Phosphorylation of EZH2 by AMPK Suppresses PRC2 Methyltransferase Activity and Oncogenic Function.

Sustained energy starvation leads to activation of AMP-activated protein kinase (AMPK), which coordinates energy status with numerous cellular processes including metabolism, protein synthesis, and autophagy. Here, we report that AMPK phosphorylates the histone methyltransferase EZH2 at T311 to disrupt the interaction between EZH2 and SUZ12, another core component of the polycomb repressive complex 2 (PRC2), leading to attenuated PRC2-dependent methylation of histone H3 at Lys27. As such, PRC2 target genes, many of which are known tumor suppressors, were upregulated upon T311-EZH2 phosphorylation, which suppressed tumor cell growth both in cell culture and mouse xenografts. Pathologically, immunohistochemical analyses uncovered a positive correlation between AMPK activity and pT311-EZH2, and higher pT311-EZH2 correlates with better survival in both ovarian and breast cancer patients. Our finding suggests that AMPK agonists might be promising sensitizers for EZH2-targeting cancer therapies.

Stretchable ionic-electronic bilayer hydrogel electronics enable in situ detection of solid-state epidermal biomarkers.

Continuous and in situ detection of biomarkers in biofluids (for example, sweat) can provide critical health data but is limited by biofluid accessibility. Here we report a sensor design that enables in situ detection of solid-state biomarkers ubiquitously present on human skin. We deploy an ionic-electronic bilayer hydrogel to facilitate the sequential dissolution, diffusion and electrochemical reaction of solid-state analytes. We demonstrate continuous monitoring of water-soluble analytes (for example, solid lactate) and water-insoluble analytes (for example, solid cholesterol) with ultralow detection limits of 0.51 and 0.26 nmol cm-2, respectively. Additionally, the bilayer hydrogel electrochemical interface reduces motion artefacts by a factor of three compared with conventional liquid-sensing electrochemical interfaces. In a clinical study, solid-state epidermal biomarkers measured by our stretchable wearable sensors showed a high correlation with biomarkers in human blood and dynamically correlated with physiological activities. These results present routes to universal platforms for biomarker monitoring without the need for biofluid acquisition.

FK228 suppress the growth of human malignant pleural mesothelioma tumor independent to epithelioid or non-epithelioid histology.

Human malignant pleural mesothelioma (hMPM) is an aggressive, rare disease with a poor prognosis. Histologically, MPM is categorized into epithelioid, biphasic, and sarcomatoid subtypes, with the epithelioid subtype generally displaying a better response to treatment. Conversely, effective therapies for the non-epithelioid subtypes are limited. This study aimed to investigate the potential role of FK228, a histone deacetylase inhibitor, in the suppression of hMPM tumor growth. We conducted a comprehensive analysis of the histological and molecular characteristics of two MPM cell lines, CRL-5820 (epithelioid) and CRL-5946 (non-epithelioid). CRL-5946 cells and non-epithelioid patient-derived xenografted mice exhibited heightened growth rates compared to those with epithelioid MPM. Both CRL-5946 cells and non-epithelioid mice displayed a poor response to cisplatin. However, FK228 markedly inhibited the growth of both epithelioid and non-epithelioid tumor cells in vitro and in vivo. Cell cycle analysis revealed FK228-induced G1/S and mitotic arrest in MPM cells. Caspase inhibitor experiments demonstrated that FK228-triggered apoptosis occurred via a caspase-dependent pathway in CRL-5946 but not in CRL-5820 cells. Additionally, a cytokine array analysis showed that FK228 reduced the release of growth factors, including platelet-derived and vascular endothelial growth factors, specifically in CRL-5946 cells. These results indicate that FK228 exhibits therapeutic potential in MPM by inducing cytotoxicity and modulating the tumor microenvironment, potentially benefiting both epithelioid and non-epithelioid subtypes.

Characterization of Mammary Tumors Arising from MMTV-PyVT Transgenic Mice.

