A novel pH-sensitive liposome formulation containing oleyl alcohol

AbstractpH-sensitive liposomes are designed to undergo acid-triggered destabilization. First generation pH-sensitive liposomes, based on the cone-shaped lipid dioleoylphosphatidylethanolamine (DOPE), have been shown to lose fusogenicity in the presence of serum. Here, we report the design and evaluation of novel serum-resistant pH-sensitive liposome formulations that are based on the composition of egg phosphatidylcholine (PC), cholesteryl hemisuccinate (CHEMS), oleyl alcohol (OAlc), and Tween-80 (T-80). When loaded with the fluorescent probe calcein, these liposomes exhibited excellent stability at pH 7.4 and underwent rapid destabilization upon acidification as shown by calcein dequenching and particle size increase. Adjusting the mole percentages of T-80 and OAlc in the formulation could regulate the stability and pH-sensitive properties of these liposomes. Liposomes with a higher T-80 content exhibited greater stability but were less sensitive to acid-induced destabilization. Meanwhile, formulations with a higher OAlc content exhibited greater content release in response to low pH. The pH-triggered liposomal destabilization did not produce membrane fusion according to an octadecylrhodamine B chloride (R18) lipid-mixing assay. Compared to DOPE-based pH-sensitive liposomes, the above formulations showed much better retention of their pH-sensitive properties in the presence of 10% serum. These liposomes were then evaluated for intracellular delivery of entrapped cytosine-β-d-arabinofuranoside (araC) in KB human oral cancer cells, which have elevated folate receptor (FR) expression. The FR, which is amplified in many types of human tumors, has been shown to mediate the internalization of folate-derivatized liposomes into an acidic intracellular compartment. FR-targeted OAlc-based pH-sensitive liposomes, entrapping 200 mM araC, showed ∼17-times greater FR-dependent cytotoxicity in KB cells compared to araC delivered via FR-targeted non-pH-sensitive liposomes. These data indicated that pH-sensitive liposomes based on OAlc, combined with FR-mediated targeting, are promising delivery vehicles for membrane impermeable therapeutic agents

Changes in practice patterns in Japan from before to after JSDT 2013 guidelines on hemodialysis prescriptions: results from the JDOPPS

Abstract Background The Japanese Society for Dialysis Therapy (JSDT) published in 2013 inaugural hemodialysis (HD) guidelines. Specific targets include 1.4 for single-pool Kt/V (spKt/V) with a minimum dose of 1.2, minimum dialysis session length of 4 hours, minimum blood flow rate (BFR) of 200 mL/min, fluid removal rate no more than 15 mL/kg/hr, and hemodiafiltration (HDF) therapy for certain identified symptoms. We evaluated the effect of these guidelines on actual practice in the years spanning 2005 – 2018. Methods Analyses were carried out to describe trends in the above HD prescription practices from December 2005 to April 2013 (before guideline publication) to August 2018 based on prevalent patient cross-sections from approximately 60 randomly selected HD facilities participating in the Japan Dialysis Outcomes and Practice Patterns Study. Results From April 2006 to August 2017 continual rises occurred in mean spKt/V (from 1.35 to 1.49), and percent of patients having spKt/V>1.2 (71% to 85%). Mean BFR increased with time from 198.3 mL/min (April 2006) to 218.4 mL/min (August 2017) , along with percent of patients with BFR >200 ml/min (65% to 85%). HDF use increased slightly from 6% (April 2006 and August 2009) to 8% by April 2013, but increased greatly thereafter to 23% by August 2017. In contrast, mean HD treatment time showed little change from 2006-2017, whereas mean UFR declined from 11.3 in 2006 to 8.4 mL/Kg/hour in 2017. Conclusions From 2006 – 2018 Japanese HD patients experienced marked improvement in reaching the spKt/V target specified by the 2013 JSDT guidelines. This may have been due to moderate increase in mean BFR even though mean HD session length did not change much. In addition, HDF use increased dramatically in this time period. Other HD delivery changes during this time, such as increased use of super high flux dialyzers, also merit study. While we cannot definitively conclude a causal relationship between the publication of the guidelines and the subsequent practice changes in Japan, those changes moved practice closer to the recommendations of the guidelines.http://deepblue.lib.umich.edu/bitstream/2027.42/173577/1/12882_2021_Article_2543.pd

Half-life extension via ABD-fusion leads to higher tumor uptake of an affibody-drug conjugate compared to PAS- and XTENylation.

