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Pedro Romero - One of the best experts on this subject based on the ideXlab platform.

  • deciphering the unusuAl hlA A2 MelAn A mArt 1 specific tcr repertoire in humAns
    European Journal of Immunology, 2014
    Co-Authors: Pedro Romero, Daniel E. Speiser, Nathalie Rufer
    Abstract:

    The MelAn-A/MART-1(26-35) Antigenic peptide is one of the best studied humAn tumor-AssociAted Antigens. It is expressed in heAlthy MelAnocytes And mAlignAnt MelAnomA And is recognized by CD8(+) T cells in the context of the MHC clAss I molecule HLA-A*0201. While An unusuAlly lArge repertoire of CD8(+) T cells specific for this Antigen hAs been documented, the reAsons for its generAtion hAve remAined elusive. In this issue of the EuropeAn JournAl of Immunology, Pinto et Al. [Eur. J. Immunol. 2014. 44: 2811-2821] uncover one importAnt mechAnism by compAring the thymic expression of the MelAn-A gene to thAt in the MelAnocyte lineAge. This study shows thAt medullAry thymic epitheliAl cells (mTECs) dominAntly express A truncAted MelAn-A trAnscript, the product of misinitiAtion of trAnscription. Consequently, the protein product in mTECs lAcks the immunodominAnt epitope spAnning residues 26-35, thus precluding centrAl tolerAnce to this Antigen. In contrAst, MelAnocytes And MelAnomA tumor cells express Almost exclusively the full-length MelAn-A trAnscript, thus providing the tArget Antigen for efficient recognition by HLA-A2-restricted CD8(+) T cells. The frequency of these AlternAtive gene trAnscription modes mAy be more common thAn previously AppreciAted And mAy represent An importAnt fActor modulAting the efficiency of centrAl tolerAnce induction in the thymus.

  • MelAn-A/MART-1-Specific CD4 T Cells in MelAnomA PAtients: IdentificAtion of New Epitopes And Ex Vivo VisuAlizAtion of Specific T Cells by MHC ClAss II TetrAmers
    Journal of Immunology, 2006
    Co-Authors: Gilles Bioley, Jean-charles Cerottini, Donata Rimoldi, Daniel E. Speiser, Camilla Jandus, Sandra Tuyaerts, William W. Kwok, Jean-marie Tiercy, Kris Thielemans, Pedro Romero
    Abstract:

    Over the pAst decAde, mAny efforts hAve been mAde to identify MHC clAss II-restricted epitopes from different tumor-AssociAted Ags. MelAn-A/MART-1 26–35 pArentAl or MelAn-A/MART-1 26–35(A27L) AnAlog epitopes hAve been widely used in MelAnomA immunotherApy to induce And boost CTL responses, but only one Th epitope is currently known (MelAn-A 51–73 , DRB1*0401 restricted). In this study, we describe two novel MelAn-A/MART-1-derived sequences recognized by CD4 T cells from MelAnomA pAtients. These epitopes cAn be mimicked by peptides MelAn-A 27–40 presented by HLA-DRB1*0101 And HLA-DRB1*0102 And MelAn-A 25–36 presented by HLA-DQB1*0602 And HLA-DRB1*0301. CD4 T cell clones specific for these epitopes recognize MelAn-A/MART-1 + tumor cells And MelAn-A/MART-1-trAnsduced EBV-B cells And recognition is reduced by inhibitors of the MHC clAss II presentAtion pAthwAy. This suggests thAt the epitopes Are nAturAlly processed And presented by EBV-B cells And MelAnomA cells. Moreover, MelAn-A-specific Abs could be detected in the serum of pAtients with meAsurAble CD4 T cell responses specific for MelAn-A/MART-1. Interestingly, even the short MelAn-A/MART-1 26–35(A27L) peptide wAs recognized by CD4 T cells from HLA-DQ6 + And HLA-DR3 + MelAnomA pAtients. Using MelAn-A/MART-1 25–36 /DQ6 tetrAmers, we could detect Ag-specific CD4 T cells directly ex vivo in circulAting lymphocytes of A MelAnomA pAtient. Together, these results provide the bAsis for monitoring of nAturAlly occurring And vAccine-induced MelAn-A/MART-1-specific CD4 T cell responses, Allowing precise And ex vivo chArActerizAtion of responding T cells.

  • MelAn-A/MART-1: Immunogenicity And vAccine strAtegies
    Cancer Immunity Archive, 2003
    Co-Authors: Jean-charles Cerottini, Pedro Romero, Daniel E. Speiser
    Abstract:

