Molecular and Cellular Pharmacology
Membrane interaction of an antitumor antibiotic, mithramycin, with anionic phospholipid vesicles

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Abstract

Small unilamellar vesicles (SUV) composed of zwitterionic phosphatidylcholine and two different anionic phospholipids, phosphatidic acid and phosphatidylserine, in different compositions, were employed to study the membrane interaction of an antitumor antibiotic, mithramycin (MTR). Binding of MTR to dimyristoylphosphatidylcholine (DMPC) liposomes containing the anionic phospholipid dimyristoylphosphatidic acid (DMPA) was estimated by measuring the increase in intensity of the intrinsic fluorescence of MTR with increasing concentrations of phospholipids. Membrane perturbations were observed in acidic SUV of DMPC/DMPA and DMPC/bovine brain phosphatidylserine by MTR and its magnesium complex as studied by monitoring the leakage of the entrapped fluorescent marker carboxyfluorescein and by electron microscopic measurements of the size of the liposomes. These results indicated a possible role of anionic phospholipids in mediating binding of MTR and its magnesium complex to the cell surface membranes before reaching the target DNA.

Section snippets

Materials and methods

MTR, MgCl2 solution (4.9 mol/L), DMPC, PS, CF, Sephadex G-50, Triton X-100, and Tris were purchased from the Sigma Chemical Co. DMPA was from Fluka. Solvents used were of spectroscopic grade. Deionized water from a Milli Q source (Millipore Corp.) was used for preparation of the buffer and all other solutions. (MTR)2Mg2+ complex was prepared by incubating 10–20 μM MTR with 10 mM Mg2+ for 1 hr.

Fluorescence measurements were performed in a Shimadzu RF-540 spectrofluorometer using both 2- and

Results

Fluorescence emission spectra of MTR both in the aqueous buffer and in two different concentrations of DMPC/DMPA SUV are shown in Fig. 1. Maximum emission of MTR was at 536 nm in aqueous buffer; it was blue-shifted by 2 nm in phospholipid vesicles, emitting maximally at 534 nm. Enhancement of the intensity of MTR fluorescence was observed in the presence of increasing concentrations of the phospholipid vesicles. Similar enhancement of fluorescence intensity also was observed when phospholipid

Discussion

The antitumor activities of antibiotics such as actinomycin, chromomycin, and the anthracycline group of antibiotics are often attributed directly to their interaction with DNA 28, 29. However, studies with other antitumor antibiotics, such as doxorubicin, indicate that the drug can be cytotoxic even without entering the cells 30, 31. A growing body of literature indicates that the cytotoxicity could be related directly to drug–membrane and in particular to drug–lipid interactions [31]. Studies

Acknowledgements

We thank Dr. Dipak Dasgupta for his keen interest in this work and a critical reading of the manuscript. We also thank Dr. Amitabha Chattopadhyay of CCMB for many fruitful suggestions in the preparation of the manuscript. We acknowledge both scientific and technical support from the electron microscope facility of SINP. Sangita Majee acknowledges a Senior Research Fellowship from CSIR, India.

References (39)

  • T.G Burke et al.

    Ligand self-association at the surface of liposomesA complication during equilibrium-binding studies

    Anal Biochem

    (1984)
  • G.S Karczmar et al.

    The interaction of adriamycin with small unilamellar vesicle liposomes. A fluorescence study

    Biochim Biophys Acta

    (1979)
  • G.F Gause

    Olivomycin, chromomycin and mithramycin

  • Calabresi P and Parks RE Jr, Chemotherapy of neoplastic diseases. In: Goodman and Gilman’s The Pharmacological Basis of...
  • Calabresi P and Chabner BA, Chemotherapy of neoplastic diseases. In: Goodman and Gilman’s The Pharmacological Basis of...
  • W.A Remers

    Aureolic acid group of antibiotics

  • D.C Ward et al.

    Base specificity in the interaction of polynucleotides with antibiotic drugs

    Science

    (1965)
  • B.J Kennedy et al.

    Effect of mithramycin on a mouse glioma

    Cancer Res

    (1968)
  • P Aich et al.

    Role of magnesium ion in mithramycin-DNA interactionBinding of mithramycin-Mg2+ complexes with DNA

    Biochemistry

    (1995)
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