We investigated the effects of chronic mineralocorticoid receptor blockade with eplerenone on the development and progression of hypertension and end organ damage in Dahl salt-sensitive rats. Eplerenone significantly attenuated the progressive rise in systolic blood pressure (SBP) (204 ± 3 vs. 179±3 mmHg, p < 0.05), reduced proteinuria (605.5 ± 29.6 vs. 479.7 ± 26.1 mg/24h, p < 0.05), improved injury scores of glomeruli, tubules, renal interstitium, and vasculature in Dahl salt-sensitive rats fed a high-salt diet. These results demonstrate that mineralocorticoid receptor antagonism provides target organ protection and attenuates the development of elevated blood pressure (BP) in a model of salt-sensitive hypertension.
mineralocorticoid receptor antagonist; hypertension; end organ protection; eplerenone; Dahl salt-sensitive rats
jaundice; bilirubin; sepsis; hepatitis; gall stones
Hysteresis loops are phenomena that sometimes are encountered in the
analysis of pharmacokinetic and pharmacodynamic relationships spanning from
pre-clinical to clinical studies. When hysteresis occurs it provides insight
into the complexity of drug action and disposition that can be encountered.
Hysteresis loops suggest that the relationship between drug concentration and
the effect being measured is not a simple direct relationship, but may have an
inherent time delay and disequilibrium, which may be the result of metabolites,
the consequence of changes in pharmacodynamics or the use of a non-specific
assay or may involve an indirect relationship. Counter-clockwise hysteresis has
been generally defined as the process in which effect can increase with time for
a given drug concentration, while in the case of clockwise hysteresis the
measured effect decreases with time for a given drug concentration. Hysteresis
loops can occur as a consequence of a number of different pharmacokinetic and
pharmacodynamic mechanisms including tolerance, distributional delay, feedback
regulation, input and output rate changes, agonistic or antagonistic active
metabolites, uptake into active site, slow receptor kinetics, delayed or
modified activity, time-dependent protein binding and the use of racemic drugs
among other factors. In this review, each of these various causes of hysteresis
loops are discussed, with incorporation of relevant examples of drugs
demonstrating these relationships for illustrative purposes. Furthermore, the
effect that pharmaceutical formulation has on the occurrence and potential
change in direction of the hysteresis loop, and the major pharmacokinetic /
pharmacodynamic modeling approaches utilized to collapse and model hysteresis
Cabozantinib, a potent pan-tyrosine kinase inhibitor, has been reported to provide enhanced antitumor efficacy by simultaneously inhibiting both MET and VEGF pathways, which are critical to tumor angiogenesis, survival and migration. It's very poor water solubility prevents its administration by the intravenous route, which may be important in patients unable to take the drug orally. In this study, we developed an efficient PEG-lipid-based polymeric micelle formulation with enhanced drug solubility and stability for cabozantinib delivery. DSPE-PEG2000 micelles encapsulating cabozantinib were prepared by a thin-film rehydration method followed by a lyophilization process to generate the dry dosage form. The average hydrodynamic diameter of freshly prepared micelles was 11 nm with a narrow size distribution, and the dry micelle cake could be fully reconstituted by rehydration. Approximately 75% of the drug was encapsulated into the lyophilized cake, and a sustained drug release profile was observed in simulated normal physiological release medium. Compared with the free cabozantinib solution, the drug-loaded micelles displayed significantly enhanced intracellular accumulation and cytotoxicity in human glioblastoma cancer cells and non-small lung cancer cells. These results suggest that the micellar formulation of cabozantinib may serve as a promising nanocarrier in anticancer treatments.
