EDITORIAL

Ligand-mediated drug delivery systems have enormous potential for improving the efficacy of cancer treatment. In particular, Arg-Gly-Asp peptides are promising ligand molecules for targeting α_vβ₃/α_vβ₅ integrins, which are overexpressed in angiogenic sites and tumors, such as intractable human glioblastoma (U87MG). We here achieved highly efficient drug delivery to U87MG tumors by using a platinum anticancer drug-incorporating polymeric micelle (PM) with cyclic Arg-Gly-Asp (cRGD) ligand molecules. Intravital confocal laser scanning microscopy revealed that the cRGD-linked polymeric micelles (cRGD/m) accumulated rapidly and had high permeability from vessels into the tumor parenchyma compared with the PM having nontargeted ligand, “cyclic-Arg-Ala-Asp” (cRAD). As both cRGD/m- and cRAD-linked polymeric micelles have similar characteristics, including their size, surface charge, and the amount of incorporated drugs, it is likely that the selective and accelerated accumulation of cRGD/m into tumors occurred <i>via</i> an active internalization pathway, possibly transcytosis, thereby producing significant antitumor effects in an orthotopic mouse model of U87MG human glioblastoma

Artificial Control of Gene Silencing Activity Based on siRNA Conjugation with Polymeric Molecule Having Coil–Globule Transition Behavior

A new strategy for controlling gene silencing activity of siRNA in the cell was developed in the present study. siRNA was linearly conjugated with PNIPAAm, where coil–globule transition of the conjugated PNIPAAm allows thermoresponsive exposure of the vicinal siRNA molecule; a coil form of PNIPAAm (<i>T</i> < LCST) inhibits siRNA interaction with gene silencing-related proteins due to the steric hindrance effect, while a globule form of PNIPAAm (<i>T</i> > LCST) allows a ready access of siRNA to gene silencing pathway. As a result, at <i>T</i> > LCST, PNIPAAm-siRNA elicited effective association of siRNA with a gene silencing-related protein of Ago2, while siRNA recruitment into the gene silencing pathway was significantly suppressed at <i>T</i> < LCST. Ultimately, gene silencing efficacy of PNIPAAm-siRNA was close to unconjugated siRNA at <i>T</i> > LCST (∼80%), while it was dramatically decreased to ∼20% at <i>T</i> < LCST, suggesting that coil–globule transition of the conjugated polymer can control the bioactivity of the vicinal siRNA molecule

Laboratory data.

<p>Laboratory data.</p

Intracellular Delivery of Charge-Converted Monoclonal Antibodies by Combinatorial Design of Block/Homo Polyion Complex Micelles

Direct intracellular delivery of antibodies has gained much attention, although only a few agents have been developed, and none of them has reached clinical stages. The main obstacles here are the insufficient characteristics of delivery systems including stability and appropriate ability for intracellular antibody release. We tailored the structure of polyion complex (PIC) micelles by loading transiently charge-converted antibody derivatives for achieving enhanced stability, delivery to cytosol, and precise antigen recognition inside cells. Citraconic anhydride was used for the charge conversion of the antibody; the optimized degree of modification was identified to balance the stability of PIC micelles in the extracellular compartment and prompt pH-triggered disintegration after their translocation into the acidic endosomal compartment of target cells. The use of a mixture of homo- and block-catiomers in an appropriate ratio to construct PIC micelles substantially enhanced the endosomal escaping efficacy of the loaded antibody, leading to improved recognition of intracellular antigens

Intravenous injection of psFlt-1-PM decreased the area of CNV.

<p>Quantification of the CNV lesion demonstrated that the neovascularized area in the mice that received psFlt-1-PM was significantly reduced by 60% than that of control mice (n = 7, p<0.01), whereas the administration of pYFP-PM had no significant effects on the neovascularized area. The lower panels show representative micrographs. Arrows indicate the CNV lesion. CNV: choroidal neovascularization, psFlt-1-PM: the PIC micelle encapsulating psFlt-1 (fms-like tyrosine kinase-1), pYFP-PM: the PIC micelles encapsulating pYFP (yellow fluorescent protein).</p

Targeted gene expression in the CNV area after intravenous injection of pYFP-PM.

<p>a: Choroidal flatmount and histological analysis demonstrated YFP fluorescence in the CNV area after intravenous injection of pYFP-PM. b: Western blotting analysis demonstrated that YFP protein was detected in the eyes with CNV, after intravenous injection of pYFP-PM. YFP expression was detected neither after generation of CNV alone nor after intravenous injection of pYFP-PM without CNV. c: The results of immunohistochemistry demonstrated that the expression of F4/80 was partially overlapping the expression of YFP, indicating that YFP protein expressed in F4/80-positive macrophage. CNV: choroidal neovascularization, pYFP-PM: the PIC micelles encapsulating pYFP (yellow fluorescent protein), PC: photocoagulation, bar:100 um.</p

Bundled Assembly of Helical Nanostructures in Polymeric Micelles Loaded with Platinum Drugs Enhancing Therapeutic Efficiency against Pancreatic Tumor

