Spironolactone Depletes the XPB Protein and Inhibits DNA Damage Responses in UVB-Irradiated Human Skin.

UVB wavelengths of light induce the formation of photoproducts in genomic DNA that are potentially mutagenic and detrimental to epidermal cell function. The mineralocorticoid and androgen receptor antagonist spironolactone (SP) was recently identified as an inhibitor of UV photoproduct removal in human cancer cells in vitro via its ability to promote the rapid proteolytic degradation of the DNA repair protein XPB. Using normal human keratinocytes in vitro and skin explants ex vivo, we found that SP rapidly depleted XPB protein in both systems and abrogated two major responses to UVB-induced DNA damage, including the removal of UV photoproducts from genomic DNA and the activation of ATR/ATM DNA damage kinase signaling. These effects were also correlated with both mutagenesis and a predisposition to UVB-induced cell death but were unique to SP, because neither the SP metabolites canrenone and 7α-thiomethylspironolactone nor the more specific mineralocorticoid receptor antagonist eplerenone affected XPB protein levels or the UVB response. Our findings provide an approach for studying XPB and its roles in the UVB DNA damage response in human skin ex vivo and indicate that SP may increase UVB mutagenesis and skin cancer risk in certain individuals.

Diagnostic Performance of Deep Learning Algorithms Applied to Three Common Diagnoses in Dermatopathology.

BACKGROUND: Artificial intelligence is advancing at an accelerated pace into clinical applications, providing opportunities for increased efficiency, improved accuracy, and cost savings through computer-aided diagnostics. Dermatopathology, with emphasis on pattern recognition, offers a unique opportunity for testing deep learning algorithms. AIMS: This study aims to determine the accuracy of deep learning algorithms to diagnose three common dermatopathology diagnoses. METHODS: Whole slide images (WSI) of previously diagnosed nodular basal cell carcinomas (BCCs), dermal nevi, and seborrheic keratoses were annotated for areas of distinct morphology. Unannotated WSIs, consisting of five distractor diagnoses of common neoplastic and inflammatory diagnoses, were included in each training set. A proprietary fully convolutional neural network was developed to train algorithms to classify test images as positive or negative relative to ground truth diagnosis. RESULTS: Artificial intelligence system accurately classified 123/124 (99.45%) BCCs (nodular), 113/114 (99.4%) dermal nevi, and 123/123 (100%) seborrheic keratoses. CONCLUSIONS: Artificial intelligence using deep learning algorithms is a potential adjunct to diagnosis and may result in improved workflow efficiencies for dermatopathologists and laboratories.

Diagnostic Accuracy of Virtual Pathology vs Traditional Microscopy in a Large Dermatopathology Study.

Importance: Digital pathology represents a transformative technology that impacts dermatologists and dermatopathologists from residency to academic and private practice. Two concerns are accuracy of interpretation from whole-slide images (WSI) and effect on workflow. Studies of considerably large series involving single-organ systems are lacking. Objective: To evaluate whether diagnosis from WSI on a digital microscope is inferior to diagnosis of glass slides from traditional microscopy (TM) in a large cohort of dermatopathology cases with attention on image resolution, specifically eosinophils in inflammatory cases and mitotic figures in melanomas, and to measure the workflow efficiency of WSI compared with TM. Design, Setting, and Participants: Three dermatopathologists established interobserver ground truth consensus (GTC) diagnosis for 499 previously diagnosed cases proportionally representing the spectrum of diagnoses seen in the laboratory. Cases were distributed to 3 different dermatopathologists who diagnosed by WSI and TM with a minimum 30-day washout between methodologies. Intraobserver WSI/TM diagnoses were compared, followed by interobserver comparison with GTC. Concordance, major discrepancies, and minor discrepancies were calculated and analyzed by paired noninferiority testing. We also measured pathologists' read rates to evaluate workflow efficiency between WSI and TM. This retrospective study was caried out in an independent, national, university-affiliated dermatopathology laboratory. Main Outcomes and Measures: Intraobserver concordance of diagnoses between WSI and TM methods and interobserver variance from GTC, following College of American Pathology guidelines. Results: Mean intraobserver concordance between WSI and TM was 94%. Mean interobserver concordance was 94% for WSI and GTC and 94% for TM and GTC. Mean interobserver concordance between WSI, TM, and GTC was 91%. Diagnoses from WSI were noninferior to those from TM. Whole-slide image read rates were commensurate with WSI experience, achieving parity with TM by the most experienced user. Conclusions and Relevance: Diagnosis from WSI was found equivalent to diagnosis from glass slides using TM in this statistically powerful study of 499 dermatopathology cases. This study supports the viability of WSI for primary diagnosis in the clinical setting.