Cabozantinib; DSPE-PEG2000; Sustained release; Micellar solubilization; Lyophilization; Cytotoxicity
Opinions about public lands and the actions of private non-industrial forest owners in the western United States play important roles in forested landscape management as both public and private forests face increasing risks from large wildfires, pests and disease. This work presents the responses from two surveys, a random-sample telephone survey of more than 1500 residents and a mail survey targeting owners of parcels with 10 or more acres of forest. These surveys were conducted in three counties (Wallowa, Union, and Baker) in northeast Oregon, USA. We analyze these survey data using structural equation models in order to assess how individual characteristics and understanding of forest management issues affect perceptions about forest conditions and risks associated with declining forest health on public lands. We test whether forest understanding is informed by background, beliefs, and experiences, and whether as an intervening variable it is associated with views about forest conditions on publicly managed forests. Individual background characteristics such as age, gender and county of residence have significant direct or indirect effects on our measurement of understanding. Controlling for background factors, we found that forest owners with higher self-assessed understanding, and more education about forest management, tend to hold more pessimistic views about forest conditions. Based on our results we argue that self-assessed understanding, interest in learning, and willingness to engage in extension activities together have leverage to affect perceptions about the risks posed by declining forest conditions on public lands, influence land owner actions, and affect support for public policies. These results also have broader implications for management of forested landscapes on public and private lands amidst changing demographics in rural communities across the Inland Northwest where migration may significantly alter the composition of forest owner goals, understanding, and support for various management actions.
Proton-coupled oligopeptide transporters belong to the major facilitator superfamily (MFS) of membrane transporters. Recent crystal structures suggest the MFS fold facilitates transport through rearrangement of their two six-helix bundles around a central ligand binding site; how this is achieved, however, is poorly understood. Using modeling, molecular dynamics, crystallography, functional assays, and site-directed spin labeling combined with double electron-electron resonance (DEER) spectroscopy, we present a detailed study of the transport dynamics of two bacterial oligopeptide transporters, PepTSo and PepTSt. Our results identify several salt bridges that stabilize outward-facing conformations and we show that, for all the current structures of MFS transporters, the first two helices of each of the four inverted-topology repeat units form half of either the periplasmic or cytoplasmic gate and that these function cooperatively in a scissor-like motion to control access to the peptide binding site during transport.
•New higher-resolution structure of PepTSo•Salt bridges stabilizing outward-facing conformations are identified•The conserved prolines in helix 8 are shown to be important•The first two helices in each inverted-topology repeat form part of a gate
Fowler et al. use biophysical and modeling approaches to identify salt bridges in two peptide transporters that stabilize their outward-facing conformations. Their results also suggest that the first two helices in each inverted-topology repeat unit forms part of either of the two gates.
The aims of this Patient Reported Outcome Measurement Information System (PROMIS®) study are (1) to conceptualize children's subjective well-being (SWB), and (2) to produce item pools with excellent content validity for calibration and use in computerized adaptive testings (CAT).
Study Design and Setting
Children's SWB was defined through semi-structured interviews with experts, children (age 8-17 years-old), parents, and a systematic literature review to identify item concepts comprehensively covering the full spectrum of SWB. Item concepts were transformed into item expressions and evaluated for comprehensibility using cognitive interviews, reading level analysis, and translatability review.
Children's SWB comprises affective (Positive Affect) and global evaluation components (Life Satisfaction). Input from experts, children, parents, and the literature indicated that the eudaimonic dimension of SWB - i.e., a sense of meaning and purpose - could be evaluated. Item pools for Life Satisfaction (56 items), Positive Affect (53 items), and Meaning and Purpose (55 items) were produced. Small differences in comprehensibility of some items were observed between children and adolescents.
The SWB measures for children are the first to assess both the hedonic and eudaimonic aspects of SWB. Both children and youth seem to understand the concepts of a meaningful life, optimism, and goal orientation.
Child; Pediatric health; Subjective Well-Being; Item Bank; Measurement; Validity
Maternal obesity is associated with unfavorable outcomes, which may be reflected in the as yet undiscovered gene expression profiles of the umbilical cord (UC).
UCs from 12 lean (pre-gravid BMI < 24.9) and 10 overweight/obese (OW/OB, pre-gravid BMI ≥25) women without gestational diabetes were collected for gene expression analysis using Human Primeview microarrays (Affymetrix). Metabolic parameters were assayed in mother’s plasma and cord blood.
Although offspring birth weight and adiposity (at 2-wk) did not differ between groups, expression of 232 transcripts was affected in UC from OW/OB compared to those of lean mothers. GSEA analysis revealed an up-regulation of genes related to metabolism, stimulus and defense response and inhibitory to insulin signaling in the OW/OB group. We confirmed that EGR1, periostin, and FOSB mRNA expression was induced in UCs from OW/OB moms, while endothelin receptor B, KFL10, PEG3 and EGLN3 expression was decreased. Messenger RNA expression of EGR1, FOSB, MEST and SOCS1 were positively correlated (p<0.05) with mother’s first trimester body fat mass (%).
