Preclinical pharmacology of the natural product anticancer agent 10-hydroxycamptothecin, an inhibitor of topoisomerase I

Purpose: 10-Hydroxycamptothecin (HCPT) is an indole alkaloid isolated from a Chinese tree, Camptotheca acuminata, and has a wide spectrum of anticancer activity in vitro and in vivo mainly through inhibitory effects on topoisomerase I. HCPT has been shown to be more potent and less toxic than camptothecin and has recently undergone clinical trials. To determine how HCPT might be best used as an anticancer agent, preclinical studies of the pharmacokinetics, tissue distribution, metabolism and elimination of HCPT in rats were undertaken. Methods: HCPT was administered to rats by i.v. bolus injection at doses of 1, 3, and 10 mg/kg body weight. HCPT (lactone and carboxylate) and its metabolites in plasma, urine, feces, and various tissues were quantitated by reversed-phase HPLC. Pharmacokinetic parameters were then estimated. Results: Following i.v. administration at doses of 3 or 10 mg/kg, the plasma concentration-time profile for lactone HCPT could be best described by a three-compartment model, with terminal elimination half-lives of 140.4 and 428.6 min, respectively. A two-compartment model was used to fit the plasma concentration-time curve at 1 mg/kg, with a terminal elimination half-life of 30.5 min. Carboxylate HCPT had a longer half-life than the lactone form of HCPT. During the initial 6 h after dosing, urinary excretion was the major route of elimination, and fecal excretion became the major route of elimination thereafter. HCPT was widely distributed to various tissues including the enterohepatic system, kidney, and bone marrow. The lactone form of HCPT was detectable in various tissues examined up to 72 h after dosing at all the three test doses. HCPT glucuronides were present in plasma, urine, feces and various tissues. No significant toxicity was observed at doses of 1 or 3 mg/kg. Polyuria and hematuria were observed only during the initial 3 h after dosing at 10 mg/kg. Conclusions: Prolonged elimination of HCPT in vivo may have a significant impact on its therapeutic effects. HCPT is metabolized to its carboxylate form and glucuronides. Dose-dependent toxicity was observed with i.v. administration of HCPT. The results of this study should be useful in the design of future human trials with this anticancer drug. Received: 6 September 1996 / Accepted: 15 July 1997     

Demonstration of ATP-Dependent,Transcellular Transport of Lipid Across the Lymphatic Endothelium Using an In Vitro Model of the Lacteal

Pharmaceutical Research - The lymphatic system plays crucial roles in tissue fluid balance, trafficking of immune cells, and the uptake of dietary lipid from the intestine. Given these roles there...     

Longer duration of homelessness is associated with a lower likelihood of non-detectable plasma HIV-1 RNA viral load among people who use illicit drugs in a Canadian setting

Homelessness is common among people who use drugs (PWUD) and, for those living with HIV/AIDS, an important contributor to sub-optimal HIV treatment outcomes. This study aims to investigate the relationship between the duration of homelessness and the likelihood of plasma HIV-1 RNA viral load (VL) non-detectability among a cohort of HIV-positive PWUD. We used data from the ACCESS study, a long-running prospective cohort study of HIV-positive PWUD linked to comprehensive HIV clinical records including systematic plasma HIV-1 RNA VL monitoring. We estimated the longitudinal relationship between the duration of homelessness and the likelihood of exhibiting a non-detectable VL (i.e., <500?copies/mL plasma) using generalized linear mixed-effects modelling. Between May 1996 and June 2014, 922 highly active antiretroviral therapy-exposed participants were recruited and contributed 8188 observations. Of these, 4800 (59%) were characterized by non-detectable VL. Participants reported they were homeless in 910 (11%) interviews (median: six months, interquartile range: 6–12 months). A longer duration of homelessness was associated with lower odds of VL non-detectability (adjusted odds ratio?=?0.71 per six-month period of homelessness, 95% confidence interval: 0.60–0.83) after adjustment for age, ancestry, drug use patterns, engagement in addiction treatment, and other potential confounders. Longer durations of episodes of homelessness in this cohort of HIV-positive illicit drug users were associated with a lower likelihood of plasma VL non-detectability. Our findings suggest that interventions that seek to promptly house homeless individuals, such as Housing First approaches, might assist in maximizing the clinical and public health benefits of antiretroviral therapy among people living with HIV/AIDS.     

