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1.
Crit Rev Oncol Hematol ; 38(2): 139-70, 2001 May.
Article in English | MEDLINE | ID: mdl-11311660

ABSTRACT

Flavopiridol inhibits phosphokinases. Its activity is strongest on cyclin dependent kinases (cdk-1, -2, -4, -6, -7) and less on receptor tyrosine kinases (EGFR), receptor associates tyrosine kinases (pp60 Src) and on signal transducing kinases (PKC and Erk-1). Although the inhibiting activity of flavopiridol is strongest for cdk, the cytotoxic activity of flavopiridol is not limited to cycling cells. Resting cells are also killed. This fact suggests that inhibition of cdks involved in the control of cell cycle is not the only mechanism of action. Inhibition of cdk's with additional functions (i.e. involved in the control of transcription or function of proteins that do not control cell cycle) may contribute to the antitumoral effect. Moreover, direct and indirect inhibition of receptor activation (EGFR) and/or a direct inhibition of kinases (pp60 Src, PKC, Erk-1) involved in the signal transduction pathway could play a role in the antiproliferative activity of flavopiridol. From pharmacokinetic data in patients it can be concluded that the inhibitory activity (IC50) of flavopiridol on these kinases is in the range of concentrations that might be achieved intracellularly after systemic application of non-toxic doses of flavopiridol. However, no in situ data from flavopiridol treated cells have been published yet that prove that by inhibition of EGFR, pp60 Src, PKC and/or Erk-1 (in addition to inhibition of cdk's) flavopiridol is able to induce apoptosis. Thus many questions regarding the detailed mechanism of antitumoral action of flavopiridol are still open. For the design of protocols for future clinical studies this review covers the essential information available on the mechanism of antitumoral activity of flavopiridol. The characteristics of this antitumoral activity include: High rate of apoptosis, especially in leukemic cells; synergy with the antitumoral activity of many cytostatics; independence of its efficacy on pRb, p53 and Bcl-2 expression; lack of interference with the most frequent multidrug resistance proteins (P-glycoprotein and MRP-190); and a strong antiangiogenic activity. Based on these pharmacological data it can be concluded that flavopiridol could be therapeutically active in tumor patients: independent on the genetic status of their tumors or leukemias (i.e. mutations of the pRb and/or p53, amplification of bcl-2); in spite of drug resistance of their tumors induced by first line treatment (and caused by enhanced expression of multidrug resistance proteins); in combination with conventional chemotherapeutics preferentially given prior to flavopiridol; and due to a complex mechanism involving cytotoxicity on cycling and on resting tumor cells, apoptosis and antiangiogenic activity. In consequence, flavopiridol is a highly attractive, new antitumoral compound and deserves further elucidation of its clinical potency.


Subject(s)
Flavonoids/pharmacology , Piperidines/pharmacology , Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Cyclin-Dependent Kinases/antagonists & inhibitors , Drug Synergism , Humans , Tumor Cells, Cultured/drug effects
2.
Crit Rev Oncol Hematol ; 37(3): 169-215, 2001 Mar.
Article in English | MEDLINE | ID: mdl-11248576

ABSTRACT

Targeting cancer gene therapy to endothelial cells seems to be a rational approach, because (a) a clear correlation exists between proliferation of tumor vessels and tumor growth and malignancy, (b) differences of cell membrane structures between tumor endothelial cells and normal endothelial cells exist which could be used for targeting of vectors and (c) tumor endothelial cells are accessible to vector vehicles in spite of the peculiarities of the transvascular and interstitial blood flow in tumors. Based on the knowledge on the pharmacokinetics of macromolecules it can be concluded that vectors targeting tumor endothelial cells should own a long blood residence time after intravascular application. This precondition seems to be fulfilled best by vectors exhibiting a slight anionic charge. A long blood residence time would allow the formation of a high amount of complexes between tumor endothelial cells and vector particles. Such high amount of complexes should enable a high transfection rate of tumor endothelial cells. In view of their pharmacokinetic behavior nonviral vectors seem to be more suitable for in vivo targeting tumor endothelial cells than viral vectors. Specific binding of nonviral vectors to tumor endothelial cells should be enhanced by multifunctional ligands and the transduction efficiency should be improved by cationic carriers. Effector genes should encode proteins potent enough to induce reactions which eliminate the tumor tissue. To be effective to that degree such proteins should induce self-amplifying antitumor reactions. Examples for proteins which have the potential to induce such self-amplifying tumor reactions are proteins endowed with antiangiogenic and antiproliferative activity, enzymes which convert prodrugs into drugs and possibly also proteins which induce embolization of tumor vessels. The pharmacological data for such examples are discussed in detail.


