Genetic Mutations 101

Understanding their connection to bleeding disorders
Author: Kadesha Thomas Smith

The story of what causes bleeding disorders starts with our body’s effort to write instructions on how it will develop and function. For these instructions to be carried out, and for the story to be completed, they have to be written with razor-sharp precision. In fact, each letter, every word must be carefully placed.

Scientists in the bleeding disorders community have discovered what happens during this “writing process” for people with hemophilia A and B. “We know that mutations lead to bleeding disorders,” says Marion Koerper, MD, pediatric hematologist at the University of California-San Francisco and medical advisor to the National Hemophilia Foundation (NHF). When genetic instructions are not written properly, people can develop bleeding disorders.

The challenge for patients is that many have forgotten how the story goes since they first learned about mutations in middle school science class, Koerper says. Then, when patients or their family members are first diagnosed with a bleeding disorder, they are thrown into a maelstrom of complex terms and decisions—on top of the emotional toll of coping with the new diagnosis. Patients may become distracted from learning what precisely caused the bleeding disorder to occur. “If you don’t understand the basis of genetic mutations, you can’t fully understand hemophilia,” Koerper adds.

Grasping the underlying genetics of hemophilia is even more important now as rapid advances are being made. Medical research is uncovering the link between specific genetic mutations and other aspects of hemophilia, like severity, the development of an inhibitor and, potentially, improved use of treatments.

Let’s review how seemingly tiny genetic changes can lead to hemophilia. As scientists continue building their knowledge base, these genetic insights will ultimately lead to more informed medical decisions now and in the future.

Back to biology class

Each person’s genetic story starts with Mom and Dad. When parents conceive a child, one of Mom’s eggs joins with one of Dad’s sperm, forming a single cell. (Multiple cells form if the couple has twins, triplets and so on.) Deep inside, the cell is carrying 46 chromosomes. These chromosomes are matched up in 23 pairs, half from Mom and the other half from Dad.

All chromosomes have sections called genes. Genes are snippets of molecule strands called deoxyribonucleic acid (DNA). These DNA strands are the written instruction manuals for the biology and characteristics of each person.

The instructions dictate how that initial cell will multiply and develop into various body parts, such as limbs and organs. They determine how the body will be assembled and how each body part will function. DNA also dictates such physical characteristics as eye color, hair texture, complexion and height. These instructions are so thorough, they even spell out details like whether you’ll have dimples or if your pinky finger will be slightly curved. Most important, the instructions give the step-by-step process for making proteins in the body—including the factor proteins needed in blood clotting.

DNA strands are tightly packed into their respective genes. If they were uncoiled, each strand would look like a helix with two adjacent bars in the middle. A single cell’s DNA would be 10 feet long if you stretched it out.

Four letters make up the entire DNA alphabet: A, T, C and G, which stand for adenine, thymine, cytosine and guanine. These nucleic acids are contained in every strand of DNA. Every line of instruction in the DNA is written with these four letters.

For a cell to function normally, there are specific rules for spelling: a T across from every A, and a G across from every C. The sequence of the A/T and G/C combinations can vary from person to person.

Boy or Girl

Among chromosome pairs in the cells, the 23rd pair is most important for hemophilia. These X and Y chromosomes are called the sex chromosomes. They determine if a baby will be a boy or a girl.

Here, mom, who is XX, can only give an X chromosome because that’s all she has. Dad, who is XY, has two options: His sperm can give either an X chromosome or a Y chromosome. If dad’s sperm provides an X chromosome, the baby has two X chromosomes. That means the baby is a girl. If dad’s sperm provides a Y chromosome, the baby has an X chromosome and a Y chromosome. That means the baby is a boy.

Another reason this pair of chromosomes is important for hemophilia is that a couple sections on the X chromosome contain instructions for producing two of the critical proteins needed for blood clotting. One section is in the middle of the X chromosome, Koerper says. It has DNA instructions on how your blood cells should produce the factor IX (FIX) protein. Another section, toward the bottom of the X chromosome, has DNA instructing your body on how to produce the factor VIII (FVIII) protein.

The X chromosome has 1,400 genes comprising more than 150 million A/T and G/C base pairs, according to the chromosome map from the National Center for Biotechnology Information, a national clearinghouse for molecular biology research based in Bethesda, Maryland. For people with hemophilia A and B, there’s a typo in the DNA instructions for producing the FVIII and FIX proteins. “The cell doesn’t understand, so it stops reading,” Koerper explains. Or it keeps reading and makes the protein incorrectly.

That’s the essence of a genetic mutation. When the body can’t properly read the DNA sequence, the instructions for producing clotting factor proteins are disrupted. Finding that abnormal DNA sequence is the important next step toward understanding why a person has hemophilia.

Misspelled instructions

Over the past few years, a push to help people with hemophilia discover their genetic mutation has been underway in the US.

