Table of Contents > Genomics > Neuroblastoma: clinical implications of real-time PCR Print

Neuroblastoma: clinical implications of real-time PCR


Related terms
Future research
Author information

Related Terms
  • Cancer, deoxyribonucleic acid, DNA, expression, gene, NDSP, neuroblastoma, primer, neuroblastoma-derived secretory protein, real-time polymerase chain reaction, ribonucleic acid, RNA, RT-PCR.

  • Neuroblastoma is a form of cancer that affects nerve cells and occurs commonly in children or infants. About 650 new cases of neuroblastoma are reported each year in the United States. Neuroblastoma can undergo metastasis, which is when cells from the original tumor break off and spread to other tissues or parts of the body. Metastasis typically puts patients at greater risk for death because it can lead to tumor growth and impaired function in other organs in the body.
  • By studying the expression of specific genes in patients with neuroblastoma, doctors may be able to make prognoses for patients. A prognosis is a doctor's prediction, based on a range of factors, of how severe a disease is likely to be and whether a patient can expect to recover.
  • Researchers use gene expression analysis to study how much of a particular gene is produced by a cell. Although all cells in the human body contain the same DNA, cell types in the body vary widely. For example, a skin cell is different from a brain cell, in terms of how it looks and what it does. Many of the differences between cells are not caused by differences in the cellular DNA (on average, DNA is 99.8% identical between two people), but rather by differences in the proteins it makes and the functions of these proteins.
  • Cells express genes by making RNA (ribonucleic acid), a molecule derived from DNA that is then converted into proteins. Each gene produces its own unique RNA. A gene is expressed if the RNA for that gene is found in a cell. Measuring the amount of different RNAs in a cell can tell researchers how much of each gene is produced by that cell.
  • DNA is located in the nucleus, a compartment of the cell, and is packaged in structures called chromosomes. Human cells contain 46 chromosomes, including 22 pairs of autosomes and one pair of sex chromosomes (females have two X chromosomes, while males have one X and one Y). Each chromosome has hundreds of genes. Genes contain the instructions for making the proteins that perform all the functions in the human body. Chromosomes also contain many other regulatory sequences that control how much of a gene will be made, when it will be made, and where in the body it will be made.
  • DNA contains four different chemical compounds called bases: cytosine, thymine, guanine, and adenine. In any given person, these bases are found in a particular order along the chromosomes, and it is the order of these bases that stores information for making genes.
  • Researchers may use a technique called real-time polymerase chain reaction, or RT-PCR, to measure the expression of genes. RT-PCR is a method that uses proteins called enzymes to detect and make many copies of genes.
  • RT-PCR may be used to determine prognoses for patients with neuroblastoma. Cancer cells frequently undergo mutations, which cause some genes to be expressed at higher or lower levels than in normal cells. It has been found that abnormal expression of some specific genes in the cancer cells of neuroblastoma patients can be used to predict how severe the cancer is and whether the patient has a good or poor chance of survival. This approach can also be used in many other types of cancer because they will also have abnormal expression of genes.

  • Real-time polymerase chain reaction (RT-PCR) is a method of multiplying a piece of RNA millions of times. It can be used to determine whether a specific RNA fragment is being expressed in a sample and how much of that RNA is present. To perform RT-PCR, RNA is first extracted from a biological sample, such as cells or a tissue. Then, a special type of protein, an enzyme called reverse transcriptase, is used to convert the RNA into DNA.
  • Next, primers, or small pieces of DNA of about 20-30 bases in length that can recognize the DNA of interest, are added to the sample. Researchers commonly select two different primers when performing PCR, and these two primers are usually designed to bind to each end of the DNA sequence being studied. A DNA molecule has two strands, and each primer is designed to recognize a different strand.
  • Another protein, DNA polymerase, is then added to the sample. DNA polymerases are enzymes that are able to synthesize DNA. In a PCR reaction, the DNA polymerase makes thousands or millions of copies of the DNA sequence located between the two primers. DNA polymerase begins copying DNA at the point where a primer is bound, and because each primer is designed to bind to a different strand of DNA, the DNA polymerase is able to duplicate both strands.
  • In RT-PCR, fluorescent dyes or probes that can measure the production of new DNA made by the polymerase are used. By detecting the amount of fluorescent signal produced during a PCR reaction, researchers may make an educated guess about how much RNA was present in the starting sample. If a large amount of an RNA fragment was present initially, more products will be synthesized during the PCR reaction, leading to a greater fluorescent signal. If a smaller amount of RNA is present, there will be fewer products synthesized and a weaker fluorescent signal.

  • Researchers have found that in neuroblastoma cells some genes become deregulated (that is, a loss of normal control), such that they are expressed at abnormally low or high levels. By studying the expression of these genes, researchers may be able to learn more about the disease in a specific patient and assess that patient's chances of survival.
  • For example, researchers have used real-time polymerase chain reaction (RT-PCR) to study the expression of RNA for a gene called neuroblastoma-derived secretory protein, or NDSP. The researchers found that patients with neuroblastoma have higher levels of NDSP expression than healthy individuals. Furthermore, the researchers found that neuroblastoma patients who showed signs of metastasis tended to have the highest levels of NDSP expression. Metastasis occurs when cells from the tumor break off and spread to other tissues or parts of the body. Metastasis typically puts patients at greater risk for death because it can lead to tumor growth and impaired organ function in other parts of the body. Therefore, high levels of NDSP expression appear to predict a poorer outcome in patients, and measuring NDSP expression levels may help predict whether a patient is at increased risk of metastasis.
  • Researchers have identified other genes using RT-PCR whose expression can be used for patient prognosis. Researchers have found that patients with high levels of expression of the genes survivin, tyrosine hydroxylase, MYCN, and ELAVL4 are more likely to have metastatic disease and poorer chances of survival than those who do not have these genes.

  • The use of real-time polymerase chain reaction (RT-PCR) to measure gene expression may help doctors determine whether a patient has advanced or metastatic disease (cancer that has spread to other tissues). RT-PCR gene expression data may also help doctors assess a patient's chances of survival after receiving treatment. Patients who still have high expression of the genes related to poor prognosis after receiving treatment may be more likely to have recurrent disease and be less likely to survive.

  • Real-time polymerase chain reaction (RT-PCR) gene expression cannot predict with absolute certainty whether a patient with neuroblastoma will survive. RT-PCR gene expression data may be used to identify those patients who appear to have more advanced disease and are at greater risk of death, but even patients with poorer prognoses may be able to survive with appropriate treatment.
  • Although the expression levels of specific genes may be measured by RT-PCR and used to formulate a patient's prognosis, it may not be clear why those genes are expressed at abnormal levels in neuroblastoma patients. Additional research is often necessary in order to understand why these specific genes are expressed at higher levels and how they may contribute the disease.
  • Even if a patient has metastatic disease, RT-PCR experiments cannot determine the organs to which the cancer has spread. Other approaches, such as biopsies, may be needed to identify where the cancer has spread.


Future research
  • Researchers have already identified several genes that may be used to develop prognoses of patients with neuroblastoma. In the future, researchers will continue to look for additional genes that are abnormally expressed in neuroblastoma patients. The identification of additional genes that can be used for patient prognosis may further increase the accuracy with which doctors are able to predict the outcome of a disease in a particular patient.

Author information
  • This information has been edited and peer-reviewed by contributors to the Natural Standard Research Collaboration (

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