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PubMed1995

Identification of HIV protein-derived cytotoxic T lymphocyte (CTL) epitopes for their possible use as synthetic vaccine.

CTL are by far the most important defence mechanisms against viral infections, and many attempts have been undertaken to induce protective CTL in vivo. In order to identify CTL epitopes for their possible use as peptide-vaccine candidates, HIV proteins were screened for peptide sequences which (i) fulfil the binding motif of the HLA-A2.1 molecule, and (ii) are involved in the natural immune response to HIV. From 73 nonameric peptides satisfying the binding motif, 20 peptides were synthesized and their binding to HLA-A2.1 was monitored by measuring the expression of HLA-A2.1 molecules on the cell surface of the mutant cell line T2. To evaluate the involvement in natural HIV infection, strongly binding peptides were used in cytotoxicity assays to assess their capacity to generate a peptide-specific CTL response in vitro. From 20 nonameric peptides synthesized, only five showed strong binding to HLA-A2.1. All five binding peptides had the secondary anchor residues, recently proposed by Ruppert et al. [1] to be required for binding to HLA-A2.1. The discrimination between bound and unbound peptides confirmed the importance of these secondary anchor residues which, beside the known binding motif, may dictate if a peptide can bind to HLA-A2.1 or not. In HIV- donors, no CTL activity against any of the HIV-derived peptides was detectable after a 12-day in vitro stimulation. In contrast, HIV-infected persons showed a cytotoxic response against peptide-labelled target cells, suggesting that they had developed upon HIV infection a cytotoxic immune response against the identified CTL epitopes.

Amino Acid Sequence↗
PubMed1988

Possible involvement of CTL granule proteases in target cell DNA breakdown.

We have carried out experiments to test whether the granule exocytosis model for lymphocyte cytotoxicity can account for the rapid target DNA breakdown seen during CTL-induced cytotoxicity. Dense granules isolated from cloned mouse CTL and from rat NK tumor cells cause target DNA breakdown during granule cytolysin-mediated lysis of tumor cells, while the purified granule cytolysin caused lysis without DNA breakdown. When target cells are permeabilized with detergent, granule extracts have the ability to release 125I-DNA from nuclei in the absence of detectable cytolysin activity. This activity formed the basis for a nuclear DNA release (NDR) assay; this activity was a property of dense granules of cytotoxic lymphocytes but generally not of other types of lymphoid cells. NDR activity in NK tumor granules had a pH optimum of 7 and was inhibited by micromolar levels of Zn+2, and could be purified away from the granule cytolysin by column chromatography. NDR activity in CTL dense granules could be inactivated by submillimolar concentrations of the protease inhibitors PMSF and DFP (but not soybean trypsin inhibitor or TLCK). In support of the relevance to CTL cytotoxicity of these findings with the NDR assay, pretreatment of CTL with PMSF in the presence of agents raising the intragranular pH inactivated 125I-DNA release from target cells (but not the 51Cr release). These results suggest that a CTL granule component(s), probably a protease, is required for target DNA breakdown.

Animals↗
PubMed1998

Quantitative and functional analysis of core-specific T-helper cell and CTL activities in acute and chronic hepatitis B.

AIMS/BACKGROUND: CD4+ T-helper cell (Th) responses to hepatitis B virus (HBV) core antigen (HBc) are increased during exacerbations in acute and chronic hepatitis B (AHB, CHB) and might influence the induction of CD8+ cytotoxic T lymphocytes (CTL) that are important for viral clearance. METHODS: HBc-specific proliferative responses and cytokine release of blood mononuclear cells (PBMC) were studied in patients with AHB or CHB, as well as responders and non-responders to interferon-alpha treatment (IFN-R, IFN-NR), by [3H]-thymidine-uptake, enzyme-linked immunosorbent assay (ELISA) and Elispot assay and were compared to peptide HBc18 27-specific CTL precursor frequencies among CD8+ T cells derived from HLA-A2+ patients. RESULTS: HBc-specific proliferative PBMC responses and Th frequencies were significantly increased in AHB patients compared with untreated CHB patients. PBMC derived from IFN-R showed stronger cellular responses than IFN-NR. Stimulated PBMC from all patient groups secreted significantly more IFN-gamma than IL-4 indicating Th1/Th0 cell responses. Furthermore, in AHB and IFN-R patients, high peptide HBc18-27-specific CTL precursor frequencies closely correlated with strong HBc-specific Th responses, whereas in untreated CHB and IFN-NR patients lower CTL frequencies were observed without correlation to Th activities. CONCLUSIONS: HBV core-specific Th-cell responses appeared to support efficient CTL induction in patients with viral clearance, whereas in chronic HBV carriers quantitatively insufficient Th and CTL responses were observed. This observation could be important for future therapeutic strategies.