NHF teamed up with the Puget Sound Blood Center, the American Thrombosis and Hemostasis Network and Biogen Idec Hemophilia to pilot genetic testing and mutation analysis at hemophilia treatment centers (HTCs) nationwide. The initiative, called My Life Our Future, launched at 11 pilot sites in November 2012. The protocol is now being opened up to other HTCs nationwide, pending institutional internal review board (IRB) approval.

The goal of My Life, Our Future is to raise awareness of genetic testing for people with hemophilia and to offer it to them free or at low cost. The goal for the larger bleeding disorders community is to help with research efforts that benefit the entire community.

A separate research initiative to build a database of genetic mutations started in 2005. Connie Miller, PhD, began leading a project at the US Centers for Disease Control and Prevention (CDC) in Atlanta to discover what mutations increases the risk of someone with hemophilia developing an inhibitor. An inhibitor is an antibody that prompts the body’s immune system to attack factor medications, which makes them ineffective.

Developing an inhibitor is considered one of the more serious complications of hemophilia. About 25% to 30% of affected individuals, usually with severe hemophilia A, develop an inhibitor within the first 3 to 50 infusions, according to Koerper. For hemophilia B, factor IX deficiency, the risk is much lower, about 3% to 4%.

It turns out that some genetic mutations increase the risk of inhibitors. But first, Miller’s team needed to identify those mutations.

“We thought it would be easy to find, like sickle cell,” says Miller, who has been a genetics researcher working with hemophilia for 30 years. Sickle cell disease is a blood disorder that causes the red blood cells to take on a curved, sickle shape, instead of the normal disc shape, according to the National Heart, Lung, and Blood Institute. Because it’s caused by a single mutation in a small gene, researchers combed through about 2,000 base pairs to find it.

In contrast, there are thousands more base pairs involved in hemophilia. There are about 186,000 DNA base pairs giving instructions for FVIII production and 34,000 DNA base pairs instructing FIX. “And it’s not like other diseases where some of the base pairs aren’t important,” says Miller. “For FVIII and FIX, all the base pairs are important.”

Finding the mutation causing hemophilia requires painstaking detective work. “When you’re looking for a mutation, say for FVIII, you may have to check out all 186,000 base pairs to find the one that’s changed,” Koerper explains.

Still, scientists have categorized some common types of mutations to guide the search:

Inversion

Inversions occur when a section of the X chromosome turns over on itself (inverts), resulting in an incomplete set of instructions. “This is the most common mutation, causing 40% of severe hemophilia,” Miller says. “This is the one we look for first.”

Deletion

A deletion results when a chunk of the DNA base pairs is missing. Picture an instruction manual that is numbered 100, 101, 102, and then skips to 200, Koerper explains.

Nonsense

In a nonsense mutation, the correct letter in one base pair has been swapped for a different letter. So there could be a T where a C should be, or an A where a G should be. This only allows the factor protein to be produced up to a certain point, then it cuts off, Miller says. The result is severe hemophilia.

Missense

Like the nonsense mutation, a missense mutation also swaps one DNA base pair letter for another. However, instead of cutting off protein production, a protein is still produced—just not the right one. About 75% of people with FIX deficiency have a missense mutation, Miller says. Because some protein is still being produced, a missense mutation usually leads to mild or moderate hemophilia.

All of these types of mutations impair the body’s ability to produce factor—if it can produce it at all.

Lengthening list

As Miller and her team of eight researchers continue to study inhibitors, they have tested blood samples from 1,200 people with hemophilia A or B from 17 HTCs. They have also compiled and published two data spreadsheets of all the known genetic mutations linked to hemophilia worldwide: the CDC Hemophilia A Mutation List and the CDC Hemophilia B Mutation List. Some of the mutations have been discovered through the blood sample mutation analyses done in Miller’s lab. Others have been reported from researchers around the world.

So far, nearly 1,100 mutations that cause FIX deficiency worldwide and more than 2,500 mutations that cause FVIII deficiency have been listed. Most mutations fall into one of the categories previously mentioned, but the spreadsheets show a list of other specific mutations, like which base pair is affected, which letter or letters in the base pair have been changed and how. And the list keeps growing.

Miller and her team are using these spreadsheets to clearly identify which mutations increase the risk of inhibitors. For example, only seven people in the study had mild hemophilia and inhibitors, but four of them had the same mutation. “That means this mutation is high risk,” Miller says.

She and her colleagues also noticed something else: African-American and Hispanic patients have twice the rate of developing inhibitors as Caucasians. To accumulate more data and understand why this disparity occurs, Miller’s study has been extended for another two years. She is hoping to get at least 300 more minority patients enrolled. If that happens, the list of mutations is expected to grow even longer.

Regardless of how many thousands of mutations are discovered and how differently the condition progresses in each person, the goal of the hemophilia mutation research is the same. “We want to tell people, ‘You have this type of mutation, which will lead to this type of hemophilia. So, here’s what you need to know going forward,’” Miller says.

Learn more

Download the CDC’s catalog of mutations for hemophilia A and hemophilia B.

University of Utah interactive video on DNA, genes and chromosomes.

 Background information on inhibitors.

My Life, Our Future.

Steps for Living: Hope for Future Treatment