Novel Antibody Exerts Antitumor Effect through Downregulation of CD147 and Activation of Multiple Stress Signals.

CD147 is an immunoglobulin-like receptor that is highly expressed in various cancers and involved in the growth, metastasis, and activation of inflammatory pathways via interactions with various functional molecules, such as integrins, CD44, and monocarboxylate transporters. Through screening of CD147-targeting antibodies with antitumor efficacy, we discovered a novel rat monoclonal antibody #147D. This humanized IgG4-formatted antibody, h4#147D, showed potent antitumor efficacy in xenograft mouse models harboring the human PDAC cell line MIA PaCa-2, HCC cell line Hep G2, and CML cell line KU812, which featured low sensitivity to the corresponding standard-of-care drugs (gemcitabine, sorafenib, and imatinib, respectively). An analysis of tumor cells derived from MIA PaCa-2 xenograft mice treated with h4#147D revealed that cell surface expression of CD147 and its binding partners, including CD44 and integrin α3ß1/α6ß1, was significantly reduced by h4#147D. Inhibition of focal adhesion kinase (FAK), activation of multiple stress responsible signal proteins such as c-JunN-terminal kinase (JNK) and mitogen-activated protein kinase p38 (p38MAPK), and expression of SMAD4, as well as activation of caspase-3 were obviously observed in the tumor cells, suggesting that h4#147D induced tumor shrinkage by inducing multiple stress responsible signals. These results suggest that the anti-CD147 antibody h4#147D offers promise as a new antibody drug candidate.

Preparation and Photocatalytic Activity of Hexagonal Plate-like ZnO Particles Using Anionic Surfactants.

Hexagonal plate-like ZnO particles with a high degree of c-face orientation have been synthesized using hydrothermal method in the presence of various anionic surfactants bearing different hydrocarbon chains. The c-face of the ZnO particles increased upon increasing the surfactant alkyl chain length. The photocatalytic activity of the as-obtained hexagonal plate-like ZnO particles was evaluated using the degradation of methylene blue (MB). Although the specific surface area of hexagonal rod-like particles is higher than that of hexagonal plate-like particles, the amount of MB adsorption on the ZnO particle surface was different for the hexagonal plate-like and rod-like particles. In addition, the hexagonal plate-like ZnO particles showed a significantly higher decrease in the MB concentration with the duration of ultraviolet light irradiation when compared to the hexagonal rod-like ZnO particles obtained in the absence of a surfactant. These results indicate that crystal-face-controlled ZnO with a high degree of c-face orientation exhibits high photocatalytic activity.

Synthesis and Photocatalytic Activity of Hexagonal Plate-like Shaped Au Nanoparticles/ZnO Composite Particles under Visible-light Irradiation.

We synthesized Au nanoparticle (AuNP)/ZnO composite particles in presence of an anionic surfactant and evaluated their photocatalytic activity under visible-light irradiation. AuNPs synthesized from HAuCl4 in the presence of amylase and the precursor solutions of ZnO were mixed, followed by a hydrothermal process, to synthesize crystal face-controlled AuNP/ZnO composite particles. X-ray diffraction (XRD) patterns and ultraviolet-visible (UV-Vis) spectra confirmed the formation of AuNP/ZnO composite particles. Furthermore, different Au to Zn concentration ratios in the precursor solutions resulted in different amounts of AuNPs in the composite particles. In addition, the average size of the AuNP/ZnO composite particles decreased with increasing Au to Zn concentration ratio. We believe AuNPs act as the nuclei for ZnO particle formation. The photocatalytic activity of the AuNP/ZnO composite particles was evaluated by the photodegradation of methylene blue (MB) under visible-light irradiation. The photodegradation rate of MB was higher with AuNP/ZnO composite particles compared to that with the ZnO particles synthesized in the absence of AuNPs. The AuNP/ZnO composite particles exhibit photocatalytic activity under visible-light irradiation owing to the enhanced charge separation efficiency and the localized surface plasmon resonance effect.

Novel anti-GARP antibody DS-1055a augments anti-tumor immunity by depleting highly suppressive GARP+ regulatory T cells.