Our data suggest a positive association between maternal obesity and changes in UC gene expression profiles favoring inflammation and insulin resistance, potentially predisposing infants to develop metabolic dysfunction later on in life.
The combination of efavirenz with HIV-1 protease inhibitors (PI) results in complex interactions secondary to mixed induction and inhibition of oxidative metabolism. ACTG A5043 was a prospective, open-label, controlled, two-period, multiple-dose study with 55 healthy volunteers. The objective of the present study was to evaluate the potential pharmacokinetic interaction between efavirenz and dual PIs. The subjects received a daily dose of 600 mg efavirenz for 10 days with amprenavir 600 mg twice daily added at day 11 and were randomized to receive nelfinavir, indinavir, ritonavir, saquinavir, or no second PI on days 15–21. Intensive pharmacokinetic studies were conducted on day 14 and 21. Efavirenz plasma concentrations were fit to candidate models using weighted non-linear regression. The disposition of efavirenz was described by a linear two-compartment model with first order absorption following a fitted lag time. Apparent clearance (CLt/F), volume of distribution at steady state (Vss/F), inter-compartmental clearance, and the central and peripheral volume of distribution were estimated. The mean CLt/F and Vss/F of efavirenz were 0.126 l/h/kg and 4.412 l/kg, respectively. Both AUC and CLt/F of efavirenz remained unchanged after 7 days of dual PI dosing. The mean Vss/F of efavirenz increased an average of 89% across arms, ranging from 52% (nelfinavir) to 115% (indinavir) relative to efavirenz with amprenavir alone. Increases were also observed in Vp/F after the addition of nelfinavir, indinavir, ritonavir and saquinavir by 85%, 170%, 162% and 111%, respectively. In conclusion, concomitant administration of dual PIs is unlikely to have any clinically significant effect on the pharmacokinetics of CYP2B6 substrates in general or oral efavirenz specifically.
pharmacokinetic modeling; drug interactions; non-nucleoside reverse transcriptase inhibitor; efavirenz; HIV-1 protease inhibitors
Treatment of locally advanced head and neck squamous cell carcinoma (HNSCC) uses a multi-disciplinary approach often limited by the toxicity and drug resistance of platinum agents.
To test whether a nanocarrier-conjugated cisplatin boosting locoregional drug delivery improves tumor efficacy while decreasing systemic toxicity over systemic cisplatin in a murine model of locally advanced HNSCC.
A randomized, controlled, in vivo study compared standard cisplatin with nanocarrier (hyaluronan [HA])–conjugated cisplatin (HA-cisplatin) each at 50% of the maximum tolerated doses in a murine model of locally advanced HNSCC (10 mice/arm, each injected with 1 × 106 MDA-1986 HNSCC cells, with phosphate-buffered saline and HA-only control arms). Mice were treated for 3 weeks and observed for 3 additional weeks.
Academic medical center.
Forty female Nu/Nu mice. Randomization and treatment arms were initiated once tumor volumes reached 30 mm3.
Injection with MDA-1986 HNSCC cells followed by 3 weeks of treatment with cisplatin, HA-cisplatin, phosphate-buffered saline, or HA only.
Main Outcomes and Measures
Animal weights and tumor volumes were measured 3 times each week (modified RECIST [Response Evaluation Criteria in Solid Tumors]). At necropsy, animal kidneys were examined for nephrotoxic effects and cochleae were examined for ototoxic effects.
The mice treated with HA-cisplatin showed superior tumor efficacy (1 with complete clinical response, 3 with partial response, 1 with stable disease, and 5 with progressive disease) compared with standard cisplatin (no animals with complete clinical response, 1 with partial response, 1 with stable disease, and 8 with progressive disease), which was statistically significant (P=.003). All control animals developed progressive disease. Weight loss and body score were surrogate measures of treatment toxicity. The HA-cisplatin group had the least weight loss (mean [SD], 10.8% [4.7%]) compared with the cisplatin group (13.6% [5.6%]; P=.25). Body score dropped to 2 or less in all cisplatin-treated mice but not in any HA-cisplatin–treated mice, which also lacked any histologic signs of nephrotoxic or ototoxic effects.
