Dept. of Immunohaematology and Blood Bank, University Hospital, P.O. Box 9600, 2300 RC Leiden, The Netherlands.

Human bone marrow transplants performed as therapeutical treatment of severe aplastic anaemia, leukaemia and immune deficiency disease became available in the seventies (Bortin, 1970). In an artificial situation, such as organ transplantation, the major Histocompatibility (H) antigens function as a major transplantation barrier and thus play an important role in the survival of transplants and patients. Consequently, improved success in bone marrow transplantation was reported when matching for the HLA antigens was taken into account (Thomas, 1975). Between 1975 and the present day, the long-term results of allogeneic bone marrow transplantation (BMT) have greatly improved due to the use of HLA-matched siblings as marrow donors, advanced pretransplant chemoradiotherapy, the use of potent immunsuppressive drugs as Graft-versus-Host-Disease (GvHD) prophylaxis, better antibiotics and isolation procedures. The results of clinical In man, the efforts of several investigators have led to the identification of a small number of mHag (Goulmy, 1977,1983; Zier, 1983; Irle, 1990; Van Eis, 1992). Here, we will briefly focuss on their possible clinical relevance for BM transplantation in both the GvHD and the Graft-versus-Leukemia (GvL) reactivities. Furthermore, we provide preliminary evidence for the evolutionary conservation of human mHag.

Clinical relevance of anti-host cytotoxic T cells (CTLs) and helper T cells (Th) in the development of GvHD

Several reports demonstrated the presence of anti-host mHag specific CTL in patients suffering from GvHD after HLA genotypically identical BMT (Goulmy, 1983; Tsoi, 1980, 1983; Irle, 1985; Van Els, 1990; Irscheck, 1992; Niederwieser, 1993) .In our laboratory , much effort was put into the further characterization of a (small) number of anti-host mHag specific CTLs. Hereto, CTL clones specific for host mHag were isolated from the peripheral blood (PBL) of patients suffering from severe GvHD. Subsequent immunogenetic analyses revealed that these CTL clones identified five non-sexlinked mHag, designated HA-1, -2, -3, -4, -5, which are recognized in a classical MHC restricted fashion (Van Els, 1992) .mHag HA-3 is recognized in the presence of HLAA 1 and mHag HA-1 , -2, -4 and 5 require the presence of HLA-A2. In order to document the effect of mH antigens in genotypically identical BMT on the occurrence of acute (grade >= 2) GvHD, we prospectively collected PBL from HLA-A 1 and HLA-A2 positive patient/donor sibling pairs. This multi center study comprised 148 HLA genotypically identical BM donor/recipient combinations, adults as well as children, grafted between 1982 and 1990. The results of the mHag typing using the CTL clones specific for five well defined mHag HA-1 to HA-5 demonstrated (table 1) a significant correlation between mHag HA-1, -2, -4 and -5 mismatch and GvHD (Goulmy, 1994).

Table 1 Correlation of mHag HA-1. -2. -4 and -5. with the occurrence of GvHD

The last few years evidence has accumulated that in addition to CTLs mH antigen specific helper T cells (Th) could be relevant in the pathogenesis of GvHD. In vitro studies reporting on host directed Th cells have been described in patients having GvHD (Tsoi, 1980; Reinsmoen, 1984; Irle, 1987). Van Els et al. reported on the long term kinetics of Th cells in response to host mH antigens in 16 patients and demonstrated that significant Th cell activity in vitro correlates with clinical acute GvHD (Van Els, 1990b). In a subsequent study, we demonstrated that these anti-host Th cells carry the CD4 phenotype and recognize mH antigens in the context of HLA-DR and -DP (Van Els, 1990c). Most recent observations support the notion that mH antigen specific Th cells are by and large likely to playa role in the pathogenesis of acute GvHD (Theobald, 1992; Schwarer, 1993) .In both latter studies the primary in vitro putative mH antigen Th activities have been measured by IL-2 production of the responding cell population.

Possible involement of mHag in Graft versus Leukemia

The hypothesis that posttransplantation of bone marrow anti-host CTL activity may have a beneficial effect is based on the assumption of the postulated anti-Ieukaemic potential as a 'desired' side-effect of the post BMT complication GvH. (Bortin, 1973; Weiden, 1981 a, 1981 b). In search for anti-host CTL and Th cell activities post BMT, we observed earlier both absence and presence of anti-host CTL in patients without any clinical signs of GvHD (see table 2).

Table 2 Anti-host T cell activities after HLA identical BMT.

