Biomedical subjects

Claude Leclerc

Publications and source records attributed to Claude Leclerc.

54 records · Page 3Linked to original sources

PubMed2003

Ontogeny and innate properties of neonatal dendritic cells.

We investigated whether a developmental immaturity of the dendritic cells (DCs) compartment could contribute to the high susceptibility to infections observed in newborns. DCs are among the first cells to colonize the spleen, but the ontogeny of DC subsets follows distinct steps. At birth, plasmacytoid DCs and CD4-CD8alpha- DCs are found in the spleen, whereas CD8alpha+ and CD4+ DCs are not present. Then, the CD8alpha+ DC compartment quickly develops and reaches an adult size by day 7, whereas the CD4+ DC compartment slowly increases to become predominant by the age of 3 weeks. The production of interleukin (IL)-12p70 by DCs is particularly efficient after birth, reflecting the stronger capacity of the neonatal CD8alpha- DCs to secrete IL-12 compared with its adult counterpart. Like-wise, neonatal DCs produced type I and II interferons. In vivo, following microbial stimulation, up-regulation of major histocompatibility complexes (MHCs) and of costimulatory molecules on DCs was induced clearly showing the activation of neonatal DCs in the neonatal environment. Therefore, despite a markedly different DC subset composition in early life compared with the adult DC compartment, neonatal DCs are fully competent in their innate immune functions.

Animals↗

PubMed2003

The shift of Th1 to Th2 immunodominance associated with the chronicity of Mycobacterium bovis bacille Calmette-Guérin infection does not affect the memory response.

In the present study we investigated the shaping and evolution of the immunodominance of the T cell response during a chronic mycobacterial infection. Using a recombinant bacille Calmette-Guérin expressing a reporter Ag, the Escherichia coli MalE protein, we analyzed the peptide specificity and the cytokine profile of the T cell response to the reporter Ag by ELISPOT. During the early steps of infection, the T cell response was focused on two dominant MalE epitopes and was characterized by a pure IFN-gamma response. Then, in the course of infection the initial IFN-gamma response to these two epitopes shifted to a mixed IFN-gamma/IL-4 response. At the same time, the peptide specificity of the T cell response was broadened to two additional MalE epitopes characterized by a unique IL-4 response resulting in the establishment of a dominant IL-4 response to the MalE protein at 16 wk postinfection. However, this phenomenon did not impair the outcome of a predominant IFN-gamma response upon subsequent MalE recall in vivo performed in the presence of CFA, a Th1-driving adjuvant. These results indicate that the Th2 nature of the immune response established during a chronic infection, which most likely reflects regulatory mechanisms to allow the return to T cell homeostasis, does not shape the Th1/Th2 nature of the memory response.

Animals↗

PubMed2003

CD4-independent protective cytotoxic T cells induced in early life by a non-replicative delivery system based on virus-like particles.

The relative immaturity of the neonatal immune system limits CD4(+) Th1 and cytotoxic T lymphocyte (CTL) responses, and represents a significant challenge for the development of vaccines against intracellular pathogens. In this report, we demonstrate the ability of a non-replicative delivery system based on parvovirus-like particles (VLP) to induce CTL responses in the neonatal period. A single immunization of 1-week-old BALB/c mice with recombinant VLP carrying a CD8(+) T cell determinant from lymphocytic choriomeningitis virus (VLP-LCMV) induced antigen-specific CD8(+) cytotoxic T cells that were similar to those elicited by adult immunization, as assessed by cytotoxic activity, interferon (IFN)-gamma secretion, cytotoxic precursor cell frequencies, in vitro avidity for antigen and protective activity against viral challenge. These CTL responses are elicited within 2 weeks of a single immunization, in the absence of adjuvant and independently of the presence and help of CD4(+) T cells, highlighting the potential of VLP as candidate vaccine vectors in early life.

Age Factors↗

PubMed2003

New approaches in vaccine development.