Conclusions and Relevance
Nanoconjugated HA-cisplatin significantly improves tumor efficacy with lower toxicity compared with standard cisplatin in locally advanced HNSCC in vivo, justifying additional translational studies.
Radiation-damaged nanodiamonds (NDs) are ideal optical contrast agents for photoacoustic (PA) imaging in biological tissues due to their good biocompatibility and high optical absorbance in the near-infrared (NIR) range. Acid treated NDs are oxidized to form carboxyl groups on the surface, functionalized with polyethylene glycol (PEG) and human epidermal growth factor receptor 2 (HER2) targeting ligand for breast cancer tumor imaging. Because of the specific binding of the ligand conjugated NDs to the HER2-overexpressing murine breast cancer cells (4T1.2 neu), the tumor tissues are significantly delineated from the surrounding normal tissue at wavelength of 820 nm under the PA imaging modality. Moreover, HER2 targeted NDs (HER2-PEG-NDs) result in higher accumulation in HER2 positive breast tumors as compared to non-targeted NDs after intravenous injection (i.v.). Longer retention time of HER-PEG-NDs is observed in HER2 overexpressing tumor model than that in negative tumor model (4T1.2). This demonstrates that targeting moiety conjugated NDs have great potential for the sensitive detection of cancer tumors and provide an attractive delivery strategy for anti-cancer drugs.
Nanodiamond; Photoacoustic Imaging; Breast Cancer; HER2
Accumulating evidence, obtained largely in vitro, indicates that opioids regulate the genesis of neurons and glia and their precursors in the nervous system. Despite this evidence, few studies have assessed opioid receptor expression in identified cells within germinal zones or examined opioid effects on gliogenesis in vivo. To address this question, the role of opioids was explored in the subventricular zone (SVZ) and/or striatum of 2-5 day-old and/or adult ICR mice. The results showed that subpopulations of neurons, astrocytes, and oligodendrocytes in the SVZ and striatum differentially express μ, δ, and/or κ receptor immunoreactivity in a cell-type-specific and developmentally regulated manner. In addition, DNA synthesis was assessed by examining 5-bromo-2′-deoxyuridine (BrdU) incorporation into glial and non-glial precursors. In GFAP+ cells, morphine (a preferential μ agonist) significantly decreased BrdU-labeled astroglia compared to controls or mice co-treated with naltrexone plus morphine. Alternatively, in S100β+ cells, morphine did not significantly decrease BrdU incorporation; however, significant differences were noted between morphine and morphine plus naltrexone-treated mice. The majority of cells were non-GFAP+/non-S100β+. When BrdU incorporation was assessed within the total population (glia and non-glia), morphine had no net effect but naltrexone alone markedly increased BrdU incorporation. This suggests DNA synthesis in non-GFAP+/non-S100β+ cells is tonically suppressed by endogenous opioids. Assuming S100β and GFAP, respectively, distinguish among younger and older astroglia, this implies that astroglial replication becomes increasingly sensitive to morphine during maturation, and suggests that opioids differentially regulate the development of distinct subpopulations of glia and glial precursors.
astrocytes; oligodendrocytes; cell division; subventricular zone; striatum; μ opioid receptors; δ opioid receptors; κ opioid receptors; central nervous system development; opiate drug abuse; glial fibrillary acidic protein; S100β; drug abuse
Targeted lymphatic delivery of nanoparticles for drug delivery and imaging is primarily dependent on size and charge. Prior studies have observed increased lymphatic uptake and retentions of over 48 hrs for negatively charged particles compared to neutral and positively charged particles. We have developed new polymeric materials that extend retention over a more pharmaceutically relevant 7-day period. We used whole body fluorescence imaging to observe in mice the lymphatic trafficking of a series of anionic star poly-(6-O-methacryloyl-D-galactose) polymer-NIR dye (IR820) conjugates. The anionic charge of polymers was increased by modifying galactose moieties in the star polymers with succinic anhydride. Increasing anionic nature was associated with enhanced lymphatic uptake up to a zeta potential of ca. -40 mV; further negative charge did not affect lymphatic uptake. Compared to the 20% acid-conjugate, the 40 to 90% acid-star-polymer conjugates exhibited a 2.5- to 3.5-fold increase in lymphatic uptake in both the popliteal and iliac nodes. The polymer conjugates exhibited node half-lives of 2 to 20 hrs in the popliteal nodes and 19 to 114 hrs in the deeper iliac nodes. These polymer conjugates can deliver drugs or imaging agents with rapid lymphatic uptake and prolonged deep-nodal retention; thus they may provide a useful vehicle for sustained intralymphatic drug delivery with low toxicity.
