Eurasian J Med 2019; 51(3): 285-92 Review

Alagebrium and Complications of Diabetes Mellitus

Cigdem Toprak , Semra Yigitaslan

ABSTRACT Glycation is the process of linking a sugar and free amino groups of proteins. Cross-linking of glycation products to proteins results in the formation of cross-linked proteins that inhibit the normal functioning of the cell. Advanced glycation end products (AGEs) are risk molecules for the cell aging process. These ends products are increasingly synthesized in diabetes and are essentially responsible for diabetic complications. They accumulate in the extracellular matrix and bind to receptors (receptor of AGE [RAGE]) to generate oxidative stress and inflammation. particularly in the cardiovascular system. Treatment methods targeting the AGE system may be of clinical importance in reducing and preventing the complications induced by AGEs in diabetes and old age. The AGE cross-link breaker alagebrium (a thiazolium derivative) is the most studied anti-AGE compound in the clinical field. Phase III clinical studies with alagebrium have been successfully conducted, and this molecule has positive effects on cardiovascular hypertrophy, diabetes, hypertension, vas- cular sclerotic pathologies, and similar processes. However, the mechanism is still not fully understood. The primary mechanism is that alagebrium removes newly formed AGEs by chemically separating α-dicarbonyl carbon–carbon bonds formed in cross-linked structures. However, it is also reported that alagebrium is a methylglyoxal effective inhibitor. It is not yet clear whether alagebrium inhibits copper-catalyzed ascorbic acid oxidation through metal chelation or destruction of the AGEs. It is not known whether alagebrium has a direct association with RAGEs. The safety profile is favorably in humans, and studies have been terminated due to financial insufficiency and inability to license as a drug. Keywords: Alagebrium, cross-link breaker, diabetes, diabetic complications

Mechanism of Glycation The first step of the glycation reaction involves a reducing sugar with the carbonyl group and the side chain ε-amino group in proteins. The Schiff base intermediate product is formed within hours, which is then converted into Amadori products within days. Cite this article as: Toprak C, Yigitaslan S. Alagebrium and Complications of Diabetes Mellitus. Eurasian J Med 2019; 51(3): 285-92. The Amadori products are then converted to dicarbonyl compounds and finally to advanced gly- ORCID IDs of the authors: cation end products (AGEs) within weeks. While the part of the formation of Amadori products C.T. 0000-0003-3718-5195 is recyclable, the later stages are irreversible (Figure 1). S.Y. 0000-0001-6722-2394 Department of Medical Pharmacology, Eskisehir Osmangazi University, School of Medicine, In a healthy state, AGEs are normally produced in a slow manner, but they are rapidly produced Eskisehir, Turkey and accumulated in hyperglycemia, diabetes, atherosclerosis, hyperlipidemia, inflammation, renal failure, and neurodegenerative diseases such as Alzheimer’s disease. AGEs contribute to the Received: November 20, 2018 Accepted: April 16, 2019 aging process because most of their primary effects are seen on long-lasting proteins such as collagen and lens crystals [1, 2]. Correspondence to: Cigdem Toprak E-mail: [email protected] AGE Pathophysiology and Mechanism of Action DOI 10.5152/eurasianjmed.2019.18434 AGE damages the cells and tissues through various mechanisms; the intracellular glycation of proteins leads to impaired cell function. Circulating AGE binds to cellular receptors and leads to the activation of the signal transduction cascade and alteration in gene expression. The conse- Content of this journal is licensed under a Creative quent accumulation of AGE in the extracellular matrix results in cross-linking of the proteins and Commons Attribution 4.0 International License. decreases the distensibility in arteries [3]. 286 • Toprak and Yigitaslan. Alagebrium and Advanced Glycation End Products Eurasian J Med 2019; 51(3): 285-92

AGE Accumulation in the Extracellular Matrix by increasing age-related arterial stiffness. AGE Chemical Properties of Alagebrium The AGE–RAGE activation increases the trans- formation begins with the classical Maillard re- Since the discovery of N-Phenacylthiazolium forming growth factor-β (TGF-β) level and action pathway, which is a non-enzymatic reac- bromide, one of the almost completely studied matrix metalloproteinase-2 (MMP-2) activity; tion of lysyl residues of proteins with reduc- analogs is 4,5-dimethyl-3-phenacylthiazolium however, RAGE signaling modulates MMP-9 ac- ing sugars. Collagen and elastin are the main chloride (ALT-711®, Alagebrium chloride). Al- tivity, which determines the changes in colla- proteins on the arterial wall targeted by this