Division of Animal Technology, Laboratories of Animal Technology, Agricultural Technology Research Institute, Taiwan
Received date: October 12, 2016; Accepted date: October 27, 2016; Published date: October 27, 2016
Citation: Chuang CK, Lin TY (2016) PTD-J-X: To Elicit Humoral and Cellular Immunity as Will. Immunother Open Acc 2:129. doi: 10.4172/2471-9552.1000129
Copyright: © 2016 Chuang CK, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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Concept proving papers upon a PTD-J-X antigen presenting system were reported recently. The X antigen segment of the recombinant protein could be simply built up to elicit humoral and/or cellular immunity, specifically and effectively.
It is conventionally pictured that extracellular antigens are engulfed by antigen presenting cells (APCs) via endocytosis. Then the antigens are digested in lysoendosome to produce peptide epitopes which are loaded to class II major histocompatibility complex (MHC-II) before recycling back onto cell membrane. On the other hand, intracellular antigens are processed by proteasome and the peptide epitope products are transported into endoplasmic reticulum where these epitopes are assembled into de novo synthesized MHC-I molecules (Figure 1). Recently, a PTD-J-X antigen presenting model which was composed of a protein transduction domain (PTD), the cell penetrating peptide of HIV Tat protein, a J-domain of Hsp40 and an X polypeptide was introduced. The X polypeptide was an assortment of either Th2 epitopes (MHC-II associating peptides) and B epitopes, which could associate with B cell receptor, or Th1 epitopes (MHC-II associating peptides) plus Tc epitopes MHC-I associating peptides .
Figure 1: The extracellular and intracelluar routes of the PTD-J-X fusion protein, the PTD-J-epitopes fusion proteins can act as extracellular antigens which are processed in APC via endocytosis route to present the Th epitopes on MHC-II molecules. By using the cell penetrating activity of the PTD, the PTD-J-epitopes fusion proteins can act as intracellular antigens, too. Cellular immunity can be elicited with the cooperation between Th1 cells which are activated by MHC-II-epitope complex and Tc cells which are activated by MHC-I-epitope complex. The cellular immunity can be further enhanced by Hsp72 protein which was found to elicit innate immunity by way of TLR2 and 4 and to stimulate IFNγ expression.
At first, the combination of the Th2 epitopes and B epitopes of FMDV VP1 was tested. Humoral immunity was elicited as expected. Hsp72, the inducible member of Hsp70 family encoded by HspA1 , which is able of associating with the J-domain  and stimulating innate immunity through Toll-like receptor 2 and 4 (TLR2/4) could enhance the FMDV VP1 specific IgG titer for more than one order . Besides the Linear B epitopes used in the above instructive case, the head domain (amino acid 361 to 543) of canine adenovirus 1 (CAdv1) capsid fiber protein was examined as a conformational epitope. High neutralization titers (1024 to 4096) against CAdv1 were detected in the canine sera after boost. The membrane penetrating activity of PTD-JDsRed recombinant protein was about 3 folds higher than that of PTD-DsRed. There were not any detectable membrane penetrating activities for the DsRed and J-DsRed controls. Nearly 7% of PTD-JDsRed recombinant protein could be transduced into Huh-7 cells , therefore, it was speculated that PTD-J could be utilized to present antigen inside a cell. An hTERTepi polypeptide which was composed of human telomerase reverse transcriptase (hTERT) Th and Tc epitopes was designed and PTD-J-hTERTepi fusion protein was prepared. Unlike CT26 colon cancer cells which could colonize in congenic BALB/c mice to form toumrs, CT26(hTERT#10), a CT26 cell line constitutivrly expressing full length hTERT, was rejected. Rest T effector cells isolated from the spleen cells BALB/c mice which had been immunized with CT26(hTERT #10) could be reactivated by PTDJ- hTERTepi as demonstrated by the expression of IFNγ. It is interested to note that much more cells were activated to express IFNγ when Hsp72 was supplemented.
Furthermore, mice pre-treated or post-treated with PTD-JhTERTepi could diminish the transplanted CT26 (hTERT#10) tumor more quickly than the untreated control. Hsp72 could enhance this effect significantly in the post-treatment cases . It is curious how could Hsp72 augment both humoral and cellular immunity? Besides hyperthermia, cells were induced to express Hsp72 under stresses, such as infection or tumorigenesis. Hsp72 could be secreted and the extracellular Hsp72 acted as a danger signal  to alarm surrounding cells and induced TLR2/4 mediated inflammation through Siglec receptors . It was also found that Hsp72 could stimulate dendritic cell maturation [9,10] so as to elevate the antigen presenting capability. IFNγ secreted by the effector cells, predominantly by Th cells , could stimulate cancer cells to release Hsp72 . Therefore, it could be imaged that the addition of Hsp72 recombinant protein was involved in the activation and maturation more APCs to augment immunity. Post-translational modifications are the other aspects on the effects of Hsp72. It is worth to mention that methylation of the lysine residue K561 in the lid domain of Hsp72 would change the localization of Hsp72 protein from cytosol to nucleus. The methylated Hsp72 was found associated with chromosomes and interacted with Aurora B kinase to promote cell proliferation [13,14]. Taken together, these studies constructively suggested that the PTD-J-X system is a versatile platform to present recombinant protein antigens for both humoral as well as cellular immunization. And Hsp72 is a fundamental adjuvant for this system.
This work was supported by the grants NSC102-2320-B-059-001- MY3 and NSC 99-2313-B-059-004-MY3 from the Ministry of Science and Technology, Taiwan.