT cell therapies without lymphodepletion or HLA matching. MiNK collaborates with leading academic and industry partners to advance novel cell-based immunotherapies.
Publication Number
US-11898165-B2
Publication Date
2024-02-13
Expiration Date
Abstract
Provided herein are engineered host cells suitable for expression of functional T cell receptors (TCR) and methods of using these cells to identify TCRs that specifically bind to a desired peptide-major histocompatibility complex (MHC) complex.
Core Innovation
The invention provides an isolated cell derived from a mouse lymphocyte. The isolated cell expresses a chimeric CD8 on the cell surface comprising chimeric CD8b1 and chimeric CD8b2, and the extracellular regions are from human CD8 while the transmembrane and cytoplasmic regions are from mouse CD8. The isolated cell expresses CD3 but does not express endogenous TCR on the cell surface.
The isolated cell further comprises one or more reporter system for measurement of TCR signaling. The reporter system includes promoter region(s) and a polynucleotide encoding a detectable marker operably linked to a promoter region, and the measured signaling includes NFAT, IL-2, AP1, and NFKB/Rel signaling, and may include calcium flux as a TCR signaling readout. In a specific example described, the IL-2-(NFAT)3-EGFP reporter is used as a detectable reporter marker for TCR signaling activities.
The disclosure also includes approaches to generate a cellular TCR library and to identify T cell receptors specific for peptideMHC complexes using engineered host cells and screening/selection concepts. Separate polynucleotide encoding TCRb1 and TCRb2 chains can be delivered to create diverse recombinant TCR expression in the engineered mouse-derived lymphocytes, including selection using peptideMHC tetramers/dimers/multimers and/or functional markers such as cytokines and activation markers. The engineered co-receptor presence can be made regulatable or abrogatable using site-specific recombination systems to compare co-receptor-present versus co-receptor-absent states for tetramer binding and signaling.
Claims Coverage
The independent claim covers an isolated mouse lymphocyte-derived cell that expresses chimeric CD8b1 and CD8b2 with human extracellular regions and mouse transmembrane/cytoplasmic regions, expresses CD3 but lacks endogenous TCR on the cell surface, and includes one or more reporter system for measurement of TCR signaling. One claim family member further specifies amino acid sequences for the chimeric CD8 chains.
Chimeric CD8b1 and CD8b2 expression without endogenous TCR
The isolated cell is derived from a mouse lymphocyte and expresses CD3 while not expressing endogenous TCR on the cell surface, and the cell expresses a chimeric CD8b1 chain and a chimeric CD8b2 chain on the cell surface.
Human extracellular / mouse transmembrane-cytoplasmic chimeric CD8 chains
The chimeric CD8b1 chain comprises an extracellular region of human CD8b1 and transmembrane and cytoplasmic regions of mouse CD8b1, and the chimeric CD8b2 chain comprises an extracellular region of human CD8b2 and transmembrane and cytoplasmic regions of mouse CD8b2.
Reporter system for measurement of TCR signaling
The isolated cell comprises one or more reporter system for measurement of TCR signaling.
TCR signaling reporter with NFAT, IL-2, AP1, and NFKB/Rel readouts
The reporter system measures TCR signaling via NFAT, IL-2, AP1, and NFKB/Rel signaling, and includes a promoter region operably linked to a polynucleotide encoding a detectable marker.
Specific chimeric CD8b1 and/or CD8b2 amino acid sequences
The chimeric CD8b1 chain comprises the amino acid sequence of SEQ ID NO: 1, and/or the chimeric CD8b2 chain comprises the amino acid sequence of SEQ ID NO: 2.
Overall claim coverage centers on an isolated mouse lymphocyte-derived cell expressing chimeric human extracellular/mouse transmembrane-cytoplasmic CD8b1 and CD8b2 with CD3 but no endogenous TCR, combined with a TCR-signaling reporter system. The claims further specify NFAT/IL-2/AP1/NFKB/Rel readouts, a detectable marker, and SEQ ID NO: 1 and SEQ ID NO: 2 for the chimeric CD8 chains.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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