Inactivation or Down-Regulation of the Overstimulated, HLA-DR-(CD2R-, CD26-, CD38-, and/or CD69-) Positive T Cell Subset Enables the Immunocompetent Cells to Restore the Pre-Disease State

Summary

A common element of diseases as different as cancer, autoimmune disorders and chronic (retro)viral infections, including HIV-infection, is the overstimulation of some immunocyte subsets (HLA-DR⁺, CD38⁺ and/or CD69⁺-cells; ß2-m, sIL-2-R/CD25, sICAM-1, sCD8, neopterin, lysozyme, cathepsinD etc.). We tested two potential approaches to down-regulate the pathologically elevated CD8⁺ and HLA-DR⁺ T cells: (a) In animal model, we tested the sensibility of these, disease inducing and maintaining T cell subsets to in vitro pretreated (cell death preprogrammed) semi-syngeneic and allogeneic donor T cells, both in tumor-bearing mice and in adjuvant arthritis (AA)-induced rats, (b) In the first clinical study, we used a novel combination of FDA-approved drugs which inhibits Ca²⁺-influx and concomitantly down-regulates cytosolic cAMP in patient’s overstimulated immunocompetent cells. We could achieve a 94,6-100% long-term survival in tumor-bearing mice and a 100% prevention both of the AA-induction and of a 3fold AA-rechallenge. In patients, large primary tumors and large metastases shrank by 80-85% and small metastases disappeared completely. Since in HIV-infected persons, the increase of HLA-DR⁺ CD38⁺T (T8) Cells is associated with a fall in CD4-level and in development of AIDS, we are looking for the elimination of these HLA-DR⁺ targets by our novel technique in two AIDS-simulating (FIV/FeLV and SIV) animal models.

By the new technique (“microimmunosurgery”), we were able -for the first time- to show (a) a complete (100%!) tumor regression by a sole attack on tumor-protecting Ts cells (instead on tumor cells), (b) the high efficacy of the GvL-effect in solid tumors (GvT-effect), and (c) the separation of the GvL-effect from the GvH-effect.

These advantages were possible since the new approach is based on an immune reaction which is — on average — 10.000 times stronger than the reaction against a “classical” antigen (more precisely: the number of participating effector cells is 10.000 times higher than in the case of the “standard” antigen). This immunological reaction retains its efficacy even in a profoundly suppressed organism and is the only one working in vitro without a preceding in vivo antigen-priming of efffector cells.

At present, we are testing the efficacy of the novel approach in a murine leukemia model and comparing the results with those of the solid tumor model (“classical” GvL versus GvT effect).

In the planned clinical studies, the advantages of a general introduction of microimmunosurgery in the treatment of solid tumors, independently of tumor type and individuality, should be elucidated.

This report deals with a novel principle in the therapy of a series of diseases which are characterized by a pathologically hyperactivated T cell subset. This technique, which we call “microimmunosurgery,” is based on a selective depletion of the pathologically over-stimulated lymphocyte subpopulation by means of a combined in vivo treatment with monoclonal antibodies (Mabs) plus alloreactive donor effector cells whose lifespan was restricted (“preprogrammed”) by a patented (P 4324877.2, P 4320878.9 and PCT/EP89/00403) ex vivo pretreatment.

By this special treatment, the alloreactive donor T cells are able to eliminate — to some extent selectively — the disease inducing and maintaining T cell subset, but are hindered in establishing GvHD (graft-versus-host disease),due to the preprogrammed cell death. In this way, a high GvL (graft-versus-leukemia) effect without an accompanying GvHD can be achieved. We tested this new techniqe in detail in a murine tumor model.

The B16-melanoma cells (2.10⁶)which were syngeneic with respect to the C57B16-strain, were inoculated into (C57B16 x DBA2) Fl-mice (“recipients”).

In the first series of experiments, the primary tumor was excised 10 days after inoculation; 24 hours before this excision, 120 mg/kg cyclophosphamide (Cy) were injected i.v. and 1-3 hrs before tumor removal, the animals were injected with Mabs, directed either against mature (CD3-positive) T cells (anti-Thy 1.2.-Mabs) or against their T 8-subpopulation (anti-Lyt 2- Mabs) (Figure 1).

In the second series of experiments, the primary tumor was not removed surgically but only irradiated with 500 R four days after its inoculation (Figure 2).

The splenocytes of the C57B16-donor mice were preactivated in a tumor-specific or non-specific way. In addition, non-preactivated donor spleen cells were tested. In one sub-group, donor splenocytes were injected into the tumor-bearing recipients 4 days, and in the other sub-group 10 days after tumor excision.

In our system, the recipients (F1-hybrids) were tolerant to both, the tumor cells and the donor splenocytes, whereas the transfused donor spleen cells were alloreactive against the recipient but tolerant (syngeneic) with respect to tumor cells.

We compared this system with a second one, in which both, the donors and the tumor-bearing recipients belonged to the same strain (C57B16). The experimental conditions, including the tumor type (B16-melanoma) were identical in both systems.