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Galkin O.Yu.

National Technical University of Ukraine “Kyiv Polytechnic Institute”, Ukraine

Monoclonal antibodies and directED Drug delivery systems

Depending on cell mechanism of macromolecules and particles absorption, endocytosis could be divided into constitutive (or liquid-phase) and receptor-dependent. In the first case a nonselective process takes place, during which the concentration of substances that are absorbed as a part of vesicles corresponds to the concentration of substances in the extracellular liquid. Receptor–dependent endocytosis is exclusively selective concentrating mechanism that allows cells to absorb big amounts of specific ligands without absorbing big volume of extracellular liquid.

Monoclonal antibodies (mAb) to different receptors on the cancer cells surface (TRFR, epithelium growth factor receptor, CD-receptors) are the most wide-used cargo molecules. Production of such mAbs underlies the oncovaccine action. Moreover, the radiolabeled mAbs, which bond with cell and cause its death because of presence of radionuclides in their structure, are sometimes used. Also mAbs are often conjugated with cytotoxic antibiotics and other bioactivity substances by different linkers (avidin-biotin, PEG, etc.). Use of conjugates based on mAb to receptors is often more effective than use of natural ligands as cargo molecules. For example, Tfr is characterized by limited ability to permeate through hematoencephalic barrier, that doesn’t allow to use Tfr in the therapy of brain oncological diseases. At the same time antibodies to TRFR freely permeate to the different brain tissues.

An original tendency in creation of directed drug delivery systems is immunoliposomes – liposomes with attached monoclonal antibodies. Monoclonal antibodies provide specific binding of liposomes with antigen-positive cells, while liposomes carry conforming hydrophobic or hydrophilic chemotherapy agent.

Nowadays three types of immunoliposomes: A, B and C are existed. In the A type immunoliposomes, mAbs are covalently bound with normal liposomes by short linker. B type is presented by PEG liposomes which are covalently bound with mAb by short linker. Type C (Pendant-type PEG-immunoliposomes) is steric stabilized PEG-liposomes with mAb attached to distal terminal PEG end.

By means of type A liposomes it was shown that immunoliposomes are more effective in delivering drugs to target cells comparing with normal liposomes in in vitro and in vivo tests. But liposomes binding with target cells in vivo was more difficult. Studying immunoliposomes in vivo showed that attaching antibodies to liposomes strengthened their absorption by RES mononuclear leukocytes. Effectivity of liposomes adhesion with target cells depended on antibodies thickness on the liposomes surface. Immunoliposomes absorbtion by RES cells and endothelial barrier compelled the scientists to create a new type of liposomes. It leaded to the steric stabilized immunoliposomes with the prolonged period of circulation in the blood construction.

In the earliest works of creating the long-persistent immunoliposomes, the antibodies were bound by short hydrophilic linker close to liposomes type B surface; these steric stabilized liposomes contained phospholipids with PEG modified main groups. The liposomes kept ability of long circulation but interaction with target cells was suppressed by PEG blockade.

Later mAb were attached to distal ends of PEG chains bound with type C liposomes. It resulted in keeping the steric stabilized liposomes ability to bind specifically with target cells surface and ability to be protected from absorption by RES mononuclears.

Nowadays for the purpose of getting the stable connection of antibodies with PEG, three conjugation methods are used for covalent binding mAb and PEG terminal ends: by thioether bond, by amid groups and by hydrazones.

It should be noticed that there are some requirements to antibodies used in immunoliposomes construction. They should keep their specifity when conjugate with liposomes, have enough affinity for low concentration liposomes binding and have low immunogenity. With that end in view chimeric and humanizated mAb and Fab antibodies fragments are used. Antibodies should be intensively internalized by target cells through endocytosis, should have biological activity and intensify antitumor response. Monoclonal antibodies have to be manufacturable and have enough shelf life.

Also there are some requirements to the antigen which is the target for the liposomes. It should greatly and homogeneously express in the tumor tissue and not disappear from the cell surface. Antigen desquamation from the tumor cell surface should be minimal to avoid immunoliposomes and soluble antigen binding or clearance intensification. “Antigene-immunoliposome” complex should pinocytate into the tumor cell. The bond between antigens and liposomes should be stable in the blood. The linker shouldn’t bind with molecule site which is recognized by antigen, should be non-immunogenic, non-toxic, should avoid opsonization, not affect on the medical agent inside the liposome and liposome membrane stability and not make steric obstacles.

Specific delivery of antitumor agents by means of immunoliposomes contribute to the better therapeutical effectiveness and to toxicity decrease comparing with normal liposomes. It was convincingly showed on the mouse solid tumor models and on the nude mice heterologous grafts of human B-cell.

Several immunoliposome preparations perspective for use in oncological practice are described to the present day. They are directed against cells that express such antigens: CD71 (transferrin receptor), Her2/neu (epidermal growth factor receptor), HLA-DR (MHC II antigens), CD19 (B-cells marker), LL2 (B-cells lymphoma antigen).