Vaccine hesitancy and emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) escaping vaccine-induced immune responses highlight the urgency for new COVID-19 therapeutics. Engineered angiotensin-converting enzyme 2 (ACE2) proteins with augmented binding affinities for SARS-CoV-2 spike (S) protein may prove to be especially efficacious against multiple variants. Using molecular dynamics simulations and functional assays, we show that three amino acid substitutions in an engineered soluble ACE2 protein markedly augmented the affinity for the S protein of the SARS-CoV-2 WA-1/2020 isolate and multiple VOCs: B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma) and B.1.617.2 (Delta). In humanized K18-hACE2 mice infected with the SARS-CoV-2 WA-1/2020 or P.1 variant, prophylactic and therapeutic injections of soluble ACE22.v2.4-IgG1 prevented lung vascular injury and edema formation, essential features of CoV-2-induced SARS, and above all improved survival. These studies demonstrate broad efficacy in vivo of an engineered ACE2 decoy against SARS-CoV-2 variants in mice and point to its therapeutic potential.

Standardized green tea extract was evaluated for exposure and toxicity in Beagle dogs following oral dosing by capsules. The main component (-)-epigallocatechin gallate (EGCG) accounted for 56-72% of the material. A 9-month chronic study (0, 200, 500, and 1000 mg/kg/day) was done in fasted dogs to take advantage of the reported improved catechin bioavailability with fasting. Extensive morbidity, mortality, and pathology of many major organs led to its early termination at 6.5 months and prevented identification of the toxicity mechanisms. A follow-up 13-week study examined the exposure to and toxicity of the extract. In general, toxicities were less severe than in the chronic study during the same interval. Dosing in a fed state resulted in considerably lower and less variable exposure than found under fasted conditions. Toxicity was less frequent and of lesser severity with lower exposure but limited sample size and large variability prevented reaching that definitive conclusion. Differences in mortality and morbidity between the preliminary terminated chronic and follow-up subchronic studies with the same dose of the same drug lot and similar exposure were not fully resolved as there may be other as yet unclear confounding factors.

Background: Angiotensin-converting enzyme (ACE) is highly expressed in renal proximal tubules, but ACE activity/levels in the urine are at least 100-fold lower than in the blood. Decreased proximal tubular ACE has been associated with renal tubular damage in both animal models and clinical studies. Because ACE is shed into urine primarily from proximal tubule epithelial cells, its urinary ACE measurement may be useful as an index of tubular damage. Objective and Methodology: We applied our novel approach—ACE phenotyping—to characterize urinary ACE in volunteer subjects. ACE phenotyping includes (1) determination of ACE activity using two substrates (ZPHL and HHL); (2) calculation of the ratio of hydrolysis of the two substrates (ZPHL/HHL ratio); (3) quantification of ACE immunoreactive protein levels; and (4) fine mapping of local ACE conformation with mAbs to ACE. Principal findings: In normal volunteers, urinary ACE activity was 140-fold less than in corresponding plasma/serum samples and did not differ between males and females. However, urinary ACE immunoreactivity (normalized binding of 25 mAbs to different epitopes) was strongly sex-dependent for the several mAbs tested, an observation likely explained by differences in tissue ACE glycosylation/sialylation between males and females. Urinary ACE phenotyping also allowed the identification of ACE outliers. In addition, daily variability of urinary ACE has potential utility as a feedback marker for dieting individuals pursuing weight loss. Conclusions/Significance: Urinary ACE phenotyping is a promising new approach with potential clinical significance to advance precision medicine screening techniques.

2,2,5,7, was administered by gavage in rats for 28 days at dose levels of 0, 100, 500, and 2000 mg/kg/day. PMCol administration induced decreases in body weight gains and food consumption, hepatotoxicity (increased TBILI, ALB, ALT, TP; increased relative liver weights; increased T4 and TSH), nephrotoxicity (increased BUN and BUN/CREAT, histopathology lesions), effect on lipid metabolism (increased CHOL), anemia, increase in WBC counts (total and differential), coagulation (FBGN↑and PT↓) and hyperkeratosis of the nonglandular stomach in the 2000 mg/kg/day dose group (in one or both sexes). In the 500 mg/kg/ day dose group, toxicity was seen to a lesser extent. In the 100 mg/kg/day dose group, only increased CHOL (females) was observed. To assess the toxicity of PMCol in male dogs it was administered orally by capsule administration for 28 days at dose levels of 0, 50, 200 and 800 mg/ kg/day (4 male dogs/dose group). PMCol treatment at 800 mg/kg/day resulted in pronounced toxicity to the male dogs. Target organs of toxicity were liver and thymus. Treatment at 200 mg/ kg/day resulted in toxicity consistent with slight adverse effect on the liver only. The results of the safety pharmacology study indicate that doses of 0, 50, 200 and 800 mg/kg administered orally did not have an effect on the QT interval, blood pressures and body temperatures following dosing over a 24-hour recording period. Under the conditions of this study, the no-observed-adverse effect level (NOAEL) for daily oral administration of PMCol by gavage for 28 days to male rats was 100 mg/kg/day and 50 mg/kg in male dogs. In female rats, the NOAEL was not established due to statistically significant and biologically meaningful increases in CHOL level seen in the 100 mg/kg/day dose group. The results of these studies indicated that administration of PMCol at higher dose levels resulted in severe toxicity in dogs and moderate toxicity in rats, however, administration at lower levels is considered to be less likely to result in toxicity following 28 days of exposure. Sex-related differences were seen in rats. Male rats appeared to have greater Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access Author ManuscriptToxicology. Author manuscript; available in PMC 2011 June 29. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript sensitivity to nephrotoxicity, while female animals had a greater incidence of hepatoxicity and changes in hematological parameters evaluated, especially at a dose of 500 mg/kg/day, which correlated to the higher plasma drug levels in female rats. I...

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