scholarly journals The Artemisinin Derivative Artemisone Is a Potent Inhibitor of Human Cytomegalovirus Replication

2018 ◽  
Vol 62 (7) ◽  
Author(s):  
E. Oiknine-Djian ◽  
Y. Weisblum ◽  
A. Panet ◽  
H. N. Wong ◽  
R. K. Haynes ◽  
...  
Keyword(s):  
Human Cytomegalovirus ◽  
Clinical Studies ◽  
Ex Vivo ◽  
Antiviral Treatment ◽  
Lung Fibroblasts ◽  
Reversible Inhibitor ◽  
Artemisinin Derivative ◽  
Viral Kinetics ◽  
Antiviral Efficacy

ABSTRACTHuman cytomegalovirus (HCMV) is a major cause of disease in immunocompromised individuals and the most common cause of congenital infection and neurosensorial disease. The expanding target populations for HCMV antiviral treatment along with the limitations of the currently available HCMV DNA polymerase inhibitors underscore the need for new antiviral agents with alternative modes of action. The antimalarial artemisinin derivative artesunate was shown to inhibit HCMVin vitroyet has demonstrated limited antiviral efficacyin vivo, prompting our search for more potent anti-HCMV artemisinin derivatives. Here we show that the innovative artemisinin derivative artemisone, which has been screened for its activity against malaria parasites in human clinical studies, is a potent and noncytotoxic inhibitor of HCMV. Artemisone exhibited an antiviral efficacy comparable to that of ganciclovir (50% effective concentration, 1.20 ± 0.46 μM) in human foreskin fibroblasts, with enhanced relative potency in lung fibroblasts and epithelial cells. Significantly, the antiviral efficacy of artemisone was consistently ≥10-fold superior to that of artesunate in all cells. Artemisone effectively inhibited both laboratory-adapted and low-passage-number clinical strains, as well as drug-resistant HCMV strains. By using quantitative viral kinetics and gene expression studies, we show that artemisone is a reversible inhibitor targeting an earlier phase of the viral replication cycle than ganciclovir. Importantly, artemisone most effectively inhibited HCMV infectionex vivoin a clinically relevant multicellular model of integral human placental tissues maintained in organ culture. Our promising findings encourage preclinical and clinical studies of artemisone as a new inhibitor against HCMV.

Ex Vivo anti-HIV Activity of Human Serum Obtained from Normal Volunteers Following Oral Administration of the HIV-1 Protease Inhibitor CGP 53437: Value as Predictor of Antiviral Efficacy

1997 ◽  
Vol 8 (1) ◽  
pp. 54-59
Author(s):  
JK Lazdins ◽  
JK Walker ◽  
RM Cozens ◽  
G Flesch ◽  
C Czendlik ◽  
...  
Keyword(s):  
Oral Administration ◽  
Protease Inhibitor ◽  
Clinical Studies ◽  
Ex Vivo ◽  
Human Lymphocytes ◽  
Normal Volunteers ◽  
Antiviral Efficacy ◽  
Anti Hiv ◽  
Hiv 1

The aim of the study was to determine whether the concentration of CGP 53437 measured in the sera of normal volunteers following oral administration of a single dose, had retained its anti-HIV activity; and whether such results could be of predictive value for future clinical antiviral efficacy studies. CGP 53437 is an inhibitor of HIV-1 protease that suppresses HIV-1 replication in human lymphocytes in vitro at 100 nM. The in vitro anti-HIV activity of human sera obtained from CGP 53437-treated individuals was compared with that of sera spiked with known concentrations of CGP 53437 (in the presence or absence of α-1 acid glycoprotein). It was found that the concentration of the compound measured in the sera from treated individuals provided the expected in vitro anti-HIV activity. These results not only validate our analytical method for detection of CGP 53437, but also support the notion that interaction of CGP 53437 with plasma proteins does not significantly affect its antiviral activity (shift of the ED90 by a factor of three). In conclusion, ex vivo anti-HIV activity determinations of sera containing an HIV protease inhibitor, in conjunction with the pharmacokinetic evaluation during Phase I clinical studies, can provide valuable information regarding the suitability of such inhibitors for further clinical studies.

