Duke University School of Medicine researchers have developed a way to grow large quantities of regulatory B cells.1 The team used an infusion of Breg cells to suppress a murine form of multiple sclerosis and now wants to test the immunomodulatory cells in other autoimmune diseases. They are hoping to launch a company based on the technology.

Breg cells produce the anti-inflammatory cytokine IL-10 and can counteract excessive immune activation in autoimmune and inflammatory diseases.2

In previous studies, a team led by Thomas Tedder, professor of research in immunology at Duke, engineered mice deficient in Breg cells and showed that the animals were more susceptible to autoimmune and inflammatory diseases than wild-type animals.3 However, because Breg cells constitute only a tiny fraction of circulating B cells, it was not possible to harvest enough cells to test whether infusing Breg-deficient animals with Breg cells would have a beneficial effect.

The new work by Tedder's team solves the Breg cell supply problem. The researchers determined the conditions that promote the development of Breg cells in the spleens of mice and reproduced those conditions ex vivo to grow large numbers of the cells.

"We can now identify subsets of Breg cell precursors, purify that subset and adoptively transfer it" to mice unable to make their own Breg cells, said Tedder. "If you want to treat autoimmune disease, being able to expand these cells is an advance."

Isolate and activate

The team began by purifying general B cell precursors from the spleens of mice and treating them in vitro with a range of cytokines suspected to influence Breg cell development.

B cell precursors activated with IL-21 secreted IL-10, whereas controls treated with media or with other cytokines did not. When transferred into Breg-deficient mice, the in vitro-activated cells decreased symptoms of experimental autoimmune encephalitis (EAE) compared with unactivated B cell precursors.

Because B cell activation typically is enhanced by T helper cells, the natural source of IL-21, the team suspected that other T cell molecules might further promote Breg cell activation and growth. Indeed, IL-21 activation of Breg cells was further increased by CD40, a B cell co-receptor that binds to CD40 ligand (CD40LG; CD40L; CD154), found on the surface of T cells.

Using this information, the researchers developed a streamlined procedure for growing Breg cells in vitro. They transfected a fibroblast cell line with CD154 and BLyS (BAFF), a general B cell-activating protein, and then cocultured B cell precursors with these fibroblasts in the presence of IL-21.

The resulting number of activated Breg cells was about four million times the initial number of inactive cells. Reinfusion of those cells prevented the initial onset of EAE and decreased disease severity in mice that already had developed the condition compared with no treatment.

Results were published in Nature.

"The novel finding here is the ability to expand murine regulatory B cells in cell culture," said Abdolmohamad Rostami, professor of neurology at Thomas Jefferson University. "The next step is to extend this observation into humans-they should take whole B cells and treat them with human IL-21."

Breg-adoon

Tedder thinks infusions of large numbers of patient-derived, ex vivo-
activated Breg cells could help to rapidly shut down autoimmune diseases.

He said Breg cells specific for autoimmunity-associated antigens already are present in patients with autoimmune disease but that there are too few of them to affect disease. The hope is that boosting overall levels of Breg cells would tip the balance of immune activity toward suppression of autoimmunity.

"Nature has already created a set of Breg cells with immunomodulatory activity," said Tedder. "If you have an autoimmune disease, these cells are already responding. Our job is just to expand them."

Tedder noted that because Breg cells act in an antigen-specific manner, there is little danger of the broad immunosuppression that is a concern with immunomodulatory therapies.

The team "did show that the mice were not immunosuppressed, which is a major problem with many medications for autoimmune disease," said Rostami. "With conventional immunosuppression, there is a risk that you would suppress the suppressor cells, too."

The challenge, Tedder said, is to grow enough of the human cells in vitro to achieve the kind of disease-suppressing effects seen in mice. Tedder said his laboratory is now working on reproducing and scaling up the Breg cell expansion procedure with human cells, which grow more slowly than mouse cells.

Both Tedder and Rostami said the technique will allow researchers to compare Breg cells head-to-head with Treg cells, another type of immumodulatory cell.

Last year, a trio of academic teams reported procedures for amplifying and reinfusing human Treg cells in humanized mouse models of autoimmune disease.4-7

Tedder thinks Treg cells might be best suited for chronic inflammatory conditions, whereas Breg cells appear to be most effective in acute conditions.

His technique is patented and is available for licensing. Tedder said he is hoping to launch a company based on the technology and is in the process of finding investors and commercial development partners.

Osherovich, L. SciBX 5(42); doi:10.1038/scibx.2012.1103
Published online Oct. 25, 2012

REFERENCES

1.   Yoshizaki, A. et al. Nature; published online Oct. 14, 2012; doi:10.1038/nature11501
Contact: Thomas F. Tedder, Duke University School of Medicine, Durham, N.C.
e-mail: thomas.tedder@duke.edu

2.   Osherovich, L. SciBX 4(24); doi:10.1038/scibx.2011.678

3.   Yanaba, K. et al. Immunity 28, 639-650 (2008)

4.   Saggoo, P. et al. Sci. Transl. Med. 3, 83ra42 (2011)

5.   Feng, G. et al. Sci. Transl. Med. 3, 83ra40 (2011)

6.   Hippen, K.L. et al. Sci. Transl. Med. 3, 83ra41 (2011)

7.   Osherovich, L. SciBX 4(23); doi:10.1038/scibx.2011.646

COMPANIES AND INSTITUTIONS MENTIONED

Duke University School of Medicine, Durham, N.C.

Thomas Jefferson University, Philadelphia, Pa.