. Labarriere, This protocol was approved by the Institutional Ethics Committee and registered with the regulatory state authority in France (Nantes). Moreover, TILs were expended in a specific genetic and cellular therapy unit, Human Tissue Specimens Tumor-infiltrating lymphocytes (TILs) were provided by Pr. F, 2002.

, France) under good manufacturing practice (GMP) conditions. Blood from healthy donors was purchased from the New York Blood Center, Nantes

, 50%]) was purchased from Biomol. The MMP-2 enzyme was inactivated either by heating to 56 C for 45min or by addition of the MMP-2 inhibitor III at 100nM (Calbiochem) for 20 min. rhPEX was purchased from Genway. rhMMP-9 (Calbiochem) was used as a mixture of the proenzyme (50%) and the active protein (50%), p.80

, Lyophilized peptides were reconstituted in DMSO and were used either individually (2 mM) or as a pool (2 mM each). rhGM-CSF was purchased from Immunex. rhIL-12, rhIL-4, rhIL-7, and rhIL-2 were from R&D Systems. The kits used for basophil isolation and IFN-g-secreting cell enrichment

, Antibodies to human Vb chains covering 70% of the repertoire (IOTest Beta Mark; Beckman Coulter) were used according to the manufacturer's intructions. Fluorescein isothiocyanate-conjugated antibody to IL-12p35/IL-12p70 (B-T21) and IFNAR1 (EP899Y) were from Abcam. Phycoerythrine-conjugated antibodies to IL-12p40/IL-12p70 (C8.6), IL-10 (B-T10), CD203c (FR3-16A11) were from Miltenyi Biotec. Blocking antibody to HLA-DQ (SPV-L3; 20 mg/ml) was from NeoMarkers. Blocking antibodies to HLA-DP (B7/21; 20 mg/ml) or HLA-DR (L243; 20 mg/ml), phycoerythrine-conjugated antibodies to phosphorylated (Y701) STAT1 (4a), TNF-a (MAb11), IFN-g (25723.11), perforin (dG9), GranzymeB (GB11), CD40 (5C3), CD80 (L307.4), CD83 (HB15e), CD86 (IT2.2), HLA-DR (TU36), fluorescein isothiocyanate-conjugated antibodies to CD45RA (HI100) and CD45RO (UCHL1), and antibody to CD4 (RPA-T4) were purchased from BD Biosciences PharMingen. Alexa fluor 488-conjugated antibody to IL-17 (eBio64DEC17)

. T-cell-culture, Stimulation and Priming Peripheral blood mononuclear cells (PBMCs) were purified from healthy donor-(HD) or cord blood (CB) donor-derived buffy coats (New York Blood Center) by Ficoll-Paque Plus (GE Healthcare) centrifugation. CD4 + /CD25 -cells were enriched (>90%) by magnetic cell sorting (Miltenyi Biotec) and primed/ stimulated for 12-15 days either with irradiated (35 Gy) autologous CD4 -cells or with autologous mature DCs in IMDM (GIBCO) supplemented with 1 mM HEPES (Life Technologies

. Godefroy, involving enrichment of IFN-g-secreting cells upon short-term culture and peptide stimulation to generate MMP-2 responsive clones. Although MMP-2-specific cells could be isolated, it was realized that IFN-g secretion was marginal compared with their secretion of IL-4 and TNF-a. IFN-gsecreting cells in response to MMP-2 peptide pool were enriched by cytokine-guided magnetic cell sorting (Miltenyi Biotec) and cloned the following day by limiting dilution in the presence of irradiated allogeneic PBMCs, 1 mg/ml phytohemagglutinin-L (Sigma) and 150 UI/ml rhIL-2. Tumor infiltrating lymphocytes (TILs) were provided by, UI/ml)/penicillin (100 mg/ml) (Sigma) and 5% heat inactivated pooled human serum (PHS; Valley Biomedical) in the presence of rhIL-2 (10 UI/ml) and IL7 (5 ng/ml) (R&D Systems), 2002.

, 10 6 cells/10 ml/dish in complete IMDM with 5% PHS. Cells were allowed to adhere for 2 hr at 37 C. Nonadherent cells were removed. The monocyteenriched fraction was supplemented with 100 UI/ml rhGM-CSF and 300 UI/ml rhIL-4 (R&D Systems) on days 0, 2, and 4. Immature DCs were harvested on day 5 and matured using poly(I:C) at 5 mg/ml/10 6 DCs (Amersham). Secretion of IL-12p70, Dendritic Cell Preparation and Activation PBMCs were purified from healthy-(HD) or cord blood-(CB) donors and plated at 40

, 000 cells/100 ml/well), polyclonal T cell populations (100,000 cells/100 ml/well), DCs (50,000 cells/100 ml/well), and basophils (10,000 cells/100 ml/well) was determined by ELISA, Enzyme-Linked Immunosorbent Assay Activation of T cell clones

R. Botella-estrada, M. Escudero, J. E. O'connor, E. Nagore, B. Fenollosa et al., Cytokine production by peripheral lymphocytes in melanoma, Eur. Cytokine Netw, vol.16, pp.47-55, 2005.

