Effects of stimbiotic supplementation on gut health, immune response, and intestinal microbiota in weaned piglets challenged with E. coli.

In order to make piglet diets more effective, it is necessary to investigate effective methods for breaking down xylan in cereal. The objective of this study was to determine the effects of dietary stimbiotic (STB) supplementation on growth performance, intestinal morphology, immune response and intestinal microbiota in weaned piglets. A total of 24 (Duroc × Yorkshire × Landrace) weaned pigs (initial body weight of 8.01 ± 0.38 kg and 28 ± 3 d old), were assigned to 4 treatments with 6 replicates per treatment. Pigs were housed in individual pens for 17 days, including 5 days adaption period and 12 days after the first Escherichia coli (E. coli) challenge. The experiment was conducted in a 2 × 2 factorial arrangement of treatments consisting of two levels of challenge (challenge and non-challenge) and two levels of STB (0 and 0.5 g/kg diet). Supplementations of STB 0.5 g/kg improved the gain to feed ratio (G:F) (P < 0.05) in piglets challenged with shiga toxigenic E. coli (STEC). STB supplementation decreased (P < 0.05) white blood cells, neutrophils, lymphocytes, and expression levels of tumor necrosis factor-alpha and interleukin-6. Supplementation of STB improved (P < 0.05) the lymphocytes and neutrophils in piglets challenged with STEC on 12 dpi. Supplementation of STB also improved (P < 0.05) the villus height to-crypt depth ratio of ileum in piglets challenged with STEC. Supplementation of STB increased (P < 0.05) the expression levels of claudin-1 of ileum. In genus level, supplementation of STB increased (P < 0.001) the abundance of Prevotella compared to non-supplementation of STB groups in pre-inoculation period. Also, supplementation of STB decreased (P < 0.05) the abundance of Faecalibacterium and Eubacterium_coprostanoligenes_group compared to non-supplementation of STB groups in post-inoculation period. In phylum level, supplementation of STB increased (P < 0.05) the abundance of Desulfobacterota and Fibrobacterota in pre-inoculation period. E. coli challenge increased the abundance of Fibrobacterota compared to non-challenged group in post-inoculation period. In conclusion, these findings indicated that STB supplementation could alleviate a decrease of the performance, immune response, and inflammatory response in piglets induced by the STEC challenge.

Topical Cream Carrying Drug-Loaded Nanogels for Melanoma Treatment.

In this study, nanogel creams carrying paclitaxel (PTX) and temozolomide (TMZ) were prepared for the topical treatment of melanoma. PTX and TMZ were first loaded in poly-(D,L-lactide-co-glycolide)-block-poly(ethylene glycol)-block-poly-(D,L-lactide-co-glycolide) (PLAG-b-PEG-b-PLGA) thermosensitive nanogels, which made a transition from a free-flowing sol (formation of micellar network) at 25°C with the z-average particle size of c.a. 96 nm to a gel (aggregation of micelles) at 33°C with the z-average particle size of c.a. 427 nm. An anhydrous absorption ointment base, aquaphor, was then added to drug-loaded nanogels to form nanogel creams carrying PTX and TMZ. Nanogel creams permitted controlled release of the payloads and improved the penetration of the payloads through the rodent skin compared to drug(s)-loaded nanogels. PTX and TMZ in a combination were synergistically effective in inhibiting SK-MEL28, A375, and B16-F10 melanoma cancer cells in vitro. Topically applied nanogel creams carrying TMZ/PTX (4 mg/1.5 mg/dose) showed a trend of tumor volume inhibition on B16-F10-bearing xenograft mice in vivo.

Stimbiotic Supplementation Alleviates Poor Performance and Gut Integrity in Weaned Piglets Induced by Challenge with E. coli.

