Publications

The mechanisms by which transcription factor haploinsufficiency alters the epigenetic and transcriptional landscape in human cells to cause disease are unknown. Here, we utilized human induced pluripotent stem cell (iPSC)-derived endothelial cells (ECs) to show that heterozygous nonsense mutations in NOTCH1 that cause aortic valve calcification disrupt the epigenetic architecture, resulting in derepression of latent pro-osteogenic and -inflammatory gene networks. Hemodynamic shear stress, which protects valves from calcification in vivo, activated anti-osteogenic and anti-inflammatory networks in NOTCH1(+/+), but not NOTCH1(+/-), iPSC-derived ECs. NOTCH1 haploinsufficiency altered H3K27ac at NOTCH1-bound enhancers, dysregulating downstream transcription of more than 1,000 genes involved in osteogenesis, inflammation, and oxidative stress. Computational predictions of the disrupted NOTCH1-dependent gene network revealed regulatory nodes that, when modulated, restored the network toward the NOTCH1(+/+) state. Our results highlight how alterations in transcription factor dosage affect gene networks leading to human disease and reveal nodes for potential therapeutic intervention.

Resistance to combined antiretroviral therapy (cART) in HIV-1-infected individuals is typically due to nonsynonymous mutations that change the protein sequence; however, the selection of synonymous or 'silent' mutations in the HIV-1 genome with cART has been reported. These silent K65K and K66K mutations in the HIV-1 reverse transcriptase (RT) occur in over 35% of drug-experienced individuals and are highly associated with the thymidine analog mutations D67N and K70R, which confer decreased susceptibility to most nucleoside and nucleotide RT inhibitors. However, the basis for selection of these silent mutations under selective drug pressure is unknown. Using Illumina next-generation sequencing, we demonstrate that the D67N/K70R substitutions in HIV-1 RT increase indel frequency by 100-fold at RT codons 65-67, consequently impairing viral fitness. Introduction of either K65K or K66K into HIV-1 containing D67N/K70R reversed the error-prone DNA synthesis at codons 65-67 in RT and improved viral replication fitness, but did not impact RT inhibitor drug susceptibility. These data provide new mechanistic insights into the role of silent mutations selected during antiretroviral therapy and have broader implications for the relevance of silent mutations in the evolution and fitness of RNA viruses.

BACKGROUND

Human immunodeficiency virus (HIV)-infected individuals are at higher risk for chronic kidney disease than HIV-uninfected individuals. We investigated whether the inflammation present in treated HIV infection contributes to kidney dysfunction among HIV-infected men receiving highly active antiretroviral therapy.

METHODS

The glomerular filtration rate (GFR) was directly measured (using iohexol) along with 12 markers of inflammation in Multicenter AIDS Cohort Study participants. Exploratory factor analysis was used to identify inflammatory processes related to kidney dysfunction. The estimated levels of these inflammatory processes were used in adjusted logistic regression analyses evaluating cross-sectional associations with kidney function outcomes.

RESULTS

There were 434 HIV-infected men receiving highly active antiretroviral therapy and 200 HIV-uninfected men. HIV-infected men were younger (median age, 51 vs 53 years) and had higher urine protein-creatinine ratios (median, 98 vs 66 mg/g) but comparable GFRs (median, 109 vs 106 mL/min|1.73 m(2)). We found an inflammatory process dominated by markers: soluble tumor necrosis factor receptor 2, soluble interleukin 2 receptor α, soluble gp130, soluble CD27, and soluble CD14. An increase of 1 standard deviation in that inflammatory process was associated with significantly greater odds of GFR ≤90 mL/min/1.73 m(2) (odds ratio, 2.0) and urine protein >200 mg/g (odds ratio, 2.3).

CONCLUSIONS

Higher circulating levels of immune activation markers among treated HIV-infected men may partially explain their higher burden of kidney dysfunction compared with uninfected men.

BACKGROUND

An accurate system for tracking of colonoscopy quality and surveillance intervals could improve the effectiveness and cost-effectiveness of colorectal cancer (CRC) screening and surveillance. The purpose of this study was to create and test such a system across multiple institutions utilizing natural language processing (NLP).

METHODS

From 42,569 colonoscopies with pathology records from 13 centers, we randomly sampled 750 paired reports. We trained (n=250) and tested (n=500) an NLP-based program with 19 measurements that encompass colonoscopy quality measures and surveillance interval determination, using blinded, paired, annotated expert manual review as the reference standard. The remaining 41,819 nonannotated documents were processed through the NLP system without manual review to assess performance consistency. The primary outcome was system accuracy across the 19 measures.

RESULTS

A total of 176 (23.5%) documents with 252 (1.8%) discrepant content points resulted from paired annotation. Error rate within the 500 test documents was 31.2% for NLP and 25.4% for the paired annotators (P=0.001). At the content point level within the test set, the error rate was 3.5% for NLP and 1.9% for the paired annotators (P=0.04). When eight vaguely worded documents were removed, 125 of 492 (25.4%) were incorrect by NLP and 104 of 492 (21.1%) by the initial annotator (P=0.07). Rates of pathologic findings calculated from NLP were similar to those calculated by annotation for the majority of measurements. Test set accuracy was 99.6% for CRC, 95% for advanced adenoma, 94.6% for nonadvanced adenoma, 99.8% for advanced sessile serrated polyps, 99.2% for nonadvanced sessile serrated polyps, 96.8% for large hyperplastic polyps, and 96.0% for small hyperplastic polyps. Lesion location showed high accuracy (87.0-99.8%). Accuracy for number of adenomas was 92%.

CONCLUSIONS

NLP can accurately report adenoma detection rate and the components for determining guideline-adherent colonoscopy surveillance intervals across multiple sites that utilize different methods for reporting colonoscopy findings.