Carmel B. Stober

IL-10 from Regulatory T Cells Determines Vaccine Efficacy in Murine Leishmania major Infection

Leishmaniasis affects 12 million people, but there are no vaccines. Immunological correlates of vaccine efficacy are unclear. Polarized Th1 vs Th2 responses in Leishmania major-infected mice suggested that a shift in balance from IL-4 to IFN-γ was the key to vaccine success. Recently, a role for IL-10 and regulatory T cells in parasite persistence was demonstrated, prompting re-evaluation of vaccine-induced immunity. We compared DNA/modified vaccinia virus Ankara heterologous prime-boost with Leishmania homolog of the receptor for activated C kinase (LACK) or tryparedoxin peroxidase (TRYP). Both induced low IL-4 and high IFN-γ prechallenge. Strikingly, high prechallenge CD4 T cell-derived IL-10 predicted vaccine failure using LACK, whereas low IL-10 predicted protection with TRYP. The ratio of IFN-γ:IL-10 was thus a clear prechallenge indicator of vaccine success. Challenge infection caused further polarization to high IL-10/low IFN-γ with LACK and low IL-10/high IFN-γ with TRYP. Ex vivo quantitative RT-PCR and in vitro depletion and suppression experiments demonstrated that Ag-driven CD4+CD25+ T regulatory 1-like cells were the primary source of IL-10 in LACK-vaccinated mice. Anti-IL-10R treatment in vivo demonstrated that IL-10 was functional in determining vaccine failure, rendering LACK protective in the presence of high IFN-γ/low IL-5 responses.

Slc11a1, Formerly Nramp1, Is Expressed in Dendritic Cells and Influences Major Histocompatibility Complex Class II Expression and Antigen-Presenting Cell Function

ABSTRACT Solute carrier family 11 member a1 (Slc11a1; formerly Nramp1) encodes a late endosomal/lysosomal protein/divalent cation transporter that regulates iron homeostasis in macrophages. During macrophage activation, Slc11a1 has multiple pleiotropic effects on gene regulation and function, including gamma interferon-induced class II expression and antigen-presenting cell function. The wild-type allele at Slc11a1 has been associated with a bias in Th1 cell function in vivo, which is beneficial in resistance to infection against intracellular macrophage pathogens but detrimental in contributing to development of type 1 diabetes. The extent to which this depends on macrophage versus dendritic cell (DC) function is not known. Here we show that Slc11a1 is expressed in late endosomes and/or lysosomes of CD11c+ DCs. DCs from mutant and congenic wild-type mice upregulate interleukin-12 (IL-12) and IL-10 mRNA in response to lipopolysaccharide (LPS) stimulation, but the ratio of IL-10 to IL-12 is higher in unstimulated DCs and DCs stimulated for 15 h with LPS from mutant mice than from wild-type mice. DCs from wild-type mice upregulate major histocompatibility complex class II in response to LPS more efficiently than DCs from mutant mice. Unstimulated DCs from wild-type and mutant mice present ovalbumin (OVA) peptide with an efficiency equivalent to that of an OVA-specific CD4 T-cell line, but DCs from wild-type mice are more efficient at processing and presenting OVA or Leishmania activator of cell kinase (LACK) protein to OVA- and LACK-specific T cells. These data indicate that wild-type Slc11a1 expressed in DCs may play a role both in determining resistance to infectious disease and in susceptibility to autoimmune disease such as type 1 diabetes.

Interleukin-4 (IL-4) and IL-10 Collude in Vaccine Failure for Novel Exacerbatory Antigens in Murine Leishmania major Infection

ABSTRACT Leishmaniasis affects 12 million people but there are no vaccines in routine use. Recently, we used DNA vaccination in a susceptible BALB/c high-dose model of infection to screen 100 novel Leishmania major genes as vaccine candidates. In addition to finding novel protective antigens, we identified several antigens that reproducibly exacerbated disease. Here we examined the immune response to two of these antigens, lmd29 and 584C, that were originally identified in an expressed sequence tag cDNA sequencing project. We show that, in addition to exacerbating disease in susceptible BALB/c mice, these antigens retain a propensity to exacerbate disease in resistant C57BL/6 mice. This ability to exacerbate disease was lost when susceptible BALB/c mice were rendered resistant by disruption of the genes encoding interleukin-4 (IL-4) alone, IL-4/IL-13, or IL-4, IL-5, IL-9, and IL-13. Failure to exacerbate disease was associated with reduced IL-5 and IL-10 production in IL-4 knockout mice. Treatment of lmd29-vaccinated mice with anti-IL-10 receptor antibody prior to challenge infection converted exacerbation in wild-type BALB/c mice into highly significant antigen-specific protection. These studies demonstrate that some highly immunogenic antigens of L. major, while having an intrinsic capacity to exacerbate disease in the context of otherwise T helper 1-promoting DNA vaccine delivery, can be rendered protective by the removal of functional IL-10.