Among genetically engineered mouse models of breast cancer, MMTV-PyVT is a mouse strain in which the oncogenic polyoma virus middle T antigen is driven by the mouse mammary tumor virus promoter. The aim of the present study was to perform morphologic and genetic analyses of mammary tumors arising from MMTV-PyVT mice. To this end, mammary tumors were obtained at 6, 9, 12, and 16 weeks of age for histology and whole-mount analyses. We conducted whole-exome sequencing to identify constitutional and tumor-specific mutations, and genetic variants were identified using the GRCm38/mm10 mouse reference genome. Using hematoxylin and eosin analysis and whole-mount carmine alum staining, we demonstrated the progressive proliferation and invasion of mammary tumors. Frameshift insertions/deletions (indels) were noted in the Muc4. Mammary tumors showed small indels and nonsynonymous single-nucleotide variants but no somatic structural alterations or copy number variations. In summary, we validated MMTV-PyVT transgenic mice as a multistage model for mammary carcinoma development and progression. Our characterization may be used as a reference for guidance in future research.

Radiotherapy versus low-dose tamoxifen following breast-conserving surgery for low-risk and estrogen receptor-positive breast ductal carcinoma in situ: an international open-label randomized non-inferiority trial (TBCC-ARO DCIS Trial).

BACKGROUND: Radiotherapy (RT) following breast-conserving surgery (BCS) is mainly used to decrease the rate of ipsilateral breast tumor recurrence (IBTR) in women with breast ductal carcinoma in situ (DCIS). Recent studies have demonstrated that low-dose tamoxifen significantly reduces IBTR in breast DCIS. Here, we aim to determine whether the administration of low-dose tamoxifen is non-inferior to RT in preventing IBTR in patients with low-risk characteristics of breast DCIS. METHODS/DESIGN: This is a prospective, international, open-label, randomized, non-inferiority trial. Patients with low-risk clinicopathologic features (> 40 years old, low risk of breast cancer susceptibility gene (BRCA) 1 and BRCA2 mutations, mammographically detected unicentric and non-mass lesions, low- or intermediate-grade without comedo or necrosis, measuring < 2.5 cm with margins ≥ 3 mm, and estrogen receptor-positive status) of DCIS who underwent BCS will be randomized at a 1:1 ratio to either receive tamoxifen (5 mg/day) for 5 years or undergo RT with conventional fractions (50 Gy in 25 fractions) or hypofractionations (40.05 Gy in 15 fractions). Randomization will be stratified by the Taiwan Breast Cancer Consortium. As approximately 5% of patients cannot tolerate the side effects of low-dose tamoxifen and will receive RT, we estimate that 405 patients will be randomized to a low-dose tamoxifen arm and 405 patients to the RT arm, according to a non-inferiority margin within 5% of IBTR difference and 90% ß-power noticing non-inferiority. The primary endpoints are breast tumor recurrence, including ipsilateral, regional, contralateral, and distant recurrence of breast DCIS or invasive cancer. The secondary endpoints are overall survival and adverse effects of RT and tamoxifen. Translational studies will also be conducted for this trial. DISCUSSION: This is the first non-inferiority trial on breast DCIS. This study will provide an important recommendation for clinical physicians on whether to use low-dose adjuvant tamoxifen for patients with low-risk breast DCIS who do not want to receive adjuvant RT. TRIAL REGISTRATION: ClinicalTrials.gov, ID: NCT04046159, Registered on April 30, 2019.

Preparation of an electrochemical sensor utilizing graphene-like biochar for the detection of tetracycline.

Tetracycline (TC), which is ubiquitous in the aquatic environment, can cause ecological imbalance and adversely affect human health. Therefore, a quick, inexpensive, and easy method for the detection of TC in water systems is highly desirable. This study reports the development of a novel electrochemical sensor from waste peanut shell for the quick detection of TC in water. Raman and TEM lattice mapping analyses confirmed the successful preparation of graphene -like biochar from waste peanut shells (PSs) via hydrothermal and pyrolysis processes. An electrochemical sensor, PS@glassy carbon electrode (PS@GCE), was then developed by coating the prepared graphene-like biochar on the surface of a glass electrode to enhance its conductivity. The feasibility of using this sensor for the detection of TC in the aqueous system was investigated. The PS@GCE sensor exhibited excellent sensitivity with a low detection limit of 3.6 × 10--9 nM and a linear range of 10-10-102 µM. These results were attributed to the large specific surface area and high conductivity, of the PS biochar. The stability of the PS@GCE sensor was also investigated in the presence of TC (10-4 M) and interfering species (10-2 M) and recovery rates in the range of 86.4%-116.0% were achieved, thus indicating the absence of an interference range of range of 84.3%-98.2% with relative standard deviation lower than 6% were achieved upon the detection of TC in natural water samples using the designed sensor, thus confirming the superior repeatability of the PS@GCE sensor. Consequently, the designed electrode has a high potential for application in the detection of TC in natural aqueous systems.

