Systemic neutralization of proinflammatory cytokines (especially TNFα) is an established strategy in the treatment of inflammatory bowel disease, however it has shown through the last years many unwanted adverse effects such as a high increase in opportunistic infections and, in some cases, of cancer. Within this doctoral dissertation we have developed recombinant lactic acid bacteria with cytokine binders displayed on their surface. We targeted interleukine17 and interleukine 23 due to the involment of Th17 immune response in inflammatory bowel disease pathology. Firstly, we used single chain variable fragments (ScFv) directed against interleukine17 (1733LH) and interleukine 23 (2350LH), which we, later on due to their poor functionality, replaced with interleukine17 binding fynomer (FIN17) and interleukine 23 binding adnectine (ADN23). Genes for binders were cloned into our proprietary surface display plasmid (pSDLBA3b) and expressed in L. lactis NZ9000 strain as fusion proteins with LysM-containing C-terminus of AcmA and Usp45 secretion signal. Additionaly we used an alternative approach in which L. lactis NZ9000 strain’s producer cells were removed, and its fusion protein-containing growth media were used for coating of genetically non-modified Lb. salivarius ATCC11741. To broaden the spectra of cytokine binders we also expressed and then used for coating a recently developed TNF-α binding »Affibody«. All three cytokine binders were then used for coating of Lb. salivarius ATCC11741 simultaneously. Simultaneous display of three different cytokine-binding proteins on the surface of lactic acid bacteria was demonstrated for the first time and suggested as a new potential treatment for inflammatory bowel disease.
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