ABSTRACT
Novel 4-aryl-1-oxa-9-thiacyclopenta[b]fluorenes were designed, synthesized, and evaluated as inhibitors of the protein tyrosine phosphatase, PTP1B. Compounds 3 (IC50 = 284 nM) and 4 (IC50 = 74 nM), showed nanomolar potency against PTP1B (TRDI(P)YETD(P)Y(P)YRK as substrate). Compound 4 also lowered insulin in the diabetic ob/ob mouse at a dose of 10 mg/kg/day, p.o.
Subject(s)
Benzofurans/chemical synthesis , Enzyme Inhibitors/chemical synthesis , Protein Tyrosine Phosphatases/antagonists & inhibitors , Thiophenes/chemical synthesis , Amino Acid Sequence , Animals , Benzofurans/chemistry , Benzofurans/pharmacology , Binding Sites , Drug Design , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Indicators and Reagents , Mice , Models, Molecular , Molecular Conformation , Oligopeptides/chemistry , Oligopeptides/metabolism , Protein Tyrosine Phosphatase, Non-Receptor Type 1 , Protein Tyrosine Phosphatases/chemistry , Structure-Activity Relationship , Substrate Specificity , Thiophenes/chemistry , Thiophenes/pharmacologyABSTRACT
Insulin resistance in the liver and peripheral tissues together with a pancreatic cell defect are the common causes of type 2 diabetes. It is now appreciated that insulin resistance can result from a defect in the insulin receptor signaling system, at a site post binding of insulin to its receptor. Protein tyrosine phosphatases (PTPases) have been shown to be negative regulators of the insulin receptor. Inhibiton of PTPases may be an effective method in the treatment of type 2 diabetes. A series of azolidinediones has been prepared as protein tyrosine phosphatase 1B (PTP1B) inhibitors. Several compounds were potent inhibitors against the recombinant rat and human PTP1B enzymes with submicromolar IC(50) values. Elongated spacers between the azolidinedione moiety and the central aromatic portion of the molecule as well as hydrophobic groups at the vicinity of this aromatic region were very important to the inhibitory activity. Oxadiazolidinediones 87 and 88 and the corresponding acetic acid analogues 119 and 120 were the best h-PTP1B inhibitors with IC(50) values in the range of 0.12-0.3 microM. Several compounds normalized plasma glucose and insulin levels in the ob/ob and db/db diabetic mouse models.
Subject(s)
Enzyme Inhibitors/chemical synthesis , Hypoglycemic Agents/chemical synthesis , Membrane Proteins/antagonists & inhibitors , Oxazoles/chemical synthesis , Protein Tyrosine Phosphatases/antagonists & inhibitors , 4-Nitrophenylphosphatase/antagonists & inhibitors , Animals , Blood Glucose/metabolism , Diabetes Mellitus, Type 2/blood , Diabetes Mellitus, Type 2/genetics , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Humans , Hypoglycemic Agents/chemistry , Hypoglycemic Agents/pharmacology , In Vitro Techniques , Insulin/blood , Liver/drug effects , Liver/enzymology , Male , Mice , Mice, Obese , Oxazoles/chemistry , Oxazoles/pharmacology , Protein Tyrosine Phosphatase, Non-Receptor Type 1 , Rats , Rats, Sprague-Dawley , Recombinant Proteins/antagonists & inhibitors , Structure-Activity RelationshipABSTRACT
Insulin resistance in the liver and peripheral tissues, together with a pancreatic cell defect, are the common causes of Type 2 diabetes. It is now appreciated that insulin resistance can result from a defect in the insulin receptor signaling system, at a site post binding of insulin to its receptor. Protein tyrosine phosphatases (PTPases) have been shown to be negative regulators of the insulin receptor. Inhibition of PTPases may be an effective method in the treatment of Type 2 diabetes. We have identified two novel series of benzofuran/benzothiophene biphenyl oxo-acetic acids and sulfonyl-salicylic acids as potent inhibitors of PTP1B with good oral antihyperglycemic activity. To assist in the design of these inhibitors, crystallographic studies have attempted to identify enzyme inhibitor interactions. Resolution of crystal complexes has suggested that the inhibitors bind to the enzyme active site and are held in place through hydrogen bonding and van der Waals interactions formed within two hydrophobic pockets. In the oxo-acetic acid series, hydrophobic substitutents at position-2 of the benzofuran/benzothiophene biphenyl framework interacted with Phe182 of the catalytic site and were very critical to the intrinsic activity of the molecule. The hydrophobic region of the catalytic-site pocket was exploited and taken advantage by hydrophobic substituents at either the alpha-carbon or the ortho aromatic positions of the oxo-acetic acid moiety. Similar ortho aromatic substitutions on the salicylic acid-type inhibitors had no effect, primarily due to the different orientation of these inhibitors in the catalytic site. The most active inhibitors of both series inhibited recombinant human PTP1B with phosphotyrosyl dodecapeptide TRDI(P)YETD(P)Y(P)YRK as the source of the substrate with IC(50) values in the range of 20-50 nM. Compound 68 was one of the most active compounds in vivo, normalizing plasma glucose levels at the 25 mg/kg dose (po) and the 1 mg/kg dose (ip). Compound 68 was also selective against several other PTPases.