Afatinib alone and in combination with vinorelbine or paclitaxel, in patients with HER2-positive breast cancer who failed or progressed on prior trastuzumab and/or lapatinib (LUX-Breast 2): an open-label, multicenter, phase II trial.

PURPOSE: Resistance to HER2 (ErbB2)-targeted therapy may be mediated by other members of the ErbB family. We investigated the efficacy and safety of the irreversible ErbB family blocker, afatinib, alone as first-line therapy in the advanced setting and in combination with vinorelbine or paclitaxel for those who progressed on afatinib monotherapy, in female patients with metastatic breast cancer who had failed or progressed on prior HER2-targeted therapy in the early disease setting. METHODS: In this phase II, single-arm, two-part study (ClinicalTrials.gov: NCT01271725), patients in part A received afatinib 40 mg/day in 21-day cycles until disease progression or intolerable adverse events (AEs). Patients with progressive disease could then receive afatinib plus weekly vinorelbine 25 mg/m2 or paclitaxel 80 mg/m2 until disease progression or intolerable AEs (part B). The primary endpoint was confirmed objective response rate (RECIST v1.1). RESULTS: Eighty-seven patients were enrolled and 74 were treated in part A (median age: 51 years [range 27-76]; 31 [42%] estrogen receptor-positive, 26 [35%] progesterone receptor-positive). Of these, 39 (53%) patients went on to receive afatinib plus vinorelbine (13 patients) or paclitaxel (26 patients) in part B. Thirteen (18%) and 12 (31%) patients achieved an objective response in parts A and B, respectively. The most common treatment-related AEs with afatinib monotherapy (any/grade ≥ 3) were diarrhea (68%/8%) and rash (49%/4%). Combination therapy was generally well tolerated, with no additive toxicity observed. CONCLUSION: Afatinib treatment, alone or in combination with vinorelbine or paclitaxel, was associated with objective responses in ≥ 18% of patients with metastatic breast cancer for whom prior HER2-targeted therapy has failed. Treatment-related AEs were generally manageable, with few grade ≥ 3 AEs reported. TRIAL REGISTRATION: ClinicalTrials.gov, NCT01271725, registered 1 July 2011.

Impact of BRCA mutation on the survival and risk of contralateral breast cancer in Asian breast cancer patients.

PURPOSE: Breast cancer is increasing around the globe, including Asia. We aimed to examine the survival and risk of contralateral breast cancer (CBC) in Asian breast cancer patients with BRCA mutations. METHODS: A total of 128 breast cancer patients with germline BRCA mutations and 4,754 control breast cancer patients were enrolled. Data on clinical-pathologic characteristics, survival, and CBC were collected from the medical record. The rates of survival and CBC were estimated by Kaplan-Meier method. RESULTS: The mean age of onset in BRCA mutation carriers was significantly younger than control patients (BRCA vs. Non-BRCA: 43.9 vs. 53.2 years old). BRCA mutation carriers had a higher proportion of triple-negative breast cancer (TNBC) (52%) than control patients (12%, p < 0.001). The risk of CBC was significantly higher in BRCA mutation patients than in control cases (hazard ratio (HR) = 3.95, 95% CI 2.71-5.75); when stratified by genotype, the HRs (95%CI) were 4.84 (3.00-7.82) for BRCA1 and 3.13 (1.78-5.49) for BRCA2 carriers, respectively. Moreover, BRCA1 mutation patients with triple-negative breast cancer (TNBC) as their first breast cancer had the highest risk of CBC (HR = 5.55, 95% CI 3.29-9.34). However, we did not observe any differences in relapse-free survival and overall survival between mutation carriers and control patients. CONCLUSION: Our study suggest that BRCA patients had a significantly higher risk of developing CBC, particularly for BRCA1 mutation carriers with TNBC as the first breast cancer.

Responses of the Soil Bacterial Community, Resistome, and Mobilome to a Decade of Annual Exposure to Macrolide Antibiotics.

