Chemical generation of small molecule-based bispecific antibody-drug conjugates for broadening the target scope

Antibody-drug conjugates (ADCs) hold great therapeutic promise for cancer indications; however, treating tumors with intratumor heterogeneity remains challenging. We hypothesized that ADCs can simultaneously target two different antigens could address this issue. Here, we report controlled production and evaluation of bispecific chemically functionalized tumor-targeting small molecules. Enzyme-mediated conjugation bi-functional branched linkers following sequential orthogonal click reactions payload tumor targeting modules (folic acid or RGD peptide) afforded homogeneous defined ligand/drug-to-antibody ratios ranging from 4 + to 16 (ligand/payload). Most were stable under physiological conditions 14 days. Functionalization the cancer-specific ligands did not impair cathepsin B-mediated release ADCs. Bispecific folate receptor (FR)/human epidermal growth factor 2 (HER2) demonstrated specific binding high cell killing potency only in cells expressing either antigen (FR HER2). Integrin/HER2 equipped peptides also showed target-specific cytotoxicity integrin- HER2-positive cells. These findings suggest our small-molecule based have potential effectively treat heterogeneous expression.