Liposomal Agents Research Articles

Preparation of AS1411 aptamer-modified PEGylated liposomal doxorubicin and evaluation of its anti-cancer effects in vitro and in vivo

Active targeting of the tumor microenvironment could be a helpful strategy to reduce toxicity and drug resistance. Encapsulating anticancer agents in liposomes has improved tumor accumulation and decreased non-specific toxicity. In this study, the potential of nucleolin-targeted PEGylated liposomal doxorubicin (PLD) to convey doxorubicin into tumor was compared to PLD. The effect of liposome coupled AS1411 aptamer (AS-PLD) was assessed via cytotoxicity, competition assay, and cellular uptake in vitro. Additionally, liposome biodistribution and pharmacokinetic analysis as well as its therapeutic efficacy were assessed in mice C26 tumor models. The cytotoxicity and cellular uptake results demonstrated that AS-PLD is more potent than PLD in killing cancer cells. The competition assay indicated the specificity of the AS-PLD in targeting C26 cells in vitro. There was more AS-PLD accumulation in the tumor after 72 h post-injection. The present study demonstrated the more therapeutic efficiency of the AS-PLD compared to PLD, making this formulation a proper active-targeted formulation for cancer treatment.

Abstract PR007: A liposomal priming agent increases the sensitivity of liquid biopsies

Abstract Liquid biopsy measurements, such as analysis of circulating tumor DNA (ctDNA) shed by cancer cells, have garnered significant attention for their potential to empower the field of precision oncology. Given that ctDNA tests could enable minimally invasive monitoring and molecular profiling of disease, they are being investigated for use in earlier detection via pan-cancer screening tests, for tracking tumor evolution to inform therapy selection, and for making treatment decisions during minimal residual disease surveillance. However, a typical blood draw carries ultra-low levels of ctDNA (as low as 1.7 copies of the tumor genome in 15mL of blood for a 1cm lung tumor), and this fundamentally limits the sensitivity and the clinical utility of ctDNA-testing in many settings. To push beyond current ctDNA detection limits, we present a first-in-class liquid biopsy priming agent that is given prior to a blood draw to increase the abundance of ctDNA in circulation. Our priming agent consists of liposomes that transiently block the uptake of cell free DNA (cfDNA) by macrophages in the liver, resulting in increased cfDNA available for diagnostic analysis in blood. Using an in vitro 2D assay, we first identified a DSPE-based liposomal formulation that inhibits the uptake of cfDNA by two independent murine macrophage cell lines. Next, we injected our liposomal agent into healthy or tumor-bearing mice and collected blood for further analysis. We found that in healthy mice priming increases the half-life and the recovery of cfDNA from a blood draw, assayed via qPCR. In tumor-bearing mice, the priming agent increases the recovery of ctDNA by up to 60-fold (P = 0.0103) and improves the sensitivity of a ctDNA diagnostic assay from 0% to 75% at low tumor burden. Importantly, cfDNA levels in mice return to baseline within five hours of agent administration and repeated dosing shows no evidence of toxicity. Our priming strategy should be of interest for precision oncology applications, including early detection, longitudinal monitoring of therapeutic response, and surveillance for minimal residual disease. This tumor-agnostic priming agent should also improve the performance of ctDNA analytical techniques other than mutational profiling and may even increase the recovery of cfDNA from other body fluids beyond plasma. Moreover, we believe that this work sets a precedent for the potential development of priming agents for liquid biopsy at large across other analytes. In summary, here we present a first-in-class liquid biopsy priming agent capable of improving the sensitivity and the robustness of ctDNA testing in tumor-bearing mice by modulating liver cfDNA clearance. We envision that further development of liquid biopsy priming agents will signify a major step forward for the successful deployment of precision oncology tools across all stages of cancer management. Citation Format: Carmen Martin-Alonso, Shervin Tabrizi, Kan Xiong, Ahmet Bekdemir Bekdemir, Sahil Patel, Zhenyi An, Timothy Blewett, Sainetra Sridhar, Douglas Shea, Ava Amini, Jesse D. Kirkpatrick, Jin Bae, Eugenia Roberts, Ruolin Liu, Justin Rhoades, Todd Golub, J. Christopher Love, Viktor A. Adalsteinsson, Sangeeta N. Bhatia. A liposomal priming agent increases the sensitivity of liquid biopsies. [abstract]. In: Proceedings of the AACR Special Conference: Precision Prevention, Early Detection, and Interception of Cancer; 2022 Nov 17-19; Austin, TX. Philadelphia (PA): AACR; Can Prev Res 2023;16(1 Suppl): Abstract nr PR007.