Biosolids that are applied to agricultural soil as an organic fertilizer are frequently contaminated with pharmaceutical residues that have persisted during wastewater treatment and partitioned into the organic phase. Macrolide antibiotics, which serve as a critically important human medicine, have been detected within biosolids. To determine the impacts of macrolide antibiotics on soil bacteria, every year for a decade, a series of replicated field plots received an application of a mixture of erythromycin, clarithromycin, and azithromycin at a realistic (0.1 mg kg soil-1) or an unrealistically high (10 mg kg soil-1) dose or were left untreated. The effects of repeated antibiotic exposure on the soil bacterial community, resistome, mobilome, and integron gene cassette content were evaluated by 16S rRNA and integron gene cassette amplicon sequencing, as well as whole-metagenome sequencing. At the unrealistically high dose, the overall diversity of the resistome and mobilome was altered, as 21 clinically important antibiotic resistance genes predicted to encode resistance to 10 different antibiotic drug classes were increased and 20 mobile genetic element variants (tnpA, intI1, tnpAN, and IS91) were increased. In contrast, at the realistic dose, no effect was observed on the overall diversity of the soil bacterial community, resistome, mobilome, or integron gene cassette-carrying genes. Overall, these results suggest that macrolide antibiotics entrained into soil at concentrations anticipated with biosolid applications would not result in major changes to these endpoints. IMPORTANCE Biosolids, produced from the treatment of sewage sludge, are rich in plant nutrients and are a valuable alternative to inorganic fertilizer when applied to agricultural soil. However, the use of biosolids in agriculture, which are frequently contaminated with pharmaceuticals, such as macrolide antibiotics, may pose a risk to human health by selecting for antibiotic resistance genes that could be transferred to plant-based food destined for human consumption. The consequences of long-term, repeated macrolide antibiotic exposure on the diversity of the soil bacterial community, resistome, and mobilome were evaluated. At unrealistically high concentrations, macrolide antibiotics alter the overall diversity of the resistome and mobilome, enriching for antibiotic resistance genes and mobile genetic elements of concern to human health. However, at realistic antibiotic concentrations, no effect on these endpoints was observed, suggesting that current biosolids land management practices are unlikely to pose a risk to human health due to macrolide antibiotic contamination alone.

Highly efficient magnetic ablation and the contrast of various imaging using biocompatible liquid-metal gallium.

BACKGROUND: Although the powerful clinical effects of radiofrequency and microwave ablation have been established, such ablation is associated with several limitations, including a small ablation size, a long ablation time, the few treatment positioning, and biosafety risks. To overcome these limitations, biosafe and efficient magnetic ablation was achieved in this study by using biocompatible liquid gallium as an ablation medium and a contrast medium for imaging. RESULTS: Magnetic fields with a frequency (f) lower than 200 kHz and an amplitude (H) × f value lower than 5.0 × 109 Am-1 s-1 were generated using the proposed method. These fields could generate an ablation size of 3 cm in rat liver lobes under a temperature of approximately 300 °C and a time of 20 s. The results of this study indicate that biomedical gallium can be used as a contrast medium for the positioning of gallium injections and the evaluation of ablated tissue around a target site. Liquid gallium can be used as an ablation medium and imaging contrast medium because of its stable retention in normal tissue for at least 3 days. Besides, the high anticancer potential of gallium ions was inferred from the self-degradation of 100 µL of liquid gallium after around 21 days of immersion in acidic solutions. CONCLUSIONS: The rapid wireless ablation of large or multiple lesions was achieved through the simple multi-injection of liquid gallium. This approach can replace the currently favoured procedure involving the use of multiple ablation probes, which is associated with limited benefits and several side effects. METHODS: Magnetic ablation was confirmed to be highly efficient by the consistent results obtained in the simulation and in vitro tests of gallium and iron oxide as well as the electromagnetic specifics and thermotherapy performance comparison detailed in this study Ultrasound imaging, X-ray imaging, and magnetic resonance imaging were found to be compatible with the proposed magnetic ablation method. Self-degradation analysis was conducted by mixing liquid gallium in acidic solutions with a pH of approximately 5-7 (to imitate a tumour-containing microenvironment). X-ray diffraction was used to identify the gallium oxides produced by degraded gallium ions.

Contamination of hay and haylage with enteric bacteria and selected antibiotic resistance genes following fertilization with dairy manure or biosolids.

