Akira Makino

Enzymatic Polymer Synthesis: An Opportunity for Green Polymer Chemistry

Polymeric materials, natural and unnatural, are indispensable to the modern society. They are widely used from everyday life usages as commodity materials to industry and technology usages in the fields such as electronics, machinery, communications, transportations, pharmacy, and medicine as highly advanced materials. Today, it is hard to think of the present society without polymeric materials. Developments of various polymeric materials have been owed to epoch-making innovative works as exemplified typically by the discovery of Ziegler-Natta catalyst,1-3 the concept of living polymerization,4 the discovery of conducting polymers,5 and the discovery of metathesis catalyst.6,7 These observations demonstrate that new polymeric materials are often brought about by new production methods including polymerization catalysts. Historically, polymerization catalysts utilized classical catalysts of acids (Brønsted acids, Lewis acids, and various cations), bases (Lewis bases and various anions), and radical generating compounds since the 1920s, the early stage of polymer chemistry. In the following * To whom correspondence should be addressed. Fax/Tel: (+81)-75-7247688. E-mail: kobayash@kit.ac.jp. † Kyoto Institute of Technology and Emeritus Professor of Kyoto University. ‡ Kyoto University. Chem. Rev. 2009, 109, 5288–5353 5288

Near-infrared fluorescent labeled peptosome for application to cancer imaging

Nonionic amphiphilic copolypeptides, which were composed of hydrophilic poly(sarcosine) and hydrophobic poly(gamma-methyl L-glutamate) blocks, were synthesized with varying chain lengths of the blocks. The polypeptides having a suitable hydrophilic and hydrophobic balance were found to form vesicular assemblies of 100 nm size in buffer, which was evidenced by the TEM observation, the DLS analysis, and the encapsulation experiment. The genuine peptide vesicles, peptosomes, were labeled with a near-infrared fluorescence (NIRF) probe. In vivo retention in blood experiment showed long circulation of the peptosome in rat blood as stable as the PEGylated liposome. NIRF imaging of a small cancer on mouse by using the peptosome as a nanocarrier was successful due to the EPR effect of the peptosome. Peptosome is shown here as a novel excellent nanocarrier for molecular imaging.

Near-infrared fluorescence tumor imaging using nanocarrier composed of poly(L-lactic acid)-block-poly(sarcosine) amphiphilic polydepsipeptide

A nanocarrier, lactosome, which is composed of poly(L-lactic acid)-block-poly(sarcosine), as a contrast agent for the liver tumor imaging was examined using the near infrared fluorescence (NIRF) optical imaging technique. Lactosome labeled with indocyanine green (ICG) showed a high escape ability from the reticulo-endothelial system (RES). Lactosome was found to be stable in a blood circulation, and gradually accumulated specifically at a model liver tumor site, which was obtained by graft of HepG2/EF-Luc cells at a mouse liver. The high tumor/liver imaging ratio is due to the enhanced permeation and retention (EPR) effect of lactosome. The fluorescence intensity at the tumor site was correlated with the degree of malignancy. Tumor imaging using lactosome as a nanocarrier is therefore a potential candidate for a facile and general tumor imaging technique.

Nanotube and Three-Way Nanotube Formation with Nonionic Amphiphilic Block Peptides

Amphiphilic block polypeptides having a helical hydrophobic block with a uniform chain length and a hydrophilic nonionic block were newly synthesized and self-assembled into homogeneous nanotubes with ca. 60 nm diameter and ca. 200 nm length. The tubular assembly was shown to be elongated by heating over micrometer length without changing the diameter. Notably, a distinctive three-way nanotube was obtained just by mixing two kinds of amphiphilic polypeptides with the same helical hydrophobic block but different chain lengths of the hydrophilic block. The morphology of the molecular assemblies was shown to be tunable from a curved sheet-shaped assembly to a long or short nanotubular assembly and a three-way nanotubular assembly by suitable molecular design of the hydrophobic block, selection of the chain length of the hydrophilic block, mixing two-type block peptides, and processing such as heating.

Transformation of peptide nanotubes into a vesicle via fusion driven by stereo-complex formation.

Two types of peptide nanotubes, one is prepared from an amphiphilic peptide having a right-handed helix segment and the other from that having a left-handed helix segment, are shown to transform the morphology into a vesicle by membrane fusion due to stereo-complex formation between these helical segments.

Suppressive immune response of poly-(sarcosine) chains in peptide-nanosheets in contrast to polymeric micelles†

Nanoparticles are expected to be applicable for the theranostics as a carrier of the diagnostic and therapeutic agents. Lactosome is a polymeric micelle composed of amphiphilic polydepsipeptide, poly(sarcosine)64‐block‐poly(l‐lactic acid)30, which was found to accumulate in solid tumors through the enhanced permeability and retention effect. However, lactosome was captured by liver on the second administration to a mouse. This phenomenon is called as the accelerated blood clearance phenomenon. On the other hand, peptide‐nanosheet composed of amphiphilic polypeptide, poly(sarcosine)60‐block‐(l‐Leu‐Aib)6, where the poly(l‐lactic acid) block in lactosome was replaced with the (l‐Leu‐Aib)6 block, abolished the accelerated blood clearance phenomenon. The ELISA and in vivo near‐infrared fluorescence imaging revealed that peptide‐nanosheets did not activate the immune system despite the same hydrophilic block being used. The high surface density of poly(sarcosine) chains on the peptide‐nanosheet may be one of the causes of the suppressive immune response. Copyright

Rational design of peptide nanotubes for varying diameters and lengths.

