WO2022162693A1 - Benzodiazepine compounds having anti-cancer activity and a process for preparation thereof - Google Patents

Benzodiazepine compounds having anti-cancer activity and a process for preparation thereof Download PDF

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WO2022162693A1
WO2022162693A1 PCT/IN2022/050063 IN2022050063W WO2022162693A1 WO 2022162693 A1 WO2022162693 A1 WO 2022162693A1 IN 2022050063 W IN2022050063 W IN 2022050063W WO 2022162693 A1 WO2022162693 A1 WO 2022162693A1
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benzodiazepine
predefined
reaction mixture
compound
preparation
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French (fr)
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Vidya Desai
Sinthiya Gawandi
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Vidya Desai
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/55—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having seven-membered rings, e.g. azelastine, pentylenetetrazole
    • A61K31/551—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having seven-membered rings, e.g. azelastine, pentylenetetrazole having two nitrogen atoms, e.g. dilazep
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D243/00—Heterocyclic compounds containing seven-membered rings having two nitrogen atoms as the only ring hetero atoms
    • C07D243/06—Heterocyclic compounds containing seven-membered rings having two nitrogen atoms as the only ring hetero atoms having the nitrogen atoms in positions 1 and 4
    • C07D243/10—Heterocyclic compounds containing seven-membered rings having two nitrogen atoms as the only ring hetero atoms having the nitrogen atoms in positions 1 and 4 condensed with carbocyclic rings or ring systems
    • C07D243/12—1,5-Benzodiazepines; Hydrogenated 1,5-benzodiazepines

Definitions

  • the present invention relates to benzodiazepine compounds used for anticancer activity and more particularly to 1,5 -benzodiazepine having specific substitution pattern.
  • the present invention also relates to a process for preparation of 1, 5-benzodiazepine derivatives,
  • Cancer is a chronic illness that has a high mortality rate. It is characterized by uncontrolled division of some cells in the body that leads to the formation of an abnormal mass of cells called as tumours. These cells can migrate to other parts of the body through metastasis and lead to the formation of new tumours. There are different types of cancers based on the tissues which they affect and organ systems where they occur.
  • the incidence of cancer in an individual can be linked to age, exposure to carcinogenic agents, genetics, etc.
  • Preventive measures for cancer include avoiding the risk factors and exposure to carcinogenic agents. Early diagnosis and early treatment of cancer can lead to high chances in the complete recovery of the patients.
  • Chemotherapy involves the use of drugs for the treatment of cancer.
  • the antineoplastic drugs are effective in the treatment of cancer as they target malignant or cancerous cells, and are not harmful to normal cells.
  • Some of the anticancer drugs include alkylating agents, antimetabolites, natural products and hormones. Over the years, a number of compounds have been researched and developed for their anti-cancer activity. Compounds such as Methotrexate, Paclitaxel, Cis-platin are routinely used in chemotherapy for the treatment of cancer.
  • Benzodiazepines are a class of compounds that are used for having a calming and sedative effect, treatment of fear, anxiety, tension, agitation and other states of mental disturbance. Benzodiazepines enhance the action of the neurotransmitter gamma-aminobutyric acid (GABA) helping in inhibition of anxiety by reducing nerve-impulse transmissions within the brain. Benzodiazepines are explored as potential anti-cancer agents for the treatment of cancer.
  • GABA neurotransmitter gamma-aminobutyric acid
  • the Chinese application CN112174901A by Henan Normal University discloses 1,3-benzodiazepine compound with anticancer activity.
  • the application describes process of synthesis of the compound from the substrates N-aryl amidines and propargyl alcohol ester compounds using rhodium or ruthenium catalyst.
  • the US patent US8426402B2 by Immunogen Inc describes benzodiazepine derivatives with antiproliferative activity, prepared using palladium catalyst.
  • Chinese Patent CN106902123B by Guo Xunxiang et al. discloses the application of 5 -hydrogen- 1,4- benzodiazepine compound for the treatment of liver cancer.
  • the 5 -hydrogen- 1,4- benzodiazepine compound has a good inhibition effect on human hepatocarcinoma (HCC), HepG2 and Huh7cell lines in seven tested 5-hydrogen-l,4-benzodiazepine compounds.
  • HCC human hepatocarcinoma
  • HepG2 HepG2
  • Huh7cell lines seven tested 5-hydrogen-l,4-benzodiazepine compounds.
  • 1.5-benzodiazepines Further they are costly, time consuming and are less environment friendly.
  • 1,5-benzodiazepines are the class of compounds that have been rarely explored for their anti-cancer activity. There is a growing interest in the research and development of 1,5-benzodiazepines as anti-cancer agents. There is a need to develop
  • the present invention relates to 1,5 -benzodiazepine derivatives used for anticancer activity and a process for preparation of 1,5-benzodiazepine derivatives.
  • Benzodiazepines are bicyclic nitrogen heterocyclic scaffold having a ring complex including a benzene ring fused with a diazepam ring.
  • the 1,5-benzodiazepine compounds have structural formula (I) as follows: wherein,
  • R1 is an aryl group
  • R2 is a methyl (-CH3) group
  • R3 is H
  • R4 is an alkyl group or -X (halogen).
  • R1 is an aryl group selected from 3-BrPh, 3-NO2Ph, 3,4-(OCH3)2Ph, 4-NO2Ph, 4- BrPh, 4-ClPh, 2-Naphthyl, 2-Pyridinyl, 4-Pyridinyl, 2-Naphthyl and R4 is selected from a methyl (-CH3) group or halogen groups such as Cl or Br.
