| 1. |
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| 2. |
- de Witte, H H, et al.
(författare)
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Prognostic significance of TP53 accumulation in human primary breast cancer : comparison between a rapid quantitative immunoassay and SSCP analysis.
- 1996
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Ingår i: International Journal of Cancer. - 0020-7136 .- 1097-0215. ; 69:2, s. 125-130
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Tidskriftsartikel (refereegranskat)abstract
- TP53 accumulation in human primary breast carcinomas was studied by a quantitative luminometric immunoassay (LIA), and TP53 gene alterations, exons 5-8, were examined by single-strand conformation polymorphism (SSCP) analysis. In 48 of 142 breast tumor samples, a TP53 gene alteration was identified. In tumor samples without a TP53 gene alteration, the median cytosolic TP53 protein level, as determined by LIA, was 0.4 ng/mg protein (range 0-70.8 ng/mg protein), whereas the median TP53 protein level for tumor samples with a TP53 gene alteration was 10 times higher, i.e., 4.1 ng/mg protein (range 0.1-176.0 ng/mg protein). Despite a significant correlation between the outcome of LIA and SSCP, a disagreement was found in 22% of cases analyzed. Significant correlations were found between TP53 protein accumulation and low estrogen receptor content, and with a shorter relapse-free as well as overall survival, with a median duration of follow-up of 100 months. Due to its rapid and easy performance on routinely prepared cytosols, the LIA for TP53 protein may be useful in evaluating the prognostic impact of TP53 protein accumulation in human primary breast cancer.
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| 3. |
- Bartlett, Sofia R., et al.
(författare)
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Sequencing of Hepatitis C Virus for Detection of Resistance to Direct-Acting Antiviral Therapy : A Systematic Review
- 2017
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Ingår i: HEPATOLOGY COMMUNICATIONS. - : JOHN WILEY & SONS LTD. - 2471-254X. ; 1:5, s. 379-390
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Forskningsöversikt (refereegranskat)abstract
- The significance of the clinical impact of direct-acting antiviral (DAA) resistance-associated substitutions (RASs) in hepatitis C virus (HCV) on treatment failure is unclear. No standardized methods or guidelines for detection of DAA RASs in HCV exist. To facilitate further evaluations of the impact of DAA RASs in HCV, we conducted a systematic review of RAS sequencing protocols, compiled a comprehensive public library of sequencing primers, and provided expert guidance on the most appropriate methods to screen and identify RASs. The development of standardized RAS sequencing protocols is complicated due to a high genetic variability and the need for genotype- and subtype-specific protocols for multiple regions. We have identified several limitations of the available methods and have highlighted areas requiring further research and development. The development, validation, and sharing of standardized methods for all genotypes and subtypes should be a priority.
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| 7. |
- Luttens, Andreas, et al.
(författare)
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Ultralarge Virtual Screening Identifies SARS-CoV-2 Main Protease Inhibitors with Broad-Spectrum Activity against Coronaviruses
- 2022
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 144:7, s. 2905-2920
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Tidskriftsartikel (refereegranskat)abstract
- Drugs targeting SARS-CoV-2 could have saved millions of lives during the COVID-19 pandemic, and it is now crucial to develop inhibitors of coronavirus replication in preparation for future outbreaks. We explored two virtual screening strategies to find inhibitors of the SARS-CoV-2 main protease in ultralarge chemical libraries. First, structure-based docking was used to screen a diverse library of 235 million virtual compounds against the active site. One hundred top-ranked compounds were tested in binding and enzymatic assays. Second, a fragment discovered by crystallographic screening was optimized guided by docking of millions of elaborated molecules and experimental testing of 93 compounds. Three inhibitors were identified in the first library screen, and five of the selected fragment elaborations showed inhibitory effects. Crystal structures of target-inhibitor complexes confirmed docking predictions and guided hit-to-lead optimization, resulting in a noncovalent main protease inhibitor with nanomolar affinity, a promising in vitro pharmacokinetic profile, and broad-spectrum antiviral effect in infected cells.
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| 8. |
- Schinazi, Raymond F., et al.
