1342 Organ-specific delivery of a mRNA-encoded bispecific T cell engager targeting Glypican-3 in hepatocellular carcinoma

Background

Bispecific T-cell engaging antibodies (BiTEs) have been clinically validated (e.g., blinatumomab, mosunetuzumab) but are mostly limited to hematological cancers. Developing BiTEs for solid tumors faces hurdles such as T cell penetration into tissue and suppressive nature of the tumor microenvironment, etc. This study aims to tackle these challenges by delivering an mRNA-encoded BiTE targeting the hepatocellular carcinoma antigen Glypican-3 (GPC3) directly to liver tissue, thus enhancing the local BiTE concentration and promoting T cell activation and anti-tumor activity.

Methods

The BiTE was engineered by fusing single-chain variable fragments (scFvs) targeting GPC3 and CD3, separated by a flexible peptide linker. The mRNA encoding the BiTE was optimized through codon usage modifications to balance the Codon Adaptation Index (CAI) and Minimal Free Energy (MFE), taking into account the entire mRNA sequence, including untranslated regions (UTRs) and the coding sequence (CDS). The mRNA was encapsulated in a novel lipid nanoparticle (LNP) for efficient liver-tropic delivery, forming the complex named MTS105. BiTE's cell-killing activity was tested using human GPC3-expressing cell lines, Hepa1-6 and Hep3B and human PBMC (T cell Dependent Cell Cytotoxicity, TDCC]). Anti-tumor efficacy was evaluated in liver orthotopic tumor-bearing mouse models (humanized CD3EDG syngeneic and human PBMC reconstituted CDX models). Safety, pharmacokinetics (PK) and biodistribution were assessed in cynomolgus monkeys.

Results

The BiTE demonstrated high affinity (Kd in single-digit nM range) for human/cynomolgus GPC3 and CD3. MTS105-translated BiTE mediated specific killing of GPC3-positive cells, T cell activation and proliferation in vitro and ex vivo. In the two liver orthotopic tumor-bearing mouse models, MTS105 achieved complete tumor regression in a dose-dependent manner at doses as low as 0.3~0.5 µg/mouse, accompanied with intra-tumoral T cell activation. MTS105, at dose levels associated with a significantly lower peripheral BiTE C_max (maximum concentration) and AUC (area under the curve), achieved a better anti-tumor efficacy than the Fc-domain-containing antibody-based BiTE (ERY974). In contrast, MTS105 showed a 100-fold higher BiTE concentration in liver tissue and tumors compared to antibody-based BiTE. This tissue-specific PK profile was confirmed in the cynomolgus monkeys showing a >10 fold liver/serum ratio in BiTE concentration. No adverse effects were observed in mice or monkeys.

Conclusion

The mRNA-encoded BiTE demonstrated optimal tissue/tumor-specific PK, safety and potent anti-tumor activity in preclinical models. MTS105 is currently in early human trials to assess its safety and preliminary efficacy.