Method for treating or preventing conditions by elevation of the adenosine level in an individual

The present application is a divisional of U.S. application Ser. No. 10/546,653, filed 27 May 2009, which in turn is a national stage filing under 35 U.S.C. §371 of PCT/FI2004/000158, filed on 19 Mar. 2004 which is a non-provisional of 60/515,425, filed 30 Oct. 2003, from which priority is claimed under 35 U.S.C. § 119(e). The present application also claims priority to Finland patent application Serial No. 20030467 filed on 28 Mar. 2003.

This invention relates to methods for inducing elevated adenosine levels in an individual and to treatment or prevention of diseases or disorders benefiting from an elevated adenosine level. The invention also concerns a method of up-regulating endothelial CD73 expression in an individual and to treatment or prevention of diseases or disorders benefiting from such CD73 up-regulation.

The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated by reference.

The interaction between lymphocytes and endothelial cells is a multi-step process. To be able to penetrate the vessel wall and to reach the target site, circulating cells use a very finely regulated set of adhesion molecules. Enhanced adhesion to endothelium and subsequent transmigration of re-circulating leukocytes through the endothelial lining of vessel wall into the tissue is characteristic for inflammation. Moreover, the release of pro- and anti-inflammatory cytokines in a high extent takes place at sites of inflammation. Those cytokines are potent regulators of the expression of adhesion molecules.

CD73 (ecto-5′-nucleotidase) is a 70-kD glycosyl-phosphatidyl-inositol-anchored cell surface molecule with ecto-enzymatic activity. It is abundantly expressed on the vascular endothelium and at a low level on certain subpopulations of human lymphocytes. It is part of the purine salvage pathway by degrading nucleoside-5′-monophosphates (AMP and IMP) into nucleotides like adenosine and inosine (1).

Adenosine, a purine nucleoside product of the CD73 enzyme activity, binds to specific receptors on the cell surface. Adenosine is reported to have a role in many physiological and pathological events. So far four different subtypes of G protein-coupled adenosine receptors A1R, A2aR, A2bR and A3R have been cloned. Due to the diversity of the receptors and their abundant localization in different tissues, adenosine-adenosine receptor interaction leads to various physiological responses. Adenosine, by binding to A1 and A2 receptors, regulates pathological consequences of inflammation by controlling leukocyte binding to endothelium and acts as an anti-inflammatory agent by binding to A2 and A3 receptors, through the inhibition of neutrophil degranulation (2). Adenosine also decreases eosinophil migration through activation of A3 receptor. This promoting effect of 5′-AMP is CD73 mediated and is followed by an increase in intracellular cAMP. Recently, a critical role for A2a receptor has been shown in decreasing systemic and tissue specific inflammatory responses in vivo.

Adenosine prevents cell damage during heart and central nervous ischemia (3-5). After hypoxia ecto-5′-nucleotidase activity increases due to phenomenon known as preconditioning. This results in release of large amounts of adenosine leading to increased resistance of cells to infarction for example in cardiac hypoxia.

So far, practically nothing is known about the regulation of endothelial CD73 expression and function. However, in inflammation there may be some inducers secreted which in vivo specifically control endothelial CD73 expression.

As adenosine, having an anti-inflammatory and cell protective effect, plays an important role in controlling the extent and consequences of inflammation, this work was designed to identify factors responsible for the regulation of CD73 expression as well as ecto-5′-nucleotidase mediated adenosine production.

Adenosine as such could be administered to patients suffering from inflammatory conditions or conditions that untreated most likely would lead to tissue inflammation. However, a serious drawback by direct administration of adenosine is the rapid elimination of adenosine in vivo. Therefore, this work offers a new way to achieve elevated levels of adenosine over a prolonged time.

The main object of the present invention is to provide a method for increase of the adenosine level in an individual and for maintaining the elevated adenosine level over a prolonged period of time and thereby prevent or treat inflammatory conditions or conditions that untreated would lead to tissue inflammation.

Another object of this invention is to provide a method for up-regulating the expression of endothelial CD73 in an individual and for preventing or treating diseases or disorders benefiting from such CD73 up-regulation, that necessary not would require an elevated level of adenosine.

Thus, according to one aspect, this invention concerns a method of inducing an elevated level of adenosine in an individual by administering to said individual an effective amount of either i) recombinant protein CD73 or ii) a cytokine or another factor being capable of inducing endothelial CD73 expression, or a combination thereof.

According to another aspect, the invention concerns a method for prevention or treatment of a disease or disorder requiring or benefiting from the elevation of the adenosine level in an individual, by administering to said individual an effective amount of either i) recombinant protein CD73 or ii) a cytokine or another factor being capable of inducing endothelial CD73 expression, or a combination thereof.

According to a third aspect, the invention concerns a method of up-regulating endothelial CD73 expression in an individual by administering to said individual an effective amount of a cytokine or another factor being capable of inducing endothelial CD73 expression.

According to a fourth aspect, the invention concerns a method for prevention or treatment of a disease or disorder requiring or benefiting from up-regulating endothelial CD73 expression in an individual, by administering to said individual an effective amount of a cytokine or another factor being capable of inducing endothelial CD73 expression.