Cross species single domain antibodies targeting mesothelin for treating solid tumors
Inventors
Ho, Mitchell • Pastan, Ira H. • Hong, Jessica D. • Li, Nan
Assignees
US Department of Health and Human Services
Publication Number
US-12180296-B2
Publication Date
2024-12-31
Expiration Date
2040-01-02
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Abstract
Camel single-domain monoclonal antibodies that specifically bind human and mouse mesothelin are described. Chimeric antigen receptor (CAR) T cells and antibody conjugates based on the mesothelin-specific antibodies are also described. The disclosed CAR T cells, mesothelin-specific antibodies and conjugates thereof can be used, for example, in the diagnosis or treatment of mesothelin-positive cancers.
Core Innovation
This disclosure concerns camel single-domain monoclonal antibodies that specifically bind both human and mouse mesothelin, and their use, such as in immunotherapy for the treatment of mesothelin-expressing tumors. The invention describes two cross species mesothelin-specific camel single-domain monoclonal antibodies, referred to as A101 and G8, which specifically bind both human and mouse mesothelin with high affinity. Chimeric antigen receptor (CAR) T cells comprised of these antibodies are capable of potently killing mesothelin-positive tumor cells both in vitro and in vivo.
The background outlines that mesothelin is a cell-surface glycoprotein expressed at low levels in normal mesothelial cells but is highly expressed in multiple solid tumors including mesotheliomas, stomach cancer, pancreatic cancer, lung cancer, breast cancer, ovarian cancer, and others. Although various mesothelin-specific antibodies and CAR T cells have been developed, none have demonstrated high binding affinity across species, particularly for both human and mouse mesothelin. There is a need for cross species mesothelin-specific antibodies to enable evaluation of therapeutic safety and efficacy in animal models.
Accordingly, the invention solves this problem by providing camel single-domain monoclonal antibodies (A101 and G8) with high affinity cross-species binding to mesothelin, nucleic acid molecules encoding these antibodies, CARs comprising these antibodies, immunoconjugates, antibody-drug conjugates, multi-specific antibodies, antibody-nanoparticle conjugates and fusion proteins. Methods of treating mesothelin-positive cancers using these antibodies, CARs or conjugates, as well as methods of diagnosing and detecting mesothelin expression using these antibodies, are also provided.
Claims Coverage
The patent claims include one independent claim directed to a single-domain monoclonal antibody specifically binding mesothelin and further claims directed to CARs with the antibody, immunoconjugates, ADCs, multi-specific antibodies, antibody-nanoparticle conjugates, fusion proteins, nucleic acid molecules encoding these constructs, isolated cells expressing the CAR, compositions, and methods of treatment, detection and diagnosis related to mesothelin-positive cancers. Below are the main inventive features extracted from the independent claims.
Single-domain monoclonal antibodies with specific cross-species mesothelin binding
Camel single-domain monoclonal antibodies that specifically bind mesothelin, comprising the complementarity determining region (CDR) 1, CDR2 and CDR3 sequences of SEQ ID NO: 2 or SEQ ID NO: 4, with CDRs defined by Kabat, IMGT, Paratome or combinations thereof.
Chimeric antigen receptors (CARs) comprising the mesothelin-specific single-domain antibodies
CARs including the mesothelin-specific monoclonal antibody, optionally further comprising a hinge region, a transmembrane domain, a co-stimulatory signaling moiety such as 4-1BB, and a CD3ξ signaling domain.
Cells expressing mesothelin-targeted CARs
Isolated cells, such as cytotoxic T lymphocytes (CTL) or natural killer (NK) cells, expressing the mesothelin-specific CARs.
Immunoconjugates comprising the antibodies and an effector molecule
Immunoconjugates that include a mesothelin-specific monoclonal antibody conjugated to an effector molecule selected from toxins, photon absorbers or detectable labels.
Antibody-drug conjugates (ADCs) with cytotoxic drugs
ADC conjugates comprising the mesothelin-specific antibody and a drug such as an anti-microtubule agent, anti-mitotic agent, or cytotoxic agent for targeted treatment.
Multi-specific antibodies including mesothelin-targeting and T cell or NK cell receptor targeting antibodies
Multi-specific antibodies containing the mesothelin-specific monoclonal antibody and one or more additional antibodies that bind components of the T cell receptor or natural killer cell activating receptor.
Antibody-nanoparticle conjugates
Conjugates of the mesothelin-specific antibodies attached to nanoparticles for targeted delivery.
Fusion proteins combining mesothelin-specific antibodies with heterologous proteins
Fusion proteins comprising the mesothelin-specific monoclonal antibody and a heterologous protein or peptide, such as an Fc protein.
Nucleic acid molecules and vectors encoding the antibodies and CAR constructs
Nucleic acid molecules encoding the mesothelin-specific monoclonal antibodies or CARs, including sequences with SEQ ID NO: 1 or 3, operably linked to promoters, and vectors (e.g., lentiviral vectors) comprising these nucleic acids.
Nucleic acid constructs encoding mesothelin-targeted CARs with huEGFRt
Nucleic acid molecules encoding in 5' to 3' sequence: GMCSFR signal sequence, mesothelin-specific antibody, extracellular hinge, transmembrane domain, intracellular co-stimulatory and signaling domains, self-cleaving 2A peptide, second GMCSFR signal sequence and truncated human epidermal growth factor receptor (huEGFRt).
Methods of treatment of mesothelin-positive cancers
Methods comprising administering the mesothelin-specific antibodies, CARs, immunoconjugates, ADCs, multi-specific antibodies, or antibody-nanoparticle conjugates in therapeutically effective amounts for treating or inhibiting tumor growth or metastasis of mesothelin-positive cancers.
Methods for detecting or diagnosing mesothelin expression and mesothelin-positive cancer
Methods comprising contacting a biological sample with the mesothelin-specific monoclonal antibody, detecting binding, and diagnosing or detecting mesothelin-positive cancers, including direct labeling or secondary antibody detection.
The independent claims cover camel single-domain monoclonal antibodies specifically binding mesothelin with defined CDRs, CAR constructs comprising these antibodies with specific domains, isolated CAR-expressing immune cells, various antibody conjugates (immunoconjugates, ADCs, multispecific antibodies, nanoparticle conjugates, fusions), nucleic acid constructs encoding these components, compositions including them, and methods for treating and diagnosing mesothelin-positive cancers using these molecules and cells.
Stated Advantages
The antibodies and CAR T cells bind mesothelin with high affinity for both human and mouse species, facilitating safety and efficacy evaluation in animal models.
The mesothelin-targeted CAR T cells demonstrate potent killing of mesothelin-positive tumor cells in vitro and exhibit antitumor activity in vivo in mouse models.
Use of camel single-domain antibodies allows generation of effective immunotherapies with potential advantages in tumor targeting and diagnostic imaging due to smaller antibody size.
Documented Applications
Treatment of mesothelin-positive cancers including mesothelioma, prostate cancer, lung cancer, stomach cancer, squamous cell carcinoma, pancreatic cancer, cholangiocarcinoma, breast cancer, and ovarian cancer using the camel single-domain antibodies, CAR T cells, immunoconjugates, ADCs, multi-specific antibodies, or antibody-nanoparticle conjugates.
Diagnostic detection and imaging of mesothelin expression in biological samples or in vivo, including immunoPET and immunohistochemical assays using labeled monoclonal antibodies.
Use of CAR T cells engineered with mesothelin-specific single-domain antibodies for adoptive cell therapy against mesothelin-expressing tumors.
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