Antiviral T-cell Research Articles

Broad and Efficient Activation of Memory CD4+ T Cells by Novel HAdV- and HCMV-Derived Peptide Pools.

Reactivation of Human Cytomegalovirus (HCMV) and Human Adenovirus (HAdV) in immunocompromised patients following stem cell transplantation (SCT) or solid organ transplantation (SOT) is associated with high morbidity and mortality. The adoptive transfer of virus-specific CD8+ and CD4+ T cells has been shown to re-establish the antiviral T-cell response and improve clinical outcome. The viral load in immunocompromised patients can efficiently be reduced solely by the infusion of virus-specific CD4+ T cells. The identification of CD4+ T-cell epitopes has mainly focused on a limited number of viral proteins that were characterized as immunodominant. Here, we used in silico prediction to determine promiscuous CD4+ T-cell epitopes from the entire proteomes of HCMV and HAdV. Immunogenicity testing with enzyme-linked immuno spot (ELISpot) assays and intracellular cytokine staining (ICS) revealed numerous novel CD4+ T-cell epitopes derived from a broad spectrum of viral antigens. We identified 17 novel HCMV-derived and seven novel HAdV-derived CD4+ T-cell epitopes that were recognized by > 50% of the assessed peripheral blood mononuclear cell (PBMC) samples. The newly identified epitopes were pooled with previously published, retested epitopes to stimulate virus-specific memory T cells in PBMCs from numerous randomly selected blood donors. Our peptide pools induced strong IFNγ secretion in 46 out of 48 (HCMV) and 31 out of 31 (HAdV) PBMC cultures. In conclusion, we applied an efficient method to screen large viral proteomes for promiscuous CD4+ T-cell epitopes to improve the detection and isolation of virus-specific T cells in a clinical setting.

Abstract 1488: Viral molecular mimicry influences the antitumor immune response in murine and human melanoma

Abstract Molecular mimicry is known to be one of the leading mechanisms by which infectious agents may induce autoimmunity. However, whether a similar mechanism triggers anti-tumor immune response is unexplored, and the role of anti-viral T-cells infiltrating the tumor has remained anecdotal. To address this question, we first developed a bioinformatic tool to identify tumor peptides with high similarity to viral epitopes. Using peptides identified by this tool, we showed that, in mice, viral pre-existing immunity enhanced the efficacy of cancer immunotherapy via molecular mimicry. Specifically, when treated with a cancer vaccine consisting of peptides with a high degree of homology with specific viral peptides, the mice with induced pre-existing immunity to these viral peptides showed significantly better anti-tumor response.To understand whether this mechanism could partly explain immunotherapy-response in humans, we analyzed a cohort of melanoma patients undergoing PD1 treatment with high IgG titer for Cytomegalovirus (CMV). In this cohort of patients, we showed that high level of CMV-antibodies was associated with a prolonged progression free survival, and found that in some cases PBMCs could cross-react with both melanoma and CMV homologous peptides. Finally, T-cell TCR sequencing revealed expansion of the same CD8+ T-cell clones, when PBMCs were pulsed with tumor- or homologous viral peptides.In conclusion, we have demonstrated that pre-existing immunity and molecular mimicry could explain part of the response observed in immunotherapy. Most importantly, we have developed a tool able to identify tumor antigens and neoantigens based on their similarity to pathogen antigens, in order to exploit molecular mimicry and cross-reactive T-cells in cancer vaccine development. Citation Format: Jacopo Chiaro, Henna Kasanen, Thomas Whalley, Cristian Capasso, Sara Feola, Mikaela Grönholm, Firas Hamdan, Karita Peltonen, Micaela Hernberg, Siru Mäkelä, Hanna Karhapää, Paul Brown, Beatriz Martins, Manlio Fusciello, Erkko Ylösmäki, Anna Kreutzman, Satu Mustjoki, Barbara Szomolay, Vincenzo Cerullo. Viral molecular mimicry influences the antitumor immune response in murine and human melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1488.

