Antidiabetogenic Features of Benzimidazoles

*Alexander A. Spasov, Pavel M. Vassiliev, Vera A. Anisimova and Olga N. Zhukovskaya*

#### **Abstract**

Literature data on the insulinogenic effect of 2-aminobenzimidazole prompted us to investigate its novel derivatives, particularly those containing an additional fused cycle in C1,2-α position, including imidazole, dihydroimidazole, or tetrahydropyrimidine ring. Consensus analysis of the hypoglycemic effect of these compounds performed with IT Microcosm and PASS system revealed that activity is mostly characteristic for N9 -2,3-dihydroimidazo[1,2-a]benzimidazole derivatives. Substructural analysis of hypoglycemic activity identified substituents that determine the greatest pharmacological effect. According to the in silico assessment of the ADME properties, RU-254 was nominated as a lead compound due to the most optimal calculated and experimental activity and pharmacokinetic parameters. Preclinical studies have shown that identified compound has a pronounced insulinogenic effect and hypoglycemic effect, both in intact animals and in animals with experimental diabetes mellitus. RU-254 also reduces the level of glycated hemoglobin upon chronic administration, slightly decreases the activity of DPP-4, and increases the average number of Langerhans islets in the pancreas. Pharmaceutical drug formulation of RU-254 was developed and investigated for pharmacokinetic, pharmacodynamic, and toxicological properties. The dosage form of the drug under the name limiglidol (compound RU-254, diabenol) was evaluated in the full cycle of clinical studies that confirmed the safety, tolerability, and prominent antidiabetic properties of the drug.

**Keywords:** in silico, IT Microcosm, consensus prediction, antidiabetic effect, aminobenzimidazoles, cyclic benzimidazoles, pharmacodynamics, pharmacokinetics, toxicology, diabenol

#### **1. Introduction**

The history of drug discovery for the treatment of diabetes mellitus was and still is strongly determined by achievements in the field of fundamental medicine. Initially, the role of the pancreas and islets of Langerhans in the development of this pathology was proved; later, the structure of insulin, insulin receptor, and glucose transporters was deciphered; the role of the liver glycogenolysis and gluconeogenic enzymes, contributing to increased glucose output and hyperglycemia, was established; molecular mechanisms for the development of insulin resistance, the importance of the incretin system and Na+ /glucose transporters in the kidneys, and intestinal α-glucosidase were revealed, which led to the introduction of novel antidiabetic drugs into clinic [1–4].

The basis of insulin resistance at the cellular level primarily resides in the disruption of insulin signaling pathway at the level of the insulin receptor and insulin receptor substrate (IRS) proteins. The underlying mechanism of this phenomenon is impaired phosphorylation of serine amino acid residues, catalyzed by a number of intracellular protein kinases. The muscles, liver, and adipose tissue are the primary target organs of concern for the development of insulin resistance [5]. It was established that the severity of insulin resistance correlates, first of all, with intracellular lipid accumulation [6]. It is intracellular lipids that hamper signal transmission from the insulin receptor and cause a decrease in insulin-dependent glucose uptake. The pivotal role of AMP-dependent protein kinase (AMPK), which is an energy "sensor" of the cell, is also established, since AMPK through TORCI, the first mTOR-based protein complex, serves as a metabolic switch between catabolic and anabolic processes of the cell. Metformin is a biguanide derivative, which is the first-line drug for the treatment of type 2 diabetes. In 2001, it was shown that the molecular mechanism of its action is at least in part mediated by AMPK [7]. It is believed that indirect activation of AMPK by metformin-induced Ser172 phosphorylation determines its pleiotropic effects [8].

At the same time, it is important to note that course of type 2 diabetes mellitus characterized by several consecutive phases. It begins with primary insulin resistance and compensatory hyperinsulinemia with the subsequent development of β-cell dysfunction, thus creating the need for administration of insulin secretagogues or insulin formulations at the late stages of the disease [9–11].

Given that the previous works described the insulinogenic effect of the antihelminthic drug mebendazole [12], which can be considered as a new scaffold (2-aminobenzimidazole or cyclic guanidine) that exhibits an insulinogenic effect, we performed an experimental study of the novel cyclic guanidine derivatives, designed by introduction of additional fused cycle (imidazole, dihydroimidazole, and tetrahydropyrimidine).

#### **2. Results**

The synthesis of novel 2-aminobenzimidazole (AmBI) [13, 14] derivatives and fused benzimidazole derivatives was carried out, including N9 -imidazo[1,2-*a*] benzimidazoles (N9 -ImBI) [15–17], N1 -imidazo[1,2-*a*]benzimidazoles (N1 - ImBI) [18, 19], N9 -2,3-dihydroimidazo[1,2-*a*]benzimidazoles (N9 -DhImBI) [20, 21], N1 -2,3-dihydroimidazo[1,2-*a*]benzimidazoles (N1 -DhImBI) [18, 19], and 2,3,4,10-tetrahydropyrimido[1,2-*a*]benzimidazoles (PrmBI) [22–24].

In order to identify the most promising antidiabetic substances using IT Microcosm [25, 26] and PASS computer systems [27, 28], a step-by-step detailed in silico analysis of the hypoglycemic properties of the new compounds was carried out. Programs DruLiTo [29] and QikProp [30] were employed to assess key ADME properties and characteristics.

Hypoglycemic effect of the newly obtained derivatives was initially studied in rats upon intraperitoneal administration at a dose of 50 mg/kg. Blood sampling was carried out 4 hours after treatment with test compounds. Blood glucose concentration was determined with the glucose oxidase method using a commercial Glucose FKD kit [31]. The ratio of glucose concentrations in the blood plasma of the experimental and control group animals served as an indicator of hypoglycemic activity [32].

It was found that among condensed benzimidazole derivatives, a number of substances exceeded hypoglycemic activity of metformin, which served as a reference drug.

**75**

*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*


Thus, it was shown that N9


[35] and supremal [36] estimates, the class of N9

than 2,3,4,10-tetrahydropyrimido[1,2-a]benzimidazole.

atom of the N9

diabenol) as the most promising highly active compound.

antidiabetic drugs metformin and glibenclamide.

experimentally studied derivatives of N9

compound RU-254 (diabenol).


IT Microcosm [25, 26] and PASS [27, 28] computer systems were employed to determine the most promising chemical class of compounds. A training set of known hypoglycemic drugs and a library of tested benzimidazole derivatives were subjected to a consensus prediction of the level of hypoglycemic activity. The average informativity coefficient *KPr* was calculated and used as a metric for comparison









value ranges from 0 for inactive compounds to 5 for highly active compounds. According to value of *KPr*, the potential of benzimidazole derivatives classes as sources of substances with hypoglycemic activity decreases in the following


2,3,4,10-tetrahydropyrimido[1,2-*a*]benzimidazole derivatives have the most promising blood glucose lowering activity. That is, these tricyclic structures containing embedded guanidine group turned out to be more active than 2-aminobenzimidazole derivatives. Subsequently, employing substructural analysis [34] and analysis via median

imidazoles was selected as the most promising for the development of hypoglycemic compounds (**Figures 1** and **2**). It was shown that this scaffold is more preferable

Substructural analysis [34] of the level of hypoglycemic activity among the N9

dihydroimidazo[1,2-*a*]benzimidazole derivatives allowed us to reveal a chemical feature (substituent) that largely determines high hypoglycemic activity—diethylaminoethyl


high hypoglycemic activity was revealed—a charge on the internal imidazole cycle of the condensed system, namely, *Q(Imid1)cs* ≥ −0.109, which is a characteristic for

Taken together, the results of a complex consensus in silico analysis of the hypoglycemic activity of six classes of benzimidazole derivatives revealed 9-diethylaminoethyl-2,3-dihydroimidazo[1,2-*a*]benzimidazole dihydrochloride (RU-254,

To assess the feasibility of a further study of the pharmacological properties of compound RU-254, we calculated parameters of drug-likeliness and ADME properties (absorption, distribution, metabolism, excretion) for RU-254 and reference

**2.1 In silico study**

of AmBI, N9

order: N9

(*KPr* = 2.00) > N1

substituent at the N9

#### **2.1 In silico study**

*Chemistry and Applications of Benzimidazole and its Derivatives*

phorylation determines its pleiotropic effects [8].

tetrahydropyrimidine).

benzimidazoles (N9

properties and characteristics.

ImBI) [18, 19], N9

**2. Results**

21], N1

The basis of insulin resistance at the cellular level primarily resides in the disruption of insulin signaling pathway at the level of the insulin receptor and insulin receptor substrate (IRS) proteins. The underlying mechanism of this phenomenon is impaired phosphorylation of serine amino acid residues, catalyzed by a number of intracellular protein kinases. The muscles, liver, and adipose tissue are the primary target organs of concern for the development of insulin resistance [5]. It was established that the severity of insulin resistance correlates, first of all, with intracellular lipid accumulation [6]. It is intracellular lipids that hamper signal transmission from the insulin receptor and cause a decrease in insulin-dependent glucose uptake. The pivotal role of AMP-dependent protein kinase (AMPK), which is an energy "sensor" of the cell, is also established, since AMPK through TORCI, the first mTOR-based protein complex, serves as a metabolic switch between catabolic and anabolic processes of the cell. Metformin is a biguanide derivative, which is the first-line drug for the treatment of type 2 diabetes. In 2001, it was shown that the molecular mechanism of its action is at least in part mediated by AMPK [7]. It is believed that indirect activation of AMPK by metformin-induced Ser172 phos-

At the same time, it is important to note that course of type 2 diabetes mellitus characterized by several consecutive phases. It begins with primary insulin resistance and compensatory hyperinsulinemia with the subsequent development of β-cell dysfunction, thus creating the need for administration of insulin secreta-

Given that the previous works described the insulinogenic effect of the antihelminthic drug mebendazole [12], which can be considered as a new scaffold (2-aminobenzimidazole or cyclic guanidine) that exhibits an insulinogenic effect, we performed an experimental study of the novel cyclic guanidine derivatives, designed by introduction of additional fused cycle (imidazole, dihydroimidazole, and

The synthesis of novel 2-aminobenzimidazole (AmBI) [13, 14] derivatives and


Hypoglycemic effect of the newly obtained derivatives was initially studied in rats upon intraperitoneal administration at a dose of 50 mg/kg. Blood sampling was carried out 4 hours after treatment with test compounds. Blood glucose concentration was determined with the glucose oxidase method using a commercial Glucose FKD kit [31]. The ratio of glucose concentrations in the blood plasma of the experimental and control group animals served as an indicator of hypoglycemic

It was found that among condensed benzimidazole derivatives, a number of substances exceeded hypoglycemic activity of metformin, which served as a refer-

In order to identify the most promising antidiabetic substances using IT Microcosm [25, 26] and PASS computer systems [27, 28], a step-by-step detailed in silico analysis of the hypoglycemic properties of the new compounds was carried out. Programs DruLiTo [29] and QikProp [30] were employed to assess key ADME






gogues or insulin formulations at the late stages of the disease [9–11].

fused benzimidazole derivatives was carried out, including N9


2,3,4,10-tetrahydropyrimido[1,2-*a*]benzimidazoles (PrmBI) [22–24].


**74**

activity [32].

ence drug.

IT Microcosm [25, 26] and PASS [27, 28] computer systems were employed to determine the most promising chemical class of compounds. A training set of known hypoglycemic drugs and a library of tested benzimidazole derivatives were subjected to a consensus prediction of the level of hypoglycemic activity. The average informativity coefficient *KPr* was calculated and used as a metric for comparison of AmBI, N9 -ImBI, N1 -ImBI, N9 -DhImBI, N1 -DhImBI, and PrmBI derivatives. *KPr* value ranges from 0 for inactive compounds to 5 for highly active compounds.

According to value of *KPr*, the potential of benzimidazole derivatives classes as sources of substances with hypoglycemic activity decreases in the following order: N9 -DhImBI (*KPr* = 4.50) > PrmBI (*KPr* = 4.25) > AmBI (*KPr* = 2.50) > N1 -ImBI (*KPr* = 2.00) > N1 -DhImBI (*KPr* = 1.25) > N9 -ImBI (*KPr* = 0.25) [33].

Thus, it was shown that N9 -2,3-dihydroimidazo[1,2-*a*]benzimidazole and 2,3,4,10-tetrahydropyrimido[1,2-*a*]benzimidazole derivatives have the most promising blood glucose lowering activity. That is, these tricyclic structures containing embedded guanidine group turned out to be more active than 2-aminobenzimidazole derivatives.