  • the compound of Formula (I) and its derivatives exhibit their anti-cancer activity by tyrosine kinase inhibition, DNA intercalation leading to apoptotic cell death.
  • the compound of Formula (I) and its derivatives act as both tyrosine kinase and DHFR inhibitor. Further the compounds of the present invention show synergistic effect with methotrexate, an anticancer drug, when combined in a ratio of 1:1.
  • a process for preparation of 1,5 -benzodi azepine and its derivatives includes the steps of adding Immol of o-phenylenediamine and 2mmol of acetophenones to a round bottom flask followed by adding 5 mol% of a biocatalyst thiamine hydrochloride to the above reaction mixture and heating the reaction mixture on a water bath at a temperature of 70-80°C for a period of 2-4 hours; monitoring of reaction mixture till the completion of the reaction followed by quenching the reaction mixture in 15 ml of cold water followed by extracting using 30 ml of ethyl acetate and drying of final crude product using Na2SO4; purifying the crude product by column chromatography using a solvent mixture of petroleum ether- ethyl acetate in a ratio of 9: 1 to obtain the compound of Formula (I) and further analyzing the product purity using high performance liquid chromat
  • the present invention relates to 1,5 -benzodiazepine derivatives used for anticancer activity and a process for preparation of 1,5-benzodiazepine derivatives.
  • Benzodiazepines are bicyclic nitrogen heterocyclic scaffold having a ring complex including a benzene ring fused with a diazepam ring.
  • the invention discloses a compound and its derivatives represented by the Formula (I) given below: wherein,
  • R1 is an aryl group
  • R2 is a methyl (-CH3) group
  • R3 is H and
  • R4 is an alkyl group or -X (halogen).
  • R1 is an aryl group selected from 3-BrPh, 3- NO 2 Ph, 3,4-(OCH 3 ) 2 Ph, 4-NO 2 Ph, 4-BrPh, 4-ClPh, 2-Naphthyl, 2-Pyridinyl, 4- Pyridinyl, 2-Naphthyl.
  • R4 is selected from a methyl (-CH3) group or halogen groups such as Cl or Br.
  • the compound of Formula (I) and its derivatives exhibit good cytotoxicity against cervical and liver carcinoma cells acting as anti-cancer agents.
  • the compound of Formula (I) and its derivatives exhibit their anti-cancer activity by tyrosine kinase inhibition, DNA intercalation leading to apoptotic cell death.
  • the compound of Formula (I) and its derivatives act as both tyrosine kinase and DHFR inhibitor. Further the compounds of the present invention show synergistic effect with methotrexate, an anticancer drug, when combined in a ratio of 1:1.
  • the process of preparation of 1,5 -benzodiazepine and its derivatives includes the steps of: i. Adding a predefined quantity of o-phenylenediamine and a predefined quantity of acetophenones to a round bottom flask; ii. Adding a predefined quantity of a biocatalyst to the above reaction mixture and heating the reaction mixture on a water bath at a predefined temperature for a predefined period of time; iii. Monitoring of reaction mixture till the completion of the reaction; iv.
  • the predefined quantity of o-phenylenediamine is Immol and the predefined quantity of acetophenones is 2mmol.
  • the reaction proceeds with the addition of a predefined quantity of 5 mol% biocatalyst thiamine hydrochloride.
  • the reaction mixture is heated on water bath for a predefined period of time ranging from 2 to 4 hours at a predefined temperature ranging from 70-80°C.
  • the reaction was monitored at constant time intervals till the completion of the reaction.
  • the quenching of the reaction takes place with a predefined quantity of 15ml of cold water.
  • the predefined solvent used for extraction is ethyl acetate in a predefined quantity of 30 ml.
  • the predefined hygroscopic agent used for drying is Na2SC>4.
  • the predefined solvent mixture used for purifying the product by column chromatography is petroleum ether- ethyl acetate in a ratio of 9:1.
  • the process of the present invention yields fourteen unique 1,5 benzodiazepine derivatives with varied substitution patterns.
  • the present invention provides 1,5 benzodiazepine derivatives having varied substitution that possess anti-cancer activity.
  • the process for the preparation of the compound of Formula (I) and its derivatives is simple, cost effective and eco-friendly. Further the catalyst is used in solvent free conditions.
  • the invention explores the synergistic nature of the compound with a preexisting anti-cancer drug and studies their anticancer activity.
  • Example 1 Process for preparation of compound of Formula (I) o-Phenylene diamine (Immol) and a substituted ketone derivative (2mmol) are added to a round bottom flask. 5 mol% of thiamine hydrochloride (Vitamin Bl) acts used as a biocatalyst is added and the reaction mixture is heated on a water bath at a temperature around 70-80 C for 2 to 4 hours. The reaction is monitored periodically till its completion. The reaction mixture is then quenched in cold water (15 ml) and is further extracted using ethyl acetate (30 ml) and is dried over anhydrous sodium sulphate (Na2SC>4).
  • thiamine hydrochloride Vitamin Bl
  • the final crude product is obtained and purified using column chromatography using the organic solvent petroleum ether (PE): ethyl acetate (EtOAc) in the ratio of 9:1.
  • the purity of the product is further analyzed using high performance liquid chromatography (HPLC).
  • HPLC high performance liquid chromatography
  • Example 2 Studies demonstrating anti-cancer activity of 1,5-Benzodiazapiene i. In-vitro Tyrosine Kinase Inhibition
  • FBS Fetal Bovine Serum
  • the cells are seeded at a density of approximately lxl0 5 cells/well in a 96-well flat-bottom micro plate and maintained at 37°C in 95% humidity and 5% CO2 for overnight.
