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Background Previous reports about risk factors for linezolid-induced thrombocytopenia have been insufficient, often due to the variability in study design and population, and some factors have not yet been studied. Aim The aims of this study are to determine potential risk factors for linezolid-induced thrombocytopenia, and to analyze the influencing factors of different thrombocytopenia definitions. Method This retrospective study involved patients who were administered intravenous linezolid for ≥ 1 day between January 1, 2015 and January 1, 2021. Their demographic and clinical data were extracted from electronic medical records. Thrombocytopenia was defined as: ①thrombocytopenia with platelet count

In recent years, immune checkpoint inhibitor has achieved remarkable success in multiple cancer treatment. However, how to pre‐judge which patients are suitable for immune checkpoint inhibitor is a difficult problem. We use the existing public bioinformatics database to comprehensively analyze the relationship between clinical data of various cancers with immune checkpoint blocking molecules and long non‐coding RNAs (lncRNAs), and try to find the potential predictive value of lncRNA for immunotherapy with checkpoint inhibitors. In this study, we found that: (a) high expression of lncRNA MIR155 host gene (MIR155HG) was closely related to better overall survival (OS) in cholangiocarcinoma (CHOL), lung adenocarcinoma (LUAD), and skin cutaneous melanoma (SKCM), and have better disease‐free survival (DFS) in CHOL. Meanwhile, the high level of MIR155HG was associated with poorer OS in glioblastoma multiforme (GBM), kidney renal clear cell carcinoma (KIRC), brain lower grade glioma (LGG), and uveal melanoma (UVM). (b) The expression of MIR155HG was significantly correlated with infiltrating levels of immune cells and immune molecules, especially with immune checkpoint molecules such as programmed cell death protein 1 (PD‐1), PD‐1 ligand 1 (PD‐L1), and cytotoxic T lymphocyte‐associated antigen 4 (CTLA4) in most kinds of cancers. (c) Detection of clinical CHOL and liver hepatocellular carcinoma tissues confirmed that there was a strong positive correlation between MIR155HG expression and the levels of CTLA4 and PD‐L1. MIR155 host gene can be used as a prognostic marker in multiple cancers, and of great value in predicting the curative effect of immune checkpoint inhibitor therapy owing to it is closely related with immune cells infiltration and immune checkpoint molecules expression., Long non‐coding RNA MIR155 host gene (MIR155HG) was closely related to overall survival in cholangiocarcinoma, lung adenocarcinoma, skin cutaneous melanoma, head and neck squamous cell carcinoma, glioblastoma multiforme, kidney renal clear cell carcinoma, brain lower grade glioma, and uveal melanoma. The expression of MIR155HG was significantly correlated with infiltrating levels of immune cells, molecules, and immune checkpoint molecules.

Human induced pluripotent stem cell (iPSC) lines are a powerful tool for studying development and disease, but the considerable phenotypic variation between lines makes it challenging to replicate key findings and integrate data across research groups. To address this issue, we sub-cloned candidate human iPSC lines and deeply characterized their genetic properties using whole genome sequencing, their genomic stability upon CRISPR-Cas9-based gene editing, and their phenotypic properties including differentiation to commonly used cell types. These studies identified KOLF2.1J as an all-around well-performing iPSC line. We then shared KOLF2.1J with groups around the world who tested its performance in head-to-head comparisons with their own preferred iPSC lines across a diverse range of differentiation protocols and functional assays. On the strength of these findings, we have made KOLF2.1J and its gene-edited derivative clones readily accessible to promote the standardization required for large-scale collaborative science in the stem cell field.

Fully automated proteomic pipelines have the potential to achieve deep coverage of cellular proteomes with high throughput and scalability. However, it is important to evaluate performance, including both reproducibility and ability to provide meaningful levels of biological insight. Here, we present an approach combining high field asymmetric waveform ion mobility spectrometer (FAIMS) interface and data independent acquisition (DIA) proteomics approach developed as part of the induced pluripotent stem cell (iPSC) Neurodegenerative Disease Initiative (iNDI), a large-scale effort to understand how inherited diseases may manifest in neuronal cells. Our FAIMS-DIA approach identified more than 8000 proteins per mass spectrometry (MS) acquisition as well as superior total identification, reproducibility, and accuracy compared to other existing DIA methods. Next, we applied this approach to perform a longitudinal proteomic profiling of the differentiation of iPSC-derived neurons from the KOLF2.1J parental line used in iNDI. This analysis demonstrated a steady increase in expression of mature cortical neuron markers over the course of neuron differentiation. We validated the performance of our proteomics pipeline by comparing it to single cell RNA-Seq datasets obtained in parallel, confirming expression of key markers and cell type annotations. An interactive webapp of this temporal data is available for aligned-UMAP visualization and data browsing (https://share.streamlit.io/anant-droid/singlecellumap). In summary, we report an extensively optimized and validated proteomic pipeline that will be suitable for large-scale studies such as iNDI.