The present study evaluated if enteric bacteria or antibiotic resistance genes carried in fecal amendments contaminate the hay at harvest, representing a potential route of exposure to ruminants that consume the hay. In the field experiments, dairy manure was applied to a hay field for three successive growing seasons, and biosolids were applied to a hay field for one growing season. Various enteric bacteria in the amendments were enumerated by viable plate count, and selected gene targets were quantified by qPCR. Key findings include the following: at harvest, hay receiving dairy manure or biosolids did not carry more viable enteric bacteria than hay from unamended control plots. The fermentation of hay did not result in a detectable increase in viable enteric bacteria. The application of dairy manure or biosolids resulted in a few gene targets being more abundant in hay during the first harvest. Fermentation of hay resulted in an increase in the abundance of gene targets, but this occurred with hay from both the amended and control plots. Overall, the application of fecal amendments resulted in an increase in the abundance of some gene targets associated with antibiotic resistance in the first cut hay.

In vitro infection of human ocular tissues by SARS-CoV-2 lineage A isolates.

BACKGROUND: The purpose of this study was: [1] to evaluate the infectivity of two SARS-CoV-2 lineage A variants on human ocular tissues in vitro, and [2] to evaluate the stability of SARS-CoV-2 lineage A variants in corneal preservation medium. METHODS: Primary cultures of donor corneal, conjunctival, and limbal epithelium were inoculated with two lineage A, GISAID clade S isolates of SARS-CoV-2 (Hong Kong/VM20001061/2020, USA-WA1/2020), to evaluate the susceptibility of the ocular tissue to infection. Flat-mounted Descemet's Stripping Automated Endothelial Keratoplasty (DSAEK) grafts were inoculated with SARS-CoV-2 to evaluate the susceptibility of the endothelium to infection. All inoculated samples were immunostained for SARS-CoV-2 nucleocapsid (N)-protein expression to confirm positive infection. SARS-CoV-2 Hong Kong was then inoculated into cornea preservation media (Life4°C, Numedis, Inc.). Inoculated media was stored at 4oC for 14 days and assayed over time for changes in infectious viral titers. RESULTS: Corneal, conjunctival, and limbal epithelial cells all demonstrated susceptibility to infection by SARS-CoV-2 lineage A variants. Conjunctiva demonstrated the highest infection rate (78% of samples infected [14/18]); however, infection rates did not differ statistically between cell types and viral isolates. After inoculation, 40% (4/10) of DSAEK grafts had active infection in the endothelium. SARS-CoV-2 lineage A demonstrated < 1 log decline in viral titers out to 14 days in corneal preservation media. CONCLUSIONS: SARS-CoV-2 lineage A variants can infect corneal, limbal, and conjunctival epithelium, as well as corneal endothelium. There was no statistical difference in infectivity between different lineage A variants. SARS-CoV-2 lineage A can survive and remain infectious in corneal preservation media out to 14 days in cold storage.

Risk factors associated with frailty in older adults with type 2 diabetes: A cross-sectional study.

AIMS AND OBJECTIVES: The aims of the study were to (i) estimate the prevalence of frailty among older adults with type 2 diabetes, and (ii) investigate risk factors associated with frailty in the study group. BACKGROUND: The high prevalence of frailty among older adults and its adverse health impacts including higher risks for physical disabilities and a higher mortality rate is quickly evolving to become an important public health issue. DESIGN: A cross-sectional design. METHODS: Data were collected from the Clinical Diabetes Centre of our study group. Frailty was assessed using the Chinese version of the Tilburg frailty indicator. Other questionnaires included Activities of Daily Living, Mini-Mental State Examination, Taiwan Geriatric Depression Scale and Instrumental Activities of Daily Living. Multivariable logistic regression was established to identify factors associated with frailty. The STrengthening the Reporting of OBservational studies in Epidemiology checklist was used preparing the manuscript. RESULTS: A total of 248 subjects were recruited in this study, a total of 66 participants had frailty (26.6%). Our results showed that factors including age, number of chronic diseases, cerebrovascular accident, renal diseases, frequency of falling, frequency of hyperglycaemia, Activities of Daily Living, depression, Mini-Mental State Examination, and Instrumental Activities of Daily Living ≧1 task disability were correlated with frailty in older adults with type 2 diabetes. After adjusting for confounding factors, multivariate logistic regression analyses showed a significant increase in the probability of frailty by 1.72-times when comparing type 2 diabetes subjects with depression to those without depression. CONCLUSION: This study revealed that depression is a significant factor associated with the occurrence of frailty among older adults with type 2 diabetes. RELEVANCE TO CLINICAL PRACTICE: Our results provided strong clinical evidence showing depression as an important factor associated with the risk of frailty in older individuals with type 2 diabetes. We recommend early detection via routine screenings for symptoms of depression during diabetes care to achieve early prevention or the delay of frailty and its implicated adverse health consequences.