A critical parameter during the development of protein therapeutics is to endow them with suitable pharmacokinetic and pharmacodynamic properties. Small protein drugs are quickly eliminated by kidney filtration, and in vivo half-life extension is therefore often desired. Here, different half-life extension technologies were studied where PAS polypeptides (PAS300, PAS600), XTEN polypeptides (XTEN288, XTEN576), and an albumin binding domain (ABD) were compared for half-life extension of an anti -human epidermal growth factor receptor 2 (HER2) affibody-drug conjugate. The results showed that extension with the PAS or XTEN polypeptides or the addition of the ABD lowered the affinity for HER2 to some extent but did not negatively affect the cytotoxic potential. The half-lives in mice ranged from 7.3 h for the construct including PAS300 to 11.6 h for the construct including PAS600. The highest absolute tumor uptake was found for the construct including the ABD, which was 60 to 160% higher than the PASylated or XTENylated constructs, even though it did not have the longest half-life (9.0 h). A comparison of the tumor -to -normal -organ ratios showed the best overall performance of the ABD-fused construct. In conclusion, PASylation, XTENylation, and the addition of an ABD are viable strategies for half-life extension of affibody-drug conjugates, with the best performance observed for the construct including the ABD

Influence of the Position and Composition of Radiometals and Radioiodine Labels on Imaging of Epcam Expression in Prostate Cancer Model Using the DARPin Ec1

Simple Summary Metastasis-targeting therapy might improve outcomes in oligometastatic prostate cancer. Epithelial cell adhesion molecule (EpCAM) is overexpressed in 40-60% of prostate cancer cases and might be used as a target for specific delivery of toxins and drugs. Radionuclide molecular imaging could enable non-invasive detection of EpCAM and stratification of patients for targeted therapy. Designed ankyrin repeat proteins (DARPins) are scaffold proteins, which can be selected for specific binding to different targets. The DARPin Ec1 binds strongly to EpCAM. To determine an optimal design of Ec1-based probes, we labeled Ec1 at two different positions with four different nuclides (Ga-68, In-111, Co-57 and I-125) and investigated the impact on Ec1 biodistribution. We found that the C-terminus is the best position for labeling and that In-111 and I-125 provide the best imaging contrast. This study might be helpful for scientists developing imaging probes based on scaffold proteins. The epithelial cell adhesion molecule (EpCAM) is intensively overexpressed in 40-60% of prostate cancer (PCa) cases and can be used as a target for the delivery of drugs and toxins. The designed ankyrin repeat protein (DARPin) Ec1 has a high affinity to EpCAM (68 pM) and a small size (18 kDa). Radiolabeled Ec1 might be used as a companion diagnostic for the selection of PCa patients for therapy. The study aimed to investigate the influence of radiolabel position (N- or C-terminal) and composition on the targeting and imaging properties of Ec1. Two variants, having an N- or C-terminal cysteine, were produced, site-specifically conjugated to a DOTA chelator and labeled with cobalt-57, gallium-68 or indium-111. Site-specific radioiodination was performed using ((4-hydroxyphenyl)-ethyl)maleimide (HPEM). Biodistribution of eight radiolabeled Ec1-probes was measured in nude mice bearing PCa DU145 xenografts. In all cases, positioning of a label at the C-terminus provided the best tumor-to-organ ratios. The non-residualizing [I-125]I-HPEM label provided the highest tumor-to-muscle and tumor-to-bone ratios and is more suitable for EpCAM imaging in early-stage PCa. Among the radiometals, indium-111 provided the highest tumor-to-blood, tumor-to-lung and tumor-to-liver ratios and could be used at late-stage PCa. In conclusion, label position and composition are important for the DARPin Ec1

Comparative Preclinical Evaluation of Peptide-Based Chelators for the Labeling of DARPin G3 with Tc-99m for Radionuclide Imaging of HER2 Expression in Cancer