The aim of this study was to investigate the effects of stimbiotic (STB), a xylanase and xylo-oligosaccharide complex. A total of 36 male weaned pigs with initial body weights of 8.49 ± 0.10 kg were used in a 3-week experiment. The experiment was conducted in a 2 × 3 factorial arrangement (six replicates/treatment) of treatments consisting of two levels of challenge (challenge and non-challenge) and three levels of STB (0, 0.5, and 1 g/kg diet). Supplementations STB 0.5 g/kg (STB5) and STB 1 g/kg (STB10) improved the G:F (p = 0.04) in piglets challenged with STEC. STB supplementation, which also decreased (p < 0.05) the white blood cells, neutrophils, lymphocytes, and expression levels of tumor necrosis factor-alpha and interleukin-6. Supplementations STB5 and STB10 improved (p < 0.01) the lymphocytes and neutrophils in piglets challenged with STEC on 14 dpi. Additionally, supplementations STB5 and STB10 improved (p < 0.01) the tumor necrosis factor-alpha in piglets challenged with STEC on 3 dpi. Supplementations STB5 and STB10 also improved the villus height-to-crypt depth ratio (p < 0.01) in piglets challenged with STEC. Supplementation with STB reduced (p < 0.05) the expression levels of calprotectin. In conclusion, STB could alleviate a decrease of the performance, immune response, and inflammatory response induced by the STEC challenge.

Nanogels as a Versatile Drug Delivery System for Brain Cancer.

Chemotherapy and radiation remain as mainstays in the treatment of a variety of cancers globally, yet some therapies exhibit limited specificity and result in harsh side effects in patients. Brain tissue differs from other tissue due to restrictions from the blood-brain barrier, thus systemic treatment options are limited. The focus of this review is on nanogels as local and systemic drug delivery systems in the treatment of brain cancer. Nanogels are a unique local or systemic drug delivery system that is tailorable and consists of a three-dimensional polymeric network formed via physical or chemical assembly. For example, thermosensitive nanogels show promise in their ability to incorporate therapeutic agents in nano-structured matrices, be applied in the forms of sprays or sols to the area from which a tumor has been removed, form adhesive gels to fill the cavity and deliver treatment locally. Their usage does come with complications, such as handling, storage, chemical stability, and degradation. Despite these limitations, the current ongoing development of nanogels allows patient-centered treatment that can be considered as a promising tool for the management of brain cancer.

Development of 3D-Printed Layered PLGA Films for Drug Delivery and Evaluation of Drug Release Behaviors.

3D printing has been widely used to rapidly manufacture a variety of solid dosage forms on-demand, without sacrificing precision. This study used extrusion-based 3D printing to prepare single-layered, tri-layered, and core-in-shell poly(lactic-co-glycolic acid) (PLGA) films carrying paclitaxel and rapamycin in combination or lidocaine alone. Each layer was composed of either low molecular weight (MW) PLGA or high MW PLGA. In vitro drug release kinetics of paclitaxel, rapamycin, and lidocaine for PLGA films were assessed and compared with PLGA-polyethylene glycol (PEG)-PLGA hydrogel discs. Regardless of the structure of PLGA film, paclitaxel (half-time: 54-63 days) was released faster than when compared with rapamycin (half-time: 74-80 days). In contrast, single-layered PLGA-PEG-PLGA discs released rapamycin (half-time 5.7 h) at a more rapid rate than paclitaxel (half-time: 7.3 h). Single-layered PLGA-PEG-PLGA discs enabled a faster drug release than PLGA films, noting that the disc matrices dissolve in water in 24 h. Similarly, lidocaine incorporated in PLGA films (half-time: 13-36 days) exhibited slower release patterns than that in PLGA-PEG-PLGA discs (half-time: 2.6 h). In vitro drug release patterns were explained using molecular models that simulate drug-polymer interactions. Analysis of models suggested that drug-polymer interactions, location of each drug in the polymeric matrix, and solubility of drugs in water were major factors that determine drug release behaviors from the polymeric films and discs.

3D Printing of Poloxamer 407 Nanogel Discs and Their Applications in Adjuvant Ovarian Cancer Therapy.

Nanogels are attractive biocompatible materials that enable local delivery of multiple drugs. In this study, we demonstrated that 3D printing technology could be used to precisely construct nanogel discs carrying paclitaxel and rapamycin. 3D-printed nanogel disc rounds (12 mm diameter × 1 mm thickness) carrying paclitaxel and rapamycin evaded premature gelation during storage and the initial burst release of the drugs in the dissolution medium. In vivo 3D-printed nanogel discs permitted successful intraperitoneal delivery of paclitaxel and rapamycin in ES-2-luc ovarian-cancer-bearing xenograft mice. They were also shown to be therapeutically effective and capable of preventing postsurgical peritoneal adhesions in the treated xenograft mice.

