The Emerging Role of Genomics in Diagnosing and Monitoring Cancer


Cancer is the result of genetic abnormalities that affect the function of particular genes. Genes determine the form, function, and growth patterns of cells. Those that accelerate or suppress growth are often involved in cancer. For example, many cancers have an abnormality in a gene that is responsible for stimulating cellular growth and/or the gene that normally prevents cancer is not working properly. Both of these genetic abnormalities can result in uncontrolled and excessive cellular growth, the hallmark trait of cancer. Genomic tests, or assays as they are called by scientists, are a tool for identifying the specific genes in a cancer that are abnormal or are not working properly. In essence, this is like identifying the genetic signature or fingerprint of a particular cancer.

Genomic testing is different from genetic testing. Genetic tests are typically used to determine whether a healthy individual has an inherited trait (gene) that predisposes them to developing cancer. Genomic tests evaluate the genes in a sample of diseased tissue from a patient who has already been diagnosed with cancer. In this way, genes that have mutated, or have developed abnormal functions, are identified in addition to those that may have been inherited.

Genomic testing can help doctors to:

  • Determine a patient’s prognosis (potential outcome)
  • Determine whether a cancer is aggressive/fast growing or slow growing
  • Choose the most effective treatment for each individual cancer
  • Monitor patients who are undergoing treatment to determine if the treatment is working
  • Monitor patients who are in remission to catch a potential disease progression early when it is more treatable

Perhaps the greatest promise of genomic testing is its potential for individualizing treatment. This means that patients with more serious conditions can be identified and offered aggressive and innovative therapies that may prolong their lives, while patients who are diagnosed with a less serious condition may be spared unnecessary treatments. For example, some women with node-negative breast cancer will relapse after being treated with surgery alone. Genomic testing has been shown to differentiate between which node-negative breast cancer patients are more likely to relapse and therefore benefit from additional chemotherapy and which patients may not need chemotherapy.

To appreciate how the science of genetics is applied to the diagnosis and monitoring of cancer, it is helpful to have an understanding of the basic principles of genetics. This includes knowing what DNA, chromosomes, and genes are, how they work, and how the information contained in DNA is transformed, through gene expression, into specific structures that dictate the functions of a cell.

With this background knowledge, it is possible to understand the promise of tests for detecting genetic abnormalities, such as:

  • Fluorescence in situ hybridization (FISH)
  • Polymerase chain reaction (PCR)
  • Reverse transcription PCR
  • Microarray technology
  • Serum proteomics

Background—Basic Principles of Genetics

The importance of genetics in heredity is well known; however, the role that genetics plays in controlling the structure and function of cells may be even more critical for an individual organism. Heredity assures that humans and all species are able to reproduce and perpetuate their unique traits and directing how cells are built, what work they do, and how they grow is necessary to ensure that an organism will survive to reproduce. An understanding of this critical role that DNA and genes have in determining the minute-by-minute life of a cell is also important for understanding how genetics are involved in cancer.

DNA: The genetic information for an entire organism is contained in the nucleus of every cell in the form of deoxyribonucleic acid, commonly known as DNA. DNA is a double-stranded helical (coiled) molecule. Each strand is composed of a structural backbone plus a sequence of nitrogen-containing compounds called nitrogenous bases, which can be thought of as the alphabet of genetics. There are four bases: adenine, guanine, thymine, and cytosine. The two strands are connected at the bases.

The genetic code, or the genetic information that controls structure and function of the cell, is contained in the sequence of bases. The base sequence eventually controls the sequence of amino acids that are connected together to make a protein molecule. Different sequences make different proteins. The proteins that are synthesized in a cell determine the structure and function of that cell.

Chromosomes: DNA is packaged in a specific number of units called chromosomes. Humans have 46 chromosomes in each cell. Most of the time, the chromosomes are packed tightly around proteins in the nucleus of the cell so that they cannot be seen. However, in the stages of the cell’s life just before cell division, the chromosomes become visible with a light microscope. They appear like a capital ‘H’ with four lengths of coiled DNA joined by a protein as the “cross” of the “H”.

