Functional Peptide-Modified Liposomes for siRNA Delivery to Rat Retinal Pigment Epithelium Cells: Effect of Peptide Sequences.

Most retinal diseases involve the degeneration of choroidal retinal pigment epithelial (RPE) cells. Because of a blood-retina barrier (tight junction formation), RPE cells restrict the entry of hydrophilic macromolecules (e.g., small interfering RNA (siRNA)) through blood stream and eye drops. A cytoplasm-responsive stearylated (STR) peptide, STR-CH2R4H2C (CH2R4) enables stable siRNA complexation, cell permeation, and intracellular dynamics control. We previously demonstrated how CH2R4-modified liposomes promoted siRNA efficacy. We investigated the influence of amino acid sequences of functional peptides on cellular uptake pathways, siRNA transfection efficacy, and the permeation of peptide-modified liposomes in rat RPE-J cells. Four STR-peptides, consisting of arginine (R), cysteine (C), histidine (H), lysine (K) or serine (S), were designed based on CH2R4. We prepared siRNA-loaded, peptide-modified cationic liposomes (CH2R4-, CH2K4-, CH2S4-, SH2R4-, and SH2S4-lipoplexes). CH2R4-, CH2K4-, and SH2R4-lipoplexes induced cellular uptake by macropinocytosis by activating cytoskeletal F-actin, possibly due to cationic amino acids (arginine, lysine). SH2R4-lipoplexes were trapped in endosomes, whereas CH2R4- and CH2K4-lipoplexes enhanced endosomal siRNA release suggesting cysteine contributes to endosomal escape. Although cationic liposome-based, CH2S4- and SH2S4-lipoplexes (not including arginine and lysine) showed lower siRNA transfection efficiency. This difference may be because siRNAs were retained on both peptide moieties and cationic liposomes in CH2R4-, CH2K4- and SH2R4-lipoplexes, whereas in CH2S4- and SH2S4-lipoplexes, siRNAs were loaded to the cationic liposomes, but not on peptides. In three-dimensional spheroids, CH2R4- and CH2K4-modified liposomes promoted permeation through tight junctions. Thus, cationic amino acids and cysteine within peptide sequences of CH2R4 could be effective for siRNA delivery to the retina using functional peptide-modified liposomes.

[A Case of Hepatocellular Carcinoma Complicated by Liver Metastasis of Colon Cancer during the Course of Treatment].

A 72‒year‒old man with hepatocellular carcinoma(HCC)was treated with transarterial chemoembolization(TACE)and radiofrequency ablation(RFA). Six months after RFA, gadolinium ethoxybenzyl diethylenetriamine pentaacetic acid(Gd‒ EOB‒DTPA)‒enhanced magnetic resonance imaging(MRI)revealed multiple metastatic recurrences in the liver. TACE was performed for the recurrent HCC. However, the treatment response on the Gd‒EOB‒DTPA‒enhanced MRI showed that the lesions had advanced and that the liver metastatic nodules had ring‒shaped contrast effects. We suspected metastatic liver cancer based on the MRI findings and performed colonoscopy. Finally, we diagnosed the patient with multiple hepatic metastases of sigmoid colon cancer based on the results of the endoscopic colon biopsy and percutaneous liver tumor biopsy. In conclusion, we had a teachable case of the treatment of HCC.

Comparison Between Linked Color Imaging and Blue Laser Imaging for Improving the Visibility of Flat Colorectal Polyps: A Multicenter Pilot Study.

