**9. Methylglyoxal and aging**

The accumulation of AGEs in extracellular tissue proteins, such as the basement membrane and matrix proteins of blood vessels and skin, is a well known phenomenon characteristic of aging and age-related diseases. Several studies demonstrate aging associated increase in AGEs. Thus, accumulation of AGEs in the vessel walls results in a gradual loss of elasticity, which makes older subjects more susceptible to cardiovascular diseases [133, 134]. MGinduced AGEs, such as CEL and CML, increased with age in human lens and cause cataract formation [90]. In a study on 172 subjects serum levels of CML, 8-isoprostanes and Creactive protein, which are markers of oxidative stress, were higher in elderly people (>60 years old) compared with younger people (<45 years old) [135]. One reason why AGEs accumulate during the aging process could be due to an age-related decrease in antioxidant enzymes. Thus, Mailankot et al. [136] reported that the activity and expression of glyoxalase I protein, which is involved in MG degradation, decreased with age in the anterior epithelial cells of human lens, which causes an accumulation of MG. Similarly, an age-dependent decrease in catalase activity in the skin may be responsible for elevated MG and peroxynitrite production [137].

However, it is doubtful whether extracellular AGEs accumulation plays a causative role in aging. On the other hand AGEs formation inside the cell, such as AGE-nucleotides in DNA, may contribute to cellular senescence [6, 60, 74, 80]. DNA integrity is an important determinant of lifespan and errors in DNA repair would lead to substitutions, deletions, insertions, and transpositions of nucleotides, with increased risk of carcinogenesis and reduced life span. Animals with a longer lifespan and more efficient DNA repair have delayed carcinogenesis [80]. In this regard MG-induced DNA damage can have a more direct effect on aging.

Studies directly implicating MG in the aging process are very few and this is one area where there is a knowledge gap. The study by Morcos et al in the worm *C. elegans* highlights the role of MG, glyoxalase I and MG-induced ROS formation in aging and life span [138]. They showed that the activity of glyoxalase I was markedly reduced with age resulting in accumulation of MG-derived adducts and oxidative stress markers, which further inhibited

Aging: Drugs to Eliminate Methylglyoxal, a Reactive

Hodge pathway

Glucose Metabolite, and Advanced Glycation Endproducts 691

**Reactive Diacrbonyls (MG, GO, 3-DG)**

**Protein + Sugar**

**Schiff base + H2O**

**Amadori product**

**Protein cross-links, advanced glycation endproducts (Pentosidine, CML, CEL, Fluorescent and Nonfluorescent AGEs)**

Based on the scheme proposed by Khalifah et al. [83]

[145, 146] and metformin [147, 148].

Fig. 4. Stages of formation of advanced glycation endproducts (AGEs) from glycation of proteins. Nonenzymatic glycation of protein leads to reversible formation of Schiff bases, which lead to further reversible formation of Amadori adducts and ultimate formation of stable irreversible AGEs. Solid vertical arrows show these steps which form the classical Hodge pathway of AGEs formation. Auto oxidation of glucose (Wolff pathway) or of the Schiff base (Namiki pathway) forms reactive dicarbonyls such as methylglyoxal (MG), glyoxal (GO) or 3-deoxyglucosone (3-DG) which is mostly seen *in vitro*, rather than *in vivo*. The dicarbonyls, which are also formed from other metabolic pathways, also contribute significantly to AGEs formation, as explained in the text. The various sites at which anti-AGEs and anti-MG compounds can act are indicated by numbers and discussed in the text.

Site 3. AGEs and reactive aldehydes also generate reactive oxygen species which adds to their damaging effects [17, 141, 142]. Antioxidants such as vitamin C or E [143] and metal

Site 4. Reactive aldehydes such as MG, glyoxal, glycoaldehyde and glucosones, which are formed during nutrient metabolism and AGEs formation, are a major source of AGEs formation. Reactive aldehydes can be neutralized by compounds such as aminoguanidine

Site 5. Amadori adducts, formed in the intermediate stages of AGEs formation can either be quenched by compounds such as aminoguanidine, or degraded enzymatically by enzymes such as amadoriase and human fructosamine-3-kinase, which belong to this group [149,

Site 6. The final group of compounds acts on formed AGEs and are therefore, known as AGE breakers or cross-link breakers. E.g. phenacylthiazolium bromide (PTB) [153] and

chelators such as penicillamine [144] can be used to quench ROS and metal ions.

