Category: 1. Genetic Testing

Circulating tumor DNA (ctDNA) is found in the bloodstream and refers to DNA that comes from cancerous cells and tumors. Most DNA is inside a cell’s nucleus. As a tumor grows, cells die and are replaced by new ones. The dead cells get broken down and their contents, including DNA, are released into the bloodstream. ctDNA are small pieces of DNA, usually comprising fewer than 200 building blocks (nucleotides) in length.

The quantity of ctDNA varies among individuals and depends on the type of tumor, its location, and for cancerous tumors, the cancer stage.

Detection of ctDNA can be helpful in the following cases:

• Detecting and diagnosing a tumor. Because tumor DNA has acquired multiple genetic changes (variants), leading to tumor development, ctDNA is not an exact match to the individual’s DNA. Finding DNA with genetic differences aids in tumor detection. Diagnosing the type of tumor using ctDNA can reduce the need for getting a sample of the tumor tissue (tumor biopsy), which can be challenging when a tumor is difficult to access, such as a tumor in the brain or lung.
• Guiding tumor-specific treatment. Analyzing the genome of tumor cells using ctDNA can help doctors determine which treatment will be most effective. Currently, however, approval from the U.S. Food and Drug Administration for ctDNA testing to personalize cancer treatment is limited.
• Monitoring treatment. A decrease in the quantity of ctDNA suggests the tumor is shrinking and treatment is successful.
• Monitoring periods with no symptoms (remission of cancer). A lack of ctDNA in the bloodstream indicates that the cancer has not returned.

Scientists have discovered that dying tumor cells release small pieces of their DNA into the bloodstream. These pieces are called cell-free circulating tumor DNA (ctDNA).

Noninvasive prenatal testing (NIPT), sometimes called noninvasive prenatal screening (NIPS), is a method of determining the risk that the fetus will be born with certain genetic abnormalities. This testing analyzes small fragments of DNA that are circulating in a pregnant woman’s blood. Unlike most DNA, which is found inside a cell’s nucleus, these fragments are free-floating and not within cells, and so are called cell-free DNA (cfDNA). These small fragments usually contain fewer than 200 DNA building blocks (base pairs) and arise when cells die off and get broken down and their contents, including DNA, are released into the bloodstream.

During pregnancy, the mother’s bloodstream contains a mix of cfDNA that comes from her cells and cells from the placenta. The placenta is tissue in the uterus that links the fetus and the mother’s blood supply. These cells are shed into the mother’s bloodstream throughout pregnancy. The DNA in placental cells is usually identical to the DNA of the fetus. Analyzing cfDNA from the placenta provides an opportunity for early detection of certain genetic abnormalities without harming the fetus.

NIPT is most often used to look for chromosomal disorders that are caused by the presence of an extra or missing copy (aneuploidy) of a chromosome. NIPT primarily looks for Down syndrome (trisomy 21, caused by an extra chromosome 21), trisomy 18 (caused by an extra chromosome 18), trisomy 13 (caused by an extra chromosome 13), and extra or missing copies of the X chromosome and Y chromosome (the sex chromosomes). The accuracy of the test varies by disorder.

NIPT may include screening for additional chromosomal disorders that are caused by missing (deleted) or copied (duplicated) sections of a chromosome. NIPT is beginning to be used to test for genetic disorders that are caused by changes (variants) in single genes. As technology improves and the cost of genetic testing decreases, researchers expect that NIPT will become available for many more genetic conditions.

NIPT is considered noninvasive because it requires drawing blood only from the pregnant woman and does not pose any risk to the fetus. NIPT is a screening test, which means that it will not give a definitive answer about whether or not a fetus has a genetic condition. The test can only estimate whether the risk of having certain conditions is increased or decreased. In some cases, NIPT results indicate an increased risk for a genetic abnormality when the fetus is actually unaffected (false positive), or the results indicate a decreased risk for a genetic abnormality when the fetus is actually affected (false negative). Because NIPT analyzes both fetal and maternal cfDNA, the test may detect a genetic condition in the mother.