Summary: The protein deacetylase SIRT1 (Sirtuin 1) regulates many cellular processes, including cell-cycle progression, DNA damage response, and metabolism. Although the centrosome is a key regulator of cell-cycle progression and genome stability, little is known concerning SIRT1 controlled centrosome-associated events. Here we report that the centrosome protein Plk2 is acetylated and undergoes deacetylation by SIRT1. Acetylation protects Plk2 from ubiquitination, and SIRT1-mediated deacetylation promotes ubiquitin-dependent degradation of Plk2. SIRT1 controls centriole duplication by temporally modulating centrosomal Plk2 levels. AURKA phosphorylates SIRT1 and promotes the SIRT1-Plk2 interaction in mitosis. In early-mid G1, phosphorylated SIRT1 deacetylates and promotes Plk2 degradation. In late G1, SIRT1 is hypophosphorylated and its affinity to Plk2 is decreased, resulting in a rapid accumulation of centrosomal Plk2, which contributes to the timely initiation of centriole duplication. Collectively, our findings uncover a critical role of SIRT1 in centriole duplication and provide a mechanistic insight into SIRT1-mediated centrosome-associated functions. : Ling et al. demonstrate that SIRT1 deacetylates and thereby destabilizes Plk2, which in turn results in the suppression of centriole duplication.

Background: Whether vancomycin (VAN) plus piperacillin-tazobactam (PTZ) could increase the risk of acute kidney injury (AKI) is still controversial in critically ill patients. The purpose of this study was to compare the risk of developing AKI and risk of developing AKI and treatment cost among this population receiving VAN/PTZ to a matched group receiving VAN/other antipseudomonal β-lactams. Methods: This multicenter, retrospective, matched study included 700 critically ill patients who received ≥48 hours of VAN/PTZ or VAN/other antipseudomonal β-lactams. The risk of developing AKI was compared between these two combination therapies using propensity-adjusted analysis. Furthermore, a pharmacoeconomic decision-analytic model was performed.Results: According to three AKI-defined criteria, VAN/PTZ was associated with significantly higher incidence of than VAN/other antipseudomonal β-lactams (all P < 0.001). In multivariate analysis, regardless of any VAN/other antipseudomonal β-lactams, VAN/PTZ was an independent predictor for stage 2 or 3 AKI. In the empiric treatment, the incremental cost-effectiveness ratios per additional nephrotoxic episode of 1147.35$, 1845.11$, and 3989.95$ were found for VAN/PTZ relative to, vancomycin plus imipenem-cilastatin, vancomycin plus meropenem, and vancomycin plus cefoperazone-sulbactam, respectively. Conclusion: In critically ill patients, VAN/PTZ was associated with both higher AKI risk and treatment cost when considering AKI occurence compared to VAN/other antipseudomonal β-lactams.Trial registration: retrospectively registered, ClinicalTrials.gov number: NCT03776409.

Human induced pluripotent stem cell (iPSC) lines are a powerful tool for studying development and disease, but the considerable phenotypic variation between lines makes it challenging to replicate key findings and integrate data across research groups. To address this issue, we sub-cloned candidate iPSC lines and deeply characterised their genetic properties using whole genome sequencing, their genomic stability upon CRISPR/Cas9-based gene editing, and their phenotypic properties including differentiation to commonly-used cell types. These studies identified KOLF2.1J as an all-around well-performing iPSC line. We then shared KOLF2.1J with groups around the world who tested its performance in head-to-head comparisons with their own preferred iPSC lines across a diverse range of differentiation protocols and functional assays. On the strength of these findings, we have made KOLF2.1J and hundreds of its gene-edited derivative clones readily accessible to promote the standardization required for large-scale collaborative science in the stem cell field.SummaryThe authors of this collaborative study deeply characterized human induced pluripotent stem cell (iPSC) lines to rationally select a clonally-derived cell line that performs well across multiple modalities. KOLF2.1J was identified as a candidate reference cell line based on single-cell analysis of its gene expression in the pluripotent state, whole genome sequencing, genomic stability after highly efficient CRISPR-mediated gene editing, integrity of the p53 pathway, and the efficiency with which it differentiated into multiple target cell populations. Since it is deeply characterized and can be readily acquired, KOLF2.1J is an attractive reference cell line for groups working with iPSCs.Graphical abstract

Second-generation H1-antihistamines are generally considered to be safe. Here we describe a healthy boy who developed left-arm convulsions after repeated exposure to a dry suspension of desloratadine combined with Huatengzi granules. The boy had no family or disease history of epilepsy, convulsions, or any other drug therapy. The Naranjo Adverse Drug Reaction Probability Scale was used to determine that the convulsions were probably related to desloratadine. Our findings suggest that desloratadine (a second-generation H1-antihistamine) can cause epileptic convulsions in healthy children, and so clinicians should be vigilant of the possibility of central side effects.