#50: Cytomegalovirus Retinitis Among Pediatric Hematopoietic Stem Cell and Solid Organ Transplantation Recipients: A Contemporary Review

Abstract Background Cytomegalovirus infection can cause significant morbidity and mortality after transplantation. Cytomegalovirus retinitis (CMVR) can be a major complication of tissue invasive CMV disease in transplant recipients. There are little data regarding the contemporary incidence and outcomes of CMVR among pediatric transplant recipients. Methods We performed a retrospective cohort analysis of allogeneic hematopoietic (allo-HCT) and solid organ transplant (SOT) recipients who received medical care at Nationwide Children’s Hospital (NCH) from 1/1/10–11/30/20 and had a diagnosis of CMVR to describe the incidence, timing post-transplant, ocular manifestations, management, and outcomes. Basic demographic and clinical data, pre-transplant donor/recipient CMV serostatus, and post-transplant CMV infection data were collected. Graft-specific CMV surveillance and prophylaxis strategies were performed according to NCH hospital guidelines. During the study period, quantitative plasma CMV PCR assays included copies/mL (2010–2013) and WHO international standard IU/mL (2013–2020). Results During the 10-year study period, 347 patients underwent allo-HCT (N=214) or SOT (N=133; heart N=46, liver N=36, kidney N= 52; 1 patient had concomitant kidney and liver transplantation); median age was 8.6 years [range 0.1–25.9] and 198 (56.5%) were male. In total, 98 patients had CMV DNAemia post-transplant, and CMVR was diagnosed in 3 (HCT=2, SOT=1), for an overall CMVR incidence of 0.86% among all transplant recipients and occurring in 3% of patients with CMV DNAemia. No CMVR was diagnosed in patients without CMV DNAemia. An ophthalmologic examination was performed in 58 (59%) of patients with any DNAemia compared with 67 (27%) of patients without DNAemia. CMVR was diagnosed on funduscopic examination a median of 183 days [range 34–512] post-transplant based on visual symptomatology (N=2) or by routine eye screening prompted by DNAemia (N=1). CMV DNAemia at the time of CMVR was 1,463 copies/mL, 3,000, and 14,204 IU/mL; all patients had prolonged (>3 months) CMV detection before CMVR. One SOT recipient had concomitant pathology confirming CMV gastrointestinal tract disease. Two of the 3 CMVR cases occurred in the setting of genotypic UL94 CMV resistance mutations (A594V, H520Q)(Figure 1). Two patients were already receiving CMV antiviral therapy (maribavir N=1; valganciclovir=1) at the time of CMVR diagnosis. After the CMVR diagnosis, all patients were prescribed systemic foscarnet. Adjunctive therapies included concurrent intravitreal antiviral therapy (ganciclovir N=1; foscarnet N=1) and adoptive immunotherapy with CMV antiviral specific T-cells (N=1, HCT). Improvement of ocular symptoms occurred in 2 patients at 15 and 20 days after starting targeted CMV antiviral therapy; one patient developed permanently impaired vision. Conclusion CMVR remains as an important complication in transplant recipients. Although CMVR incidence was low in our cohort of transplant recipients, it occurred most frequently in cases of CMV resistance, prolonged DNAemia, and led to permanent visual impairment in 1 of 3 patients. The development of resistant CMV may have a role in the occurrence, progression, and severity of CMVR. Additional pediatric-specific data are needed to better characterize CMVR and inform optimal prevention strategies.

T-cell engager antibodies enable T cells to control HBV infection and to target HBsAg-positive hepatoma in mice