DNA-Salmonella enterica Serovar Typhimurium Primer-Booster Vaccination Biases towards T Helper 1 Responses and Enhances Protection against Leishmania major Infection in Mice

ABSTRACT Successful resolution of infections by intracellular pathogens requires gamma interferon (IFN-γ). DNA vaccines promote T helper 1 (Th1) responses by triggering interleukin-12 (IL-12) release by dendritic cells (DC) through Toll-like receptor 9 (TLR9). In humans TLR9 is restricted to plasmacytoid DC. Here we show that DNA-Salmonella enterica serovar Typhimurium primer-booster vaccination, which provides alternative ligands to bind TLR4 on myeloid DC, strongly biases towards Th1 responses compared to vaccination with DNA alone. This results in higher immunoglobulin G2a (IgG2a) responses compared to IgG1 responses, higher IFN-γ responses compared to IL-10 CD4+-T-cell responses, and enhanced protection against Leishmania major infection in susceptible BALB/c mice.

Cytokine Responses to Novel Antigens in an Indian Population Living in an Area Endemic for Visceral Leishmaniasis

Background There are no effective vaccines for visceral leishmaniasis (VL), a neglected parasitic disease second only to malaria in global mortality. We previously identified 14 protective candidates in a screen of 100 Leishmania antigens as DNA vaccines in mice. Here we employ whole blood assays to evaluate human cytokine responses to 11 of these antigens, in comparison to known defined and crude antigen preparations. Methods Whole blood assays were employed to measure IFN-γ, TNF-α and IL-10 responses to peptide pools of the novel antigens R71, Q51, L37, N52, L302.06, J89, M18, J41, M22, M63, M57, as well as to recombinant proteins of tryparedoxin peroxidase (TRYP), Leishmania homolog of the receptor for activated C kinase (LACK) and to crude soluble Leishmania antigen (SLA), in Indian patients with active (n = 8) or cured (n = 16) VL, and in modified Quantiferon positive (EHC+ve, n = 20) or modified Quantiferon negative (EHC−ve, n = 9) endemic healthy controls (EHC). Results Active VL, cured VL and EHC+ve groups showed elevated SLA-specific IFN-γ, but only active VL patients produced IL-10 and EHC+ve did not make TNF-α. IFN-γ to IL-10 and TNF-α to IL-10 ratios in response to TRYP and LACK antigens were higher in cured VL and EHC+ve exposed individuals compared to active VL. Five of the eleven novel candidates (R71, L37, N52, J41, and M22) elicited IFN-γ and TNF-α, but not IL-10, responses in cured VL (55–87.5% responders) and EHC+ve (40–65% responders) subjects. Conclusions Our results are consistent with an important balance between pro-inflammatory IFNγ and TNFγ cytokine responses and anti-inflammatory IL-10 in determining outcome of VL in India, as highlighted by response to both crude and defined protein antigens. Importantly, cured VL patients and endemic Quantiferon positive individuals recognise 5 novel vaccine candidate antigens, confirming our recent data for L. chagasi in Brazil, and their potential as cross-species vaccine candidates.

Heterologous Priming-Boosting with DNA and Modified Vaccinia Virus Ankara Expressing Tryparedoxin Peroxidase Promotes Long-Term Memory against Leishmania major in Susceptible BALB/c Mice

ABSTRACT Leishmaniasis affects 12 million people, but there are no vaccines in routine clinical use. Th1 polarizing vaccines that elicit long-term protection are required to prevent disease in susceptible populations. We recently showed that heterologous priming-boosting with tryparedoxin peroxidase (TRYP) DNA followed by TRYP-modified vaccinia virus Ankara (TRYP MVA) protected susceptible BALB/c mice from Leishmania major. Here we compared treatment with TRYP DNA with treatment with TRYP DNA/TRYP MVA. We found that equivalent levels of protection during the postvaccination effector phase correlated with equivalent levels of serum immunoglobulin G2a and gamma interferon (IFN-γ) in draining lymph nodes. In contrast, challenge infection during the memory phase revealed that there was enhanced clinical efficacy with TRYP DNA/TRYP MVA. This correlated with higher levels of effector phase splenic IFN-γ, sustained prechallenge levels of memory phase IFN-γ, and a more polarized post-L. major challenge Th1 response compared to the Th2/Treg response. Thus, TRYP DNA/TRYP MVA, but not TRYP DNA alone, provides long-term protection against murine leishmaniasis.