Amphiphilic helical peptides (Sar)m‐b‐(L‐Leu‐Aib)n (m = 22–25; n = 7, 8, 10) with a hydrophobic block as a right‐handed helix were synthesized and their mixtures with (Sar)25‐b‐(D‐Leu‐Aib)6 containing the hydrophobic block as a left‐handed helix were examined in their molecular assembly formation. The single component (Sar)25‐b‐(D‐Leu‐Aib)6 forms peptide nanotubes of 70 nm diameter and 200 nm length. The two‐component mixtures of (Sar)25‐b‐(D‐Leu‐Aib)6 with (Sar)24‐b‐(L‐Leu‐Aib)7, (Sar)22‐b‐(L‐Leu‐Aib)8, and (Sar)25‐b‐(L‐Leu‐Aib)10 yield peptide nanotubes of varying dimensions with 200 nm diameter and 400 nm length, 70 nm diameter and several micrometer length (maximum 30 µm), and 70 nm diameter and 100–600 nm length, respectively. The right‐handed and the left‐handed helix were thus found to be molecularly mixed due to the stereo‐complex formation and to generate nanotubes of different sizes. When the mismatch of the hydrophobic helical length between the two components was of four residues, the longest nanotube was generated. Correspondingly, the hydrophobic helical segments have to interdigitate with an anti‐parallel orientation at the hydrophobic core region of the nanotube. Copyright

Pharmacokinetic change of nanoparticulate formulation "Lactosome" on multiple administrations.

Lactosome, which is a polymer micelle composed of poly(lactic acid)-b-poly(sarcosine), was applied successfully for solid tumor imaging. Lactosome is considered to escape from the reticuloendothelial system recognition, and shows prolonged in vivo blood clearance time. In vivo disposition of Lactosome, however, changed upon multiple dosages. Lactosome at the 2nd dosage was cleared from the blood stream by trapping at liver. This accelerated blood clearance (ABC) phenomenon is explained by production of anti-Lactosome IgM and IgG(3) through the immune response related with B-lymphocyte cells. The memory effect of B-lymphocyte cells lasted nearly for six months in mouse. The epitope moiety of Lactosome is concluded to be poly(sarcosine) based on the competitive inhibition assay. Since the ABC phenomenon was also reported with PEGylated liposome, nanoparticles in general may be potential in triggering the immune system.

Morphology Control between Twisted Ribbon, Helical Ribbon, and Nanotube Self-Assemblies with His-Containing Helical Peptides in Response to pH Change

pH-Responsive molecular assemblies with a variation in morphology ranging from a twisted ribbon, a helical ribbon, to a nanotube were prepared from a novel A3B-type amphiphilic peptide having three hydrophilic poly(sarcosine) (A block) chains, a hydrophobic helical dodecapeptide (B block), and two histidine (His) residues between the A3 and B blocks. The A3B-type peptide adopted morphologies of the twisted ribbon at pH 3.0, the helical ribbon at pH 5.0, and the nanotube at pH 7.4, depending upon the protonation states of the two His residues. On the other hand, another A3B-type peptide having one His residue between the A3 and B blocks showed a morphology change only between the helical ribbon and the relatively planar sheets with pH variation in this range. The morphology change is thus induced by one- or two-charge generation at the linking site of the hydrophilic and hydrophobic blocks of the component amphiphiles but in different ways.

Micelle-based activatable probe for in vivo near-infrared optical imaging of cancer biomolecules

UNLABELLED Near-infrared (NIR: 800-1000 nm) fluorescent probes, which activate their fluorescence following interaction with functional biomolecules, are desirable for noninvasive and sensitive tumor diagnosis due to minimal tissue interference. Focusing on bioavailability and applicability, we developed a probe with a self-assembling polymer micelle, a lactosome, encapsulating various quantities of NIR dye (IC7-1). We also conjugated anti-HER2 single chain antibodies to the lactosome surface and examined the probes capacity to detect HER2 in cells and in vivo. Micelles encapsulating 20mol% IC7-1 (hIC7L) showed 30-fold higher fluorescence (λem: 858 nm) after micelle denaturation compared to aqueous buffer. Furthermore, antibody modification allowed specific activation of the probe (HER2-hIC7L) following internalization by HER2-positive cells, with the probe concentrating in lysosomes. HER2-hIC7L intravenously administered to mice clearly and specifically visualized HER2-positive tumors by in vivo optical imaging. These results indicate that HER2-hIC7L is a potential activatable NIR probe for sensitive tumor diagnosis. FROM THE CLINICAL EDITOR Near-infrared probes that activate their fluorescence following interaction with specific biomolecules are desirable for noninvasive and sensitive tumor detection due to minimal tissue interference. This team of authors developed a probe termed hIC7L and demonstrate its potential in HER2 tumor diagnosis.