Subsequently, employing substructural analysis [34] and analysis via median [35] and supremal [36] estimates, the class of N9 -2,3-dihydroimidazo[1,2-*a*]benzimidazoles was selected as the most promising for the development of hypoglycemic compounds (**Figures 1** and **2**). It was shown that this scaffold is more preferable than 2,3,4,10-tetrahydropyrimido[1,2-a]benzimidazole.

Substructural analysis [34] of the level of hypoglycemic activity among the N9 -2,3 dihydroimidazo[1,2-*a*]benzimidazole derivatives allowed us to reveal a chemical feature (substituent) that largely determines high hypoglycemic activity—diethylaminoethyl substituent at the N9 atom of the N9 -DhImBI scaffold.

According to the frequency analysis of physicochemical parameters [37] of experimentally studied derivatives of N9 -DhImBI scaffold, a significant feature of high hypoglycemic activity was revealed—a charge on the internal imidazole cycle of the condensed system, namely, *Q(Imid1)cs* ≥ −0.109, which is a characteristic for compound RU-254 (diabenol).

Taken together, the results of a complex consensus in silico analysis of the hypoglycemic activity of six classes of benzimidazole derivatives revealed 9-diethylaminoethyl-2,3-dihydroimidazo[1,2-*a*]benzimidazole dihydrochloride (RU-254, diabenol) as the most promising highly active compound.

To assess the feasibility of a further study of the pharmacological properties of compound RU-254, we calculated parameters of drug-likeliness and ADME properties (absorption, distribution, metabolism, excretion) for RU-254 and reference antidiabetic drugs metformin and glibenclamide.

#### **Figure 1.**

*Informativity coefficients describing the influence of basic benzimidazole structure on high hypoglycemic activity level (according to the substructural analysis).*

#### **Figure 2.**

Using the DruLiTo program [29], it was found that diabenol satisfies the boundary conditions of all eight drug-likeliness filters, while metformin and glibenclamide correspond only for two of them.

Water solubility, serum albumin binding parameters, cellular permeability, and absorbability through the gastrointestinal tract for the three aforementioned substances were calculated with QikProp program [30]. A comparative analysis of the obtained characteristics showed that water solubility and the degree of binding to serum albumin of diabenol are higher than that of glibenclamide and lower than that of metformin. Indicators of bioavailability and absorbability through the gastrointestinal tract in diabenol are higher than that of glibenclamide and metformin. Thus, in terms of the total pharmacokinetic characteristics calculated in the QikProp program, diabenol is superior to metformin and glibenclamide. It should be noted that the calculated values of pharmacokinetic parameters of all three compounds are in the ranges that are recognized as appropriate for drug molecules.

Summarizing the results of the evaluation of ADME properties obtained using two computational approaches, it can be argued that diabenol in regard of its calculated drug-like and pharmacokinetic characteristics is not inferior to metformin and glibenclamide and is a very promising substance for performing advanced preclinical studies.

**77**

**2.3 Preclinical studies**

*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

desired dihydrochloride [21].

Synthesis of 9-diethylaminoethyl-2,3-dihydroimidazo[1,2-*a*]benzimidazole dihydrochloride (RU-254, diabenol) is readily realized through condensation of 2-amino-1-diethylaminoethylbenzimidazole with an excess of dibromoethane and

**Example.** A stirred suspension of 69.6 g (0.3 mol) of 2-amino-1-diethylaminoethylbenzimidazole (I) in 104 ml (1.2 mol) of dibromoethane is gently heated in a glycerin bath. At 60–70°C, the initial amine dissolves completely, and at 100–105°C, an exothermic cyclization reaction occurs (the bath is set aside at the beginning), accompanied by strong boiling up of the reaction mass while temperature rises to 140°C and a heavy colorless precipitate begins to form. After 5–7 minutes, the reaction virtually ends, and, in order to complete it, the mixture is heated for an additional 20 minutes at 140–145°C. After that, 80 ml of DMF are added to the thick mass with vigorous stirring, and the mixture is heated for another 10–15 minutes. Cooling to 20–25°C, filtering the precipitate, and washing with DMF (3 × 20 ml) and acetone (3 × 25 ml) afford 106 g of dihydrobromide (II) in 84% yield. The latter is dissolved in 230 ml of water and boiled for 10 minutes with 3–5 g of activated carbon. Carbon is filtered off, and the filtrate after cooling is brought to pH 10 with 40% sodium hydroxide solution. The light yellow oil (III) which separates on the surface is extracted with toluene. The toluene extracts are washed with water and dried with anhydrous potassium carbonate. The desiccant is filtered off and washed with toluene. Combined toluene fractions are acidified by gradual addition of a saturated solution of hydrogen chloride in 2-propanol to pH 1. The heavy colorless precipitate of dihydrochloride (IV) is filtered off after 4–5 hours at 20–25°C, washed with acetone, and dried at 100–110°С for 2–3 hours to a constant weight. Yield is 75 g (90.9%) from dihydrobromide (III) and 75% from the initial amine (II). The crude product can be recrystallized from 2-propanol to pharmacopeia grade purity.

Diabenol had a pronounced hypoglycemic effect and antihyperglycemic activity in carbohydrate tolerance tests performed on white outbred rats and rabbits. The compound studied showed a marked decrease of glycemia in animals with impaired glucose tolerance (in rats with severe streptozotocin diabetes and insulin resistance syndrome), in rats with alloxan diabetes, and in rabbits with acute insulin deficiency, induced with administration of anti-insulin guinea pig serum. In experiments on pancreatomic dogs, diabenol did not reduce blood glucose but enhanced the hypoglycemic effect of exogenously administered insulin [38–40],

Detailed study of antidiabetic action revealed not only pancreatotropic but also extrapancreatic components of diabenol action. Its pancreatotropic effect is determined by the enhancement of phase 1 insulin secretion, especially in glucose-

thus confirming insulin-mediated mechanism of action.

stimulated conditions (**Figures 3** and **4**).

2,3-dihydroimidazo[1,2-*a*]benzimidazole dihydrobromide to the base and the

subsequent transformation of the resulting of 9-diethylaminoethyl-

**2.2 Synthesis**

*Supremal evaluations of the effect of basic benzimidazole structure on high hypoglycemic activity level.*

#### **2.2 Synthesis**

*Chemistry and Applications of Benzimidazole and its Derivatives*

Using the DruLiTo program [29], it was found that diabenol satisfies the bound-

ary conditions of all eight drug-likeliness filters, while metformin and gliben-

*Supremal evaluations of the effect of basic benzimidazole structure on high hypoglycemic activity level.*

*Informativity coefficients describing the influence of basic benzimidazole structure on high hypoglycemic* 

Water solubility, serum albumin binding parameters, cellular permeability, and absorbability through the gastrointestinal tract for the three aforementioned substances were calculated with QikProp program [30]. A comparative analysis of the obtained characteristics showed that water solubility and the degree of binding to serum albumin of diabenol are higher than that of glibenclamide and lower than that of metformin. Indicators of bioavailability and absorbability through the gastrointestinal tract in diabenol are higher than that of glibenclamide and metformin. Thus, in terms of the total pharmacokinetic characteristics calculated in the QikProp program, diabenol is superior to metformin and glibenclamide. It should be noted that the calculated values of pharmacokinetic parameters of all three compounds are in the ranges that are recognized as appropriate for drug molecules. Summarizing the results of the evaluation of ADME properties obtained using two computational approaches, it can be argued that diabenol in regard of its calculated drug-like and pharmacokinetic characteristics is not inferior to metformin and glibenclamide and is a very promising substance for performing advanced preclinical studies.

clamide correspond only for two of them.

*activity level (according to the substructural analysis).*

**76**

**Figure 2.**

**Figure 1.**

Synthesis of 9-diethylaminoethyl-2,3-dihydroimidazo[1,2-*a*]benzimidazole dihydrochloride (RU-254, diabenol) is readily realized through condensation of 2-amino-1-diethylaminoethylbenzimidazole with an excess of dibromoethane and subsequent transformation of the resulting of 9-diethylaminoethyl-2,3-dihydroimidazo[1,2-*a*]benzimidazole dihydrobromide to the base and the desired dihydrochloride [21].

**Example.** A stirred suspension of 69.6 g (0.3 mol) of 2-amino-1-diethylaminoethylbenzimidazole (I) in 104 ml (1.2 mol) of dibromoethane is gently heated in a glycerin bath. At 60–70°C, the initial amine dissolves completely, and at 100–105°C, an exothermic cyclization reaction occurs (the bath is set aside at the beginning), accompanied by strong boiling up of the reaction mass while temperature rises to 140°C and a heavy colorless precipitate begins to form. After 5–7 minutes, the reaction virtually ends, and, in order to complete it, the mixture is heated for an additional 20 minutes at 140–145°C. After that, 80 ml of DMF are added to the thick mass with vigorous stirring, and the mixture is heated for another 10–15 minutes. Cooling to 20–25°C, filtering the precipitate, and washing with DMF (3 × 20 ml) and acetone (3 × 25 ml) afford 106 g of dihydrobromide (II) in 84% yield. The latter is dissolved in 230 ml of water and boiled for 10 minutes with 3–5 g of activated carbon. Carbon is filtered off, and the filtrate after cooling is brought to pH 10 with 40% sodium hydroxide solution. The light yellow oil (III) which separates on the surface is extracted with toluene. The toluene extracts are washed with water and dried with anhydrous potassium carbonate. The desiccant is filtered off and washed with toluene. Combined toluene fractions are acidified by gradual addition of a saturated solution of hydrogen chloride in 2-propanol to pH 1. The heavy colorless precipitate of dihydrochloride (IV) is filtered off after 4–5 hours at 20–25°C, washed with acetone, and dried at 100–110°С for 2–3 hours to a constant weight. Yield is 75 g (90.9%) from dihydrobromide (III) and 75% from the initial amine (II). The crude product can be recrystallized from 2-propanol to pharmacopeia grade purity.

#### **2.3 Preclinical studies**

Diabenol had a pronounced hypoglycemic effect and antihyperglycemic activity in carbohydrate tolerance tests performed on white outbred rats and rabbits. The compound studied showed a marked decrease of glycemia in animals with impaired glucose tolerance (in rats with severe streptozotocin diabetes and insulin resistance syndrome), in rats with alloxan diabetes, and in rabbits with acute insulin deficiency, induced with administration of anti-insulin guinea pig serum. In experiments on pancreatomic dogs, diabenol did not reduce blood glucose but enhanced the hypoglycemic effect of exogenously administered insulin [38–40], thus confirming insulin-mediated mechanism of action.

Detailed study of antidiabetic action revealed not only pancreatotropic but also extrapancreatic components of diabenol action. Its pancreatotropic effect is determined by the enhancement of phase 1 insulin secretion, especially in glucosestimulated conditions (**Figures 3** and **4**).

#### **Figure 3.**

*Effect of diabenol (10 mg/kg, intravenously) in blood insulin levels during glucose tolerance test (1 g/kg) in cats.*

Diabenol increases the insulin-dependent glucose uptake in muscles of rat diaphragm. Under conditions of alloxan-induced diabetes in rats, diabenol restored liver glycogen content and glycolysis rate and inhibited glycogenolysis in insulindependent organs and tissues (liver, striated muscles) while having no significant effect on these parameters in kidneys, which are insulin-independent organs [38].

It could be assumed that increased insulinotropic effect of diabenol is associated with a possible incretinomimetic effect. Studies [41] showed the ability of diabenol to inhibit the incretin-degrading DPP-4 enzyme, leading to a modulation of the insulin response. In our studies, diabenol also inhibited DPP-4, but in substantially higher concentrations (IC50 1.35–2.05 mM), which cannot be achieved in the animal's body. Along with that, a 28-day administration of diabenol to rats with streptozotocin-induced diabetes was found to slightly and statistically insignificantly decrease the plasma activity of DPP-4 [42], which could be attributed to the action of its metabolites.

**Figure 4.** *Effect of diabenol (10 mg/kg, intravenously) on the basal portal vein blood glucose levels in cats [38].*

**79**

*\**

**Table 1.**

**Experimental groups**

Streptozotocinnicotinamideinduced diabetes

Streptozotocinnicotinamide induced diabetes +25 mg/kg diabenol

*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

area of the pancreatic β-cells (**Table 1**) [44].

activity of β-cells under the influence of diabenol [44].

