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1.
J Am Chem Soc ; 131(36): 13031-6, 2009 Sep 16.
Article in English | MEDLINE | ID: mdl-19697908

ABSTRACT

Circular dichroism (CD) spectroscopy is a well-known method for the analysis of chiral chemical compounds and is often used for studying the structure and interaction of proteins, DNA and bioactive compounds in solution. Here we demonstrate that CD spectroscopy is also a powerful tool for investigating the cellular uptake and mode of action of drugs in live cells. By means of CD spectroscopy, we identified DNA as the cellular target of several novel anticancer agents based on the highly cytotoxic natural antibiotic CC-1065. Furthermore, time-dependent changes in the CD spectra of drug-treated cells enabled us to rationalize differences in drug cytotoxicity. The anticancer agents rapidly penetrate the cell membrane and bind to cellular DNA as their intracellular target. Thereby, the formation of a reversible noncovalent complex with the DNA is followed by a covalent binding of the drugs to the DNA and the more toxic compounds show a higher stability and a lower alkylation rate. Since no drug manipulation is necessary for this kind of investigation and achiral compounds bound to chiral biomolecules may also show induced CD signals, CD spectroscopy of live cells is not limited to the study of analogues of CC-1065. Thus, it constitutes a general approach for studying the mode of action of bioactive compounds on the cellular and molecular level.


Subject(s)
Antibiotics, Antineoplastic/pharmacology , Circular Dichroism , DNA/metabolism , Indoles/pharmacology , Antibiotics, Antineoplastic/analysis , Antibiotics, Antineoplastic/pharmacokinetics , Cell Line, Tumor , Duocarmycins , Humans , Indoles/analysis , Indoles/pharmacokinetics , Molecular Structure
2.
ChemMedChem ; 3(12): 1946-55, 2008 Dec.
Article in English | MEDLINE | ID: mdl-19021160

ABSTRACT

A severe limitation in cancer therapy is the often insufficient differentiation between malign and benign tissue using known chemotherapeutics. One approach to decrease side effects is antibody-directed enzyme prodrug therapy (ADEPT). We have developed new glycosidic prodrugs such as (-)-(1S)-26 b based on the antibiotic (+)-duocarmycin SA ((+)-1) with a QIC(50) value of 3500 (QIC(50)=IC(50) of prodrug/IC(50) of prodrug+enzyme) and an IC(50) value for the corresponding drug (prodrug+enzyme) of 16 pM. The asymmetric synthesis of the precursor (-)-(1S)-19 was performed by arylation of the enantiomerically pure epoxide (+)-(S)-29 (> or = 98 % ee).


Subject(s)
Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Glycosides/chemical synthesis , Glycosides/pharmacology , Prodrugs/chemical synthesis , Prodrugs/pharmacology , Antineoplastic Agents/chemistry , Cell Line, Tumor , Duocarmycins , Glycosides/chemistry , Humans , Indoles/chemical synthesis , Indoles/chemistry , Indoles/pharmacology , Inhibitory Concentration 50 , Neoplasms/drug therapy , Prodrugs/chemistry , Pyrroles/chemical synthesis , Pyrroles/chemistry , Pyrroles/pharmacology , Stereoisomerism , beta-Galactosidase/metabolism
3.
Chemistry ; 14(9): 2811-8, 2008.
Article in English | MEDLINE | ID: mdl-18214880

ABSTRACT

The synthesis of the novel pentagastrin seco-CBI conjugate 3, which is based on the highly cytotoxic antitumor antibiotic (+)-duocarmycin SA (1), is reported. A key step in the synthesis is the palladium-catalyzed carbonylation of aryl bromide 7 to give the benzyl ester 16, which is transformed into the new seco-CBI derivative 21 bearing a carboxylic acid ester moiety. Subsequent transformation of 21 into an activated ester followed by the introduction of beta-alanine and tetragastrin led to the new pentagastrin drug 3 that contains a peptide moiety for targeting cancer cells expressing CCK-B/gastrin receptors.


Subject(s)
Antineoplastic Agents/chemical synthesis , Indoles/chemistry , Indoles/chemical synthesis , Pentagastrin/analogs & derivatives , Pentagastrin/chemistry , Antineoplastic Agents/chemistry , Catalysis , Cell Proliferation/drug effects , Drug Delivery Systems , Duocarmycins , Esters/chemistry , Molecular Structure , Palladium/chemistry , Pentagastrin/chemical synthesis , Pyrroles/chemistry , Receptor, Cholecystokinin B/biosynthesis , Receptor, Cholecystokinin B/drug effects
4.
Int J Mol Sci ; 9(5): 821-837, 2008 May.
Article in English | MEDLINE | ID: mdl-19325786

ABSTRACT

A novel carbamate prodrug 2 containing a pentagastrin moiety was synthesized. 2 was designed as a detoxified analogue of the highly cytotoxic natural antibiotic duocarmycin SA (1) for the use in a targeted prodrug monotherapy of cancers expressing cholecystokinin (CCK-B)/gastrin receptors. The synthesis of prodrug 2 was performed using a palladium-catalyzed carbonylation of bromide 6, followed by a radical cyclisation to give the pharmacophoric unit 10, coupling of 10 to the DNA-binding subunit 15 and transformation of the resulting seco-drug 3b into the carbamate 2 via addition of a pentagastrin moiety.