Subject(s)
Endothelium, Vascular , Genetic Therapy , Neoplasms/genetics , Neoplasms/pathology , Neoplasms/therapy , Neovascularization, Pathologic/genetics , Animals , Endothelium, Vascular/pathology , Endothelium, Vascular/physiology , Gene Expression Regulation, Neoplastic , Gene Targeting , Humans , Neoplasms/blood supply
3.
Drugs ; 59(3): 435-76, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10776829

ABSTRACT

The most essential kinases involved in cell membrane receptor activation, signal transduction and cell cycle control or programmed cell death and their interconnections are reviewed. In tumours, the genes of many of those kinases are mutated or amplified or the proteins are overexpressed. The use of key kinases offers the possibility to screen in vitro for synthetic small molecule kinase inhibitors. In view of the many interconnections of cellular kinases, their role in preventing or inducing programmed cell death and the possibility that a considerable number of signal transducing proteins are still unknown, cellular test systems are recommended in which the respective key kinase or one of its main partner molecules are overexpressed.


Subject(s)
Cell Membrane/drug effects , ErbB Receptors/drug effects , Neoplasms/drug therapy , Phosphotransferases/antagonists & inhibitors , Signal Transduction/drug effects , Cell Cycle/drug effects , Humans , Neoplasms/etiology , Phosphotransferases/adverse effects , Phosphotransferases/genetics , Phosphotransferases/metabolism , Protein Kinase Inhibitors , Protein Kinases/adverse effects , Protein Kinases/metabolism , Protein Serine-Threonine Kinases/adverse effects , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein Serine-Threonine Kinases/metabolism , Protein-Tyrosine Kinases/adverse effects , Protein-Tyrosine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/metabolism , Receptor, IGF Type 1/drug effects , Transcription, Genetic , src-Family Kinases/adverse effects , src-Family Kinases/metabolism
4.
Gene Ther ; 7(3): 224-31, 2000 Feb.
Article in English | MEDLINE | ID: mdl-10694799

ABSTRACT

A major problem of tumor gene therapy is the low transduction efficiency of the currently available vectors. One way to circumvent this problem is the delivery of therapeutic genes encoding intracellular enzymes for the conversion of a prodrug to a cytotoxic drug which can then spread to neighboring non-transduced cells (bystander effect). One possibility to improve the bystander effect could be the extracellular conversion of a hydrophilic prodrug to a lipophilic, cell-permeable cytotoxic drug. Toward this end, we have used a secreted form of the normally lysosomal human beta-glucuronidase (s-betaGluc) to establish an extracellular cytotoxic effector system that converts an inactivated glucuronidated derivative of doxorubicin (HMR 1826) to the cytotoxic drug. We demonstrate that s-betaGluc-transduced tumor cells convert HMR 1826 to doxorubicin which is taken up by both transduced and non-transduced cells. s-betaGluc in combination with HMR 1826 efficiently induces tumor cell killing both in cell culture and in vivo. This effect is mediated through a pronounced bystander effect of the generated cytotoxic drug. Most notably, this gene therapeutic strategy is shown to be clearly superior to conventional chemotherapy with doxorubicin. Gene Therapy (2000) 7, 224-231.


Subject(s)
Choriocarcinoma/therapy , Doxorubicin/analogs & derivatives , Glucuronates/therapeutic use , Glucuronidase/genetics , Prodrugs/therapeutic use , Choriocarcinoma/enzymology , Dose-Response Relationship, Drug , Doxorubicin/therapeutic use , Gene Transfer Techniques , Glucuronates/genetics , Glucuronidase/metabolism , Humans , Immunohistochemistry , Transduction, Genetic/genetics , Tumor Cells, Cultured
5.
Int J Cancer ; 81(4): 568-72, 1999 May 17.
Article in English | MEDLINE | ID: mdl-10225446