scholarly journals Prevalence of Mutations in the pfcoronin Gene and Association with Ex Vivo Susceptibility to Common Quinoline Drugs against Plasmodium falciparum

Pharmaceutics ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1273
Author(s):  
Océane Delandre ◽  
Mathieu Gendrot ◽  
Isabelle Fonta ◽  
Joel Mosnier ◽  
Nicolas Benoit ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Sanger Sequencing ◽  
Ex Vivo ◽  
Central Africa ◽  
Artemisinin Resistance ◽  
Current Data ◽  
Artemisinin Derivative ◽  
African Countries ◽  
Low Prevalence

Background: Artemisinin-based combination therapy (ACT) was recommended to treat uncomplicated falciparum malaria. Unlike the situation in Asia where resistance to ACT has been reported, artemisinin resistance has not yet emerged in Africa. However, some rare failures with ACT or patients continuing to be parasitaemic on day 3 after ACT treatment have been reported in Africa or in travellers returning from Africa. Three mutations (G50E, R100K, and E107V) in the pfcoronin gene could be responsible for artemisinin resistance in Africa. Methods: The aims of this study were first to determine the prevalence of mutations in the pfcoronin gene in African P. falciparum isolates by Sanger sequencing, by targeting the 874 samples collected from patients hospitalised in France after returning from endemic areas in Africa between 2018 and 2019, and secondly to evaluate their association with in vitro reduced susceptibility to standard quinoline antimalarial drugs, including chloroquine, quinine, mefloquine, desethylamodiaquine, lumefantrine, piperaquine, and pyronaridine. Results: The three mutations in the pfcoronin gene (50E, 100K, and 107V) were not detected in the 874 P. falciparum isolates. Current data show that another polymorphism (P76S) is present in many countries of West Africa (mean prevalence of 20.7%) and Central Africa (11.9%) and, rarely, in East Africa (4.2%). This mutation does not appear to be predictive of in vitro reduced susceptibility to quinolines, including artemisinin derivative partners in ACT such as amodiaquine, lumefantrine, piperaquine, pyronaridine, and mefloquine. Another mutation (V62M) was identified at low prevalence (overall prevalence of 1%). Conclusions: The 76S mutation is present in many African countries with a prevalence above 10%. It is reassuring that this mutation does not confer in vitro resistance to ACT partners.

scholarly journals Human Cytomegalovirus Replication and Infection-Induced Syncytia Formation in Labial, Foreskin, and Fetal Lung Fibroblasts

Viruses ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2355
Author(s):  
Alexis Aguiar ◽  
Melissa Galinato ◽  
Maite’ Bradley Silva ◽  
Bryant Toth ◽  
Michael A. McVoy ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Human Cytomegalovirus ◽  
Fetal Lung ◽  
Lung Fibroblasts ◽  
Anatomical Site ◽  
Cmv Infection ◽  
Viral Spread ◽  
Syncytia Formation

Only a handful of cell types, including fibroblasts, epithelial, and endothelial cells, can support human cytomegalovirus (CMV) replication in vitro, in striking contrast to the situation in vivo. While the susceptibility of epithelial and endothelial cells to CMV infection is strongly modulated by their anatomical site of origin, multiple CMV strains have been successfully isolated and propagated on fibroblasts derived from different organs. As oral mucosal cells are likely involved in CMV acquisition, we sought to evaluate the ability of infant labial fibroblasts to support CMV replication, compared to that of commonly used foreskin and fetal lung fibroblasts. No differences were found in the proportion of cells initiating infection, or in the amounts of viral progeny produced after exposure to the fibroblast-adapted CMV strain AD169 or to the endothelial cell-adapted strain TB40/E. Syncytia formation was, however, significantly enhanced in infected labial and lung fibroblasts compared to foreskin-derived cells, and did not occur after infection with AD169. Together, these data indicate that fibroblast populations derived from different tissues are uniformly permissive to CMV infection but retain phenotypic differences of potential importance for infection-induced cell–cell fusion, and ensuing viral spread and pathogenesis in different organs.