Lymphatics; imaging; polymer trafficking
Computer-based clinical decision support (CDS) is an important component of the electronic health record (EHR). As an increasing amount of CDS is implemented, it will be important that this be accomplished in a fashion that assists in clinical decision making without imposing unacceptable demands and burdens upon the provider's practice.
The objective of our study was to explore an approach that allows CDS to be clinician-friendly from a variety of perspectives, to build a prototype implementation that illustrates features of the approach, and to gain experience with a pilot framework for assessment.
The paper first discusses the project's design philosophy and goals. It then describes a prototype implementation (Neuropath/CDS) that explores the approach in the domain of neuropathic pain and in the context of the US Veterans Administration EHR. Finally, the paper discusses a framework for assessing the approach, illustrated by a pilot assessment of Neuropath/CDS.
The paper describes the operation and technical design of Neuropath/CDS, as well as the results of the pilot assessment, which emphasize the four areas of focus, scope, content, and presentation.
The work to date has allowed us to explore various design and implementation issues relating to the approach illustrated in Neuropath/CDS, as well as the development and pilot application of a framework for assessment.
Internet; clinical decision support systems; electronic health records; neuropathic pain; therapeutics
The FANTOM5 project investigates transcription initiation activities in more than 1,000 human and mouse primary cells, cell lines and tissues using CAGE. Based on manual curation of sample information and development of an ontology for sample classification, we assemble the resulting data into a centralized data resource (http://fantom.gsc.riken.jp/5/). This resource contains web-based tools and data-access points for the research community to search and extract data related to samples, genes, promoter activities, transcription factors and enhancers across the FANTOM5 atlas.
Electronic supplementary material
The online version of this article (doi:10.1186/s13059-014-0560-6) contains supplementary material, which is available to authorized users.
Alphaherpesvirus reactivation from thoracic sympathetic ganglia (TSG) and transaxonal spread to target organs cause human visceral disease. Yet alphaherpesvirus latency in TSG has not been well characterized. In this study, quantitative PCR detected varicella-zoster virus (VZV), herpes simplex virus 1 (HSV-1), and HSV-2 DNA in 117 fresh TSG obtained postmortem from 15 subjects. VZV DNA was found in 76 (65%) ganglia from all subjects, HSV-1 DNA was found in 5 (4%) ganglia from 3 subjects, and no HSV-2 was found.
Background: An in-depth understanding of the fundamental principles that regulate thyroid hormone homeostasis is critical for the development of new diagnostic and treatment approaches for patients with thyroid disease.
Summary: Important clinical practices in use today for the treatment of patients with hypothyroidism, hyperthyroidism, or thyroid cancer are the result of laboratory discoveries made by scientists investigating the most basic aspects of thyroid structure and molecular biology. In this document, a panel of experts commissioned by the American Thyroid Association makes a series of recommendations related to the study of thyroid hormone economy and action. These recommendations are intended to promote standardization of study design, which should in turn increase the comparability and reproducibility of experimental findings.
Conclusions: It is expected that adherence to these recommendations by investigators in the field will facilitate progress towards a better understanding of the thyroid gland and thyroid hormone dependent processes.
Children's physical activity (PA) levels are commonly assessed in pediatric clinical research, but rigorous self-report assessment tools for children are scarce, and computer adaptive test implementations are rare. Our objective was to improve pediatric self-report measures of activity using semi-structured interviews with experts and children for conceptualization of a child-informed framework.
