Serum and Salivary IgG and IgA Response After COVID-19 Messenger RNA Vaccination.

Importance: There is still considerable controversy in the literature regarding the capacity of intramuscular messenger RNA (mRNA) vaccination to induce a mucosal immune response. Objective: To compare serum and salivary IgG and IgA levels among mRNA-vaccinated individuals with or without previous SARS-CoV-2 infection. Design, Setting, and Participants: In this cohort study, SARS-CoV-2-naive participants and those with previous infection were consecutively included in the CoviCompare P and CoviCompare M mRNA vaccination trials and followed up to day 180 after vaccination with either the BNT162b2 (Pfizer-BioNTech) vaccine or the mRNA-1273 (Moderna) vaccine at the beginning of the COVID-19 vaccination campaign (from February 19 to June 8, 2021) in France. Data were analyzed from October 25, 2022, to July 13, 2023. Main Outcomes and Measures: An ultrasensitive digital enzyme-linked immunosorbent assay was used for the comparison of SARS-CoV-2 spike-specific serum and salivary IgG and IgA levels. Spike-specific secretory IgA level was also quantified at selected times. Results: A total of 427 individuals were included in 3 groups: participants with SARS-CoV-2 prior to vaccination who received 1 single dose of BNT162b2 (Pfizer-BioNTech) (n = 120) and SARS-CoV-2-naive individuals who received 2 doses of mRNA-1273 (Moderna) (n = 172) or 2 doses of BNT162b2 (Pfizer-BioNTech) (n = 135). The median age was 68 (IQR, 39-75) years, and 228 (53.4%) were men. SARS-CoV-2 spike-specific IgG saliva levels increased after 1 or 2 vaccine injections in individuals with previous infection and SARS-CoV-2-naive individuals. After vaccination, SARS-CoV-2-specific saliva IgA levels, normalized with respect to total IgA levels, were significantly higher in participants with previous infection, as compared with the most responsive mRNA-1273 (Moderna) recipients (median normalized levels, 155 × 10-5 vs 37 × 10-5 at day 29; 107 × 10-5 vs 54 × 10-5 at day 57; and 104 × 10-5 vs 70 × 10-5 at day 180 [P < .001]). In contrast, compared with day 1, spike-specific IgA levels in the BNT162b2-vaccinated SARS-CoV-2-naive group increased only at day 57 (36 × 10-5 vs 49 × 10-5 [P = .01]). Bona fide multimeric secretory IgA levels were significantly higher in individuals with previous infection compared with SARS-CoV-2-naive individuals after 2 antigenic stimulations (median optical density, 0.36 [IQR, 0.16-0.63] vs 0.16 [IQR, 0.10-0.22]; P < .001). Conclusions and Relevance: The findings of this cohort study suggest that mRNA vaccination was associated with mucosal immunity in individuals without prior SARS-CoV-2 infection, but at much lower levels than in previously infected individuals. Further studies are needed to determine the association between specific saliva IgA levels and prevention of infection or transmission.

Incidence of COVID-19 mRNA vaccine symptomatic breakthrough infections during Omicron circulation in adults with or without infection prior to vaccination.

OBJECTIVES: COVID-19 vaccine breakthrough infections were frequently reported during circulation of the Omicron variant. The ANRS|MIE CoviCompareP study investigated these infections in adults vaccinated and boosted with BNT162b2 [Pfizer-BioNTech] and with/without SARS-CoV-2 infection before vaccination. METHODS: In the first half of 2021, healthy adults (aged 18-45, 65-74 and 75 or older) received either one dose of BNT162b2 (n = 120) if they had a documented history of SARS-CoV-2 infection at least five months previously, or two doses (n = 147) if they had no history confirmed by negative serological tests. A first booster dose was administered at least 6 months after the primary vaccination, and a second booster dose, if any, was reported in the database. Neutralizing antibodies (NAbs) against the European (D614G) strain and the Omicron BA.1 variant were assessed up to 28 days after the first booster dose. A case-control analysis was performed for the 252 participants who were followed up in 2022, during the Omicron waves. RESULTS: From January to October 2022, 78/252 (31%) had a documented symptomatic breakthrough infection after full vaccination: 21/117 (18%) in those who had been infected before vaccination vs. 57/135 (42%) in those who had not. In a multivariate logistic regression model, factors associated with a lower risk of breakthrough infection were older age, a higher number of booster doses, and higher levels of Omicron BA.1 NAb titers in adults with infection before vaccination, but not in those without prior infection. CONCLUSION: Our results highlight the need to consider immune markers of protection in association with infection and vaccination history.

