Bacterial expression of a designed single‐chain IL‐10 prevents severe lung inflammation

Interleukin‐10 (IL‐10) is an anti‐inflammatory cytokine that is active as a swapped domain dimer and is used in bacterial therapy of gut inflammation. IL‐10 can be used as treatment of a wide range of pulmonary diseases. Here we have developed a non‐pathogenic chassis (CV8) of the human lung bacterium Mycoplasma pneumoniae (MPN) to treat lung diseases. We find that IL‐10 expression by MPN has a limited impact on the lung inflammatory response in mice. To solve these issues, we rationally designed a single‐chain IL‐10 (SC‐IL10) with or without surface mutations, using our protein design software (ModelX and FoldX). As compared to the IL‐10 WT, the designed SC‐IL10 molecules increase the effective expression in MPN four‐fold, and the activity in mouse and human cell lines between 10 and 60 times, depending on the cell line. The SC‐IL10 molecules expressed in the mouse lung by CV8 in vivo have a powerful anti‐inflammatory effect on Pseudomonas aeruginosa lung infection. This rational design strategy could be used to other molecules with immunomodulatory properties used in bacterial therapy.