P. C. Brooks, S. Stromblad, L. C. Sanders, T. L. Von-schalscha, R. T. Aimes et al., Localization of matrix metalloproteinase MMP-2 to the surface of invasive cells by interaction with integrin alpha v beta 3, Cell, vol.85, pp.683-693, 1996.

P. C. Brooks, S. Silletti, T. L. Von-schalscha, M. Friedlander, and D. A. Cheresh, Disruption of angiogenesis by PEX, a noncatalytic metalloproteinase fragment with integrin binding activity, Cell, vol.92, pp.391-400, 1998.

W. S. Choi, O. H. Jeon, H. H. Kim, and D. S. Kim, MMP-2 regulates human platelet activation by interacting with integrin alphaIIbbeta3, J. Thromb. Haemost, vol.6, pp.517-523, 2008.

P. G. Coulie and P. Van-der-bruggen, T-cell responses of vaccinated cancer patients, Curr. Opin. Immunol, vol.15, pp.131-137, 2003.

T. Decker, M. Muller, and S. Stockinger, The yin and yang of type I interferon activity in bacterial infection, Nat. Rev. Immunol, vol.5, pp.675-687, 2005.

H. Emonard, G. Bellon, L. Troeberg, A. Berton, A. Robinet et al., Low density lipoprotein receptor-related protein mediates endocytic clearance of pro-MMP-2.TIMP-2 complex through a thrombospondin-independent mechanism, J. Biol. Chem, vol.279, pp.54944-54951, 2004.

B. Everts, G. Perona-wright, H. H. Smits, C. H. Hokke, A. J. Van-der-ham et al., Omega-1, a glycoprotein secreted by Schistosoma mansoni eggs, drives Th2 responses, J. Exp. Med, vol.206, pp.1673-1680, 2009.

F. H. Falcone, C. A. Dahinden, B. F. Gibbs, T. Noll, U. Amon et al., Human basophils release interleukin-4 after stimulation with Schistosoma mansoni egg antigen, Eur. J. Immunol, vol.26, pp.1147-1155, 1996.

G. Gautier, M. Humbert, F. Deauvieau, M. Scuiller, J. Hiscott et al., A type I interferon autocrineparacrine loop is involved in Toll-like receptor-induced interleukin-12p70 secretion by dendritic cells, J. Exp. Med, vol.201, pp.1435-1446, 2005.

E. Godefroy, A. Moreau-aubry, E. Diez, B. Dreno, F. Jotereau et al., alpha v beta3-dependent cross-presentation of matrix metalloproteinase-2 by melanoma cells gives rise to a new tumor antigen, J. Exp. Med, vol.202, pp.61-72, 2005.

E. Godefroy, L. Scotto, N. E. Souleimanian, G. Ritter, L. J. Old et al., Identification of two Melan-A CD4+ T cell epitopes presented by frequently expressed MHC class II alleles, Clin. Immunol, vol.121, pp.54-62, 2006.

E. Godefroy, Y. Wang, N. E. Souleimanian, L. Scotto, S. Stevanovic et al., Assessment of CD4+ T cells specific for the tumor antigen SSX-1 in cancer-free individuals, Cancer Immunol. Immunother, vol.56, pp.1183-1192, 2007.

K. Grobe, W. M. Becker, M. Schlaak, and A. Petersen, Grass group I allergens (beta-expansins) are novel, papain-related proteinases, Eur. J. Biochem, vol.263, pp.33-40, 1999.

Y. Hirohashi, T. Torigoe, S. Inoda, J. Kobayasi, M. Nakatsugawa et al., The functioning antigens: beyond just as the immunological targets, Cancer Sci, vol.100, pp.798-806, 2009.

T. Ito, Y. H. Wang, O. Duramad, T. Hori, G. J. Delespesse et al., TSLP-activated dendritic cells induce an inflammatory T helper type 2 cell response through OX40 ligand, J. Exp. Med, vol.202, pp.1213-1223, 2005.

