Advances of Benzimidazole Derivatives as Anticancer Agents: Bench to Bedside

Benzimidazole is one of the privileged nitrogen-containing scaffolds known for its versatile diversified role in insecticides, pesticides, dyes, pigments and phar-maceuticals. Due to its electron-rich environment, structural features and binding potency of various therapeutic targets, benzimidazole derivatives exhibit a broad spectrum of biological activity that majorly includes antimicrobial, antifungal, analgesics, anti-diabetic and anticancer agents. Several benzimidazole scaffolds bearing drugs are clinically approved; they are used for various indications. For example, Bilastine, Lerisetron, Maribavir and Nocodazole are the most widely used benzimidazole-based marketed drugs available as an antihistamine, antiviral and antimitotic agent, respectively. Another example is the recently approved anticancer drug Binimetinib and Selumetinib, which are indicated for BRAF mutated melanoma and plexiform neurofibromas. Not only this, many benzimidazole-based anticancer drugs are in late phases of clinical development. Due to the vast therapeutic potential of benzimidazole scaffold in cancer research, medicinal chemists have gained a lot of attraction to explore it more and develop novel, highly effective and target-specific benzimidazole-based potential anticancer drugs.


Introduction
Cancer is a complex, severe class of diseases that involves a group of cells that exhibit abnormal and uncontrolled division and proliferation. It is one of the primary health concerns which accounts for the second major cause of death globally. As per the recent statistics of the world health organization (WHO), in 2020, around 10 million people succumbed to death due to cancer. However, every year the number of incidences is increasing day by day. According to WHO, around 0.3 million new cases are diagnosed each year among the age group of 0-19 years. Cancer can affect a person of any age; however, with age, the risk increases. Globally, steady increases in cancer cases every year are taking a toll on the health care system [1][2][3][4][5]. To combat cancer, identification of potential drugs and potential drugs combination is essential. Potential research has been carried out to counter such problems by addressing novel drug design and discovery approaches. In medicinal chemistry, heterocyclic rings have played a significant role in the search for potential therapeutic agents. Various drugs are currently in use and in development that widely addresses such problems. However, due to changes in cancer forms and mutations, current therapy faces challenges of poor selectivity and specificity towards certain types of cancer cells, which narrows down their effectiveness. Generally, cancer cells act by disrupting and disturbing the cell signaling pathways; therefore, it is crucial to design novel target-based heterocyclic

Advances of benzimidazole based anticancer agents
Benzimidazole based compounds have got much attention due to exhibiting significant cytotoxic activity. In last one decade a lot of benzimidazole based anticancer drugs have received status of US FDA global approval. Recently, Binimetinib, Selumetinib and Abemaciclib got approval for treatment of various mutated forms of cancer. Here, we have discussed some of benzimidazole based anticancer drugs which are recently approved, under development and in pipeline.

Binimetinib (1)
Binimetinib (1) is chemically 5-((4-bromo-2-fluorophenyl)amino)-4-fluoro-N-(2-hydroxyethoxy)-1-methyl-1H-benzimidazole-6-carboxamide approved by US FDA recently in 2018. It is an orally available, potent selective inhibitor of mitogen activated protein kinase (MEK 1/2). Binimetinib is developed by Array Biopharma, commercially available by the name of Mektovi. It is indicated for patients having metastatic melanoma with BRAF mutation as combination therapy with BRAF inhibitors Encorafenib [32]. Presently, Binimetinib is in various phases of clinical development as monotherapy or in combination for conditions like KRAS mutated cancer, mutated non-small cell lung cancer [33,34]. Structures of all the drugs are presented in Figure 2. More details of clinical trials are enlisted in Table 1.

Nazartinib (9)
Nazartinib (9) it is an orally available third generation EGFR kinase inhibitor under development for treatment of conditions like non-small cell lung cancer (NSCLC) and EGFR mutated NSCLC [55]. Nazartinib have demonstrated favorable safety profile and efficacy in a Phase-I study when administered to adult patients with EGFR mutated lung carcinoma [56]. However, clinical development of Nazartinib is progress for different forms of mutated carcinomas as monotherapy or in combination [57,58].

Benzimidazole based derivatives as potent kinase inhibitors
Commonly the mechanism behind action of anticancer agents involve DNA intercalation, gene regulation, microtubule inhibition, transcription regulation, DNA synthesis inhibition, enzyme inhibition and so on. Nowadays in cancer treatment, target therapy emerged as one of the acknowledged strategies. Most of the available anticancer drugs acts by targeting structural proteins, tyrosine kinases, phosphoinositide 3 kinase and protein kinases for example Binimetinib acts by inhibiting mitogen activated kinase as discussed in earlier section. In this section we have included some recent examples of benzimidazole based enzyme inhibitors as potent anticancer agents.

EGFR inhibitors
Akhter et al. have reported a novel series of benzimidazole based oxadiazole derivatives as potential EGFR inhibitors. The target compound 10 and 11 demonstrated significant binding to EGFR with an IC 50  respectively. Further findings of these derivatives showed that compound 10 cause cell cycle arrest of MCF7 cells in a dose dependent manner at G2/M phase. Docking analysis of target compound 10 and 11 showed that both the compound made strong interactions within the active site of protein kinase, the binding pattern of target compounds resembles as that of standard drug erlotinib, which is a potent EGFR inhibitor. In vivo acute toxicity of target compound showed that both compounds 10 and 11 are nontoxic and safe with oral LD 50 value >500 < 2000 mg/kg which is recommended by OECD guidelines [59]. Srour    Bax/Bcl-2 and caspase-3 expression and remarkable decrease in levels of PARP-1 enzyme. Molecular docking analysis of compound 12 and 13 showed that both the compounds embedded tightly by hydrogen bond formed between the Nitrogen of benzimidazole with amino acid residue Lys721 and Phe699 respectively [60].