Hypothesis of a CD137/Eomes activating axis for effector T cells in HPV oropharyngeal cancers.

Chronic Human Papilloma Virus (HPV) infection is supplanting alcohol and tobacco intoxications as the leading cause of oropharyngeal cancer in developed countries. HPV-related squamous cell carcinomas of the oropharynx (HPV + OSC) present better survival and respond better to radiotherapy and chemotherapy. Regulatory T cells (TREG) are mainly described as immunosuppressive and protumoral in most solid cancers. However, TREG are paradoxically associated with a better prognosis in HPV + OSCs. The transcription factor FoxP3 is the basis for the identification of TREG. Among CD4 + FoxP3 + T cells, some have effector functions. A medical hypothesis is formulated here: the existence of a CD137 (4.1BB)-Eomesodermin (Eomes) activated pathway downstream of TCR-specific activation in a subpopulation of CD4 + FoxP3 + T cells may explain this effector function. Evidence suggest that this axis may exist either in CD4 + FoxP3 + T cells or CD8 + T cells. This pathway could lead T cells to strong antitumor cytotoxic activity in a tumor-specific manner. Furthermore, CD137 is one of the most expected targets for the development of agonist immunotherapies. The identification of CD137 + Eomes + FoxP3+/- T cells could be a key element in the selective activation of the most anti-tumor cells in the HPV + OSC microenvironment.

A Validated LC-MS/MS Method for Performing Belatacept Drug Monitoring in Renal Transplantation.

Belatacept, a CTLA4-Ig, was designed to prevent rejection and graft loss in kidney transplant recipients. This immunotherapy showed a long-term clinical benefit mainly on renal function and better glycemic control but was also associated with a higher number of severe infectious diseases, particularly CMV disease, and lymphoproliferative disease. Therapeutic drug monitoring usually guides the benefit-risk assessment of long-term immunosuppression. In this study, an analytical method by LC-MS/MS was developed in 20 microL of plasma for the belatacept quantification. Intra- and inter-assay precision and accuracy were lower than 20% for the limit of quantification, and 15% for higher concentrations. The method was implemented in our lab and provided data about the inter-variability (N = 108) and intra-variability (N = 33) of belatacept concentrations in kidney transplant recipients with a stable renal function, after conversion from a CNI- to a belatacept-based regimen.

Enteral citrulline supplementation versus placebo on SOFA score on day 7 in mechanically ventilated critically ill patients: the IMMUNOCITRE randomized clinical trial.

BACKGROUND: Restoring plasma arginine levels through enteral administration of L-citrulline in critically ill patients may improve outcomes. We aimed to evaluate whether enteral L-citrulline administration reduced organ dysfunction based on the Sequential Organ Failure Assessment (SOFA) score and affected selected immune parameters in mechanically ventilated medical intensive care unit (ICU) patients. METHODS: A randomized, double-blind, multicenter clinical trial of enteral administration of L-citrulline versus placebo for critically ill adult patients under invasive mechanical ventilation without sepsis or septic shock was conducted in four ICUs in France between September 2016 and February 2019. Patients were randomly assigned to receive enteral L-citrulline (5 g) every 12 h for 5 days or isonitrogenous, isocaloric placebo. The primary outcome was the SOFA score on day 7. Secondary outcomes included SOFA score improvement (defined as a decrease in total SOFA score by 2 points or more between day 1 and day 7), secondary infection acquisition, ICU length of stay, plasma amino acid levels, and immune biomarkers on day 3 and day 7 (HLA-DR expression on monocytes and interleukin-6). RESULTS: Of 120 randomized patients (mean age, 60 ± 17 years; 44 [36.7%] women; ICU stay 10 days [IQR, 7-16]; incidence of secondary infections 25 patients (20.8%)), 60 were allocated to L-citrulline and 60 were allocated to placebo. Overall, there was no significant difference in organ dysfunction as assessed by the SOFA score on day 7 after enrollment (4 [IQR, 2-6] in the L-citrulline group vs. 4 [IQR, 2-7] in the placebo group; Mann‒Whitney U test, p = 0.9). Plasma arginine was significantly increased on day 3 in the treatment group, while immune parameters remained unaffected. CONCLUSION: Among mechanically ventilated ICU patients without sepsis or septic shock, enteral L-citrulline administration did not result in a significant difference in SOFA score on day 7 compared to placebo. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT02864017 (date of registration: 11 August 2016).

A synthetic delivery vector for mucosal vaccination.

