ART TK Cancer and the chapter 22 Immune system S THE DEATH TOLL FROM INFECTIOUS DISEASE has declined in the Western world, cancer has become the second-ranking cause of death there. led only by heart disease. Current estimates project that one person in three in the United States will develop cancer, that one in five will die from it. From an immunologic per spective, cancer cells can be viewed as altered self-cells that have escaped normal growth-regulating mechanisms. This chapter examines the unique properties of cancer cells, pay. ing particular attention to those properties that can be recog- Cancerous melanoma cells nized by the immune system. The immune responses that develop to cancer cells, as well as the methods by which can cers manage to evade those responses, are then described. Cancer: Origin and Terminolo The final section describes current clinical and experimental a Malignant Transformation of Cells unotherapies for cancer. Oncogenes and Cancer Induction a Tumors of the Immune System Cancer: Origin and Terminology s Tumor Antigens In most organs and tissues of a mature animal, a balance is Immune Response to Tumors usually maintained between cell renewal and cell death. The Tumor Evasion of the Immune System various types of mature cells in the body have a given life span: as these cells die, new cells are generated by the prolif- Cancer Immunotherapy eration and differentiation of various types of stem cells. Under normal circumstances, the production of new cells is regulated so that the number of any particular type of cell constant. Occasionally, though, cells arise that longer respond to normal growth-control mechanisms. These cells give rise to clones of cells that can expand to a consider- of cancers of the colon, breast, prostate, and lung are carci- able size, producing a tumor, or neoplasm. nomas. The leukemias and lymphomas are malignant tu- A tumor that is not capable of indefinite growth and does mors of hematopoietic cells of the bone marrow and ac- not invade the healthy surrounding tissue extensively is be- count for about 9% of cancer incidence in the United States nign. a tumor that continues to grow and becomes progres- Leukemias proliferate as single cells, whereas lymphomas sively invasive is malignant; the term cancer refers speci- tend to grow as tumor masses. Sarcomas, which arise less fically to a malignant tumor. In addition to uncontrolled frequently (around 1% of the incidence in the United States) growth, malignant tumors exhibit metastasis; in this pro- are derived from mesodermal connective tissues such as ess, small clusters of cancerous cells dislodge from a tumor, bone, fat, and cartilage evade the blood or lymphatic vessels, and are carried to other tissues, where they continue to proliferate. In this way a primary tumor at one site can give rise to a secondary Malignant Transformation of Cells Malignant tumors or cancers are classified according to Treatment of normal cultured cells with chemical carcino- he embryonic origin of the tissue from which the tumor is gens, irradiation, and certain viruses can alter their mor derived. Most(>80%)are carcinomas, tumors that arise phology and growth properties. In some cases this process from endodermal or ectodermal tissues such as skin or the referred to as transformation, makes the cells able to pro- epithelial lining of internal organs and glands. The majority duce tumors when they are injected into animals. Such cells
■ Cancer: Origin and Terminology ■ Malignant Transformation of Cells ■ Oncogenes and Cancer Induction ■ Tumors of the Immune System ■ Tumor Antigens ■ Immune Response to Tumors ■ Tumor Evasion of the Immune System ■ Cancer Immunotherapy Cancerous melanoma cells. Cancer and the Immune System A has declined in the Western world, cancer has become the second-ranking cause of death there, led only by heart disease. Current estimates project that one person in three in the United States will develop cancer, and that one in five will die from it. From an immunologic perspective, cancer cells can be viewed as altered self-cells that have escaped normal growth-regulating mechanisms. This chapter examines the unique properties of cancer cells, paying particular attention to those properties that can be recognized by the immune system. The immune responses that develop to cancer cells, as well as the methods by which cancers manage to evade those responses, are then described. The final section describes current clinical and experimental immunotherapies for cancer. Cancer: Origin and Terminology In most organs and tissues of a mature animal, a balance is usually maintained between cell renewal and cell death. The various types of mature cells in the body have a given life span; as these cells die, new cells are generated by the proliferation and differentiation of various types of stem cells. Under normal circumstances, the production of new cells is regulated so that the number of any particular type of cell remains constant. Occasionally, though, cells arise that no longer respond to normal growth-control mechanisms. These cells give rise to clones of cells that can expand to a considerable size, producing a tumor, or neoplasm. A tumor that is not capable of indefinite growth and does not invade the healthy surrounding tissue extensively is benign. A tumor that continues to grow and becomes progressively invasive is malignant; the term cancer refers specifically to a malignant tumor. In addition to uncontrolled growth, malignant tumors exhibit metastasis; in this process, small clusters of cancerous cells dislodge from a tumor, invade the blood or lymphatic vessels, and are carried to other tissues, where they continue to proliferate. In this way a primary tumor at one site can give rise to a secondary tumor at another site (Figure 22-1). Malignant tumors or cancers are classified according to the embryonic origin of the tissue from which the tumor is derived. Most (>80%) are carcinomas, tumors that arise from endodermal or ectodermal tissues such as skin or the epithelial lining of internal organs and glands. The majority of cancers of the colon, breast, prostate, and lung are carcinomas. The leukemias and lymphomas are malignant tumors of hematopoietic cells of the bone marrow and account for about 9% of cancer incidence in the United States. Leukemias proliferate as single cells, whereas lymphomas tend to grow as tumor masses. Sarcomas, which arise less frequently (around 1% of the incidence in the United States), are derived from mesodermal connective tissues such as bone, fat, and cartilage. Malignant Transformation of Cells Treatment of normal cultured cells with chemical carcinogens, irradiation, and certain viruses can alter their morphology and growth properties. In some cases this process, referred to as transformation, makes the cells able to produce tumors when they are injected into animals. Such cells chapter 22 ART TK
502 RT Iv The Immune System in Health and Disease VISUALIZING CONCEPTS Initially modified tumor cell (b) Mass of tumor cells (localized benign Tumor cells metastasis to occur carried by the blood or lymph to other sites in the ign tumor. ( c) The tumor cells become progressively more inva- body /Adapted from 1 Amell et al., 1990, Molecular Cell Biology sive, invading the underlying basal lamina. The tumor is now 2d ed, Scientific American Books] are said to have undergone malignant transformation, and physical carcinogens appears to involve multiple steps and at they often exhibit properties in vitro similar to those of can- least two distinct phases: initiation and promotion. Initiation cer cells. For example, they have decreased requirements for involves changes in the genome but does not, in itself, lead to rowth factors and serum, are no longer anchorage-dependent, malignant transformation. After initiation, promoters stimu and grow in a density-independent fashion. Moreover, both late cell division and lead to malignant transformation. cancer cells and transformed cells can be subcultured indefi- The importance of mutagenesis in the induction of cancer nitely, that is, for all practical purposes, they are immortal. is illustrated by diseases such as xeroderma pigmentosum. Because of the similar properties of cancer and transformed This rare disorder is caused by a defect in the gene that en- cells, the process of malignant transformation has been stud- codes a DNA-repair enzyme called UV-specific endonuclease. ied extensively as a model of cancer induction. Individuals with this disease are unable to repair UV-induced Various chemical agents(e.g, DNA-alkylating reagents)and mutations and consequently develop skin cancers. physical agents (e.g, ultraviolet ight and ionizing radiation) A number of DNA and RNa viruses have been shown to that cause mutations have been shown to induce transforma- induce malignant transformation. Two of the best-studied tion. Induction of malignant transformation with chemical or are SV40 and polyoma. In both cases the viral genomes