Desloratadine, a second generation H1-antihistamine, is generally considered to be safe. We found only one article reporting four children with a family or disease history of epilepsy who developed the condition after desloratadine treatment, with all four patients recovering well. Here we describe a healthy boy who developed left-arm convulsions on day 68 after taking desloratadine, at which point the desloratadine treatment was immediately stopped. Investigations were completed on day 83 and the patient was diagnosed with epilepsy. He was prescribed sodium valproate combined with oxcarbazepine, topiramate, lamotrigine and clonazepam for 15 months, which did not control the convulsions. During the following 3 months the patient received sodium valproate combined with lacosamide, and on day 615 the seizures stopped and no further convulsions occurred. At the follow-up, his father reported that the boy's memory was not as good as it had been previously. The convulsions continued after the withdrawal of desloratadine; therefore, the pathological mechanism of convulsion and the treatment plan need further research.

Tumor penetration of nanocarriers is still an unresolved challenge for effective drug delivery. Herein, we described a size-switchable nanoplatform in response to an external near-infrared (NIR) laser for transcellular drug delivery. The nanoplatform was constructed with a poly(N-isopropylacrylamide) (PNIPAM)-based nanogel encapsulating chitosan-coated single-walled carbon nanotubes, followed by loading a chemotherapeutic drug, doxorubicin (DOX). In mice bearing orthotopic breast tumors, the photothermal effect from single-walled carbon nanotubes upon NIR irradiation potently inhibited tumor growth. The antitumor effect of the nanomedicine with NIR irradiation might be attributed to its capability of transcellular transport and tumor penetration in mice. In addition, the nanomedicine with NIR irradiation could elicit an antitumor response by increasing cytotoxic T cells and decreasing myeloid-derived suppressor cells. These results validated the application of photo/thermo-responsive nanomedicine in the orthotopic model of breast cancer.

Cardiac hypertrophy is a one of common type of CHD, responsible for cardiac mortality worldwide. The present study designed to investigate the effect of muscarinic receptors agonist in the rat model of cardiac hypertrophy. A total of 30 male adult Wistar rats having body weight 300-400 gram were equally distributed in two groups (Test group: Rats with Angiotensin II + M3 receptor agonist [acetylcholine]; Reference group: Rats with cardiac hypertrophy induced by Angiotensin II). Rat model of cardiac hypertrophy were induced by Angiotensin II. Effect of M3 receptor agonist on cardiac hypertrophy was evaluated by electrocardiography, hemodynamic and histological assessment. Also, expression of M3 receptor was analyzed using by real-time-PCR and Western blot analysis. Also, vital signs such as pulse rate, and blood pressure were measured. Echocardiographic related variable including ejection fraction were also assessed in both the groups. The results of this study showed acetylcholine attenuates the hypertrophic response triggered by Angiotensin II, by upregulation of M3 receptor. Upregulation of M3 receptor after administration of acetylcholine ameliorates hypertrophic responses induced by angiotensin II. Also acetylcholine treatment prevents Angiotensin II induced increase in level of ANP and β-myosin, which are responsible for inducing cardiac hypertrophic responses. Moreover, acetylcholine ameliorates Angiotensin II induced cell enlargement by reducing the surface area of cells. Overall finding suggested that acetylcholine improves left ventricle hypertrophy and ejection fraction by activating M3 receptor in heart. The finding of this study gives the new vision to cardiovascular researchers to develop anti- hypertrophy therapy based on M3 receptor.

We describe the first case of Stevens-Johnson syndrome (SJS) occurring 8 days after the first dose of a three-dose rabies vaccination series. She had no history of vaccine-related rash or other adverse drug reactions, nor had she received any other drug therapy. The temporal relationship between the development of SJS and the vaccination suggests that the rabies vaccination probably was the causal agent. This case serves as a warning of a distinct cutaneous reaction of rabies vaccination.