Safety and efficacy of everolimus (EVE) plus exemestane (EXE) in postmenopausal women with locally advanced or metastatic breast cancer: final results from EVEREXES.

BACKGROUND: This study was conducted to collect clinical safety, tolerability, and efficacy data with the use of everolimus (EVE) combined with exemestane (EXE) in patients with advanced breast cancer (ABC). METHODS: The EVEREXES trial initiated in 2012, provided early access to the first dual blockade treatment with EVE + EXE in patients with HR+, HER2 - ABC in Asia and other emerging growth countries. Postmenopausal women with HR+, HER2 - ABC who had documented recurrence or progression, following a nonsteroidal aromatase inhibitor therapy, were treated with EVE (10 mg/day) + EXE (25 mg/day) orally. RESULTS: A total of 235 patients received ≥ 1 dose of study medication. At the end of the study, all patients ceased the treatment. Disease progression (66.0%) was the primary reason of discontinuation. The most common AEs (≥ 20%) were stomatitis, decreased appetite, hyperglycemia, rash, aspartate aminotransferase increased, anemia, alanine aminotransferase increased, cough, and fatigue. No new safety concerns were identified in the current study. Median progression-free survival (PFS) in the Asian subset was similar to that of the overall population (9.3 months in both groups). Confirmed overall response rate (ORR) was achieved for 19.6% of the patients. Efficacy of EVE + EXE across subgroups (prior CT, line of treatment, and presence of visceral metastases) was maintained. CONCLUSION: The safety and efficacy results from EVEREXES trial are consistent to data previously reported in BOLERO-2. These results support that EVE + EXE could be a viable treatment option for the postmenopausal women with HR+, HER2 - ABC in Asian region.

On-Farm Anaerobic Digestion of Dairy Manure Reduces the Abundance of Antibiotic Resistance-Associated Gene Targets and the Potential for Plasmid Transfer.

The present study investigated the impact of on-farm anaerobic digestion on the abundance of enteric bacteria, antibiotic resistance-associated gene targets, and the horizontal transfer potential of extended-spectrum ß-lactamase (ESBL) genes. Samples of raw and digested manure were obtained from six commercial dairy farms in Ontario, Canada. Digestion significantly abated populations of viable coliforms in all six farms. Conjugative transfer of plasmids carrying ß-lactamase genes from manure bacteria enriched overnight with buffered peptone containing 4 mg/liter cefotaxime into a ß-lactam-sensitive green fluorescent protein (GFP)-labeled Escherichia coli recipient strain was evaluated in patch matings. Digestion significantly decreased the frequency of the horizontal transfer of ESBL genes. Twenty-five transconjugants were sequenced, revealing six distinct plasmids, ranging in size from 40 to 180 kb. A variety of ESBL genes were identified: blaCTX-M-1, blaCTX-M-14, blaCTX-M-15, blaCTX-M-27, blaCTX-M-55, and blaPER-1. blaCTX-M-15 was the most prevalent ESBL gene detected on plasmids harbored by transconjugants. Various mobile genetic elements were found located proximal to resistance genes. Ten gene targets, including sul1, str(A), str(B), erm(B), erm(F), intI1, aadA, incW, blaPSE, and blaOXA-20, were quantified by quantitative PCR on a subset of 18 raw and 18 digested samples. Most targets were significantly more abundant in raw manure; however, erm(B) and erm(F) targets were more abundant in digested samples. Overall, on-farm digestion of dairy manure abated coliform bacteria, a number of antibiotic resistance-associated gene targets, and the potential for in vitro conjugation of plasmids conferring resistance to extended-spectrum ß-lactams and other classes of antibiotics into E. coli CV601. IMPORTANCE Using livestock manure for fertilization can entrain antibiotic-resistant bacteria into soil. Manure on some dairy farms is anaerobically digested before being land applied. Recommending the widespread implementation of the practice should be founded on understanding the impact of this treatment on various endpoints of human health concern. Although lab-scale anaerobic treatments have shown potential for reducing the abundance of antibiotic resistance genes, there are very few data from commercial farms. Anaerobic digestion of manure on six dairy farms efficiently abated coliform bacteria, E. coli, and a majority of antibiotic resistance-associated gene targets. In addition, the conjugation potential of plasmids carrying ESBL genes into introduced E. coli strain CV601 was reduced. Overall, anaerobic digestion abated coliform bacteria, the genes that they carry, and the potential for ESBL-carrying plasmid transfer.