It was notable that in 3 out of 4 cases without anti-host CTL activity (see table 2), leukemia relapse was manifested. On the other hand, the presence of anti-host CTLs in the absence of GvHD (N-5) argues for the possible role of these CTLs in the antileukemic activity .In an attempt to study the post BMT anti-host CTL responses for their putative anti-leukemic activity in vitro, we observed "GvHD" related and "GvL" related acitivities. The latter type of CTL clones recognized patient's neoplastic cells only. The former type of CTL clones were reactive with ligands, like mH antigens, shared by host PBL and leukemic cells (Van Lochem, 1992). Another line of investigation support the notion that anti-host mHag specific CTL may playa role in the anti-leukemic effect of allogeneic BMT. Namely, mHag specific CTLs are capable of inhibiting in vitro outgrowth of clonogeneic leukemic precursor cells as well as lyse freshly obtained myeloid and lymphoid leukemic cells (Fakenburg, 1991; Van der Harst, 1994).

Human mHag are conserved in evolution

To substantiate the importance of the human mH antigenic systems, we investigated whether the mHag are conserved in evolution between man and chimpansee. Hereto, cells from chimpansees were transfected with the human HLA-A2.1 gene. Subsequent analyses with our human allo HLA-A2.1 and four mHag HLA-A2.1 restricted CTL clones revealed the presentation of chimpansees' allo and mHag peptides in the context of the transfected human HLA-A2.1 molecule by chimpansees' target cells (table 3) .These results implicate that the chimpansee cell derived allo and mHag peptides investiged in this study are very similar to the human allo HLA-A2 and HLAA2 restricted mHag peptides.

Table 3 Human mHag are evolutionary conserved

In conclusion, although lots of information was gathered during the past decades on the murine and human mHag, still many questions remain to be answered. Besides identification of the mHag and the genes they are encoded by, we must be able to dissect the majors from the minor minors. To achieve this, more information is needed on the Th and CTL defined human mHag repertoire, and to establish the immunodominant ones. To understand their biological role in bone marrow transplantation information on their cytokine secretion profile is essential.

Acknowledgements

I am indebted to Els Blokland, Ronald Bontrop, Cecile van Els, Fred Falkenburg, Ellen van Lochem and Jos Pool for their great contributions. Ingrid Curiel for typing the manuscript. This work was supported in part by grants from the Dutch Organisation for Scientific Research (NWO), the J.A. Cohen Institute for Radiopathology and Radiation Protection IRS) and the Dutch Cancer Society.

References

Beatty PG, Herve P (1989) Immunogenetic factors relevant to acute GvHD. In: S.J. Burakoff, D.H.J. Deeg, S. Ferrara, K. Atkinson (eds): Graft-versus-Host-Disease, Immunology, Pathophysiology and Treatment. New York, Dekker, 415-23.

Bortin MM (1970) A compendium of reported human bone marrow transplants. Transplantation, 9: 571-587.

Bortin MM, Rimm AA, Salzstein EC, Rodey GE (1973) Graft versus leukemia. III. Apparent independent anti-host and anti-leukemic activity of transplanted immunocompetent cells. Transplantation 16: 182-188.

Bortin MM, Horowitz MM, Ursic M, Rimm AA and Sobocinskym KA (1991 ). Progress in bone marrow transplantation for leukemia: a preliminary report from the advisory committee of the international Bone Marrow Transplant Registry. Transplant Proc. 23: 61-62.

Falkenburg F, Goselink H, van der Harst D et al (1991 ). Growth inhibition of clonogenic leukemic precursor cells by minor histocompatibility antigen-specific cytotoxic T lymphocytes. J. Exp. Med. 174: 27-33.

Goulmy E, Schipper R, Pool Jet al. Minor histocompatibility antigen mismatches influence the development of GvHD after HLA genotypically identical bone marrow transplantation. Manuscript subm. for publication 1994.

Goulmy E, Termijtelen A, Bradley BA, Van Rood JJ (1977). Y-antigen killing by T cells of women is restricted by HLA. Nature 266: 544-545.

Goulmy E, Gratama JW, Blokland E, Zwaan FE, van Rood JJ (1983) A Minor transplantation antigen detected by MHC restricted cytotoxic T lymphocytes during graft-versus-host-disease. Nature 302: 159-161.

Irle C, Beatty PG, Mickelson E, Thomas ED, Hansen JA (1985) Alloreactive T cell responses between HLA identical siblings. Transplantation 40: 329-333.