Association between neuromelanin-sensitive MRI signal and psychomotor slowing in late-life depression

Late-life depression (LLD) is a prevalent and disabling condition in older adults that is often accompanied by slowed processing and gait speed. These symptoms are related to impaired dopamine function and sometimes remedied by levodopa (L-DOPA). In this study, we recruited 33 older adults with LLD to determine the association between a proxy measure of dopamine function—neuromelanin-sensitive magnetic resonance imaging (NM-MRI)—and baseline slowing measured by the Digit Symbol test and a gait speed paradigm. In secondary analyses, we also assessed the ability of NM-MRI to predict L-DOPA treatment response in a subset of these patients (N = 15) who received 3 weeks of L-DOPA. We scanned a further subset of these patients (N = 6) with NM-MRI at baseline and after treatment to preliminarily evaluate the effects of L-DOPA treatment on the NM-MRI signal. We found that lower baseline NM-MRI correlated with slower baseline gait speed (346 of 1807 substantia nigra–ventral tegmental area (SN-VTA) voxels, P_corrected = 0.038), particularly in the more medial, anterior, and dorsal SN-VTA. Secondary analyses failed to show an association between baseline NM-MRI and treatment-related changes in gait speed, processing speed, or depression severity (all P_corrected > 0.361); we however found preliminary evidence of increases in the NM-MRI signal 3 weeks post-treatment with L-DOPA compared to baseline (200 of 1807 SN-VTA voxels; P_corrected = 0.046), although the small sample size of these preliminary analyses warrants caution in their interpretation and future replications. Overall, our findings indicate that NM-MRI is sensitive to variability in gait speed in patients with LLD, suggesting this non-invasive MRI measure may provide a promising marker for dopamine-related psychomotor slowing in geriatric neuropsychiatry.Subject terms: Cognitive neuroscience, Depression, Biomarkers     

SARS-CoV-2 Spike Protein Disrupts Blood–Brain Barrier Integrity via RhoA Activation

The SARS-CoV-2 spike protein has been shown to disrupt blood–brain barrier (BBB) function, but its pathogenic mechanism of action is unknown. Whether angiotensin converting enzyme 2 (ACE2), the viral binding site for SARS-CoV-2, contributes to the spike protein-induced barrier disruption also remains unclear. Here, a 3D-BBB microfluidic model was used to interrogate mechanisms by which the spike protein may facilitate barrier dysfunction. The spike protein upregulated the expression of ACE2 in response to laminar shear stress. Moreover, interrogating the role of ACE2 showed that knock-down affected endothelial barrier properties. These results identify a possible role of ACE2 in barrier homeostasis. Analysis of RhoA, a key molecule in regulating endothelial cytoskeleton and tight junction complex dynamics, reveals that the spike protein triggers RhoA activation. Inhibition of RhoA with C3 transferase rescues its effect on tight junction disassembly. Overall, these results indicate a possible means by which the engagement of SARS-CoV-2 with ACE2 facilitates disruption of the BBB via RhoA activation. Understanding how SARS-CoV-2 dysregulates the BBB may lead to strategies to prevent the neurological deficits seen in COVID-19 patients.

