scholarly journals Reversion of Ebolavirus Disease from a Single Intramuscular Injection of a pan-Ebolavirus Immunotherapeutic

2022 ◽  
Author(s):  
Erin Kuang ◽  
Robert W. Cross ◽  
Maria McCavitt-Malvido ◽  
Dafna M Abelson ◽  
Viktoriya Borisevich ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Half Life ◽  
Intramuscular Injection ◽  
Life Extension ◽  
Limited Resources ◽  
Significant Protection ◽  
Post Exposure ◽  
Single Intramuscular Injection ◽  
Rapid Treatment

Intravenous administration (IV) of antiviral monoclonal antibodies (mAbs) is challenging due to limited resources for performing infusions during an ongoing epidemic. An ebolavirus therapeutic administered via intramuscular (IM) injection would reduce these burdens and allow rapid treatment of exposed individuals during an outbreak. Here, we demonstrate how MBP134, a two mAb pan-ebolavirus cocktail, reverses the course of Sudan ebolavirus (SUDV/Gulu) disease with a single IV or IM dose in non-human primates (NHPs) as far as five days post-exposure. Furthermore, we investigated the utility of adding half-life extension mutations to the MBP134 mAbs, ultimately creating a half-life extended cocktail designated MBP431. MBP431 demonstrated an extended serum half-life in vivo and offered complete or significant protection with a single IM dose delivered as a post-exposure prophylactic (PEP) or therapeutic in NHPs challenged with EBOV. These results support the use of MBP431 as a rapidly deployable IM medical countermeasure against every known ebolavirus.

116 INCORPORATION AND DEVELOPMENTAL TOXICITY OF QUANTUM DOT NANOPARTICLES IN AMPHIBIAN LARVAE

2017 ◽  
Vol 29 (1) ◽  
pp. 166 ◽  
Author(s):  
A. R. Julien ◽  
S. B. Park ◽  
C. K. Vance ◽  
P. L. Ryan ◽  
S. T. Willard ◽  
...  
Keyword(s):  
Quantum Dot ◽  
In Vivo Imaging ◽  
Half Life ◽  
Imaging System ◽  
Fluorescence Emission ◽  
Total Flux ◽  
Intestinal Lumen ◽  
Post Exposure ◽  
Amphibian Larvae

The use of nanoparticles both commercially and pharmaceutically has increased over the past decade, including fluorescent quantum dot nanoparticles (QD) in biochemical research for in vivo imaging. Previous studies have reported the toxic effects of nanoparticles, but their effects on larval metamorphosis and animal development and growth have not been thoroughly examined. Additionally, the method of uptake of nanoparticles by larval systems is unknown. Amphibian larvae are an ideal model for assessing toxicity because of their sensitivity to environmental contaminants and rapid and easily observable developmental stages. We used Anaxyrus fowleri tadpoles to investigate QD (≤ 25 nm diameter) integration into larvae and possible deleterious effects on their growth and development. Tadpoles (A. fowleri; n = 5/group) were placed in 24-well plates containing 1 mL of distilled water and increasing concentrations of QD (0, 1, and 2 nM) 72 h post-hatch. The fluorescence emission of QD in wells was detected at various time points (1, 2, 24, 48, and 72 h) using the in vivo imaging system (IVIS). A subset of tadpoles was killed (MS-222) and sectioned for histopathology. Remaining tadpoles were monitored throughout development. Fluorescence emission of QD in sectioned tadpoles was visualised using an EVOS Cell Imaging System. Developmental metrics of living tadpoles were recorded until metamorphosis. Fluorescence intensity between controls and dosage groups were analysed by ANOVA-1, followed by Student’s l.s.d. test to evaluate the effects of QD concentration and exposure time. The threshold of significance was P < 0.05. The rate of incorporation of QD into tadpoles was determined using the equation y = C + Ao*2(–x/t1/2), where t1/2 is the half-life of QD remaining in solution. The IVIS imaging revealed a rapid decrease of QD fluorescence (total flux) signals from the aqueous tadpole environment. Decreases in fluorescence occurred within 1 h post-exposure and appeared dose and time dependent, with signal nearly gone within 48 h. Half-life of total flux (time necessary for tadpoles to absorb half of the QD in solution) is 20.75 h (R2 = 0.92) and 2.54 h (R2 = 0.96) for 1 nm and 2 nm QD in solution, respectively. The EVOS imaging revealed integration of QD and localization into tadpole tissues. Fluorescence was exclusively found within the mouth, gills, and sections of the intestinal lumen of exposed tadpoles within the first hour. Dose-dependent increases in fluorescence within tissue were observed at each time-point. No signal was observed in controls. In remaining live tadpoles, QD treated tadpoles were smaller in size [t(34) = 2.35, P = 0.024] than controls. Findings reveal that (1) A. fowleri tadpoles integrate and accumulate nanoparticles, without detectable excretion within 72 h post-exposure, and (2) nanoparticles impede normal tadpole development. Ongoing studies are determining the effects of QD exposure on complete tadpole metamorphosis. The work was supported by USDA-ARS Biophotonics Initiative grant #58–6402–3-018.

