Laboratory of lipid chemistry

Laboratory studies a relationship between a structure and a function of lipids – the main component of the cell membrane. Basing on the study researchers create lipid conjugates with biologically active molecules (including drugs), surfactants, lipids which have reporter groups (lipid probes are unique instruments for scientific research) etc.

Fluorescent and photoaffinity lipid probes, designed taking into account the properties of lipid membranes, are developing by use of effective synthetic schemes. Today researchers are creating advanced drug delivery systems for nanomedicine.

The laboratory is equipped with everything necessary for analysis (HPLC and GC) and chemical synthesis of lipids (including rare devices for oxidation by singlet oxygen and ozonation).

In the field of synthesis, isolation and purification, study, and most importantly - the use of substances derived Lipid Laboratory collaborates with other laboratories of the IBCh RAS, as well as with the Moscow Technological University (MTU), Moscow State University (MSU), The Pirogov Russian National Research Medical University  (RNRMU), Saratov and Nizhny Novgorod Universities, N.F. Gamaleya Research Institute for Epidemiology and Microbiology, Gel'mgol'ts Institute of Ocular Diseases, N.N. Blokhin Russian Cancer Research Center, University of Umea (Sweden), Hormel Institute (USA), University of Liège (Belgium) and others.

Laboratory was created in 1963, just four years after the founding of the Institute. Lev D. Bergelson headed a new Laboratory. He was able to hold a tremendous amount of work to systematize methods of isolation of lipids from natural sources, study the structure and develop the chemical synthesis of lipids and their analogues. He also made a serious progress in the field of studying the structure and function of biological membranes, the role of lipids in various pathologies. In 1991 the Laboratory was headed by Julian G. Molotkovsky who contributed greatly to the development of fluorescent lipid probes. Since 2009 the Laboratory headed by Elena L. Vodovozova is mainly engaged in a pharmaceutical development.

Experience accumulated over 50 years, in fact, created over the years a scientific school, allows to finding unconventional solutions in the field of lipid chemistry and successfully deal with the challenges.

• Creating nanoscale systems of targeted delivery drugs based on liposomes, lipophilic prodrugs and lipophilic glycoconjugates (Figs. 1 and 2). Targeted liposomes surpassed original drugs and liposomes free of a carbohydrate ligand, in the antitumor effect (Fig. 3).

• Synthesis of new fluorescent lipid probes; studying structure and functions of membranes, and patterns of excitation energy transfer between the fluorophores using these probes (Figs. 4 and 5).

• Was realized the synthesis of unsaturated natural fatty acid based on stereochemistry of the Wittig reaction.

• Was found the phenomenon of dedifferentiation the lipid composition of organelles of cancer cells (L.D. Bergelson, E.V. Dyatlovitskaya).

• Was studied a spread and a role of a new class of diol lipid (L.D. Bergelson and V.A. Wawer).

• Was synthesized and proven a structure of bacterial lipoamino acids and performed the synthesis of unsaturated phosphatidylinositol (L.D. Bergelson and J.G. Molotkovsky).

• Was studied topology of membranes using NMR-spectroscopy (L.D. Bergelson and L.I. Barsukov in collaboration with V.F. Bystrov).

• Was discovered and proved the structure of a new class of bacterial ornithine-containing lipids (L.D. Bergelson and S.G. Batrakov).

• Was synthesized and used in biological studies a large range of fluorescent and photoaffinity lipid probes (L.D. Bergelson, J.G. Molotkovsky, E.L. Vodovozova, I.I. Mikhalyov, I.A. Boldyrev).

• Was developed a targeted liposomal delivery to the tumor of lipid-modified anti-cancer agents (J.G. Molotkovsky, E.L. Vodovozova, G.P. Gayenko in cooperation with N.V. Bovin).

• Was shown that the developed liposomal drugs are hemocompatible (N.R. Kuznetsova, E.L. Vodovozova in collaboration with the University of Liège and Nanomedicine Center in Budapest).

Was shown in vivo (in the model of Lewis lung carcinoma) that the antitumor effect of the 100 nm liposomes designed on the basis of natural phospholipids and lipophilic melphalan prodrug and bearing selectin ligand sialyl Lewis X (SiaLeX) tetrasaccharide, was caused by the specific antivasсular effect in tumor tissue (E.L. Vodovozova, N.R. Kuznetsova in cooperation with N.V. Bovin and N.N. Blokhin Russian Cancer Research Center).

• Was shown (on the model of human umbilical vein endothelial cells) that SiaLeX-liposomes loaded with a lipophilic melphalan prodrug selectively deliver drug to cells activated by tumor necrosis factor alpha (TNF-α) (A.S. Alekseev, E.L. Vodovozova in cooperation with the Institute N.F. Gamaleya Research Institute for Epidemiology and Microbiology).