Enhanced Efficacy of Radiopharmaceuticals When Using Technetium-99m-Labeled Liposomal Agents: Synthesis and Pharmacokinetic Properties.

Challenges posed by the retention of radiopharmaceuticals in unintended organs affect the quality of patient procedures when undergoing diagnostics and therapeutics. The aim of this study was to formulate a suitable tracer encapsulated in liposomes using different techniques and compounds to enhance the stability, uptake, clearance, and cytotoxic effect of the radiopharmaceutical. Cationic liposomes were prepared by a thin-film method using dipalmitoyl phosphatidylcholine (DPPC) and cholesterol. Whole-body gamma camera images were acquired of intravenously injected New Zealand rabbits. Additionally, liposomes were assessed using stability, toxicity, zeta potential, and particle size tests. In the control cases, Technetium-99m (99mTc)-sestamibi exhibited the lowest heart uptake the blood pool and delayed images compared to both 99mTc-liposomal agents. Liver and spleen uptake in the control samples with 99mTc-sestamibi increased in 1-h-delayed images, unlike with 99mTc-liposomal agents, which were decreased in delayed images. The mean maximum count in the bladder for 99mTc-sestamibi loaded liposomes 1 h post-injection was 2354.6 (±2.6%) compared to 178.4 (±0.54%) for 99mTc-sestamibi without liposomes. Liposomal encapsulation reduced the cytotoxic effect of the sestamibi. 99mTc-MIBI-cationic liposomes exhibited excellent early uptake and clearance compared to 99mTc-MIBI without liposomes. Adding cholesterol during liposome formation enhanced the stability and specificity of the targeted organs.

Redox-Responsive MRI Probes Based on First-Row Transition-Metal Complexes.

The presence of multiple oxidation and spin states of first-row transition-metal complexes facilitates the development of switchable MRI probes. Redox-responsive probes capitalize on a change in the magnetic properties of the different oxidation states of the paramagnetic metal ion center upon exposure to biological oxidants and reductants. Transition-metal complexes that are useful for MRI can be categorized according to whether they accelerate water proton relaxation (T1 or T2 agents), induce paramagnetic shifts of 1H or 19F resonances (paraSHIFT agents), or are chemical exchange saturation transfer (CEST) agents. The various oxidation state couples and their properties as MRI probes are summarized with a focus on Co(II)/Co(III) or Fe(II)/Fe(III) complexes as small molecules or as liposomal agents. Solution studies of these MRI probes are reviewed with an emphasis on redox changes upon treatment with oxidants or with enzymes that are physiologically important in inflammation and disease. Finally, we outline the challenges of developing these probes further for in vivo MRI applications.

Дослідження токсикологічних властивостей нового ліпосомального засобу «Ліпохром»

A preclinical study of a new pharmacological agent Lipochrome (a lyophilized powder for injection, produced by Nano Technologies in Medicine, Ukraine) specific activity showed that its administration in an isovolemic volume of saline 30 min after acute massive blood loss caused a moderate hemodynamic effect and prevented the development of decompensated metabolic acidosis and disseminated intravascular blood coagulation.An important stage in new potential drugs compex preclinical studies is their toxicological parameters investigation. The aim of the study was an experimental investigation of new pharmacological agent Lipochrome toxicological properties to substantiate its safety for clinical use.Lipochrome test sample acute toxicity was studied in two laboratory animals species: female white mice (18–20 g body weight) and female rats (200–210 g body weight). Intravenous injection of the pharmacological agent Lipochrome in a single dose 50 mg/kg body weight (mice) and 20 mg/kg (rats) (maximum allowable volumes for these species with intravenous injection) did not cause experimental animals death, as soon as the development of any clinical signs of toxic effects, changes in the dynamics of body weight or absolute and relative masses of internal organs. External examination, autopsy and macroscopic examination of female white mice and rats 14 days after Lipochrome test sample single injection did not detect alternate changes, signs of impaired blood circulation, inflammation or dysregenerative changes. No signs of pathological changes in the surrounding tissues were found at the Lipochrame injection site. Repeated intraperitoneal injections of the Lipochrome test sample to rats of both sexes for 14 days at a dose 10 mg/kg (supposed therapeutic) and 50 mg/kg (five times higher than supposed therapeutic) caused no death of animals, no clinical signs of toxic action, had no influence on the body weight dynamicsand general behavioral reactions. There were no negative effects on the peripheral blood hematological parameters, general metabolism violations as well as no changes in the liver and kidneys structural and functional properties in comparison with physiological norm. Under these conditions, there were no macro- and microscopic changes in the main integral indices, no hemodynamic disorders, no inflammatory reactions and degenerative lesions of internal organs and tissues.Thus, the results obtained confirm the advisability and prospectiveness of further investigation new liposomal agent Lipochrome pharmacological properties and therapeutic efficacy.