agebrium (Synvista Therapeutics) is a new gen- gen-4 transformation [4]. reaction [13]. eration of cross-link breaker and a thiazolium compound (Figure 2) [18]. The extracellular matrix is ​​highly sensitive to gly- Glycation induces AGE formation and cross- cation because of its slow conversion rate, and linking for collagen, which leads to an increase This compound has enzymatic properties that AGE accumulation is responsible for cross-link- in the stiffness of the vessel wall. It is still un- breakdown covalent bonds formed in cross- ing. The subsequent mechanical changes include clear whether AGEs and cross-links occur in all linked proteins, which allows the release of the reduced elasticity, increased vessel myocardial major glycation sites or in preferential regions of protein and thereby maintain its normal func- stiffness and thickness, decreased vessel lumen, the collagen. However, similar to ribonuclease, tion. It has been suggested that it non-enzy- and development of glomerular sclerosis and glucosepane cross-links have been also found in matically breaks down the cross-linked AGEs atherosclerosis [5-9]. the lysine and arginine domains in the globular formed on long-lived proteins such as collagen model proteins [14]. and elastin. Importance of Metal-catalyzed Oxidation in AGE Formation Compared to collagen, elastin has a low num- Its molecular weight (MA) is 232.32, and the

Trace levels of redox active transition metals, ber of glycation sites due to low lysine content. chemical formula is C13H14NOS. such as copper(Cu2+), iron (Fe2+), manganese Importantly, elastin glycation has serious con- (Mn2+), and zinc (Zn2+), are mandatory to main- sequences for optimal functioning of the car- The AGE cross-link breaker and methyl glyoxal tain biological reactions, but they are cytotoxic diovascular system during aging and diabetes. inhibitor alagebrium (ALT-711) was developed when present in excess and result in cellular However, data are scarce on the relationship by Alteon Corporation. dysfunction via oxidative stress and free radical between AGE and elastin cross-linking. production (reactive oxygen products). In addi- Alagebrium is a molecule in the Phase III clini- tion, these metals can catalyze carbonyl forma- Therefore, AGE formation and cross-linking cal trials, which showed positive effects in the tion in proteins. Furthermore, the inhibition of are very important in systemic hypertension in treatment of diabetes, hypertension, vascular AGE formation reduces arterial stiffness and the which elastin synthesis is stimulated, known as sclerotic pathologies, cardiovascular hypertro- ability to chelate metals [10, 11]. tropoelastin and contains a high amount of ly- phy, and similar diseases. In early studies, after sine residues [15]. 3 weeks of treatment, diabetic rats showed im- Particularly, for connective tissue, the ability of proved vascular function, decreased IgG binding metals to directly oxidize proteins is mediated According to Santa et al., elastin is prone to the to red blood cells, and decreased tail-induced by a hydroxyl radical, and a metal catalyst oxida- Fenton reaction, whereas it has been shown to collagen cross-linking and ischemia [19]. tion (MCO) reaction results in the deamination cause a vicious cycle of CML, including protein of carbonyl, α-aminoadipic acid δ-semialdehyde aging with glycation, producing CML formation, In humans, this compound exhibits a good safety lysine residue known as alizine. The physiologi- binding to redox active metals, such as Cu+2 and profile; however, the studies have been discon- cal cross-linking of collagen and elastin by lysis Fe+3, and resulting in the formation of the CML- tinued by Alteon Corporation at the beginning oxidase is also similar. Other mechanisms for metal complex, and inducing lipid peroxidation of 2009 due to financial difficulties, and it could alizine formation could be α-dicarbonyl sugar (providing CML formation by glyoxal) [16]. not be licensed as a drug [20, 21]. MCO-mediated Strecker degradation of lysine residues [12]. In addition, myeloperoxidase mediates the for- The clinical trial results indicate that this com- mation of CML through oxidation of serine and pound might be the first-line specific treatment Mechanism of Protein Damage by Glycation production of the CML precursor glycolaldehyde for cardiovascular disorders caused by AGEs on the Arterial Wall at the inflammation sites. The reactive oxygen [20, 21]. When the total carbonyl stress increases in a species (ROS) produced during this cycle induces carbonyl-rich environment, such as diabetes elastolysis in the arterial wall. Moreover, glycated and end-stage renal disease, glycation and AGE elastin and collagen show greater affinity for ions formation are known to show a theory effect of metals such as copper and iron [17].

