In ER-Positive, HER2-Negative Breast Cancers, HER2 mRNA Levels Correlate Better with Clinicopathologic Features and Oncotype DX Recurrence Score than HER2 Immunohistochemistry.

With the approval of trastuzumab deruxtecan for treating advanced human epidermal growth factor receptor-2 (HER2) low breast cancer (BC), it has become increasingly important to develop more accurate and reliable methods to identify HER2-low BC. In addition, HER2 immunohistochemistry (IHC) has limitations for quantification of HER2. We explored the relationship between HER2 IHC and mRNA levels and evaluated whether HER2 IHC scores and mRNA levels are associated with clinicopathologic features and Oncotype DX Recurrence Score (RS) in estrogen receptor (ER)-positive, HER2-negative BCs. A total of 750 BCs sent for Oncotype DX (ODX) testing were included in this study, and 559 with HER2 mRNA levels were available. There were no statistically significant differences between HER2 0 and HER2-low BC in clinicopathologic variables or ODX RS using HER2 IHC. There was a significant difference in median HER2 mRNA values between HER2 0 and HER2-low (8.7 vs 9.3, P < .001); however, the HER2 mRNA distribution had substantial overlap between these 2 groups with a suboptimal area under the receiver operating characteristic curve (area under the receiver operating characteristic curve = 0.68). A HER2 mRNA value of 9.2 was generated as the optimal cutoff for distinguishing HER2 0 and HER2-low BC. Comparing ER+ BCs with HER2 mRNA high (>9.2) and low (≤9.2) revealed a statistically significant difference in most clinicopathologic variables and ODX RS. From this large cohort of ER-positive, HER2-negative BC, our results demonstrated that HER2 mRNA levels correlated better with clinicopathologic features and recurrence risk as assessed by ODX RS than HER2 IHC scores. Our findings suggest that HER2 mRNA-detecting methods could potentially serve as a quantitative and reliable method for identifying a biologically meaningful group of HER2-low BC. Further study is needed to determine whether HER2 mRNA levels could be more reliable than IHC for identifying which patients will be most likely to benefit from trastuzumab deruxtecan.

Associations of Single Versus Multiple Human Papillomavirus Infections With the Prevalence of Cervical Intraepithelial Neoplasia 2/3 and Squamous Cell Carcinoma Lesions: Human Papillomavirus Type-Specific Attribution.

The risk of developing cervical squamous lesions in women with multiple high-risk human papillomavirus (hrHPV) infections is uncertain. The aim of this retrospective study was to investigate the type-specific attribution and phylogenetic effects of single and multiple hrHPV subtypes in cervical squamous lesions. All cases with cervical histopathologic diagnosis and human papillomavirus (HPV) genotyping results in the 6 months preceding biopsy from October 2018 to December 2022 were studied and analyzed. Over the study period, 70,361 cases with histopathologic follow-up and prior HPV genotyping were identified. The hrHPV-positive rate was 55.6% (39,104/70,361), including single hrHPV detected in 27,182 (38.6%), 2 types of hrHPV detected in 8158 (11.6%), and 3 types of hrHPV detected in 2486 (3.5%). Among 16,457 cases with a histologically diagnosed squamous lesion (cervical intraepithelial neoplasia 1: 11411; cervical intraepithelial neoplasia 2/3: 4192; squamous cell carcinoma: 854 cases), the prevalence of single hrHPV infection increased, but the rate of multiple concomitant hrHPV infections showed negative association as the degree of squamous lesions increased. Among women with a single HPV16 infection, cervical intraepithelial neoplasia 2/3 and squamous cell carcinoma (CIN2+) diagnostic rate was 30.6%, and it increased to 47.6% when coinfected with HPV33 (P < .001) but significantly decreased when coinfected with all other hrHPV types (P < .05). By comparing CIN2+ diagnostic rates in 40 most common 2 types of hrHPV infections with related single hrHPV infection, CIN2+ rates were decreased in 12 combinations (30.0%), equivalent in 26 combinations (65.0%), and increased in 2 combinations (5.0%). The cases with 3 types of HPV infections reduced the risk for CIN2+ compared with related single HPV infections. HPV16+52+53, HPV16+52+68, HPV16+52+51, HPV16+39+52, and HPV16+58+53 significantly decreased the risk of CIN2+ compared with HPV16 single infection (P < .05). This study demonstrates that multiple hrHPV infections are not associated with cumulatively higher risk for CIN2+ development, suggesting that oncogenic progression of multiple hrHPV-associated cervical squamous lesions is neither synergistic nor a cumulative effect at the phylogenetic level, possibly a way of competitive interference.