Current antiviral therapies control but rarely eliminate HBV, leaving chronic HBV carriers at risk of developing hepatocellular carcinoma (HCC). Lacking or dysfunctional virus-specific adaptive immunity prevents control of HBV and allows the virus to persist. Restoring antiviral T-cell immunity could lead to HBV elimination and cure of chronically infected patients. We constructed bispecific T-cell engager antibodies that are designed to induce antiviral immunity through simultaneous binding of HBV envelope proteins (HBVenv) on infected hepatocytes and CD3 or CD28 on T cells. T-cell engager antibodies were employed in co-cultures with healthy donor lymphocytes and HBV-infected target cells. Activation of the T-cell response was determined by detection of pro-inflammatory cytokines, effector function (by cytotoxicity) and antiviral effects. To study invivo efficacy, immune-deficient mice were transplanted with HBVenv-positive and -negative hepatoma cells. The 2T-cell engager antibodies synergistically activated T cells to become polyfunctional effectors that in turn elicited potent antiviral effects by killing infected cells and in addition controlled HBV via non-cytolytic, cytokine-mediated antiviral mechanisms. Invivo in mice, the antibodies attracted T cells specifically to the tumors expressing HBVenv resulting in T-cell activation, tumor infiltration and reduction of tumor burden. This study demonstrates that the administration of HBVenv-targeting T-cell engager antibodies facilitates a robust T-cell redirection towards HBV-positive target cells and provides a feasible and promising approach for the treatment of chronic viral hepatitis and HBV-associated HCC. T-cell engager antibodies are an interesting, novel therapeutic tool to restore immunity in patients with chronic hepatitis B. As bispecific antibodies, they bind envelope proteins on the surface of the hepatitis B virus (HBV) and CD3 or CD28 on T cells. This way, they induce a potent antiviral and cytotoxic T-cell response that leads to the elimination of HBV-positive cells. These bispecific T-cell engager antibodies are exciting therapeutic candidates for chronic hepatitis B and HBV-associated hepatocellular carcinoma.

Roles of the Fc Receptor γ-Chain in Inducing Protective Immune Responses after Heterologous Vaccination against Respiratory Syncytial Virus Infection.

The roles of the Fc receptor (FcR) in protection or inflammatory disease after respiratory syncytial virus (RSV) vaccination and infection remain unknown. Virus-like particles containing RSV fusion proteins (RSV F-VLPs) induce T-helper type 1 antibody responses and protection against RSV. Heterologous RSV F-VLP prime and formalin-inactivated RSV (FI-RSV) boost vaccination has been reported to be effective in providing protection without inflammatory disease. Here, we investigated whether the FcRγ-chain is important for immune protection by the heterologous F-VLP and FI-RSV vaccination using FcRγ-chain knockout (−/−) mice. RSV F-VLP-primed and FI-RSV-boosted FcRγ −/− mice displayed less protective efficacy, as shown by higher lung viral titers upon RSV challenge, compared to RSV F-VLP-primed and FI-RSV-boosted immunized wild-type mice. RSV F-VLP and FI-RSV immunization induced lower levels of neutralizing activity and interferon-γ-producing CD8 T-cells in the bronchoalveolar lavage cells of FcRγ −/− mice than in those of wild-type mice. In addition, FcRγ −/− mice displayed a trend of enhancing lung histopathology after RSV vaccination and infection. This study suggests that the FcRγ-chain plays an important role in inducing antiviral protection and CD8 T-cell responses in RSV F-VLP prime and FI-RSV boost vaccination after RSV infections.

Severe acute respiratory syndrome coronavirus 2 for physicians: Molecular characteristics and host immunity (Review).

Recently, severe acute respiratory syndrome (SARS) coronavirus (CoV) 2 (SARS-CoV-2)-causing CoV disease 2019 (COVID-19) emerged in China and has become a global pandemic. SARS-CoV-2 is a novel CoV originating from β-CoVs. Major distinctions in the gene sequences between SARS-CoV and SARS-CoV-2 include the spike gene, open reading frame (ORF) 3b and ORF 8. SARS-CoV-2 infection is initiated when the virus interacts with angiotensin-converting enzyme 2 (ACE2) receptors on host cells. Through this mechanism, the virus infects the alveolar, esophageal epithelial, ileum, colon and other cells on which ACE2 is highly expressed, causing damage to target organs. To date, host innate immunity may be the only identified direct factor associated with viral replication. However, increased ACE2 expression may upregulate the viral load indirectly by increasing the baseline level of infectious virus particles. The peak viral load of SARS-CoV-2 is estimated to occur ~10 days following fever onset, causing patients in the acute stage to be the primary infection source. However, patients in the recovery stage or with occult infections can also be contagious. The host immune response in patients with COVID-19 remains to be elucidated. By studying other SARS and Middle East respiratory syndrome coronaviruses, it is hypothesized that patients with COVID-19 may lack sufficient antiviral T-cell responses, which consequently present with innate immune response disorders. This may to a certain degree explain why this type of CoV triggers severe inflammatory responses and immune damage and its associated complications.

Immune checkpoint inhibitors for progressive multifocal leukoencephalopathy: a new gold standard?

