Computer Based Design and Synthesis of Some Novel Thiazole Derivatives Bearing a Sulfonamide Moiety and Studying Their Potential Synergistic Anticancer Effect With γ-Irradiation
Thesis presented by
Aiten Mahmoud Mohamed Soliman
Bachelor Degree in Pharmaceutical Sciences Faculty of Pharmacy Ain Shams University (2006)
Submitted for the partial fulfillment of M.Sc. Degree in Pharmaceutical Sciences (Pharmaceutical Chemistry)
Prof. Dr. Dalal Abdel-Rahman Prof. Dr. Mostafa Mohamed Ghorab Abou El Ella Professor of Pharmaceutical Chemistry Professor of Applied Organic Chemistry Head of Pharmaceutical Chemistry Department Drug Radiation Research Department, Faculty of Pharmacy National Centre for Radiation Research and Ain Shams University Technology, Atomic Energy Authority
Prof. Dr. Helmy Ismail Heiba Assoc. Professor of Applied Organic Chemistry, Drug Radiation Research Department National Centre for Radiation Research and Technology Atomic Energy Authority
Faculty of Pharmacy Ain Shams University (2011)
Acknowledgement
Firstly, I want to thank Allah who gave me the patience and support to achieve my goal.
I wish to express my unlimited thanks and gratitude to Prof. Dr. Dalal Abdel-Rahman Abou El-Ella, Prof. of Pharmaceutical chemistry, Faculty of Pharmacy, Ain Shams University, for her support, kind supervision and encouragement throughout the work.
I am also deeply grateful to Prof. Dr. Mostafa Mohamed Soliman Ghorab, Prof. of Applied Organic Chemistry, National Center for Radiation Research and Technology(NCRRT), for suggesting the research subject, kind supervision, valuable advices and continuous support and guidance which helped me to complete this work.
I would like to express my deep thanks and appreciation to Dr. Helmy Ismail Heiba, Assist. Prof. of Applied Organic Chemistry(NCRRT), for his continuous support and supervision, valuable advices and fruitful opinions through the whole work.
I would like to give my special thanks to my whole family for the patience and moral support they provided throughout my research work.
Finally, my deep thanks to my colleagues in Drug Chemistry Lab (NCRRT), and to all who gave me help and support for the fulfillment of this work.
Contents
Abstract……………………………………...…………………... i - v 1. Introduction…………………………………………...……….. 1 1.1. Cancer ……………………………………………….…….…. 2 1.2. Rationale for combining chemotherapy and radiotherapy...... 6 1.3. Sulfonamides …..……………………………………………. 12 1.4. Thiazoles and fused thiazoles as anticancer agents …...….…. 24 1.5. Synthesis of thiazoles and fused thiazoles.…………..………. 31 2. Aim of the present investigation……………………...…...... 38 3. Theoretical Discussion……………………………...……...... 49 4. Experimental…………………………………………………... 68 5. Biological Activity………………………………...………….... 95 5.1. In-vitro anticancer activity………………………………….... 96 5.2. Radiosensitizing evaluation……………...…….…………...... 104 6. Molecular Modeling………………………………….....…..... 110 7. References…………………………………...……………...... 120
أ …………………..………………………………Arabic summary
List of Figures
Figure (1): Schematic representation of the catalytic mechanism for the α-
CA catalysed CO2 hydration………………………………………………14 Figure (2): Zinc binding groups…………………………………….…….15
Figure (3): The cell cycle…………………………………………………20
Figure (4): Tiazofurin and its analogues………………………………….25
Figure (5): Chemical structure of six hybrid compounds (MSt)……….…26
Figure (6): Polar interactions of the inhibitor 45 docked into MMP-9-A16 model……………………………………………………………………....27
Figure (7): Structural elements of CA inhibitors in the CA enzymatic active site………………………………………………………………….………40 Figure (8): Survival curve for HEPG2 cell line for compound III………..98
Figure (9): Survival curve for HEPG2 cell line for compound IVa………98
Figure (10): Survival curve for HEPG2 cell line for compound IVb…….98
Figure (11): Survival curve for HEPG2 cell line for compound VI…...…98
Figure (12): Survival curve for HEPG2 cell line for compound VII..……99
Figure (13): Survival curve for HEPG2 cell line for compound IXa….....99
Figure (14): Survival curve for HEPG2 cell line for compound IXb….....99
Figure (15): Survival curve for HEPG2 cell line for compound IXc…...... 99
Figure (16): Survival curve for HEPG2 cell line for compound IXd……..99
Figure (17): Survival curve for HEPG2 cell line for compound IXe……...99
Figure (18): Survival curve for HEPG2 cell line for compound IXf...... 100
Figure (19): Survival curve for HEPG2 cell line for compound IXg…....100
Figure (20): Survival curve for HEPG2 cell line for compound IXh…....100
Figure (21): Survival curve for HEPG2 cell line for compound XIa…....100
Figure (22): Survival curve for HEPG2 cell line for compound XIb……100
Figure (23): Survival curve for HEPG2 cell line for compound XIc….....100
Figure (24): Survival curve for HEPG2 cell line for compound XId……101
Figure (25): Survival curve for HEPG2 cell line for compound XII…....101
Figure (26): Survival curve for HEPG2 cell line for compound XIIIa.…101
Figure (27): Survival curve for HEPG2 cell line for compound XIIIb….101
Figure (28): Survival curve for HEPG2 cell line for compound XIVa.....101
Figure (29): Survival curve for HEPG2 cell line for compound XIVb….101
Figure (30): Survival curve for HEPG2 cell line for compound XV…….102
Figure (31): Survival curve for HEPG2 cell line for compound XVI…...102
Figure (32): Survival curve for HEPG2 cell line for compound XVII…..102
Figure (33): Survival curve for HEPG2 cell line for compound XVIII....102
Figure (34): Survival curve for HEPG2 cell line for compound XIX…...102
Figure (35): Survival curve for HEPG2 cell line for compound XX…….102
Figure (36): Survival curve for HEPG2 cell line for compound III alone and in combination with γ-irradiation (8 Gy)…………………………………106
Figure (37): Survival curve for HEPG2 cell line for compound IVb alone and in combination with γ-irradiation (8Gy)….………………………….106
Figure (38): Survival curve for HEPG2 cell line for compound IXb alone and in combination with γ-irradiation (8Gy)…………………………..…107
Figure (39): Survival curve for HEPG2 cell line for compound XIIIa alone and in combination with γ-irradiation (8 Gy)…………………………….107
Figure (40): Survival curve for HEPG2 cell line for compound XIVb alone and in combination with γ-irradiation (8 Gy)……………….…………....108
Figure (41): Survival curve for HEPG2 cell line for compound XVII alone and in combination with γ-irradiation (8 Gy)……………………………108
Figure (42): Survival curve for HEPG2 cell line for compound XX alone and in combination with γ-irradiation (8 Gy)……………………………109
Figure (43): Superimposition of hCA II-inhibitor adducts………………111 Figure (44): CA inhibition mechanism by sulfonamides………………...112 Figure(45): Interaction map of N-(2,3,4,5,6-pentafluorobenzyl)-4 sulfamoylbenzamide with hCA II………………………………………...114 Figure (46): Interaction map of E7070 with the active site of hCA II…...115
Figure (47): 2D and 3D Interaction map of compound IVb with hCA II .116
Figure (48): 2D and 3D Interaction map of compound XIVb with hCA II……………..……………………………………………………………116
Figure (49): 2D and 3D Interaction map of compound XVII with hCA II……………………………………………………………………..……117
Figure (50): 2D and 3D Interaction map of compound XX with hCA II ..117
Figure (51): Superimposition of E7070 and compound XIVb in the active site of hCA II……………………………………………………..……….118
Figure (52): Superimposition of E7070 and compound XIc in the active site of hCA II……………………………………………………….…….…...119
List of Tables
Table 1, Physico-chemical and analytical data for compounds IVa,b …....72
Table 2, Spectral data for compounds IVa,b………..………………..……73
Table 3, Physico-chemical and analytical data for compound VI……..…..75
Table 4, Spectral data for compound VI……………………………..…....75
Table 5, Physico-chemical and analytical data for compound VII………..76
Table 6, Spectral data for compound VII…………………….…………....76
Table 7, Physico-chemical and analytical data for compounds IX(a-h).….78
Table 8, Spectral data for compounds IX(a-h)……………………….……79
Table 9, Physico-chemical and analytical data for compounds XI(a-d)…..82
Table 10, Spectral data for compounds XI(a-d)…………………….….…83
Table 11, Physico-chemical and analytical data for compound XII...….…84
Table 12, Spectral data for compound XII……………..……………….....84
Table 13, Physico-chemical and analytical data for compound XIIIa,b….85
Table 14, Spectral data for compound XIIIa,b……………………….…...86
Table 15, Physico-chemical and analytical data for compound XIVa,b….87
Table 16, Spectral data for compound XIVa,b…………….……………..88
Table 17, Physico-chemical and analytical data for compound XV……...89
Table 18, Spectral data for compound XV……..…………………………89
Table 19, Physico-chemical and analytical data for compound XVI…...... 90
Table 20, Spectral data for compound XVI…………………………….…90
Table 21, Physico-chemical and analytical data for compound XVII…….91
Table 22, Spectral data for compound XVII…………………………….. 91
Table 23, Physico-chemical and analytical data for compound XVIII…... 92
Table 24, Spectral data for compound XVIII…………..………………... 92
Table 25, Physico-chemical and analytical data for compound XIX.….... 93
Table 26, Spectral data for compound XIX……….……………….…….. 93
Table 27, Physico-chemical and analytical data for compound XX…...….94
Table 28, Spectral data for compound XX………………...…………..….94
Table 29, In vitro anticancer screening of the synthesized compounds against (HEPG2)………………………………………………………….103
Table 30, In vitro anticancer screening of compounds against (HEPG2) in combination with radiation……………………………………………….105
Abbreviations
AMP: Ammonium salt of heteropoly acid. ANOVA: Analysis of variance. bFEF: basic Fibroblast growth factor. Boc: t-butoxy carbonyl. CA: Carbonic anhydrase. CAI: Carbonic anhydrase inhibitor. CAMS: Cell adhesion molecules. CARP: Carbonic anhydrase related protein. CDK: Cyclin dependent kinase. CNS: Central nervous system. Conc.: concentrated. CRT: Chemoradiotherapy. CT: Chemotherapy. Cys: Cysteine. dGTP: deoxy Guanosine triphosphate. Dil.: Diluted. DME: 1, 2-Dimethoxyethane. DMF: Dimethyl formamide. DMSO: Dimethyl sulfoxide. DNA: Deoxyribonucleic acid. E-cadherin: Epithelial cadherin. EGFR: Epidermal growth factor receptor. EI/MS: Electron impact mass spectrometry. ELISA: Enzyme-linked immunosorbent assay. EPG: Epithelial growth factor. 5-FU: 5-Fluorouracil. Gln: Glutamine. Glu: Glutamic acid. GSK-3: Glycogen synthase kinase-3. GTP: Guanosine triphosphate. Gy: Gray.
h: hour. hCA: human carbonic anhydrase. HEPG2: Human hepatocellular liver carcinoma cell line. His: Histidine. HMDO: Hexamethyldisiloxane. 1H-NMR: Proton nuclear magnetic resonance. IMP: Inosine monophosphate. IMPDH: Inosine monophosphate dehydrogenase. IR: Infrared. Leu: Leucine. min: minute. MMFF94x: Merck Molecular Force Field 94x. MMPIs: Matrix metalloproteinase inhibitors. MMPs: Matrix metalloproteinases. MOE: Molecular operating environment. NAD: nicotinamide adenine dinucleotide. NSCLC: Non-small cell lung cancer. OTT: Overall treatment time. PABA: p-amino benzoic acid. PI3: Phosphoionositide 3. pRb: Retinoblastoma protein. RNA: Ribonucleic acid. RT: Radiotherapy. SAR: Structure activity relationship. SE: Standard error. SRB: Sulfo-rhodamine B. TAD: Thiazole carboxamide adenosine dinucleotide. TFAA: Trifluoroacetic anhydride. THF: Tetrahydrofuran. Thr: Threonine. TIMPs: Tissue inhibitors metalloproteinases. TLC: Thin layer chromatography. UK: United Kingdom.
USA: United States of America. VCR: Vincristine. VEGFs: Vascular endothelial growth factors. XMP: Xanthosine monophosphate. ZBG: Zinc binding group.
Abstract| i
Abstract In a search for new cytotoxic agents with improved antitumor activity and selectivity, some new thiazole, thiazolopyrimidine, thiazolopyrane and thiazolopyranopyrimidine derivatives bearing sulfonamide moiety were synthesized. The newly synthesized compounds were evaluated for their antitumor activity alone and in combination with γ-irradiation. These new compounds were docked inside the active site of carbonic anhydrase II to predict their mechanism of action.
The thesis includes the following parts:
1. Introduction:
This part includes a brief literature review on cancer, chemotherapy and radiotherapy, rationale of combining chemotherapy and radiotherapy, the expected anticancer activity of new thiazole, thiazolopyrimidine and tricyclic pyrimidine derivatives, and different methods for the synthesis of these new compounds.
2. Aim of the present investigation:
This part includes the biological bases on which the synthesized compounds were designed. Schemes illustrate the synthetic pathways adopted in the preparation of the target compounds.
3. Theoretical Discussion:
This part deals with the discussion of experimental methods adopted for the synthesis of the target compounds.
Abstract| ii
4. Experimental:
This part includes the detailed practical methods for the synthesis of twenty seven new compounds and five known intermediates that are listed below with their elemental analyses and spectral data (IR, 1H-NMR and mass spectroscopy).
Known intermediates:
2-Chloro-N-(4-sulfamoylphenyl) acetamide (II) 4-(4-Oxo-4,5-dihydrothiazol-2-ylamino)benzenesulfonamide (III) 4-Isothiocyanato benzenesulfonamide (V) 4-(4-Methylbenzylideneamino)benzenesulfonamide ( VIIIa) 4-(4-Nitrobenzylideneamino) benzenesulfonamide (VIIIb).
New compounds:
4-(7-Phenyl-5-thioxo-4,5,6,7-tetrahydrothiazolo[4,5-d]pyrimidin-2- ylamino) benzenesulfonamide (IVa) 4-(5-Thioxo-7-p-tolyl-4,5,6,7-tetrahydrothiazolo[4,5-d]pyrimidin-2- ylamino) benzenesulfonamide (IVb) Ethyl 2-cyano-3-(4-sulfamoylphenylamino)-3-thioxopropanoate (VI) Ethyl 2-(4-oxo-4,5-dihydrothiazol-2-yl)-3-(4-sulfamoylphenyl amino)-3-thioxopropanoate (VII) 4-(4-Oxo-2-p-tolylthiazolidin-3-yl) benzenesulfonamide (IXa) Abstract| iii
4-(2-(4-Nitrophenyl)-4-oxothiazolidin-3-yl) benzenesulfonamide (IXb) 4-(4-Oxo-2-phenylthiazolidin-3-yl) benzenesulfonamide (IXc) 4-(2-(2-Hydroxyphenyl)-4-oxothiazolidin-3-yl) benzenesulfonamide (IXd) 4-(2-(4-Hydroxyphenyl)-4-oxothiazolidin-3-yl) benzenesulfonamide (IXe) 4-(2-(Benzo[d][1,3]dioxol-5-yl)-4-oxothiazolidin-3-yl) benzenesulfonamide (IXf) 4-(2-(2-Chlorophenyl)-4-oxothiazolidin-3-yl) benzenesulfonamide (IXg) 4-(2-(3-Bromophenyl)-4-oxothiazolidin-3-yl) benzenesulfonamide (IXh) 4-(5-Amino-6-cyano-7-(4-nitrophenyl)-7H-thiazolo[4,5-b]pyran-2- ylamino) benzenesulfonamide (XIa) 4-(5-Amino-6-cyano-7-(3,4-dimethoxyphenyl)-7H- thiazolo[4,5- b]pyran -2- ylamino) benzenesulfonamide (XIb) 4-(5-Amino-7-(2-chlorophenyl)-6-cyano-7H- thiazolo[4,5-b]pyran-2- ylamino) benzenesulfonamide(XIc) 4-(5-Amino-7-(3-bromophenyl)-6-cyano-7H- thiazolo[4,5-b]pyran-2- ylamino) benzenesulfonamide (XId) 8-Amino,9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano [2,3-d] pyrimidine (XII) 8-Oxo, 9-(4-nitro phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano [2,3-d] pyrimidine (XIIIa) Abstract| iv
8-Oxo,9-(3,4-dimethoxy phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b] pyrano [2,3-d] pyrimidine (XIIIb) 6-Methyl,8- oxo,9-(4-nitro phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b] pyrano [2,3-d] pyrimidine (XIVa) 6-Methyl,8- oxo,9-(3,4-dimethoxy phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b] pyrano [2,3-d] pyrimidine (XIVb) 8-Amino, 6-oxo ,9-(4-nitro phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano [2,3-d] pyrimidine (XV) 8-Amino, 6-thioxo ,9-(4-nitro phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b] pyrano [2,3-d] pyrimidine (XVI) 4-(6-Cyano-7-(4-nitrophenyl)-2-(4-sulfamoylphenylamino)-7H- thiazolo[4,5-b]pyran-5-ylamino)-4-oxobutanoic acid (XVII) 4-(6-Cyano-5-(2,5-dioxopyrrolidinyl)-7-(4-nitrophenyl)-7H- thiazolo[4,5-b]pyran-2-ylamino) benzenesulfonamide (XVIII) N-(6-cyano-7-(4-nitrophenyl)-2-(4-sulfamoylphenylamino)-7H- pyrano[2,3-d]thiazol-5-yl)-3-oxobutanamide (XIX) 8-Imino, 6-thioxo, 9-(3,4-dimethoxy phenyl) 9-H,1N phenyl,[2-(4- sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano [2,3-d] pyrimidine (XX)
5. Biological activity:
Twenty seven new compounds were screened for their in-vitro anticancer activity against human carcinoma liver cell line (HEPG2) while seven promising compounds were screened for their anticancer activity in combination with gamma irradiation to study the synergestic effect of combining chemotherapy and radiotherapy. Abstract| v
6. Molecular modeling:
All the newly synthesized compounds are docked in the active site of carbonic anhydrase II to predict the expected mechanism of action.
7. References:
This part includes 127 references.
8. Arabic summary
INTRODUCTION
Introduction | 2
1. Introduction
1.1. Cancer
Most human neoplasms arise from genetic mutations within a single affected cell and over subsequent divisions heterogeneity develops through the accumulation of further abnormalities. The genes most commonly affected can be characterized as those controlling cell cycle check points, DNA repair and DNA damage recognition, apoptosis, differentiation and growth signaling. Mutations are common in the genes controlling a series of intracellular proteins, such as the cyclins and the cyclin-dependent kinases that regulates proliferation. Proliferation may also be abnormal due to defects in the nuclear enzyme tolemerase. Tolemerase is an enzyme that prevents the normal shortening of DNA with each cell division that leads to senescence. Persistant tolemerase activity helps to maintain the neoplastic state in cancer cells. Epithelial growth factor (EPG) and its receptors are over expressed in many human epithelial tumors to maintain the growth of those tumors.1
1.1.1. Apoptosis and growth
Normal cells usually die by an active and tightly regulated process known as apoptosis or programmed cell death. Apoptosis can occur in response to a number of physiological or pathological stimuli (tumor necrosis factor and DNA- damaging cytotoxic drugs) and is mediated by a family of proteins known as caspases. 1
Introduction | 3
1.1.2. Angiogenesis
Angiogenesis, i.e. new vessel formation is stimulated by a variety of peptides produced by both tumor cells and host inflammatory cells, such as the vascular endothelial growth factors (VEGFs) and basic fibroblast growth factor (bFGF). Inhibition of angiogenesis is a potentially novel method of cancer therapy, as new vessel formation within and around tumors not only provides the cancer with nutrients and oxygen, but permits hematogenous spread, or metastasis. 1
1.1.3. Invasion and metastasis
Cancers spread by both invasion and metastasis in vessels of the blood or lymphatic system. Infiltration into surrounding tissues is associated with loss of cell-cell cohesion. Cohesion is mediated by active homotypic cell adhesion molecules (CAMS). The cadherin molecules are Epithelial cadherin (E-cadherin) is expressed by many carcinomas and loss of E- cadherin expression is associated with an increase in invasion of the tumor. Invasion is partly determined by the balance of activators to inhibitors of proteolysis. Secretion of proteolytic enzymes, including the matrix metalloproteinases, occurs from adjacent fibroblasts. The balance between the expression and activity of the matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) is important for tumor growth, invasion, metastasis and angiogenesis. 1
1.1.4. Aims of cancer treatment
Cancer treatment requires the cooperation of a multi-disciplinary team to coordinate the delivery of the appropriate treatment (surgery, chemotherapy Introduction | 4 and biological therapy), supportive and symptomatic care and physiological support.1
1.1.5. Principles of Chemotherapy
The therapeutic effect on the cancer is achieved by a variety of mechanisms which seek to differentiate between normal and transformed cells. Chemotherapy employs systematically administered drugs that directly damage cellular DNA (and RNA). It kills cells by promoting apoptosis. There is a narrow therapeutic window between effective treatment of the cancer and normal tissue toxicity, because the drugs are not cancer specific (unlike some of the biological agents), and the increased proliferation in cancers is not much greater than in normal tissues. Most of the drugs have been derived in the past by empirical testing of many different compounds, e.g. alkylating agents, the new molecular biology is leading top renewed attempts to target particular genetic defects in cancer, e.g. tyrosine kinase inhibitors. Most tumors rapidely develop resistance to single agents given on their own. For this reason the principle of combination chemotherapy was developed. Several drugs are combined together, chosen on the basis of differing mechanisms of action and non-overlapping toxicities. These drugs are given over a period of a few days followed by a rest of a few weeks, during which time the normal tissues have the opportunity for regrowth .1
1.1.6. Principles of radiation therapy
Radiation delivers energy to tissues, causing ionization and excitation of atoms and molecules. The biological effect is exerted through the generation of single – and double –strand DNA breaks, including apoptosis of cells as Introduction | 5 they progress through the cell cycle, and through the generation of short – lived free radicals, particularly from oxygen, which damage proteins and membranes. The most commonly used form of radiotherapy is external beam or teletherapy from a linear accelerator source which provides x-rays, the energy of which is transmitted as photons. Cobalt-60 generators can also provide gamma rays and high energy photons. Brachytherapy is the use of radiation sources in close contact with the tissue to provide intense exposure over a short distance to a restricted volume. Systemic radionuclides, e.g. iodine-131, or radioisotope–labelled monoclonal antibodies and hormones can be administered by intravenous routes to provide radiation targeted to particular tissue uptake via surface antigens or receptors.
