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In this study, we examined the roles of anonymity and social closeness in predicting young women’s perceptions of “sex talk” (i.e., communication about sexual interests, enjoyment, and experiences) and intentions to post such content in cyberspace. We also examined cultural differences among Asian, Latina, and European Americans. A total of 466 undergraduate women from the three cultural groups participated in the online experiment. Participants were randomly assigned to either a low anonymity condition (i.e., Facebook) or a high anonymity condition (i.e., an anonymous online forum) where they were exposed to identical sex talk stimuli. The main findings showed that greater anonymity increased both the level of perceived appropriateness of sex talk posted by other female users and participants’ intentions to post sex talk online themselves. Compared to European American women, Asian and Latina Americans reported greater intentions to post sex talk online and perceived other female users’ sex talk posts as more appropriate. The results of this study prompt educators and practitioners to help young women strategically manage their impressions of sex talk online while being sensitive to women’s cultural backgrounds. They also suggest the need for further support from practitioners, educators, and parents to construct safe spaces for young women to engage open conversations about sexual matters in the digital space.

Chromobox 6 (CBX6) is a subunit of Polycomb Repressive Complex 1 (PRC1) that mediates epigenetic gene repression and acts as an oncogene or tumor suppressor in a cancer type-dependent manner. The specific function of CBX6 in breast cancer is currently undefined. In this study, a comprehensive analysis of The Cancer Genome Atlas (TCGA) dataset led to the identification of CBX6 as a consistently downregulated gene in breast cancer. We provided evidence showing enhancer of zeste homolog 2 (EZH2) negatively regulated CBX6 expression in a Polycomb Repressive Complex 2 (PRC2)-dependent manner. Exogenous overexpression of CBX6 inhibited cell proliferation and colony formation, and induced cell cycle arrest along with suppression of migration and invasion of breast cancer cells in vitro. Microarray analyses revealed that CBX6 governs a complex gene expression program. Moreover, CBX6 induced significant downregulation of bone marrow stromal cell antigen-2 (BST2), a potential therapeutic target, via interactions with its promoter region. Our collective findings support a tumor suppressor role of CBX6 in breast cancer.

The megalin/cubilin complex is responsible for the majority of serum protein reclamation in the proximal tubules. The current study examined if decreases in their renal expression, along with the albumin recycling protein neonatal Fc receptor (FcRn) could account for proteinuria/albuminuria in the Zucker diabetic fatty rat model of type 2 diabetes. Immunoblots of renal cortex samples obtained at worsening disease stages demonstrated no loss in megalin, cubilin, or FcRn, even when proteinuria was measured. Additionally, early diabetic rats exhibited significantly increased renal megalin expression when compared with controls (adjusted P < 0.01). Based on these results, the ability of insulin to increase megalin was examined in a clonal subpopulation of the opossum kidney proximal tubule cell line. Insulin treatments (24 h, 100 nM) under high glucose conditions significantly increased megalin protein ( P < 0.0001), mRNA ( P < 0.0001), and albumin endocytosis. The effect on megalin expression was prevented with inhibitors against key effectors of insulin intracellular signaling, phosphatidylinositide 3-kinase and Akt. Studies using rapamycin to inhibit the mechanistic target of rapamycin complex 1 (mTORC1) resulted in a loss of insulin-induced megalin expression. However, subsequent evaluation demonstrated these effects were independent of initial mTORC1 suppression. The presented results provide insight into the expression of megalin, cubilin, and FcRn in type 2 diabetes, which may be impacted by elevated insulin and glucose. Furthermore, proximal tubule endocytic activity in early diabetics may be enhanced, a process that could have a significant role in proteinuria-induced renal damage.

Monocarboxylate transporter 1 (MCT1) represents a potential therapeutic target in cancer. The objective of this study was to determine the efficacy of AZD3965 (a specific inhibitor of MCT1) and α-cyano-4-hydroxycinnamic acid (CHC, a nonspecific inhibitor of MCTs) in the murine 4T1 tumor model of triple-negative breast cancer (TNBC). Expression of MCT1 and MCT4 in 4T1 and mouse mammary epithelial cells were determined by Western blot. Inhibition of MCT1-mediated l-lactate uptake and cellular proliferation by AZD3965 and CHC was determined. Mice bearing 4T1 breast tumors were treated with AZD3965 100 mg/kg i.p. twice-daily or CHC 200 mg/kg i.p. once-daily. Tumor growth, metastasis, intra-tumor lactate concentration, immune function, tumor MCT expression, and concentration-effect relationships were determined. AZD3965 and CHC inhibited cell growth and l-lactate uptake in 4T1 cells. AZD3965 treatment resulted in trough plasma and tumor concentrations of 29.1 ± 13.9 and 1670 ± 946 nM, respectively. AZD3965 decreased the tumor proliferation biomarker Ki67 expression, increased intra-tumor lactate concentration, and decreased tumor volume, although tumor weight was not different from untreated controls. CHC had no effect on tumor volume and weight, or intra-tumor lactate concentration. AZD3965 treatment reduced the blood leukocyte count and spleen weight and increased lung metastasis, while CHC did not. These findings indicate AZD3965 is a potent MCT1 inhibitor that accumulates to high concentrations in 4T1 xenograft tumors, where it increases tumor lactate concentrations and produces beneficial effects on markers of TNBC; however, overall effects on tumor growth were minimal and lung metastases increased.

