Gene Therapy & Stem Cell Cure for Anemia?

Novel combo stem cell-and-gene therapy may cure sickle-cell anemia, beta-thalassemia. A new experimental therapy that combines advanced stem cell techniques with the latest in gene therapy promises to cure sickle cell anemia and other debilitating and life-threatening genetic disorders that arise in the blood.

Such a cure is the goal of a US$6.7-million, five-year research project headed by University of California in San Francisco scientist Dr. Y. W. Kan, a pioneer of modern genetics and the diagnosis of genetic diseases before birth. The research began last December (2011).

The new pre-clinical research that combines stem cell and gene therapies is funded through a competitively awarded grant from the United States National Institutes of Health (NIH) and will be undertaken by Dr. Kan and other UCSF collaborators including Dr. Dieter Gruenert and Dr. Marcus Muench.

“This project offers a possibility of curing both newborn and adult patients with their own cells that have been reprogrammed, corrected and converted to cells that will regenerate all of their blood cells, including (that in) the immune system,” Dr. Kan explains.

The UCSF researchers aren’t working with controversial human embryonic stem cells (hESCs), but with stem cells derived by manipulating and reprogramming cells from human adults.

Put simply, the experiment involves:
• First, deriving stem cells from a patient’s own tissue by converting the patients’ blood cells into powerful inducted pluripotent stem cells or iPSCs.
• Second, correcting the inborn genetic error in these cells through genetic engineering or replacing a defective portion of the genes in the stem cells using experimental techniques.
• Third, coaxing the genetically “corrected” iPSCs to become hematopoietic stem cells—the type that can specifically regenerate the entire range of blood cells.
• Fourth, returning the “corrected” stem cells to the patient through transfusion, where they are expected to regenerate and replace the patients’ damaged blood cells.

By developing this procedure, the UCSF scientists are hoping to hit two birds with one stone:
• Circumvent the need to find immunologically compatible bone marrow donors for blood disease patients since such compatible donors are often very difficult to find.
• Eliminate the threat of graft versus host disease, since the use of stem cells derived from the patient don’t cause unwanted immune responses when put back into the same person.

Other UCSF scientists who are playing a leading role in this research include:

• Dr. Muench, a senior scientist at Blood System Research Institute, whose research for two decades has focused on understanding the development of hematopoietic cells and the development of new stem cell transplantation strategies
• Dr. R. Geoffrey Sargent, who studies new ways to genetically modify stem cells
• Dr. Long-Cheng Li, a pioneer in the use of another tool, called small activating RNA, used to develop iPSCs

Expected to benefit first from the experimental treatment are people who suffer from sickle-cell anemia and beta thalassemia—blood-forming system disorders that are caused by defects in a single gene, the gene for beta globin. Part of hemoglobin molecule in red blood cells, beta globin incorporates iron to pick up oxygen in the lungs and deliver it to tissues throughout the body.

What it means for sufferers
It’s a rare, inherited blood disorder, but sickle cell anemia still affects millions throughout the world.

Most of the people who suffer this disease are in Africa, but still about 80,000 people in the United States have the disease and thousands in Canada, as well as the United Kingdom, France, Italy and other European countries.

Found mainly in people whose ancestors come from sub-Saharan Africa, it occurs to a lesser degree in people from South and Central America, Saudi Arabia, India and Mediterranean countries like Turkey, Greece, and Italy.

Anemia often develops in people with the disease because sickle cells die off quickly and bone marrow doesn’t make new ones fast enough. But while serious, anemia is only one common symptom.

When affected red blood cells lose oxygen, they collapse into a sickle or C shape, become stiff and sticky and can clump together and block small blood vessels—causing episodes of severe pain that can last hours or days and can recur repeatedly.

Sufferers describe the pain like a jackhammer on the back, the stomach, in the head and sometimes in the whole body that is relieved only by powerful, addictive painkillers.

Sickle cell anemia isn’t only incurable, it’s chronic—and the pain and complications associated with the disease impact a patients’ quality of life profoundly, cutting down their ability to work, and long-term health and well-being.

Beta-thalassemia is another rare, inherited blood disorder that is most common in Mediterranean and Asian populations.

People with this ailment suffer from severe anemia that doesn’t only make them weak and tired but may even require them to undergo frequent blood transfusions. If untreated, severe anemia results in bone thinning and heart failure.

Right now, these diseases are treated with transplantation of bone marrow or cord blood. But compatible donors are needed—and aren’t often readily available.

Patients who cannot receive bone marrow transplants to cure the disease require frequent blood transfusions.

But iron can eventually build up in the body and damage organs as a result of continual transfusions. Increased deposits of iron can cause thyroid and liver failure — and even heart failure. Organ failure is, in fact, a common cause of death in people with blood-forming diseases.

Nowadays, patients are treated with chelation to remove excess iron. But even with these treatments, patients survive only up to middle age.

And treatments are costly, too — making them out of reach for many sufferers. Direct medical costs for these diseases with treatment that includes chelation are estimated by studies to average more than US$60,000 dollars per year for each patient.

Another main obstacle to treating a larger number of African-Americans with sickle cell disease is the relative lack of a compatible donor.

Combining stem cell and gene therapy
This isn’t the first time that Dr. Kan and Dr. Gruenert, respected pioneers in their own fields, have collaborated.