The radiation dose is measured in grays (Gy), where 1 gray= 1 joule absorbed per kilogram of absorbing tissue and 1 centigray= 1 rad. The biological effect is dependent upon the dose rate, duration, volume irradiated and the tissue sensitivity. Fractionation is the delivery of the radiation dose in increments separated by at least 4-6 hours. Radiation dose is thus described by three factors:
-Total dose in cGy (centigray). -Number of fractions. -Time of completion. Most treatments are delivered in 150-200 cGy fractions daily for 5 days per week; although a regimen of two fractions daily (hyper fractionation) has improved survival benefit in a recent lung cancer trial .1 Introduction | 6
1.2. Rationale for combining chemotherapy (CT) and radiotherapy (RT)
The rationale for combining CT and RT is mainly based on two ideas, one being spatial cooperation, and the other is the enhancement of radiation effects.2-4 Spatial cooperation is effective if CT is sufficiently active to eradicate subclinical metastases and if the primary local tumor is effectively treated by RT. In this regard, no interaction between RT and CT is required. A major limitation is the relatively poor efficacy of anticancer drugs against common solid tumors in adults. It is often difficult to eradicate even small subclinical metastases by CT. Also, local failure rates of a primary tumor following RT are high for many tumor sites. To decrease the local failure rate, the enhancement of RT effects is necessary. In the presence of chemotherapeutic drugs, an increased response such as enhancement occurs within the irradiated volume. However, virtually all chemotherapeutic agents enhance radiation damage to normal tissues as well. Consequently, a therapeutic benefit is only achieved if enhancement of the tumor response is greater than that for normal tissues. Among the many chemotherapeutic agents used, cisplatin is one of the best agents for yielding a therapeutic benefit. An enhancing effect by the additional use of daily cisplatin before each RT fraction was observed in an in-vivo animal study.5
1.2.1- Mechanisms responsible for CT-RT interactions
Recent clinical trials have shown that CT given concurrently with RT results in improved local control and survival, implying interactions between Introduction | 7
CT and RT. 6-10 Five major mechanisms responsible for CT-RT interactions are discussed in the following paragraphs.2-4
1.2.1.1. Initial radiation damage
The first mechanism responsible for CT-RT interaction is the direct enhancement of the initial radiation damage, resulting from the incorporation of the chemotherapeutic drugs into DNA. The primary target for radiation injury is DNA, where halogenated pyrimidines such as 5- fluorouracil (5-FU) are incorporated, making the DNA more susceptible to RT. Cisplatin interacts with nucleophilic sites on DNA or RNA to form intra- and interstrand cross-links. When cisplatin-DNA cross-links are formed during RT, radioenhancement by cisplatin may occur. This has been observed in both hypoxic and oxygenated cells.4
1.2.1.2. Inhibition of radiation damage repair
Secondly, the inhibition of cellular repair increases radiation damage. Cells have the ability to repair sublethal and potentially lethal radiation damage.2 Halogenated pyrimidines, nucleoside analogs, and cisplatin interfere with cellular repair mechanisms. This inhibition of cellular repair can be effective when drugs are administered following fractionated RT. In general, nucleoside analogs such as fludarabine and gemcitabine are potent radiosensitizers. In animal experiments, the effect of fludarabine on radiocurability was greater when fludarabine was combined with fractionated RT than when it was combined with single-dose RT.11 This implies that the inhibition of sublethal or potentially lethal damage repair is Introduction | 8 a significant mechanism responsible for the enhancement of the tumor radioresponse to fludarabine.
1.2.1.3. Cell-cycle effects
The third mechanism focuses on a cell-cycle effect. The cytotoxicity of most chemotherapeutic agents and that of radiation is highly dependent on the phase of the cell cycle. Both chemotherapeutic agents and radiation are more effective against proliferating cells than against nonproliferating cells. Among proliferating cells, cells in the G2 and M phases are the most radiosensitive, and the cells in the S phase are the most radioresistant.2 Based on this variation in radiosensitivity over the cell cycle, there exist two strategies for CRT, the use of chemotherapeutic agents that accumulate cells in a radiosensitive phase or those that eliminate radioresistant S-phase cells. The latter strategy is related to the mode of action of nucleoside analogs. Fludarabine and gemcitabine are incorporated into radioresistant S phase cells, many of which die by apoptosis. This preferential removal of S phase cells therefore contributes to the radioenhancement effects.
1.2.1.4. Hypoxic cells
Hypoxic cells are 2.5–3.0 times less sensitive to radiation than well oxygenated cells.2,3 Tumors often include hypoxic areas, which is a cause of radioresistance. Chemotherapeutic agents can improve the RT effect by eliminating well-oxygenated tumor cells, which lead to tumor reoxygenation, selectively eliminating hypoxic cells, or sensitizing the hypoxic cells to radiation. Introduction | 9
1.2.1.5. Repopulation of tumor cells
The importance of the overall treatment time (OTT) for local tumor control by RT has been documented in a number of studies of head and neck cancers, uterine cervical cancer, and esophageal cancer.12-14 Analysis of esophageal and laryngeal squamous cell carcinomas treated by RT alone showed that the prolongation of OTT significantly reduced the local control rate.12,13 One mechanism responsible for this may be the accelerated repopulation of tumor cells during fractionated RT. Any approach that reduces or eliminates the accelerated repopulation of tumor cells improves the efficacy of RT. This is likely to be one of the major mechanisms by which CT improves local tumor control when given concurrently with RT. Even a small decrease in repopulation between radiation fractions can significantly improve the tumor response to fractionated RT. However, most chemotherapeutic drugs inhibit repopulation not only in the tumor, but also in the compensatory cell regeneration of normal tissues that occurs during fractionated RT. Thus, a therapeutic benefit is expected if drugs are tumor selective. Recently, various molecular targeting drugs have become clinically available. Several drugs, such as epidermal growth factor receptor (EGFR) inhibitors, block the membrane receptors of growth factors or interfere with the signaling pathways involved in cell proliferation. These agents offer another possible method for inhibiting the accelerated repopulation of tumor cells during fractionated RT.15,16
Introduction | 10
1.2.2. Sequencing of CT and RT
According to the sequencing of CT and RT, CT is designated as induction (neoadjuvant) CT, concurrent CT, or adjuvant CT, when it is given before, during, or after the course of RT, respectively. As clinical trials of adjuvant CT following RT have not been systematically studied, the aims and clinical results of induction CT and concurrent CT are described in the following paragraphs;
1.2.2.1. Induction chemotherapy
Induction CT has two main objectives,2,3 one being the eradication of micrometastases while they contain small number of tumor cells, and the other being to reduce the size of the primary tumor that is to be irradiated. Reducing the number of cells in the tumor increases the probability of tumor control by RT. In addition, CT induced tumor shrinkage may provide a smaller target volume for RT, thereby limiting normal tissue damage.
1.2.2.2. Concurrent chemotherapy
For concurrent CRT, CT can act on both systemic and primary lesions. However, the main objective of concurrent CT is to use CT-RT interactions to maximize the antitumor effect, even though it may increase the acute toxicity of the treatment.2,4 Therefore, it should be remembered that the therapeutic benefit of concurrent CRT only occurs when enhancement of the tumor response is greater than the toxic effects on critical normal tissues. Introduction | 11
An alternating schedule of CT and RT can be used without a treatment gap, to minimize excessive toxic effects on normal tissues and to enhance the tumor response by perturbing cell cycling or reoxygenation. Several clinical trials involving alternating schedules have yielded promising results.9,17
Introduction | 12
1.3. Sulfonamides
1.3.1. Sulfonamides as anticancer agents
Sulfonamides constitute an important class of drugs, with several types of pharmacological activities including antibacterial,18 anti-carbonic anhydrase,19 diuretic,20 hypoglycemic21 and antithyroid activity.22 Also, some structurally novel sulfonamide derivatives have recently been reported to show substantial antitumor activity in-vitro and/or in-vivo. (E7010) 1, (ER-34410) 2 and (E7070, Indisulam) 3 and QBS are examples for antitumor sulfonamides in advanced clinical trials.23
There are a variety of mechanisms describing the antitumor action of sulfonamides, such as carbonic anhydrase (CA) inhibition, cell cycle arrest in the G1 phase, disruption of microtubules, angiogenesis and matrix metalloproteinase (MMP) inhibition. The most prominent mechanism was the inhibition of carbonic anhydrase isozymes (CAs).23
Introduction | 13
1.3.2. Sulfonamides as carbonic anhydrase inhibitors
1.3.2.1. Carbonic anhydrases
The carbonic anhydrases are metalloenzymes containing zinc ion in their active site. Carbonic anyhdrases are present in prokaryotes and eukaryotes, and are encoded by four distinct gene families: the α-CAs, β-CAs, γ-CAs and δ-CAs. In mammals the α-CA is present and 16 different α-CA isozymes or CA-related proteins (CARP) were described with different subcellular localization and tissue distribution.24,25 Basically there are several cytosolic forms (CA I-III, CA VII and CA XIII), five membrane-bound isozymes (CA IV, CA IX, CA XII, CA XIV and CA XV), two mitochondrial isoforms (CA VA and VB) and CA VI is secreted in the saliva and milk. Three cytosolic acatalytic forms are also known (CARP VIII, CARP X and CARP XI).26
All these CAs are able to catalyze the hydration of CO2 to bicarbonate at physiological pH (Figure 1).19, 26, 27 This chemical interconversion is crucial since bicarbonate is the substrate for several carboxylation steps in a number of fundamental metabolic pathways such as gluconeogenesis, biosynthesis of several amino acids, lipogenesis, ureagenesis and pyrimidine synthesis.28 Apart from these biosynthetic reactions, some of the CAs are involved in many physiological processes related to respiration and transport of CO2/ bicarbonate between metabolizing tissues and the lungs, pH homeostasis and electrolyte secretion in a variety of tissues/ organs.19,27
Introduction | 14
Figure (1): Schematic representation of the catalytic mechanism for the α-
CA catalysed CO2 hydration. The hydrophobic pocket for the binding of substrate(s) is shown schematically at step (B)
1.3.2.2. Role of carbonic anhydrases in cancer
The importance of this family of enzymes for the uptake of bicarbonate by many organisms, and the presence of a large number of isoforms (which can be distinguished from each other in activity and are located in different areas inside the cell) make the CAs undoubtedly involved in cell growth. Furthermore, CA isozymes (CAII, CAV, CA IX, CA XII and CA XIV) have close connections with tumors.28,29 The role of CAs in cancer can be Introduction | 15 explained in light of the metabolic processes required by growing cancer cells that develop with a higher rate of replication than normal cells. Such a circumstance requires a high flux of bicarbonate into the cell in order to provide substrate for the synthesis of either nutritionally essential components (nucleotides) or cell structural components (membrane lipids).28
1.3.2.3. Carbonic anhydrases inhibition
CA inhibitors are mainly used as antiglaucoma agents, 20, 27, 30 antithyroid drugs,30 hypoglycemic agents21 and, ultimately, some novel types of anticancer agents.31 Sulfonamides are known to possess high affinity for CAs as such compounds possess a zinc binding group (ZBG) by which they interact with the metal ion in the active site of the enzyme and the residues Thr 199 and Glu 106 in its neighborhood.32 In recent years, an entire range of new ZBGs were reported as shown in Figure 2. These new ZBGs include, in addition to the classical sulfonamides A, sulfamates B, sulfamides C, substituted sulfonamide D, Schiff’s base E, urea F and hydroxyurea derivatives G, as well as hydroxamates H.25, 32
Figure (2): Zinc binding groups: sulfonamides A, sulfamates B, sulfamides C, substituted sulfonamides D, Schiff’s base E, urea F, hydroxyurea G and hydroxamates H. Introduction | 16
1.3.2.4. Sulfonamides as anticancer agents through CA inhibition
Sulfonamides CA inhibitors reduce the provision of bicarbonate for the synthesis of nucleotides (mediated by carbamoyl phosphate synthetase II) and other cell components such as membrane lipids (mediated by pyruvate carboxylase). Such mechanism would likely involve CA II and CA V.33 An alternative, or additional mechanism, may involve the acidification of intracellular milieu as a consequence of CA inhibition by these potent CA inhibitors.34 It is also possible that the sulfonamides interfere with the activity of the CA isozymes known to be present predominantly in tumor cells, CA II, IX, XII and XIV.19, 27, 29 A combination of these mechanisms proposed above is also possible.
1.3.2.5. Examples of sulfonamides acting as anticancer agents by CA inhibition
Several potent, clinically used sulfonamide CA inhibitors such as acetazolamide 4, methazolamide 5, or ethoxzolamide 6 have shown to inhibit the growth of human lymphoma cells.33
A program of screening several hundred sulfonamide CA inhibitors (both aromatic as well as heterocyclic derivatives) for their tumor cell growth inhibitory effects against a panel of 60 cancer cell lines of the National Cancer Institute of the USA, has identified derivatives 7-11 as interesting Introduction | 17 leads. These derivatives showed potent activities, against a wide variety of cancer cell lines including leukemia, non-small cell lung cancer, ovarian, melanoma, colon, CNS, renal, prostate and breast cancer cell lines.35,36
1.3.2.6. Examples of Sulfonamides acting as selective CA inhibitiors
Several miscellaneous sulfonamides have been synthesized and screened by Vullo et al.37 in an attempt to obtain selective CA inhibitors especially towards CA II CA IX and CA XII. From these compounds, compounds 3, 12-14 were found to behave as very potent CA II and IX inhibitors. Introduction | 18
Casey et al.38 prepared a series of positively charged, membrane impermeant sulfonamide CA inhibitors 15-20 and showed high affinity for the cytosolic isozymes CA I and CA II, as well as for the membrane-bound ones CA IV and CA IX. In vitro studies showed that this new class of inhibitors is able to discriminate between the membrane-bound versus the cytosolic isozymes. These compounds were proved to be unable to penetrate through the membranes, obviously due to their cationic nature. These compounds constitute the basis of selectively inhibiting only the target, tumor-associated CA IX in-vivo, whereas the cytosolic isozymes would remain unaffected.38
Introduction | 19
CA XII is present in a wide range of tumors and it appears probable that inhibition of this isozyme with potent and possibly specific inhibitors may have clinical relevance for the development of novel antitumor therapies. The most selective hCA XII over hCA II inhibitors were compounds 12, 21 and 22.
A series of indanesulfonamide derivatives 23 and 24 were synthesized by Thiry et al.39 and tested for their inhibitory activity against CA IX, and against CA I and II and two other physiologically relevant CA isozymes, to measure the selectivity of the compounds. Nearly all the derivatives show a high inhibitory potency against CA IX and CA II and a weak inhibition against CA I.
1.3.3. Sulfonamides targeting G1 phase of the cell cycle
G1 phase of the cell cycle is an important period where various complex signals interact to decide a cell’s fate: proliferation, quiescence, differentiation, or apoptosis (Figure 3).40 Introduction | 20
Figure (3): The cell cycle
It is now well-recognized that malfunctioning of cell cycle control in G1 phase is among the most critical molecular bases for tumorigenesis and tumor progression. Thus, there is a growing possibility that a small molecule targeting the control machinery in the G1 phase can be a new type of drug efficacious against refractory clinical cancers.41
(E7070) 3 was found to block the entry of human NSCLC A549 cells into the S phase, leading to the accumulation of cells in the late G1 phase. In addition, treatment of A549 cells with (E7070) 3 resulted in the inhibition of pRb phosphorylation, a crucial step in the G1/S transition. Down regulation of CDK2 and cyclin A expressions as well as suppression of CDK2 catalytic activity with the induction of p53 and p21, may account for the inhibition of pRb phosphorylation by E7070.42
In addition, (E7070) 3 was shown to inhibit the phosphorylation of CDK2 itself, leading to the inhibiton of CDK2 catalytic activity. These Introduction | 21 results suggest that E7070 may target the late G1 phase of the cell cycle and restore the pRb dependent growth-inhibitory pathway disrupted in human NSCLC cells. This is accomplished by inhibiting CDK2 catalytic activity.42
1.3.4. Sulfonamides causing disruption of microtubules
The effects of (E7010) 1 on microtubule structure in tumor cells were shown to cause the disappearance of cytoplasmic microtubules and mitotic spindles. The experiments clearly demonstrated that the growth-inhibitory activity of (E7010) 1 is caused by the inhibition of microtubule assembly.43 A novel series of 7-aroylaminoindoline-1-benzenesulfonamides have been identified by Chang et al.44 as a novel class of highly potent antitubulin agents. The lead compounds 25 and 26 exhibit antiproliferative activity. The SAR information of the 7-aminoindoline-substitution pattern revealed that the 7-amide bond formation in the indoline-1-sulfonamides contributed to a significant extent for maximal activity rather than the carbamate, carbonate, urea, alkyl linkers.
Hu et al.45 have synthesized two series of carbazole sulfonamide compounds. Compounds such as 27 and 28 exhibited strong activities against human leukemia cells. Preliminary studies demonstrated that the lead compound 27 arrests tumor cell cycle at M-phase and induces apoptotic cell Introduction | 22 death by increasing expression of p53 and promoting bcl-2 phosphorylation. Lead compounds such as 27 and 28 merit further studies as novel promising antimitotic agents against solid tumors.
1.3.5. Sulfonamides as matrix metalloproteinase (MMP) inhibitors
The matrix metalloproteinases (MMPs), a family of zinc-containing endopeptidases, were shown to play a central role in several physiological and physiopathological processes and in our main interest regards the involvement of these enzymes in angiogenesis and tumor invasion.46,47 They consist of a Zn(II) ion coordinated by three histidines, with the fourth ligand being a water molecule/hydroxide ion which is the nucleophile intervening in the catalytic cycle of the enzyme.48 Inhibition of MMPs is correlated with the coordination of the inhibitor molecule (in neutral or ionized state) to the catalytic metal ion, with or without replacement of the metal-bound water molecule.27, 47, 49 Thus, MMP inhibitors (MMPIs) must contain a zinc-binding function attached to a scaffold that will interact with other binding regions of the enzymes.48
Sulfonylated amino acid hydroxamates were only recently discovered to act as efficient MMPIs.50,51 The first compounds from this class to be developed for clinical trials are (CGS 27023A) 29 and (CGS 25966) 30.50 Introduction | 23
Further developments in this field have led to some strong and relatively selective inhibitors of this type such as 31.51 Then a large number of arylsulfonyl hydroxamates derived from glycine, L-alanine, L-valine and L- leucine possessing N-benzyl- or N-benzyl-substituted moieties, of types 32, were reported.52 Also, some hydroxamates structurally related to the MMPIs were shown to act as CAIs 33.48
Introduction | 24
1.4. Thiazoles and fused thiazoles as anticancer agents
Thiazoles and fused thiazole derivatives are known to possess several biological activities53 including anticancer activity. 54-57 There are a variety of mechanisms for the antitumor action of thiazole and fused thiazole derivatives, they act on cancer biotargets, such as tumor necrosis factor TNF- α,58 inosine monophosphate dehydrogenase (IMPDH),59 and apoptosis inducers.60
Tiazofurin 34, 61 the synthetic nucleoside analogue, is a potent inhibitor of inosine 5’ monophosphate dehydrogenase (IMPDH), a rate-limiting enzyme of de novo guanine nucleotide synthesis.59 which catalyzes the NAD-dependent conversion of inosine 5’-monophosphate (IMP) to xanthosine 5’-monophosphate (XMP), was shown to be significantly increased in highly proliferative cells. Inhibition of this enzyme results in a decrease in GTP and dGTP biosynthesis, producing inhibition of tumor cell proliferation.62 Tiazofurin is a high-priority candidate for clinical trials with potential importance for treatment of lung tumours and metastases.63,64 Tiazofurin has shown a significant reduction in leukaemic cell burden in acute myelogenous leukaemia patients.65 Despite the efficacy achieved in the clinical trials of tiazofurin, lack of specificity and occasional neuro- and cardiovascular toxicity remain a problem in its clinical use.64,66 To improve its biological properties, many analogues 35-37 have been prepared, including a number of those with variations in the furanose ring (Figure 4).67-69 Introduction | 25
Figure (4): Tiazofurin and its analogues
Epothilones 38, The recent class of natural products used for tubulin binding,70 and stabilization of microtubule dynamics.71 Epothilones 38, are apoptosis inducers, most apoptotic agents are known to induce apoptosis through one of two pathways, namely a receptor-mediated and a non receptor-mediated or chemical-induced pathway,72,73 epothilones 38, have much greater activity against multi-drug resistant cell lines.74
Thiazole-containing side chain of epothilones 39, was investigated. Among a tested series of hybrid compounds the one containing thiazole side chain at C15 (MSt-2) showed the maximum potency to induce apoptosis, while another containing thiazole side chain at C3 (MSt-6) was less potent.75 Introduction | 26
It was reported that the 16-membered trilactone core structure of macrosphelides and the thiazole-containing side chain of epothilones are promising apoptosis inducers (Figure 5). 76
39
Figure(5): Chemical structure of six hybrid compounds (MSt) 39 composed of 16 membered trilactone core structure of macrosphelides and the thiazole- containing side chain of epothilones.
Benzenesulfonamides and thiazolyl benzenesulfonamides constitute an important class of MMPs inhibitors.77,78 MMPs are associated with cancer- cell invasion and metastasis.79,80 They are involved in the massive up regulation in malignant tissues and characterized by their unique ability to degrade all components of the extracellular matrix.81 Degradation of extracellular matrix is crucial for malignant tumour growth, invasion, Introduction | 27 metastatis and angiogenesis.82,83 Kiyama et al.84 described a thiazolyl benzenesulfonamide as MMPI 40a (Figure 6).
Figure (6): Polar interactions of the inhibitor 40a docked into MMP-9-A16 model.
Recently, 84 it has been reported that several C-nucleosides, sulfonamide- schieff’s bases and sulfonamide- thiazolidinones 40b possess considerable cytotoxic effect against breast carcinoma cell line MCF7 and cervix carcinoma cell line HELA.
