The disadvantages of protein drugs lie in short circulation time (blood) and immunological side effects ranging in severity from mild allergic reaction to anaphylactic shock. Chemical modification of protein drugs with synthetic macromolecules can eliminate some of drawbacks of the proteins and give them new functions. Non toxic and non-immunogenic synthetic polymer, polyethylene glycol (PEG) such as 2-[O-methoxypoly(ethylene glycol)]-4, 6-dichloro-s-triazine (activated PEG₁) or 2, 4-bis[O-methoxy-poly(ethylene glycol)]-6-chloro-s-triazine(activated PEG₂), has been extensively used as a modifying reagent. Recently, PEG-derivatives with a comb-shaped form were explored by us, which are copolymers of polyethylene glycol derivative and maleic anhydride, with the molecular weight of 13, 000 and 100, 000 (activated PM₁₃ and activated PM₁₀₀). The modifications of asparaginase (136 kD) and bovine serum albumin (BSA, 67 kD) by PEGs (activated PEG₁ and activated PEG₂) with chain shaped-form and PMs(activated PM₁₃ and activate PM₁₀₀) with comb-shaped form are introduced in relation to the physiological and immunological functions. Both PM₁₀₀-asparaginase and PM₁₀₀-BSA have the reduced immunoreactivities with lower degree of modifications and with higher enzymic activities in comparison with PM₁₃- or PEG-asparaginases and PM₁₃- or PEG-BSAs. Furthermore, the half-life of asparaginases modified with both activated PEG₂ and PM₁₀₀ is prolonged about 20 times in comparison with non-modified asparaginase. General advantages of PEG-protein drugs seem to be the reduction of immunogenicity or antigenicity, prolongation of clearance time, increasing of solubility, and high stability in biomedical field. Now, more than 50 proteins have been modified with PEG or PEG derivatives for the purpose of therapy of various diseases.

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Superoxide dismutase (SOD), an enzyme that scavenges superoxide anion (O_2^-), has been suggested as a possible therapeutic tool, for treating diseases related to active oxygen spiecies. However, its low cell membrane affinity and rapid metabolic clearance limit its clinical application. We synthesized SOD, in which cytotropic phosphatidylcholine derivatives were covalently bound to recombinant human SOD, in order to target SOD to cell membrane. Lecithinized SOD was shown to have a high cell membrane affinity using laser confocal imaging technique, showed a longer plasma half-life, and efficiently scavenged O_2^- produced by phorbol myristate acetate(PMA). Lecithinized SOD showed more potent inhibitory effect on human endothelial cells injury by O_2^- produced by PMA stimulated neutrophils and on ischemia-reperfusion mouse paw edema than, that of unmodified SOD. Moreover, other potent pharmacological activities of lecithinized SOD has been reported, and toxicological and antigenic findings of lecithinized SOD had no particullar in comparison with unmodified SOD. Accordingly, lecithinized SOD seems to have the potential for clinical applications.

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Generally, the oral absorption of peptide and protein drugs is typically poor. This has been attributed to their extensive hydrolysis and poor membrane permeability characteristics. To overcome these delivery problems, we synthesized lipophific derivatives of peptides and proteins such as tetragastrin, insulin, human calcitonin and lysozyme by chemical modification with various fatty acids. A significant increase in the intestinal absorption of these lipophilic peptides was observed in comparison with native peptides. In addition, these derivatives were more stable than the parent peptides in homogenates of the various intestinal mucosae Furthermore, the mechanisms whereby intestinal absorption and stability of these peptides was improved by acylation were discussed in this review article. In summary, this chemical modification approach may be useful to improve the intestinai absorption of peptides and proteins.

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“On-off” regulation of drug release in response to temperature changes as a command from the exterior of body has been achieved using temperature-responsive copolymer geis of N-isopropylacrylamide(IPAAm). In the design of the “on-off” regulating system using these thermosensitive gels, the phase transition must be maintained within temperature values which meet external temperature changes. Design of the gel structures with phase transition temperatures greater than that of poly (IPAAm) (32°C) are required for application in the human body. In this study, both phase transition and thermosensitivity of poly (IPAAm) gels have been controlled by introduction of comonomers into the gel. Hydrophobic alkyl methacrylate, hydrophilic acrylamide and N, N'-dimethylacrylamide have been used as comonomers. Effects of these comonomers on the phase transition temperature, the thermosensitivity and the skin formation process with increasing temperature have been investigated. By controlling these gel properties, the switching temperature of drug release has been controlled to achieve “on-off” regulation in response to small temperature changes around body temperature. Actually, complete “on-off” drug release for the temperature changes between 36 °C and 38 °C was achieved.

