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Current gene delivery methods for maize are limited to specific genotypes and depend on time-consuming and labor-intensive tissue culture techniques. Here, we report a new method to transfect maize that is culture-free and genotype independent. To enhance efficiency of DNA entry and maintain high pollen viability of 32%-55%, transfection was performed at cool temperature using pollen pretreated to open the germination aperture (40%-55%). Magnetic nanoparticles (MNPs) coated with DNA encoding either red fluorescent protein (RFP), β-glucuronidase gene (GUS), enhanced green fluorescent protein (EGFP) or bialaphos resistance (bar) was delivered into pollen grains, and female florets of maize inbred lines were pollinated. Red fluorescence was detected in 22% transfected pollen grains, and GUS stained 55% embryos at 18 d after pollination. Green fluorescence was detected in both silk filaments and immature kernels. The T1 generation of six inbred lines showed considerable EGFP or GUS transcripts (29%-74%) quantitated by polymerase chain reaction, and 5%-16% of the T1 seedlings showed immunologically active EGFP or GUS protein. Moreover, 1.41% of the bar transfected T1 plants were glufosinate resistant, and heritable bar gene was integrated into the maize genome effectively as verified by DNA hybridization. These results demonstrate that exogenous DNA could be delivered efficiently into elite maize inbred lines recalcitrant to tissue culture-mediated transformation and expressed normally through our genotype-independent pollen transfection system.

Calreticulin (CRT) is a Ca2+-binding chaperone protein that alters cellular Ca2+-homeostasis in the endoplasmic reticulum (ER). Previously it was shown that CRT was overexpressed in esophageal squamous cell carcinoma (ESCC), and elevated CRT expression promoted the migration and invasion of ESCC cells. In the present study, the mechanisms underlying the role of CRT in esophageal carcinoma progression were investigated. Critically, depletion of CRT or protein-tyrosine phosphatase 1B (PTP1B) reduced ESCC cell migration and metastasis to the lung, whereas restoration of PTP1B protein levels rescued cell migration in CRT-silenced cells. Knockdown of CRT decreased PTP1B protein expression by reducing phosphorylation at the Y694 site of STAT5A, whereas knockdown of PTP1B reduced ERK1/2 phosphorylation at T204. Immunohistochemical analysis of CRT and PTP1B expression in ESCC patient tissues was strongly correlated. Importantly, PTP1B expression was associated with poor survival in patients with CRT overexpression. Overall, these data indicate a novel signaling pathway connecting CRT, STAT5A, PTP1B, and ERK1/2 in the regulation of ESCC cell migration.Implications: These findings suggest that PTP1B is a downstream effector of CRT signaling, promotes tumor progression, and can potentially be used as a new drug target for ESCC. Mol Cancer Res; 11(9); 986–94. ©2013 AACR.

PDF file - 1061K, S Figure 1. Down-regulation of CRT expression reduces PTP1B mRNA and protein expression in KYSE510 cells. S2. PTP1B promotes ESCC cell invasion and migration in KYSE510 cells. KYSE510 cells were transfected with PTP1B-specific siRNA (siRNA PTP1B) or non-targeting siRNA control (siRNA cont). S3. CRT regulates cell motility through PTP1B in ESCC cells. (A-C) 1x105 KYSE150 cells were seeded on 24-well plates with three replicates. S4. Cancer-inflammatory response was minimal in mouse model. S5. CRT regulates PTP1B transcription through Stat5a. S6. The expression of CRT and PTP1B in 176 ESCC tissues was evaluated by immunohistochemistry. S7. Correlation of PTP1B expression with overall survival in patients with CRT high-expressing tumors. S8. Model of CRT-dependent signaling pathway in ESCC cell metastasis. Supplementary Table 1. The candidate genes altered in CRT-siRNA KYSE150 cells as compared with non-targeting siRNA cells detected by cDNA array

Protein kinase Cι (PKCι) is an atypical PKC isoform and participates in multiple aspects of the transformed phenotype in human cancer cells. We previously reported that frequent amplification and overexpression of PKCι were correlated with lymph node metastasis in primary esophageal squamous cell carcinomas (ESCC). In the present study, short interfering RNA–mediated silencing of PKCι revealed that this enzyme was required for cell migration, invasion, and resistance to anoikis. In vivo experiments showed that PKCι suppression decreased tumor growth in esophageal cancer xenografts and lung metastases in nude mice. At the molecular level, knockdown of PKCι in suspended ESCC cells caused a decrease in S-phase kinase-associated protein 2 (SKP2) that had been reported to promote resistance to anoikis via the PI3K/AKT pathway. AKT phosphorylation was abolished after PKCι suppression, but AKT activation could be refreshed by PKCι upregulation, suggesting that PKCι enhanced cell resistance to anoikis via the PKCι-SKP2-PI3K/AKT pathway. Addition of the proteasome inhibitor MG132 prevented the decrease of SKP2 in PKCι silenced cells, and polyubiquitin-SKP2 was elevated after PKCι depletion, showing that PKCι might regulate the expression of SKP2 through the ubiquitin-proteasome pathway in suspended cells. Furthermore, overexpression of SKP2 in PKCι-downregulated cells restored cell resistance to anoikis. Most importantly, PKCι expression significantly correlated with SKP2 in 133 ESCC tissues (P = 0.031). Taken together, our data show that PKCι promotes tumorigenicity and metastasis of human esophageal cancer and that SKP2 is a candidate downstream effector of PKCι signaling in ESCC. Mol Cancer Res; 9(4); 390–402. ©2011 AACR.

