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Tumor‐specific T‐cell clones recognize different protein determinants of autologous human malignant melanoma cells
Author(s) -
Notter Michael,
Schirrmacher Volker
Publication year - 1990
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910450508
Subject(s) - lymphoblast , antigen , microbiology and biotechnology , biology , tumor antigen , clone (java method) , lymphokine , cell culture , immunology , immune system , immunotherapy , biochemistry , dna , genetics
This study attempts to characterize human melanomaassociated tumor antigens that are recognized by autologous T‐lymphocyte clones. Peripheral blood lymphocytes of a melanoma patient (AV) were activated in an autologous‐mixedlymphocyte tumor culture (AMLTC) and responding cells were cloned using irradiated autologous melanoma cells (AV‐Mel) as antigen source, EBV‐transformed B‐lymphoblasts as feeder cells and recombinant Interleukin 2. T‐cell clones were isolated which proliferated in response to autologous tumor cells, but not to autologous B‐lymphoblasts (AV‐B), and which were cytolytic for AV‐Mel cells. In an attempt to identify the antigens recognized, an in vitro test system was used, in which 3 H‐Thymidine ( 3 H‐TdR) incorporation by T lymphocytes was measured in the presence of protein from AV‐Mel cells presented by irradiated autologous accessory cells. Antigenbearing particles of AV‐Mel or AV‐B cells were prepared by spotting cell lysates onto nitrocellulose (NC) followed by dissolution with DMSO and precipitation with an aqueous buffer. T cells sensitized against autologous melanoma cells were specifically stimulated by NC‐bound AV‐Mel protein. Stimulation required the presence of AV‐B accessory cells, indicating that B‐lymphoblasts are able to present tumor antigens. This approach facilitates screening of polypeptide fractions of AV‐Mel cells separated by polyacrylamide gel electrophoresis followed by Western blotting for their capacity to stimulate antigen‐dependent T‐cells. CD4 + and CD8 + tumorspecific clones appear to recognize different antigens on the tumor cell: proliferation of an antigen‐dependent CD8 + clone was stimulated by 240‐ and 24‐kDa protein fractions, while proliferation of 2 antigen‐dependent CD4 + clones was observed either with an 84‐ or with 140‐ and 55‐kDa fractions. Molecular definition of the different antigens of tumor cells recognized by autologous T cells may be a prerequisite in attempts to manipulate T‐cell‐mediated anti‐tumor responses in a controlled way.