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Amino acids are amphoteric electolytes and water-soluble compounds. In general, a water-soluble drug such as amino acid shows low skin permeability due to the barrier effects of the stratum corneum. To attempt to improve the percutaneous absorption of amino acid, we investigated the skin permeability of L-Phenylalanine and its ester derivatives as model compounds. L-Phenylalanine derivatives used in this study were L-Phenylalanine methyl ester, ethyl ester, t-butyl ester and benzyl ester. Excised hairless mouse skin was mounted between the donor and the receptor chamber of a skin-permeation apparatus. The compound that permeate through the skin was analyzed by HPLC. The permeability of L-Phenylalanine and its ester derivatives was calculated from the accumulated amount of the sample in the receptor chamber for the specific time period (2, 4, 6, 8, 24, and 28hr). The results showed that (1) the ester derivatives which permeated through the skin were detected as Phenylalanine and (2) the permeation rate of L-Phenylalanine, the methyl ester, the ethyl ester, the t-butyl ester, and the berizyl ester were 0.10, 7.22, 13.63, 22.51, and 16.88 μg/cm²/hr, respectively. All the ester derivatives showed higher permeability through the skin than that of L-Phenylalanine (71∼220 times). The results indicated that (1) the permeability of the derivatives depended on its ester type and (2) the esterification of L-Phenylalanine could improve its permeability through the skin. Since no derivative could be detected in the receptor chamber, the derivatives, which could permeate the stratum corneum, were hydrolyzed quickly by the esterase in skin tissue. Because the esterification of amino acids suppresses its ionization, enhances its hydrophobicity, and improves its permeability to skin tissue, the amino acid derivatives (e.g. ester form) is useful a.a prodrug that could be applied to skin.

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Cepharanthine (CEP), a biscoclaurine alkaloid, is reported to be effective in prevention of immune suppression and leukopenia induced by the administration of antitumor agents. It is known that CEP inhibits a function of P-glycoprotein, and expected to reduce the multiple drug resistance of tumor cells. We reported some physico-chemical characteristics of CEP containing liposomes already. In this paper, we studied whether the efficacy of immuno-modulating action of CEP will be enhanced by using liposome as a drug carrier. Effects of liposome type and composition which contained CEP on the antigen-specific IgM production in mice were investigated. Liposomes composed of egg phosphatidylcholine and cholesterol in a molar ratio of 7 : 2. Multilamellar liposomes(MLV) were prepared by vortexing dried lipid films, and small unilamellar liposomes (SUV) were prepared by ultrasonication of MLV. Antigen(pneumococcal polysaccharides) and CEP-containing liposomes were injected i.p. to BALB/C mice. Blood was collected to obtain plasma, and the primary immune responsed was examined. The specific IgM antibody against antigen in plasma was measured by ELISA. Liposomes containing CEP enhanced the antigen-specific IgM antibody production compared to free CEP or vehicle liposomes. CEP contained in MLV enhanced specific IgM antibody production against pneumococcal polysaccharides more effectively than CEP contained in SUV. The increase in specific IgM antibody production was influenced by both the type and composition of liposomes. These results suggest that the efficacy of immuno-modulating action of CEP is enhanced by using appropriate liposome as a drug delivery system.

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Colon delivery capsule which disintegrates by inner pressure in the colon has been developed. Its efficiency was evaluated in beagle dogs using recombinant human granulocyte-colony stimulating factor(G-CSF) and 5-aminosalicylic acid (5-ASA) as model drugs. An oral gelatin capsule of which inside surface was coated with ethylcellulose (EC) was prepared for this purpose. Several kinds of EC-gelatin capsules having the thickness of EC layer from 46 to 221 μm, were prepared. As a content, propylene glycol solution containing polyoxyethylated, 60 μmol, castor oil derivative, citric acid and fluorescein, as an absorption marker, was fulfilled in the capsule. To measure the arrival time to the colon, the experiment using suifasalazine was performed. It was found that our new capsule arrived at the colon within 4∼5hr after oral administration. When EC capsule of thin thickness was administered to beagle dogs, it disintegrated within 2∼3hr after administration. In the case of 84 μm thickness capsule, it disintegrated within 4∼5hr after administration, which suggests its disintegration in the colon. After the administration of this capsule filled with G-CSF solution, 25 μg/kg, the maximum blood total leukocytes count (BTL) appeared at 10hr. Thereafter, the BTL count gradually decreased and returned to its normal level at 48hr. This result suggests the possibility of new oral dosage form of protein drug. Furthermore, after the administration of 5-ASA, 5-ASA began to appear in the blood at 4∼5hr after administration. With this dosage form, it will be easy to control the dosage of 5-ASA without suffering from the side effect of suifapyridine. Therefore, this new capsule is thought to be safe and useful for protein drugs and drugs for colonic disease.

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This study explored the effect of temperature on the diffusion and partition of nitroglycerin in the hairless mouse skin in vitro. The diffusion coefficient of the drug in the stratum corneum and viable skin agreed with the Arrhenius relationship with the activation energy of 69.1 (kJ/mol) and 46.1 (kJ/mol), respectively. The partition coefficient remained almost constant within the temperature range varied (12∼42 °C). The step change in the eenvironmental temperature from +10 °C to -20 °C range required about 0.6 hours before the skin permeation reached a steady state.

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