Breast Cancer With HER2 Immunohistochemical Score 2 and Average HER2 Signals/Cell 6 or More and HER2/CEP17 Ratio Less Than 2 ('ISH Group 3'): A Multiinstitutional Cohort Analysis Emphasizing Outcome and Molecular Subtype.

Breast cancer (BC) with average human epidermal growth factor receptor 2 (HER2) signals/cell ≥6 and HER2/chromosome enumeration probe 17 (CEP17) ratio <2 (in situ hybridization [ISH] group 3) is very rare, accounting for 0.4% to 3.0% of cases sent for the dual-probe ISH assay. Although such patients are currently eligible for treatment with HER2-targeted therapy, their characteristics and outcomes remain poorly understood. Sixty-two BCs with equivocal HER2 immunohistochemical score (2+) and reflex ISH group 3 results were identified across 4 institutions. Available clinicopathologic characteristics, MammaPrint and BluePrint molecular results, and follow-up information were retrospectively analyzed. Most BCs with HER2 equivocal immunohistochemical and ISH group 3 results were histologic grade 2 or 3 (100%), estrogen receptor (ER) positive (90.3%), with an average HER2 signals/cell of 7.3. Molecular profiles revealed that 80% (16/20) of tumors were luminal subtypes, and HER2 molecular subtype was identified in 10% of tumors (2/20). Twelve (19.4%) out of 62 patients developed local recurrence and/or distant metastasis with a median follow-up of 50 months. One (10%) of 10 patients achieved pathologic complete response after neoadjuvant chemotherapy. Forty-nine (79%) out of 62 patients completed anti-HER2 agents, and exploratory analysis showed no statistically significant difference in disease outcomes between patients who completed anti-HER2 treatment and those who did not. Univariate analysis revealed advanced clinical stage, and ER/progesterone receptor negativity was associated with unfavorable disease outcomes, and exploratory multivariate analysis demonstrated that clinical stage was the most significant factor associated with disease outcomes in the studied population. These findings increase our understanding of this rare, but clinically important HER2 category. Large-scale prospective randomized studies are needed to further evaluate the role of perioperative HER2-targeted therapy in this patient population.

Correlation of HER2 Protein Level With mRNA Level Quantified by RNAscope in Breast Cancer.

Trastuzumab deruxtecan (T-DXd) has been approved by the US Food and Drug Administration (FDA) to treat patients with metastatic HER2-positive and HER2-low breast cancer, and clinical trials are examining its efficacy against early-stage breast cancer. Current HER2 immunohistochemical (IHC) assays are suboptimal in evaluating HER2-low breast cancers and identifying which patients would benefit from T-DXd. HER2 expression in 526 breast cancer tissue microarray (TMA) cores was measured using the FDA-approved PATHWAY and HercepTest IHC assays, and the corresponding RNA levels were evaluated by RNAscope. HER2 protein levels by regression analysis using a quantitative immunofluorescence score against cell line arrays with known HER2 protein levels determined by mass spectrometry were available in 48 of the cores. RNAscope was also performed in 32 metastatic biopsies from 23 patients who were subsequently treated with T-DXd, and the results were correlated with response rate. HER2 RNA levels by RNAscope strongly correlated with HER2 protein levels (P < .0001) and with HER2 IHC H-scores from the PATHWAY and HercepTest assays (P < .0001). However, neither protein levels nor RNA levels significantly differed between cases scored 0, ultralow, and 1+ by PATHWAY and HercepTest. The RNA levels were significantly higher (P = .030) in responders (6.4 ± 8.2 dots/cell, n = 12) than those in nonresponders (2.6 ± 2.2, n = 20) to T-DXd. RNAscope is a simple assay that can be objectively quantified and is a promising alternative to current IHC assays in evaluating HER2 expression in breast cancers, especially HER2-low cases, and may identify patients who would benefit from T-DXd.

HER2 evaluation for clinical decision making in human solid tumours: pearls and pitfalls.

The significant clinical benefits of human epidermal growth factor receptor 2 (HER2)-targeted therapeutic agents have revolutionized the clinical treatment landscape in a variety of human solid tumours. Accordingly, accurate evaluation of HER2 status in these different tumour types is critical for clinical decision making to select appropriate patients who may benefit from life-saving HER2-targeted therapies. HER2 biomarker scoring criteria is different in different organ systems, and close adherence to the corresponding HER2 biomarker testing guidelines and their updates, if available, is essential for accurate evaluation. In addition, knowing the unusual patterns of HER2 expression is also important to avoid inaccurate evaluation. In this review, we discuss the key considerations when evaluating HER2 status in solid tumours for clinical decision making, including tissue handling and preparation for HER2 biomarker testing, as well as pathologist's readout of HER2 testing results in breast carcinomas, gastroesophageal adenocarcinomas, colorectal adenocarcinomas, gynaecologic carcinomas, and non-small cell lung carcinomas.