**Islet area (μm2**

*Statistically significant compared with the intact control group.*

*induced diabetic rats after administration of diabenol for 21 days (M ± m) [44].*

**) Volume** 

**fraction of islets (%)**

Intact control 15,448.2 ± 9819.4 11.0 ± 1.2 63.8 ± 7.2 74.2 ± 5.6 26.4 ± 3.7

*Morphometric parameters of pancreatic islets in splenic region of the pancreas of streptozotocin-nicotinamide-*

**Relative number of β-cells (%)**

12,801.5 ± 11,252.3 5.1 ± 2.3\* 47.1 ± 3.5\* 55.3 ± 6.1\* 30.5 ± 6.2

9559.6 ± 11,513.8 7.5 ± 1.5 54.3 ± 9.5 63.1 ± 4.6 25.4 ± 6.4

**Volume fraction of β-cells (%)**

**Nuclear area of β-cells (μm2 )**

Long-term administration of diabenol to streptozotocin-nicotinamide-induced diabetic rats allowed us to obtain interesting and valuable results. Oral administration of diabenol in a dose of 25 mg/kg for 4 weeks reduced blood glucose levels and volume of consumed liquid by more than 2 times, and level of glycated hemoglobin by 2.2%, and increased the content of C-peptide [43]. Moreover, diabenol administration resulted in a significant increase in the average number of islets of Langerhans in the splenic region of the pancreas and a significant increase in the

The studied compound did not affect the apoptosis index (fraction of caspase-3 positive cells) and the proliferation index (PCNA-positive and Ki-67-positive cells) of the endocrinocytes of Langerhans islets. That is, diabenol had a cytoprotective effect on the cells. These data confirm the possibility of increasing the synthetic

Given the complex nature of type 2 diabetes mellitus and aiming to increase the effectiveness of antidiabetic therapy in clinical practice, combination drugs (fixed combinations) are actively used to simultaneously target several key pathogenesis factors of the underlying disease or its complications [7, 9]. The optimal ratios of diabenol with metformin (1:4) and glibenclamide (5:1) were determined in experiments on rats with streptozotocin-nicotinamide-induced diabetes. Administration of these fixed combinations proved to be effective in terms of key metabolic markers, including blood glucose level, dynamics of glycated hemoglobin reduction, C-peptide level, and recovery of pancreatic β-cells, and has a positive impact on carbohydrate metabolism—liver glycogen content and glycogenolysis [45]. A very important aspect of diabetes pathogenesis is the activation of lipid peroxidation, which facilitates development of β-cell dysfunction and peripheral insulin resistance [11]. In order to address this issue in clinical practice, combination therapy regimens for diabetes have begun to include an antioxidant, for example, lipoic acid [46]. At the first stage of our study, the direct effect of some antidiabetic agents on free radical processes was studied in vitro. It was established that diabenol is a scavenger of superoxide anion, hydroxyl, and peroxyl radicals; rosiglitazone is active only against the superoxide anion, gliclazide has an antiradical effect in experiments with DPPH, and metformin and glibenclamide were unable to interfere with these processes. At the same time, the established direct antioxidant properties of some studied drugs are difficult to be expected in vivo, since they require relatively high concentrations to exert antiradical activity in vitro [47, 48].

#### *Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

*Chemistry and Applications of Benzimidazole and its Derivatives*

**78**

**Figure 4.**

action of its metabolites.

**Figure 3.**

*cats.*

*Effect of diabenol (10 mg/kg, intravenously) on the basal portal vein blood glucose levels in cats [38].*

Diabenol increases the insulin-dependent glucose uptake in muscles of rat diaphragm. Under conditions of alloxan-induced diabetes in rats, diabenol restored liver glycogen content and glycolysis rate and inhibited glycogenolysis in insulindependent organs and tissues (liver, striated muscles) while having no significant effect on these parameters in kidneys, which are insulin-independent organs [38]. It could be assumed that increased insulinotropic effect of diabenol is associated with a possible incretinomimetic effect. Studies [41] showed the ability of diabenol to inhibit the incretin-degrading DPP-4 enzyme, leading to a modulation of the insulin response. In our studies, diabenol also inhibited DPP-4, but in substantially higher concentrations (IC50 1.35–2.05 mM), which cannot be achieved in the animal's body. Along with that, a 28-day administration of diabenol to rats with streptozotocin-induced diabetes was found to slightly and statistically insignificantly decrease the plasma activity of DPP-4 [42], which could be attributed to the

*Effect of diabenol (10 mg/kg, intravenously) in blood insulin levels during glucose tolerance test (1 g/kg) in* 

Long-term administration of diabenol to streptozotocin-nicotinamide-induced diabetic rats allowed us to obtain interesting and valuable results. Oral administration of diabenol in a dose of 25 mg/kg for 4 weeks reduced blood glucose levels and volume of consumed liquid by more than 2 times, and level of glycated hemoglobin by 2.2%, and increased the content of C-peptide [43]. Moreover, diabenol administration resulted in a significant increase in the average number of islets of Langerhans in the splenic region of the pancreas and a significant increase in the area of the pancreatic β-cells (**Table 1**) [44].

The studied compound did not affect the apoptosis index (fraction of caspase-3 positive cells) and the proliferation index (PCNA-positive and Ki-67-positive cells) of the endocrinocytes of Langerhans islets. That is, diabenol had a cytoprotective effect on the cells. These data confirm the possibility of increasing the synthetic activity of β-cells under the influence of diabenol [44].

Given the complex nature of type 2 diabetes mellitus and aiming to increase the effectiveness of antidiabetic therapy in clinical practice, combination drugs (fixed combinations) are actively used to simultaneously target several key pathogenesis factors of the underlying disease or its complications [7, 9]. The optimal ratios of diabenol with metformin (1:4) and glibenclamide (5:1) were determined in experiments on rats with streptozotocin-nicotinamide-induced diabetes. Administration of these fixed combinations proved to be effective in terms of key metabolic markers, including blood glucose level, dynamics of glycated hemoglobin reduction, C-peptide level, and recovery of pancreatic β-cells, and has a positive impact on carbohydrate metabolism—liver glycogen content and glycogenolysis [45].

A very important aspect of diabetes pathogenesis is the activation of lipid peroxidation, which facilitates development of β-cell dysfunction and peripheral insulin resistance [11]. In order to address this issue in clinical practice, combination therapy regimens for diabetes have begun to include an antioxidant, for example, lipoic acid [46]. At the first stage of our study, the direct effect of some antidiabetic agents on free radical processes was studied in vitro. It was established that diabenol is a scavenger of superoxide anion, hydroxyl, and peroxyl radicals; rosiglitazone is active only against the superoxide anion, gliclazide has an antiradical effect in experiments with DPPH, and metformin and glibenclamide were unable to interfere with these processes. At the same time, the established direct antioxidant properties of some studied drugs are difficult to be expected in vivo, since they require relatively high concentrations to exert antiradical activity in vitro [47, 48].


*\* Statistically significant compared with the intact control group.*

#### **Table 1.**

*Morphometric parameters of pancreatic islets in splenic region of the pancreas of streptozotocin-nicotinamideinduced diabetic rats after administration of diabenol for 21 days (M ± m) [44].*

A further study [48] determined the optimal ratios for a combination of diabenol and lipoic acid (2.8: 1 and 5.6: 1). Its activity was studied in a streptozotocinnicotinamide-induced diabetic rat model. It was established that this combination possesses a more pronounced antidiabetic effect than monotherapy with diabenol. The more important finding of this study is a significantly reduced content of lipid peroxidation products in the liver, pancreas, and kidneys. In the pancreas under streptozotocin intoxication conditions, β-cells were significantly preserved by the combined treatment with diabenol and lipoic acid.

It is known that diabetes is associated with the increased thrombogenic potential of the blood and impaired rheology properties [49]. This effect is attributed not only to hyperosmolarity of the blood due to hyperglycemia but also to an increased aggregation of platelets and red blood cells. Among the antidiabetic agents used in clinical practice, only gliclazide has a direct inhibitory effect on platelets [8]. For other drugs, a similar effect is observed only with prolonged therapy. Given the fact that diabetes increases the frequency of thrombosis events, we studied the effect of diabenol on aggregation properties of platelets and red blood cells and its influence on microcirculation in experimental diabetes.

It was established that diabenol, both in vitro and in the conditions of the whole organism, has an antiplatelet activity. Probably, the effect on functional activity of platelets is determined by the influence of diabenol on balance of prostacyclin and thromboxane A2 systems. Diabenol showed an antithrombogenic effect on the model of thrombosis of the carotid induced by electric current and in systemic adrenaline-collagen thrombosis, exceeding the activity of gliclazide [50–52].

Diabenol reduced the aggregability and increased the erythrocyte deformability in normal conditions and, more profoundly, in experimental diabetes. Using fluorescent probes, it has been shown that diabenol was able to increase electronegativity and reduces the microviscosity of the red blood cells membrane, which results in the increase in their deformability [53–56].

Amelioration of thrombogenic potential and blood viscosity gives diabenol ability to enhance the survival of skin graft (a model of the diabetic foot) in both intact and alloxan-induced diabetic animals [57].

Toxicological study of the diabenol pharmaceutical substance and the dosage form (tablets containing 0.2 g of the active ingredient) involved acute and chronic toxicity, examination of cumulative properties, immunotoxicity, effects on carcinogenesis, and transplacental action. The therapeutic dose of the drug had no adverse effects. Subtoxic doses of diabenol lead to a sharp decrease in pancreatic β-cell secretion along with platelet hemorrhages.

Upon long-term administration of diabenol to low-cancer NMRI mice, transgenic HER-2/neu mice, and LIO rats with drinking water, no toxic or carcinogenic effect was observed. An interesting fact has been demonstrated, that is, in NMRI mice, diabenol delayed the development of age-related disorders of the extra function and increased the life span of animals. The drug inhibited the occurrence of spontaneous tumors, reduced the incidence of malignant lymphomas, and inhibited the onset and development of colon cancer induced with 1,2-dimethylhydrazine in rats. The authors of this study conclude that diabenol has an anticarcinogenic and geroprotective effect. Hence, both diabenol and metformin contain a guanidine group in their structure and exert experimentally proven antitumor effect and geroprotective activity [58–60].

The data on the pharmacokinetic study of diabenol established the values of the drug half-life and average retention time, which suggest that the substance undergoes significantly rapid elimination. The drug penetrates well into organs and tissues, especially in those with a high degree of vascularization. An important role in the processes of elimination of a compound is played by processes of its metabolism [61].

**81**

*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

izing coagulation hemostasis [62, 63].

and geroprotective effect in rodents.

to tricyclic structures led us to the identification of N9

agents with high glucose lowering activity were identified.

multi-target action but are pleiotropic effects of diabenol.

Clinical studies of diabenol were conducted on 180 patients with type 2 diabetes.

Clinical and laboratory parameters of patients allowed to conclude that diabenol administered in a dose of 0.4 g per day for 3 months had no adverse or toxic effects [63].

Based on the studies performed, it can be stated that aminobenzimidazole is a universal privileged substructure that can be used as a source for development of novel antidiabetic agents. Introduction of structural modifications by means of transition

benzimidazole as an optimal scaffold, and in this chemical class of compounds,

As a result, RU-254, or diabenol, was developed as the most active compound. Both in the experimental and clinical settings, it restores insulin secretion and ameliorates peripheral tissue glucose uptake. Another important effect of diabenol that has been established experimentally and confirmed in clinical studies is a reduction of thrombogenic potential and viscosity of blood. It has been demonstrated that diabenol, much like the biguanide derivative metformin, exerts an anticarcinogenic

Thus, a new cyclic aminobenzimidazole derivative, diabenol, containing a guanidine group, combines pharmacological effects characteristic for both biguanide derivatives (reduction of hyperglycemia and liver glycogenolysis, improved glucose tolerance, anticarcinogenic and geroprotective effects) and for sulfonylurea derivative gliclazide (restoring insulin secretion, antiplatelet, and antiradical activity). It is quite possible that all of the identified effects are not a manifestation of the


The drug was administered orally in solid dosage form (0.2 g tablets) 2 times a day. The study design was a randomized controlled comparative study of efficacy, tolerability, and safety. Glidiab (gliclazide) served as a reference drug. The course of diabenol administration ameliorated both fasting and postprandial hyperglycemia, reduced glycated hemoglobin level by 1.1% at the end of the third month, and increased postprandial insulin levels. Diabenol reduced platelet aggregation, increased erythrocyte deformability, and reduced their aggregability, thus normal-

**2.4 Clinical studies**

**3. Conclusions**

#### **2.4 Clinical studies**

*Chemistry and Applications of Benzimidazole and its Derivatives*

combined treatment with diabenol and lipoic acid.

on microcirculation in experimental diabetes.

the increase in their deformability [53–56].

and alloxan-induced diabetic animals [57].

secretion along with platelet hemorrhages.

geroprotective activity [58–60].

