Influence of Phenyl Ring Disubstitution on Bisbenzimidazole and Terbenzimidazole Cytotoxicity:  Synthesis and Biological Evaluation as Radioprotectors

Abstract

DNA minor groove binders, Hoechst 33258 and Hoechst 33342, have been reported to protect against radiation-induced DNA-strand breakage, but their mutagenicity and cytotoxicity limit their use as protectors of normal tissue during radiotherapy and as biological radioprotectors during accidental radiation exposure. On the basis of these observations, two new nontoxic disubstituted benzimidazoles were synthesized, one having two methoxy groups (5-(4-methylpiperazin-1-yl)-2-[2'-(3,4-dimethoxyphenyl)-5'-benzimidazolyl]benzimidazole, 5) and another having a methoxy and a hydroxyl group (5-(4-methylpiperazin-1-yl)-2-[2'[2''-(4-hydroxy-3-methoxyphenyl)-5' '-benzimidazolyl]-5'-benzimidazolyl]benzimidazole, 6) ortho to each other on the phenyl ring. The radiomodifying effects of these nontoxic ligands were investigated with a human glioma cell line exposed to low linear energy transfer radiation by determining cell survival and cell proliferation compared with effects of the parent compound, Hoechst 33342. Cytotoxicity assayed by analyzing clonogenicity, cell growth, and metabolic viability showed that both 5 and 6 were nontoxic at 100 microM after 72 h of exposure, whereas Hoechst 33342 resulted in lysis of 77% of these cells in 24 h. Macrocolony assay (clonogenicity) showed that 73%, 92%, and 10% of the cells survived when treated with 100 microM 5, 6, and Hoechst 33342, respectively. Both 5 and 6 did not affect the growth of BMG-1 cells. At 10 microM, 5 and 6 showed 82% and 37% protection against radiation-induced cell death (macrocolony assay) while 100% protection was observed against growth inhibition. Disubstitution of the phenyl ring has not only reduced cytotoxicity but also enhanced DNA-ligand stability, conferring high degree of radioprotection.

Preclinical Evaluation of DMA, a Bisbenzimidazole, as Radioprotector: Toxicity, Pharmacokinetics, and Biodistribution Studies in Balb/c Mice

“…In order to understand the mechanism involved for in vivo radioprotection, an ORAC-FL assay was performed to study the antioxidant capacity of DMA to cement the already proven free radical scavenging property of DMA (Tawar et al, 2003). This assay measures the loss of fluorescein fluorescence over time due to the peroxyl-radical formation by the breakdown of 2,2ʹ-azobis(2-amidino-propane) dihydrochloride (AAPH).…”

“…DMA was synthesized, characterized, and converted to its hydrochloride salt to make it water soluble and purified by highperformance liquid chromatography (HPLC) as per the procedure reported elsewhere (Tawar et al, 2003) (Fig. 1).…”

“…We have proved that DMA acts mechanistically as a radioprotector by reducing DNA damage and free-radical scavenging in mammalian cells. DMA was observed to be nonmutagenic and noncytotoxic to mammalian cells in comparison with the parent analog (Tawar et al, 2003(Tawar et al, , 2007. It showed inhibitory action at micromolar concentrations in a human epithelial cancer cell line (HeLa), a human breast cancer cell line (MCF7), a human glioma cell line (U87), and a human embryonic kidney cell line (HEK293) (Singh and Tandon, 2011).…”

3,4-Dimethoxyphenyl bis-benzimidazole, a novel DNA topoisomerase inhibitor that preferentially targets Escherichia coli topoisomerase I

“…13 We have synthesized several analogues of Hoechst 33342 with dichloro {DCA, 2-(2,4-dichlorophenyl)-5-[5-(4-methylpiperazin-1-yl)-1H-benzimidazol-2-yl]-1H-benzimidazole} and difluoro {DFA, 2-(2,4-difluorophenyl)-5-[5-(4-methylpiperazinyl)-1H-benzimidazol-2-yl]-1H-benzimidazole} substitutions on the phenyl ring and evaluated their activity against HuTOPI (Figure 1). 14 In the present study, we have evaluated DCA, DFA and DMA for their antibacterial activity against standard E. coli and resistant, clinical strains of E. coli isolated from UTI patients. The MIC and MBC study showed that DMA is more active against most of the resistant E. coli strains.…”

Interaction of DNA Minor Groove Binder Hoechst 33258 with Bovine Serum Albumin

“…8,9) Our group has also studied cellular responses of a few synthesized bisbenzimidazole derivatives in in-vitro systems. 10,11) Hoechst 33258 is a bisbenzimidazole molecule that binds strongly in the minor groove region of duplex DNA with predominant specificity for AT rich sequences. [12][13][14] Bisbenzimidazole compounds have also been reported to increase radiation induced cytotoxicity in several cancerous cell lines 15) and thus have an ability to improve radiotherapy.…”