    MelAn-A/MART-1 is A representAtive member of the cAtegory of differentiAtion tumor Antigens. It is expressed in normAl MelAnocytes And in the mAjority of primAry And metAstAtic MelAnomAs. The MelAn-A/MART-1 gene encodes A type III protein of 118 Amino Acids thAt is locAlized mAinly in the endoplAsmic reticulum And the trAnsgolgi network (1). Its function remAins unknown. An immunodominAnt HLA-A2 restricted epitope hAs been mApped to residues 26 - 35, which Are locAted within the putAtive trAnsmembrAne region. We hAve designed single Amino Acid substituted peptide AnAlogs with enhAnced Antigenicity And immunogenicity (2) As well As doubly substituted non-nAturAl Amino Acid peptide AnAlogs thAt Are stAble to serum proteAses (3). DetAiled studies of the cytolytic T lymphocyte (CTL) response to the MelAn-A/MART-1 Antigen with A vAriety of AssAys, including the use of fluorescent HLA-A2/peptide AnAlog multimers, hAve reveAled the positive selection of A broAd repertoire of MelAn-A/MART-1 specific CD8 T cells in the humAn thymus And the mAintenAnce of A high frequency of these T cells in the periphery of the mAjority of heAlthy HLA-A2 individuAls, which remAin in A nAive stAte despite the expression of Antigen in skin MelAnocytes, A phenomenon Akin to immunologicAl ignorAnce (4). In contrAst, MelAn-A/MART-1 specific T cells undergo in vivo proliferAtion And differentiAtion in two thirds of HLA-A2 metAstAtic MelAnomA pAtients. High Avidity, Antigen-experienced specific CTLs migrAte And AccumulAte At the tumor sites. However, we hAve observed thAt, while these cells Are functionAlly competent when sAmpled from the peripherAl blood mononucleAr lymphocyte pool, those freshly AssAyed from the tumor sites displAy reduced IFN-gAmmA production in response to Antigen. In Addition, the MelAn-A/MART-1 specific T cells residing in metAstAtic lymph nodes, but not those residing in blood or subcutAneous or viscerAl metAstAses, exhibit virtuAlly no perforin messAge And protein. Of interest, once releAsed from the tumor mAss And trAnsferred to in vitro culture in the presence of tumor cells And cytokines, they rApidly recover normAl functionAl Activity before undergoing proliferAtion (5). We hAve cArried out phAse I clinicAl triAls of vAccinAtion with nAturAl or substituted peptide AnAlogs together with immunologicAl AdjuvAnts in pAtients with metAstAtic MelAnomA. Our strAtegy, which is to rApidly test vArious pArAmeters such As dose, route of AdministrAtion, type of moleculAr AdjuvAnt, with the Aim of selecting strongly immunogenic vAccines, is bAsed on quAntitAtive And quAlitAtive monitoring of MelAn-A/MART-1 Antigen-specific CD8 T-cell responses. A unique AdvAntAge offered by the MelAn-A/MART-1 Antigenic system is the existence of A bAseline frequency of specific T cells thAt is meAsurAble directly ex vivo with fluorescent multimers in the mAjority of the pAtient populAtion. Thus, there is no need to use Any in vitro expAnsion step prior to meAsuring the response. We stAndArdized lAborAtory immune monitoring to reAch stAndArds for medicAl diAgnostic procedures (high reproducibility, low intrA-individuAl vAriAtions, thresholds to distinguish responder pAtients from non-responders). We AnAlyzed four pArAmeters for Antigen specific CD8 T cells in peripherAl blood: frequency of IFN-gAmmA Elispot forming cells, frequency of multimer lAbeled CD8 T cells And the proportion of these T cells expressing the cell surfAce CD45RA And CD28 molecules (6, 7). The shifts of post-vAccine vAlues relAtive to pre-vAccine vAlues for eAch pArAmeter were converted into A combined score thAt weights the mAgnitude of the increAse of the specific T-cell Activity in responder pAtients. SignificAnt ActivAtion of Antigen-specific CD8 T cells wAs detected in PBMCs from 12/17 pAtients vAccinAted with peptides emulsified in minerAl oil-bAsed AdjuvAnt. T-cell responses correlAted significAntly with inflAmmAtory skin reActions At vAccine injection sites, demonstrAting thAt immune monitoring dAtA were relevAnt for in vivo ongoing immunity. RepeAted vAccinAtion resulted in enhAnced T-cell Activity in >90 % of the responder pAtients, indicAting thAt vAccine induced Activity wAs sustAined. Although All pAtients were vAccinAted with A mixture of MelAn-A And influenzA peptides, responses to the two Antigens occurred independently. Thus, non-responsiveness wAs Antigen-specific rAther thAn pAtient specific. Interestingly, vAccine responsiveness wAs significAntly more frequent in pAtients with Antigen-specific CD8 T cells thAt were pre-ActivAted before vAccinAtion. Therefore, endogenous stimulAtion by tumor derived Antigen contributed to T-cell ActivAtion observed After immunotherApy. This is in line with T-cell ActivAtion being A process requiring severAl triggering steps, And indicAtes thAt vAccinAtion with peptide in conjunction with minerAl oil-bAsed AdjuvAnt is cApAble to promote some, but not All, of these steps. An essentiAl pArAmeter thAt remAins to be Assessed in the context of vAccine-induced MelAn-A/MART-1 specific T-cell responses is thAt of the Avidity of responding T cells. Thus fAr the estimAtion of TCR Avidity hAs relied on indirect functionAl AssAys. The AvAilAbility of mutAnt multimers offers the possibility to develop direct AssAys of TCR Avidity. We hAve developed new multimeric MHC clAss I/peptide reAgents Allowing to visuAlize low, intermediAte And high Avidity T cells (8). In conclusion, ex vivo monitoring of the response elicited by MelAn-A/MART-1 peptide-bAsed vAccines provides precise quAntitAtive dAtA on T-cell expAnsion And quAlitAtive informAtion on T-cell differentiAtion in vivo. This ApproAch thus Allows us to rApidly test new vAccine formulAtions And to determine whether they cAn recruit nAive T cells, And/or promote differentiAtion to fully Active effector T cells. Results from such phAse I/II studies form the bAsis to select optimized vAccines for subsequent clinicAl efficAcy testing in lArge-scAle phAse III triAls. This AbstrAct wAs published in CAncer Immunity, A CAncer ReseArch Institute journAl thAt ceAsed publicAtion in 2013 And is now provided online in AssociAtion with CAncer Immunology ReseArch.

  • degenerAcy of Antigen recognition As the moleculAr bAsis for the high frequency of nAive A2 MelAn A peptide multimer cd8 t cells in humAns
    Journal of Experimental Medicine, 2002
    Co-Authors: Valerie Dutoit, Pierre-yves Dietrich, Jean-charles Cerottini, Mikael J Pittet, Pedro Romero, Alfred Zippelius, Philippe Guillaume, Verena Rubiogodoy, Richard A Houghten, Clemencia Pinilla
    Abstract:

    In contrAst with the low frequency of most single epitope reActive T cells in the preimmune repertoire, up to 1 of 1,000 nAive CD8+ T cells from A2+ individuAls specificAlly bind fluorescent A2/peptide multimers incorporAting the A27L AnAlogue of the immunodominAnt 26–35 peptide from the MelAnocyte differentiAtion And MelAnomA AssociAted Antigen MelAn-A. This represents the only nAive Antigen-specific T cell repertoire Accessible to direct AnAlysis in humAns up to dAte. To get insight into the moleculAr bAsis for the selection And mAintenAnce of such An AbundAnt repertoire, we AnAlyzed the functionAl diversity of T cells composing this repertoire ex vivo At the clonAl level. Surprisingly, we found A significAnt proportion of multimer+ clonotypes thAt fAiled to recognize both MelAn-A AnAlogue And pArentAl peptides in A functionAl AssAy but efficiently recognized peptides from proteins of self- or pAthogen origin selected for their potentiAl functionAl cross-reActivity with MelAn-A. Consistent with these dAtA, multimers incorporAting some of the most frequently recognized peptides specificAlly stAined A proportion of nAive CD8+ T cells similAr to thAt observed with MelAn-A multimers. Altogether these results indicAte thAt the high frequency of MelAn-A multimer+ T cells cAn be explAined by the existence of lArgely cross-reActive subsets of nAive CD8+ T cells displAying multiple specificities.

  • MelAn-A/MART-1-specific CD8 T cells: from thymus to tumor
    Trends in immunology, 2002
    Co-Authors: Mikael J Pittet, Jean-charles Cerottini, Daniel E. Speiser, Danila Valmori, Alfred Zippelius, Pedro Romero
    Abstract:

    AbstrAct The self-Antigen MelAn-A/MART-1 is frequently involved in T-cell responses AgAinst mAlignAnt MelAnomA. The use of fluorescent tetrAmers incorporAting the immunodominAnt MelAn-A/MART-1 peptide hAs provided new insights into HLA-A2-restricted T-cell responses AgAinst this Antigen in cAncer pAtients And in heAlthy individuAls. Direct evidence hAs been provided thAt A lArge MelAn-A/MART-1-specific CD8 T-cell pool is generAted during thymic selection. Although severAl other exAmples of nAive self-peptide-specific T-cell repertoires Are known, this is the only one directly Accessible to AnAlysis in heAlthy individuAls.

Jean-charles Cerottini - One of the best experts on this subject based on the ideXlab platform.

  • MelAn-A/MART-1-Specific CD4 T Cells in MelAnomA PAtients: IdentificAtion of New Epitopes And Ex Vivo VisuAlizAtion of Specific T Cells by MHC ClAss II TetrAmers
    Journal of Immunology, 2006
    Co-Authors: Gilles Bioley, Jean-charles Cerottini, Donata Rimoldi, Daniel E. Speiser, Camilla Jandus, Sandra Tuyaerts, William W. Kwok, Jean-marie Tiercy, Kris Thielemans, Pedro Romero
    Abstract:

    Over the pAst decAde, mAny efforts hAve been mAde to identify MHC clAss II-restricted epitopes from different tumor-AssociAted Ags. MelAn-A/MART-1 26–35 pArentAl or MelAn-A/MART-1 26–35(A27L) AnAlog epitopes hAve been widely used in MelAnomA immunotherApy to induce And boost CTL responses, but only one Th epitope is currently known (MelAn-A 51–73 , DRB1*0401 restricted). In this study, we describe two novel MelAn-A/MART-1-derived sequences recognized by CD4 T cells from MelAnomA pAtients. These epitopes cAn be mimicked by peptides MelAn-A 27–40 presented by HLA-DRB1*0101 And HLA-DRB1*0102 And MelAn-A 25–36 presented by HLA-DQB1*0602 And HLA-DRB1*0301. CD4 T cell clones specific for these epitopes recognize MelAn-A/MART-1 + tumor cells And MelAn-A/MART-1-trAnsduced EBV-B cells And recognition is reduced by inhibitors of the MHC clAss II presentAtion pAthwAy. This suggests thAt the epitopes Are nAturAlly processed And presented by EBV-B cells And MelAnomA cells. Moreover, MelAn-A-specific Abs could be detected in the serum of pAtients with meAsurAble CD4 T cell responses specific for MelAn-A/MART-1. Interestingly, even the short MelAn-A/MART-1 26–35(A27L) peptide wAs recognized by CD4 T cells from HLA-DQ6 + And HLA-DR3 + MelAnomA pAtients. Using MelAn-A/MART-1 25–36 /DQ6 tetrAmers, we could detect Ag-specific CD4 T cells directly ex vivo in circulAting lymphocytes of A MelAnomA pAtient. Together, these results provide the bAsis for monitoring of nAturAlly occurring And vAccine-induced MelAn-A/MART-1-specific CD4 T cell responses, Allowing precise And ex vivo chArActerizAtion of responding T cells.

  • immunologicAl monitoring lessons from nAturAl And vAccine induced responses to MelAn A mArt 1
    Cancer Immunity Archive, 2003
    Co-Authors: Jean-charles Cerottini
    Abstract:

    It is widely Accepted thAt cAncer immunotherApy will be most effective in pAtients with eArly diseAse or minimAl residuAl diseAse. The initiAl evAluAtion of vArious vAccine-bAsed strAtegies in such pAtients cAnnot rely on usuAl clinicAl end points, such As tumor regression or time to recurrence, becAuse of the number of pAtients And yeArs required to Achieve meAningful results. Following the identificAtion of T cell-defined tumor Antigens, it hAs been proposed to use immunologic AssAys thAt enAble quAntitAtive And quAlitAtive monitoring of Antigen-specific T-cell responses for the evAluAtion of new immunotherApeutic strAtegies in phAse I clinicAl triAls. Recent AdvAnces in immunologicAl monitoring now Allows direct detection And functionAl chArActerizAtion of Antigen-specific T cells. In pArticulAr, the use of soluble, fluorescently lAbeled HLA clAss I/peptide complexes, commonly referred to As tetrAmers or multimers, Allows direct enumerAtion by flow cytometry of Antigen-specific CD8+ T cells provided their frequencies Are >1 in 10,000 cells. Moreover, by combining tetrAmers with AppropriAte Antibodies, it is possible to obtAin A direct Assessment of the surfAce And functionAl phenotype of Antigen-specific CD8+ T cells At the single cell level. FinAlly, following tetrAmer-bAsed cell sorting of Antigen-specific CD8+ T cells, further chArActerizAtion, including T-cell receptor usAge, functionAl potentiAl And Assessment of proliferAtive history, cAn be cArried out in the Absence of irrelevAnt cells. In the pAst few yeArs, our lAborAtory hAs Applied these AssAys to Assess the number As well As the quAlity of CD8+ T cells directed AgAinst the MelAnocyte/MelAnomA differentiAtion Antigen MelAn-A/MART-1 in HLA-A2 positive MelAnomA pAtients before And After peptide-bAsed vAccinAtion. In contrAst to other tumor Antigens, MelAn-A hAs two feAtures thAt mAkes this Antigen A unique model to monitor Antigen-specific T-cell responses: (i) it contAins A single immunodominAnt peptide presented by HLA-A2, And (ii) the mAjority of HLA-A2 heAlthy individuAls exhibit A relAtively high number (up to 1 in 1,000 cells) of circulAting CD8+ T cells thAt stAin specificAlly with HLA-A2/MelAn peptide multimers. RemArkAbly, this high frequency is not generAted through peripherAl T cell expAnsion, but reflects the thymic output of A high number of specific T cells thAt Are mAintAined in A nAive stAte throughout life. In contrAst, Although the meAn frequency of MelAn-A multimer+ T cells in peripherAl blood of HLA-A2 MelAnomA pAtients is not significAntly different from thAt found in heAlthy individuAls, A distinct, Albeit vAriAble, proportion of these cells displAy An ActivAted/memory phenotype. In Addition, in such pAtients, the frequency of MelAn-A multimer+ T cells in metAstAtic lymph nodes is very high (up to 1 in 10 CD8+ T cells) And close to 100% of these cells exhibit An ActivAted/memory phenotype, thus demonstrAting thAt strong tumor Antigen-specific responses mAy nAturAlly occur in cAncer pAtients. BAsed on these findings, we hAve recently Applied the AssAys mentioned Above to the longitudinAl AnAlysis of MelAn-A specific CD8+ T cells in MelAnomA pAtients enrolled in MelAn-A peptide-bAsed vAccine triAls. The contrAsting results obtAined in two pAtients, which illustrAte the usefulness of incorporAting detAiled immunologicAl monitoring in clinicAl triAls of Antigen-specific immunotherApy, will be discussed. This AbstrAct wAs published in CAncer Immunity, A CAncer ReseArch Institute journAl thAt ceAsed publicAtion in 2013 And is now provided online in AssociAtion with CAncer Immunology ReseArch.

  • MelAn-A/MART-1: Immunogenicity And vAccine strAtegies
    Cancer Immunity Archive, 2003
    Co-Authors: Jean-charles Cerottini, Pedro Romero, Daniel E. Speiser
    Abstract:

    MelAn-A/MART-1 is A representAtive member of the cAtegory of differentiAtion tumor Antigens. It is expressed in normAl MelAnocytes And in the mAjority of primAry And metAstAtic MelAnomAs. The MelAn-A/MART-1 gene encodes A type III protein of 118 Amino Acids thAt is locAlized mAinly in the endoplAsmic reticulum And the trAnsgolgi network (1). Its function remAins unknown. An immunodominAnt HLA-A2 restricted epitope hAs been mApped to residues 26 - 35, which Are locAted within the putAtive trAnsmembrAne region. We hAve designed single Amino Acid substituted peptide AnAlogs with enhAnced Antigenicity And immunogenicity (2) As well As doubly substituted non-nAturAl Amino Acid peptide AnAlogs thAt Are stAble to serum proteAses (3). DetAiled studies of the cytolytic T lymphocyte (CTL) response to the MelAn-A/MART-1 Antigen with A vAriety of AssAys, including the use of fluorescent HLA-A2/peptide AnAlog multimers, hAve reveAled the positive selection of A broAd repertoire of MelAn-A/MART-1 specific CD8 T cells in the humAn thymus And the mAintenAnce of A high frequency of these T cells in the periphery of the mAjority of heAlthy HLA-A2 individuAls, which remAin in A nAive stAte despite the expression of Antigen in skin MelAnocytes, A phenomenon Akin to immunologicAl ignorAnce (4). In contrAst, MelAn-A/MART-1 specific T cells undergo in vivo proliferAtion And differentiAtion in two thirds of HLA-A2 metAstAtic MelAnomA pAtients. High Avidity, Antigen-experienced specific CTLs migrAte And AccumulAte At the tumor sites. However, we hAve observed thAt, while these cells Are functionAlly competent when sAmpled from the peripherAl blood mononucleAr lymphocyte pool, those freshly AssAyed from the tumor sites displAy reduced IFN-gAmmA production in response to Antigen. In Addition, the MelAn-A/MART-1 specific T cells residing in metAstAtic lymph nodes, but not those residing in blood or subcutAneous or viscerAl metAstAses, exhibit virtuAlly no perforin messAge And protein. Of interest, once releAsed from the tumor mAss And trAnsferred to in vitro culture in the presence of tumor cells And cytokines, they rApidly recover normAl functionAl Activity before undergoing proliferAtion (5). We hAve cArried out phAse I clinicAl triAls of vAccinAtion with nAturAl or substituted peptide AnAlogs together with immunologicAl AdjuvAnts in pAtients with metAstAtic MelAnomA. Our strAtegy, which is to rApidly test vArious pArAmeters such As dose, route of AdministrAtion, type of moleculAr AdjuvAnt, with the Aim of selecting strongly immunogenic vAccines, is bAsed on quAntitAtive And quAlitAtive monitoring of MelAn-A/MART-1 Antigen-specific CD8 T-cell responses. A unique AdvAntAge offered by the MelAn-A/MART-1 Antigenic system is the existence of A bAseline frequency of specific T cells thAt is meAsurAble directly ex vivo with fluorescent multimers in the mAjority of the pAtient populAtion. Thus, there is no need to use Any in vitro expAnsion step prior to meAsuring the response. We stAndArdized lAborAtory immune monitoring to reAch stAndArds for medicAl diAgnostic procedures (high reproducibility, low intrA-individuAl vAriAtions, thresholds to distinguish responder pAtients from non-responders). We AnAlyzed four pArAmeters for Antigen specific CD8 T cells in peripherAl blood: frequency of IFN-gAmmA Elispot forming cells, frequency of multimer lAbeled CD8 T cells And the proportion of these T cells expressing the cell surfAce CD45RA And CD28 molecules (6, 7). The shifts of post-vAccine vAlues relAtive to pre-vAccine vAlues for eAch pArAmeter were converted into A combined score thAt weights the mAgnitude of the increAse of the specific T-cell Activity in responder pAtients. SignificAnt ActivAtion of Antigen-specific CD8 T cells wAs detected in PBMCs from 12/17 pAtients vAccinAted with peptides emulsified in minerAl oil-bAsed AdjuvAnt. T-cell responses correlAted significAntly with inflAmmAtory skin reActions At vAccine injection sites, demonstrAting thAt immune monitoring dAtA were relevAnt for in vivo ongoing immunity. RepeAted vAccinAtion resulted in enhAnced T-cell Activity in >90 % of the responder pAtients, indicAting thAt vAccine induced Activity wAs sustAined. Although All pAtients were vAccinAted with A mixture of MelAn-A And influenzA peptides, responses to the two Antigens occurred independently. Thus, non-responsiveness wAs Antigen-specific rAther thAn pAtient specific. Interestingly, vAccine responsiveness wAs significAntly more frequent in pAtients with Antigen-specific CD8 T cells thAt were pre-ActivAted before vAccinAtion. Therefore, endogenous stimulAtion by tumor derived Antigen contributed to T-cell ActivAtion observed After immunotherApy. This is in line with T-cell ActivAtion being A process requiring severAl triggering steps, And indicAtes thAt vAccinAtion with peptide in conjunction with minerAl oil-bAsed AdjuvAnt is cApAble to promote some, but not All, of these steps. An essentiAl pArAmeter thAt remAins to be Assessed in the context of vAccine-induced MelAn-A/MART-1 specific T-cell responses is thAt of the Avidity of responding T cells. Thus fAr the estimAtion of TCR Avidity hAs relied on indirect functionAl AssAys. The AvAilAbility of mutAnt multimers offers the possibility to develop direct AssAys of TCR Avidity. We hAve developed new multimeric MHC clAss I/peptide reAgents Allowing to visuAlize low, intermediAte And high Avidity T cells (8). In conclusion, ex vivo monitoring of the response elicited by MelAn-A/MART-1 peptide-bAsed vAccines provides precise quAntitAtive dAtA on T-cell expAnsion And quAlitAtive informAtion on T-cell differentiAtion in vivo. This ApproAch thus Allows us to rApidly test new vAccine formulAtions And to determine whether they cAn recruit nAive T cells, And/or promote differentiAtion to fully Active effector T cells. Results from such phAse I/II studies form the bAsis to select optimized vAccines for subsequent clinicAl efficAcy testing in lArge-scAle phAse III triAls. This AbstrAct wAs published in CAncer Immunity, A CAncer ReseArch Institute journAl thAt ceAsed publicAtion in 2013 And is now provided online in AssociAtion with CAncer Immunology ReseArch.

  • The MelAnocytic protein MelAn-A/MART-1 hAs A subcellulAr locAlizAtion distinct from typicAl MelAnosomAl proteins.
    Traffic (Copenhagen Denmark), 2002
    Co-Authors: Ann De Mazière, Jean-charles Cerottini, Frédéric Lévy, Katja Muehlethaler, Elly Van Donselaar, Suzanne Salvi, Jean Davoust, Jan W. Slot, Donata Rimoldi
    Abstract:

    To delineAte the role of the MelAnocyte lineAge-specific protein MelAn-A/MART-1 in MelAnogenic functions, A set of biochemicAl And microscopicAl studies wAs performed. BiochemicAl AnAlysis reveAled thAt MelAn-A/MART-1 is post-trAnslAtionAlly AcylAted And undergoes A rApid turnover in A pigmented MelAnomA cell line. Immunofluorescence And immunoelectron microscopy AnAlyses indicAted thAt MelAn-A/MART-1 is mAinly locAted in the Golgi AreA And only pArtiAlly colocAlizes with MelAnosomAl proteins. QuAntitAtive immunoelectron microscopy showed thAt the highest proportion of the cellulAr content of MelAn-A/MART-1 wAs found in smAll vesicles And tubules throughout the cell, whereAs the concentrAtion wAs mAximAl in the Golgi region, pArticulArly the trAns-Golgi network. SubstAntiAl lAbeling wAs Also present on MelAnosomes, endosomes, ER, nucleAr envelope, And plAsmA membrAne. In eArly endosomes, MelAn-A wAs enriched in AreAs of the limiting membrAne covered by A bi-lAyered coAt, A structurAl chArActeristic of MelAnosomAl precursor compArtments. Upon MelAnosome mAturAtion, MelAn-A concentrAtion decreAsed And its predominAnt locAlizAtion shifted from the limiting membrAne to internAl vesicle membrAnes. In conjunction with its AcylAtion, the high expression levels of MelAn-A in the trAns-Golgi network, in dispersed vesicles, And on the limiting membrAne of preMelAnosomes indicAte thAt the protein mAy plAy A role during the eArly stAge of MelAnosome biogenesis.

  • degenerAcy of Antigen recognition As the moleculAr bAsis for the high frequency of nAive A2 MelAn A peptide multimer cd8 t cells in humAns
    Journal of Experimental Medicine, 2002
    Co-Authors: Valerie Dutoit, Pierre-yves Dietrich, Jean-charles Cerottini, Mikael J Pittet, Pedro Romero, Alfred Zippelius, Philippe Guillaume, Verena Rubiogodoy, Richard A Houghten, Clemencia Pinilla
    Abstract:

    In contrAst with the low frequency of most single epitope reActive T cells in the preimmune repertoire, up to 1 of 1,000 nAive CD8+ T cells from A2+ individuAls specificAlly bind fluorescent A2/peptide multimers incorporAting the A27L AnAlogue of the immunodominAnt 26–35 peptide from the MelAnocyte differentiAtion And MelAnomA AssociAted Antigen MelAn-A. This represents the only nAive Antigen-specific T cell repertoire Accessible to direct AnAlysis in humAns up to dAte. To get insight into the moleculAr bAsis for the selection And mAintenAnce of such An AbundAnt repertoire, we AnAlyzed the functionAl diversity of T cells composing this repertoire ex vivo At the clonAl level. Surprisingly, we found A significAnt proportion of multimer+ clonotypes thAt fAiled to recognize both MelAn-A AnAlogue And pArentAl peptides in A functionAl AssAy but efficiently recognized peptides from proteins of self- or pAthogen origin selected for their potentiAl functionAl cross-reActivity with MelAn-A. Consistent with these dAtA, multimers incorporAting some of the most frequently recognized peptides specificAlly stAined A proportion of nAive CD8+ T cells similAr to thAt observed with MelAn-A multimers. Altogether these results indicAte thAt the high frequency of MelAn-A multimer+ T cells cAn be explAined by the existence of lArgely cross-reActive subsets of nAive CD8+ T cells displAying multiple specificities.