scholarly journals Silver Nanoparticles Alter Cell Viability Ex Vivo and in Vitro and Induce Proinflammatory Effects in Human Lung Fibroblasts

Nanomaterials ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1868
Author(s):  
Anna Löfdahl ◽  
Andreas Jern ◽  
Samuel Flyman ◽  
Monica Kåredal ◽  
Hanna L Karlsson ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Cell Viability ◽  
Human Lung ◽  
Inflammatory Responses ◽  
Ex Vivo ◽  
Lung Fibroblasts ◽  
Human Lung Fibroblasts ◽  
Metabolic Activities ◽  
Precision Cut Lung Slices

Silver nanoparticles (AgNPs) are commonly used in commercial and medical applications. However, AgNPs may induce toxicity, extracellular matrix (ECM) changes and inflammatory responses. Fibroblasts are key players in remodeling processes and major producers of the ECM. The aims of this study were to explore the effect of AgNPs on cell viability, both ex vivo in murine precision cut lung slices (PCLS) and in vitro in human lung fibroblasts (HFL-1), and immunomodulatory responses in fibroblasts. PCLS and HFL-1 were exposed to AgNPs with different sizes, 10 nm and 75 nm, at concentrations 2 µg/mL and 10 μg/mL. Changes in synthesis of ECM proteins, growth factors and cytokines were analyzed in HFL-1. Ag10 and Ag75 affected cell viability, with significantly reduced metabolic activities at 10 μg/mL in both PCLS and HFL-1 after 48 h. AgNPs significantly increased procollagen I synthesis and release of IL-8, prostaglandin E2, RANTES and eotaxin, whereas reduced IL-6 release was observed in HFL-1 after 72 h. Our data indicate toxic effects of AgNP exposure on cell viability ex vivo and in vitro with altered procollagen and proinflammatory cytokine secretion in fibroblasts over time. Hence, careful characterizations of AgNPs are of importance, and future studies should include timepoints beyond 24 h.

scholarly journals Antigen sensitivity is a major determinant of CD8+ T-cell polyfunctionality and HIV-suppressive activity

Blood ◽  
2009 ◽  
Vol 113 (25) ◽  
pp. 6351-6360 ◽  
Author(s):  
Jorge R. Almeida ◽  
Delphine Sauce ◽  
David A. Price ◽  
Laura Papagno ◽  
So Youn Shin ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Ex Vivo ◽  
Cd8 T Cell ◽  
Basic Knowledge ◽  
Suppressive Activity ◽  
Antiviral Efficacy ◽  
Hiv 1

Abstract CD8+ T cells are major players in the immune response against HIV. However, recent failures in the development of T cell–based vaccines against HIV-1 have emphasized the need to reassess our basic knowledge of T cell–mediated efficacy. CD8+ T cells from HIV-1–infected patients with slow disease progression exhibit potent polyfunctionality and HIV-suppressive activity, yet the factors that unify these properties are incompletely understood. We performed a detailed study of the interplay between T-cell functional attributes using a bank of HIV-specific CD8+ T-cell clones isolated in vitro; this approach enabled us to overcome inherent difficulties related to the in vivo heterogeneity of T-cell populations and address the underlying determinants that synthesize the qualities required for antiviral efficacy. Conclusions were supported by ex vivo analysis of HIV-specific CD8+ T cells from infected donors. We report that attributes of CD8+ T-cell efficacy against HIV are linked at the level of antigen sensitivity. Highly sensitive CD8+ T cells display polyfunctional profiles and potent HIV-suppressive activity. These data provide new insights into the mechanisms underlying CD8+ T-cell efficacy against HIV, and indicate that vaccine strategies should focus on the induction of HIV-specific T cells with high levels of antigen sensitivity to elicit potent antiviral efficacy.