Acute Disease↗
PubMedDate not supplied

Presence of circulating CTL induced by infection with wild-type or attenuated SIV and their correlation with protection from pathogenic SHIV challenge.

The aim of this study was to evaluate the role of CTLs in the protection from challenge with pathogenic SHIV in macaques vaccinated with attenuated virus. More specifically, we have analyzed the CTL response in cynomolgus macaques vaccinated/infected with the attenuated SIVmacC8 or the wild-type SIVmacJ5 and correlated these responses to the protection from SHIV89.6P challenge. SIVmacC8-vaccinated monkeys demonstrated a broader CTL response than the SIVmacJ5-infected animals. Nevertheless, CTL against some proteins in SIVmacC8-vaccinated monkeys became progressively more difficult to detect through the day of challenge. In regards to protection from superinfection with SHIV89.6P, neither the presence of circulating CTL nor the CTL precursor frequency against any of the tested proteins correlated with the outcome of the challenge when SIVmacJ5- and SIVmacC8-infected animals were analyzed together. By analyzing the SIVmacC8-vaccinated animals separately, only the protected animal had detectable CTL precursors with moderate frequencies against all three tested proteins at the day of challenge.

AIDS Vaccines↗
PubMed1988

Differential expression of the ASGM1 antigen on anti-reovirus and alloreactive cytotoxic T lymphocytes (CTL).

Anti-reovirus cytotoxic effectors were found to be: (i) H-2 restricted; (ii) virus specific; (iii) non-lytic (in 4 h) for natural killer (NK)-sensitive YAC-1 cells; and (iv) positive for the Thy-1 and Lyt-2 lymphocyte markers. Thus, anti-reovirus cytotoxic effectors have the functional and phenotype characteristics of cytotoxic T lymphocyte (CTL). A significant fraction of anti-viral CTL, as well as alloreactive CTL, were also found to be positive for the asialo GM1 (ASGM1) cell surface antigen, generally considered to be a NK cell marker. ASGM1 expression on these CTL, as determined by sensitivity to antibody plus complement (C), appeared to be highly variable and dependent on two factors-the nature of the antigenic stimulus (viral vs. alloantigen), and the mouse strain from which the CTL originated. Thus, ASGM1 antigen expression on CTL appears to be regulated and may be under the control of lymphokines, development differentiation signals and/or other strain-dependent genetic factors.

Animals↗
PubMed2000

Cytotoxic T lymphocyte responses and CTL epitope escape mutation in HBsAg, anti-HBe positive individuals.

BACKGROUND/AIMS: Clearance of hepatitis B virus (HBV) is characterised by a strong cytotoxic T cell response. Persistence of HBV in chronic hepatitis B carriers may be related to failure of this response. The aim of this study was to determine whether HLA class I restricted cytotoxic T lymphocyte (CTL) responses persist in anti-hepatitis B e (HBe) positive / HBV DNA negative individuals, and to correlate the presence of viral CTL epitope mutation with clinical outcome. METHODS: An HLA/HBV dual transfectant model was used to demonstrate these CTL responses in individuals chronically infected with HBV. Subsequently, a known hepatitis B core (HBc) CTL epitope was sequenced in a family of five chronically infected individuals all sharing a HLA allele (HLA-A68.1). RESULTS: Low level HLA class I restricted cytotoxic T cell responses were detected in the peripheral blood of five of eight anti-HBe positive individuals. In the family of HLA-A68.1 positive chronically infected individuals, mutation of the HLA-A68.1 restricted hepatitis B core antigen (HBcAg) CTL epitope STLPETTVVRR was found in all four anti-HBe positive individuals but not in the sole hepatitis B e antigen (HBeAg) positive patient. CONCLUSION: These data are consistent with a continued immune selection pressure on HBV in anti-HBe positive chronically infected individuals with low replicating HBV infection and suggest that mutation of a CTL epitope may be a consequence of the immune response, as opposed to the cause of viral persistence.

