Cancer and the Immune System CHAPTER 22 509 OO Tamplin bs Tum+ cells do not express TSTA Tu TSTAs oo ad Clone #l(tum* Clone #2(tum) Clone #3(tum*) Tumor growth No tumor growth Tumor growth CTLs from mouse TSTA-specific CTls Prepare CDNA Transfect Incubate transfected Observe tum gene tum+ cells with TSTA tum- cells into tum+ cells specific CTL clone do not express Tunmr gene群 TSTA TSTA TSTA) FIGURE 22-7 One procedure for identifying genes encoding tumor- cell-mediated response against it. To isolate the gene encoding the specific transplantation antigens(TSTAs). Most TSTAs can be detected TSTA a cosmid gene library is prepared from the tum" cell line, the only by the cell-mediated rejection they elicit. In the first part of this pro- genes are transfected into tumorigenic tum cells, and the transfected cedure, a nontumorigenic(tum")cell line is generated; this cell line ex. cells are incubated with TSTA-specific CTLS. sses a TSTa that is recognized by syngeneic mice, which mount a tumor in syngeneic mice. These mutant tumor cells are desig- the genes encoding the TSTAs that are expressed on a tum" cell nated as tum variants. Most tum" variants have been shown line, a cosmid dna library is prepared from the tum cells. to express TSTAs that are not expressed by the original tum Genes from the tum cells are transfected into the original tumor-cell line. When tum cells are injected into syngeneic tum* cells. The transfected tum" cells are tested for the expres- mice, the unique TSTAs that the tum" cells express are recog- sion of the tum TSTAs by their ability to activate cloned nized by specific CTIs. The TSTA-specific CTLs destroy the CTLs specific for the tum TSTA. A number of diverse TSTAs tum tumor cells, thus preventing tumor growth. To identify have been identified by this method.tumor in syngeneic mice. These mutant tumor cells are desig￾nated as tum– variants. Most tum– variants have been shown to express TSTAs that are not expressed by the original tum+ tumor-cell line. When tum– cells are injected into syngeneic mice, the unique TSTAs that the tum– cells express are recog￾nized by specific CTLs. The TSTA-specific CTLs destroy the tum– tumor cells, thus preventing tumor growth. To identify the genes encoding the TSTAs that are expressed on a tum– cell line, a cosmid DNA library is prepared from the tum– cells. Genes from the tum– cells are transfected into the original tum+ cells. The transfected tum+ cells are tested for the expres￾sion of the tum– TSTAs by their ability to activate cloned CTLs specific for the tum– TSTA. A number of diverse TSTAs have been identified by this method. Cancer and the Immune System CHAPTER 22 509 Clone #1 (tum+) No tumor growth Clone #2 (tum–) Clone #3 (tum+) TSTAs Mutagenize and clone Tum+ cells Tumor growth Tumor growth Isolate and clone CTLs from mouse TSTA–specific CTLs Incubate transfected tum+ cells with TSTA specific CTL clone Prepare cDNA library from tum– cells Transfect tum– gene into tum+ cells Tum– gene #1 Tum– gene #2 Tum– TSTA gene Tumor growth Tum+ cells do not express TSTA Transplant Observe for lysis No lysis (cells do not express TSTA) No lysis (cells do not express TSTA) Lysis (cells express TSTA) FIGURE 22-7 One procedure for identifying genes encoding tumor￾specific transplantation antigens (TSTAs). Most TSTAs can be detected only by the cell-mediated rejection they elicit. In the first part of this pro￾cedure, a nontumorigenic (tum– ) cell line is generated; this cell line ex￾presses a TSTA that is recognized by syngeneic mice, which mount a cell-mediated response against it. To isolate the gene encoding the TSTA, a cosmid gene library is prepared from the tum– cell line, the genes are transfected into tumorigenic tum+ cells, and the transfected cells are incubated with TSTA-specific CTLs