Trilostane, a 3ß-hydroxysteroid dehydrogenase inhibitor, suppresses growth of hepatocellular carcinoma and enhances anti-cancer effects of sorafenib.

Background Human 3ß-hydroxysteroid dehydrogenase type 1 (HSD3B1) is an enzyme associated with steroidogenesis, however its' role in hepatocellular carcinoma (HCC) biology is unknown. Trilostane is an inhibitor of HSD3B1 and has been tested as a treatment for patients with breast cancer but has not been studied in patients with HCC. Methods and Results The expression of HSD3B1 in HCC tumors in 57 patients were examined. A total of 44 out of 57 tumors (77.2%) showed increased HSD3B1 expression. The increased HSD3B1 in tumors was significantly associated with advanced HCC. In vitro, the knockdown of HSD3B1 expression in Mahlavu HCC cells by a short hairpin RNA (shRNA) led to significant decreases in colony formation and cell migration. The suppression of clonogenicity in the HSD3B1-knockdown HCC cells was reversed by testosterone and 17ß-estradiol. Trilostane-mediated inhibition of HSD3B1 in different HCC cells also caused significant inhibition of clonogenicity and cell migration. In subcutaneous HCC Mahlavu xenografts, trilostane (30 or 60 mg/kg, intraperitoneal injection) significantly inhibited tumor growth in a dose-dependent manner. Furthermore, the combination of trilostane and sorafenib significantly enhanced the inhibition of clonogenicity and xenograft growth, surpassing the effects of each drug used alone, with no documented additional toxicity to animals. HSD3B1 blockade was found to suppress the phosphorylation of extracellular signal-regulated kinase (ERK). The decreased ERK phosphorylation was reversed by testosterone or 17b-estradiol. Conclusions Trilostane significantly inhibited the growth of HCC by inhibiting HSD3B1 function and augmenting the efficacy of sorafenib.

Overexpression of chitinase-3-like protein 1 is associated with structural recurrence in patients with differentiated thyroid cancer.

We previously identified that the expression of chitinase-3-like protein 1 (CHI3L1) was upregulated during thyroid cancer progression. Here, we investigated the prognostic significance of CHI3L1 expression in thyroid neoplasms and examined the potential oncogenic roles. CHI3L1 immunochemical staining was performed on tissue microarrays of benign and malignant thyroid tumours. Compared with normal thyroid tissue and benign thyroid lesions that had low or no detectable CHI3L1 expression, CHI3L1 was overexpressed in both differentiated and undifferentiated thyroid cancer. High CHI3L1 expression was associated with extrathyroidal extension, lymph node metastasis, and shorter recurrence-free survival in differentiated thyroid cancer. The biological roles of CHI3L1 were further investigated by gain- and loss-of-function assays. CHI3L1 silencing suppressed clonogenicity, migration, invasion, anoikis resistance, and angiogenesis in thyroid cancer cells, although exogenous CHI3L1 treatment promoted these malignant phenotypes. Cysteine-rich angiogenic inducer 61 (CYR61) was identified as a downstream target of CHI3L1 by RNA-seq analysis. CYR61 silencing or treatment reversed the alterations induced by CHI3L1 modulation. Our results demonstrate that CHI3L1 is overexpressed in thyroid cancer and is associated with an increased risk of disease recurrence. Additionally, CYR61 may participate in CHI3L1-mediated tumour progression. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Inhibition of 3ß-Hydroxysteroid Dehydrogenase Type 1 Suppresses Interleukin-6 in Breast Cancer.