Figure 1. AGE formation. Figure 2. Alagebrium chemical structure. Eurasian J Med 2019; 51(3): 285-92 Toprak and Yigitaslan. Alagebrium and Advanced Glycation End Products • 287

Mechanism of Action of Alagebrium Clinical Studies sumption (VO2), was measured. Compared with The mechanism of action of alagebrium re- In several clinical studies sponsored by Syn- baseline, alagebrium reduced left ventricular mass mains controversial. Similar to PTB, initially, vista Therapeutics, between 2002 and 2010 and corrected diastolic function but did not alter alagebrium primarily results in the chemical (DIAMOND [NCT00043836], SAPPHIRE blood pressure, VO2, or aortic distensibility [35]. cleavage of α-dicarbonyl carbon–carbon bonds [NCT00045981], SILVER [NCT00045994], in cross-linked structures, thereby removing SPECTRA [NCT00089713], BREAK- The effect of 8-week alagebrium treatment on newly formed AGEs. A recent study has sup- DHF-I [NCT00661611], and BENEFICIAL vascular inflammation, inflammation, and col- ported that alagebrium is an effective inhibitor [NCT00516646]), alagebrium has been exten- lagen turnover was assessed in a double-blind of MG, reducing α-dicarbonyl compounds [22, sively studied. study on individuals with systolic hypertension. 23]. Alagebrium improved vascular function and in- Data have been published, and other studies versely correlated with collagen turnover and Zinc, iron, and copper are required for normal were terminated early due to the financial in- serum markers of inflammation [36]. function integrity but are abundantly present in sufficiency. In 2001, the data of 93 hypertensive diabetes and the related complications. Alage- patients randomized to receive placebo or al- In the BENEFICIAL study, diabetic and non-di- brium is a potent inhibitor of copper-catalyzed agebrium were recorded in a pilot study. After abetic heart failure patients were treated with ascorbic acid oxidation. Although alagebrium 2 weeks of treatment, the patients in the alage- placebo or alagebrium to assess their aerobic exerts an antioxidant capacity in preclinical ani- brium group had decreased arterial pulse rate capacity and diastolic/systolic function after 36 mal models and the in vitro hydrolyzed product with increased vessel compliance [8]. weeks. The primary (aerobic capacity) and sec- shows a strong chelating activity, it is unclear ondary (diastolic/systolic function) results were whether it is a direct or indirect effect. There- A 2-year toxicity study of alagebrium on rats unchanged. The analysis of diabetes subgroups fore, it is difficult to distinguish whether the ef- showed liver alterations. Since the data were also revealed no significant interaction [37]. fect of alagebrium is through metal chelation or published in 2004, Synvista Therapeutics Inc., AGEs destruction. [24-29]. which synthesized alagebrium, did not include In 4 clinical studies (ClinicalTrials.gov Identifier: new patients in clinical trials until the preclinical NCT00739687, NCT00557518, NCT00662116, work for alagebrium was completed [34]. NTC00089713), the effects of alagebrium on Also, it is unknown whether alagebrium has a chronic heart disease and renal dysfunction in indi- direct link to RAGE. The correlation of the In the subsequent Phase I study, high-dose alage- viduals with and without diabetes were evaluated reduction in RAGE protein expression with brium was used to evaluate its effect on triglyc- by Synvista Therapeutics Inc. between 2004 and alagebrium was found in preclinical studies. In eride, total cholesterol, high-density lipoprotein 2009. Unfortunately, these studies were termi- transgenic mice without cell surface RAGE, al- cholesterol (HDL-C), and low-density lipoprotein nated in the early period due to the bankruptcy of agebrium treatment has been shown to improve cholesterol (LDL-C), and no harmful effects on Synvista Therapeutics Inc. and global financial crisis. renal function and pathology and cardiovascu- liver function were detected [34]. In 2013, a randomized, double-blind, placebo-con- lar disease. Therefore, it is unknown whether trolled cardiovascular clinic trial aimed to evaluate the deterioration of AGE pathway is related to In the Phase I studies on healthy subjects, no the effect