• Was studied the function of the reopened ceramide-1-phosphate-transporting protein (C1P), widely distributed in mammalian tissues using a set of fluorescently-labeled phospholipids and glycolipids. Was shown that C1P modifies phospholipase A2 activity and mediates the biosynthesis of eicosanoids (J.G. Molotkovsky in collaboration with the Institute of Hormel and others).

Fig. 1. Structures of lipophilic prodrugs and glycoconjugates.

Fig. 2. Electron micrographs of the replicas of freeze fracture surfaces of (A, a) MTX-DG- and (B, b) Mlph-DG-containing liposomes.

Fig. 3. Weekly survival dynamics of BLRB mice with grafted mammary adenocarcinoma in different experimental groups (n=10). Mice were treated iv on the 3rd and 7th days after tumor cells inoculation. Groups: 1 — merphalan (sarcolysine); 2 — empty liposomes; 3 — liposomes + prodrug; 4 — liposomes + prodrug + SiaLeX-conjugate; 5 — liposomes + SiaLeX-conjugate; Control — physiological solution.

Fig. 4. Set of fluorescent probes for membrane studies across the bilayer. Plots in grey — order parameter profiles for the set in bilayers of different composition.

lipchem

Fig. 5. Structure of the BODIPY–FL–C3–GM1 ganglioside, the fluorescent raft marker.

Current Grants:

RNF № 14-15-00128 “«Gate» of blood-brain barrier: mechanisms of regulation, their dependence on the state of the body and age, methods of correction by supramolecular transport systems". Performers:  E.L. Vodovozova; leader institution: Saratov State University.

RFBR № 15-04-07415-a "Study of lipid-protein interactions in biological membranes and cells with a fluorescent lipid probes". Head: J.G. Molotkovsky.

RFBR № 16-04-01585-a "Studies of the fate of antitumor liposomes loaded in the bilayer with lipophilic prodrug of methotrexate in human plasma". Head: E.L. Vodovozova.

RFBR № 15-33-20523 "New fluorescent indicators of activity and substrate specificity of different types of phospholipase A2". Head: I.A. Boldyrev.

RFBR № 16-34-01237 "Study the mechanisms of penetration into cells and intracellular trafficking of liposomal forms of lipophilic methotrexate prodrug". Head: A.S. Alekseeva.

NamePositionContacts
Elena Vodovozova, D.ScheadElvod@ibch.ru+7(495)330-66-10
Julian Molotkovsky, D.Sc, professorpr. r. f.jgmol@ibch.ru+7(495)330-66-01
Ivan Boldyrev, Ph.D.s. r. f.ivan@lipids.ibch.ru+7(495)330-66-10, +7(926)224-68-06
Galina Gayenko, Ph.D.s. r. f.GPG008@mail.ru+7(495)330-66-83
Ilya Mikhalyov, Ph.D.s. r. f.Ilya.Mikhalyov@gmail.com+7(495)330-66-10
Anna Vostrova, Ph.D.r. f.anna.vostrova@gmail.com+7(495)330-69-74
Natalia Onishchenko (Kuznetsova), Ph.D.r. f.natalia@lipids.ibch.ru+7(495)330-66-10
Anna Alekseeva, Ph.D.j. r. f.anna@lipids.ibch.ru+7(495)330-66-10
Daria TretiakovaPhD stud.daria@lipids.ibch.ru+7(495)330-66-10
Ekaterina RyabukhinaPhD stud.miss.katringet@yandex.ru
Svetlana Volkovat. q. - lab. as.+7(495)335-32-00

Former members:

Lev Bergelson, corresponding member of the academy of scienceshead
Galina Zhukovat. q. - lab. as.

All publications (show selected)

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Elena Vodovozova

  • Russia, Moscow, Ul. Miklukho-Maklaya 16/10 — On the map
  • IBCh RAS, build. 34, office. 532
  • Phone: +7(495)330-66-10
  • E-mail: Elvod@ibch.ru

Enzyme-responsive liposomes with phospholipidic colchicinoids

New phospholipidic prodrugs of colchicinoids were synthesized for incorporation into bilayer of enzyme-responsive liposomes. The prodrug design takes into account the structure of the substrate-binding site of target enzyme, phospholipase A2 (PLA2), and the lateral pressure profile inside the bilayer. This allowed for minimal distortions of the lipid packing by the colchicinoid prodrugs and submicromolar cytotoxicity of the liposome carriers.