The two scientists previously worked together on an NIH-funded Gene Therapy Core Center at UCSF in the 1990s. That and other research collaborations have made them familiar with a broad range of stem-cell and gene-therapy techniques.

Both scientists are members of the UCSF Institute for Human Genetics and the Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research.

In early studies done four decades ago, Dr. Kan identified DNA mutations responsible for diseases, leading to some of the first prenatal, DNA-based, diagnostics tests.

For his part Dr. Gruenert, co-director of the new project, is an innovator in the development of gene therapy strategies. A method he’s developed and patented will be used in the current project to replace the defective beta globin genes in induced pluripotent stem cells derived from the blood cells taken from patients.

After the UCSF researchers take blood cells from people with sickle-cell anemia and beta thalassemia, they will create induced pluripotent stem cells—iPSCs that can be grown indefinitely and induced to generate into specialized cells of almost any part of the body.

Once they’re created the pluripotent stem cell lines, they will replace the defective stretch of the beta globin gene using a technique called small fragment homologous replacement — first developed by Dr. Gruenert in the 1990s and now used in gene therapy studies. In previously research, he’s been able to use the technique to correct physiological defects caused by mutations in the cystic fibrosis gene.

With small fragment homologous replacement, researchers will be able to perform genetic “surgery” that will get rid of the genetic defect that causes the blood-forming diseases.

Genetic “surgery” involves swapping mutation-containing DNA regions that are hundreds of nucleic acid base pairs long.

But Dr. Gruenert also expects to combine this small fragment homologous replacement technique with other new methods used to make target-gene modification more efficient.

Another major objective of the multimillion USCF research project is to find a way to safely reprogram cells and generate pluripotent stem cells without introducing unnecessary DNA that pose the risk of creating cancers or other disruptions to normal cellular functioning.

Since 2007, when Dr. Shinya Yamanaka first succeeded in creating immortal pluripotent stem cells from skin cells, many of the techniques used by scientists to generate stem cells allowed foreign reprogramming DNA to be incorporated randomly into the cellular genome—leading to tumors and other problems.

The new research holds great promise, but even Dr. Kan advises caution. Much more work needs to be done before the approach can be applied to patients, he says.

Blood Stem-Cell Transplant Regimen Reverses Sickle Cell Disease in Adults
About two years ago, another study showed another form of stem-cell transplants effectively reversing sickle cell anemia in nine out of 10 adults suffering severe forms of the disease.

The treatment, which involved a modified form of adult blood stem cells, was published in the Dec. 10, 2011 issue of the New England Journal of Medicine. It was conducted by NIH researchers from the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), the National Heart, Lung and Blood Institute (NHLBI), and the National Institute of Allergy and Infectious Diseases, at the NIH Clinical Center in Bethesda, Maryland.

In previous trials by other researchers, nearly 200 children with severe sickle cell disease were cured when their bone marrow was first completely destroyed with chemotherapy then replaced with bone marrow transplants.

But adults who had suffered years of accumulated organ damage from the disease are less able to tolerate complete marrow transplantation and chemotherapy had proven too toxic.

Instead of completely replacing the bone marrow, as is the established method for children with sickle cell anemia, the new adult trial sought to reduce toxicity by only partially replacing the bone marrow.

This was possible since normal red blood cells have a much longer life span compared to sickle red blood cells, allowing the healthy cells to outlast — and after several partial transplants — completely replace the disease-causing cells.

But transplants of tissue or organs — even bone marrow — pose the risk of graft-versus-host disease, in which the recipient’s immune system rejects the transplants from the donor. GVHD is a common complication of stem cell transplantation and can lead to rash, diarrhea and nausea, and even serious problems like liver disease and death.

To replace the disease-causing red blood cells completely without causing GVHD, the NIH researchers used two drugs, alemtuzumab and sirolimus coupled with a low dose of radiation directed at the whole body.

Alemtuzumab depletes immune cells, but doesn’t adversely affect blood stem cells, while sirolimus doesn’t block the activation of immune cells but inhibits their proliferation. By using the two drugs in tandem, investigators sought to create a balance to help prevent rejection of the new stem cells.

Meanwhile, the radiation favorably conditions the bone marrow, where donor stem cells move in and begin producing new, healthy red blood cells.

Examining the patients after an average of two and one half years, researcher found that all 10 recipients were alive and sickle cell disease was eliminated in nine.

“Our patients have had a remarkable change in their lives,” Dr. John F. Tisdale, the trial’s principal investigator in the NIH Molecular and Clinical Hematology Branch says in a press release last year.

“They’re no longer being admitted to the hospital for frequent pain crises. They have been able to stop chronic pain medications, go back to school and work, get married and have children,” he says.

“Given these results, our regimen will likely have broad application to other nonmalignant diseases and can be performed at most transplant centers,” he concludes.

For the transplants, bone marrow was taken from donors, mostly siblings of patients, who were matched immunologically with the patients through human leukocyte antigen (HLA) typing.

Hemacord approved by FDA
In November last year, a new therapy using human cord blood iNPCs to treat patients with blood-forming system disorders, was approved by the U.S. Food and Drug Administration.

The new product, called HEMACORD [see: Hemacord: Stem Cell Cure for Hemophilia Gains FDA Approval], is made from self-recreating “progenitor” cells that are similar to stem cells and was also approved for use to treat blood cancers, some inherited metabolic and immune system disorders. It’s manufactured by the New York Blood Center, Inc., based in New York, NY.

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