Adult↗
PubMed2005

Determination of cellularly processed HLA-A2402-restricted novel CTL epitopes derived from two cancer germ line genes, MAGE-A4 and SAGE.

PURPOSE: For identification of CTL epitopes useful for cancer vaccines, it is crucial to determine whether cognate epitopes are presented on the cell surface of target cancer cells through natural processing of endogenous proteins. For this purpose, we tried to use the cellular machinery of both mice and human to define naturally processed CTL epitopes derived from two "cancer germ line" genes, MAGE-A4 and SAGE. EXPERIMENTAL DESIGN: We vaccinated newly produced HLA-A2402 transgenic mice with DNA plasmids encoding target antigens. Following screening of synthesized peptides by splenic CD8(+) T cells of vaccinated mice, we selected candidate epitopes bound to HLA-A2402. We then examined whether human CD8(+) T cells sensitized with autologous CD4(+) PHA blasts transduced by mRNA for the cognate antigens could react with these selected peptides in an HLA-A2402-restricted manner. RESULTS: After DNA vaccination, murine CD8(+) T cells recognizing MAGE-A4(143-151) or SAGE(715-723) in an HLA-A2402-restricted manner became detectable. Human CTLs specific for these two peptides were generated after sensitization of HLA-A2402-positive CD8(+) T cells with autologous CD4(+) PHA blasts transduced with respective mRNA. CTL clones were cytotoxic toward tumor cell lines expressing HLA-A2402 and cognate genes. Taken together, these CTL epitopes defined in HLA-A24 transgenic mice are also processed and expressed with HLA-A2402 in human cells. The presence of SAGE(715-723)-specific precursors was observed in HLA-A2402-positive healthy individuals. CONCLUSIONS: Two novel HLA-A2402-restricted CTL epitopes, MAGE-A4(143-151) and SAGE(715-723), were identified. Our approach assisted by cellular machinery of both mice and human could be widely applicable to identify naturally processed CTL epitopes.

Animals↗
PubMed2004

Therapeutic polypeptides based on HBV core 18-27 epitope can induce CD8+ CTL-mediated cytotoxicity in HLA-A2+ human PBMCs.

AIM: To explore how to improve the immunogenicity of HBcAg CTL epitope based polypeptides and to trigger an HBV-specific HLA I-restricted CD8+ T cell response in vitro. METHODS: A new panel of mimetic therapeutic peptides based on the immunodominant B cell epitope of HBV PreS2 18-24 region, the CTL epitope of HBcAg18-27 and the universal T helper epitope of tetanus toxoid (TT) 830-843 was designed using computerized molecular design method and synthesized by Merrifield's solid-phase peptide synthesis. Their immunological properties of stimulating activation and proliferation of lymphocytes, of inducing T( H1) polarization, CD8+ T cell magnification and HBV-specific CD8+ CTL mediated cytotoxicity were investigated in vitro using HLA-A2+ human peripheral blood mononuclear cells (PBMCs) from healthy donors and chronic hepatitis B patients. RESULTS: Results demonstrated that the therapeutic polypeptides based on immunodominant HBcAg18-27 CTL, PreS2 B- and universal T(H) epitopes could stimulate the activation and proliferation of lymphocytes, induce specifically and effectively CD8+ T cell expansion and vigorous HBV-specific CTL-mediated cytotoxicity in human PBMCs. CONCLUSION: It indicated that the introduction of immunodominant T helper plus B-epitopes with short and flexible linkers could dramatically improve the immunogenicity of short CTL epitopes in vitro.

