1032 Preclinical characterization of a highly potent TLR8 agonist antibody conjugate for the treatment of solid tumor

Background Toll-like receptors (TLRs) are an important family of receptors that senses ssRNA fragments from pathogens. TLR8 activation can initiate both innate and adaptive immune responses and reverse Treg and Myeloid-derived suppressor cells (MDSC) mediated immune suppression by tumors. Immune-stimulating antibody conjugates (ISAC) combining tumor-targeting monoclonal antibodies with immunostimulatory agents allow localized delivery of immune activators to the tumor microenvironment. Here, we report the Denovo preclinical candidate DN027262, a novel ISAC comprising a highly potent anti-tumor linker-payload conjugated to an anti-HER2 antibody. The tumor growth was inhibited by TLR mediated activation of myeloid cells and subsequent T-cell-mediated antitumor immunity against HER2-expressing tumors. Epitope spreading and endogenous T cell activation within the tumor can be promoted by cDC1 in the tumor, led to enhanced inhibition of antigen-negative tumor and long-term immunological memory. Methods In vivo efficacy studies were performed in TLR8 humanized Syngeneic models. Safety, immunogenicity, pharmacokinetics, and pharmacodynamics were assessed in single- and repeat-dose studies using cynomolgus monkeys. Results Through structure-based drug design, we discovered a novel selective small molecule TLR8 agonist, which exhibited a balanced potency and hydrophilicity characteristic. A series of drug-linkers were synthesized and conjugated to the HER2-targeting antibodies. In vitro, these ISACs were found to induce high cytokines release in a PMBC assay when co-cultured with the HER2-high expressing cancer cell lines. In vivo, tumor regression was observed when dosing with the DeNovo preclinical candidate DN027262 in hTLR8 syngeneic HER2-expressing tumors models. Tumor is completely eradicated in all groups at a single dose of 5 mg/kg via subcutaneous administration in immunocompetent murine models, with prolonged inhibition effect on tumor growth after drug withdrawal. At lower dose, the TLR8-ISAC performed encouraging synergies when combined with mPd-1 or HER2-ADC (DS8201). The innate and adaptive cytokine and chemokine markers are upregulated, showed beneficial for tumor clearance and immunological memory. Consistently, in vivo non-GLP cynomolgus monkey studies by consecutive treatments with DN027262 resulted in a significant increase of immune system cells including monocytes, cDCs and natural killer (NK) cells, correlated with TLR8 activation. Cmax increases proportionally with ascending dose levels, up to 12mg/kg. All these changes were back to normal after a period of recovery (2–3 weeks). Conclusions DN027262 exhibited a remarkable anti-tumor activity both in vitro and in vivo and provided a favorable safety window, supporting novel therapeutic option for conventional therapy-resistant cancer.