of multi-dose alagebrium with individual RAGE protein translation/transcription through side effects or deaths were observed after al- exercise on diastolic heart failure (ClinicalTrials.gov direct feedback or the reduction of metal ions agebrium treatment. However, it is unknown Identifier: NCT01913301). This study was also and oxidative stress [30, 31]. exactly at which time period these Phase I ex- discontinued because of financial causes. In the last periments were made. Between 2001 and 2003, two randomized, double-blind, placebo-controlled Alagebrium Clinical Trials Phase IIb clinical trials were conducted in many studies, exercise was combined with 200 mg/day regions of North America. Individuals were di- alagebrium for 1 year in the elderly population. Preclinical Studies vided into two cohorts based on left ventricular Alagebrium was mostly well-tolerated except for Many animal model studies of ALT-711 have hypertrophy (LVH). The Systolic Hypertension the gastrointestinal symptoms observed in two in- shown the potential benefit in reversing cardio- Interaction with Left Ventricular Remodeling dividuals [38, 39]. vascular complications in aging and diabetes. (SILVER) study was based on LVH hypertrophy, and there was no study except for Systolic and In one study, when compared to controls, no In a study by the Alton Ochsner Clinic (Hyper- Pulse Pressure Hemodynamic Improvement by independent effect of alagebrium was observed tension Research Laboratory, Alton Ochsner Restriction Elasticity (SAPPHIRE) [34]. on VO2 on endothelial function and arterial Medical Foundation, New Orleans, LA, USA) stiffness. This effect simultaneously disappeared on non-diabetic, spontaneous hypertensive rats, Overall, in these studies, there was no significant with exercise intervention. ALT-711 treatment reversed aortic stiffness by change in systolic and pulse pressures when al- regulating left ventricular elasticity. The oral ad- agebrium therapy was compared with placebo. Similar to other pharmaceutical agents reported ministration of ALT-711 improved late diastolic When individual basal measurements were to date, it is impossible to note the beneficial ef- and stroke volumetric index, decreased left ven- compared with 3- and 6-month measurements, fects of alagebrium observed in diabetic animal tricular stiffness, and improved cardiac function some changes were observed in the systolic models at the same time in humans due to in- in aged dogs [32]. blood pressure [34]. complete clinical trials. To date, clinical trials with alagebrium have not been continued. In another study on aged monkeys, the pulse In the Phase II Distensibility Improvement with wave velocity and augmentation index de- ALT-711 Remodeling in Diastolic Heart Failure Diabetes and Alagebrium creased significantly and aortic stiffness -de (DIAMOND) study, the effect of alagebrium on Cardiovascular complications associated with creased continuously [33]. many parameters, including exercise oxygen con- diabetes mellitus are one of the major causes of 288 • Toprak and Yigitaslan. Alagebrium and Advanced Glycation End Products Eurasian J Med 2019; 51(3): 285-92 morbidity and mortality worldwide. Diabetes is nificantly increased in the aorta and kidney of on TGF-β or RAGE expression; different AGE- a common disease affecting 346 million people the methylglyoxal-administered rats [47]. mediated pathways were identified as mediating worldwide. the local NH response [52]. It has been shown that long-term alagebrium It is expected that this number will increase by treatment is effective in many chronic diseases, AGEs have recently shown to be associated with 552 million in 2030. In diabetes, micro- and mac- such as atherosclerosis, erectile dysfunction, vascular calcification through a RAGE-mediated rovascular diseases cause increased mortality cardiovascular disease, hypertension and kidney process. Despite the correlation between AGE and morbidity. More specifically, the risk of car- damage in diabetic animal models [48-51]. levels and vascular calcification, there is no evi- diovascular diseases and cardiovascular deaths in dence that reducing in vivo AGEs or inhibiting diabetic individuals is 2-fold higher than in those The mechanism of