Supplementary Figures S1-S5.

Purpose: Our aim was to identify frequent genomic aberrations in both esophageal squamous cell carcinoma (ESCC) and esophageal dysplasia and to discover important copy number-driving genes and microRNAs (miRNA) in ESCC.Experimental Design: We conducted array-based comparative genomic hybridization (array CGH) on 59 ESCC resection samples and 16 dysplasia biopsy samples. Expression of genes at 11q13.3 was analyzed by real-time PCR (RT-PCR) and immunohistochemistry (IHC). Integrated analysis was conducted to identify genes or miRNAs with copy number-expression correlations.Results: Array CGH identified 11 amplifications and eight homozygous deletions in ESCC. Integrated analysis of array CGH data with matched gene expression microarray data showed that 90 overexpressed genes and 24 underexpressed genes were consistent with DNA copy number changes, including 12 copy number-driving miRNAs. In esophageal dysplasia, six gains, four losses, 12 amplifications, and four homozygous deletions were detected. Amplifications of 7p11.2 and 11q13.2–11q13.3 (CCND1) and homozygous deletion at 9p21.3 (CDKN2A) were consistent genomic changes in both dysplasia and carcinoma. ANO1 at 11q13.3 was overexpressed at the mRNA and protein levels in tumors, and higher mRNA expression was correlated with the copy number increase. In particular, ANO1 expression was elevated in moderate dysplasia compared with normal esophageal epithelium. IHC revealed that ANO1 overexpression was positively correlated with lymph node metastasis and advanced clinical stage. Knockdown of ANO1 significantly inhibited the proliferation of KYSE30 and KYSE510 cells.Conclusion: Copy number aberrations in both esophageal dysplasia and ESCC may be useful as potential biomarkers for early detection. In addition, ANO1 may be a candidate target gene in esophageal tumorigenesis. Clin Cancer Res; 19(21); 5867–78. ©2013 AACR.

Effect of ANO1 on cell proliferation. Knock-down the expression of ANO1 inhibited the cell proliferation of KYSE30 (A) and KYSE510 cells (B). The absorbance at 450 nm was measured.

Supplementary Figures S1-S8; Supplementary Tables S1-S4.

Purpose: To investigate the molecular mechanisms through which polo-like kinase-1 (PLK1) takes part in anoikis resistance of esophageal squamous cell carcinoma (ESCC) cells.Experimental Design: The role of PLK1 in cell anoikis resistance was examined by ectopic gene expression and siRNA-mediated knockdown. Glutathione S-transferase pull-down and co-immunoprecipitation assays were utilized to investigate PLK1-interacting proteins. Electrophoretic mobility shift assay, chromatin immunoprecipitation, and reporter gene assays were carried out to identify the transcription factors responsible for PLK1 expression during anoikis resistance.Results: We found that detachment of ESCC cells triggers the upregulation of PLK1. Elevated PLK1 expression contributes to protection against anoikis in cancer cells through the regulation of β-catenin expression. Moreover, we showed that, through direct binding to the PLK1 promoter, the NF-κB subunit RelA transcriptionally activates PLK1, which inhibits the ubiquitination and degradation of β-catenin. Inhibition of the NF-κB pathway restores the sensitivity of cancer cells to anoikis by downregulating PLK1/β-catenin expression. In addition, RelA gene amplification and protein overexpression was significantly correlated with PLK1 expression in ESCC tissues.Conclusions: Our findings suggest that upregulation of PLK1 triggered by cell detachment is regulated by RelA at the transcriptional level. PLK1 protects esophageal carcinoma cells from anoikis through modulation of β-catenin protein levels by inhibiting their degradation. Taken together, this study reveals critical mechanisms involved in the role of RelA/PLK1/β-catenin in anoikis resistance of ESCC cells. Clin Cancer Res; 17(13); 4285–95. ©2011 AACR.