Benzo[a]pyrene (B[a]P) is a carcinogen in cigarette smoke. We found that B[a]P induced SIRT1 in human bronchial epithelial BEAS-2B cell. SIRT1 was overexpressed in the lung of B[a]P-exposed mice and in human lung cancer biopsies. SIRT1 up-regulated TNF-α and β-catenin and down-regulated the membrane fraction of E-cadherin. In addition, SIRT1 promoted invasion, migration and tumorigenesis of BEAS-2B cells in nude mice upon B[a]P exposure. Thus, SIRT1 is involved in B[a]P-induced transformation associated with activation of the TNF-α/β-catenin axis and is as a potential therapeutic target for lung cancer.

Downstream signaling of physiological and pathological cell responses depends on post-translational modification such as ubiquitination. The mechanisms regulating downstream DNA damage response (DDR) signaling are not completely elucidated. Sirtuin 1 (SIRT1), the founding member of Class III histone deacetylases, regulates multiple steps in DDR and is closely associated with many physiological and pathological processes. However, the role of post-translational modification or ubiquitination of SIRT1 during DDR is unclear. We show that SIRT1 is dynamically and distinctly ubiquitinated in response to DNA damage. SIRT1 was ubiquitinated by the MDM2 E3 ligase in vitro and in vivo. SIRT1 ubiquitination under normal conditions had no effect on its enzymatic activity or rate of degradation; hypo-ubiquitination, however, reduced SIRT1 nuclear localization. Ubiquitination of SIRT1 affected its function in cell death and survival in response to DNA damage. Our results suggest that ubiquitination is required for SIRT1 function during DDR.

ObjectiveThe aims of this study were to identify the influencing factors such as gender, age, dose and combinations of other antiepileptic drugs (AEDs), especially in triple combinations on the pharmacokinetic of Lamotrigine (LTG) in epilepsy patients of Northwest Chinese Han population.MethodsData of the LTG concentration and clinical information were analyzed retrospectively from a therapeutic drug monitoring (TDM) database at the Clinical Pharmacy Laboratory of Xi'an Central Hospital between January 1, 2016 and January 1, 2018. The independent-sample t-test, one-way ANOVA analysis and Bonferroni and Tamhane T3 post-hoc test, the stepwise multivariate regression analysis were adopted by IBM SPSS, version 22.0.Results226 serum samples met the inclusion criteria and were evaluated. The mean LTG serum concentration was 5.48±3.83 μg/mL. There were no gender differences (P = 0.64), and there were no significant effects by age on LTG serum concentration after age stratification (3-14 years old, 14-45 years old, 45-59 years old) (P = 0.05). Multiple regression analysis showed that the daily LTG dose and co-administration of other AEDs significantly affected LTG serum concentrations. Combination with enzyme-inducer AEDs, the mean steady-state LTG concentration could be decreased by 30.73% compared with LTG monotherapy. Among enzyme-inducer AEDs, particularly strong inducer Carbamazepine (CBZ) could decrease the mean LTG concentration by 53.65%, but weak inducer AEDs such as Oxcarbazepine (OXC) and Topiramate (TPM) had no effect, Valproic acid (VPA) could increase the mean LTG concentration by 93.95%, and the inducer only partially compensated for the inhibitory effect of VPA in triple combination.ConclusionsThere were no significant gender and age effects, but the LTG daily dose and co-administration of other AEDs significantly affected LTG serum concentration. Combination with enzyme-inducer AEDs, especially CBZ could significantly decrease LTG serum concentrations, VPA could significantly increase LTG serum concentrations, and the inducer only partially compensated for the inhibitory effect of VPA in triple combination. In the clinical setting, these findings can help to estimate LTG concentrations and adjust dosage and evaluate adverse drug reactions.

Centrosome duplication is controlled both negatively and positively by a number of proteins. The activities and stabilities of those regulatory proteins are in many cases controlled by posttranslational modifications. Although acetylation and deacetylation are highly common posttranslational modifications, their roles in the regulation of centrosome duplication had not been closely examined. Here, through focusing on the deacetylases, we investigated the role of acetylation/deacetylation in the regulation of centrosome duplication and induction of abnormal amplification of centrosomes. We found that the deacetylation event negatively controls centrosome duplication and amplification. Of the 18 total known deacetylases (HDAC1-11, SIRT1-7), ten deacetylases possess the activity to suppress centrosome amplification, and their centrosome amplification suppressing activities are strongly associated with their abilities to localize to centrosomes. Among them, HDAC1, HDAC5 and SIRT1 show the highest suppressing activities, but each of them suppresses centrosome duplication and/or amplification with its unique mechanism.