Triolimus: A Multi-Drug Loaded Polymeric Micelle Containing Paclitaxel, 17-AAG, and Rapamycin as a Novel Radiosensitizer.

Triolimus is a multi-drug loaded polymeric micelle containing paclitaxel (PTX), 17-allylamino-17-demethoxygeldanamycin (17-AAG), and rapamycin (RAP). This study examines the radiosensitizing effect of Triolimus in vitro and in vivo. Radiosensitizing effects of Triolimus on A549 cells are dose dependent and at 2 × 10-9 m, Triolimus shows significant radiosensitization even at low radiation doses (2 Gy). By sensitivity enhancement ratio, PTX alone, dual drug combinations, and Triolimus treatment at 2 × 10-9 m have radiosensitizing effects with potency as follows: PTX alone (PTX) > PTX and RAP (P/R) > Triolimus (TRIO) > PTX and 17-AAG (P/17) >17-AAG and RAP (17/R). In vivo, fractionated radiation of 15 Gy preceded by infusion of PTX alone, dual drug combinations, or an intermediate dose of Triolimus (Int. TRIO: PTX/17-AAG/RAP at 15/15/7.5 mg kg-1 ) strongly inhibits A549 tumor growth. Notably, pretreatment with high dose of Triolimus (High TRIO: PTX/17-AAG/RAP at 60/60/30 mg kg-1 ) before the fractionated radiation leads to tumor control for up to 24 weeks. An enhanced radiosensitizing effect is observed without an increase in acute toxicity compared to PTX alone or radiation alone. These results suggest that further investigations of Triolimus in combination with radiation therapy are merited.

Proof-of-Concept of Polymeric Sol-Gels in Multi-Drug Delivery and Intraoperative Image-Guided Surgery for Peritoneal Ovarian Cancer.

PURPOSE: The purpose of this study is to investigate a sol-gel transition property and content release profiles for thermosensitive poly-(D,L-lactide-co-glycolide)-block-poly-(ethylene glycol)-block-poly-(D,L-lactide-co-glycolide) (PLGA-b-PEG-b-PLGA) hydrogels carrying paclitaxel, rapamycin, and LS301, and to present a proof-of-concept that PLGA-b-PEG-b-PLGA hydrogels carrying paclitaxel, rapamycin, and LS301, called TheranoGel, exhibit excellent theranostic activity in peritoneal ES-2-luc ovarian cancer xenograft mice. METHODS: Thermosensitive PLGA-b-PEG-b-PLGA hydrogels carrying paclitaxel, rapamycin, and LS301, individually or in combination, were prepared via a lyophilization method, characterized with content release kinetics, and assessed with theranostic activity in ES-2-luc xenograft mice. RESULTS: A thermosensitive PLGA-b-PEG-b-PLGA sol-gel system was able to entrain 3 poorly water-soluble payloads, paclitaxel, rapamycin, and LS301 (TheranoGel). TheranoGel made a sol-to-gel transition at 37°C and slowly released 3 drugs at a simultaneous release rate in response to the physical dissociation of hydrogels in vitro. TheranoGel enabled loco-regional delivery of multi-drugs by forming a gel-depot in the peritoneal cavity of ES-2-luc xenograft mice. An intraperitoneal (IP) administration of TheranoGel resulted in excellent therapeutic and diagnostic activities, leading to the improved peritoneal surgery in ES-2-luc xenograft mice. CONCLUSIONS: TheranoGel prepared via a facile lyophiliation method enabled successful IP delivery of multi-drugs and exhibited excellent theranostic activity in vivo.

PEG-b-PLA micelles and PLGA-b-PEG-b-PLGA sol-gels for drug delivery.