Sirtuin 2 deficiency aggravates ageing-induced vascular remodelling in humans and mice.

AIMS: The mechanisms underlying ageing-induced vascular remodelling remain unclear. This study investigates the role and underlying mechanisms of the cytoplasmic deacetylase sirtuin 2 (SIRT2) in ageing-induced vascular remodelling. METHODS AND RESULTS: Transcriptome and quantitative real-time PCR data were used to analyse sirtuin expression. Young and old wild-type and Sirt2 knockout mice were used to explore vascular function and pathological remodelling. RNA-seq, histochemical staining, and biochemical assays were used to evaluate the effects of Sirt2 knockout on the vascular transcriptome and pathological remodelling and explore the underlying biochemical mechanisms. Among the sirtuins, SIRT2 had the highest levels in human and mouse aortas. Sirtuin 2 activity was reduced in aged aortas, and loss of SIRT2 accelerated vascular ageing. In old mice, SIRT2 deficiency aggravated ageing-induced arterial stiffness and constriction-relaxation dysfunction, accompanied by aortic remodelling (thickened vascular medial layers, breakage of elastin fibres, collagen deposition, and inflammation). Transcriptome and biochemical analyses revealed that the ageing-controlling protein p66Shc and metabolism of mitochondrial reactive oxygen species (mROS) contributed to SIRT2 function in vascular ageing. Sirtuin 2 repressed p66Shc activation and mROS production by deacetylating p66Shc at lysine 81. Elimination of reactive oxygen species by MnTBAP repressed the SIRT2 deficiency-mediated aggravation of vascular remodelling and dysfunction in angiotensin II-challenged and aged mice. The SIRT2 coexpression module in aortas was reduced with ageing across species and was a significant predictor of age-related aortic diseases in humans. CONCLUSION: The deacetylase SIRT2 is a response to ageing that delays vascular ageing, and the cytoplasm-mitochondria axis (SIRT2-p66Shc-mROS) is important for vascular ageing. Therefore, SIRT2 may serve as a potential therapeutic target for vascular rejuvenation.

Single and Multiple High-Risk Human Papillomavirus Infections in Histopathologically Confirmed Cervical Squamous Lesions: Incidences, Distribution, and Associated Detection Rates for Precancerous and Cancerous Lesions.

Coinfection with multiple high-risk human papillomavirus (hrHPV) is frequently observed in cervical specimens; however, the clinical significance of concomitant multiple hrHPV infections is poorly understood, and the published results remain inconsistent. A retrospective study at a tertiary care institution was performed, evaluating Tellgenplex human papillomavirus (HPV) 27 genotyping or YanengBio HPV 23 genotyping results and immediate cervical histologic diagnosis (within 6 months after HPV genotyping), between November 2015 and October 2022. Among 49,299 cases with hrHPV genotyping and histologic diagnosis, 24,361 cases were diagnosed as cervical intraepithelial neoplasia (CIN) and squamous cell carcinoma. Among women with cervical squamous lesions, 86.5% (21,070/24,361) had hrHPV infections, and concomitant multiple hrHPV infections accounted for 24.7% of hrHPV-positive cases (5210/21,070). The hrHPV-positive rates in these cervical squamous lesions increased progressively with disease severity; however, the percentages of concomitant multiple hrHPV infection rates among hrHPV-positive cases decreased significantly with increasing degree of squamous abnormalities. There was no increased detection rate of CIN3+ (CIN3 and squamous cell carcinoma) in cases with concomitant 2 or 3 hrHPV genotype infections when compared with those with corresponding single hrHPV infections. Conversely, some combinations of multiple hrHPV infections demonstrated a decrease in the detection rates of CIN3+ lesions. In this large cohort, our results demonstrated that multiple hrHPV infections do not carry an increased risk for developing CIN3+ lesions when compared to the corresponding single-genotype infection. The reduced risk of CIN3+ in women infected with some combinations of hrHPV genotypes compared to those with single-genotype infections supports the concept of intergenotypic competition of hrHPV genotypes in cervical squamous lesions.

Is HER2 ultra-low breast cancer different from HER2 null or HER2 low breast cancer? A study of 1363 patients.

