Chapter 25: Cancer Genetics

Cancer arises through a series of somatic alterations in DNA that result in unrestrained cellular proliferation. Most of these alterations involve actual sequence changes in DNA (i.e., mutations). They may originate as a consequence of random replication errors, exposure to carcinogens (e.g., radiation), or faulty DNA repair processes. While most cancers arise sporadically, familial clustering of cancers occurs in certain families that carry a germline mutation in a cancer gene.

The idea that cancer progression is driven by sequential somatic mutations in specific genes has only gained general acceptance in the past 25 years. Before the advent of the microscope, cancer was believed to be composed of aggregates of mucus or other noncellular matter. By the middle of the nineteenth century, it became clear that tumors were masses of cells and that these cells arose from the normal cells of the tissue from which the cancer originated. However, the molecular basis for the uncontrolled proliferation of cancer cells was to remain a mystery for another century. During that time, a number of theories for the origin of cancer were postulated. The great biochemist Otto Warburg proposed the combustion theory of cancer, which stipulated that cancer was due to abnormal oxygen metabolism. In addition, some believed that all cancers were caused by viruses, and that cancer was in fact a contagious disease.

In the end, observations of cancer occurring in chimney sweeps, studies of x-rays, and the overwhelming data demonstrating cigarette smoke as a causative agent in lung cancer, together with Ames’s work on chemical mutagenesis, provided convincing evidence that cancer originated through changes in DNA. Although the viral theory of cancer did not prove to be generally accurate (with the exception of human papillomaviruses, which can cause cervical and other cancers in human), the study of retroviruses led to the discovery of the first human oncogenes in the late 1970s. Soon after, the study of families with genetic predisposition to cancer was instrumental in the discovery of tumor-suppressor genes. The field that studies the type of mutations, as well as the consequence of these mutations in tumor cells, is now known as cancer genetics.

Nearly all cancers originate from a single cell; this clonal origin is a critical discriminating feature between neoplasia and hyperplasia. Multiple cumulative mutational events are invariably required for the progression of a tumor from normal to fully malignant phenotype. The process can be seen as Darwinian microevolution in which, at each successive step, the mutated cells gain a growth advantage resulting in an increased representation relative to their neighbors (Fig. 25-1). Based on observations of cancer frequency increases during aging, as well as molecular genetics work, it is believed that 5 to 10 accumulated mutations are necessary for a cell to progress from the normal to the fully malignant phenotype.

We are beginning to understand the precise nature of the genetic alterations responsible for some malignancies and to get a sense of the order in which they occur. The best-studied example is colon cancer, in which analyses of DNA from tissues extending from normal colon epithelium through adenoma to carcinoma have identified some of the genes mutated in the process (Fig. 25-2). Other malignancies are believed to progress in a similar stepwise fashion, although the order and identity of genes affected may be different.

There are two major types of cancer genes. The first type comprises genes that positively influence tumor formation and are known as oncogenes. The second type of cancer genes negatively impact tumor growth and have been named tumor-suppressor genes. Both oncogenes and tumor-suppressor genes exert their effects on tumor growth through their ability to control cell division (cell birth) or cell death (apoptosis), although the mechanisms can be extremely complex. While tightly regulated in normal cells, oncogenes acquire mutations in cancer cells, and the mutations typically relieve this control and lead to increased activity of the gene products. This mutational event typically occurs in a single allele of the oncogene and acts in a dominant fashion. In contrast, the normal function of tumor-suppressor genes is usually to restrain cell growth, and this function is lost in cancer. Because of the diploid nature of mammalian cells, both alleles must be inactivated for a cell to completely lose the function of a tumor-suppressor gene, leading to a recessive mechanism at the cellular level. From these ideas and studies on the inherited form of retinoblastoma, Knudson and others formulated the two-hit hypothesis, which in its modern version states that both copies of a tumor-suppressor gene must be inactivated in cancer.

