[show abstract][hide abstract] ABSTRACT: Certain peptide analogs that carry substitutions at residues other than the main major histocompatibility complex anchors and are surprisingly much more antigenic than wild-type peptide (heteroclitic analogs). To date, it was unknown how frequently wild-type epitopes could be modified to obtain heteroclitic activity. In this study, we analyzed a large panel of analogs of two different human histocompatibility leukocyte antigen (HLA)-A2.1-restricted epitopes and found that heteroclitic analogs were associated with higher magnitude responses and increased (up to 10(7)-fold) sensitivity to antigen, and corresponded to conservative or semiconservative substitutions at odd-numbered positions in the middle of the peptide (positions 3, 5, or 7). These findings were validated by performing additional immunogenicity studies in murine and human systems with four additional epitopes. The biological relevance of heteroclitic analogs was underlined when predicted analogs of the p53.261 epitope was shown to induce cytotoxic T lymphocytes (CTLs) that recognize low concentrations of peptide (high avidity) in vivo and demonstrate in vitro antitumor recognition. Finally, in vitro immunization of human peripheral blood mononuclear cells with two heteroclitic analogs resulted in recruitment of more numerous CTLs which were associated with increased antigen sensitivity. In conclusion, heteroclitic analogs were identified in each of the six cases studied and structural features were defined which allow identification of such analogs. The strong CTL immunity elicited by heteroclitic epitopes suggest that they could be of significant value in vaccination against tolerant or weakly immunogenic tumor-associated and viral antigens.
Journal of Experimental Medicine 10/2001; 194(6):833-46. · 13.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: Cytotoxic T lymphocytes (CTL) recognize and kill virus-infected cells and contribute to immunologic control of viral replication. For many herpesviruses (e.g., Epstein-Barr and cytomegalovirus), virus-specific CTL responses can be readily detected in infected persons, but CTL responses against Kaposi's sarcoma-associated herpesvirus (KSHV) appear to be weak and remain poorly characterized. Using a human leukocyte antigen (HLA) binding motif-based epitope prediction algorithm, we identified 37 HLA-A*0201 binding peptides from 8 KSHV open-reading frames (ORFs). After in vitro stimulation of peripheral blood mononuclear cells from KSHV-infected persons, CTL responses against 1 peptide in the KSHV kaposin protein (ORF K12) were detected in 2 HLA-A*0201-positive subjects. The optimal CTL epitope was identified by HLA restriction analysis and peptide titration assays. These data describe a latent phase viral gene product targeted by CTL that may be relevant for KSHV immunopathogenesis.
The Journal of Infectious Diseases 08/2001; 184(2):119-26. · 5.85 Impact Factor
[show abstract][hide abstract] ABSTRACT: Forty-two wild-type and analogue peptides derived from p53, carcinoembryonic Ag, Her2/neu, and MAGE2/3 were screened for their capacity to induce CTLs, in vitro, capable of recognizing tumor target lines. All the peptides bound HLA-A*0201 and two or more additional A2 supertype alleles with an IC(50) of 500 nM or less. A total of 20 of 22 wild-type and 9 of 12 single amino acid substitution analogues were found to be immunogenic in primary in vitro CTL induction assays, using normal PBMCs and GM-CSF/IL-4-induced dendritic cells. These results suggest that peripheral T cell tolerance does not prevent, in this system, induction of CTL responses against tumor-associated Ag peptides, and confirm that an HLA class I affinity of 500 nM or less is associated with CTL epitope immunogenicity. CTLs generated by 13 of 20 of the wild-type epitopes, 6 of 9 of the single, and 2 of 5 of the double substitution analogues tested recognized epitopes generated by endogenous processing of tumor-associated Ags and expressed by HLA-matched cancer cell lines. Further analysis revealed that recognition of naturally processed Ag was correlated with high HLA-A2.1-binding affinity (IC(50) = 200 nM or less; p = 0.008), suggesting that high binding affinity epitopes are frequently generated and can be recognized as a result of natural Ag processing. These results have implications for the development of cancer vaccines, in particular, and for the process of epitope selection in general.
