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Tolerance Induction Research Shows Success

Exciting news infused the room Monday afternoon as attendees of “Medical: Innovating Immune Tolerance Induction for Hemophilia” learned what is on the horizon for tolerance induction. Shannon Meeks, Atlanta, Georgia, USA; Roland Herzog, Gainesville, Florida, USA and Sebastien Lacroix-Desmazes, Paris, France, comprised a panel presenting their research in this field.

Meeks provided a quick overview of the immune response to FVIII. “Most FVIII is released into the circulation as a set of heterodimeric proteins and is made up of a heavy and a light chain,” said Meeks. “VWF inhibits FVIII binding to phospholipids.”

She reminded the audience that most inhibitors—IgG antibodies directed against the missing FVIII—are developed in the first 10 to 20 exposure days. In those with severe hemophilia A, 20 percent to 30 percent develop inhibitors. In research with mice, she noted the following conclusions:

  • There is no statistical difference in titers between inactive construct and FVIII
  • Thrombin generated by FVIII is not necessary for immune response
  • FVIII thrombin generation does not account for immunogenicity of FVIII
  • Patients with point mutations have more limited spectrum of potential immunogenicity peptides

“More research is needed on if it is possible to engineer less immunogenic FVIII and also on the impact of whether the VWF presence or absence makes a difference,” Shannon Meeks concluded.

Based on the premise that we tend to be tolerant to things we eat and that we activate regulatory T cells in this manner, Herzog introduced the concept of oral tolerance induction using transgenic plants. Traditionally oral agents were hampered by cost and may no longer be useful by the time they get to the small intestine. “Plants have cell walls that protect the proteins inside,” said Herzog.

The first plant-based oral tolerance experiment for hemophilia involved tobacco plants and mice.  The results showed suppressed inhibitor formation against FIX in hemophilia B mice. “This also protected them from anaphylaxis,” said Herzog. “While our control mice were dying, the ones eating the infused plants were protected.” This works as the plant cell protects the protein until it gets to the small intestine and then the antigen is released. The epithelial cells in the small intestine then deliver it to the immune system.

Since Herzog and his team felt it might not be ethical to feed tobacco plants to children, they concentrated their next efforts on lettuce. “Lettuce appears to remain stable,” he stated.

The research was then scaled up to include dogs. “Of course we had to trick the dogs into eating lettuce and we wondered if dogs would have the enzymes in their guts they needed. Not only was the oral administration of chloroplast transgenic leaf cells expressing CTB fusion proteins efficient in prevention, and to some extent reversal of antibody responses to FVIII and fix in hemophilic mice, the first successful oral tolerance studies in hemophilic dogs were carried out indicating that the approach is applicable to the nonrodent immune system.”

Lacroix-Desmazes addressed tolerance induction through transplacental delivery. “We know that maternal IgG is transferred in the third trimester, so that the infant has some protection when born,” he said. “We wanted to see if we can transfer FVIII to induce tolerance in utero when we know the baby is to be born with hemophilia A.”

In certain experimental mice models transplacental transfer has been successful with specific T cell receptors. “There are issues, however,” he said. “Would mothers-to-be of hemophilic babies accept this treatment? Would the VWF protein be too large to cross the placenta? And would humans behave as mice in coagulation and immunogenicity?”

In order to answer these questions, more research in this area will need to be done.