OBJECTIVE: This study aims to analyze whether there are any differences in clinicopathological features and prognosis between HER2 ultra-low, HER2-null, and HER2-low expression in Chinese breast cancer (BC) patients. METHODS: The clinicopathological data of 1363 HER2-negative BC patients were retrospectively collected (from January 2018 to December 2019). HER2 status was further classified into HER2-null, HER2 ultra-low, and HER2-low. HER2-null expression is defined as infiltrating cancer cells completely free of staining. HER2 ultra-low expression is defined as ≤10% of infiltrating cancer cells showing incomplete and faint/weak membrane staining. HER2-low expression is defined as HER2 immunohistochemistry (IHC) 1+ or 2+ with negative in situ hybridization (ISH) assay. RESULTS: Of 1363 patients, there were 86 (6.3%) HER2-null patients, 395 (29.0%) HER2 ultra-low patients, and 882 (64.7%) HER2-low patients. HER2 ultra-low patients were different from HER2-low patients in terms of N stage, hormone receptor (HR) status, Ki-67 expression, and type of surgery. There were also significant differences in histologic type and postoperative endocrine therapy between HER2 ultra-low and HER2-null patients. HR+ (81.0%) tumors was more common than HR- (19.0%) in HER2 ultra-low patients. In addition, there was a significant difference in HR status between HER2 ultra-low and HER2-low patients (P = 0.001). The survival analysis showed that HER2 status had no effect on disease-free survival (DFS) in HER2-negative patients (all P > 0.05). However, regardless of HER2 status, HR+ patients had better DFS than HR- patients (P = 0.003). Cox multivariate analysis revealed that age (HR [95% CI] = 0.950 [0.928, 0.972], P < 0.001), HR status (HR [95% CI] = 3.342 [1.658, 6.736], P = 0.001), and postoperative endocrine therapy (HR [95% CI] = 0.048 [0.048, 0.023], P < 0.001) were important influencing factors of DFS in HER2-negative BC patients. CONCLUSION: HER2 ultra-low BC patients demonstrated distinct clinicopathological features from HER2-null and HER2-low tumors; while, HER2 status (null, ultra-low, or low) had no prognostic value in these HER2-negative BC population. Consistent with the published literature, HR status was an independent prognostic factor for DFS in HER2-negative BC patients.

The Role of Artificial Intelligence in Accurate Interpretation of HER2 Immunohistochemical Scores 0 and 1+ in Breast Cancer.

The new human epidermal growth factor receptor (HER)2-targeting antibody-drug conjugate offers the opportunity to treat patients with HER2-low breast cancer. Distinguishing HER2 immunohistochemical (IHC) scores of 0 and 1+ is not only critical but also challenging owing to HER2 heterogeneity and variability of observers. In this study, we aimed to increase the interpretation accuracy and consistency of HER2 IHC 0 and 1+ evaluation through assistance from an artificial intelligence (AI) algorithm. In addition, we examined the value of our AI algorithm in evaluating HER2 IHC scores in tumors with heterogeneity. AI-assisted interpretation consisted of AI algorithms and an augmenting reality module with a microscope. Fifteen pathologists (5 junior, 5 midlevel, and 5 senior) participated in this multi-institutional 2-round ring study that included 246 infiltrating duct carcinoma cases that were not otherwise specified. In round 1, pathologists analyzed 246 HER2 IHC slides by microscope without AI assistance. After a 2-week washout period, the pathologists read the same slides with AI algorithm assistance and rendered the definitive results by adjusting to the AI algorithm. The accuracy of interpretation accuracy with AI assistance (0.93 vs 0.80), thereby the evaluation precision of HER2 0 and the recall of HER2 1+. In addition, the AI algorithm improved the total consistency (intraclass correlation coefficient = 0.542-0.812), especially in HER2 1+ cases. In cases with heterogeneity, accuracy improved significantly (0.68 to 0.89) and to a similar level as in cases without heterogeneity (accuracy, 0.97). Both accuracy and consistency improved more for junior pathologists than those for the midlevel and senior pathologists. To the best of our knowledge, this is the first study to show that the accuracy and consistency of HER2 IHC 0 and 1+ evaluation and the accuracy of HER2 IHC evaluation in breast cancers with heterogeneity can be significantly improved using AI-assisted interpretation.

Low-grade adenosquamous carcinoma of the breast: a clinical, morphological and immunohistochemical analysis of 25 patients.