Progressive multifocal leukoencephalopathy (PML) is a very rare and opportunistic encephalitis caused by JC polyomavirus that is linked to profound immunosuppression and is usually fatal unless immune function can be restored. Immune checkpoint inhibitors (ICI) are monoclonal antibodies (mAbs) that block either CTLA-4 or PD-1 inhibitor receptors, thus enhancing antiviral T-cell activity. Successful treatment of PML by ICI has recently generated some enthusiasm in case reports/small series of patients. However, the initial enthusiasm was mitigated by some individual case reports that did not show any benefit. More data are thus warranted about efficacy of immune checkpoint inhibitors in the specific context of PML. We report here the outcomes of six PML patients treated by ICI between 2017 and 2019. Underlying causes of immunosuppression consisted in hematologic malignancies (n = 4), primary immune deficiency (n = 1) and use of immunosuppressive therapies for myasthenia gravis (n = 1). Three patients were alive with a mean follow-up of 21months (14-33) after first ICI infusion, including one patient with frank clinical response, one with stabilization, and one with initial worsening and further stabilization of PML. The three other patients rapidly died from PML. Our data suggest that ICI may be effective for PML treatment but were less impressive than the ones previously reported. Larger studies are thus warranted to confirm this efficacy and to identify the predictive factors of response.

Lipophagy confers a key metabolic advantage that ensures protective CD8A T-cell responses against HIV-1

ABSTRACT Although macroautophagy/autophagy has been proposed as a critical defense mechanism against HIV-1 by targeting viral components for degradation, its contribution as a catabolic process in providing optimal anti-HIV-1 immunity has never been addressed. The failure to restore proper antiviral CD8A/CD8 T-cell immunity, especially against HIV-1, is still the major limitation of current antiretroviral therapies. Consequently, it is of clinical imperative to provide new strategies to enhance the function of HIV-1-specific CD8A T-cells in patients under antiretroviral treatments (ART). Here, we investigated whether targeting autophagy activity could be an optional solution to make this possible. Our data show that, after both polyclonal and HIV-1-specific activation, CD8A T-cells from ART displayed reduced autophagy-dependent degradation of lysosomal contents when compared to naturally HIV-1 protected elite controllers (EC). We further confirmed in EC, by using specific BECN1 gene silencing and lysosomal inhibitors, the critical role of active autophagy in superior CD8A T-cell protection against HIV-1. More importantly, we found that an IL21 treatment was effective in rescuing the antiviral CD8A T-cell immunity from ART in an autophagy-dependent manner. Finally, we established that IL21-dependent rescue occurred due to the enhanced degradation of endogenous lipids via autophagy, referred to as lipophagy, which fueled the cellular rates of mitochondrial beta-oxidation. In summary, our data show that autophagy/lipophagy can be considered as a therapeutic tool to elicit functional antiviral CD8 T-cell responses. Our results also provide additional insights toward the development of improved T-cell-based prevention and cure strategies against HIV-1. Abbreviations: ART: patients under antiretroviral therapy; BaF: bafilomycin A1; BECN1: beclin 1; CEF: cytomegalo-, Epstein-Barr- and flu-virus peptide pool; Chloro.: chloroquine; EC: elite controllers; FAO: fatty acid beta-oxidation; HIVneg: HIV-1-uninfected control donors; IFNG/IFN-γ: interferon gamma; IL21: interleukin 21; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; PBMC: peripheral blood mononuclear cells; SQSTM1: sequestosome 1; ULK1: unc-51 like autophagy activating kinase 1.

Differential and sequential immunomodulatory role of neutrophils and Ly6Chi inflammatory monocytes during antiviral antibody therapy