Subject(s)
Benzofurans/chemical synthesis , Enzyme Inhibitors/chemical synthesis , Hypoglycemic Agents/chemical synthesis , Phenylpropionates/chemical synthesis , Protein Tyrosine Phosphatases/antagonists & inhibitors , Thiophenes/chemical synthesis , Administration, Oral , Animals , Benzofurans/chemistry , Benzofurans/pharmacology , Blood Glucose/analysis , Catalytic Domain , Crystallography, X-Ray , Diabetes Mellitus/blood , Diabetes Mellitus/genetics , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Humans , Hydrogen Bonding , Hypoglycemic Agents/chemistry , Hypoglycemic Agents/pharmacology , Mice , Models, Molecular , Phenylpropionates/chemistry , Phenylpropionates/pharmacology , Protein Tyrosine Phosphatase, Non-Receptor Type 1 , Protein Tyrosine Phosphatases/chemistry , Structure-Activity Relationship , Thiophenes/chemistry , Thiophenes/pharmacologySubject(s)
Enzyme Inhibitors/chemical synthesis , Furans/chemical synthesis , Hypoglycemic Agents/chemical synthesis , Protein Tyrosine Phosphatases/antagonists & inhibitors , Thiophenes/chemical synthesis , Animals , Enzyme Inhibitors/chemistry , Furans/chemistry , Humans , Hypoglycemic Agents/chemistry , Mice , Mice, Obese , Models, Molecular , Protein Tyrosine Phosphatases/chemistry , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/chemistry , Structure-Activity Relationship , Thiophenes/chemistryABSTRACT
Resistance of Walker 256 rat mammary carcinoma cells to chlorambucil has been shown to be accompanied by a specific increase in the A2-2 subunit of glutathione S-transferase (GST) (Buller et al., Mol Pharmacol 31: 575-578, 1987). Analysis of the time course of GST activity following chlorambucil exposure revealed a 7.5- and 3-fold elevation on day 7 post-treatment in Walker-sensitive (WS) and Walker-resistant (WR) cells, respectively. Flow activated cell sorting (FACS) analyses using antibodies specific for rat liver cytosolic GST supported these results and demonstrated the heterogeneous response of WS cells to chlorambucil exposure. The range of GST levels in drug-treated cells was very broad as compared to that of untreated cells. Transcripts for each class of GST (alpha, mu and pi) were quantified for days 1-9 post-treatment from densitometric scans of RNA slot blots. Elevations in GST alpha RNA preceded increases in GST activity (day 7) in both WS and WR cells. Because fluctuations in GSTA1-1 transcripts were not observed, it was concluded that the increased expression of the alpha class must be attributed to increases in GSTA2-2 transcripts. Amplification of the GST genes in drug-treated cells was not present. These results support the role of GSTA2-2 in the detoxification of chlorambucil. The time course of the cellular response to chlorambucil suggests that the elevation of GSTA2-2 transcripts following alkylating agent exposure may represent only one component of a series of events which collectively confer protection and lead to the establishment of drug resistance.
Subject(s)
Chlorambucil/pharmacology , Glutathione Transferase/metabolism , Animals , Carcinoma 256, Walker , Drug Resistance , Enzyme Activation/drug effects , RNA/isolation & purification , RNA/metabolism , Rats , Time Factors , Tumor Cells, Cultured/drug effects , Tumor Cells, Cultured/enzymologyABSTRACT
Overexpression of the glutathione S-transferases (GSTs) and their involvement in the detoxification of anticancer agents has prompted numerous investigations of the enzyme activity of human tumor tissue. This study represents an in-depth evaluation of the contribution of patient history and pathological status to the GST activity of various human tissues. GST activity was elevated significantly in tumors of the lung, breast and colon as compared to unmatched and matched normal tissue from the same organ. The GST activity of primary breast tumors varied significantly with the stage of the tumor. Breast tumors previously treated with both radiation and chemotherapy had significantly lower levels of GST activity than untreated tumors. Neither progesterone nor estrogen receptor content was associated with the GST activity in primary breast tumors. Colon metastases possessed higher levels of GST activity than primary colon tumors but enzyme activity was independent of the Duke's classification of the tumor. Only tumors of the left colon had levels of GST activity that were higher than those of adjacent normal mucosa. No relationship was evident between either age or sex and the GST activity of any of the tissues examined. GST activity levels may reflect the site-specific ability of tissues to provide cellular protection against xenobiotics.
