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 M. pneumoniae (MPN) to treat lung diseases. We found 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). The designed SC-IL10 molecules increase in comparison with IL-10 WT, the effective expression in MPN four-fold, and the activity in mouse and human cell lines between 10 to 60 times. In vivo, the SC-IL10 molecules expressed in the mouse lung by CV8 showed a powerful anti-inflammatory effect upon Pseudomonas aeruginosa lung infection. This rational design strategy could be used to other molecules with immunomodulatory properties used in bacterial therapy.