Efficacy of liposomal bupivacaine vs. traditional anaesthetic infiltration for pain management in total hip arthroplasty: a systematic review and meta-analysis.

In this review, we assess the effectiveness of liposomal bupivacaine against the traditional bupivacaine infiltration in the postoperative management of total hip arthroplasty (THA). Various databases including PubMed Central, Medline, Scopus, Embase, Google Scholar, Cochrane library and ScienceDirect (inception date till August 2020) were searched. The quality of published trials was assessed using Cochrane risk of bias tool, and a random-effects model was used for meta-analysis. We report pooled Risk ratios (RR) or pooled Standardized Mean difference (SMD) with 95% confidence intervals (CIs). We analyzed a total of 13 studies with 62,582 participants. The majority of the studies were retrospective with lower bias risks. Liposomal bupivacaine was significantly associated with the reduction in opioid requirement at 48 hours (SMD = -0.25; 95% CI: -0.40 to -0.09; p=0.002) and length of hospital stay (SMD = -0.25; 95% CI: -0.43 to -0.07, p=0.006) following THA compared with the control group. However, there was no statistically significant difference between the effect of liposomal bupivacaine and other agents for pain score (24 and 48 hours), opioid requirement at 24 hours and incidence of nausea. Liposomal bupivacaine has selective benefits in terms of opioid consumption and length of hospital stay against the traditional bupivacaine among the patients undergoing THA.

Fast processes of nanoparticle blood clearance: Comprehensive study

Blood circulation is the key parameter that determines the in vivo efficiency of nanoagents. Despite clinical success of the stealth liposomal agents with their inert and shielded surfaces, a great number of non-stealth nanomaterials is being developed due to their potential of enhanced functionality. By harnessing surface phenomena, such agents can offer advanced control over drug release through intricately designed nanopores, catalysis-propelled motion, computer-like analysis of several disease markers for precise target identification, etc. However, investigation of pharmacokinetic behavior of these agents becomes a great challenge due to ultra-short circulation (usually around several minutes) and impossibility to use the invasive blood-sampling techniques. Accordingly, the data on circulation of such agents has been scarce and irregular. Here, we demonstrate high-throughput capabilities of the developed magnetic particle quantification technique for nanoparticle circulation measurements and present a comprehensive investigation of factors that affect blood circulation of the non-stealth nanoparticles. Namely, we studied the following 9 factors: particle size, zeta-potential, coating, injection dose, repetitive administration, induction of anesthesia, mice strain, absence/presence of tumors, tumor size. Our fundamental findings demonstrate potential ways to extend the half-life of the agents in blood thereby giving them a better chance of achieving their goal in the organism. The study will be valuable for design of the next generation nanomaterials with advanced biomedical functionality.

Immunological and Toxicological Considerations for the Design of Liposomes

Liposomes hold great potential as gene and drug delivery vehicles due to their biocompatibility and modular properties, coupled with the major advantage of attenuating the risk of systemic toxicity from the encapsulated therapeutic agent. Decades of research have been dedicated to studying and optimizing liposomal formulations for a variety of medical applications, ranging from cancer therapeutics to analgesics. Some effort has also been made to elucidate the toxicities and immune responses that these drug formulations may elicit. Notably, intravenously injected liposomes can interact with plasma proteins, leading to opsonization, thereby altering the healthy cells they come into contact with during circulation and removal. Additionally, due to the pharmacokinetics of liposomes in circulation, drugs can end up sequestered in organs of the mononuclear phagocyte system, affecting liver and spleen function. Importantly, liposomal agents can also stimulate or suppress the immune system depending on their physiochemical properties, such as size, lipid composition, pegylation, and surface charge. Despite the surge in the clinical use of liposomal agents since 1995, there are still several drawbacks that limit their range of applications. This review presents a focused analysis of these limitations, with an emphasis on toxicity to healthy tissues and unfavorable immune responses, to shed light on key considerations that should be factored into the design and clinical use of liposomal formulations.