are said to have undergone malignant transformation, and they often exhibit properties in vitro similar to those of cancer cells. For example, they have decreased requirements for growth factors and serum, are no longer anchorage-dependent, and grow in a density-independent fashion. Moreover, both cancer cells and transformed cells can be subcultured indefinitely; that is, for all practical purposes, they are immortal. Because of the similar properties of cancer and transformed cells, the process of malignant transformation has been studied extensively as a model of cancer induction. Various chemical agents (e.g., DNA-alkylating reagents) and physical agents (e.g., ultraviolet light and ionizing radiation) that cause mutations have been shown to induce transformation. Induction of malignant transformation with chemical or physical carcinogens appears to involve multiple steps and at least two distinct phases: initiation and promotion. Initiation involves changes in the genome but does not, in itself, lead to malignant transformation. After initiation, promoters stimulate cell division and lead to malignant transformation. The importance of mutagenesis in the induction of cancer is illustrated by diseases such as xeroderma pigmentosum. This rare disorder is caused by a defect in the gene that encodes a DNA-repair enzyme called UV-specific endonuclease. Individuals with this disease are unable to repair UV-induced mutations and consequently develop skin cancers. A number of DNA and RNA viruses have been shown to induce malignant transformation. Two of the best-studied are SV40 and polyoma. In both cases the viral genomes, 502 PART IV The Immune System in Health and Disease VISUALIZING CONCEPTS (a) (b) (c) (d) Initially modified tumor cell Invasive tumor cells Mass of tumor cells (localized benign tumor) Basal lamina Blood vessel Tumor cells invade blood vessels, allowing metastasis to occur FIGURE 22-1 Tumor growth and metastasis. (a) A single cell develops altered growth properties at a tissue site. (b) The altered cell proliferates, forming a mass of localized tumor cells, or benign tumor. (c) The tumor cells become progressively more invasive, invading the underlying basal lamina. The tumor is now classified as malignant. (d) The malignant tumor metastasizes by generating small clusters of cancer cells that dislodge from the tumor and are carried by the blood or lymph to other sites in the body. [Adapted from J. Darnell et al., 1990, Molecular Cell Biology, 2d ed., Scientific American Books.]
Cancer and the Immune System CHAPTER 22 503 which integrate randomly into the host chromosomal dna, gene and its corresponding proto-oncogene appear to have include several genes that are expressed early in the course of very similar functions. As described below, the conversion of viral replication. SV40 encodes two early proteins called large a proto-oncogene into an oncogene appears in many cases to Tand little T, and polyoma encodes three early proteins called accompany a change in the level of expression of a normal large T, middle T, and little T. Each of these proteins plays a growth-controlling protein. role in the malignant transformation of virus-infected cells Most RNa viruses replicate in the cytosol and do not Cancer-Associated Genes Have induce malignant transtormation. The exceptions are retro. Many Functions reverse-transcriptase enzyme and then integrate the tran- Homeostasis in normal tissue is maintained by a highly reg- script into the host's DNA. This process is similar in the cyto- ulated process of cellular proliferation balanced by cell death. pathic retroviruses such as HIV-1 and HIV-2 and in the If there is an imbalance, either at the stage of cellular prolif- transforming retroviruses, which induce changes in the host eration or at the stage of cell death, then a cancerous state will cell that lead to malignant transformation. In some cases, develop Oncogenes and tumor suppressor genes have been retrovirus-induced transformation is related to the presence shown to play an important role in this process, by regulating of oncogenes, or"cancer genes "carried by the retrovirus. either cellular proliferation or cell death. Cancer-associated One of the best-studied transforming retroviruses is the genes can be divided into three categories that reflect these Rous sarcoma virus. This virus carries an oncogene called different activities, summarized in Table 22-1 v-src, which encodes a 60-kDa protein kinase(v-Src)that cat- alyzes the addition of phosphate to tyrosine residues on pro- INDUCTION OF CELLULAR PROLIFERATION teins. The first evidence that oncogenes alone could induce One category of proto-oncogenes and their oncogenic coun- malignant transformation came from studies of the v-src on- terparts encodes proteins that induce cellular proliferation. gene from Rous Sarcoma virus When this oncogene was Some of these proteins function as growth factors or growth cloned and transfected into normal cells in culture, the cells factor receptors. Included among these are sis, which encodes underwent malignant transformation. a form of platelet-derived growth factor, and fms, erbB, and neu,which encode growth-factor receptors. In normal cells, the expression of growth factors and their receptors is care- Oncogenes and Cancer Induction fully regulated. Usually, one population of cells secretes a growth factor that acts on another population of cells that In 1971, Howard Temin suggested that oncogenes might not tion of the second population. Inappropriate expression of rries the receptor for the factor, thus stimulating prolifera- normal cells; indeed, he proposed that a virus might acquire either a growth factor or its receptor can result in uncon- oncogenes from the genome of an infected cell. He called trolled proliferation. these cellular genes proto-oncogenes, or cellular oncogenes Other oncogenes in this category encode products tha (c-onc, to distinguish them from their viral counterparts function in signal-transduction pathways or as transcription (v-onc). In the mid-1970s, I.M. Bishop and H. E Varmus factors. The src and abl oncogenes encode tyrosine kinases, identified a DNA sequence in normal chicken cells that is and the ras oncogene encodes a GTP-binding protein.The homologous to v-src from Rous sarcoma virus. This cellular products of these genes act as signal transducers. The myc, oncogene was designated c-src. Since these early discoveri jun, and fos oncogenes encode transcription factors. Overac numerous cellular oncogenes have been identified. tivity of any of these oncogenes may result in unregulated uence comparisons of viral and cellular oncogenes proliferation. reveal that they are highly conserved in evolution. Although most cellular oncogenes consist of a series of exons and in- INHIBITION OF CELLULAR PROLIFERATION trons, their viral counterparts consist of uninterrupted cod- A second category of cancer-associated genes-called tumor ing sequences, suggesting that the virus might have acquired suppressor genes, or anti-oncogenes-encodes proteins that the oncogene through an intermediate RNA transcript from hich the intron sequences had been removed during RNa sults in unregulated proliferation. The prototype of this cate processing. The actual coding sequences of viral oncogenes gory of oncogenes is Rh, the retinoblastoma gene Hereditary and the corresponding proto-oncogenes exhibit a high de- retinoblastoma is a rare childhood cancer, in which tumors gree of homology; in some cases, a single point mutation is develop from neural precursor cells in the immature retina. all that distinguishes a viral oncogene from the correspond- The affected child has inherited a mutated Rb allele; somatic ing proto-oncogene. It has now become apparent that most, inactivation of the remaining Rballele leads to tumor growth. if not all, oncogenes(both viral and cellular)are derived from Probably the single most frequent genetic abnormality in cellular genes that encode various growth-controlling pro- human cancer is mutation in p53, which encodes a nuclear teins. In addition, the proteins encoded by a particular onco- phosphoprotein. Over 90% of small-cell lung cancers and