Indolequinone substituted by a thiazole ring was shown to be inhibitors of topoisomerase II with marked cytotoxic activities towards human tumour cell lines. The indolequinone antibiotic (BE 10988) 41 does not covalently bind to DNA but triggers double stranded cleavage of DNA Via the inhibition of the enzyme topoisomerase II.85 This DNA relaxing enzyme is Introduction | 28 the primary target of several clinically used anticancer drugs including etoposide and mitoxantrone. 86
Ruthenium compounds form a class of metallopharmaceuticals, of potential wide application in various disease areas.87 Michael Clarke described the possible medical uses of ruthenium compounds. 88 A number of studies have exploited the use of ruthenium compounds as experimental anticancer agents. 89 An example on these is the novel compound, trans–cis– cis-[Ru(II) Cl2(DMSO)2(2-amino-5-methyl-thiazole)2] 42.
Also, thiazole -4- carboxamide adenine dinucleotide (TAD) analogue 43 displaces NAD binding at the cofactor site. Therefore, TAD analogues are expected to provide probes of the stereochemical requirements of the adenine end of the cofactor site of IMPDH.90 Introduction | 29
The most promising new approaches in cancer treatment is blocking angiogenesis.91 Tie-2, an endothelium specific receptor tyrosine kinase, promotes tumour angiogenesis through interaction with angiopoietin, 92 examples of this group are the bicyclic pyrimidines in which, the 93 thiazolopyrimidine are the most potent.
Also, thiazolo[4,5-d]pyrimidines 44a,b are related to 2,4-diamino- thiazoles that have been previously reported to be potent inhibitors of both CDK 94,95and GSK-3 (glycogen synthase kinase-3), which belongs to the same family of serine-threonine protein kinases as CDKs.96
Introduction | 30
On the other hand, it has been reported that several pyrano[2,3- d]thiazolo[2,3-b]pyrimidines 45c were prepared for anticancer evaluation starting with 2H-5-amino-6-cyano-3,7-dioxothiazole[2,3-b]pyrimidine 45a and its phenyl methylene derivative 45b. 96
Introduction | 31
1.5. Synthesis of thiazoles and fused thiazoles
1.5.1. From Aldehydes:
Substituted aldehyde 46 reacted directly with thiosemicarbazide, and the obtained thiosemicarbazones 47 subsequently reacted with suitable 2- halogenoketone to yield the 2,4- disubstituted thiazole derivative 48.97
1.5.2. From thioimides:
Dimethyl cyanodithioimidocarbonate 49 obtained from cyanamide in presence of KOH, CS2 and MeI, was engaged in a one-pot three step procedure and was reacted successively with sodium sulfide, chloroacetonitrile, and potassium carbonate to obtain the substituted thiazole derivative 50.98
Introduction | 32
1.5.3. From L-proline:
N-Boc protected L-proline thiazole amino acid ester 52 was first synthesized starting from commercially available Boc-L-proline 51 via a modified Hantzsch reaction99 with ethyl bromopyruvate in presence of ◦ KHCO3 and 1,2-dimethoxyethane (DME) at -12 C, followed by trifluoroacetic anhydride (TFAA), the next step is N-Boc deprotection of the resulting carboxylic ester by HCl in 1, 4-dioxane, to yield the corresponding thiazole derivative 53.99
1.5.4. From methyl benzoate:
The first step consisted in converting the methyl benzoate 54 into a thionoester 55 using a mixture of P4S10/ hexamethyldisiloxane (HMDO) under microwave irradiation at 150 °C, which provided the desired thionoester in approximately 1 h. The purified thionoester is then subjected to phenylglycine to afford the desired 2,4- disubstituted-5-acetoxythiazoles 56.100
Introduction | 33
1.5.5. From phenacyl bromide:
A mixture of a phenacyl bromide 57, thioacetamide/ thiourea in MeOH and the catalyst AMP (ammonium salt of a heteropoly acid). The mixture was stirred at room temperature to obtain the thiazole derivative 58.101
1.5.6. From α-halo-β-ketoesters:
Thiazole 5-carboxylic acid derivative 61 was obtained in good yields by the condensation of α-chloro-β-ketoester 59 with 2-(2,6-dichlorophenyl) ethanethioamide 60 in EtOH and pyridine, followed by a basic hydrolysis of the resulting esters in NaOH, MeOH/ THF.102
1.5.7. From bis (aroylmethyl) sulfides:
2-Aroyl methyl-2,4-diaryl-5-benzylidine thiazole 63 was obtained from the reaction of aromatic aldehydes and ammonium acetate with bis (aroylmethyl) sulfides 62 which were thoroughly mixed in a borosilicate boiling tube kept partially immersed in a silica bath, which, in turn, was placed in a microwave oven.103 Introduction | 34
1.5.8. From isothiocyanate:
An equimolar mixture of isothiocyanate 64, malononitrile and sulfur powder in DMF was stirred in ice bath. After 15 min, triethylamine was added drop-wise to the mixture, the reaction was continued for 4 h to give 4- amino-3-substituted-2-thioxo-2,3-dihydro-thiazole-5-carbonitrile derivative 65 .104
1.5.9. From 2- aminothiazole:
Thiazolidinone derivative 68 was produced from stirring of 2- aminothiazole 66 with chloroacetylchloride in DMF at room temperature for 2h, then the produced compound 67 was refluxed with ammonium thiocyanate in ethanol for 1 hr.105
Introduction | 35
1.5.10. From sulfanilamide:
A suitable aldehyde was refluxed with aminocompound in absolute ethanol for 6 h until the formation of Schiff’s base 69, then the separated Schiff base was refluxed with mercaptoacetic acid for additional 12 h in dry benzene to obtain the corresponding thiazolidinone 70 through a two step reaction.106
A one pot reaction can be conducted by refluxing a suitable aldehyde with sulfanilamide in dry benzene in a flask connected to Dean-Stark tap for 12 h to form the azomethine, then a solution of α- mercaptoacetic acid in benzene was added to the reaction mixture and refluxed for additional 12 h to obtain the corresponding thiazolidinone 70.106
Introduction | 36
1.5.11. From Schiff’s base of cyanoacetic acid hydrazide:
5-Amino-N’-[1-chloro-3,4-dihydronaphthalen-2-yl)methylene]-3-phenyl- 2-thioxo-2,3-dihydrothiazole-4-carbohydrazide 72 was prepared by the reaction of Schiff’s base of cyanoacetic acid hydrazide 71 with sulfur and phenyl isothiocyanate in the presence of triethylamine as a basic catalyst. The thiazolo[5,4-d]pyrimidinone derivative 73 was prepared by heating 72 with a mixture of triethylorthoformate and acetic anhydride.107
1.5.12. From 4-aminothiazole-5-carbonitrile:
4-Amino-3-alkyl-2-thioxo-2,3-dihydrothiazole-5-carbonitrile 74 was treated with formamide and heated at 110 °C for 6 h to yield the 7-amino-3- ethyl thiazolo[4,5-d]pyrimidine-2(3H)-thione 75.108 Introduction | 37
1.5.13. From 1,2,3,4 -tetrahydropyrimidine -2- thione derivative:
1,2,3,4-Tetrahydropyrimidine- 2-thione 76 was heated under reflux with chloroacetic acid and benzaldehyde in the presence of acetic anhydride and sodium acetate to yield the corresponding thiazolo[3,2 - a]pyrimidine derivative 77. 109
1.5.14. From arylidenemalononitrile:
To a solution of arylidenemalononitrile 78 obtained from reacting malononitrile with aldehyde, the active methylene thiazolin-2,4-dione 79 was added in methanol with drops of morpholine, and heated until boiling to obtain the corresponding thiazolopyrane derivative 80. 110
AIM OF THE PRESENT INVESTIGATION
Aim of the present investigation| 39
2. Aim of the present investigation
A series of structurally novel sulfonamide derivatives have recently been reported to show substantial antitumor activity in-vitro and/or in-vivo.37 In order to explain this antitumor activity, several mechanisms were adopted including carbonic anhydrase inhibition, cell cycle arrest at the G1 phase, disruption of microtubules, and angiogenesis inhibition. The most prominent among these mechanisms was carbonic anhydrase inhibition.23
On the other hand, thiazoles, and thiazolopyrimidines are known to possess several biological activities including anticancer activity. 53-57,92-96
Based on the above information, the present investigation deals with the design and synthesis of some novel thiazole and thiazolopyrimidine derivatives. Moreover, our goal is to introduce new heterocyclic entities in the field of anticancer agents, such as thiazolopyrane and thiazolopyranopyrimidine derivatives having a biologically active sulfonamide moiety in order to explore the effect of combining different heterocyclic moieties with the biologically active sulfonamide group on the antitumor activity and to study their SAR. In addition, the synergism with gamma radiation was studied. As the carbonic anhydrase inhibition is the most prominent mechanism of the antitumor activity of sulfonamide derivatives, the synthesized compounds were designed to comply with the pharmacophore of compounds that may act as CA inhibitors. The most general structure features of a CA inhibitor complexed to the enzyme active site include: A zinc binding group (ZBG), which corresponds to SO2NH2 group. The sulfonamide is attached to a scaffold, which is usually a benzene Aim of the present investigation| 40 ring, a tail attached to the scaffold (such as thiazole, thiazolopyrimidine, thiazolopyrane and thiazolopyrano-pyrimidine) and this corresponds to the side chain that possess a hydrophilic link able to interact with the hydrophilic part of the active site and a hydrophobic moiety which can interact with the hydrophobic part of the CA active site (Figure 7).39 This pharmacophore originated from the analysis of the CAs active site and from the structure of inhibitors described in literature.111
Figure (7): Structural elements of CA inhibitors in the CA enzymatic active site.
The target compounds include the following classes: Aim of the present investigation| 41
1. Sulfonamide derivatives bearing a thiazole moiety III,VII:
2. Sulfonamide derivatives bearing a thiazolopyrimidine moiety IVa, b:
No. R IVa H IVb CH3
3. Sulfonamide derivatives bearing a thiazole moiety with various substitutions on the attached phenyl ring IX(a-h): Aim of the present investigation| 42
No. R1 R2 R3
IXa H H CH3
IXb H H NO2 IXc H H H IXd OH H H IXe H H OH IXf H
IXg Cl H H IXh H Br H
4. Sulfonamide derivatives bearing a thiazolopyrane moiety XIa-d, XVII- XIX:
No. R1 R2 R3 R4
XIa H H NO2 NH2
XIb H OCH3 OCH3 NH2
XIc Cl H H NH2
XId H Br H NH2
XVII H H NO2
XVIII H H NO2
XIX H H NO2
5. Sulfonamide derivatives bearing a thiazolopyranopyrimidine moiety XII- XVI, XIIIb, XIVb: Aim of the present investigation| 43
No. X XII H XV O XVI S
No. R1 R2 R3
XIIIa NO2 H H
XIIIb OCH3 OCH3 H
XIVa NO2 H CH3
XIVb OCH3 OCH3 CH3
Aim of the present investigation| 44
On the other hand, the rationale for combining chemotherapy and radiotherapy is based mainly on two ideas, one being spatial cooperation, which is effective if chemotherapy is sufficiently active to eradicate subclinical metastases and if the primary local tumor is effectively treated by radiotherapy. In this regard, no interaction between radiotherapy and chemotherapy is required. The other idea is the enhancement of radiation effects by direct enhancement of the initial radiation damage by incorporating drugs into DNA, inhibiting cellular repair, accumulating cells in a radiosensitive phase or eliminating radioresistant phase cells, eliminating hypoxic cells, or inhibiting the accelerated repopulation of tumor cells. Virtually, all chemotherapeutic agents have the ability to sensitize cancer cells to the lethal effects of ionizing radiation.112 Therefore, the most active synthesized compounds; will be selected to be evaluated for their ability to enhance the cell killing effect of γ-irradiation. Aim of the present investigation| 45
Accordingly, the designed compounds were synthesized adopting the chemical pathways outlined in schemes 1-5 inorder to evaluate their anticancer activity. Scheme 1
Scheme1: Synthesis of the target compounds IVa,b
Aim of the present investigation| 46
Scheme 2
Scheme 2: Synthesis of the target compounds VI, VII.
Scheme 3
Scheme 3: Synthesis of the target compounds VIIIa,b, IX a-h. Aim of the present investigation| 47
Scheme 4
Scheme 4: Synthesis of the target compounds XI a-d.
Aim of the present investigation| 48
Scheme 5
Scheme 5: Synthesis of the target compounds XII-XX.
THEORETICAL DISCUSSION
Theoretical discussion| 50
3. Theoretical discussion
2-Chloro-N-(4-sulfamoylphenyl) acetamide (II)113
Compound II was reported to be obtained by slow addition of chloroacetyl chloride to a stirred solution of sulfanilamide I in N,N- dimethylformamide for 2 h, at room temperature. The reaction proceeded via a simple nucleophilic substitution reaction with elimination of one molecule of HCl.105,113
The formation of compound II was supported by its micro-analytical and spectral data. The IR spectrum showed the presence of (C=O) band at 1705 cm-1, and (C-Cl) band at 790 cm-1.
While 1H-NMR spectrum revealed a singlet at 4.0 corresponding to the
NH group, singlet at 4.2 ppm corresponding to CH2, two doublets for the
AB-system and a singlet for SO2NH2 at the range of 7.2-7.9 ppm.
4-(4-Oxo-4,5-dihydrothiazol-2-ylamino) benzenesulfonamide (III)114
Treatment of compound II with ammonium thiocyanate in ethanol under reflux for 1 h, yielded the corresponding thiazolidinone derivative III, via intramolecular cyclization.105 Theoretical discussion| 51
The formation of compound III was supported by its micro-analytical and spectral data. IR spectrum of compound III showed the absence of (C-Cl) band of compound II, and the appearance of a two bands at 1710 cm-1 and -1 1 1602 cm attributed to the presence of (C=O) and (C=N). Additionally, H-
NMR spectrum showed CH2 protons of the thiazole ring as a singlet at 4.1 ppm, and multiplet for aromatic protons and SO2NH2 in the range of 7.0-8.0 ppm. Finally, mass spectrum of compound III exhibited molecular ion peak at m/z 271 [M+, 100%]. Also, Compound III displays amino-imino tautomerism as shown.
Theoretical discussion| 52
4-(7-Phenyl-5-thioxo-4,5,6,7-tetrahydrothiazolo[4,5-d]pyrimidin-2- ylamino) benzenesulfonamide (IVa) and 4-(5-thioxo-7-p-tolyl-4,5,6,7- tetrahydrothiazolo[4,5-d]pyrimidin-2-ylamino) benzenesulfonamide (IVb)
Compounds IVa,b were obtained via refluxing of compound III with thiourea and substituted benzaldehyde in ethanol containing few drops of conc. HCl for 3 h, through bignelli’s cyclocondensation reaction.115 This reaction proceeded via condensation followed by elimination of two moles of water to give the thiazolo[4,5-d]pyrimidine IVa,b.
IR spectra of compounds IVa,b revealed the absence of the band corresponding to the carbonyl group of compound III and the presence of Theoretical discussion| 53 two bands at 1650,1630 cm-1 and 1270,1290 cm-1 corresponding to (C=N) and (C=S), respectively. Additionally, 1H-NMR spectra of compounds IVa,b showed the absence of a singlet corresponding to the CH2 group of the thiazole ring. 1H-NMR spectrum of compound IVa showed the appearance of a singlet at 4.2 ppm corresponding to the CH of the pyrimidine, two singlets at 11.9 and 12.3 ppm attributed to the two NH groups of the pyrimidine ring which are exchangeable with D2O, and Mass spectrum of compound IVa exhibited a molecular ion peak at m/z 417 [M+, 18.5%], with a base peak at m/z 134. 1H-NMR spectrum of compound IVb showed the presence of two singlets at 2.3 ppm and 5.0 ppm corresponding to the CH3 group and the CH of the pyrimidine, two singlets at 11.9 and 12.5 ppm attributed to the two
NH groups of the pyrimidine ring which are exchangeable with D2O.
4-Isothiocyanato benzenesulfonamide (V)116
Compound V was prepared according to the published procedure.116
Conversion of the amine into isothiocyanate was processed via reaction with thiocarbonylating agent thiophosgene, and elimination of 2 moles of HCl.
Theoretical discussion| 54
Ethyl 2-cyano-3-(4-sulfamoylphenylamino)-3-thioxopropanoate (VI)
A solution of compound V in tetrahydrofuran (THF), was added to ethyl cyanoacetate sodium salt, to obtain the corresponding thioxopropanoic acid ethyl ester VI. The suggested reaction mechanism for the formation of compound VI involves a nucleophilic addition of CH2 group of ethyl cyanoacetate (an active methylene) on the isothiocyanate group.
IR spectrum of compound VI revealed the absence of (NCS) band at 2106 cm-1 and the presence of two bands at 2274 cm-1 and 1689 cm-1 corresponding to (C≡N) and (C=O), respectively. Additionally, 1H-NMR spectrum of compound VI showed the presence of a triplet at 1.2 ppm corresponding to the CH3, singlet at 2.1 ppm corresponding to CH, quartet at 4.1 ppm corresponding to CH2, and a singlet at 12.4 corresponding to NH which is exchangeable with D2O.
Theoretical discussion| 55
Ethyl 2-(4-oxo-4,5-dihydrothiazol-2-yl)-3-(4-sulfamoylphenylamino)-3- thioxopropanoate (VII)
The thiazolidinone derivative VII was obtained by refluxing compound VI with thioglycolic acid. The reaction proceeded by addition followed by intramolecular cyclization.
IR spectrum of compound VII revealed the absence of the band corresponding to the carbonitrile group and the presence of a band at 1710 cm-1 corresponding to (C=O) attached to the thiazole ring and 1625 cm-1 corresponding to (C=N). Additionally, 1H-NMR spectrum of compound VII revealed the presence of a singlet at 3.9 ppm corresponding to the CH2 of the thiazole ring. Mass spectrum of compound VII exhibited a molecular ion peak at m/z 401 [M+, 12.23%], with a base peak at m/z 44.
Theoretical discussion| 56
4-(4-Methylbenzylideneamino)benzenesulfonamide (VIIIa)106 and 4-(4- nitrobenzylideneamino) benzenesulfonamide (VIIIb). 117
Compounds VIIIa, b were prepared according to the reported method.106
4-(2-(Substituted phenyl)-4-oxothiazolidin-3-yl)benzenesulfonamide IXa-h
Compounds IXa-h were prepared according to the published procedure.106
The formation of thiazolidinone IXa-h was preceded by a one pot reaction via refluxing sulfanilamide I with the required aromatic aldehyde and thioglycolic acid in dry benzene for 48 hours. In addition, a two step Theoretical discussion| 57 reaction can be conducted by refluxing Schiff’s base with thioglycolic acid in dry benzene for additional 18 hours.
The IR spectra of compounds IXa-h showed the absence of the band corresponding to (C=N) of the schiff base and the appearance of (C=O) band attached to the thiazole ring ranging from 1691-1705 cm-1.
Additionally, the 1H-NMR spectra of compounds IXa-h showed the appearance of two singlets ranging from 3.6- 4.1 ppm and 5.9- 6.4 ppm,
attributed to the presence of CH2 and CH groups of the thiazole ring, and multiplet ranging from 6.2- 8.0 ppm attributed to the aromatic protons and
SO2NH2.
1H-NMR spectrum of compound IXa showed a singlet at 2.3 ppm
attributed to the presence of CH3 group.
-1 IR spectrum of compound IXb showed NO2 bands at 1514, 1310 cm .
IR spectra of compounds IXd, e showed OH band at 3450, 3456 cm-1, respectively. Also, 1H-NMR spectra of compounds IXd, e showed a singlet at 10.5 and 9.1 ppm, respectively, attributed to the presence of OH.
1H-NMR spectrum of compound IXf showed a singlet at 6.0 ppm attributed to the presence of O-CH2-O group.
IR spectrum of compound IXg showed (C-Cl) band at 790 cm-1.
Theoretical discussion| 58
4-(5-Amino-7-( substituted phenyl)-6-cyano-7H-thiazolo[4,5-b]pyrane-2- ylamino) benzenesulfonamides (XIa-d).
Treatment of thiazolidinone III with arylidine malononitrile derivatives Xa-d in presence of a catalytic amount of piperidine, as a basic catalyst, yielded the corresponding Thiazolopyrane derivatives XIa-d, via the formation of the intermediates followed by intramolecular cyclization. The arylidenes were prepared by just stirring the corresponding aldehyde with malononitrile in ethanol containing few drops of TEA for about 20 min at room temperature.118
The formation of compounds XIa-d was proved from their micro- analytical and spectral data. IR spectra of compounds XIa-d showed the Theoretical discussion| 59 disappearance of (C=O) band, and the appearance of (C≡N) band at 2220- 2194 cm-1 and (C=N) at 1620-1610 cm-1. Additionally, the 1H-NMR spectra showed two singlets one at 4.0 - 4.1 ppm and the other at 12.0-12.5 ppm attributed to the presence of CH of pyrane ring and NH2 of the orthoaminocarbonitrile group, respectively.
IR spectrum of compound XIa showed two bands at 1514, 1310 cm-1 corresponding to (NO2). Also, IR spectrum of compound XIc showed a band at 750 cm-1 corresponding to (C-Cl).
8-Amino, 9-(4-nitrophenyl) 9-H,[2-(4-sulfamoylphenyl amino)] thiazolo- [4,5-b ]pyrano [2,3-d] pyrimidine (XII)
The thiazolopyranopyrimidine derivative XII was obtained by the reaction of compound XIa with formamide, where cyclization occurred through elimination of one molecule of water, followed by intramolecular cyclization.119
IR spectrum of compound XII revealed the absence of the band corresponding to the (C≡N) group. Additionally, 1H-NMR spectrum of compound XII showed two singlets corresponding to the NH2 group Theoretical discussion| 60
attached to the pyrimidine at 7.0 ppm which is exchangeable with D2O, and the CH of pyrimidine at 8.3 ppm.
8-Oxo 9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo- [4,5-b ]pyrano [2,3-d] pyrimidine (XIIIa) and 8-oxo, 9-(3,4-dimethoxy phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo [4,5-b ]pyrano [2,3- d] pyrimidine(XIIIb)
The thiazolopyranopyrimidine derivatives XIIIa,b were obtained by refluxing compound XIa,b in formic acid. This reaction proceeded via condensation followed by elimination of two moles of water .120
Theoretical discussion| 61
IR spectra of compounds XIIIa,b revealed the absence of the band corresponding to the (C≡N) group.
IR spectrum of compound XIIIa showed a band at 1705 cm-1 attributed to the presence of (C=O) group. Additionally, 1H-NMR spectrum of compound XIIIa exhibited two singlets corresponding to the NH group of pyrimidine at 8.2 ppm and the CH of pyrimidine at 8.3 ppm. Mass spectrum of compound XIIIa exhibited a molecular ion peak at m/z 498 [M+, 12.15%], with a base peak at m/z 179.