Amino Acid Sequence↗
PubMed2000

Bovine and human insulin activate CD8+-autoreactive CTL expressing both type 1 and type 2 cytokines in C57BL/6 mice.

CD8+ T cells down-regulate a variety of immune responses. For example, porcine and human insulin do not stimulate Abs in C57BL/6 mice because CD8+ T cells inhibit CD4+ helper T cells. By contrast, bovine insulin induces Ab in C57BL/6 mice, and removal of CD8+ T cells does not alter this response. This raises the question of whether porcine, but not bovine, insulin activates CD8+ T cells or whether both insulins activate CD8+ T cells but CD4+ helper T cells are differentially inhibited by them. In this study, we show that insulin-specific CD8+ CTL can be cultured from C57BL/6 mice primed with either bovine or human insulin in CFA. Thus, exogenous Ags, besides OVA, induce CD8+ CTL when administered in an adjuvant, suggesting this is a typical response. These CTL are H-2Kb restricted and produce IL-5, IL-10, IFN-gamma, and small amounts of IL-4, which is distinct from IFN-gamma and TNF-alpha that are typically secreted by virus-specific CTL. Moreover, the CTL primed with either bovine or human insulin recognize an A-chain peptide that is identical to the mouse insulin sequence. That foreign proteins, which are closely related to self-proteins, activated autoreactive, CD8+ T cells in vivo is a novel finding. It raises the possibility that self-reactive CTL may be activated by cross-reacting Ags and once activated they might participate in autoimmunity. These results also suggest that down-regulation of insulin-specific responses by autoreactive CD8+ T cells is most likely due to the differential sensitivity of bovine and human insulin-specific CD4+ T cells.

Amino Acid Sequence↗
PubMed2000

Differential role of Fas/Fas ligand interactions in cytolysis of primary and metastatic colon carcinoma cell lines by human antigen-specific CD8+ CTL.

We have previously identified mutated ras peptides reflecting the glycine to valine substitution at position 12 as HLA-A2-restricted, CD8+ CTL neo-epitopes. CTL lines produced against these peptide epitopes lysed the HLA-A2+ Ag-bearing SW480 primary colon adenocarcinoma cell line, although IFN-gamma treatment of the targets was necessary to achieve efficient cytotoxicity. Here, we compared the lytic phenotype of the SW480 cell line to its metastatic derivative, SW620, as an in vitro paradigm to further characterize the nature of a HLA class I-restricted, Ag-specific CTL response against neoplastic cell lines of primary and metastatic origin. Although both colon carcinoma cell lines were lysed by these Ag-specific CTL following IFN-gamma pretreatment, the mechanisms of lysis were distinct, which reflected differential levels of sensitivity to the Fas pathway. Whereas IFN-gamma pretreatment rendered SW480 cells sensitive to both Fas-dependent and -independent (perforin) pathways, SW620 cells displayed lytic susceptibility to Fas-independent mechanisms only. Moreover, pretreatment of SW480 cells with the anti-colon cancer agent, 5-fluorouracil (5-FU), led to enhanced Fas and ICAM-1 expression and triggered Ag-specific CTL-mediated lysis via Fas- and perforin-based pathways. In contrast, these phenotypic and functional responses were not observed with SW620 cells. Overall, these data suggested that 1) IFN-gamma and 5-FU may enhance the lytic sensitivity of responsive colon carcinoma cells to immune effector mechanisms, including Fas-induced lysis; 2) the malignant phenotype may associate with resistance to Fas-mediated lysis in response to Ag-specific T cell attack; and 3) if Ag-specific CTL possess diverse lytic capabilities, this may overcome, to some extent, the potential "escape" of Fas-resistant carcinoma cells.

