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* Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Lausanne Branch, University Hospital, Lausanne, Switzerland;
Molecular Statistics and Bioinformatics Section, Biometric Research Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892;
Ludwig Institute for Cancer Research, Lausanne Branch, Lausanne, Switzerland; and
Torrey Pines Institute for Molecular Studies and Mixture Sciences, San Diego, CA 92121
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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TCR. The number of different TCR that
the immune system is able to generate through a sophisticated
recombination process is theoretically large enough to ensure
recognition of any potential complex (1). However, several
factors limit both the composition and the size of the actual TCR
repertoire. Important constraints to its composition are imposed during
thymic selection. Immature thymocytes are "educated" by antigenic
self-peptide/MHC (pMHC) complexes presented on thymic APC
(2). Thymocytes recognizing self complexes with high
affinity are deleted, whereas those recognizing them with low affinity
are selected to undergo further maturation (3). Thus,
recognition of a limited number of self complexes tremendously
restricts and shapes the TCR repertoire, with the proportion of
positive-selected thymocytes not higher than 5% of the total
(4). The size of the TCR repertoire is first limited by
the number of T cells present in an animal
(
1012 in an adult human being). The number of
distinct specificities in the TCR repertoire is further limited by the
fact that, to mount an appropriate immune response to a given antigenic
determinant, a certain number of specific T cells have to be present in
the repertoire. Indeed, it has been shown that, in a single individual,
many different clonotypes usually contribute to immune responses
against an antigenic determinant (5). Therefore, it is becoming increasingly clear, both from theoretical considerations and experimental evidence, that a high degree of cross-reactivity is a normal feature of the TCR and constitutes an essential characteristic of T cell recognition (6). However, both the molecular nature and the extent of the cross-reactivity between the pool of natural antigenic determinants and the T cell repertoire are not immediately obvious and most likely vary widely for each antigenic determinant and for each TCR. Indeed, even if flexible, T cell recognition remains exquisitely specific and very small changes can profoundly affect it (7). These changes do not necessarily need to be located in TCR contact residues of the antigenic peptide but can be due to more-or-less subtle modifications of the MHC/peptide complex conformation, such as variations in the conformation that the peptide itself adopts in the MHC molecule, or even small peptide-dependent variations of the MHC molecule (8, 9, 10). Moreover, as the relevant conformation can be mimicked by apparently completely unrelated ligands, cross-reactivity does not necessarily imply sequence homology (11, 12). As a result of this complexity, it is very difficult to predict whether, and to what extent, one antigenic determinant will cross-react with another.
An empirical approach for the identification of antigenic determinants cross-recognized by Ag-specific T cells is based on the analysis of the recognition by clonal T cells of synthetic combinatorial peptide libraries in positional scanning format (PS-SCL5; Ref. 13). We have recently elaborated a biometric analysis that uses the data generated by screening PS-SCL with T cell clones to analyze and rank all peptides in public protein databases for sequences with predicted reactivity (14). In this study, we have analyzed the recognition by tumor-reactive CTL specific for an immunodominant peptide from the melanocyte differentiation and tumor-associated Ag Melan-A (15), of peptides retrieved in proteins of self or pathogen origin by using biometric analysis of the data generated by screening of PS-SCL with a Melan-A-specific clone.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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A scoring matrix was generated by using data previously obtained by screening an amidated C terminus decapeptide PS-SCL (16) with the Melan-A-specific CTL clone LAU 203/1.5 in a functional chromium-release assay. A Z-scoring matrix was generated using the average and SD of the percentage of specific-lysis values of multiple experimental data obtained for each mixture defined with one of the 20 L-amino acids in each of the 10 positions of the decamer library (14). Based on the assumption of independent and additive contribution of the individual amino acids at each position of a peptide to the peptide’s activity, the score of each individual peptide was calculated by adding individual stimulatory values of the composing amino acids. A program was designed to use the matrix to score all overlapping decapeptides contained in the GenPept protein database (ftp://ftp.ncicfr.gov/pub/genpept) and thus identify sequences with the highest predicted stimulatory scores (14).
