A B7-H3-targeted CD28 bispecific antibody enhances the activity of anti-PD1 and CD3 T-cell engager immunotherapies.

T-cell activation is a multistep process requiring T-cell receptor engagement by peptide-major histocompatibility complexes (Signal 1) coupled with CD28-mediated costimulation (Signal 2). Tumors typically lack expression of CD28 ligands, so tumor-specific Signal 1 (e.g.

Translational PK/PD and the first-in-human dose selection of a PD1/IL15: an engineered recombinant targeted cytokine for cancer immunotherapy.

Introduction: Interleukin 15 (IL-15) is a potential anticancer agent and numerous engineered IL-15 agonists are currently under clinical investigation. Selective targeting of IL-15 to specific lymphocytes may enhance therapeutic effects while helping to minimize toxicities.

Methods: We designed and built a heterodimeric targeted cytokine (TaCk) that consists of an anti-programmed cell death 1 receptor antibody (anti-PD-1) and an engineered IL-15.

IL-15/IL-15Rα-Fc-Fusion Protein XmAb24306 Potentiates Activity of CD3 Bispecific Antibodies through Enhancing T-Cell Expansion.

An insufficient quantity of functional T cells is a likely factor limiting the clinical activity of T-cell bispecific antibodies, especially in solid tumor indications. We hypothesized that XmAb24306 (efbalropendekin alfa), a lymphoproliferative interleukin (IL)-15/IL-15 receptor α (IL-15Rα) Fc-fusion protein, may potentiate the activity of T-cell dependent (TDB) antibodies. The activation of human peripheral T cells by cevostamab, an anti-FcRH5/CD3 TDB, or anti-HER2/CD3 TDB resulted in the upregulation of the IL-2/15Rβ (CD122) receptor subunit in nearly all CD8+ and majority of CD4+ T cells, suggesting that TDB treatment may sensitize T cells to IL-15.

Using quantitative single molecule localization microscopy to optimize multivalent HER2-targeting ligands.

Introduction: The progression-free survival of patients with HER2-positive metastatic breast cancer is significantly extended by a combination of two monoclonal antibodies, trastuzumab and pertuzumab, which target independent epitopes of the extracellular domain of HER2. The improved efficacy of the combination over individual antibody therapies targeting HER2 is still being investigated, and several molecular mechanisms may be in play: the combination downregulates HER2, improves antibody-dependent cell mediated cytotoxicity, and/or affects the organization of surface-expressed antigens, which may attenuate downstream signaling.

Methods: By combining protein engineering and quantitative single molecule localization microscopy (qSMLM), here we both assessed and optimized clustering of HER2 in cultured breast cancer cells.

Meditope-Fab interaction: threading the hole.

Meditope, a cyclic 12-residue peptide, binds to a unique binding side between the light and heavy chains of the cetuximab Fab. In an effort to improve the affinity of the interaction, it was sought to extend the side chain of Arg8 in the meditope, a residue that is accessible from the other side of the meditope binding site, in order to increase the number of interactions. These modifications included an n-butyl and n-octyl extension as well as hydroxyl, amine and carboxyl substitutions.

Engineering a high-affinity peptide binding site into the anti-CEA mAb M5A.

We have previously identified a cyclic peptide called meditope which binds to the central cavity of the Fab portion of cetuximab and shown that this peptide binding site can be grafted, or 'meditope-enabled', onto trastuzumab. This peptide has been shown to act as a hitch for the non-covalent attachment of imaging agents to meditope-enabled antibodies. Herein, we explore the process of grafting this peptide binding site onto M5A, an anti-CEA antibody in clinical trials for cancer diagnostics.

Development of a high affinity, non-covalent biologic to add functionality to Fabs.

Functionalization of monoclonal antibodies (mAbs) requires chemical derivatization and/or genetic manipulation. Inherent in these methods are challenges with protein heterogeneity, stability and solubility. Such perturbations could potentially be avoided by using a high affinity, non-covalent intermediate to bridge the desired functionality to a stable mAb.

Activating mutations of STAT5B and STAT3 in lymphomas derived from γδ-T or NK cells.

Lymphomas arising from NK or γδ-T cells are very aggressive diseases and little is known regarding their pathogenesis. Here we report frequent activating mutations of STAT3 and STAT5B in NK/T-cell lymphomas (n=51), γδ-T-cell lymphomas (n=43) and their cell lines (n=9) through next generation and/or Sanger sequencing. STAT5B N642H is particularly frequent in all forms of γδ-T-cell lymphomas.

Identification and grafting of a unique peptide-binding site in the Fab framework of monoclonal antibodies.

Capitalizing on their extraordinary specificity, monoclonal antibodies (mAbs) have become one of the most reengineered classes of biological molecules. A major goal in many of these engineering efforts is to add new functionality to the parental mAb, including the addition of cytotoxins and imaging agents for medical applications. Herein, we present a unique peptide-binding site within the central cavity of the fragment antigen binding framework region of the chimeric, anti-epidermal growth factor receptor mAb cetuximab.