(författare)
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Approaches for the development of antiviral compounds : the case of hepatitis C virus
- 2009
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Ingår i: Antiviral Strategies. - Berlin, Heidelberg : Springer Berlin/Heidelberg. ; , s. 25-51
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Bokkapitel (refereegranskat)abstract
- Traditional methods for general drug discovery typically include evaluating random compound libraries for activity in relevant cell-free or cell-based assays. Success in antiviral development has emerged from the discovery of more focused libraries that provide clues about structure activity relationships. Combining these with more recent approaches including structural biology and computational modeling can work efficiently to hasten discovery of active molecules, but that is not enough. There are issues related to biology, toxicology, pharmacology, and metabolism that have to be addressed before a hit compound becomes nominated for clinical development. The objective of gaining early preclinical knowledge is to reduce the risk of failure in Phases 1, 2, and 3, leading to the goal of approved drugs that benefit the infected individual. This review uses hepatitis C virus (HCV), for which we still do not have an ideal therapeutic modality, as an example of the multidisciplinary efforts needed to discover new antiviral drugs for the benefit of humanity.
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| 9. |
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| 10. |
- Borg, A, et al.
(författare)
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Prognostic significance of p53 overexpression in primary breast cancer : a novel luminometric immunoassay applicable on steroid receptor cytosols.
- 1995
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Ingår i: British Journal of Cancer. - : Springer Science and Business Media LLC. - 0007-0920 .- 1532-1827. ; 71:5, s. 1013-1017
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Tidskriftsartikel (refereegranskat)abstract
- A novel quantitative luminometric immunoassay (LIA) has been developed for the measurement of wild-type and mutant p53 protein in extracts from breast tumour tissue. The LIA was found to yield reliable estimates of p53 expression in cytosol samples routinely prepared for steroid receptor analysis as compared with results obtained with immunohistochemical analysis. The LIA was evaluated on 205 primary breast tumour cytosols prepared for steroid receptor analysis and stored frozen at -80 degrees C for 6-8 years, p53 protein being detected in 65% of the samples (range 0.01-23 ng mg-1 protein). Using an arbitrary cut-off value of 0.15 ng mg-1 protein, 30% of the tumours were classified as manifesting p53 overexpression. Significant and independent correlations were found to exist between p53 overexpression and shorter disease-free (P < 0.001) and overall survival (P = 0.039) at a median duration of follow-up of 50 months. p53 overexpression was related to low oestrogen receptor content and high proliferation rate (S-phase fraction). No relationship was found to tumour size or the presence of lymph node metastasis. Three tumours possessed an extremely high p53 content (> 10 ng mg-1 protein), all of which were of medullary or high-grade ductal type, oestrogen and progesterone receptor negative, DNA non-diploid, had S-phase fractions of > 22% and recurred within 1-2 years. In summary, a new sensitive and quantitative LIA suitable for routine analysis of p53 protein in steroid receptor cytosol preparations from breast tumours has been developed to confirm the prognostic importance of p53 protein accumulation in human breast cancer.
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| 11. |
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| 12. |
- Gronowitz, J S, et al.
(författare)
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Carrier bound templates for single tube reverse transcriptase assays and for combined purification and activity analyses, with special reference to HIV
- 1991
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Ingår i: Biotechnology and applied biochemistry. - 0885-4513 .- 1470-8744. ; 13:1, s. 127-142
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Tidskriftsartikel (refereegranskat)abstract
- Polyriboadenosine (prA) was coupled to polycarbonate macrobeads or magnetic beads. The efficiency of the beads and of prA-Sepharose, after priming with odT, as templates in activity assays of purified AMV- and HIV-reverse transcriptase (RT), using [125I]iododeoxyuridine-triphosphate as substrate, was studied. Although the use of immobilized templates, compared with soluble template, resulted in a decreased total molar turnover, it did not affect the sensitivity of the assay for detecting RT. The utility of the new assay was analyzed by mixing purified AMV- or HIV-Rt with different dilutions of the untreated clinical specimen. This showed that RT activity was unaffected by 100 microliters of an extract of whole blood cells resuspended to their original blood volume and diluted 1/64, and also by 100 microliters of serum diluted 1/64. To improve the utility of the assay at the inhibitory concentrations of clinical specimens, the following procedure was adopted: the sample to be analyzed was incubated with the carrier bound template in order to allow the RT to bind, the carrier was washed to remove inhibitory factors, and the reaction components were then added to determine the amount of bound RT. This procedure greatly enhanced the recovery of RT activity from crude specimens and made the direct detection of HIV-RT possible. The assay is easily automated and useful for RT determination in multiple samples and for determining RT-inhibiting substances such as substrate analogs and antibodies.
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| 13. |
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| 14. |
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| 15. |
- Imamura, K, et al.
(författare)
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Reduced activity in the extrastriate visual cortex of individuals with strabismic amblyopia
- 1997
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Ingår i: NEUROSCIENCE LETTERS. - : ELSEVIER SCI IRELAND LTD. - 0304-3940. ; 225:3, s. 173-176
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- In order to examine the relationship between reduced visual acuity in human strabismic amblyopia and the cortical activation pattern, we studied, by use of positron emission tomography (PET) and the (H2O)-O-15 bolus technique, changes in the regional cer
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| 16. |
- Larkin, Cormac J.K., et al.