which integrate randomly into the host chromosomal DNA, include several genes that are expressed early in the course of viral replication. SV40 encodes two early proteins called large T and little T, and polyoma encodes three early proteins called large T, middle T, and little T. Each of these proteins plays a role in the malignant transformation of virus-infected cells. Most RNA viruses replicate in the cytosol and do not induce malignant transformation. The exceptions are retroviruses, which transcribe their RNA into DNA by means of a reverse-transcriptase enzyme and then integrate the transcript into the host’s DNA. This process is similar in the cytopathic retroviruses such as HIV-1 and HIV-2 and in the transforming retroviruses, which induce changes in the host cell that lead to malignant transformation. In some cases, retrovirus-induced transformation is related to the presence of oncogenes, or “cancer genes,” carried by the retrovirus. One of the best-studied transforming retroviruses is the Rous sarcoma virus. This virus carries an oncogene called v-src,which encodes a 60-kDa protein kinase (v-Src) that catalyzes the addition of phosphate to tyrosine residues on proteins. The first evidence that oncogenes alone could induce malignant transformation came from studies of the v-src oncogene from Rous sarcoma virus. When this oncogene was cloned and transfected into normal cells in culture, the cells underwent malignant transformation. Oncogenes and Cancer Induction In 1971, Howard Temin suggested that oncogenes might not be unique to transforming viruses but might also be found in normal cells; indeed, he proposed that a virus might acquire oncogenes from the genome of an infected cell. He called these cellular genes proto-oncogenes, or cellular oncogenes (c-onc), to distinguish them from their viral counterparts (v-onc). In the mid-1970s, J. M. Bishop and H. E. Varmus identified a DNA sequence in normal chicken cells that is homologous to v-src from Rous sarcoma virus. This cellular oncogene was designated c-src. Since these early discoveries, numerous cellular oncogenes have been identified. Sequence comparisons of viral and cellular oncogenes reveal that they are highly conserved in evolution. Although most cellular oncogenes consist of a series of exons and introns, their viral counterparts consist of uninterrupted coding sequences, suggesting that the virus might have acquired the oncogene through an intermediate RNA transcript from which the intron sequences had been removed during RNA processing. The actual coding sequences of viral oncogenes and the corresponding proto-oncogenes exhibit a high degree of homology; in some cases, a single point mutation is all that distinguishes a viral oncogene from the corresponding proto-oncogene. It has now become apparent that most, if not all, oncogenes (both viral and cellular) are derived from cellular genes that encode various growth-controlling proteins. In addition, the proteins encoded by a particular oncogene and its corresponding proto-oncogene appear to have very similar functions. As described below, the conversion of a proto-oncogene into an oncogene appears in many cases to accompany a change in the level of expression of a normal growth-controlling protein. Cancer-Associated Genes Have Many Functions Homeostasis in normal tissue is maintained by a highly regulated process of cellular proliferation balanced by cell death. If there is an imbalance, either at the stage of cellular proliferation or at the stage of cell death, then a cancerous state will develop. Oncogenes and tumor suppressor genes have been shown to play an important role in this process, by regulating either cellular proliferation or cell death. Cancer-associated genes can be divided into three categories that reflect these different activities, summarized in Table 22-1. INDUCTION OF CELLULAR PROLIFERATION One category of proto-oncogenes and their oncogenic counterparts encodes proteins that induce cellular proliferation. Some of these proteins function as growth factors or growthfactor receptors. Included among these are sis, which encodes a form of platelet-derived growth factor, and fms, erbB, and neu, which encode growth-factor receptors. In normal cells, the expression of growth factors and their receptors is carefully regulated. Usually, one population of cells secretes a growth factor that acts on another population of cells that carries the receptor for the factor, thus stimulating proliferation of the second population. Inappropriate expression of either a growth factor or its receptor can result in uncontrolled proliferation. Other oncogenes in this category encode products that function in signal-transduction pathways or as transcription factors. The src and abl oncogenes encode tyrosine kinases, and the ras oncogene encodes a GTP-binding protein. The products of these genes act as signal transducers. The myc, jun, and fos oncogenes encode transcription factors. Overactivity of any of these oncogenes may result in unregulated proliferation. INHIBITION OF CELLULAR PROLIFERATION A second category of cancer-associated genes—called tumorsuppressor genes, or anti-oncogenes—encodes proteins that inhibit excessive cell proliferation. Inactivation of these results in unregulated proliferation. The prototype of this category of oncogenes is Rb, the retinoblastoma gene. Hereditary retinoblastoma is a rare childhood cancer, in which tumors develop from neural precursor cells in the immature retina. The affected child has inherited a mutated Rb allele; somatic inactivation of the remaining Rb allele leads to tumor growth. Probably the single most frequent genetic abnormality in human cancer is mutation in p53, which encodes a nuclear phosphoprotein. Over 90% of small-cell lung cancers and Cancer and the Immune System CHAPTER 22 503