Also, IR spectrum of compound XIIIb showed a band at 1714 cm-1 attributed to the presence of (C=O) group. Additionally, 1H-NMR spectrum of compound XIIIb exhibited three singlets at 4.0, 8.2 and 8.3 ppm corresponding to 2OCH3, NH of pyrimidine, and CH of pyrimidine, respectively. Mass spectrum of compound XIIIb exhibited a molecular ion peak at m/z 513 [M+ , 0.2%], with a base peak at m/z 194.
6-Methyl, 8-oxo,9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano[2,3-d]pyrimidine (XIVa) and 6-methyl, 8-oxo,9- (3,4-dimethoxyphenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5- b] pyrano [2,3-d] pyrimidine (XIVb)
When compound XIa,b were refluxed in acetic anhydride for short or long time (5 h and 10 h), the pyrimidine derivatives XIVa,b was isolated via the formation of monoacetyl derivative C rather than the diacetyl derivative D on the bases of IR and 1H-NMR spectroscopy.
Theoretical discussion| 62
The reaction proceeded via acetylation of the amino group affording the corresponding fused pyrimido derivative XIVa,b via Dimroth rearrangement.121
IR spectra of compounds XIVa,b revealed the absence of the band corresponding to the (C≡N) group.
IR spectrum of compound XIVa showed a band at 1696 cm-1 attributed to the presence of (C=O) group. Additionally, 1H-NMR spectrum of compound
XIVa exhibited two singlets corresponding to the CH3 group at 1.9 ppm and the NH of pyrimidine at 8.1 ppm which is exchangeable with D2O. Mass spectrum of compound XIVa exhibited a molecular ion peak at m/z 512 [M+, 2.43%], with a base peak at m/z 78. Theoretical discussion| 63
IR spectrum of compound XIVb showed a band at 1719 cm-1 attributed to the presence of (C=O) group. Additionally, 1H-NMR spectrum of compound XIVb exhibited three singlets corresponding to the CH3 group at
1.8 ppm, 2 OCH3 at 4.0 ppm and the NH of pyrimidine at 8.2 ppm which is exchangeable with D2O. Mass spectrum of compound XIVb exhibited a molecular ion peak at m/z 527 [M+, 4.4%], with a base peak at m/z 100.
8-Amino, 6-oxo ,9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b ]pyrano [2,3-d] pyrimidine (XV) and 8-amino, 6-thioxo, 9- (4-nitro phenyl) 9-H,[2-(4-sulphamoyl phenyl amino)] thiazolo[4,5-b ] pyrano [2,3-d] pyrimidine (XVI)
Compounds XV and XVI were obtained by fusion of compound XIa with urea and/ or thiourea, respectively. This reaction proceeded by nucleophilic substitution followed by intramolecular cyclization to give the corresponding 2-oxo- and 2-thioxothiazolopyranopyrimidine, respectively.
IR spectra of compounds XV, XVI revealed the absence of the band corresponding to the (C≡N) group.
IR spectrum of compound XV showed a band at 1697 cm-1 attributed to the presence of (C=O) group. Additionally, 1H-NMR spectrum of compound Theoretical discussion| 64
XV exhibited two singlets at 8.0 and 8.2 ppm corresponding to the NH2 group attached to the pyrimidine ring and NH group of the pyrimidine, respectively (exchangeable with D2O). Mass spectrum of compound XV exhibited a molecular ion peak at m/z 513 [M+, 16%], with a base peak at m/z 78.
IR spectrum of compound XVI showed a band at 1280 cm-1 attributed to the presence of (C=S) group. Additionally, 1H-NMR spectrum of compound
XVI exhibited two singlets at 8.0 and 8.2 ppm corresponding to the NH2 group attached to the pyrimidine ring and NH group of the pyrimidine, respectively (exchangeable with D2O). Mass spectrum of compound XVI exhibited a molecular ion peak at m/z 529 [M+, 3.22%], with a base peak at m/z 194.
4-(6-Cyano-7-(4-nitrophenyl)-2-(4-sulfamoylphenylamino)-7H-thiazolo [4,5-b] pyran-5-ylamino)-4-oxobutanoic acid (XVII) and 4-(6-cyano-5- (2,5-dioxopyrrolidinyl)-7-(4-nitrophenyl)-7H- thiazolo [4,5-b] pyran--2- ylamino) benzenesulfonamide (XVIII)
When compound XIa and succinic anhydride were refluxed together in presence of ethanol the oxobutanoic acid derivative XVII was formed. While, when they were fused together, intramolecular cyclization occurred via the elimination of one mole of water to obtain the thiazolopyrane derivative bearing dioxopyrrolidine ring XVIII.119 Theoretical discussion| 65
IR spectrum of compound XVII showed two bands at 1710 and 1685 cm- 1 attributed to the presence of 2(C=O) group. Additionally, 1H-NMR spectrum of compound XVII exhibited two triplets at 1.2 and 1.6 ppm attributed to the presence of (CH2-CH2) group, and two singlets at 8.2 and 10.5 ppm corresponding to the NH group of the orthoaminocarbonitrile and the OH group, respectively. Mass spectrum of compound XVII exhibited a molecular ion peak at m/z 570 [M+, 2.5%], with a base peak at m/z 63.
Also, IR spectrum of compound XVIII showed two bands at 1700 and 1680 cm-1 attributed to the presence of 2(C=O) group. Additionally, 1H- NMR spectrum of compound XVIII exhibited two triplets at 1.5 and 1.6 ppm attributed to the presence of 2CH2 group. Mass spectrum of compound XVIII exhibited a molecular ion peak at m/z 552 [M+, 4.09%], with a base peak at m/z 92.
Theoretical discussion| 66
N-(6-Cyano-7-(4-nitrophenyl)-2-(4-sulfamoylphenylamino)-7H- thiazolo [4,5-b] pyran-5-yl)-3-oxobutanamide (XIX)
Compound XIX was obtained by reaction of compound XIa with ethyl acetoacetate under fusion. The reaction proceeded via the elimination of one mole of ethanol.121
IR spectrum of compound XIX showed two bands at 1717 and 1710 cm-1 attributed to the presence of (2C=O) group. Additionally, 1H-NMR spectrum of compound XIX exhibited three singlets at 1.6, 2.2 and 8.2 ppm corresponding to the COCH3, CH2 and the NH group of the orthoaminocarbonitrile (exchangeable with D2O), respectively. Mass spectrum of compound XIX exhibited a molecular ion peak at m/z 554 [M+, 7.28%], with a base peak at m/z 129.
Theoretical discussion| 67
8-Imino, 6-thioxo, 9-(3,4-dimethoxy phenyl) 9-H,1N phenyl,[2-(4- sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano [2,3-d] pyrimidine (XX)
Refluxing compound XIb with phenyl isothiocyanate yielded the corresponding thiazolopyranopyrimidine derivative XX.119
IR spectrum of compound XX revealed the absence of (C≡N) band and the presence of a band at 1270 cm-1 corresponding to (C=S) group. Additionally, 1H-NMR spectrum of compound XX exhibited three singlets at 4.0, 8.2 and 11.6 ppm corresponding to the 2OCH3, NH of the pyrimidine ring (exchangeable with D2O) and the NH imino group (exchangeable with
D2O), respectively. Mass spectrum of compound XX exhibited a molecular ion peak at m/z 620 [M+ -2, 10%], with a base peak at m/z 271.
EXPERIMENTAL
Experimental| 69
4. Experimental
Remarks:
All melting points are uncorrected and were determined on a Stuart melting point apparatus (Stuart Scientific, Redhill, UK) at the National Centre for Radiation Research & Technology, Atomic Energy Authority. Elemental analyses (C, H, N) were performed on Perkin-Elmer 2400 analyser (Perkin-Elmer, Norwalk, CT, USA) at the Microanalytical Laboratories of the Faculty of Science, Cairo University. Infrared spectra (KBr) were determined using Shimadzu IR-110 spectrophotometer (Shimadzu, Koyoto, Japan) at the National Centre for Radiation Research & Technology, Atomic Energy Authority. 1H-NMR spectra were carried out using BRUCKER proton NMR-Avance
300 (300, MHz), in DMSO-d6 as a solvent, using tetramethylsilane (TMS) as internal standard, at the NMR Laboratory of the Faculty of Science, Cairo University. Mass spectra were run on JEOL JMS AX-500 mass spectrometer (JEOL JMS, Tokyo, Japan) at the Microanalytical Laboratories of the National Research centre. All compounds were chemically named using ChemDraw Ultra software (V 11.0). All reactions were monitored by thin layer chromatograph (TLC) using precoated Aluminium sheets Silica gel Merck 60 F254. Ethyl acetate/Cyclohexane (2.5:7.5 mL) mixture was used as eluting solvent and TLC sheets were visualized by UV lamp (Merck, Darmstadt, Germany). Experimental| 70
2-Chloro-N-(4-sulfamoylphenyl) acetamide (II)113
Prepared according to the reported method.105
A mixture of sulfanilamide I (1.72 g, 0.01 mol) and chloro acetyl chloride (1.1 ml, 0.01 ml) in dimethyl formamide (30 ml) was stirred at room temperature for 2 h. The reaction mixture was poured onto iced water. The solid obtained was crystallized from ethanol to give II.
Melting point: 214-216 °C (as reported). Yield: 85% (as reported).
Experimental| 71
4-(4-Oxo-4,5-dihydrothiazol-2-ylamino)benzenesulfonamide (III)114
Prepared according to the reported procedure.105
A mixture of II (2.34 g, 0.01 mol) and ammonium thiocyanate (0.76 g, 0.01 mol) in ethanol (30 mL) was refluxed for 1 h. The reaction mixture was filtered on hot and crystallized from dioxane
Melting point: 244-246°C (as reported). Yield: 78% (as reported).
Experimental| 72
4-(7-Phenyl-5-thioxo-4,5,6,7-tetrahydrothiazolo[4,5-d]pyrimidin-2- ylamino) benzenesulfonamide (IVa) and 4-(5-thioxo-7-p-tolyl-4,5,6,7- tetrahydrothiazolo[4,5-d]pyrimidin-2-ylamino) benzenesulfonamide (IVb)
A mixture of compound III (2.71 g, 0.01 mol), thiourea (0.76 g , 0.01 mol) and benzaldehyde (1 g, 0.01 mol) or 4-methyl benzaldehyde (1.2 g, 0.01 mol) in ethanol (50 mL) containing few drops of Conc. HCl was refluxed for 3 h. The reaction mixture was poured onto iced water. The solid obtained was crystallized from dioxane to give IVa and IVb, respectively.
Table 1, Physico-chemical and analytical data for compounds IVa,b.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% IVa > 300 55 C17H15N5O2S3 48.90 3.62 16.77 (417.53) 48.80 3.75 16.40 IVb >300 60 C18H17N5O2S3 50.10 3.97 16.23 (431.55) 50.30 3.88 16.05
Experimental| 73
Table 2, Spectral data for compounds IVa,b:
No. Type Data
IR 3330, 3280, 3120 (NH, NH2), 3065 (CH arom.), (KBr, cm -1) 2970, 2930 (CH aliph.), 1650 (C=N), 1350, 1180
(SO2), 1270(C=S). 1H-NMR δ: 4.2 (s, 1H, CH pyrimidine), 7.2-7.9 (m, 12 H, Ar-
(DMSO-d6) H+ SO2NH2+ NH sulfanilamide), 11.9, 12.3 (2s, 2H,
IVa ppm 2NH exchangeable with D2O). EI/MS 417 [M+] (18.5), 134 (100). (m/z) (relative abundance)
IR 3313, 3246, 3120 (NH, NH2), 3070 (CH arom.), (KBr, cm -1) 2976, 2930 (CH aliph.), 1630 (C=N), 1324, 1180
IVb (SO2), 1290 (C=S). 1 H-NMR δ: 2.3 (s, 3H, CH3), 5.0 (s, 1H, CH pyrimidine), 7.2-
(DMSO-d6) 7.9 (m, 11 H, Ar-H+ SO2NH2 +NH sulfanilamide),
ppm 11.9, 12.5 (2s, 2H, 2NH exchangeable with D2O).
Experimental| 74
4-Isothiocyanato benzenesulfonamide (V) 116
A solution of sulfanilamide I (1.72 g, 0.01 mol) in water (200 mL) was added to thiophosgene (1.15 g, 0.01mol), and stirred at room tempreture for 1 h (until the disappearance of red color), filter and the solid obtained was collected and crystallized from dioxane to give V.
Melting point: 208-210°C (as reported). Yield: 78% (as reported).
Experimental| 75
Ethyl 2-cyano-3-(4-sulfamoylphenylamino)-3-thioxopropanoate (VI)
A solution of compound V (2.14 g, 0.01 mol) in THF (20 mL) was added to ethyl cyanoacetate sodium salt, prepared by addition of (2.3 g, 0.1 mol) sodium metal to a solution of ethyl cyanoacetate (11.3 g, 0.1 mol) in THF (50 mL), the reaction mixture was poured onto acidified water (20 mL), where an oily product was obtained, extracted with diethyl ether, then evaporated and the residual solid was crystallized from ethanol to give VI.
Table 3, Physico-chemical and analytical data for compound VI.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N%
VI 168-170 62 C12H13N3O4S2 44.02 4.00 12.84 (327.38) 44.26 4.17 13.00
Table 4, Spectral data for compound VI:
No. Type Data IR 3410, 3367, 3257 (NH, NH2), 3070 (CH arom.), (KBr, cm -1) 2984, 2927 (CH aliph.), 2274(C≡N), 1689 (C=O), 1380, 1130 (SO2), 1274(C=S). 1 VI H-NMR δ: 1.2 (t, J=6 Hz,3H, CH3), 2.1 (s, 1H, CH), 4.1 (q, (DMSO-d6) J=6 Hz, 2H, CH2), 7.3 (s, 2H, SO2NH2 exchangeable ppm with D2O), 7.7, 8.1 (2d, 4H,AB system), 12.4(s, 1H, Experimental| 76
NH exchangeable with D2O).
Ethyl 2-(4-oxo-4,5-dihydrothiazol-2-yl)-3-(4-sulfamoylphenylamino)-3- thioxopropanoate (VII)
A mixture of compound VI (3.27 g, 0.01 mol) and thioglycolic acid (0.92 g, 0.01 mol) in glacial acetic acid (40 mL) was refluxed for 5 h. The reaction mixture was concentrated and the residue was triturated with ethanol. The solid obtained was crystallized from ethanol.
Table 5, Physico-chemical and analytical data for compound VII.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% VII >300 50 C14H15N3O5S3 41.88 3.77 10.47 (403.50) 41.78 3.82 10.27 Table 6, Spectral data for compound VII:
No. Type Data IR 3470, 3300, 3240 (NH, NH2), 2950, 2907 (CH (KBr, cm -1) aliph.) 1710, 1693(2 C=O), 1625 (C=N), 1396, 1156 (SO2), 1284 (C=S).
1 H-NMR δ: 1.2 (t, J=7 Hz 3H, CH3), 2.0 (s, 1H, CH), 3.9 (s, VII (DMSO-d6) 2H, CH2 thiazole), 4.0 (q, J=7 Hz 2H, CH2), 7.3 (s, ppm 2H, SO2NH2 exchangeable with D2O), 7.7-8.0 (2d, 4H, AB system, Ar-H), 12.1 (s, 1H, NH exchangeable with D2O). Experimental| 77
EI/MS(m/z) 401 [M+-2] (12.23), 44 (100). (relative abundance) 4-(4-Methylbenzylideneamino)benzenesulfonamide ( VIIIa)106 and 4-(4- nitrobenzylideneamino) benzenesulfonamide (VIIIb). 117
Prepared according to the reported procedure.106
A mixture of sulfanilamide I (1.72 g, 0.01 mol) and 4-methyl benzaldehyde or 4-nitro benzaldehyde (0.01 mol) was refluxed in ethanol (50 mL) for 12 h. The reaction mixture was poured onto iced water and filtered. The solid obtained was crystallized from dioxane to give VIIIa, b, respectively.
Melting point of VIIIa: 193-195 °C (as reported).
Yield of VIIIa: 69% (as reported).
Melting point of VIIIb: 182-184 °C (as reported).
Yield of VIIIb: 87% (as reported).
Experimental| 78
4-(2-(Substituted phenyl)-4-oxothiazolidin-3-yl) benzenesulfonamides (IXa-h).
A mixture of sulfanilamide I (1.72 g, 0.01 mol) was refluxed with mercaptoacetic acid (0.92 g, 0.01 mol) and the appropriate aldehyde (0.01 mol) in dry benzene for 48 hrs. Benzene was decanted and the sticky product was triturated with sodium carbonate to remove the excess mercaptoacetic acid. The product was then dissolved in ethanol and precipitated by addition of diethyl ether. The precipitated product was filtered and crystallized from dioxane to give IXa-h, respectively.
Table 7, Physico-chemical and analytical data for compounds IX(a-h).
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% IXa 264-266 63 C16H16N2O3S2 55.15 4.63 7.88 (347.98) 55.00 4.43 8.04 IXb 190-194 67 C15H13N3O5S2 47.48 3.45 11.08 (379.41) 47.53 3.28 10.78 IXc 154-156 65 C15H14N2O3S2 53.87 4.22 8.38 (334.41) 53.49 4.03 7.98 IXd >300 61 C15H14N2O4S2 51.41 4.03 7.99 (350.04) 51.85 3.92 8.23 IXe 204-206 63 C15H14N2O4S2 51.41 4.03 7.99 Experimental| 79
(350.04) 51.30 3.98 7.78 IXf 192-194 72 C16H14N2O5S2 50.78 3.73 7.40 (378.42) 50.44 3.63 7.35 IXg 188-190 59 C15H13ClN2O3S2 48.84 3.55 7.59 (368.32) 48.54 3.65 7.34 IXh 206-208 68 C15H13BrN2O3S2 43.59 3.17 6.78 (411.90) 43.50 3.35 6.36
Table 8, Spectral data for compounds IX(a-h).
No. Type Data
IR 3312, 3211 (NH2), 3043 (CH arom.), 2933, 2910 - 1 (KBr, cm ) (CH aliph.), 1705 (C=O), 1324, 1148(SO2). 1 H-NMR δ: 2.3 (s, 3H, CH3), 3.6 (s, 2H, CH2 thiazole), 5.9 (s,
IXa (DMSO-d6) 1H, CH thiazole), 7.1-7.9 (m, 10 H, Ar-H+
ppm SO2NH2). EI/MS 348 [M+] (16.19), 120 (100). (m/z) (relative abundance)
IR 3300, 3221 (NH2), 3040 (CH arom.), 2937, 2910 - 1 (KBr, cm ) (CH aliph.), 1705(C=O), 1514, 1310 (NO2), 1380,
1130 (SO2). 1 IXb H-NMR δ: 4.0 (s, 2H, CH2 thiazole), 6.4 (s, 1H, CH
(DMSO-d6) thiazole), 7.1-8.0 (m, 10 H, Ar-H+ SO2NH2). ppm EI/MS(m/z) 379 [M+] (12.05), 79 (100). (relative abundance)
IR 3340, 3252 (NH2), 3104 (CH arom.), 2920, 2910 - 1 (KBr, cm ) (CH aliph.), 1694 (C=O), 1328, 1155 (SO2). 1 H-NMR δ: 4.0 (s, 2H, CH2 thiazole), 5.9 (s, 1H, CH
IXc (DMSO-d6) thiazole), 7.1-8.0 (m, 11 H, Ar-H+ SO2NH2 ). ppm EI/MS(m/z) 334 [M+] (12.82), 87 (100). Experimental| 80
(relative abundance)
IR 3450 (OH), 3269, 3188 (NH2), 3077(CH arom.), (KBr, cm- 1) 2925, 2906 (CH aliph.), 1704 (C=O), 1328, 1154
(SO2). 1 IXd H-NMR δ: 4.0 (s, 2H, CH2 thiazole), 5.9 (s, 1H, CH
(DMSO-d6) thiazole), 7.2-7.7 (m, 10 H, Ar-H+ SO2NH2), 10.5 ppm (s, 1H, OH). EI/MS 348 [M• -2] (11.79), 230 (100). (m/z) (relative abundance)
IR 3456(OH), 3363, 3252 (NH2), 3067 (CH arom.), (KBr, cm- 1) 2920, 2907 (CH aliph.), 1701(C=O), 1322, 1170
(SO2). 1 IXe H-NMR δ: 4.0 (s, 2H, CH2 thiazole), 5.9 (s, 1H, CH
(DMSO-d6) thiazole), 6.6-7.5 (m, 10 H, Ar-H+ SO2NH2 ), 9.1 (s, ppm 1H, OH). EI/MS(m/z) 348 [M• -2] (13.08), 87 (100). (relative abundance)
IR 3480, 3269(NH2), 3105(CH arom.), 2917, 2903 (CH - 1 (KBr, cm ) aliph.), 1698 (C=O), 1329, 1158(SO2). 1 H-NMR δ: 4.0 (s, 2H, CH2 thiazole), 5.9 (s, 1H, CH
IXf (DMSO-d6) thiazole), 6.0 (s, 2H, O-CH2 -O), 6.2-7.8 (m, 9 H, Ar
ppm –H+ SO2NH2 ). EI/MS(m/z) 378 [M+] (12.07), 87 (100). (relative abundance)
IR 3300, 3242 (NH2), 3118 (CH arom.), 2923, 2908 - 1 (KBr, cm ) (CH aliph.), 1691 (C=O), 1377, 1158 (SO2), 790 (C- Cl). 1 IXg H-NMR δ: 4.0 (s, 2H, CH2 thiazole), 5.9 (s, 1H, CH
(DMSO-d6) thiazole), 6.6-8 (m, 10H, Ar –H+ SO2NH2). Experimental| 81
ppm EI/MS(m/z) [M+] at 368 (11.26), 370 (3.82), 87(100). (relative abundance)
IR 3269, 3201 (NH2), 3120 (CH arom.), 2928, 2912 - 1 (KBr, cm ) (CH aliph.), 1691 (C=O), 1354, 1166 (SO2). 1 H-NMR δ: 4.1 (s, 2H, CH2 thiazole), 6.2 (s, H, CH thiazole),
IXh (DMSO-d6) 7.1-7.9 (m, 10 H, Ar-H+ SO2NH2). ppm EI/MS (m/z) [M+] at 412 (12.57), 414 (12.8), 87 (100). (relative abundance)
Experimental| 82
4-(5-Amino-7-( substituted phenyl)-6-cyano-7H-thiazolo[4,5-b]pyran-2- ylamino) benzenesulfonamides (XIa-d).