The reversible acetylation of histones and non-histone proteins by histone acetyltransferases and deacetylases (HDACs) plays a critical role in many cellular processes in eukaryotic cells. HDAC6 is a unique histone deacetylase with two deacetylase domains and a C-terminal zinc finger domain. HDAC6 resides mainly in the cytoplasm and regulates many important biological processes, including cell migration and degradation of misfold proteins. HDAC6 has also been shown to localize in the nucleus to regulate transcription. However, how HDAC6 shuttles between the nucleus and cytoplasm is largely unknown. In addition, it is not clear how HDAC6 enzymatic activity is modulated. Here, we show that HDAC6 can be acetylated by p300 on five clusters of lysine residues. One cluster (site B) of acetylated lysine is in the N-terminal nuclear localization signal region. These lysine residues in site B were converted to glutamine to mimic acetylated lysines. The mutations significantly reduced HDAC6 tubulin deacetylase activity and further impaired cell motility, but had no effect on histone deacetylase activity. More interestingly, these mutations retained HDAC6 in the cytoplasm by blocking the interaction with the nuclear import protein importin-α. The retention of HDAC6 in the cytoplasm by acetylation eventually affects histone deacetylation. Thus, we conclude that acetylation is an important post-translational modification that regulates HDAC6 tubulin deacetylase activity and nuclear import.

DNA methylation and histone acetylation/deacetylation are distinct biochemical processes that control gene expression. While DNA methylation is a common epigenetic signal that inhibits gene transcription, histone deacetylation similarly represses transcription but can be both an epigenetic and nonepigenetic phenomenon. Here we report that the histone deacetylase SIRT1 regulates the activities of DNMT1, a key enzyme responsible for DNA methylation. In mass spectrometry analysis, 12 new acetylated lysine sites were identified in DNMT1. SIRT1 physically associates with DNMT1 and can deacetylate acetylated DNMT1 in vitro and in vivo. Interestingly, deacetylation of different lysines on DNMT1 has different effects on the functions of DNMT1. For example, deacetylation of Lys1349 and Lys1415 in the catalytic domain of DNMT1 enhances DNMT1's methyltransferase activity, while deacetylation of lysine residues in the GK linker decreases DNMT1's methyltransferase-independent transcriptional repression function. Furthermore, deacetylation of all identified acetylated lysine sites in DNMT1 abrogates its binding to SIRT1 and impairs its capability to regulate cell cycle G(2)/M transition. Finally, inhibition of SIRT1 strengthens the silencing effects of DNMT1 on the expression of tumor suppressor genes ER-α and CDH1 in MDA-MB-231 breast cancer cells. Together, these results suggest that SIRT1-mediated deacetylation of DNMT1 is crucial for DNMT1's multiple effects in gene silencing.

The ataxia telangiectasia group D-complementing (ATDC) gene product, also known as TRIM29, is a member of the tripartite motif (TRIM) protein family. ATDC has been proposed to form homo- or heterodimers and to bind nucleic acids. In cell cultures, ATDC expression leads to rapid growth and resistance to ionizing radiation (IR), whereas silencing of ATDC expression decreases growth rates and increases sensitivity to IR. Although ATDC is overexpressed in many human cancers, the biological significance of ATDC overexpression remains obscure. We report here that ATDC increases cell proliferation via inhibition of p53 nuclear activities. ATDC represses the expression of p53-regulated genes, including p21 and NOXA. Mechanistically, ATDC binds p53, and this interaction is potentially fine-tuned by posttranslational acetylation of lysine 116 on ATDC. The association of p53 and ATDC results in p53 sequestration outside of the nucleus. Together, these results provide novel mechanistic insights into the function of ATDC and offer an explanation for how ATDC promotes cancer cell proliferation.

RATIONALE Diaphragmatic Hernia is rare as complication of Colonoscopy. Diaphragmatic hernia as a complication of colonoscopy has been reported only few cases. Additionally, it is often misdiagnosed as other disease by clinicians due to their lack of related knowledge, which delays diagnosis of Diaphragmatic hernia and thus exacerbates the prognosis. PATIENT CONCERNS We report the case of a 66-year-old man with fecal occult blood. In the case, sudden epigastric pain after colonoscopy owing to diaphragmatic hernia in a left hemidiaphragm. DIAGNOSES The diagnoses made by a CT scan without delay. It showed marked protrusion of the large bowel into the left thoracic cavity along with elevation of the left diaphragm. INTERVENTIONS The diaphragmatic defect was repaired by simple closure and intestinal adhesions release surgery. OUTCOMES Five days after surgery, the patient was discharged in good condition. LESSONS Most of diaphragmatic hernia is congenital with high mortality. However, there are a few cases of Diaphragmatic hernia caused by previous trauma or surgery. We herein report an unusual case of diaphragmatic hernia related to colonoscopy but usually life-threatening complication.