In the last century, vaccines have been one of the most powerful tools for preventing infectious diseases. Smallpox has been eradicated and other diseases such as poliomyelitis or measles have been reduced to very low levels in many regions of the world. However, infectious diseases remain the leading cause of death worldwide. Thus, the development of vaccines to prevent diseases for which no vaccine currently exists such as AIDS or malaria as well as the improvement of efficacy and safety of existing vaccines remains a high priority. Achieving such ambitious goals in a near future will certainly require a strong modification of the methods that have been used so far to identify vaccine candidates. In particular, modern vaccinology could strongly benefit of the latest developments of molecular biology and immunology. Here, we will discuss some potential applications of the increasing knowledge of pathogen genomes as well as the immune system for the discovery of new antigenic targets and the development of new strategies of vaccination.

Acquired Immunodeficiency Syndrome↗

PubMed2003

The B subunit of Shiga toxin coupled to full-size antigenic protein elicits humoral and cell-mediated immune responses associated with a Th1-dominant polarization.

A number of studies in animal models and humans have shown that both humoral andcell-mediated immune responses play an important role in the control of viral infection and tumor development. In most cases, vaccination with non-vectorized peptides or proteins induces low antibody responses and fails to elicit specific cytotoxic T lymphocytes (CTL). In order to make vaccination more efficient, we chemically coupled the non-toxic B subunit of Shiga toxin (STxB) to a full-size antigenic model protein, ovalbumin (OVA), yielding STxB-OVA. We found that STxB-OVA delivers OVA-derived peptides into both the MHC class I- and II-restricted presentation pathways in mouse dendritic cells. Accordingly, the study of STxB trafficking in these cells revealed that, after internalization, a fraction of STxB followed the retrograde transport pathway to the endoplasmic reticulum, while another fraction was targeted to late endosomes/lysosomes. Vaccination of mice with STxB-OVA primed a specific anti-OVA CTL response without the use of adjuvants. Splenocytes and, particularly, CD4(+) T cells from mice immunized with STxB-OVA also produced higher amounts of the T(h)1 cytokine IFN-gamma and IgG2a-type antibodies than mice immunized with non-vectorized ovalbumin. In conclusion, this study identifies a unique non-live vaccine delivery system for polyepitopic antigens that elicits antigen-specific CTL, a humoral immune response and a T(h)1-type polarization without the use of adjuvant.

Animals↗

PubMed2003

Recombinant adenylate cyclase toxin of Bordetella pertussis induces cytotoxic T lymphocyte responses against HLA*0201-restricted melanoma epitopes.

The adenylate cyclase (CyaA) of Bordetella pertussis is able to deliver CD8(+) T cell epitopes into the cytosol of CD11b(+) dendritic cells (DC) following its specific interaction with the alpha(M)beta(2) integrin (CD11b/CD18). This delivery results in intracellular processing and presentation by MHC class I molecules of the CD8(+) T cell epitopes inserted into CyaA. Indeed, we previously showed that CyaA toxins carrying a single cytotoxic T lymphocyte (CTL) epitope can induce efficient protective and therapeutic antitumor immunity in mice. With a view to elaborating cancer immunotherapy in humans using CyaA, we constructed two recombinant CyaA carrying HLA*0201-restricted melanoma epitopes. Here we show that these recombinant CyaA induce strong anti-melanoma CTL responses in HLA*0201 transgenic mice, even after a single i.v. immunization without adjuvant. These responses are long lasting, since they were also detected 5 months after the last injection. Finally, human DC treated with the recombinant CyaA were shown to process and present efficiently the melanoma epitopes to human CTL clones. Altogether, our results demonstrate that tumoral epitopes inserted into CyaA are efficiently processed and presented in association with human MHC molecules. These observations suggest that CyaA is capable of activating antitumoral CTL in humans and highlight the potential of CyaA for use in cancer immunotherapy.

Adenylate Cyclase Toxin↗

PubMed2003

CD8+-T-cell responses of Mycobacterium-infected mice to a newly identified major histocompatibility complex class I-restricted epitope shared by proteins of the ESAT-6 family.

