IMPORTANCE: Improved screening methods for women with dense breasts are needed because of their increased risk of breast cancer and of failed early diagnosis by screening mammography. OBJECTIVE: To ...compare the screening performance of abbreviated breast magnetic resonance imaging (MRI) and digital breast tomosynthesis (DBT) in women with dense breasts. DESIGN, SETTING, AND PARTICIPANTS: Cross-sectional study with longitudinal follow-up at 48 academic, community hospital, and private practice sites in the United States and Germany, conducted between December 2016 and November 2017 among average-risk women aged 40 to 75 years with heterogeneously dense or extremely dense breasts undergoing routine screening. Follow-up ascertainment of cancer diagnoses was complete through September 12, 2019. EXPOSURES: All women underwent screening by both DBT and abbreviated breast MRI, performed in randomized order and read independently to avoid interpretation bias. MAIN OUTCOMES AND MEASURES: The primary end point was the invasive cancer detection rate. Secondary outcomes included sensitivity, specificity, additional imaging recommendation rate, and positive predictive value (PPV) of biopsy, using invasive cancer and ductal carcinoma in situ (DCIS) to define a positive reference standard. All outcomes are reported at the participant level. Pathology of core or surgical biopsy was the reference standard for cancer detection rate and PPV; interval cancers reported until the next annual screen were included in the reference standard for sensitivity and specificity. RESULTS: Among 1516 enrolled women, 1444 (median age, 54 range, 40-75 years) completed both examinations and were included in the analysis. The reference standard was positive for invasive cancer with or without DCIS in 17 women and for DCIS alone in another 6. No interval cancers were observed during follow-up. Abbreviated breast MRI detected all 17 women with invasive cancer and 5 of 6 women with DCIS. Digital breast tomosynthesis detected 7 of 17 women with invasive cancer and 2 of 6 women with DCIS. The invasive cancer detection rate was 11.8 (95% CI, 7.4-18.8) per 1000 women for abbreviated breast MRI vs 4.8 (95% CI, 2.4-10.0) per 1000 women for DBT, a difference of 7 (95% CI, 2.2-11.6) per 1000 women (exact McNemar P = .002). For detection of invasive cancer and DCIS, sensitivity was 95.7% (95% CI, 79.0%-99.2%) with abbreviated breast MRI vs 39.1% (95% CI, 22.2%-59.2%) with DBT (P = .001) and specificity was 86.7% (95% CI, 84.8%-88.4%) vs 97.4% (95% CI, 96.5%-98.1%), respectively (P < .001). The additional imaging recommendation rate was 7.5% (95% CI, 6.2%-9.0%) with abbreviated breast MRI vs 10.1% (95% CI, 8.7%-11.8%) with DBT (P = .02) and the PPV was 19.6% (95% CI, 13.2%-28.2%) vs 31.0% (95% CI, 17.0%-49.7%), respectively (P = .15). CONCLUSIONS AND RELEVANCE: Among women with dense breasts undergoing screening, abbreviated breast MRI, compared with DBT, was associated with a significantly higher rate of invasive breast cancer detection. Further research is needed to better understand the relationship between screening methods and clinical outcome. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02933489
Abstract
Cancer chemopreventive agents inhibit the formation of precursor lesions and/or the progression of these lesions to late stage disease. This approach to disease control has the potential to ...reduce the physical and financial costs of cancer in society. Several drugs that have been approved by the FDA for other diseases and have been extensively evaluated for their safety and pharmacokinetic/pharmacodynamic characteristics have the potential to be repurposed for use as cancer chemopreventive agents. These agents often mechanistically inhibit signaling molecules that play key roles in the carcinogenic process. The safety profile of agents is a primary concern when considering the administration of drugs for chemoprevention, as the drugs will be given chronically to high-risk, asymptomatic individuals. To decrease drug toxicity while retaining efficacy, several approaches are currently being explored. In this short review, we describe studies that use preclinical in vivo models to assess efficacy of alternative drug dosing strategies and routes of drug administration on chemopreventive drug efficacy. In vivo drug dosing strategies that reduce toxicity while retaining efficacy will pave the way for future cancer prevention clinical trials.
