In previous posts, I have discussed a group of inherited diseases known as lysosomal storage diseases. To briefly review, the garbage disposal of the cells is a small membrane-enclosed vesicle known as the lysosome. Lysosomes take up materials that need to be degrades and degrade them, and it does this by means of an array of enzymes called acid hydrolases. These enzymes are not active unless they are present inside the lysosome, since the lysosome acidifies its interior. Acid hydrolases are only active in an acid environment, and this prevents acid hydrolases from degrading the cell if they escape from the lysosome.
If the genes that encode acid hydrolases contain a loss-of-function mutation that causes them to produce nonfunctional proteins, then the lysosome will be unable to degrade particular molecules. Those undegraded molecules will build up to toxic levels and kill cells. These genetic diseases that result from nonfunctional lysosomal acid hydrolases are called lysosomal storage diseases. Fortunately, some new treatment strategies, such as enzyme replacement therapies and stem cell treatments have provided new hope for these diseases.
One particular lysosomal storage disease is called Gaucher disease and this disease results from mutations in the gene that encodes an enzyme called β-glucocerebrosidase. β-Glucocerebrosidase degrades a molecule called glucosylceramide, and in the absence of a functional copy of β-glucocerebrosidase, glucosylceramide builds up and accumulates especially in a type of white blood cell called a macrophage. Glucosylceramidase also tends to accumulate in the spleen, liver, kidneys, lungs, brain and bone marrow.
Babies that have Gaucher disease show enlarged spleens and livers. Their livers do not work properly and they also have bone problems, nervous system problems, swelling of their lymph nodes and their joints too, a bloated, swollen abdomen, a brownish tint to the skin, messed up blood work (too few red and white blood cells and platelets), and yellow fatty deposits on the white of the eye.
Gaucher disease is caused by a recessive mutation in a gene located on chromosome 1 and it equally affects both males and females. About 1 in 100 people in the United States are carriers of the most common type of Gaucher disease, but the carrier rate among Ashkenazi Jews is 8.9% while the birth incidence is 1 in 450.
Some forms of Gaucher disease can be treated by enzyme replacement treatment in which recombinant glucocerebrosidase (imiglucerase) is administered intravenously. Such treatments dramatically decrease liver and spleen size, reduce skeletal abnormalities, and reverse other manifestations of Gaucher disease. Unfortunately, this treatment is rather expensive (approximately US $200,000 per year for a single patient), has to be continued for life. Another treatment, Velaglucerase alfa, was approved by the Food and Drug Administration (FDA) as an alternative treatment in February 2010, and in May 2012 the FDA approved an additional treatment called Taliglucerase alfa, or Elelyso.
Is there a better way to treat this disease? To determine that, we need to know more about the disease, and that requires a better model for Gaucher disease. Fortunately, induced pluripotent stem cells (iPSCs) can provide such a model system, and some enterprising stem cell biologists have just made such a model system for Gaucher disease. Leelamma Panicker, and colleagues in the laboratory of Ricardo Feldman at the University of Maryland School of Medicine, in Baltimore, Maryland have made iPSCs from patients with Gaucher disease. These cells show many of the pathologies of Gaucher disease patients.
Panicker and her colleagues made human iPSCs from skin fibroblasts from patients with all three types of Gaucher disease. Gaucher disease comes in three different types, type 1, 2, and 3. Type I is a milder form of the disease and tends occur in Ashkenazi Jews, at 100 times the occurrence in the general populace, and the median age at diagnosis is 28 years of age. The life expectancy is mildly decreased, but there are no neurological symptoms. Type II is more severe and is characterized by neurological problems in small children. The prognosis is poor and most of these children die before the age of 3. Type III is moderately severe and occurs in Swedish patients from the Norrbotten region, and this disease is manifested somewhat later in life but most die before their 30th birthday.
The iPSCs made from Gaucher disease patients were differentiated in all kinds of cell types and tissues. They used the iPSCs to form a special type of tumor called teratomas in laboratory animals. Teratomas grow quickly and form a wild pastiche of tissues and cell types as they overgrow. Most usefully, the teratomas formed a whole wade of cell types that show pathologies in Gaucher disease. For example the teratomas differentiated into macrophages and nerve cells. The macrophages were able to gobble up materials responded to and were capable of responding to foreign molecules. This demonstrated that these macrophages are real macrophages.
The really cool part about these iPSC-derived cells is that they show many of the pathological hallmarks of Gaucher disease. First, the macrophages exhibited low levels of glucocerebrosidase enzymatic activity. They also accumulated glucosylceramide and their lysosomes worked very poorly to say the least. These macrophages were largely unable to digest red blood cells they had gobbled up. This is a feature of macrophages from patients with Gaucher disease. Even more interestingly, the speed with which each cultured macrophage population was able to digest red blood cells tightly correlated with the type of Gaucher disease that afflicted the patient from whom the original cells were isolated. Thus, the macrophages made from iPSCs derived from type I patients digested red blood cells the fastest of three lines, and those from type II patients digested them the slowest, if at all. To ensure that a lack of beta-glucosylceramidase was the problem with these cells, they Incubated the macrophages with recombinant beta-glucosylceramidase, and this completely reversed the delay in red blood cell digestion. if these macrophages were treated with a drug called isofagomine (Plicera), which is made by Amicus Therapeutics and has been approved as a treatment for Gaucher disease, the digestion of red blood cells was only partially restored. Isofagomine binds to beta-glucosylceramidase and locks it into a conformation that increases its activity by three-fold.
These experiments show that these cell types that were derived from iPSCs made from Gaucher’s disease patients effectively recapitulates the pathologic hallmarks of the disease.
According to senior author, Ricardo Feldman, associate professor of microbiology and immunology at the University of Maryland and a research scientist at the University of Maryland Center for Stem Cell Biology and Regenerative Medicine: “We are confident that this will allow us to test more drugs faster, more accurately, and more safely, bringing us closer to new treatments for Gaucher disease. Our findings have potential to help patients with other neurodegenerative diseases as well. For example, about 10 percent of Parkinson’s patients carry mutations in the recessive gene for Gaucher disease, making our research possible significant for Parkinson’s disease as well.”
This strategy will revolutionize the use of model systems to study and design and test drugs for genetic and metabolic diseases. This also demonstrate why Shinya Yamanaka’s Nobel Prize this year in Medicine for designing the technology that gave us iPSCs is so well deserved.