HLA-A2 ASSOCIATED PEPTIDE EPITOPES FOR CTL ON MELANOMA

拟议项目的长期目标是确定 以人黑色素瘤细胞为表位表达的抗原(S)(S) 用于黑色素瘤特异性细胞毒性T淋巴细胞(CTL)。临床部 这些抗原的鉴定和合成的意义 包括(1)创造安全的重组疫苗的可能性 黑色素瘤，(2)体外刺激人淋巴细胞的可能性 使用一种特定的刺激剂，对黑色素瘤患者进行收养转移 黑色素瘤特异性CTL的可能性，以及(3)识别 蛋白质)和产生这些抗原的遗传物质， 这本身就会对理解恶性 黑色素瘤的转化。 将从人白血病细胞表面和细胞质中提纯人类白细胞抗原A2 至少10(9)个人黑色素瘤细胞。用作以下用途的制剂 对照将从A2()非黑色素瘤细胞中提纯。这些多肽 结合在A2分子缝隙中的分子将被酸化洗脱 和过滤离心法。得到的多肽混合物将是 电喷雾电离条件下的质谱分析。 质谱学鉴定黑色素瘤上存在的多肽 非黑色素瘤细胞中没有的细胞将按质量进行测序 光谱分析方法。测序后的多肽将被合成并 测试他们使A2()非黑色素瘤对 人类白细胞抗原A2限制性黑色素瘤特异性细胞毒性T淋巴细胞的裂解。

The long-term objective of the proposed project is to identify the antigen(s) expressed by human melanoma cells that serve as epitope(s) for melanoma-specific cytotoxic T-lymphocytes (CTL). The clinical implications of the identification and synthesis of these antigens include (1) the possibility of creating a safe recombinant vaccine for melanoma, (2) the possibility of stimulating human lymphocytes in vitro for adoptive transfer to melanoma patients, using a specific stimulant of melanoma-specific CTL, and (3) the possibility of identifying the proteins) and genetic material from which these antigens are derived, which would in itself have implications for understanding malignant transformation of melanoma. HLA-A2 antigens will be purified from the cell surface and cytoplasm of a least 10(9) HLA-A2(+) human melanoma cells. Preparations serving as controls will be purified from A2(+) non-melanoma cells. The peptides bound in the cleft of the A2 molecules will be eluted by acidification and filtration centrifugation. The resulting peptide mixtures will be analyzed by mass spectrometry under electrospray ionization conditions. Peptides identified by mass spectrometry that are present on melanoma cells but absent on non-melanoma cells will be sequenced by mass spectrometric methods. The sequenced peptides will be synthesized and tested for their ability to render A2(+) non-melanomas sensitive to lysis by HLA-A2-restricted melanoma-specific cytotoxic T lymphocytes.