scholarly journals Engineering of Long-Circulating Peptidoglycan Hydrolases Enables Efficient Treatment of Systemic Staphylococcus aureus Infection

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Anna M. Sobieraj ◽  
Markus Huemer ◽  
Léa V. Zinsli ◽  
Susanne Meile ◽  
Anja P. Keller ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Human Blood ◽  
Half Life ◽  
Multidrug Resistant ◽  
Life Extension ◽  
Drug Resistant ◽  
Content Type ◽  
Peptidoglycan Hydrolases ◽  
Life Threatening

ABSTRACT Staphylococcus aureus is a human pathogen causing life-threatening diseases. The increasing prevalence of multidrug-resistant S. aureus infections is a global health concern, requiring development of novel therapeutic options. Peptidoglycan-degrading enzymes (peptidoglycan hydrolases, PGHs) have emerged as a highly effective class of antimicrobial proteins against S. aureus and other pathogens. When applied to Gram-positive bacteria, PGHs hydrolyze bonds within the peptidoglycan layer, leading to rapid bacterial death by lysis. This activity is highly specific and independent of the metabolic activity of the cell or its antibiotic resistance patterns. However, systemic application of PGHs is limited by their often low activity in vivo and by an insufficient serum circulation half-life. To address this problem, we aimed to extend the half-life of PGHs selected for high activity against S. aureus in human serum. Half-life extension and increased serum circulation were achieved through fusion of PGHs to an albumin-binding domain (ABD), resulting in high-affinity recruitment of human serum albumin and formation of large protein complexes. Importantly, the ABD-fused PGHs maintained high killing activity against multiple drug-resistant S. aureus strains, as determined by ex vivo testing in human blood. The top candidate, termed ABD_M23, was tested in vivo to treat S. aureus-induced murine bacteremia. Our findings demonstrate a significantly higher efficacy of ABD_M23 than of the parental M23 enzyme. We conclude that fusion with ABD represents a powerful approach for half-life extension of PGHs, expanding the therapeutic potential of these enzybiotics for treatment of multidrug-resistant bacterial infections. IMPORTANCE Life-threatening infections with Staphylococcus aureus are often difficult to treat due to the increasing prevalence of antibiotic-resistant bacteria and their ability to persist in protected niches in the body. Bacteriolytic enzymes are promising new antimicrobials because they rapidly kill bacteria, including drug-resistant and persisting cells, by destroying their cell wall. However, when injected into the bloodstream, these enzymes are not retained long enough to clear an infection. Here, we describe a modification to increase blood circulation time of the enzymes and enhance treatment efficacy against S. aureus-induced bloodstream infections. This was achieved by preselecting enzyme candidates for high activity in human blood and coupling them to serum albumin, thereby preventing their elimination by kidney filtration and blood vessel cells.