T. Itoh, M. Tanioka, H. Yoshida, T. Yoshioka, H. Nishimoto et al., Reduced angiogenesis and tumor progression in gelatinase A-deficient mice, Cancer Res, vol.58, pp.1048-1051, 1998.

D. Jankovic, M. C. Kullberg, N. Noben-trauth, P. Caspar, W. E. Paul et al., Single cell analysis reveals that IL-4 receptor/Stat6 signaling is not required for the in vivo or in vitro development of CD4+ lymphocytes with a Th2 cytokine profile, J. Immunol, vol.164, pp.3047-3055, 2000.

K. Kessenbrock, V. Plaks, and Z. Werb, Matrix metalloproteinases: regulators of the tumor microenvironment, Cell, vol.141, pp.52-67, 2010.

A. Khammari, J. M. Nguyen, M. C. Pandolfino, G. Quereux, A. Brocard et al., Long-term follow-up of patients treated by adoptive transfer of melanoma tumor-infiltrating lymphocytes as adjuvant therapy for stage III melanoma, Cancer Immunol. Immunother, vol.56, pp.1853-1860, 2007.

F. Kheradmand, A. Kiss, J. Xu, S. H. Lee, P. E. Kolattukudy et al., A protease-activated pathway underlying Th cell type 2 activation and allergic lung disease, J. Immunol, vol.169, pp.5904-5911, 2002.

N. Labarriere, M. C. Pandolfino, N. Gervois, A. Khammari, M. H. Tessier et al., Therapeutic efficacy of melanoma-reactive TIL injected in stage III melanoma patients, Cancer Immunol. Immunother, vol.51, pp.532-538, 2002.
URL : https://hal.archives-ouvertes.fr/inserm-02482115

L. Lauerova, L. Dusek, M. Simickova, I. Kocak, M. Vagundova et al., Malignant melanoma associates with Th1/Th2 imbalance that coincides with disease progression and immunotherapy response, Neoplasma, vol.49, pp.159-166, 2002.

L. A. Liotta, K. Tryggvason, S. Garbisa, I. Hart, C. M. Foltz et al., Metastatic potential correlates with enzymatic degradation of basement membrane collagen, Nature, vol.284, pp.67-68, 1980.

Y. J. Liu, Thymic stromal lymphopoietin: master switch for allergic inflammation, J. Exp. Med, vol.203, pp.269-273, 2006.

Y. J. Liu, V. Soumelis, N. Watanabe, T. Ito, Y. H. Wang et al., TSLP: an epithelial cell cytokine that regulates T cell differentiation by conditioning dendritic cell maturation, Annu. Rev. Immunol, vol.25, pp.193-219, 2007.

D. Loose and C. Van-de-wiele, The immune system and cancer, Cancer Biother. Radiopharm, vol.24, pp.369-376, 2009.

M. Mccarter, J. Clarke, D. Richter, W. , and C. , Melanoma skews dendritic cells to facilitate a T helper 2 profile, Surgery, vol.138, pp.321-328, 2005.

, Cancer Cell MMP-2-Dependent Immune Deviation in Melanoma Cancer Cell, vol.19, pp.333-346, 20112011-03-15.

J. H. Mckerrow, C. Caffrey, B. Kelly, P. Loke, and M. Sajid, Proteases in parasitic diseases, Annu. Rev. Pathol, vol.1, pp.497-536, 2006.

K. Minkis, D. G. Kavanagh, G. Alter, D. Bogunovic, D. O'neill et al., Type 2 Bias of T cells expanded from the blood of melanoma patients switched to type 1 by IL-12p70 mRNA-transfected dendritic cells, Cancer Res, vol.68, pp.9441-9450, 2008.

M. Moser and K. M. Murphy, Dendritic cell regulation of TH1-TH2 development, Nat. Immunol, vol.1, pp.199-205, 2000.

T. R. Mosmann and R. L. Coffman, TH1 and TH2 cells: different patterns of lymphokine secretion lead to different functional properties, Annu. Rev. Immunol, vol.7, pp.145-173, 1989.

T. Nishimura, K. Iwakabe, M. Sekimoto, Y. Ohmi, T. Yahata et al., Distinct role of antigen-specific T helper type 1 (Th1) and Th2 cells in tumor eradication in vivo, J. Exp. Med, vol.190, pp.617-627, 1999.

T. Nishimura, M. Nakui, M. Sato, K. Iwakabe, H. Kitamura et al., The critical role of Th1-dominant immunity in tumor immunology, Cancer Chemother. Pharmacol. Suppl, vol.46, pp.52-61, 2000.