ABSTRACT Antiviral monoclonal antibodies (mAbs) can generate protective immunity through Fc-FcγRs interactions. We previously showed a role for immune complexes (ICs) in the enhancement of antiviral T-cell responses through FcγR-mediated activation of dendritic cells (DCs). Here we addressed how mAb therapy in retrovirus-infected mice affects the activation of neutrophils and inflammatory monocytes, two FcγR-expressing innate effector cells rapidly recruited to sites of infection. We found that both cell-types activated in vitro by viral ICs secreted chemokines able to recruit monocytes and neutrophils themselves. Moreover, inflammatory cytokines potentiated chemokines and cytokines release by IC-activated cells and induced FcγRIV upregulation. Similarly, infection and mAb-treatment upregulated FcγRIV on neutrophils and inflammatory monocytes and enhanced their cytokines/chemokines secretion. Notably, upon antibody therapy neutrophils and inflammatory monocytes displayed distinct functional activation states and sequentially modulated the antiviral immune response by secreting Th1-type polarizing cytokines and chemokines, which occurred in a FcγRIV-dependent manner. Consistently, FcγRIV- blocking in mAb-treated, infected mice led to reduced immune protection. Our work provides new findings on the immunomodulatory role of neutrophils and monocytes in the enhancement of immune responses upon antiviral mAb therapy.

Phylodynamic Analysis and Implication of HCV Genotype 4 Variability on Antiviral Drug Response and T-Cell Recognition.

Therapies for HCV care could change the prevalence and the geographic distribution of genotypes due to differences in Sustained Virologic Response (SVR). In this scenario, uncommon genotypes/subtypes, such as genotype 4, could spread from high-risk groups, replacing genotypes eradicated by antiviral drugs. Genotype eradication is also strongly influenced by the CD8+ T cell response. In this study, the genetic variability in HCV genotype 4 strains obtained from a cohort of 67 patients naïve to DAA therapy was evaluated. We found that the presence of resistance-associated substitutions (RAS) was able to affect drug responses. Next, using a prediction tool, viral mutations were identified by their ability, or lack thereof, to reduce the binding affinity with HLA, which affects T cell recognition. The Bayesian coalescent analysis suggested two different circulation clusters, one in risk groups (IDUs and MSM) and the other due to migration flows, dated to 1940 and 1915, respectively. Most of the RAS overlapped with HLA and a lack of binding mutations was observed in 96% of strains. This study describes the introduction of HCV genotype 4 in a region of the Mediterranean basin and evaluates how HCV genotype 4’s genetic variability could affect the response of antiviral drugs and CD8+ T cell recognition.

T-cell responses and therapies against SARS-CoV-2 infection.

Coronavirus disease 2019 (COVID‐19) is caused by SARS‐CoV‐2, a novel coronavirus strain. Some studies suggest that COVID‐19 could be an immune‐related disease, and failure of effective immune responses in initial stages of viral infection could contribute to systemic inflammation and tissue damage, leading to worse disease outcomes. T cells can act as a double‐edge sword with both pro‐ and anti‐roles in the progression of COVID‐19. Thus, better understanding of their roles in immune responses to SARS‐CoV‐2 infection is crucial. T cells primarily react to the spike protein on the coronavirus to initiate antiviral immunity; however, T‐cell responses can be suboptimal, impaired or excessive in severe COVID‐19 patients. This review focuses on the multifaceted roles of T cells in COVID‐19 pathogenesis and rationalizes their significance in eliciting appropriate antiviral immune responses in COVID‐19 patients and unexposed individuals. In addition, we summarize the potential therapeutic approaches related to T cells to treat COVID‐19 patients. These include adoptive T‐cell therapies, vaccines activating T‐cell responses, recombinant cytokines, Th1 activators and Th17 blockers, and potential utilization of immune checkpoint inhibitors alone or in combination with anti‐inflammatory drugs to improve antiviral T‐cell responses against SARS‐CoV‐2.