There is a subset of tumor-suppressor genes, the caretaker genes, that do not affect cell growth directly, but rather control the ability of the cell to maintain the integrity of its genome. Cells with a deficiency in these genes have an increased rate of mutations throughout their genomes, including in oncogenes and tumor-suppressor genes. This “mutator” phenotype was first hypothesized by Loeb to explain how the multiple mutational events required for tumorigenesis can occur in the lifetime of an individual. A mutator phenotype has now been observed in some forms of cancer, such as those associated with deficiencies in DNA mismatch repair. The great majority of cancers, however, do not harbor repair deficiencies, and their rate of mutation is similar to that observed in normal cells. Many of these cancers, however, appear to harbor a different kind of genetic instability, affecting the loss or gains of whole chromosomes or large parts thereof (as explained in more detail below).

Work by Peyton Rous in the early 1900s revealed that a chicken sarcoma could be transmitted from animal to animal in cell-free extracts, suggesting that cancer could be induced by an agent acting positively to promote tumor formation. The agent responsible for the transmission of the cancer was a retrovirus (Rous sarcoma virus, RSV) and the oncogene responsible was identified 75 years later as v-src. Other oncogenes were also discovered through their presence in the genomes of retroviruses that are capable of causing cancers in chickens, mice, and rats. The cellular homologues of these viral genes are called protooncogenes and are often targets of mutation or aberrant regulation in human cancer. Whereas many oncogenes were discovered because of their presence in retroviruses, other oncogenes, particularly those involved in translocations characteristic of particular leukemias and lymphomas, were isolated through genomic approaches. Investigators cloned the sequences surrounding the chromosomal translocations observed cytogenetically and then deduced the nature of the genes that were the targets of these translocations (see below). Some of these were oncogenes known from retroviruses (like ABL, involved in chronic myeloid leukemia [CML]), whereas others were new (like BCL2, involved in B cell lymphoma). In the normal cellular environment, protooncogenes have crucial roles in cell proliferation and differentiation. Table 25-1 is a partial list of oncogenes known to be involved in human cancer.

The normal growth and differentiation of cells is controlled by growth factors that bind to receptors on the surface of the cell. The signals generated by the membrane receptors are transmitted inside the cells through signaling cascades involving kinases, G proteins, and other regulatory proteins. Ultimately, these signals affect the activity of transcription factors in the nucleus, which regulate the expression of genes crucial in cell proliferation, cell differentiation, and cell death. Oncogene products have been found to function at critical steps in these pathways (Chap. 26), and inappropriate activation of these pathways can lead to tumorigenesis.

POINT MUTATION

Point mutation is a common mechanism of oncogene activation. For example, mutations in one of the RAS genes (HRAS, KRAS, or NRAS) are present in up to 85% of pancreatic cancers and 45% of colon cancers but are less common in other cancer types, although they can occur at significant frequencies in leukemia, lung, and thyroid cancers. Remarkably—and in contrast to the diversity of mutations found in tumor-suppressor genes (see below)—most of the activated RAS genes contain point mutations in codons 12, 13, or 61 (these mutations reduce RAS GTPase activity, leading to constitutive activation of the mutant RAS protein). The restricted pattern of mutations observed in oncogenes compared to that of tumor-suppressor genes reflects the fact that gain-of-function mutations are less likely to occur than mutations that simply lead to loss of activity. Indeed, inactivation of a gene can in theory be accomplished through the introduction of a stop codon anywhere in the coding sequence, whereas activations require precise substitutions at residues that can somehow lead to an increase in the activity of the encoded protein. Importantly, the specificity of oncogene mutations provides diagnostic opportunities, as tests that identify mutations at defined positions are easier to design than tests aimed at detecting random changes in a gene.

