Stimulated by the growing interest of The Pulse readers in the Pfizer/BioNTech and Moderna vaccines that use messenger RNA (mRNA) to carry a recipe for body cells to make the spike protein SARS-CoV2 (the virus that causes COVID-19) in order to teach the immune system to fight the virus, our opening article of this series introduced you to CRISPR gene editing technology. Developed from components of an immune system employed by microorganisms to fight invading viruses, CRISPR now constitutes an expanding set of technologies being developed and tested as therapeutics against particular medical conditions. There is so much that is happening, too much for just one post, so this post will focus on just one application of CRISPR, namely revving up the production of fetal hemoglobin in order to treat certain hereditary blood diseases. For background, in order to appreciate the clinical applications of the new technology, we need to take a trip, as we often do, through the beautiful world of biology, this time looking at hemoglobin — its structure and function and how it is produced in the precursor cells that become red blood cells (RBCs), the blood cells that carry oxygen from your lungs to your body tissues and that pick up a portion of the carbon dioxide (CO2) that your body cells give off and carry to the lungs to be exhaled.
Like so many of the important substances in your body, hemoglobin is a protein, meaning that it consists, mostly, of building blocks called amino acids that are linked together in chains. In the case of hemoglobin, each molecule consists of four chains of amino acids. I wrote that hemoglobin is mostly amino acids, because each chain also has another part, called a heme group, which has at its center an iron atom whose job is to hold onto a molecule of oxygen (O2) while traveling in a RBC through the blood, and then release the O2 in the tissues where it is needed. During pregnancy, one of the places where the O2 is needed is in the placenta, so that it can transport from RBCs within the mother’s blood vessels to RBCs within the fetal circulation. This is possible, because fetal hemoglobin (the hemoglobin inside the fetal RBCs) is a little bit different from the adult hemoglobin that fills the mother’s RBCs. While O2 sticks both to adult hemoglobin and to fetal hemoglobin, it sticks a little more tightly to fetal hemoglobin. Think of fetal hemoglobin being a slightly stronger magnet than adult hemoglobin and you have the idea. The extra binding strength causes the O2 from the mother’s lungs to move from the mother’s blood to the fetal blood.
When it comes to certain hemoglobin diseases, what’s important is not so much that fetal hemoglobin attracts O2 more strongly than adult hemoglobin does, but that fetal hemoglobin can take the place of the adult hemoglobin that normally fills the RBCs of both adults and children too. As we noted above, a hemoglobin molecule contains four chains. In adult hemoglobin, two of these chains are called alpha chains. While embryonic hemoglobin (hemoglobin that is made early in pregnancy in the developing child) has slightly different chains in place of alpha chains, fetal hemoglobin has two alpha chains identical to the alpha chains of adult hemoglobin. As for the other two chains, they are what make fetal hemoglobin different from the adult hemoglobin that replaces most fetal hemoglobin by age one, or sooner, in most people. Most adult hemoglobin has what are called beta chains, two of them to partner up with the two alpha chains, creating the four chain molecule of hemoglobin. A small percentage of adult hemoglobin (called hemoglobin A2) has similar chains called delta chains in place of the beta chains, but to function properly hemoglobin must have either two beta chains, two delta chains, or two gamma chains partnering up with two alpha chains.
What is interesting from a genetics perspective is that each of these different hemoglobin chains is encoded by a different gene and that the beta gene, the delta gene, and the gamma gene all are located together on the same chromosome (chromosome 11), whereas the alpha gene is located on a different chromosome (chromosome 16) together with genes for the chains that play the role of the alpha chains during the embryonic period. Studies suggest that such an arrangement is because the versions evolved by way of duplication of an ancestral hemoglobin chain gene, followed by the multiple copies each changing in a slightly different way.
At present, the main emphasis in CRISPR therapy is on hereditary disorders of the beta chain, what doctors call beta hemoglobinopathies, because these are the lower hanging fruit on account of what CRISPR editing in its early forms has been able to do. Two of these beta hemoglobinopathies in particular —beta thalassemia and sickle cell disease— are fairly common, so these two have been a focus. CRISPR gene therapy for both is advancing, with beta thalassemia leading the way for reasons that will become clear over the next several sentences.