INTRODUCTION: Linked color imaging (LCI) and blue laser imaging-bright (BLI-b) improve the visibility of gastrointestinal lesions. In this multicenter study, we compared the effects of LCI and BLI-b on the visibility of flat polyps with visibility scores and color difference (CD) values, including fast-withdrawal and large-monitor observation. METHODS: We recorded 120 videos of 40 consecutive flat polyps (2-20 mm), adenoma, and sessile serrated adenoma and polyp (SSA/P), using white light imaging (WLI), BLI-b, and LCI from July 2017 to December 2017. All videos were evaluated by eight endoscopists according to a published polyp visibility score of 4 (excellent) to 1 (poor). Additionally, 1.5 ×faster and 1.7 ×sized videos were evaluated. Moreover, we calculated the CD values for each polyp in three modes. RESULTS: The mean LCI scores (3.1 ± 0.9) were significantly higher than the WLI scores (2.5 ± 1.0, p < 0.001) but not significantly higher than the BLI-b scores (3.0 ± 1.0). The scores of faster videos on LCI (3.0 ± 1.1) were significantly higher than WLI (2.0 ± 1.0, p < 0.001) and BLI-b (2.8 ± 1.1, p = 0.03). The scores of larger-sized videos on LCI were not significantly higher than those of WLI or BLI-b. The CD value of LCI (18.0 ± 7.7) was higher than that of WLI (11.7 ± 7.0, p < 0.001), but was not significantly higher than that of BLI-b (16.6 ± 9.6). The CD value of LCI was significantly higher than that of BLI-b for adenoma, but the CD value of BLI-b was significantly higher than that of LCI for SSA/P. CONCLUSIONS: The superiority of LCI to BLI-b was proven for the visibility of adenoma and fast observation.

Kinetics of cytokine mRNA and protein expression by plastic adherent cells in the thymus after split-dose irradiation.

Whole body irradiation causes significant apoptosis in various tissues such as the thymus. If apoptotic cells outnumber the phagocytic capacity of macrophages, apoptosis becomes secondary necrosis, inducing inflammatory cytokine expression in macrophages. Radiation also induces thymic lymphomas in C57BL/6 mice after four consecutive irradiations with 1.6 Gy X-rays with nearly 100% incidence. Since cancer development is modulated by a microenvironment involving macrophages, we examined the kinetics of thymocyte number and plastic adherent cell number in the thymus as well as cytokine mRNA expression by plastic adherent cells in the thymus after split-dose irradiation. Upon split-dose irradiation, thymocyte number changed dramatically, whereas plastic adherent cell number did not. Among cytokine mRNAs tested, IL-1ß, IL-11 and IL-12p40 mRNAs were up regulated 2 days after the 1st and 2nd, 3rd and 4th, and 2nd and 3rd irradiations, respectively. On the other hand, TNF-α mRNA was up regulated 2 days after the 3rd irradiation and 2 weeks after the 4th irradiation. The level of IL-11 protein was also increased 2 days after 3rd and 4th irradiations. These results suggest that, upon split-dose irradiation, macrophages in the thymus produce various cytokines in a time-dependent manner, thereby contributing to induction of thymic lymphomas.

Simulation of Stimuli-Responsive and Stoichiometrically Controlled Release Rate of Doxorubicin from Liposomes in Tumor Interstitial Fluid.

PURPOSE: To simulate the stimuli-responsive and stoichiometrically controlled doxorubicin (DOX) release from liposomes in in vivo tumor interstitial fluid (TIF), the effect of ammonia concentration and pH on the DOX release from liposomes in human plasma at 37°C was quantitatively evaluated in vitro and the release rate was calculated as a function of ammonia concentration and pH. METHODS: Human plasma samples spiked with DOX-loaded PEGylated liposomes (PLD) or Doxil®, containing ammonia (0.3-50 mM) at different pH values, were incubated at 37°C for 24 h. After incubation, the concentration of encapsulated DOX in the samples was determined by validated solid-phase extraction (SPE)-SPE-high performance liquid chromatography. RESULTS: Accelerated DOX release (%) from liposomes was observed as the increase of ammonia concentration and pH of the matrix, and the decrease of encapsulated DOX concentration. The release rate was expressed as a function of the ammonia concentration and pH by using Henderson-Hasselbalch equation. CONCLUSIONS: The DOX release from PLD in TIF was expressed as a function ammonia concentration and pH at various DOX concentrations. Further, it was found that the DOX release from liposomes in a simulated TIF was more than 15 times higher than in normal plasma.

