CROI 2015 Program and Abstracts

Abstract Listing

Poster Abstracts

immune responses which may eventually lead to autoimmune disease and immunopathology. Activated T-Regs on the other hand, as they limit host immunity they can inhibit pathogen clearance hence facilitating pathogen multiplication and dissemination. However, their role in the regulation of Latent TB infection (LTBI) in household contacts is not yet fully defined. Methods: This was a retrospective cross sectional study. Household contacts of adult smear positive TB patients in Kampala, Uganda were enrolled and investigated for LTBI using Tuberculin Skin Test (TST) and QuantiFERON ®-TB Gold In-Tube (QFN). LTBI was defined as a positive result of both TST and QFN. Cryo-preserved peripheral blood mononuclear cells (PBMCs) were used to determine the number and phenotypic markers of T-Regs among household contacts (HHC) with or without LTBI using multi-color flow cytometry. The difference between the two groups was compared using the Mann Whitney test for non-parametric tests. Results: The study analyzed samples from 18 HHC with LTBI and 22 HHC without LTBI and found that the natural (naïve) T-Regs were increased in the HHC with no LTBI (median 70 cells, (Inter-quartile range (IQR) 25) as compared to HHCs with LTBI (60 cells (IQR) 18), ( P =0.0278). On the other hand, the induced (memory) T-Regs were higher in HHC with LTBI (median 40 cells, (IQR) 18.2) than those without LTBI (median 30 cells, (IQR) 25), ( P =0.0045). In the Phenotypic analysis, the natural T-Regs were CD4 + CD127 low/- CD45RO - while the induced T-Regs were CD4 + CD127 low/- CD45RO + as expected. Conclusions: These results suggest that there is a relationship between T-Regs and TB latency. It is anticipated that the MTB antigenic persistence in the individuals with LTBI induces a continuous generation of T-Regs resulting from the rapid turnover of induced T-Regs increasing their number in this select population. Further studies are needed to access for the function of these raised T-Regs. 818 Antiretroviral Therapy Fails to Restore Mycobacterium Tuberculosis-specific Th1 and Th17 CD4 Responses LyleW. Murray 1 ; Dominique Goedhals 2 ; Iman Satti 1 ; Rodney Phillips 1 ; Helen McShane 1 ; John Frater 1 Phillips/Frater 1 University of Oxford, Oxford, United Kingdom; 2 University of the Free State, Bloemfontein, South Africa Background: HIV-1 is the biggest risk factor for the development of tuberculosis (TB). Antiretroviral therapy (ART) substantially reduces the risk of TB in HIV+ve individuals, but incidence remains greater than for those who are HIV-ve. We hypothesized that this lack of protection results from the failure of key components of Mtb-specific Th1 and Th17 CD4 T cell responses to restore in individuals on ART. Methods: 115 subjects were recruited in a TB-endemic setting in Bloemfontein, South Africa. The Mtb-specific T cell responses of HIV+ve (n=71) subjects were evaluated before initiation of ART, and at 6 and 12 months thereafter. HIV-ve (n=44) subjects were recruited as a comparative group. The expression on CD4 and CD8 T cells of IFN γ , TNF α , IL2 and IL17 in response to Mtb antigens (PPD, ESAT-6/CFP-10 (EC) and α -crystallin (Rv2031c)) was measured using a whole-blood flow cytometry assay. Results: CD4 count, CD4:CD8 ratio and body mass index all increased after 6 and 12 months of ART (P<0.05 for all comparisons). There was no increase in either the proportion of CD4 responders or in the frequency of IFN γ + CD4 cells to EC, PPD or Rv2031c at 6 or 12 months. On ART, there was an increase in the absolute number of IFN γ , TNF α and IL2 expressing CD4 cells responding to EC, PPD and Rv2031c at 6 and 12 months (P<0.05 for all comparisons). However, there was no increase in the number of IL17+ CD4 cells responding to EC and PPD (EC, P=0.35; PPD, P=0.10), although there was to Rv2031c (P<0.0001). When comparing the contribution of IL17+ CD4 cells to the overall cytokine+ CD4 response, this declined by 6 months