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Pathogenesis and Immunity

Human Papillomavirus Type 33 E7 Peptides Presented by HLA-DR*0402 to Tumor-Infiltrating T Cells in Cervical Cancer

Hanni Höhn, Henryk Pilch, Susanne Günzel, Claudia Neukirch, Kirsten Freitag, Antje Necker, Markus J. Maeurer
Hanni Höhn
Department of Medical Microbiology and
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Henryk Pilch
Department of Gynecology, Johannes Gutenberg University, Mainz, Germany, and
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Susanne Günzel
Department of Gynecology, Johannes Gutenberg University, Mainz, Germany, and
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Claudia Neukirch
Department of Medical Microbiology and
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Kirsten Freitag
Department of Medical Microbiology and
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Antje Necker
Antibody Department, Beckman/Coulter, Marseille, France
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Markus J. Maeurer
Department of Medical Microbiology and
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DOI: 10.1128/JVI.74.14.6632-6636.2000
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  • Fig. 1.
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    Fig. 1.

    TCR VB families in PBL and TIL. PBL were gated on CD4+ or CD8+ T cells and tested for expression of individual TCR VB chains by flow cytometry. No major TCR VB expansion could be detected in PBL. In contrast, the majority of TIL (>98% CD4+) stained positive for the TCR VB16. +, detection of monoclonal TCR VB chains as listed in Table 2.

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    Fig. 2.

    Detection of TCR VB16+ T cells infiltrating into cervical cancer. Serial sections from tumor tissue were stained for CD3, CD4, CD8, and TCR VB16+ T cells. Note that CD4+ T cells represent the majority of T cells and that TCR VB16+ T cells directly infiltrate the tumor lesion.

  • Fig. 3.
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    Fig. 3.

    Titration of HPV33 E773–87 peptides onto autologous antigen-presenting cells. Peptides were serially diluted and pulsed onto antigen-presenting cells, incubated for 2 h at room temperature, and tested for CD4+VB16+ T-cell recognition, as determined by IFN-γ secretion. The peptide ASDLRTIQQLLMGTV represents the dominant epitope.

Tables

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  • Table 1.

    TIL recognize autologous tumor cellsa

    TargetBlocking MAbIFN-γ (pg/ml/106 cells)
    TumorNone0
    TILNone23
    Tumor + TILNone400
    W6/32400
    L24320
    • ↵a Autologous cervical cancer cells harboring HPV33 were tested for T-cell recognition as determined by IFN-γ and IL-2 secretion by ELISA. TIL secreted IFN-γ, but not IL-2 in response to tumor cells, which could be blocked with the anti-DR-directed MAb L243. Blocking with the anti-DR-directed MAb clone B8.12.2 showed a similar result in a parallel experiment.

  • Table 2.

    Deduced TCR amino acid sequencesa

    T-cell populationTCR chainVariable region sequenceCDR3 sequenceSequence of joining region
    PBL-VA CD8+ VA2DSQPSDSATYLCAVSGNDMRFGAG (AJ43)
    VA3* ADTASYFCASRGGDNTDKLIFGTG (AJ34)
    VA4TLRDAAVYYCILSFASAGGTSYGKLTFGQG (AJ52)
    VA9* FAQEEDSAMYYCAYGNNRLAFGKG (AJ7)
    VA16SALVSDSALYFCAVRHRIDTGRRALTFGSG (AJ5)
    VA21QPGDSAVYFCAASESGTSYGKLTFGQG (AJ52)
    VA23ASQPGDSATYLCAVRYLLSSGSARQLTFGSG (AJ22)
    VA25STYLCAVGPSSSGSARQLTFGSG (AJ22)
    PBL-VB CD8+ VB2SAHPEDSSFYICSARGGGSGNQPQHFGDG (BJ1-5)
    VB3SASTNQTSMYLCASSLGVGYEQYFGPG (BJ2-7)
    VB9LELGDSAVYFCASSQLWTSGAPPSTDTQYFGPG (BJ2-3)
    CD4+TIL-VAVA2FNTSCGTDYLCAMSSDRGSTLGRLYFGRG (AJ18)
    VA3* ADTASYFCASRGGDNTDKLIFGTG (AJ34)
    VA6ASQLGDSAMYFCAMREGWAAGNKLTFGGG (AJ17)
    VA9* FAQEEDSAMYYCAYGNNRLAFGKG (AJ7)
    VA10 (a)AAQPGDTGLYLCA GG SDGQKLLFARG (AJ16)
    VA10 (b)AAQPGDTGLYLCA GGPMDSSYKLIFGSG (AJ12)
    VA21PSQPGDSAVYFCA ASGG KLVFGKG (AJ57)
    VA26IYLCA GG SDGQKLLFARG (AJ16)
    CD4+TIL-VB16VB16PAELDSGVYFCASSRGTSGTLKQYFGPG (BJ2-7)
    TIL VB16 sortedVA10AAQPGDTGLYLCA GGPMDSSYKLIFGSG (AJ12)
    VA21PSQPGDSAVYFCA AS GGKLVFGKG (AJ57)
    VA22AVYFCALS YGCCRLAFGKG (AJ7)
    • ↵a Monoclonal TCRs in CD8+ or CD4+ T-cell populations were identified by DNA sequence analysis as described previously (18). Exclusively the CDR3 region and the adjacent variable and joining TCR segments are listed. Note that the CD4+ VB16+ population represents 96% of all T cells in TIL (Fig. 1). Sorting of these VB16+T cells yields a TCR VB16+ VA10+VA21+ T-cell line. Note that in unsorted TIL, two TCR VA10 chains could be identified, but only one of these was present in the VB16-sorted T-cell population [TCR VA10(b)]. *, identical TCR sequences in PBL and TIL. TCR VA chains combined with VB16 exhibit a common CDR3 motif: TCR VA10 and TCR VA21 share glycine residues (bold), and TCR VA21 shares a serine residue (underlined) with VA22.