There must be enough fetal cfDNA in the mother’s bloodstream to be able to identify fetal chromosome abnormalities. The proportion of cfDNA in maternal blood that comes from the placenta is known as the fetal fraction. Generally, the fetal fraction must be above 4 percent, which typically occurs around the tenth week of pregnancy. Low fetal fractions can lead to an inability to perform the test or a false negative result. Reasons for low fetal fractions include testing too early in the pregnancy, sampling errors, maternal obesity, and fetal abnormality.

There are multiple NIPT methods to analyze fetal cfDNA. To determine chromosomal aneuploidy, the most common method is to count all cfDNA fragments (both fetal and maternal). If the percentage of cfDNA fragments from each chromosome is as expected, then the fetus has a decreased risk of having a chromosomal condition (negative test result). If the percentage of cfDNA fragments from a particular chromosome is more than expected, then the fetus has an increased likelihood of having a trisomy condition (positive test result). A positive screening result indicates that further testing (called diagnostic testing, because it is used to diagnose a disease) should be performed to confirm the result.

Secondary findings are genetic test results that provide information about changes (variants) in a gene unrelated to the primary purpose for the testing.

When a clinician orders a genetic test to discover the genetic cause of a particular condition, the test will often sequence one or a few genes that seem most likely to be associated with that individual’s set of signs and symptoms. However, if the individual’s signs and symptoms do not have an obvious genetic cause, a clinician might order a test that sequences all of the pieces of an individual’s DNA that provide instructions for making proteins (called an exome) or a test that sequences all of an individual’s DNA building blocks (nucleotides), called a genome. These tests are called whole exome sequencing and whole genome sequencing, respectively.

Many more genetic changes can be identified with whole exome and whole genome sequencing than by sequencing just one or a few genes. Sometimes, testing finds a variant that is associated with a condition other than the one for which testing was originally indicated. This is called a secondary finding. Some individuals with a secondary finding may not yet have any of the symptoms associated with the condition, but may be at risk of developing it later in life. For example, a person with a variant in the BRCA1 gene, which is associated with an increased risk of breast and ovarian cancer, may not have developed cancer. Other individuals with secondary findings may have a known medical condition, such as extremely high cholesterol, but receive results that indicate a genetic cause for that condition, such as a variant in the LDLR gene.

In 2013, then again in 2017 and 2021, the American College of Medical Genetics and Genomics (ACMG) recommended that all labs performing whole exome and whole genome sequencing tests report particular secondary findings, in addition to any variants that are found related to the primary purpose of the testing. In the 2021 updated recommendations, ACMG proposed a list of 73 genes that are associated with a variety of conditions, from cancer to heart disease. The 73 genes for which secondary findings are reported were chosen because they are associated with conditions that have a definable set of clinical features, the possibility of early diagnosis, a reliable clinical genetic test, and effective intervention or treatment. The goal of reporting these secondary findings to an individual is to provide medical benefit by preventing or better managing health conditions. The variants that are reported are known to cause disease. Variants of unknown significance, whose involvement in disease at the current time is unclear, are not reported.

The information provided by secondary findings can be very important because it may help prevent a disease from occurring or guide the management of signs and symptoms if the disease develops or is already present. However, as with any type of medical diagnosis, the news of an unexpected potential health problem may lead to additional health costs and stress for individuals and their families. On the basis of secondary findings, additional testing to confirm results, ongoing screening tests, or preventive care may be advised. Individuals receiving whole exome or whole genome sequencing can choose to “opt out” of analysis of the 73 secondary finding genes and not receive variant results. As whole exome and whole genome sequencing become more common, it is important for individuals to understand what type of information they may learn and how it can impact their medical care.

Determining the order of DNA building blocks (nucleotides) in an individual’s genetic code, called DNA sequencing, has advanced the study of genetics and is one technique used to test for genetic disorders. Two methods, whole exome sequencing and whole genome sequencing, are increasingly used in healthcare and research to identify genetic variations; both methods rely on new technologies that allow rapid sequencing of large amounts of DNA. These approaches are known as next-generation sequencing (or next-gen sequencing).

The original sequencing technology, called Sanger sequencing (named after the scientist who developed it, Frederick Sanger), was a breakthrough that helped scientists determine the human genetic code, but it is time-consuming and expensive. The Sanger method has been automated to make it faster and is still used in laboratories today to sequence short pieces of DNA, but it would take years to sequence all of a person’s DNA (known as the person’s genome). Next-generation sequencing has sped up the process (taking only days to weeks to sequence a human genome) while reducing the cost.