Supramolecular assemblies of amphiphilic block copolymers having polypeptide segments offer significant advantages for tailoring spatial arrangement based on secondary structures in their optically active backbones. Here, we demonstrated the critical effect of α-helix bundles in cisplatin-conjugated poly(l- (or d-)glutamate) [P(l(or d)Glu)-CDDP] segment on the packaging of poly(ethylene glycol) (PEG)-P(l(or d)Glu)-CDDP block copolymers in the core of polymeric micelles (CDDP/m) and enhanced micelle tolerability to harsh <i>in vivo</i> conditions for accomplishing appreciable antitumor efficacy against intractable pancreatic tumor by systemic injection. CDDP/m prepared from optically inactive PEG-poly(d,l-glutamate) (P(d,lGlu)), gradually disintegrated in the bloodstream, resulting in increased accumulation in liver and spleen and reduced antitumor efficacy. Alternatively, CDDP/m from optically active PEG-P(l(or d)Glu) maintained micelle structure during circulation, and eventually attained selective tumor accumulation while reducing nonspecific distribution to liver and spleen. Circular dichroism and small-angle X-ray scattering measurements indicated regular bundled assembly of α-helices in the core of CDDP/m from PEG-P(l(or d)Glu), which is suggested to stabilize the micelle structure against dilution in physiological condition. CDDP/m suffered corrosion by chlorides in medium, yet the optically active micelles with α-helix bundles kept the micelle structure for prolonged time, with slowly releasing unimers and dimers from the surface of the bundled core in an erosion-like process, as verified by ultracentrifugation analysis. This is in sharp contrast with the abrupt disintegration of CDDP/m from PEG-P(d,lGlu) without secondary structures. The tailored assembly in the core of the polymeric micelles through regular arrangement of constituting segments is key to overcome their undesirable disintegration in bloodstream, thereby achieving efficient delivery of loaded drugs into target tissues

Evaluation of Differences in Automated QT/QTc Measurements between Fukuda Denshi and Nihon Koden Systems

<div><p>Background</p><p>Automatic measurement becomes a preference, and indeed a necessity, when analyzing 1000 s of ECGs in the setting of either drug-inducing QT prolongation screening or genome-wide association studies of QT interval. The problem is that individual manufacturers apply different computerized algorithms to measure QT interval. We conducted a comparative study to assess the outcomes with different automated measurements of QT interval between ECG machine manufacturers and validated the related heart rate correction methods.</p><p>Methods and Results</p><p>Herein, we directly compared these different commercial systems using 10,529 Fukuda Denshi ECGs and 72,754 Nihon Kohden ECGs taken in healthy Japanese volunteers. Log-transformed data revealed an equal optimal heart rate correction formula of QT interval for Fukuda Denshi and Nihon Kohden, in the form of QTc = QT/RR^−0.347. However, with the raw data, the optimal heart rate correction formula of QT interval was in the form of QTc = QT+0.156×(1-RR) for Fukuda Denshi and QTc = QT+0.152×(1-RR) for Nihon Kohden. After optimization of heart rate correction of QT interval by the linear regression model using either log-transformed data or raw data, QTc interval was ∼10 ms longer in Nihon Kohden ECGs than in those recorded on Fukuda Denshi machines. Indeed, regression analysis revealed that differences in the ECG machine used had up to a two-fold larger impact on QT variation than gender difference. Such an impact is likely to be of considerable importance when ECGs for a given individual are recorded on different machines in the setting of multi-institutional joint research.</p><p>Conclusions</p><p>We recommend that ECG machines of the same manufacturer should be used to measure QT and RR intervals in the setting of multi-institutional joint research. It is desirable to unify the computer algorithm for automatic QT and RR measurements from an ECG.</p></div

Analysis of resting Fukuda Denshi ECGs.

<p>(A) Histograms of QT, log-transformed QT, RR, and log-transformed RR intervals. (B) Scatter plots of log QT versus log RR and log QTc_ours log versus log RR. (C) QT versus RR and QTc_ours raw versus RR. Units of all variables are ms.</p

Effect of Knockout of <i>Mdr1a</i> and <i>Mdr1b</i> ABCB1 Genes on the Systemic Exposure of a Doxorubicin-Conjugated Block Copolymer in Mice

We previously elucidated that ATP-binding cassette subfamily B member 1 (ABCB1) mediates the efflux of doxorubicin-conjugated block copolymers from HeLa cells. Here, we investigated the role of ABCB1 in the in vivo behavior of a doxorubicin-conjugated polymer in <i>Mdr1<i>a</i>/1b­(−/−)</i> mice. The area under the curve for intravenously administered polymer in <i>Mdr1<i>a</i>/1b­(−/−)</i> mice was 2.2-fold greater than that in wild-type mice. The polymer was mostly distributed in the liver followed by spleen and less so in the brain, heart, kidney, and lung. The amount of polymer excreted in the urine was significantly decreased in <i>Mdr1<i>a</i>/1b­(−/−)</i> mice. The amounts of polymers excreted in the feces were similar in both groups despite the higher systemic exposure in <i>Mdr1<i>a</i>/1b­(−/−)</i> mice. Confocal microscopy images showed polymer localized in CD68⁺ macrophages in the liver. These results show that knockout of ABCB1 prolonged systemic exposure of the doxorubicin-conjugated polymer in mice. Our results suggest that ABCB1 mediated the excretion of doxorubicin-conjugated polymer in urine and feces. Our results provide valuable information about the behavior of block copolymers in vivo, which is important for evaluating the pharmacokinetics of active substances conjugated to block copolymers or the accumulation of block copolymers in vivo