br Acknowledgements br Introduction Diabetic complications

Acknowledgements
Introduction Diabetic complications are responsible for increased morbidity and mortality of diabetic patients. Increased flow of Phenyl sulfate through the polyol pathway under conditions of hyperglycemia contributes to the development of diabetic complications. Aldose reductase (ALR2), the first enzyme of the polyol pathway, thus represents an important target of a pharmacotherapy of pathologies related to glucose toxicity. Besides attenuating the activated polyol pathway under hyperglycemic conditions, inhibition of ALR2 was found to be beneficial in pathologies related to inflammation1, 2 which shifted searching for efficient aldose reductase inhibitors (ARIs) upwards to the top of health research issues. Thus in targeting long-term diabetic complications, as well as inflammatory pathologies, ARIs have been gaining increased attention. Concurrently, a multi-substrate specificity of ALR2 brought a new perspective in the designing of new ARIs. A variety of substrates may interact with the enzyme in multiple interactive modes, which offers a possibility of identifying ARIs, thus allowing to discriminate among different substrates.4, 5 In our previous study, tetrahydropyridoindoles carboxymethylated in position 8 (Fig. 1) were identified as aldose reductase inhibitors with mild efficacy and selectivity yet with significant antioxidant effect as an additional biological activity. Moreover, the compounds were found interesting from the point of their acidobasic behavior. The presence of abasicity center at the tertiary nitrogen, in addition to the acidic carboxylic function, predisposes these compounds to form double charged zwitterionic species, a characteristic which favorably affected their pH-lipophilicity profile. Substituted pyridoindoles represent an interesting group of compounds with aplethora of biological activities.8, 9, 10, 11, 12 We proceeded with optimization of the tetrahydropyridoindole scaffold by shifting the carboxymethyl pharmacophore from position 8 to position 5, with the aim to improve aldose reductase inhibitory efficacy and selectivity. Compounds from the group of 5-carboxymethyl-1,2,3,4-tetrahydro-1H-pyrido[4,3-b]indoles used to be characterized as antagonists of prostaglandin D2 (CRTH2) receptor13, 14, 15, 16, 17 and as modulators of cannabinoid (CB1) receptor. They have been patented as promising remedies to treat asthma, allergy, androgenic alopecia and pain. Structurally related fused tricyclic compounds comprising indole-1-acetic acid fragment were characterized as aldose reductase inhibitors.17, 19, 20 In the present study, an experimental set of 5-carboxymethyl-1,2,3,4-tetrahydro-1H-pyrido[4,3-b]indoles was created and their inhibitory potency against aldose reductase was tested. Selectivity in relation to the closely related rat kidney aldehyde reductase (ALR1) was determined. Structure−activity relationships supported by molecular docking simulations into the ALR2 binding site along with a computer-based physicochemical profiling of the compounds are discussed. For the most efficient compound, enzyme kinetics was assessed, along with its effect on sorbitol accumulation in isolated rat eye lenses incubated in the presence of high glucose as well as in selected organs in streptozotocin-induced diabetic rats in vivo.
Results and discussion
Conclusions In an effort to improve aldose reductase inhibitory efficacy and selectivity of 8-carboxymethylated tetrahydropyridoindoles we proceeded with optimization of the tetrahydropyridoindole scaffold by shifting the carboxymethyl pharmacophore from position 8 to position 5. Screening of commercial databases for the presence of tetrahydropyridoindoles carboxymethylated in position 5 yielded an experimental set of eight compounds of which five derivatives revealed markedly increased ALR2 inhibitory activity and selectivity compared to 8-substituted counterparts, reported previously. The most active and selective compounds, 3 and 4 respectively, were also potent inhibitors of sorbitol accumulation in rat lenses ex vivo. Moreover, in streptozotocin-induced diabetic rats, compound 3 administered by daily dose of 50 mg/kg/day intragastrically for five consecutive days significantly inhibited sorbitol accumulation in red blood cells and the sciatic nerve pointing to effective bioavailability of the compound. Molecular obesity indices predicted along with reasonable water solubility point to an excellent “drug-likeness” of compound 3, with prospects of further structure optimizations.