Cloning, Expression and Characterization of a Peptibody to Deplete Myeloid Derived Suppressor Cells in a Murine Mammary Carcinoma Model Publisher Pubmed



Ramezaniali Akbari K¹ ; Khakibakhtiarvand V¹ ; Mahmoudian J² ; Asgarianomran H³ ; Shokri F⁴ ; Hojjatfarsangi M⁵ ; Jedditehrani M² ; Shabani M¹ Show Affiliations
Source: Protein Expression and Purification Published:2022

Abstract

Background: Myeloid derived suppressor cells (MDSCs) are an immature heterogeneous population of myeloid lineage that attenuate the anti-tumor immune responses. Depletion of MDSCs has been shown to improve efficacy of cancer immunotherapeutic approaches. Here, we expressed and characterized a peptibody which had previously been defined by phage display technique capable of recognizing and depleting murine MDSCs. Materials and methods: Using splicing by overlap extension (SOE) PCR, the coding sequence of the MDSC binding peptide and linker were synthesized and then ligated into a home-made expression plasmid containing mouse IgG2a Fc. The peptibody construct was transfected into CHO–K1 cells by lipofectamine 3000 reagent and the resulting fusion protein was purified with protein G column and subsequently characterized by ELISA, SDS-PAGE and immunoblotting. The binding profile of the peptibody to splenic MDSCs and its MDSC depletion ability were then tested by flow cytometry. Results: The purified peptibody appeared as a 70 KDa band in Western blot. It could bind to 98.8% of splenic CD11b+/Gr-1+ MDSCs. In addition, the intratumoral MDSCs were significantly depleted after peptibody treatment compared to their PBS-treated negative control counterparts (P < 0.05). Conclusion: In this study, a peptibody capable of depleting intratumoral MDSCs, was successfully expressed and purified. Our results imply that it could be considered as a potential tool for research on cancer immunotherapy. © 2022