Abstract S03-01: Pan-HLA prediction of SARS-CoV-2 epitopes

Abstract Introduction: Delineating antiviral T-cell responses to SARS-CoV-2 may shed light on the heterogeneity of clinical outcomes and inform vaccine or therapeutic approaches. Viral antigens can be predicted using computational tools that calculate the binding affinity between viral peptides and antigen presentation machinery. However, in order to account for the role of host genetics in the diversity of responses, this analysis must be performed with consideration of the global diversity of the human leukocyte antigen (HLA) proteins responsible for antigen presentation. Methods: We deployed binding predictions across the SARS-CoV-2 peptidome for 9,360 Class I HLA alleles (2,987 HLA-A; 3,707 HLA-B; 2,666 HLA-C; 9-mers) using a consensus approach of 7 algorithms and 3,486 Class II HLA alleles (15-mers) using a consensus approach of 4 algorithms. All pMHC predictions were filtered to include only those with consensus binding less than 500 nM. Results: There were 368,145 unique combinations of peptides and HLA alleles (pMHCs) with a predicted binding affinity of less than 500nM, including 1,103 unique 9-mer and 2,547 15-mer peptides and 1,022 MHC Class I and 8,075 MHC Class II HLA proteins. Of these pMHCs, 82% of 9-mers overlapped with 15-mers, suggesting cross-presentation to both CD4 and CD8 T cells in a subset of individuals. We evaluated this filtered dataset with respect to the population frequency of HLA haplotypes. While the predicted susceptibility of SARS-CoV-2 antigen presentation differed greatly across countries, there was a subset of 21 Class I antigens shared by common HLA types across 30 or more countries (out of 79 countries with reported population frequency data). Our database has been made publicly available, and we have developed a user interface to explore the results based upon viral proteins, HLA alleles, or country populations of interest. Conclusions: With the ongoing SARS-CoV-2 pandemic, there are worldwide efforts to generate a successful vaccine and to evaluate clinical samples to understand the viral pathogenesis and diverse outcomes in patients. This application can serve as a guide to identify responses of putative SARS-CoV-2-specific T cells across patients with a broad range of HLA haplotypes internationally. Citation Format: Katie M. Campbell, Gabriela Steiner, Daniel K. Wells, Antoni Ribas, Anusha Kalbasi. Pan-HLA prediction of SARS-CoV-2 epitopes [abstract]. In: Proceedings of the AACR Virtual Meeting: COVID-19 and Cancer; 2020 Jul 20-22. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(18_Suppl):Abstract nr S03-01.

Review on the coronavirus disease (COVID-19) pandemic: Its outbreak and current status.

BackgroundIn late December 2019 and on 1st January 2020, the coronavirus (COVID‐19) infecting humans was first identified in Wuhan, Hubei Province, China. Later cases have also been confirmed worldwide. Coronaviruses are RNA viruses that are phenotypically and genotypically diverse. Globally, as of 6th April 2020, laboratory confirmed cases of COVID‐19 reported to the World Health Organisation (WHO) amounted to 1 211 214, including 67 666 deaths.AimIn the current study, we performed a literature review on coronavirus outbreak to summarise details about the pathogenesis, epidemiology, diagnosis and the management strategies for the disease control.PathogenesisCoronaviruses are tremendously precise and mature only in differentiated respiratory epithelial cells, as seen in both organ cultures as well as human volunteers. This virus will cause the antiviral T‐cell response to be erratic, owing to the T‐cell apoptosis activation, triggering the immune system to collapse.TransmissionThe understanding of the transmission of COVID‐19 risk is incomplete. The transmission mainly occurs through the respiratory droplets once an infected person sneezes, like the spread of flu and other respiratory infectious agents.Clinical presentationPresentations of COVID‐19 includes fever, cough, shortness of breath, malaise and respiratory distress.TreatmentThere have been no approved vaccines available for COVID‐19 until today. The Ministry of Science and Technology in the People’s Republic of China declared three potential antiviral medicines suitable for treating COVID‐19. Those three medicines are, namely, favilavir, chloroquine phosphate and remdesivir. Hydroxychloroquine combined with azithromycin enhances the reduction of the viral load in COVID‐19 patients.ConclusionThe corona virus transmits quicker than its two predecessors the MERS‐CoV and SARS‐CoV, but has reduced casualty. The global effects of this latest pandemic are still unclear. Nevertheless, considering that so far no vaccine has been available; preventive approaches are the best way to fight against the virus.

