Specific depletion of alloreactive cytotoxic lymphocyte precursors

Abstract

THE major barrier to replacement therapy in various clinical immunodeficiency states is the likelihood that the transferred immunocompetent T lymphocytes would react against alloantigens of the recipient and produce graft-versus-host (GVH) disease¹. The logical resolution of this problem would be to delete or inactivate those clones of T cells which are specifically coded to react against the histocompatibility antigens of the recipient tissues. The remaining cell population could then fully reconstitute the recipient without danger of GVH reactions. Specific clonal deletion of this kind has been obtained in experimental animal situations by pre-circulation through a third-party host^2,3 or by induction of specific anti-idiotype immunity⁴. These studies show the validity of this approach but are technically complicated and have limited application. A simpler method of obtaining this result has been sought based on the following strategy. To reconstitute a strain A mouse by transfer of lymphocytes from an allogeneic strain B, the strain B lymphocytes are first incubated on a monolayer of cells of strain A. Those clones which recognise the strain A alloantigens should adhere to the monolayer; the non-adherent populations of cells should now be unable to react against alloantigens of strain A but should still respond to alloantigens of other haplotypes. Although this kind of technique has been widely effective in removing fully activated cytotoxic lymphocytes (T_c)^5–9, its use to deplete alloreactive T_c precursors has given variable results^8,10–16. Significant depletion of precursors has been reported primarily when the non-adherent cells have been assayed in vivo^13–16, but this has been claimed to be due to change in the responder cells, not depletion, as the same population assayed by in vitro generation of T_c shows no evidence of depletion¹⁶. We present here a simple depletion method developed from our finding that when splenic lymphocytes are incubated for a few hours before being formed into a monolayer, they can effectively absorb alloreactive T_c precursors even when assayed by the sensitive in vitro assays.