ImmYOUnology

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The most promising treatments for ebola are based on basic immunology-part 2

One of the more peculiar, historic and almost cinematic treatments being discussed in the midst of the ebola crises is the use of blood transfusions. In movies, the blood of a survivor or someone special is often supposed to have some sort of mystical effect on the (usually villainous) recipient. It turns out, blood transfusions from people who have survived ebola are nearly as mystical.

It seemed obvious to me at first that the active components in blood transfusions from ebola survivors must be anti-ebola antibodies. Such antibodies would neutralize the virus and help the immune system clear it out. And in fact, a 1999 study reported that seven patients who survived the 1995 outbreak in the Democratic Republic of the Congo after receiving transfusions from survivors had anti-ebola antibodies circulating in their blood. One kind of antibody, called IgM, was absent in another patient who received a transfusion and died. This very small study seemed to indicate that transfusions could work against ebola and that antibodies are key to making them work. (You may be wondering why, if the transfusions worked, more patients weren’t treated this way during the 1995 outbreak. One important reason is that that blood cannot be transfused unless the donor and recipient blood types are compatible.  So the treatment is limited by the number of willing survivors and their blood types.)

This idea was challenged by a 2007 study done in a nonhuman primate species, rhesus macaques. For this study, researchers drew blood from a rare few monkeys who had survived an ebola infection four or five years earlier and a second “boost” infection 30 days earlier. They transferred the blood into other, recently-infected animals, and none of them survived…even those that made a lot of antibody. There are, of course, caveats. Monkeys are not humans, after all. It is possible that they fight the virus differently. And in the discussion of the paper, the researchers admit the experiments that had successfully transferred antibody-mediated immunity in guinea pigs had not worked in rhesus macaques.

There are also caveats to the human study though. The main one being that it can’t account for the better treatment transfusion patients received compared to other patients. The seven may have survived simply because they received better care in the clinics. The boost of cells, fluids, proteins and electrolytes that come along with blood transfusions may also have helped.

Scanning electron micrograph of red and white blood cells (National Cancer Institute)

Scanning electron micrograph of red and white blood cells (National Cancer Institute)

In spite of it all, the World Health Organization is behind blood transfusions and transfer of plasma from ebola survivors.  Plasma is the liquid part of blood that contains proteins like antibodies, along with other things like electrolytes and hormones. The first of the eight original transfusion patients, a 27 year-old nurse, was originally supposed to receive plasma and not whole blood. This was because, in 1978, a researcher who received plasma survived an ebola infection brought on by a finger prick in the lab. The nurses’ doctors settled for blood because they didn’t have the right tools to separate the plasma (a process called plasmapheresis).

The seven transfusion patients who followed the nurse ranged from a 54 year-old woman to a 12 year-old girl who caught the virus by kissing her newborn niece just days before the infant died. Antibodies seem to be the most likely explanation for the high rate of survival, but it is still not clear whether they were. Well-controlled human trials to determine whether blood transfusions work for ebola will probably never be possible. But, more and more people may be receiving them, so there may soon be more information about whether and how they work.


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The most promising ebola treatments are based on basic immunology

Though for many of us, the ebola crises is oceans away, the epidemic still weighs heavily on the hearts and minds of people all over the world. For some researchers, public health officials and drug developers, it is the driving force of all daily activity. Right now, there are two vaccines and eight treatments being developed or tested for their effectiveness against controlling infection or stopping the  virus’ spread. The most encouraging results have come from treatments that rely on a very basic aspect of immunology: antibodies neutralize viruses.

Antibodies are proteins made by immune cells called B cells. Each one of your millions of B cells is capable of producing antibodies specific for one thing, and when a B cell comes into contact with that one thing, it secretes lots of antibodies. The antibodies then tag invading pathogens, like viruses, to make other immune cells aware of the invader’s presence. If enough antibodies stick to a virus, they can cover it up, or neutralize it, and prevent it from infecting cells.

Ebola infection does trigger an antibody response, but for reasons that are still being studied, those antibodies are not usually enough to stop the virus before it spreads throughout the body. The concept behind ebola treatments like Zmapp, blood transfusions, vaccines and even supportive care, is to help the immune system outpace the growing virus.

 

Zmapp

Over the summer, this product was on headlines everywhere. Zmapp sparked a controversy over who should get the most cutting-edge treatments when it was given to two missionary doctors who flown to Atlanta for care.  Zmapp is not really a drug; it’s a combination of three kinds of antibodies that bind to the surface of ebola virus particles. Because each type was originally produced by one individual B cell they are called monoclonal antibodies. Monoclonal antibodies are used for treating cancer, autoimmune diseases and other infections.

Identifying the right monoclonal antibodies can be a painstaking and years-long process. Researchers collect B cells from a person or animal in the midst of an active immune response, in this case, against ebola. Then they seed individual antibody-making B cells into tiny wells on a cell culture dish. Later they test the culture media from each well for the presence of antibodies and select the cells making the best antibody to be “immortalized.” Cells are immortalized by altering their genes or fusing them with cancer cells that are already immortal. Usually the cells are frozen and stored for later use. They can be thawed anytime and grown to large quantities to make antibody.

Forget cigarettes...tobacco plants have lots of potential for "pharming" biological drugs like the monoclonal antibodies in Zmapp (From Wikemedia Commons)

Forget cigarettes…tobacco plants have lots of potential for “pharming” biological drugs like the monoclonal antibodies in Zmapp (From Wikemedia Commons)

It seems simple, but getting the process right can take years. The monoclonal antibodies in Zmapp were originally derived in mice back in 2000.

From there, the antibodies have to be purified. It can take liters and liters of cell media to purify enough antibody to treat one person one time. As a therapeutic, monoclonal antibodies are typically dosed over multiple treatments. In a recently published study showing the effectiveness of Zmapp against ebola infected monkeys, the animals were treated three to five times a day.

There are alternative ways to do this, however. Because antibodies are proteins they are coded by specific genes. So instead of fusing selected B cells with cancer cells, researchers could copy the gene coding for the cell’s antibody and put it into something else, like bacteria or, in the case of Zmapp, tobacco plants. Many biological products, like insulin have been produced in bacteria since the 1980s. Plant production of human proteins is a bit more recent.  The first human protein produced in plants in 2012 for a medical purpose was an enzyme injected into patients who can’t make it themselves. Some insulin is also now produced in plants.

Unlike cell-based or bacteria-based approaches, plants don’t have to be genetically manipulated and then grown up and harvested. Instead, adult plants are infected with viruses engineered to express the antibody-coding genes. The viruses introduce the genes, and the plants make the antibody. For some proteins, this results in much higher yields than cell-based methods. The monoclonal antibodies in Zmapp are being made by three different companies using a variety of these methods.

But there is a catch. When any kind of cell (plant, animal, bacteria) produces a protein, it adds little sugar labels to keep track of it during each stage of production. This process is called glycosylation. These glyo-labels vary by species and they can affect the way a protein functions. Because of this, the plants being used to grow Zmapp are not your run-of-mill tobacco. They are genetically modified so that they can give the anti-ebola antibodies more human-looking labels. That adds another layer of complexity to be addressed as these companies start to make large quantities of Zmapp.

It’s fascinating how this technology was developed step by step—often in obscurity—over the course of many decades.  Hopefully, it will be scaled up successfully in the coming months to provide more much-needed doses.

Next time…Blood transfusions.