Escaping antibody responses to bacteriolytic enzymes Pal and Cpl-1 by epitope scanning and engineering

Harhala, M. A.; Gembara, K.; Rybicka, I.; Kazmierczak, Z.; Miernikiewicz, P.; Majewska, J.; Budziar, W.; Nasulewicz-Goldeman, A.; Nelson, D. C.; Owczarek, B.; Dabrowska, K.

doi:10.1101/2022.10.06.511193

bioRxiv · 10.1101/2022.10.06.511193

Escaping antibody responses to bacteriolytic enzymes Pal and Cpl-1 by epitope scanning and engineering

Harhala, M. A.·Gembara, K.·Rybicka, I.·Kazmierczak, Z.·Miernikiewicz, P.·Majewska, J.·Budziar, W.·Nasulewicz-Goldeman, A.·Nelson, D. C.·Owczarek, B.·Dabrowska, K.

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Abstract

Bacteriolytic enzymes are promising antibacterial agents, but they can cause a typical immune response in vivo. In this study, we used a targeted modification of two antibacterial endolysins, Pal and Cpl-1. We identified the key immunogenic amino-acids, and designed and tested new, bacteriolytic variants with altered immunogenicity. One new variant of Pal (257-259 MKS [->] TFG) demonstrated decreased immunogenicity while a similar mutant (257-259 MKS [->] TFK) demonstrated increased immunogenicity. A third variant (280-282 DKP [->] GGA) demonstrated significantly increased antibacterial activity and it was not cross-neutralized by antibodies induced by the wild-type enzyme. We propose this variant as a new engineered endolysin with increased antibacterial activity that is capable of escaping cross-neutralization by antibodies induced by wild-type Pal. We show that efficient antibacterial enzymes that avoid cross-neutralization by IgG can be developed by epitope scanning, in silico modelling, and substitutions of identified key amino acids with a high rate of success. Importantly, this universal approach can be applied to many proteins beyond endolysins and has the potential for design of numerous biological drugs.

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Harhala, M. A., Gembara, K., Rybicka, I., Kazmierczak, Z., Miernikiewicz, P., Majewska, J., Budziar, W., Nasulewicz-Goldeman, A., Nelson, D. C., Owczarek, B., Dabrowska, K.. 2022-10-10. Escaping antibody responses to bacteriolytic enzymes Pal and Cpl-1 by epitope scanning and engineering. https://doi.org/10.1101/2022.10.06.511193

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