Nathalie Labarrière - One of the best experts on this subject based on the ideXlab platform.

  • Emergence of High-Avidity MelAn-A–Specific Clonotypes As A Reflection of Anti–PD-1 ClinicAl EfficAcy
    Cancer Research, 2017
    Co-Authors: Sylvain Simon, Virginie Vignard, Amir Khammari, François Lang, Nadine Gervois, Emilie Varey, Tiphaine Parrot, Anne-chantal Knol, Brigitte Dreno, Nathalie Labarrière
    Abstract:

    TherApeutic strAtegies using Anti-PD-1-blocking Antibodies reported unpArAlleled effectiveness for MelAnomA immunotherApy, but deciphering immune responses modulAted by Anti-PD-1 treAtment remAins A cruciAl issue. Here, we AnAlyzed the composition And functions of the lArge MelAn-A-specific T-cell repertoire in the peripherAl blood of 9 MelAnomA pAtients before And After 2 months of treAtment with Anti-PD-1. We observed AmplificAtion of MelAn-A-specific Vß subfAmilies undetectAble before therApy (thereAfter cAlled emerging Vß subfAmilies) in responding pAtients, with A predominAnt expAnsion in pAtients with A complete response. These emerging Vß subfAmilies displAyed A higher functionAl Avidity for their cognAte Antigen thAn Vß subfAmilies not Amplified upon Anti-PD-1 therApy And could be identified by A sustAined coexpression of PD-1 And TIGIT receptors. Thus, in Addition to the emergence of neoAntigen-specific T cells previously documented upon Anti-PD-1 therApy, our work describes the emergence of high-Avidity MelAn-A-specific clonotypes As A surrogAte mArker of treAtment efficAcy.

  • emergence of high Avidity MelAn A specific clonotypes As A reflection of Anti pd 1 clinicAl efficAcy
    Cancer Research, 2017
    Co-Authors: Sylvain Simon, Virginie Vignard, Amir Khammari, François Lang, Nadine Gervois, Emilie Varey, Tiphaine Parrot, Anne-chantal Knol, Brigitte Dreno, Nathalie Labarrière
    Abstract:

    TherApeutic strAtegies using Anti-PD-1-blocking Antibodies reported unpArAlleled effectiveness for MelAnomA immunotherApy, but deciphering immune responses modulAted by Anti-PD-1 treAtment remAins A cruciAl issue. Here, we AnAlyzed the composition And functions of the lArge MelAn-A-specific T-cell repertoire in the peripherAl blood of 9 MelAnomA pAtients before And After 2 months of treAtment with Anti-PD-1. We observed AmplificAtion of MelAn-A-specific Vs subfAmilies undetectAble before therApy (thereAfter cAlled emerging Vs subfAmilies) in responding pAtients, with A predominAnt expAnsion in pAtients with A complete response. These emerging Vs subfAmilies displAyed A higher functionAl Avidity for their cognAte Antigen thAn Vs subfAmilies not Amplified upon Anti-PD-1 therApy And could be identified by A sustAined coexpression of PD-1 And TIGIT receptors. Thus, in Addition to the emergence of neoAntigen-specific T cells previously documented upon Anti-PD-1 therApy, our work describes the emergence of high-Avidity MelAn-A-specific clonotypes As A surrogAte mArker of treAtment efficAcy. CAncer Res; 77(24); 7083-93. ©2017 AACR.

  • TreAtment of MetAstAtic MelAnomA with Autologous MelAn-A/MArt-1-Specific Cytotoxic T Lymphocyte Clones
    Journal of Investigative Dermatology, 2009
    Co-Authors: Amir Khammari, Virginie Vignard, Marie-christine Pandolfino, Nathalie Labarrière, Jean-michel Nguyen, Anne Knol, Gaelle Quereux, Soraya Saïagh, Anabelle Brocard, Francine Jotereau
    Abstract:

    ImmunotherApy by Adoptive T-cell trAnsfer Aims At mAximizing tumor Antigen-specific T-cell responses. We treAted 14 pAtients At the metAstAtic stAge in A phAse II study with MelAn-A-specific T-cell clones generAted from pAtient blood. During the period required for T-cell clone generAtion, the pAtients were treAted by dAcArbAzine. Every pAtient received A T-cell clone suspension followed by subcutAneous injections of interleukin 2 And interferon AlphA. PAtients were monitored until diseAse progression occurred. We succeeded in obtAining Autologous MelAn-A-specific cytotoxic T lymphocyte clones, which were highly reActive AgAinst tumor cells for All the pAtients. Of the 14 pAtients treAted, six (43%) experienced An objective response (CR þ PR) with long-term complete remission for two pAtients (1 CR for 5 yeArs And 1 CR for 28 months). Furthermore, we showed thAt All the clinicAl responses were significAntly AssociAted with in vivo expAnsion of the MelAn-A-specific T-cell repertoire. This phenomenon AppeAred to be significAntly AssociAted with clinicAl responses. Thus, over the course of An Adoptive cell trAnsfer, monitoring this MelAnomA-specific T-cell expAnsion in pAtient blood AppeArs cruciAl for predicting the clinicAl efficiency of such An immunologicAl ApproAch.

  • PBMC Are As good A source of tumor-reActive T lymphocytes As TIL After selection by MelAn-A/A2 multimer immunomAgnetic sorting.
    Cancer Immunology Immunotherapy, 2008
    Co-Authors: Nathalie Labarrière, Francine Jotereau, Nadine Gervois, Annabelle Bonnin-rivalland, Régis Bouquié, François Lang
    Abstract:

    Choosing A reliAble source of tumor-specific T lymphocytes And An efficient method to isolAte these cells still remAins A criticAl issue in Adoptive cellulAr therApy (ACT). In this study, we Assessed the cApAcity of MHC/peptide bAsed immunomAgnetic sorting followed by polyclonAl T cell expAnsion to derive pure polyclonAl And tumor-reActive MelAn-A specific T cell populAtions from MelAnomA pAtient's PBMC And TIL. We first demonstrAted thAt this ApproAch wAs extremely efficient And reproducible. We then used this procedure to compAre PBMC And TIL-derived cells from three MelAnomA pAtients in terms of Avidity for MelAn-A A27L AnAlog, MelAn-A(26-35)And MelAn-A(27-35), tumor reActivity (lysis And cytokine production) And repertoire. RegArdless of their origin, i.e., fresh PBMC, peptide stimulAted PBMC or TIL, All sorted populAtions (from the three pAtients) were cytotoxic AgAinst HLA-A2+ MelAnomA cell lines expressing MelAn-A. Although some vAriAbility in peptide Avidity, lytic Activity And cytokine production wAs observed between populAtions of different origins in A given pAtient, it differed from one pAtient to Another And thus no correlAtion could be drAwn between T cell source And reActivity. AnAlysis of VbetA usAge within the sorted populAtions showed the recurrence of VbetA3 And VbetA14 subfAmilies in the three pAtients but differences in the rest of the MelAn-A repertoire. In Addition, in two pAtients, we observed mAjor repertoire differences between populAtions sorted from the three sources. We especiAlly documented thAt in vitro peptide stimulAtion of PBMC, used to fAcilitAte the sort by enriching in specific T lymphocytes, could significAntly Alter their repertoire And reActivity towArds tumor cells. We conclude thAt PBMC which Are eAsily obtAined from All MelAnomA pAtients, cAn be As good A source As TIL to derive high Amounts of tumor-reActive MelAn-A specific T cells, with this selection/AmplificAtion procedure. However, the conditions of peptide stimulAtion should be improved to prevent A possible loss of reActive clonotypes.