With next-generation sequencing, it is now feasible to sequence large amounts of DNA, for instance all the pieces of an individual’s DNA that provide instructions for making proteins. These pieces, called exons, are thought to make up 1 percent of a person’s genome. Together, all the exons in a genome are known as the exome, and the method of sequencing them is known as whole exome sequencing. This method allows variations in the protein-coding region of any gene to be identified, rather than in only a select few genes. Because most known mutations that cause disease occur in exons, whole exome sequencing is thought to be an efficient method to identify possible disease-causing mutations.

However, researchers have found that DNA variations outside the exons can affect gene activity and protein production and lead to genetic disorders–variations that whole exome sequencing would miss. Another method, called whole genome sequencing, determines the order of all the nucleotides in an individual’s DNA and can determine variations in any part of the genome.

While many more genetic changes can be identified with whole exome and whole genome sequencing than with select gene sequencing, the significance of much of this information is unknown. Because not all genetic changes affect health, it is difficult to know whether identified variants are involved in the condition of interest. Sometimes, an identified variant is associated with a different genetic disorder that has not yet been diagnosed (these are called incidental or secondary findings).

In addition to being used in the clinic, whole exome and whole genome sequencing are valuable methods for researchers. Continued study of exome and genome sequences can help determine whether new genetic variations are associated with health conditions, which will aid disease diagnosis in the future.

The main differences between clinical genetic testing and research testing are the purpose of the test and who receives the results. The goals of research testing include finding unknown genes, learning how genes work, developing tests for future clinical use, and advancing our understanding of genetic conditions. The results of testing done as part of a research study are usually not available to patients or their healthcare providers. Clinical testing, on the other hand, is done to find out about an inherited disorder in an individual patient or family. People receive the results of a clinical test and can use them to help them make decisions about medical care or reproductive issues.

It is important for people considering genetic testing to know whether the test is available on a clinical or research basis. Clinical and research testing both involve a process of informed consent in which patients learn about the testing procedure, the risks and benefits of the test, and the potential consequences of testing.

Screening tests evaluate an individual’s risk of developing a genetic condition, while diagnostic tests identify genetic conditions. All genetic tests have both benefits and limitations.

Genetic screening tests are generally used in people who do not have signs or symptoms of a disorder. These tests estimate whether an individual’s risk of having a certain condition is increased or decreased compared with the risk in other people in a similar population. A positive result means that a person’s risk of developing the condition is higher than average. A negative screening test means that a person’s risk is lower than average. However, having a positive screening result does not mean the individual has the condition. Because screening tests are only estimates, in some cases the results indicate an increased risk for a genetic abnormality when the person is actually unaffected (false positive), or the results indicate a decreased risk for a genetic abnormality when the person is really affected (false negative). While genetic screening tests do not provide a conclusive answer, they can help guide next steps, such as whether additional, diagnostic testing is needed.  

Genetic diagnostic tests are often used in people who have signs and symptoms. These tests are used to confirm or rule out suspected genetic conditions. Diagnostic tests can also help inform a person’s chance of developing a genetic condition or of passing on a genetic condition to their children. Diagnostic testing can be performed before birth or at any time during a person’s life, but it is not available for all genes or all genetic conditions. The results of a diagnostic test can be used to guide a person’s choices about health care and the management of the disorder.

Examples of genetic screening tests include:

Noninvasive prenatal testing/screening (NIPT/NIPS): This screening test is performed before birth to help determine the risk that a fetus will be born with certain genetic abnormalities, such as Down syndrome and other chromosomal disorders. 

Newborn screening: In the United States, a screening test is performed on all newborns shortly after birth. This test can assess the risk for developing more than 35 genetic conditions. For many of these conditions, the test analyzes various protein and enzyme levels, which would be abnormal in affected individuals.

Examples of genetic diagnostic tests include:

Molecular gene tests: These tests determine the order of DNA building blocks (nucleotides) in an individual’s genetic code, a process called DNA sequencing. The purpose of these tests is to identify genetic changes that can cause disease.