  • the cells are treated with different concentration of test samples and the cells are incubated for another 24 hours.
  • the cells in well are washed twice with phosphate buffer solution, and 1 ml of extraction buffer was added. By using cell scraper, the cells were recovered carefully and further the cells are centrifuged at 4°C for 10 min at 10,000 rpm. Collect the supernatant and store as further analysis.
  • Tyrosine Kinase Assay The collected supernatant was diluted approximately 25 times with the kinase reacting solution provided along with kit. The diluted control and treated sample are added to each well in duplicates. Further 10 pl of 40 mM ATP-2Na solution is added into each well and mixed. The solution is incubated for 30 min at 37°C. The samples are removed and the wells are washed thrice with wash buffer. A lOOpl of blocking solution is added into each well and incubated for 30 min at 37°C. The blocking solution is then discarded and 50 pl of an anti-phospho tyrosine - HRP solution is added into each well and incubated for 30 min at 37°C.
  • EFGR epidermal growth factor receptor
  • the in vitro cytotoxicity of the derivatives of 1,5-benzodiazapene is determined by MTT [(3-(4,5-dimethyl-2- thiazolyl) 2,5-diphenyl-2H-tetrazolium bromide)] assay against a panel of three different human cancer cell lines namely; HEPG2 (human liver carcinoma), HeLa (human cervical) and HEK-293T (Human embryonic kidney cells) and the normal cell line.
  • the cell lines as disclosed are seeded in a ninety-six (96) well flat-bottom micro plate containing Dulbecco’s Modified Eagle Media (DMEM) supplemented with 10% heat inactivated Fetal Bovine Serum (FBS) and 1% antibiotic- antimycotic 100X solution.
  • DMEM Modified Eagle Media
  • FBS Fetal Bovine Serum
  • antibiotic- antimycotic 100X solution The cells are maintained overnight at 37°C at 95% humidity in 5% CO2 and were seeded in separate ninety-six (96 well plates.
  • the compound concentration of 400, 200, 100, 50, 25, 12.5 pg/mL was treated and cells were incubated for 48 hours before the addition of MTT solution.
  • the wells are washed twice with phosphate buffer solution followed by addition of 20 p L of MTT solution and then they are incubated at 37°C. After 4 hours, formazan crystals are dissolved using 100 pL DMSO and absorbance was recorded at 570 nm
  • IC50 values of the screened compounds and the reference drugs, Paclitaxel and Methotrexate show significant antitumor activities with IC50 values in the range of 0.067-1.65 pM and this indicated that the 1,5 -benzodiazepine group can play a significant role in intensifying the antitumor effects. All the screened compounds showed better activity (0.067-0.21 pM) against HeLa cell lines compared to one of the reference compounds Paclitaxel (IC50 values 0.23 pM). One of the derivatives of 1,5-benzodiazpene showed an IC50 value in close proximity to the other standard drug Methotrexate (0.057 pM and 0.083 pM). iii.
  • the 1,5-benzodiazepine molecule was tested to check its activity as an intercalating agent between the DNA helix strands.
  • the intercalation between the DNA helix strands is further also proved by DNA binding studies assay with good results for 1,5- benzodiazepine. iv. Apoptosis
  • the mode of tumor cell death analysis was done by fluorescence activated cell sorter (FACS) caliber on HepG2 liver cancer cells treated with 20pM of selected compounds for 24 hours.
  • FACS fluorescence activated cell sorter
  • the flow cytometry analysis revealed that HepG2 cells treated with 1,5-benzodiazepine compounds showed apoptosis.
  • the most prevailing technique used to study cell apoptosis/ necrosis is flow cytometry using propidium iodide (PI) and annexin V- FITC as dyes to spot viable and dead cells. A negative control was used for the analysis.
  • This assay facilitates the detection of live cells (Ql-LL; AV-/PI-), early apoptotic cells (Ql-LR; AV+/PI-), late apoptotic cells (Ql-UR; AV+/PI+) and necrotic cells (Ql-UL; AV-/PI+).
  • the apoptosis is carried out by a flow cytometer.
  • the cell lines used for the assay are Hep G2 (Liver cancer) DMEM with low glucose, Fetal Bovine Serum (FBS) (Gibco, Invitrogen), Antibiotic - Antimycotic 100X solution (Thermo Fisher Scientific) and Annexin V-FITC Kit (Beckman Coulter) Tests Propidium Iodide (Sigma-Aldrich Methodology.
  • the cells were seeded in a 24-well flat bottom micro plate containing cover slips and maintained at 37 °C in a CO2 incubator for overnight.
  • the cells 20pg/ml of each sample compound were treated for 24 hrs.
  • the cells were washed with phosphate buffer solution (PBS) and centrifuged for 5 minutes at 500 x g at 4°C. After centrifugation the supernatant was discarded and the cell pellets were resuspended in ice-cold IX Binding Buffer to 1 x 10 5 per mL. Tubes are kept on ice and then 1 pL of Annexin V-FITC solution and 5 L PI were added and mixed gently. Tubes are kept on ice and incubated for 15 minutes in the dark. 400 pL of ice-cold IX binding buffer is added and mixed gently. The cell preparations are analyzed within 30 minutes by flowcytometry.
  • PBS phosphate buffer solution
  • the synthesized 1,5-benzodiazepines derivatives can effectively function on targeted enzymes such as tyrosine kinase and dihydrofolate reductase (DHFR).
  • DHFR dihydrofolate reductase
  • the experiment studies the efficiency /bioactivity of 1,5 -benzodiazepine by checking the enhancement in the activity and reduction of toxicity of the drug Methotrexate.