  • Table 3.

    HPV33 E773–87 represents the dominant epitope defined by CD4+VB16+ TILa

    PeptideSequenceIFN-γ secretion (pg/ml per 106 cells per 24 h)
    Autologous cells (DR*0402)T2T2-DR*0401
    E7000
    E719–3 PTDLYCYEQLSDSSD000
    E764–78 TVRLCVNSTASDLRT000
    E773–87 ASDLRTIQQLLMGTV4,00004,800
    E762–76 NTTVRLCVNSTASDL000
    E75–19 KPTLKEYVLDLYPEP000
    E79–23 KEYVLDLYPEPTDLY000
    E776–90 LRTIQQLLMGTVNIV35000
    E779–93 IQQLLMGTVNIVVPT000
    TTQYIKANSKFIGITE000
    • ↵a Peptides were pulsed onto autologous antigen-presenting cells (1 μM), T2 cells, or T2 cells expressing the DR*0401 molecule and tested for T-cell recognition by IFN-γ secretion determined by ELISA. The HPV33 E773–87 peptide could be presented by either autologous antigen-presenting cells or T2 cells expressing DR*0401. Nontransfected T2 cells served as a negative control. A tetanus toxin peptide (TT) (28) was not recognized.

  • Table 4.

    HLA-DR-restricted recognition of HPV33 E773–87 epitope by CD4+ TILa

    Target cellsBlocking MAbIFN-γ secretion (pg/ml/2 × 106 cells per 24 h)
    Autologous macrophagesIgG4,800
    Anti-DR400
    T2-DR4IgG3,800
    Anti-DR0
    • ↵a The HPV33 E773–87 peptide was pulsed onto autologous antigen-presenting cells or T2 cells expressing HLA-DR*0401. Antigen-presenting cells were incubated with IgG or anti-DR-directed MAbs (5 μg/well), and an IFN-γ release assay was performed. Different anti-DR-directed MAbs (clone L243 and clone B8.12.2) exhibited similar results. Data are from a blocking experiment with the L243 MAb.

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Human Papillomavirus Type 33 E7 Peptides Presented by HLA-DR*0402 to Tumor-Infiltrating T Cells in Cervical Cancer
Hanni Höhn, Henryk Pilch, Susanne Günzel, Claudia Neukirch, Kirsten Freitag, Antje Necker, Markus J. Maeurer
Journal of Virology Jul 2000, 74 (14) 6632-6636; DOI: 10.1128/JVI.74.14.6632-6636.2000

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Human Papillomavirus Type 33 E7 Peptides Presented by HLA-DR*0402 to Tumor-Infiltrating T Cells in Cervical Cancer
Hanni Höhn, Henryk Pilch, Susanne Günzel, Claudia Neukirch, Kirsten Freitag, Antje Necker, Markus J. Maeurer
Journal of Virology Jul 2000, 74 (14) 6632-6636; DOI: 10.1128/JVI.74.14.6632-6636.2000
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KEYWORDS

HLA-DR Antigens
Lymphocytes, Tumor-Infiltrating
Oncogene Proteins, Viral
Papillomaviridae
Peptide Fragments
T-Lymphocytes
Uterine Cervical Neoplasms

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