The Journal of Immunology 08/2001; 167(2):787-96. · 5.52 Impact Factor
[show abstract][hide abstract] ABSTRACT: This unit describes a technique for the direct and quantitative measurement of the capacity of peptide ligands to bind Class I and Class II MHC molecules. The binding of a peptide of interest to MHC is assessed based on its ability to inhibit the binding of a radiolabeled probe peptide to MHC molecules. The establishment of an MHC/peptide binding assay, and its subsequent use in determining the MHC binding capacities of peptide ligands, requires sufficient stocks of purified MHC and both labeled and unlabeled peptides. Accordingly, this unit includes protocols for the purification of Class I and Class II MHC molecules by affinity chromatography, and for the radiolabeling of peptides using the chloramine T method. A support protocol describes alterations in the basic protocol that are necessary when performing direct binding assays, which are required for (1) selecting appropriate high-affinity, assay-specific, radiolabeled ligands and (2) determining the amount of MHC necessary to yield assays with the highest sensitivity. After a 2-day incubation, the bound and unbound radiolabeled species are separated, and their relative amounts are determined. Two methods for separation by size-exclusion gel-filtration chromatography are described, as is data analysis.
Current protocols in immunology / edited by John E. Coligan ... [et al.] 06/2001; Chapter 18:Unit 18.3.
[show abstract][hide abstract] ABSTRACT: Human immunodeficiency virus (HIV)-specific helper T lymphocytes (HTL) play a key role in the immune control of HIV type 1 (HIV-1) infection, and as such are an important target of potential HIV-1 vaccines. In order to identify HTL epitopes in HIV-1 that might serve as vaccine targets, conserved HIV-1-derived peptides bearing an HLA-DR binding supermotif were tested for binding to a panel of the most representative HLA-DR molecules. Eleven highly cross-reactive binding peptides were identified: three in Gag and eight in Pol. Lymphoproliferative responses to this panel of peptides, as well as to the HIV-1 p24 and p66 proteins, were evaluated with a cohort of 31 HIV-1-infected patients. All 11 peptides were recognized by peripheral blood mononuclear cells from multiple HIV-infected donors. Many of the responsive HIV-infected subjects showed recognition of multiple peptides, indicating that HIV-1-specific T-helper responses may be broadly directed in certain individuals. A strong association existed between recognition of the parental recombinant HIV-1 protein and the corresponding HTL peptides, suggesting that these peptides represent epitopes that are processed and presented during the course of HIV-1 infection. Lastly, responses to the supermotif peptides were mediated by CD4(+) T cells and were restricted by major histocompatibility complex class II molecules. The epitopes described herein are potentially important components of HIV-1 therapeutic and prophylactic vaccines.
Journal of Virology 06/2001; 75(9):4195-207. · 5.08 Impact Factor
[show abstract][hide abstract] ABSTRACT: The 'classical' myelin basic protein (MBP) exons belong to a much larger unit, termed the 'Golli-MBP' gene. Here we have examined the T-cell determinant structure of the Golli protein region in the BALB/c mouse. Golli p10-24, which was shown to have the strongest affinity for I-A(d), could not induce T-cell activation. Paradoxically, the poorer binding, overlapping p5-19 was effective at inducing T-cell proliferation. Thus, immunogenicity is not necessarily related to the MHC-binding affinity of self-peptides. In addition, MBP: p151-168-specific T cell clones responded only poorly to J37, a Golli-MBP protein, while MBP: 59-76-specific clones responded well to J37.
Journal of Autoimmunity 12/2000; 15(3):315-22. · 8.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: Studies of mouse models of tuberculosis (TB) infection have indicated a central role for MHC class I-restricted CD8+ T cells in protective immunity. To define antigens and epitopes of Mycobacterium tuberculosis (MTB) proteins that are presented by infected cells to CD8+ T cells, we screened 40 MTB proteins for HLA class I A*0201-binding motifs. Peptides that bound with high affinity to purified HLA molecules were subsequently analyzed for recognition by CD8+ cytotoxic T lymphocytes. We identified three epitopes recognized by CD8+ T cells from patients recovering from TB infection. Those three epitopes were derived from three different antigens: thymidylate synthase (ThyA(30-38)), RNA polymerase beta-subunit (RpoB(127-135)), and a putative phosphate transport system permease protein A-1 (PstA1(75-83)). In addition, CD8+ T cell lines specific for three peptides (ThyA(30-38), PstA1(75-83), and 85B(15-23)) were generated from peripheral blood mononuclear cells of normal HLA-A*0201 donors. These CD8+ T cell lines specifically recognized MTB-infected macrophages, as demonstrated by production of IFN-gamma and lysis of the infected target cells. Finally, CD8+ cytotoxic T lymphocytes reduced the viability of the intracellular MTB, providing evidence that CD8+ T cell recognition of MHC class I-restricted epitopes of these MTB antigens can contribute to effective immunity against the pathogen.
