Recently competitive inhibitors of arginase have been develo

Recently, competitive inhibitors of arginase have been developed which have greater specificity for the enzyme. Initial development of these compounds involved determination of the crystal structure of arginase. Christianson and colleagues found that a binuclear manganese cluster was required for catalytic activity [92]. They also identified the full structures of human A1 and A2 [93]. Boronic rho inhibitor analogs of L-arginine [S-(2-boronoethyl)-L-cysteine (BEC) and 2(S)-amino-6-boronohexanoic acid (ABH)] are highly-selective arginase inhibitors. Both contain a boronic acid or N-hydroxyguanidinium head which binds to the manganese cluster, the active catalytic site in arginase [49]. Several inhibitors of arginase are commercially available, including nor-NOHA, BEC, and ABH. Although several specific arginase inhibitors have been developed, none of those currently available are isoform-selective. This is a significant limitation of the field in light of the growing evidence that arginase activity can be damaging in some contexts and protective in others. For example, the activity of A2-positive macrophages has been implicated in the development and progression of atherosclerosis, whereas A1-positive macrophages may promote plaque resolution 33, 34, 35, 36, 94. A1-positive macrophages are clearly involved in repair functions after tissue injury, whereas excessive A2 activity appears to be damaging in several CNS disease/injury conditions. In the absence of isoform-selective inhibitors, RNA interference has been used to specifically inhibit expression of A1 or 2. For example, administration of short hairpin RNA (shRNA) against A1 greatly reduced interleukin (IL)-13-induced airway hyper-responsiveness, with a concomitant decrease in A1 mRNA and protein levels in the lungs of mice [95]. Direct delivery of antisense A1 using an adenovirus vector into the corpus cavernosum improved erectile function in aged mice [96]. Antisense A1 also increased NO production in endothelial cells exposed to high glucose [12]. The search for more potent and isoform selective inhibitors is ongoing and involves structure-based design and plant extracts [38]. Studies using arginase inhibitors in humans with cardiovascular disease have so far been limited to small-scale clinical ‘proof-of-concept’ studies involving local administration of arginase inhibitors by cutaneous microdialysis or intra-arterial infusion. However, promising results have been reported in patients with coronary artery disease and type 2 diabetes 30, 97, heart failure [98], hypertension [99], or following resuscitation after cardiac arrest [100]. These observations suggest that inhibiting arginase activity could be beneficial in the treatment of cardiovascular disease. Systemic treatment with arginase inhibitors is used in the treatment of parasitic disease without significant adverse effects. However, further study is needed to demonstrate the safety of long-term treatment in patients with cardiovascular disease. Given the important role that A1 plays in detoxification of ammonia in the urea cycle, it will be important to make sure that the treatment does not suppress hepatic arginase activity enough to adversely impact on the urea cycle. Another potential limitation is that suppression of A1 may further limit tissue repair function which is already suppressed in many cardiovascular diseases.
Concluding remarks The role of excessive expression and activity of arginase and its downstream targets in cardiovascular dysfunction and injury has been well established by studies in both animal models and human disease conditions. Studies have also clearly demonstrated the involvement of these pathways in CNS disease and injury. Targeting specific components of the arginase/ornithine pathway holds great promise as a therapy for both cardiovascular and CNS diseases. Recent proof-of-concept studies in cardiovascular disease patients have shown beneficial effects of local delivery of arginase inhibitors. As has been noted above, the two arginase isoforms share 100% homology in areas crucial for enzyme function. Moreover, both have been shown to be fundamentally involved in dysregulation of NOS function. However, the two isoforms are encoded by different genes, localized to different intracellular compartments, expressed in different cell types and tissues, and are involved in different cellular functions. Analyses using knockout mice and specific gene knockdown have been informative about the differential involvement of increases in A1 and A2 in various disease conditions. However, there is also considerable evidence to support a positive impact of arginase on cell growth, collagen synthesis, and neuronal development during physiological conditions as well as during tissue repair following injury. Thus, there is some risk associated with global inhibition of arginase activity. Studies are beginning to examine the subcellular and molecular regulation of arginase activity, and this information should facilitate design of new approaches to specifically limit its pathological effects. However, there are many questions that need to be answered to better understand the specific role of the two arginase isoforms and their downstream targets in both health and disease and to identify better therapies (Box 3).