Genes: DNA is organized into genes, which are long segments of DNA that include regions that contain codes for proteins called exons, as well as non-coding regions called introns. Genes are defined as the basic unit of heredity because they are passed to offspring and then replicated and passed on to individual cells during cell division. Replication involves using both strands of DNA as templates to synthesize complimentary DNA (cDNA), which is a matching strand. The result is two identical copies of DNA for each cell after cell division is complete. Under normal conditions, the structure of DNA, and thus genes, remains relatively constant through replication and cell division.

Gene expression: The genetic information contained in genes is translated into cellular structure and function through a process called gene expression. Genes can be thought of as codes, or recipes, for making proteins. Proteins are the basic component of cell structure and function. When a gene is “expressed,” the protein or proteins that it codes for are actively being built in the cell and the function that those proteins serve are being performed. For example, when the HER-2/neu gene is expressed in breast cancer, there are more epidermal growth factor receptors (EGFRs) present, which are proteins on the cell surface that HER-2/neu codes for. Furthermore, the function of EGFR is to stimulate cell growth; so a cell that is expressing HER-2/neu has many EGFRs and is actively growing.

Gene expression occurs through a complex system that involves the following steps:

  • Temporary separation of the two strands of the DNA molecule at a particular gene.
  • Transcription of the segment of DNA, which is the synthesis of a single-stranded copy of the DNA sequence that is exposed; this copy is called messenger RNA (mRNA).
  • Protein synthesis, or building new proteins in the cell, based on the information contained in the mRNA.

Genetic abnormalities: Genetic abnormalities are alterations in the DNA of a cell that may occur by chance or due to an environmental influence. These alterations lend the affected cell some advantage over normal cells that helps them grow. As a result, the cell is able to divide rapidly, becoming a cancer growth. However, this growth advantage only benefits the individual cell, and not necessarily the whole organism (human).

Types of genetic abnormalities include:

Translocations—the changing places of a gene from one chromosome with a gene on another chromosome; this type of abnormality defines the many different leukemias

Deletions—a gene or sequence of nucleotides is missing in the DNA

Polymorphisms—variations in nucleotide sequence

Tests for Detecting Genetic Abnormalities

A variety of new laboratory tests can detect genetic abnormalities. Finding a disease-causing mutation in a gene can confirm a suspected diagnosis of cancer or identify those predisposed to certain cancers. Some of these techniques that are currently used in the clinical setting include:

  • Fluorescence in situ hybridization (FISH)
  • Polymerase chain reaction (PCR)
  • Reverse transcription PCR

Furthermore, the following laboratory techniques are being used in cancer research and may be available for clinical use in the future:

  • Microarray

Fluorescence in situ hybridization (FISH)

FISH is a laboratory technique that is used to detect genetic abnormalities at the single-cell and single-gene level, such as numerical abnormalities (gains and losses of nucleotides), and translocations (the changing places of a gene or segment of genes on one chromosome with gene or a segment on another chromosome). These abnormalities play a role in the development and progression of some cancers, such as leukemias and lymphomas[1].

How does FISH work? FISH is performed on sample cells whose DNA has unraveled so that the individual chromosomes are visible. This happens during cell phases just before cell division, called metaphase or interphase. The sample DNA is first denatured using heat and the chemical formamide so that the individual strands separate, exposing the base sequence. Next, specific DNA sequences, called probes, that are attached to colored fluoros are incubated, or combined, with the sample DNA. The probes hybridize (connect) with the DNA in the chromosomes that is the compliment to the base sequence in the probe. The presence or absence of fluorescence from the hybridized DNA and probe are visible with a specialized microscope and indicate whether the DNA sequence of interest is present in the sample. Furthermore, specialized FISH techniques can be used to detect translocations, inversions, and amplifications that are involved in cancer.[2]

FISH in breast and ovarian cancer: A common use if FISH is to determine whether patients with breast and ovarian cancer overexpress the HER2/neu oncogene, a gene that is commonly involved in cancer. HER2/neu carries the genetic code for the HER2 receptor, a protein on the surface of some cancer cells. HER2 binds with growth factors in the blood, thereby stimulating cancer cells to grow.

HER2/neu is amplified in approximately 20% to 30% of breast and ovarian cancers and this amplification and/or overexpression indicates a poor prognosis.[3] FISH can be used to observe whether the HER2/neu oncogene is sending multiple signals at the level of the individual cells, which indicates gene amplification.