Synthesis of Cationic and Ionizable Amphiphiles Based on Heminal Aminodiol as Potential siRNA Delivery Systems

Research methods based on the use of RNA interference mechanisms are now included among the basic methods of molecular biology. Drugs based on siRNA are being developed for the treatment of cancer, infectious diseases and other pathologies that are associated with impaired Junctions of specific genes. One of the main problems of siRNA-based drug development is their efficient and safe delivery to target cells. Modern delivery strategies are based on the use of chemical compounds or biological carriers, such as viruses. Lipid nanoparticles (liposomal agents) are the most advanced platform among non-viral vectors for the delivery of gene materials into cells. In this paper, a scheme has been proposed and the synthesis of new cationic amphiphiles has been carried out as the basis for the means of delivering siRNA to target cells. Synthesized amphiphiles belong to two types of cationic lipids: with a permanently charged amino group in the form of a salt and with an ionizable polar block. Ionized amphiphiles are a new generation of cationic lipids that exhibit reduced toxicity and immunogenicity. They undergo ionization only in the acidic environment of endosomes during intracellular transport in the cytosol, which leads to the release of the encapsulated cargo. The structure of the target compounds is based on diethanolamine derivatives, which is a spacer between the hydrophobic block and the polar fragment. The hydrophobic block contains palmitoyl residues, and the polar one - ethylenediamine and 3-diethylaminopropylamine derivatives. The developed synthetic scheme is distinguished by the simplicity and versatility of the proposed approach, which allows it to be used in the preparation of a series of samples in preparative quantities necessary for the subsequent physicochemical and biochemical studies.

Efficacy of liposomal dosage forms and hyperosmolar salines in experimental pharmacotherapy of acute lung injury

Introduction: Hypertonic sodium chloride solutions and liposomal drugs with pulmotropic effect are of great interest for the treatment of acute lung injury (ALI). The results of the studies on the efficacy of hypertonic solutions and lipo[1]somes in ALI treatment are currently controversial.
 Materials and methods: For the experiment, liposomes with dexamethasone, N-acetylcysteine (NAC), aprotinin and dye Cyanine-7 (Cy-7) were obtained. A liposome analysis was performed by means of spectrophotometry. ALI was modeled in rats by the administration of the damaging agents into the trachea. The experimental agents were injected once intravenously after the modeling of ALI. For experimental therapy used liposomal agents, 7.5% hypertonic saline (HS) and HyperHAES solutions in the respective groups. The efficacy of the therapy was assessed by the survival of animals, functional indicators of the cardiovascular and respiratory systems, and by the lung-body ratio. The biodistri[1]bution of liposomes after intravenous administration was investigated in mice through using a fluorescent dye Cy-7. The biodistribution of liposomes with Cy-7 was assessed using bioimaging according to the fluorescence intensity of internal organs (lungs, liver, and kidneys) and blood, expressed as dye concentration according to the calibration de[1]pendence of dye concentrarion on fluorescence intensity.
 Results and discussion: All the studied liposomal drugs were effective for the pharmacological correction of ALI. Hypertonic solutions, unlike liposomal drugs, were less likely to prevent the development of pulmonary edema. All the studied therapeutic agents increased the survival rate of the laboratory animals with ALI. The most effective experimen[1]tal agent was liposomal dexamethasone. The use of drugs in form of simple liposomes with average diameter of 350 nm provided for a higher concentration of the drug in the lungs within the first 40 minutes after intravenous administration.
 Conclusion: Intravenous administration of liposomal forms is promising for the pharmacotherapy of acute lung injury.

Liposomes Co- encapsulating Anticancer Drugs in Synergistic Ratios as an Approach to Promote Increased Efficacy and Greater Safety.