  • the combination of standard drug Methotrexate with the 1,5 -benzodiazepine resulted in improved activity with IC50 values of 0.046 ⁇ 0.002 against HeLa cell line and 0.057 ⁇ 0.002 against HEPG2 cell line.
  • IC50 values 0.046 ⁇ 0.002 against HeLa cell line
  • the IC50 values obtained for 1,5-benzodiazepine in synergism with drug Methotrexate show a difference of 0.015 micromolar as compared to the IC50 value of the drug Methotrexate alone.
  • the percentage of late apoptotic cells induced by the combination was 67.6%.
  • the molecular docking studies revealed that the compounds are good DHFR inhibitors.
  • the obtained results from the studies were encouraging and 1,5- benzodiazepines and the derivatives were used with Methotrexate (acts as an DHFR enzyme inhibitor) synergistically.
  • the docking results were screened thoroughly and screening of the best hits resulted in testing the compounds for in-vitro anti-cancer activity against two cancer cell lines and one human cell line.
  • the organic compounds exhibited good cytotoxicity against cervical and liver carcinoma cells.
  • the apoptotic cell pathway studies indicated late apoptotic cell death.
  • the organic compound proves its potency as an anticancer agent.

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Abstract

The present invention relates to a class of benzodiazepine compounds and a process for preparation thereof. The 1,5-benzodiazepine compounds effectively 5 function on targeted enzymes such as tyrosine kinase and dihydrofolate reductase (DHFR) and also acts as a DNA intercalating agent therefore demonstrating its anti-cancer activity. The 1,5-benzodiazepine compounds also act synergistically with a conventional anti-cancer drug such as Methotrexate. The process of preparation of 1,5-benzodiazepine and its derivatives includes the steps of reacting o-10 phenylenediamine and acetophenones in presence of a biocatalyst thiamine hydrochloride to obtain the compound of Formula (I). The process for preparation of the 1,5-benzodiazepine compounds is simple, cost effective and eco-friendly approach as it takes place in the absence of a solvent and uses a biocatalyst.

Description

“BENZODIAZEPINE COMPOUNDS HAVING ANTI-CANCER ACTIVITY AND A PROCESS FOR PREPARATION THEREOF”
FIELD OF THE INVENTION
The present invention relates to benzodiazepine compounds used for anticancer activity and more particularly to 1,5 -benzodiazepine having specific substitution pattern. The present invention also relates to a process for preparation of 1, 5-benzodiazepine derivatives,
BACKGROUND OF THE INVENTION
Cancer is a chronic illness that has a high mortality rate. It is characterized by uncontrolled division of some cells in the body that leads to the formation of an abnormal mass of cells called as tumours. These cells can migrate to other parts of the body through metastasis and lead to the formation of new tumours. There are different types of cancers based on the tissues which they affect and organ systems where they occur.
The incidence of cancer in an individual can be linked to age, exposure to carcinogenic agents, genetics, etc. Preventive measures for cancer include avoiding the risk factors and exposure to carcinogenic agents. Early diagnosis and early treatment of cancer can lead to high chances in the complete recovery of the patients.
Various new age treatments are available for cancer. These include radiation therapy, surgery, chemotherapy and immunotherapy. Chemotherapy involves the use of drugs for the treatment of cancer. The antineoplastic drugs are effective in the treatment of cancer as they target malignant or cancerous cells, and are not harmful to normal cells. Some of the anticancer drugs include alkylating agents, antimetabolites, natural products and hormones. Over the years, a number of compounds have been researched and developed for their anti-cancer activity. Compounds such as Methotrexate, Paclitaxel, Cis-platin are routinely used in chemotherapy for the treatment of cancer.
Benzodiazepines are a class of compounds that are used for having a calming and sedative effect, treatment of fear, anxiety, tension, agitation and other states of mental disturbance. Benzodiazepines enhance the action of the neurotransmitter gamma-aminobutyric acid (GABA) helping in inhibition of anxiety by reducing nerve-impulse transmissions within the brain. Benzodiazepines are explored as potential anti-cancer agents for the treatment of cancer.
The Chinese application CN112174901A by Henan Normal University discloses 1,3-benzodiazepine compound with anticancer activity. The application describes process of synthesis of the compound from the substrates N-aryl amidines and propargyl alcohol ester compounds using rhodium or ruthenium catalyst. The US patent US8426402B2 by Immunogen Inc describes benzodiazepine derivatives with antiproliferative activity, prepared using palladium catalyst. Chinese Patent CN106902123B by Guo Xunxiang et al. discloses the application of 5 -hydrogen- 1,4- benzodiazepine compound for the treatment of liver cancer. The 5 -hydrogen- 1,4- benzodiazepine compound has a good inhibition effect on human hepatocarcinoma (HCC), HepG2 and Huh7cell lines in seven tested 5-hydrogen-l,4-benzodiazepine compounds. The processes disclosed in the prior art result in end products other than
1.5-benzodiazepines. Further they are costly, time consuming and are less environment friendly.
1,5-benzodiazepines are the class of compounds that have been rarely explored for their anti-cancer activity. There is a growing interest in the research and development of 1,5-benzodiazepines as anti-cancer agents. There is a need to develop
1.5-benzodiazepines of varied substitution pattern and a process of synthesis of benzodiazepine derivatives that is cost effective, time saving, environment friendly and causes minimum hazard to the living beings. SUMMARY OF THE INVENTION
The present invention relates to 1,5 -benzodiazepine derivatives used for anticancer activity and a process for preparation of 1,5-benzodiazepine derivatives. Benzodiazepines are bicyclic nitrogen heterocyclic scaffold having a ring complex including a benzene ring fused with a diazepam ring. The 1,5-benzodiazepine compounds have structural formula (I) as follows:
Figure imgf000004_0001
wherein,
R1 is an aryl group;
R2 is a methyl (-CH3) group; R3 is H and
R4 is an alkyl group or -X (halogen).