DNA AMPLIFICATION

The second mechanism for activation of oncogenes is DNA sequence amplification, leading to overexpression of the gene product. This increase in DNA copy number may cause cytologically recognizable chromosome alterations referred to as homogeneous staining regions (HSRs) if integrated within chromosomes, or double minutes (dmins) if extrachromosomal. The recognition of DNA amplification is accomplished through various cytogenetic techniques such as comparative genomic hybridization (CGH) or fluorescence in situ hybridization (FISH), which allow the visualization of chromosomal aberrations using fluorescent dyes. In addition, noncytogenetic, microarray-based approaches are now available for identifying changes in copy number at high resolution. Newer short-tag–based sequencing approaches have been used to evaluate amplifications. When paired with next-generation sequencing, this approach offers the highest degree of resolution and quantification available. With both microarray and sequencing technologies, the entire genome can be surveyed for gains and losses of DNA sequences, thus pinpointing chromosomal regions likely to contain genes important in the development or progression of cancer.

Numerous genes have been reported to be amplified in cancer. Several of these genes, including NMYC and LMYC, were identified through their presence within the amplified DNA sequences of a tumor and had homology to known oncogenes. Because the region amplified often includes hundreds of thousands of base pairs, multiple oncogenes may be amplified in a single amplicon in some cancers (particularly in sarcomas). Indeed, MDM2, GLI, CDK4, and SAS at chromosomal location 12q13-15 have been shown to be simultaneously amplified in several types of sarcomas and other tumors. Amplification of a cellular gene is often a predictor of poor prognosis; for example, ERBB2/HER2 and NMYC are often amplified in aggressive breast cancers and neuroblastoma, respectively.

CHROMOSOMAL REARRANGEMENT

Chromosomal alterations provide important clues to the genetic changes in cancer. The chromosomal alterations in human solid tumors such as carcinomas are heterogeneous and complex and occur as a result of the frequent chromosomal instability (CIN) observed in these tumors (see below). In contrast, the chromosome alterations in myeloid and lymphoid tumors are often simple translocations, i.e., reciprocal transfers of chromosome arms from one chromosome to another. Consequently, many detailed and informative chromosome analyses have been performed on hematopoietic cancers. The breakpoints of recurring chromosome abnormalities usually occur at the site of cellular oncogenes. Table 25-2 lists representative examples of recurring chromosome alterations in malignancy and the associated gene(s) rearranged or deregulated by the chromosomal rearrangement. Translocations are particularly common in lymphoid tumors, probably because these cell types have the capability to rearrange their DNA to generate antigen receptors. Indeed, antigen receptor genes are commonly involved in the translocations, implying that an imperfect regulation of receptor gene rearrangement may be involved in the pathogenesis. An interesting example is Burkitt’s lymphoma, a B cell tumor characterized by a reciprocal translocation between chromosomes 8 and 14. Molecular analysis of Burkitt’s lymphomas demonstrated that the breakpoints occurred within or near the MYC locus on chromosome 8 and within the immunoglobulin heavy chain locus on chromosome 14, resulting in the transcriptional activation of MYC. Enhancer activation by translocation, although not universal, appears to play an important role in malignant progression. In addition to transcription factors and signal transduction molecules, translocation may result in the overexpression of cell cycle regulatory proteins or proteins such as cyclins and of proteins that regulate cell death.

The first reproducible chromosome abnormality detected in human malignancy was the Philadelphia chromosome detected in CML. This cytogenetic abnormality is generated by reciprocal translocation involving the ABL oncogene on chromosome 9, encoding a tyrosine kinase, being placed in proximity to the BCR (breakpoint cluster region) gene on chromosome 22. Figure 25-3 illustrates the generation of the translocation and its protein product. The consequence of expression of the BCR-ABL gene product is the activation of signal transduction pathways leading to cell growth independent of normal external signals. Imatinib (marketed as Gleevec), a drug that specifically blocks the activity of Abl tyrosine kinase, has shown remarkable efficacy with little toxicity in patients with CML. It is hoped that knowledge of genetic alterations in other cancers will likewise lead to mechanism-based design and development of a new generation of chemotherapeutic agents.