AIMS: Due to its rarity and non-specific clinical and pathological features, low-grade adenosquamous carcinoma (LGASC) of the breast continues to pose diagnostic challenges. Unlike other triple-negative breast carcinomas, LGASC tends to have an indolent clinical behaviour. It is essential to recognise this lesion for accurate diagnosis and appropriate management. METHODS AND RESULTS: Twenty-five cases of LGASC were identified in our archives and collaborating institutes. Cases of LGASC with dominant coexisting other type carcinomas were excluded. We studied the clinical presentation, morphological features, patterns of the commonly used immunohistochemical stains and follow-up. In our cohort, LGASC was commonly located at the outer aspect of the breast and associated with intraductal papilloma. The morphology of LGASC is characterised by infiltrating small glands and nests with variable squamous differentiation. We also found cuffing desmoplastic (fibrolamellar) stromal change in 75% of patients and peripheral lymphocytic aggregates in 87.5% of patients. P63 and smooth muscle myosin (SMM) were the most common myoepithelial markers used to assist in diagnosis. P63 often stained peripheral tumour cells surrounding invasive glands (circumferential staining in 80% of the cases), mimicking myoepithelial cells. It also stained the small nests with squamous differentiation. However, SMM was negative in 63% of the cases. The vast majority of our cases were triple-negative; only a few had focal and weak expressions of ER and PR. One patient who did not have excision developed lymph node metastasis. Most patients underwent excision or mastectomy with negative margins as surgical treatment; there were no recurrences or metastases in these patients with clinical follow-ups up to 108 months. CONCLUSIONS: LGASC has some unique, although not entirely specific, morphological features and immunohistochemical staining patterns. Fibrolamellar stromal change, peripheral lymphocytic aggregates and variable staining of p63 and SMM are valuable features to facilitate the diagnosis.

PRDX1 negatively regulates bleomycin-induced pulmonary fibrosis via inhibiting the epithelial-mesenchymal transition and lung fibroblast proliferation in vitro and in vivo.

BACKGROUND: Pulmonary fibrosis is a major category of end-stage changes in lung diseases, characterized by lung epithelial cell damage, proliferation of fibroblasts, and accumulation of extracellular matrix. Peroxiredoxin 1 (PRDX1), a member of the peroxiredoxin protein family, participates in the regulation of the levels of reactive oxygen species in cells and various other physiological activities, as well as the occurrence and development of diseases by functioning as a chaperonin. METHODS: Experimental methods including MTT assay, morphological observation of fibrosis, wound healing assay, fluorescence microscopy, flow cytometry, ELISA, western blot, transcriptome sequencing, and histopathological analysis were used in this study. RESULTS: PRDX1 knockdown increased ROS levels in lung epithelial cells and promoted epithelial-mesenchymal transition (EMT) through the PI3K/Akt and JNK/Smad signalling pathways. PRDX1 knockout significantly increased TGF-ß secretion, ROS production, and cell migration in primary lung fibroblasts. PRDX1 deficiency also increased cell proliferation, cell cycle circulation, and fibrosis progression through the PI3K/Akt and JNK/Smad signalling pathways. BLM treatment induced more severe pulmonary fibrosis in PRDX1-knockout mice, mainly through the PI3K/Akt and JNK/Smad signalling pathways. CONCLUSIONS: Our findings strongly suggest that PRDX1 is a key molecule in BLM-induced lung fibrosis progression and acts through modulating EMT and lung fibroblast proliferation; therefore, it may be a therapeutic target for the treatment of BLM-induced lung fibrosis.

Determination of rebamipide in human plasma by a validated liquid chromatography-tandem mass spectrometry: Application in pharmacokinetics research.

A new method for the determination of rebamipide in human heparin sodium plasma by LC-MS was established and its methodology was validated. In this method, protein precipitation method was used to pretreat the samples and the rebamipide-d4 isotope of rebamipide was used as the internal standard. In the multi reaction monitoring mode, the electrospray ion source was used as the ionization technolog and LC-MS was used for detection and analysis. The liquid chromatographic conditions were: 00B-4605-AN (Kinetex® XB-C18 100A 50mm × 2.1mm, 5µm); mobile phase A: 0.1% FA and 1 mM NH4FA aqueous solution, mobile phase B: 0.1% FA and 1mM NH4FA 90% ACN solution, flow rate: 0.300mL/min, injection volume: 10uL, column temperature: 30oC, collection time: 3 min, injector temperature control: 5oC. The retention time of rebamipide and rebamipide-d4 were 1.32min and 1.31min, respectively. The lower limit of quantification was 1ng/mL and the calibration map of rebamipide in the concentration range of 1 to 800ng/mL was linear (R2 >0.990, n=11). The CV% values of the inter and intra batch precision of the method were both less than 15.0%. This method has been successfully applied to pharmacokinetic studies to evaluate the main pharmacokinetic parameters of rebamipide.

A validated method for the determination of quetiapine fumarate tablets in human plasma by UPLC-MS/MS and its application to a pharmacokinetic study in healthy Chinese subjects.