restenosis in type 2 diabetes the AGE–RAGE signaling pathway reduces me- without diabetes [40-43]. mellitus (T2DM) is not fully understood; how- dial calcification. In this study, the effect of the ever, AGEs most likely contribute to vascular inhibition of AGE formation by pyridoxamine Diabetes affects the cardiac tissue in many ways remodeling. AGE-induced collagen cross-linking and elimination of AGEs by alagebrium on dia- and one of them is the direct exposure of endo- (ARCC) increases intravenous resistance and betic medial calcification was assessed. When thelial cells to high blood sugar. Toxic products induces neointimal hyperplasia (NH) in T2DM the AGE–RAGE signaling pathway was inhibited, of non-enzymatic glycolysis, particularly MG, by altering intra-stent hemodynamics. Because calcification was observed to be prevented. Pre- induce endothelial cell damage. MG is the reac- Alagebrium reduces ARCC, these responses vious studies have shown that AGE inhibitors tive ketoaldehyde occurring from glucose me- will also decrease. Abdominal aortic stents were prevent time-dependent AGE accumulation in tabolism and is also the precursor for AGE and implanted to Zucker lean (ZL), Obese (ZO), the femoral arteries of diabetic rats. This ef- triggers many pathological events, such as type 2 and diabetic (ZD) rats. Blood pressure, vessel fect is accompanied by a decrease in acceler- diabetes, pancreatic cell dysfunction, endothelial diameter, and vessel wall stress were calculated ated calcification in diabetes. In ex-vivo experi- dysfunction, and hypertension [44]. after 21 days, and the amount of NH was de- ments, it was observed that the RAGE agonist termined [52]. N-methylpyridinium induced diabetic femoral The administration of ALT-711 for 4 weeks in artery calcification, and the inhibition of antioxi- rats with MG-induced diabetes improved beta- ARCC and TGF-β and RAGE expressions were dants and inhibitors of different signaling path- cell dysfunction. Therefore, selective and spe- determined in the arterial segments (the aorta, ways were associated with RAGE activation. The cific MG inhibitors may be effective against high carotid, iliac, femoral, and arterioles). In the ZD physiological significance of oxidative stress has glucose, high blood pressure, and AGEs. How- and ZL rats, the flow-direction resistance in- been demonstrated by the reduction of femoral ever, currently, there are no selective MG inhibi- creased by 60%, but flow and wall shear stress artery calcification in diabetic rats treated with tors; most compounds, such as aminoguanidine (WSS) decreased significantly (44% and 56%, the inhibitor of ROS apocynin. AGE inhibitors and metformin, act non-specifically [44]. respectively) [52]. have limited or prevented medial calcification. Therefore, the inhibition of AGE–RAGE signal- In a study, 4-week MG treatment significantly There was no significant difference in RAGE and ing probably reduces medial calcification [53]. increased blood pressure, total cholesterol, and TGF-β expressions between arterial segments. triglyceride levels and decreased HDL level. The In decellular matrix cells, ALT-711 modified the Diabetic Cardiomyopathy effect of the novel ALT-711 analog (compound lectin-type oxidized LDL receptor 1 but not Diabetic heart disease is a clinical condition that Nos. 1-15) on MG was investigated; the inhibi- RAGE expression. In conclusion, ALT-711 re- can progress to heart failure and sudden death. tory effect of compound No. 13 on MG was duced ARCC in ZD and ZO rats via a RAGE- However, the mechanisms responsible for the found. It has been observed that compound No. independent pathway, increased intra-stent flow changes in excitation–contraction coupling lead- 13 reduces MG-induced metabolic parameters rate, and reduced NH. This study aimed to ing to cardiac dysfunction during diabetes are significantly. Alagebrium had a protective effect emphasize the important role of NH in AGE- unknown. Hyperglycemia causes the formation against the harmful effects of high glucose and induced remodeling in T2DM [52]. of AGEs in longlived proteins, including sarco- MG. ALT-711, which is protective against dicar- plasmic reticulum (SR) Ca2+ regulatory proteins. bonyl-induced AGEs and