R1 is an aryl group selected from 3-BrPh, 3-NO2Ph, 3,4-(OCH3)2Ph, 4-NO2Ph, 4- BrPh, 4-ClPh, 2-Naphthyl, 2-Pyridinyl, 4-Pyridinyl, 2-Naphthyl and R4 is selected from a methyl (-CH3) group or halogen groups such as Cl or Br. The compound of Formula (I) and its derivatives exhibit their anti-cancer activity by tyrosine kinase inhibition, DNA intercalation leading to apoptotic cell death. The compound of Formula (I) and its derivatives act as both tyrosine kinase and DHFR inhibitor. Further the compounds of the present invention show synergistic effect with methotrexate, an anticancer drug, when combined in a ratio of 1:1.
In a preferred embodiment, a process for preparation of 1,5 -benzodi azepine and its derivatives is disclosed. The process of preparation of 1,5 -benzodiazepine and its derivatives includes the steps of adding Immol of o-phenylenediamine and 2mmol of acetophenones to a round bottom flask followed by adding 5 mol% of a biocatalyst thiamine hydrochloride to the above reaction mixture and heating the reaction mixture on a water bath at a temperature of 70-80°C for a period of 2-4 hours; monitoring of reaction mixture till the completion of the reaction followed by quenching the reaction mixture in 15 ml of cold water followed by extracting using 30 ml of ethyl acetate and drying of final crude product using Na2SO4; purifying the crude product by column chromatography using a solvent mixture of petroleum ether- ethyl acetate in a ratio of 9: 1 to obtain the compound of Formula (I) and further analyzing the product purity using high performance liquid chromatography (HPLC). The process of the present invention yields fourteen unique 1,5 benzodiazepine derivatives with varied substitution patterns.
DETAILED DESCRIPTION OF THE INVENTION
The invention described herein is explained using specific exemplary details for better understanding. However, the invention disclosed can be worked on by a person skilled in the art without the use of these specific details.
References in the specification to "one embodiment" or "an embodiment"means that particular feature, structure, characteristic, or function described in connection with the embodiment is included in at least one embodiment of the invention. The appearances of the phrase “in one embodiment” in various places in the specification are not necessarily all referring to the same embodiment.
References in the specification to “preferred embodiment” means that a particular feature, structure, characteristic, or function described in detail thereby omitting known constructions and functions for clear description of the present invention.
The foregoing description of specific embodiments of the present invention has been presented for purposes of illustration and description. They are not intended to be exhaustive or to limit the present invention to the precise forms disclosed, and obviously many modifications and variations are possible in light of the above teaching.
The present invention relates to 1,5 -benzodiazepine derivatives used for anticancer activity and a process for preparation of 1,5-benzodiazepine derivatives. Benzodiazepines are bicyclic nitrogen heterocyclic scaffold having a ring complex including a benzene ring fused with a diazepam ring.
In an embodiment of the present invention, the invention discloses a compound and its derivatives represented by the Formula (I) given below:
Figure imgf000006_0001
wherein,
R1 is an aryl group; R2 is a methyl (-CH3) group;
R3 is H and
R4 is an alkyl group or -X (halogen).
In this embodiment, R1 is an aryl group selected from 3-BrPh, 3- NO2Ph, 3,4-(OCH3)2Ph, 4-NO2Ph, 4-BrPh, 4-ClPh, 2-Naphthyl, 2-Pyridinyl, 4- Pyridinyl, 2-Naphthyl. R4 is selected from a methyl (-CH3) group or halogen groups such as Cl or Br.
In accordance with the embodiment of the present invention, the compound of Formula (I) and its derivatives exhibit good cytotoxicity against cervical and liver carcinoma cells acting as anti-cancer agents. The compound of Formula (I) and its derivatives exhibit their anti-cancer activity by tyrosine kinase inhibition, DNA intercalation leading to apoptotic cell death.
The compound of Formula (I) and its derivatives act as both tyrosine kinase and DHFR inhibitor. Further the compounds of the present invention show synergistic effect with methotrexate, an anticancer drug, when combined in a ratio of 1:1.
Now a preferred process for the preparation of compound of Formula (I) and its derivatives in accordance with the present invention is disclosed. The process of preparation of 1,5 -benzodiazepine and its derivatives includes the steps of: i. Adding a predefined quantity of o-phenylenediamine and a predefined quantity of acetophenones to a round bottom flask; ii. Adding a predefined quantity of a biocatalyst to the above reaction mixture and heating the reaction mixture on a water bath at a predefined temperature for a predefined period of time; iii. Monitoring of reaction mixture till the completion of the reaction; iv. Quenching the reaction mixture in a predefined quantity of cold water followed by extracting using a predefined quantity of a predefined solvent and drying of final crude product with the use of a predefined hygroscopic agent; v. Purifying the crude product by column chromatography using a predefined solvent mixture to obtain the compound of Formula (I); and vi. Further analyzing the product purity using high performance liquid chromatography (HPLC)
Figure imgf000008_0001
In accordance with this embodiment of the present invention the predefined quantity of o-phenylenediamine is Immol and the predefined quantity of acetophenones is 2mmol. The reaction proceeds with the addition of a predefined quantity of 5 mol% biocatalyst thiamine hydrochloride. The reaction mixture is heated on water bath for a predefined period of time ranging from 2 to 4 hours at a predefined temperature ranging from 70-80°C. The reaction was monitored at constant time intervals till the completion of the reaction. The quenching of the reaction takes place with a predefined quantity of 15ml of cold water. The predefined solvent used for extraction is ethyl acetate in a predefined quantity of 30 ml. The predefined hygroscopic agent used for drying is Na2SC>4. The predefined solvent mixture used for purifying the product by column chromatography is petroleum ether- ethyl acetate in a ratio of 9:1.