Circadian Clock in a Mouse Colon Tumor Regulates Intracellular Iron Levels to Promote Tumor Progression.

Iron is an important biological catalyst and is critical for DNA synthesis during cell proliferation. Cellular iron uptake is enhanced in tumor cells to support increased DNA synthesis. Circadian variations in DNA synthesis and proliferation have been identified in tumor cells, but their relationship with intracellular iron levels is unclear. In this study, we identified a 24-h rhythm in iron regulatory protein 2 (IRP2) levels in colon-26 tumors implanted in mice. Our findings suggest that IRP2 regulates the 24-h rhythm of transferrin receptor 1 (Tfr1) mRNA expression post-transcriptionally, by binding to RNA stem-loop structures known as iron-response elements. We also found thatIrp2mRNA transcription is promoted by circadian clock genes, including brain and muscle Arnt-like 1 (BMAL1) and the circadian locomotor output cycles kaput (CLOCK) heterodimer. Moreover, growth in colon-26(Δ19) tumors expressing the clock-mutant protein (CLOCK(Δ19)) was low compared with that in wild-type colon-26 tumor. The time-dependent variation of cellular iron levels, and the proliferation rate in wild-type colon-26 tumor was decreased by CLOCK(Δ19)expression. Our findings suggest that circadian organization contributes to tumor cell proliferation by regulating iron metabolism in the tumor.

Detectability of colorectal neoplastic lesions using a novel endoscopic system with blue laser imaging: a multicenter randomized controlled trial.

BACKGROUND AND AIMS: Most studies have not reported an improvement in the detection of adenomas with the use of image-enhanced colonoscopy methods, possibly because of the darkness of the images. To overcome this limitation, a new-generation endoscopic system has been developed. This system has 2 blue-laser imaging (BLI) observation modes. The BLI observation was set to BLI-bright mode to detect lesions. We aimed to evaluate the efficacy of BLI in detecting lesions. METHODS: This study was designed as a randomized controlled trial with participants from 8 institutions. We enrolled patients aged ≥40 years. The participants were randomly assigned to 2 groups: observation by using white-light imaging (WLI) with a conventional xenon light source (WLI group) or observation by using BLI-bright mode with a laser light source (BLI group). All of the detected lesions were resected or had a biopsy taken for histopathologic analysis. The primary outcome was the mean number of adenomas per patient (MAP) that were detected per procedure. RESULTS: The WLI and BLI groups consisted of 474 and 489 patients, respectively. The MAP was significantly higher in the BLI group than in the WLI group (mean ± standard deviation [SD] WLI 1.01 ± 1.36, BLI 1.27 ± 1.73; P = .008). Adenoma detection rate in the BLI group was not significantly higher than in the WLI group. Observation times differed significantly, with BLI (9.48 minutes) being longer than WLI (8.42; P < .001). The mean (± SD) number of polyps per patient was significantly higher in the BLI group compared with the WLI group (WLI 1.43 ± 1.64, BLI 1.84 ± 2.09; P = .001). CONCLUSIONS: A newly developed system that uses BLI improves the detection of adenomatous lesions compared with WLI. (Clinical trial registration number: UMIN 000014555.).

Association between Skeletal Muscle Depletion and Sorafenib Treatment in Male Patients with Hepatocellular Carcinoma: A Retrospective Cohort Study.

