The fact that the mean tumor size of the
The fact that the mean tumor size of the E1202-654 + α-GalCer group was much smaller than that of the α-GalCer inoculated group in both protocols indicates that immunization with E1202-654 plus α-GalCer, which induces a strong CD8+ T cell E1 antigen-specific immune response, is involved in the elimination of B16-F0/E1244-550 cells. It should be noted that the percentage of CD8+ T Cy5.5 NHS ester actively cytotoxic is greater than the percentage of NK cells actively cytotoxic.
The current results could be the basis to propose that E1-specific CD8+ adaptive immune response could play a role in the clearance of HPV infected cells in patients with early HPV-infection, including those with normal cytology or with low-grade cervical intraepithelial lesions, where E1 is necessary for viral replication. Moreover, as the most conserved of early HPV proteins, E1 could potentially induce immunity against various HPV types. This remains to be tested in models inoculated with tumors bearing E1 from other HPV types. It is important to note that the finding that tumor cells transiently expressing E1244-550 antigen were eliminated, strengthens the potential role of E1 as an immunizing antigen for broad-spectrum therapeutic vaccines against HPV. Additionally, α-GalCer served as a potent adjuvant to elicit E1244-550 antigen-specific CD8+ T cell response also inducing a strong innate immune response through NK cells.
With these results, it is now important to predict whether a similar response could be obtained in humans immunized with the same E1 sequences. Thus, we used the same analysis criteria to predict peptides for HLA-A2, which is the largest and most diverse allele family in humans . This revealed that the E1 full-length protein contains 303 highest-scoring predicted peptides and E1244-550 sequence contains 243 (Supplementary Table 2).
Finally, although we did not address CD4 + T cell help in these studies, the E1202-654 fragment is long enough and contains potential epitopes that, together with the adjuvant effect of α-GalCer, should have induced CD4 help to enhance CD8 T cell responses. This will be the subject of further studies. This is an important issue because, depending on the phenotype of the CD4 + T cells induced, they would lead to distinct types of responses, including help or suppression, which could affect the outcome of tumor-infiltrating CTLs.
Conflicts of interest
Introduction Ubiquitin fold modifier 1 (UFM1) is a ubiquitin-like protein that shares the β-grasp fold of ubiquitin, although it has only 16% sequence identity with ubiquitin , , . UFM1 is involved in several cellular processes, including endoplasmic reticulum homeostasis, erythropoiesis, fatty acid metabolism and cell division , , , , , , , . Moreover, protein modification by UFM1 (ufmylation) is associated with breast cancer and neurological disorders , , , , . Notably, prevention of protein modification by UFM1 is embryonic lethal in mice due to severe anemia . Similarly to ubiquitination, ufmylation is mediated through an enzymatic cascade involving E1, E2 and E3 enzymes , , . First, the E1-activating enzyme, UBA5 (ubiquitin-like modifier-activating enzyme 5), binds ATP, magnesium and UFM1 and forms, upon the release of a pyrophosphate, an acyl adenylate intermediate. This intermediate is then subjected to attack by the UBA5 active-site Cys, and with the release of AMP, a high-energy thioester bond with UFM1 C-terminal glycine is formed. At that point, UFM1 is activated and ready to be transferred in a transthioesterification reaction from UBA5 to the active-site Cys of the E2, UFC1 (Ubiquitin-fold modifier-conjugating enzyme 1). Finally, in the presence of the E3, UFL1 (ubiquitin-fold modifier-protein ligase 1), UFM1 is transferred from UFC1 to a lysine residue on a target protein, forming an isopeptide bond , , , .