(författare)
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M5 — Mars Magnetospheric Multipoint Measurement Mission: A multi-spacecraft plasma physics mission to Mars
- 2024
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Ingår i: Advances in Space Research. - : Elsevier. - 0273-1177 .- 1879-1948. ; 73:6, s. 3235-3255
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Tidskriftsartikel (refereegranskat)abstract
- Mars, lacking an intrinsic dynamo, is an ideal laboratory to comparatively study induced magnetospheres, which can be found in other terrestrial bodies as well as comets. Additionally, Mars is of particular interest to further exploration due to its loss of habitability by atmospheric escape and possible future human exploration. In this context, we propose the Mars Magnetospheric Multipoint Measurement Mission (M5), a multi-spacecraft mission to study the dynamics and energy transport of the Martian induced magnetosphere comprehensively. Particular focus is dedicated to the largely unexplored magnetotail region, where signatures of magnetic reconnection have been found. Furthermore, a reliable knowledge of the upstream solar wind conditions is needed to study the dynamics of the Martian magnetosphere, especially the different dayside boundary regions but also for energy transport phenomena like the current system and plasma waves. This will aid the study of atmospheric escape processes of planets with induced magnetospheres. In order to resolve the three-dimensional structures varying both in time and space, multi-point measurements are required. Thus, M5 is a five spacecraft mission, with one solar wind monitor orbiting Mars in a circular orbit at 5 Martian radii, and four smaller spacecraft in a tetrahedral configuration orbiting Mars in an elliptical orbit, spanning the far magnetotail up to 6 Mars radii with a periapsis just outside the Martian magnetosphere of 1.8 Mars radii. We not only present a detailed assessment of the scientific need for such a mission but also show the resulting mission and spacecraft design taking into account all aspects of the mission requirements and constraints such as mass, power, and link budgets. Additionally, different aspects of the mission programmatics like a possible mission timeline, cost estimates, or public outreach are shown. The common requirements for acceptance for an ESA mission are considered. The mission outlined in this paper was developed during the Alpbach Summer School 2022 on the topic of “Comparative Plasma Physics in the Universe”.
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| 17. |
- Lennerstrand, Johan, et al.
(författare)
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A Method for Combined Immunoaffinity Purification and Assay of HIV-1 Reverse Transcriptase Activity Useful for Crude Samples
- 1996
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Ingår i: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 235:2, s. 141-152
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Tidskriftsartikel (refereegranskat)abstract
- Detection of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) activity in crude specimens was greatly enhanced using a novel capture RT assay. Eighteen different monoclonal antibodies (Mabs) raised against purified HIV-1 RT were tested for their ability to bind to HIV-1 RT without affecting its activity. The anti-HIV-1 RT Mabs were immobilized on plastic macrobeads and used as solid carriers in the capture RT assay. The assay system first involved RT's adherence to the immobilized Mabs. Nonspecific enzymes and other impurities were removed by a simple wash after which the RT reaction mixture was added. Substrate and product were finally separated by a wash of the beads. Practically all radioactivity incorporated into DNA (>98%) was recovered on the bead. The Michaelis-Menten constants and the saturation velocity values for the nucleotide substrate were similar for free and immobilized RT. The reaction mechanism for the immobilized RT is discussed. When comparing the function of this assay with more conventional soluble RT assays for samples consisting of recombinant HIV-1 RT mixed with an extract of peripheral blood lymphocytes (PBL), an almost 100-fold higher sensitivity was found. The capture RT assay had the capacity to recover approximately 80% of the RT activity added to an extract of 1 x 10(7) PBL cells/ ml. A strong correlation (r = 0.947) between the results obtained with this assay and a HIV-1 p24 enzyme-linked immunosorbent assay was found, when samples from a collection of 16 HIV strains propagated in cell culture were analyzed.
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| 18. |
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| 19. |
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| 20. |
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| 21. |
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| 22. |
- Linderholm, BK, et al.