Initiation of protein-primed (-) strand DNA synthesis in hepatitis B virus (HBV) requires interaction of the viral reverse transcriptase with epsilon (ε), a
-acting regulatory signal located at the ...5' terminus of pre-genomic RNA (pgRNA), and several host-encoded chaperone proteins. Binding of the viral polymerase (P protein) to ε is necessary for pgRNA encapsidation and synthesis of a short primer covalently attached to its terminal domain. Although we identified small molecules that recognize HBV ε RNA, these failed to inhibit protein-primed DNA synthesis. However, since initiation of HBV (-) strand DNA synthesis occurs within a complex of viral and host components (e.g., Hsp90, DDX3 and APOBEC3G), we considered an alternative therapeutic strategy of allosteric inhibition by disrupting the initiation complex or modifying its topology. To this end, we show here that 3,7-dihydroxytropolones (3,7-dHTs) can inhibit HBV protein-primed DNA synthesis. Since DNA polymerase activity of a ribonuclease (RNase H)-deficient HBV reverse transcriptase that otherwise retains DNA polymerase function is also abrogated, this eliminates direct involvement of RNase (ribonuclease) H activity of HBV reverse transcriptase and supports the notion that the HBV initiation complex might be therapeutically targeted. Modeling studies also provide a rationale for preferential activity of 3,7-dHTs over structurally related α-hydroxytropolones (α-HTs).
Kaposi's sarcoma-associated herpes virus (KSHV) polyadenylated nuclear (PAN) RNA facilitates lytic infection, modulating the cellular immune response by interacting with viral and cellular proteins ...and DNA. Although a number nucleoprotein interactions involving PAN have been implicated, our understanding of binding partners and PAN RNA binding motifs remains incomplete. Herein, we used SHAPE-mutational profiling (SHAPE-MaP) to probe PAN in its nuclear, cytoplasmic or viral environments or following cell/virion lysis and removal of proteins. We thus characterized and put into context discrete RNA structural elements, including the cis-acting Mta responsive element and expression and nuclear retention element (1,2). By comparing mutational profiles in different biological contexts, we identified sites on PAN either protected from chemical modification by protein binding or characterized by a loss of structure. While some protein binding sites were selectively localized, others were occupied in all three biological contexts. Individual binding sites of select KSHV gene products on PAN RNA were also identified in in vitro experiments. This work constitutes the most extensive structural characterization of a viral lncRNA and interactions with its protein partners in discrete biological contexts, providing a broad framework for understanding the roles of PAN RNA in KSHV infection.
Therapeutic targeting of RAS-mutated cancers is difficult, whereas prevention or interception (treatment before or in the presence of preinvasive lesions) preclinically has proven easier. In the A/J ...mouse lung model, where different carcinogens induce tumors with different KRAS mutations, glucocorticoids and RXR agonists are effective agents in prevention and interception studies, irrespective of specific KRAS mutations. In rat azoxymethane-induced colon tumors (45% KRAS mutations), cyclooxygenase 1/2 inhibitors and difluoromethylornithine are effective in preventing or intercepting KRAS-mutated or wild-type tumors. In two KRAS mutant pancreatic models multiple COX 1/2 inhibitors are effective. Furthermore, combining a COX and an EGFR inhibitor prevented the development of virtually all pancreatic tumors in transgenic mice. In the N-nitroso-N-methylurea-induced ER positive rat breast model (50%HRAS mutations) various selective estrogen receptor modulators, aromatase inhibitors, epidermal growth factor receptor inhibitors, and retinoid X receptor agonists are profoundly effective in prevention and interception of tumors with wild type or mutant HRAS, while the farnesyltransferase inhibitor tipifarnib preferentially inhibits HRAS-mutant breast tumors. Thus, many agents not known to specifically inhibit the RAS pathway, are effective in an organ specific manner in preventing or intercepting RAS-mutated tumors. Finally, we discuss an alternative prevention and interception approach, employing vaccines to target KRAS.