The effect of skeletal muscle mass (SMM) on the outcomes of sorafenib treatment for hepatocellular carcinoma (HCC) has not been established. We measured the SMM in HCC patients treated with sorafenib, evaluated the patients' survival, and evaluated the association between skeletal muscle depletion and sorafenib treatment. Of the 97 HCC patients treated with sorafenib at our institution in the period from July 2009 to February 2015, our study included 69 patients (51 males, 18 females) who had received sorafenib for ≥ 8 weeks and whose follow-up data were available. SMM was calculated from computed tomography images at the mid-L3 level (cm2) and normalized to height (m2) to yield the L3 skeletal muscle index (L3-SMI, cm2/m2). The median L3-SMI value was higher in the males (43 cm2/m2) compared to the females (36 cm2/m2). In the males only, the multivariate Cox regression identified an L3-SMI <43 cm2/m2 as independently associated with higher mortality compared to an L3-SMI ≥43 cm2/m2 (hazard ratio 2.315, 95% confidence interval: 1.125-4.765, p=0.023). Skeletal muscle depletion is a factor predicting poor prognosis for male patients with advanced HCC treated with sorafenib.

Novel Endoscopic Findings as Visualized by Magnifying Endoscopy with Narrow-Band Imaging: White Opaque Substance Is Present in Colorectal Hyperplastic Polyps.

BACKGROUND: Magnifying endoscopy (ME) with narrow-band imaging (NBI) can visualize a white opaque substance (WOS) in gastric epithelial neoplasms, gastric intestinal metaplasias, and colorectal epithelial neoplasms. Histological examination showed the WOS to be lipid droplets accumulated in the epithelium. The white appearance of colorectal hyperplastic polyps suggests that they may contain WOS, but this has not been investigated as yet. AIMS: The purpose of this study was to determine whether WOS is present in colorectal hyperplastic polyps. METHODS: We retrospectively evaluated endoscopic images of 26 consecutive lesions investigated by ME with NBI and subsequently endoscopically resected and confirmed to be hyperplastic polyps. RESULTS: WOS was present in 21 of the 26 colorectal hyperplastic polyps (80.8%) based on the findings of ME with NBI. Adipophilin was present in 24 of the 26 colorectal hyperplastic polyps (92.3%). CONCLUSIONS: This study is the first to demonstrate that WOS (i.e. lipid droplets) accumulates in the epithelium of colorectal hyperplastic polyps.

Comparison of Absorbents and Drugs for Internal Decorporation of Radiocesium: Advances of Polyvinyl Alcohol Hydrogel Microsphere Preparations Containing Magnetite and Prussian Blue.

Radiocesium nuclides, used as a gamma ray source in various types of industrial equipments and found in nuclear waste, are strictly controlled to avoid their leakage into the environment. When large amounts of radiocesium are accidentally incorporated into the human body, decorporation therapy should be considered. Although standard decorporation methods have been studied since the 1960s and were established in the 1970s with the drug Radiogardase(®) (a Prussian blue preparation), application of recent advances in pharmacokinetics and ethical standards could improve these methods. Here we designed a modern dosage form of hydrogel containing cesium-absorbents to alleviate intestinal mucosa irritation due to the cesium-binding capacity of the absorbents. The effectiveness of the dosage form on fecal excretion was confirmed by quantitative mouse experiments. The total cesium excretion rate of the crystal form (1.37±0.09) was improved by the hydrogel form (1.52±0.10) at the same dose of Prussian blue, with a longer gastrointestinal tract transit time. Using a mouse model, we compared the effects of several drugs on fecal and urinary excretion of internal cesium, without the use of absorbents. Only phenylephrine hydrochloride significantly enhanced cesium excretion (excretion rate of 1.17±0.08) via the urinary pathway, whereas none of the diuretic drugs tested had this effect. These findings indicate that modifying the dosage form of cesium absorbents is important for the decorporation of internal radiocesium contamination.

A case of idiopathic portal hypertension associated with nodular regenerative hyperplasia-like nodule of the liver and mixed connective tissue disease.