The process of the present invention yields fourteen unique 1,5 benzodiazepine derivatives with varied substitution patterns.
Advantageously, the present invention provides 1,5 benzodiazepine derivatives having varied substitution that possess anti-cancer activity. The process for the preparation of the compound of Formula (I) and its derivatives is simple, cost effective and eco-friendly. Further the catalyst is used in solvent free conditions.
The invention explores the synergistic nature of the compound with a preexisting anti-cancer drug and studies their anticancer activity.
EXAMPLES
Only a few examples and implementations are disclosed. Variations, modifications and enhancements to the described examples and implementations and other implementations can be made based on what is disclosed.
Examples are set forth herein below and are illustrative of different amounts and types of reactants and reaction conditions that can be utilized in practicing the disclosure. It will be apparent, however, that the disclosure can be practiced with other amounts and types of reactants and reaction conditions than those used in the examples, and the resulting devices various different properties and uses in accordance with the disclosure above and as pointed out hereinafter.
Example 1: Process for preparation of compound of Formula (I) o-Phenylene diamine (Immol) and a substituted ketone derivative (2mmol) are added to a round bottom flask. 5 mol% of thiamine hydrochloride (Vitamin Bl) acts used as a biocatalyst is added and the reaction mixture is heated on a water bath at a temperature around 70-80 C for 2 to 4 hours. The reaction is monitored periodically till its completion. The reaction mixture is then quenched in cold water (15 ml) and is further extracted using ethyl acetate (30 ml) and is dried over anhydrous sodium sulphate (Na2SC>4). The final crude product is obtained and purified using column chromatography using the organic solvent petroleum ether (PE): ethyl acetate (EtOAc) in the ratio of 9:1. The purity of the product is further analyzed using high performance liquid chromatography (HPLC). The reaction provides fourteen (14) derivatives of 1,5 -benzodiazepine (BZ) with different substituents in them.
Example 2: Studies demonstrating anti-cancer activity of 1,5-Benzodiazapiene i. In-vitro Tyrosine Kinase Inhibition
In silico molecular docking study was performed on the compounds with Molegro Virtual Docker (MVD-2007, 6.0). The molecular docking study reveals that 1,5-benzodiazepines act as good inhibitors of tyrosine kinase due to characteristic features. This was validated by in-vitro EGFR tyrosine kinase assay.
Materials used for the in-vitro Tyrosine Kinase Inhibition
1. Fetal Bovine Serum (FBS)
2. Antibiotic-Antimycotic 100X solution
3. Well plates
4. Universal Tyrosine Kinase Assay kit
Methodology
Cell Treatment: The cells are seeded at a density of approximately lxl05cells/well in a 96-well flat-bottom micro plate and maintained at 37°C in 95% humidity and 5% CO2 for overnight. The cells are treated with different concentration of test samples and the cells are incubated for another 24 hours. The cells in well are washed twice with phosphate buffer solution, and 1 ml of extraction buffer was added. By using cell scraper, the cells were recovered carefully and further the cells are centrifuged at 4°C for 10 min at 10,000 rpm. Collect the supernatant and store as further analysis.
Tyrosine Kinase Assay: The collected supernatant was diluted approximately 25 times with the kinase reacting solution provided along with kit. The diluted control and treated sample are added to each well in duplicates. Further 10 pl of 40 mM ATP-2Na solution is added into each well and mixed. The solution is incubated for 30 min at 37°C. The samples are removed and the wells are washed thrice with wash buffer. A lOOpl of blocking solution is added into each well and incubated for 30 min at 37°C. The blocking solution is then discarded and 50 pl of an anti-phospho tyrosine - HRP solution is added into each well and incubated for 30 min at 37°C. The antibody solution is discarded and each well is then washed 4 times with washing buffer. 100 pl of HRP substrate solution (TMBZ) is added into each well. The solution is incubated for 30 min at 37°C. Lastly, a 100 pl of stop solution is added into each well in the same order as HRP substrate solution. Absorbance is measured at 450 nm with a plate reader.
The docking studies of the compounds against EFGR-Tyrosine kinase proved the action of the compounds by inhibiting protein tyrosine kinase. The enzyme inhibition assay was also performed against epidermal growth factor receptor (EFGR) tyrosine kinase and led to promising results for the molecules.
Results: The IC50 values of the synthesized compounds ranged from 0.156 to 0.514 pM, which further proved that 1,5-benzodiazepines have the potential to be good anticancer agents with tyrosine kinase inhibitory action. ii. In-vitro MTT assay:
The in vitro cytotoxicity of the derivatives of 1,5-benzodiazapene is determined by MTT [(3-(4,5-dimethyl-2- thiazolyl) 2,5-diphenyl-2H-tetrazolium bromide)] assay against a panel of three different human cancer cell lines namely; HEPG2 (human liver carcinoma), HeLa (human cervical) and HEK-293T (Human embryonic kidney cells) and the normal cell line.