ABSTRACT

Endoglin/CD105 is a membrane protein involved in the TGF-beta receptor signalling pathway. Endoglin expression has been reported to be selective for a few cell types, in particular endothelial cells, although a number of conflicting reports have been published. In this study, we performed a detailed analysis of endoglin expression in human lung tumors and different tumor and endothelial cell lines, employing reverse-transcriptase-polymerase-chain reaction as well as immunoblotting and immunohistochemistry using verified antibodies to endoglin. Our data show a clearly preferential expression of both endoglin mRNA and protein in endothelial cells. In tumors, endoglin expression was strongly elevated in the angiogenic endothelium at the tumor edges. In agreement with this observation, we find a clear correlation between endoglin expression and markers of proliferation, such as cyclin A and Ki-67, suggesting that endoglin expression is linked to cell-cycle regulation. These findings not only resolve some of the discrepancies in the literature, but also provide the basis for further applications making use of its selective localization and expression in the tumor vasculature.


Subject(s)
Endothelium, Vascular/metabolism , Lung Neoplasms/blood supply , Lung/physiology , Transcription, Genetic , Vascular Cell Adhesion Molecule-1/genetics , Antigens, CD , Cells, Cultured , Endoglin , Endothelium, Vascular/cytology , Endothelium, Vascular/pathology , Humans , Lung/cytology , Lung Neoplasms/pathology , RNA, Messenger/genetics , Receptors, Cell Surface , Receptors, Transforming Growth Factor beta/genetics , Receptors, Transforming Growth Factor beta/physiology , Reverse Transcriptase Polymerase Chain Reaction , Umbilical Veins
6.
Int J Cancer ; 77(1): 146-52, 1998 Jul 03.
Article in English | MEDLINE | ID: mdl-9639406

ABSTRACT

The anti-tumor drug Flavopiridol is a potent inhibitor of cyclin-dependent kinases (cdks). As a consequence, Flavopiridol-treated cells arrest in both G1 and G2, but Flavopiridol has also been shown to be cytotoxic for some tumor cell lines. The underlying molecular events are, however, unclear. We now show that Flavopiridol induces apoptosis in human umbilical vein endothelial cells (HUVECs), as judged by the occurrence of classical apoptotic markers, including chromatin condensation, internucleosomal cleavage, DNA fragmentation (TUNEL assay), annexin V binding and poly(ADP-ribose) polymerase (PARP)-cleavage. Such induction of apoptosis occurs with equal efficiency in both proliferating and G0/G1-arrested cells. Because growth-arrested HUVECs lack cdk2 activity and contain high levels of the cdk inhibitor p27, our observations suggest that cell cycle regulated cdks may not be the only critical target for Flavopiridol-induced apoptosis. Surprisingly, A549 lung carcinoma cells were clearly dependent on cell proliferation for the induction of cell death, pointing to cell type-related differences in the mechanism of Flavopiridol action.


Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Endothelium, Vascular/drug effects , Endothelium, Vascular/pathology , Flavonoids/pharmacology , Piperidines/pharmacology , Cell Cycle/drug effects , Cells, Cultured , Humans
7.
Life Sci ; 62(20): 1861-73, 1998.
Article in English | MEDLINE | ID: mdl-9600328

ABSTRACT

Flavopiridol (FLAP) is a promising novel chemotherapeutic agent currently undergoing clinical phase I trials. To examine hepatic metabolism and biliary disposition of FLAP we applied the isolated perfused rat liver system. Besides FLAP two metabolites were detected by high performance liquid chromatography in bile and perfusate. Twenty-five min after FLAP (30 microM) addition to the perfusion medium, biliary secretion of metabolite 1 and 2 reached a maximum of 1.04 +/- 0.52 and 11.34 +/- 4.72 nmol/g.liver.min, respectively. Biliary excretion of parent FLAP, however, continuously increased for 60 min up to 406 +/- 134 pmol/g liver.min. In the perfusate, metabolite 1 was below detection limit and release of metabolite 2 was low (2.8 +/- 0.7 pmol/g liver.min after 60 min). Enzymatic hydrolysis with beta-glucuronidase, mass spectroscopy and electron absorption spectroscopy revealed that both metabolites are monoglucuronides with the glucuronide in position 5 and 7 of the flavonoid core, respectively. The amount of FLAP, metabolite 1 and metabolite 2 excreted into bile during the 60 min of perfusion was 1.94 +/- 0.91, 5.15 +/- 1.95 and 57.29 +/- 23.60% of FLAP cleared from the perfusate during 60 min, respectively. In contrast to the structurally similar flavonoids genistein and daidzein, no inhibition of UDP-glucuronyltransferase with methylumbelliferone as a substrate was observed indicating that different UDP-glucuronyltransferase isoforms are involved in FLAP metabolism. In conclusion, we find that glucuronidation is the major mechanism of hepatic FLAP biotransformation. Metabolites are mainly excreted into bile but also released into systemic circulation. The pharmacological and toxicological effects of these metabolites remain to be elucidated.