504 art Iv The Immune System in Health and disease TABLE 22-1 Functional classification of cancer-associated genes lype/name Nature af gene product ATEGORY L: GENES THAT INDUCE CELLULAR PROLIFERATION Growth factors A form of platelet-derived growth factor(PDGF) Growth-factor receptors Receptor for colony-stimulating factor 1(CSF-1) Receptor for epidermal growth factor(EGF) Protein(HER2)related to EGF receptor Receptor for thyroid hormone lyrosine kinase Ha-ras GTP-binding protein with GTPase activity N-ras GTP-binding protein with GTPase activity K-ras GTP-binding protein with GTPase activity Transcription factors Component of transcription factor AP fos Component of transcription factor AP ayc DNA-binding protein CATEGORY II: TUMOR SUPRESSOR GENES. INHIBITORS OF CELLULAR PROLIFERATION Rb Suppressor of retinoblastoma Nuclear phosphoprotein that inhibits formation of small-cell lung cander and colon cancers DCC pressor of colon carcinoma pressor of adenomatous polyposis NFl Suppressor of neurofibromatosis WIT Suppressor of Wilms tumor CATEGORY III: GENES THAT REGULATE PROGRAMMED CELL DEATH The activity of the normal products of the category ll genes inhibits progression of the cell cycle. Loss of a gene or its inactivation by mutation in an indicated tumor-suppressor gene is associated with development of the indicated cancers. 50% of breast and colon cancers have been shown to be Proto-Oncogenes Can Be Converted ed with mutations in p5. to Oncogenes REGULATION OF PROGRAMMED CELL DEATH In 1972, R J. Huebner and G I Todaro suggested that muta A third category of cancer-associated genes regulates pro- tions or genetic rearrangements of proto-oncogenes by car- grammed cell death. These genes encode proteins that either cinogens or viruses might alter the normally regulated function block or induce apoptosis. Included in this category of onco- of these genes, converting them into potent cancer-causing genes is bcl-2, an anti-apoptosis gene. This oncogene was oncogenes( Figure 22-2). Considerable evidence supporting originally discovered because of its association with B-cell fol- this hypothesis accumulated in subsequent years. For example licular lymphoma. Since its discovery, bd-2 has been shown to some malignantly transformed cells contain multiple copies of lay an important role in regulating cell survival cellular oncogenes, resulting in increased production of onco hematopoiesis and in the survival of selected B cells and gene products. Such amplification of cellular oncogenes has T cells during maturation. Interestingly, the Epstein-Barr been observed in cells from various types of human cancers. virus contains a gene that has sequence homology to bd-2 Several groups have identified c-myc oncogenes in homo- and may act in a similar manner to suppress apoptosis. geneously staining regions(HSRs)of chromosomes from can
over 50% of breast and colon cancers have been shown to be associated with mutations in p53. REGULATION OF PROGRAMMED CELL DEATH A third category of cancer-associated genes regulates programmed cell death. These genes encode proteins that either block or induce apoptosis. Included in this category of oncogenes is bcl-2, an anti-apoptosis gene. This oncogene was originally discovered because of its association with B-cell follicular lymphoma. Since its discovery, bcl-2 has been shown to play an important role in regulating cell survival during hematopoiesis and in the survival of selected B cells and T cells during maturation. Interestingly, the Epstein-Barr virus contains a gene that has sequence homology to bcl-2 and may act in a similar manner to suppress apoptosis. Proto-Oncogenes Can Be Converted to Oncogenes In 1972, R. J. Huebner and G. J. Todaro suggested that mutations or genetic rearrangements of proto-oncogenes by carcinogens or viruses might alter the normally regulated function of these genes, converting them into potent cancer-causing oncogenes (Figure 22-2). Considerable evidence supporting this hypothesis accumulated in subsequent years. For example, some malignantly transformed cells contain multiple copies of cellular oncogenes, resulting in increased production of oncogene products. Such amplification of cellular oncogenes has been observed in cells from various types of human cancers. Several groups have identified c-myc oncogenes in homogeneously staining regions (HSRs) of chromosomes from can- 504 PART IV The Immune System in Health and Disease TABLE 22-1 Functional classification of cancer-associated genes Type/name Nature of gene product CATEGORY I: GENES THAT INDUCE CELLULAR PROLIFERATION Growth factors sis A form of platelet-derived growth factor (PDGF) Growth-factor receptors fms Receptor for colony-stimulating factor 1 (CSF-1) erbB Receptor for epidermal growth factor (EGF) neu Protein (HER2) related to EGF receptor erbA Receptor for thyroid hormone Signal transducers src Tyrosine kinase abl Tyrosine kinase Ha-ras GTP-binding protein with GTPase activity N-ras GTP-binding protein with GTPase activity K-ras GTP-binding protein with GTPase activity Transcription factors jun Component of transcription factor AP1 fos Component of transcription factor AP1 myc DNA-binding protein CATEGORY II: TUMOR-SUPRESSOR GENES, INHIBITORS OF CELLULAR PROLIFERATION* Rb Suppressor of retinoblastoma p53 Nuclear phosphoprotein that inhibits formation of small-cell lung cander and colon cancers DCC Suppressor of colon carcinoma APC Suppressor of adenomatous polyposis NF1 Suppressor of neurofibromatosis WT1 Suppressor of Wilm’s tumor CATEGORY III: GENES THAT REGULATE PROGRAMMED CELL DEATH bcl-2 Suppressor of apoptosis * The activity of the normal products of the category II genes inhibits progression of the cell cycle. Loss of a gene or its inactivation by mutation in an indicated tumor-suppressor gene is associated with development of the indicated cancers
Cancer and the Immune System CHAPTER 22 505 Normal cells Transformed cells gene. For example, avian leukosis virus(ALv) is a retrovirus that does not carry any viral oncogenes and yet is able to trans form B cells into lymphomas. This particular retrovirus has Retroviral been shown to integrate within the c-myc proto-oncogene, transduction which contains three exons. Exon 1 of c-myc has an unknown function; exons 2 and 3 encode the Myc protein Insertion of avl between exon i and exon 2 has been shown in some cases allow the provirus promoter to increase transcription of Mutagens, viruses. exons 2 and 3, resulting in increased synthesis of c-Myc. Expression radiation, and genetic A variety of tumors have been shown to express signifi- cantly increased levels of growth factors or growth-factor sential growth- Cellular oncogenes Growth-factor Expression (a) Chronic myelogenous leukemia Signal transducers trinuclear factors gelators of programmed ① Qualitatively altered, ell death hyperactive pro chromosome Quantitative alterations (gene amplification or translocation) or decreased levels FIGURE 22-2 Conversion of proto-oncogenes into oncogenes 22 involve mutation, resulting in production of qualitatively different gene products, or DNA amplification or translocation, resulting in increased or decreased expression of gene products. cer cells; these HSRs represent long tandem arrays of amplified 9q+ In addition, some cancer cells exhibit chromosomal trans (b)Burkitt's lymphor locations, usually the movement of a proto-oncogene fror one chromosomal site to another(Figure 22-3). In many cases of Burkitt's lymphoma, for example, c-myc is moved from its normal position on chromosome 8 to a position near the immunoglobulin heavy-chain enhancer on chro mosome 14. As a result of this translocation, synthesis of the c-Myc protein, which functions as a transcription factor Increases Mutation in proto-oncogenes also has been associated ∪cmyr with cellular transformation, and it may be a major mecha- nism by which chemical carcinogens or x-irradiation convert FIGURE 22-3 Chromosomal translocations in(a)chronic myeloge. a proto-oncogene into a cancer-inducing oncogene. For in- nous leukemia(CML) and(b)Burkitts lymphoma. Leukemic cells stance, single-point mutations in c-ras have been detected in from all patients with CML contain the so-called Philadelphia chromo- a significant fraction of several human cancers, including car- some, which results from a translocation between chromosomes 9 cinomas of the bladder, colon, and lung. Some of these muta- and 22. Cancer cells from some patients with Burkitts lymphoma ex- tions appear to reduce the ability of Ras to associate with hibit a translocation that moves part of chromosome 8 to chromo- GTPase-stimulating proteins, thus prolonging the growth- some 14. It is now known that this translocation involves c-myc,a activated state of Ras cellular oncogene. Abnormalities such as these are detected by band. pe Viral integration into the host-cell genome may in itself ing analysis of metaphase chromosomes. Normal chromosomes are rve to convert a proto-oncogene into a transforming onco- shown on the left, and translocated chromosomes on the right