Semi-structured interviews were conducted to conceptualize physical activity, sedentary behaviors, and strengthening activities. We performed systematic literature reviews to identify item-level concepts used to assess these 3 domains.
We developed conceptual frameworks for each domain using words and phrases identified by children as relevant.
Semi-structured interview methods provide valuable information of children's perspectives and the ways children recall previous activities. Conceptualized domains of physical activity are based on the literature and expert views that also reflect children's experiences and understanding providing a basis for pediatric self-report instruments.
adolescent; child; female; health status; humans; information systems; male; outcome assessment (health care)/methods; parents; pediatrics/methods; physical activities; sedentary lifestyle; strengthening, quality of life
Children's activity level is commonly assessed in clinical research, but rigorous assessment tools for children are scarce. Our objectives were to improve pediatric activity self-report measures using qualitative methods to develop item pools that measure these concepts.
Based on the items generated from our conceptual framework development, we applied cognitive interviews and comprehensibility reviews to ensure children readily understood the items.
Our methods resulted in 129 unique items, physical activity (80 items), sedentary behaviors (23 items), and strengthening activities (26 items), that were comprehensible to children between the ages of 8 – 18 years. Comprehensibility review resulted in the deletion of 4 items.
The resultant item pools reflect children's experiences and understanding of the concepts of physical activity, sedentary behavior and strengthening activities. The item pools will undergo calibration using item response theory to support computer adaptive test administration of self and proxy reported outcomes.
adolescent; child; female; health status; humans; information systems; male; outcome assessment (health care)/methods; parents; pediatrics/methods; physical activities; sedentary lifestyle; strengthening, quality of life
sodium-potassium pump; Purkinje cell; neuron model; cerebellum; neural code; Na+/K+ pump; Na+/K+-ATPase
Efforts to elucidate the cellular and molecular mechanisms of regeneration have required the application of methods to detect specific cell types and tissues in a growing cohort of experimental animal models. For example, in the planarian Schmidtea mediterranea, substantial improvements to nucleic acid hybridization and electron microscopy protocols have facilitated the visualization of regenerative events at the cellular level. By contrast, immunological resources have been slower to emerge. Specifically, the repertoire of antibodies recognizing planarian antigens remains limited, and a more systematic approach is needed to evaluate the effects of processing steps required during sample preparation for immunolabeling.
To address these issues and to facilitate studies of planarian digestive system regeneration, we conducted a monoclonal antibody (mAb) screen using phagocytic intestinal cells purified from the digestive tracts of living planarians as immunogens. This approach yielded ten antibodies that recognized intestinal epitopes, as well as markers for the central nervous system, musculature, secretory cells, and epidermis. In order to improve signal intensity and reduce non-specific background for a subset of mAbs, we evaluated the effects of fixation and other steps during sample processing. We found that fixative choice, treatments to remove mucus and bleach pigment, as well as methods for tissue permeabilization and antigen retrieval profoundly influenced labeling by individual antibodies. These experiments led to the development of a step-by-step workflow for determining optimal specimen preparation for labeling whole planarians as well as unbleached histological sections.
We generated a collection of monoclonal antibodies recognizing the planarian intestine and other tissues; these antibodies will facilitate studies of planarian tissue morphogenesis. We also developed a protocol for optimizing specimen processing that will accelerate future efforts to generate planarian-specific antibodies, and to extend functional genetic studies of regeneration to post-transcriptional aspects of gene expression, such as protein localization or modification. Our efforts demonstrate the importance of systematically testing multiple approaches to species-specific idiosyncracies, such as mucus removal and pigment bleaching, and may serve as a template for the development of immunological resources in other emerging model organisms.
Electronic supplementary material
The online version of this article (doi:10.1186/s12861-014-0045-6) contains supplementary material, which is available to authorized users.
Planarian; Regeneration; Intestine; Monoclonal antibody screen; Immunohistochemistry; Immunofluorescence
To examine temporal trends in plasma viral load (pVL) suppression and antiretroviral resistance from 1997-2010 in British Columbia (BC), Canada, and determine characteristics, pVL ranges, and resistance profiles of HIV-positive individuals with unsuppressed pVL in 2010.