Subject(s)
Antineoplastic Agents/metabolism , Cyclin-Dependent Kinases/antagonists & inhibitors , Enzyme Inhibitors/metabolism , Flavonoids/metabolism , Liver/metabolism , Piperidines/metabolism , Animals , Bile/metabolism , Biotransformation , Chromatography, High Pressure Liquid , Electron Spin Resonance Spectroscopy , Enzyme Inhibitors/pharmacology , Flavonoids/pharmacology , Glucuronates/metabolism , Glucuronosyltransferase/metabolism , Hydrolysis , Liver/drug effects , Male , Mass Spectrometry , Piperidines/pharmacology , Rats , Rats, Wistar
8.
Biochem J ; 301 ( Pt 3): 821-8, 1994 Aug 01.
Article in English | MEDLINE | ID: mdl-8053907

ABSTRACT

The cDNA sequence encoding human beta-glucuronidase [Oshima, Kyle, Miller, Hoffmann, Powell, Grubb, Sly, Troplak, Guise and Gravel (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 685-689] was expressed in baby hamster kidney (BHK) cells. After purification from the culture supernatant in one step by use of immunoaffinity chromatography, the biochemical properties of the enzyme were examined. With a pH optimum of 4.0, a Km of 1.3 mM and thermal stability up to 68 degrees C, this protein has characteristics very similar to those described for beta-glucuronidase from human placenta [Brot, Bell and Sly (1978) Biochemistry 17, 385-391. However, the recombinant product has several structural properties not previously reported for beta-glucuronidase isolated from natural sources. First, recombinant beta-glucuronidase is synthesized as a tetramer consisting of two disulphide-linked dimers. As can be inferred from the cDNA sequence, the enzyme possesses five cysteine residues after cleavage of the signal peptide. By introducing a C-terminal truncation, we eliminated the last cysteine at position 644. In the mutant, covalent linkage between two monomers is no longer observed, indicating that Cys-644 is involved in intermolecular disulphide-bond formation. The functional role of the disulphide bond remains elusive, as it was shown that (i) intracellular transport of the mutant is not impaired and (ii) it is still able to form an enzymically active tetramer. A second feature that has not previously been observed for beta-glucuronidase from any origin is the existence of two enzymically active species for recombinant beta-glucuronidase, when examined by gel filtration on a TSK 3000 column. With apparent molecular masses of 380 kDa and 190 kDa we propose that they represent tetramers and dimers respectively. Partial N-terminal sequencing and electrophoresis under denaturing conditions revealed that the dimers consist of subunits that have been proteolytically processed at their C-terminus losing 3-4 kDa in peptide mass. Controlled proteolysis demonstrates that the enzyme's overall protein backbone as well as its activity are resistant to a number of proteases. Only the C-terminal portion is susceptible to protease action, and the disulphide-linked form is readily converted into non-disulphide-bonded subunits. Pulse-chase analysis shows that human beta-glucuronidase remaining intracellular in BHK cells after synthesis undergoes a similar proteolytic processing event, i.e. a reduction in mass of 3-4 kDa.(ABSTRACT TRUNCATED AT 400 WORDS)


Subject(s)
Glucuronidase/metabolism , Recombinant Proteins/metabolism , Animals , Base Sequence , Biological Transport , Cell Line , Chromatography, Affinity , Chromatography, Gel , Cricetinae , DNA, Complementary/chemistry , Disulfides/metabolism , Endopeptidases/metabolism , Enzyme Stability , Gene Expression , Glucuronidase/chemistry , Glucuronidase/genetics , Humans , Hydrogen-Ion Concentration , Kidney , Macromolecular Substances , Molecular Sequence Data , Mutation , Transfection
9.
Crit Rev Oncog ; 5(6): 555-87, 1994.
Article in English | MEDLINE | ID: mdl-8519811