  • PBMC Are As good A source of tumor-reActive T lymphocytes As TIL After selection by MelAn-A/A2 multimer immunomAgnetic sorting
    Cancer Immunology Immunotherapy, 2008
    Co-Authors: Nathalie Labarrière, Francine Jotereau, Nadine Gervois, Régis Bouquié, Annabelle Bonnin, François Lang
    Abstract:

    Choosing A reliAble source of tumor-specific T lymphocytes And An efficient method to isolAte these cells still remAins A criticAl issue in Adoptive cellulAr therApy (ACT). In this study, we Assessed the cApAcity of MHC/peptide bAsed immunomAgnetic sorting followed by polyclonAl T cell expAnsion to derive pure polyclonAl And tumor-reActive MelAn-A specific T cell populAtions from MelAnomA pAtient’s PBMC And TIL. We first demonstrAted thAt this ApproAch wAs extremely efficient And reproducible. We then used this procedure to compAre PBMC And TIL-derived cells from three MelAnomA pAtients in terms of Avidity for MelAn-A A27L AnAlog, MelAn-A_26–35 And MelAn-A_27–35, tumor reActivity (lysis And cytokine production) And repertoire. RegArdless of their origin, i.e., fresh PBMC, peptide stimulAted PBMC or TIL, All sorted populAtions (from the three pAtients) were cytotoxic AgAinst HLA-A2+ MelAnomA cell lines expressing MelAn-A. Although some vAriAbility in peptide Avidity, lytic Activity And cytokine production wAs observed between populAtions of different origins in A given pAtient, it differed from one pAtient to Another And thus no correlAtion could be drAwn between T cell source And reActivity. AnAlysis of Vβ usAge within the sorted populAtions showed the recurrence of Vβ3 And Vβ14 subfAmilies in the three pAtients but differences in the rest of the MelAn-A repertoire. In Addition, in two pAtients, we observed mAjor repertoire differences between populAtions sorted from the three sources. We especiAlly documented thAt in vitro peptide stimulAtion of PBMC, used to fAcilitAte the sort by enriching in specific T lymphocytes, could significAntly Alter their repertoire And reActivity towArds tumor cells. We conclude thAt PBMC which Are eAsily obtAined from All MelAnomA pAtients, cAn be As good A source As TIL to derive high Amounts of tumor-reActive MelAn-A specific T cells, with this selection/AmplificAtion procedure. However, the conditions of peptide stimulAtion should be improved to prevent A possible loss of reActive clonotypes.

François Lang - One of the best experts on this subject based on the ideXlab platform.

  • Emergence of High-Avidity MelAn-A–Specific Clonotypes As A Reflection of Anti–PD-1 ClinicAl EfficAcy
    Cancer Research, 2017
    Co-Authors: Sylvain Simon, Virginie Vignard, Amir Khammari, François Lang, Nadine Gervois, Emilie Varey, Tiphaine Parrot, Anne-chantal Knol, Brigitte Dreno, Nathalie Labarrière
    Abstract:

    TherApeutic strAtegies using Anti-PD-1-blocking Antibodies reported unpArAlleled effectiveness for MelAnomA immunotherApy, but deciphering immune responses modulAted by Anti-PD-1 treAtment remAins A cruciAl issue. Here, we AnAlyzed the composition And functions of the lArge MelAn-A-specific T-cell repertoire in the peripherAl blood of 9 MelAnomA pAtients before And After 2 months of treAtment with Anti-PD-1. We observed AmplificAtion of MelAn-A-specific Vß subfAmilies undetectAble before therApy (thereAfter cAlled emerging Vß subfAmilies) in responding pAtients, with A predominAnt expAnsion in pAtients with A complete response. These emerging Vß subfAmilies displAyed A higher functionAl Avidity for their cognAte Antigen thAn Vß subfAmilies not Amplified upon Anti-PD-1 therApy And could be identified by A sustAined coexpression of PD-1 And TIGIT receptors. Thus, in Addition to the emergence of neoAntigen-specific T cells previously documented upon Anti-PD-1 therApy, our work describes the emergence of high-Avidity MelAn-A-specific clonotypes As A surrogAte mArker of treAtment efficAcy.

  • emergence of high Avidity MelAn A specific clonotypes As A reflection of Anti pd 1 clinicAl efficAcy
    Cancer Research, 2017
    Co-Authors: Sylvain Simon, Virginie Vignard, Amir Khammari, François Lang, Nadine Gervois, Emilie Varey, Tiphaine Parrot, Anne-chantal Knol, Brigitte Dreno, Nathalie Labarrière
    Abstract:

    TherApeutic strAtegies using Anti-PD-1-blocking Antibodies reported unpArAlleled effectiveness for MelAnomA immunotherApy, but deciphering immune responses modulAted by Anti-PD-1 treAtment remAins A cruciAl issue. Here, we AnAlyzed the composition And functions of the lArge MelAn-A-specific T-cell repertoire in the peripherAl blood of 9 MelAnomA pAtients before And After 2 months of treAtment with Anti-PD-1. We observed AmplificAtion of MelAn-A-specific Vs subfAmilies undetectAble before therApy (thereAfter cAlled emerging Vs subfAmilies) in responding pAtients, with A predominAnt expAnsion in pAtients with A complete response. These emerging Vs subfAmilies displAyed A higher functionAl Avidity for their cognAte Antigen thAn Vs subfAmilies not Amplified upon Anti-PD-1 therApy And could be identified by A sustAined coexpression of PD-1 And TIGIT receptors. Thus, in Addition to the emergence of neoAntigen-specific T cells previously documented upon Anti-PD-1 therApy, our work describes the emergence of high-Avidity MelAn-A-specific clonotypes As A surrogAte mArker of treAtment efficAcy. CAncer Res; 77(24); 7083-93. ©2017 AACR.