Activity of Amikacin, Gentamicin and Kanamycin against Pseudomonas Aeruginosa

1976 ◽  
Vol 4 (3) ◽  
pp. 165-175 ◽  
Author(s):  
Jose Ximenes ◽  
Orlando Natale Bassoi ◽  
Jairo Perche de Menezes ◽  
Wilson Fry
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Half Life ◽  
Intramuscular Injection ◽  
Saline Solution ◽  
Serum Levels ◽  
Post Dose ◽  
Mueller Hinton ◽  
Single Intramuscular Injection ◽  
Mueller Hinton Broth

The activity of amikacin, gentamicin and kanamycin was tested in vitro against clinical isolates of Pseudomonas aeruginosa. Concentrations of the antibiotics in serum and in saline solution were prepared according to serum levels produced in volunteers 15 minutes, 1, 2, and 6 hours after a single intramuscular injection of 500 mg amikacin, 80 mg gentamicin and 500 mg kanamycin. Following isolation of the Pseudomonas strains in cultures, they were incubated and seeded in Mueller-Hinton broth, then 107 dilutions of the organisms were kept in contact with the prepared antibiotic solutions in serum and in saline solution for three hours, the approximate half-life of the antibiotics in serum. Amikacin was active at concentrations seen six hours post-dose, inhibiting growth in a total of 72·5% of seeded plates. Gentamicin was active for only two hours and inhibited growth in 2·5% of the plates. Kanamycin showed no anti-pseudomonal activity.

scholarly journals The extended recovery ring-stage survival assay provides superior prediction of patient clearance half-life and increases throughput

2019 ◽  
Author(s):  
Sage Z. Davis ◽  
Puspendra P. Singh ◽  
Katelyn M. Vendrely ◽  
Douglas A. Shoue ◽  
Lisa A. Checkley ◽  
...  
Keyword(s):  
Southeast Asia ◽  
Half Life ◽  
Artemisinin Resistance ◽  
Pcr Amplification ◽  
Fold Change ◽  
Ring Stage ◽  
Post Exposure ◽  
Survival Assay

Abstract Background Tracking and understanding artemisinin resistance is key for preventing global setbacks in malaria eradication efforts. The ring-stage survival assay (RSA) is the current gold standard for in vitro artemisinin resistance phenotyping. However, the RSA has several drawbacks: it is relatively low throughput, has high variance due to microscopy readout, and correlates poorly with the current benchmark for in vivo resistance, patient clearance half-life post-artemisinin treatment. Here a modified RSA is presented, the extended Recovery Ring-stage Survival Assay (eRRSA), using 15 cloned patient isolates from Southeast Asia with a range of patient clearance half-lives, including parasite isolates with and without kelch13 mutations. Methods P. falciparum cultures were synchronized with single layer Percoll during the schizont stage of the erythrocytic cycle. Cultures were left to reinvade to early ring-stage and parasitemia was quantified using flow cytometry. Cultures were diluted to 2% hematocrit and 0.5% parasitemia in a 96-well plate to start the assay, allowing for increased throughput and decreased variability between biological replicates. Parasites were treated with 700nM of dihydroartemisinin or an equivalent amount of dimethyl sulfoxide (DMSO) for 6 h, washed three times in drug-free media, and incubated for 66 or 114 h, when samples were collected and frozen for PCR amplification. A SYBR Green-based quantitative PCR method was used to quantify the fold-change between treated and untreated samples. Results 15 cloned patient isolates from Southeast Asia with a range of patient clearance half-lives were assayed using the eRRSA. Due to the large number of pyknotic and dying parasites at 66 h post-exposure (72 h sample), parasites were grown for an additional cell cycle (114 h post-exposure, 120 h sample), which drastically improved correlation with patient clearance half-life compared to the 66 h post-exposure sample. A Spearman correlation of 0.8393 between fold change and patient clearance half-life was identified in these 15 isolates from Southeast Asia, which is the strongest correlation reported to date. Conclusions eRRSA drastically increases the efficiency and accuracy of in vitro artemisinin resistance phenotyping compared to the traditional RSA, which paves the way for extensive in vitro phenotyping of hundreds of artemisinin resistant parasites.