A further study [48] determined the optimal ratios for a combination of diabenol and lipoic acid (2.8: 1 and 5.6: 1). Its activity was studied in a streptozotocinnicotinamide-induced diabetic rat model. It was established that this combination possesses a more pronounced antidiabetic effect than monotherapy with diabenol. The more important finding of this study is a significantly reduced content of lipid peroxidation products in the liver, pancreas, and kidneys. In the pancreas under streptozotocin intoxication conditions, β-cells were significantly preserved by the

It is known that diabetes is associated with the increased thrombogenic potential

It was established that diabenol, both in vitro and in the conditions of the whole organism, has an antiplatelet activity. Probably, the effect on functional activity of platelets is determined by the influence of diabenol on balance of prostacyclin and thromboxane A2 systems. Diabenol showed an antithrombogenic effect on the model of thrombosis of the carotid induced by electric current and in systemic adrenaline-collagen thrombosis, exceeding the activity of gliclazide [50–52].

Diabenol reduced the aggregability and increased the erythrocyte deformability in normal conditions and, more profoundly, in experimental diabetes. Using fluorescent probes, it has been shown that diabenol was able to increase electronegativity and reduces the microviscosity of the red blood cells membrane, which results in

Amelioration of thrombogenic potential and blood viscosity gives diabenol ability to enhance the survival of skin graft (a model of the diabetic foot) in both intact

Toxicological study of the diabenol pharmaceutical substance and the dosage form (tablets containing 0.2 g of the active ingredient) involved acute and chronic toxicity, examination of cumulative properties, immunotoxicity, effects on carcinogenesis, and transplacental action. The therapeutic dose of the drug had no adverse effects. Subtoxic doses of diabenol lead to a sharp decrease in pancreatic β-cell

Upon long-term administration of diabenol to low-cancer NMRI mice, transgenic HER-2/neu mice, and LIO rats with drinking water, no toxic or carcinogenic effect was observed. An interesting fact has been demonstrated, that is, in NMRI mice, diabenol delayed the development of age-related disorders of the extra function and increased the life span of animals. The drug inhibited the occurrence of spontaneous tumors, reduced the incidence of malignant lymphomas, and inhibited the onset and development of colon cancer induced with 1,2-dimethylhydrazine in rats. The authors of this study conclude that diabenol has an anticarcinogenic and geroprotective effect. Hence, both diabenol and metformin contain a guanidine group in their structure and exert experimentally proven antitumor effect and

The data on the pharmacokinetic study of diabenol established the values of the drug half-life and average retention time, which suggest that the substance undergoes significantly rapid elimination. The drug penetrates well into organs and tissues, especially in those with a high degree of vascularization. An important role in the processes of elimination of a compound is played by processes of its metabolism [61].

of the blood and impaired rheology properties [49]. This effect is attributed not only to hyperosmolarity of the blood due to hyperglycemia but also to an increased aggregation of platelets and red blood cells. Among the antidiabetic agents used in clinical practice, only gliclazide has a direct inhibitory effect on platelets [8]. For other drugs, a similar effect is observed only with prolonged therapy. Given the fact that diabetes increases the frequency of thrombosis events, we studied the effect of diabenol on aggregation properties of platelets and red blood cells and its influence

**80**

Clinical studies of diabenol were conducted on 180 patients with type 2 diabetes. The drug was administered orally in solid dosage form (0.2 g tablets) 2 times a day. The study design was a randomized controlled comparative study of efficacy, tolerability, and safety. Glidiab (gliclazide) served as a reference drug. The course of diabenol administration ameliorated both fasting and postprandial hyperglycemia, reduced glycated hemoglobin level by 1.1% at the end of the third month, and increased postprandial insulin levels. Diabenol reduced platelet aggregation, increased erythrocyte deformability, and reduced their aggregability, thus normalizing coagulation hemostasis [62, 63].

Clinical and laboratory parameters of patients allowed to conclude that diabenol administered in a dose of 0.4 g per day for 3 months had no adverse or toxic effects [63].

#### **3. Conclusions**

Based on the studies performed, it can be stated that aminobenzimidazole is a universal privileged substructure that can be used as a source for development of novel antidiabetic agents. Introduction of structural modifications by means of transition to tricyclic structures led us to the identification of N9 -2,3-dihydroimidazo[1,2-*a*] benzimidazole as an optimal scaffold, and in this chemical class of compounds, agents with high glucose lowering activity were identified.

As a result, RU-254, or diabenol, was developed as the most active compound. Both in the experimental and clinical settings, it restores insulin secretion and ameliorates peripheral tissue glucose uptake. Another important effect of diabenol that has been established experimentally and confirmed in clinical studies is a reduction of thrombogenic potential and viscosity of blood. It has been demonstrated that diabenol, much like the biguanide derivative metformin, exerts an anticarcinogenic and geroprotective effect in rodents.

Thus, a new cyclic aminobenzimidazole derivative, diabenol, containing a guanidine group, combines pharmacological effects characteristic for both biguanide derivatives (reduction of hyperglycemia and liver glycogenolysis, improved glucose tolerance, anticarcinogenic and geroprotective effects) and for sulfonylurea derivative gliclazide (restoring insulin secretion, antiplatelet, and antiradical activity). It is quite possible that all of the identified effects are not a manifestation of the multi-target action but are pleiotropic effects of diabenol.

#### **Author details**

Alexander A. Spasov1 , Pavel M. Vassiliev1 \*, Vera A. Anisimova<sup>2</sup> and Olga N. Zhukovskaya<sup>2</sup>

1 Volgograd State Medical University, Volgograd, Russia

2 Research Institute for Physical and Organic Chemistry, South Federal University, Rostov-on-Don, Russia

\*Address all correspondence to: pvassiliev@mail.ru

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**83**

*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

[1] Blaslov K, Naranda FS, Kruljac I, Renar IP. Treatment approach to type 2 diabetes: Past, present and future. World Journal of Diabetes. 2018;**9**:209-219.

[10] Ametov AS, Kondratyeva LV, Prudnikova MA. Metformin—More than the gold standard. In: Ametov AS, editor. Type 2 Diabetes. Problems and Solutions. Moscow: GEOTAR-Media;

[11] Balabolkin MI, Klebanova EM, Kreminskaya VM. Treatment of

Diabetes Mellitus and Its Complications. Moscow: Medicine; 2005. pp. 288-304

[12] Caprio S, Ray TK, Boden G, et al. Improvement of metabolic control in diabetic patients during mebendazole administration: Preliminary studies.

[13] Anisimova VA, Koshchienko YV, Pyatin BM et al. The Method of Obtaining Derivatives of 2-Amino-1-Aminoalkylbenzimidazoles. USSR Authorship certificate №. 1149592.

Diabetologia. 1984;**27**:52-55

Bulletin of Inventions. 1984

[14] Anisimova VA, Spasov AA, Kosolapov VA, et al. Synthesis and pharmacological activity of 3-(N,N-disubstituted)acetamide-1-R-2-aminobenzimidazolium

Zhurnal. 2012;**46**:6-10. DOI:

benzimidazoles. Khimiya Geterotsiklicheskikh Soedinenii.

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imidazo[1,2-a]benzimidazoles. Khimiko-Farmatsevticheskii Zhurnal.

[17] Simonov AM, Anisimova VA, Borisova TA. Preparation of imidazo[1,2-a]benzimidazole derivatives from 1-alkyl-,

Kucheryavenko AF, et al. Synthesis and pharmacological activity of 2-(hetaryl)

2015. pp. 202-256

[2] Dedov II, Shestakova MV. Diabetes Mellitus: Diagnosis, Treatment, Prevention. Moscow: Medical Information Agency; 2011. p. 808

combination therapy of type 2 diabetes mellitus. Bulletin of Volgograd State Medical University. 2011;**1**:8-12

[4] Spasov AA, Petrov VI, Chepliaeva NI, Lenskaya KV. Fundamental bases of search of medicines for therapy of a diabetes mellitus type 2. Vestnik Rossiĭskoĭ Akademii Meditsinskikh Nauk. 2013;**2**:43-49. DOI: 10.15690/

[5] Tkachuk VA, Vorotnikov AV. Molecular mechanisms of insulin resistance. Diabetes Mellitus. 2014;**2**: 29-40. DOI: 10.14341/DM2014229-40

[6] Snel M, Jonker JT, Schoones J, Lamb H, de Roos A, Pijl H, et al. Ectopic fat and insulin resistance: Pathophysiology and effect of diet and lifestyle interventions. International Journal of Endocrinology. 2012;**2012**:1-18. DOI: 10.1155/2012/983814

[7] Steinberg GR, Kemp BE. AMPK in health and disease. Physiological Reviews. 2009;**89**:1025-1078. DOI: 10.1152/physrev.00011.2008

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vramn.v68i2.548

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DOI: 10.4239/wjd.v9.i12.209

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*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

#### **References**

*Chemistry and Applications of Benzimidazole and its Derivatives*

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

2 Research Institute for Physical and Organic Chemistry, South Federal University,

\*, Vera A. Anisimova<sup>2</sup>

, Pavel M. Vassiliev1

1 Volgograd State Medical University, Volgograd, Russia

\*Address all correspondence to: pvassiliev@mail.ru

**82**

**Author details**

Alexander A. Spasov1

Rostov-on-Don, Russia

and Olga N. Zhukovskaya<sup>2</sup>

provided the original work is properly cited.

[1] Blaslov K, Naranda FS, Kruljac I, Renar IP. Treatment approach to type 2 diabetes: Past, present and future. World Journal of Diabetes. 2018;**9**:209-219. DOI: 10.4239/wjd.v9.i12.209

[2] Dedov II, Shestakova MV. Diabetes Mellitus: Diagnosis, Treatment, Prevention. Moscow: Medical Information Agency; 2011. p. 808

[3] Spasov AA, Chepurnova MV. Scientific approaches to the combination therapy of type 2 diabetes mellitus. Bulletin of Volgograd State Medical University. 2011;**1**:8-12

[4] Spasov AA, Petrov VI, Chepliaeva NI, Lenskaya KV. Fundamental bases of search of medicines for therapy of a diabetes mellitus type 2. Vestnik Rossiĭskoĭ Akademii Meditsinskikh Nauk. 2013;**2**:43-49. DOI: 10.15690/ vramn.v68i2.548

[5] Tkachuk VA, Vorotnikov AV. Molecular mechanisms of insulin resistance. Diabetes Mellitus. 2014;**2**: 29-40. DOI: 10.14341/DM2014229-40

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[11] Balabolkin MI, Klebanova EM, Kreminskaya VM. Treatment of Diabetes Mellitus and Its Complications. Moscow: Medicine; 2005. pp. 288-304

[12] Caprio S, Ray TK, Boden G, et al. Improvement of metabolic control in diabetic patients during mebendazole administration: Preliminary studies. Diabetologia. 1984;**27**:52-55

[13] Anisimova VA, Koshchienko YV, Pyatin BM et al. The Method of Obtaining Derivatives of 2-Amino-1-Aminoalkylbenzimidazoles. USSR Authorship certificate №. 1149592. Bulletin of Inventions. 1984

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[43] Spasov AA, Voronkova MP, Snigur GL. An experimental model of type 2 diabetes mellitus. Biomedicine. 2011;**3**:12-18

[44] Cheplyaeva NI. Pharmacological properties of the combination of hypoclycemic substance diabenol and α-lipoic acid [thesis]. Volgograd: Volgograd State Medical University; 2012

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in patient with type 2 diabetes. Diabetes Technology & Therapeutics. 2000;**2**:401-413

[47] Spasov AA, Kosolapov VA, Cheplyaeva NI. Antioxidant activity of oral hypoglycemic agents. Problems of Endocrinology. 2011;**4**:21-24

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[49] Morel O, Kossler L, Ohimann P. Diabetes and the platelets: Toward new therapentic paradigms for diabetic atherothrombosis. Atheroscherosis. 2010;**2**:367-376