The mechanism underlying EZH2 overexpression in breast cancer and its involvement in tumorigenesis remain poorly understood. In this study, we developed an approach to systematically identify the trans-acting factors regulating the EZH2 expression, and identified more than 20 such factors. We revealed reciprocal regulation of early growth response 1 (EGR1) and EZH2: EGR1 activates the expression of EZH2, and EZH2 represses EGR1 expression. Using CRISPR-mediated genome/epigenome editing, we demonstrated that EHZ2 represses EGR1 expression through a silencer downstream of the EGR1 gene. Deletion of the EGR1 silencer resulted in reduced cell growth, invasion, tumorigenicity of breast cancer cells, and extensive changes in gene expression, such as upregulation of GADD45, DDIT3, and RND1; and downregulation of genes encoding cholesterol biosynthesis pathway enzymes. We hypothesize that EZH2/PRC2 acts as a "brake" for EGR1 expression by targeting the EGR1 silencer, and EZH2 overexpression dampens tumor-suppressive signals mediated by EGR1 to drive breast tumorigenesis.

PURPOSE: To evaluate the pharmacokinetics (PK) of the monocarboxylate transporter 1 (MCT1) inhibitor AZD3965 in mice after IV and oral administration and to develop mechanistic PK models to assess the potential enterohepatic circulation (EHC) and target-mediated drug disposition (TMDD) of AZD3965. METHODS: Female BALB/c mice were administered AZD3965 by IV injection (10, 50 and 100 mg/kg) or oral gavage (100 mg/kg). Plasma samples were analyzed using LC/MS/MS, and PK parameters determined by compartmental and non-compartmental analyses. RESULTS: AZD3965 exhibited a large volume of distribution and rapid oral absorption, with a high oral bioavailability. Prominent reentry peaks were observed after both oral and IV administration, suggesting potential EHC of AZD3965 or of a potential glucuronide conjugate. The dose-dependent studies indicated greater than proportional increases in exposure, an increase in the terminal half-life, and decrease in clearance and volume of distribution with increasing IV doses, indicating nonlinear pharmacokinetics and potential TMDD of AZD3965. Mechanistic compartmental models were developed to characterize the complex pharmacokinetics of AZD3965. CONCLUSIONS: The current study represents the first comprehensive report of the pharmacokinetics of AZD3965 in mice, indicating the potential contribution of EHC and TMDD in the disposition of AZD3965.

Background TET-mediated oxidation of 5-mC participates in both passive and active DNA demethylation, which exerts a significant influence on diverse biological processes. Mass spectrometry has identified multiple phosphorylation sites of TET2. However, the functions of these phosphosites and their corresponding kinases are mostly unknown. Results Here, we showed that AMP-activated protein kinase (AMPK) phosphorylates murine TET2 at the serine residue 97 (S97), and the phosphorylation enhances TET2 stability through promoting its binding to 14-3-3β. AMPK ablation resulted in decreased global 5-hmC levels at the myotube stages, severe differentiation defects of C2C12 cells and significantly, total loss of expression of Pax7. Genome-wide analyses revealed increased DNA methylation at genic and enhancer regions of AMPK-null myoblasts and myotubes. Using CRISPR/Cas9 technology, we showed that a novel enhancer, which is hypermethylated in AMPK-null cells, regulates Pax7 expression. The phospho-mimicking mutant, TET2-S97E, could partly rescue the differentiation defect in AMPK-ablated C2C12 cells. Conclusions Together, our data demonstrated that AMPK is a critical regulator of myogenesis, partly through phosphorylating TET2. Electronic supplementary material The online version of this article (10.1186/s13072-019-0281-x) contains supplementary material, which is available to authorized users.