A rapid, highly specific and sensitive UPLC-MS/MS method was developed for the determination of Quetiapine Fumarate, a therapeutic drug for various psychiatric disorders, in human plasma. The samples were pretreated using a protein precipitation method, followed by chromatographic separation using a column (Kinetex C18, 2.6µm 50*2.1mm) equipped with an ESI source and MRM mode mass spectrometer. In the validation results of the method, the analyte quetiapine showed a peak at approximately 1.0 minute and exhibited good linearity within the concentration from 2.5 to 2000ng/mL. The intra- and inter-batch precision CV% were within the range of -1.3% to 7.7% and precision of intra- and inter-batch were below 15.0%. Furthermore, this method demonstrated low matrix effects and high recovery rates. The quetiapine plasma sample solution remained stable at room temperature for 25 hours and following 4 freeze-thaw cycles. The prepared samples remained stable in the autosampler (The temperature control of the autosampler was 5oC) for 185 hours and after four freeze-thaw cycles at -20oC and -70oC for 40 days. The present work effectively employed this approach to investigate the pharmacokinetics of orally administered quetiapine fumarate tablets in a cohort of healthy Chinese individuals, both in a fasting state and after a meal.

HER2-low breast cancers: incidence, HER2 staining patterns, clinicopathologic features, MammaPrint and BluePrint genomic profiles.

Recently, clinical trials have demonstrated promising efficacy for novel HER2-targeted therapies in HER2-low breast cancers, raising the prospect of including a HER2-low category (immunohistochemical [IHC] score of 1+, or 2+ with non-amplified in-situ hybridization [ISH]) in the HER2 evaluation of breast cancers. In order to better understand this newly-proposed HER2 category, we investigated the incidence, HER2 staining patterns, clinicopathologic features, and genomic profile of HER2-low breast cancers. HER2-stained slides of 281 consecutive breast cancers were re-reviewed and the clinicopathologic information, MammaPrint, and BluePrint results of these cases were retrospectively analyzed. HER2-low breast cancers were identified in 31% of cases and were more common in estrogen receptor (ER)-positive than ER-negative breast cancers (33.6% vs 15%, p = 0.017). HER2-low cancers were generally clinical stages I-II (79%), ER-positive (93.1%), had homogenous HER2 staining (59.2%), HER2 IHC score of 1+ (87.4%), ductal phenotype (81.6%), histologic grades of 1 or 2 (94.2%) and luminal molecular subtypes (94.3%). Three HER2-low patients received neoadjuvant chemotherapy and none of them achieved pathologic complete response. When compared to HER2-negative (IHC of 0+) and HER2-positive (IHC of 3+ or IHC of 2+ with amplified ISH) cancers, HER2-low breast cancers had significantly lower Ki-67 (p = 0.03 and p < 0.01, respectively) and higher ER positivity (p = 0.01 and p = 0.03, respectively). HER2-low breast cancers were less likely to be basal molecular subtype when compared to HER2-negative cancers (p < 0.01) and were less likely to have a HER2 molecular subtype when compared to HER2-positive cancers (p < 0.01). When adjusted for ER status, there was no significant difference on all the examined variables between HER2-low and HER2-negative groups. Our study provides valuable baseline characteristics of HER2-low breast cancers deriving from consecutive, real-world cases with a consensus confirmation of HER2 status, and would help to increase our understanding of this newly-proposed HER2 category in breast cancers.

Downregulation of NEDD4L by EGFR signaling promotes the development of lung adenocarcinoma.

Cumulative evidence indicates that the abnormal regulation of the NEDD4 family of E3-ubiquitin ligases participates in the tumorigenesis and development of cancer. However, their role in lung adenocarcinoma (LUAD) remains unclear. This study comprehensively analyzed the NEDD4 family in LUAD data sets from public databases and found only NEDD4L was associated with the overall survival of LUAD patients. Gene set enrichment analysis (GSEA) indicated that NEDD4L might be involved in the regulation of mTORC1 pathway. Both cytological and clinical assays showed that NEDD4L inhibited the activity of the mTOR signaling pathway. In vivo and in vitro experiments showed that NEDD4L could significantly inhibit the proliferation of LUAD cells. In addition, this study also found that the expression of NEDD4L was regulated by EGFR signaling. These findings firstly revealed that NEDD4L mediates an interplay between EGFR and mTOR pathways in LUAD, and suggest that NEDD4L held great potential as a novel biomarker and therapeutic target for LUAD.

Associations of RBC and Serum Folate Concentrations with Cervical Intraepithelial Neoplasia and High-Risk Human Papillomavirus Genotypes in Female Chinese Adults.