ABSTRACT

Tumors are resistant to the immune response as evidenced by both their progressive growth in patients despite specific humoral and cellular immune responses to tumor antigens and by the moderate clinical effect of active specific immunotherapy with tumor vaccines tested to date. This "immune resistance" may be due to various reasons, among which the most important ones are: (1) in the afferent pathway of the immune response, (a) expression of major histocompatibility complex (MHC)-class II molecules without coexpression of costimulatory B7 molecules on tumor cells, which impairs activation of T- and B-cells and, (b) release of prostaglandins and other factors from tumor cells, that may inhibit proliferation and function of helper T-cells; and (2) in the efferent pathway; (c) release of tumor antigens, which blocks cytotoxic cells and antibodies and release of proteolytic enzymes, which degrades specific antibodies; (d) reduced expression of MHC-class I molecules by tumor cells, which inhibits their recognition by cytotoxic T-lymphocytes (CTL); and (e) cell membrane-associated inhibitors of complement factors that block complement-mediated lysis. Altogether, the chance for a successful tumor therapy by tumor vaccines has to be estimated to be low. Alternatives would be to use tumor antigens as tumor cell targets for cytotoxic compounds with differing action from the cytotoxic mechanisms used by the immune system. The problems of low tumor localization rates of tumor-specific monoclonal antibodies, immune resistance of the tumor cells, and general toxicity of cytotoxic drugs may be solved by a biphasic therapeutic approach called immune specific enzyme-mediated chemotherapy. It uses, in a first phase, an appropriate antibody-enzyme fusion protein and, in the second phase, a nontoxic prodrug that is cleaved at the tumor site by the enzyme of the fusion protein into the cytotoxic drug.


Subject(s)
Immunotherapy , Neoplasms/therapy , Antibodies, Neoplasm/biosynthesis , Antibodies, Neoplasm/immunology , Humans , Immunity, Cellular , Neoplasms/immunology , Vaccination
10.
Exp Toxicol Pathol ; 44(6): 317-24, 1992 Oct.
Article in English | MEDLINE | ID: mdl-1280497

ABSTRACT

Two monoclonal antibodies (mabs) were raised against ganglioside GD2 (Gtri2) and tested on human intracranial tumors and normal brain by immunohistochemical methods both in frozen and paraffin embedded sections. In normal brain structures, astrocytes were visualized with both mabs (BW 625 and BW 704) almost exclusively in the subventricular and subpial layer. A minor amount of myelin sheaths in well defined localisation was only recognized in frozen sections. Consequently in astrocytic tumors of different grades of malignancy (WHO I-IV), astrocytes were depicted in their characteristic shape and arrangement around vessels. In addition, staining was observed in meningiomas and schwannomas, but not in pituitary adenomas or metastatic carcinomas. In meningioma und schwannoma the staining was restricted to the cellular periphery and was again present in frozen section material and absent in paraffin embedded tissue. In astrocytes, reactive and neoplastic, obviously fibrous processes and cytoplasm were distinctly stained both in frozen and paraffin embedded sections. It is concluded that some neuroectodermal derived cells as well as myelin of defined localisation express GD2 on the membrane surfaces and in the cytoplasm. The latter may primarily be the case in fibrous astrocytes, which were stained in reactive and pathologic conditions. The reaction can be used as diagnostic tool in astrocytic tumors; its positive therapeutic significance is hampered by the fact that (1) not all cells in heterogeneous tumor populations express the epitope and (2) there are normal structures which do so.


Subject(s)
Brain Chemistry , Brain Neoplasms/chemistry , Epitopes/analysis , Gangliosides/analysis , Glioma/chemistry , Meningioma/chemistry , Neurilemmoma/chemistry , Antibodies, Monoclonal , Humans , Immunohistochemistry
11.
Br J Cancer ; 65(2): 234-8, 1992 Feb.
Article in English | MEDLINE | ID: mdl-1739623

ABSTRACT

A fusion protein consisting of the humanised Fab fragment of the anti CEA MAb BW 431 and the human beta-glucuronidase was expressed in BHK cells. Functional testing revealed that the specificity and avidity of the humanised V region was similar to the original murine MAb BW 431. Furthermore, the enzymatic activity, pH sensitivity and stability of the human beta-glucuronidase in the fusion protein was comparable to the activity of recombinant human beta-glucuronidase. Using anti-idiotype affinity chromatography, two molecules of a molecular weight of 125 kDa or 250 kDa could be visualized under nonreducing conditions in SDS-PAGE. Reducing conditions revealed a 25 kDa light and 100 kDa heavy chain. Due to its suitable biological characteristics this fusion protein might be an appropriate molecule allowing a site specific antibody directed enzyme prodrug therapy (ADEPT) in vivo.