Vesicular Stomatitis Virus Encoding a Destabilized Tumor Antigen Improves Activation of Anti-tumor T Cell Responses

Enhancing the immunogenicity of tumor-associated antigens would represent a major advance for anti-tumor vaccination strategies. Here, we investigated structure-directed antigen destabilization as a strategy to improve the degradation, immunogenic epitope presentation, and Tcell activation against a vesicularstomatitis virus (VSV)-encoded tumor antigen. We used the crystal structure of the model antigen ovalbumin to identify charge-disrupting amino acid mutations that were predicted to decrease the stability of the protein. One mutation, OVA-C12R, significantly reduced the half-life of the protein and was preferentially degraded in a 26-S proteasomal-dependent manner. The destabilized ovalbumin protein exhibited enhanced presentation of the major histocompatibility complex (MHC) class I immunogenic epitope, SIINFEKL, on the surface of B16F10 cells or murine bone marrow-derived dendritic cells (BMDCs) invitro. Enhanced presentation correlated with better recognition by cognate CD8 OT-I Tcells as measured by activation, proliferation, and effector cytokine production. Finally, VSV encoding the degradation-prone antigen was better able to prime an antigen ovalbumin-specific CD8 Tcell response invivo without altering the anti-viral CD8 Tcell response. Our studies highlight that not only is the choice of antigen in cancer vaccines of importance, but that emphasis should be placed on modifying antigen quality to ensure optimal priming of anti-tumor responses.

Fibroblast-derived IL-33 is dispensable for lymph node homeostasis but critical for CD8 T-cell responses to acute and chronic viral infection.

Upon viral infection, stressed or damaged cells can release alarmins like IL-33 that act as endogenous danger signals alerting innate and adaptive immune cells. IL-33 coming from nonhematopoietic cells has been identified as important factor triggering the expansion of antiviral CD8+ T cells. In LN the critical cellular source of IL-33 is unknown, as is its potential cell-intrinsic function as a chromatin-associated factor. Using IL-33-GFP reporter mice, we identify fibroblastic reticular cells (FRC) and lymphatic endothelial cells (LEC) as the main IL-33 source. In homeostasis, IL-33 is dispensable as a transcriptional regulator in FRC, indicating it functions mainly as released cytokine. Early during infection with lymphocytic choriomeningitis virus (LCMV) clone 13, both FRC and LEC lose IL-33 protein expression suggesting cytokine release, correlating timewise with IL-33 receptor expression by reactive CD8+ T cells and their greatly augmented expansion in WT versus ll33-/- mice. Using mice lacking IL-33 selectively in FRC versus LEC, we identify FRC as key IL-33 source driving acute and chronic antiviral T-cell responses. Collectively, these findings show that LN T-zone FRC not only regulate the homeostasis of naïve T cells but also their expansion and differentiation several days into an antiviral response.

Host CD8α+ and CD103+ dendritic cells prime transplant antigen-specific CD8+ T cells via cross-dressing.

The participation of dendritic cells (DCs) in CD8+ T-cell-mediated allograft rejection is a long-standing question of great clinical relevance for tissue transplantation. Here, we show that Batf3-/- mice demonstrate delayed allo-skin graft rejection and are deficient in priming allo-specific CD8+ T cells. Batf3-/- mouse priming is restored by transferring either purified CD8α+ or CD103+ DCs, demonstrating the critical role of these cells in alloreactivity. Using Db -restricted antiviral F5 transgenic T-cell receptor T cells, which we demonstrate are alloreactive with H-2Kd , we show that cross-dressing of CD8α+ and CD103+ primes CD8+ T-cell or allo-specific responses in vitro and in vivo. These findings suggest novel strategies for moderating tissue rejection based on interfering with DC cross-dressing or subsequent interaction with T cells.

Moving in for the kill: natural killer cell immunoregulation requires CXCR3

Abstract Natural killer (NK) cells dictate the pathogenic outcomes of infection via both direct killing of virus-infected cells and indirect immunoregulatory killing of antiviral T cells. The latter involves a perforin-dependent mechanism targeting activated CD4 T cells during the first three days of infection. Given that perforin-dependent killing involves cell-cell contact, we speculated that NK cells must re-locate proximal to recently activated T cells within the white pulp (WP) follicles of secondary lymphoid tissue during this critical temporal window of immunoregulatory activity. As expected, virus infection prompted a gradual accumulation of NK cells within T cell rich regions of splenic WP that peaked by the third day of infection. NK cells deficient in the chemokine receptor CXCR3 exhibited impaired localization to the WP and a markedly diminished capacity to suppress antiviral T-cell and germinal center B-cell responses. Our results reveal a critical role for CXCR3 in properly positioning NK cells to prune developing antiviral T cell responses, which potentially explains loss of tolerance and enhanced immune-mediated organ damage in absence of CXCR3. Strategies to curtail localization of NK cells in the WP during immunization may be an effective means to enhance vaccine-elicited immune responses.