The success of mRNA-based vaccines during the Covid-19 pandemic has highlighted the value of this new platform for vaccine development against infectious disease. However, the CD8+ T cell response remains modest with mRNA vaccines, and these do not induce mucosal immunity, which would be needed to prevent viral spread in the healthy population. To address this drawback, we developed a dendritic cell targeting mucosal vaccination vector, the homopentameric STxB. Here, we describe the highly efficient chemical synthesis of the protein, and its in vitro folding. This straightforward preparation led to a synthetic delivery tool whose biophysical and intracellular trafficking characteristics were largely indistinguishable from recombinant STxB. The chemical approach allowed for the generation of new variants with bioorthogonal handles. Selected variants were chemically coupled to several types of antigens derived from the mucosal viruses SARS-CoV-2 and type 16 human papillomavirus. Upon intranasal administration in mice, mucosal immunity, including resident memory CD8+ T cells and IgA antibodies was induced against these antigens. Our study thereby identifies a novel synthetic antigen delivery tool for mucosal vaccination with an unmatched potential to respond to an urgent medical need.

Engineered Synthetic STxB for Enhanced Cytosolic Delivery.

Many molecular targets for cancer therapy are located in the cytosol. Therapeutic macromolecules are generally not able to spontaneously translocate across membranes to reach these cytosolic targets. Therefore a strong need exists for tools that enhance cytosolic delivery. Shiga toxin B-subunit (STxB) is used to deliver therapeutic principles to disease-relevant cells that express its receptor, the glycolipid Gb3. Based on its naturally existing membrane translocation capacity, STxB delivers antigens to the cytosol of Gb3-positive dendritic cells, leading to the induction of CD8+ T cells. Here, we have explored the possibility of further increasing the membrane translocation of STxB to enable other therapeutic applications. For this, our capacity to synthesize STxB chemically was exploited to introduce unnatural amino acids at different positions of the protein. These were then functionalized with hydrophobic entities to locally destabilize endosomal membranes. Intracellular trafficking of these functionalized STxB was measured by confocal microscopy and their cytosolic arrival with a recently developed highly robust, sensitive, and quantitative translocation assay. From different types of hydrophobic moieties that were linked to STxB, the most efficient configuration was determined. STxB translocation was increased by a factor of 2.5, paving the path for new biomedical opportunities.

One-month humoral response following two or three doses of messenger RNA coronavirus disease 2019 vaccines as primary vaccination in specific populations in France: first results from the Agence Nationale Recherche contre le Sida (ANRS)0001S COV-POPART cohort.

OBJECTIVES: We aimed to investigate the 1-month humoral response to two or three doses of a messenger RNA coronavirus disease 2019 (COVID-19) vaccine as a primary vaccination regimen in specific populations compared with that in healthy adults. METHODS: Agence Nationale Recherche contre le Sida (ANRS)0001S-COV-POPART (NCT04824651) is a French nation-wide, multi-centre, prospective, observational cohort study assessing the immune response to COVID-19 vaccines routinely administered to 11 sub-groups of patients with chronic conditions and two control groups. Patients and controls who received at least two vaccine doses and whose results 1 month after the second dose were available were included. The humoral response was assessed 1 month after the first, second and third doses (if applicable) based on the percentage of responders (positive for anti-Spike severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2] IgG antibodies), geometric means of anti-Spike SARS-CoV-2 IgG antibodies (enzyme-linked immunosorbent assay) and proportion of participants with anti-SARS-CoV-2-specific neutralizing antibodies (in vitro neutralization assay for the original SARS-CoV-2 strain). All analyses were centralized. RESULTS: We included 4091 participants in this analysis: 2979 participants from specific sub-populations and 1112 controls. Only 522 (17.5%) participants from the specific populations received three doses as a primary vaccination regimen. Patients living with human immunodeficiency virus, cancer and diabetes had high percentages of responders after two doses, whereas patients with solid organ transplants, allogeneic hematopoietic stem cell transplants and hypogammaglobulinaemia had the lowest percentage of responders (35.9% [95% CI, 29.2-43.0], 57.4% [95% CI, 48.1-66.3] and 77.1% [95% CI, 65.6-86.3], respectively). In those who received the third dose, the percentage of responders reached 54.2% (95% CI, 42.9-65.2) (vs. 32.3% [95% CI, 16.7-51.4] after 2 doses) among those with solid organ transplants and 73.9% (95% CI, 58.9-85.7) (vs. 56.1% [95% CI, 46.2-65.7] after 2 doses) among those with hematopoietic stem cell transplants. Similar results were found with anti-SARS-CoV-2-specific neutralizing antibodies. CONCLUSIONS: A lower humoral response to COVID-19 vaccines was observed in the specific populations compared with that in the controls. The third dose of this vaccine in the primary regimen had a positive effect on the percentages of patients who developed anti-Spike IgG antibodies and specific neutralizing antibodies.