safe and specific for Several previous findings have suggested that However, their pathogenic role in the use of MG, is therapeutically attractive [45,46]. there are many complex relationships between SR Ca2+ in cardiac myocytes is unknown. Thus, biomechanics and biological processes regulat- it was investigated whether an AGE cross-link In the majority of diabetic patients, hyperten- ing NH development in diabetes and obesity. breaker could prevent the changes in SR Ca2+ sion develops with increased renin–angiotensin In ZD rats, AGEs are localized in all vascular cycle leading to in vivo cardiac dysfunction during system activity. Methylglyoxal is reactive glucose locations and their high presence in T2DM has diabetes. Streptozotocin (STZ)-induced diabetic metabolite and is increased in diabetic patients. been confirmed. In ZD rats, increased ARCC- rats were treated with ALT-711 for 8 weeks and In a previous study, the effect of methylglyoxal induced stiffness in the arterioles was alleviated compared with age-matched placebo-treated on the renin–angiotensin system and blood with ALT-711 treatment. The mean blood pres- diabetic rats and healthy rats. Cardiac function pressure was investigated. sure decreased in ZD rats and at the same time was assessed using echocardiographic examina- arteriolar resistance increased. Compared with tion. Ventricular myocytes were isolated to as- There was a significant increase in the blood ZL rats receiving ALT-711 treatment, the stent sess the SR Ca2+ cycle using confocal imaging and pressure and plasma levels of aldosterone, renin, area WSS was lower in untreated ZD rats. NH quantitative western blot. Diabetes resulted in angiotensin, and catecholamine in methylglyoxal- was increased in the stent area in ZO rats but in vivo cardiac dysfunction, and ALT-711 treat- administered rats. The levels of methylglyoxal not in ZD rats; however, ALT-711 treatment re- ment partially alleviated diastolic dysfunction and angiotensin, angiotensin receptor 1, adren- duced NH in all groups. LOX-1 expression was (27% vs. 41%, P <0.05 versus untreated diabetic ergic α1D receptor, and renin mRNA were sig- increased with ALT-711, but there was no effect rats, respectively) by reducing the isovolumetric Eurasian J Med 2019; 51(3): 285-92 Toprak and Yigitaslan. Alagebrium and Advanced Glycation End Products • 289 relaxation time and myocardial performance in- In mesangial cells, in collaboration with cellular apoE-KO mice. Diabetic animals were left un- dex. Collectively, that study showed that AGE ROS, ALA effectively prevented the NADPH treated or treated with AGE formation inhibi- accumulation in the type 1 diabetes model sub- oxidase subunit (p47 phox, p67 phox, and rac1), tor aminoguanidine (1 g/kg per d) or cross-link stantially impaired SR Ca2+ re-uptake in cardiac protein kinase C (PKC) isoform (α, βI, and βII), breaker ALT-711. Non-diabetic apo-E mice are myocytes and that long-term treatment with an and Nox4 messenger RNA expression induced characterized by accelerated renal injury, albu-

AGE cross-link breaker partially improved SR by not only high glucose but also H2O2. The di- minuria, and glomerular tubulointerstitial dam- Ca2+ utilization and diabetic cardiomyopathy rect and indirect antioxidant effect plays an im- age compared to diabetes-induced apo-E mice. [54]. portant role in the renoprotective effect of ALA These abnormalities include increased type I in diabetic kidneys [57]. and type III collagen, TGF-β expression, alpha- Diabetic Nephropathy smooth muscle actin immunohistochemistry, Diabetic nephropathy (DN) is a global issue AGEs play a role in the development of both and macrophage infiltration and serum and renal that increases in number along with the increas- nephropathy and major arterial diseases. How- AGE increase. Both treatments reduced renal ing number of people with diabetes every year, ever, it is still unclear which AGE subtypes play AGE accumulation, i.e., resulted in less albumin- leading to an increase in the incidence of end- a pathogenic role and which RAGEs are associ- uria, structural damage, and macrophage infiltra- stage renal disease [55]. ated with the effects of AGE on the cell [58]. tion and TGF-β1 and collagen expression [60].