Chromosomal tests: These tests analyze whole chromosomes or long lengths of DNA to identify large-scale changes, such as an extra or missing copy of a chromosome (trisomy or monosomy, respectively) or abnormalities of large segments of chromosomes, that underlie certain genetic conditions.

A gene patent is the exclusive rights to a specific sequence of DNA (a gene) given by a government to the individual, organization, or corporation who claims to have first identified the gene. Once granted a gene patent, the holder of the patent dictates how the gene can be used, in both commercial settings, such as clinical genetic testing, and in noncommercial settings, including research, for 20 years from the date of the patent. Gene patents have often resulted in companies having sole ownership of genetic testing for patented genes.

On June 13, 2013, in the case of the Association for Molecular Pathology v. Myriad Genetics, Inc., the Supreme Court of the United States ruled that human genes cannot be patented in the U.S. because DNA is a “product of nature.” The Court decided that because nothing new is created when discovering a gene, there is no intellectual property to protect, so patents cannot be granted. Prior to this ruling, more than 4,300 human genes were patented. The Supreme Court’s decision invalidated those gene patents, making the genes accessible for research and for commercial genetic testing.

The Supreme Court’s ruling did allow that DNA manipulated in a lab is eligible to be patented because DNA sequences altered by humans are not found in nature. The Court specifically mentioned the ability to patent a type of DNA known as complementary DNA (cDNA). This synthetic DNA is produced from the molecule that serves as the instructions for making proteins (called messenger RNA).

Genetic discrimination occurs when people are treated differently by their employer or insurance company because they have a gene mutation that causes or increases the risk of an inherited disorder. Fear of discrimination is a common concern among people considering genetic testing.

Several laws at the federal and state levels help protect people against genetic discrimination. In particular, a federal law called the Genetic Information Nondiscrimination Act (GINA) is designed to protect people from this form of discrimination.

GINA has two parts: Title I, which prohibits genetic discrimination in health insurance, and Title II, which prohibits genetic discrimination in employment. Title I makes it illegal for health insurance providers to use or require genetic information to make decisions about a person’s insurance eligibility or coverage. This part of the law went into effect on May 21, 2009. Title II makes it illegal for employers to use a person’s genetic information when making decisions about hiring, promotion, and several other terms of employment. This part of the law went into effect on November 21, 2009.

GINA and other laws do not protect people from genetic discrimination in every circumstance. For example, GINA does not apply when an employer has fewer than 15 employees. GINA also does not protect against genetic discrimination in forms of insurance other than health insurance, such as life, disability, or long-term care insurance.

The physical risks associated with most genetic tests are very small, particularly for those tests that require only a blood sample or buccal smear (a method that samples cells from the inside surface of the cheek). The procedures used for prenatal diagnostic testing (called amniocentesis and chorionic villus sampling) carry a small but real risk of losing the pregnancy (miscarriage) because they require a sample of amniotic fluid or tissue from around the fetus.

Many of the risks associated with genetic testing involve the emotional, social, or financial consequences of the test results. People may feel angry, depressed, anxious, or guilty about their results. In some cases, genetic testing creates tension within a family because the results can reveal information about other family members in addition to the person who is tested. The possibility of genetic discrimination in employment or insurance is also a concern. (Refer to What is genetic discrimination? for additional information.)

Genetic testing can provide only limited information about an inherited condition. The test often can’t determine if a person will show symptoms of a disorder, how severe the symptoms will be, or whether the disorder will progress over time. Another major limitation is the lack of treatment strategies for many genetic disorders once they are diagnosed.

A genetics professional can explain in detail the benefits, risks, and limitations of a particular test. It is important that any person who is considering genetic testing understand and weigh these factors before making a decision.

Genetic testing has potential benefits whether the results are positive or negative for a gene mutation. Test results can provide a sense of relief from uncertainty and help people make informed decisions about managing their health care. For example, a negative result can eliminate the need for unnecessary checkups and screening tests in some cases. A positive result can direct a person toward available prevention, monitoring, and treatment options. Some test results can also help people make decisions about having children. Newborn screening can identify genetic disorders early in life so treatment can be started as early as possible.