Subject(s)
Genetic Engineering/methods , Prodrugs/metabolism , Recombinant Fusion Proteins/metabolism , Carcinoembryonic Antigen/metabolism , Cell Line , Hydrogen-Ion Concentration , Immunoglobulin G/genetics , Promoter Regions, Genetic , Recombinant Fusion Proteins/isolation & purification , Transfection
12.
Br J Cancer ; 63(5): 681-6, 1991 May.
Article in English | MEDLINE | ID: mdl-2039692

ABSTRACT

A two phase radioimmunotherapy based on bispecific MAbs in which one arm recognises a tumour antigen and the other a radiolabelled chelate, may prove more effective in the treatment of carcinomas than currently available immunotherapies. To establish this system we first showed that penetration into human carcinoma xenografts as well as long term retention of intact MAb outside the carcinoma cells can be obtained. Epitope saturation was not obtained however, despite the large MAb doses injected i.v. for 10 days. We then generated hybridomas producing high avidity anti-metal chelate MAbs (anti-DTPA-Y). These hybridomas were fused with hybridomas producing MAbs against CEA or GIT-mucin, and stable bispecific MAb producing quadromas were obtained. For the anti-GIT-mucin x anti-chelate MAb a purification procedure based on double anti-idiotype affinity chromatography was shown to result in greater than 95% pure bispecific immunoreactive MAb. Comparative in vivo stability studies profiled DTPA-Y as the chelate of choice for in vivo application.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Colonic Neoplasms/radiotherapy , Pancreatic Neoplasms/radiotherapy , Pentetic Acid/therapeutic use , Yttrium Radioisotopes/therapeutic use , Animals , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/metabolism , Colonic Neoplasms/metabolism , Humans , Injections, Intravenous , Mice , Mice, Nude , Pancreatic Neoplasms/metabolism , Pentetic Acid/metabolism , Tumor Cells, Cultured , Yttrium Radioisotopes/metabolism
13.
Int J Immunopharmacol ; 13(5): 541-8, 1991.
Article in English | MEDLINE | ID: mdl-1783467

ABSTRACT

The isoxazoline derivative HAB 439 was tested for its enzyme inhibiting potency and was found to be an inhibitor of aminopeptidase B (IC50 = 22.5 micrograms/ml). In further immunopharmacological experiments its efficacy to stimulate cell-mediated immunity was evaluated. HAB 439 was shown to stimulate DTH-reaction against Salmonella typhimurium and Listeria monocytogenes. HAB 439 protected animals against infection by reducing the bacterial load in livers and spleens and by decreasing the mortality rate. Treatment with the antibiotic ampicillin induced a decreased DTH-reaction in mice which was demonstrated to be due to a reduction of the antigen to be presented to the immune system and not to immune suppression. HAB 439 restored the impaired immune response to S. typhimurium and L. monocytogenes in a dose-dependent way. Restoration of DTH was shown to lead to an improvement of protection in ampicillin-treated mice which were challenged with the intracellular bacteria.


Subject(s)
Immunity, Cellular/drug effects , Isoxazoles/pharmacology , Listeriosis/immunology , Organophosphorus Compounds/pharmacology , Salmonella Infections/immunology , T-Lymphocytes/immunology , Aminopeptidases/antagonists & inhibitors , Ampicillin/pharmacology , Animals , Female , Hypersensitivity, Delayed , Listeria monocytogenes/immunology , Mice , Prognosis , Salmonella typhimurium/immunology
14.
Behring Inst Mitt ; (87): 33-47, 1990 Dec.
Article in English | MEDLINE | ID: mdl-2096819