Phosphatase of regenerating liver-3 (PRL-3) is overexpressed in several human cancers and associated with tumor progression, invasion and metastasis. However, the correlation between PRL-3 expression and clinical outcome in ovarian cancer has not been studied. In the present study, we investigated the expression of PRL-3 in 119 ovarian cancers and 30 normal ovarian tissues by immunohistochemistry with an anti-PRL-3 mouse monoclonal antibody 3B6, and analyzed its relationship with clinicopathologic factors and survival. The results demonstrated that PRL-3 expression was significantly higher in ovarian cancers compared to normal ovarian tissues (P < 0.001). PRL-3 expression is not correlated with patient age, menstruation, tumor size, histological type, residual tumor, or other clinical findings. The patients with PRL-3-positive tumors had a significant poor prognosis than those with PRL-3-negative tumors. Univariate analysis identified PRL-3 expression as a poor outcome predictor (HR 1.925, 95% CI, 1.046-3.544, P = 0.035). Multivariate analysis indicated that PRL-3 expression was an independent prognostic factor of borderline significance (HR 1.695, 95% CI, 0.914-3.145, P = 0.094). Our results suggest that PRL-3 may serve as a valuable marker for diagnosis of ovarian cancer and as a potential independent prognostic factor for ovarian cancer.

Histone deacetylase (HDAC) inhibition leads to cell cycle arrest in G1 and G2, suggesting HDACs as therapeutic targets for cancer and diseases linked to abnormal cell growth and proliferation. Many HDACs are transcriptional repressors. Some may alter cell cycle progression by deacetylating histones and repressing transcription of key cell cycle regulatory genes. Here, we report that HDAC10 regulates the cell cycle via modulation of cyclin A2 expression, and cyclin A2 overexpression rescues HDAC10 knockdown-induced G2/M transition arrest. HDAC10 regulates cyclin A2 expression by deacetylating histones near the let-7 promoter, thereby repressing transcription. In HDAC10 knockdown cells, let-7f and microRNA 98 (miR-98) were upregulated and the let-7 family target, HMGA2, was downregulated. HMGA2 loss resulted in enrichment of the transcriptional repressor E4F at the cyclin A2 promoter. These findings support a role for HDACs in cell cycle regulation, reveal a novel mechanism of HDAC10 action, and extend the potential of HDACs as targets in diseases of cell cycle dysregulation.

Cancer and embryo expression protein 65 (CEP65) is a centrosomal protein that is expressed at relatively high levels in embryonic tissue and different cancerous tissues, but its role in tumorigenesis remains unknown. In the present study, CEP65 was stably expressed in AGS gastric cancer cells. CEP65 was found to promote cell growth in the MTT assay and to enhance cell migration and invasion in Transwell chamber assays. To validate results from the in vitro experiments, CEP65 was stably expressed in BICR-H1 breast cancer cells through adenovirus-mediated transduction. By inoculating BICR-H1 cells on chick chorioallantoic membrane (CAM), it was found that CEP65 promotes cell growth on the CAM and increases cell metastasis to the lungs of the chicken. By utilizing a xenograft severe combined immunodeficiency mouse model, CEP65 was also found to accelerate BICR-H1 cell growth and metastasis to the lungs. Furthermore, it was shown that CEP65 increases matrix metalloproteinase (MMP)2 activity in zymographic assays, however, microarray screening and reverse transcription polymerase chain reaction validation revealed that CEP65 had no effect on the expression levels of MMP2 or MMP9, but decreased the expression levels of metastasis-associated genes, TIMP2, RAP and VTN. Taken together, the results of the present study demonstrated the oncogenic function of CEP65 in promoting cancer cell growth and metastasis.