Non-invasive radionuclide imaging of human epidermal growth factor receptor type 2 (HER2) expression in breast, gastroesophageal, and ovarian cancers may stratify patients for treatment using HER2-targeted therapeutics. Designed ankyrin repeat proteins (DARPins) are a promising type of targeting probe for radionuclide imaging. In clinical studies, the DARPin [Tc-99m]Tc-(HE)(3)-G3 labeled using a peptide-based chelator His-Glu-His-Glu-His-Glu ((HE)(3)), provided clear imaging of HER2 expressing breast cancer 2-4 h after injection. The goal of this study was to evaluate if the use of cysteine-containing peptide-based chelators Glu-Glu-Glu-Cys (E3C), Gly-Gly-Gly-Cys (G(3)C), and Gly-Gly-Gly-Ser-Cys connected via a (Gly-Gly-Gly-Ser)(3)-linker (designated as G3-(G(3)S)(3)C) would further improve the contrast of imaging using Tc-99m-labeled derivatives of G3. The labeling of the new variants of G3 provided a radiochemical yield of over 95%. Labeled G3 variants bound specifically to human HER2-expressing cancer cell lines with affinities in the range of 1.9-5 nM. Biodistribution of [Tc-99m]Tc-G3-G(3)C, [Tc-99m]Tc-G3-(G(3)S)(3)C, and [Tc-99m]Tc-G3-E3C in mice was compared with the biodistribution of [Tc-99m]Tc-(HE)(3)-G3. It was found that the novel variants provide specific accumulation in HER2-expressing human xenografts and enable discrimination between tumors with high and low HER2 expression. However, [Tc-99m]Tc-(HE)(3)-G3 provided better contrast between tumors and the most frequent metastatic sites of HER2-expressing cancers and is therefore more suitable for clinical applications

International variation in dialysis discontinuation in patients with advanced kidney disease

BACKGROUND: Decisions about dialysis for advanced kidney disease are often strongly shaped by sociocultural and system-level factors rather than the priorities and values of individual patients. We examined international variation in the uptake of conservative approaches to the care of patients with advanced kidney disease, in particular discontinuation of dialysis.METHODS: We employed an observational cohort study design using data collected from patients maintained on long-term hemodialysis between 1996 and 2015 in facilities across 12 developed countries participating in the Dialysis Outcomes and Practice Patterns Study (DOPPS). The main outcome was discontinuation of dialysis therapy. We analyzed the association between several patient characteristics and time to dialysis discontinuation by country and phase of study entry.RESULTS: A total of 259 343 DOPPS patients contributed data to the study, of whom 48 519 (18.7%) died during the study period. Of the decedents, 5808 (12.0%) discontinued dialysis before death. Rates of discontinuation were higher within the first few months after initiation of dialysis, among older adults, among those with a greater number of comorbidities and among those living in an institution. After adjustment for age, sex, dialysis duration, diabetes and dialysis era, rates of discontinuation were highest in Canada, the United States and Australia/New Zealand (33.8, 31.4 and 21.5 per 1000/yr, respectively) and lowest in Japan and Italy (< 0.1 per 1000/yr). Crude discontinuation rates were highest in dialysis facilities that were more likely to offer comprehensive conservative renal care to older adults.INTERPRETATION: We found persistent international variation in average rates of dialysis discontinuation not explained by differences in patient case-mix. These differences may reflect physician-, facility- and society-level differences in clinical practice. There may be opportunities for international cross-collaboration to improve support for patients with end-stage renal disease who prefer a more conservative approach

The development of a highly specific radiochemical compound based on labeled 99mtc recombinant molecules for targeted imaging of cells with the overexpression of Her-2 / neu

Currently, there is a urgent need to search for new diagnostic methods that allow us to reveal malignant tumors with the overexpression of Her-2/neu with high accuracy. In recent years radioisotope methods have been actively developing to identify specific tumor targets, with antibodies being the “targeting” module.The purpose of the study. Creation of a chemically stable radiochemical compound for the imaging of cells with the overexpression of Her-2/neu.Materials and methods. The study was conducted using two human adenocarcinoma cell lines with expression (BT-474) and without expression (MCF-7) Her-2/neu. The specificity of the binding of the test complex with Her-2/neu receptor was determined by direct radiometric and planar scintigraphy. To evaluate the differences in quantitative characteristics between the groups a non-parametric Mann – Whitney test was used.Results. The yield of the labeled complex was more than 91% and the radiochemical frequency was more than 94%. When performing a visual scintigraphic evaluation, a much higher accumulation rate of the studied radiopharmaceutical preparation (RFP) was observed in the culture of cells with overexpression of the surface Her-2/neu receptor. Direct radiometric results also demonstrated a higher accumulation of RFPs in the human BT-474 mammary adenocarcinoma cell line with Her-2/neu overexpression in comparison with the control group.Conclusion. Preclinical studies demonstrated high stability of the test compound, as well as its accumulation in the group of cells with Her-2/neu overexpressio