ABSTRACT

The application of monoclonal antibodies (MAb) of non human origin in the treatment of human diseases is hampered by the immunogenicity of those molecules in patients. The development of molecular biology provides us with the opportunity to overcome the problem of immunogenicity and even allows the development of new immunotherapeutic approaches. These approaches use the MAb as carriers to deliver certain properties (MAb + X) to a specific site in the patient. These properties can be toxic principles like radioactivity or cytostatically active substances, enzymatic activities or specific receptor functions. In addition to the molecular biology of MAb a detailed analysis of the pharmacokinetic behaviour of molecules of MAb size (150 KD) and of small molecules (less than 1KD) in solid tissues gave rise to a new therapeutical concept called two phase immunotherapy. This concept is based on the separation of the localisation phase, the phase in which the MAb + X molecule localises at the position where therapy is desired, from the toxic phase. During the toxic phase, a untoxic "substance" develops a toxic activity by the action of the MAb + X only at the desired localisation. This concept is presently converted into practice in numerous different approaches, for example in a two phase tumor radioimmunotherapy, in a immune specific enzyme mediated chemotherapy (ISEC) and in a concept using Major Histocompatibility (MHC) antigen specific CTL as toxic agents to destroy MHC class I labeled target cells.


Subject(s)
Antibodies, Monoclonal/genetics , Genes, Immunoglobulin , Neoplasms/therapy , Animals , Antibodies, Monoclonal/therapeutic use , Gene Rearrangement , Genetic Engineering/methods , Humans , Immunoglobulin Variable Region/genetics , Immunotherapy , Mice , Models, Molecular , Protein Conformation , Recombinant Proteins/therapeutic use
15.
Behring Inst Mitt ; (87): 68-75, 1990 Dec.
Article in English | MEDLINE | ID: mdl-2096821

ABSTRACT

The distribution of the monoclonal antibody (MAb) BW494 in human pancreatic carcinoma biopsies during high dose i.v. immunotherapy was investigated. Using immunohistochemical techniques combined with anti-idiotypic, endothelial cell specific and bispecific MAbs it was shown that 3 days after onset of immunotherapy, MAb BW494 was bivalently bound to tumor cells in some highly vascualized areas near capillaries. No binding was observed in other highly vascularized tumor cell areas although the epitope detected by MAb BW494 was present. In contrast to our expectation the majority of the tumor cells was not yet saturated by the antibody, probably due to diffusion barriers in the solid tumor tissue.


Subject(s)
Antibodies, Monoclonal , Immunotherapy , Pancreatic Neoplasms/pathology , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/therapeutic use , Humans , Immunohistochemistry , Pancreatic Neoplasms/immunology , Pancreatic Neoplasms/therapy
17.
Br J Cancer Suppl ; 10: 37-9, 1990 Jul.
Article in English | MEDLINE | ID: mdl-2200494

ABSTRACT

The distribution of the monoclonal antibody (MAb) BW494 in human pancreatic carcinoma biopsies during high dose intravenous immunotherapy was investigated. Using immunohistochemical techniques combined with anti-idiotypic, endothelial cell-specific and bispecific MAbs, it was shown that 3 days after onset of immunotherapy, MAb BW494 was bivalently bound to tumour cells in some highly vascularised areas near capillaries. No binding was observed in other highly vascularised tumour cell areas although the epitope detected by MAb BW494 was present. In contrast to our expectation the majority of the tumour cells were not yet saturated by the antibody, probably due to diffusion barriers in the solid tumour tissue.


Subject(s)
Antibodies, Monoclonal/metabolism , Pancreatic Neoplasms/metabolism , Antibodies, Monoclonal/therapeutic use , Humans , Immunoenzyme Techniques , Pancreatic Neoplasms/therapy
18.
J Med Chem ; 33(1): 166-71, 1990 Jan.
Article in English | MEDLINE | ID: mdl-2296017

ABSTRACT

The cytotoxic activities of several natural and semisynthetic anthracyclines against L1210 leukemia and two human colon tumor cells (Colon 4, HT 29) in vitro were examined after short (1 h) and long (7 days) incubation times and correlated with the water/octanol partition coefficients and the DNA-binding affinity of the compounds. Analysis of equation in which cytotoxicity against L1210 (1-h incubation) was parabolically related to the partition coefficient revealed an almost exclusive correlation (r = 0.80) between the cytotoxicity and the parameters, and this correlation was only slightly improved by addition of DNA-binding affinity (r = 0.85). On the other hand, cytotoxic activities displayed after continuous incubation were partially related to both partition coefficients (parabolic dependence) and DNA-binding affinities (linear dependence). In this case the correlation between the activity and partition coefficient (r = 0.67) was significantly improved by addition of DNA-binding affinity (r = 0.90). Similar results were also obtained for human colon tumor cells although the corresponding correlation coefficients were generally of lower value, indicating that cytotoxic activity of anthracyclines against these primary resistant cells may be influenced by additional factors not yet determined.