Solid tumors are generally highly aneuploid, containing an abnormal number of chromosomes; these chromosomes also exhibit structural alterations such as translocations, deletions, and amplifications. These abnormalities are collectively referred to as chromosomal instability (CIN). Normal cells possess several cell cycle checkpoints, essentially quality-control requirements that have to be met before subsequent events are allowed to take place. The mitotic checkpoint, which ensures proper chromosome attachment to the mitotic spindle before allowing the sister chromatids to separate, is altered in certain cancers. The molecular basis of CIN remains unclear, although a number of mitotic checkpoint genes are found mutated or abnormally expressed in various tumors. The exact effects of these changes on the mitotic checkpoint are unknown, and both weakening and overactivation of the checkpoint have been proposed. The identification of the cause of CIN in tumors will likely be a formidable task, considering that several hundred genes are thought to control the mitotic checkpoint and other cellular processes ensuring proper chromosome segregation. Regardless of the mechanisms underlying CIN, the measurement of the number of chromosomal alterations present in tumors is now possible with both cytogenetic and molecular techniques, and several studies have shown that this information can be useful for prognostic purposes. In addition, because the mitotic checkpoint is essential for cellular viability, it may become a target for novel therapeutic approaches.

The first indication of the existence of tumor-suppressor genes came from experiments showing that fusion of mouse cancer cells with normal mouse fibroblasts led to a nonmalignant phenotype in the fused cells. The normal role of tumor-suppressor genes is to restrain cell growth, and the function of these genes is inactivated in cancer. The two major types of somatic lesions observed in tumor-suppressor genes during tumor development are point mutations and large deletions. Point mutations in the coding region of tumor-suppressor genes will frequently lead to truncated protein products or otherwise nonfunctional proteins. Similarly, deletions lead to the loss of a functional product and sometimes encompass the entire gene or even the entire chromosome arm, leading to loss of heterozygosity (LOH) in the tumor DNA compared to the corresponding normal tissue DNA (Fig. 25-4). LOH in tumor DNA is considered a hallmark for the presence of a tumor-suppressor gene at a particular chromosomal location, and LOH studies have been useful in the positional cloning of many tumor-suppressor genes.

Gene silencing, an epigenetic change that leads to the loss of gene expression and occurs in conjunction with hypermethylation of the promoter and histone deacetylation, is another mechanism of tumor-suppressor gene inactivation. (An epigenetic modification refers to a change in the genome, heritable by cell progeny, that does not involve a change in the DNA sequence. The inactivation of the second X chromosome in female cells is an example of an epigenetic silencing that prevents gene expression from the inactivated chromosome.) During embryologic development, regions of chromosomes from one parent are silenced and gene expression proceeds from the chromosome of the other parent. For most genes, expression occurs from both alleles or randomly from one allele or the other. The preferential expression of a particular gene exclusively from the allele contributed by one parent is called parental imprinting and is thought to be regulated by covalent modifications of chromatin protein and DNA (often methylation) of the silenced allele.

The role of epigenetic control mechanisms in the development of human cancer is unclear. However, a general decrease in the level of DNA methylation has been noted as a common change in cancer. In addition, numerous genes, including some tumor-suppressor genes, appear to become hypermethylated and silenced during tumorigenesis. VHL and p16INK4 are well-studied examples of such tumor-suppressor genes. Overall, epigenetic mechanisms may be responsible for reprogramming the expression of a large number of genes in cancer and, together with the mutation of specific genes, are likely to be crucial in the development of human malignancies. The use of drugs that can reverse epigenetic changes in cancer cells may represent a novel therapeutic option in certain cancers or premalignant conditions. For example, demethylating agents (azacitidine or decitabine) are now approved by the U.S. Food and Drug Administration (FDA) for the treatment of patients with high-risk myelodysplastic syndrome (MDS).

