An engineered IL-2 reprogrammed for anti-tumor therapy using a semi-synthetic organism

Abstract

The implementation of applied engineering principles to create synthetic biological systems promises to revolutionize medicine, but application of fundamentally redesigned organisms has thus far not impacted practical drug development. Here we utilize an engineered microbial organism with a six-letter semi-synthetic DNA code to generate a library of site-specific, click chemistry compatible amino acid substitutions in the human cytokine IL-2. Targeted covalent modification of IL-2 variants with PEG polymers and screening identifies compounds with distinct IL-2 receptor specificities and improved pharmacological properties. One variant, termed THOR-707, selectively engages the IL-2 receptor beta/gamma complex without engagement of the IL-2 receptor alpha. In mice, administration of THOR-707 results in large-scale activation and amplification of CD8+ T cells and NK cells, without Treg expansion characteristic of IL-2. In syngeneic B16-F10 tumor-bearing mice, THOR-707 enhances drug accumulation in the tumor tissue, stimulates tumor-infiltrating CD8+ T and NK cells, and leads to a dose-dependent reduction of tumor growth. These results support further characterization of the immune modulatory, anti-tumor properties of THOR-707 and represent a fundamental advance in the application of synthetic biology to medicine, leveraging engineered semi-synthetic organisms as cellular factories to facilitate discovery and production of differentiated classes of chemically modified biologics.

Yeast Display Enables Identification of Covalent Single Domain Antibodies Against Botulinum Neurotoxin Light Chain A

“…While there are now several demonstrations of covalent target engagement 35,36,38,39,68 and the improvement of additional biomolecule properties such as potency, selectivity, and stability, 33,42,[95][96][97][98][99][100][101] few studies have sought to extend these strategies to higher throughput approaches. 42 The streamlined characterizations demonstrated here are expected to support the preparation of dozens to hundreds of chemically augmented candidate proteins, all without laborious expression and purification steps.…”

Advancements of Common Gamma-Chain Family Cytokines in Cancer Immunotherapy

“…Next-generation IL-2 candidates have been developed to bypass IL-2Rα binding, a strategy that lowers the risk of toxicity by lowering the therapeutic dose. THOR-707 (SAR444245, Sanofi, Paris, France) is a recombinant IL-2 variant with a polyethylene glycol (PEG) moiety irreversibly bound to a novel amino acid via click chemistry ( 13 ), restraining the binding of the α chain while retaining native binding affinity of the βγ receptor ( 14 ). Functionally, THOR-707 elicits NK and CD8 T-cell-mediated anti-tumor activity while limiting proliferation of Tregs and other lymphoid cells that trigger life-threatening complications ( 15 ).…”

Genetic Code Engineering by Natural and Unnatural Base Pair Systems for the Site-Specific Incorporation of Non-Standard Amino Acids Into Proteins

“…Scientists at Synthorx (founded by Romesberg in 2014, acquired by Sanofi in 2019) used Romesberg’s SSO (YZ3 strain) to successfully identify a suitable PEGylated position (P65) in IL-2 from 10 candidates (K35, R38, T41, F42, K43, Y45, F62, P65, E68, and V69) and addressed the above two issues. These analyses resulted in the development of THOR-707, the IL-2 compound with an unAA at position 65, followed by further modification with a 30 kDa mPEG, which retained the binding ability to IL-2 Rβγ but lacked that to the undesired IL-2 Rαβγ, and showed an extended half-life ( Manandhar et al, 2021 ; Ptacin et al, 2021 ; Romesberg, 2021 ). THOR-707 is currently in a phase I/II study, not only as a monotherapeutic, but also in combination with a checkpoint inhibitor (pembrolizumab or cemiplimab) ( Romesberg, 2021 ).…”