AUTHOR=Zhang Qing , Lu Xiong , Gao Liang , Tao Siyu , Ge Yinghua , Cui Daocheng , Zhu Renying , Lu Wenshu , Wang Jian , Jiang Shisong 

TITLE=In Vitro and In Vivo Antigen Presentation and Diagnosis Development of Recombinant Overlapping Peptides Corresponding to Mtb ESAT-6/CFP-10

JOURNAL=Frontiers in Immunology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2022.872676

DOI=10.3389/fimmu.2022.872676

ISSN=1664-3224

ABSTRACT=<p>Cellular immunity in <italic>Mycobacteria tuberculosis</italic> (<italic>Mtb</italic>) infection is important for the pathogenesis and final clearance of intracellular <italic>Mtb</italic> infection. In addition, it is valuable for the diagnosis of tuberculosis. In this pioneering work, we tested <italic>in vitro</italic> and <italic>in vivo</italic> antigen presentation and diagnostic application of a recombinant overlapping peptide-protein derived from two <italic>Mtb</italic> RD1 antigens ESAT-6 and CFP-10 (ROP-TB). The overlapping peptide sequence of ROP-TB is cleaved by the cathepsin S enzyme and covers the entire length of the two proteins. ROP-TB can be expressed and purified from <italic>E. coli</italic>. Once taken in by antigen-presenting cells, ROP-TB can be cleaved into a peptide pool by cathepsin S within the cells. We found that in dendritic cells, ROP-TB can be processed in 6 hours of co-culture, while the ESAT-6/CFP-10 fusion protein remained in the endosomal compartment. In <italic>Mtb</italic>-infected mice, ROP-TB stimulated stronger specific T cell responses than pooled synthetic peptides derived from ESAT-6 and CFP-10. With regard to the presentation of <italic>in vivo</italic> antigens, in a guinea pig model infected with <italic>Mtb</italic>, ROP-TB induced delayed type hypersensitivity (DTH) responses comparable to those of the tuberculin purified protein derivative (PPD) and ESAT-6/CFP-10 fusion protein. In <italic>Mycobacterium bovis (Bovine TB)</italic>-infected cattle, ROP-TB elicited DTH responses. Finally, in <italic>Mtb</italic> infected patients, ROP-TB stimulated cellular immune responses in majority of patients (16/18) of different HLA phenotypes while a single peptide derived from the same proteins did not elicit the immune responses in all patients. In summary, <italic>in vitro</italic> and <italic>in vivo</italic> data suggest that ROP-TB stimulates a strong cellular immune response irrespective of HLA phenotypes and is therefore suitable for use <italic>in vitro</italic> and <italic>in vivo</italic> diagnostics.</p>