Poly(ethylene glycol)-block-poly(D,L-lactic acid) (PEG-b-PLA) micelles and poly(D,L-lactic-co-glycolic acid)-block-polyethylene glycol)-block-poly(D,L-lactic-co-glycolic acid) (PLGA-b-PEG-b-PLGA) sol-gels have been extensively researched for systemic and localized drug delivery applications, respectively, and they have both progressed into humans for paclitaxel, an important yet poorly water-soluble chemotherapeutic agent. In this review article, preclinical and clinical research on PEG-b-PLA micelles and PLGA-b-PEG-b-PLGA sol-gels that has focused on paclitaxel will be updated, and recent research on other poorly water-soluble anticancer agents and delivery of drug combinations (i.e. multi-drug delivery) that seeks synergistic anticancer efficacy will be summarized. PEG-b-PLA micelles are a first-generation platform for the systemic multi-delivery of poorly water soluble anticancer agents. PLGA-b-PEG-b-PLGA sol-gels are a first-generation platform for the localized multi-drug delivery of water-soluble and/or poorly water-soluble anticancer agents. In summary, PEG-b-PLA micelles and PLGA-b-PEG-b-PLGA sol-gels may safely enable pre-clinical evaluation and clinical translation of poorly water-soluble anticancer agents, especially for promising, rapidly emerging anticancer combinations.

Hydrogel-Based Drug Delivery Systems for Poorly Water-Soluble Drugs.

Hydrogels are three-dimensional materials that can withstand a great amount of water incorporation while maintaining integrity. This allows hydrogels to be very unique biomedical materials, especially for drug delivery. Much effort has been made to incorporate hydrophilic molecules in hydrogels in the field of drug delivery, while loading of hydrophobic drugs has not been vastly studied. However, in recent years, research has also been conducted on incorporating hydrophobic molecules within hydrogel matrices for achieving a steady release of drugs to treat various ailments. Here, we summarize the types of hydrogels used as drug delivery vehicles, various methods to incorporate hydrophobic molecules in hydrogel matrices, and the potential therapeutic applications of hydrogels in cancer.

Polymeric micelles for multi-drug delivery in cancer.

Drug combinations are common in cancer treatment and are rapidly evolving, moving beyond chemotherapy combinations to combinations of signal transduction inhibitors. For the delivery of drug combinations, i.e., multi-drug delivery, major considerations are synergy, dose regimen (concurrent versus sequential), pharmacokinetics, toxicity, and safety. In this contribution, we review recent research on polymeric micelles for multi-drug delivery in cancer. In concurrent drug delivery, polymeric micelles deliver multi-poorly water-soluble anticancer agents, satisfying strict requirements in solubility, stability, and safety. In sequential drug delivery, polymeric micelles participate in pretreatment strategies that "prime" solid tumors and enhance the penetration of secondarily administered anticancer agent or nanocarrier. The improved delivery of multiple poorly water-soluble anticancer agents by polymeric micelles via concurrent or sequential regimens offers novel and interesting strategies for drug combinations in cancer treatment.

Thermosensitive poly-(d,l-lactide-co-glycolide)-block-poly(ethylene glycol)-block-poly-(d,l-lactide-co-glycolide) hydrogels for multi-drug delivery.

A current treatment strategy for peritoneal ovarian cancer is a combination of peritoneal surgery and multi-drug-based chemotherapy that often involves intraperitoneal (IP) injection. A thermosensitive poly-(D,L-lactide-co-glycolide)-block-poly(ethylene glycol)-block-poly-(D,L-lactide-co-glycolide) (PLGA-b-PEG-b-PLGA) hydrogel platform (thermogels) enabled gel loading of poorly work-soluble paclitaxel (cytotoxic agent), 17-allylamino-17-demethoxygeldanamycin (17-AAG, heat shock protein inhibitor), and rapamycin (mammalian target of rapamycin protein inhibitor). PLGA-b-PEG-b-PLGA thermogels (15%) carrying paclitaxel, 17-AAG, and rapamycin (named Triogel) made a successful transition from a free-flowing solution below ambient temperature to a gel depot at body temperature. Triogel gradually released paclitaxel, 17-AAG, and rapamycin at an equal release rate in response to the physical gel erosion. In an ES-2-luc ovarian cancer xenograft model, a single IP injection of Triogel (60, 60, and 30 mg/kg of paclitaxel, 17-AAG, and rapamycin, respectively) significantly reduced tumor burden and prolonged survival of ES-2-luc-bearing nude mice without notable systemic toxicity relative to those delivered by poly(ethylene glycol)-block-poly(d,l-lactic acid) (PEG-b-PLA) micelles in solution via IP or intravenous (IV) injection route. These results show a great potential of a biodegradable thermogel platform carrying multi-drugs for IP chemotherapy in peritoneal ovarian cancer.