HIV-positive individuals ≥19 years old in the provincial database at the BC Centre for Excellence in HIV/AIDS were included. Virologic suppression was defined as two consecutive pVL <500 copies/mL within each calendar year. Temporal trends were evaluated using the Cochran-Armitage test. Persons with suppressed vs. unsuppressed pVL in 2010 were compared using the Pearson χ2 or Fisher’s exact test (categorical variables) and the Wilcoxon rank-sum test (quantitative variables), including unsuppressed individuals only if they were on antiretroviral therapy (ART) in 2010 or their baseline CD4 count was <350 cells/mm3 or <500 cells/mm3, in separate analyses.
The proportion of individuals with suppressed pVL increased from 24% to 80% (p<0.001). In comparative analyses, individuals with unsuppressed pVL (877 of 6142) were more likely to be female (30% vs. 16%), younger (median 43 vs. 48 years), have injection drug use history (38% vs. 30%), report Aboriginal ancestry (30% vs. 16%), and have hepatitis C co-infection (57% vs. 34%) (all p<0.001). Similar patterns were observed using the <500 cells/mm3 CD4 cut-off. The median pVL of all unsuppressed individuals in 2010 was 12,896 copies/mL (IQR 1,495-47,763).
The proportion of individuals achieving pVL suppression in BC has increased markedly since 1997, however further efforts are needed to maximize the individual and societal benefits of modern ART.
HIV; viral suppression; antiretroviral therapy; treatment as prevention; antiretroviral resistance; Canada
Hepatocellular carcinoma (HCC) is an increasingly lethal malignancy for which management is critically dependent on accurate imaging. Glypican-3 (GPC3) is a cell surface receptor overexpressed in most HCCs and provides a unique target for molecular diagnostics. The use of monoclonal antibodies (mAbs) that target GPC3 (αGPC3) in PET imaging has shown promise but comes with inherent limitations associated with mAbs such as long circulation times. This study used 89Zr-conjugated F(ab′)2 fragments directed against GPC3 (89Zr-αGPC3-F(ab′)2) to evaluate the feasibility of the fragments as a diagnostic immuno-PET imaging probe.
Immobilized ficin was used to digest αGPC3, creating αGPC3-F(ab′)2 fragments subsequently conjugated to 89Zr. In vivo biodistribution and PET studies were performed on GPC3-expressing HepG2 and GPC3-nonexpressing RH7777 orthotopic xenografts.
Reliable αGPC3-F(ab′)2 production via immobilized ficin digestion was verified by high-performance liquid chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. 89Zr-αGPC3-F(ab′)2 demonstrated F(ab′)2-dependent, antigen-specific cell binding. HepG2 tumor uptake was higher than any other tissue, peaking at 100 ± 21 percentage injected dose per gram (%ID/g) 24 h after injection, a value 33- to 38-fold higher than GPC3-nonexpressing RH7777 tumors. The blood half-life of the 89Zr-αGPC3-F(ab′)2 conjugate was approximately 11 h, compared with approximately 115 h for historic mAb controls. This shorter half-life enabled clear tumor visualization on PET 4 h after administration, with a resultant peak tumor-to-liver contrast ratio of 23.3. Blocking antigen-expressing tumors with an excess of nonradiolabeled αGPC3 resulted in decreased tumor uptake similar to native liver. The kidneys exhibited high tissue uptake, peaking at 24 h with 83 ± 12 %ID/g. HepG2 tumors ranging from 1.5 to 7 mm were clearly visible on PET, whereas larger RH7777 tumors displayed signal lower than background liver tissue.
This study demonstrates the feasibility of using 89Zr-αGPC3-F(ab′) 2 for intrahepatic tumor localization with small-animal PET. Faster blood clearance and lower background liver uptake enable excellent signal-to-noise ratios at early time points. Increased renal uptake is similar to that as has been seen with clinical radioactive peptide imaging. 89Zr-αGPC3-F(ab′)2 addresses some of the shortcomings of whole-antibody immuno-PET probes. Further optimization is warranted to maximize probe sensitivity and specificity in the process of clinical translation.
F(ab′)2; glypican-3; 89Zr; positron emission tomography (PET); hepatocellular carcinoma (HCC)