scholarly journals Saracatinib, a Selective Src Kinase Inhibitor, Blocks Fibrotic Responses in In Vitro, In Vivo and Ex Vivo Models of Pulmonary Fibrosis

2022 ◽  
Author(s):  
Farida Ahangari ◽  
Christine Becker ◽  
Daniel G Foster ◽  
Maurizio Chioccioli ◽  
Meghan Nelson ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Kinase Inhibitor ◽  
Recombinant Adenovirus ◽  
Ex Vivo ◽  
Src Kinase ◽  
Lung Fibroblasts

Idiopathic Pulmonary Fibrosis (IPF) is a chronic, progressive, and often fatal disorder. Two FDA approved anti-fibrotic drugs, nintedanib and pirfenidone, slow the rate of decline in lung function, but responses are variable and side effects are common. Using an in-silico data-driven approach, we identified a robust connection between the transcriptomic perturbations in IPF disease and those induced by saracatinib, a selective Src kinase inhibitor, originally developed for oncological indications. Based on these observations, we hypothesized that saracatinib would be effective at attenuating pulmonary fibrosis. We investigated the anti-fibrotic efficacy of saracatinib relative to nintedanib and pirfenidone in three preclinical models: (i) in vitro in normal human lung fibroblasts (NHLFs); (ii) in vivo in bleomycin and recombinant adenovirus transforming growth factor-beta (Ad-TGF-β) murine models of pulmonary fibrosis; and (iii) ex vivo in precision cut lung slices from these mouse models. In each model, the effectiveness of saracatinib in blocking fibrogenic responses was equal or superior to nintedanib and pirfenidone.

CMV Specific Immunotherapy: How Many Patients Will Benefit?.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2244-2244
Author(s):  
Russell J. Garland ◽  
Paraskevi Diamanti ◽  
Samantha E. West ◽  
Ann Green ◽  
David Carrington ◽  
...  
Keyword(s):  
Stem Cell ◽  
Ex Vivo ◽  
Antiviral Treatment ◽  
Antiviral Drug ◽  
Disease Free Survival ◽  
Group B ◽  
Selection Factors ◽  
Cmv Reactivation

Abstract Cytomegalovirus (CMV) infection is associated with increased transplant related mortality and decreased overall survival after stem cell transplantation (SCT) despite major advances in early detection, prophylaxis and treatment. This effect is most marked in CMV seropositive patients who have a statistically significant decrement in overall or disease free survival of 20–46% when compared to low risk (−/−) transplants (Boeckh Blood 2004 103: 2003–8). Transfer of cellular immunity from a seropositive donor results in reconstitution of T cells to CMV and protection from CMV disease post-SCT, and this underlies the recent development of cellular immunotherapeutic manoeuvres. As a centre looking to develop such therapy, we have carried out a retrospective analysis of a patient cohort in order to estimate how many patients would require this intervention. We used twice weekly quantitative PCR surveillance to analyse the CMV reactivation profiles of 104 recipients of 106 allogeneic SCT, transplanted between April 2002 and April 2004. Hence follow up was 5–27 months. The cohort comprised 41 adults and 65 children. Transplants were from related (42), unrelated (52) and haploidentical donors (12). T cell depletion was performed either (i) in vivo using either CAMPATH-1H (44 transplants) or ATG (5), (ii) ex vivo by either CD34 (8) or CAMPATH-1H in vitro (1), or (iii) by combinations of (i) and (ii) (26). SCTs were for malignancies in 92 cases and non-neoplastic disorders in 14. Many patients were considered high risk, necessitating short search to transplant times. A hierarchy of donor selection factors was considered: HLA matching was the primary determinant, with other factors including stem cell dose, age, gender and CMV status. Seventy-one patients were at risk of CMV reactivation. There were 28 episodes of CMV reactivation in 27 of these patients, with the following Recipient/Donor serostatus combinations: 0 (of 16) −/+, 22 (of 38) +/− and 6 (of 17) +/+. Seven patients (Group A) resolved their infections without intervention. Nine patients (Group B) resolved infections after antiviral drug treatment, whilst the remaining 12 reactivators (Group C) did not clear their CMV DNA load despite antiviral treatment (of whom 3 died, 1 relapsed and the remainder have ongoing CMV PCR positivity). As a minimum a CMV immunotherapy programme should allow prophylaxis or early treatment of all patients in groups B and C. However, using our current selection criteria only 5/21 cases had a CMV seropositive donor. Logistical problems e.g. CTL precursor frequency or availability of an immunodominant tetramer might have rendered some donors inappropriate providers of anti-CMV CTL. Thus a maximum of 5 out of 106 transplanted patients in our unit could have benefited, although this figure could be slightly improved by deliberate selection of CMV positive donors for CMV positive patients. Such numbers should be borne in mind by any centre contemplating the development of antiviral immunotherapy programmes.