for PPD and Rv2031c and by 12 months for EC. This failure of restoration was dominant for the Th17 Mtb-specific CD4 responses and was not seen for the Th1 CD4 populations. ART did increase the polyfunctionality of CD4 responses to EC, PPD and Rv2031c, possibly reflecting functional restoration of the Mtb-specific CD4 response (EC 6m, P=0.03; PPD 12m, P=0.01; Rv2013c, P=0.04). However, despite evidence for reconstitution of Mtb-specific CD4 quantity and function, the absolute magnitude of all cytokine+ CD4 responses for EC, PPD and Rv2013c in HIV+ve subjects remained significantly lower after 12 months of ART than HIV-ve subjects (P<0.0001 for all cytokines). Conclusions: Over a 12-month period, ART only partially restored the Mtb-specific CD4 T cell response, with a diminished effect on Th17 responses. This lack of full immune reconstitution may contribute to the observed elevated TB risk despite ART. 819 Treg/Th17 and T-Cell Effector Responses in Tuberculosis Patients Coinfected with HIV Christine Lacabaratz 1 ; AurelieWiedemann 1 ; Celine Manier 1 ; Laure Bourdery 1 ; Mathieu Surenaud 1 ; Jean-Daniel Lelièvre 2 ; Giovanna Melica 3 ;Yves Lévy 2 1 INSERM U955, Université Paris-Est Créteil, Vaccine Research Institute, Créteil, France; 2 INSERM U955, Université Paris-Est Créteil, Vaccine Research Institute, CHU H. Mondor-A.Chenevier, Créteil, France; 3 CHU H. Mondor-A.Chenevier, Créteil, France Background: Whether the balance between Effector and Regulatory T cell (Treg) responses against Mycobacterial Tuberculosis (MTb) may impact the outcome of clinical status of Tuberculosis (TB) infection has not been fully explored. Moreover, how these responses are perturbed and correlate with the clinical status of TB in patients coinfected with HIV is unknown. Methods: Ex vivo frequency and function of Treg (CD25 high CD127 low FoxP3+) and Th1/Th2/Th17 cells characterized by expression of surface markers (CD45RA/CCR4/CCR6/CXCR3) among CD4 T cells, were analyzed from a cohort of TB patients, coinfected or not with HIV (n=11 TB+HIV+; n=25 TB+HIV-) and healthy donors (n=16 HD). MTb-specific responses were detected after stimulation of patients and HD PBMC with PPD and/or ESAT6-CFP10 by intracellular cytokine production (ICS) or secretion (Multiplex assays). Results: As compared to HD, the ex vivo mean ( ± SD) frequency of Treg, Th2 and Th17 was significantly higher in TB patients (6.3 ± 1.6%; 16.4 ± 6.7%; 13.6 ± 3.6%, respectively vs. 4.9 ± 1.2%; 10.0 ± 2.8%; 10.3 ± 2.5% in HD) whereas the frequency of Th1 was lower (19.9 ± 10.4% and 33.6 ± 10.2%, respectively) (P<0.05 for all comparisons). Accordingly, cytokine production by PBMC stimulated with PPD was mainly oriented towards a Treg-, Th2- and Th17-related cytokine pattern (IL-10, IL-27, IL-13, IL-17A and IL-22) in TB patients as compared to HD. This profile was highly perturbed in TB+HIV+ patients who exhibited a significantly lower frequency of Treg (4.9 ± 1.6%), Th2 (5.4 ± 1.1%) and Th17 (7.0 ± 3.2%) as compared to TB+HIV- patients. This was confirmed by a lower secretion of IL-5, IL-9, IL-13, IL-17 in HIV+ TB+ patients (P<0.05). Correlation between these immune responses, HIV status and the presence of sputum bacillary load (SBL+) showed: i) Mono-infected TB+HIV- patients were at a higher frequency SBL- (83%); ii) the production (mean ± SD) of IL-1 β and IL-27 by ESAT6 stimulated PBMC was significantly higher in SBL- patients either coinfected or not with HIV (13138 ± 5913 and 1235 ± 461 pg/ml, respectively) as compared to SBL+ patients (5037 ± 6929 and 534 ± 399) (P<0.05). Conclusions: TB infection, characterized by an increased frequency of peripheral Treg and a MTb response-biased towards Th2/Th17, is impacted in patients coinfected with HIV. This pattern is frequently associated with a SBL+ status. Finally, patients without SBL are characterized by a higher IL-1 β and IL-27 response to TB antigens, which may represent a signature of TB infection control in the pulmonary site.

Poster Abstracts

497

CROI 2015