A mixture of compound III (2.71 g, 0.01 mol) and benzylidene malononitrile Xa-d (0.01 mol) in ethanol (50 mL) containing piperidine (1 mL) was refluxed for 8 h. The reaction mixture was filtered and the filtrate was poured onto iced water. The solid obtained was crystallized from dioxane to give XIa-d.
Table 9, Physico-chemical and analytical data for compounds XI(a-d).
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N%
XIa 178-180 67 C19H14N6O5S2 48.50 3.00 17.86 (470.48) 48.38 2.95 17.95
XIb 216-218 64 C21H19N5O5S2 51.95 3.94 14.42 (485.54) 51.93 3.82 14.27
XIc >300 64 C19H14ClN5O3S2 49.62 3.07 15.23 (459.93) 49.47 3.22 15.12
XId 250-252 65 C19H14BrN5O3S2 45.24 2.80 13.89 (503.30) 45.35 2.71 13.78
Experimental| 83
Table 10, Spectral data for compounds XI(a-d)
No. Type Data
IR 3354, 3237, 3077 (NH, NH2), 3067 (CH arom.), (KBr, cm- 1) 2943, 2974 (CH aliph.), 2194 (C≡N), 1620 (C=N),
1514, 1310 (NO2), 1360, 1190(SO2). 1H-NMR δ: 4.0 (s, 1H, CH pyrane), 6.5 -8.0 (m, 10 H, Ar-H+
XIa (DMSO-d6) SO2NH2), 8.2 (s, 1H, NH exchangeable with D2O),
ppm 12.0 (s, 2H, NH2 exchangeable with D2O) EI/MS(m/z) 470 [M+] (2.91), 63(100). (relative abundance)
IR 3390, 3300, 3280 (NH, NH2), 3060 (CH arom.), (KBr, cm- 1) 2960, 2980 (CH aliph.), 2220 (C≡N), 1617 (C=N),
1380, 1130(SO2). 1 XIb H-NMR δ: 4.0 (s, 6H, 2 OCH3), 4.1 (s, 1H, CH pyrane), 7.1-
(DMSO-d6) 8.0 (m, 9H, Ar-H+ SO2NH2), 8.1 (s, 1H, NH ppm exchangeable with D2O), 12.0 (s, 2H, NH2 exchangeable with D2O). EI/MS(m/z) 485 [M+] (2.54), 209(100). (relative abundance)
IR 3453, 3230, 3113 (NH, NH2), 3070 (CH arom.), (KBr, cm- 1) 2960, 2984 (CH aliph.), 2202 (C≡N), 1610 (C=N), XIc 1340, 1170 (SO2), 750 (C-Cl). 1H-NMR δ: 4.1 (s, 1H, CH pyrane), 7.2-7.9 (m, 10 H, Ar-H+
(DMSO-d6) SO2NH2), 8.0 (s, 1H, NH exchangeable with D2O), ppm 12.5 (s, 2H, NH2 exchangeable with D2O).
IR 3343, 3251, 3150 (NH, NH2), 3080 (CH arom.), (KBr, cm- 1) 2942, 2974 (CH aliph.), 2199 (C≡N), 1613 (C=N),
XId 1340, 1170 (SO2). 1H-NMR δ: 4.0(s, 1H, CH pyrane), 6.6- 8.0 (m, 10 H, Ar-H+
(DMSO-d6) SO2NH2), 8.2 (s, 1H, NH exchangeable with D2O),
ppm 12.0(s, 2H, NH2 exchangeable with D2O).
Experimental| 84
8-Amino,9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b ]pyrano [2,3-d] pyrimidine (XII)
A solution of compound XIa (4.7 g, 0.01 mol) in formamide (30 mL), was refluxed for 3 h. The reaction mixture was poured onto iced water. The solid obtained was crystallized from ethanol to give XII.
Table 11, Physico-chemical and analytical data for compound XII.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XII 252-254 65 C20H15N7O5S2 48.28 3.04 19.71 (499.21) 48.32 3.10 19.83 Table 12, Spectral data for compound XII:
No. Type Data IR 3222 (br NH, NH2), 3050 (CH arom.), 2948, 2929 -1 (KBr, cm ) (CH aliph.), 1625 (C=N), 1510, 1350 (NO2), 1380, 1130 (SO2). 1 H-NMR δ: 4.0 (s, 1H, CH pyrane), 7.0 (s, 2H, NH2 XII (DMSO-d6) exchangeable with D2O), 7.1- 8.0 (m, 10H, Ar-H+ ppm SO2NH2), 8.2 (s, 1H, NH exchangeable with D2O), 8.3 (s, 1H, CH pyrimidine). EI/MS(m/z) 499 [M+] (13.05), 194 (100). (relative abundance) Experimental| 85
8-Oxo, 9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo- [4,5-b ]pyrano [2,3-d] pyrimidine (XIIIa) and 8-Oxo, 9-(3,4-dimethoxy phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo [4,5-b ]pyrano [2,3- d] pyrimidine (XIIIb)
A solution of compound XIa,b (0.01 mol) in formic acid (30 mL), was refluxed for 3 h. The reaction mixture was poured onto ice cold water. The solid obtained was crystallized from dioxane to give XIIIa,b.
Table 13, Physico-chemical and analytical data for compound XIIIa,b.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N%
XIIIa 212-214 73 C20H14N6O6S2 48.19 2.83 16.86 (498.04) 47.98 2.92 16.79
XIIIb 250-252 64 C22H19N5O6S2 51.45 3.73 13.64 (513.08) 51.38 3.79 13.66
Experimental| 86
Table 14, Spectral data for compound XIIIa,b:
No. Type Data
IR 3480, 3263, 3200 (NH, NH2), 3050 (CH arom.), (KBr, cm -1) 2929, 2950 (CH aliph.), 1705 (C=O), 1625(C=N),
1524, 1304 (NO2), 1329, 1158(SO2). 1H-NMR δ: 4.0(s, 1H, CH pyrane), 7.2-8.0 (m, 10H, Ar-H+
XIIIa (DMSO-d6) SO2NH2), 8.2 (s, 2H, 2NH exchangeable with
ppm D2O), 8.3 (s, 1H, CH pyrimidine). EI/MS(m/z) 498 [M+] (12.15), 179(100). (relative abundance)
IR 3217 (br NH, NH2), 3085 (CH arom.), 2913, 2924 (KBr, cm -1) (CH aliph), 1714 (C=O), 1608 (C=N), 1380, 1130
(SO2). 1 H-NMR δ: 4.0 (s, 6H, 2 OCH3), 4.1 (s, 1H, CH pyrane), 7.0-
XIIIb (DMSO-d6) 7.9 (m, 9 H, Ar-H+ SO2NH2), 8.2 (s, 2H, 2NH
ppm exchangeable with D2O), 8.3 (s, 1H, CH pyrimidine). EI/MS(m/z) 513 [M+] (0.2), 194 (100). (relative abundance)
Experimental| 87
6-Methyl,8- oxo,9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b ] pyrano [2,3-d] pyrimidine (XIVa) and 6-Methyl, 8- oxo, 9-(3,4-dimethoxy phenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo- [4,5-b] pyrano [2,3-d] pyrimidine (XIVb)
A solution of compound XIa,b (0.01 mol) in acetic anhydride (20 mL) was refluxed for 10 h, the reaction mixture was then concentrated, and the solid separated was crystallized from ethanol to give XIVa,b, respectively. Table 15, Physico-chemical and analytical data for compound XIVa,b.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N%
XIVa 185-187 75 C21H16N6O6S2 49.21 3.15 16.40 (512.52) 49.38 3.25 16.33
XIVb 188-190 66 C23H21N5O6S2 52.36 4.01 13.27 (527.57) 52.22 4.13 13.12
Experimental| 88
Table 16, Spectral data for compound XIVa,b:
No. Type Data
IR 3460, 3351, 3310 (NH, NH2), 3077(CH arom.), (KBr, cm -1) 2930, 2953 (CH aliph.), 1696 (C=O), 1600 (C=N),
1520, 1314 (NO2), 1377, 1158 (SO2). 1 H-NMR δ: 1.9 (s, 3H, CH3), 4.0(s, 1H, CH pyrane),7.1-8.0
XIVa (DMSO-d6) (m, 10 H, Ar-H+ SO2NH2), 8.1 (s, 2H, 2 NH
ppm exchangeable with D2O). EI/MS(m/z) 512 [M+] (2.43), 78(100). (relative abundance)
IR 3490, 3300, 3242 (NH, NH2), 3078 (CH arom.), (KBr, cm -1) 2945, 2910 (CH aliph.), 1719 (C=O), 1628 (C=N), 1380, 1130 (SO2). 1 XIVb H-NMR 1.8 (s, 3H, CH3), 4.0 (s, 6H, 2 OCH3), 4.1 (s, 1H,
(DMSO-d6) CH pyrane), 7.1-8.0 (m, 9H, Ar-H+ SO2NH2), ppm 8.2(s, 2H, 2NH exchangeable with D2O). EI/MS(m/z) 527 [M+] (4.4), 100 (100). (relative abundance)
Experimental| 89
8-Amino, 6-oxo ,9-(4-nitrophenyl) 9-H,[2-(4-sulfamoyl phenyl amino)] thiazolo[4,5-b]pyrano [2,3-d] pyrimidine (XV)
A mixture of compound XIa (4.7 g, 0.01 mol) and urea (0.6 g, 0.01 mol) was fused at 220 ºC for 15 min, then ethanol was added to the reaction mixture while hot and filtered. The solid obtained was crystallized from dioxane to give XV.
Table 17, Physico-chemical and analytical data for compound XV.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XV >300 50 C20H15N7O6S2 46.78 2.94 19.09 (513.51) 46.49 2.92 19.10
Table 18, Spectral data for compound XV:
No. Type Data IR 3400, 3337, 3200 (NH, NH2), 3060 (CH arom.), (KBr, cm -1) 2950, 2928 (CH aliph.), 1697 (C=O), 1612 (C=N), 1539, 1335 (NO2), 1329, 1158 (SO2). 1H-NMR δ: 4.0 (s, 1H, CH pyrane), 6.8-8.0 (m, 12H, Ar-H, XV (DMSO-d6) 2NH2), 8.2 (s, 2H, 2NH exchangeable with D2O). ppm EI/MS(m/z) 513 [M+] (16), 78(100). (relative abundance) Experimental| 90
8-Amino, 6-thioxo ,9-(4-nitrophenyl) 9-H,[2-(4-sulphamoyl phenyl amino)] thiazolo[4,5-b] pyrano [2,3-d] pyrimidine (XVI)
A mixture of compound XIa (4.7 g, 0.01 mol) and thiourea (0.76 g, 0.01 mol) was fused at 220 ºC for 15 min. Ethanol was added to the reaction mixture while hot and filtered. The solid obtained was crystallized from dioxane to give XVI.
Table 19, Physico-chemical and analytical data for compound XVI.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XVI >300 59 C20H15N7O5S3 45.36 2.85 18.51 (529.57) 45.30 2.77 18.65
Table 20, Spectral data for compound XVI:
No. Type Data IR 3314, 3320, 3220 (NH, NH2), 3080 (CH arom.), (KBr, cm -1) 2945, 2920 (CH aliph.), 1615 (C=N), 1280 (C=S), 1530, 1345 (NO2), 1329, 1158(SO2). 1H-NMR δ: 4.0 (s, 1H, CH pyrane), 6.8-8.0 (m, 12H, Ar-H, XVI (DMSO-d6) 2NH2), 8.2 (s, 2H, 2NH exchangeable with D2O). ppm EI/MS(m/z) 529 [M+] (3.22), 194(100). (relative abundance) Experimental| 91
4-(6-Cyano-7-(4-nitrophenyl)-2-(4-sulfamoylphenylamino)-7H-thiazolo [4,5-b]pyran-5-ylamino)-4-oxobutanoic acid (XVII)
A mixture of compound XIa (4.7 g, 0.01 mol) and succinic anhydride (1.0 g, 0.01 mol) in ethanol (50 mL) was refluxed for 3h. The reaction mixture was poured onto iced water and filtered. The solid obtained was crystallized from dioxane to give XVII.
Table 21, Physico-chemical and analytical data for compound XVII.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XVII 201-203 63 C23H16N6O7S2 50.00 2.92 15.21 (570.06) 50.14 2.89 15.11 Table 22, Spectral data for compound XVII:
No. Type Data IR 3460 (OH), 3400, 3320, 3200 (NH, NH2), 3080 (KBr, cm -1) (CH arom.), 2900, 2916 (CH aliph), 2203 (C≡N), 1710, 1685 (2C=O), 1620 (C=N), 1329, 1158(SO2). 1 H-NMR δ: 1.2, 1.6 (2t, 4H, CH2- CH2), 4.0 (s, 1H, CH XVII (DMSO-d6) pyrane), 7.2-8.0 (m, 10 H, Ar-H+ SO2NH2), 8.2 (s, ppm 2H, 2NH exchangeable with D2O), 10.5 (s, 1H, OH exchangeable with D2O). EI/MS(m/z) 570 [M+] (2.5), 63(100). (relative abundance) Experimental| 92
4-(6-Cyano-5-(2,5-dioxopyrrolidinyl)-7-(4-nitrophenyl)-7H-thiazolo[4,5- b]pyran-2-ylamino) benzenesulfonamide (XVIII)
A mixture of compound XIa (4.7 g, 0.01 mol) and succinic anhydride (1.0 g, 0.01 mol) was fused at 220 ºC for 15 min. Ethanol was added to the reaction mixture while hot and filtered. The solid obtained was crystallized from dioxane to give XVIII.
Table 23, Physico-chemical and analytical data for compound XVIII.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XVIII >300 61 C23H16N6O7S2 50.00 2.92 15.21 (552.04) 50.12 2.75 15.38 Table 24, Spectral data for compound XVIII:
No. Type Data IR 3480, 3290, 3200 (NH, NH2), 3065 (CH arom.), (KBr, cm -1) 2925, 2912 (CH aliph.), 2200 (C≡N), 1700, 1680 (2C=O), 1610(C=N), 1544, 1340 (NO2), 1329, 1158 (SO2). 1 XVIII H-NMR δ: 1.5, 1.6 (2t, 4H, 2CH2), 4.1 (s, 1H, CH pyrane), (DMSO-d6) 7.2-8.0 (m, 10H, Ar-H + SO2NH2 exchangeable ppm with D2O), 8.2 (s, 1H, NH exchangeable with D2O). EI/MS(m/z) 552 [M+] (4.09), 92(100). (relative abundance) Experimental| 93
N-(6-cyano-7-(4-nitrophenyl)-2-(4-sulfamoylphenylamino)-7H- thiazolo[4,5-b]pyran-5-yl)-3-oxobutanamide (XIX)
A solution of compound XIa (4.7 g, 0.01 mol) in ethyl acetoacetate (10 mL) was fused at 220 ºC for 1 h. The fused product was dissolved in ethanol then poured onto cold water. The precipitated solid was filtered and crystallized from dioxane to give XIX.
Table 25, Physico-chemical and analytical data for compound XIX.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XIX 121-123 53 C23H18N6O7S2 49.81 3.27 15.15 (554.5) 49.76 3.33 15.32 Table 26, Spectral data for compound XIX:
No. Type Data IR 3443, 3362, 3237 (NH, NH2), 3050 (CH arom.) (KBr, cm -1) 2939, 2923 (CH aliph), 2195 (C ≡N), 1717, 1710 (C=O), 1627 (C=N), 1530, 1312 (NO2), 1346, 1160 (SO2). 1 XIX H-NMR δ: 1.6 (s, 3H, COCH3), 2.2 (s, 2H, CH2), 4.0 (s, 1H, (DMSO-d6) CH pyrane), 7.2-8.0 (m, 10H, Ar-H+ SO2NH2), 8.2 ppm (s, 2H, 2NH exchangeable with D2O). EI/MS(m/z) 554 [M+] (7.28), 129(100). (relative abundance) Experimental| 94
8-Imino, 6-thioxo, 9-(3,4-dimethoxy phenyl) 9-H,1N phenyl,[2-(4- sulfamoyl phenyl amino)] thiazolo[4,5-b ]pyrano [2,3-d] pyrimidine (XX)
A mixture of compound XIb (4.85g, 0.01 mol) with phenyl isothiocyanate (1.35 g, 0.01 mol) in ethanol or in pyridine (50 ml) was refluxed for 5 h. The reaction mixture was poured onto iced water. The solid obtained was crystallized from dioxane to give XX.
Table 27, Physico-chemical and analytical data for compound XX.
Compd. M.P.(°C) Yield Mol. Formula Analysis No. % (Mol.Wt.) Calculated/Found C% H% N% XX 244-246 45 C28H24N6O5S3 54.18 3.90 13.54 (622.11) 54.25 3.87 13.48 Table 28, Spectral data for compound XX:
No. Type Data IR 3337, 3255, 3123 (NH, NH2), 3050 (CH arom.), (KBr, cm -1) 2968, 2929 (CH aliph.), 1618 (C=N), 1380, 1130 (SO2), 1270 (C=S). 1 XX H-NMR δ : 4.0 (s, 6H, 2 OCH3), 4.1 (s, 1H, CH pyrane), 7.0- (DMSO-d6) 7.9 (m, 14H, Ar-H+ SO2NH2), 8.2 (s, 2H, 2NH ppm exchangeable with D2O), 11.6 (s, 1H, NH imino) . EI/MS(m/z) 620 [M+-2] (10), 271(100). (relative abundance)
BIOLOGICAL ACTIVITY
Biological activity| 96
5. Biological Activity
5. 1. In vitro anticancer activity
The in vitro cytotoxic activity was measured for twenty seven new compound and doxorubicin on liver human tumor cell line (HEPG2) using the Sulfo-Rhodamine-B stain (SRB) assay. The in vitro anticancer screening was done by the pharmacology unit at the National Cancer Institute, Cairo University.
5.1.1. Introduction
The sulfo-rhodamine B (SRB) assay,122 which was developed in 1990, remains one of the most widely used methods for in vitro cytotoxic screening. The assay relies on the ability of SRB to bind to protein components of cells that have been fixed to tissue-culture plates by trichloroacetic acid (TCA). SRB is a bright-pink aminoxanthene dye with two sulfonic groups that bind to basic amino-acid residues under mild acidic conditions, and dissociate under basic conditions. The amount of dye extracted from stained cells is directly proportional to the cell mass.122-124 The strong intensity of SRB staining allows the assay to be carried out in a 96-well format. Skehen et al.122 showed that the assay can detect densities as low as 1,000-2,000 cells per well. Furthermore, the SRB method has proven to be practical, because after the TCA-fixed and SRB-stained cell monolayers are dried they can be stored indefinitely. Color extracted from SRB-stained cells is also stable. With its high level of sensitivity, adaptability to the 96-well format and endpoint stability, the SRB assay is well suited to large-scale screening applications, as well as research. This Biological activity| 97 assay has been widely used for drug-toxicity testing against different types of cancerous and non-cancerous cell lines. 122-124 The application of the SRB assay is limited to the manual or semiautomatic screening due to the multiple washing and drying steps, which, at present, are not amenable to automation. This method nevertheless provides an efficient and sensitive tool for screening, especially for use in less well-equipped laboratories.125 5.1.2. Procedure
- Cells were plated in 96-multiwell plate (104 cells/ well) for 24 hrs before treatment with the compounds to allow attachment of cell to the wall of the plate. - Test compounds were dissolved in dimethylsulfoxide (DMSO) and diluted with saline to the appropriate volume. - Different concentrations of the compounds under test (10, 25, 50 and 100μM) were added to the cell monolayer. Triplicate wells were prepared for each individual dose. - Monolayer cells were incubated with the compounds for 48 hrs at 0 37 C and in atmosphere of 5% CO2. - After 48 hrs, cells were fixed, washed and stained for 30 min. with 0.4% (wt/ vol) SRB dissolved in 1% acetic acid. - Unbounded dye was removed by four washes with 1% acetic acid, and attached stain was recovered with Tris-EDTA buffer. - Color intensity was measured in an enzyme linked immunosorbent assay (ELISA) reader. Biological activity| 98
- The relation between surviving fraction and drug concentration is plotted to get the survival curve of each tumor cell line after the specified time.
- The concentration required for 50% inhibition of cell viability (IC50) was calculated and compared with the reference drug Doxorubicin and the results are given in table 29. 5.1.3. Results
The results are shown in Table 29 are compared with the reference drug doxorubicin (IC50= 32 µM). The most potent compounds are the pyrimidine analogue XIVb (IC50= 30 µM), the pyrimidine derivative with phenyl ring substitution XX (IC50= 31 µM) then the thiazolopyrane derivative XVII with oxobutanoic acid substitution and XIIIb which also contain a pyrimidine ring and are equipotent (IC50=32 μM) followed by compound XIIIa (IC50= 36 µM).