AGEs are important mediators of DN, and they In experimental diabetes, similar findings were AGE accumulation was suppressed with al- trigger renal inflammation and glomeruloscle- observed with the use of aminoguanidine and agebrium and ramipril to assess the effect of rosis via RAGE and several other mechanisms. ramipril (ACEI) in fluorescence and immunos- mediators having a role in renal function and The contribution of RAGE-dependent and inde- table AGE levels, renal PKC activity, nitrotyrosine diabetic renal injury on renin–angiotensin, AGE, pendent signaling pathways was investigated in expression, lysosomal function, and protein utili- mitochondrial and cytosolic oxidative stress, vivo. STZ-induced diabetic RAGE apoE double zation. These findings suggest that the inhibition and intracellular signaling molecules. The treat- knockout (KO) mice were treated with ALT-711 of the renin–angiotensin system blocks the path- ment groups received ramipril (1 mg/kg/d, wk (1 mg/kg/day) or the angiotensin-converting en- ways leading to tissue injury. Chemical pathways 0-32), alagebrium (10 mg/kg/d, wk 16-32), or zyme inhibitor (ACEI) quinapril (30 mg/kg/day) leading to the formation of AGEs and the renin– combined treatment. A significant effect on -al for 20 weeks, and the renal parameters were angiotensin systems may be induced by glucose buminuria was observed in the monotherapy evaluated. RAGE deletion reduced mesangial and angiotensin II to interact with the formation groups, but this effect was not observed in the expansion, glomerular matrix accumulation, and of free radicals. AGE-dependent pathways play combined treatment group. renal oxidative stress associated with diabetes. In an important role in tubulointerstitial fibrosis contrast, it has not prevented the inflammation in the diabetic kidney. This effect is dependent A change in the urinary vascular endothelial and AGE accumulation associated with diabetes. on RAGE, TGF-β, and connective TGF (CTGF) growth factor (VEGF) excretion was observed However, the treatment with alagebrium in dia- [58]. in albuminuria. In diabetes, circulatory angioten- betic RAGE apoE-KO mice reduced renal AGE sin II suppression is associated with increased levels and glomerular matrix accumulation. Even STZ-induced diabetic rats were treated with levels of AGE and CML in circulation and the in the absence of RAGE expression, alagebrium ALT-711 and divided into groups according to kidneys. All treatments reduced circulatory reduced cortical inflammation, as indicated by treatment periods of 16-32 weeks (early) and CML but not the renal CML. RAGE 1 renal the reduced expression of monocyte chemoat- 24-32 weeks (late). Treatment with ALT-711 gene expression and soluble receptor AGE have tractant protein-1, intracellular adhesion mol- significantly reduced diabetes-induced serum been significantly reduced in diabetes, which ecule-1, and macrophage marker cluster of dif- and AGE peptide fluorescence, resulting in re- returned to the normal level with alagebrium. ferentiation molecule-11b. These novel findings duced renal CML and RAGE. The decrease in Diabetes-induced renal mitochondrial oxidative confirm the presence of RAGE-independent tail tendon collagen cross-linking was observed stress reduced with alagebrium. In cytosol, both signaling pathways as well as RAGE-dependent only in the group treated for 16 weeks. ALT-711 treatments have been shown to be equally ef- signaling pathways that may be active in the kid- delayed albumin excretion rate and reduced fective in reducing ROS. Membrane PKC activity neys through AGEs [56]. blood pressure and renal hypertrophy regard- decreased in all treatment groups, but diabetes- less of the treatment duration. It also reduced associated nuclear factor kappa B p65 transloca- Despite the beneficial effects of ALT-711, the diabetes-induced TGF-β1, CTGF, and collagen tion remained unchanged [61]. renoprotective mechanisms in DN are not fully IV. However, glomerulosclerotic index, tubuloin- understood. Since oxidative stress exacerbates terstitial area, total renal collagen, nitrotyrosine, Another study investigated the effect of AGE diabetic renal damage through the interac- collagen protein expression, and TGF-β1 only on PKC isoform expression in DN. STZ-induced tion with AGE, a previous study examined the improved in the early treatment group. This diabetic rats were divided into groups that re- antioxidant properties of ALA in diabetic db/ study also explains the effect of cross-link break- ceived no treatment, ALT-711, or aminoguani- db