The era of chemotherapy began in the 1940s, but it was in the 1960s that it was seen as really promising when the first patients with childhood acute lymphoblastic leukemia were cured with combination chemotherapy. Today, it is known that due to resistance to single agents, combination therapy is essential for tumor eradication and cure. In the last decade, studies have shown that anticancer drug combinations can act synergistically or antagonistically against tumor cells in vitro, depending on the ratios of the individual drugs forming the combination. From this observation and facing the possibility of maintaining the in vivo synergistic ratio of combinations came the idea of co-encapsulating anticancer agents in nanosystems. In vivo studies validated this idea by showing that the co-encapsulation of anticancer agents in liposomes allows the maintenance of drug ratios in the plasma and the delivery of fixed drug ratios directly to tumor tissue, leading to a better efficacy compared to the administration of the free drugs combination. Liposomes co-encapsulating irinotecan/floxuridine are now in Phase II trial, and liposomes co-encapsulating cytarabine/daunorubicin were recently approved by the FDA for treatment of patients with acute myeloid leukemia.

CAL02, a novel antitoxin liposomal agent, in severe pneumococcal pneumonia: a first-in-human, double-blind, placebo-controlled, randomised trial

Severe community-acquired pneumonia caused by Streptococcus pneumoniae is associated with high morbidity and mortality rates. CAL02, a novel antitoxin agent with an unprecedented mode of action, consists of liposomes that capture bacterial toxins known to dysregulate inflammation, cause organ damage, and impede immune defence. We aimed to assess the safety of CAL02 as an add-on therapy to antibiotics. This randomised, double-blind, multicentre, placebo-controlled trial was done in ten intensive care units (ICUs) in France and Belgium (but only six units enrolled patients), in patients with severe community-acquired pneumococcal pneumonia who required ICU admission and had been identified as being infected with S pneumoniae. We randomly assigned participants in two stages-the first stage randomly assigned six patients (1:1) to either low-dose CAL02 or placebo, and the second stage randomly assigned 18 patients (14:4) to either high-dose CAL02 or placebo, and stratified in four blocks (4:1, 4:1, 3:1, and 3:1), in addition to standard of care. Block randomisation was done with a computer-generated random number list. Participants, investigators, other site study personnel, the sponsor, and the sponsor's designees involved in study management and monitoring were masked to the randomisation list and treatment assignment. Patients were treated with low-dose (4 mg/kg) or high-dose (16 mg/kg) CAL02 or placebo (saline), in addition to standard antibiotic therapy. Two intravenous doses of study treatment were infused, with a 24 h interval, at a concentration of 10 mg/mL, stepwise, over a maximum of 2 h on days 1 and 2. The primary objective of the study was to assess the safety and tolerability of low-dose and high-dose CAL02 in patients with severe community-acquired pneumonia treated with standard antibiotic therapy, and the primary analysis was done on the safety population (all patients who received at least one dose of the study treatment). Efficacy was a secondary outcome. This trial is registered with ClinicalTrials.gov, number NCT02583373. Between March 21, 2016, and Jan 13, 2018, we screened 280 patients with community-acquired pneumonia. 19 patients were enrolled and randomly assigned, resulting in 13 patients in the CAL02 groups (three assigned to low-dose CAL02 and ten assigned to high-dose CAL02) and six in the placebo group. One patient randomly assigned to placebo was allocated to the wrong treatment group and received high-dose CAL02 instead of placebo. Thus, 14 patients received CAL02 (three received low-dose CAL02 and 11 received high-dose CAL02) and five patients received placebo, constituting the safety population. At baseline, the mean APACHE II score for the total study population was 21·5 (SD 4·9; 95% CI 19·3-23·7) and 11 (58%) of 19 patients had septic shock. Adverse events occurred in 12 (86%) of 14 patients in the CAL02 treatment groups combined and all five (100%) patients in the placebo group. Serious adverse events occurred in four (29%) of 14 patients in the CAL02 treatment groups combined and two (40%) of five patients in the placebo group. One non-serious adverse event (mild increase in triglycerides) in a patient in the high-dose CAL02 group was reported as related to study drug. However, analysis of the changes in triglyceride levels in the CAL02 groups compared with the placebo group revealed no correlation with administration of CAL02. No adverse events were linked to local tolerability events. All patients, apart from one who died in the low CAL02 group (death not related to the study drug) achieved clinical cure at the test of cure visit between days 15 and 22. The sequential organ failure assessment score decreased by mean 65·0% (95% CI 50·7-79·4) in the combined CAL02 groups compared with 29·2% (12·8-45·5) in the placebo group between baseline and day 8. The nature of adverse events was consistent with the profile of the study population and CAL02 showed a promising safety profile and tolerability. However, the difference between high-dose and low-dose CAL02 could not be assessed in this study. Efficacy was in line with the expected benefits of neutralising toxins. The results of this study support further clinical development of CAL02 and provide a solid basis for a larger clinical study. Combioxin.