Polymeric micelles for apoptosis-targeted optical imaging of cancer and intraoperative surgical guidance.

In a two-step strategy, an intraperitoneal (IP) injection of poly(ethylene glycol)-block-poly(ε-caprolactone) (PEG-b-PCL) micelles containing paclitaxel (PTX), cyclopamine (CYP), and gossypol (GSP) at 30, 30, and 30 mg/kg, respectively, debulked tumor tissues by 1.3-fold, based on loss of bioluminescence with <10% body weight change, and induced apoptosis in peritoneal tumors when used as neoadjuvant chemotherapy (NACT) in an ES-2-luc-bearing xenograft model for ovarian cancer. In a second step, a single intravenous (i.v.) injection of apoptosis-targeting GFNFRLKAGAKIRFGS-PEG-b-PCL micelles containing a near-infrared (NIR) fluorescence probe, DiR (1,1'-dioctadecyltetramethyl indotricarbocyanine iodide), resulted in increased peritoneal DiR accumulation in apoptosis-induced ES-2-luc tumor tissues (ex vivo) by 1.5-fold compared with DiR molecules delivered by methoxy PEG-b-PCL micelles (non-targeted) at 48 h after i.v. injection in a second step. As a result, a tandem of PEG-b-PCL micelles enabled high-resolution detection of ca. 1 mm diameter tumors, resulting in resection of approximately 90% of tumors, and a low peritoneal cancer index (PCI) of ca. 7. Thus, a tandem of PEG-b-PCL micelles used for NCAT and NIR fluorescence imaging of therapy-induced apoptosis for intraoperative surgical guidance may be a promising treatment strategy for metastatic ovarian cancer.

Poly(ethylene glycol)-block-poly(ε-caprolactone) micelles for combination drug delivery: evaluation of paclitaxel, cyclopamine and gossypol in intraperitoneal xenograft models of ovarian cancer.

Ovarian cancer is the most lethal gynecological malignancy, characterized by a high rate of chemoresistance. Current treatment strategies for ovarian cancer focus on novel drug combinations of cytotoxic agents and molecular targeted agents or novel drug delivery strategies that often involve intraperitoneal (IP) injection. Poly(ethylene glycol)-block-poly(ε-caprolactone) (PEG-b-PCL) micelles were loaded with paclitaxel (cytotoxic agent), cyclopamine (hedgehog inhibitor), and gossypol (Bcl-2 inhibitor). After physicochemical studies focusing on combination drug solubilization, 3-drug PEG-b-PCL micelles were evaluated in vitro in 2-D and 3-D cell culture and in vivo in xenograft models of ovarian cancer, tracking bioluminescence signals from ES-2 and SKOV3 human ovarian cancer cell lines after IP injection. 3-Drug PEG-b-PCL micelles were not significantly more potent in 2-D cell culture in comparison to paclitaxel; however, they disaggregated ES-2 tumor spheroids, whereas single drugs or 2-drug combinations only slowed growth of ES-2 tumor spheroids or had no noticeable effects. In ES-2 and SKOV3 xenograft models, 3-drug PEG-b-PCL micelles had significantly less tumor burden than paclitaxel based on bioluminescence imaging, 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) PET imaging, and overall survival. (18)F-FLT-PET images clearly showed that 3-drug PEG-b-PCL micelles dramatically reduce tumor volumes over paclitaxel and vehicle controls. In summary, PEG-b-PCL micelles enable the IP combination drug delivery of paclitaxel, cyclopamine and gossypol, resulting in tumor growth inhibition and prolonged survival over paclitaxel alone. These results validate a novel treatment strategy for ovarian cancer based on drug combinations of cytotoxic agents and molecular targeted agents, delivered concurrently by a nanoscale drug delivery system, e.g. PEG-b-PCL micelles.

Pharmacokinetic study of 3-in-1 poly(ethylene glycol)-block-poly(D, L-lactic acid) micelles carrying paclitaxel, 17-allylamino-17-demethoxygeldanamycin, and rapamycin.