Procedure for the MTT assay:
The cell lines as disclosed are seeded in a ninety-six (96) well flat-bottom micro plate containing Dulbecco’s Modified Eagle Media (DMEM) supplemented with 10% heat inactivated Fetal Bovine Serum (FBS) and 1% antibiotic- antimycotic 100X solution. The cells are maintained overnight at 37°C at 95% humidity in 5% CO2 and were seeded in separate ninety-six (96 well plates. The compound concentration of 400, 200, 100, 50, 25, 12.5 pg/mL was treated and cells were incubated for 48 hours before the addition of MTT solution. The wells are washed twice with phosphate buffer solution followed by addition of 20 p L of MTT solution and then they are incubated at 37°C. After 4 hours, formazan crystals are dissolved using 100 pL DMSO and absorbance was recorded at 570 nm using microplate reader. The calculation of the IC50 values was carried out using graph pad prism version 5.1.
Results: The IC50 values of the screened compounds and the reference drugs, Paclitaxel and Methotrexate show significant antitumor activities with IC50 values in the range of 0.067-1.65 pM and this indicated that the 1,5 -benzodiazepine group can play a significant role in intensifying the antitumor effects. All the screened compounds showed better activity (0.067-0.21 pM) against HeLa cell lines compared to one of the reference compounds Paclitaxel (IC50 values 0.23 pM). One of the derivatives of 1,5-benzodiazpene showed an IC50 value in close proximity to the other standard drug Methotrexate (0.057 pM and 0.083 pM). iii. Intercalation of 1,5-benzodiazepine between the DNA helix strands The molecular docking studies revealed that the ligands get intercalated in the minor grove of the DNA fragment. N 1 atom of benzodiazepine forms hydrogen bond with Thiamine while N atom of 4-pyridine ring forms hydrogen bond with adenine of the chain B of the DNA fragment. The 2-pyridine ring exhibits pi-stacked in-between two adenine rings of the DNA chain A. DNA binding assay also proved the same.
After achieving promising results and increase in the late apoptotic cells, the 1,5-benzodiazepine molecule was tested to check its activity as an intercalating agent between the DNA helix strands. The intercalation between the DNA helix strands is further also proved by DNA binding studies assay with good results for 1,5- benzodiazepine. iv. Apoptosis
For further investigative studies relating to the mechanism underlying the antiproliferative effect of these potent compounds, the mode of tumor cell death analysis was done by fluorescence activated cell sorter (FACS) caliber on HepG2 liver cancer cells treated with 20pM of selected compounds for 24 hours. The flow cytometry analysis revealed that HepG2 cells treated with 1,5-benzodiazepine compounds showed apoptosis. The most prevailing technique used to study cell apoptosis/ necrosis is flow cytometry using propidium iodide (PI) and annexin V- FITC as dyes to spot viable and dead cells. A negative control was used for the analysis. This assay facilitates the detection of live cells (Ql-LL; AV-/PI-), early apoptotic cells (Ql-LR; AV+/PI-), late apoptotic cells (Ql-UR; AV+/PI+) and necrotic cells (Ql-UL; AV-/PI+).
Procedure for Apoptosis:
Annexin V- FITC/ Propidium iodide dual staining assay
The apoptosis is carried out by a flow cytometer. The cell lines used for the assay are Hep G2 (Liver cancer) DMEM with low glucose, Fetal Bovine Serum (FBS) (Gibco, Invitrogen), Antibiotic - Antimycotic 100X solution (Thermo Fisher Scientific) and Annexin V-FITC Kit (Beckman Coulter) Tests Propidium Iodide (Sigma-Aldrich Methodology. The cells were seeded in a 24-well flat bottom micro plate containing cover slips and maintained at 37 °C in a CO2 incubator for overnight. The cells 20pg/ml of each sample compound were treated for 24 hrs. After the incubation, the cells were washed with phosphate buffer solution (PBS) and centrifuged for 5 minutes at 500 x g at 4°C. After centrifugation the supernatant was discarded and the cell pellets were resuspended in ice-cold IX Binding Buffer to 1 x 105 per mL. Tubes are kept on ice and then 1 pL of Annexin V-FITC solution and 5 L PI were added and mixed gently. Tubes are kept on ice and incubated for 15 minutes in the dark. 400 pL of ice-cold IX binding buffer is added and mixed gently. The cell preparations are analyzed within 30 minutes by flowcytometry.
Results: The percentage of late apoptotic cells induced by compounds was in the range (10-65%).
Example 3: Studies demonstrating synergism with other anti-cancer drugs
The synthesized 1,5-benzodiazepines derivatives can effectively function on targeted enzymes such as tyrosine kinase and dihydrofolate reductase (DHFR). The experiment studies the efficiency /bioactivity of 1,5 -benzodiazepine by checking the enhancement in the activity and reduction of toxicity of the drug Methotrexate. The combination of standard drug Methotrexate with the 1,5 -benzodiazepine resulted in improved activity with IC50 values of 0.046 ± 0.002 against HeLa cell line and 0.057 ± 0.002 against HEPG2 cell line. Thus, proving the potency of the compound to be good leads for synergistic anticancer activity.
The IC50 values obtained for 1,5-benzodiazepine in synergism with drug Methotrexate show a difference of 0.015 micromolar as compared to the IC50 value of the drug Methotrexate alone. The percentage of late apoptotic cells induced by the combination was 67.6%. The molecular docking studies revealed that the compounds are good DHFR inhibitors. The obtained results from the studies were encouraging and 1,5- benzodiazepines and the derivatives were used with Methotrexate (acts as an DHFR enzyme inhibitor) synergistically. The docking results were screened thoroughly and screening of the best hits resulted in testing the compounds for in-vitro anti-cancer activity against two cancer cell lines and one human cell line. The organic compounds exhibited good cytotoxicity against cervical and liver carcinoma cells. The apoptotic cell pathway studies indicated late apoptotic cell death. The organic compound proves its potency as an anticancer agent.