150]. Amadoriases have not been detected in higher organisms [151, 152].

the expression and the activity of glyoxalase I. Over expression of glyoxalase I decreased MG-induced modification of mitochondrial proteins and ROS production, and prolonged the lifespan of *C. elegans*; whereas CeGly knock-out produced the opposite effect [138].

Scheckhuber et al. studied the degradation of MG by the glyoxalase system enzymes and its effect on growth and lifespan in filamentous ascomycete and a model of aging, Podospora anserine (*P. anserina*) [139]. Using genetic manipulation of the two enzymes of the glyoxalase system, they found that up-regulation of both components of the glyoxalase system was effective in increasing lifespan in P. anserina.

Oxidative stress-induced cellular senescence was demonstrated in the study by Sejeresen and Rattan [140]. They treated human skin fibroblasts with MG (400 μM), or glyoxal (1.0 mM), and found the appearance of various senescent phenotypes within three days. These phenotypes showed growth arrest, had increased hydrogen peroxide and the glyoxalinduced AGE, Nε-carboxymethyl lysine (CML) protein levels, and altered SOD and catalase antioxidant enzyme activities [140]. Sejeresen and Rattan proposed this model to study cellular senescence *in vitro* [140].

In this review we have highlighted some important facts that oxidative stress is a major factor in the cellular and whole body aging processes, AGEs are strongly associated with aging and MG is a major precursor of AGEs formation, and both MG and AGEs are potent inducers of oxidative stress. Based on these facts, it is highly likely that MG may have a major role in the aging process through induction of oxidative stress as depicted in the scheme in Fig. 3. In fact, elevated MG levels may be responsible for causing accelerated aging in many tissues and organs of the cardiovascular system, nervous system, and other systems of the human body. A strategy to prevent aging should include targeting MG by reducing excessive formation, inhibit AGEs formation, and remove excessively formed MG and AGEs from the body.

#### **10. Anti-MG and anti-AGEs compounds**

The deleterious effects of MG and AGEs can be prevented by compounds that can do one or all of the following: (i) bind and neutralize reactive aldehydes, especially MG, (ii) prevent the formation of AGEs, (iii) neutralize formed AGEs, (iv) break down formed AGEs.

Considering these multiple ways of preventing the effects of MG or AGEs these compounds will be termed as "anti-MG" or "anti-AGE". Unfortunately, specific anti-MG or anti-AGE compounds are not yet available. The ones available are non-specific and have one or more other effects, which limits their usefulness. A number of the available compounds happen to have anti-MG as well as anti-AGE effects.

The possible sites at which anti-AGEs and anti-MG drugs can work are shown in Fig. 4, which outlines the various stages of AGEs formation.

Site 1. The first step in glycation is the binding of a sugar to the free amino groups of a protein. Drugs that bind to the amino group of proteins, such as aspirin, can prevent the binding of the sugar to the amino group. This group of compounds is likely to produce non-specific effects.

Site 2. Compounds can be used to bind aldose and ketose sugars to neutralize them and prevent them from reacting with proteins. E.g. Aminoguanidine reacts with the carbonyl group of glucose and prevents AGEs formation.

the expression and the activity of glyoxalase I. Over expression of glyoxalase I decreased MG-induced modification of mitochondrial proteins and ROS production, and prolonged the lifespan of *C. elegans*; whereas CeGly knock-out produced the opposite effect [138].

Scheckhuber et al. studied the degradation of MG by the glyoxalase system enzymes and its effect on growth and lifespan in filamentous ascomycete and a model of aging, Podospora anserine (*P. anserina*) [139]. Using genetic manipulation of the two enzymes of the glyoxalase system, they found that up-regulation of both components of the glyoxalase

Oxidative stress-induced cellular senescence was demonstrated in the study by Sejeresen and Rattan [140]. They treated human skin fibroblasts with MG (400 μM), or glyoxal (1.0 mM), and found the appearance of various senescent phenotypes within three days. These phenotypes showed growth arrest, had increased hydrogen peroxide and the glyoxalinduced AGE, Nε-carboxymethyl lysine (CML) protein levels, and altered SOD and catalase antioxidant enzyme activities [140]. Sejeresen and Rattan proposed this model to study

