title: A screening assay to identify agents that enhance T-cell recognition of human melanomas.
abstract: Although a series of melanoma differentiation antigens for immunotherapeutic targeting has been described, heterogeneous expression of antigens such as Melan-A/MART-1 and gp100 results from a loss of antigenic expression in many late stage tumors. Antigen loss can represent a means for tumor escape from immune recognition, and a barrier to immunotherapy. However, since antigen-negative tumor phenotypes frequently result from reversible gene regulatory events, antigen enhancement represents a potential therapeutic opportunity. Accordingly, we have developed a cell-based assay to screen for compounds with the ability to enhance T-cell recognition of melanoma cells. This assay is dependent on augmentation of MelanA/MART-1 antigen presentation by a melanoma cell line (MU89). T-cell recognition is detected as interleukin-2 production by a Jurkat T cell transduced to express a T-cell receptor specific for an HLA-A2 restricted epitope of the Melan-A/MART-1 protein. This cellular assay was used to perform a pilot screen by using 480 compounds of known biological activity. From the initial proof-of-principle primary screen, eight compounds were identified as positive hits. A panel of secondary screens, including orthogonal assays, was used to validate the primary hits and eliminate false positives, and also to measure the comparative efficacy of the identified compounds. This cell-based assay, thus, yields consistent results applicable to the screening of larger libraries of compounds that can potentially reveal novel molecules which allow better recognition of treated tumors by T cells.
authors.affiliation: CytoCure LLC , Beverly, Massachusetts 01915, USA. thaggerty@cytocure.com
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Haggerty
authors.forename: Ian S
authors.initials: IS
authors.lastname: Dunn
authors.forename: Lenora B
authors.initials: LB
authors.lastname: Rose
authors.forename: Estelle E
authors.initials: EE
authors.lastname: Newton
authors.forename: James T
authors.initials: JT
authors.lastname: Kurnick
mesh_terms: D000918:Antibody Specificity; D000951:Antigens, Neoplasm / Q000096:biosynthesis; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D016207:Cytokines / Q000378:metabolism; D004354:Drug Screening Assays, Antitumor / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D007167:Immunotherapy; D019169:Jurkat Cells; D016086:Lentivirus / Q000235:genetics; D058965:MART-1 Antigen / Q000276:immunology; D008545:Melanoma / Q000276:immunology* / Q000628:therapy; D011948:Receptors, Antigen, T-Cell / Q000187:drug effects; D013601:T-Lymphocytes / Q000276:immunology*; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7 / Q000032:analysis
chemical_list: D000951:Antigens, Neoplasm; D016207:Cytokines; D058965:MART-1 Antigen; D011948:Receptors, Antigen, T-Cell; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins
references.citation: J Immunol. 2005 Jun 1;174(11):6863-71
references.pmid: 15905528
references.citation: J Virol. 1998 Nov;72(11):8463-71
references.pmid: 9765382
references.citation: Curr Opin Chem Biol. 2010 Jun;14(3):412-20
references.pmid: 20409745
references.citation: Clin Exp Immunol. 2000 Jan;119(1):11-8
references.pmid: 10606959
references.citation: J Immunol. 2008 Oct 1;181(7):5128-36
references.pmid: 18802117
references.citation: J Immunol. 2005 Nov 1;175(9):6169-76
references.pmid: 16237114
references.citation: J Immunol. 1995 Mar 1;154(5):2257-65
references.pmid: 7868898
references.citation: Nat Rev Drug Discov. 2004 Aug;3(8):673-83
references.pmid: 15286734
references.citation: Nature. 1985 Aug 15-21;316(6029):606-9
references.pmid: 4033759
references.citation: Lab Invest. 1996 Jan;74(1):43-7
references.pmid: 8569196
references.citation: J Biomol Screen. 1999;4(2):67-73
references.pmid: 10838414
references.citation: Nat Rev Cancer. 2005 Oct;5(10):761-72
references.pmid: 16175177
references.citation: Blood. 2007 Nov 15;110(10):3564-72
references.pmid: 17660381
references.citation: Eur J Cancer. 1990;26(11-12):1152-6
references.pmid: 2149997
references.citation: J Clin Invest. 1986 May;77(5):1501-6
references.pmid: 2422208
references.citation: Pigment Cell Melanoma Res. 2010 Feb;23(1):27-40
references.pmid: 19995375
references.citation: Cancer Res. 1994 Feb 1;54(3):724-9
references.pmid: 8306334
references.citation: Biochem Biophys Res Commun. 1979 Jun 27;88(4):1194-202
references.pmid: 224865
references.citation: Mol Cancer Ther. 2008 Apr;7(4):737-9
references.pmid: 18375819
references.citation: Oncology. 1987;44(1):17-23
references.pmid: 3561923
references.citation: Melanoma Res. 1995 Oct;5(5):311-9
references.pmid: 8541721
references.citation: Biochem Pharmacol. 1992 Aug 18;44(4):805-13
references.pmid: 1324685
references.citation: Mol Cancer Res. 2003 Apr;1(6):411-9