Proceedings of the National Academy of Sciences 11/2000; 97(22):12210-5. · 9.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: Previously, we identified and established the antigenicity of 17 CD8+ T cell epitopes from five P. falciparum Ags that are restricted by multiple common HLA class I alleles. Here, we report the identification of 11 peptides from the same Ags, cicumsporozoite protein, sporozoite surface protein 2, exported protein-1, and liver-stage Ag-1, that bind between at least five and up to 11 different HLA-DR molecules representative of the most common HLA-DR Ags worldwide. These peptides recall lymphoproliferative and cytokine responses in immune individuals experimentally immunized with radiation-attenuated Plasmodium falciparum sporozoites (irradiated sporozoites) or semi-immune individuals naturally exposed to malaria in Irian Jaya or Kenya. We establish that all peptides are recognized by individuals of each of the three populations, and that the frequency and magnitude of helper T lymphocyte responses to each peptide is influenced by the intensity of exposure to P. falciparum sporozoites. Mean frequencies of lymphoproliferative responses are 53.2% (irradiated sporozoites) vs 22.4% (Kenyan) vs 5.8% (Javanese), and mean frequencies of IFN-gamma responses are 66.3% (irradiated sporozoites) vs 27.3% (Kenyan) vs 8. 7% (Javanese). The identification of HLA class II degenerate T cell epitopes from P. falciparum validates our predictive strategy in a biologically relevant system and supports the potential for developing a broadly efficacious epitope-based vaccine against malaria focused on a limited number of peptide specificities.
The Journal of Immunology 08/2000; 165(2):1123-37. · 5.52 Impact Factor
[show abstract][hide abstract] ABSTRACT: Previous studies indicated that the Plasmodium yoelii circumsporozoite protein (PyCSP) 57-70 region elicits T cells capable of eliminating infected hepatocytes in vitro. Herein, we report that the PyCSP58-67 sequence contains an H-2(d) binding motif, which binds purified K(d) molecules in vitro with low affinity (3, 267 nM) and encodes an H-2(d)-restricted cytotoxic T lymphocyte (CTL) epitope. Immunization of BALB/c mice with three doses of a multiple antigen peptide (MAP) construct containing four branches of amino acids 57 to 70 linked to a lysine-glycine core [MAP4(PyCSP57-70)] and Lipofectin as the adjuvant induced both T-cell proliferation and a peptide-specific CTL response that was PyCSP59-67 specific, H-2(d) restricted, and CD8(+) T cell dependent. Immunization with either DNA encoding the PyCSP or irradiated sporozoites demonstrated that this CTL epitope is subdominant since it is not recognized in the context of whole CSP immunization. The biological relevance of this CTL response was underlined by the demonstration that it could mediate genetically restricted, CD8(+)- and nitric-oxide-dependent elimination of infected hepatocytes in vitro, as well as partial protection of BALB/c mice against sporozoite challenge. These findings indicate that subdominant epitopes with low major histocompatibility complex affinity can be used to engineer epitope-based vaccines and have implications for the selection of epitopes for subunit-based vaccines.
Infection and Immunity 07/2000; 68(6):3403-11. · 4.07 Impact Factor
[show abstract][hide abstract] ABSTRACT: The HLA class II-restricted T-cell response to hepatitis C virus (HCV) antigens is believed to influence the final outcome of hepatitis C, because it is vigorous in patients who recover from acute hepatitis C, but it is weak in those who develop a chronic infection. For this reason, exogenous stimulation of T-cell responses in chronic HCV infection may represent a strategy to cure patients with chronic hepatitis C by approximating the vigor of their T-cell reactivity to that of patients who succeed in recovering from hepatitis. It may also be a preventive approach to avoid spread of the virus by facilitating the development of a vigorous protective response at the very early stages of infection. T-cell-based vaccines composed of immunodominant, promiscuous, and conserved T-cell epitopes may represent a powerful tool to achieve optimal stimulation of the T-cell reactivity. To identify HLA class II-restricted T-cell epitopes useful for this purpose, 22 subjects with acute HCV infection were studied and followed for an average time of 29 months. Eight of them recovered from hepatitis, and 14 developed a chronic infection. Overlapping 20-mer peptides covering the entire core and NS4 antigens and a panel of peptides representing highly conserved regions of core, NS3, NS4, and NS5 were used. By direct peripheral blood T-cell stimulation and by fine-specificity analysis of HCV-specific T-cell lines and clones, highly immunogenic T-cell epitopes were identified within core, NS3, and NS4. All these epitopes are immunodominant and highly conserved among the known HCV isolates. Moreover, they are promiscuous, because they can be presented to T cells by different HLA class II molecules. Immunodominance, sequence conservation, and promiscuity make these epitopes ideal components of preventive or therapeutic T-cell-based vaccines against HCV.