Here we describe the identification of a new CD8(+)-T-cell epitope, the GYAGTLQSL nonamer, shared by the TB10.3 and TB10.4 proteins of the Mycobacterium tuberculosis ESAT-6 family. Cytotoxic T cells from mycobacterium-infected mice efficiently recognized this epitope. GYAGTLQSL-specific T-cell hybridomas, which were able to recognize Mycobacterium bovis BCG-infected macrophages, were generated and now allow investigation of mycobacterial-antigen processing through the major histocompatibility complex class I pathway.

Amino Acid Sequence↗

PubMed2003

[In-vivo dynamic changes of antigen presenting cells infected with recombinant attenuated Salmonella typhimurium expressing green fluorescent protein].

AIM: To clarify in-vivo early dynamic changes of antigen-presenting cells (APCs) infected with recombinant attenuated Salmonella typhimurium. METHODS: BALB/c mice were orally infected with recombinant attenuated Salmonella typhimurium expressing green fluorescent protein (GFP) X4550 (pYAGFP). Peritoneal macrophages were taken out 3 days after infection, cultured for 24 hours, and observed under the fluorescent microscope. Moreover, BALB/c mice were also infected intravenously with X4550 (pYAGFP) and low density cells (LDCs) were isolated and prepared from mouse spleen and liver 3,6 and 12 hours after infection. Infection rates of macrophages and dendritic cells (DCs) were analyzed by flow cytometry. RESULTS: About 50% peritoneal macrophages were X4550 (pYAGFP) positive. Infection rates of macrophages in spleen and liver were about 20%-40%. As for the DCs, the infection rates in spleen and liver were about 4%-10% and 10%-20%, respectively. CONCLUSION: Recombinant attenuated Salmonella can be captured by APCs in-vivo in early infection, which provides a precondition for inducing effective immune responses.

Animals↗

PubMed2002

Induction of HLA-A2-restricted CTL responses by a tubular structure carrying human melanoma epitopes.

Epitope-based vaccination strategies designed to induce strong tumor-specific CD8(+) T cell responses are being widely considered for cancer immunotherapy. Here, two recombinant tubular structures, NS1-Mela 1 and NS1-Mela 2, carrying, respectively two HLA-A2 epitopes derived from human melanoma antigens were constructed and their capability to induce CTL responses in vivo were studied in HLA-A2 transgenic mice. Strong CTL responses specific for GnT-V/NA 17-A and gp100 (154-162) epitopes were generated in HLA-A2 transgenic mice immunized by the construct NS1-Mela l carrying these two epitopes. The second construct NS1-Mela 2 carrying both Tyrosinase (369-377Da) and Melan-A/Mart-1 (27-35) epitopes induced a weak Tyrosinase-specific CTL response in mice but failed to induce specific CTL responses against the Melan-A/Mart-1 (27-35) epitope in the tested mice. Thus, recombinant tubular structures containing multiple tumoral epitopes may lead to new strategies for the induction of strong tumor-specific CTL responses in cancer patients.

Amino Acid Sequence↗

PubMed2002

CD8alpha- CD11b+ dendritic cells present exogenous virus-like particles to CD8+ T cells and subsequently express CD8alpha and CD205 molecules.

Recombinant porcine parvovirus virus-like particles (PPV-VLPs) are particulate exogenous antigens that induce a strong, specific cytotoxic T lymphocyte (CTL) response in the absence of adjuvant. In the present report, we demonstrate in vivo that dendritic cells (DCs) present PPV-VLPs to CD8+ T cells after intracellular processing. PPV-VLPs are captured by DCs with a high efficacy, which results in the delivery of these exogenous antigens to 50% of the whole spleen DC population. In vivo, a few hours after injection, PPV-VLPs are presented exclusively to CD8+ T cells by CD8alpha- DCs, whereas 15 hours later they are presented mainly by CD8alpha+ DCs. After PPV-VLPs processing, a fraction of CD11b+ DCs undergo phenotypic changes, i.e., the up-regulation of CD8alpha and CD205 and the loss of CD4 molecules on their surface. The failure to detect mRNA coding for CD8alpha in CD11b+ DCs suggests that CD8alpha expression by these cells is not due to de novo synthesis. In recombination-activating gene knockout mice (Rag-/-), CD11b+ DCs did not express CD8alpha and PPV-VLPs presentation by CD8alpha+ DCs was severely diminished. These results indicate that both CD8alpha- and CD8alpha+ DCs play an important role in the induction of CTL responses by exogenous antigens, such as VLP.