A 51-year-old woman was diagnosed with mixed connective tissue disease (MCTD) in 2011. She underwent treatment with prednisolone. Her hepatobiliary enzyme level increased, and multiple nodules were found in both liver lobes in abdominal imaging studies. Ultrasonography revealed large and small hyperechoic lesions with indistinct or well-defined borders. No findings of classic hepatocellular carcinoma or liver cirrhosis were observed on contrast-enhanced computed tomography, but some nodules showed an enhanced effect of the central lesion that was characteristic of focal nodular hyperplasia (FNH) in an arterial phase. On gadolinium-ethoxybenzyl-diethylenetriamine penta-acetic acid (Gd-EOB-DTPA)-enhanced magnetic resonance imaging, slightly high-intensity nodules, 10-40mm in size, were observed on T1- and T2-weighted images. The nodules showed highest intensities in the hepatocyte phase and were enhanced with the uptake of Gd-EOB-DTPA as compared with the background liver. FNH was suspected based on the imaging findings, but we performed a liver tumor biopsy for differential diagnosis of the malignant lesion. Based on the immunohistopathological examination results, the final diagnosis was idiopathic portal hypertension associated with nodular regenerative hyperplasia (NRH)-like nodule of the liver. Benign nodular hepatocellular lesions are caused by abnormal hepatic circulation and were previously known as anomalous portal tract syndrome. Our case of atypical NRH with large nodules may be included in this disease entity. Here, we report a rare case of MCTD with NRH-like nodules and idiopathic portal hypertension with a review of literature.

[A case of a simple ulcer associated with ulcerative colitis that demonstrated a good clinical response to infliximab].

We present a case of a man in his 60s who had been in clinical remission of ulcerative colitis (UC) after treatment with 5ASA. Over the clinical course, he developed an isolated deep ulcer at the end of the ileum. There were moderate active UC findings in the rectum. We diagnosed a simple ulcer associated with UC and started treatment with azathioprine and infliximab (IFX). Shortly after the treatment, the ulcer began to scar. We report a rare case of a simple ulcer that accompanied UC, and for which IFX was effective.

Cell-penetrating peptide-conjugated lipid nanoparticles for siRNA delivery.

Lipid nanoparticles (LNP) modified with cell-penetrating peptides (CPP) were prepared for the delivery of small interfering RNA (siRNA) into cells. Lipid derivatives of CPP derived from protamine were newly synthesized and used to prepare CPP-decorated LNP (CPP-LNP). Encapsulation of siRNA into CPP-LNP improved the stability of the siRNA in serum. Fluorescence-labeled siRNA formulated in CPP-LNP was efficiently internalized into B16F10 murine melanoma cells in a time-dependent manner, although that in LNP without CPP was hardly internalized into these cells. In cells transfected with siRNA in CPP-LNP, most of the siRNA was distributed in the cytoplasm of these cells and did not localize in the lysosomes. Analysis of the endocytotic pathway indicated that CPP-LNP were mainly internalized via macropinocytosis and heparan sulfate-mediated endocytosis. CPP-LNP encapsulating siRNA effectively induced RNA interference-mediated silencing of reporter genes in B16F10 cells expressing luciferase and in HT1080 human fibrosarcoma cells expressing enhanced green fluorescent protein. These data suggest that modification of LNP with the protamine-derived CPP was effective to facilitate internalization of siRNA in the cytoplasm and thereby to enhance gene silencing.

Radiation increases the cellular uptake of exosomes through CD29/CD81 complex formation.

Exosomes mediate intercellular communication, and mesenchymal stem cells (MSC) or their secreted exosomes affect a number of pathophysiologic states. Clinical applications of MSC and exosomes are increasingly anticipated. Radiation therapy is the main therapeutic tool for a number of various conditions. The cellular uptake mechanisms of exosomes and the effects of radiation on exosome-cell interactions are crucial, but they are not well understood. Here we examined the basic mechanisms and effects of radiation on exosome uptake processes in MSC. Radiation increased the cellular uptake of exosomes. Radiation markedly enhanced the initial cellular attachment to exosomes and induced the colocalization of integrin CD29 and tetraspanin CD81 on the cell surface without affecting their expression levels. Exosomes dominantly bound to the CD29/CD81 complex. Knockdown of CD29 completely inhibited the radiation-induced uptake, and additional or single knockdown of CD81 inhibited basal uptake as well as the increase in radiation-induced uptake. We also examined possible exosome uptake processes affected by radiation. Radiation-induced changes did not involve dynamin2, reactive oxygen species, or their evoked p38 mitogen-activated protein kinase-dependent endocytic or pinocytic pathways. Radiation increased the cellular uptake of exosomes through CD29/CD81 complex formation. These findings provide essential basic insights for potential therapeutic applications of exosomes or MSC in combination with radiation.

