quarta-feira, 10 de setembro de 2014

Plant-based research prevents complication of hemophilia treatment in mice

Date: September 4, 2014

Source: University of Pennsylvania

Summary:
A strategy to prevent one of the most serious complications of hemophilia treatment is underway at an American university. The approach, which uses plant cells to teach the immune system to tolerate rather than attack the clotting factor protein, offers hope for preventing one of the most serious complications of hemophilia treatment.

While healthy people have proteins in their blood called clotting factors that act quickly to plug wounds, hemophiliacs lack these proteins, making even minor bleeds difficult to stop.

The main treatment option for people with severe hemophilia is to receive regular infusions of clotting factor. But 20 to 30 percent of people who get these infusions develop antibodies, called inhibitors, against the clotting factor. Once these inhibitors develop, it can be very difficult to treat or prevent future bleeding episodes.

In a new study, researchers from the University of Pennsylvania School of Dental Medicine and the University of Florida College of Medicine teamed up to develop a strategy to prevent these antibodies from forming. Their approach, which uses plant cells to teach the immune system to tolerate rather than attack the clotting factor protein, offers hope for preventing one of the most serious complications of hemophilia treatment.

Henry Daniell, a professor in Penn Dental Medicine's departments of biochemistry and pathology and director of translational research, was the senior author on the research, which is featured on the cover of the journal Blood. Other Penn Dental Medicine authors included Jin Su and Shina Lin. Daniell's lab teamed with University of Florida researchers Roland Herzog, who completed postdoctoral training and held a faculty position at Penn; Alexandra Sherman; and Xiaomei Wang.

"The only current treatments for inhibitor formation cost $1 million and are risky for patients," Daniell said. "Our technique, which uses plant-based capsules, has the potential to be a cost-effective and safe alternative."

"This could potentially be a way to prevent antibodies from forming or to lower the incidence of it," Herzog said. "This is a major step forward."

Their study focused on hemophilia A, in which the clotting factor VIII, or FVIII, is mutated, causing a defect in clotting. Worldwide, one in 7,500 males are born with this disease. After receiving infusions of FVIII, some patients develop antibodies against it. Their immune system responds to this foreign protein as an invader that must be attacked and eliminated.

"These antibodies, in the hemophilia world, are known as inhibitors," Herzog said. "That is what patients are all scared of, because they render their standard therapy ineffective and inhibit the blood from clotting."

To prevent the immune system's attack on clotting factors, the researchers looked to previous studies that had found that exposing the immune system to individual components of the clotting factor protein could induce tolerance to the whole protein. FVIII is composed of a heavy chain and a light chain, with each containing three domains. For their study, the researchers used the whole heavy chain and the C2 domain of the light chain.

Daniell and colleagues have developed a platform for delivering drugs and bio-therapeutics that relies on genetically modifying plants so that they express the protein of interest. Trying that same method with the components of the FVIII molecule, they first fused the heavy chain DNA with DNA encoding a cholera toxin subunit, a protein that can cross the intestinal wall and enter the bloodstream, and did the same with the C2 DNA. They introduced the fused genes into tobacco chloroplasts, so that some plants expressed the heavy chain and cholera toxin proteins and others expressed the C2 and cholera toxin proteins. They then ground up the plant leaves and suspended them in a solution, mixing the heavy chain and C2 solutions together.

The researchers fed the mixed solution to mice with hemophilia A twice a week for two months and compared them to mice that were fed unmodified plant material. They then gave the mice infusions of FVIII, just as human hemophilia patients would receive. As expected, the control group of mice formed high levels of inhibitors. In contrast, the mice fed the experimental plant material formed much lower levels of inhibitors, on average seven times lower.

To find out how this was being accomplished, the research team looked for particular types of signaling molecules called cytokines, which send messages to the immune system's T cells. They found that mice that had been fed the experimental plants had more cytokines associated with suppressing or regulating immune responses, while mice in the control group had more cytokines associated with triggering an immune response.

By transferring subsets of regulatory T cells taken from the mice fed the experimental plants into normal mice, the team was able to suppress inhibitor formation, suggesting that the T cells were able to carry tolerance-inducing characteristics to the new population of animals.

"This gives us an explanation for the mechanism of how this tolerance is being created," Daniell said.

Finally, the researchers tried to reverse inhibitor formation. They fed the experimental plant material to mice that had already developed inhibitors. Compared to a control group, the mice given the FVIII-containing plant material had their inhibitor formation slow and then reverse, decreasing three- to seven-fold over two or three months of feeding.

This new treatment strategy holds promise for preventing and even reversing inhibitor formation in hemophiliacs receiving FVIII infusions. The researchers noted that their experiments showed that inhibitor levels could rise again as time passes, however.

"After some time, antibodies do develop if you stop giving them the plant material," Daniell said. "This is not a one-time treatment. You need to do it repetitively to maintain the tolerance."

Daniell, Herzog and the Penn Center for Innovation are now working with a pharmaceutical company to test this oral tolerance strategy in other animal species, with plans to begin human trials shortly thereafter.

"With multi-million dollar funding from a global pharmaceutical company and their decades of expertise in bringing numerous protein therapeutics to the clinic, we're excited to take lettuce capsules producing human blood-clotting factors to the clinic soon," Daniell said.

Story Source:

The above story is based on materials provided by University of Pennsylvania. Note: Materials may be edited for content and length.

Journal Reference:
A. Sherman, J. Su, S. Lin, X. Wang, R. W. Herzog, H. Daniell. Suppression of inhibitor formation against factor VIII in hemophilia A mice by oral delivery of antigens bioencapsulated in plant cells. Blood, 2014; DOI: 10.1182/blood-2013-10-528737

Cite This Page:

University of Pennsylvania. "Plant-based research prevents complication of hemophilia treatment in mice." ScienceDaily. ScienceDaily, 4 September 2014. <www.sciencedaily.com/releases/2014/09/140904131637.htm>.

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