Inference: The above studies demonstrate that 1,5-benzodiazepine act as a tyrosine kinase inhibitor, DNA intercalating agent and also acts synergistically with standard drug Methotrexate.
The embodiments were chosen and described in order to best explain the principles of the present invention and its practical application, to thereby enable others, skilled in the art to best utilize the present invention and various embodiments with various modifications as are suited to the particular use contemplated.
It is understood that various omission and substitutions of equivalents are contemplated as circumstance may suggest or render expedient, but such are intended to cover the application or implementation without departing from the spirit or scope of the present invention.

Claims

Claims:
1. A 1,5-benzodiazepine compound comprising structural formula (I) as follows:
Figure imgf000016_0001
wherein,
R1 is an aryl group;
R2 is a methyl (-CH3) group;
R3 is H and
R4 is an alkyl group or -X (halogen).
2. A 1,5-benzodiazepine compound as claimed in Claim 1, wherein R1 is an aryl group selected from 3-BrPh, 3-NO2Ph, 3,4-(OCH3)2Ph, 4-NO2Ph, 4-BrPh, 4-ClPh, 2-Naphthyl, 2-Pyridinyl, 4-Pyridinyl, 2-Naphthyl.
3. A 1,5-benzodiazepine compound as claimed in Claim 1, wherein R4 is selected from a methyl (-CH3) group or halogen groups such as Cl or Br.
4. A process for the preparation of 1,5-benzodiazepine compound as claimed in Claim 1, comprising the steps of: a) adding predefined quantity of o-phenylenediamine and predefined quantity of acetophenones to a round bottom flask; b) adding predefined quantity of a biocatalyst to the above reaction mixture and heating the reaction mixture on a water bath at a predefined temperature for a predefined period of time; c) monitoring of reaction mixture till the completion of the reaction; d) quenching the reaction mixture in predefined quantity of cold water followed by extracting using predefined quantity of ethyl acetate and drying of final crude product using a predefined hygroscopic agent; e) purifying the crude product by column chromatography using a predefined solvent mixture in a predefined ratio to obtain the compound of Formula (I); and f) further analyzing the product purity using high performance liquid chromatography (HPLC). A process for the preparation of 1,5-benzodiazepine compound as claimed in Claim 4, comprising the steps of: a) adding Immol of o-phenylenediamine and 2mmol of acetophenones to a round bottom flask; b) adding 5 mol% of a biocatalyst thiamine hydrochloride to the above reaction mixture and heating the reaction mixture on a water bath at a temperature of 70-80°C for a period of 2-4 hours; c) monitoring of reaction mixture till the completion of the reaction; d) quenching the reaction mixture in 15 ml of cold water followed by extracting using 30 ml of ethyl acetate and drying of final crude product using Na2SC>4; e) purifying the crude product by column chromatography using a solvent mixture of petroleum ether- ethyl acetate in a ratio of 9:1 to obtain the compound of Formula (I); and f) further analyzing the product purity using high performance liquid chromatography (HPLC). A combination including 1,5-benzodiazepine compound of Claim 1 along with an anticancer agent in a predefined ratio, wherein the 1,5-benzodiazepine compounds show synergistic effect with the anticancer agent. A combination including 1,5-benzodiazepine compound of Claim 1 along with Methotrexate in a ratio of 1:1, wherein the 1,5-benzodiazepine compounds show synergistic effect with methotrexate.
17
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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
MX2007008753A (en) * 2005-01-19 2007-09-26 Zeria Pharm Co Ltd Antitumor agent.
WO2014128283A1 (en) * 2013-02-22 2014-08-28 Centre National De La Recherche Scientifique Use of compositions obtained by calcining particular metal-accumulating plants for implementing catalytical reactions
IN202021007824A (en) * 2020-02-24 2021-08-27 A. Peerzade Nargisbano

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
MX2007008753A (en) * 2005-01-19 2007-09-26 Zeria Pharm Co Ltd Antitumor agent.
WO2014128283A1 (en) * 2013-02-22 2014-08-28 Centre National De La Recherche Scientifique Use of compositions obtained by calcining particular metal-accumulating plants for implementing catalytical reactions
IN202021007824A (en) * 2020-02-24 2021-08-27 A. Peerzade Nargisbano

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
NEOCHORITIS CONSTANTINOS G., NEOCHORITIS CONSTANTINOS, TSOLERIDIS CONSTANTINOS, STEPHANIDOU STEPHANATOU JULIA, KONTOGIORGIS CHRIST: "1,5-Benzoxazepines vs 1,5-Benzodiazepines. One-Pot Microwave-Assisted Synthesis and Evaluation for Antioxidant Activity and Lipid Peroxidation Inhibition", JOURNAL OF MEDICINAL CHEMISTRY, AMERICAN CHEMICAL SOCIETY, US, vol. 53, no. 23, 9 December 2010 (2010-12-09), US , pages 8409 - 8420, XP055960497, ISSN: 0022-2623, DOI: 10.1021/jm100739n *
ORLOV V. D., ORLOV, DESENKO, KIROGA: "Aromatic derivatives of 2,3-dihydro-1H-1,5-benzodiazepine", CHEMISTRY OF HETEROCYCLIC COMPOUNDS, SPRINGER US, NEW YORK, vol. 23, no. 3, 1 March 1987 (1987-03-01), New York , pages 311 - 315, XP055960496, ISSN: 0009-3122, DOI: 10.1007/BF00761991 *

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