cer cells; these HSRs represent long tandem arrays of amplified genes. In addition, some cancer cells exhibit chromosomal translocations, usually the movement of a proto-oncogene from one chromosomal site to another (Figure 22-3). In many cases of Burkitt’s lymphoma, for example, c-myc is moved from its normal position on chromosome 8 to a position near the immunoglobulin heavy-chain enhancer on chromosome 14. As a result of this translocation, synthesis of the c-Myc protein, which functions as a transcription factor, increases. Mutation in proto-oncogenes also has been associated with cellular transformation, and it may be a major mechanism by which chemical carcinogens or x-irradiation convert a proto-oncogene into a cancer-inducing oncogene. For instance, single-point mutations in c-ras have been detected in a significant fraction of several human cancers, including carcinomas of the bladder, colon, and lung. Some of these mutations appear to reduce the ability of Ras to associate with GTPase-stimulating proteins, thus prolonging the growthactivated state of Ras. Viral integration into the host-cell genome may in itself serve to convert a proto-oncogene into a transforming oncogene. For example, avian leukosis virus (ALV) is a retrovirus that does not carry any viral oncogenes and yet is able to transform B cells into lymphomas. This particular retrovirus has been shown to integrate within the c-myc proto-oncogene, which contains three exons. Exon 1 of c-myc has an unknown function; exons 2 and 3 encode the Myc protein. Insertion of AVL between exon 1 and exon 2 has been shown in some cases to allow the provirus promoter to increase transcription of exons 2 and 3, resulting in increased synthesis of c-Myc. A variety of tumors have been shown to express significantly increased levels of growth factors or growth-factor Cancer and the Immune System CHAPTER 22 505 Normal cells Transformed cells Proto–oncogenes Expression Retroviral transduction Mutagens, viruses, radiation, and genetic predisposition Cellular oncogenes Expression Viral oncogenes Essential growth– controlling proteins Growth factors Growth–factor receptors Signal transducers Intranuclear factors Regulators of programmed cell death 1 Qualitatively altered, hyperactive proteins 2 Quantitative alterations (gene amplification or translocation) resulting in increased or decreased levels of products FIGURE 22-2 Conversion of proto-oncogenes into oncogenes can involve mutation, resulting in production of qualitatively different gene products, or DNA amplification or translocation, resulting in increased or decreased expression of gene products. (a) Chronic myelogenous leukemia 9 22 Philadelphia chromosome (b) Burkitt's lymphoma 8 14 9 q+ 22 q– CH VH CH c–myc c–myc VH 8 q– 14 q+ FIGURE 22-3 Chromosomal translocations in (a) chronic myelogenous leukemia (CML) and (b) Burkitt’s lymphoma. Leukemic cells from all patients with CML contain the so-called Philadelphia chromosome, which results from a translocation between chromosomes 9 and 22. Cancer cells from some patients with Burkitt’s lymphoma exhibit a translocation that moves part of chromosome 8 to chromosome 14. It is now known that this translocation involves c-myc, a cellular oncogene. Abnormalities such as these are detected by banding analysis of metaphase chromosomes. Normal chromosomes are shown on the left, and translocated chromosomes on the right
506 PARt IV The Immune System in Health and Disease receptors. Expression of the receptor for epidermal growth by mating the bd-2 transgenic mice with myc*transgenic factor, which is encoded by c-erbB, has been shown to be mice. These mice develop leukemia very rapidly amplified in many cancer cells And in breast cancer, increased synthesis of the growth-factor receptor encoded by c-neu has Tumors of the Immune System been linked with a poor prognosis. of the immune system are classified as lymphomas The Induction of Cancer Is a Multiste mias. Lymphomas proliferate as solid tumors within Process hoid tissue such as the bone marrow, lymph nodes, or thymus; they include Hodgkins and non-Hodgkins lym- The development from a normal cell to a cancerous cell is phomas Leukemias tend to proliferate as single cells and are ally a multistep process of clonal evolution driven by a detected by increased cell numbers in the blood or lymph. series of somatic mutations that progressively convert the cell Leukemia can develop in lymphoid or myeloid lineages from normal growth to a precancerous state and finally Historically, the leukemias were classified as acute or cancerous state hronic according to the clinical progression of the disease The presence of myriad chromosomal abnormalities in The acute leukemias appeared suddenly and progressed precancerous and cancerous cells lends support to the role of rapidly, whereas the chronic leukemias were much less ag. multiple mutations in the development of cancer. This has gressive and developed slowly as mild, barely symptomatic been demonstrated in human colon cancer, which progresses diseases. These clinical distinctions apply to untreated leuke- in a series of well-defined morphologic stages(Figure 22-4). mias; with current treatments, the acute leukemias often have lon cancer begins as small, benign tumors called adeno- a good prognosis, and permanent remission can often be mas in the colorectal epithelium. These precancerous tumors achieved. Now the major distinction between acute and grow, gradually becoming increasingly disorganized in their chronic leukemias is the maturity of the cell involved. Acute intracellular organization until they acquire the malignant leukemias tend to arise in less mature cells, whereas chronic phenotype. These well-defined morphologic stages of colon leukemias arise in mature cells. The acute leukemias include cancer have been correlated with a sequence of gene changes acute lymphocytic leukemia (ALL) and acute my involving inactivation or loss of three tumor-suppressor genes leukemia(AML); these diseases can develop at (APC, DCC, and p53) and activation of one cellular prolifer- have a rapid onset. The chronic leukemia ation oncogene(K-ras lymphocytic leukemia(CLL) and chronic myelogenous Studies with transgenic mice also support the role of multi- leukemia( CML); these diseases develop slowly and are seen in ple steps in the induction of cancer. Transgenic mice express- adults. ing high levels of Bcl-2 develop a population of small resting A number of B-and T-cell leukemias and lymphomas in B cells, derived from secondary lymphoid follicles, that have volve a proto-oncogene that has been translocated into the greatly extended life spans. Gradually these transgenic mice immunoglobulin genes or T-cell receptor genes. One of the develop lymphomas. Analysis of lymphomas from these mice best characterized is the translocation of c-myc in Burkitt has shown that approximately half have a c-myc translocation lymphoma and in mouse plasmacytomas. In 75% of Burkitt's to the immunoglobulin H-chain locus. The synergism of Myc lymphoma patients, c-myc is translocated from chromosome 8 and Bcl-2 is highlighted in double-transgenic mice produced to the Ig heavy-chain gene cluster on chromosome 14(see Chromosomal 7p eration Loss hypomethylation alterations epithelium epithelium FIGURE 22-4 Model of sequential genetic alterations leading to quence of genetic alterations. Adapted from B. Vogelstein and K. W. metastatic colon cancer. Each of the stages indicated at the bottom is Kinzler, 1993, Trends Genet. 9: 138 morphologically distinct, allowing researchers to determine the se