Adenocarcinoma↗
PubMed2000

Efficient simultaneous presentation of NY-ESO-1/LAGE-1 primary and nonprimary open reading frame-derived CTL epitopes in melanoma.

Recent studies have shown that CTL epitopes derived from tumor-associated Ags can be encoded by both primary and nonprimary open reading frames (ORF). In this study we have analyzed the HLA-A2-restricted CD8(+) T cell response to a recently identified CTL epitope derived from an alternative ORF product of gene LAGE-1 (named CAMEL), and the highly homologous gene NY-ESO-1 in melanoma patients. Using MHC/peptide tetramers we detected CAMEL(1-11)-specific CD8(+) T cells in peptide-stimulated PBMC as well as among tumor-infiltrated lymph node cells from several patients. Sorting and expansion of tetramer(+) CD8(+) T cells allowed the isolation of tetramer(bright) and tetramer(dull) populations that specifically recognized the peptide Ag with high and low avidity, respectively. Remarkably, only high avidity CAMEL-specific CTL were able to recognize Ag-expressing tumor cells. A large series of HLA-A2-positive melanoma cell lines was characterized for the expression of LAGE-1 and NY-ESO-1 mRNA and protein and tested for recognition by CAMEL-specific CTL as well as CTL that recognize a peptide (NY-ESO-1(157-165)) encoded by the primary ORF products of the LAGE-1 and NY-ESO-1 genes. This analysis revealed that tumor-associated CD8(+) T cell epitopes are simultaneously and efficiently generated from both primary and nonprimary ORF products of LAGE-1 and NY-ESO-1 genes and, importantly, that this occurs in the majority of melanoma tumors. These findings underscore the in vivo immunological relevance of CTL epitopes derived from nonprimary ORF products and support their use as candidate vaccines for inducing tumor specific cell-mediated immunity against cancer.

Animals↗
PubMed2000

MHC class Ib-restricted CTL provide protection against primary and secondary Listeria monocytogenes infection.

Infection of B6 mice with the intracellular pathogen Listeria monocytogenes (LM) results in the activation of CD8(+) T cells that respond to Ag presented by both MHC class Ia and class Ib molecules. Enzyme-linked immunospot analysis reveals that these CTL populations expand and contract at different times following a primary sublethal LM infection. Between days 4 and 6 postinfection, class Ib-restricted CTL exhibit a rapid proliferative response that is primarily H2-M3 restricted. The peak response of class Ia-restricted CD8(+) T cells occurs a few days later, after the majority of bacteria have been cleared. Although class Ia-restricted CTL exhibit a vigorous recall response to secondary LM infection, we observe limited expansion of class Ib-restricted memory CTL, even in MHC class Ia-deficient mice (B6.K(b-/-)D(b-/-)). Despite this lack of enhanced expansion in vivo, class Ib-restricted memory CTL retain the ability to proliferate and expand when provided with Ag in vitro. Furthermore, we demonstrate that in vivo depletion of CD8(+) T cells in LM-immune B6.K(b-/-)D(b-/-) mice severely impairs memory protection. Together, these data demonstrate that class Ib-restricted CTL play an important role in clearing a primary LM infection and generate a memory population capable of providing significant protection against subsequent infection.

Animals↗
PubMed2001

Mucosal delivery of a respiratory syncytial virus CTL peptide with enterotoxin-based adjuvants elicits protective, immunopathogenic, and immunoregulatory antiviral CD8+ T cell responses.

