In the stroma subjacent to persistent CIN2/3 from subjects in the unvaccinated cohort, we found down-regulated expression of genes associated with immune cell activation (CXCR3), TH1 polarization such as Tbet (TBX21), and antiviral activity (IFN- and IFN-). 6). Peripheral blood mononuclear cells (PBMCs) were obtained at baseline and at weeks 8, 15, and 19. Rates and severity of injection site reactions during the 30 days after each vaccination and frequency of adverse events (AEs) are summarized in furniture S1 and S2. All reported AEs were mild, and injection site reactions resolved without sequelae or intervention. One of three patients in each of the DDV1 and DDV2 cohorts Deracoxib experienced total histologic regression at time of resection. In the highest dose cohort (DDV3), three of six patients experienced a total histologic response. Table 1 Treatment cohorts. CR, total regression. = 0.0020) (Fig. 2). These infiltrates were Deracoxib localized in foci of residual dysplasia, not in immediately Deracoxib adjacent normal mucosa. Within-subject increases in tissue CD8+ T cells were significantly greater than the increases we have reported previously in unvaccinated subjects followed over the same time frame (= 0.0300, fig. S2) (11). These infiltrates included increased absolute numbers of Tbet+ cells, suggestive of an effector T cell response. Intraepithelial CD8+ infiltrates were associated Deracoxib with histologic features of apoptosis in lesional epithelial cells. In contrast, the intensity of Foxp3+ infiltrates did not switch significantly, resulting in an increased ratio of effector to Foxp3+ cells (= 0.0488). Open in a separate windows Fig. 2 Tissue CD8+ T cell infiltrates in the target lesion increase after vaccination(A) Representative immunohistochemical (IHC) staining for CD8 in lesional tissue before (left column) and after (right column) vaccination (patient 3009). (B) These infiltrates are Tbet+. (C) In contrast, the intensity of Foxp3+ infiltrates does not switch substantially. (D) Bar graphs depicting quantitated CD8+ and Foxp3+ infiltrates, and the ratio of CD8/Foxp3+ cells in epithelium (e) and stroma (s) of CIN3, before and after vaccination, in all study subjects. Data from bar graphs are means of 3 to 10 regions of interest (ROIs) quantitated per tissue compartment per subject. Error bars show SEM. < 0.05, **< 0.01, Wilcoxon signed rank test. Scale bars, 50 m. To explore the association between the intensity of tissue T cell infiltrates and immune responses in the blood, we calculated the Pearson correlations between lesional epithelial and stromal CD8 infiltrates before and after vaccination, and peripheral blood immune responses to HPV16 E6 and E7 at baseline (before vaccination), at 8 weeks (T8), at the time of resection at week 15 (T15), and postoperatively at week 19 (T19). We found a strong association between intraepithelial CD8 infiltrates at baseline (T0) and the magnitude of T cell response to E6 in the blood after vaccination, at week 15 (= 0.742, = 0.0057) and at week 19 (= 0.751, = 0.0049). These comparisons also recognized a strong correlation between the intensity of lesional stromal CD8 infiltrates at baseline and peripheral blood T cell response to E7 at week 19 (= Deracoxib 0.755, = 0.0045). Finally, in subjects who experienced foci of residual disease at week 15, we found that peripheral blood responses to E6 at weeks 15 and 19 correlated with increased intraepithelial CD8 infiltrates compared to baseline (week 15: = 0.788, = 0.0023; week 19: = 0.76, = 0.004). These findings suggest that detectable peripheral blood responses to vaccination in the setting of established preinvasive disease may reflect potentially effective, endogenous priming at the site of the lesion. In vaccinated patients, the cervix is usually infiltrated by activated effector memory T cells with potent effector functions NR4A1 We used circulation cytometry phenotyping to compare the frequencies of T cell subsets isolated from new tissue explants as explained previously (15), in explants from normal cervical mucosa, from unvaccinated HPV16+ prolonged.