receptors. Expression of the receptor for epidermal growth factor, which is encoded by c-erbB, has been shown to be amplified in many cancer cells. And in breast cancer, increased synthesis of the growth-factor receptor encoded by c-neu has been linked with a poor prognosis. The Induction of Cancer Is a Multistep Process The development from a normal cell to a cancerous cell is usually a multistep process of clonal evolution driven by a series of somatic mutations that progressively convert the cell from normal growth to a precancerous state and finally a cancerous state. The presence of myriad chromosomal abnormalities in precancerous and cancerous cells lends support to the role of multiple mutations in the development of cancer. This has been demonstrated in human colon cancer, which progresses in a series of well-defined morphologic stages (Figure 22-4). Colon cancer begins as small, benign tumors called adenomas in the colorectal epithelium. These precancerous tumors grow, gradually becoming increasingly disorganized in their intracellular organization until they acquire the malignant phenotype. These well-defined morphologic stages of colon cancer have been correlated with a sequence of gene changes involving inactivation or loss of three tumor-suppressor genes (APC, DCC, and p53) and activation of one cellular proliferation oncogene (K-ras). Studies with transgenic mice also support the role of multiple steps in the induction of cancer. Transgenic mice expressing high levels of Bcl-2 develop a population of small resting B cells, derived from secondary lymphoid follicles, that have greatly extended life spans. Gradually these transgenic mice develop lymphomas. Analysis of lymphomas from these mice has shown that approximately half have a c-myc translocation to the immunoglobulin H-chain locus. The synergism of Myc and Bcl-2 is highlighted in double-transgenic mice produced by mating the bcl-2+ transgenic mice with myc+ transgenic mice. These mice develop leukemia very rapidly. Tumors of the Immune System Tumors of the immune system are classified as lymphomas or leukemias. Lymphomas proliferate as solid tumors within a lymphoid tissue such as the bone marrow, lymph nodes, or thymus; they include Hodgkin’s and non-Hodgkin’s lymphomas. Leukemias tend to proliferate as single cells and are detected by increased cell numbers in the blood or lymph. Leukemia can develop in lymphoid or myeloid lineages. Historically, the leukemias were classified as acute or chronic according to the clinical progression of the disease. The acute leukemias appeared suddenly and progressed rapidly, whereas the chronic leukemias were much less aggressive and developed slowly as mild, barely symptomatic diseases. These clinical distinctions apply to untreated leukemias; with current treatments, the acute leukemias often have a good prognosis, and permanent remission can often be achieved. Now the major distinction between acute and chronic leukemias is the maturity of the cell involved. Acute leukemias tend to arise in less mature cells, whereas chronic leukemias arise in mature cells. The acute leukemias include acute lymphocytic leukemia (ALL) and acute myelogenous leukemia (AML); these diseases can develop at any age and have a rapid onset. The chronic leukemias include chronic lymphocytic leukemia (CLL) and chronic myelogenous leukemia (CML); these diseases develop slowly and are seen in adults. A number of B- and T-cell leukemias and lymphomas involve a proto-oncogene that has been translocated into the immunoglobulin genes or T-cell receptor genes. One of the best characterized is the translocation of c-myc in Burkitt’s lymphoma and in mouse plasmacytomas. In 75% of Burkitt’s lymphoma patients, c-myc is translocated from chromosome 8 to the Ig heavy-chain gene cluster on chromosome 14 (see 506 PART IV The Immune System in Health and Disease Chromosomal site Alteration Gene 5q Loss APC 18q Loss DCC 12p Activation K-ras 17p Loss p53 DNA hypomethylation Other alterations Normal epithelium Hyperproliferative epithelium Early adenoma Intermediate adenoma Late adenoma Carcinoma Metastasis FIGURE 22-4 Model of sequential genetic alterations leading to metastatic colon cancer. Each of the stages indicated at the bottom is morphologically distinct, allowing researchers to determine the sequence of genetic alterations. [Adapted from B. Vogelstein and K. W. Kinzler, 1993, Trends Genet. 9:138.]
Cancer and the Immune System CHAPTER 22 507 l Promoter Switch region 5●◆Lxm0mm JH Enhancer (a) 5吧y如Mmb增m Enhancer C exons C-lmycexons FIGURE 22-5 In many patients with Burkitt's lymphoma, the c-myc exons(2 and 3)of c-myc are inserted at the S, switch site(b).Only gene is translocated to the immunoglobulin heavy -chain gene cluster exons 2 and 3 of c-myc are coding exons. Translocation may lead to on chromosome 14. In some cases, the entire c-mycgene is inserted overexpression of C-Myc. near the heavy-chain enhancer (a), but in other cases, only the coding Figure 22-3b) In the remaining patients, c-myc remains on (TATAs). Tumor-specific antigens are unique to tumor cells chromosome 8 and the k or y light-chain genes are translo- and do not occur on normal cells in the body. They may cated to a region 3'of c-myc. Kappa-gene translocations result from mutations in tumor cells that generate altered from chromosome 2 to chromosome 8 occur 9% of the time, cellular proteins; cytosolic processing of these proteins and y-gene translocations from chromosome 22 to chromo- would give rise to novel peptides that are presented with class some 8 occur 16% of the time I MHC molecules, inducing a cell-mediated response by Translocations of c-myc to the Ig heavy-chain gene cluster tumor-specific CTLs (Figure 22-6). Tumor-associated anti on chromosome 14 have been analyzed, and, in some cases, gens, which are not unique to tumor cells, may be proteins the entire c-myc gene is translocated head-to-head to a re- that are expressed on normal cells during fetal development gion near the heavy-chain enhancer. In other cases, exons 1, when the immune system is immature and unable to respond 2, and 3 or exons 2 and 3 of c-myc are translocated head-to- but that normally are not expressed in the adult Reactivation head to the Su or Sa switch site( Figure 22-5). In each case, of the embryonic genes that encode these proteins in tumor the translocation removes the myc coding exons from the cells results in their expression on the fully differentiated regulatory mechanisms operating in chromosome 8 and tumor cells. Tumor-associated antigens may also be proteins places them in the immunoglobulin-gene region, a very ac- that are normally expressed at extremely low levels on normal tive region that is expressed constitutively in these cells. The cells but are expressed at much higher levels on tumor cells. It consequences of enhancer-mediated high levels of constitu- is now clear that the tumor antigens recognized by human T sion ive been investigated cells fall into one of four major categories in transgenic mice. In one study, mice containing transgene Antigens encoded by genes exclusively expressed by isting of all three c-myc exons and the immunoglobulin tumors heavy-chain enhancer were produced. Of 15 transgenic pups born, 13 developed lymphomas of the B-cell lineage within a Antigens encoded by variant forms of normal genes that few months of birth have been altered by mutation Antigens normally expressed only at certain stages of Tumor Antigens differentiation or only by certain differentiation lineages a Antigens that are overexpressed in particular tumors The subdiscipline of tumor immunology involves the stud of antigens on tumor cells and the immune response to these Many tumor antigens are cellular proteins that give rise to antigens. Two types of tumor antigens have been identified peptides presented with MHC molecules; typically, these an- on tumor cells: tumor-specific transplantation antigens tigens have been identified by their ability to induce the pro- (TSTAs) and tumor-associated transplantation antigens liferation of antigen-specific CTLs or helper T cells