[show abstract][hide abstract] ABSTRACT: The T cell repertoire is shaped by the processes of positive and negative selection. We have previously shown that mice are tolerant to a native self-Ag, mouse lysozyme (ML), but they respond vigorously when challenged with different ML peptides ("cryptic" self-determinants). In this study, we have addressed the issue of the physiological significance of both the hierarchy (dominance/crypticity) of self-determinants within ML and the anti-cryptic, self (ML)-directed T cell repertoire. Our results demonstrate that there are several ML peptides that bind well to MHC but are totally nonimmunogenic when tested for proliferative T cell response and cytokine secretion: a subset of these peptides presumably represent the originally dominant self-determinants of ML, which have rendered the T cells tolerant during thymic selection. Other ML peptides, which bind well to MHC and are immunogenic, correspond to the cryptic determinants of ML: T cells against cryptic ML determinants escape tolerance induction. Thus, the mature T cell repertoire against ML bears the direct imprint of the hierarchy of self (ML)-determinants. Interestingly, hen egg white lysozyme could prime T cells in vivo that were cross-reactive with certain cryptic ML determinants, and vice versa, without requiring any coimmunization with the foreign lysozyme and ML peptide(s). Moreover, repeated, deliberate priming and expansion of T cells by hen egg white lysozyme immunization concomitantly enhanced T cell response to such cross-reactive ML determinants. This reciprocal self-foreign determinant cross-reactivity may play a previously unrecognized, but crucial, role in the expansion and diversification of self-reactive clones in the autoimmune response.
The Journal of Immunology 11/1999; 163(8):4232-7. · 5.52 Impact Factor
[show abstract][hide abstract] ABSTRACT: Celiac disease (CD) patients usually express a DQ2 heterodimer, whose chains DQalpha1*0501/DQbeta1*0201, are encoded by the genes HLA-DQA1*0501 and DQB1*0201, respectively. Among the DQ2 carriers, the risk of developing disease was shown to correlate with the number of DQbeta1*0201 chains encoded. Studying two separate cohorts of Italian and Tunisian patients, we now show a significant association of celiac disease with expression of either the DQ2 or DR53 heterodimers. The risk is maximal for individuals that carry both DQ2 and DR53 heterodimers. When twenty synthetic peptides overlapping most of A-gliadin sequence were tested for the binding to various purified DR molecules, it was found that DR53 molecules bind selectively and with high affinity (IC50<1 microM) to A-gliadin-derived peptides. These data suggest that both HLA DQ2 and DR53 molecules are associated with increased genetic risk for CD, and provide a possible biochemical basis for this complex association.
[show abstract][hide abstract] ABSTRACT: The normal functioning immune system is programmed to attack foreign pathogens and other foreign proteins while maintaining tolerance to self-proteins. The mechanisms by which tolerance is broken in the initiation of autoimmunity are not completely understood. In the present study, mice immunized with the murine cytochrome c peptide 90-104 showed no response by the B or T cell compartments. However, immunization with the isoaspartyl form of this peptide, where the linkage of Asp(93) to Leu(94) occurs through the beta-carboxyl group, resulted in strong B and T cell autoimmune responses. Antibodies elicited by immunization with the isoaspartyl form of self-peptide were cross-reactive in binding to both isoforms of cytochrome c peptide and to native cytochrome c self-protein. In a similar manner, immunization of mice with the isoaspartyl form of a peptide autoantigen of human systemic lupus erythematosus (SLE) resulted in strong B and T cell responses while mice maintained tolerance to the normal aspartyl form of self-antigen. Isoaspartyl linkages within proteins are enhanced in aging and stressed cells and arise under physiological conditions. These post-translationally modified peptides may serve as an early immunologic stimulus in autoimmune disease.