Cells
The Melan-A-specific CTL clone LAU 203/1.5 was derived from tumor-infiltrated lymph node cells of patient LAU 203 by limiting dilution culture in the presence of irradiated allogeneic PBMC, PHA, and human (h)rIL-2 (150 IU/ml; Glaxo, Geneva, Switzerland; kindly provided by Dr. M. Nabholz, Institut Suisse de Recherches Experimentales sur le Cancer, Epalinges, Switzerland) as previously described (17). Cells were subsequently expanded by periodic (3- to 4-wk) restimulation in microtiter plates together with irradiated feeder cells in the presence of PHA and hrIL-2. Polyclonal Melan-A monospecific CTL lines and other Melan-A specific clones were similarly generated from Melan-A multimer+-sorted cells as previously described (18). All selected populations were able to specifically lyse Melan-A-expressing melanoma tumor cells. For peptide stimulation of PBMC, CD8+ lymphocytes were positively selected by magnetic cell sorting from PBMC of A2 melanoma patients using a miniMACS device (Miltenyi Biotec, Sunnyvale, CA). Cells of the CD8- fraction were irradiated (3000 rad) and used as autologous APC. CD8+ highly enriched lymphocytes (1 x 106) were stimulated with peptide (1 µg/ml) and irradiated autologous APC in 2 ml of IMDM (Life Technologies, Basel, Switzerland) containing 8% pooled human serum, hrIL-2 (100 IU/ml), and hrIL-7 (10 ng/ml, R&D Systems, Oxon, U.K.). Cells underwent an additional cycle of stimulation with peptide-pulsed APC before analysis.
Ag-recognition assay
Ag recognition was assessed in chromium-release experiments. Briefly, target cells were labeled with 51Cr for 1 h at 37°C and washed three times. Labeled target cells (1000 cells/well) were incubated in the presence of peptide libraries (100 µg/ml final) or, in peptide titration experiments, with various peptide doses, for 15 min at room temperature before the addition of effector cells at a lymphocyte-to-target cell ratio of 10:1. Chromium release was measured in supernatants harvested after 4-h incubation at 37°C. The percentage of specific lysis was calculated as: 100 x [(experimental - spontaneous release)/(total - spontaneous release)]
Multimers, mAbs, and flow cytometry analysis
A2/peptide multimers incorporating the Melan-A26–35 A27L analog (8) or peptides MSI-44–10 (PT 178–187), MSI-44–25 (PV 454–463), and MSI-44–56 (CT 51–60) were synthesized as described previously (19). Cells were stained with multimersPE (4.5 µg/ml in 20 µl) for 1 h at room temperature, before the addition of anti-CD8FITC mAb (BD Biosciences, San Jose, CA) and further incubated for an additional 30 min at 4°C. At the end of the incubation period, cells were washed and analyzed by flow cytometry on a FACScan (BD Biosciences). Data analysis was performed using the CellQuest software.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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In a previous study, we used the Melan-A-specific and
tumor-reactive CTL clone LAU 203/1.5 to screen an amidated C terminus
decapeptide PS-SCL, (16) in a functional chromium release
assay (CTL assay). Based on those results, in this study, we performed
a biometric data analysis (as recently reported (14) and
further detailed in Materials and Methods) that allows the
use of the results of the PS-SCL screening to search for potentially
cross-reactive sequences in public protein databases. As illustrated in
Table I
, our search retrieved 21 peptides
of self origin and 25 peptides of viral origin with higher predicted T
cell stimulatory values (scores) than the parental
Melan-A26–35 decapeptide (MSI-44–22, Table I).
These peptides were selected for analysis. Among peptides of bacterial
origin, a high number (280) scored higher than
Melan-A26–35. Of those, the first 25 peptides
were selected for analysis. For consistency with the PS-SCL used for
the screening, peptides were synthesized with amidated C termini.
Recognition of the selected peptides by the CTL clone LAU 203/1.5 was
initially tested at a single peptide dose (1 µg/ml) in a CTL assay.