[50] Kucheryavenko AF, Spasov AA, Petrov VI, Anisimova VA. Antiaggregant activity of a new benzimidazole derivative. Bulletin of Experimental Biology and Medicine. 2014;**156**:796- 798. DOI: 10.1007/s10517-014-2453-9

[51] Kucheryavenko AF, Spasov AA, Smirnov AV. Antithrombotic activity of a new hypoglycemic compound limiglidole in mouse model of cell thrombosis. Bulletin of Experimental Biology and Medicine. 2015;**159**:41-43. DOI: 10.1007/s10517-015-2885-x

[52] Spasov AA, Kucheryavenko AF, Chepurnova MV, Lenskaya KV. Antithrombotic activity of hypoglycemic agents. Regional Blood Circulation and Microcirculation. 2011;**10**:95-98

[53] Degtyarev AN, Kucheryavenko AF, Spasov AA, Ostrovskiy OV. Benzimidazole derivatives as the basis for the creation of drugs that affect the rheological properties of blood. In: Proceedings of Actual Problems of Experimental and Clinical Pharmacology; May 1999;

Saint-Petersburg. Saint-Petersburg: Politekhnika; 1999. p. 120

[54] Degtyarev AN, Ostrovskiy OV, Shakhova NI, Anisimova VA. Evaluation of the corrective effect of new compounds, benzimidazole derivatives, with the "increased viscosity syndrome". In: Proceedings of Actual Problems of Experimental and Clinical Pharmacology; May 1999; Saint-Petersburg. Saint-Petersburg: Politekhnika; 1999. p. 59

[55] Spasov AA, Petrov VI, Anisimova VA. New hypoglycemic agent with hemorheological properties. In: Proceedings of IVth Russian Diabetological Congress; 19-22 May 2008; Moscow. Moscow; 2008. p. 336

[56] Kucheryavenko AF, Spasov AA, Naumenko LV. The influence of new hypoglycemic agent limiglidol on the parameters of hemostasis in experimental diabetes. Problems of Endocrinology. 2015;**61**:51-56. DOI: 10.14341/probl201561151-56

[57] Vasilyeva SV, Galenko-Yaroshevskiy VP, Khropova TN, Tegay AV, Uvarov AV. The effect of imidazobenzimidazole derivatives RU-185 and RU-254 on the viability of the skin graft. Bulletin of Experimental Biology and Medicine. 2002;**2**:90

[58] Popovich IG, Zabezhinskiĭ MA, Anikin IV, Tyndyk ML, Spasov AA, Anisimov VN. Inhibition of 1,2-dimethylhydrazine-induced carcinogenesis in rat gut by the antidiabetic drug Diabenol. Voprosy Onkologii. 2004;**50**:562-566

[59] Popovich IG, Zabezhinskiĭ MA, Egormin PA, Anikin IV, Tyndyk ML, Semenchenko AV, et al. Effect of antidiabetic drug diabenol on parameters of biological age, life span and tumor development in NMRI and HER-2/neu mice. Advances in Gerontology. 2004;**15**:80-90

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*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

[60] Popovich IG, Zabezhinski MA, Egormin PA, Tyndyk ML, Anikin IV, Spasov AA, et al. Insulin in aging and cancer: Antidiabetic drug diabenol as geroprotector and anticarcinogen. The International Journal of Biochemistry & Cell Biology. 2005;**37**:1117-1129. DOI:

10.1016/j.biocel.2004.08.002

University. 1999;**5**:26-34

2008. p. 268

[62] Dedov II, Balabolkin MI, Spasov AA, Petrov VI. New domestic hypoglycemic agent with hemorheological properties—diabenol (clinical studies). In: Proceedings of IVth Russian Diabetological Congress; 19-22 May 2008; Moscow. Moscow;

[61] Spasov AA, Dudchenko GP, Smirnova LA, Gavrilova ES.

Pharmacodynamic and pharmacokinetic properties of the compound RU-254. Bulletin of Volgograd State Medical

[63] Petrov VI, Spasov AA, Nedogoda SV, Voronkova MP. Clinical efficacy of the drug diabenol in type 2 diabetes mellitus. In: Spasov AA, Petrov VI, Minkin VI, editors. Antidiabetic

Potential of Benzimidazoles. Volgograd:

VolgSMU; 2016. pp. 451-464

*Antidiabetogenic Features of Benzimidazoles DOI: http://dx.doi.org/10.5772/intechopen.84802*

*Chemistry and Applications of Benzimidazole and its Derivatives*

Saint-Petersburg. Saint-Petersburg:

[54] Degtyarev AN, Ostrovskiy OV, Shakhova NI, Anisimova VA. Evaluation of the corrective effect of new compounds, benzimidazole derivatives, with the "increased viscosity syndrome". In: Proceedings of Actual Problems of Experimental and Clinical Pharmacology; May 1999; Saint-Petersburg. Saint-Petersburg:

[55] Spasov AA, Petrov VI, Anisimova

[56] Kucheryavenko AF, Spasov AA, Naumenko LV. The influence of new hypoglycemic agent limiglidol on the parameters of hemostasis in experimental diabetes. Problems of Endocrinology. 2015;**61**:51-56. DOI:

[57] Vasilyeva SV, Galenko-Yaroshevskiy VP, Khropova TN, Tegay AV, Uvarov AV. The effect of imidazobenzimidazole derivatives RU-185 and RU-254 on the viability of the skin graft. Bulletin of Experimental Biology and Medicine.

[58] Popovich IG, Zabezhinskiĭ MA, Anikin IV, Tyndyk ML, Spasov AA,

[59] Popovich IG, Zabezhinskiĭ MA, Egormin PA, Anikin IV, Tyndyk ML, Semenchenko AV, et al. Effect of antidiabetic drug diabenol on parameters of biological age, life span and tumor development in NMRI and HER-2/neu mice. Advances in Gerontology. 2004;**15**:80-90

Anisimov VN. Inhibition of 1,2-dimethylhydrazine-induced carcinogenesis in rat gut by the antidiabetic drug Diabenol. Voprosy

Onkologii. 2004;**50**:562-566

10.14341/probl201561151-56

2002;**2**:90

Politekhnika; 1999. p. 120

Politekhnika; 1999. p. 59

VA. New hypoglycemic agent with hemorheological properties. In: Proceedings of IVth Russian Diabetological Congress; 19-22 May 2008; Moscow. Moscow; 2008. p. 336

in patient with type 2 diabetes. Diabetes Technology & Therapeutics.

[47] Spasov AA, Kosolapov VA,

Endocrinology. 2011;**4**:21-24

[48] Spasov AA, Kosolapov VA, Cheplyaeva NI. Comparative

Cheplyaeva NI. Antioxidant activity of oral hypoglycemic agents. Problems of

characteristics of antioxidant properties of hypoglycemic agents diabenol and gliclazide. Eksperimental'naia i Klinicheskaia Farmakologiia.

[49] Morel O, Kossler L, Ohimann P. Diabetes and the platelets: Toward new therapentic paradigms for diabetic atherothrombosis. Atheroscherosis.

[50] Kucheryavenko AF, Spasov AA, Petrov VI, Anisimova VA. Antiaggregant

[51] Kucheryavenko AF, Spasov AA, Smirnov AV. Antithrombotic activity of a new hypoglycemic compound limiglidole in mouse model of cell thrombosis. Bulletin of Experimental Biology and Medicine. 2015;**159**:41-43. DOI: 10.1007/s10517-015-2885-x

[52] Spasov AA, Kucheryavenko AF, Chepurnova MV, Lenskaya KV. Antithrombotic activity of hypoglycemic agents. Regional Blood Circulation and Microcirculation.

[53] Degtyarev AN, Kucheryavenko AF,

Spasov AA, Ostrovskiy OV. Benzimidazole derivatives as the basis for the creation of drugs that affect the rheological properties of blood. In: Proceedings of Actual Problems of Experimental and Clinical Pharmacology; May 1999;

2011;**10**:95-98

activity of a new benzimidazole derivative. Bulletin of Experimental Biology and Medicine. 2014;**156**:796- 798. DOI: 10.1007/s10517-014-2453-9

2000;**2**:401-413

2011;**74**:14-16

2010;**2**:367-376

**86**

[60] Popovich IG, Zabezhinski MA, Egormin PA, Tyndyk ML, Anikin IV, Spasov AA, et al. Insulin in aging and cancer: Antidiabetic drug diabenol as geroprotector and anticarcinogen. The International Journal of Biochemistry & Cell Biology. 2005;**37**:1117-1129. DOI: 10.1016/j.biocel.2004.08.002

[61] Spasov AA, Dudchenko GP, Smirnova LA, Gavrilova ES. Pharmacodynamic and pharmacokinetic properties of the compound RU-254. Bulletin of Volgograd State Medical University. 1999;**5**:26-34

[62] Dedov II, Balabolkin MI, Spasov AA, Petrov VI. New domestic hypoglycemic agent with hemorheological properties—diabenol (clinical studies). In: Proceedings of IVth Russian Diabetological Congress; 19-22 May 2008; Moscow. Moscow; 2008. p. 268

[63] Petrov VI, Spasov AA, Nedogoda SV, Voronkova MP. Clinical efficacy of the drug diabenol in type 2 diabetes mellitus. In: Spasov AA, Petrov VI, Minkin VI, editors. Antidiabetic Potential of Benzimidazoles. Volgograd: VolgSMU; 2016. pp. 451-464

Chapter 6

Abstract

anticancer drugs/agents

1. Introduction

in vitamin B12 [7].

be discussed.

89

Puranik Purushottamachar,

Development of Benzimidazole

Compounds for Cancer Therapy

A fact that is largely unknown in the lay press and even the scientific community is that today cancer kills more people worldwide than tuberculosis (TB), malaria,

heterocyclic aromatic organic compound considered to be a useful pharmacophore in a variety of impactful drugs. The purpose of this review is to highlight the benzimidazole-containing agents that are currently in clinical use or in clinical development as anticancer drugs. It is hoped that this review would function as comprehensive working reference of research accomplishment in the field of discovery and development of benzimidazole-based anticancer drugs.

Benzimidazole (1) (Figure 1) is used as the major scaffold or as a moiety on other scaffolds for the development of a variety of drugs [1–4]. The wide range of pharmacological activities of benzimidazole-containing agents are attributed to the unique fused benzene and imidazole rings, which can interact in a noncovalent manner with a range of biological targets due to the presence of an electron-rich aromatic system and the two hetero-nitrogen atoms [5, 6]. Because of the ability of benzimidazole derivative to interact with a variety of unrelated molecular targets, the term "privileged substructure/moiety" is ascribed to this unique azole agent. It is believed that the interest in benzimidazole chemistry and as a scaffold/moiety in the discovery and development of drugs arose from the discovery of the rare

Senthilmurugan Ramalingam and Vincent C.O. Njar

and human immunodeficiency virus (HIV) combined. Benzimidazole is a

Keywords: benzimidazole derivatives, privileged pharmacophore,

and most prominent benzimidazole compound in nature, N-ribosyl-

received FDA approval [8–10]. Two prominent benzimidazole agents,

dimethylbenzimidazole (2) (Figure 1), which serves as an axial ligand for cobalt

Although several benzimidazole derivatives have been approved for clinical use, including antiparasitic, antiulcer, antihypertensive, antihistaminic, and

antiemetic drugs [1–4], only one anticancer drug, bendamustine (3) (Figure 2), has

selumetinib (4) (Figure 2) [1, 6] and galeterone (5) (Figure 2) [11], that advanced to phase III clinical trials, but are yet to be approved as anticancer drugs, will also

#### Chapter 6

## Development of Benzimidazole Compounds for Cancer Therapy

Puranik Purushottamachar, Senthilmurugan Ramalingam and Vincent C.O. Njar

#### Abstract

A fact that is largely unknown in the lay press and even the scientific community is that today cancer kills more people worldwide than tuberculosis (TB), malaria, and human immunodeficiency virus (HIV) combined. Benzimidazole is a heterocyclic aromatic organic compound considered to be a useful pharmacophore in a variety of impactful drugs. The purpose of this review is to highlight the benzimidazole-containing agents that are currently in clinical use or in clinical development as anticancer drugs. It is hoped that this review would function as comprehensive working reference of research accomplishment in the field of discovery and development of benzimidazole-based anticancer drugs.