A. Ocmant, Y. Peignois, S. Mulier, L. Hanssens, A. Michils et al., Flow cytometry for basophil activation markers: the measurement of CD203c up-regulation is as reliable as CD63 expression in the diagnosis of cat allergy, J. Immunol. Methods, vol.320, pp.40-48, 2007.

K. Oh, T. Shen, G. Le-gros, M. , and B. , Induction of Th2 type immunity in a mouse system reveals a novel immunoregulatory role of basophils, Blood, vol.109, pp.2921-2927, 2007.

S. A. Rosenberg, Shedding light on immunotherapy for cancer, N. Engl. J. Med, vol.350, pp.1461-1463, 2004.

M. Severa, M. E. Remoli, E. Giacomini, J. Ragimbeau, R. Lande et al., Differential responsiveness to IFNalpha and IFN-beta of human mature DC through modulation of IFNAR expression, J. Leukoc. Biol, vol.79, pp.1286-1294, 2006.
URL : https://hal.archives-ouvertes.fr/pasteur-00161981

T. So, J. Song, K. Sugie, A. Altman, and M. Croft, Signals from OX40 regulate nuclear factor of activated T cells c1 and T cell helper 2 lineage commitment, Proc. Natl. Acad. Sci. USA, vol.103, pp.3740-3745, 2006.

C. L. Sokol, G. M. Barton, A. G. Farr, and R. Medzhitov, A mechanism for the initiation of allergen-induced T helper type 2 responses, Nat. Immunol, vol.9, pp.310-318, 2008.

C. L. Sokol, N. Q. Chu, S. Yu, S. A. Nish, T. M. Laufer et al., Basophils function as antigen-presenting cells for an allergen-induced T helper type 2 response, Nat. Immunol, vol.10, pp.713-720, 2009.

V. Soumelis and Y. J. Liu, Human thymic stromal lymphopoietin: a novel epithelial cell-derived cytokine and a potential key player in the induction of allergic inflammation, Springer Semin. Immunopathol, vol.25, pp.325-333, 2004.

V. Soumelis, P. A. Reche, H. Kanzler, W. Yuan, G. Edward et al., Human epithelial cells trigger dendritic cell mediated allergic inflammation by producing TSLP, Nat. Immunol, vol.3, pp.673-680, 2002.

M. Stefanidakis, M. Bjorklund, E. Ihanus, C. G. Gahmberg, and E. Koivunen, Identification of a negatively charged peptide motif within the catalytic domain of progelatinases that mediates binding to leukocyte beta 2 integrins, J. Biol. Chem, vol.278, pp.34674-34684, 2003.

S. Steinfelder, J. F. Andersen, J. L. Cannons, C. G. Feng, M. Joshi et al., The major component in schistosome eggs responsible for conditioning dendritic cells for Th2 polarization is a T2 ribonuclease (omega-1), J. Exp. Med, vol.206, pp.1681-1690, 2009.

S. L. Swain, A. D. Weinberg, M. English, and G. Huston, IL-4 directs the development of Th2-like helper effectors, J. Immunol, vol.145, pp.3796-3806, 1990.

S. J. Szabo, S. T. Kim, G. L. Costa, X. Zhang, C. G. Fathman et al., A novel transcription factor, T-bet, directs Th1 lineage commitment, Cell, vol.100, pp.655-669, 2000.

T. Tatsumi, L. S. Kierstead, E. Ranieri, L. Gesualdo, F. P. Schena et al., Disease-associated bias in T helper type 1 (Th1)/Th2 CD4(+) T cell responses against MAGE-6 in HLA-DRB10401(+) patients with renal cell carcinoma or melanoma, J. Exp. Med, vol.196, pp.619-628, 2002.

N. Watanabe, S. Hanabuchi, V. Soumelis, W. Yuan, S. Ho et al., Human thymic stromal lymphopoietin promotes dendritic cell-mediated CD4+ T cell homeostatic expansion, Nat. Immunol, vol.5, pp.426-434, 2004.

J. Westermarck and V. M. Kahari, Regulation of matrix metalloproteinase expression in tumor invasion, FASEB J, vol.13, pp.781-792, 1999.

C. Yee, J. A. Thompson, P. Roche, D. R. Byrd, P. P. Lee et al., , 2000.

, Melanocyte destruction after antigen-specific immunotherapy of melanoma: direct evidence of t cell-mediated vitiligo, J. Exp. Med, vol.192, pp.1637-1644

Q. Yu, A. Sharma, S. Y. Oh, H. G. Moon, M. Z. Hossain et al., T cell factor 1 initiates the T helper type 2 fate by inducing the transcription factor GATA-3 and repressing interferon-gamma, Nat. Immunol, vol.10, pp.992-999, 2009.