An example of the data obtained is illustrated in Fig. 1A. Data obtained for all
peptides tested and a summary thereof are reported in Fig. 1, B and C, respectively. Specific lysis of >10%
was considered significant, because values <10% were consistently
obtained for a group of 100 unrelated peptides similarly tested (data
not shown). Thirty-nine percent of the selected peptides were
significantly recognized by clone LAU 203/1.5 (Fig. 1C). It
is noteworthy that the proportion of peptides recognized was higher
among those of bacterial origin than those of self or viral origin. The
relative efficiency of recognition of the selected peptides was further
assessed by performing peptide titrations as illustrated in Fig. 2A. Results are summarized in
Fig. 2B. Of the 28 peptides recognized by clone LAU 203/1.5,
12 were recognized with roughly comparable efficiency (relative
recognition <10 but >0.1), as compared with Melan-A
26–35; 4 were recognized significantly more
efficiently (relative recognition of 
10); and the remaining 12 were
recognized significantly less efficiently (relative recognition of
0.1). Cross-reactivity between amidated and free C termini peptides
was tested for a group of peptides selected among the most active ones.
As summarized in Fig. 2C and consistent with our previous
data (20), peptides bearing free C termini were generally
recognized as well as or more efficiently than their amidated C termini
counterparts, although to a variable extent.
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To elucidate the factors impacting on the recognition of the
selected peptides by clone LAU 203 1.5, their potential binding
capacity to A2 was first determined using an algorithm that calculates
the theoretical dissociation rate of the corresponding
A2/peptide complexes based on their amino acid
composition (available at the Bioinformatics and Molecular analysis
section of the National Institutes of Health website,
http://bimas.dcrt.nih.gov/molbio/hla_bind/) (21). As
expected from both the absence of canonical "anchor residues" and
our previous experimental data (8, 17, 22), peptide
Melan-A26–35 was predicted as a relatively poor
binder (theoretical t1/2 of
2m dissociation was around 3300-fold lower
than for a known good A2 binder, peptide
Flu-MA58–66, supplemental data, Table II). Among the selected peptides, 35%
had a relative predicted binding lower than that of Melan-A
26–35, whereas the majority (65%) had a similar
or higher (up to >20,000-fold) predicted binding value. Interestingly,
the frequency of peptides recognized by clone LAU 203/1.5 was high
(89%; Fig. 3A) among peptides
with relative predicted binding equal to or greater than that of
Melan-A26–35. Indeed, only 13% of peptides with
predicted binding lower than that of
Melan-A26–35 were significantly recognized,
whereas about half (47%) of peptides with predicted binding equal to
or greater than that of Melan-A26–35 were
significantly recognized (Fig. 3A). In addition, all of the
peptides that were recognized more efficiently than the parental
Melan-A peptide had a higher predicted A2 binding value (Fig. 3B). Relative predicted binding was independent from peptide
score (the mean score was similar in the two groups, Fig. 3C), consistent with the peptide selection procedure used
that was, in this sense, completely unbiased. However, when peptides
were divided into two groups according to whether their scores were
higher or lower than the mean score of all peptides tested (Fig. 3D), 70% of the peptides in the high scoring group were
recognized. In addition, 75% of the peptides that were recognized more
efficiently than the parental Melan-A peptide belonged to the group
with the highest scores (Fig. 3E). To compare predicted to
experimental binding, experimental binding was assessed in a functional
competition assay as described previously (17) for
peptides available in the terminal C-free form (Fig. 2
C).
The experimental values obtained were consistent with predicted binding
values (not shown).
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We next investigated whether cross-recognition of the selected
peptides was found only for clone LAU 203/1.5 or also for other
Melan-A-specific CTL. To this end, we tested their recognition by a
large panel of Melan-A-monospecific CTL lines (18) and
clones derived from four melanoma patients (LAU 203, LAU 337, LAU 455,
and LAU 465) and one healthy donor (HD 421). Our previous studies of
the TCR repertoire of Melan-A-specific CTL have shown that the
latter is extremely large and diverse and mostly nonoverlapping
between different individuals (5, 23). Thus, the majority
of Melan-A-specific CTL clones used in this study are likely to express
different TCR. In addition, TCR sequences were previously determined in
the case of CTL clones from LAU 337 (24) and HD 421
(25), and all were found to be distinct. Results obtained
are shown in Table III and summarized in
Tables IV and V. Surprisingly, we found a
degree of cross-reactivity that was
higher than that of clone LAU 203/1.5.