We have talked about both of these hereditary blood conditions in previous posts, as they are a concern to parents-to-be who may be carriers, depending on their genetic background. Both conditions, or at least being a carrier (having one gene copy for the condition and but also one normal gene copy), occur relatively frequently in people of Mediterranean heritage, although sickle cell disease is particularly common in people of African lineage. But the basic thing to review is that, in beta thalassemia, you don’t make enough beta chain (there actually are two versions of the defective gene, one making a tiny amount of beta chain, the other making none at all), whereas in sickle cell disease the wrong kind of beta chain is made and the chain does damage. Since there are only two copies of the beta gene —one on the chromosome 11 from your mother, the other on the chromosome 11 from your father, beta thalassemia and sickle cell disease are almost the classic recessive, Mendelian diseases, meaning that having one bad gene copy and one normal copy makes you a carrier and usually normal healthwise, while having two genes gives you the disease. There is some variation from this simplified scenario, however for a few reasons, one being what I wrote above about different versions of the bad gene for beta thalassemia, another being that people with sickle cell trait, which is to say carriers, can develop what is called sickle cell crises, under certain circumstances (that’s why they are said to have the trait, rather than just being carriers). And one’s tendency to get sick depends on how much of the bad hemoglobin one’s cells make compared with how much normal hemoglobin.
This means that beta thalassemia is the lower hanging fruit, because something only needs to be supplied and nothing bad needs to be turned off. Meanwhile, in the case of sickle cell disease, it would be nice to turn off production of the something bad, the defective beta chain, but turning on something good to replace it could also be helpful. And what is that something good to be turned on?
It is the gamma chain, the chain that fetal hemoglobin has instead of beta chains. It turns out that some people have a genetic ‘defect’ that causes a certain protein not to be made, or not to function well, and the job of that protein is to turn off the gene for the gamma chain beginning in the months after you are born. There is variation in the timing, even when the gene works, causing some people to start replacing fetal hemoglobin with adult hemoglobin, not until years into life, or even decades into life. And when such people have beta thalassemia or sickle cell disease, they tend to have more mild disease than others whose fetal hemoglobin turns off earlier in life. Given all of this, the early gene therapy utilizing CRISPR has focussed on turning the gene for the gamma chain back on by turning off the gene for the protein that otherwise keeps the gamma chain off. This strategy is the low handing fruit, because the easier thing to do with CRISPR is to design an RNA molecule that delivers a special protein called a CAS to a particular gene and turn it off, generally by cutting it. Since turning off the protein that we mentioned consequently restarts production of the gamma chain, clinical trials of CRISPR therapy have been advancing successfully, particularly for cases of beta thalassemia, where simply supplying a gamma chain to create fetal hemoglobin means that the person will have functional hemoglobin.
But the same treatment is also being tested for sickle cell disease, where it may prove to be effective for some patients, because, even despite the presence of defective beta chain that can cause cells to sickle, revving up the production of gamma chain will produce some normal fetal hemoglobin, thereby preventing at least some of the defective beta chains from partnering up with alpha chains. In other words, making gamma chains might drown out the defective beta chains. Even so, it would be even better to go to the next step and not only turn on the gamma chain production, but also turn off production of the faulty beta chain. If you think that this would make a CRISPR therapy more complicated, then your thinking is correct, yet the technology is getting increasingly more sophisticated anyway.
In a very real sense, there a CRISPR 2.0, a CRISPR 3.0, showing promise for still more disease situations that we will discuss in future posts. It’s going to get very interesting, because the Nobel Prize-winning two scientists whom we mentioned in the introductory post of this series —Emmanuelle Charpentier and Jennifer Doudna— not only have founded different companies developing different forms of CRISPR for different applications, but they have competitors who also have contributed greatly to the discovery of the technology and have founded a bunch of their own companies too. As for this post though, we can certainly say that the fetus comes out as the hero.