Proteins binding HER2, NKG2D and CD16
Inventors
Chang, Gregory P. • Cheung, Ann F. • Haney, William • LUNDE, Bradley M. • Prinz, Bianka
Assignees
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Publication Number
US-11884732-B2
Publication Date
2024-01-30
Expiration Date
Abstract
Multi-specific binding proteins that bind HER2, the NKG2D receptor, and CD 16 are described, as well as pharmaceutical compositions and therapeutic methods useful for the treatment of cancer.
Core Innovation
The invention provides multispecific TriNKET proteins as polypeptides that bind NKG2D and HER2 using two antigen-binding sites. A first antigen-binding site binds NKG2D, and a second antigen-binding site binds HER2 on cancer cells. The proteins further include an Fc-mediated component configured to bind CD16, thereby enabling enhanced NK activation and tumor killing against HER2+ targets.
The TriNKET design combines an NKG2D-binding variable-domain set and a HER2-binding variable-domain set within an IgG1 Fc-based architecture. The document describes protein formats including heterodimeric IgG-like constructs and Fc-engineered heterodimeric forms such as Triomab, KiH, DVD-Ig, Ortho-Fab, and other Fc-engineered variants. The Fc domains include hinge/CH2 and are configured for CD16 binding, while different amino acid mutations are included on the two human IgG1 Fc domains to promote heterodimerization.
The disclosed HER2-binding and NKG2D-binding components are tied to defined sequence identities in the variable domains and/or CDRs, including sequences identified by SEQ ID NOs. The Fc heterodimerization mutations are described using Kabat EU indexing and include named mutation positions and mutation sets. The document reports that these multispecific proteins enhance NKG2D and NK-cell activation and increase cytotoxicity against HER2+ cancer cell targets compared with trastuzumab or SC2.2, including improved performance for medium/low HER2 expression.
Claims Coverage
The partial claim set centers on one independent claim covering a multispecific protein with defined NKG2D and HER2 binding sites and a dual human IgG1 Fc architecture that promotes heterodimerization while enabling CD16 binding. The remaining provided claim excerpts relate to dependent refinements that further specify Fc sub-regions, heterodimerization mutation positions, and the HER2-binding CDR or variable-domain sequence identities.
NKG2D and HER2 dual antigen-binding sites with CD16-binding heterodimeric Fc
A protein comprising a first antigen-binding site that binds NKG2D, a second antigen-binding site that binds HER2, and a first antibody Fc domain of human IgG1 or a portion thereof and a second antibody Fc domain of human IgG1 or a portion thereof that together are sufficient to bind CD16, wherein the first antibody Fc domain or the portion thereof and the second antibody Fc domain or the portion thereof comprise different amino acid mutations to promote heterodimerization.
Defined NKG2D antigen-binding variable-domain sequences
The first antigen-binding site comprises a heavy chain variable domain comprising the amino acid sequence of SEQ ID NO:1 and a light chain variable domain comprising the amino acid sequence of SEQ ID NO:2.
Hinge and CH2 Fc sub-regions
The first and second antibody Fc domains or portions each comprise hinge and CH2 domains.
Heterodimerization-promoting Fc mutation positions using Kabat EU index
The first antibody Fc domain or the portion thereof and/or the second antibody Fc domain or the portion thereof include one or more amino-acid changes at Kabat EU-indexed positions Q347, Y349, T350, L351, S354, E356, E357, K360, Q362, S364, T366, L368, K370, N390, K392, T394, D399, S400, D401, F405, Y407, K409, T411, and/or K439 to promote heterodimerization.
HER2 CDR sequence identities
The second antigen-binding site includes heavy chain CDR1/2/3 and light chain CDR1/2/3 sequences that are each identical to SEQ ID NOs 83–88.
HER2 variable-domain sequence identity thresholds
The second antigen-binding site includes a heavy chain variable domain at least 90% identical to SEQ ID NO:59 and a light chain variable domain at least 90% identical to SEQ ID NO:60.
Overall, the claim coverage requires a protein with dual antigen-binding sites for NKG2D and HER2, a dual human IgG1 Fc configuration sufficient for CD16 binding, heterodimerization-promoting different Fc mutations, and specific NKG2D variable-domain sequences. The dependent claim excerpts further constrain Fc sub-regions, enumerate Kabat EU-indexed Fc mutation positions, and define HER2 binding via exact CDR sequence identities and/or ≥90% identity thresholds for HER2 variable domains.
Stated Advantages
Enhanced NK activation and tumor killing via CD16-mediated engagement in addition to NKG2D and HER2 binding.
Enhanced NKG2D and NK-cell activation, including TNFα, CD107a, and IFNγ responses.
Increased cytotoxicity against HER2+ targets versus trastuzumab/SC2.2.
Improved efficacy for medium/low HER2 expression.
Improved long-term PBMC cytotoxicity.
Safety indicators via reduced killing of normal B cells while sparing myeloid cells.
Documented Applications
Therapeutic cancer treatment indicated for cancers including breast, ovarian, gastric, and other listed cancer treatment indications.
Combination-therapy context is described alongside the therapeutic use of the multispecific TriNKET proteins.
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