Chemistry: analytical and immunological testing – For preexisting immune complex or auto-immune disease
Reexamination Certificate
1999-01-13
2001-11-20
Wortman, Donna C. (Department: 1648)
Chemistry: analytical and immunological testing
For preexisting immune complex or auto-immune disease
C436S513000, C436S518000, C435S007940
Reexamination Certificate
active
06319726
ABSTRACT:
SPECIFICATION
The invention relates to a method for detecting antibodies from body fluids by means of an immune reaction with tissue transglutaminase (tTG), the antigenic structures, immunoreactive sequences or analogues thereof, and with tTG-containing compounds, the antigenic structures, immunoreactive sequences or analogues thereof. The method may be used in the diagnosis and therapy control of diseases associated with an immune reaction against tTG, tTG-containing compounds, and antigenic structures, immunoreactive sequences or analogues thereof. Therefore, the invention is also directed to the use of tTG and the above-mentioned substances in diagnosis and therapy control, preferably in the diagnosis and therapy control of chronically inflammatory diseases or autoimmune diseases, and more preferably in the diagnosis and therapy control of sprue or coeliac disease. The invention is also directed to an oral pharmaceutical agent which includes tTG, tTG-containing compounds, the antigenic structures, immunoreactive sequences or analogues thereof as active ingredients and may be employed in the treatment of diseases accompanied by an immune reaction against these substances, because oral administration of the above-mentioned compounds results in an immune tolerance.
The present invention is based on the discovery that tissue transglutaminase (tTG, EC 2.3.2.13) is the autoantigen of sprue or coeliac disease.
On the basis of the above finding, the immunological method of the invention for detecting antibodies against tTG and tTG-containing compounds has been developed.
Coeliac disease is a disease of the small intestine mucosa, the first manifestation predominantly occurring during the late infant and toddler ages. If the corresponding clinical picture does not occur before the adult age, it is termed non-tropical sprue. Thus, both of these terms describe the same disease. Sprue is accompanied by an inflammatory change of the mucosa and a general malabsorption as a result thereof. In most of the cases, there is a morphological and clinical response to a treatment using a diet free of gluten.
Well-known as pathogenic factors are glutens from wheat, barley, rye and, to some extent, oats, while those from plant types having a lower degree of phylogenetic relation, such as corn, rice and soy are non-pathogenic. Amongst said glutens, the role of the pathogenic agent is ascribed to the alcohol-soluble prolamins, specifically &agr;-gliadin.
For this reason, sprue preferentially occurs in countries where wheat is used as major source of food (Europe, U.S.A, Australia) and has an incidence rate of 0.14/1,000 newborns in Denmark, 0.7/1,000 in Spain, 1/1,000 in Italy, 0.45/1000 in Germany, and 2.42/1,000 in Sweden, for example.
However, more recent investigations demonstrate that a subclinical pattern, i.e., a morphological change of the mucosa without massive symptoms is more widespread than believed so far. Thus, a study carried out in Italy in 1994 revealed an incidence of 3.28/1,000 among school children. The risk of latent sprue in the next of kin of sprue patients ranges up to 50%.
The predominantly latent sprue is frequently accompanied by a polymorphic dermatosis, i.e.,
Dermatitis herpetiformis,
where characteristic subepidermal small blisters with granular IgA deposits in the dermal papillae tips can be observed. Biopsies of the small intestine show an irregular, more or less severely damaged mucosa.
Another well-established association can be observed between sprue and insulin-dependent
Diabetes mellitus,
thyroid gland diseases, and a selective IgA deficiency.
In addition to numerous concomitant clinical symptoms of sprue, such as anemia which, among other things, has been ascribed to a vitamin B
12
malabsorption, and a vitamin K deficiency representing the reason for an increased hemorrhage tendency, the massively increased risk of gastrointestinal malignant tumors plays a special role. Up to 15% of the sprue patients, mostly at an age of more than 50 years, develop neoplastic diseases, about 50% of which involving intestinal T cell lymphomas and another 25% involving esophagus, oropharyngeal and small intestine tumors.
The therapy of sprue comprises strict observance of a lifelong gluten-free diet, where not only gluten-containing products made of wheat but also those made of rye, barley and oats must be excluded. As for the patients, this represents a grave restriction in both eating habits and social interactions.
If diagnosis and therapy of sprue are effected in time, there is a good prognosis. However, complications once having occurred are frequently not completely reversible. Conversely, if the disease remains unrecognized and untreated, severe symptoms may arise as a result of malabsorption. Ultimately, there is an increased risk of developing intestinal lymphomas and other gastrointestinal neoplasias.
At present, biopsy of the small intestine represents the top level standard in the diagnosis of sprue and the follow-up under gluten-free diet, but also non-invasive diagnostic methods based on immunological markers become more and more important. Because IgA and IgG class antibodies are present in the serums of sprue patients, which, on the one hand, are directed against gliadin and, on the other hand, against an autoantigen of the endomysium which is a special connective tissue which, among other things, contains the collagens I, III and V, elastic fibers, noncollagenic proteins such as fibronectin and proteoglycans, the serums may be tested for IgG and IgA antibodies against gliadin using ELISA, and for IgG and IgA antibodies against endomysium using indirect immunofluorescence. While antibodies against gliadin are not sufficiently specific for sprue, high sensitivity and specificity (97-100%) are reported for the IgA Ab against endomysium. However, esophagus sections from primates are required for the immunofluorescence detection. At present, attempts are made to detect the endomysium antibodies on umbilical cord material as well.
With a timely diagnosis and a strict observance of a gluten-free diet, the disease may be maintained in remission and thus, the increased risk of malignant tumors of the patients can also be lowered to a normal value. Therefore, there is great interest in developing a suitable detection assay for sprue. Because the group of individuals bearing a latent sprue also belong to the high-risk group, all of the individuals in question (especially, the next of kin), and ultimately, all the school children, as is presently taken into consideration in Italy, should be examined using a sensitive, specific, easily feasible, and low-cost assay.
To date, however, large-scale screening programs failed as a result of the following problems:
The invasive duodenal biopsies of symptom-free persons are unconscionable and exceedingly expensive.
An ELISA detection based on antibodies against gliadin is scarcely useful as a result of its poor specificity.
The immunofluorescence detection of IgA class endomysium antibodies, which is based on primate esophagus, is too expensive as a general screening method. Furthermore, the assessment is subjective and does not permit identification of sprue patients having an IgA deficiency (2% of the patients).
To date, therefore, a non-invasive, specific, quantitative, rapid, easily and inexpensively feasible detection assay for sprue/coeliac disease and therapy control thereof does not exist.
This problem is solved by the present invention. Based on the surprising finding that tissue transglutaminase (tTG, EC 2.3.2.13) is the autoantigen of sprue, an immunological method according to claims
1
through
6
for detecting antibodies against tTG and tTG-containing compounds from body fluids, particularly from serum was developed, which method not only permits diagnosing sprue or coeliac disease, but all diseases accompanied by an immune reaction against tTG, tTG-containing compounds, the antigenic structures, immunoreactive sequences or analogues thereof.
The tissue transglutaminase belongs to the class of transglutaminases. T
Dieterich Walburga
Ehnis Tobias
Schuppan Detlef
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