Hans Brachwitz

Analogs of alkyllysophospholipids: Chemistry, effects on the molecular level and their consequences for normal and malignant cells

In the search for new approaches to cancer therapy, the first alkyllysophospholipid (ALP) analogs were designed and studied about two decades ago, either as potential immunomodulators or as antimetabolites of phospholipid metabolism. In the meantime, it has been demonstrated that they really act in this way. However, their special importance is based on the fact that, in addition, they interfere with key events of signal transduction, such as hormone (or cytokine)-receptor binding or processing, protein kinase C or phospholipase C function and phosphatidylinositol and calcium metabolism. There are no strict structural requirements for their activity. Differences in the cellular uptake or the state of cellular differentiation seem to be mainly responsible for higher or lower sensitivities of cells towards ALP analogs. Consequences of the molecular effects mentioned on the cellular level are cytostasis, induction of differentiation (while in contrast the effects of known inducers of differentiation such as 12-O-tetradecanoylphorbol-13-acetate are inhibited, probably as a consequence of protein kinase C inhibition) and loss of invasive properties. Already in sublytic concentrations, alterations in the membrane structure were observed, and lysis may begin at concentrations not much higher than those causing the other effects described. Few ALP analogs have already entered clinical studies or are in clinical use. ALP analogs are the only antineoplastic agents that do not act directly on the formation and function of the cellular replication machinery. Therefore, their effects are independent of the proliferative state of the target cells. Because of their interference with cellular regulatory events, including those failing in cancer cells, ALP analogs, beyond their clinical importance, are interesting model compounds for the development of new, more selective drugs for cancer therapy.

Halo lipids. V. Synthesis, nuclear magnetic resonance spectra and cytostatic properties of halo analogues of alkyllysophospholipids☆☆☆

Abstract A series of fluorine- and chlorine-containing analogues of alkyllysophospholipids has been synthesised as potential antimetabolites of phospholipids. These compounds include various structural isomers of racemic long-chain alkyldeoxyhaloglycerophosphocholines and N,N-dimethylethanolamines, alkyldeoxyhaloglycerophosphoric acids, and alkyl esters. 1H- and 19F-NMR spectroscopic data are presented and analysed. Alkyldeoxyhaloglycerophosphocholines were found to exhibit a strong inhibitory effect on the proliferation of Ehrlich ascites carcinoma cells in vitro.

Alteration of DNA topoisomerase II activity during infection of H9 cells by human immunodeficiency virus type 1 in vitro: A target for potential therapeutic agents

Infection of H9 cells with human immunodeficiency virus type 1 (HIV-1) was found to decrease the phosphorylation of DNA topoisomerase II during the initial phase of infection. Simultaneously, with a later overshoot of phosphorylation and the subsequent activation of DNA topoisomerase II, the production of HIV-1 started. Applying three new protein kinase C inhibitors from the class of O-alkylglycerophospholipids we demonstrated that inhibition of protein kinase C-mediated phosphorylation of DNA topoisomerase II resulted in an inhibition of HIV-1 production. Based on the differential effect of the two protein kinase C activators, phorbol ester and bryostatin, we conclude that phosphorylation of DNA topoisomerase II is mediated by the form alpha and gamma of protein kinase C. These data suggest that agents which inhibit these two forms of protein kinase C are also potential candidates for an anti-HIV therapy.

Cytostatic Activity of Synthetic O-Alkylglycerolipids'

A series of halogen-containing alkylglycerolipid analogs has been checked for their cytostatic activity both in vitro and in vivo. The compounds included alkyldeoxyhaloglycerols (I), alkyldeoxyhaloglycerophosphocholines (II), and alkyldeoxyhaloglycerophosphoric acids and alkyl esters (III). While compounds I and III were moderately active, compounds II were found to have a strong inhibitory effect on the proliferation of Ehrlich ascites tumor cells in vitro. Cell growth inhibition of 50% or more was found mainly in the late S- or G2-phase of the cell cycle as revealed by flow cytometry. Alkyl lysophospholipid analogs II and cholesterol form liposomes with high encapsulation efficiency and low permeability for entrapped substances. Compounds II were active against Lewis lung carcinoma in mice when applied in free form or as liposomes.

Halo lipids. 7. Synthesis of rac 1-chloro-1-deoxy-2-O-hexadecylglycero-3-phosphocholine

Abstract In order to obtain detailed data on the structure-activity relationship of cytostatically active alkylglycerophosphocholine analogues, rac 1-chloro-1-deoxy-2-O-hexadecylglycero-3-phosphocholine was synthesised via 2-O-alkyl-1-chloro-1-deoxyglycerol and (2-O-alkyl-1-chloro-1-deoxyglycero-3)-2-bromoethyl hydrogen phosphate.

Halo lipids, 8. Synthesis of O-alkyl-O-alkyl-O-(2,2,2-trifluoroethyl)glycerols and O-alkyl-O-(2,2,2-trifluoroethyl)glycero-phosphocholines☆

Abstract The syntheses and structure determination of 3 positional isomers of O-alkyl-O-(2,2,2,-trifluoroethyl)glycerols and their conversion into the corresponding O-alkyl-O-(2,2,2-trifluoroethyl)glycerophosphocholines via the [O-alkyl-O-(2,2,2-trifluoroethyl)glycero]-2-bromoethyl hydrogen phosphates are described.

Modulation of the cytosolic Ca+ + concentration by alkylphospho-l-serine analogs: relation to their antiproliferative action

Antiproliferative alkyllysophospholipid (ALP) analogs produced multiple effects on the cytosolic Ca++ concentration ([Ca++]i) in an immortalized human breast epithelial cell line (H 184). The addition of small concentrations resulted in a short transient [Ca++]i response. With higher concentrations the transient rise was followed by a sustained increase. Pretreatment of cells with the ALP analogs for two minutes inhibited the transient [Ca++] response. Increases in [Ca++]i and inhibition of the transient increase were studied in relation to the dose and structure of several ALP analogs. In a series of alkylphospho-L-serine analogs with different lengths of the alkyl chain we found different dependencies of the stimulatory and inhibitory effects on the dose and the structure. The ability to increase [Ca++]i is absent with the C14 and C15 analogs, is low with the C16 and high with the C18 analog. With the exception of the C12 analog, a dose-related inhibition was observed with all derivatives but the effective concentrations differed very strongly and the maximal potency was reached with the C15 and C16 analogs. The antiproliferative action seems to correlate rather with the potency to inhibit the transient [Ca++]i response than with its stimulation.

SYNTHESIS AND CYTOSTATICAL EVALUATION OF CYTIDINE- AND ADENOSINE-5'-HEXADECYLPHOSPHATE AND THEIR PHOSPHONATE ANALOGS

Abstract Four phospholipid conjugates containing the non-cytotoxic nucleosides cytidine and adenosine were prepared by condensation reactions, and their cytotoxic activity was tested in vitro against the human immortalized mammary epithelial cell H184 A1N4, the human mammary tumor cells MaTu and MCF7 and the B lymphoblast cell line Daudi. The synthesized compounds showed considerable activity towards H184 A1N4, MaTu and Daudi cells, but they were not effective against MCF7 cells. The phosphorus moiety—either monophosphate or monophosphonate—does not influence the effectiveness of the phospholipid derivatives in the case of the solid tumor cell lines and H184 A1N4. The leukemic Daudi cell line is strongly sensitive towards the different types of ester as well as to the type of the nucleoside component. Adenosine-5′-hexadecylphosphate proved to be the most potent compound among the substances prepared (IC50: 9.0 μmol).