A small fraction of cancers occur in patients with a genetic predisposition. In these families, the affected individuals have a predisposing loss-of-function mutation in one allele of a tumor-suppressor gene. The tumors in these patients show a loss of the remaining normal allele as a result of somatic events (point mutations or deletions), in agreement with the two-hit hypothesis (Fig. 25-4). Thus, most cells of an individual with an inherited loss-of-function mutation in a tumor-suppressor gene are functionally normal, and only the rare cells that develop a mutation in the remaining normal allele will exhibit uncontrolled regulation.

Roughly 100 syndromes of familial cancer have been reported, although many are rare. The majority are inherited as autosomal dominant traits, although some of those associated with DNA repair abnormalities (xeroderma pigmentosum, Fanconi’s anemia, ataxia telangiectasia) are autosomal recessive. Table 25-3 shows a number of cancer predisposition syndromes and the responsible genes. The current paradigm states that the genes mutated in familial syndromes can also be targets for somatic mutations in sporadic (noninherited) tumors. The study of cancer syndromes has thus provided invaluable insights into the mechanisms of progression for many tumor types. This section examines the case of inherited colon cancer in detail, but similar lessons can be applied to many of the cancer syndromes listed in Table 25-3. In particular, the study of inherited colon cancer will clearly illustrate the difference between two types of tumor-suppressor genes: the gatekeepers, which directly regulate the growth of tumors, and the caretakers, which, when mutated, lead to genetic instability and therefore act indirectly on tumor growth.

Familial adenomatous polyposis (FAP) is a dominantly inherited colon cancer syndrome due to germline mutations in the adenomatous polyposis coli (APC) tumor-suppressor gene on chromosome 5. Patients with this syndrome develop hundreds to thousands of adenomas in the colon. Each of these adenomas has lost the normal remaining allele of APC but has not yet accumulated the required additional mutations to generate fully malignant cells (Fig. 25-2). The loss of the second functional APC allele in tumors from FAP families often occurs through loss of heterozygosity. However, out of these thousands of benign adenomas, several will invariably acquire further abnormalities and a subset will even develop into fully malignant cancers. APC is thus considered to be a gatekeeper for colon tumorigenesis: in the absence of mutation of this gatekeeper (or a gene acting within the same pathway), a colorectal tumor simply cannot form. Figure 25-5 shows germline and somatic mutations found in the APC gene. The function of the APC protein is still not completely understood, but it likely provides differentiation and apoptotic cues to colonic cells as they migrate up the crypts. Defects in this process may lead to abnormal accumulation of cells that should normally undergo apoptosis.

In contrast to patients with FAP, patients with hereditary nonpolyposis colon cancer (HNPCC, or Lynch’s syndrome) do not develop multiple polyposis, but instead develop only one or a small number of adenomas that rapidly progress to cancer. Most HNPCC cases are due to mutations in one of four DNA mismatch repair genes (Table 25-3), which are components of a repair system that is normally responsible for correcting errors in freshly replicated DNA. Germline mutations in MSH2 and MLH1 account for more than 90% of HNPCC cases, whereas mutations in MSH6 and PMS2 are much less frequent. When a somatic mutation inactivates the remaining wild-type allele of a mismatch repair gene, the cell develops a hypermutable phenotype characterized by profound genomic instability, especially for the short repeated sequences called microsatellites. This microsatellite instability (MSI) favors the development of cancer by increasing the rate of mutations in many genes, including oncogenes and tumor-suppressor genes (Fig. 25-2). These genes can thus be considered caretakers. Interestingly, CIN can also be found in colon cancer, but MSI and CIN appear to be mutually exclusive, suggesting that they represent alternative mechanisms for the generation of a mutator phenotype in this cancer (Fig. 25-2). Other cancer types rarely exhibit MSI, but most exhibit CIN.