Animals↗

PubMed2002

Efficient in vivo priming of specific cytotoxic T cell responses by neonatal dendritic cells.

In early life, a high susceptibility to infectious diseases as well as a poor capacity to respond to vaccines are generally observed as compared with observations in adults. The mechanisms underlying immune immaturity have not been fully elucidated and could be due to the immaturity of the T/B cell responses and/or to a defect in the nature and quality of Ag presentation by the APC. This prompted us to phenotypically and functionally characterize early life murine dendritic cells (DC) purified from spleens of 7-day-old mice. We showed that neonatal CD11c(+) DC express levels of costimulatory molecules and MHC molecules similar to those of adult DC and are able to fully maturate after LPS activation. Furthermore, we demonstrated that neonatal DC can efficiently take up, process, and present Ag to T cells in vitro and induce specific CTL responses in vivo. Although a reduced number of these cells was observed in the spleen of neonatal mice as compared with adults, this study clearly shows that neonatal DC have full functional capacity and may well prime Ag-specific naive T cells in vivo.

Animals↗

PubMed2002

Dendritic cells are host cells for mycobacteria in vivo that trigger innate and acquired immunity.

In the present study, we investigated in vivo the infection and APC functions of dendritic cells (DC) and macrophages (Mphi) after administration of live mycobacteria to mice. Experiments were conducted with Mycobacterium bovis bacillus Calmette-Guerin (BCG) or a rBCG expressing a reporter Ag. Following infection of mice, DC and Mphi were purified and the presence of immunogenic peptide/MHC class II complexes was detected ex vivo on sorted cells, as was the secretion of IL-12 p40. We show in this study that DC is a host cell for mycobacteria, and we provide an in vivo detailed picture of the role of Mphi and DC in the mobilization of immunity during the early stages of a bacterial infection. Strikingly, BCG bacilli survive but remain stable in number in the DC leukocyte subset during the first 2 wk of infection. As Ag presentation by DC is rapidly lost, this suggests that DC may represent a hidden reservoir for mycobacteria.

Animals↗

PubMed2002

Induction of protective antiviral cytotoxic T cells by a tubular structure capable of carrying large foreign sequences.

Bluetongue virus (BTV) produces large numbers of tubules during infection which are formed by a single virus coded non-structural protein, NS1. The NS1 protein has been fused with full length green fluorescent protein (GFP) and was shown to retain the capacity to form tubules when expressed in heterologous expression systems. Moreover, recombinant purified chimeric tubules were demonstrated to be internalized by macrophages and dendritic cells. The ability of such chimeric tubules to induce protective cytotoxic T lymphocytes (CTL) responses has been assessed by generating chimeric tubules carrying a single CD8(+) T cell epitope from the lymphocytic choriomeningitis virus (LCMV) nucleoprotein. These chimeric tubules were recognized by MHC class I restricted T cell hybridoma in vitro and induced in vivo strong CD8(+) class I-restricted CTL responses in immunized mice. Further, the immunized mice were protected when challenged with a lethal dose of LCMV. This is the first study that demonstrates that the virus derived tubules synthesized by a recombinant non-structural protein carrying a single viral CTL epitope could induce protective immunity against a lethal viral challenge. Since recombinant tubules carrying large inserts can be purified at a large quantity from insect cells, they have potential to develop as safe multi-CTL vaccine delivery systems.

Animals↗

PubMed2002

In vivo receptor-mediated delivery of a recombinant invasive bacterial toxoid to CD11c + CD8 alpha -CD11bhigh dendritic cells.