1.2 1.2
1 1 0.8 0.8 0.6 IC50=43 uM 0.6 IC50=52 uM 0.4 0.4
0.2 0.2
Surviving Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (8): Survival curve for HEPG2 cell Figure (9): Survival curve for HEPG2 cell line for compound III line for compound IVa
Biological activity| 99
1.2 1.2 1 1 0.8 0.8 IC50=43 uM 0.6 0.6 IC50=89 0.4 0.4
0.2 0.2 Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM)
Figure (10): Survival curve for HEPG2 cell Figure (11): Survival curve for HEPG2 cell
line for compound IVb line for compound VI
1.2 1.2 1 1 0.8 0.8 0.6 IC50=52 uM 0.6 IC50=50 uM 0.4 0.4
0.2 0.2 Surviving Surviving Fraction 0 Surviving Fraction 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM)
Figure (12): Survival curve for HEPG2 cell Figure (13): Survival curve for HEPG2 cell line for compound VII line for compound IXa
1.2
1.2 1 1 0.8 0.8 0.6 IC50=46 uM 0.6 IC50=52 uM 0.4 0.4
0.2 0.2
Surviving Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (14): Survival curve for HEPG2 cell Figure (15): Survival curve for HEPG2 cell line for compound IXb line for compound IXc
Biological activity| 100
1.2 1.2 1 1 0.8 0.8 IC50=63 uM 0.6 0.6 IC50=52 uM 0.4 0.4
0.2 0.2 Surviving Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (16): Survival curve for HEPG2 cell Figure (17): Survival curve for HEPG2 cell line for compound IXd line for compound IXe
1.2
1.2 1 1 0.8 0.8 IC50=51 uM 0.6 0.6 IC50=47 uM 0.4 0.4
0.2 0.2 Surviving Surviving Fraction 0 Fraction Surviving 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (18): Survival curve for HEPG2 cell Figure (19): Survival curve for HEPG2 cell line for compound IXf line for compound IXg
1.2
1.2 1 1 IC50=65 uM 0.8 0.8 IC50=60 uM 0.6 0.6 0.4 0.4
0.2 0.2
Surviving Surviving Fraction Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (20): Survival curve for HEPG2 cell Figure (21): Survival curve for HEPG2 cell line for compound IXh line for compound XIa
Biological activity| 101
1.2 1.2
1 1 0.8 0.8 0.6 0.6 0.4 0.4 0.2
0.2 Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (22): Survival curve for HEPG2 cell Figure (23): Survival curve for HEPG2 cell line for compound XIb line for compound XIc
1.2 1.2 1 1 0.8 0.8 IC50=72 uM IC50=65 uM 0.6 0.6 0.4 0.4
0.2 0.2 Surviving Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (24): Survival curve for HEPG2 cell Figure (25): Survival curve for HEPG2 cell line for compound XId line for compound XII
1.2 1.2
1 1 0.8 0.8 0.6 IC50=36 uM 0.6 IC50=32 uM 0.4 0.4
0.2 0.2 Surviving Surviving Fraction
0 Surviving Fraction 0 0 50 100 150 -0.2 0 50 100 150 Concentration (uM) Concentration (uM)
Figure (26): Survival curve for HEPG2 cell Figure (27): Survival curve for HEPG2 cell line for compound XIIIa line for compound XIIIb
Biological activity| 102
1.2 1.2
1 1 0.8 0.8 0.6 0.6 IC50= 30 uM IC50=45 uM 0.4 0.4 0.2 0.2 Surviving Surviving Fraction 0 Surviving Surviving Fraction 0 0 50 100 150 -0.2 0 50 100 150 Concentration (uM) Concentration (uM)
Figure (28): Survival curve for HEPG2 cell Figure (29): Survival curve for HEPG2 cell line for compound XIVa line for compound XIVb
1.2 1.2
1 1 0.8 0.8 IC50=45 uM 0.6 0.6 IC50=39 uM 0.4 0.4
0.2 0.2
Surviving Surviving Fraction Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (30): Survival curve for HEPG2 cell Figure (31): Survival curve for HEPG2 cell line for compound XV line for compound XVI
1.2 1.2
1 1 0.8 0.8 IC50=32 uM 0.6 0.6 IC50=45 uM 0.4 0.4 0.2
Surviving Surviving Fraction 0.2 Surviving Surviving Fraction 0 0 0 50 100 150 0 50 100 150 Concentration (uM) Concentration (uM) Figure (32): Survival curve for HEPG2 cell Figure (33): Survival curve for HEPG2 cell line for compound XVII line for compound XVIII
Biological activity| 103
1.2 1.2
1 1 0.8 0.8 IC50= 31 uM 0.6 IC50=37 uM 0.6 0.4 0.4 0.2
Surviving Surviving Fraction 0.2 0 0 50 100 150 Surviving Fraction 0 0 50 100 150 Concentration (uM) -0.2 Concentration (uM) Figure (34): Survival curve for HEPG2 cell Figure (35): Survival curve for HEPG2 cell line for compound XIX line for compound XX
Table 29: In vitro anticancer screening of the synthesized compounds against human liver cell line (HEPG2).
Compound concentration (µM) Cpd. No. 10 (µM) 25 (µM) 50 (µM) 100 (µM) IC 50 (µM) Surviving fraction (mean ± SE) a DOX. 0.551 ± 0.026 0.480 ± 0.003 0.139 ± 0.005 0.130 ± 0.016 32 III 0.848 ± 0.012 0.501 ± 0.025 0.168 ± 0.008 0.231 ± 0.022 43 IVa 0.849 ± 0.022 0.619 ± 0.049 0.317 ± 0.013 0.271 ± 0.015 52 IVb 0.687 ± 0.020 0.451 ± 0.019 0.274 ±0.013 0.289 ± 0.026 43 VI 0.942 ± 0.035 0.627 ± 0.016 0.407 ± 0.030 0.606 ± 0.005 89 VII 0.850 ± 0.008 0.581 ± 0.045 0.320 ± 0.023 0.239 ± 0.006 52 IXa 0.978 ± 0.040 0.553 ± 0.004 0.271 ± 0.017 0.244 ± 0.010 50 IXb 0.796 ± 0.032 0.562 ± 0.014 0.264 ± 0.016 0.213 ± 0.006 46 IXc 0.835 ± 0.015 0.653 ± 0.006 0.309 ± 0.010 0.258 ± 0.006 52 IXd 0.824 ± 0.023 0.757 ± 0.020 0.402 ± 0.018 0.355 ± 0.015 63 IXe 0.862 ± 0.027 0.625 ± 0.032 0.333 ± 0.020 0.249 ± 0.009 52 IXf 0.873 ± 0.009 0.651 ± 0.006 0.338 ± 0.018 0.197 ± 0.011 51 IXg 0.808 ± 0.026 0.523 ± 0.098 0.275 ±0.003 0.244 ± 0.034 47 IXh 0.823± 0.023 0.542± 0.033 0.377 ± 0.010 0.463 ± 0.015 65 XIa 0.846 ± 0.010 0.767 ± 0.016 0.376 ± 0.005 0.312 ± 0.007 60 XIb 0.628 ± 0.049 0.564 ± 0.061 0.640 ± 0.017 0.672 ± 0.018 - XIc 0.909 ± 0.022 0.861 ± 0.036 0.582 ± 0.015 0.655 ± 0.061 - XId 0.846 ± 0.015 0.564 ± 0.046 0.388 ± 0.010 0.509 ± 0.066 72 XII 0.814 ± 0.029 0.605 ± 0.030 0.287 ± 0.022 0.489± 0.020 65 XIIIa 0.625 ± 0.005 0.498 ± 0.025 0.168 ± 0.007 0.015 ± 0.015 36 XIIIb 0.561 ± 0.008 0.446 ± 0.035 0.164 ± 0.019 0.127 ± 0.020 32 XIVa 0.764 ± 0.005 0.498 ± 0.062 0.301 ± 0.008 0.213 ± 0.017 45 XIVb 0.529 ± 0.018 0.477 ± 0.036 0.118 ± 0.018 0.115 ± 0.08 30 XV 0.779 ± 0.027 0.487 ± 0.004 0.244 ± 0.005 0.262 ± 0.014 45 XVI 0.615 ± 0.045 0.432 ± 0.006 0.284 ± 0.018 0.246 ± 0.014 39 Biological activity| 104
XVII 0.626 ± 0.007 0.382 ± 0.031 0.127 ± 0.015 0.166 ± 0.005 32 XVIII 0.710 ± 0.006 0.474 ± 0.033 0.342 ± 0.018 0.231 ± 0.004 45 XIX 0.546 ± 0.012 0.456 ± 0.015 0.206 ± 0.006 0.299 ± 0.018 37 XX 0.593 ± 0.019 0.434 ± 0.012 0.128 ± 0.006 0.137 ± 0.036 31
a Each value is the mean of three experiments ± standard error.
5.2. Radiosensitizing evaluation
This study was conducted to evaluate the ability of the most active compounds to enhance the cell killing effect of γ-radiation.
5.2.1. Procedure
Irradiation was performed at the National Center for Radiation Research and Technology (Atomic Energy Authority), using Gamma cell-40 (137Cs) source. Seven compounds III, IVb, IXb, XIIIa, XIVb, XVII and XX were selected to be evaluated again for their in vitro anticancer activity in combination with γ-irradiation.
Cells were plated in 96-multiwell plate (104 cells/ well) for 24 hours before γ-irradiation with a single dose of 8 Gy. Cells were incubated for 48 h at 37 ˚C in atmosphere of 5% CO2. After 48 h, cells were fixed, washed and stained with 0.4% (wt/vol) SRB dissolved in 1% acetic acid, for 30 minutes. Excess unbound dye was removed by four washes with 1% acetic acid and attached stain was recovered with Tris–EDTA buffer. Color intensity was measured in an ELISA reader at a wave length of 570 nm. Biological activity| 105
In another multiwell plate, cells were incubated with the previously mentioned selected compounds; III, IVb, IXb, XIIIa, XIVb, XVII and XX in molar concentrations of 10, 25, 50, 100 µM. After 2 hours, cells were subjected to a single dose of γ-radiation at a dose level of 8 Gy with a dose rate of 2 Gy/min. After 48 h, the surviving fractions were measured using the above mentioned procedures by ELISA reader.
The surviving fractions were expressed as means ± standard error. The results were analyzed using 1-way ANOVA test and given in Table 30.
5.2.2. Results
From the results obtained in Table 29, compound XIVb showed an in vitro cytotoxic activity with IC50 value of 30 µM, when the cells were subjected to different concentrations of the compound alone. While when the cells were subjected to the same concentrations of compound XIVb, and irradiated with a single dose of γ-radiation at a dose level of 8 Gy, as shown in Table 30, the IC50 value was synergistically decreased to 8 µM (Figure
40). Similarly, compounds III, IVb, IXb, XIIIa, XVII and XX showed IC50 values of 43 µM, 43 µM, 46 µM 36 µM, 32 µM and 31 µM, respectively, when used alone (Table 29). The IC50 values were decreased to 30 µM, 30 µM, 32µM 18 µM, 15µM and 17 µM, when the cells were treated with compounds, in combination with γ-radiation (Figures 36-42) (Table 30).
Table 30: In vitro anticancer screening of compounds III, IVb, IXb, XIIIa, XIVb, XVII and XX against human liver cell line (HEPG2) in combination with γ-radiation.
Irradiated Compound concentration (µM) + irradiation (8 Gy) IC 50 Cpd. Control a (8 Gy) Surviving fraction (mean ± SE) (µM) Biological activity| 106
No. 10 25 50 100 0.53 ± 0.48± 0.12 ± 0.12± III 1.000 0.927 ±0.02 30 0.08* 0.01* 0.01* 0.01* 0.52 ± 0.47± 0.11 ± 0.13 ± IVb 1.000 0.927 ±0.02 30 0.01* 0.02* 0.01* 0.01* 0.56 ± 0.45 ± 0.16± 0.13 ± IXb 1.000 0.927 ±0.02 32 0.02* 0.01* 0.01* 0.01* 0.41 ± 0.22 ± 0.12 ± 0.14 ± XIIIa 1.000 0.927 ±0.02 18 0.01* 0.02* 0.05* 0.06* 0.31± 0.12 ± 0.10 ± 0.11 ± XIVb 1.000 0.927 ±0.02 8 0.07* 0.01* 0.01* 0.08* 0.36 ± 0.23 ± 0.11 ± 0.13 ± XVII 1.000 0.927 ±0.02 15 0.07* 0.01* 0.01* 0.08* 0.40± 0.21 ± 0.12 ± 0.13± XX 1.000 0.927 ±0.02 17 0.01* 0.02* 0.05* 0.06* a: Each value is the mean of three values ± Standard Error *: Significant difference from control group at p<0.001
1.2
1
CpdIII 0.8 Cpd III+ 8 Gy 0.6 Linear (CpdIII) 0.4 Linear (Cpd III+ 8 Gy)
Surviving Fraction Surviving 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (36): Survival curve for HEPG2 cell line for compound III alone and in combination with γ-irradiation (8 Gy) Biological activity| 107
1.2
1 Cpd IVb Cpd IVb + 8 Gy 0.8 Linear (Cpd IVb) 0.6 Linear (Cpd IVb + 8 Gy) 0.4
Surviving Surviving Fraction 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (37): Survival curve for HEPG2 cell line for compound IVb alone and in combination with γ-irradiation (8 Gy)
1.2
1 Cpd IXb Cpd IXb+ 8 Gy 0.8 Linear (Cpd IXb) 0.6 Linear (Cpd IXb+ 8 Gy) 0.4
Surviving Surviving Fraction 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (38): Survival curve for HEPG2 cell line for compound IXb alone and in combination with γ-irradiation (8 Gy) Biological activity| 108
1.2
1 Cpd XIIIa Cpd XIIIa + 8 Gy
0.8 Linear (Cpd XIIIa) 0.6 Linear (Cpd XIIIa + 8 Gy)
0.4
Surviving Surviving Fraction 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (39): Survival curve for HEPG2 cell line for compound XIIIa alone and in combination with γ-irradiation (8 Gy)
1.2
1 Cpd XIVb
0.8 Cpd XIVb + 8 Gy
0.6 Linear (Cpd XIVb) Linear (Cpd XIVb + 8 Gy) 0.4
Surviving Surviving Fraction 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (40): Survival curve for HEPG2 cell line for compound XIVb alone and in combination with γ-irradiation (8 Gy)
Biological activity| 109
1.2
1 Cpd XVII
0.8 Cpd XVII+ 8 Gy Linear (Cpd XVII) 0.6 Linear (Cpd XVII+ 8 Gy) 0.4
Surviving Surviving Fraction 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (41): Survival curve for HEPG2 cell line for compound XVII alone and in combination with γ-irradiation (8 Gy)
1.2
1 Cpd XX Cpd XX + 8 Gy 0.8 Linear (Cpd XX) 0.6 Linear (Cpd XX + 8 Gy)
0.4
Surviving Surviving Fraction 0.2
0 0 20 40 60 80 100 120 -0.2 Concentration
Figure (42): Survival curve for HEPG2 cell line for compound XX alone and in combination with γ-irradiation (8 Gy)
MOLECULAR MODELING
Molecular Modeling| 111
6. Molecular Modeling
Previous literature shows that carbonic anhydrase inhibition is one of the anticancer mechanisms of sulfonamides, and this is clearly reported by Abbate et al.,126 who stated that the potent anticancer sulfonamide drug (E7070) 3, currently undergoing clinical developments for the treatment of several types of cancer, also acts as a strong carbonic anhydrase inhibitor, and this may contribute at least in part, to its in vivo efficacy.
The X-ray crystal structure of the adduct of hCA II with (E7070) 3 revealed unique interactions between the inhibitor and the active site, with the compound showing three different conformation in the active site of the enzyme. A superimposition of two of these conformations with those of other sulfonamide CA inhibitors indicated that similar regions of the hCA II active site are involved in the interaction with such inhibitors (Figure 43). So these data may be used for the design of novel CA inhibitors with antitumor activity.126
Figure (43): Superimposition of hCA II-inhibitor adducts: (magenta), (green), (yellow), (dark blue, two conformations of E7070). Molecular Modeling| 112
As reported by Supuran et al.26 and Pastorekova et al.32 which is confirmed from the X-ray crystallographic structure for many adducts of sulfonamide inhibitors with isozymes CA I, II and IV, that the sulfonamides which are CA inhibitors share the following: i. They bind to the Zn(II) ion in a tetrahedral geometry which is a stable geometry for the metal ion. ii. In all these adducts, the nitrogen of the sulfonamide is coordinated to the Zn(II) ion of the enzyme. iii. The NH moiety participates in a hydrogen bond with Thr199. iv. One of the oxygen atoms of the SO2NH2 moiety also participates in a hydrogen bond with the backbone NH moiety of Thr199 (Figure 44).
Figure (44): CA inhibition mechanism by sulfonamides.
Since the newly synthesized compounds are sulfonamide derivatives and also, comply with the general pharmacophore of the carbonic anhydrase Molecular Modeling| 113 inhibitors, it would be interesting to perform docking studies on the synthesized compounds to hCA II and to compare their docking interactions with previously reported interactions of (E7070) 3.
Docking procedures
All molecular modeling calculations and docking studies were performed 127 using “Molecular Operating Environment (MOE) version 2007.09”. The ligands were drawn on ChemDraw and imported in MOE. The structures were subjected to energy minimization using Merck Molecular Force Field 94x (MMFF94x) and the partial charges were computed using the same forcefield.
The X-ray crystallographic structure of hCA II complexed with N- (2,3,4,5,6- pentafluorobenzyl)-4-sulfamoyl-benzamide (1G54) was obtained from the Protein Data Bank and its two-dimensions interaction map is shown in Figure 45.
The enzyme was prepared for the docking studies where: i. The ligand molecule was removed from the enzyme active site. ii. Hydrogen atoms were added to the structure with their standard geometry. iii. Partial charges were computed using Amber99 forcefield.
Molecular Modeling| 114
Figure (45): Interaction map of N-(2,3,4,5,6-pentafluorobenzyl)-4 sulfamoylbenzamide with hCA II.
Docking calculations were done using Alpha triangle placement method and poses were prioritized based on affinity dG scoring method.
In order to validate our docking procedure, E7070 –with known reported interactions- was docked into the active site of hCA II. The docking results clearly showed that indeed, the compound exhibits similar interactions as those previously reported in the literature and stated above (Figure 46). Molecular Modeling| 115
Figure (46): Interaction map of (E7070) 3 with the active site of hCA II, binding score= -9.34
Results
Docking results indicate that all the newly synthesized compounds exhibited similar interactions to that previously reported for E7070. Figures 47-51 are the interaction maps of compounds IVb, XIVb, XVII and XX, in which they all share the same requirements including: i. They all bind to the Zn(II) ion in a tetrahedral geometry, a stable geometry for the metal ion. ii. In all instances, the nitrogen of the sulfonamide moiety is coordinated to the Zn(II) ion of the enzyme. iii. Thr199 participates in a hydrogen bond with the NH moiety and with one of the oxygen atoms of the sulfonamide. Interestingly, the remaining parts of the compounds showed a variety of interactions with the amino acids of the binding site. Molecular Modeling| 116
Figure (47): 2D and 3D Interaction maps of compound IVb with the active site of hCA II, binding score= -9.93
Figure (48): 2D and 3D Interaction maps of compound XIVb with the active site of hCAII, binding score= -9.78
Molecular Modeling| 117
Figure (49): 2D and 3D Interaction maps of compound XVII with the active site of hCA II, binding score= -10.33
Figure (50): 2D and 3D Interaction maps of compound XX with the active site of hCA II, binding score= -10.35
Molecular Modeling| 118
In order to give a clearer comparison of the docked poses of E7070 and the synthesized compounds with free sulfonamide moiety, a three- dimensional superimposition was done for the docked poses of compound XIVb which is the most potent compound and E7070 (Figure 51).
The superimposition shows that the benzenesulfonamide moieties bound to hCA II overlap each other completely, while the tails adopt slightly different conformations.
Figure (51): Superimposition of E7070 (red) and compound XIVb (yellow) in the active site of hCA II shows that the benzenesulfonamide moieties bound to hCA II overlap each other completely while the tails adopt different conformations. A comparison was done between the above three dimensional interaction map and that of compound XIc which is the least potent compound and Molecular Modeling| 119
E7070, in which the two compounds do not completely overlap at the benzenesulfonamide moieties (Figure 52).
Figure (52): Superimposition of E7070 (red) and compound XIc (green) in the active site of hCA II shows that the benzenesulfonamide moieties bound to hCA II do not completely overlap.
Conclusion
Finally, it can be seen from our docking study that the synthesized compounds having a free sulfonamide moiety exhibit similar conformations and binding interactions with hCA II similar to those previously reported for other sulfonamide compounds that act as CA inhibitors. This suggests that these compounds might possibly act as CA inhibitors, and this may contribute at least in part, to their anticancer activity.
REFERENCES
References| 121
7. References
1. Kumar P.; Clark M., In Clinical medicine . WB Saunders , London. 2005; p 485-500.
2. Milas, L.; Cox, J.D., “Principles of combining radiation therapy and chemotherapy”, in Radiation oncology, rationale, technique, results. Cox, J.D.; Ang, K.K., 2003, Mosby, St. Louis, p. 108–124.
3. Tannock, I.F., “Treatment of cancer with radiation and drugs”. J. Clin. Oncol. 1996, 14, 3156–3174.
4. Vokes, E.E.; Weichselbaum, R.R., “Concomitant chemoradiotherapy: rationale and clinical experience in patients with solid tumors”. J. Clin. Oncol. 1990, 8, 911–934.
5. Bartelink, H.; Kallman, R.F.; Rapacchietta, D.; Hart, G.A., “Therapeutic enhancement in mice by clinically relevant dose and fractionation schedules of cis-diamminedichloroplatinum (II) and irradiation”. Radiother. Oncol. 1986, 6, 61–74.
6. Al-Sarraf, M.; Martz, K.; Herskovic, A.; Leichman, L.; Brindle, J.S.; Vaitkevicius, V.K.; Cooper, J.; Byhardt, R.; Davis, L.; Emami, B., “Progress report of combined chemoradiotherapy versus radiotherapy alone in patients with esophageal cancer: an Intergroup study”. J. Clin. Oncol. Res. 1997, 15, 277–284.
7. Furuse, K.; Fukuoka, M.; Kawahara, M.; Nishikawa, H.; Takada, Y.; Kudoh, S.; Katagami, N.; Ariyoshi, Y., “Phase III study of concurrent versus sequential thoracic radiotherapy in combination with mitomycin, vindesine, and cisplatin in unresectable stage III non-small-cell lung caner”. J. Clin. Oncol. 1999, 17, 2692–2699.
8. Green, J.A.; Kirwan, J.M.; Tierney, J.F.; Symonds, P.; Fresco, L.; Collingwood, M.; Williams, C.J., “Survival and recurrence after concomitant chemotherapy and radiotherapy for cancer of the uterine cervix: a systematic review and metaanalysis”. Lancet. 2001, 358, 781– 786.
References| 122
9. Komaki, R., “Managemant of limited small-cell lung cancer”. Int. J. Clin. Oncol. 2000, 5, 205–216.
10. Pignon, J.P.; Bourhis, J.; Domenge C.; Designe, L., “Chemotherapy added to locoregional treatment for head and neck squamous-cell carcinoma: three meta-analyses of updated individual data”. Lancet. 2000, 355, 949– 955.
11. Gregoire, V.; Hunter, N.; Brock, W.A.; Hittelman, W.N.; Plunkett, W.; Milas, L., “Improvement in the therapeutic ratio of radiotherapy for a murine sarcoma by indomethacin plus fludarabine”. Radiat. Res.1996, 146, 548– 553.
12. Nishimura, Y.; Ono, K.; Tsutsui, K.; Oya, N.; Okajima, K.; Hiraoka, M.; Abe, M., “Esophageal cancer treated with radiotherapy: impact of total treatment time and fractionation”. Int. J. Radiat. Oncol. Biol. Phys. 1994, 30, 1099–1105.