Subject(s)
Antibiotics, Antineoplastic/therapeutic use , Neoplasms/drug therapy , Animals , Antibiotics, Antineoplastic/metabolism , Chemical Phenomena , Chemistry, Physical , Colonic Neoplasms/drug therapy , DNA/metabolism , Humans , Leukemia L1210/drug therapy , Molecular Structure , Regression Analysis , Structure-Activity Relationship , Tumor Cells, Cultured
19.
Arzneimittelforschung ; 39(10): 1235-9, 1989 Oct.
Article in English | MEDLINE | ID: mdl-2692570

ABSTRACT

To evaluate the influence of [2-(3-carboxy-1-probylthio)-4-methyl-1,3-thiazole]acetic acid (tiprotimod, HBW 538) on the host defense mechanisms, a number of experimental studies in different animal models were performed. The prophylactic treatment of NMRI mice with tiprotimod significantly prolonged the mean survival time of the animals after intravenous infection with Candida albicans 200/175 and increased the resistance to the fungal infection to 180% in comparison to controls. In vitro the drug showed no direct fungistatic or fungicidal activity. In an experimental model of persistent systemic candidiasis Balb/c mice infected intravenously with Candida albicans were treated with the immunomodulator tiprotimod after the fungal colonization of kidney was manifested (3 days post infection). The treatment of the mice after the infection resulted in a reduction of the infectious load and the abscess formation in kidney as well as in a decrease of numbers of yeasts in the urine. In the syngeneic B16 melanoma tumor model tiprotimod significantly prolonged the medium survival time and reduced the number of visuable metastases in the lungs even when applied after resection of the primary tumor graft. Tiprotimod also beneficially influenced the course of the disease in two murine graft-vs-host models (hemolytic anemia and immune complex glomerulonephritis) which lead to a B cell mediated autoimmune disease with fatal outcome. The application of the drug in the induction phase mitigated the development of the diseases and prevented animals from dying.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Adjuvants, Immunologic/pharmacology , Thiazoles/pharmacology , Anemia, Hemolytic, Autoimmune/immunology , Anemia, Hemolytic, Autoimmune/microbiology , Animals , Candida albicans/drug effects , Candidiasis/immunology , Candidiasis/microbiology , Female , Graft vs Host Reaction/drug effects , Male , Melanoma, Experimental/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Inbred DBA , Mice, Inbred Strains , Rats , Sepsis/immunology , Sepsis/microbiology , Skin Transplantation
20.
Arzneimittelforschung ; 39(9): 1085-9, 1989 Sep.
Article in English | MEDLINE | ID: mdl-2590257

ABSTRACT

The thiazole derivative [2-(3-carboxy-1-propylthio)-4-methyl-1,3-thiazole] acetic acid (tiprotimod, HBW 538) a new synthetic immunopotentiator of low molecular weight, has been tested in vivo and ex vivo in various experimental models. Its influence on parameters of macrophage functions, on DTH (delayed type hypersensitivity)-reaction and antibodies to sheep erythrocytes (SRBC), Tetanus toxoid and heatkilled E. coli bacteria in mice, and in the popliteal lymph node assay in rats was investigated. When mice were treated with the test substance i.v., i.p., or p.o. in a dose range from 1-100 mg/kg, a time and dose-dependent stimulation of macrophage activity was observed. The drug was able to enhance the DTH-response against SRBC and to stimulate the humoral immune response against Tetanus toxoid and heat-killed E. coli. In the popliteal lymph node assay, a murine graft-vs-host model, a stimulating effect of the substance was observed when it was administered at the same time of the grafts to rats. These results demonstrate that tiprotimod is a potent immunopotentiator for both humoral and cell mediated immune response in experimental animals.


Subject(s)
Adjuvants, Immunologic/pharmacology , Animals , Antibody Formation/drug effects , Cell Survival/drug effects , Culture Techniques , Female , Hypersensitivity, Delayed/immunology , Immunoglobulin G/immunology , Luminescent Measurements , Lymph Nodes/drug effects , Lysosomes/drug effects , Lysosomes/enzymology , Male , Mice , Nitroblue Tetrazolium , Oxidation-Reduction , Rats , Thiazoles/pharmacology
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