Monoclonal Antibodies to Tissue-Type Plasminogen Activator which Prolong its Clearance In Vivo

1989 ◽  
Vol 61 (02) ◽  
pp. 259-261
Author(s):  
Thomas M Reilly ◽  
Robert M Knabb ◽  
Andrew T Chiu ◽  
David L Bradfute ◽  
Pieter B M W M Timmermans
Keyword(s):  
Monoclonal Antibodies ◽  
Plasminogen Activator ◽  
Half Life ◽  
Cell Receptor ◽  
Tissue Type ◽  
Human Hepatoma Cells ◽  
Tissue Type Plasminogen Activator ◽  
Type Plasminogen Activator

SummaryThree murine monoclonal antibodies to tissue-type plasminogen activator (t-PA) were evaluated for their effects on the binding of iodinated t-PA to cultured human hepatoma cells (Hep G2), and on extending the half-life of t-PA injected into rabbits. Two of the antibodies, AE5 and EG2, significantly inhibited t-PA binding in vitro, and extended the in vivo half-life of t-PA four to five-fold. A third antibody, BA10, which had a much smaller inhibitory effect on t-PA binding, had no influence in extending t-PA’s half-life. MOPC-21, a control antibody not directed to t-PA, had no effect on either test. Our results are the first to correlate different compounds’ effects on t-PA binding with their ability to retard t-PA clearance in vivo, and provide additional evidence for the importance of a liver cell receptor in the t-PA clearance process.

scholarly journals Improved in vivo anti-tumor effects of IgA-Her2 antibodies through half-life extension and serum exposure enhancement by FcRn targeting

mAbs ◽  
2015 ◽  
Vol 8 (1) ◽  
pp. 87-98 ◽  
Author(s):  
Saskia Meyer ◽  
Maaike Nederend ◽  
J.H. Marco Jansen ◽  
Karli R. Reiding ◽  
Shamir R. Jacobino ◽  
...  
Keyword(s):  
Half Life ◽  
Life Extension

scholarly journals Fc-GDF15 glyco-engineering and receptor binding affinity optimization for body weight regulation

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ella Fung ◽  
Liya Kang ◽  
Diana Sapashnik ◽  
Susan Benard ◽  
Annette Sievers ◽  
...  
Keyword(s):  
Weight Loss ◽  
Binding Affinity ◽  
Receptor Binding ◽  
Half Life ◽  
Metabolic Diseases ◽  
Single Point ◽  
Life Extension ◽  
Single Point Mutation ◽  
Receptor Binding Site

AbstractGDF15 is a distant TGF-β family member that induces anorexia and weight loss. Due to its function, GDF15 has attracted attention as a potential therapeutic for the treatment of obesity and its associated metabolic diseases. However, the pharmacokinetic and physicochemical properties of GDF15 present several challenges for its development as a therapeutic, including a short half-life, high aggregation propensity, and protease susceptibility in serum. Here, we report the design, characterization and optimization of GDF15 in an Fc-fusion protein format with improved therapeutic properties. Using a structure-based engineering approach, we combined knob-into-hole Fc technology and N-linked glycosylation site mutagenesis for half-life extension, improved solubility and protease resistance. In addition, we identified a set of mutations at the receptor binding site of GDF15 that show increased GFRAL binding affinity and led to significant half-life extension. We also identified a single point mutation that increases p-ERK signaling activity and results in improved weight loss efficacy in vivo. Taken together, our findings allowed us to develop GDF15 in a new therapeutic format that demonstrates better efficacy and potential for improved manufacturability.

scholarly journals Engineering a Monomeric Fc Domain Modality by N-Glycosylation for the Half-life Extension of Biotherapeutics

2013 ◽  
Vol 288 (23) ◽  
pp. 16529-16537 ◽  
Author(s):  
Tetsuya Ishino ◽  
Mengmeng Wang ◽  
Lidia Mosyak ◽  
Amy Tam ◽  
Weili Duan ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Tandem Repeat ◽  
Half Life ◽  
Fusion Proteins ◽  
Life Extension ◽  
Dependent Manner ◽  
Neonatal Fc Receptor ◽  
Dimer Interface ◽  
Ph Dependent

