This invention relates to DNA sequences that code for the DR xcex2-chain locus of the human lymphocyte antigen complex. More particularly, it relates to the use of those DNA sequences in diagnostic typing processes and products. Such processes and products are useful in determining an individual""s susceptibility to a wide variety of diseases and an individual""s characteristics as a donor or acceptor of a tissue or organ transplant. The DNA sequences of this invention are also useful in the expression of polypeptides encoded by them.
The human lymphocyte antigen (xe2x80x9cHLAxe2x80x9d) system is the major histocompatibility complex in man. It, therefore, constitutes the strongest barrier for tissue and organ transplants between individuals, apparently distinguishing between self and non-self. In addition, HLA factors have been demonstrated to be associated with increased susceptibility to a wide variety of diseases. Therefore, the antigens of the HLA system have found use in diagnostic typing processes and products for determining an individual""s susceptibility to a wide variety of diseases and his characteristics as a donor or acceptor of a tissue or organ transplant [F. H. Bach and J. J. Van Rood, N. Engl. J. Med., 295, pp. 806-13 (1976)].
From a genetic point of view the HLA system is fairly well characterized. See e.g., L. P. Ryder et al., xe2x80x9cGenetics Of HLA Disease Associationxe2x80x9d, Ann. Rev. Genet., 15, pp. 169-87 (1981); J. L. Strominger et al., in The Role of the Major Histocompatibility Complex in Immunobiology, M. Dorf, ed., Garland SPTM Press, pp. 115-172 (1981); T. Sasazuki et al., xe2x80x9cThe Association Between Genes In The Major Histocompatibility Complex And Disease Susceptibilityxe2x80x9d, Ann. Rev. Med., 28, pp. 425-52 (1977). It consists of a series of more or less highly polymorphic loci situated within an interval of about 2 centimorgan (cM) on the short arm of chromosome 6. Three loci in that system (HLA-A, B and C) encode one class of codominantly expressed alloantigens (Class 1). Another locus (HLA-D/DR) encodes a second class of codominant alloantigens with a high degree of recognized polymorphism (Class 2). Three other loci, controlling some of the initial components (C2, C4 and factor Bf) of the complement cascade, also belong to the HLA system (Class 3). Finally, there is an non-specific region in the HLA complex designated Ia. Region Ia appears related to, but different than, the DR locus.
The biology of the HLA system is less well understood. Class 1 factors are distributed in all tissues except erythrocytes. Class 2 factors are substantially restricted to xcex2-lymphocytes and mononuclear phagocytic cells and the Class 3 complement factors are directly involved in the activation of the C3 factor, key component in the complement system. The HLA-DR antigens appear to be involved in immunological phenomenaxe2x80x94immune responsiveness, T-cell suppression, T-cell and xcex2-cell cooperation and T-cell and macrophage presentation [B. Benacerraf in xe2x80x9cThe Role Of The Major Histocompatibility Complex In Immunobiologyxe2x80x9d, M. E. Dorf, ed., Garland SPTM Press, pp. 255-69 (1981)].
The HLA-DR antigens are composed of two non-covalently-linked glycosylated peptide chains, a heavy or xcex1-chain of about 35000 molecular weight and a light or xcex2-chain of about 29000 molecular weight, that span the cellular membrane [Strominger et al., supra; and Ryder et al., supra]. Intracellularly, a third peptide chain of about 32000 molecular weight is associated with the xcex1- and xcex2-chains [D. J. Charron and H. O. McDevitt, J. Exp. Med., 152, pp. 180-365 (1980); Strominger, supra]. It appears that the light or xcex2-chain carries the polymorphism of the HLA-DR antigens, while the xcex1-chain and third chain appear identical in different individuals [G. Corte et al., Proc. Natl. Acad. Sci. USA, 78, pp. 534-38 (1981); Charron and McDevitt, supra]. Several serologically distinct HLA-DR antigens have been identifiedxe2x80x94HLA-DR1 through HLA-DR8xe2x80x94and monoclonal antibodies have defined subparts of DR antigens within homozygous cell lines [V. Quaranta et al., J. Immunol., 125, pp. 1421-25 (1980); S. Carrel et al., Mol. Immunol., 18, pp. 403-11 (1981)]. At least two DR xcex2-chains can also be distinguished in several homozygous cell lines by peptide analysis [R. S. Accolla et al., Proc. Natl. Acad. Sci. USA, 78, pp. 4549-51 (1981)].
Several other loci also exist that encode polymorphic Ia-like antigens that are closely linked but not identical, to HLA-DR [G. Corte et al., Nature, 292, pp. 357-60 (1981); Nadler et al., Nature, 290, pp. 591-93 (1981)]. These distinct subregions are called DC [R. Tosi et al., J. Exp. Med., 148, pp. 1592-1611 (1978); D. A. Shackelford et al., Proc. Natl. Acad. Sci. USA, 78, pp. 4566-70 (1981)] and SB [S. Shaw et al., J. Exp. Med., 156, pp. 731-43 (1982)]. The DC antigens are in strong linkage disequilibrium with the DR antigens. SB antigens control a secondary lymphocyte reaction and are encoded in a region centromeric to the DR loci.
At present the HLA-DR antigens are isolated serologically by precipitation with antisera. Therefore, the exact nature of the HLA-DR determinants is uncertain. However, these antigens have found use in typing processes and products to determine the compatibility of donors and acceptors for tissue or organ transplants and to determine susceptibility of an individual to a wide variety of diseases. For example, Ryder et al., supra, has reported the following disease susceptibilities based on DR1 through DR8 typing:
From these typings, it can be seen that an individual typed positive for D/DR4 has a 6.4 times higher risk of developing insulin-dependent diabetes than individuals typed negative for D/DR4.
In some cases it has also been demonstrated that a disease is more severe in patients having the disease-associated antigen than in those who do not have that antigen. For example, multiple sclerosis progresses more rapidly in D/DR2-positive patients than in D/DR2-negative patients. Moreover, relapses in certain diseases are more common in patients positive for the disease-associated antigens. Plainly, then, HLA-DR typing has great diagnostic and prognostic value.
However, the use of such typing processes and products and, therefore, the attainment of the important advantages that they would provide in identifying acceptable transplant donors and recipients and disease-susceptible individuals, has been severely restricted because the present typing procedure is complex and time consuming and because there are not sufficient HLA-DR antigens available to provide a useful and economical source for such processes and products.
The present invention solves the problems referred to by providing DNA sequences coding for the DR-xcex2-chains, the major polymorphic regions of the DR locus of the human lymphocyte antigen complex, and diagnostic typing processes and products related thereto.
By virtue of this invention, the DNA sequences encoding the HLA-DR light or xcex2-chains are now for the first time made available for use in HLA-DR typing processes and products. Not only are the DNA sequences of this invention able to be produced economically and in large amount, their use in typing processes and products substantially simplifies and reduces the cost of the former HLA-DR antigen-based typing processes and products. For example, the DNA typing process of this invention is simple, can be performed with as little as 10-20 ml of blood and can easily be scaled-up to several thousand typings.
Finally, the DNA sequences of this invention permit the expression of those sequences in appropriate hosts and the production of the specific DR xcex2-chain antigens encoded by them, uncontaminated by other HLA-DR factors, for use as diagnostic, preventive or therapeutic agents.