13. Nishimura, Y.; Nagata, Y.; Okajima, K.; Mitsumori, M.; Hiraoka, M.; Masunaga, S.; Ono, K.; Shoji, K.; Kojima, H., “Radiation therapy for T1,2 glottic carcinoma: impact of overall treatment time on local control”. Radiother. Oncol. 1996, 40, 225–232.
14. Withers, H.R.; Taylor, J.M.G.; Maciejewski, B., “The hazard of accelerated tumor clonogen repopulation during radiotherapy”. Acta. Oncol. 1988, 27, 131–146.
15. Ang, K.K.; Andratschke, N.H.; Milas, L., “Epidermal growth factor receptor and response of head-and-neck carcinoma to therapy”. Int. J. Radiat. Oncol. Biol. Phys. 2004, 58, 959–965.
16. Milas, L.; Fan, Z.; Andratschke, N.H.; Ang, K.K., “Epidermal growth factor receptor and tumor response to radiation: in vivo preclinical studies”. Int. J. Radiat. Oncol. Biol. Phys. 2004, 58, 966–971.
17. Fuwa, N.; Kano, M.; Toita, T.; Shikama, N.; Kodaira, T.; Matsumoto, A.; Kamata, M.; Furutani, K.; Tatibana, H., “Alternating chemoradiotherapy for nasopharyngeal cancer using cisplatin and 5-fluorouracil: a preliminary report of phase II study”. Radiother. Oncol. 2001, 61, 257–260.
References| 123
18. Drews, J., "Drug discovery: a historical perspective". Science 2000, 287, 1960-1964.
19. Supuran, C.T.; Scozzafava, A., “Carbonic anhydrase inhibitors”. Curr. Med. Chem. 2001, 1, 61-97.
20. Maren, T.H., “Relations between structure and biological activity of sulfonamides”. Annu. Rev. Pharmacol. Toxicol. 1976, 16, 309-327.
21. Boyd, A.E. “Sulfonylurea receptors, ion channels, and fruit flies”. Diabetes 1988, 37, 847-850.
22. Thornber, C.W., “Isosterism and molecular modification in drug design”. Chem. Soc. Rev. 1979, 8, 563-580.
23. a) Casini, A.; Scozzafava, A.; Mastrolorenzo, A.; Supuran, C.T., “Sulfonamides and sulfonylated derivatives as anticancer agents”. Curr. Cancer Drug Targets 2002, 2, 55-75. b) Kim, Y.H.; Shin, K.J.; Lee, T.G.; Kim. E.; Lee, M.S.; Ryu, S.H.; Suh, P.G., “ G2 arrest and apoptosis by 2-amino-N-quinoline-8-yl- benzenesulfonamide (QBS), a novel cytotoxic compound”. Biochem. Pharmacol. 2005, 69, 1333-1341.
24. Hilvo, M.; Tolvanen, M.; Clark, A.; Shen, B.; Shah, G.N.; Waheed, A.; Halmi, P.; Hanninen, M.; Hamalainen, J.M.; Vihinen, M.; Sly, W.S.; Parkkila, S., “Characterization of CA XV, a new GPI-anchored form of carbonic anhydrase”. Biochem. J. 2005, 392, 83-92.
25. Scozzafava, A.; Owa, T.; Mastrolorenzo, A.; Supuran, C.T., “Anticancer and antiviral sulfonamides”. Curr. Med. Chem. 2003, 10, 925-953.
26. Supuran, C.T.; Scozzafava, A., “Carbonic anhydrases as targets for medicinal chemistry”. Bioorg. Med. Chem. 2007, 15, 4336-4350.
27. Supuran, C.T.; Scozzafava, A., “Carbonic anhydrase inhibitors and their therapeutic potential”. Expert Opin. Ther. Pat. 2000, 10, 575-600.
28. Chegwidden, W.R.; Dodgson, S.J.; Spencer, I.M., “The roles of carbonic anhydrase in metabolism, cell growth and cancer in animals”. Exs. 2000, 90, 343-363. References| 124
29. Pastorekova, S.; Parkkila, S.; Parkkila, A.K.; Opavsky, R.; Zelnik, V.; Saarnio, J.; Pastorek, J., “Carbonic anhydrase IX, MN/CA IX: analysis of stomach complementary DNA sequence and expression in human and rat alimentary tracts”. Gastroenterology 1997, 112, 398-408.
30. Maren, T.H., “Carbonic anhydrase: chemistry, physiology, and inhibition”. Physiol. Rev. 1967, 47, 595-781.
31. Supuran, C.T.; Scozzafava, A., “Indisulam”. IDrugs 2002, 5, 1075-1079.
32. Pastorekova, S.; Parkkila, S.; Pastorek, J.; Supuran, C.T., “Carbonic anhydrases: current state of the art, therapeutic applications and future prospects”. J. Enzyme Inhib. Med. Chem. 2004, 19, 199-229.
33. Chegwidden, W.R.; Spencer, I.M., “Sulphonamide inhibitors of carbonic anhydrase inhibit the growth of human lymphoma cells in culture”. Inflammopharmacology 1995, 3, 231-239.
34. Teicher, B.A.; Liu, S.D.; Liu, J.T.; Holden, S.A.; Herman, T.S., “A carbonic anhydrase inhibitor as a potential modulator of cancer therapies”. Anticancer Res. 1993, 13, 1549-1556.
35. Supuran, C.T.; Scozzafava, A., “Carbonic anhydrase inhibitors: aromatic sulfonamides and disulfonamides act as efficient tumor growth inhibitors”. J. Enzyme Inhib. 2000, 15, 597-610.
36. Supuran, C.T.; Scozzafava, A., “Carbonic anhydrase inhibitors - Part 94. 1,3,4-thiadiazole-2-sulfonamidederivatives as antitumor agents”. Eur. J. Med. Chem. 2000, 35, 867-874.
37. Vullo, D.; Franchi, M.; Gallori, E.; Pastorek, J.; Scozzafava, A.; Pastorekova, S.; Supuran, C. T., “Carbonic anhydrase inhibitors: inhibition of the tumor-associated isozyme IX with aromatic and heterocyclic sulfonamides”. Bioorg. Med. Chem. Lett. 2003, 13, 1005-1009.
38. Casey, J.R.; Morgan, P.E.; Vullo, D.; Scozzafava, A.; Mastrolorenzo, A.; Supuran, C.T., “Carbonic anhydrase inhibitors. Design of selective, membrane-impermeant inhibitors targeting the human tumor-associated isozyme IX”. J. Med. Chem. 2004, 47, 2337-2347.
References| 125
39. Thiry, A.; Ledecq, M.; Cecchi, A.; Dogne, J.M.; Wouters, J.; Supuran, C.T.; Masereel, B., “Indanesulfonamides as carbonic anhydrase inhibitors. Toward structure-based design of selective inhibitors of the tumorassociated isozyme CA IX”. J. Med. Chem. 2006, 49, 2743-2749.
40. Patrick, S.C.; Peter, M.G., “Cancer and Cancer Chemotherapy, in Foye's principles of medicinal chemistry”, Thomas, L.L.; David, A.W., 2002, Eds. Lippincott Williams & Wilkins: Philadelphia, fifth ed. p. 928-947.
41. Owa, T.; Yoshino, H.; Okauchi, T.; Yoshimatsu, K.; Ozawa, Y.; Sugi, N.H.; Nagasu, T.; Koyanagi, N.; Kitoh, K., “Discovery of novel antitumor sulfonamides targeting G1 phase of the cell cycle”. J. Med. Chem. 1999, 42, 3789-3799.
42. Fukuoka, K.; Usuda, J.; Iwamoto, Y.; Fukumoto, H.; Nakamura, T.; Yoneda, T.; Narita, N.; Saijo, N.; Nishio, K., “Mechanisms of action of the novel sulfonamide anticancer agent E7070 on cell cycle progression in human non-small cell lung cancer cells”. Invest. New Drugs 2001, 19, 219- 227.
43. Yoshimatsu, K.; Yamaguchi, A.; Yoshino, H.; Koyanagi, N.; Kitoh, K., “Mechanism of action of E7010, an orally active sulfonamide antitumor agent: inhibition of mitosis by binding to the colchicine site of tubulin”. Cancer Res. 1997, 57, 3208-3213.
44. Chang, J.Y.; Hsieh, H.P.; Chang, C.Y.; Hsu, K.S.; Chiang, Y.F.; Chen, C.M.; Kuo, C.C.; Liou, J.P., “7-Aroyl-aminoindoline-1-sulfonamides as a novel class of potent antitubulin agents”. J. Med. Chem. 2006, 49, 6656- 6659.
45. Hu, L.; Li, Z.R.; Li, Y.; Qu, J.; Ling, Y.H.; Jiang, J.D.; Boykin, D.W., “Synthesis and structure-activity relationships of carbazole sulfonamidesas a novel class of antimitotic agents against solid tumors”. J. Med. Chem. 2006, 49, 6273-6282.
46. Supuran, C.T.; Scozzafava, A., “Matrix Metalloproteinase Inhibitors”, in Proteinase and Peptidase Inhibition: Recent Potential Targets for Drug Development, Smith, H.J.; Simons, C.E., 2001, Francis & Taylor: London. p. 35-52. References| 126
47. Wossner, J.F.; Nagase, H., “Matrix Metalloproteinases and TIMPs”. Oxford University Press, Oxford, 2000, 245-253.
48. Scozzafava, A.; Supuran, C.T., “Carbonic anhydrase and matrix metalloproteinase inhibitors: sulfonylated amino acid hydroxamates with MMP inhibitory properties act as efficient inhibitors of CA isozymes I, II, and IV, and N-hydroxysulfonamides inhibit both these zinc enzymes”. J. Med. Chem. 2000, 43, 3677-3687.
49. Supuran, C.T.; Scozzafava, A.; Mastrolorenzo, A., “Bacterial proteases: Current therapeutic use and future prospects for the development of new antibiotics”. Expert Opin. Ther. Pat. 2001, 11, 221-259.
50. MacPherson, L.J.; Bayburt, E.K.; Capparelli, M.P.; Carroll, B.J.; Goldstein, R.; Justice, M.R.; Zhu, L.; Hu, S.; Melton, R.A.; Fryer, L.; Goldberg, R.L.; Doughty, J.R.; Spirito, S.; Blancuzzi, V.; Wilson, D.; O'Byrne, E.M.; Ganu, V.; Parker, D.T., “Discovery of CGS 27023A, a non- peptidic, potent, and orally active stromelysin inhibitor that blocks cartilage degradation in rabbits”. J. Med. Chem. 1997, 40, 2525-2532.
51. Whittaker, M.; Floyd, C.D.; Brown, P.; Gearing, A.J., “Design and therapeutic application of matrix metalloproteinase inhibitors”. Chem. Rev. 1999, 99, 2735-2776.
52. Clare, B.W.; Scozzafava, A.; Supuran, C.T., “Protease inhibitors: synthesis of a series of bacterial collagenase inhibitors of the sulfonyl amino acyl hydroxamate type”. J. Med. Chem. 2001, 44, 2253-2258.
53. Lesyk, R.; Zimenkovsky, B., “4-Thiazolidones: centenarian history, current status and perspectives for modern organic and medicinal chemistry”, Curr. Org. Chem.2004, 8, 1547-1577.
54. Lesyk, R.; Vladzimirska, O.; Holota, S.; Zaprutko, L.; Gzella, A., “New 5-substituted thiazolo[2,3-d][1,2,4]triazol-6-ones. Synthesis and anticancer evaluation”, Eur. J. Med. Chem. 2007, 42, 641-648.
55. Havrylyuk, D.; Zimenkovsky, B.; Vasylenko, O.; Zaprutko, L.; Gzella, A.; Lesyk, R., “Synthesis of novel thiazolone-based compounds containing References| 127 pyrazoline moiety and evaluation of their anticancer activity”, Eur. J. Med. Chem. 2009, 44, 1396-1404.
56. Havrylyuk, D.; Zimenkovsky, B.; Lesyk, R., “Synthesis and anticancer activity of novel nonfused bicyclic thiazolidinone derivatives”, Phosphorus, Sulfur, and Silicon. 2009, 184, 638-650.
57. Kaminskyy, D.; Zimenkovsky, B.; Lesyk, R., “Synthesis and in vitro anticancer activity of 2,4-azolidinedione-acetic acids derivatives”, Eur. J. Med. Chem. 2009,44, 3627-3636.
58. Carter, P.H.; Scherle, P.A.; Muckelbauer, J.A.; Voss, M.E.; Liu, R.Q.; Thompson, L.A.; Tebben, A.J.; Solomon, K.A.; Lo, Y.C.; Li, Z.; Strzemienski, P.; Yang, G.; Falahatpisheh, N.; Xu, M.; Wu, Z.; Farrow, N.A.; Ramnarayan, K.; Wang, J.; Rideout, D.; Yalamoori, V.; Domaille, P.; Underwood, D.J.; Trzaskos, J.M.; Friedman, S.M.; Newton, R.C.; Decicco, C.P.; “Photochemically enhanced binding of small molecules to the tumor necrosis factor receptor-1 inhibits the binding of TNF-α”, Proc. Natl. Acad. Sci. U.S.A. 2001, 98 (21), 11879-11884.
59. Franchetti, P.; Grifantini, M., “Potent and selective inhibitors of human immunodeficiency virus protease structurally related to L-964, 746”, Curr. Med. Chem. 1999, 6, 599-614.
60. Matsuya, Y.; Kawaguchi, T.; Ishihara, K.; Ahmed, K.; Zhao, Q.L.; Kondo, T.; Nemoto, H.; “Synthesis of macrosphelides with a thiazole side chain, new antitumor candidates having apoptosis inducing property”, Org. Lett. 2006, 8, 4609– 4612.
61. Srivastava, P.C.; Pickering, M.V.; Allen, L.B.; Streeter, D.G.; Campbell, M.T.; Witkowski, J.T.; Sidwell, R.W.; Robins, R.K.,”Synthesis and antiviral activity of certainthiazole C- nucleosides”, J. Med. Chem. 1977, 20, 256- 262.
62. Franchetti, P.; Cappellacci, L.; Marchetti, S.; Trincavelli, T.; Martini, C.; Mazzoni, M.R.; Lucacchini, A.; Grifantini, M., "2’-C-Methyl analogues of selective adenosine receptor agonists: synthesis and binding studies”, J. Med. Chem. 1998, 41, 1708–1715.
References| 128
63. Robins, R.K.; Srivastava, P.C.; Narayanan, V.L.; Plowman, L.; Paull, K. D.; “2-Beta- D-ribofuranosyl thiazole-4-carboxamide, a novel potential antitumor agent for lung tumors and metastases”, J. Med. Chem. 1982, 25, 107-108.
64. Weber, G.; Shen, F.; Orban, T. I.; Kokeny, S.; Olah, E., "Targeting signal transduction”, Adv. Enzyme Regul. 2003, 43, 47-56.
65. Tricot, G.J.; Jayaram, H.N.; Heerema, N.; Weber, G.; Hoffman, R. “Biochemically directed therapy of leukemia with tiazofurin, a selective blocker of inosine 5’-phosphate dehydrogenase activity ”, Cancer Res. 1989, 49, 3696-3701.
66. Wright, D.G.; Boosalis, M.S.; Waraska, K.; Oshry, L.J.; Weintraub, L. R.; Vosburgh, E., “Tiazofurin effects on IMP-dehydrogenase activity and expression in the leukemia cells of patients with CML blast crisis”, Anticancer Res. 1996, 16, 3349-3354.
67. Merino, P.; Tejero, T.; Unzurrunzaga, F.J.; Franco, S.; Chiacchio, U.; Saita, M.G.; Iannazzo, D.; Pipernoc, A.; Romeo, G., "An efficient approach to enantiomeric isoxazolidinyl analogues of tiazofurin based on nitrone cycloadditions ", Tetrahedron: Asym. 2005, 16, 3865-3876.
68. Popsavin, M.; Spaic, S.; Svircev, M.; Kojic, V.; Bogdanovicb G.; Popsavina, V., “2-(3-Amino-3-deoxy-b-D-xylofuranosyl) thiazole 4- carboxamide: A new tiazofurin analogue with potent antitumour activity”, Bioorg. Med. Chem. Lett. 2006, 16, 5317–5320.
69. Popsavin, M.; Spaic, S.; Svircev, M.; Kojic, V.; Bogdanovicb G.; Popsavina, V., “Synthesis and antitumour activity of new tiazofurin analogues bearing a 2,3-anhydro functionality in the furanose ring”, Bioorg. Med. Chem. Lett. 2007, 17, 4123–4127.
70. Schiff, P.B.; Fant, J.; Horwitz, S. B., "Promotion of microtubule assembly invitro by taxol”, Nature 1979, 22, 665-667.
71. Wilson, L.; Jordan, M.A., “Microtubule dynamics: taking aim at a moving target”, Chem. Biol. 1995, 2,569-642.
References| 129
72. Ashkenazi, S.; Cleary, T.G., “A method for detecting Shiga toxin and Shiga-like toxin-1 in pure and mixed culture”, J. Med. Microbiol. 1990, 32, 255–261.
73. Sun, X.M.; MacFarlane, M.; Zhuang, J.; Wolf, B.B.; Green, D.R.; Cohen, G.M., “Distinct caspase cascade are initiated receptor-mediated and chemical-induced apoptosis”, J. Biol. Chem. 1999, 274, 5053–5060.
74. Bollag, D.M.; McQueney, P.A. ;Zhu, J.; Hensens, O.; Koupal, L.; Liesch, J.; Goetz, M., Lazaides, E.; Woods, C.M., "Epothilones, a new class of microtubule-stabilizing agents with a taxol like mechanism of action”, Cancer Res. 1995, 55, 2325-2333.
75. Ahmed, K.; Matsuya,Y.; Nemoto, H.; Faisal, S.; Zaidi, H.; Sugiyama, T.; Yoshihisa, Y.; Shimizu, T.; Kondo, T., “Mechanism of apoptosis induced by a newly synthesized derivative of macrosphelides with a thiazole side chain”, Chem. Biol. Inter. 2009,177, 218–226.
76. Matsuya, Y.; Kawaguchi, T.; Ishihara, K.; Ahmed, K.; Zhao, Q.L.; Kondo, T.; Nemoto, H., “Synthesis of macrosphelides with a thiazole side chain, new antitumor candidates having apoptosis inducing property”, Org. Lett. 2006, 8, 4609– 4612.
77. Tamura, Y.; Watanabe, F.; Nakatani, T.; Yasui, K.; Fuji, M.; Komurasaki, T.; Tsuzuki, H.; Maekawa, R.; Yoshioka, T.; Kawada, K.; Sugita, K.; Ohtani, M., “Highly selective and orally active inhibitors of type IV collagenase (MMP-9 and MMP-2): N-sulfonyl amino acid derivatives”, J. Med. Chem. 1998, 41, 640-649.
78. Kiyama, R.; Tamura, Y.; Watanabe, F.; Tsuzuki, H.; Ohtani, M.; Yodo, M., “Homology modeling of gelatinase catalytic domains and docking simulations of novel sulfonamide inhibitors”, J. Med. Chem. 1999, 42, 1723- 1738.
79. Sternlicht, M.; Werb, Z., “How matrix metalloproteinases regulate cell behavior”, Annu. Rev. Cell. Dev. Biol. 2001, 17, 463-516.
80. Chang, C.; Werb, Z., “The many faces of metalloproteases: cell growth, invasion, angiogenesis and metastasis”, Trends Cell. Biol. 2001, 11, 537- 543. References| 130
81. Vihinen, P.; Kahari, V., “Involvement of matrix metalloproteinase type-3 in hepatocyte growth factor–induced invasion of human hepatocellular carcinoma cells”, Int. J. Cancer. 2002, 99, 157-162. 82. Tosetti, F.; Ferrari, N.; DeFlora, S.; Albini, A., "Angioprevention: angiogenesis is a common and key target for cancer chemopreventive agents”, FASEB J. 2002, 16, 2-14.
83. Visse, R.; Nagase, H., "Matrix metalloprotinases and tissue inhibitors of metalloproteinases”, Circ. Res. 2003, 92, 827-839.
84. a) Watanabe, F.; Tamura, Y., “Sulfonamide derivative selectively inhibiting MMP-13”, (Shionogi & Co., Ltd, Japan) Patent WO 02/28844 Chem. Abstr. 2002, 136, 355251. b) Kamel, M.M.; Ali, H.I.; Anwar, M.M.; Mohamed, N.A., “Synthesis, antitumor activity and molecular docking study of novel sulfonamide- schiff’s bases, thiazolidinones, benzothiazinones and their C- nucleoside derivatives”, Eur. J. Med. Chem. 2010, 45, 572-580.
85. Suda, H.; Matsunaga, K.; Yamamura, S.; Shizuri, Y., “Structure of a new topoisomerase II inhibitor BE 10988", Tetrahedron. Lett.1991, 32, 2791- 2792.
86. Burden, D.A.; Osheroff, N., "Mechanism of action of eukaryotic topoisomerase II and drugs targeted to the enzyme", Biochim. Biophys. Acta 1998, 1400, 139-154.
87. Clarke, M.J.; Zhu, F.; Frasca, D.R., "Non-platinum chemotherapeutic metallopharmaceuticals”, Chem. Rev. 1999, 99, 2511-2534.
88. Clarke, M.J., "Ruthenium metallopharmaceuticals”, Chem. Rev. 2002, 232, 69-93.
89. Kostova, I., “Ruthenium complexes as anticancer agents” Curr. Med. Chem. 2006, 13, 1085-1107.
90. Krzysztof W.P., "Novel Nicotinamide Adenine Dinucleotide Analogues as Potential Anticancer Agents: Quest for Specific Inhibition of Inosine Monophosphate Dehydrogenase”, Pharmacol. Ther. 1997, 76, 89-100.
References| 131
91. Folkman, “Angiogensis” J. Annu. Rev. Med. 2006, 57, 1-18.
92. Kim, I.; Kim, H.G.; Moon, S.O.; Chac, S.W.; So, J.N.; Koh, K.N.; Ahn, B.C.; Koh, G. Y., “Angiopoietin-1 induces endothelial cell sprouting through the activation of focal adhesion kinase and plasmin secretion”, Circ. Res. 2000, 86, 952-959.
93. Thurston, G., “Role of angiopoietins and Tie receptor tyrosine kinases in angiogensis and lymphangiogensis”, Cell Tissue Res. 2003, 314, 61-68.