FISH in hematological (blood) cancers: FISH may also be used to diagnose and manage various hematological malignancies. The genetic abnormality that underlies many hematological malignancies is chromosomal translocation, or the changing places of gene from one chromosome with a gene on another chromosome.

Polymerase chain reaction (PCR)

PCR is an in vitro laboratory method that is useful for genetic testing for disease and detecting minimal residual disease, which is a small amount of disease left after treatment that may lead to recurrence and is typically not detectable with other techniques. This procedure amplifies a segment of DNA from a small sample, making it detectable. With PCR, relatively small sequences of known DNA can be replicated into millions of copies over a short period of time.

How does PCR work? This method requires four principle components: 1) the sample DNA, 2) an ample supply of nucleotides, 3) a heat-stable polymerase enzyme which is responsible for copying DNA, and 4) primers, short sequence of nucleotides that lie on either side of the DNA fragment of interest and signal the polymerase to begin replication of the specific DNA segment.

PCR is a three-step process, each occurring at a different temperature. The sample DNA is first heated to approximately 90ºC in order to separate the 2 paired DNA strands. Once separated, it is cooled to a temperature that allows the primers to hybridize to their complementary sequence on the target DNA, approximately 40ºC. Lastly, DNA replication occurs at approximately 70ºC, the temperature at which DNA polymerase is most active. This process is repeated 20 to 30 times, resulting in approximately 1 million-fold amplification of the DNA fragment of interest.[4]

Reverse transcription PCR

Reverse transcription (RT)-PCR is a technique that detects the degree to which genes are expressed. Complicated processes control which segment of DNA separates, gets transcribed (copied) into mRNA, and then expressed as proteins in the cell. Not all genes are transcribed and then expressed equally. Due to many controls in the cell, some genes are over-expressed, which means they are transcribed and expressed at a higher rate than normal, while other genes are now expressed, or “turned off” so that certain functions are not manifested in the cell.

How does RT-PCR work? RT-PCR uses the same steps as PCR to amplify a segment of DNA, but the sample is a complimentary copy of mRNA. By starting with mRNA, this test measures only the DNA that is expressed, making it possible to determine the degree to which certain genes are expressed. Recent uses of RT-PCR in clinical oncology include detection of lymph node micrometastases in prostate cancer and bone metastases in breast cancer.[5]

RT-PCR in breast cancer: The breast cancer test Oncotype DX utilizes RT-PCR to determine the individual risk of recurrence in women with node-negative estrogen receptor (ER)-positive breast cancer. This test evaluates expression of 21 genes in breast cancer. Overexpression of some of these genes indicates a worse prognosis, while expression of others may indicate a better prognosis. The expression of all 21 genes is used to calculate a “Recurrence Score™”, or the likelihood that that cancer will recur. A large clinical trial showed that Recurrence Score™ was more effective for predicting prognosis of women with node-negative, ER-positive breast cancer than standard measures such as patient age, cancer size, and cancer stage.[6]

Strategies to Improve Detection of Genetic Abnormalities

Several methods for detecting genetic abnormalities are being utilized for cancer research. While they are not yet routinely used in the clinical setting, the following appear to be promising and may be used in the future for diagnosing, testing, and monitoring cancer.

Microarrays: Microarray analysis is a technique that combines biology with computer science to generate a genetic profile for a given tissue sample that reflects the activity of thousands of genes. This technology has advantages over FISH or PCR because, in a single analysis, it can evaluate the expression of all of the genes that may be involved in a cancer, rather than just a few. By graphically showing how all of the genes are involved in a cancer, microarrays can generate a “genetic signature” for a particular cancer. This makes the identification of cancer subtype more precise. The ability to take a snapshot of a cancer’s genetic signature may lead to a better understanding of how that cancer develops and how to design individualized treatment.

How do microarrays work? While different microarray methods are utilized, each consists of five basic steps:

  • Preparation of the sample
  • Combining the sample with the computer chip
  • Scanning the computer chip
  • Normalization
  • Computer analysis of the results.