Indeed, the proportion of peptides recognized by at least one of the
CTL populations analyzed accounted for 94% of the total (Table IV).
Melan-A-specific clones largely differed in terms of their ability to
cross-recognize the peptides, ranging from no detectable
cross-recognition by clone LAU 337/6C3 to recognition of 68% (48/71)
of the peptides by clone LAU 337/2A5 (Table V). Polyclonal monospecific
lines generally displayed a higher degree of cross-recognition as
compared with CTL clones, possibly because of the combined reactivity
of clonal populations composing these lines. The polyclonal
monospecific population derived from patient LAU 203 cross-recognized a
higher proportion of peptides (68%) than did clone LAU 203/1.5. It is
of note that, in contrast with the data obtained for clone LAU 203/1.5
(Fig. 3A), there was no correlation between the ability of
the peptides to be recognized by different Melan-A-specific CTL and
their A2-binding capacity (Fig. 3F). Interestingly, some
peptides were recognized by the majority of CTL (i.e., MSI-44–25,
Table IV). To further explore the extent of cross-reactivity between
these peptides (in particular those most frequently recognized by
Melan-A-specific CTL) and Melan-A, we synthesized A2/peptide multimers
incorporating three of the most frequently recognized peptides, one of
self origin (peptide PT 178–187 from PG transporter), one of viral
origin (peptide PV 454–463 from the glycoprotein GIII precursor of
pseudorabies virus), and one of bacterial origin (peptide CT 51–60
from the arginine/ornithine antiporter of Chlamydia
trachomatis). These multimers specifically stained clone LAU
203/1.5 but not clones of unrelated specificity; in addition, they were
able to specifically stain ex vivo a significant proportion of
circulating CD8+ T cells from patient LAU 203,
similar to what had been observed previously with multimers
incorporating a Melan-A26–35 analog A27L (Ref.
26 and data not shown).
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To determine whether the peptides frequently cross-recognized by
Melan-A-specific CTL could elicit CTL responses cross-reactive with
Melan-A, we used three of the most frequently cross-recognized peptides
to stimulate in vitro highly enriched CD8+ T
lymphocytes from melanoma patients. As internal controls, we included
CD8+ T cells cultured under the same conditions,
but in the absence of any exogenously added peptide, or stimulated with
Melan-A26–35 parental peptide or with the analog
Melan-A26–35 A27L. Consistent with our previous
data (27), stimulation with the analog
Melan-A26–35 A27L elicited a specific response
significantly higher than that observed with the Melan-A26–3
5 parental peptide for all patients (Fig. 4, A and B).
Importantly, as illustrated in Fig. 4A for patient LAU 241,
each of the three selected peptides elicited an even stronger response
when tested with multimers incorporating the peptide used for in vitro
stimulation (autologous multimers), whereas a lower, but very
significant, proportion of CD8+
Melan-A26–35 A27L
multimer+ cells were detected in each of these
cultures. Similar results were obtained in the case of two additional
melanoma patients (Fig. 4B) and two healthy donors (not
shown). To further dissect the heterogeneity of the CTL response
elicited by Melan-A related peptides, CD8+
autologous multimer+ cells from each culture were
isolated by cell sorting, expanded in vitro with PHA, and stained again
with multimers incorporating either the autologous peptide or peptide
Melan-A26–35 A27L. In each case, the resulting
populations were homogeneously stained by autologous multimers, whereas
only part of these cells were stained by multimers incorporating
peptide Melan-A26–35 A27L (Fig. 5A).
CD8+ Melan-A26–35 A27L
multimer+ and multimer-
cell fractions were further isolated from each culture by cell sorting,
and their activity was analyzed in CTL assay. As illustrated in Fig. 5B for one of the cultures and summarized in Fig. 5C for all tested cultures from two melanoma patients,
CD8+ Melan-A26–35 A27L
multimer+ fractions cross-recognized peptide
Melan-A26–35 A27L much more efficiently than
CD8+ Melan-A26–35 A27L
multimer- fractions. In contrast, the peptide
used for the in vitro stimulation was generally recognized by both
fractions with roughly similar efficiency (Fig. 5C).