mice (C57BL/KsJ Jc1 db/db) mesangial cells er treatment on DN not only with renal AGEs, dine. Diabetes also induced beta-I, beta-II, and cultured with high glucose or hydrogen perox- but also with presklorotic cytokine and oxidative epsilon isoforms as well as PKC-alpha. ide (H2O2). ALA not only significantly reduced stress [59]. urinary albumin excretion and renal pathologic ALT-711 and aminoguanidine treatment re- changes in db/db mice, but also reduced pen- A previous study was designed to elucidate the duced renal AGE accumulation and abolished tosidine and nitrotyrosine accumulation and ex- role of AGEs in the development and progres- the increase in PKC expression. However, the pression of nicotinamide adenine dinucleotide sion of renal disease and the renal damage in translocation of phosphorylated PKC-alpha phosphate hydrogenase (NADPH) oxidase sub- diabetic apolipoprotein E-KO (apoE-KO) mice. from the cytoplasm to the membrane de- units independent of the treatment protocol. Diabetes was induced with STZ in 6-week-old creased only with ALT-711. ALT-711 treatment 290 • Toprak and Yigitaslan. Alagebrium and Advanced Glycation End Products Eurasian J Med 2019; 51(3): 285-92 attenuated the expression of VEGF, extracel- limit neuropathic diabetic cutaneous pressure tion End Products (AGEs) in Diabetic Cardio- lular matrix proteins, fibronectin, and laminin ulcers [64]. vascular Disease. Res Cardiovasc Med 2015; 4: with decreased albuminuria. Despite no effect e26949. on VEGF expression, aminoguanidine treatment Diabetic Retinopathy 4. Serban AI, Stanca L, Geicu OI, Munteanu MC, resulted in a slight decrease in fibronectin and AGE accumulation is associated with many Costache M, Dinischiotu A. Extracellular matrix laminin. These findings suggest that AGEs are a complications of diabetes mellitus, including is modulated in advanced glycation end products major stimulus for PKC activation in the diabetic diabetic retinopathy. AGE-breakers, such as N- milieu via a RAGE receptor dependent pathway kidney and that PKC-alpha can be directly inhib- phenylacylthiazolium and alagebrium, have been boosted by transforming growth factor-beta1 RAGE. J Diabetes 2015; 7: 114-24. [CrossRef] ited by ALT-711 [62]. proposed as therapeutic agents for reversing 5. Sell DR, Monnier VM,Molecular basis of arte- the increase in protein cross-linking. Epicatechin rial stiffening: role of glycation - a mini-review. To evaluate the effect of ALT-711 on DN, ALT- is the main diet flavonoid with a wide range of Gerontology 2012; 58: 227-37. 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Sell DR, Strauch CM, Shen W, Monnier VM. against pressure ulcers on return. Peer-review: Externally peer-reviewed. 2-Aminoadipic acid is a marker of protein car- bonyl oxidation in the aging human skin: effects Author Contributions: Concept – C.T., S.Y.; Design In diabetic rats, severe neuropathy can signifi- of diabetes, renal failure and sepsis. Biochem J – C.T., S.Y.; Supervision - C.T., S.Y.; Resources - C.T., cantly affect vasodilatation induced by pressure. 2007; 404: 269-77. [CrossRef] S.Y.; Materials - C.T., S.Y.; Data Collection and/or An animal study aimed at determining if there is 13. Baynes JW, Thorpe SR. Role of oxidative stress in Processing - C.T., S.Y.; Analysis and/or Interpretation a pressure-induced vasodilatation change in dia- diabetic complications: a new perspective on an old - C.T., S.Y.; Literature Search - C.T., S.Y.; M.A.A.; Writ- paradigm. Diabetes 1999; 48: 1-9. [CrossRef] betic mice at 8 weeks. Control and diabetic mice ing Manuscript - C.T., S.Y.; Critical Review - C.T., S.Y.; were left untreated or treated with sorbinil (al- Other - C.T., S.Y.; 14. Reiser KM, Amigable MA, Last JA. Nonenzy- dose reductase inhibitor) or alagebrium in the matic glycation of type I collagen: the effects of last 2 weeks. Acknowledgements: We kindly thank the all investiga- aging on preferential glycation sites. J Biol Chem tors for their participation in this review. 1992; 267: 24207-16. Nerve functions were evaluated using the 15. Winlove CP, Parker KH, Avery NC, Bailey AJ. Inter- motor neuron speed (MNCV) as well as Conflict of Interest: The authors have no conflict of actions of elastin and aorta with sugars in vitro and the C-fiber-mediated nociception threshold. interest to declare. their effects on biochemical and physical properties. Diabetologia 1996; 39: 1131-9. [CrossRef] Pressure-induced vasodilatation, endothelial Financial Disclosure: The authors declared that this 16. 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