Preparation of the sample: In the initial step, cDNA is synthesized from RNA by reverse transcription (remember transcription involves copying DNA to make RNA, so reverse transcription is generating DNA from RNA) from RNA that has been extracted from both a test and a reference sample. The sample DNA segments are labeled with fluorochromes, or radioactive chemicals, so that they can be detected after they combine with the computer chip.

Combining the sample with the computer chip: Next, the sample is combined with the computer chip, which is a rectangular grid of spots. Each spot has many copies of a particular DNA sequence. These sequences are derived from public databases of DNA sequences that were generated through the Human Genome Project, the scientific endeavor that identified virtually all of the DNA sequences in the human species.

When the sample is added to the computer chip, a process called hybridization occurs. This means that the sample DNA segment binds (hybridizes) to the segment on the computer chip that has the exact complimentary sequence of nucleotides (the four compounds that are the alphabet of genetics).

Scanning the computer chip: Once hybridization is complete, scanners are used to detect the fluorescence and create a digital image that reflects where the sample DNA combined with spots on the microarray chip.

Normalization: Because raw signal intensities may vary between individual chips from many patients or experiments, individual chip intensity must be adjusted to a common standard, or normalized. For example, subtraction of background noise is a common normalization method that is applied to all samples. Normalization makes it possible to compare gene expression profiles from many patients or experiments.

Computer analysis: The final step in a microarray experiment is computer analysis. The thousands of raw data points that result from microarray analyses are essentially unintelligible unless they are evaluated in the context of other results. For example, the gene expression profile (microarray results) of normal and diseased tissue can be compared to identify genes that vary in their expression and also identify a pattern (profile) that may indicate a distinct class or stage of disease.[7]

Microarrays in oncology: Microarray analysis has contributed to oncology by increasing an understanding of the genetic basis of several types of cancer, including B-cell non-Hodgkin’s lymphoma (BCNHL), acute leukemia, and breast cancer.

  • Considerable knowledge regarding the pathology of BCNHL has been gained by comparing gene expression patterns of diseased and normal tissue. Two different disease categories display distinct gene expression profiles. Microarrays have helped establish these expression profiles and, in the future, they may help accurately classify new cases of BCNHL.
  • In the case of acute leukemia, microarrays have helped establish distinct gene expression patterns that have helped differentiate acute lymphocytic leukemia (ALL) and acute myeloid leukemia (AML). Using these profiles, 29 of 34 new cases of leukemia were correctly predicted.
  • Furthermore, microarrays have helped identify two distinct gene expression profiles in breast cancer, BCRA1 and BCRA2. This finding suggests different ways that breast cancer develops and provides clues that promote further understanding of the cause of breast cancer.[7]


[1] Spagnolo SD, Ellis DW, Juneja S, Leong AS, et al. The role of molecular studies in lymphoma diagnosis: a review. Pathology 2004; 36 (1)19-44.

[2] Spurbeck JL, Adams SA, Stupca PJ, Dewald GW. Primer on Medical Genomics Part XI: Visualizing Human Chromosomes. Mayo Clinic Proceedings 2004:79:58-75.

[3] Paik S, Hazan R, Fisher ER, et al. Pathological findings from the national surgical adjuvant breast and bowel project: prognostic significance of erb B-2 protein overexpression in primary breast cancer. J Clin Oncol 1990;8:103-112.

[4] Tefferi A, Wieben ED, Dewald GW, et al. Primer on Medical Genomics Part II: Background Principles and Methods in Molecular Genetics. Mayo Clinic Proceedings 2002;77:785-808.

[5] Tefferi A, Wieben ED, Dewald GW, et al. Primer on Medical Genomics Part II: Background Principles and Methods in Molecular Genetics. Mayo Clinic Proceedings 2002;77:785-808.

[6] Paik S, Shak S, Tang G, et al. Multi-gene PT-PCR assay for predicting recurrence in node negative breast cancer patients—NSABP studies B-20 and B-14. Proc of the 26th Annual San Antonio Breast Cancer Symposium. December 3-8k, 2003; San Antonio, TX, Abstract #16.

[7] Tefferi A, Bolander ME, Ansell SM, et al. Primer on Medical Genomics Part III: Microarray Experiments and Data Analysis. Mayo Clinic Proceedings 2002;77:927-940.