In this review we have highlighted some important facts that oxidative stress is a major factor in the cellular and whole body aging processes, AGEs are strongly associated with aging and MG is a major precursor of AGEs formation, and both MG and AGEs are potent inducers of oxidative stress. Based on these facts, it is highly likely that MG may have a major role in the aging process through induction of oxidative stress as depicted in the scheme in Fig. 3. In fact, elevated MG levels may be responsible for causing accelerated aging in many tissues and organs of the cardiovascular system, nervous system, and other systems of the human body. A strategy to prevent aging should include targeting MG by reducing excessive formation, inhibit AGEs formation, and remove excessively formed MG

The deleterious effects of MG and AGEs can be prevented by compounds that can do one or all of the following: (i) bind and neutralize reactive aldehydes, especially MG, (ii) prevent

Considering these multiple ways of preventing the effects of MG or AGEs these compounds will be termed as "anti-MG" or "anti-AGE". Unfortunately, specific anti-MG or anti-AGE compounds are not yet available. The ones available are non-specific and have one or more other effects, which limits their usefulness. A number of the available compounds happen to

The possible sites at which anti-AGEs and anti-MG drugs can work are shown in Fig. 4,

Site 1. The first step in glycation is the binding of a sugar to the free amino groups of a protein. Drugs that bind to the amino group of proteins, such as aspirin, can prevent the binding of the sugar to the amino group. This group of compounds is likely to produce non-specific effects. Site 2. Compounds can be used to bind aldose and ketose sugars to neutralize them and prevent them from reacting with proteins. E.g. Aminoguanidine reacts with the carbonyl

the formation of AGEs, (iii) neutralize formed AGEs, (iv) break down formed AGEs.

system was effective in increasing lifespan in P. anserina.

cellular senescence *in vitro* [140].

and AGEs from the body.

**10. Anti-MG and anti-AGEs compounds** 

have anti-MG as well as anti-AGE effects.

which outlines the various stages of AGEs formation.

group of glucose and prevents AGEs formation.

**(Pentosidine, CML, CEL, Fluorescent and Nonfluorescent AGEs)**

Fig. 4. Stages of formation of advanced glycation endproducts (AGEs) from glycation of proteins. Nonenzymatic glycation of protein leads to reversible formation of Schiff bases, which lead to further reversible formation of Amadori adducts and ultimate formation of stable irreversible AGEs. Solid vertical arrows show these steps which form the classical Hodge pathway of AGEs formation. Auto oxidation of glucose (Wolff pathway) or of the Schiff base (Namiki pathway) forms reactive dicarbonyls such as methylglyoxal (MG), glyoxal (GO) or 3-deoxyglucosone (3-DG) which is mostly seen *in vitro*, rather than *in vivo*. The dicarbonyls, which are also formed from other metabolic pathways, also contribute significantly to AGEs formation, as explained in the text. The various sites at which anti-AGEs and anti-MG compounds can act are indicated by numbers and discussed in the text. Based on the scheme proposed by Khalifah et al. [83]

Site 3. AGEs and reactive aldehydes also generate reactive oxygen species which adds to their damaging effects [17, 141, 142]. Antioxidants such as vitamin C or E [143] and metal chelators such as penicillamine [144] can be used to quench ROS and metal ions.

Site 4. Reactive aldehydes such as MG, glyoxal, glycoaldehyde and glucosones, which are formed during nutrient metabolism and AGEs formation, are a major source of AGEs formation. Reactive aldehydes can be neutralized by compounds such as aminoguanidine [145, 146] and metformin [147, 148].

Site 5. Amadori adducts, formed in the intermediate stages of AGEs formation can either be quenched by compounds such as aminoguanidine, or degraded enzymatically by enzymes such as amadoriase and human fructosamine-3-kinase, which belong to this group [149, 150]. Amadoriases have not been detected in higher organisms [151, 152].

Site 6. The final group of compounds acts on formed AGEs and are therefore, known as AGE breakers or cross-link breakers. E.g. phenacylthiazolium bromide (PTB) [153] and

Aging: Drugs to Eliminate Methylglyoxal, a Reactive

as an insulin sensitizing agent.

their anti-MG or anti-AGE effects.