Background: Endometriosis is a benign gynecologic condition that is known to have transformation potential. Women with ovarian endometriosis have twice the risk for developing ovarian cancer of the endometrioid or clear cell types. However, the carcinogenic pathways by which endometriosis associated ovarian carcinoma (EAOC) develops remain poorly understood. Epigenetics are emerging as an important player in the progression from a pre-malignant to a malignant phenotype. Previously, we showed that endometriosis cells and tissues are characterized by increased levels of HDAC1 and 2, histone acetylation at lysine 9 and 16 (H3K9, H4K16), and that promoter sequences of candidate genes for endometriosis are enriched in these histone marks. Aim: We sought to investigate the role of these histone modifications in modulation of gene expression in ovarian endometriosis (OE), endometriosis-associated ovarian cancer (EAOC), and adjacent histologically normal tissues (normal adjacent, NA). Methods: Series of formalin-fixed paraffin-embedded tissue (FFPE) blocks from EAOC patients containing these three histological tissues were studied. DNA extracted from these three tissues were subjected to chromatin immunoprecipitation using either anti-Ac-H3K9 or anti-Ac-H4K16, and then next generation sequencing. We are analyzing if these three histological tissues are characterized by a different pattern of histone modification enrichment in gene promoters and gene expression profile and identify the genes involved. Also, we assessed the effects of HDAC inhibitors (HDACis) in endometriotic and normal endometrial cells, by measuring its effects in proliferation, invasion and cell viability. Results: The analysis of ChIP-seq data was mapped of the short reads. Unmapped reads: raw reads were mapped to human genome using BWA. Many unmapped reads (have 30%) were found. Further analysis of unmapped reads using blast against NCBI notation database. Peaks (binding sites of histone): MACS2 software was used to call peaks. Most samples (P018-OEH3K9Ac P018-OEH4K16Ac) have less than 10k peaks detected, indicating a non-successful experiment. Code for IDR analysis was done to check how well biological replicates are reproducible. The p104-AN-H3K09 and p018-AN-H3K09 have good reproducible results, the rests have poor correlations. Based in this find NanoString molecular technology the assay will be necessary to validate our ChIP-Seq results and screening large sample sets against focused sets of loci involved in chromatin remodeling and gene expression. Citation Format: Janice Barros Monteiro, Alexandra M. Maldonado López, Lirong Peng, Idhaliz Flores, Ed Seto. Understanding of the ovarian endometriosis and gene expression functions in neoplastic transformation. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Sep 24-27, 2015; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2016;76(2 Suppl):Abstract nr B06.

Histone deacetylase 9 (HDAC9), like most Class II HDACs, catalyzes the removal of acetyl moieties from the ϵ-amino groups of conserved lysine residues in the N-terminal tail of histones. Biologically, HDAC9 regulates a wide variety of normal and abnormal physiological functions, including cardiac growth, T-regulatory cell function, neuronal disorders, muscle differentiation, development, and cancer. In a biochemical approach to identify non-histone substrates of HDAC9, we found that HDAC9 co-purifies specifically with the ataxia telangiectasia group D-complementing (ATDC; also called TRIM29) protein. HDAC9 deacetylates ATDC, alters the ability of ATDC to associate with p53, and consequently inhibits the cell proliferation-promoting activity of ATDC. These results implicate the importance of non-histone deacetylation by HDAC9 and confirm and further extend the multifunctions of this Class II deacetylase.

Background: Increasing evidence has suggested that phosphatase of regenerating liver-3 (PRL-3) plays an important role in cancer cell migration, invasion and metastasis. However, the correlation between the PRL-3 expression and clinical outcome in breast cancer has not been investigated. Patients and methods: Using a PRL-3-specific monoclonal antibody 3B6, PRL-3 expression was assessed by immunohistochemistry in tumor tissues from 382 breast cancer patients with a median follow-up of 65 months. Results: We found that 34.8% patients expressed a high level of PRL-3 protein in their tumors. Patients with a high level of PRL-3 in the tumor had a worse disease-specific survival (DSS) rate than those with a low level of PRL-3 (74.0% versus 84.9%, P = 0.011), and PRL-3 remained an independent prognostic marker for DSS (HR 1.8, 95% CI 1.1–2.9, P = 0.019) in multivariate analysis. More importantly, in 219 node-negative patients, PRL-3 showed a significant correlation with DSS in univariate analysis (P = 0.014) and retained a borderline significance (HR 2.65, 95% CI 0.92–7.64, P = 0.071) in multivariate analysis. Conclusions: Our results suggest that PRL-3 may serve as an unfavorable prognostic marker in breast cancer, especially for patients with node-negative diseases. Thus, our findings may provide useful information for individualized therapy in the clinical setting.