Copy number variation in archival melanoma biopsies versus benign melanocytic lesions.

BACKGROUND: Skin melanocytes can give rise to benign and malignant neoplasms. Discrimination of an early melanoma from an unusual/atypical benign nevus can represent a significant challenge. However, previous studies have shown that in contrast to benign nevi, melanoma demonstrates pervasive chromosomal aberrations. OBJECTIVE: This substantial difference between melanoma and benign nevi can be exploited to discriminate between melanoma and benign nevi. METHODS: Array-comparative genomic hybridization (aCGH) is an approach that can be used on DNA extracted from formalin-fixed paraffin-embedded (FFPE) tissues to assess the entire genome for the presence of changes in DNA copy number. In this study, high resolution, genome-wide single-nucleotide polymorphism (SNP) arrays were utilized to perform comprehensive and detailed analyses of recurrent copy number aberrations in 41 melanoma samples in comparison with 21 benign nevi. RESULTS: We found statistically significant copy number gains and losses within melanoma samples. Some of the identified aberrations are previously implicated in melanoma. Moreover, novel regions of copy number alterations were identified, revealing new candidate genes potentially involved in melanoma pathogenesis. CONCLUSIONS: Taken together, these findings can help improve melanoma diagnosis and introduce novel melanoma therapeutic targets.

Genomic DNA extraction methods using formalin-fixed paraffin-embedded tissue.

As new technologies come within reach for the average cytogenetic laboratory, the study of chromosome structure has become increasingly more sophisticated. Resolution has improved from karyotyping (in which whole chromosomes are discernible) to fluorescence in situ hybridization and comparative genomic hybridization (CGH, with which specific megabase regions are visualized), array-based CGH (aCGH, examining hundreds of base pairs), and next-generation sequencing (providing single base pair resolution). Whole genome next-generation sequencing remains a cost-prohibitive method for many investigators. Meanwhile, the cost of aCGH has been reduced during recent years, even as resolution has increased and protocols have simplified. However, aCGH presents its own set of unique challenges. DNA of sufficient quantity and quality to hybridize to arrays and provide meaningful results is required. This is especially difficult for DNA from formalin-fixed paraffin-embedded (FFPE) tissues. Here, we compare three different methods for acquiring DNA of sufficient length, purity, and "amplifiability" for aCGH and other downstream applications. Phenol-chloroform extraction and column-based commercial kits were compared with adaptive focused acoustics (AFA). Of the three extraction methods, AFA samples showed increased amplicon length and decreased polymerase chain reaction (PCR) failure rate. These findings support AFA as an improvement over previous DNA extraction methods for FFPE tissues.

Comparative metabolomic and genomic analyses of TCDD-elicited metabolic disruption in mouse and rat liver.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) elicits a broad spectrum of species-specific effects that have not yet been fully characterized. This study compares the temporal effects of TCDD on hepatic aqueous and lipid metabolite extracts from immature ovariectomized C57BL/6 mice and Sprague-Dawley rats using gas chromatography-mass spectrometry and nuclear magnetic resonance-based metabolomic approaches and integrates published gene expression data to identify species-specific pathways affected by treatment. TCDD elicited metabolite and gene expression changes associated with lipid metabolism and transport, choline metabolism, bile acid metabolism, glycolysis, and glycerophospholipid metabolism. Lipid metabolism is altered in mice resulting in increased hepatic triacylglycerol as well as mono- and polyunsaturated fatty acid (FA) levels. Mouse-specific changes included the induction of CD36 and other cell surface receptors as well as lipases- and FA-binding proteins consistent with hepatic triglyceride and FA accumulation. In contrast, there was minimal hepatic fat accumulation in rats and decreased CD36 expression. However, choline metabolism was altered in rats, as indicated by decreases in betaine and increases in phosphocholine with the concomitant induction of betaine-homocysteine methyltransferase and choline kinase gene expression. Results from these studies show that aryl hydrocarbon receptor-mediated differential gene expression could be linked to metabolite changes and species-specific alterations of biochemical pathways.