Figure 22-3b). In the remaining patients, c-myc remains on chromosome 8 and the or light-chain genes are translocated to a region 3 of c-myc. Kappa-gene translocations from chromosome 2 to chromosome 8 occur 9% of the time, and -gene translocations from chromosome 22 to chromosome 8 occur 16% of the time. Translocations of c-myc to the Ig heavy-chain gene cluster on chromosome 14 have been analyzed, and, in some cases, the entire c-myc gene is translocated head-to-head to a region near the heavy-chain enhancer. In other cases, exons 1, 2, and 3 or exons 2 and 3 of c-myc are translocated head-tohead to the S or S switch site (Figure 22-5). In each case, the translocation removes the myc coding exons from the regulatory mechanisms operating in chromosome 8 and places them in the immunoglobulin-gene region, a very active region that is expressed constitutively in these cells. The consequences of enhancer-mediated high levels of constitutive myc expression in lymphoid cells have been investigated in transgenic mice. In one study, mice containing a transgene consisting of all three c-myc exons and the immunoglobulin heavy-chain enhancer were produced. Of 15 transgenic pups born, 13 developed lymphomas of the B-cell lineage within a few months of birth. Tumor Antigens The subdiscipline of tumor immunology involves the study of antigens on tumor cells and the immune response to these antigens. Two types of tumor antigens have been identified on tumor cells: tumor-specific transplantation antigens (TSTAs) and tumor-associated transplantation antigens (TATAs). Tumor-specific antigens are unique to tumor cells and do not occur on normal cells in the body. They may result from mutations in tumor cells that generate altered cellular proteins; cytosolic processing of these proteins would give rise to novel peptides that are presented with class I MHC molecules, inducing a cell-mediated response by tumor-specific CTLs (Figure 22-6). Tumor-associated antigens, which are not unique to tumor cells, may be proteins that are expressed on normal cells during fetal development when the immune system is immature and unable to respond but that normally are not expressed in the adult. Reactivation of the embryonic genes that encode these proteins in tumor cells results in their expression on the fully differentiated tumor cells. Tumor-associated antigens may also be proteins that are normally expressed at extremely low levels on normal cells but are expressed at much higher levels on tumor cells. It is now clear that the tumor antigens recognized by human T cells fall into one of four major categories: ■ Antigens encoded by genes exclusively expressed by tumors ■ Antigens encoded by variant forms of normal genes that have been altered by mutation ■ Antigens normally expressed only at certain stages of differentiation or only by certain differentiation lineages ■ Antigens that are overexpressed in particular tumors Many tumor antigens are cellular proteins that give rise to peptides presented with MHC molecules; typically, these antigens have been identified by their ability to induce the proliferation of antigen-specific CTLs or helper T cells. Cancer and the Immune System CHAPTER 22 507 5′ JH 3′ Cµ exons Enhancer D Switch region V JH H Promoter Sµ 5′ 3′ Cµ exons Enhancer Sµ 3 2 1 c–myc exons (a) Rearranged Ig heavy–chain gene on chromosome 14 Translocated c–myc gene in some Burkitt's lymphomas 5′ 3′ Cµ exons S 3 2 µ c–myc exons (b) Translocated c–myc gene in other Burkitt's lymphomas L FIGURE 22-5 In many patients with Burkitt’s lymphoma, the c-myc gene is translocated to the immunoglobulin heavy-chain gene cluster on chromosome 14. In some cases, the entire c-myc gene is inserted near the heavy-chain enhancer (a), but in other cases, only the coding exons (2 and 3) of c-myc are inserted at the S switch site (b). Only exons 2 and 3 of c-myc are coding exons. Translocation may lead to overexpression of c-Myc.
508 PARt IV The Immune System in Health and Disease Normal cell Self-peptide Self-peptic Class I mhc Class I MHC Altered se Mutation generates new peptide in class I MHC molecule(TSTa) embryonic gene(TATA) ormal protein (TATA FIGURE 22-6 Different mechanisms generate tumor-specific transplantation antigens(TSTAs) and tumor-associated transplantation antigens(TATAs). The latter are more common. Some Antigens Are Tumor-Specific (tum"), which gives rise to progressively growing tumors, is Tumor-specific antigens have been identified on tumors in- treated in vitro with a chemical cells duced with chemical or physical carcinogens and on some they no longer virally induced tumors. Demonstrating the presence of tumor specific antigens on spontaneously occurring tumors is pal ticularly difficult because the immune response to such tu Immune response to mors eliminates all of the tumor cells bearing sufficient TABLE 22-2 methyl-cholanthrene(MCA) numbers of the antigens and in this way selects for cells bear or polyoma virus(P ing low levels of the antigens. transplanted Live tumor cells Tumor CHEMICALLY OR PHYSICALLY INDUCED lled tumor cells for challenge growth TUMOR ANTIGENS CHEMICALLY INDUCED methylcholanthrene and ultraviolet light are two carcinogens that have been used extensively to generate lines of tumor cells. MCA-induced sarcoma A MCA-induced sarcoma When syngeneic animals are injected with killed cells from a MCA-induced sarcoma A MCA-induced sarcoma carcinogen-induced tumor-cell line, the animals develop a specific immunologic response that can protect against later VIRALLY INDUCED challenge by live cells of the same line but not other tumor-cel lines(Table 22-2). Even when the same chemical carcinogen PV-induced sarcoma A PV-induced sarcoma A duces two separate tumors at different sites in the same ani- PV-induced sarcoma A PV-induced sarcoma B the tumor antigens are distinct and the immune resp PV-induced sarcoma a SV40-induced sarcoma+ to one tumor does not protect against the other tumor. The tumor-specific transplantation antigens of chemically"Tumors were induced either with MCA or PV, and kiled cells from the induced induced tumors have been difficult to characterize because tumors were injected into syngeneic animal, which were then challenged with they cannot be identified by induced antibodies but only by lve cell from the indicated tumor -cell ines. The absence of tumor growth after their T-cell-mediated rejection. One experimental approach Iive challenge indicates tha nmune response induced by tumor antigens that has allowed identification of genes encoding some TSTAs on the killed olls provided protection against the live cells. is outlined in Figure 22-7. When a mouse tumorigenic cell line
Some Antigens Are Tumor-Specific Tumor-specific antigens have been identified on tumors induced with chemical or physical carcinogens and on some virally induced tumors. Demonstrating the presence of tumorspecific antigens on spontaneously occurring tumors is particularly difficult because the immune response to such tumors eliminates all of the tumor cells bearing sufficient numbers of the antigens and in this way selects for cells bearing low levels of the antigens. CHEMICALLY OR PHYSICALLY INDUCED TUMOR ANTIGENS Methylcholanthrene and ultraviolet light are two carcinogens that have been used extensively to generate lines of tumor cells. When syngeneic animals are injected with killed cells from a carcinogen-induced tumor-cell line, the animals develop a specific immunologic response that can protect against later challenge by live cells of the same line but not other tumor-cell lines (Table 22-2). Even when the same chemical carcinogen induces two separate tumors at different sites in the same animal, the tumor antigens are distinct and the immune response to one tumor does not protect against the other tumor. The tumor-specific transplantation antigens of chemically induced tumors have been difficult to characterize because they cannot be identified by induced antibodies but only by their T-cell–mediated rejection. One experimental approach that has allowed identification of genes encoding some TSTAs is outlined in Figure 22-7. When a mouse tumorigenic cell line (tum+ ), which gives rise to progressively growing tumors, is treated in vitro with a chemical mutagen, some cells are mutated so that they no longer are capable of growing into a 508 PART IV The Immune System in Health and Disease Altered self-peptide Mutation generates new peptide in class I MHC molecule (TSTA) Oncofetal peptide Normal cell Inappropriate expression of embryonic gene (TATA) Self-peptide Self-peptide Class I MHC Class I MHC Overexpression of normal protein (TATA) FIGURE 22-6 Different mechanisms generate tumor-specific transplantation antigens (TSTAs) and tumor-associated transplantation antigens (TATAs). The latter are more common. TABLE 22-2 Immune response to methyl-cholanthrene (MCA) or polyoma virus (PV)* Transplanted Live tumor cells Tumor killed tumor cells for challenge growth CHEMICALLY INDUCED MCA-induced sarcoma A MCA-induced sarcoma A – MCA-induced sarcoma A MCA-induced sarcoma B + VIRALLY INDUCED PV-induced sarcoma A PV-induced sarcoma A – PV-induced sarcoma A PV-induced sarcoma B – PV-induced sarcoma A SV40-induced sarcoma C + *Tumors were induced either with MCA or PV, and killed cells from the induced tumors were injected into syngeneic animals, which were then challenged with live cells from the indicated tumor-cell lines. The absence of tumor growth after live challenge indicates that the immune response induced by tumor antigens on the killed cells provided protection against the live cells