Journal of Biological Chemistry 09/1999; 274(32):22321-7. · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: CTL responses against multiple hepatitis C virus (HCV) epitopes were detected in 7 of 29 (24.1%) healthy family members (HFM) persistently exposed to chronically HCV-infected patients (HCV-HFM). These precursor CTL were at very low or undetectable frequencies, as determined by limiting dilution analysis. However, when HCV-specific effector CD8+ T cells, freshly isolated from PBMC of HCV-HFM, were assessed by a sensitive enzyme-linked immunospot assay, their frequencies were severalfold higher than those of precursor CTL. These results indicate that the two assays detect two functionally distinct T cell populations and that the effector cells are not assayed by the 51Cr-release assay. Furthermore, the combination of cell depletion and enzyme-linked immunospot analyses showed that the effector cells were confined into a CD8+ CD45RO+ CD28- population. The persistence of effector CD8+ T cells specific for both the structural and nonstructural viral proteins in uninfected HCV-HFM, suggest that: 1) an immunological memory is established upon a subclinical infection without any evidence of hepatitis, in a large cohort of HCV-exposed individuals; 2) because these cells required neither restimulation nor the addition of particular cytokines in vitro for differentiating in effectors, they should be capable of prompt HCV-specific effector function in vivo, possibly providing antiviral protection; and 3) the maintenance of effector T cell responses may be sustained by persisting low-level stimulation induced by inapparent infections.
The Journal of Immunology 07/1999; 162(11):6681-9. · 5.52 Impact Factor
[show abstract][hide abstract] ABSTRACT: Molecular mimicry refers to structural homologies between a self-protein and a microbial protein. A major epitope of myelin basic protein (MBP), p87-99 (VHFFKNIVTPRTP), induces experimental autoimmune encephalomyelitis (EAE). VHFFK contains the major residues for binding of this self-molecule to T cell receptor (TCR) and to the major histocompatibility complex. Peptides from papilloma virus strains containing the motif VHFFK induce EAE. A peptide from human papilloma virus type 40 (HPV 40) containing VHFFR, and one from HPV 32 containing VHFFH, prevented EAE. A sequence from Bacillus subtilis (RKVVTDFFKNIPQRI) also prevented EAE. T cell lines, producing IL-4 and specific for these microbial peptides, suppressed EAE. Thus, microbial peptides, differing from the core motif of the self-antigen, MBPp87-99, function as altered peptide ligands, and behave as TCR antagonists, in the modulation of autoimmune disease.
Journal of Experimental Medicine 05/1999; 189(8):1275-84. · 13.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: Experimental leishmaniasis offers a well characterized model of T helper type 1 cell (Th1)-mediated control of infection by an intracellular organism. Susceptible BALB/c mice aberrantly develop Th2 cells in response to infection and are unable to control parasite dissemination. The early CD4(+) T cell response in these mice is oligoclonal and reflects the expansion of Vbeta4/ Valpha8-bearing T cells in response to a single epitope from the parasite Leishmania homologue of mammalian RACK1 (LACK) antigen. Interleukin 4 (IL-4) generated by these cells is believed to direct the subsequent Th2 response. We used T cells from T cell receptor-transgenic mice expressing such a Vbeta4/Valpha8 receptor to characterize altered peptide ligands with similar affinity for I-Ad. Such altered ligands failed to activate IL-4 production from transgenic LACK-specific T cells or following injection into BALB/c mice. Pretreatment of susceptible mice with altered peptide ligands substantially altered the course of subsequent infection. The ability to confer a healer phenotype on otherwise susceptible mice using altered peptides that differed by a single amino acid suggests limited diversity in the endogenous T cell repertoire recognizing this antigen.
Journal of Experimental Medicine 05/1999; 189(7):1111-20. · 13.21 Impact Factor
[show abstract][hide abstract] ABSTRACT: Using a bipalmitoylated lipopeptide consisting of an ovalbumin helper T-cell epitope covalently linked to an influenza virus cytotoxic T-lymphocyte (CTL) epitope, we addressed possible factors that may be critical for CTL induction. Antigen processing of lipopeptide appears to be required for T-cell induction since there was virtually no in vitro binding of lipopeptide to purified MHC molecules. A major portion of lipopeptide immunogenicity was due to its particulate nature inasmuch as CTL induction in mice correlated with insoluble lipopeptide constructs, whereas more soluble analogs were significantly less immunogenic. Immunohistological analysis of tissue from immunized animals revealed that lipopeptide migration from the s.c. injection site to the spleen could be detected as early as 1 h after immunization and cell-associated lipopeptide was observed on macrophages and dendritic cells, implicating both cell populations in the processing and presentation of lipopeptide particles to CTLs.