Human immunodeficiency virus type 1 (HIV-1) replication requires reverse transcription of its RNA genome into a double-stranded cDNA copy, which is then integrated into the host cell chromosome. The ...essential steps of reverse transcription and integration are catalyzed by the viral enzymes reverse transcriptase (RT) and integrase (IN), respectively. In vitro, HIV-1 RT can bind with IN, and the C-terminal domain (CTD) of IN is necessary and sufficient for this binding. To better define the RT-IN interaction, we performed nuclear magnetic resonance (NMR) spectroscopy experiments to map a binding surface on the IN CTD in the presence of RT prebound to a duplex DNA construct that mimics the primer-binding site in the HIV-1 genome. To determine the biological significance of the RT-IN interaction during viral replication, we used the NMR chemical shift mapping information as a guide to introduce single amino acid substitutions of nine different residues on the putative RT-binding surface in the IN CTD. We found that six viral clones bearing such IN substitutions (R231E, W243E, G247E, A248E, V250E, and I251E) were noninfectious. Further analyses of the replication-defective IN mutants indicated that the block in replication took place specifically during early reverse transcription. The recombinant INs purified from these mutants, though retaining enzymatic activities, had diminished ability to bind RT in a cosedimentation assay. The results indicate that the RT-IN interaction is functionally relevant during the reverse transcription step of the HIV-1 life cycle.
To establish a productive infection, human immunodeficiency virus type 1 (HIV-1) needs to reverse transcribe its RNA genome to create a double-stranded DNA copy and then integrate this viral DNA genome into the chromosome of the host cell. These two essential steps are catalyzed by the HIV-1 enzymes reverse transcriptase (RT) and integrase (IN), respectively. We have shown previously that IN physically interacts with RT, but the importance of this interaction during HIV-1 replication has not been fully characterized. In this study, we have established the biological significance of the HIV-1 RT-IN interaction during the viral life cycle by demonstrating that altering the RT-binding surface on IN disrupts both reverse transcription and viral replication. These findings contribute to our understanding of the RT-IN binding mechanism, as well as indicate that the RT-IN interaction can be exploited as a new antiviral drug target.
•aCFLD had higher S. salivarius and V. parvula fecal abundances compared to CFnoLD.•Both species decreased after ETI in subjects with aCFLD and calprotectin decrease.•This supports the gut-liver axis ...linking dysbiosis, intestinal inflammation & aCFLD.
Cystic fibrosis associated liver disease (CFLD) carries a significant disease burden with no effective preventive therapies. According to the gut-liver axis hypothesis for CFLD pathogenesis, dysbiosis and increased intestinal inflammation and permeability permit pathogenic bacterial translocation into the portal circulation, leading to hepatic inflammation and fibrosis. Evaluating the effect of CFTR (cystic fibrosis transmembrane conductance regulator) modulation with elexacaftor/tezacaftor/ivacaftor (ETI) may help determine the role of CFTR in CFLD and increase understanding of CFLD pathogenesis, which is critical for developing therapies. We aimed to characterize the fecal microbiota in participants with CF with and without advanced CFLD (aCFLD) before and after ETI.
This is an ancillary analysis of stool samples from participants ages ≥12 y/o enrolled in PROMISE (NCT04038047). Included participants had aCFLD (cirrhosis with or without portal hypertension, or non-cirrhotic portal hypertension) or CF without liver disease (CFnoLD). Fecal microbiota were defined by shotgun metagenomic sequencing at baseline and 1 and 6 months post-ETI.
We analyzed 93 samples from 34 participants (11 aCFLD and 23 CFnoLD). Compared to CFnoLD, aCFLD had significantly higher baseline relative abundances of potential pathogens Streptococcus salivarius and Veillonella parvula. Four of 11 aCFLD participants had an initially abnormal fecal calprotectin that normalized 6 months post-ETI, correlating with a significant decrease in S. salivarius and a trend towards decreasing V. parvula.