(författare)
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The expression of vascular endothelial growth factor correlates with mutant p53 and poor prognosis in human breast cancer
- 2001
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Ingår i: Cancer Research. - 0008-5472 .- 1538-7445. ; 61:5, s. 2256-2260
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Tidskriftsartikel (refereegranskat)abstract
- Wild-type p53 protein has been shown to inhibit angiogenesis through thrombospondin in thepreclinical setting, Here, we determined the associations between the expression of the angiogenic factorvascular endothelial growth factor (VEGF) and the p53 status, including different mutation sites and types,in primary breast cancer. Cytosols from 224 primary breast cancer patients were analyzed with an enzyme immunoassay for determination of human VEGF(165) protein content. p53 status was determined hv cDNA-based sequencing of the entire coding region, by immunohistochemistry (IHC). and by a p53luminometric immunoassay (I,IA) method. Statistically significant associations was found between higher VEGF content and non-wild-type p53 status for all methods; sequence-based data (P = 0.0019), IHC data (P = 0.0068), and the I,IA method (r = 0.427: P > 0.001), Highest VEGF values were detected in tumorswith p53 insertions, deletions, and stop codon mutations (P = 0.0043). Combining p53 status and VEGF content resulted in additional prognostic information, relapse-free survival (RFS: P = 0.0377), overall survival (OS; P = 0.0319), and breast cancer corrected survival (BCCS: P = 0.0292). In multivariate analysis, the relative hazard increased when the VEGF data were added to the p53 status, with a relative hazard of 1.7 for RFS and 3.0 for BCCS, compared with 1.1 fur RFS and 1.4 for BCCS among the patientswith either high VEGF content or p53 mutation. Higher VEGF content was statistically significantly correlated with a worse outcome fur patients with estrogen receptor-positive tumors receiving adjuvant tamoxifen: RFS (P = 0.0471), OS (P = 0.0134), BCCS (P = 0.0064), as well as in multivariate analysis withpoint estimates of 3.4 and 2.1 fur BCCS and RFS, respectively, VEGF expression is related to p53 statusin human breast cancer patients. Combining VEGF with p53 status resulted in better prognostic prediction.
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| 23. |
- Neumüller, M, et al.
(författare)
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HIV-1 reverse transcriptase inhibiting antibody titer in serum : relation to disease progression and to core-antibody levels.
- 1992
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Ingår i: Journal of Medical Virology. - : Wiley. - 0146-6615 .- 1096-9071. ; 36:4, s. 283-291
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Tidskriftsartikel (refereegranskat)abstract
- A new assay for detecting inhibition of reverse transcriptase activity (the RT-i REA) was developed. This assay was standardized for screening serum samples for reverse transcriptase inhibiting antibodies (RT-iAb). High specificity (100%) and sensitivity (greater than 98%) were achieved with samples from HIV-negative individuals and HIV-infected individuals. The RT-i REA was also used in a study of the titers of RT-iAb in serum samples obtained from 33 HIV-infected homosexual men. The results confirmed the relation between decreasing RT-iAb levels and progression to late stages of the disease. Furthermore, a falling RT-iAb titer was observed in 14 of 15 individuals experiencing periods of severe clinical symptoms attributed to HIV-activity. In 7 of the patients the decline in RT-iAb titer began prior to severe clinical symptoms. The fall in RT-iAb titer also correlated with a reduction in core Ab level. The core Ab level has previously been reported to be a disease progression marker with considerable prognostic value. However, whereas all patients were positive for RT-iAb, 8 of the 33 patients did not have detectable core Ab. The use of RT-iAb titer as a marker of disease progression is discussed.
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| 24. |
- Neumüller, Magnus, et al.
(författare)
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HIV reverse transcriptase inhibiting antibodies detected by a new technique : relation to p24 and gp41 antibodies, HIV antigenemia and clinical variables.
- 1991
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Ingår i: Journal of Medical Virology. - : Wiley. - 0146-6615 .- 1096-9071. ; 34:1, s. 55-63
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Tidskriftsartikel (refereegranskat)abstract
- A new assay for HIV reverse transcriptase activity inhibiting antibodies (RTI-ab) was used for the analysis of a large collection of sera sampled before and after confirmation of HIV infection. In this assay HIV-RT was preincubated with diluted serum, after which residual RT activity was determined by a technique using a template coupled to macrobeads and 125I-lodo-deoxyuridine-triphosphate as the tracer-substrate. Of the 936 sera analysed, 818 were found positive for RTI-ab, and 824 were positive in Western blot (Wb). The prevalence of RTI-ab compared to Wb was therefore 99.3%. The corresponding figure for 930 sera analysed for envelope-ab, i.e., gp41-ab, was 823 positive, and of these 930 sera 815 were Wb positive, giving a comparative prevalence of 101%. In contrast, only 678 samples of 993 analyzed for core ab, i.e., p24, were positive, giving a prevalence of 77.0% as 880 of these samples were Wb positive. Thus, RTI-ab was as prevalent as gp41-ab, and although the analyses of RTI-ab amounts in different stages showed decreasing levels in stage IV compared to stages II or III, all of the sera except 1 were found positive in stages III and IV. Further, it was found that both the few RTI-ab negative samples in stage II and the few RTI-ab positive samples among Wb negative sera were sampled in connection with seroconversion. The specificity of the RTI-ab assay was 100% in a test of 200 serum samples from HIV negative blood donors. It was concluded that RTI-ab analyses can be made highly sensitive and specific and useful for studies of HIV infection.