Publications

  1. Shchegravina ES, Tretiakova DS, Alekseeva AS, Galimzyanov TR, Utkin YN, Ermakov YA, Svirshchevskaya EV, Negrebetsky VV, Karpechenko NY, Chernikov VP, Onishchenko NR, Vodovozova EL, Fedorov AY, Boldyrev IA (2019). Phospholipidic Colchicinoids as Promising Prodrugs Incorporated into Enzyme-Responsive Liposomes: Chemical, Biophysical, and Enzymological Aspects. Bioconjug Chem 30 (4), 1098–1113

Influence of stabilizing components in the lipid bilayer on the integrity of antitumor liposomes loaded with lipophilic prodrug, in human serum

In collaboration with Laboratory of Carbohydrates

We compared the effect of different amphiphiles in lipid bilayer on the integrity of 100-nm-liposomes loaded with lipophilic prodrug of chemotherapeutic agent melphalan, in human serum. Using fluorescence methods phosphatidylinositol was shown to protect fluid phase lipid bilayer based upon egg phosphatidylcholine at least for 4 hours, while ganglioside GM1 or a conjugate of carboxylated oligoglycine with phosphatidylethanolamine up to 24 hours. At the same time, polyethylene glycol (2000 Da) conjugated with dipalmitoylphosphatidylethanolamine (PEG-lipid) promoted degradation of liposomes, so that lipids began to exit fluid phase membrane, while gel phase membrane with less than 10 mol % of PEG-lipid was immediately cracked. Cholesterol-containing bilayers of condenced liquid ordered phase, supplemented with sufficient amounts of the PEG-lipid, showed good stability in serum. The above effects should be accounted when using lipophilic conjugates of PEG in the composition of supramolecular drug delivery systems devoid of covalent bonds, such as liposomes, lipid nanospheres, or micelles.

New fluorescent probes to research the structure and functions of membranes

In collaboration with Laboratory of molecular toxinology

The use of anthrylvinyl-perylenoyl FRET-pair of phospholipid probes revealed the existence of regulatory interaction site(s) on the surface of ceramide-1 phosphate transfer protein that are specific to the polar head groups of phosphoglycerides in the lipid membrane. This finding delineates new differences between Glycolipid Transfer Proteins superfamily members that are specific for C1P versus glycolipid [1]. By means of new BODIPY FRET-pair of phosphatidylcholine probes, it was shown that heterodimeric V. nikolskii phospholipases A2 induce aggregation and stacking of negatively charged lipid bilayers [2]; this may be one of the mechanisms of PLA2 biological activity. A novel combination of FRET between BODIPY-ganglioside probes and Monte Carlo simulations (MC-FRET) identified directly 10 nm large nanodomains (rafts) composed of sphingomyelin and cholesterol in liquid-disordered model membranes that mimic the cytoplasmic membrane; the nanodomains are also fluid and disordered [3].

Publications

  1. Alekseeva AS, Tretiakova DS, Chernikov VP, Utkin YN, Molotkovsky JG, Vodovozova EL, Boldyrev IA (2017). Heterodimeric V. nikolskii phospholipases A2 induce aggregation of the lipid bilayer. Toxicon 133, 169–179
  2. Zhai X, Gao YG, Mishra SK, Simanshu DK, Boldyrev IA, Benson LM, Bergen HR, Malinina L, Mundy J, Molotkovsky JG, Patel DJ, Brown RE (2017). Phosphatidylserine stimulates ceramide 1-phosphate (C1P) intermembrane transfer by C1P transfer proteins. J Biol Chem 292 (6), 2531–2541
  3. Koukalová A, Amaro M, Aydogan G, Gröbner G, Williamson PTF, Mikhalyov I, Hof M, Šachl R (2017). Lipid Driven Nanodomains in Giant Lipid Vesicles are Fluid and Disordered. Sci Rep 7 (1), 5460

Liposomal formulation of a methotrexate lipophilic prodrug: interactions with tumor cells and studies in vivo

In collaboration with Laboratory of biotechnology

Previously, we developed a formulation of widely used cytostatic agent methotrexate incorporated in the lipid bilayer of 100-nm liposomes in the form of diglyceride ester (MTX-DG, lipophilic prodrug). Here, we first studied interactions of MTX-DG liposomes with various human and mouse tumor cell lines using fluorescence techniques. Liposomes were labeled with fluorescent analogues of phosphatidylcholine and MTX-DG. Carcinoma cells accumulated 5 times more MTX-DG liposomes than the empty liposomes. Studies with inhibitors of liposome uptake and processing by cells demonstrated that the formulation utilized multiple mechanisms to deliver the prodrug inside the cell. According to our data, undamaged liposomes fuse with the cell membrane only 1.5–2 h after binding to the cell surface and then the components of liposomal bilayer enter the cell separately. The study of the time course of plasma concentration in mice showed that the AUC (area under the curve) of methotrexate generated upon intravenous injection of MTX-DG liposomes exceeded that of intact methotrexate 2.5-fold. These data suggested the advantage of using liposomal formulation to treat systemic manifestation of hematological malignancies. Indeed, administration of MTX-DG liposomes to recipient mice bearing T-cell leukemic lymphoma using a dose-sparing regimen (only four low to middle-dose injections) resulted in lower toxicity and retarded lymphoma growth rate as compared to methotrexate.