The precise contribution of mouse dendritic cells (DC) CD8 alpha +CD11blow and CD8 alpha -CD11bhigh subsets to CTL priming is not fully defined. Here we show that CyaA, the adenylate cyclase toxin of Bordetella pertussis, an invasive bacterial toxin that binds cells through CD11b/CD18 can be exploited for the targeted delivery of an exogenous peptide to the CD8 alpha -CD11bhigh subset in vivo. Antigen (Ag) genetically inserted in the N-terminal domain of mutant CyaA devoid of catalytic activity, are targeted to CD8 alpha -CD11bhigh DC by the CD11b/CD18-dependent binding of CyaA to the cell surface. Ag is then presented by MHC class I molecules of CD8 alpha -CD11bhigh DC after a TAP-dependent, cytosolic processing. As a result, CTL are primed after a single injection, bypassing requirement for adjuvant, CD4+ T cell help and CD40 signaling. Beside the interest of the CyaA vector for vaccine development, these results show that Ag presentation focused on CD8 alpha -CD11bhigh DC in vivo is sufficient for eliciting a vigorous CTL response and that CD11b/CD18 could be a suitable surface molecule for targeting Ag to DC.

Adenylate Cyclase Toxin↗

PubMed2002

Challenges and issues in new vaccine development.

The Euroconference on Vaccines of the Future: from Rational Design to Clinical Development was held at the Pasteur Institute, Paris, France from 17-19 October 2001.

Acquired Immunodeficiency Syndrome↗

PubMed2002

Delivery of a MalE CD4(+)-T-cell epitope into the major histocompatibility complex class II antigen presentation pathway by Bordetella pertussis adenylate cyclase.

Recombinant adenylate cyclase toxoids are shown to deliver inserted foreign CD4(+)-T-cell epitopes into the major histocompatibility complex class II presentation pathway, inducing a specific CD4(+)-T-cell response in vivo and yielding in vitro stimulation of specific CD4(+) T cells at a 100-times-higher molar efficiency than the free peptide containing the epitope.

Adenylate Cyclase Toxin↗

PubMed2002

Virus-like particles: a new family of delivery systems.

The development of antiviral vaccines has almost exclusively been based on live attenuated vaccines up until now. However, the efficacy of HBsAg particles as an antiHBV vaccine has clearly demonstrated that protective antiviral immunity can be achieved by other strategies. Virus-like particles formed by structural proteins were proven to be highly immunogenic and capable of inducing protective immunity against various viral infections in preclinical studies. Clinical trials using virus-like particles confirmed their safety and immunogenicity. Moreover, chimeric virus-like particles carrying foreign peptidic sequences were shown to elicit potent B- and T-cell responses. Virus-like particles formed by a fusion protein between the HBsAg and the circumsporozoïte surface protein are safe and immunogenic in volunteers and induce a partial protection against natural Plasmodium falciparum infection.

Animals↗

PubMedDate not supplied

Chemoselective assembly and immunological evaluation of multiepitopic glycoconjugates bearing clustered Tn antigen as synthetic anticancer vaccines.

In this paper we investigated the use of regioselectively addressable functionalized templates (RAFTs) as new scaffolds for the design of anticancer vaccine candidates. We report the synthesis of well-defined multiepitopic RAFT scaffolds and their immunological evaluation. These conjugates exhibit clustered Tn analogue as tumor-associated carbohydrate antigen (TACA, B-cell epitope) and the CD4+ helper T-cell peptide from the type 1 poliovirus. The saccharidic and peptidic epitopes were both synthesized separately and combined regioselectively to the RAFT core using a sequential oxime bond formation strategy. B- and T-antigenicity and immunogenicity of the vaccine candidates were investigated in vitro and in vivo. These studies clearly demonstrate that the saccharidic part of the conjugates is recognized by Tn-specific monoclonal antibodies. Moreover, the antibodies elicited by immunization of mice with our vaccine candidates recognize the native form of Tn epitope expressed on human tumor cells. Together with oxime ligation technique, these results suggest that the RAFT scaffold provides a promising and suitable tool for engineering potent synthetic anticancer vaccine.

Animals↗