[show abstract][hide abstract] ABSTRACT: The inverse relationship between peripheral blood CTL responsiveness to multiple hepatitis C virus (HCV) epitopes and viral titer in patients with persistent HCV infection suggests that enhancement of the CTL response might result in viral clearance. Since several HLA-A2-restricted HCV CTL epitopes are already known, we aimed to identify CTL epitopes restricted by other HLA types in an effort to expand the epitope repertoire available for T cell-mediated therapeutic vaccine development. Scanning of 14 different HCV genome sequences for the presence of conserved peptides containing the HLA-A3 and -B7 motifs revealed 9- to 10-mer peptides that were synthesized and assayed for binding to HLA-A3, -B7 supertype molecules. Peptides with good HLA-binding affinities and cross-reactivities with at least three of five most common molecules of each supertype were tested for the ability to stimulate a memory CTL response in the peripheral blood from selected HCV-infected patients and normal seronegative donors in vitro. We identified eight HLA-A3 supertype-restricted CTL epitopes and one HLA-B7 supertype-restricted CTL epitope that were recognized by infected patients but not by healthy seronegative donors. HLA class I serotyping of 158 chronically infected patients revealed that 80% expressed one or more of HLA molecules belong to either the A2, A3, or B7 supertypes. In conclusion, the epitopes, herein identified combined with previously defined HLA-A2-restricted CTL epitopes, should be useful for the design of an ethnically unbiased, therapeutic CTL vaccine for the treatment of patients with chronic HCV infection.
The Journal of Immunology 02/1999; 162(2):1156-64. · 5.52 Impact Factor
[show abstract][hide abstract] ABSTRACT: The human melanocyte lineage-specific antigen gp100 contains several epitopes recognized by cytotoxic T lymphocytes (CTL). However, most of the epitopes reported to date are HLA-A2.1-restricted. Despite the high frequency of HLA-A2.1 in melanoma patients, effective population coverage requires the identification of epitopes restricted by other frequent HLA alleles. Herein, HLA-A3 binding, gp100-derived synthetic peptides were tested for their capacity to elicit anti-melanoma CTL in vitro using CD8+ T cells from healthy donors as responders and peptide-pulsed autologous dendritic cells as antigen-presenting cells. Of 7 peptides tested, 2 (gp100[9(87)] and gp100[10(86)]) induced CTLs that killed melanoma cell lines expressing HLA-A3 and gp100. Additional MHC-binding studies to various HLA molecules belonging to the HLA-A3 superfamily (HLA-A*1101, -A*3101, -A*3301 and -A*6801) were performed to determine whether these CTL epitopes could further increase potential population coverage. Further experiments indicated that the peptide gp100[9(87)], which bound to HLA-A11 with high affinity, was capable of inducing specific CTLs that killed melanoma cells expressing gp100 and HLA-A11 molecules. Our results indicate that the gp100[9(87)] peptide corresponds to a CTL epitope which may be restricted by either the HLA-A3 or HLA-A11 allele, emphasizing its utility for the design and development of epitope-based therapies for melanoma.
International Journal of Cancer 12/1998; 78(4):518-24. · 6.20 Impact Factor
[show abstract][hide abstract] ABSTRACT: Tyrosinase-related protein 2 (TRP2) is a melanosomal enzyme expressed in most mammalian melanocytes and melanomas. This protein has been identified as a melanoma antigen recognized by tumor reactive CTLs derived from tumor infiltrating lymphocytes in the context of HLA-A31 and HLA-A33. The frequencies of these HLA-A alleles among melanoma patients in the United States is low (approximately 6% for HLA-A31 and approximately 2% for HLA-A33) compared with that of HLA-A*0201 (approximately 46%). Therefore, to extend significantly the use of TRP2-based immunotherapies for the treatment of patients with melanoma, we searched for new HLA-A*0201-restricted epitopes from this protein by screening TRP2-derived peptides for the induction of melanoma-reactive CTL. Fifty-one peptides were selected from TRP2 based on a permissive HLA-A*0201 binding motif, and the 21 peptides with the highest experimentally determined binding affinities were used to stimulate peripheral blood lymphocytes from HLA-A*0201+ melanoma patients in vitro. One peptide, TRP2(180-188) (SVYDFFVWL), induced CTLs from three of four patients that specifically recognized peptide-pulsed T2 cells, COS-7 cells expressing HLA-A*0201 and TRP2, and HLA-A2+ TRP2+ melanomas. TRP2(180-188) is identical to a previously identified TRP2 epitope recognized by murine melanoma-reactive CTLs in the context of H-2Kb. These results suggest that TRP2 may be useful for the development of murine tumor immunotherapy models and for the treatment of melanoma patients who are diverse in HLA expression.
Cancer Research 12/1998; 58(21):4895-901. · 8.65 Impact Factor