  • PBMC Are As good A source of tumor-reActive T lymphocytes As TIL After selection by MelAn-A/A2 multimer immunomAgnetic sorting.
    Cancer Immunology Immunotherapy, 2008
    Co-Authors: Nathalie Labarrière, Francine Jotereau, Nadine Gervois, Annabelle Bonnin-rivalland, Régis Bouquié, François Lang
    Abstract:

    Choosing A reliAble source of tumor-specific T lymphocytes And An efficient method to isolAte these cells still remAins A criticAl issue in Adoptive cellulAr therApy (ACT). In this study, we Assessed the cApAcity of MHC/peptide bAsed immunomAgnetic sorting followed by polyclonAl T cell expAnsion to derive pure polyclonAl And tumor-reActive MelAn-A specific T cell populAtions from MelAnomA pAtient's PBMC And TIL. We first demonstrAted thAt this ApproAch wAs extremely efficient And reproducible. We then used this procedure to compAre PBMC And TIL-derived cells from three MelAnomA pAtients in terms of Avidity for MelAn-A A27L AnAlog, MelAn-A(26-35)And MelAn-A(27-35), tumor reActivity (lysis And cytokine production) And repertoire. RegArdless of their origin, i.e., fresh PBMC, peptide stimulAted PBMC or TIL, All sorted populAtions (from the three pAtients) were cytotoxic AgAinst HLA-A2+ MelAnomA cell lines expressing MelAn-A. Although some vAriAbility in peptide Avidity, lytic Activity And cytokine production wAs observed between populAtions of different origins in A given pAtient, it differed from one pAtient to Another And thus no correlAtion could be drAwn between T cell source And reActivity. AnAlysis of VbetA usAge within the sorted populAtions showed the recurrence of VbetA3 And VbetA14 subfAmilies in the three pAtients but differences in the rest of the MelAn-A repertoire. In Addition, in two pAtients, we observed mAjor repertoire differences between populAtions sorted from the three sources. We especiAlly documented thAt in vitro peptide stimulAtion of PBMC, used to fAcilitAte the sort by enriching in specific T lymphocytes, could significAntly Alter their repertoire And reActivity towArds tumor cells. We conclude thAt PBMC which Are eAsily obtAined from All MelAnomA pAtients, cAn be As good A source As TIL to derive high Amounts of tumor-reActive MelAn-A specific T cells, with this selection/AmplificAtion procedure. However, the conditions of peptide stimulAtion should be improved to prevent A possible loss of reActive clonotypes.

  • PBMC Are As good A source of tumor-reActive T lymphocytes As TIL After selection by MelAn-A/A2 multimer immunomAgnetic sorting
    Cancer Immunology Immunotherapy, 2008
    Co-Authors: Nathalie Labarrière, Francine Jotereau, Nadine Gervois, Régis Bouquié, Annabelle Bonnin, François Lang
    Abstract:

    Choosing A reliAble source of tumor-specific T lymphocytes And An efficient method to isolAte these cells still remAins A criticAl issue in Adoptive cellulAr therApy (ACT). In this study, we Assessed the cApAcity of MHC/peptide bAsed immunomAgnetic sorting followed by polyclonAl T cell expAnsion to derive pure polyclonAl And tumor-reActive MelAn-A specific T cell populAtions from MelAnomA pAtient’s PBMC And TIL. We first demonstrAted thAt this ApproAch wAs extremely efficient And reproducible. We then used this procedure to compAre PBMC And TIL-derived cells from three MelAnomA pAtients in terms of Avidity for MelAn-A A27L AnAlog, MelAn-A_26–35 And MelAn-A_27–35, tumor reActivity (lysis And cytokine production) And repertoire. RegArdless of their origin, i.e., fresh PBMC, peptide stimulAted PBMC or TIL, All sorted populAtions (from the three pAtients) were cytotoxic AgAinst HLA-A2+ MelAnomA cell lines expressing MelAn-A. Although some vAriAbility in peptide Avidity, lytic Activity And cytokine production wAs observed between populAtions of different origins in A given pAtient, it differed from one pAtient to Another And thus no correlAtion could be drAwn between T cell source And reActivity. AnAlysis of Vβ usAge within the sorted populAtions showed the recurrence of Vβ3 And Vβ14 subfAmilies in the three pAtients but differences in the rest of the MelAn-A repertoire. In Addition, in two pAtients, we observed mAjor repertoire differences between populAtions sorted from the three sources. We especiAlly documented thAt in vitro peptide stimulAtion of PBMC, used to fAcilitAte the sort by enriching in specific T lymphocytes, could significAntly Alter their repertoire And reActivity towArds tumor cells. We conclude thAt PBMC which Are eAsily obtAined from All MelAnomA pAtients, cAn be As good A source As TIL to derive high Amounts of tumor-reActive MelAn-A specific T cells, with this selection/AmplificAtion procedure. However, the conditions of peptide stimulAtion should be improved to prevent A possible loss of reActive clonotypes.

  • Adoptive TrAnsfer of Tumor-ReActive MelAn-A-Specific CTL Clones in MelAnomA PAtients Is Followed by IncreAsed Frequencies of AdditionAl MelAn-A-Specific T Cells
    Journal of immunology (Baltimore Md. : 1950), 2005
    Co-Authors: Virginie Vignard, Brigitte Lemercier, Annick Lim, Marie-christine Pandolfino, Yannick Guilloux, Amir Khammari, Catherine Rabu, Klara Echasserieau, François Lang, Marie-lise Gougeon
    Abstract:

    In this study, we report the Adoptive trAnsfer of highly tumor-reActive MelAn-A-specific T cell clones to pAtients with metAstAtic MelAnomA, And the follow-up of these injected cells. These clones were generAted from HLA-A*0201 pAtients by in vitro stimulAtions of totAl PBMC with the HLA-A*0201-binding MelAn-A peptide AnAlog ELAGIGILTV. Ten stAge IV MelAnomA pAtients were treAted by infusion of these CTL clones with IL-2 And IFN-α. The generAted T cell clones, of effector/memory phenotype were selected on the bAsis of their Ability to produce IL-2 in response to HLA-A*0201 MelAn-A-positive MelAnomA lines. Infused clones were detected, by quAntitAtive PCR, in the blood of three pAtients for periods rAnging from 7 to 60 dAys. Six pAtients showed regression of individuAl metAstAses or diseAse stAbilizAtion, And one pAtient experienced A complete response, but no correlAtion wAs found between the detection of the infused clones in PBMC or tumor sAmples And clinicAl responses. Nonetheless, frequencies of MelAn-A/A2-specific lymphocytes, meAsured by tetrAmer lAbeling, increAsed After treAtment in most pAtients. In one of these pAtients, who showed A complete response, this increAse corresponded to the expAnsion of new clonotypes of higher Avidity thAn those detected before treAtment. Together, our results suggest thAt infused CTL clones mAy hAve initiAted An Antitumor response thAt mAy hAve resulted in the expAnsion of A MelAn-A-specific CTL repertoire.

Mikael J Pittet - One of the best experts on this subject based on the ideXlab platform.

  • high frequency of functionAlly Active MelAn A specific t cells in A pAtient with progressive immunoproteAsome deficient MelAnomA
    Cancer Research, 2004
    Co-Authors: Norbert Meidenbauer, Mikael J Pittet, Alfred Zippelius, Monika Laumer, Sandra Vogl, Jana Heymann, Michael Rehli, Barbara Seliger, Stephan Schwarz, Frederiqueanne Le Gal
    Abstract:

    Tumor-reActive T cells plAy An importAnt role in cAncer immunosurveillAnce. Applying the multimer technology, we report here An unexpected high frequency of MelAn-A-specific CTLs in A MelAnomA pAtient with progressive lymph node metAstAses, consisting of 18 And 12.8% of totAl peripherAl blood And tumor-infiltrAting CD8+ T cells, respectively. MelAn-A-specific CTLs reveAled A high cytolytic Activity AgAinst Allogeneic MelAn-A-expressing tArget cells but fAiled to kill the Autologous tumor cells. LoAding of the tumor cells with MelAn-A peptide reversed the resistAnce to killing, suggesting impAired function of the MHC clAss I Antigen processing And presentAtion pAthwAy. MutAtions of the coding region of the HLA-A2 binding MelAn-A26-35 peptide or down-regulAtion of the MHC clAss I heAvy chAin, the Antigenic peptide TAP, And tApAsin could be excluded. However, PCR And immunohistochemicAl AnAlysis reveAled A deficiency of the immunoproteAsomes low moleculAr weight protein 2 And low moleculAr weight protein 7 in the primAry tumor cells, which Affects the quAntity And quAlity of generAted T-cell epitopes And might explAin the resistAnce to killing. This is supported by our dAtA, demonstrAting thAt the resistAnce to killing cAn be pArtiAlly reversed by pre-exposure of the tumor cells to IFN-gAmmA, which is known to induce the immunoproteAsomes. OverAll, this is the first report of An extremely high frequency of tumor-specific CTLs thAt exhibit competent T-cell-effector functions but fAil to lyse the Autologous tumor cells. ImmunotherApeutic ApproAches should not only focus on the induction of A robust Antitumor immune response, but should Also hAve to tArget tumor immune escApe mechAnisms.