Regulatory T (Treg) cells, which are essential for maintaining self-tolerance, inhibit anti-tumor immunity, consequently hindering protective cancer immunosurveillance, and hampering effective anti-tumor immune responses in tumor-bearing hosts. Here, we show that depletion of Treg cells via targeting glycoprotein A repetitions predominant (GARP) induces effective anti-tumor immune responses. GARP was specifically expressed by highly suppressive Treg cells in the tumor microenvironment (TME) of multiple cancer types in humans. In the periphery, GARP was selectively induced in Treg cells, but not in effector T cells, by polyclonal stimulation. DS-1055a, a novel afucosylated anti-human GARP monoclonal antibody, efficiently depleted GARP+ Treg cells, leading to the activation of effector T cells. Moreover, DS-1055a decreased FoxP3+CD4+ T cells in the TME and exhibited remarkable anti-tumor activity in humanized mice bearing HT-29 tumors. We propose that DS-1055a is a new Treg-cell-targeted cancer immunotherapy agent with augmentation of anti-tumor immunity.

Effect of Anionic Amphiphiles on the Morphology of Hexagonal Plate-like ZnO Particles.

Zinc oxide (ZnO) particles were synthesized in the presence of anionic surfactants (ASs). The effect of ASs on the morphology of the ZnO particles was investigated by using ASs with various alkyl chain lengths and changing the molar ratio of AS/ZnSO4. Hexagonal plate-like ZnO particles were formed in the presence of ASs. Adsorption of the AS on the c face of the ZnO crystals inhibited (promoted) crystal growth along the c-axis (ab-axis) direction. Increasing the molar ratio of AS/ZnSO4 decreased the particle thickness, owing to the resulting increase in the coverage of the c face with AS. The particle diameter of the hexagonal plate-like ZnO particles (the diagonal length of the hexagonal plate) increased with increasing alkyl chain length of AS as a result of the van der Waals interactions between the alkyl chains. The data indicate that the particle diameter and thickness can be controlled by fine-tuning the van der Waals interactions between the alkyl chains and the coverage of the c face of the ZnO particles with AS.

Preparation of Hexagonal Plate-like ZnO Single-crystal Particles in the Presence of Anionic Amphiphiles.

In this study, we synthesized ZnO particles using anionic amphiphiles as an additive. While the single-crystal particles prepared in the absence of such amphiphiles had a hexagonal rod-like shape, those fabricated using anionic amphiphilic molecules had a hexagonal plate-like shape. The anionic amphiphiles inhibited crystal growth in the c-axis direction of ZnO. This demonstrated that the anionic surfactants served as crystal-growth-directing agents, controlling the shape of the ZnO particles.

Detection of histidine oxidation in a monoclonal immunoglobulin gamma (IgG) 1 antibody.

Although oxidation of methionine and tryptophan are known as popular chemical modifications that occur in monoclonal antibody (mAb) molecules, oxidation of other amino acids in mAbs has not been reported to date. In this study, oxidation of the histidine residue in a human immunoglobulin gamma (IgG) 1 molecule was discovered for the first time by mass spectrometry. The oxidation of a specific histidine located at the CH2 domain of IgG1 occurred under light stress, but it was not observed under heat stress. With the forced degradation study using several reactive oxygen species, the singlet oxygen was attributed to a reactive source of the histidine oxidation. The reaction mechanism of the histidine oxidation was proposed on the basis of the mass spectrometric analysis of IgG1 oxidized in deuterium oxide and hydrogen heavy oxide.

Specific racemization of heavy-chain cysteine-220 in the hinge region of immunoglobulin gamma 1 as a possible cause of degradation during storage.

Therapeutic antibodies often suffer from degradation due to various modifications during storage. We detected a degradation of immunoglobulin gamma 1 (IgG1) stored for 6 month at 40 °C, and identified the modification as the racemization of Cys220 in the hinge sequence S(219)CDKTHT(225) of the heavy chain by tryptic peptide mapping and tandem mass spectrometry. The rate of racemization at Cys220 was enhanced deliberately by incubating the protein at 50 °C and pH 9.0, while all the other cysteine residues were not affected. The racemization of Cys220 was confirmed by mass spectrometry in conjunction with extracted ion chromatography of the tryptic digest of IgG1 forced to degrade in D(2)O and a series of synthetic hinge fragments containing D-amino acid, as well as the detection of D-cysteine in the acid hydrolysate. To rationalize the possible relationship between the racemization of Cys220 and isopeptide formation at the neighboring residue of Asp221, we suggest a new reaction mechanism that assumes a base catalyst to initiate these reactions by activating the amide nitrogen of Lys222. Due to the highly flexible nature of the hinge region, Lys222 can participate in either the formation of a cyclic imidazoline intermediate involving the α-carbonyl carbon of Cys220 to facilitate the racemization of Cys220 or that of a succinimide structure leading to the isomerization of Asp221.