A light-responsive liposomal agent for MRI contrast enhancement and monitoring of cargo delivery.

Medical magnetic resonance imaging (MRI) produces high-resolution anatomical images of the human body, but has limited capacity to provide useful molecular information. The light-responsive, liposomal MRI contrast agent described herein could be used to provide an intrinsic theranostic aspect to MRI and enable tracking the distribution and cargo release of drug delivery systems upon light-triggered activation.

Curcumin inhibits autocrine growth hormone-mediated invasion and metastasis by targeting NF-κB signaling and polyamine metabolism in breast cancer cells.

Curcumin is assumed to be a plant-derived therapeutic drug that triggers apoptotic cell death in vitro and in vivo by affecting different molecular targets such as NF-κB. Phase I/II trial of curcumin alone or with chemotherapeutic drugs has been accomplished in pancreatic, colon, prostate and breast cancer cases. Recently, autocrine growth hormone (GH) signaling-induced cell growth, metastasis and drug resistance have been demonstrated in breast cancer. In this study, our aim was to investigate the potential therapeutic effect of curcumin by evaluating the molecular machinery of curcumin-triggered apoptotic cell death via focusing on NF-κB signaling and polyamine (PA) metabolism in autocrine GH-expressing MCF-7, MDA-MB-453 and MDA-MB-231 breast cancer cells. For this purpose, a pcDNA3.1 (+) vector with a GH gene insert was transfected by a liposomal agent in all breast cancer cells and then selection was conducted in neomycin (G418) included media. Autocrine GH-induced curcumin resistance was overcome in a dose-dependent manner and curcumin inhibited cell proliferation, invasion-metastasis and phosphorylation of p65 (Ser536), and thereby partly prevented its DNA binding activity in breast cancer cells. Moreover, curcumin induced caspase-mediated apoptotic cell death by activating the PA catabolic enzyme expressions, which led to generation of toxic by-products such as H2O2 in MCF-7, MDA-MB-453 and MDA-MB-231 GH+ breast cancer cells. In addition, transient silencing of SSAT prevented curcumin-induced cell viability loss and apoptotic cell death in each breast cancer cells. In conclusion, curcumin could overcome the GH-mediated resistant phenotype via modulating cell survival, death-related signaling routes and activating PA catabolic pathway.

Abstract 5927: Eribulin modulates tumor microenvironment through vascular remodeling for antitumor effect in multiple mouse xenograft models

Abstract Eribulin mesilate (Eribulin), a first-in-class halichondrin B-based microtubule dynamics inhibitor, has been reported to remodel tumor vasculature (i.e., improvement of tumor vessels and perfusion) in human breast cancer xenograft models; however, the role of this vascular remodeling in anti-tumor effects is not fully understood. Here, we investigated the effects of eribulin-induced vascular remodeling on anti-tumor activities in multiple human cancer xenograft models. Microvessel densities (MVDs) were evaluated by immunohistochemistry (CD31 staining), and anti-tumor effects were examined in 10 human cancer xenograft models treated with eribulin. Eribulin treatment significantly increased MVDs compared to the corresponding control groups in 7 out of 10 models with a correlation between enhanced MVD levels and anti-tumor effects (R2=0.55). Because of observed increases in MVDs, we next utilized 111In-labeled PEGylated liposomes to examine if eribulin treatment would result in increased tumoral accumulation levels of such liposomes, and indeed, we found that eribulin, unlike vinorelbine another tubulin inhibitor, enhanced them. Since eribulin treatment increased accumulation of 111In-labeled PEGylated liposomes, we postulated that this treatment might enhance anti-tumor effect of Doxil (a liposomal anti-cancer agent). As expected, eribulin enhanced anti-tumor activity of Doxil in the PE-H1650 xenograft model. Then, we evaluated whether eribulin facilitated the recruitment of immune cells into the tumor. Infiltrating CD11b-positive immune cells were significantly increased in the multiple eribulin-treated xenografted tumors, and natural killer (NK) cell depletion reduced anti-tumor effects of eribulin. Collectively, these findings suggested a contribution of the immune cells for anti-tumor activities of eribulin. Taken together, our results obtained from multiple human cancer xenograft models suggested that vascular remodeling induced by eribulin therapy would act as a microenvironment modulator, and contributed to show anti-tumor effects of eribulin. Citation Format: Ken Ito, Shusei Hamamichi, Takanori Abe, Tsuyoshi Akagi, Hiroshi Shirota, Satoshi Kawano, Makoto Asano, Osamu Asano, Akira Yokoi, Junji Matsui, Izumi O. Umeda, Hirofumi Fujii. Eribulin modulates tumor microenvironment through vascular remodeling for antitumor effect in multiple mouse xenograft models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5927. doi:10.1158/1538-7445.AM2017-5927