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 designated 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 recognized 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 expression 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 tumorspecific transplantation antigens (TSTAs). Most TSTAs can be detected only by the cell-mediated rejection they elicit. In the first part of this procedure, a nontumorigenic (tum– ) cell line is generated; this cell line expresses 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
510 PART I The Immune System in Health and Disease In the past few years, two methods have facilitated the char- by the same virus. For example, when syngeneic mice are acterization of TSTAs(Figure 22-8). In one method, peptides injected with killed cells from a particular polyoma-induced bound to class I MHC molecules on the membranes of the tumor, the recipients are protected against subsequent chal- tumor cells are eluted with acid and purified by high-pressure lenge with live cells from any polyoma-induced tumors(see liquid chromatography(HPLC). In some cases, sufficient pep- Table 22-2). Likewise, when lymphocytes are transferred from tide is eluted to allow its sequence to be deduced by Edman mice with a virus-induced tumor into normal syngeneic degradation. In a second approach, cDNA libraries are pre- recipients, the recipients reject subsequent transplants of all pared from tumor cells. These cDNA libraries are transfected syngeneic tumors induced by the same virus. In the case of transiently into COS cells, which are monkey kidney cells both SV40-and polyoma-induced tumors, the presence of transfected with the gene that codes for the Sv40 large-T anti- tumor antigens is related to the neoplastic state of the cell. In gen. When these cells are later transfected with plasmids con- humans, Burkitt's-lymphoma cells have been shown to express taining both the tumor-cell cDNA and an SV40 origin of repli- a nuclear antigen of the Epstein-Barr virus that may indeed be cation, the large-T antigen stimulates plasmid replication, so a tumor-specific antigen for this type of tumor. Human papil that up to 104-105 plasmid copies are produced per cell. This loma virus(HPV) E6 and E7 proteins are found in more than results in high-level expression of the tumor-cell DNA. 80%of invasive cervical cancers-the clearest example of a The genes that encode some TSTAs have been shown to virally encoded tumor antigen. Consequently, there is great differ from normal cellular genes by a single point mutation. interest in testing as vaccine candidates the HPVs that Further characterization of TSTAs has demonstrated that strongly linked to cervical cancer, such as HPV-16 many of them are not cell-membrane proteins; rather, as indi- The potential value of these virally induced tumor antigens cated already, they are short peptides derived from cytosolic can be seen in animal models. In one experiment, mice immu- proteins that have been processed and presented together with nized with a preparation of genetically engineered polyoma class i mhc molecules virus tumor antigen were shown to be immune to subsequent injections of live polyoma-induced tumor cells. In another Tumor Antigens May Be Induced by Viruses experiment, mice were immunized with a vaccinia-virus vac- cine engineered with the gene encoding the polyoma-tumol In contrast to chemically induced tumors, virally induced antigen. These mice also developed immunity, rejecting later tumors express tumor antigens shared by all tumors induced injections of live polyoma-induced tumor cells( Figure 22-9) Elute HPLC peptides peptide O Plasmids replicate Tumor-cell cdnA libl of cDNA CDNA SV40 large-T FIGURE 22-8 Two methods used to isolate tumor antigens that induce tumor-specific CTLs. See text for details
In the past few years, two methods have facilitated the characterization of TSTAs (Figure 22-8). In one method, peptides bound to class I MHC molecules on the membranes of the tumor cells are eluted with acid and purified by high-pressure liquid chromatography (HPLC). In some cases, sufficient peptide is eluted to allow its sequence to be deduced by Edman degradation. In a second approach, cDNA libraries are prepared from tumor cells. These cDNA libraries are transfected transiently into COS cells, which are monkey kidney cells transfected with the gene that codes for the SV40 large-T antigen. When these cells are later transfected with plasmids containing both the tumor-cell cDNA and an SV40 origin of replication, the large-T antigen stimulates plasmid replication, so that up to 104–105 plasmid copies are produced per cell. This results in high-level expression of the tumor-cell DNA. The genes that encode some TSTAs have been shown to differ from normal cellular genes by a single point mutation. Further characterization of TSTAs has demonstrated that many of them are not cell-membrane proteins; rather, as indicated already, they are short peptides derived from cytosolic proteins that have been processed and presented together with class I MHC molecules. Tumor Antigens May Be Induced by Viruses In contrast to chemically induced tumors, virally induced tumors express tumor antigens shared by all tumors induced by the same virus. For example, when syngeneic mice are injected with killed cells from a particular polyoma-induced tumor, the recipients are protected against subsequent challenge with live cells from any polyoma-induced tumors (see Table 22-2). Likewise, when lymphocytes are transferred from mice with a virus-induced tumor into normal syngeneic recipients, the recipients reject subsequent transplants of all syngeneic tumors induced by the same virus. In the case of both SV40- and polyoma-induced tumors, the presence of tumor antigens is related to the neoplastic state of the cell. In humans, Burkitt’s-lymphoma cells have been shown to express a nuclear antigen of the Epstein-Barr virus that may indeed be a tumor-specific antigen for this type of tumor. Human papilloma virus (HPV) E6 and E7 proteins are found in more than 80% of invasive cervical cancers—the clearest example of a virally encoded tumor antigen. Consequently, there is great interest in testing as vaccine candidates the HPVs that are strongly linked to cervical cancer, such as HPV-16. The potential value of these virally induced tumor antigens can be seen in animal models. In one experiment, mice immunized with a preparation of genetically engineered polyoma virus tumor antigen were shown to be immune to subsequent injections of live polyoma-induced tumor cells. In another experiment, mice were immunized with a vaccinia-virus vaccine engineered with the gene encoding the polyoma-tumor antigen. These mice also developed immunity, rejecting later injections of live polyoma-induced tumor cells (Figure 22-9). 510 PART IV The Immune System in Health and Disease Monkey kidney cos cells (a) Acid Elute peptide Purify peptides HPLC Melanoma tumor cells Tumor-cell cDNA library Tumor cell cDNA SV40 origin of replication Plasmid DNA (b) Sequence peptides SV40 large-T antigen Plasmids replicate (104–105 copies of cDNA) FIGURE 22-8 Two methods used to isolate tumor antigens that induce tumor-specific CTLs. See text for details