Irle C, Chapuis B, Jeannet Met al (1987) Transplant Proc. suppl. 1, 19: 2674.

Irscheck E, Hladik T, Niederwieser D et al (1992) Studies on the mechanism of tolerance or Graft-versus-Host Disease in allogeneic bone marrow recipients at the level of cytotoxic T cell precursor frequencies. Blood 79: 1622-1628.

Martin PJ (1991 ). Increased disparity for minor Histocompatibility antigens as a potential cause of increased GvHD risk in marrow transplantations from unrelated donors compared with related donors. Bone Marrow Transplantation. 8: 217-223.

Niederwieser D, Grassegger A, Auböck J, Herold M, Nachbaur D, Rosenmayr A, Gächter A, Nussbaumer W, Gaggl S, Ritter M and Huber C (1993) Correlation of minor histocompatibility antigen specific cytotoxic T lymphocytes with Graftversus-Host Disease status and analyses of tissue distribution of their target antigens. Blood, 81: 2200-2208.

Reinsmoen NL, Kersey JH, Bach FH (1984). Detection of HLA restricted anti minor histocompatibility antigen(s) reactive cells from skin GvHD lesions. Human Immunol. 1, 11: 249-257.

Schwarer AP, Jiang JZ, Barrett JM et al (1993). Helper T -lymphocyte precursor (HTLp) frequency predicts the occurrence and severity of acute GvHD and survival after allogeneic BMT in both recepients of genotypically HLA-identical sibling (SIB) and phenotypically HLA-matched unrelated donor (MUD) marrow. Lancet 341: 203-205.

Theobald M, Nierle T, Bunjes D et al (1992). Host-specific interleukin-2-secreting donor T cell precursors as predictors of acute Graft-versus-Host Disease in bone marrow transplantation between HLA-identical siblings. N. Engl. J. Med. 327: 1613-1617.

Tsoi M-S, Storb R, Dobbs S, Medill I, Thomas ED (1980). Cell mediated immunity to non-HLA antigens of the host by donor lymphocytes in patients with chronic graft-vs-host disease. J. Immunol. 125: 2258-2262.

Tsoi M-S, Storb R, Santos E, Thomas ED (1983) Anti-host cytotoxic cells in patients with acute graft-versus-host disease after HLA identical marrow grafting. Transplant Proc. 15: 1484-1486.

Van der Harst D, Goulmy E, Falkenburg JHF et al (1994). Recognition of minor histocompatibility antigens on lymphocytic and myeloid leukemic cells by cytotoxic T -cell clones. Blood 83: 1060-1 066.

Van Els C, Bakker A, Zwinderman AH, Zwaan FE, van Rood JJ, Goulmy E (1990a) Effector mechanisms in GvHD in response to minor Histocompatibility antigens. I. Absence of correlation with CTLs. Transplantation 50: 62-66.

Van Els CACM, Bakker A, Zwinderman AH, Zwaan FE, Van Rood JJ, Goulmy E (1990b) Effector mechanisms in GvHD in response to minor histocompatibility antigens. II: Evidence for a possible involvement of proliferative T cells. Transplantation 50: 67-71.

Van Els C, Zantvoort E, Jacobs Net al (1990c). Graft-versus-host disease associated T helper cell responses specific for minor histocompatibility antigens are mainly restricted by HLA-DR molecules. Bone Marrow Transplantation 5: 365-372.

Van Els C, D'Amaro J, Pool J, Bakker A, van den Elsen PJ et al (1992) Immunogenetics of human minor Histocompatibility antigens: their polymorphism and immunodominance. Immunogenetics 35: 161-165.

Van Lochem E, De Gast Band Goulmy E (1992) .In vitro separation of host specific graft-versus-host and graft-versus-Ieukemia cytotoxic T cell activities. Bone Marrow Transplantation 10: 181-183.

Weiden PL, Flournoy N, Sanders JE, Sullivan KM, Thomas ED (1981 a) Antileukemic effect of graft-versus-host disease contributes to improved survival after allogeneic marrow transplantation. Transplant. Proc. 18: 248-251.

Weiden PL, Sullivan KM, Flournoy N, Storb R, Thomas ED (1981 b) The Seattle marrow transplant team. Antileukemic effect of chronic graft-versus-host disease. N. Engl. J. Med. 304: 1529-1533.

Zier KS, Elkins WL, Pierson GR, Leo MM (1983) The use of cytotoxic T cell lines to detect the segregation of human minor alloantigen within families. Hum. Immunol. 7: 117-129.