These results support an association between dysbiosis and intestinal inflammation in CFLD with improvements in both post-ETI, lending further support to the gut-liver axis in aCFLD.
Airway smooth muscle cells (ASM) are fundamental to asthma pathogenesis, influencing bronchoconstriction, airway hyper-responsiveness, and airway remodelling. Extracellular matrix (ECM) can influence ...tissue remodelling pathways, however, to date no study has investigated the effect of ASM ECM stiffness and crosslinking on the development of asthmatic airway remodelling. We hypothesised that TGFβ activation by ASM is influenced by ECM in asthma and sought to investigate the mechanisms involved. This study combines
and
approaches: human ASM cells were used
to investigate basal TGFβ activation and expression of ECM crosslinking enzymes. Human bronchial biopsies from asthmatic and non-asthmatic donors were used to confirm LOXL2 expression ASM. A chronic ovalbumin model of asthma was used to study the effect of LOXL2 inhibition on airway remodelling. We found that ASM cells from asthmatics activated more TGFβ basally than non-asthmatic controls and that diseased cell-derived ECM influences levels of TGFβ activated. Our data demonstrate that the ECM crosslinking enzyme LOXL2 is increased in asthmatic ASM cells and in bronchial biopsies. Crucially, we show that LOXL2 inhibition reduces ECM stiffness and TGFβ activation
, and can reduce subepithelial collagen deposition and ASM thickness, two features of airway remodelling, in an ovalbumin mouse model of asthma. These data are the first to highlight a role for LOXL2 in the development of asthmatic airway remodelling and suggest that LOXL2 inhibition warrants further investigation as a potential therapy to reduce remodelling of the airways in severe asthma.
Hundreds of structures of type 1 human immunodeficiency virus (HIV-1) reverse transcriptase (RT) have been determined, but only one contains an RNA/DNA hybrid. Here we report three structures of ...HIV-1 RT complexed with a non-nucleotide RT inhibitor (NNRTI) and an RNA/DNA hybrid. In the presence of an NNRTI, the RNA/DNA structure differs from all prior nucleic acid-RT structures including the RNA/DNA hybrid. The enzyme structure also differs from all previous RT-DNA complexes. Thus, the hybrid has ready access to the RNase-H active site. These observations indicate that an RT-nucleic acid complex may adopt two structural states, one competent for DNA polymerization and the other for RNA degradation. RT mutations that confer drug resistance but are distant from the inhibitor-binding sites often map to the unique RT-hybrid interface that undergoes conformational changes between two catalytic states.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Kaposi's sarcoma-associated herpesvirus (KSHV), the etiological agent of Kaposi's sarcoma, belongs to the
family, whose members employ a multicomponent terminase to resolve nonparametric viral DNA ...into genome-length units prior to their packaging. Homology modeling of the ORF29 C-terminal nuclease domain (pORF29C) and bacteriophage Sf6 gp2 have suggested an active site clustered with four acidic residues, D
, E
, D
, and D
, that collectively sequester the catalytic divalent metal (Mn
) and also provided important insight into a potential inhibitor binding mode. Using this model, we have expressed, purified, and characterized the wild-type pORF29C and variants with substitutions at the proposed active-site residues. Differential scanning calorimetry demonstrated divalent metal-induced stabilization of wild-type (WT) and D
A pORF29C, consistent with which these two enzymes exhibited Mn
-dependent nuclease activity, although the latter mutant was significantly impaired. Thermal stability of WT and D
A pORF29C was also enhanced by binding of an α-hydroxytropolone (α-HT) inhibitor shown to replace divalent metal at the active site. For the remaining mutants, thermal stability was unaffected by divalent metal or α-HT binding, supporting their role in catalysis. pORF29C nuclease activity was also inhibited by two classes of small molecules reported to inhibit HIV RNase H and integrase, both of which belong to the superfamily of nucleotidyltransferases. Finally, α-HT inhibition of KSHV replication suggests ORF29 nuclease function as an antiviral target that could be combined with latency-activating compounds as a shock-and-kill antiviral strategy.
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