Keywords: benzimidazole derivatives, privileged pharmacophore, anticancer drugs/agents

#### 1. Introduction

Benzimidazole (1) (Figure 1) is used as the major scaffold or as a moiety on other scaffolds for the development of a variety of drugs [1–4]. The wide range of pharmacological activities of benzimidazole-containing agents are attributed to the unique fused benzene and imidazole rings, which can interact in a noncovalent manner with a range of biological targets due to the presence of an electron-rich aromatic system and the two hetero-nitrogen atoms [5, 6]. Because of the ability of benzimidazole derivative to interact with a variety of unrelated molecular targets, the term "privileged substructure/moiety" is ascribed to this unique azole agent. It is believed that the interest in benzimidazole chemistry and as a scaffold/moiety in the discovery and development of drugs arose from the discovery of the rare and most prominent benzimidazole compound in nature, N-ribosyldimethylbenzimidazole (2) (Figure 1), which serves as an axial ligand for cobalt in vitamin B12 [7].

Although several benzimidazole derivatives have been approved for clinical use, including antiparasitic, antiulcer, antihypertensive, antihistaminic, and antiemetic drugs [1–4], only one anticancer drug, bendamustine (3) (Figure 2), has received FDA approval [8–10]. Two prominent benzimidazole agents, selumetinib (4) (Figure 2) [1, 6] and galeterone (5) (Figure 2) [11], that advanced to phase III clinical trials, but are yet to be approved as anticancer drugs, will also be discussed.

2.1.1 Chemistry

cial production of 3.

Figure 4.

91

Bendamustine (3) 4-{5-[bis(2-chloroethyl)amino]-1-methylbenzimidazol-2-yl} butanoic acid was first synthesized via eight synthetic steps with an overall yield of 12% [13, 17]. However, Chen and colleagues have developed a new, more efficient, and cost-effective route focused on the use of sustainable chemistry for the synthesis of bendamustine hydrochloride, with the overall yield improved from 12 to 45% as outlined in Figure 4 [18]. This new synthesis is currently used for the commer-

Even though bendamustine is an alkylating agent, due to its ability to cause intra-strand and inter-strand cross-links between DNA bases, it has been reported that the DNA breaks induced by bendamustine are more extensive/durable than those induced by other alkylating agents, such as chlorambucil, cyclophosphamide, or carmustine [19–21]. In addition, the drug was shown to exhibit partial crossresistance to other alkylating agents. These data suggested that bendamustine may possess additional mechanisms of action. Indeed, a comprehensive study by Leoni and colleagues clearly demonstrated that bendamustine exhibits a distinct pattern of activities unrelated to other alkylating drugs. Using a variety of lymphoid cancer cell lines, the study concluded that mechanisms of action include induction of mitotic catastrophe, inhibition of mitotic checkpoints, and activation of DNAdamage stress response and apoptosis. Compared to other alkylating agents, bendamustine was shown to activate the base excision DNA repair pathway rather

Although bendamustine is approved for the treatment of a variety of lymphoid cancers, its activity has also been reported in several cancers, including cancers of small cell lung, breast, hepatic, bile duct, and head and neck. The studies by Chow and colleagues using leukemic cell lines in vitro or ex vivo cells from patients with

2.1.2 Summary of bendamustine's preclinical and clinical pharmacology

Development of Benzimidazole Compounds for Cancer Therapy

DOI: http://dx.doi.org/10.5772/intechopen.86691

than the alkyl transferase DNA repair mechanism [20].

New optimized synthesis of bendamustine hydrochloride (3).

Figure 1. Chemical structures of benzimidazole (1) and N-ribosyl-dimethylbenzimidazole (2).

Figure 2. Chemical structures of bendamustine (3), selumetinib (4), and galeterone (5).

#### 2. Benzimidazole agents in the clinic and in clinical development

#### 2.1 Bendamustine (3)

Bendamustine (3) (Figure 2) was discovered in a structure-activity relationship (SAR) campaign directed to obtain more effective and safer water-soluble analogs of chlorambucil (6) (Figure 3), a nitrogen mustard, which is used clinically against chronic lymphatic leukemia, lymphomas, and advanced ovarian and breast carcinomas [12]. The strategy was replacement of the benzene ring in compound 6 with purine-like N-methylbenzimidazole moiety in the hope of obtaining an anticancer agent with antimetabolite and DNA-alkylating activities. Although bendamustine was first synthesized in the early 1960s [13], it was approved under the trade name Treanda® by the US Food and Drug Administration (FDA) in 2008 for the treatment of chronic lymphocytic leukemia, multiple myeloma, and non-Hodgkin's lymphoma [10, 14–16].

Figure 3. Replacement of aromatic benzene ring of chlorambucil (6) to produce bendamustine (3).

#### 2.1.1 Chemistry

Bendamustine (3) 4-{5-[bis(2-chloroethyl)amino]-1-methylbenzimidazol-2-yl} butanoic acid was first synthesized via eight synthetic steps with an overall yield of 12% [13, 17]. However, Chen and colleagues have developed a new, more efficient, and cost-effective route focused on the use of sustainable chemistry for the synthesis of bendamustine hydrochloride, with the overall yield improved from 12 to 45% as outlined in Figure 4 [18]. This new synthesis is currently used for the commercial production of 3.

#### 2.1.2 Summary of bendamustine's preclinical and clinical pharmacology

Even though bendamustine is an alkylating agent, due to its ability to cause intra-strand and inter-strand cross-links between DNA bases, it has been reported that the DNA breaks induced by bendamustine are more extensive/durable than those induced by other alkylating agents, such as chlorambucil, cyclophosphamide, or carmustine [19–21]. In addition, the drug was shown to exhibit partial crossresistance to other alkylating agents. These data suggested that bendamustine may possess additional mechanisms of action. Indeed, a comprehensive study by Leoni and colleagues clearly demonstrated that bendamustine exhibits a distinct pattern of activities unrelated to other alkylating drugs. Using a variety of lymphoid cancer cell lines, the study concluded that mechanisms of action include induction of mitotic catastrophe, inhibition of mitotic checkpoints, and activation of DNAdamage stress response and apoptosis. Compared to other alkylating agents, bendamustine was shown to activate the base excision DNA repair pathway rather than the alkyl transferase DNA repair mechanism [20].

Although bendamustine is approved for the treatment of a variety of lymphoid cancers, its activity has also been reported in several cancers, including cancers of small cell lung, breast, hepatic, bile duct, and head and neck. The studies by Chow and colleagues using leukemic cell lines in vitro or ex vivo cells from patients with

Figure 4. New optimized synthesis of bendamustine hydrochloride (3).

2. Benzimidazole agents in the clinic and in clinical development

Replacement of aromatic benzene ring of chlorambucil (6) to produce bendamustine (3).

Chemical structures of benzimidazole (1) and N-ribosyl-dimethylbenzimidazole (2).

Chemistry and Applications of Benzimidazole and its Derivatives

Chemical structures of bendamustine (3), selumetinib (4), and galeterone (5).

Bendamustine (3) (Figure 2) was discovered in a structure-activity relationship (SAR) campaign directed to obtain more effective and safer water-soluble analogs of chlorambucil (6) (Figure 3), a nitrogen mustard, which is used clinically against chronic lymphatic leukemia, lymphomas, and advanced ovarian and breast carcinomas [12]. The strategy was replacement of the benzene ring in compound 6 with purine-like N-methylbenzimidazole moiety in the hope of obtaining an anticancer agent with antimetabolite and DNA-alkylating activities. Although bendamustine was first synthesized in the early 1960s [13], it was approved under the trade name Treanda® by the US Food and Drug Administration (FDA) in 2008 for the treatment of chronic lymphocytic leukemia, multiple myeloma, and non-Hodgkin's

2.1 Bendamustine (3)

Figure 1.

Figure 2.

lymphoma [10, 14–16].

Figure 3.

90

leukemic progression to clarify interactions between bendamustine and other chemotherapeutic drugs unraveled synergy with cladribine, in contrast to observed antagonism with mitoxantrone or doxorubicin. The observation of synergism between bendamustine and rituximab (an anti-CD20 antibody) in in vitro CD20 positive DOHH-2 and WSU-NHL cell lines and ex vivo B-cell chronic lymphocytic leukemia (CLL) cells [22] and in mice with Daudi xenografts [23] provided the impetus for clinical trials combining these two drugs [24, 25].

The promising preclinical in vitro and in vivo data provided the rationale for multiple clinical trials in cancers with activated Raf-MEK-ERK signaling. In preparation for clinical evaluation of selumetinib, it was originally developed as a free base and administered as a liquid suspension, but subsequently a capsule formulation of the hydrogen sulfate salt was found to be more suitable for further development [38]. Several phase I and II clinical trials conducted against solid tumors to test the impact of selumetinib as a monotherapy were unsuccessful [41–44]. This led to the conduct of several clinical trials with selumetinib in combination with other cancer drugs. A notable trial was the randomized phase II study of selumetinib in combination with docetaxel, as a second-line treatment for patients with KRASmutant advanced NSCLC which showed very promising results [45]. The median progression-free survival was 5.3 months with selumetinib + docetaxel and 2.1 months with docetaxel alone. The objective response rate was 37% for selumetinib + docetaxel vs. 0% for docetaxel alone (p < 0.001), and the median overall survival was 9.4 months for selumetinib + docetaxel vs. 5.2 months for docetaxel alone (HR for death, 0.80 [80% CI, 0.56–1.14]; one-sided p = 0.21). Unfortunately, in a multinational 510 randomized patients with previously treated

Development of Benzimidazole Compounds for Cancer Therapy

DOI: http://dx.doi.org/10.5772/intechopen.86691

advanced KRAS-mutant NSCLC trial, the combination of selumetinib with

variety of cancers [47, 48].

2.3 Galeterone (5)

clinical studies [50].

but is yet to appear in the literature.

2.3.1 Chemistry

93

docetaxel did not improve progression-free survival compared with docetaxel alone [46]. Clearly, addition clinical studies are required to realize the potential impact of selumetinib alone and in combination with other drugs for the treatment of a

Galeterone (also called VN/124-1 or TOK-001) is an orally available anticancer agent. It was rationally designed as an inhibitor of androgen biosynthesis via inhibition of 17α-hydroxylase/17,20-lyase (CYP17), the key enzyme which catalyzes the biosynthesis of androgens from the progestins. Through extensive and rigorous preclinical studies, galeterone was shown to modulate two other targets in the androgen/androgen receptor (AR) signaling pathway [11] and shown to inhibit the eukaryotic initiation factor 4E (eIF4E) protein translational machinery [49]. Galeterone advanced successfully through phases I and II clinical trials in prostate cancer patients but was unsuccessful in the pivotal phase III clinical trial in men with castration-resistant prostate cancer (CRPC), harboring AR splice variants (e.g., AR-V7). We present a summary of the chemistry, preclinical studies, and

Galeterone, 3β-hydroxy-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (5), is one of a series of novel Δ16-17-azolyl steroid, which, unlike previously known 17 heteroaryl steroids, the azole moiety is attached to the steroid nucleus at C-17 via a nitrogen of the azole. The synthesis of galeterone from commercially available 3βacetoxyandrosta-5-en-17-one (12) is presented in Figure 5 [11, 51], and a facile and large-scale preparation (commercial process) of the compound has been developed

Using intact CYP17 expressing Escherichia coli, galeterone was shown to be a potent inhibitor of the enzyme with an IC50 value of 300 nM and was shown to be more potent than abiraterone (IC50 value of 800 nM) [52]. Additional studies by

2.3.2 Summary of galeterone's preclinical and clinical pharmacology

Based on the discussion above, it is obvious that bendamustine is an "old drug rediscovered." For over 30 years, bendamustine was used in Eastern Germany as monotherapy for several cancers, including breast cancer, chronic lymphocytic leukemia (CLL), Hodgkin's lymphoma, non-Hodgkin's lymphoma (NHL), and multiple myeloma (MM) [26–37]. However, following the reunification of Germany, other countries initiated clinical trials of bendamustine as a single agent and in combination with other drugs. Bendamustine has achieved worldwide regulatory approval and is a standard-of-care drug for the treatment on many lymphoid malignancies. Several articles that provide comprehensive reviews of the discovery and development of this unique drug are available [8–10].