Although most autosomal dominant inherited cancer syndromes are due to mutations in tumor-suppressor genes (Table 25-3), there are a few interesting exceptions. Multiple endocrine neoplasia type 2, a dominant disorder characterized by pituitary adenomas, medullary carcinoma of the thyroid, and (in some pedigrees) pheochromocytoma, is due to gain-of-function mutations in the protooncogene RET on chromosome 10. Similarly, gain-of-function mutations in the tyrosine kinase domain of the MET oncogene lead to hereditary papillary renal carcinoma. Interestingly, loss-of-function mutations in the RET gene cause a completely different disease, Hirschsprung’s disease (aganglionic megacolon [Chap. 52]).

Although the Mendelian forms of cancer have taught us much about the mechanisms of growth control, most forms of cancer do not follow simple patterns of inheritance. In many instances (e.g., lung cancer), a strong environmental contribution is at work. Even in such circumstances, however, some individuals may be more genetically susceptible to developing cancer, given the appropriate exposure, due to the presence of modifier alleles.

The discovery of cancer susceptibility genes raises the possibility of DNA testing to predict the risk of cancer in individuals of affected families. An algorithm for cancer risk assessment and decision making in high-risk families using genetic testing is shown in Fig. 25-6. Once a mutation is discovered in a family, subsequent testing of asymptomatic family members can be crucial in patient management. A negative gene test in these individuals can prevent years of anxiety in the knowledge that their cancer risk is no higher than that of the general population. On the other hand, a positive test may lead to alteration of clinical management, such as increased frequency of cancer screening and, when feasible and appropriate, prophylactic surgery. Potential negative consequences of a positive test result include psychological distress (anxiety, depression) and discrimination, although the Genetic Information Nondiscrimination Act (GINA) makes it illegal for predictive genetic information to be used to discriminate in health insurance or employment. Testing should therefore not be conducted without counseling before and after disclosure of the test result. In addition, the decision to test should depend on whether effective interventions exist for the particular type of cancer to be tested. Despite these caveats, genetic cancer testing for some cancer syndromes already appears to have greater benefits than risks. Companies offer genetic testing for many of the cancer syndromes listed in Table 25-3, including FAP (APC gene), hereditary breast and ovarian cancer syndrome (BRCA1 and BRCA2 genes), Lynch’s syndrome (mismatch repair genes), Li-Fraumeni syndrome (TP53 gene), Cowden syndrome (PTEN gene), hereditary retinoblastoma (RB1 gene), and others.

Because of the inherent problems of genetic testing such as cost, specificity, and sensitivity, it is not yet appropriate to offer these tests to the general population. However, testing may be appropriate in some subpopulations with a known increased risk, even without a defined family history. For example, two mutations in the breast cancer susceptibility gene BRCA1, 185delAG and 5382insC, exhibit a sufficiently high frequency in the Ashkenazi Jewish population that genetic testing of an individual of this ethnic group may be warranted.

As noted above, it is important that genetic test results be communicated to families by trained genetic counselors, especially for high-risk high-penetrance conditions such as the hereditary breast and ovarian cancer syndrome (BRCA1/BRCA2). To ensure that the families clearly understand its advantages and disadvantages and the impact it may have on disease management and psyche, genetic testing should never be done before counseling. Significant expertise is needed to communicate the results of genetic testing to families. For example, one common mistake is to misinterpret the result of negative genetic tests. For many cancer predisposition genes, the sensitivity of genetic testing is less than 70% (i.e., of 100 kindreds tested, disease-causing mutations can be identified in 70 at most). Therefore, such testing should in general begin with an affected member of the kindred (the youngest family member still alive who has had the cancer of interest). If a mutation is not identified in this individual, then the test should be reported as noninformative (Fig. 25-6) rather than negative (because it is possible that, for technical reasons, the mutation in this individual is not detectable by standard genetic assays). On the other hand, if a mutation can be identified in this individual, then testing of other family members can be performed, and the sensitivity of such subsequent tests will be 100% (because the mutation in the family is in this case known to be detectable by the method used).