White opaque substance visualized using magnifying endoscopy with narrow-band imaging in colorectal epithelial neoplasms.

BACKGROUND: The presence of a white opaque substance (WOS) on magnifying endoscopy (ME) with narrow-band imaging (NBI) has been reported for gastric epithelial neoplasms, but the presence of WOS in colorectal epithelial neoplasms has not been investigated. AIMS: The purpose of this study was to determine whether WOS is present in colorectal epithelial neoplasms and to clarify its clinical significance. METHODS: A total of 590 colorectal epithelial neoplasms from 368 consecutive patients were retrospectively analyzed using prospectively collected data. Presence or absence of WOS in colorectal epithelial neoplasms was recorded based on the findings of ME with NBI. RESULTS: White opaque substance was present in 236 of the 590 (40 %) colorectal epithelial neoplasms. Compared with WOS-negative patients, WOS-positive patients showed significantly larger tumors (p < 0.0001) and significantly more tumors in the proximal colon (p = 0.0003). WOS was more frequently present in carcinomas (66.0 %) than in adenomas (31.8%; p < 0.0001). WOS was also more frequent in submucosal carcinomas (75.9%) than in intramucosal carcinomas (59.0%; p = 0.0380). CONCLUSIONS: This study confirmed the presence of WOS in colorectal epithelial neoplasms, and prevalence increased with the progression of cancer, from adenoma to carcinoma and from intramucosal carcinoma to submucosal carcinoma.

High-energy SR-XRF Imaging of Cesium and Trace Elements in Mouse Kidneys: Short Communication.

For understanding trace element dynamics in tissues, methods for analyzing elemental distribution and localization without destroying tissue structures and cell arrangements are desired. Synchrotron radiation X-ray fluorescence (SR-XRF) analysis is one of the non-destructive and multi-element simultaneous analyses. The kidney is the major excretion pathway of cesium (Cs) taken into the body, and an understanding of cesium distribution in the kidney would be useful for establishing technology to facilitate the excretion of radioactive Cs from the body due to nuclear disasters. In the present study, the distribution of cesium and trace elements, such as iron (Fe) and zinc (Zn), corresponding to the kidney structure was examined in Cs-administered mice by SR-XRF imaging with high-energy excitation X-rays (40 keV). By beam scanning with a 200-µm square beam, clear Cs images corresponding to the renal layer structure were obtained for the renal specimen at the early phase after Cs administration with the mean renal Cs concentration of 24.1 ± 3.2 µg/g. Cs was distributed mainly in the medulla and the outer stripe of the outer medulla located in the center area of the kidney. Unlike the Cs distribution, endogenous Fe and Zn tended to be lower in the medulla than in the outer stripe of the outer medulla and the cortex. This method is effective for analyzing Cs distribution because it can simultaneously analyze the distribution of endogenous trace elements.

Effectiveness of concomitant enteral nutrition therapy and infliximab for maintenance treatment of Crohn's disease in adults.

