INDUCTION OF TOLERANCE TO GAL a(1,3) GAL USING AN OCTAMERIC GAL a(1,3) GAL CONJUGATE, LJP920.

Mary-Ann Campbell, David S. Ramirez and Richard M. Jack.

La Jolla Pharmaceutical Company, San Diego.

Naturally occurring xenoreactive antibodies (Ab) specific for the gal a(1,3) gal epitope (agal) mediate both hyperacute and acute vascular rejection of xenografts in humans. The temporary removal of anti-agal Ab has been shown to protect against hyperacute rejection. Therefore we sought to develop a therapeutic that would specifically turn off (tolerize) for extended periods the B cells producing anti-agal Ab. This strategy is based upon a toleragen, LJP 394, which we are testing in phase II/III human clinical trials for anti-dsDNA Ab in lupus nephritis.

Mice bearing a homozygous deletion of the a1,3-galactosyl transferase gene (GalT KO mice) express anti-agal Ab. GalT KO mice were treated daily i.p. for 15 weeks with either PBS or LJP 920, a therapeutic compound consisting of eight copies of agal conjugated to an immunologically inert carrier. Levels of anti-agal Ab in serum were measured by ELISA. To measure anti-aGal specific B cell tolerance, ELISPOT was used to determine whether LJP 920 treatment decreased the number of agal-specific antibody-forming cells (AFC).

LJP 920 treatment significantly reduced the levels of both IgM and IgG anti-agal in GalT KO mice when compared with placebo-treated GalT KO mice (p<0.0001). Levels of anti-agal Ab in LJP 920 treated GalT KO mice were not significantly different from those in wild-type mice. LJP 920 also significantly reduced the levels of anti-agal Ab in GalT KO immunized with rabbit red cells (rRBC).

LJP 920 treatment resulted in a significant diminution of IgM secreting, agal-specific splenic AFC in GalT KO mice immunized with rRBC when compared with those treated with placebo (p=0.02). We were unable to detect any agal-specific AFC in peritoneal cells, neither were we able to detect IgG secreting, agal-specific AFC.

These results indicate that treatment with LJP 920 is highly effective at decreasing the level of both IgM and IgG anti-agal antibodies. That this reflects tolerance induced by the toleragen is supported by the fact that there is a significant decrease in agal-specific AFC following LJP 920 treatment.

Presented at the 5th Congress of the International Xenotransplantation Association, Oct 24-28, 1999, Nagoya, Japan, and IBC's 5th Annual Conference on Xenotransplantation, Dec. 6-7, 1999.