#### 2.2 Selumetinib (4)

Selumetinib (4) (AZD6244: ARRY-142866) is an orally available, potent, selective inhibitor of mitogen-activated protein kinase (MAPK)/extracellular signalrelated kinase (ERK) kinases 1 and 2 (MEK1 and MEK2) [6]. This agent has been extensively studied in many preclinical and clinical studies in several tumor types with mixed results. Here, we will summarize the chemistry, preclinical studies, and clinical studies.

#### 2.2.1 Chemistry

Selumetinib (6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3 methyl benzimidazole-5-carboxamide) (4) is a diarylamine hydroxamide, containing mono-methylated benzimidazole subunit [38, 39]. It is a secondgeneration, orally active small molecule that acts as a selective and ATPuncompetitive inhibitor of MEK1 and MEK2, binding to the allosteric binding site [38, 39]. The synthesis of selumetinib is yet to be reported in the literature.

#### 2.2.2 Summary of selumetinib's preclinical and clinical pharmacology

Selumetinib inhibits the enzymatic activity of purified constitutively active MEK1 with a half maximal inhibitory concentration (IC50) of 14 nM and was shown to be highly selective for inhibition of these targets compared to other related kinases [39]. Using several human cancer cell lines such as NSCLC, melanoma, and pancreatic and colorectal cell lines, it was shown that selumetinib was a potent antiproliferative agent. Analysis of the data revealed that cell lines with mutant BRAF and RAS were sensitive to selumetinib [40]. Selumetinib had little effect on the growth of Malme-3, the control cell line to the melanoma. Additional studies suggested that the growth inhibitory effects of selumetinib was not due to wideranging cytotoxicity [39], and it was also established that selumetinib effectively inhibits the phosphorylation of ERK 1 and ERK 2, which are substrates of MEK1 and MEK2 in the MAP kinase pathway. This mechanism of action was confirmed in tumor xenografts. Additionally, increased markers of apoptosis such as cleaved caspase 3 and decreased cell proliferation were seen in response to treatment with selumetinib in the xenograft models [39, 40].

Development of Benzimidazole Compounds for Cancer Therapy DOI: http://dx.doi.org/10.5772/intechopen.86691

The promising preclinical in vitro and in vivo data provided the rationale for multiple clinical trials in cancers with activated Raf-MEK-ERK signaling. In preparation for clinical evaluation of selumetinib, it was originally developed as a free base and administered as a liquid suspension, but subsequently a capsule formulation of the hydrogen sulfate salt was found to be more suitable for further development [38]. Several phase I and II clinical trials conducted against solid tumors to test the impact of selumetinib as a monotherapy were unsuccessful [41–44]. This led to the conduct of several clinical trials with selumetinib in combination with other cancer drugs. A notable trial was the randomized phase II study of selumetinib in combination with docetaxel, as a second-line treatment for patients with KRASmutant advanced NSCLC which showed very promising results [45]. The median progression-free survival was 5.3 months with selumetinib + docetaxel and 2.1 months with docetaxel alone. The objective response rate was 37% for selumetinib + docetaxel vs. 0% for docetaxel alone (p < 0.001), and the median overall survival was 9.4 months for selumetinib + docetaxel vs. 5.2 months for docetaxel alone (HR for death, 0.80 [80% CI, 0.56–1.14]; one-sided p = 0.21). Unfortunately, in a multinational 510 randomized patients with previously treated advanced KRAS-mutant NSCLC trial, the combination of selumetinib with docetaxel did not improve progression-free survival compared with docetaxel alone [46]. Clearly, addition clinical studies are required to realize the potential impact of selumetinib alone and in combination with other drugs for the treatment of a variety of cancers [47, 48].

#### 2.3 Galeterone (5)

leukemic progression to clarify interactions between bendamustine and other chemotherapeutic drugs unraveled synergy with cladribine, in contrast to observed antagonism with mitoxantrone or doxorubicin. The observation of synergism between bendamustine and rituximab (an anti-CD20 antibody) in in vitro CD20 positive DOHH-2 and WSU-NHL cell lines and ex vivo B-cell chronic lymphocytic leukemia (CLL) cells [22] and in mice with Daudi xenografts [23] provided the

Based on the discussion above, it is obvious that bendamustine is an "old drug rediscovered." For over 30 years, bendamustine was used in Eastern Germany as monotherapy for several cancers, including breast cancer, chronic lymphocytic leukemia (CLL), Hodgkin's lymphoma, non-Hodgkin's lymphoma (NHL), and multiple myeloma (MM) [26–37]. However, following the reunification of Germany, other countries initiated clinical trials of bendamustine as a single agent and in combination with other drugs. Bendamustine has achieved worldwide regulatory approval and is a standard-of-care drug for the treatment on many lymphoid malignancies. Several articles that provide comprehensive reviews of the discovery

Selumetinib (4) (AZD6244: ARRY-142866) is an orally available, potent, selective inhibitor of mitogen-activated protein kinase (MAPK)/extracellular signalrelated kinase (ERK) kinases 1 and 2 (MEK1 and MEK2) [6]. This agent has been extensively studied in many preclinical and clinical studies in several tumor types with mixed results. Here, we will summarize the chemistry, preclinical studies, and

Selumetinib (6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3-

uncompetitive inhibitor of MEK1 and MEK2, binding to the allosteric binding site [38, 39]. The synthesis of selumetinib is yet to be reported in the literature.

Selumetinib inhibits the enzymatic activity of purified constitutively active MEK1 with a half maximal inhibitory concentration (IC50) of 14 nM and was shown to be highly selective for inhibition of these targets compared to other related kinases [39]. Using several human cancer cell lines such as NSCLC, melanoma, and pancreatic and colorectal cell lines, it was shown that selumetinib was a potent antiproliferative agent. Analysis of the data revealed that cell lines with mutant BRAF and RAS were sensitive to selumetinib [40]. Selumetinib had little effect on the growth of Malme-3, the control cell line to the melanoma. Additional studies suggested that the growth inhibitory effects of selumetinib was not due to wideranging cytotoxicity [39], and it was also established that selumetinib effectively inhibits the phosphorylation of ERK 1 and ERK 2, which are substrates of MEK1 and MEK2 in the MAP kinase pathway. This mechanism of action was confirmed in tumor xenografts. Additionally, increased markers of apoptosis such as cleaved caspase 3 and decreased cell proliferation were seen in response to treatment with

methyl benzimidazole-5-carboxamide) (4) is a diarylamine hydroxamide, containing mono-methylated benzimidazole subunit [38, 39]. It is a secondgeneration, orally active small molecule that acts as a selective and ATP-

2.2.2 Summary of selumetinib's preclinical and clinical pharmacology

selumetinib in the xenograft models [39, 40].

impetus for clinical trials combining these two drugs [24, 25].

Chemistry and Applications of Benzimidazole and its Derivatives

and development of this unique drug are available [8–10].

2.2 Selumetinib (4)

clinical studies.

2.2.1 Chemistry

92

Galeterone (also called VN/124-1 or TOK-001) is an orally available anticancer agent. It was rationally designed as an inhibitor of androgen biosynthesis via inhibition of 17α-hydroxylase/17,20-lyase (CYP17), the key enzyme which catalyzes the biosynthesis of androgens from the progestins. Through extensive and rigorous preclinical studies, galeterone was shown to modulate two other targets in the androgen/androgen receptor (AR) signaling pathway [11] and shown to inhibit the eukaryotic initiation factor 4E (eIF4E) protein translational machinery [49]. Galeterone advanced successfully through phases I and II clinical trials in prostate cancer patients but was unsuccessful in the pivotal phase III clinical trial in men with castration-resistant prostate cancer (CRPC), harboring AR splice variants (e.g., AR-V7). We present a summary of the chemistry, preclinical studies, and clinical studies [50].

#### 2.3.1 Chemistry

Galeterone, 3β-hydroxy-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (5), is one of a series of novel Δ16-17-azolyl steroid, which, unlike previously known 17 heteroaryl steroids, the azole moiety is attached to the steroid nucleus at C-17 via a nitrogen of the azole. The synthesis of galeterone from commercially available 3βacetoxyandrosta-5-en-17-one (12) is presented in Figure 5 [11, 51], and a facile and large-scale preparation (commercial process) of the compound has been developed but is yet to appear in the literature.

#### 2.3.2 Summary of galeterone's preclinical and clinical pharmacology

Using intact CYP17 expressing Escherichia coli, galeterone was shown to be a potent inhibitor of the enzyme with an IC50 value of 300 nM and was shown to be more potent than abiraterone (IC50 value of 800 nM) [52]. Additional studies by

Based on these impressive preclinical data, galeterone was licensed by the Uni-

versity of Maryland, Baltimore, to Tokai Pharmaceuticals, Inc., who initiated Androgen Receptor Modulation Optimized for Response 1 (ARMOR1) phase 1/ phase 2 trials in castrate-resistant prostate cancer patients on November 5, 2009 [11]. The ARMOR phase I and phase II studies conducted with galeterone demonstrated that galeterone is well tolerated with promising clinical activity in patients with CRCP [62, 63]. To determine whether galeterone has clinical activity in patients with C-terminal loss of the androgen receptor, circulating tumor cells were retrospectively tested for C-terminal loss. Of the seven patients identified, six had PSA50 responses. These promising phases I and II studies enabled the selection of

Development of Benzimidazole Compounds for Cancer Therapy

DOI: http://dx.doi.org/10.5772/intechopen.86691

galeterone 2500 mg/day dose for the pivotal phase III trial (ARMOR3-SV, NCT02438007). The retrospective data of patients with C-terminal loss of the androgen receptor supported the design of ARMOR3-SV pivotal trial in which patients with AR-V7 were randomized to receive either galeterone or enzalutamide. Regrettably, the trial was discontinued following review by the independent Data Monitoring Committee, though no safety concerns were cited regarding this recommendation [50]. Gratifyingly, Educational and Scientific LLC (ESL), Baltimore, announced (December 17, 2018) that the University of Maryland, Baltimore (UMB), has granted ESL an exclusive license for the development of galeterone for the treatment of patients with CRPC. We eagerly await the initiation of a new phase

Despite the enormous literature on the synthesis and preclinical evaluation of numerous benzimidazole-containing compounds, it is unclear why very few of this class of compounds have entered clinical trials for evaluation as potential anticancer drugs. Given the fact that many benzimidazole-containing drugs have achieved blockbuster status for other diseases, it may be reasonable to suggest that the researchers interested in the development of benzimidazole-contained anticancer drugs should carefully study the process that have resulted in successful non-cancer benzimidazole drugs. We hope that this review will stimulate research activities

This work was supported in part by a grant from the National Institutes of Health (NIH) and the National Cancer Institute (NCI) (R01CA224696) to VCON.

VCON is the lead inventor of galeterone; the patents and technologies thereof are owned by the University of Maryland, Baltimore. The other authors declare no

During the review of this manuscript, it was reported that an analog of Selumetinib called Binimetinib (Mektovi) in combination with a BRAF inhibitor

III clinical study of galeterone in men with prostate cancer.

that would eventually produce new anticancer benzimidazole drugs.

3. Concluding remarks

Acknowledgements

Conflict of interest

potential conflict of interest.

Note added in proof

95

Figure 5. Synthesis of galeterone.

our group revealed that galeterone could disrupt androgen signaling through multiple targets [51–54].

We strongly believe that the increased efficacy of galeterone in several prostate cancer models both in vitro and in vivo is due to its ability to downregulate the AR and block androgen binding to AR. Using well-established AR-competitive binding assays (against the synthetic androgen [3 H]R1881), galeterone was equipotent to Casodex in LNCaP cells but had a slightly higher affinity for the wild-type receptor in PC3-AR cells. In transcriptional activation assays (utilizing a luciferase reporter), galeterone was shown to be a pure AR antagonist of the wild-type AR and the T877A mutation found in LNCaP cells [53]. In prostate cancer cell lines, galeterone inhibited the growth of CRPCs, which had increased AR and were no longer sensitive to Casodex [53] and was also shown to inhibit the growth of AR-negative prostate cancer cells [54]. In addition, galeterone demonstrated superior synergy for growth inhibition in combination with everolimus or gefitinib compared with Casodex [55].

Recent in vitro studies have shown additional activities of galeterone, including proteasomal degradation of AR and its splice variants [56, 57] and inhibition of the eukaryotic initiation factor 4E (eIF4E) protein translational machinery via induction of proteasomal degradation of mitogen-activated protein kinase-interacting kinases 1 and 2 (Mnk1 and Mnk2) [58, 59].