AR-C155858 and AZD3965, pyrrole pyrimidine derivatives, represent potent monocarboxylate transporter 1 (MCT1) inhibitors, with potential immunomodulatory and chemotherapeutic properties. Currently, there is limited information on the inhibitory properties of this new class of MCT1 inhibitors. The purpose of this study was to characterize the concentration- and time-dependent inhibition of L-lactate transport and the membrane permeability properties of AR-C155858 and AZD3965 in the murine 4T1 breast tumor cells that express MCT1. Our results demonstrated time-dependent inhibition of L-lactate uptake by AR-C155858 and AZD3965 with maximal inhibition occurring after a 5-min pre-incubation period and prolonged inhibition. Following removal of AR-C155858 or AZD3965 from the incubation buffer, inhibition of L-lactate uptake was only fully reversed after 3 and 12 h, respectively, indicating that these inhibitors are slowly reversible. The uptake of AR-C155858 was concentration-dependent in 4T1 cells, whereas the uptake of AZD3965 exhibited no concentration dependence over the range of concentrations examined. The uptake kinetics of AR-C155858 was best fitted to a Michaelis-Menten equation with a diffusional clearance component, P (K(m) = 0.399 ± 0.067 μM, V(max) = 4.79 ± 0.58 pmol/mg/min, and P = 0.330 ± 0.088 μL/mg/min). AR-C155858 uptake, but not AZD3965 uptake, was significantly inhibited by alpha-cyano-4-hydroxycinnamic acid, a known nonspecific inhibitor of MCTs 1, 2, and 4. AR-C155858 demonstrated a trend toward higher uptake at lower pH, a characteristic of proton-dependent MCT1. These findings provide evidence that AR-C155858 and AZD3965 exert slowly reversible inhibition of MCT1-mediated L-lactate uptake in 4T1 cells, with AR-C155858 representing a potential substrate of MCT1.

Monocarboxylate transporter 1 (MCT1), also known as a l-lactate transporter, is a potential therapeutic target in cancer. The objectives of this study were to evaluate efficacy and assess concentration-effect relationships of AR-C155858 (a selective and potent MCT1 inhibitor) in murine 4T1 breast cancer cells and in the 4T1 tumor xenograft model. Western blotting of 4T1 cells demonstrated triple negative breast cancer (TNBC) characteristics and overexpression of MCT1 and CD147 (a MCT1 accessory protein), but absence of MCT4 expression. AR-C155858 inhibited the cellular l-lactate uptake and cellular proliferation at low nanomolar potencies (IC(50) values of 25.0 ± 4.2 and 20.2 ± 0.2 nM, respectively). In the xenograft 4T1 mouse model of immunocompetent animals, AR-C155858 (10 mg/kg i.p. once daily) had no effect on tumor volume and weight. Treatment with AR-C155858 resulted in slightly increased tumor lactate concentrations; however, the changes were not statistically significant. AR-C155858 was well tolerated, as demonstrated by the unchanged body weight and blood lactate concentrations. Average blood and tumor AR-C155858 concentrations (110 ± 22 and 574 ± 245 nM, respectively), 24 h after the last dose, were well above the IC(50) values. These data indicate that AR-C155858 penetrated 4T1 xenograft tumors and was present at high concentrations but was ineffective in decreasing tumor growth. Evaluations of AR-C155858 in other preclinical models of breast cancer are needed to further assess its efficacy.

AZD3965, a pyrole pyrimidine derivative, is a potent and orally bioavailable inhibitor of monocarboxylate transporter 1 (MCT1), currently in a Phase I clinical trial in UK for lymphomas and solid tumors. There is currently no published assay for AZD3965. The objectives of this study were to develop and validate a LC/MS/MS assay for quantifying AZD3965 in mouse plasma and tumor tissue. Protein precipitation with 0.1% formic acid in acetonitrile was used for sample preparation. Chromatographic separation was achieved on a C18 column followed by tandem mass spectrometry detection in multiple reaction monitoring mode with utilizing Atmospheric Pressure Chemical Ionization. AR-C155858 was used as the internal standard. The inter-day and intra-day precision and accuracy of quality control samples evaluated in plasma and tumor tissue were less than ±7% of the nominal concentrations. The extraction recovery, matrix effect and stability values were all within acceptable levels. Sample dilution integrity, accessed by diluting plasma spiked with AZD3965 10-fold with blank plasma, was 101%. The lower limit of quantification (LLOQ) and upper limit of quantification (ULOQ) were 0.15 ng/mL and 12 μg/mL, respectively, in plasma. The assay of AZD3965 in tumor tissue was also validated with good precision and accuracy. The LLOQ was 0.15 ng/mL in tumor tissue. This assay was successfully applied to pharmacokinetic and murine 4T1 breast tumor xenograft studies of AZD3965 in mice.