Cytokine Responses to Novel Antigens in a Peri-Urban Population in Brazil Exposed to Leishmania infantum chagasi

Visceral leishmaniasis (VL) is fatal if untreated, and there are no vaccines for this disease. High levels of CD4-derived interferon-γ (IFN-γ) in the presence of low levels of interleukin-10 (IL-10) predicts vaccine success. Tumor necrosis factor-α (TNF-α) is also important in this process. We characterized human immune responses in three groups exposed to Leishmania infantum chagasi in Brazil: 1) drug-cured VL patients (recovered VL); 2) asymptomatic persons with positive Leishmania-specific delayed-type hypersensitivity skin reactions (DTH+); and 3) DTH-negative household contacts. Magnitude of DTH correlated with crude Leishmania antigen-driven IFN-γ, TNF-α, and IL-5, but not IL-10. DTH+ persons showed equivalent levels of IFN-γ, but higher levels of IL-10, to tryparedoxin peroxidase and Leishmania homolog of receptor for activated C kinase compared with recovered VL patients. The IFN-γ:IL-10 and TNF-α:IL-10 ratios were higher in recovered VL patients than in DTH+ persons. Seven of 11 novel candidates (R71, L37, N52, L302.06, M18, J41, and M22) elicited cytokine responses (36-71% of responders) in recovered VL patients and DTH+ persons. This result confirmed their putative status as cross-species vaccine/immunotherapeutic candidates.

Proceedings of the 2017 GRAPPA Collaborative Research Network Meeting

The Group for Research and Assessment of Psoriasis and Psoriatic Arthritis (GRAPPA) Collaborative Research Network (CRN) is an endeavor that aims to address gaps in the knowledge of the etiopathogenesis and management of psoriatic disease by best using the large community of experienced investigators who are already collecting rich clinical phenotype data and biologic samples using validated techniques. Exemplar rheumatology and dermatology projects will inform strategies to implement the CRN, while input and funding from government organizations, charities, and industry will shape the CRN. The key immediate priorities to establish the CRN are discussed herein and include (1) strategies for building infrastructure to collect and store biosamples and associated clinical data, (2) best practices for sample collection and storage, (3) approaches to engage the GRAPPA community of investigators and industry to collaborate most effectively on shared priorities, and (4) agreement on a funding strategy. The following 4 CRN candidate flagship research areas were identified: (1) predictors of treatment response in psoriatic arthritis (PsA) and cutaneous psoriasis (PsC) to permit personalized and stratified medicine approaches; (2) predictors of structural damage and disease severity, linking with the existing PsA BioDAM project; (3) predictors of PsC progressing to PsA to enable earlier intervention and possibly halt progression to PsA; and (4) comorbidity prevalence and effect on clinical outcomes in psoriatic disease. The collaboration and momentum provided by a GRAPPA-CRN will offer more than the sum of its individual contributing centers. A CRN will permit high-quality research that can more effectively address questions pertinent to patients, clinicians, scientists, industry, and governments.

Prevalence and predictors of tumour necrosis factor inhibitor persistence in psoriatic arthritis

Objectives To evaluate TNF-α inhibitor (TNFi) persistence when used as first- or second-line biologic therapy for the management of PsA, and to determine baseline clinical and laboratory parameters associated with TNFi persistence. Methods A retrospective single-centre cohort study was performed on all patients with PsA initiated on TNFi therapy between 2003 and 2015. Demographic, clinical and laboratory characteristics were compared with TNFi persistence, using Kaplan-Meier survival and Cox proportional hazards models. Results One hundred and eighty-eight patients with PsA were prescribed TNFi therapy as first-line biologic therapy over a period of 635 person-years [46% male, mean (s.d.) age 47.3 (11.4) years; median (interquartile range) disease duration 11 (7-16) years]. At 12 months of follow-up 79% of patients persisted with TNFi therapy, and 73% at 24 months. Of those discontinuing TNFi, 35% stopped due to primary inefficacy, 22% secondary inefficacy and 43% adverse events. Multivariable analysis identified female sex (hazard ratio (HR) 2.57; 95% CI: 1.26, 5.24; P = 0.01) and the presence of metabolic syndrome-related co-morbidities (HR = 2.65, 95% CI: 1.24, 5.69; P = 0.01) as predictors of lower persistence. Of 32 cases treated with a second TNFi, persistence at 12 months was 56%. TNFi persistence was 2-fold less likely in these 32 cases compared with first-line TNFi users (HR = 2.02, 95% CI: 1.20, 3.42; P = 0.01). Conclusion Patients with PsA who are female and have metabolic syndrome-related co-morbidities have lower TNFi persistence. Although persistence was lower in patients who had switched to a second TNFi, a substantial proportion of these cases responded, advocating switching to a second TNFi as a valid therapeutic strategy.

GRAPPA Trainees Symposium 2016: A Report from the GRAPPA 2016 Annual Meeting

At the 2016 annual meeting of the Group for Research and Assessment of Psoriasis and Psoriatic Arthritis (GRAPPA) in Miami, Florida, USA, a trainees symposium was held. Rheumatology and dermatology trainees, engaged in psoriasis or psoriatic arthritis research, presented their work. This report briefly reviews 6 oral presentations and 21 posters presented at that meeting.