Because of the short half-life (t<sup>½</sup> = 40 min) in mice, galeterone was administered twice daily in our antitumor efficacy studies. Galeterone (0.13 mmol/kg twice daily) caused a 93.8% reduction (p = 0.00065) in the mean final LAPC-4 xenograft volume compared with controls, and this efficacy was significantly more effective than castration or our most potent CYP17 inhibitor, VN/85-1 [51]. In another antitumor efficacy study, treatment of galeterone (0.13 mmol twice daily) was very effective in preventing the formation of LAPC4 tumors (6.94 vs. 2410.28 mm<sup>3</sup> in the control group). Galeterone (0.13 mmol/kg twice daily) and VN/124-1 (0.13 mmol/kg twice daily) + castration induced regression of LAPC4 tumor xenografts by 26.55 and 60.67%, respectively [53]. Using castration-resistant prostate cancer (CRPC) HP-LNCaP tumor xenografts, we showed that galeterone + everolimus (m-TORC1 inhibitor) acted in concert to reduce tumor growth [60]. Utilizing the androgen-dependent LAPC-4 prostate cancer xenograft model, we have shown galeterone is more efficacious than the blockbuster prostate cancer drug abiraterone (Zytiga®) [61]. We also reported that galeterone potently inhibits the growth of CRPC CWR22Rv1 tumor xenografts [56].

Development of Benzimidazole Compounds for Cancer Therapy DOI: http://dx.doi.org/10.5772/intechopen.86691

Based on these impressive preclinical data, galeterone was licensed by the University of Maryland, Baltimore, to Tokai Pharmaceuticals, Inc., who initiated Androgen Receptor Modulation Optimized for Response 1 (ARMOR1) phase 1/ phase 2 trials in castrate-resistant prostate cancer patients on November 5, 2009 [11]. The ARMOR phase I and phase II studies conducted with galeterone demonstrated that galeterone is well tolerated with promising clinical activity in patients with CRCP [62, 63]. To determine whether galeterone has clinical activity in patients with C-terminal loss of the androgen receptor, circulating tumor cells were retrospectively tested for C-terminal loss. Of the seven patients identified, six had PSA50 responses. These promising phases I and II studies enabled the selection of galeterone 2500 mg/day dose for the pivotal phase III trial (ARMOR3-SV, NCT02438007). The retrospective data of patients with C-terminal loss of the androgen receptor supported the design of ARMOR3-SV pivotal trial in which patients with AR-V7 were randomized to receive either galeterone or enzalutamide. Regrettably, the trial was discontinued following review by the independent Data Monitoring Committee, though no safety concerns were cited regarding this recommendation [50]. Gratifyingly, Educational and Scientific LLC (ESL), Baltimore, announced (December 17, 2018) that the University of Maryland, Baltimore (UMB), has granted ESL an exclusive license for the development of galeterone for the treatment of patients with CRPC. We eagerly await the initiation of a new phase III clinical study of galeterone in men with prostate cancer.

#### 3. Concluding remarks

our group revealed that galeterone could disrupt androgen signaling through mul-

We strongly believe that the increased efficacy of galeterone in several prostate cancer models both in vitro and in vivo is due to its ability to downregulate the AR and block androgen binding to AR. Using well-established AR-competitive binding

Casodex in LNCaP cells but had a slightly higher affinity for the wild-type receptor in PC3-AR cells. In transcriptional activation assays (utilizing a luciferase reporter), galeterone was shown to be a pure AR antagonist of the wild-type AR and the T877A

Recent in vitro studies have shown additional activities of galeterone, including proteasomal degradation of AR and its splice variants [56, 57] and inhibition of the eukaryotic initiation factor 4E (eIF4E) protein translational machinery via induction of proteasomal degradation of mitogen-activated protein kinase-interacting

Because of the short half-life (t<sup>½</sup> = 40 min) in mice, galeterone was administered twice daily in our antitumor efficacy studies. Galeterone (0.13 mmol/kg twice daily) caused a 93.8% reduction (p = 0.00065) in the mean final LAPC-4 xenograft volume compared with controls, and this efficacy was significantly more effective than castration or our most potent CYP17 inhibitor, VN/85-1 [51]. In another antitumor efficacy study, treatment of galeterone (0.13 mmol twice daily) was very effective in preventing the formation of LAPC4 tumors (6.94 vs. 2410.28 mm<sup>3</sup> in

the control group). Galeterone (0.13 mmol/kg twice daily) and VN/124-1

cancer (CRPC) HP-LNCaP tumor xenografts, we showed that

inhibits the growth of CRPC CWR22Rv1 tumor xenografts [56].

(0.13 mmol/kg twice daily) + castration induced regression of LAPC4 tumor xenografts by 26.55 and 60.67%, respectively [53]. Using castration-resistant prostate

galeterone + everolimus (m-TORC1 inhibitor) acted in concert to reduce tumor growth [60]. Utilizing the androgen-dependent LAPC-4 prostate cancer xenograft model, we have shown galeterone is more efficacious than the blockbuster prostate cancer drug abiraterone (Zytiga®) [61]. We also reported that galeterone potently

mutation found in LNCaP cells [53]. In prostate cancer cell lines, galeterone inhibited the growth of CRPCs, which had increased AR and were no longer sensitive to Casodex [53] and was also shown to inhibit the growth of AR-negative prostate cancer cells [54]. In addition, galeterone demonstrated superior synergy for growth inhibition in combination with everolimus or gefitinib compared with

H]R1881), galeterone was equipotent to

tiple targets [51–54].

Synthesis of galeterone.

Figure 5.

Casodex [55].

94

assays (against the synthetic androgen [3

Chemistry and Applications of Benzimidazole and its Derivatives

kinases 1 and 2 (Mnk1 and Mnk2) [58, 59].

Despite the enormous literature on the synthesis and preclinical evaluation of numerous benzimidazole-containing compounds, it is unclear why very few of this class of compounds have entered clinical trials for evaluation as potential anticancer drugs. Given the fact that many benzimidazole-containing drugs have achieved blockbuster status for other diseases, it may be reasonable to suggest that the researchers interested in the development of benzimidazole-contained anticancer drugs should carefully study the process that have resulted in successful non-cancer benzimidazole drugs. We hope that this review will stimulate research activities that would eventually produce new anticancer benzimidazole drugs.

#### Acknowledgements

This work was supported in part by a grant from the National Institutes of Health (NIH) and the National Cancer Institute (NCI) (R01CA224696) to VCON.

#### Conflict of interest

VCON is the lead inventor of galeterone; the patents and technologies thereof are owned by the University of Maryland, Baltimore. The other authors declare no potential conflict of interest.

#### Note added in proof

During the review of this manuscript, it was reported that an analog of Selumetinib called Binimetinib (Mektovi) in combination with a BRAF inhibitor (Encorafenib, Braftovi) was approved by US Food and Drug Administration (FDA) for the treatment of unresectable or metastatic melanoma with BRAF mutations [64].

## List of abbreviations


Author details

and Vincent C.O. Njar1,2,3\*

Medicine, Baltimore, MD, USA

Baltimore, MD, USA

97

Puranik Purushottamachar1,2, Senthilmurugan Ramalingam1,2

Development of Benzimidazole Compounds for Cancer Therapy

DOI: http://dx.doi.org/10.5772/intechopen.86691

Maryland School of Medicine, Baltimore, MD, USA

provided the original work is properly cited.

\*Address all correspondence to: vnjar@som.umaryland.edu

1 Department of Pharmacology, University of Maryland School of Medicine,

2 Center for Biomolecular Therapeutics, University of Maryland School of

3 Marlene and Stewart Greenebaum Comprehensive Cancer Center, University of

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

Development of Benzimidazole Compounds for Cancer Therapy DOI: http://dx.doi.org/10.5772/intechopen.86691

#### Author details

(Encorafenib, Braftovi) was approved by US Food and Drug Administration (FDA) for the treatment of unresectable or metastatic melanoma with BRAF

ARMOR androgen receptor modulation optimized for response

CWR22Rv1 a type of AR/AR-splice variants positive human prostate can-

BRAF a human gene that encodes a protein called B-Raf

CYP17 cytochrome P450 17α-hydroxylase/17,20-lyase

ERK1 and ERK2 extracellular signal-regulated kinases 1 and 2

enzyme activity by 50% KRAS Kirsten retrovirus-associated DNA sequences

IC50 is the concentration of inhibitor required to inhibit the

LAPC-4 a type of AR-positive human prostate cancer cell line LNCaP a type of AR-positive human prostate cancer cell line

Mnk1 and Mnk2 mitogen-activated protein kinase-interacting kinases 1 and 2

PC3-AR a type of AR-negative prostate cancer cell line transfected

VN/85-1 code name for a CYP17 inhibitor/AR antagonist/AR degrader

AME apparent mineralocorticoid excess

Chemistry and Applications of Benzimidazole and its Derivatives

CLL chronic lymphocytic leukemia CRPC castration-resistant prostate cancer

cer cell line

eIF4E eukaryotic initiation factor 4E

ESL Educational and Scientific LLC FDA Food and Drug Administration HIV human immunodeficiency virus

MAPK mitogen-activated protein kinase

AR-V7 a type of androgen receptor splice variant

AR androgen receptor

CI confidence interval

DNA deoxyribonucleic acid

HR hazard ratio

MEK1 and MEK2 MAPK/ERK kinase MM multiple myeloma

TB tuberculosis

96

NHL non-Hodgkin's lymphoma NSCLC non-small cell lung carcinoma

with AR PSA prostate-specific antigen

RAF rapidly accelerated fibrosarcoma RAS retrovirus-associated DNA sequences SAR structure-activity relationship

TOK-001 another code name of galeterone UMB University of Maryland, Baltimore

VN/124-1 original code name of galeterone

mutations [64].

List of abbreviations

Puranik Purushottamachar1,2, Senthilmurugan Ramalingam1,2 and Vincent C.O. Njar1,2,3\*

1 Department of Pharmacology, University of Maryland School of Medicine, Baltimore, MD, USA

2 Center for Biomolecular Therapeutics, University of Maryland School of Medicine, Baltimore, MD, USA

3 Marlene and Stewart Greenebaum Comprehensive Cancer Center, University of Maryland School of Medicine, Baltimore, MD, USA

\*Address all correspondence to: vnjar@som.umaryland.edu

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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**105**

**Chapter 7**

*Yousef Najajreh*

**Abstract**

Benzimidazoles: From

Anticancer Agents

non-kinase target (as shown recently for dovitinib).

chloroacetamide, amidine, binding, mode

**1.1 Antiproliferative action of benzimidazoles**

**1.2 Benzimidazoles act on numerous biological targets**

**1. Introduction**

**Keywords:** benzimidazole, selective, cytotoxic, inhibitor, multitargeting, multikinase, polypharmacology, antiproliferative, quinolinone, carbamate, quinolinone, pyrazole, urea, aniline, anilinobenzimidazolylpyrimidine,

Benzimidazole, a heterocyclic moiety comprising six-membered benzene ring fused with five-membered imidazole ring, containing molecules, was known for its ability to induce antiproliferative effects (named as antineoplastic, anticancer, or antitumor agents). Numerous structures were reported as effective inhibitors of cell growth and division, thus acting as antiviral, antibacterial, antifungal, anthelmintic (or antihelminthics), and anticancer agents. Over the years, several published scripts have reviewed the synthetic approaches, medicinal chemistry, SAR, bioactivities, and preclinical and clinical studies of such "gifted" fragment [1–8].

A wide range of activities and medical situations benzimidazole containing compounds have been used for. That includes antihypertensive [9–12], anti-inflammatory [13–15], antibacterial [16–18], antiviral [19–21], antifungal [22–24], antihelmintic [25–28], anticancer [29–32], antiulcer [33–35], antioxidant [36–38], and

Antiproliferative to Multitargeted

Benzimidazole derivatives are known to act against a range of biological targets and thus gained clinical applications in a wide spectrum of diseases. Few examples of multitargeted benzimidazole derivatives that were reported during the last decade will be described in this chapter. Multitargeting agents for serving the polypharmacology approach to combat shortcomings of the main one-drug-one target main dogma will be briefly explored. In that context, the multitargeting benzimidazole derivatives gain a special attention. This includes discovery (hit-to-lead), structureactivity relationship (SAR), and binding mode of at least one lead (or hit) in each group. Special attention will be given to two structures dovitinib and AT9283 that are reported to exhibit potent in vitro and in vivo activities against a group of kinases and

#### **Chapter 7**