Concurrent delivery of multiple poorly water-soluble anticancer drugs has been a great challenge due to the toxicities exerted by different surfactants or organic solvents used in solubilizing individual drugs. We previously found that poly(ethylene glycol)-block-poly(D, L-lactic acid) (PEG-b-PLA) micelles can serve as a safe delivery platform for simultaneous delivery of paclitaxel (PTX), 17-allylamino-17-demethoxygeldanamycin (17-AAG), and rapamycin (RAP) to mice. The high tolerance of this polymeric micelle formulation by mice allowed us to investigate the pharmacokinetics of the 3 co-delivered drugs. In this study, it was shown that 3-in-1 PEG-b-PLA micelle delivering high doses of PTX, 17-AAG, and RAP (60, 60, and 30 mg/kg, respectively) significantly increased the values of the area under the plasma concentration-time curves (AUC) of PTX and RAP in mice compared to the drugs delivered individually, while the pharmacokinetic parameters of 17-AAG were similar in both 3-in-1 and single drug-loaded PEG-b-PLA micelle formulations. Moreover, pharmacokinetic study using 2-in-1 micelles indicated that the augmented AUC value of RAP was due to the co-delivery of 17-AAG, while the increase in AUC of PTX was more likely caused by the co-delivery of RAP. In contrast, when 3-in-1 and single drug-loaded PEG-b-PLA micelles were administrated at modest dose (PTX, 17-AAG, and RAP at 10, 10, and 5 mg/kg, respectively), pharmacokinetic differences of individual drugs between 3-in-1 and single drug formulations were eliminated. These results suggest that 3-in-1 PEG-b-PLA micelles can concurrently deliver PTX, 17-AAG, and RAP without changing the pharmacokinetics of each drug at modest doses, but altered pharmacokinetic profiles emerge when drugs are delivered at higher doses.

In vivo cancer imaging by poly(ethylene glycol)-b-poly(ɛ-caprolactone) micelles containing a near-infrared probe.

Noninvasive near-infrared (NIR) fluorescence imaging is a promising technique for the intraoperative assessment of solid tumor removal. We incorporated a lipophilic NIR probe, 1,1'-dioctadecyltetramethyl indotricarbocyanine iodide (DiR), in poly(ethylene glycol)-b-poly(ɛ-caprolactone) (PEG-b-PCL) micelles, resulting in DiR solubilization in water, occupying nanoscopic PEG-b-PCL micelles. DiR in a self-quenched or nonquenched state showed different kinetics of release from PEG-b-PCL micelles in vitro; however, both obtained high tumor delineation (tumor-to-muscle ratio of 30-43 from collected organs). These results suggest that PEG-b-PCL micelles with DiR are a promising nanosized imaging agent that will provide a basis for enhanced surgical guidance via NIR visualization of tumors. FROM THE CLINICAL EDITOR: In this paper, noninvasive near-infrared fluorescence imaging coupled with specific lipophilic probes is discussed as a promising technique for intraoperative assessment of solid tumor removal, leading to optimized outcomes for in toto removal of tumors.

Polymeric micelles for neoadjuvant cancer therapy and tumor-primed optical imaging.

Poly(ethylene glycol)-block-poly(d,l-lactic acid) (PEG-b-PLA) micelles act as a 3-in-1 nanocontainer for three poorly water-soluble drugs-paclitaxel, 17-allylamino-17-demethoxygeldanamycin, and rapamycin (PTX/17-AAG/RAPA)-for cancer therapy. In a LS180 human colon xenograft model, a single intravenous (IV) injection of 3-in-1 PEG-b-PLA micelles reduced tumor volume by 1.6-fold with <10% body weight change. In a second step, IV injection of poly(ethylene glycol)-block-poly(ε-caprolactone) (PEG-b-PCL) micelles carrying a carbocyanine dye (1,1'-dioctadecyl tetramethyl indotricarbocyanine iodide (DiR)) after 48 h resulted in a 2.1-fold higher near-infrared (NIR) optical signal from excised solid tumors versus a negative control, presumably due to a reduction in tumor cell density and interstitial tumor pressure. Thus, a tandem of 3-in-1 PEG-b-PLA and PEG-b-PCL micelles could potentially be used for neoadjuvant cancer therapy and tumor-primed NIR optical imaging for intraoperative surgical guidance in oncology, offering a promising multimodal strategy for cancer therapy and imaging.

A 3-in-1 polymeric micelle nanocontainer for poorly water-soluble drugs.