Glucose Metabolite, and Advanced Glycation Endproducts 693

which binds with MG to form an inactive product, triazepinone [147, 148, 174, 175]. The MG-scavenging effect of metformin resulted in significantly reduced elevated MG levels in type 2 diabetes patients treated with high doses of metformin, between 1,500 and 2,500 mg/day [172]. We have shown that fructose-fed Sprague-Dawley rats had significantly elevated serum MG and blood pressure, and increased levels of MG, hydrogen peroxide and the MG-derived AGE, CEL, in the aorta, all of which were attenuated by metformin [65]. Metformin has been proposed to protect against MG-induced increased atherogenicity of low density lipoprotein (LDL) [176]. The use of metformin as a MG scavenger and AGEs inhibitor is limited, and hopefully more studies showing its anti-MG, anti-AGEs effectiveness will promote its use for this purpose. Metformin can be considered to have a good therapeutic potential in this regard since it is already in clinical use for type 2 diabetes

Pioglitazone, a thiazolidinedione, is a peroxisome proliferator-activated receptor gamma (PPARγ) agonist which acts as an insulin sensitizer and is used in type 2 diabetes. Pioglitazone has been proposed to have anti-AGEs effect by inhibiting glycation, AGEs formation and protein cross-linking [177, 178]. Studies employing pioglitazone as an anti-MG or anti-AGEs compound are not forthcoming and more evidence is needed to make

N-acetylcysteine (NAC) is a MG scavenging and antioxidant compound [179, 180]. There are good reasons for using NAC as an anti-MG compound: NAC can increase GSH levels [181], which is an efficient MG scavenger and antioxidant [70, 179, 180], NAC is a cysteine containing thiol compound and MG binds with high affinity to cysteine [180, 181], and NAC is already used clinically for other conditions such as acetaminophen overdose [179, 181]. More studies employing NAC as an anti-MG drug should provide interesting results and

A widely used class of antihypertensive drugs has been proposed to have anti-AGE effects. Angiotensin receptor blockers (ARBs) and angiotensin converting enzyme inhibitors (ACEIs) have been shown to protect against kidney damage, an effect claimed to be independent of their blood pressure lowering action [182-185], but proposed to be due to an anti-AGE effect. Clinical trials evaluating AGEs lowering effects of ARBs and ACEIs at

A number of compounds have been investigated for anti-AGE effects in limited studies, but have not been widely used as such in experimental studies in animals or humans. These compounds include the cycloxygenase inhibitors, aspirin [186-188], ibuprofen [189], diclofenac [190], xanthine derivative, pentoxifylline [191, 192], which is used for claudication in peripheral vascular disease, metal chelators, D-penicillamine [144] and desferoxamine [83, 144], thiamine pyrophosphate and pyridoxamine [193-195]. Many more studies are necessary for these compounds in order to draw definitive conclusions about

A number of deglycating enzymes have been discovered, especially in microorganisms, which remove the sugar bound to the protein molecule, and possibly provide these bacteria with energy substrates derived from glycated products [196]. These enzymes, known as amadoriases, include fructosylamine oxidases [197, 198], fructosamine-3-kinase [149, 150], fructoselysine-6-kinase [199], fructoselysine-3-epimerase (FrlC) [200], and glucoselysine-6-

blood pressure lowering doses can add to the therapeutic utility of these drugs.

definitive statements about the therapeutic potential of pioglitazone in this regard.

may help to establish the potential of NAC in this regard.

alagebrium (previously known as ALT-711), which are thiazolium compounds. AGEs which do not have cross-links such as pentosidine [154], GOLD and MOLD [155] will not be affected by these drugs.

Some of the more common compounds with anti-MG or anti-AGEs effects are described below.