In an effort to develop a safe and effective vaccine against respiratory syncytial virus (RSV), we used Escherichia coli heat-labile toxin (LT), and LTK63 (an LT mutant devoid of ADP-ribosyltransferase activity) to elicit murine CD8(+) CTL responses to an intranasally codelivered CTL peptide from the second matrix protein (M2) of RSV. M2(82-90)-specific CD8(+) T cells were detected by IFN-gamma enzyme-linked immunospot and (51)Cr release assay in local and systemic lymph nodes, and their induction was dependent on the use of a mucosal adjuvant. CTL elicited by peptide immunization afforded protection against RSV challenge, but also enhanced weight loss. CTL-mediated viral clearance was not dependent on IFN-gamma since depletion using specific mAb during RSV challenge did not affect cellular recruitment or viral clearance. Depletion of IFN-gamma did, however, reduce the concentration of TNF detected in lung homogenates of challenged mice and largely prevented the weight loss associated with CTL-mediated viral clearance. Mice primed with the attachment glycoprotein (G) develop lung eosinophilia after intranasal RSV challenge. Mucosal peptide vaccination reduced pulmonary eosinophilia in mice subsequently immunized with G and challenged with RSV. These studies emphasize that protective and immunoregulatory CD8(+) CTL responses can be mucosally elicited using enterotoxin-based mucosal adjuvants but that resistance against viral infection may be accompanied by enhanced disease.

Adjuvants, Immunologic↗
PubMed2001

Induction of polyclonal prostate cancer-specific CTL using dendritic cells transfected with amplified tumor RNA.

Polyvalent cancer vaccines targeting the entire antigenic spectrum on tumor cells may represent a superior therapeutic strategy for cancer patients than vaccines solely directed against single Ags. In this study, we show that autologous dendritic cells (DC) transfected with RNA amplified from microdissected tumor cells are capable of stimulating CTL against a broad set of unidentified and critical prostate-specific Ags. Although the polyclonal CTL responses generated with amplified tumor RNA-transfected DC encompassed as a subcomponent a response against prostate-specific Ag (PSA) as well as against telomerase reverse transcriptase, the tumor-specific CTL were consistently more effective than PSA or telomerase reverse transcriptase CTL to lyse tumor targets, suggesting the superiority of the polyclonal response. Although tumor RNA-transfected DC stimulated CTL, which recognized not only tumor but also self-Ags expressed by benign prostate tissue, these cross-reactive CTL were exclusively specific for the PSA, indicating an immunodominant role of PSA in the prostate cancer-specific immune response. Our data suggest that tumor RNA-transfected DC may represent a broadly applicable, potentially clinically effective vaccine strategy for prostate cancer patients, which is not limited by tumor tissue availability for Ag preparation and may minimize the risk of clonal tumor escape.

Adjuvants, Immunologic↗
PubMed2001

Cancer vaccine design: a novel bacterial adjuvant for peptide-specific CTL induction.

The recent identification of tumor Ags as potential vaccines has prompted the search for efficient adjuvants and delivery systems, especially in the case of peptide-based vaccination protocols. Here, we investigated the adjuvant potential of the recombinant 40-kDa outer membrane protein of Klebsiella pneumoniae (P40) for specific CTL induction. We studied the CTL response induced in HLA-A*0201/K(b) transgenic mice immunized with peptides derived from two melanoma-associated differentiation Ags, the HLA-A*0201-restricted decapeptide Melan-A(26--35) substituted at position 2 and the K(b)-restricted tyrosinase-related protein 2(181--188) T cell epitope. We found that both peptides are able to generate a specific CTL response when mixed with the protein in the absence of conventional adjuvant. This CTL response is a function of the amount of P40 used for immunization. Moreover, the CTL response generated against the tyrosinase-related protein 2(181-188) peptide in presence of P40 is associated with tumor protection in two different experimental models and is independent of the presence of CD4(+) T lymphocytes. Thus, the recombinant bacterial protein P40 functions as a potent immunological adjuvant for specific CTL induction.

Adjuvants, Immunologic↗
PubMed2001

Efficient delivery of Antennapedia homeodomain fused to CTL epitope with liposomes into dendritic cells results in the activation of CD8+ T cells.