Poly(ethylene glycol)-block-poly(D,L-lactic acid) (PEG-b-PLA) micelles have a proven capacity for drug solubilization and have entered phase III clinical trials as a substitute for Cremophor EL in the delivery of paclitaxel in cancer therapy. PEG-b-PLA is less toxic than Cremophor EL, enabling a doubling of paclitaxel dose in clinical trials. We show that PEG-b-PLA micelles act as a 3-in-1 nanocontainer for paclitaxel, 17-allylamino-17-demethoxygeldanamycin (17-AAG), and rapamycin for multiple drug solubilization. 3-in-1 PEG-b-PLA micelles were ca. 40 nm in diameter; dissolved paclitaxel, 17-AAG, and rapamycin in water at 9.0 mg/mL; and were stable for 24 h at 25 °C. The half-life for in vitro drug release (t(1/2)) for 3-in-1 PEG-b-PLA micelles was 1-15 h under sink conditions and increased in the order of 17-AAG, paclitaxel, and rapamycin. The t(1/2) values correlated with log P(o/w) values, implicating a diffusion-controlled mechanism for drug release. The IC(50) value of 3-in-1 PEG-b-PLA micelles for MCF-7 and 4T1 breast cancer cell lines was 114 ± 10 and 25 ± 1 nM, respectively; combination index (CI) analysis showed that 3-in-1 PEG-b-PLA micelles exert strong synergy in MCF-7 and 4T1 breast cancer cell lines. Notably, concurrent intravenous (iv) injection of paclitaxel, 17-AAG, and rapamycin using 3-in-1 PEG-b-PLA micelles was well-tolerated by FVB albino mice. Collectively, these results suggest that PEG-b-PLA micelles carrying paclitaxel, 17-AAG, and rapamycin will provide a simple yet safe and efficacious 3-in-1 nanomedicine for cancer therapy.

Suppression of hepatitis B virus-derived human hepatocellular carcinoma by NF-kappaB-inducing kinase-specific siRNA using liver-targeting liposomes.

Hepatitis B virus triggers an increase of NF-kappaB inducing kinase (NIK)-dependent NF-kappaB activation, followed by the promotion of hepatocellular carcinoma (HCC). Here, we examined the inhibitory effect of NIK-specific siRNA on NF-kappaB signaling and HCC. The results of this study indicated that these siRNAs suppressed HBV-derived HCC by regulating NIK activation. To exert a protective effect from degradation enzyme, cationic liposomes were contrived and modified to contain beta-sitosterol glucoside to target the asialoglycoprotein receptors in liver cancer cells. The cationic dimyristoyl diacyltrimethylammonium propane liposomes were prepared by a reverse-phase evaporation method with slight modification. beta-Sitosterol glucoside was added to the lipid mixture at the beginning of the liposome preparation for the purpose of liver targeting. These liposomes assisted the delivery of the siRNA to specific cells and protected it from various lyases, followed by the ultimate suppression of HCC.

Preparation of pH-sensitive, long-circulating and EGFR-targeted immunoliposomes.

A long-circulating formulation of pH-sensitive liposomes (PSLs) with antibodies against epidermal growth factor receptor (EGFR) attached was designed, expecting an increase in binding and delivery of liposomes to the target cells including non-small cell lung cancer (NSCLC) cells. Physicochemical properties of the PSLs were measured by SEM and DLS. Leakage of a self-quenching fluorescent probe, calcein, from the liposome was studied for the evaluation of pH-sensitivity. Encapsulation efficiency of gemcitabine (an anti-cancer drug) in PSLs was about 67%. Average size of liposomes was 88 nm in diameter. The PSL of DOPE/CHEMS (6:4 molar ratio) formulation showed a dramatic pH-sensitivity at/around pH 5.5, whereas non-PSL of DPPC/Chol or PC/CHEMS formulation did not. Anti-proliferation effect of gemcitabine-encapsulating PSLs & Ab-PSLs in A549 cells was 2-fold higher than the free drug, which was further elucidated by the apoptosis of the cells by gemcitabine (approximately 10% apoptosis for PSL or Ab-PSL formulation vs. approximately 1% for free drug or non-PSL formulation) using FACS analysis. These data demonstrate delivery of gemcitabine to tumor cells can be improved by long-circulating PSLs or Ab-PSLs formulations in vitro.