BACKGROUND: Although folate status is associated with cervical carcinogenesis, it is not clear whether folate deficiency is associated with risk of cervical intraepithelial neoplasia (CIN) progression and infection with high-risk human-papillomavirus (hrHPV). OBJECTIVES: To evaluate the associations of RBC and serum folate concentrations with prevalence of CIN grades and hrHPV infection, their interactions with prevalence of CIN grades, and RBC folate with the risk of CIN1 progressing to CIN2. METHODS: Using data from the Shanxi CIN cohort of 2304 female Chinese adults, we used logistic-regression model to estimate ORs and prevalence ratios (PRs) of RBC and serum folate concentrations with prevalence of CIN grades and hrHPV infection. Categoric and spline analyses were used to evaluate the dose-response relations. We estimated the association of RBC folate with risk of CIN1 progressing to CIN2 in the nested case-control cohort. RESULTS: An inverse association was observed between increased RBC folate concentration and the odds of all CIN grades [quartile 1 (Q1) compared with Q4: OR: 2.28; 95% CI: 1.77, 2.93; Ptrend < 0.001]. Significant interaction of RBC folate and hrHPV infection was observed for prevalence of CIN2 or above (Pinteraction < 0.01). No associations were found between RBC and serum folate with PRs of hrHPV in each CIN grade. Over a median follow-up of 21.0 mo, RBC folate was associated with increased risk of CIN1 progressing to CIN2 (Q1 compared with Q4: OR: 3.86; 95% CI: 1.01, 14.76). CONCLUSIONS: Our study indicates that RBC folate concentration is associated with prevalence of CIN grades and CIN1 progression in female Chinese adults. Maintenance of normal folate status is important for reducing the risk of CIN and its progression in women with or without hrHPV infection.

miRNA-Mediated Suppression of a Cardioprotective Cardiokine as a Novel Mechanism Exacerbating Post-MI Remodeling by Sleep Breathing Disorders.

RATIONALE: Obstructive sleep apnea-hypopnea syndrome, a sleep breathing disorder in which chronic intermittent hypoxia (CIH) is the primary pathology, is associated with multiple cardiovascular diseases. However, whether and how CIH may affect cardiac remodeling following myocardial infarction (MI) remains unknown. OBJECTIVE: To determine whether CIH exposure at different periods of MI may exacerbate post-MI heart failure and to identify the mechanisms underlying CIH-exacerbated post-MI remodeling. METHODS AND RESULTS: Adult male mice were subjected to MI (4 weeks) with and without CIH (4 or 8 weeks). CIH before MI (CIH+MI) had no significant effect on post-MI remodeling. However, double CIH exposure (CIH+MI+CIH) or CIH only during the MI period (MI+CIH) significantly exacerbated pathological remodeling and reduced survival rate. Mechanistically, CIH activated TGF-ß (tumor growth factor-ß)/Smad (homologs of both the Drosophila protein MAD and the C. elegans protein SMA) signaling and enhanced cardiac epithelial to mesenchymal transition, markedly increasing post-MI cardiac fibrosis. Transcriptome analysis revealed that, among 15 genes significantly downregulated (MI+CIH versus MI), Ctrp9 (a novel cardioprotective cardiokine) was one of the most significantly inhibited genes. Real-time polymerase chain reaction/Western analysis confirmed that cardiomyocyte CTRP9 expression was significantly reduced in MI+CIH mice. RNA-sequencing, real-time polymerase chain reaction, and dual-luciferase reporter assays identified that microRNA-214-3p is a novel Ctrp9 targeting miRNA. Its upregulation is responsible for Ctrp9 gene suppression in MI+CIH. Finally, AAV9 (adeno-associated virus 9)-mediated cardiac-specific CTRP9 overexpression or rCTRP9 (recombinated CTRP9) administration inhibited TGF-ß/Smad and Wnt/ß-catenin pathways, attenuated interstitial fibrosis, improved cardiac function, and enhanced survival rate in MI+CIH animals. CONCLUSIONS: This study provides the first evidence that MI+CIH upregulates miR-214-3p, suppresses cardiac CTRP9 (C1q tumor necrosis factor-related protein-9) expression, and exacerbates cardiac remodeling, suggesting that CTRP9 may be a novel therapeutic target against pathological remodeling in MI patients with obstructive sleep apnea-hypopnea syndrome.

Oxidation Flow Reactor Results in a Chinese Megacity Emphasize the Important Contribution of S/IVOCs to Ambient SOA Formation.