Synthetic Melanin Acts as Efficient Peptide Carrier in Cancer Vaccine Strategy.

We previously reported that a novel peptide vaccine platform, based on synthetic melanin nanoaggregates, triggers strong cytotoxic immune responses and significantly suppresses tumor growth in mice. However, the mechanisms underlying such an efficacy remained poorly described. Herein, we investigated the role of dendritic cells (DCs) in presenting the antigen embedded in the vaccine formulation, as well as the potential stimulatory effect of melanin upon these cells, in vitro by coculture experiments and ELISA/flow cytometry analysis. The vaccine efficiency was evaluated in FLT3-L-/- mice constitutively deficient in DC1, DC2, and pDCs, in Zbtb46DTR chimera mice deficient in DC1 and DC2, and in LangerinDTR mice deficient in dermal DC1 and Langerhans cells. We concluded that DCs, and especially migratory conventional type 1 dendritic cells, seem crucial for mounting the immune response after melanin-based vaccination. We also assessed the protective effect of L-DOPA melanin on peptides from enzymatic digestion, as well as the biodistribution of melanin-peptide nanoaggregates, after subcutaneous injection using [18F]MEL050 PET imaging in mice. L-DOPA melanin proved to act as an efficient carrier for peptides by fully protecting them from enzymatic degradation. L-DOPA melanin did not display any direct stimulatory effects on dendritic cells in vitro. Using PET imaging, we detected melanin-peptide nanoaggregates up to three weeks after subcutaneous injections within the secondary lymphoid tissues, which could explain the sustained immune response observed (up to 4 months) with this vaccine technology.

Neutralizing antibodies against SARS-CoV-2 variants following mRNA booster vaccination in adults older than 65 years.

Immune response induced by COVID-19 vaccine booster against delta and omicron variants was assessed in 65 adults (65-84 years old) early aftesr a first booster dose. An increase in SARS-CoV-2 neutralizing antibodies was shown in individuals not previously infected without evidence of an age-related effect, with lower increase in those infected before a single dose of primary vaccination. Of note, humoral response was observed only starting from the 5th day after the boost.

CXCR6 expressing T cells: Functions and role in the control of tumors.

CXCR6 is a receptor for the chemokine CXCL16, which exists as a membrane or soluble form. CXCR6 is a marker for resident memory T (TRM) cells that plays a role in immunosurveillance through their interaction with epithelial cells. The interaction of CXCR6 with CXCL16 expressed at the membrane of certain subpopulations of intratumor dendritic cells (DC) called DC3, ideally positions these CXCR6+ T cells to receive a proliferation signal from IL-15 also presented by DC3. Mice deficient in cxcr6 or blocking the interaction of CXCR6 with its ligand, experience a poorer control of tumor proliferation by CD8+ T cells, but also by NKT cells especially in the liver. Intranasal vaccination induces CXCL16 production in the lungs and is associated with infiltration by TRM expressing CXCR6, which are then required for the efficacy of anti-tumor vaccination. Therapeutically, the addition of CXCR6 to specific CAR-T cells enhances their intratumoral accumulation and prolongs survival in animal models of pancreatic, ovarian and lung cancer. Finally, CXCR6 is part of immunological signatures that predict response to immunotherapy based on anti-PD-(L)1 in various cancers. In contrast, a protumoral role of CXCR6+T cells has also been reported mainly in Non-alcoholic steatohepatitis (NASH) due to a non-antigen specific mechanism. The targeting and amplification of antigen-specific TRM expressing CXCR6 and its potential use as a biomarker of response to immunotherapy opens new perspectives in cancer treatment.

Plasma CD27, a Surrogate of the Intratumoral CD27-CD70 Interaction, Correlates with Immunotherapy Resistance in Renal Cell Carcinoma.