Aminoguanidine is one of the popular and widely used AGEs inhibitor [145] and MG scavenger. Despite being a guanidine derivative it shares many common properties with hydrazine and is classified as a hydrazine [156]. As described above, aminoguanidine acts at site 2 as well as site 4 (Fig. 4) meaning that it prevents AGEs formation by combining with the carbonyl group of glucose as well as by scavenging reactive dicarbonyls formed during various metabolic processes [146]. The inhibitory effect of aminoguanidine is mainly at the Amadori stage [83]. Aminoguanidine is not a specific AGEs inhibitor or MG scavenger and it has other actions. Aminoguanidine potently inhibits histaminases [157, 158] and prevents deamination of histamine and putrescine. Aminoguanidine also inhibits nitric oxide synthase (NOS) [159, 160] and prevents the formation of nitric oxide (NO), a dynamic signaling molecule in the body [161], from L-arginine. Aminoguanidine also binds to the enzyme S-adenosylmethionine decarboxylase and increases synthesis of polyamines such as spermidine and spermine from ornithine [162]. Aminoguanidine can also bind pyridoxal and cause vitamin B6 deficiency, which in turn can result in adverse reactions to aminoguanidine [163]. A number of *in vitro* and *in vivo* studies have described the inhibitory effects of aminoguanidine on AGEs formation [145, 146, 164-167]. The doses used *in vivo* range from 25 mg/kg/day [145, 166] to 50 mg/kg/day [165] and up to 100 mg/kg/day [167]. Thus, aminoguanidine is far from an ideal MG scavenger and AGEs inhibitor. In clinical trials aminoguanidine was found to be too toxic for use in patients. Two doublemasked, multiple-dose, placebo-controlled, randomized clinical trials, ACTION I and ACTION II [168, 169] investigated the therapeutic potential of aminoguanidine in preventing the progression of renal damage in patients with diabetic nephropathy. The ACTION I trial involving 690 participants did not show a statistically significant difference between the placebo group and the combined aminoguanidine dose groups, even though patients treated with aminoguanidine showed a tendency of having a lower risk of doubling of serum creatinine [168]. Due to safety concerns and an apparent lack of efficacy, the External Safety Monitoring Committee for the ACTION II trial involving 599 participants recommended early termination [169]. Patients with diabetes may have impaired red blood cell-deformability, which could cause microvascular and kidney damage. A one year trial with aminoguanidine and erythropoietin on 12 patients on dialysis restored red blood celldeformability to near-normal levels, an effect attributed to inhibition of AGEs formation by aminoguanidine [170]. As mentioned earlier the toxic effects of aminoguanidine have limited its therapeutic potential. For example, like hydrazine, aminoguanidine may be associated with drug-induced systemic lupus erythematosus and abnormal liver function tests, and it can cause flu-like syndromes and vasculitis [169]. Aminoguanidine can also cause damage to DNA through hydroxyl- and hydrogen peroxide-formation in the presence of Fe+3 [171].

Metformin is an oral dimethylbiguanide antihyperglycemic agent, which can also inhibit AGEs formation [172], through its action in the post-Amadori stages [83, 173]. Metformin has also been proposed to have a MG scavenging effect attributed to its guanidino group,

alagebrium (previously known as ALT-711), which are thiazolium compounds. AGEs which do not have cross-links such as pentosidine [154], GOLD and MOLD [155] will not be

Some of the more common compounds with anti-MG or anti-AGEs effects are described

Aminoguanidine is one of the popular and widely used AGEs inhibitor [145] and MG scavenger. Despite being a guanidine derivative it shares many common properties with hydrazine and is classified as a hydrazine [156]. As described above, aminoguanidine acts at site 2 as well as site 4 (Fig. 4) meaning that it prevents AGEs formation by combining with the carbonyl group of glucose as well as by scavenging reactive dicarbonyls formed during various metabolic processes [146]. The inhibitory effect of aminoguanidine is mainly at the Amadori stage [83]. Aminoguanidine is not a specific AGEs inhibitor or MG scavenger and it has other actions. Aminoguanidine potently inhibits histaminases [157, 158] and prevents deamination of histamine and putrescine. Aminoguanidine also inhibits nitric oxide synthase (NOS) [159, 160] and prevents the formation of nitric oxide (NO), a dynamic signaling molecule in the body [161], from L-arginine. Aminoguanidine also binds to the enzyme S-adenosylmethionine decarboxylase and increases synthesis of polyamines such as spermidine and spermine from ornithine [162]. Aminoguanidine can also bind pyridoxal and cause vitamin B6 deficiency, which in turn can result in adverse reactions to aminoguanidine [163]. A number of *in vitro* and *in vivo* studies have described the inhibitory effects of aminoguanidine on AGEs formation [145, 146, 164-167]. The doses used *in vivo* range from 25 mg/kg/day [145, 166] to 50 mg/kg/day [165] and up to 100 mg/kg/day [167]. Thus, aminoguanidine is far from an ideal MG scavenger and AGEs inhibitor. In clinical trials aminoguanidine was found to be too toxic for use in patients. Two doublemasked, multiple-dose, placebo-controlled, randomized clinical trials, ACTION I and ACTION II [168, 169] investigated the therapeutic potential of aminoguanidine in preventing the progression of renal damage in patients with diabetic nephropathy. The ACTION I trial involving 690 participants did not show a statistically significant difference between the placebo group and the combined aminoguanidine dose groups, even though patients treated with aminoguanidine showed a tendency of having a lower risk of doubling of serum creatinine [168]. Due to safety concerns and an apparent lack of efficacy, the External Safety Monitoring Committee for the ACTION II trial involving 599 participants recommended early termination [169]. Patients with diabetes may have impaired red blood cell-deformability, which could cause microvascular and kidney damage. A one year trial with aminoguanidine and erythropoietin on 12 patients on dialysis restored red blood celldeformability to near-normal levels, an effect attributed to inhibition of AGEs formation by aminoguanidine [170]. As mentioned earlier the toxic effects of aminoguanidine have limited its therapeutic potential. For example, like hydrazine, aminoguanidine may be associated with drug-induced systemic lupus erythematosus and abnormal liver function tests, and it can cause flu-like syndromes and vasculitis [169]. Aminoguanidine can also cause damage to DNA through hydroxyl- and hydrogen peroxide-formation in the presence