Pre-clinical evaluation of a nanoparticle-based blood-pool contrast agent for MR imaging of the placenta

IntroductionNon-invasive 3D imaging that enables clear visualization of placental margins is of interest in the accurate diagnosis of placental pathologies. This study investigated if contrast-enhanced MRI performed using a liposomal gadolinium blood-pool contrast agent (liposomal-Gd) enables clear visualization of the placental margins and the placental-myometrial interface (retroplacental space). Non-contrast MRI and contrast-enhanced MRI using a clinically approved conventional contrast agent were used as comparators. Materials and methodsStudies were performed in pregnant rats under an approved protocol. MRI was performed at 1T using a permanent magnet small animal scanner. Pre-contrast and post-liposomal-Gd contrast images were acquired using T1-weighted and T2-weighted sequences. Dynamic Contrast enhanced MRI (DCE-MRI) was performed using gadoterate meglumine (Gd-DOTA, Dotarem®). Visualization of the retroplacental clear space, a marker of normal placentation, was judged by a trained radiologist. Signal-to-noise (SNR) and contrast-to-noise (CNR) ratios were calculated for both single and averaged acquisitions. Images were reviewed by a radiologist and scored for the visualization of placental features. Contrast-enhanced CT (CE-CT) imaging using a liposomal CT agent was performed for confirmation of the MR findings. Transplacental transport of liposomal-Gd was evaluated by post-mortem elemental analysis of tissues. Ex-vivo studies in perfused human placentae from normal, GDM, and IUGR pregnancies evaluated the transport of liposomal agent across the human placental barrier. ResultsPost-contrast T1w images acquired with liposomal-Gd demonstrated significantly higher SNR (p = 0.0002) in the placenta compared to pre-contrast images (28.0 ± 4.7 vs. 6.9 ± 1.8). No significant differences (p = 0.39) were noted between SNR in pre-contrast and post-contrast liposomal-Gd images of the amniotic fluid, indicating absence of transplacental passage of the agent. The placental margins were significantly (p < 0.001) better visualized on post-contrast liposomal-Gd images. DCE-MRI with the conventional Gd agent demonstrated retrograde opacification of the placenta from fetal edge to the myometrium, consistent with the anatomy of the rat placenta. However, no consistent and reproducible visualization of the retroplacental space was demonstrated on the conventional Gd-enhanced images. The retroplacental space was only visualized on post-contrast T1w images acquired using the liposomal agent (SNR = 15.5 ± 3.4) as a sharply defined, hypo-enhanced interface. The retroplacental space was also visible as a similar hypo-enhancing interface on CE-CT images acquired using a liposomal CT contrast agent. Tissue analysis demonstrated undetectably low transplacental permeation of liposomal-Gd, and was confirmed by lack of permeation through a perfused human placental model. ConclusionsContrast-enhanced T1w-MRI performed using liposomal-Gd enabled clear visualization of placental margins and delineation of the retroplacental space from the rest of the placenta; the space is undetectable on non-contrast imaging and on post-contrast T1w images acquired using a conventional, clinically approved Gd chelate contrast agent.

Profiling the relationship between tumor-associated macrophages and pharmacokinetics of liposomal agents in preclinical murine models

The mononuclear phagocyte system (MPS) has previously been shown to significantly affect the clearance, tumor delivery, and efficacy of nanoparticles (NPs). This study profiled MPS cell infiltration in murine preclinical tumor models and evaluated how these differences may affect tumor disposition of PEGylated liposomal doxorubicin (PLD) in models sensitive and resistant to PLD. Significant differences in MPS presence existed between tumor types (e.g. ovarian versus endometrial), cell lines within the same tumor type, and location of tumor implantation (i.e. flank versus orthotopic xenografts). Further, the differences in MPS presence of SKOV-3 ovarian and HEC1A endometrial orthotopic cancer models may account for the 2.6-fold greater PLD tumor exposure in SKOV-3, despite similar plasma, liver and spleen exposures. These findings suggest that profiling the presence of MPS cells within and between tumor types is important in tumor model selection and in tumor types and patients likely to respond to NP treatment.