BACKGROUND: Recently, we demonstrated that the expression of 3ß-hydroxysteroid dehydrogenase type 1 (HSD3B1) in breast cancer is associated with shorter recurrence-free survival, and genetic or pharmacologic inhibition of HSD3B1 reduced colony formation and xenograft growth. However, the mechanisms are unclear. METHODS: Triple-negative MDA-MB-231 and BT-20 breast cancer cells underwent HSD3B1 silencing. Microarray and bioinformatic analysis were performed. The interleukin-6 (IL-6) expression and secretion were evaluated using real-time quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. Clonogenic ability and cell viability were determined in the absence or presence of recombinant IL-6. RESULTS: Functional and pathway enrichment analyses showed that HSD3B1 silencing modulates the expression of several growth factors and cytokines. Cells transfected with HSD3B1-targeting small interfering RNA or treated with an HSD3B1 inhibitor (trilostane) had decreased IL-6 expression and secretion. HSD3B1 inhibition reduced colony formation, which was partially rescued by IL-6 supplementation. The HSD3B1 knockdown enhanced paclitaxel sensitivity, and IL-6 treatment partially reversed the augmented cytotoxicity. CONCLUSIONS: Our findings suggest that the therapeutic potential of targeting HSD3B1 is in part mediated by IL-6 suppression.

A preliminary report of head-to-head comparison of 18-gene-based clinical-genomic model and oncotype DX 21-gene assay for predicting recurrence of early-stage breast cancer.

BACKGROUND: The information of Oncotype DX applied in Asian breast cancer patients is limited. A recurrence index for distant recurrence (RI-DR) has been developed for early-stage breast cancer (EBC) from tumor samples in Chinese patients. In this study, we compared the prognostic performance of the Oncotype DX (ODx) recurrence score (RS) with the RI-DR for any recurrence risk type. MATERIALS AND METHODS: One hundred thirty-eight (138) patients with hormone receptor-positive and human epidermal growth factor receptor 2-negative EBC who were previously tested with ODx were included for testing with the RI-DR. The cutoff score to partition the low- and high-risk patients was 26 for RS and 36 for RI-DR. The primary endpoint was recurrence-free survival (RFS). RESULTS: The concordance between the RI-DR and RS was 83% in N0 patients and 81% in node-positive patients when the RS score cutoff was set at 26. With a median follow-up interval of 36.8 months, the 4-year RFS for the high- and low-risk groups categorized by the RS were 61.9% and 95.0%, respectively (hazard ratio: 10.6, 95.0% confidence interval [CI]: 1.8-62.9). The 4-year RFS in the high- and low-risk groups categorized by the RI-DR were 72.6% and 98.5%, respectively (hazard ratio: 18.9, 95% CI: 1.8-138.8). CONCLUSION: This paper illustrated the performance of RI-DR and ODx RS in breast cancer women in Taiwan. There was high concordance between the RI-DR and RS. The RI-DR is not inferior to the RS in predicting RFS in EBC patients. This study will fill the gap between the current and best practice in Chinese patients.

Specific drug delivery efficiently induced human breast tumor regression using a lipoplex by non-covalent association with anti-tumor antibodies.

BACKGROUND: A cationic liposome-PEG-PEI complex (LPPC) was employed as a carrier for achieving targeted delivery of drug to human epidermal growth factor receptor-2 (HER2/neu)-expressing breast cancer cells. LPPC can be easily loaded with an anti-tumor drug and non-covalently associated with an anti-tumor antibody such as Herceptin that is clinically used to rapidly form immunoparticles within 1 h. RESULTS: Drug-loaded LPPC have an average size about 250 nm and a zeta potential of about 40 mV. Herceptin was complexed onto surface of the LPPC to form the drug/LPPC/Herceptin complexes. The size of curcumin/LPPC/Herceptin complexes were 280 nm and the zeta potentials were about 23 mV. Targeting ability of this delivery system was demonstrated through specific binding on surface of cells and IVIS images in vivo, which showed specific binding in HER2-positive SKBR3 cells as compared to HER2-negative Hs578T cells. Only the drug/LPPC/Herceptin complexes displayed dramatically increased the cytotoxic activity in cancer cells. Both in vitro and in vivo results indicated that Herceptin adsorbed on LPPC directed the immunocomplex towards HER2/neu-positive cells but not HER2/neu-negative cells. The complexes with either component (curcumin or doxorubicin) used in the LPPC-delivery system provided a better therapeutic efficacy compared to the drug treatment alone and other treatment groups, including clinical dosages of Herceptin and LipoDox, in a xenografted model. CONCLUSIONS: LPPC displays important clinical implications by easily introducing a specific targeting characteristic to drugs utilized for breast cancer therapy.