scholarly journals Pentoxifylline Promotes Replication of Human Cytomegalovirus In Vivo and In Vitro

Blood ◽  
1997 ◽  
Vol 89 (10) ◽  
pp. 3682-3690 ◽  
Author(s):  
Kerstin Staak ◽  
Susanna Prösch ◽  
Joachim Stein ◽  
Christina Priemer ◽  
Ralf Ewert ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Human Cytomegalovirus ◽  
Peak Plasma ◽  
Immediate Early ◽  
Lung Fibroblasts ◽  
Sequence Motif ◽  
Peak Plasma Level ◽  
Hcmv Disease

Abstract OKT3 monoclonal antibody (MoAb) therapy is well established in the prevention and therapy of acute rejection in transplant patients. Unfortunately, this therapy is associated with several short-term (cytokine release syndrome) and long-term (infections, EBV-related lymphoma) side effects. Recently, we were able to demonstrate an association between the TNFα release following the first OKT3 MoAb infusions and the appearance of human cytomegalovirus (HCMV) reactivation several days later. In order to prevent this TNFα associated HCMV reactivation patients were additionally treated with pentoxifylline (PTX), a methylxanthine derivative that has been shown to suppress TNFα induction. Although the TNFα peak plasma level following OKT3 MoAb treatment was markedly reduced, the incidence of HCMV reactivation and HCMV disease was not influenced. In transient transfection experiments using HCMV immediate early enhancer/promoter CAT reporter gene constructs PTX enhanced the promoter activity independently from TNFα in premonocytic cells. Furthermore, PTX acted synergistically with TNFα. In virus-infected human embryonal lung fibroblasts HCMV replication was triggered in the presence of both PTX and TNFα, while either substance alone had only marginal effects. The stimulatory effect of PTX on the immediate early (IE) enhancer/promoter was mediated via CREB/ATF, a eukaryotic transcription factor that binds to the 19 bp sequence motif in the enhancer region, while TNFα stimulation was mediated by activation of the transcription factor NF-kB and its binding to the 18 bp sequence motif in the enhancer. These data suggest a potential side effect of cAMP-elevating drugs such as PTX.

scholarly journals Human Cytomegalovirus Selectively Suppresses the Megakaryo/Thrombopoiesis of PDGFR+ and αvβ3+ Megakaryocytes Via the TPO/c-Mpl Pathway after Allo-HSCT

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 25-25
Author(s):  
Feng-qi Liu ◽  
Fei-er Feng ◽  
Gao-chao Zhang ◽  
Yan Su ◽  
Xue-yan Sun ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Human Cytomegalovirus ◽  
Hcmv Infection ◽  
Conflicts Of Interest ◽  
Antiviral Treatment ◽  
Infection Model ◽  
Hematopoietic Stem ◽  
The Impact