5.
Chemistry ; 13(16): 4396-409, 2007.
Article in English | MEDLINE | ID: mdl-17455190

ABSTRACT

Novel diastereomerically pure beta-D-galactosidic prodrugs (+)-12 a-e of the cytotoxic antibiotics CC-1065 and the duocarmycins were prepared for an antibody directed enzyme prodrug therapy (ADEPT) using 4 as a substrate via a radical cyclization to give rac-5 and rac-6 followed by a chromatographic resolution of the enantiomers of rac-5, glycosidation and linkage to the DNA-binding units 10 a-e. These only slightly toxic compounds can be toxified enzymatically by an antibody-beta-D-galactosidase conjugate at the surface of malignant cells to give the cytotoxic drugs, which then alkylate DNA. The new prodrugs were tested in in vitro cytotoxicity assays showing excellent QIC(50) values of 4800 and 4300 for (+)-12 a and (+)-12 b, respectively. The absolute configuration of precursor (+)-5 was determined by comparison of the experimental CD spectrum with the theoretically predicted CD spectra and by X-ray structure analysis.


Subject(s)
Antibiotics, Antineoplastic/pharmacology , Indoles/pharmacology , Lung Neoplasms/drug therapy , Prodrugs/pharmacology , Antibiotics, Antineoplastic/chemical synthesis , Antibiotics, Antineoplastic/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Design , Drug Screening Assays, Antitumor , Duocarmycins , Humans , Indoles/chemical synthesis , Indoles/chemistry , Inhibitory Concentration 50 , Molecular Structure , Prodrugs/chemical synthesis , Prodrugs/chemistry , Pyrroles/chemical synthesis , Pyrroles/chemistry , Pyrroles/pharmacology , Stereoisomerism
8.
J Pharmacol Exp Ther ; 315(1): 352-62, 2005 Oct.
Article in English | MEDLINE | ID: mdl-16014573

ABSTRACT

Haloperidol is known as an antagonist of dopamine D2 receptors. However, it also blocks a variety of ion channels at concentrations above the therapeutic range. Reduced haloperidol (R-haloperidol), one of the main metabolites of haloperidol, has been reported to accumulate in certain tissues, particularly in brain cortex, and it may produce the pharmacological effects associated with haloperidol treatment. In this study, we assessed the effect of R-haloperidol and other related compounds on native delayed-rectifier potassium channels (K(DR)) in mouse cortical neurons by using the whole-cell patch-clamp technique. Although R-haloperidol has much lower affinity to D2 receptors than haloperidol, the IC50 of R-haloperidol to block K(DR) currents was 4.4 microM, similar to its parent compound. The binding site of R-haloperidol is on the cytoplasmic side of the channel because its quaternary derivative preferentially inhibited the currents from intracellular side. 4-Chlorophenyl-4-hydroxypiperidine (4C4HP) is the active fragment of haloperidol because other compounds containing this moiety, including L-741,626 (3-[4-(4-chlorophenyl)-4-hydroxypiperidin-L-yl]-methyl-1H-indole) and loperamide, also blocked K(DR) channels. The potency of the 4C4HP fragment positively correlated with the hydrophobicity index (clogP) of the compounds tested. We conclude that R-haloperidol is a K(DR) channel blocker, although it does not interfere with the normal channel function at a clinically relevant concentration.


Subject(s)
Cerebral Cortex/drug effects , Haloperidol/pharmacology , Neurons/drug effects , Potassium Channel Blockers/pharmacology , Animals , Binding Sites , Female , Haloperidol/metabolism , Loperamide/pharmacology , Mice , Mice, Inbred C57BL , Oxidation-Reduction
9.
J Gen Physiol ; 124(4): 301-17, 2004 Oct.
Article in English | MEDLINE | ID: mdl-15365094

ABSTRACT

Ether à go-go (Eag; KV10.1) voltage-gated K+ channels have been detected in cancer cell lines of diverse origin and shown to influence their rate of proliferation. The tricyclic antidepressant imipramine and the antihistamine astemizole inhibit the current through Eag1 channels and reduce the proliferation of cancer cells. Here we describe the mechanism by which both drugs block human Eag1 (hEag1) channels. Even if both drugs differ in their affinity for hEag1 channels (IC50s are approximately 2 microM for imipramine and approximately 200 nM for astemizole) and in their blocking kinetics, both drugs permeate the membrane and inhibit the hEag1 current by selectively binding to open channels. Furthermore, both drugs are weak bases and the IC50s depend on both internal an external pH, suggesting that both substances cross the membrane in their uncharged form and act from inside the cell in their charged forms. Accordingly, the block by imipramine is voltage dependent and antagonized by intracellular TEA, consistent with imipramine binding in its charged form to a site located close to the inner end of the selectivity filter. Using inside- and outside-out patch recordings, we found that a permanently charged, quaternary derivative of imipramine (N-methyl-imipramine) only blocks channels from the intracellular side of the membrane. In contrast, the block by astemizole is voltage independent. However, as astemizole competes with imipramine and intracellular TEA for binding to the channel, it is proposed to interact with an overlapping intracellular binding site. The significance of these findings, in the context of structure-function of channels of the eag family is discussed.


Subject(s)
Astemizole/pharmacology , Imipramine/pharmacology , Ion Channel Gating/drug effects , Ion Channel Gating/physiology , Kidney/drug effects , Kidney/physiology , Potassium Channels, Voltage-Gated/drug effects , Potassium Channels, Voltage-Gated/physiology , Cell Membrane/drug effects , Cell Membrane/physiology , Cells, Cultured , Dose-Response Relationship, Drug , Drug Interactions , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels , Humans , Hydrogen-Ion Concentration , Membrane Potentials/drug effects , Membrane Potentials/physiology , Models, Biological , Recombinant Proteins/metabolism , Structure-Activity Relationship
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