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| 29. |
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| 31. |
- Shao, X-W, et al.
(författare)
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Application of a colorimetric chain-termination assay for characterization of reverse transcriptase from 3-azido-2,3-deoxythymidine-resistant HIV isolates
- 2002
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Ingår i: Biotechnology and applied biochemistry. - 0885-4513 .- 1470-8744. ; 35, s. 155-164
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Tidskriftsartikel (refereegranskat)abstract
- wo different enzyme assays, both based on the interaction of native reverse transcriptase(IRT) and 3'-azido-2',3'-deoxythymidine triphosphate (AZT-TP), were used to characterize the enzymesfrom 18 HIV-I isolates with decreased sensitivity to AZT in cell culture. The first assay, which measures the balance between incorporation and excision of AZT monophosphate in the presence of dNTP substrate (in terms of IC50), gave an approx. 9-fold variation in sensitivity to AZT-TP. There was a correlation between the IC50 values and the sensitivity of the corresponding virus to AZT in cell culture (r = 0.60, P < 0.01). The second assay, which was designed specifically for measurement of chain termination in the absence of dNTP substrate (as the concentration of AZT-TP giving 50% residual primer function, or CT50), revealed a more than 600-fold difference between the different isolate RTs. For the majority ofenzymes there was a strict correlation between the results from the two assays; however, four isolatesexhibited significantly higher CT50/IC50 ratios than the other isolates. These differences were not related to sensitivity of the corresponding viruses to AZT but to the occurrence of certain mutations in their pol gene. The four deviating isolates contained either a minimum of four AZT-specific substitutions, including Thr-215 --> Tyr (isolates 134 and 143), or some of the known specific substitutions combined with Thr-39 --> Ala (isolates 80 and 157). The Thr-39 Ala substitution has previously been recorded in connection with AZT/Foscarnet combination therapy.
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| 32. |
- Shao, Xing-Wu, et al.
(författare)
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Use of HIV-1 reverse transcriptase recovered from human plasma for phenotypic drug susceptibility testing
- 2003
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Ingår i: AIDS. ; 17, s. 1463-1471
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Tidskriftsartikel (refereegranskat)abstract
- Objective: To demonstrate the use of HIV-1 reverse transcriplase (RT) recovered directly fromplasma for phenotypic drug susceptibility testing. Methods: Plasma from HIV-1 infected individuals with and without drug resistance-associated mutations were selected for the study. The blind coded plasmas were treated to inactivate cellular enzymes. The virions were immobilized on a gel and washed to remove antiretroviral drugs and RT activity blocking antibodies. The immobilized virions were lysed; the viral RTeluted and quantified, all according to the ExaVir Load procedure. The drug sensitivity profiles of each RT were determined using serially diluted drugs and modified Cavidi HS Lenti RT kits. Results: The phenotypic drug sensitivity profiles of the RT and the patterns of drug resistance mutations were highly concordant. Plasma RT from virions devoid of mutations associated with drug resistance had average 50% inhibitory concentrations (IC50) of 1.5 +/- 0.93 muM for nevirapine, 0.21 +/- 0.099 muM forefavirenz, 7.1 +/- 3.2 muM for delavirdine, 0.42 +/- 0.15 muM for azidothymidine triphosphate and 0.059 +/- 0.018 muM for didehydrothymidine triphosphate. The increase in IC50 value for RT with drugresistance associated substitutions was from 3- to more than 65-fold for non-nucleoside inhibitors and between 2- and 30-fold for thymidine analogue drugs. Conclusion: RT derived from virions recovered from the plasma of HIV infected individuals can be used foranalysis of phenotypic drug susceptibility. The methods presented provide rapid alternatives for analysingphenotypic drug susceptibility especially when the therapy is based on non-nucleoside RT inhibitors and thymidine-analogue drugs.
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| 33. |
- Ygge, J, et al.
(författare)
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Visual impairment and dyslexia in childhood
- 1997
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Ingår i: Current opinion in ophthalmology. - : Ovid Technologies (Wolters Kluwer Health). - 1040-8738. ; 8:5, s. 40-4
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Tidskriftsartikel (refereegranskat)
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