Human IgG is a bivalent molecule that has two identical Fab domains connected by a dimeric Fc domain. For therapeutic purposes, however, the bivalency of IgG and Fc fusion proteins could cause undesired properties. We therefore engineered the conversion of the natural dimeric Fc domain to a highly soluble monomer by introducing two Asn-linked glycans onto the hydrophobic CH3-CH3 dimer interface. The monomeric Fc (monoFc) maintained the binding affinity for neonatal Fc receptor (FcRn) in a pH-dependent manner. We solved the crystal structure of monoFc, which explains how the carbohydrates can stabilize the protein surface and provides the rationale for molecular recognition between monoFc and FcRn. The monoFc prolonged the in vivo half-life of an antibody Fab domain, and a tandem repeat of the monoFc further prolonged the half-life. This monoFc modality can be used to improve the pharmacokinetics of monomeric therapeutic proteins with an option to modulate the degree of half-life extension.

UNTERSUCHUNGEN ÜBER DEN STOFFWECHSEL VON 17α-HYDROXY-19-NOR-PROGESTERON-CAPRONAT BEIM MENSCHEN IN VIVO UND VON 17α-HYDROXY-19-NOR-PROGESTERON BEI DER RATTE IN VITRO

1966 ◽  
Vol 51 (1) ◽  
pp. 114-130 ◽  
Author(s):  
H. Breuer ◽  
B. P. Lisboa
Keyword(s):  
Urinary Excretion ◽  
Comparative Studies ◽  
Intramuscular Injection ◽  
Pregnant Patient ◽  
Main Metabolite ◽  
Postmenopausal Patient ◽  
Enzyme Preparations ◽  
Single Intramuscular Injection

ABSTRACT The effect of a single intramuscular injection of 300 mg of 17α-hydroxy-19-nor-progesterone caproate on the urinary excretion of oestrogens, 17-ketosteroids, 17-hydroxycorticosteroids and the »pregnanetriol«-fraction has been studied in two pregnant patients and in two postmenopausal patients. In one postmenopausal patient, small and irregular increases in the excretion of oestrogens, 17-hydroxycorticosteroids and »pregnanetriol«-fraction were observed; in one pregnant patient, there was also an increase in the excretion of oestrone and 17β-oestradiol. Apart from these instances, the excretion of the steroid fractions investigated remained unchanged after administration of 17α-hydroxy-19-nor-progesterone caproate. In addition to a 19-nor-pregnanetriol isomer, a 19-nor-pregnanediol-20-one was found as main metabolite in the »pregnanetriol«-fraction. In comparative studies, the metabolism of 17α-hydroxyprogesterone and 17α-hydroxy-19-nor-progesterone was investigated, using the cytoplasmic and microsomal fractions of rat liver as enzyme preparations. With NADPH2 as cofactor the two steroids yielded predominantly ring A saturated compounds, whereas with NADH2 only 20α- and 20β-hydroxysteroids were formed. After incubation of 17α-hydroxyprogesterone with the microsomal and/or cytoplasmic fractions, the following metabolites were identified: 17α-hydroxy-5α-pregnane-3,20-dione; 17α,20β-dihydroxy-pregn-4-en-3-one; 17α,20α-dihydroxy-pregn-4-en-3-one; 3β,17α-dihydroxy-5α-pregnan-20-one; 3β,17α-dihydroxy-5β-pregnan-20-one; 3α, 17α-dihydroxy-5β-pregnan-20-one. After incubation of 17α-hydroxy-19-nor-progesterone the following metabolites were characterized: 17α-hydroxy-19-nor-5α-pregnane-3,20-dione; 17α,20β-dihydroxy-19-nor-pregn4-en-3-one; 17α,20α-dihydroxy-19-nor-pregn-4-en-3-one; 3β,17α-dihydroxy-19-nor-5α-pregnan-20-one; 3α,17α-dihydroxy-19-nor-5β-pregnan-20-one. These results shows that no qualitative differences exist in the metabolism of 17α-hydroxyprogesterone and 17α-hydroxy-19-nor-progesterone.

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