الفهرس 
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Abstract
Cancer is one of the most common health problems worldwide. It is defined as the abnormal and excessive growth of the tissue and it is classified into benign and malignant tumors. Breast cancer is a tumor formed in breast tissue including; lobules (milk producing glands), ducts (transport milk from glands to nipples) and the cell surrounding them. It is also the second cause of cancer death in women worldwide. Benzimidazole is one of the most important heterocyclic compounds due to its numerous biological activity as known in the literature. Herein, we focus our attention on the anti-breast cancer activity of benzimidazoles on cellular and molecular levels. Our work aims to design new hybrids structurally containing benzimidazole and other pharmacophores to develop new, selective and more effective anti-breast cancer agents. In order to characterize targeting breast cancer on a molecular level, binding to 15N-labeled Pin1 enzyme was conducted using state-of-the-art 2D NMR 15N-1H HSQC (Heteronuclear Single Quantum Coherence Spectroscopy) NMR experiments. Human Pin1 (protein interacting with NIMA “never in mitosis gene a”) is drawing increasing attention as an auspicious target for cancer treatment as it is involved in diverse processes in cancer development, also it is able to bind to estrogen receptor so that, misregulation and overexpression of Pin1 can cause a various cancers, particularly breast cancer, making Pin1 a potential target for anticancer therapy. In order to form our target new compounds the following different four schemes were adopted: The first scheme involved preparation of a novel series of benzimidazole dithiocarbamates through the reaction of different secondary amines with dry Na3PO4 and carbon disulfide in dry DMF to form dithiocarbamate salt, followed by addition of 2-chloromethylbenzimidazole derivatives 66 or 67 to give the desired hybrids 68-78 in excellent yields. The second scheme described the preparation of a series of benzimidazole thiopropyl compounds through reaction of 2-thiobenzimidazole derivatives 87 and 88 with bromochloropropane and anhydrous K2CO3 in dry DMF to give the intermediates 89 and 90, then stirring of these intermediates with appropriate secondary amine and anhydrous K2CO3 in dry DMF to yield the desired compounds 91-100. The third scheme included the synthesis of chalcone intermediate followed by its cyclization to give five and six membered heterocyclic rings through the following reactions: 1. Reaction of 2-acetyl-5,6-dichloro-benzimidazole (102) with 4- dimethylaminobenzaldehyde and NaOH in ethanol to give chalcone intermediate 103. 2. Heating chalcone 103 with hydrazine hydrate in ethanol give compound 104. 3. Refluxing of chalcone 103 and malononitrile in presence of ammonium acetate or piperidine in ethanol to give compound 105 or 106, respectively. 4. Cyclization reaction of chalcone 103 with hydroxylamine hydrochloride in the presence of potassium hydroxide produced isoxazole derivative 107. Finally the fourth scheme described the synthesis of pyrazanobenzimidazoles or pyridinobenzimidazoles through the following reactions: 1. Stirring of 2-acetyl-5,6-dichloro-benzimidazole (102) with bromophenacylbromide and anhydrous K2CO3 in acetone to give the intermediate 108. 2. The new benzimidazopyrazine derivatives 109, 110, 111 and 112 were obtained by heating the intermediate 108 with p-toluidine, o-toluidine, benzylamine and ammonium acetate, respectively in gl acetic acid. 3. Refluxing of intermediate 108 with K2CO3 in n-butanol to yield the pyridinobenzimidazole derivative 113. The previously mentioned reactions resulted in thirty three new compounds (thirty new finals and three new intermediates) that were not mentioned in the literature. Structure elucidation of the newly synthesized compounds was achieved through different spectroscopic techniques (1H, 13C 1D & 2D NMR such as 13C-APT and 1H-1H COSY, ESI-MS and IR) and microanalytical analyses. The preparation of the previously mentioned new compounds (Schemes 1-4) needed the formation of the following intermediates as guided in published literatures: 1. 2-(Chloromethyl)-5,6-dimethyl-(1H)-benzo[d]imidazole (66) and 5,6-dichloro- 2-(chloromethyl)-(1H)-benzo[d]imidazole (67). 2. 1H-Benzo[d]imidazole-2-thiol (87) or 5-chloro-(1H)-benzo[d]imidazole-2-thiol (88). 3. 2-((3-Chloropropyl)thio)-(1H)-benzo[d]imidazole (89). 4. 1-(5,6-Dichloro-(1H)-benzo[d]imidazol-2-yl)ethan-1-ol (101). 5. 1-(5,6-Dichloro-(1H)-benzo[d]imidazol-2-yl)ethan-1-one (102). Biological study for the newly synthesized compounds was performed against breast cancer cell line (MCF-7), and the most active compounds were further subjected to Normal Human Lung Fibroblast (WI38) that gives an indication of their safety. It was found that most of them exhibit high cytotoxic activity against breast cancer (MCF-7) and low cytotoxic activity against normal (WI38) cell line. Compounds 68, 78, 97, 104, 107 and 111 possess the highest anti-breast cancer activity against MCF-7 cell line were selected for Pin1 inhibition assay against tannic acid as reference drug. Compounds 78 and 107 were examined for their effect on cell cycle progression and their ability to apoptosis induction. The mechanistic evaluation of apoptosis induction was demonstrated by triggering intrinsic apoptotic pathway via inducing ROS accumulation, increasing Bax, decreasing Bcl-2 and activation of caspases 6, 7 and 9. In order to characterize the interaction between some of the newly synthesized compounds and Pin1, a new NMR multidimensional technique for drug design was adopted where a 2D 15N-1H HSQC NMR experiments were conducted. This state-of-the-art NMR experiment allowed us to monitor the interaction potentially targeting breast cancer on molecular level. Compounds 68, 75, 94 and 111 showed chemical shift perturbation (peak shifting or peak disappearance) indicating binding to Pin1 target. Molecular docking experiments were performed using ‘Molecular Operating Environment’ software and it was quite valuable to explain the binding mode of active derivatives via hydrogen bonding. Also, additional contact preferences and surface mapping studies stated the similarity pattern between active newly synthesized candidates which may pave the way for more precise anti-breast cancer target optimization.