94. Chong, W.M.; Chu, S.; Duvadie, R.K.; Li, L.; Na, J.; Schaffer, L.; Yang, Y., PCT Int. Appl.2004, WO2004072070.
95. Chu, X.; Ding, Q.; Jiang, N.; Kim, K.; Lovey, A.J.; McComas, W.M.; Mullin, J.G., Tilley, J.W., “Diaminothiazoles”, PCT Int. Appl. 2003, WO2003097048.
96. a) Bowler, A.N.; Olesen, P.H.; Sorensen, A.R.; Hansen, B.F.; Worsaae, H.; Kurtzhals, P., "Use of 2, 4-diaminothiazole derivatives”, PCT Int. Appl. 2001, WO2001056567. b) Abdel razik, H.H., “Synthesis of new thiazolo[2,3-b]pyrimidine derivatives of pharmaceutical interest”, Chem. Mater. Sci. 2006, 60, 268- 273.
97. Lv, P.C.; Wang, K.R.; Yang, Y.; Mao, W.J.; Chen, J.; Xiong, J.; Zhu, H.L., "Design, synthesis and biological evaluation of novel thiazole derivatives as potent FabH inhibitors", Bioorg. Med. Chem. Lett. 2009, 19, 6750–6754.
98. Thomae, D.; Perspicace, E.; Xu, Z.; Henryon, D.; Schneider, S.; Hesse, S.; Kirsch, G.; Seck, P., "One-pot synthesis of new 2,4,5-trisubstituted 1,3- thiazoles and 1,3-selenazoles", Tetrahedron 2009, 65, 2982–2988.
99. Jayaprakash, S.; Pattenden, G.; Viljoen, M.S.; Wilson, G., "Synthesis of novel proline–thiazole based cyclic hexa- and octapeptides", Tetrahedron 2003, 59, 6637–6646.
100. Qiao, Q.; Dominique, R.; Goodnow, R., "2,4-Disubstituted-5- acetoxythiazoles:useful intermediates forthe synthesis of thiazolones and 2,4,5-trisubstituted thiazoles", Tetrahedron Lett. 2008, 49, 3682–3686. References| 132
101. Das, B.; Reddy, V.S.; Ramu, R., "A rapid and high-yielding synthesis of thiazoles and aminothiazoles using ammonium-12-molybdophosphate", J. Mol. Cataly. A: Chem. 2006, 252, 235–237.
102. Xi, N.; Bo, Y.; Doherty, E.M.; Fotsch, C.; Gavva, N.R.; Han, N.; Hungate, R.W., Klionsky, L.; Liu, Q.; Tamir, R.; Xu, S.; Treanorb, J.S.; Norman, M.H., "Synthesis and evaluation of thiazole carboxamides as vanilloid receptor 1 (TRPV1) antagonists", Bioorg. Med. Chem. Lett. 2005, 15, 5211–5217.
103. Renuga, S.; Gnanadeebam, M.; Vinosha, B.M.; Perumal, S.; "A novel four-component tandem protocol for the stereoselective synthesis of highly functionalised thiazoles", Tetrahedron 2007, 63, 10054–10058.
104. Luthra, P.M.; Mishra, C.B.; Jha, P.K.; Barodia, S.K.; "Synthesis of novel 7-imino-2-thioxo-3,7-dihydro-2H-thiazolo [4,5-d] pyrimidine derivatives as adenosine A2A receptor antagonists", Bioorg. Med. Chem. Lett. 2010, 20, 1214–1218.
105. Vicini, P.; Geronikaki, A.; Anastasia, K.; Incerti, M.; Zani, F., "Synthesis and antimicrobial activity of novel 2-thiazolylimino-5-arylidene- 4-thiazolidinones", Bioorg. Med. Chem. 2006, 14, 3859-3864.
106. Vazzana, I.; Terranova, E.; Mattioli, F.; Sparatore, F., "Aromatic sciff bases and 2,3-disubstituted -1,3-Thiazolidin-4-one derivatives as anti- inflammatory agents", ARKIVOC 2004, 5, 364-374.
107. Bondock, S.; Khalifa, W.; Fadda, A.A., "Synthesis and antimicrobial evaluation of some new thiazole, thiazolidinone and thiazoline derivatives starting from 1-chloro-3,4-dihydronaphthalene-2-carboxaldehyde", Eur. J. Med. Chem. 2007, 42, 948-954.
108. Azam, F.; Alkskas, I.A.; Ahmed, M.A., "Synthesis of some urea and thiourea derivatives of 3-phenyl/ethyl-2-thioxo-2,3-dihydrothiazolo[4,5- d]pyrimidine and their antagonistic effects on haloperidol-induced catalepsy and oxidative stress in mice", Eur. J. Med. Chem. 2009, 44, 3889–3897
References| 133
109. Tozkoparan, B.; Ertan, M.; Kelicen, P.; Demirdamar, R., "Synthesis and anti-inflammatory activities of some thiazolo[3,2-a]pyrimidine derivatives", Il Farmaco 1999, 54, 588–593.
110. AL-Hazimi, H.M.; Al-Alshaikh, M.A., "Microwave assisted synthesis of substituted furan-2-carboxaldehydes and their reactions", J. Saudi. Chem. Soc. 2010, 14, 373-382.
111. Supuran, C.T.; Scozzafava, A.; Casini, A., “Carbonic anhydrase inhibitors”. Med. Res. Rev. 2003, 23, 146-189.
112. Nishimura, Y., “Rationale for chemoradiotherapy”. Int. J. Clin. Oncol. 2004, 9, 414–420.
113. Donkor, I.O.; Abdel-Ghany, Y.S.; Kador, P.F.; Mizoguchi, T.; Malik, E.A.; Miller, D.D.,"Synthesis and biological activities of aldose reductase inhibitors bearing acyl benzenesulfonamides as carboxylic acid surrogates", Eur. J. Med. Chem. 1998, 33, 15-22.
114. Siavosh, M.; Andreas, S.; Heymo, H.; Emerich, E.; Thomas, B.; Mathias, S.; Thomas, M.; Josef, S.F.; Thomas, B.L.," [4-(Imidazol-1-yl)- thiazol-2-yl]phenylamines – A novel class of highly potent colchicine site binding tubulin polymerization inhibitors: Synthesis and cytotoxic activity on some human cancer cell lines", J. Med. Chem. 2006, 49, 5769- 5776.
115. Amr, A.E.; Mohamed, A.M.; Mohamed, S.F.; Abdel-Hafez, N.A.; Hammam, A.G., "Anticancer activities of some newly synthesized pyridine, pyrane, and pyrimidine derivatives", Bioorg. Med. Chem. 2006, 14, 5481– 5488
116. Casini, A.; Scozzafava, A.; Mincione, F.; Menabuoni, L.; Ilies, M.A.; Supuran, C.T., "Carbonic anhydrase inhibitors: water-soluble 4- sulfamoylphenylthioureas as topical intraocular pressure-lowering agents with long lasting effect", J.Med. Chem. 2000, 43, 4884-4892.
117. Supuran, C.T.; Nicolae, A.; Popescu, A., "Carbonic anhydrase inhibitors, Synthesis of Schiff bases derived from sulfanilamide and aromatic aldehydes: the first inhibitors with equally high affinity towards References| 134 cytosolic and membrane-bound isoenzymes", Eur. J. Med. Chem. 1996, 31,431-438.
118. Ghorab, M.M.; Ragab, F.A.; Noaman, E.; Heiba, H.I.; El-Hossary, E.M., "Utility of 4-(5,5-dimethyl-3-oxo-cyclohex-1-enylamino)- benzenesulfonamide in the synthesis of novel quinolines as possible anticancer and radioprotective agents”. Arzneim. Forsch. 2008, 58, 35-41.
119. Ghorab, M.M.; Noaman, E.; Ismail, M.F.; Heiba, H.I.; Ammar, Y.A.; Sayed, M.Y., “Novel antitumor and radioprotective sulfonamides containing pyrrolo[2,3-d]pyrimidines”. Arzneim. Forsch. 2006, 56, 405- 413.
120. Ismail, M.F.; Ghorab, M.M.; Noaman, E.; Ammar, Y.A.; Heiba, H.I.; Sayed, M.Y., “Novel synthesis of pyrrolo[2,3-d]pyrimidines bearing sulfonamide moieties as potential antitumor and radioprotective agents”. Arzneim. Forsch. 2006, 56, 301-308.
121. Abdel-Gawad, S.M.; El-Gaby, M.A.; Heiba, H.I.; Aly, H.M.; Ghorab, M.M., “Synthesis and radiation stability of some new biologically active hydroquinoline and pyrimido[4,5-b]quinoline derivatives”. J. Chin. Chem. Soc. 2005, 52, 1227-1236.
122. Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; McMahon, J.; Vistica, D.; Warren, J. T.; Bokesch, H.; Kenney, S.; Boyd, M. R., "New colorimetric cytotoxicity assay for anticancer-drug screening". J. Natl. Cancer Inst. 1990, 82, (13), 1107-1112.
123. McCaffrey, T. A.; Agarwal, L. A.; Weksler, B. B., "A rapid fluorometric DNA assay for the measurement of cell density and proliferation in vitro". In Vitro Cell Dev. Biol. 1988, 24, 247-252.
124. Monks, A.; Scudiero, D.; Skehan, P.; Shoemaker, R.; Paull, K.; Vistica, D.; Hose, C.; Langley, J.; Cronise, P.; Vaigro-Wolff, A., "Feasibility of a high-flux anticancer drug screen using a diverse panel of cultured human tumor cell lines". J. Natl. Cancer Inst. 1991, 83, 757- 766.
125. Keepers, Y. P.; Pizao, P. E.; Peters, G. J.; van Ark-Otte, J.; Winograd, B.; Pinedo, H. M., "Comparison of the sulforhodamine B protein and tetrazolium (MTT) assays for in vitro chemosensitivity testing". Eur. J. Cancer 1991, 27, 897-900. References| 135
126. Abbate, F.; Casini, A.; Owa, T.; Scozzafava, A.; Supuran, C.T., “Carbonic anhydrase inhibitors: E7070, a sulfonamide anticancer agent, potently inhibits cytosolic isozymes I and II, and transmembrane, tumor- associated isozyme IX”. Bioorg. Med. Chem. Lett. 2004, 14, 217-223.
127. Chemical Computing Group Inc., S.S.W., Suite 910, Montreal, H3A 2R7, Canada. Molecular Operating Environment (MOE), 2007.09.
ARABIC SUMMARY
ب | الولخص العربى
الولخص العربً
ٚٓذف ْزا انجحش إنٗ رشٛٛذ يشكجبد عذٚذح راد فبعهٛخ يعبدح نهغشغبٌ ,رى رحعٛش ثعط يشزمبد انضٛبصٔل, انضٛبصٔنٕثٛشاٌ, انضٛبصٔنٕثٛشيٛذٍٚ ٔ انضٛبصٔنٕثٛشإَثٛشًٚذٍٚ انغذٚذح ٔانزٙ رحزٕ٘ عهٙ يغًٕعخ انغهفَٕبيٛذ . رى إخزجبس فبعهٛخ ْزِ انًشكجبد عهٗ خالٚب األٔساو انغشغبَٛخ األديٛخ نهكجذ ٔ رى دساعخ انزأصٛش انزحفٛضٖ نهزشعٛع انغبيٗ كًب رى أٚعب إسعبء ْزح انًشكجبد فٙ انًٕلع انُشػ إلَضٚى انكشثَٕٛك أَٛٓذساص II.
ٔ رحزٕٖ انشعبنخ عهٗ األعضاء انزبنٛخ:
الوقذهت
ٚحزٕٖ ْزا انغضء عهٗ عشد يخزصش عٍ انغشغبٌ ٔ األعظ انًغزخذيخ فٙ عالعّ ٔ انعالط ثبإلشعبع ثبإلظبفخ انٙ أعجبة اعزخذاو انعالط انضُبئٙ ثبنكًٛٛبء ٔ اإلشعبع يعب, ْزا ثغبَت يُبلشخ أًْٛخ كم يٍ يشزمبد انغهفَٕبيٛذ, انضٛبصٔل , انضٛبصٔنٕثٛشيٛذٍٚ ٔانجٛشإَ صٛبصٔنٕثٛشًٛٚذٍٚ انغذٚذح كًعبداد نهغشغبٌ. ثبإلظبفخ انٙ اعزعشاض ثعط انطشق انًغزخذيخ نزحعٛش يشزمبد انضٛبصٔل, انضٛبصٔنٕثٛشيٛذٍٚ ٔانضٛبصٔنٕثٛشإَثٛشًٚذٍٚ. الغرض هي البحث
ٚحزٕ٘ ْزا انغضء عهٙ األعظ انجٕٛنٕعّٛ انزٙ ألزشػ عهٙ أعبعٓب رحعٛش ْزح انًشكجبد انغذٚذح ٔيحبٔنخ رٕلع آنٛخ عًم ْزح انًشكجبد عٍ غشٚك اإلسعبء انغضٚئٙ. هٌبقشت الجسء العولً
ُٚبلش ْزا انغضء انطشق انًغزخذيخ فٙ رشٛٛذ انًشكجبد انغذٚذح ٔانطشق انًزجعّ فٙ رحعٛشْب ٔكزنك يُبلشخ غشق انزأكذ يٍ انزشكٛت انجُبئٙ انكًٛٛبئٙ نٓب.
الجسء العولً
ٚزعًٍ ْزا انغضء ٔصفب رفصٛهٛب نهطشق انًعًهّٛ انزٙ أرجعذ نزحعٛش انًشكجبد انغذٚذح ٔرى انزحمك يٍ انزشكٛت انجُبئٙ نٓزح انًشكجبد ٔرنك عٍ غشٚك انزحبنٛم انذلٛمخ نهعُبصش ٔ األشعخ رحذ انحًشاء ٔ انشٍَٛ انُٕٔ٘ انًغُبغٛغٙ ٔكزنك يطٛبف انكزهخ.
رزعًٍ انشعبنخ رحعٛش انًشكجبد األرٛخ:
٢- كهٕسٔ-N-)٤- عهفبيٕٚم فُٛٛم( اعٛزبيٛذ )II(. ٤- )٤-اوكسو-٥,٤- داٛٓٚذسٔ صٛبصٔل -٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )III(. ت | الولخص العربى
٤- )٧- فُٛٛم -٥-ثيوكسو- ٧,٦,٥,٤ – تيتراهيدروثيازول [ ٥,٤- د] ثٛشيٛذٍٚ - ٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )IVa(. ٤- )٥-ثيوكسو- ٧- p - توليل- ٧,٦,٥,٤ – تيتراهيدروثيازول [ ٥,٤- د] ثٛشيٛذٍٚ - ٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )IVb(. ٤- أيزوثيوسيانيتو ثُضٍٚ عهفَٕبيٛذ )V(. اٚضٛم ٢- عٛبَٕ- ۳- )٤- عهفبيٕٚم فُٛٛم ايُٕٛ(-۳- صٕٛكغٕثشٔثبَٕاد )VI(. اٚضٛم ٢-)٤- اكغٕ-٥,٤- داٚٓبٚذسٔ صٛبصٔل -٢-ٚم( -۳- )٤- عهفبيٕٚم فُٛٛم ايُٕٛ( -۳- صٕٛكغٕثشٔثبَٕاد )VII(. ٤- )٤- يٛضٛم ثُضٚهٛذٍٚ ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )VIIIa(. ٤- )٤- َٛزشٔ ثُضٚهٛذٍٚ ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )VIIIb(. ٤- )٤-اكغٕ -٢- p- رٕنٛم صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXa(. ٤- )٢-)٤- َٛزشٔفُٛٛم( -٤ - أكغٕ صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXb(. ٤- )٤-اكغٕ -٢-فينيل صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXc(. ٤- )٢-)٢- ْٛذسٔكغٙ فُٛٛم( -٤ - أكغٕ صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXd(. ٤- )٢-)٤- ْٛذسٔكغٙ فُٛٛم( -٤ - أكغٕ صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXe(. ٤-)ثُضٔ [د] [۱, ۳] داٚكغٕل-٥- ٚم( -٤ - أكغٕ صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ .)IXf( ٤- )٢-)٢- كهٕسٔ فُٛٛم( -٤ - أكغٕ صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXg(. ٤- )٢-)۳- ثشٔيٕ فُٛٛم( -٤ - أكغٕ صٛبصٔنٛذٚ -۳-ٍٚم( ثُضٍٚ عهفَٕبيٛذ )IXh(. ٤- )٥-أيُٕٛ-٦- عٛبَٕ-٧-)٤- َٛزشٔفُٛٛم(-7H – صٛبصٔنٕ[5،4- ب] بيران-٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )XIa(. ٤- )٥-أيُٕٛ-٦- عٛبَٕ-٧-)٤,۳- دا٘ يٛضٕكغٙ فُٛٛم(-7H - صٛبصٔنٕ[5،4- ب] بيران - ٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )XIb(. ٤- )٥-أيُٕٛ -٧- )٢- كهٕسٔفُٛٛم( -٦- عٛبَٕ-7H - صٛبصٔنٕ[5،4- ب] بيران -٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )XIc(. ٤- )٥-أيُٕٛ -٧- )۳- ثشٔيٕفُٛٛم( -٦- عٛبَٕ-7H - صٛبصٔنٕ[5،4- ب] بيران -٢- ٚم ايُٕٛ( ثُضٍٚ عهفَٕبيٛذ )XId(. ۸- أمينو,9-)٤- نيترو فينيل( H-9, [٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XII(. ۸- أكسو,9-)٤- نيترو فينيل( H-9,[٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين (XIIIa) ۸- أكسو,9-)٤,۳- دا٘ يٛضٕكغٙ فُٛٛم( H-9 ,[٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XIIIb(. ث | الولخص العربى
6-ميثيل, ۸- أكسو,9- )٤- نيترو فينيل( H-9,[٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XIVa(. 6-ميثيل, ۸- أكسو,9 -)٤,۳- دا٘ يٛضٕكغٙ فُٛٛم( H-9,[٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XIVb(. ۸- أمينو,6- أكسو,9-)٤- نيترو فينيل( H-9,[٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XV(. ۸- أمينو,6- ثايوكسو,9-)٤- نيترو فينيل( H-9 ,[٢- )4- سلفامويل فُٛٛم أيُٕٛ( ] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XVI(. ٤- )٦-سيانو-٧-)٤- نيترو فينيل(-٢- )٤- سلفامويل فُٛٛم أيُٕٛ(- ٧-H- صٛبصٔنٕ[5،4- ب ] بيران -٥- ٚم أيُٕٛ(-٤- أكغٕثٕٛربَٕٚك أعٛذ )XVII(. ٤-)٦-سيانو-٥- )٥,٢- داي أكسوبيروليدينيل (-٧- )٤-نيتروفينيل( ٧-H- صٛبصٔنٕ[-5،4 ب] بيران -٢- ٚم أيُٕٛ( ثُضٍٚ عهفَٕبيٛذ)XVIII(. N - )٦-سيانو-٧-)٤- نيترو فينيل(-٢- )٤- سلفامويل فُٛٛم أيُٕٛ(- ٧-H- صٛبصٔنٕ[-5،4 ب] بيران -٥- ٚم(-۳- أكغٕثٕٛربٌ أيٛذ)XIX(. ۸- أيمينو,6-ثيوكسو,9 -)٤,۳- دا٘ يٛضٕكغٙ فُٛٛم( N1,H-9 فينيل [٢- )4- سلفامويل فُٛٛم أيُٕٛ(] صٛبصٔنٕ[٥,٤-ة] بيرانو[۳,٢- د] بيريميدين )XX(.
اإلختببراث البٍولوجٍت
رى إخزجبسانًشكجبد انغذٚذح كًعبداد نهغشغبٌ عهٙ خالٚب األٔساو انغشغبَّٛ األديّٛ نهكجذ )HEPG2) انًًُبِ فٙ ٔعػ خبسعٙ ثبإلظبفخ انٙ اخزجبس ٧ يشكجبد نهكشف عٍ صٚبدح حغبعٛخ انخالٚب انغشغبَٛخ رغبِ انزأصٛش انزحفٛض٘ انًحزًم نهزشعٛع انغبيٙ ٔرى عشض انُزبئظ ٔ يُبلشزٓب.
اإلرسبء الجسٌئً
ٚشزًم ْزا انغضء عهٙ إسعبء انًشكجبد انغذٚذح فٙ يٕلع اإلسرجبغ إلَضٚى انكشثَٕٛك أَٛٓذساص II إلعزٛعبػ إيكبَٛخ اعزخذاو ْزح انًشكجبد كًعبداد إلَضٚى انكشثَٕٛك أَٛٓذساص ٔثبنزبنٙ إيكبَٛخ اعزخذايٓب كًعبداد نهغشغبٌ.
الوراجع
ٚحزٕ٘ ْزا انغضء عهٙ 721 يشععب عهًٛب.
تصوٍن بواسطت الحبسوة و تشٍٍذ بعط هشتقبث الثٍبزول الجذٌذة التً تحتوي علً هجووعت السلفوًبهٍذ ودراست التأثٍر التحفٍسي الوتوقع كوضبداث لألورام السرطبًٍت هع التشعٍع الجبهً
سعبنخ يمذيخ يٍ أٌتي هحوود هحوذ سلٍوبى
ثكبنٕسٕٚط انعهٕو انصٛذنٛخ كهٛخ انصٛذنخ- عبيعخ عٍٛ شًظ ٢٠٠٦
نهحصٕل عهٙ دسعخ انًبعغزٛش فٙ انعهٕو انصٛذنٛخ )كٍوٍبء صٍذلٍت(
أ. د/ دالل عبد الرحمن أبو العال أ. د/ مصطفي محمد سليمان غراب
أعزبر انكًٛٛبء انصٛذنٛخ أعزبر انكًٛٛبء انععٕٚخ انزطجٛمٛخ سئٛظ لغى انكًٛٛبء انصٛذنٛخ لغى انجحٕس انذٔائٛخ االشعبعٛخ كهٛخ انصٛذنخ انًشكض انمٕيٙ نجحٕس ٔ ركُٕنٕعٛب األشعبع عبيعخ عٍٛ شًظ ْٛئخ انطبلخ انزسٚخ
أ.د/ حلوً إسوبعٍل هٍبت
أعزبر يغبعذ انكًٛٛبء انععٕٚخ انزطجٛمٛخ لغى انجحٕس انذٔائٛخ االشعبعٛخ انًشكض انمٕيٙ نجحٕس ٔ ركُٕنٕعٛب األشعبع ْٛئخ انطبلخ انزسٚخ
كلية الصيدلة جبهعت عٍي شوس ۲١٠٠