Preserved specific anti-viral T-cell response but associated with decreased lupus activity in SLE patients with cytomegalovirus infection

SLE is an autoimmune disease characterized by aberrant autoantibody production and immune dysfunctions. Whether the anti-CMV immunity is impaired in SLE patients is poorly understood. We investigated the specific anti-viral T-cell response in SLE patients with CMV infection and its possible impacts on clinical manifestations in lupus. CD28 null T-cell percentages were measured by flow cytometry in 89 SLE patients and 58 healthy controls. A specific anti-CMV CD8 T-cell response was assessed ex vivo by the production of intracellular cytokines in response to CMV phosphoprotein 65 (pp65) by flow cytometry. Clinical manifestations and immune parameters were analysed in SLE patients according to their CMV serostatus. CD28 null T cells were significantly expanded in SLE patients. When the anti-CMV pp65 CD8 polyfunctional T cell response was analysed, as defined by production of at least three of four functional cytokines or effectors (intracellular IFN-γ, IL-2, TNF-α and surface CD107a), the results demonstrated that it was not impaired in SLE patients. In contrast, when comparing clinical manifestations, there were lower anti-ds-DNA levels and decreased SLEDAI in SLE patients with CMV infection. Furthermore, the expansion of CD4+CD28 null T cells was negatively associated with anti-ds-DNA levels and SLEDAI in these lupus patients. In SLE patients with CMV infection, the specific anti-CMV CD8 T-cell response is preserved but is associated with decreased disease activity and lower anti-DNA levels among these patients, suggesting CMV infection may mitigate lupus activity.

Covid-19 Immune Signatures Reveal Stable Antiviral T-Cell Function Despite Declining Humoral Responses

To investigate the role of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific T-cell immunity and its relationship with antibody levels and pre-existing immunity against endemic human coronaviruses (huCoV) during disease and beyond, we analyzed patients with recovered (RC, n=178) and active Coronavirus Disease-2019 (COVID-19; AC, n=10) and healthy donors (HD, n=58). Overall, ACs had highest SARS-CoV-2 antibody levels against nucleocapsid (N) and spike (S) proteins but reduced antiviral T-cell immunity, whereas in RCs, antibody levels partially correlated with SARS-CoV-2-specific T-cell frequencies. Interestingly, humoral responses declined throughout convalescence, whereas T-cell immunity remained stable. RCs exhibited polyfunctional, mainly IFN-γ-secreting CD4 + effector memory T-cell responses. Humoral and cellular response towards huCoV strains in RCs with strong SARS-CoV-2 T-cell immunity implies a protective role of pre-existing immunity against huCoV. This study provides essential evidence-based data about stable protective T-cell immunity during disease and recovery which is essential to guide diagnosis and treatment of COVID-19.

Type I IFN ineffectively activates neonatal dendritic cells limiting respiratory antiviral T-cell responses.

Insufficient T cell responses contribute to the increased burden of viral respiratory disease in infancy. Neonatal dendritic cells (DCs) often provide defective activation of pathogen-specific T cells through mechanisms that are incompletely understood, which hinders vaccine design for this vulnerable age group. Enhancing our characterization of neonatal DC sub-specialization and function is therefore critical to developing their potential for immunomodulation of T cell responses. In this study, we engineered respiratory syncytial virus (RSV) to express a model protein, ovalbumin, to track antigen-presenting DCs in vivo. We found that murine neonatal conventional DC1s (cDC1s) efficiently migrated and presented RSV-derived antigen, challenging the paradigm that neonatal DCs are globally immature. In a key observation, however, we discovered that during infection neonatal cDC1s presenting viral antigen were unable to upregulate costimulatory molecules in response to type I interferons (IFN-I), contributing to poor antiviral T cell responses. Importantly, we showed that the deficient response to IFN-I was also exhibited by human neonatal cDC1s, independent of infection. These findings reveal a functionally distinct response to IFN-I by neonatal cDC1s that may leave young infants susceptible to viral infections, and provide a new target for exploration, in light of failed efforts to design neonatal RSV vaccines.