Radiolabeled liposome imaging determines an indication for liposomal anticancer agent in ovarian cancer mouse xenograft models.

Liposomal anticancer agents can effectively deliver drugs to tumor lesions, but their therapeutic effects are enhanced in only limited number of patients. Appropriate biomarkers to identify responder patients to these liposomal agents will improve their treatment efficacies. We carried out pharmacological and histopathological analyses of mouse xenograft models bearing human ovarian cancers (Caov-3, SK-OV-3, KURAMOCHI, and TOV-112D) to correlate the therapeutic effects of doxorubicin-encapsulated liposome (Doxil(®) ) and histological characteristics linked to the enhanced permeability and retention effect. We next generated (111) In-encapsulated liposomes to examine their capacities to determine indications for Doxil(®) treatment by single-photon emission computed tomography (SPECT)/CT imaging. Antitumor activities of Doxil(®) were drastically enhanced in Caov-3, moderately in SK-OV-3, and minimally in KURAMOCHI and TOV-112D when compared to doxorubicin. Microvessel density and vascular perfusion were high in Caov-3 and SK-OV-3, indicating a close relation with the enhanced antitumor effects. Next, (111) In-encapsulated liposomes were given i.v. to the animals. Their tumor accumulation and area under the curve values over 72 h were high in Caov-3, relatively high in SK-OV-3, and low in two other tumors. Importantly, as both Doxil(®) effects and liposomal accumulation varied in the SK-OV-3 group, we individually obtained SPECT/CT images of SK-OV-3-bearing mouse (n = 11) before Doxil(®) treatment. Clear correlation between liposomal tumor accumulation and effects of Doxil(®) was confirmed (R(2) = 0.73). Taken together, our experiments definitely verified that enhanced therapeutic effects through liposomal formulations of anticancer agents depend on tumor accumulation of liposomes. Tumor accumulation of the radiolabeled liposomes evaluated by SPECT/CT imaging is applicable to appropriately determine indications for liposomal antitumor agents.

A multimodal nano agent for image-guided cancer surgery

Intraoperative imaging technologies including computed tomography and fluorescence optical imaging are becoming routine tools in the cancer surgery operating room. They constitute an enabling platform for high performance surgical resections that assure local control while minimizing morbidity. New contrast agents that can increase the sensitivity and visualization power of existing intraoperative imaging techniques will further enhance their clinical benefit. We report here the development, detection and visualization of a dual-modality computed tomography and near-infrared fluorescence nano liposomal agent (CF800) in multiple preclinical animal models of cancer. We describe the successful application of this agent for combined preoperative computed tomography based three-dimensional surgical planning and intraoperative target mapping (>200 Hounsfield Units enhancement), as well as near-infrared fluorescence guided resection (>5-fold tumor-to-background ratio). These results strongly support the clinical advancement of this agent for image-guided surgery with potential to improve lesion localization, margin delineation and metastatic lymph node detection.

Improved oral bioavailability of capsaicin via liposomal nanoformulation: preparation, in vitro drug release and pharmacokinetics in rats.

This study innovatively prepared an effective capsaicin-loaded liposome, a nanoformulation with fewer irritants, for oral administration. The in vitro and in vivo properties of the liposomal encapsulation were investigated and the potential possibility of oral administration evaluated. The liposomal agent composed of phospholipid, cholesterol, sodium cholate and isopropyl myristate was prepared using film-dispersion method. A level A in vitro-in vivo correlation (IVIVC) was established for the first time, which demonstrated an excellent IVIVC of both formulated and free capsaicin in oral administration. Physicochemical characterizations including mean particle size, zeta (ζ) potential and average encapsulation efficiency of capsaicin-loaded liposome were found to be 52.2±1.3nm, -41.5±2.71mv and 81.9±2.43%, respectively. In vivo, liposomal encapsulation allowed a 3.34-fold increase in relative bioavailability compared to free capsaicin. The gastric mucosa irritation studies indicated that the liposomal system was a safe carrier for oral administration. These results support the fact that capsaicin, an effective drug for the treatment of neuropathic pain, could be encapsulated in liposome for improved oral bioavailability. The excellent IVIVC of capsaicin-loaded liposome could also be a promising tool in liposomal formulation development with an added advantage of reduced animal testing.