Metformin is an oral dimethylbiguanide antihyperglycemic agent, which can also inhibit AGEs formation [172], through its action in the post-Amadori stages [83, 173]. Metformin has also been proposed to have a MG scavenging effect attributed to its guanidino group,

affected by these drugs.

below.

of Fe+3 [171].

which binds with MG to form an inactive product, triazepinone [147, 148, 174, 175]. The MG-scavenging effect of metformin resulted in significantly reduced elevated MG levels in type 2 diabetes patients treated with high doses of metformin, between 1,500 and 2,500 mg/day [172]. We have shown that fructose-fed Sprague-Dawley rats had significantly elevated serum MG and blood pressure, and increased levels of MG, hydrogen peroxide and the MG-derived AGE, CEL, in the aorta, all of which were attenuated by metformin [65]. Metformin has been proposed to protect against MG-induced increased atherogenicity of low density lipoprotein (LDL) [176]. The use of metformin as a MG scavenger and AGEs inhibitor is limited, and hopefully more studies showing its anti-MG, anti-AGEs effectiveness will promote its use for this purpose. Metformin can be considered to have a good therapeutic potential in this regard since it is already in clinical use for type 2 diabetes as an insulin sensitizing agent.

Pioglitazone, a thiazolidinedione, is a peroxisome proliferator-activated receptor gamma (PPARγ) agonist which acts as an insulin sensitizer and is used in type 2 diabetes. Pioglitazone has been proposed to have anti-AGEs effect by inhibiting glycation, AGEs formation and protein cross-linking [177, 178]. Studies employing pioglitazone as an anti-MG or anti-AGEs compound are not forthcoming and more evidence is needed to make definitive statements about the therapeutic potential of pioglitazone in this regard.

N-acetylcysteine (NAC) is a MG scavenging and antioxidant compound [179, 180]. There are good reasons for using NAC as an anti-MG compound: NAC can increase GSH levels [181], which is an efficient MG scavenger and antioxidant [70, 179, 180], NAC is a cysteine containing thiol compound and MG binds with high affinity to cysteine [180, 181], and NAC is already used clinically for other conditions such as acetaminophen overdose [179, 181]. More studies employing NAC as an anti-MG drug should provide interesting results and may help to establish the potential of NAC in this regard.

A widely used class of antihypertensive drugs has been proposed to have anti-AGE effects. Angiotensin receptor blockers (ARBs) and angiotensin converting enzyme inhibitors (ACEIs) have been shown to protect against kidney damage, an effect claimed to be independent of their blood pressure lowering action [182-185], but proposed to be due to an anti-AGE effect. Clinical trials evaluating AGEs lowering effects of ARBs and ACEIs at blood pressure lowering doses can add to the therapeutic utility of these drugs.

A number of compounds have been investigated for anti-AGE effects in limited studies, but have not been widely used as such in experimental studies in animals or humans. These compounds include the cycloxygenase inhibitors, aspirin [186-188], ibuprofen [189], diclofenac [190], xanthine derivative, pentoxifylline [191, 192], which is used for claudication in peripheral vascular disease, metal chelators, D-penicillamine [144] and desferoxamine [83, 144], thiamine pyrophosphate and pyridoxamine [193-195]. Many more studies are necessary for these compounds in order to draw definitive conclusions about their anti-MG or anti-AGE effects.