BACKGROUND: One of the problems associated with infliximab (IFX) treatment for Crohn's disease (CD) is loss of response during maintenance therapy. AIMS: The aim of this multicenter, retrospective, cohort study was to determine whether enteral nutrition (EN) added to the IFX therapy regimen is effective for maintaining remission in adult CD patients. METHODS: Patients with CD who had started IFX therapy between April 2003 and March 2008 at any one of the seven participating medical centers and who met the following inclusion criteria were enrolled in the study: remission after triple infusions of IFX followed by IFX maintenance therapy every 8 weeks, and follow-up data available for ≥ 1 year. Remission was defined as a C-reactive protein (CRP) level of <0.3 mg/dL, and recurrence was defined as an increase in CRP to ≥ 1.5 mg/dL or shortening of the IFX interval. Patients were classified by EN dosage into two groups (EN group and non-EN group). The cumulative remission period and related factors were analyzed. RESULTS: Of the 102 adult CD patients who met the inclusion criteria, 45 were in the EN group and 57 were in the non-EN group. The cumulative remission rate was significantly higher in the EN group than in the non-EN group (P = 0.009). Multivariate analysis revealed that EN was the only suppressive factor for disease recurrence (P = 0.01). CONCLUSIONS: The results demonstrate that among this CD patient cohort, EN combined with IFX maintenance treatment was clinically useful for maintaining remission.

Current status and prospects of polyethyleneglycol-modified medicines.

Polyethyleneglycol (PEG) compounds have a large hydrogen bonding capability and possess various functions that depend on the length of the chain and conformational diversity. Modification of a drug with PEG is a well-known technology for improving the physicochemical properties and biological responses of a drug. There are many reports about the modification of small molecules with PEG, however, there are no modified small molecule products currently on the market. Several protein products for medical use are commercially available. In this review, the effects of modification with PEG on biopharmaceuticals are described through the comparison of two interferon-α products modified with PEG, one with 12 kDa linear PEG and the other 40 kDa branched PEG. There is one original drug delivery system product, Doxil(®)/Caelyx(®), on the market in which liposomes modified with PEG and encapsulating doxorubicin are stabilized sterically and invisible to the reticuloendothelial system. The benefits of modification with PEG are described here with examples of modified products on the market and used in clinical trials.

Development of liposomal anticancer drugs.

Liposomes are drug delivery systems that can alter the pharmacokinetic properties of compounds. The adverse effects of anticancer agents are a limiting factor for cancer chemotherapy, therefore, liposomal formulations have the potential to improve the therapeutic efficacy of anticancer agents by enhancing their accumulation in tumors and reducing non-selective distribution to normal tissues, which is known as the enhanced permeability and retention effect. To develop a liposomal anticancer agent as a drug product, its formulation must be designed to ensure its quality until it is administered to patients and to exert maximum potency in clinical use rather than in animal experiments. The chemical stability and physicochemical stability of the ingredients are key factors in the design of liposomal formulations. Drug release rates are critical factors in the therapeutic efficacy of liposomal drug products because the encapsulated drug has no pharmacological activity, and only released drug can exert antitumor/toxic activities. Liposomes should maintain the drug in a stable state in the circulation and then promptly release it after accumulation in the target tissue in order to achieve a sufficient drug concentration. To understand the profile of the formulation and to guarantee the quality of drug product, a reliable analytical method that can determine the released and encapsulated drugs in biological fluids is required. Simple online solid phase extractions of the released and encapsulated drugs using a column-switching HPLC system meet the requirements and this system enables accurate in vitro release testing and in vivo pharmacokinetic evaluation. This review introduces the process of liposomal drug product development from various viewpoints.

[A case of so-called carcinosarcoma of the middle portion of the bile duct diagnosed preoperatively].

A 73-year-old man was admitted to a hospital with a complaint of epigastralgia and for evaluation of liver dysfunction. After hospitalization, he experienced disturbance of consciousness with septic shock, and was then transferred to our hospital. Computed tomography revealed dilatation of the intrahepatic bile duct and tumor of the middle bile duct. We diagnosed acute obstructive suppurative cholangitis. As a result, endoscopic nasobiliary drainage was performed, and the patient recovered. Based on pathological examinations of the bile duct biopsy specimen, the tumor was diagnosed as a carcinosarcoma. Consequently, the patient underwent pylorus-preserving pancreatoduodenectomy. However, 4 months after surgery, the patient died due to widespread metastasis of the carcinosarcoma. Preoperative diagnosis of carcinosarcoma of the bile duct is extremely rare. Our study suggests the efficacy of bile duct biopsy in such cases.