Abstract 359FN2 T-cells are an essential component of immune mediated tumor rejection. Adoptive transfer of T-cells has resulted in durable antitumor responses in some patients with hematologic malignancies. Further improvement in the efficacy of this treatment modality will require a better understanding of the regulatory checkpoint(s) limiting T-cell expansion and/or reactivity in vivo. Our group has been especially interested in the epigenetic regulatory mechanisms affecting T-cells, particularly those involving histone deacetylases (HDACs). HDACs are enzymes recruited to gene promoters where they regulate transcription through histone modifications. The role of HDACs in cell biology, initially limited to their effects upon histones, now encompasses more complex regulatory functions that are dependent on their tissue expression, subcellular compartment distribution and the stage of cellular differentiation. For instance, HDAC11, the most recently discovered member of the HDAC family, has been found to be predominantly expressed in the brain, kidney, testis as well as in T-cells. Recently, HDAC11 knock out (KO) mice have been generated by targeted deletion of floxed exon 3 of the HDAC11 gene. Although analysis of the T-cell compartment in these mice revealed no significant differences in the absolute number and/or percent of T-cells in lymphoid organs as compared to wildtype mice, striking functional differences were observed in HDAC11KO T cells. First, in response to in vitro stimulation with anti-CD3 plus anti-CD28 antibodies, HDAC11KO T-cells display an enhanced proliferation, produce significantly higher levels of the pro-inflammatory cytokines IL-2, IFN-gamma and TNF-alpha and are less susceptible to Treg-mediated suppression. Studies performed in HDAC11KO T-cells expressing a transgenic receptor (TCR) specific for ovalbumin (OTII;HDAC11KO mice) confirm that these T-cells are hyperreactive in an antigen-specific manner. In vivo, HDAC11KO T cells induced significantly more severe graft-versus-host disease (GVHD) than wildtype T-cells after allogeneic bone marrow transplantation. Enhanced GVHD mediated by HDAC11KO T cells was associated with increased levels of T-cell expansion and secreted Th1-cytokines such as IFN-gamma. Further demonstration of the intrinsic potency of HDAC11KO T-cells was provided by the finding that when the number of adoptively transferred T-cells was titrated down such that wildtype T-cells no longer induced GVHD, T-cells from HDAC11KO still potently did so. HDAC11KO T-cells are also endowed with a stronger antitumor effect given the prolonged survival observed in mice challenged with C1498 leukemic cells. Mechanistically, using ChIP analysis, we have found that HDAC11 is recruited to the T-bet promoter, which may explain, at least in part, the Th1 phenotype displayed by T-cells in the absence of the repressive effect of HDAC11. Taken together, we have unveiled for the first time that HDAC11 is a regulatory checkpoint in T-cells and represents a novel molecular target to improve the efficacy of T-cell adoptive immunotherapy for the treatment of hematologic malignancies. Disclosures: No relevant conflicts of interest to declare.

Abstract 1466 Background: Histone acetyltransferases and histone deacetylases (HDAC) regulate gene expression through acetylation-deacetylation of histones. HDACs are the target of a family of compounds known as HDAC inhibitors, which have been shown to suppress pro-inflammatory cytokines and reduce acute graft-versus-host disease (GVHD) while preserving the graft-versus-leukemia (GVL) effect after allogeneic bone marrow transplantation (BMT) in mice. However, the role of individual HDAC members in the development of GVHD is not clear. Recently, HDAC11, the newest member of the HDAC family has emerged as an important transcriptional regulator of inflammatory responses in antigen-presenting cells (APCs)1. Here, we evaluated the role of HDAC11 on APCs and T cells in the allogeneic BMT setting in mice with genetic disruption of HDAC11. Method: Proliferation of wild-type (WT) and HDAC11 knock-out (KO) T cells in response to allogeneic antigens was compared by [H3] thymidine incorporation assay. Using the same method, we also tested the antigen presentation ability of WT and HDAC11 KO APCs. For in vivo studies, we used a clinical relevant mouse model of BMT: C57BL/6 (B6) ® BALB/c. To evaluate the role of HDAC11 in the function of T cells and APCs, WT and KO mice on B6 background were used as donors and recipients, respectively. Recipient survival was monitored daily and GVHD symptom was evaluated at least twice a week. HDAC11 KO mice were supplied by Merck and Co., Inc. Results: In vitro, HDAC11 KO T cells proliferated stronger than WT T cells under the stimulation of allogeneic APCs. Recipients of HDAC11 KO T cells lost significantly more body weight (p < 0.05), and died significantly sooner than those of WT T cells (p < 0.01). The pathologic score of KO recipients was higher than that of WT recipients in each of GVHD target organs including lung, liver, small intestine and colon. Mechanistically, we found that there were significantly more total and IFNγ-producing donor T cells in the recipients of KO cells than those of WT cells (p < 0.05). Collectively, HDAC11 KO T cells have higher activity in response to alloantigens in vitro and induced more severe GVHD in vivo compared to WT T cells. In contrast, KO and WT APCs had a similar ability to stimulate allogeneic T cells in vitro, and no significant difference in GVHD development was observed in WT or KO recipients after allogeneic BMT. Conclusion: HDAC11 negatively regulates T-cell function, but has no significant effect on APC function. This finding provides a rationale to promote T-cell immunity or tolerance by inhibiting or enhancing HDAC11, respectively. 1 Villagra et al. Nature Immunology, 10:92-100, 2009. Disclosures: No relevant conflicts of interest to declare.