Oxygenated volatile organic compounds (OVOCs) and secondary organic aerosol (SOA) formation potential of ambient air in Guangzhou, China was investigated using a field-deployed oxidation flow reactor (OFR). The OFR was used to mimic hours to weeks of atmospheric exposure to hydroxyl (OH) radicals within the 2-3 min residence time. A comprehensive investigation on the variation of VOCs and OVOCs as a function of OH exposure is shown. Substantial formation of organic acids and nitrogen-containing OVOC species were observed. Maximum SOA formation in the OFR was observed following 1-4 equiv days' OH exposure. SOA produced from known/measured VOC/IVOC precursors such as single-ring aromatics and long-chain alkanes can account for 52-75% of measured SOA under low NOx and 26-60% under high NOx conditions based on laboratory SOA yield parametrizations. To our knowledge, this is the first time that the contribution (8-20%) of long-chain (C8-C20) alkane oxidation to OFR SOA formation was quantified from direct measurement. By additionally estimating contribution from unmeasured semivolatile and intermediate volatility compounds (S/IVOCs) that are committed with C8-C20 alkanes, 64-100% of the SOA formation observed in the OFR can be explained, signifying the important contribution of S/IVOCs such as large cyclic alkanes to ambient SOA.

Berberine Reverses Nitroglycerin Tolerance through Suppressing Protein Kinase C Alpha Activity in Vascular Smooth Muscle Cells.

PURPOSE: The aim of this study was to evaluate the effects of berberine on nitroglycerin (NTG) tolerance and explore the underlying mechanism involved. METHODS: NTG tolerance was induced by pre-exposure of Sprague-Dawley rat aortas to NTG in vitro or by pretreating Sprague-Dawley rats with an NTG patch in vivo. The aortas were pre-treated with berberine or PKC inhibitors for different durations of time before induction of NTG tolerance. NTG-induced vasorelaxations was measured on wire myograph. Primary vascular smooth cells (VSMCs) were used to dissect the underlying mechanism of berberine-induced inhibition of NTG tolerance. RESULTS: NTG tolerance induced by either prior exposure of rat aortas to NTG in vitro or pretreatment with an NTG patch in vivo was reversed by co-treatment with berberine, as well as the inhibitors of protein kinase C (PKC) and protein kinase C alpha (PKCα). The mechanistic study revealed that PKCα participated in the development of NTG tolerance as NTG increased the activity of PKCα with enriched PKCα membrane localization and elevated phosphorylation of PKCα in VSMCs, which was reversed by berberine or PKCα inhibitors. CONCLUSION: This study is probably the first demonstration that berberine reverses NTG tolerance through inhibiting PKCα activity in VSMCs and PKCα is an important contributor to the development of NTG tolerance. These new findings suggest that berberine could become a promising drug for prevention of NTG tolerance and that targeting PKCα in VSMCs is likely to be a potential therapeutic strategy for reversal of NTG tolerance in blood vessels.

Mirabegron Ameliorated Atherosclerosis of ApoE-/- Mice in Chronic Intermittent Hypoxia but Not in Normoxia.

PURPOSE: It has been established that obstructive sleep apnea (OSA) is an independent risk factor for atherosclerosis. Chronic intermittent hypoxia (CIH) activates sympathoadrenal system and upregulates ß3 adrenergic receptor (ß3 AR). However, the effect of selective ß3 AR agonist mirabegron in CIH-induced atherosclerosis remains unknown. METHODS: We generated a CIH-induced atherosclerosis model through exposing ApoE-/- mice to CIH (8 h per day, cyclic inspiratory oxygen fraction 5-21%, 60-s cycle) for 6 weeks after 4-week high-fat dieting and investigated the effects of mirabegron, a selective ß3 AR agonist, on CIH-induced atherosclerosis. The coronary endarterectomy (CE) specimens from coronary artery disease patients with OSA and without OSA were collected. RESULTS: The expression of ß3 AR was significantly elevated in CIH-induced atherosclerosis model. Furthermore, treatment with mirabegron (10mg/kg per day by oral administration for 6 weeks) ameliorated atherosclerosis in ApoE-/- mice in CIH but not in normoxia. Mechanistically, mirabegron activated ß3 AR and ameliorated intraplaque oxidative stress by suppressing p22phox expression and reactive oxygen species (ROS) level. In addition, in human CE specimens, ß3 AR was also upregulated associated with increased p22phox expression and ROS level both in the lumen and in the plaque of coronary artery in OSA subjects. CONCLUSION: This study first demonstrated that mirabegron impeded the progression of CIH-induced atherosclerosis, at least in part, via ß3 AR-mediated oxidative stress, suggesting a promising therapeutic strategy for protecting against atherosclerosis induced by CIH.