The in vivo induction of a CTL response using Antennapedia homeodomain (AntpHD) fused to a poorly immunogenic CTL epitope requires that the Ag is given in presence of SDS, an unacceptable adjuvant for human use. In the present report, we developed a hybrid CTL epitope delivery system consisting of AntpHD peptide vector formulated in liposomes as an alternative approach to bypass the need for SDS. It is proposed that liposomes will prevent degradation of the Ag in vivo and will deliver AntpHD recombinant peptide to the cytosol of APCs. We show in this work that dendritic cells incubated with AntpHD-fused peptide in liposomes can present MHC class I-restricted peptide and induce CTL response with a minimal amount of Ag. Intracellular processing studies have shown that encapsulated AntpHD recombinant peptide is endocytized before entering the cytosol, where it is processed by the proteasome complex. The processed liposomal peptides are then transported to the endoplasmic reticulum. The increase of the CTL response induced by AntpHD-fused peptide in liposomes correlates with this active transport to the class I-processing pathway. In vivo studies demonstrated that positively charged liposomes increase the immunogenicity of AntpHD-Cw3 when injected s.c. in mice in comparison to SDS. Moreover, addition of CpG oligodeoxynucleotide immunostimulatory sequences further increase the CD8+ T cell response. This strategy combining lipid-based carriers with AntpHD peptide to target poorly immunogenic Ags into the MHC class I processing pathway represents a novel approach for CTL vaccines that may have important applications for development of cancer vaccines.

Animals↗
PubMed2001

Clonally diverse CTL response to a dominant viral epitope recognizes potential epitope variants.

RNA viruses undergo rapid sequence variation as the result of error-prone RNA replication mechanisms. When viable mutations arise in RNA regions encoding B or T cell epitopes, mutant viruses that can evade immune detection may be selected. In the carefully studied CTL response to the Gag p11C(C-M) epitope in SIVmac-infected Mamu-A*01(+) rhesus monkeys, it has been shown that CTL recognition of that epitope can occur even in the face of accruing mutations. To explore the underlying mechanism for this breadth of recognition, we have constructed Mamu-A*01 tetramers which discriminate T cells specific for epitope variants. Using these reagents we have defined discrete subsets of p11C(C-M)-specific T cells that cross-react with cells presenting variant peptides. We have found that individual Mamu-A*01(+) monkeys differ functionally in their ability to recognize epitope variants despite consistently strong recognition of the p11C(C-M) epitope. This functional difference is accounted for by the relative number of variant-specific T cells and by differences in the functionally relevant TCR repertoire of the infected monkeys. We have also found that monkeys immunized with DNA vaccine constructs encoding only the wild-type epitope sequence develop p11C(C-M)-specific CTL cross-reactive with variant peptides. Thus, cross-reactive CTL do not merely arise secondary to the emergence and immune presentation of viral CTL escape mutants but rather arise de novo following priming with a dominant epitope peptide sequence. Taken together, our results support the concept that the CTL response to a dominant viral epitope, although highly focused, can be clonally diverse and recognize potential epitope variants.

Animals↗
PubMed2002

Association of prolonged survival in HLA-A2+ progressive multifocal leukoencephalopathy patients with a CTL response specific for a commonly recognized JC virus epitope.

The role of JC virus (JCV)-specific CTL was explored in the immunopathogenesis of progressive multifocal leukoencephalopathy (PML). We identified a 9-aa epitope of the JCV capsid protein VP1, the VP1(p100) peptide ILMWEAVTL, which is recognized by CTL of HLA-A2+ HIV+/PML survivors. We then constructed an HLA-A*0201/VP1(p100) tetrameric complex that allowed us to assess by flow cytometry the PBMC of 13 PML patients and 11 control subjects for the presence of JCV-specific CTL. VP1(p100)-specific CTL were detected by tetramer binding in VP1(p100)-stimulated PBMC of five of seven (71%) PML survivors and zero of six PML progressors (p = 0.02). Two of three HIV+ patients with a leukoencephalopathy resembling PML, but with no virologic evidence of JCV infection, also had detectable VP1(p100)-specific CTL in their PBMC. PBMC of eight HIV+ patients with other neurologic diseases and healthy control subjects had no detectable JCV-specific CTL. These data suggest that the JCV-specific cellular immune response may be important in the containment of PML, and the tetramer-staining assay may provide a useful prognostic tool in the clinical management of these patients.

Antigens, Viral↗