PI3K inhibitors
GSK2636771 (23) is a novel, potent, orally available benzimidazole derivatives. It demonstrated selective PI3K beta inhibitor with antineoplastic activity.   Preclinical study of GSK2636771 demonstrated selective inhibition of PTENdeficient cancer cell growth along with inhibition of protein kinase B in a dose and time dependent manner. First in human trial study of GSK2636771 in patients of advanced solid tumors on oral administration as monotherapy demonstrated significant exposure, inhibition of target and excellent safety profile [67,68].
Jin et al. have reported novel benzimidazole derivatives as potent PI3K inhibitor. Target compound 24 was found most potent against PI3Kα with 36% and 86% inhibition compare to reference drug Alpelisib, which showed an inhibition of 110% and 109% at 50 nM and 500 nM respectively. Further, molecular docking analysis of target compound 24 demonstrated strong binding with six strong hydrogen bond with GLN-859, SER-854 and VAL-851 amino acid residues. Further, HUMO-LUMO calculation which is studied by using Gaussian 09 software target compound 24 showed presence of thiazole core and amide bonds which played an important role in its biological activity [69].
Recently In vivo pharmacokinetic profile of target compound was found good with oral bioavailability of 45% and 41% respectively. In vivo studied of compound 28 and 29 suggested that both the compounds can inhibit KLH-specific antibodies [73].
He et al. has reported benzimidazole-isoquinolinone derivatives which inhibits the cell growth via inhibiting PI3K/mTOR/Akt pathway. Target compound 30 demonstrated excellent inhibitory activity against SW620 and HT29 cancer cell line with an GI 50  Further western blot analysis study of compound 32 confirmed that it completely prevented the phosphorylation of Akt and p70S6K in HCT116 cells, thus target compound was determined as potential dual inhibitor of PI3K and mTOR kinase. Molecular docking analysis of compound 32 displayed that good binding interaction within the active site of PI3Kα [75] (Figure 6).

CDK inhibitors
Ibrahim et al. have reported a novel series of flavopiridol-benzimidazole as potent inhibitor potent inhibitor of CDK2 and CDK9 kinase. Target compound 33 exhibits potential inhibitory activity with an IC 50 value of 0.064 and 1.725 μM against CDK2 and CDK9 kinases respectively. Furthermore, compound 33 also displayed potential antiproliferative activity against selected cancer cell line SKOV3, PC3 and K562 with an IC 50 value of 94.0 μM, 85.0 μM and 50.8 μM respectively. Cell cycle analysis study of target compound revealed that it arrests the cell cycle of K562 cancer cell at G1 and G2 phase in a dose dependent manner [76] (Figure 7).     Both hybrid derivative 38 and 39 demonstrated significant inhibition of A549 cells migration through disruption of F-actin assembly, further treatment with 38 and 39 also showed increase in level of ROS in A549 cells by collapsing the mitochondrial membrane potential [80].

Benzimidazole based hybrid derivatives as potent anticancer agents
Bistrovic et al. have reported novel hybrid derivatives of benzimidazole-1,2,3triazole as potent anticancer agents. Target compound 40 and 41 demonstrated excellent inhibitory activity with IC50 value of 0.05 and 6.18 against A549 cancer cell line and an IC50 value of 17.53 and 8.80 against HeLa cancer cell line respectively. Furthermore, apoptosis detection study by annexin assay of compound 40 showed significant reduction of viable cell population by 70.59%, with increase in early necrotic cell population by 27.81% and late apoptotic cells by 40%. Similarly compound 41 also displayed markable decrease in cell population by 49.77%. Molecular docking analysis of compound 40 and 41 demonstrated that both the compound bind to the active site of p38 complex strongly [81] (Figure 8).

Conclusion
Many benzimidazole-containing compounds as anticancer agents are studied and available, involving various mechanisms in inhibiting mutated cancerous cells, in which kinases inhibitors play a significant role. However, in targeted therapy, benzimidazole-based derivatives are still widely explored. Due to the challenge of target specificity and poor selectivity, very few compounds have been approved to treat mutated cancers. The search for a novel benzimidazole-based next generation kinase inhibitor is going to subside such challenges. Benzimidazole-based target therapies such as enzyme inhibitors have gained a lot of attraction; owing to this, recently US FDA has approved EGFR inhibitor Abemaciclib and MEK inhibitor Binimetinib and Selumetinib as potent anticancer compounds against mutated forms of cancer. Apart from this, many benzimidazole-containing compounds are in the developmental phase as EGFR, VEGFR-2, CDK and PI3K inhibitors. However, some of the compounds demonstrated excellent kinase inhibitory activity but failed to provide a strong safety profile; these compounds will pave a path as lead compounds; further modifications, designing, and developing such compounds will give potent compounds with maximum efficiency and minimal side effects. The presented chapter mainly focuses on benzimidazole-based kinase inhibitors and their advances; the pivotal information catered here can be regarded as noteworthy and crucial by medicinal chemists for drug design, discovery and development of novel, potent and safe, target-based anticancer agents. carcinoma: A review of clinical features, treatment targets and advancement in improving the immune response to monoclonal antibody therapy. Biochimica et Biophysica Acta (BBA)-Reviews on Cancer. 2021;1875:188523