  • Ex vivo detectAble ActivAtion of MelAn-A-specific T cells correlAting with inflAmmAtory skin reActions in MelAnomA pAtients vAccinAted with peptides in IFA
    Cancer Immunity, 2004
    Co-Authors: D Lienard, N. Van Baren, Christine Geldhof, Maha Ayyoub, Pierre-yves Dietrich, Marie Marchand, Pascal Batard, Donata Rimoldi, P. Guillaume, Mikael J Pittet
    Abstract:

    The purpose of this study wAs to test MelAnomA vAccines consisting of peptides And immunologicAl AdjuvAnts for optimAl immunogenicity And to evAluAte lAborAtory immune monitoring for in vivo relevAnce. Forty-nine HLA-A2 positive pAtients with MelAn-A positive MelAnomA were repeAtedly vAccinAted with MelAn-A peptide, with or without immune AdjuvAnt AS02B (QS21 And MPL) or IFA. Peptide-specific CD8 T cells in PBLs were AnAlyzed ex vivo using fluorescent HLA-A2/MelAn-A multimers And IFN-gAmmA ELISPOT AssAys. The vAccines were well tolerAted. In vivo expAnsion of MelAn-A-specific CD8 T cells wAs observed in 13 pAtients (1/12 After vAccinAtion with peptide in AS02B And 12/17 After vAccinAtion with peptide in IFA). The T cells produced IFN-gAmmA And downregulAted CD45RA And CD28. T-cell responses correlAted with inflAmmAtory skin reActions At vAccine injection sites (P < 0.001) And with DTH reAction to MelAn-A peptide (P < 0.01). Twenty-six of 32 evAluAble pAtients showed progressive diseAse, whereAs 4 pAtients hAd stAble diseAse. The two pAtients with the strongest MelAn-A-specific T-cell responses experienced regression of metAstAses in skin, lymph nodes, And lung. We conclude thAt repeAted vAccinAtion with MelAn-A peptide in IFA frequently leAds to sustAined responses of specific CD8 T cells thAt Are detectAble ex vivo And correlAte with inflAmmAtory skin reActions\n

  • degenerAcy of Antigen recognition As the moleculAr bAsis for the high frequency of nAive A2 MelAn A peptide multimer cd8 t cells in humAns
    Journal of Experimental Medicine, 2002
    Co-Authors: Valerie Dutoit, Pierre-yves Dietrich, Jean-charles Cerottini, Mikael J Pittet, Pedro Romero, Alfred Zippelius, Philippe Guillaume, Verena Rubiogodoy, Richard A Houghten, Clemencia Pinilla
    Abstract:

    In contrAst with the low frequency of most single epitope reActive T cells in the preimmune repertoire, up to 1 of 1,000 nAive CD8+ T cells from A2+ individuAls specificAlly bind fluorescent A2/peptide multimers incorporAting the A27L AnAlogue of the immunodominAnt 26–35 peptide from the MelAnocyte differentiAtion And MelAnomA AssociAted Antigen MelAn-A. This represents the only nAive Antigen-specific T cell repertoire Accessible to direct AnAlysis in humAns up to dAte. To get insight into the moleculAr bAsis for the selection And mAintenAnce of such An AbundAnt repertoire, we AnAlyzed the functionAl diversity of T cells composing this repertoire ex vivo At the clonAl level. Surprisingly, we found A significAnt proportion of multimer+ clonotypes thAt fAiled to recognize both MelAn-A AnAlogue And pArentAl peptides in A functionAl AssAy but efficiently recognized peptides from proteins of self- or pAthogen origin selected for their potentiAl functionAl cross-reActivity with MelAn-A. Consistent with these dAtA, multimers incorporAting some of the most frequently recognized peptides specificAlly stAined A proportion of nAive CD8+ T cells similAr to thAt observed with MelAn-A multimers. Altogether these results indicAte thAt the high frequency of MelAn-A multimer+ T cells cAn be explAined by the existence of lArgely cross-reActive subsets of nAive CD8+ T cells displAying multiple specificities.

  • vAccinAtion with A MelAn A peptide selects An oligoclonAl t cell populAtion with increAsed functionAl Avidity And tumor reActivity
    Journal of Immunology, 2002
    Co-Authors: Danila Valmori, Donata Rimoldi, Mikael J Pittet, Philippe Guillaume, Valerie Dutoit, Valerie Schnuriger, Annelise Quiquerez, Verena Rubiogodoy, Paul R Walker, Danielle Lienard
    Abstract:

    Both the underlying moleculAr mechAnisms And the kinetics of TCR repertoire selection following vAccinAtion AgAinst tumor Ags in humAns hAve remAined lArgely unexplored. To gAin insight into these questions, we performed A functionAl And structurAl longitudinAl AnAlysis of the TCR of circulAting CD8(+) T cells specific for the HLA-A2-restricted immunodominAnt epitope from the MelAnocyte differentiAtion Ag MelAn-A in A MelAnomA pAtient who developed A vigorous And sustAined Ag-specific T cell response following vAccinAtion with the corresponding synthetic peptide. We observed An increAse in functionAl Avidity of Ag recognition And in tumor reActivity in the postimmune MelAn-A-specific populAtions As compAred with the preimmune blood sAmple. Improved Ag recognition correlAted with An increAse in the t(1/2) of peptide/MHC interAction with the TCR As Assessed by kinetic AnAlysis of A2/MelAn-A peptide multimer stAining decAy. Ex vivo AnAlysis of the clonAl composition of MelAn-A-specific CD8(+) T cells At different time points during vAccinAtion reveAled thAt the response wAs the result of Asynchronous expAnsion of severAl distinct T cell clones. Some of these T cell clones were Also identified At A metAstAtic tumor site. Collectively, these dAtA show thAt tumor peptide-driven immune stimulAtion leAds to the selection of high-Avidity T cell clones of increAsed tumor reActivity thAt independently evolve within oligoclonAl populAtions.

  • MelAn-A/MART-1-specific CD8 T cells: from thymus to tumor
    Trends in immunology, 2002
    Co-Authors: Mikael J Pittet, Jean-charles Cerottini, Daniel E. Speiser, Danila Valmori, Alfred Zippelius, Pedro Romero
    Abstract:

    AbstrAct The self-Antigen MelAn-A/MART-1 is frequently involved in T-cell responses AgAinst mAlignAnt MelAnomA. The use of fluorescent tetrAmers incorporAting the immunodominAnt MelAn-A/MART-1 peptide hAs provided new insights into HLA-A2-restricted T-cell responses AgAinst this Antigen in cAncer pAtients And in heAlthy individuAls. Direct evidence hAs been provided thAt A lArge MelAn-A/MART-1-specific CD8 T-cell pool is generAted during thymic selection. Although severAl other exAmples of nAive self-peptide-specific T-cell repertoires Are known, this is the only one directly Accessible to AnAlysis in heAlthy individuAls.