Importantly, a low but significant level of specific lysis of
Melan-A+ A2+ tumor cells
was detected for the majority of the tested cultures and mostly
segregated with the CD8+
Melan-A26–35 A27L
multimer+ fractions (Fig. 5D).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Validation of the high predictive value of our analytical approach came from the finding that a high proportion of the peptides analyzed, including sequences with the highest stimulatory scores of either self, viral, or bacterial origin, were recognized by the CTL clone used for the initial PS-SCL screening. It is noteworthy that recognition was highly specific as none of 100 unrelated peptides was significantly recognized (not shown). Reactive peptides were recognized with variable efficiency; remarkably, however, more than half of them were recognized with similar or higher efficiency as compared with the Melan-A parental decapeptide. Further support for the validity of our approach came from the correlation found between predicted stimulatory scores and peptide recognition. It is of note that the most active peptides were identified among peptides of bacterial origin, in good agreement with the higher predicted stimulatory score of this group. In addition, and interestingly, we also found a clear correlation between the relative predicted A2 binding efficiency of the retrieved peptides and their recognition. Thus, although a good A2 binding did not necessarily predict cross-recognition, omitting from the screening peptides with extremely poor predicted binding capacity would have further improved the likelihood of identifying peptides cross-recognized by the CTL clone used for the PS-SCL screening.
A surprising finding of this study was that a much higher proportion of peptides were cross-recognized by Melan-A-specific tumor-reactive CTL other than clone LAU 203/1.5 as compared with clone LAU 203/1.5 itself. However, the extent of cross-recognition was extremely variable among clones, ranging from no detectable cross-recognition (i.e., for clone LAU 337/6C3) to cross-recognition of 68% of the analyzed peptides (i.e., for clone LAU 337/2A5). Remarkably, these two clones recognized Melan-A peptides with comparable efficiency (24); thus, a difference in functional avidity of Ag recognition did not account for the different extent of cross-recognition, which could be due to differences in the fine specificity of Ag recognition of the clones analyzed. However, an alternative explanation could be that the different Melan-A-specific clones analyzed differ in terms of "degeneracy" of Ag recognition, e.g., in their capability to productively interact with a more-or-less large number of MHC/peptide complexes. The concept of "degeneracy" of Ag recognition (13) originates from the elucidation of the molecular interactions leading to T cell activation. It is based on the fact that, as underlined by the close inspection of crystal structures of TCR complexes with their MHC/peptide ligands (28, 29), most of the TCR contacts with the MHC/peptide complex are made with the MHC molecule rather than with the peptide. Thus, the majority of the TCR-MHC/peptide complex binding energy is determined by the affinity of a given TCR for the MHC molecule, the role of the peptide being only to modulate the latter and possibly to raise it beyond the threshold required for T cell activation. It follows that, if the TCR affinity for the MHC molecule is relatively low, the threshold can be attained with only few peptides and Ag recognition appears as highly selective. If, instead, the TCR affinity for the MHC is relatively high, activation is achieved by a relatively high number of different peptides and the TCR appears more "degenerate." Whether, and to what extent, peptides cross-recognized by the more degenerate TCR need to be structurally related is yet undetermined; however, it can be expected that at least a fraction of these peptides share some common structural and/or sequence features. In agreement with this hypothesis, while the strategy used to find Melan-A cross-reactive peptides was not conducted by searching for sequence homologies, the search appears to have yielded a group of cross-reactive peptides, the majority of which share significant internal homology, particularly in the central part of the peptide (position 3 to 7). Thus, our search may have retrieved a "family" of Melan-A-related sequences, only part of which achieve the threshold of activation for clone LAU 203/1.5, whereas others achieve it only for more degenerate Melan-A-specific CTL.