references.pmid: 12692260
references.citation: J Immunol. 2003 Jul 1;171(1):166-74
references.pmid: 12816995
references.citation: J Immunol. 2003 May 15;170(10):5103-9
references.pmid: 12734356
references.citation: Nature. 1978 Oct 5;275(5679):458-60
references.pmid: 692726
references.citation: Annu Rev Immunol. 1994;12:337-65
references.pmid: 8011285
references.citation: Eur J Immunol. 2003 Feb;33(2):556-66
references.pmid: 12645955
references.citation: Cancer Res. 2007 Apr 15;67(8):3898-903
references.pmid: 17440104
references.citation: Int J Cancer. 1996 May 16;66(4):470-6
references.pmid: 8635862
references.citation: Hematol Oncol Clin North Am. 2009 Jun;23(3):447-65, viii
references.pmid: 19464596
references.citation: Methods Mol Biol. 2009;486:1-12
references.pmid: 19347612
references.citation: Melanoma Res. 1996 Oct;6(5):351-62
references.pmid: 8908595
references.citation: Cancer Res. 2005 Dec 1;65(23):10686-91
references.pmid: 16322212
references.citation: Trends Mol Med. 2006 Sep;12(9):406-14
references.pmid: 16899407
references.citation: Cell Growth Differ. 1997 Sep;8(9):935-40
references.pmid: 9300176
references.citation: Int J Cancer. 1999 Mar 1;80(5):781-90
references.pmid: 10048982
references.citation: Int Rev Immunol. 1997;14(2-3):173-92
references.pmid: 9131386
references.citation: Dermatol Res Pract. 2010;2010:
references.pmid: 20936153
references.citation: Int J Cancer. 2010 Oct 1;127(7):1625-36
references.pmid: 20099279
references.citation: J Immunol. 2007 Aug 15;179(4):2134-42
references.pmid: 17675472
references.citation: Pharmacol Rev. 1986 Dec;38(4):321-416
references.pmid: 2432624
references.citation: Nature. 2003 Sep 25;425(6956):407-10
references.pmid: 14508491
references.citation: Cancer Res. 1997 Aug 1;57(15):3223-9
references.pmid: 9242453
references.citation: J Exp Med. 2008 Jul 7;205(7):1687-700
references.pmid: 18573907
references.citation: Int J Cancer. 2002 May 1;99(1):7-13
references.pmid: 11948485
references.citation: J Immunol. 1995 Jan 15;154(2):762-71
references.pmid: 7814882
references.citation: Blood. 2007 Mar 15;109(6):2331-8
references.pmid: 17082316
references.citation: Immunity. 2004 Aug;21(2):137-48
references.pmid: 15308095
references.citation: Cancer Chemother Pharmacol. 1998;42(4):273-9
references.pmid: 9744771
references.citation: J Immunol. 1990 Jun 1;144(11):4487-95
references.pmid: 2160503
references.citation: Mol Cancer Res. 2006 Oct;4(10):779-92
references.pmid: 17050671
references.citation: J Biochem. 1997 Mar;121(3):612-8
references.pmid: 9133633
references.citation: J Exp Med. 1999 Sep 6;190(5):651-67
references.pmid: 10477550
references.citation: Oncogene. 2003 May 19;22(20):3035-41
references.pmid: 12789278
references.citation: Anticancer Res. 1984 Jul-Oct;4(4-5):229-34
references.pmid: 6091526
references.citation: J Exp Med. 1999 Dec 6;190(11):1669-78
references.pmid: 10587357
references.citation: J Biol Chem. 1982 Jul 10;257(13):7847-51
references.pmid: 7085651
references.citation: Int J Cancer. 1998 Feb 9;75(4):517-24
references.pmid: 9466650
references.citation: J Immunol. 1984 Jul;133(1):123-8
references.pmid: 6327821
references.citation: Nucleic Acids Res. 1993 Aug 25;21(17):3997-4004
references.pmid: 8371975
references.citation: Cancer Immunol Immunother. 2011 Jan;60(1):133-44
references.pmid: 21052994
references.citation: J Immunol. 1996 Jan 15;156(2):700-10
references.pmid: 8543823
references.citation: Clin Cancer Res. 2010 Feb 1;16(3):1042-8
references.pmid: 20086001
references.citation: Int J Cancer. 1997 Apr 10;71(2):142-7
references.pmid: 9139833
references.citation: Proc Natl Acad Sci U S A. 2006 Jan 3;103(1):57-62
references.pmid: 16371460
references.citation: Clin Immunol. 2006 May;119(2):135-45
references.pmid: 16458072
references.citation: Biochemistry. 1995 Sep 26;34(38):12404-10
references.pmid: 7547985
references.citation: Nature. 2001 May 17;411(6835):380-4
references.pmid: 11357146
references.citation: Invest Ophthalmol Vis Sci. 2008 Jun;49(6):2348-56
references.pmid: 18281615
references.citation: J Immunol. 2001 Aug 1;167(3):1204-11
references.pmid: 11466335
references.citation: Surgery. 1999 Aug;126(2):112-20
references.pmid: 10455872
references.citation: Proc Natl Acad Sci U S A. 1993 May 1;90(9):3933-7
references.pmid: 8483913
references.citation: Curr Opin Mol Ther. 2004 Feb;6(1):17-26
references.pmid: 15011777
references.citation: Nat Chem Biol. 2007 Aug;3(8):466-79
references.pmid: 17637779
references.citation: Clin Cancer Res. 2008 Dec 15;14(24):8302-7
references.pmid: 19088048
references.citation: Nat Med. 2008 Dec;14(12):1390-5
references.pmid: 18997777
references.citation: FEBS Lett. 1991 Aug 19;288(1-2):5-9
references.pmid: 1879564
references.citation: Nat Med. 2003 Oct;9(10):1269-74
references.pmid: 14502282
references.citation: Pharmacotherapy. 1985 Mar-Apr;5(2):43-62
references.pmid: 3923453