MicroRNAs (miRNAs) are small noncoding RNAs 20–22 nucleotides in length that are involved in posttranscriptional gene regulation. Studies in chronic lymphocytic leukemia first suggested a link between miRNAs and cancer when miR-15 and miR-16 were found to be deleted or downregulated in the vast majority of tumors. Various miRNAs have since been found abnormally expressed in several human malignancies. Aberrant expression of miRNAs in cancer has been attributed to several mechanisms, such as chromosomal rearrangements, genomic copy number change, epigenetic modifications, defects in miRNA biogenesis pathway, and regulation by transcriptional factors. Somatic mutations of miRNAs have been identified in many cancers, but the exact functional consequences of these changes on cancer development remain to be determined. The SomaMir database (http://compbio.uthsc.edu/SomamiR) catalogs somatic and germline miRNA mutations that have been identified in cancer.

Functionally, miRNAs have been suggested to contribute to tumorigenesis through their ability to regulate oncogenic signaling pathways. For example, miR-15 and miR-16 have been shown to target the BCL2 oncogene, leading to its downregulation in leukemic cells and apoptosis. As another example of miRNAs’ involvement in oncogenic pathways, the p53 tumor suppressor can transcriptionally induce miR-34 following genotoxic stress, and this induction is important in mediating p53 function. The expression of miRNAs is extremely specific, and there is evidence that miRNA expression patterns may be useful in distinguishing lineage and differentiation state, as well as cancer diagnosis and outcome prediction.

Certain human malignancies are associated with viruses. Examples include Burkitt’s lymphoma (Epstein-Barr virus), hepatocellular carcinoma (hepatitis viruses), cervical cancer (human papillomavirus [HPV]), and T cell leukemia (retroviruses). The mechanisms of action of these viruses are varied but always involve activation of growth-promoting pathways or inhibition of tumor-suppressor products in the infected cells. For example, HPV proteins E6 and E7 bind and inactivate cellular tumor suppressors p53 and pRB, respectively. There are several HPV types, and some of these types have been associated with the development of several malignancies, including cervical, vulvar, vaginal, penile, anal, and oropharyngeal cancer. Viruses are not sufficient for cancer development, but constitute one alteration in the multistep process of cancer progression.

The tumorigenesis process, driven by alterations in tumor suppressors, oncogenes, and epigenetic regulation, is accompanied by changes in gene expression. The advent of powerful techniques for high-throughput gene expression profiling, based on sequencing or microarrays, has allowed the comprehensive study of gene expression in neoplastic cells. It is indeed possible to identify the expression levels of thousands of genes expressed in normal and cancer tissues. Figure 25-7 shows a typical microarray experiment examining gene expression in cancer. This global knowledge of gene expression allows the identification of differentially expressed genes and, in principle, the understanding of the complex molecular circuitry regulating normal and neoplastic behaviors. Such studies have led to molecular profiling of tumors, which has suggested general methods for distinguishing tumors of various biologic behaviors (molecular classification), elucidating pathways relevant to the development of tumors, and identifying molecular targets for the detection and therapy of cancer. The first practical applications of this technology have suggested that global gene expression profiling can provide prognostic information not evident from other clinical or laboratory tests. The Gene Expression Omnibus (GEO, http://www.ncbi.nlm.nih.gov/geo/) is a searchable online repository for expression profiling data.