Related to this is another important question on the modality of T cell cross-recognition. It has indeed not yet been established whether, and to what extent, T cell cross-recognition is focused or unfocused (6), i.e., whether or not distinct T cell clonotypes specific for the same Ag cross-recognize the same set of peptides. Whether cross-reactivity is focused or unfocused has important implications for the potential breaking of self-tolerance as the result of immune responses to foreign Ags. Indeed, if cross-reactivity is unfocused, the fortuitous cross-reaction between a foreign and a self peptide would lead to the expansion of a single or of a limited number of T cell clonotypes, whereas, in the opposite case, it could rapidly lead to a generalized autoimmune reaction. In this regard, the data obtained in this study support the latter hypothesis, as cross-reactivity was, in this case, at least partially focused. Indeed, although some peptides were recognized by only a few clonotypes, others were cross-recognized by the majority of Melan-A-specific CTL. The more frequently recognized peptides shared significant internal homology with Melan-A, particularly in the central part of the peptide. In contrast with the data obtained with the CTL clone used for the initial PS-SCL screening, there was no obvious correlation between the capacity of Melan-A-related sequences to be recognized by different Melan-A-specific CTL and the efficiency of their binding to A2. This indicates that cross-recognition of Melan-A-related peptides by different Melan-A-specific CTL is impacted more by the presence of TCR contact residues in the core region of the peptide than by their A2 binding efficiency. In addition, and importantly, in good agreement with recognition data, stimulation of PBMC from melanoma patients with the more frequently cross-recognized peptides elicited vigorous CTL responses that were at least partially cross-reactive with Melan-A.
It has been previously reported that Melan-A-like sequences are relatively frequent in proteins (30), possibly because the localization of this peptide in the transmembrane region of the Melan-A protein results in an amino acid composition that may be frequent in hydrophobic regions from other proteins. Thus, it could be argued that the high rate of cross-reactive sequences identified in this study is related to the high frequency of Melan-A-like sequences in proteins and that a very different result could be obtained for other epitopes. To address this point we have recently performed a similar analysis for an HLA-A2-restricted epitope from the cancer testis Ag SSX-2 (31). Interestingly, we found that the majority of retrieved sequences were cross-recognized by the CTL clone used for the PS-SCL screening (D. Valmori, manuscript in preparation), thus supporting the applicability of this approach to epitopes other than Melan-A.
What is the physiologic relevance of the Melan-A cross-reactive sequences? Melan-A cross-reactive peptides of self origin that are recognized by Melan-A-specific CTL with comparable or higher efficiency than Melan-A parental peptides are most likely not presented by thymic APC, as this would result in the deletion of Melan-A-specific thymocytes. In contrast, Melan-A-related self sequences that are recognized weakly or are able, when associated to an MHC molecule, to engage the TCR without leading to T cell activation (antagonists) may play a role in thymic positive selection of Melan-A-specific T cell precursors as well in the maintenance of the peripheral pool of Melan-A-specific T cells. Melan-A-related sequences of pathogen origin could be involved in the pathogenesis of vitiligo, a common progressive depigmentary condition of unknown etiology that is due to the autoimmune destruction of skin melanocytes. The spontaneous appearance of vitiligo has been associated with a favorable prognosis in melanoma patients (32, 33). Interestingly, higher frequencies of A2/Melan-A multimer+ cells have been found in individuals with autoimmune vitiligo as compared with healthy controls (34). In this respect, it is noteworthy that our search retrieved 25 viral and 280 bacterial peptide sequences with higher predicted stimulatory values than Melan-A.. Interestingly, some of these sequences were derived from common human pathogens and are thus of clear interest for investigating a potential pathogen-related etiology of vitiligo. Finally, it is of note that the Melan-A cross-reactive peptides identified in this study most likely represent only a small subgroup of the existing ones and are not necessarily among the most relevant. Depending on the likely rapid improvement of relevant technologies, it will be of great interest, in future studies, to extend this analysis to a large number of peptides, possibly selected from the ones that are naturally processed and presented, to unveil the most relevant cross-reactivity between Melan-A and the universe of peptides encoded by human and pathogen genomes.
In conclusion, the findings reported in this study strongly encourage the use of this methodology, which combines PS-SCL screening with T cell clones of interest and includes biometric analysis of the generated data, to further elucidate the molecular basis of T cell cross-reactivity. In addition, the reported data validate the predictive value of this approach for identifying in public protein databases the sequence of natural peptides cross-recognized by clinically relevant T cells.