PURPOSE: CD70 is a costimulatory molecule known to activate CD27-expressing T cells. CD27-CD70 interaction leads to the release of soluble CD27 (sCD27). Clear-cell renal cell carcinoma (ccRCC) expresses the highest levels of CD70 among all solid tumors; however, the clinical consequences of CD70 expression remain unclear. EXPERIMENTAL DESIGN: Tumor tissue from 25 patients with ccRCC was assessed for the expression of CD27 and CD70 in situ using multiplex immunofluorescence. CD27+ T-cell phenotypes in tumors were analyzed by flow cytometry and their gene expression profile were analyzed by single-cell RNA sequencing then confirmed with public data. Baseline sCD27 was measured in 81 patients with renal cell carcinoma (RCC) treated with immunotherapy (35 for training cohort and 46 for validation cohort). RESULTS: In the tumor microenvironment, CD27+ T cells interacted with CD70-expressing tumor cells. Compared with CD27- T cells, CD27+ T cells exhibited an apoptotic and dysfunctional signature. In patients with RCC, the intratumoral CD27-CD70 interaction was significantly correlated with the plasma sCD27 concentration. High sCD27 levels predicted poor overall survival in patients with RCC treated with anti-programmed cell death protein 1 in both the training and validation cohorts but not in patients treated with antiangiogenic therapy. CONCLUSIONS: In conclusion, we demonstrated that sCD27, a surrogate marker of T-cell dysfunction, is a predictive biomarker of resistance to immunotherapy in RCC. Given the frequent expression of CD70 and CD27 in solid tumors, our findings may be extended to other tumors.

Mild dyslipidemia accelerates tumorigenesis through expansion of Ly6Chi monocytes and differentiation to pro-angiogenic myeloid cells.

Cancer and cardiovascular disease (CVD) share common risk factors such as dyslipidemia, obesity and inflammation. However, the role of pro-atherogenic environment and its associated low-grade inflammation in tumor progression remains underexplored. Here we show that feeding C57BL/6J mice with a non-obesogenic high fat high cholesterol diet (HFHCD) for two weeks to induce mild dyslipidemia, increases the pool of circulating Ly6Chi monocytes available for initial melanoma development, in an IL-1ß-dependent manner. Descendants of circulating myeloid cells, which accumulate in the tumor microenvironment of mice under HFHCD, heighten pro-angiogenic and immunosuppressive activities locally. Limiting myeloid cell accumulation or targeting VEGF-A production by myeloid cells decrease HFHCD-induced tumor growth acceleration. Reverting the HFHCD to a chow diet at the time of tumor implantation protects against tumor growth. Together, these data shed light on cross-disease communication between cardiovascular pathologies and cancer.

Vaccination-based immunotherapy to target profibrotic cells in liver and lung.

Fibrosis is the final path of nearly every form of chronic disease, regardless of the pathogenesis. Upon chronic injury, activated, fibrogenic fibroblasts deposit excess extracellular matrix, and severe tissue fibrosis can occur in virtually any organ. However, antifibrotic therapies that target fibrogenic cells, while sparing homeostatic fibroblasts in healthy tissues, are limited. We tested whether specific immunization against endogenous proteins, strongly expressed in fibrogenic cells but highly restricted in quiescent fibroblasts, can elicit an antigen-specific cytotoxic T cell response to ameliorate organ fibrosis. In silico epitope prediction revealed that activation of the genes Adam12 and Gli1 in profibrotic cells and the resulting "self-peptides" can be exploited for T cell vaccines to ablate fibrogenic cells. We demonstrate the efficacy of a vaccination approach to mount CD8+ T cell responses that reduce fibroblasts and fibrosis in the liver and lungs in mice. These results provide proof of principle for vaccination-based immunotherapies to treat fibrosis.

Synergistic effect of combining sunitinib with a peptide-based vaccine in cancer treatment after microenvironment remodeling.

Although it has proven difficult to demonstrate the clinical efficacy of therapeutic vaccination as a monotherapy in advanced cancers, its combination with an immunomodulatory treatment to reduce intra-tumor immunosuppression and improve vaccine efficacy is a very promising strategy. In this context, we are studying the combination of a vaccine composed of peptides of the tumor antigen survivin (SVX vaccine) with the anti-angiogenic agent sunitinib in a colorectal carcinoma model. To this end, we have been focusing on administration scheduling and have highlighted a therapeutic synergy between SVX vaccine and sunitinib when the vaccine was administered at the end of anti-angiogenic treatment. In this setting, a prolonged control of tumor growth associated with an important percentage of complete tumor regression was observed. Studying the remodeling induced by each therapy on the immunological and angiogenic tumor microenvironment over time we observed, during sunitinib treatment, a transient increase in polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) and a decrease in NK cells in the tumor microenvironment. In contrast, after sunitinib treatment was stopped, a decrease in PMN-MDSC populations has been observed in the tumor, associated with an increase in NK cells, pericyte coverage of tumor vessels and CD8+ T cell population and functionality. In conclusion, sunitinib treatment results in the promotion of an immune-favorable tumor microenvironment that can guide the optimal sequence of vaccine and anti-angiogenic combination to reinforce their synergy.