Introduction Virus-induced thrombocytopenia is a severe complication in immunocompromised hosts. Among patients following allogeneic hematopoietic stem cell transplantation (allo-HSCT), human cytomegalovirus (HCMV) infection contributes to a variety of end-organ diseases and hematological complications, leading to increased mortality. Even with antiviral treatment, HCMV remains a potentially lethal infection due to the lack of understanding of the underlying mechanisms of host-virus interactions. The key to solving this problem is to identify the factors that predispose patients to HCMV infection and carry out targeted therapy. Here, we investigated the megakaryo/thrombopoiesis process, including the thrombopoietin (TPO)/c-Mpl pathway, after HCMV infection in vivo and in vitro, screened for susceptible subsets of megakaryocytes (MKs) and explored novel therapeutic targets for HCMV infection. Methods To test whether thrombocytopenia induced by HCMV results from an impaired megakaryo/thrombopoiesis process, we studied the impact of HCMV in an in vivo model of HCMV DNAemia patients following allo-HSCT and an in vitro model of bone marrow CD34+-derived MKs infected with serum from HCMV DNAemia patients. Forty patients who had received allo-HSCT were enrolled in this study, among whom 18 recipients had HCMV DNAemia and 22 were HCMV negative, and bone marrow-derived mononuclear cells (MNCs) from patients were tested for CD41, vWF, pp65, c-Mpl, PDGFR, αvβ3 and TLR2 using flow cytometry (FCM). Transmission electron microscopy (TEM) was used to detect HCMV capsids inside MKs. Cell apoptosis was measured by Annexin V. MK ploidy was determined by FCM for propidium iodide (PI) staining. Finally, inhibitors of PDGFR (IMC-3G3 and Gleevec), αvβ3 and TLR2 were cocultured with MKs. Results Our data showed that pp65+ cells accounted for 40.59±6.12% of total CD41+vWF+ MKs from HCMV DNAemia patients, and there was a significant increase in the expression of αvβ3, PDGFR and TLR2 in pp65+ MKs compared with that in control patients. Furthermore, the percentage of PDGFR+αvβ3+ MKs emerged as an independent factor associated with HCMV infection in multivariate analysis (p = 0.008). MKs in HCMV-infected patients showed increased apoptosis and necrosis and different patterns of MK ploidy distribution compared with those in HCMV-negative patients, with a decreased proportion from 16N to 64N and a peak at 8N. Meanwhile, the expression of TPO receptor c-Mpl was lower in pp65+ MKs from HCMV DNAemia patients (0.77±0.38% in pp65+ MKs from HCMV DNAemia patients, 1.75±0.40% in pp65- MKs from HCMV DNAemia patients, 1.97±0.67% in MKs from HCMV-negative patients, and 2.06±0.29% in MKs from healthy controls, p<0.01) while the TPO level in serum was increased compared with that in controls. Next, we established an in vitro HCMV infection model of CD34+-derived MKs with serum from HCMV DNAemia patients, and the laboratory HCMV strain Towne was used as a positive control. After 9 days of coculturing, the viral capsids of HCMV were observed in the nuclei of MKs (Figure 1A), and HCMV infection increased the apoptosis of MKs and shifted them to low ploidy, with a significant decrease in platelet release. As with the in vivo results, c-Mpl was downregulated in HCMV-infected MKs. The expression levels of PDGFR, TLR2 and αvβ3 on MKs were increased in coculture with HCMV DNAemia serum, and pp65-positive MKs were decreased compared with the control after treatment with inhibitors of PDGFR and αvβ3 (Figure 1B). However, neither Gleevec nor anti-TLR2 altered the HCMV infection rate. Conclusions Our study showed that HCMV could impair megakaryopoiesis throughout maturation, apoptosis, and platelet generation via the TPO/c-Mpl pathway both in vivo and in vitro. MKs with PDGFR+ and αvβ3+ phenotypes are susceptible to HCMV infection and we proposed PDGFR and αvβ3 inhibitors as potential therapeutic alternatives for allo-HSCT patients with HCMV infection. Disclosures No relevant conflicts of interest to declare.

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