A number of deglycating enzymes have been discovered, especially in microorganisms, which remove the sugar bound to the protein molecule, and possibly provide these bacteria with energy substrates derived from glycated products [196]. These enzymes, known as amadoriases, include fructosylamine oxidases [197, 198], fructosamine-3-kinase [149, 150], fructoselysine-6-kinase [199], fructoselysine-3-epimerase (FrlC) [200], and glucoselysine-6-

Aging: Drugs to Eliminate Methylglyoxal, a Reactive

pancreatic islet β-cell function in male Sprague-Dawley rats [72].

more can be done about their therapeutic potential.

the elderly. *Circulation* 123:1900-1910; 2011.

cells. *Nat. Rev. Mol. Cell Biol.* 8:729-740; 2007.

*Proc. Natl. Acad. Sci. U. S. A.* 81:105-109; 1984.

**12. References** 

2011.

164; 2006.

614-636; 1965.

Glucose Metabolite, and Advanced Glycation Endproducts 695

In the studies described above alagebrium has been studied mainly for its chronic effects on AGEs as an AGEs breaker. We investigated whether alagebrium also has acute preventive effects against the reactive dicarbonyl, MG, in 12 wk old male Sprague-Dawley rats [66]. Our results showed that alagebrium also has acute (< 6 h) MG scavenging ability [66]. AGEs are formed slowly over a time ranging from 24 h to up to 7 days and more. Therefore, the attenuation of MG-induced acute effects (seen within 6 h of MG administration) is most likely due to scavenging of MG by alagebrium. Thus, alagebrium significantly attenuated the significant increases in MG levels in the plasma, and different organs (measured 2 h after administration), and also attenuated MG-induced impaired glucose tolerance and the reduced insulin-stimulated glucose uptake by adipose tissue. In an *in vitro* assay in which MG (10 μM) was incubated with or without alagebrium (100 μM) for different times at 37° C, alagebrium significantly reduced the amount of detectable MG [66]. Our results strongly indicate an acute MG scavenging effect of alagebrium which can add to its AGEs breaking ability. More direct evidence of interaction of alagebrium and MG using mass spectrometry would be very useful. We have also recently shown that alagebrium significantly attenuated the deleterious effects of chronic MG administration for 4 wks on glucose tolerance and

In conclusion, MG and AGEs are very likely to be involved in the initiation and or progression of the aging process. Commonly available pharmacological compounds to investigate these roles of MG and AGEs, such as aminoguanidine, are non-specific, whereas some of the newer compounds appear promising in inhibiting AGEs formation at multiple steps in the pathway in *in vitro* studies. However, more *in vivo* studies are required before their therapeutic potential can be established. A more dedicated effort is necessary to identify newer anti-MG and anti-AGEs compounds which are more specific and safer before

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[2] Kovacic, J. C.; Moreno, P.; Hachinski, V.; Nabel, E. G.; Fuster, V. Cellular senescence,

[3] Kovacic, J. C.; Moreno, P.; Nabel, E. G.; Hachinski, V.; Fuster, V. Cellular senescence,

[4] Matthews, C.; Gorenne, I.; Scott, S.; Figg, N.; Kirkpatrick, P.; Ritchie, A.; Goddard, M.;

[5] Campisi, J.; d'Adda di Fagagna, F. Cellular senescence: When bad things happen to good

[6] Bucala, R.; Model, P.; Cerami, A. Modification of DNA by reducing sugars: A possible

vascular disease, and aging: Part 1 of a 2-part review. *Circulation* 123:1650-1660;

vascular disease, and aging: Part 2 of a 2-part review: Clinical vascular disease in

Bennett, M. Vascular smooth muscle cells undergo telomere-based senescence in human atherosclerosis: Effects of telomerase and oxidative stress. *Circ. Res.* 99:156-

mechanism for nucleic acid aging and age-related dysfunction in gene expression.

phosphate deglycase [201]. The use of these enzymes or development of stable analogues for deglycation therapy remains speculative .

A number of AGE inhibitors were synthesized and screened for their AGEs inhibitory effects by Rahbar et al. [173, 202, 203]. These are derivatives of aryl ureido and aryl carboxaminido phenoxy isobutyric acids and were derived from some known AGEs inhibitors. These compounds act at multiple stages of the AGEs formation process. Some of these compounds have AGEs breaking properties. More studies are needed for these compounds to establish their specificity and safety for therapeutic use.