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Acknowledgments |
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Footnotes |
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2 V.R.-G. and V.D. contributed equally to this work.
3 Address correspondence about the use of combinatorial peptide libraries and biometric analysis to Dr. Clemencia Pinilla, Torrey Pines Institute for Molecular Studies and Mixture Sciences, 3550 General Atomics Court, San Diego, CA 92121. E-mail address: cpinilla{at}tpims.org
4 Address correspondence and reprint requests to Dr. Danila Valmori, Division of Clinical Onco-Immunology, Hôpital Orthopédique, Avenue Pierre-Decker 4, CH-1005 Lausanne, Switzerland. E-mail address: danila.valmori{at}inst.hospvd.ch
5 Abbreviations used in this paper: PS-SCL, synthetic combinatorial peptide libraries in positional scanning format; h, human; A2, HLA-A*0201
Received for publication June 12, 2002. Accepted for publication September 5, 2002.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain usage reveals a large repertoire of Melan-A-specific CD8+ T cells in melanoma patients. J. Immunol. 165:533. T cell receptor structure at 2.5 Å and its orientation in the TCR-MHC complex. Science 274:209.This article has been cited by other articles:
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  V. Voelter, N. Rufer, S. Reynard, G. Greub, R. Brookes, P. Guillaume, F. Grosjean, T. Fagerberg, O. Michelin, S. Rowland-Jones, et al. Characterization of Melan-A reactive memory CD8+ T cells in a healthy donor Int. Immunol., August 1, 2008; 20(8): 1087 - 1096. [Abstract] [Full Text] [PDF]
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 K. Fujio, A. Okamoto, Y. Araki, H. Shoda, H. Tahara, N. H. Tsuno, K. Takahashi, T. Kitamura, and K. Yamamoto Gene Therapy of Arthritis with TCR Isolated from the Inflamed Paw J. Immunol., December 1, 2006; 177(11): 8140 - 8147. [Abstract] [Full Text] [PDF]
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S. Colombetti, T. Fagerberg, P. Baumgartner, L. Chapatte, D. E. Speiser, N. Rufer, O. Michielin, and F. Levy Impact of Orthologous Melan-A Peptide Immunizations on the Anti-Self Melan-A/HLA-A2 T Cell Cross-Reactivity. J. Immunol., June 1, 2006; 176(11): 6560 - 6567. [Abstract] [Full Text] [PDF]
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 L. Chapatte, M. Ayyoub, S. Morel, A.-L. Peitrequin, N. Levy, C. Servis, B. J. Van den Eynde, D. Valmori, and F. Levy Processing of Tumor-Associated Antigen by the Proteasomes of Dendritic Cells Controls In vivo T-Cell Responses. Cancer Res., May 15, 2006; 66(10): 5461 - 5468. [Abstract] [Full Text] [PDF]
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J. Lustgarten, A. L. Dominguez, and C. Pinilla Identification of Cross-Reactive Peptides Using Combinatorial Libraries Circumvents Tolerance against Her-2/neu-Immunodominant Epitope J. Immunol., February 1, 2006; 176(3): 1796 - 1805. [Abstract] [Full Text] [PDF]
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M. Sospedra, P. A. Muraro, I. Stefanova, Y. Zhao, K. Chung, Y. Li, M. Giulianotti, R. Simon, R. Mariuzza, C. Pinilla, et al. Redundancy in Antigen-Presenting Function of the HLA-DR and -DQ Molecules in the Multiple Sclerosis-Associated HLA-DR2 Haplotype J. Immunol., February 1, 2006; 176(3): 1951 - 1961. [Abstract] [Full Text] [PDF]
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 V. A. Judkowski, G. M. Allicotti, N. Sarvetnick, and C. Pinilla Peptides From Common Viral and Bacterial Pathogens Can Efficiently Activate Diabetogenic T-Cells Diabetes, September 1, 2004; 53(9): 2301 - 2309. [Abstract] [Full Text] [PDF]
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