With the completion of the Human Genome Project and advances in sequencing technologies, systematic mutational analysis of the cancer genome has become possible. In fact, whole genome sequencing of cancer cells is now possible, and this technology has the potential to revolutionize our approach to cancer prevention, diagnosis, and treatment. The International Cancer Genome Consortium (http://icgc.org/) was developed by leading cancer agencies worldwide, genome and cancer scientists, and statisticians with the goal to launch and coordinate cancer genomics research projects worldwide and to disseminate the data. Hundreds of cancer genomes from at least 25 cancer types have been sequenced through various collaborative efforts. In addition, exome sequencing (sequencing all the coding regions of the genome) has also been performed on a large number of tumors. These sequencing data have been used to elucidate the mutational profile of cancer, including the identification of driver mutations that are functionally involved in tumor development. There are generally 40 to 100 genetic alterations that affect protein sequence in a typical cancer, although statistical analyses suggest that only 8–15 are functionally involved in tumorigenesis. The picture that emerges from these studies is that most genes found mutated in tumors are actually mutated at relatively low frequencies (<5%), whereas a small number of genes (such as p53, KRAS) are mutated in a large proportion of tumors (Fig. 25-8). In the past, the focus of research has been on the frequently mutated genes, but it appears that the large number of genes that are infrequently mutated in cancer are major contributors to the cancer phenotype. Understanding the signaling pathways altered by mutations in these genes, as well as the functional relevance of these different mutations, represents the next challenge in the field. Moreover, a detailed knowledge of the genes altered in a particular tumor may allow for a new era of personalized treatment in cancer medicine (see below). A major effort in the United States, The Cancer Genome Atlas (http://cancergenome.nih.gov) is a coordinated effort from the National Cancer Institute and the National Human Genome Research Institute to systematically characterize the entire spectrum of genomic changes involved in human cancers. Similarly, COSMIC (Catalogue of Somatic Mutations in Cancer) is an initiative from the Welcome Trust Sanger Institute to store and display somatic mutation information and related details regarding human cancers (http://cancer.sanger.ac.uk/).

Gene expression profiling and genomewide sequencing approaches have allowed for an unprecedented understanding of cancer at the molecular level. It has been suggested that individualized knowledge of pathways or genes deregulated in a given tumor (personalized genomics) may provide a guide for therapeutic options on the tumor, thus leading to personalized therapy (also called precision medicine). Because tumor behavior is highly heterogeneous, even within a tumor type, personalized information-based medicine will likely supplement or perhaps one day supplant the current histology-based therapy, especially in the case of tumors resistant to conventional therapeutic approaches. Molecular nosology has revealed similarities in tumors of diverse histotype. The success of this approach will be dependent on the identification of sufficient actionable changes (mutations or pathways that can be targeted with a specific drug). Examples of currently actionable changes include mutations in BRAF (targeted by the drug vemurafenib) and RET (targeted by sunitinib and sorafenib), and ALK rearrangements (targeted by crizotinib). Interestingly, studies have reported that 20% of triple-negative breast cancers and 60% of lung cancers have potentially actionable genetic changes. Gene expression also offers the potential to predict drug sensitivities as well as provide prognostic information. Commercial diagnostic tests, such as Mammaprint and Oncotype DX for breast cancer, are available to help the patients and their physicians make treatment decisions. Personalized medicine is an exciting new avenue for cancer treatment based on matching the unique features of a tumor to an effective therapy, and this concept is in the process of changing our approach to cancer therapy in fundamental ways. On a cautionary note, gene expression can vary enormously within a single person’s cancer and at different anatomic sites in the patient. We have not yet determined whether such clonal variation within an individual tumor will interfere with the goal tailoring therapy to a particular patient’s tumor.

A revolution in cancer genetics has occurred in the past 25 years. Identification of cancer genes has led to a deep understanding of the tumorigenesis process and has had important repercussions on all fields of cancer biology. In particular, the advancement of powerful techniques for genomewide expression profiling and mutation analyses has provided a detailed picture of the molecular defects present in individual tumors. Individualized treatment based on the specific genetic alterations within a given tumor has already become possible. Although these advances have not yet translated into overall changes in cancer prevention, prognosis, or treatment, it is expected that breakthroughs in these areas will continue to emerge and be applicable to an ever-increasing number of cancers.