Graphic Abstract

     

Effects of differential privacy techniques: Considerations for end users

We study the effects of differentially private (DP) noise injection techniques in a survey data setting, using the release of cost of early care and education estimates from the National Survey of Early Care and Education as a motivating example. As an example of how DP noise injection affects statistical estimates, our analysis compares the relative performance of DP techniques in the context of releasing estimates of means, medians, and regression coefficients. The results show that for many statistics, basic DP techniques show good performance provided that the privacy budget does not need to be split over too many estimates. Throughout, we show that small decisions, such as the number of bins in a histogram or the scaling of a variable in a regression equation, can have sometimes dramatic effects on the end results. Because of this, it is important to develop DP techniques with an eye towards the most important aspects of the data for end users.     

Endogenous theta stimulation during meditation predicts reduced opioid dosing following treatment with Mindfulness-Oriented Recovery Enhancement

Veterans experience chronic pain at greater rates than the rest of society and are more likely to receive long-term opioid therapy (LTOT), which, at high doses, is theorized to induce maladaptive neuroplastic changes that attenuate self-regulatory capacity and exacerbate opioid dose escalation. Mindfulness meditation has been shown to modulate frontal midline theta (FMT) and alpha oscillations that are linked with marked alterations in self-referential processing. These adaptive neural oscillatory changes may promote reduced opioid use and remediate the neural dysfunction occasioned by LTOT. In this study, we used electroencephalography (EEG) to assess the effects of a mindfulness-based, cognitive training intervention for opioid misuse, Mindfulness-Oriented Recovery Enhancement (MORE), on alpha and theta power and FMT coherence during meditation. We then examined whether these neural effects were associated with reduced opioid dosing and changes in self-referential processing. Before and after 8 weeks of MORE or a supportive psychotherapy control, veterans receiving LTOT (N = 62) practiced mindfulness meditation while EEG was recorded. Participants treated with MORE demonstrated significantly increased alpha and theta power (with larger theta power effect sizes) as well as increased FMT coherence relative to those in the control condition—neural changes that were associated with altered self-referential processing. Crucially, MORE significantly reduced opioid dose over time, and this dose reduction was partially statistically mediated by changes in frontal theta power. Study results suggest that mindfulness meditation practice may produce endogenous theta stimulation in the prefrontal cortex, thereby enhancing inhibitory control over opioid dose escalation behaviors.Subject terms: Human behaviour, Addiction     

High Proportion of Heteroresistance in gyrA and gyrB in Fluoroquinolone-Resistant Mycobacterium tuberculosis Clinical Isolates

Heteroresistance is the coexistence of populations with differing nucleotides at a drug resistance locus within a sample of organisms. Although Sanger sequencing is the gold standard for sequencing, it may be less sensitive than deep sequencing for detecting fluoroquinolone heteroresistance in Mycobacterium tuberculosis. Twenty-seven fluoroquinolone monoresistant and 11 fluoroquinolone-susceptible M. tuberculosis isolates were analyzed by Sanger and Illumina deep sequencing. Individual sequencing reads were analyzed to detect heteroresistance in the gyrA and gyrB genes. Heteroresistance to fluoroquinolones was identified in 10/26 (38%) phenotypically fluoroquinolone-resistant samples and 0/11 (P = 0.02) fluoroquinolone-susceptible controls. One resistant sample was excluded because of contamination with the laboratory strain M. tuberculosis H37Rv. Sanger sequencing revealed resistance-conferring mutations in 15 isolates, while deep sequencing revealed mutations in 20 isolates. Isolates with fluoroquinolone resistance-conferring mutations by Sanger sequencing all had at least those same mutations identified by deep sequencing. By deep sequencing, 10 isolates had a single fixed (defined as >95% frequency) mutation, while 10 were heteroresistant, 5 of which had a single unfixed (defined as <95% frequency) mutation and 5 had multiple unfixed mutations. Illumina deep sequencing identified a higher proportion of fluoroquinolone-resistant M. tuberculosis isolates with heteroresistance than did Sanger sequencing. The heteroresistant isolates frequently demonstrated multiple mutations, but resistant isolates with fixed mutations each had only a single mutation.