Supplement file: Materials and Methods

We conducted a single-arm, open-label, single-center phase I study (Clinicaltrials.gov NCT03984968) to assess the safety and efficacy of multicycle-sequential anti-CD19 CAR T-cell therapy in combination with autologous CD19+ feeding T cells (FTCs) and TKI as consolidation therapy in patients under the age of 65 years with de novo Ph-positive CD19+ B-ALL who are not eligible for allo-HSCT. Participants were given induction chemotherapy as well as systemic chemotherapy with TKI. Afterward, they received a single cycle of CD19 CAR T-cell infusion and another three cycles of CD19 CAR T-cell and CD19+ FTC infusions, followed by TKI as consolidation therapy. CD19+ FTCs were given at three different doses: (2×106/kg, 3.25×106/kg, and 5×106/kg). The phase I results of the first 15 patients, including 2 withdrawals, are presented. Phase II research is still ongoing. The most common adverse events were cytopenia (13/13) and hypogammaglobinemia (12/13). There were no CRS above grade 2 or ICANS, or grade 4 nonhematologic toxicities. All 13 patients achieved CR, including 12 patients with CMR at the data cutoff on Mar 31, 2022. The RFS was 84% (95% CI, 66%-100%), and the OS was 83% (95% CI, 58%-100%) with a median follow-up of 27 months (7-57 months). The total number of CD19-expressing cells decreased with an increasing CMR rate. CD19 CAR T cells survived for up to 40 months, whereas CD19+ FTCs vanished in 8 patients 3 months after the last infusion. These findings merit further evaluation and could form the basis for the development of an allo-HSCT-free consolidation paradigm.

Decidualization is necessary for the successful establishment of early pregnancy in rodents and humans. Disturbed decidualization results in recurrent implantation failure, recurrent spontaneous abortion, and preeclampsia. Tryptophan (Trp), one of the essential amino acids in humans, has a positive effect on mammalian pregnancy. Interleukin 4-induced gene 1 (IL4I1) is a recently identified enzyme that can metabolize L-Trp to activate aryl hydrocarbon receptor (AHR). Although IDO1-catalyzed kynurenine (Kyn) from Trp has been shown to enhance human in vitro decidualization via activating AHR, whether IL4I1-catalyzed metabolites of Trp are involved in human decidualization is still unknown. In our study, human chorionic gonadotropin stimulates IL4I1 expression and secretion from human endometrial epithelial cells through ornithine decarboxylase-induced putrescine production. Either IL4I1-catalyzed indole-3-pyruvic acid (I3P) or its metabolite indole-3-aldehyde (I3A) from Trp is able to induce human in vitro decidualization by activating AHR. As a target gene of AHR, Epiregulin induced by I3P and I3A promotes human in vitro decidualization. Our study indicates that IL4I1-catalyzed metabolites from Trp can enhance human in vitro decidualization through AHR-Epiregulin pathway.

In the recent decades, obesity rates among children and adolescents, especially males, have increased significantly. This worldwide phenomenon is thought to significantly affect the levels of sex hormones. However, the association between waist circumference (a marker of abdominal obesity) and sex hormone levels in children and adolescents is unknown. In this study, 4031 participants aged 6-19 years from the United States National Health and Nutrition Examination Survey (NHANES) in the USA were enrolled in this study. The common confounders of age, race, body mass index, educational level, family income, diabetes, and time of sample collection were also collected. The participants missing any of the above information were excluded from the study. We used multiple linear regression and other multiple statistics to assess the associations between waist circumference and serum testosterone, estradiol, sex hormone-binding globulin (SHBG), free androgen index (FAI), and testosterone/estradiol ratio (T/E

The current classification of acute myeloid leukemia (AML) relies largely on genomic alterations. Robust identification of clinically and biologically relevant molecular subtypes from nongenomic high-throughput sequencing data remains challenging. We established the largest multicenter AML cohort (n = 655) in China, with all patients subjected to RNA sequencing (RNA-Seq) and 619 (94.5%) to targeted or whole-exome sequencing (TES/WES). Based on an enhanced consensus clustering, eight stable gene expression subgroups (G1–G8) with unique clinical and biological significance were identified, including two unreported (G5 and G8) and three redefined ones (G4, G6, and G7). Apart from four well-known low-risk subgroups including PML::RARA (G1), CBFB::MYH11 (G2), RUNX1::RUNX1T1 (G3), biallelic CEBPA mutations or -like (G4), four meta-subgroups with poor outcomes were recognized. The G5 (myelodysplasia-related/-like) subgroup enriched clinical, cytogenetic and genetic features mimicking secondary AML, and hotspot mutations of IKZF1 (p.N159S) (n = 7). In contrast, most NPM1 mutations and KMT2A and NUP98 fusions clustered into G6–G8, showing high expression of HOXA / B genes and diverse differentiation stages, from hematopoietic stem/progenitor cell down to monocyte, namely HOX -primitive (G7) , HOX -mixed (G8), and HOX -committed (G6). Through constructing prediction models, the eight gene expression subgroups could be reproduced in the Cancer Genome Atlas (TCGA) and Beat AML cohorts. Each subgroup was associated with distinct prognosis and drug sensitivities, supporting the clinical applicability of this transcriptome-based classification of AML. These molecular subgroups illuminate the complex molecular network of AML, which may promote systematic studies of disease pathogenesis and foster the screening of targeted agents based on omics.

Mixed phenotype acute leukemia (MPAL) is a subtype of leukemia in which lymphoid and myeloid markers are co-expressed. Knowledge regarding the genetic features of MPAL is lacking due to its rarity and heterogeneity. Here, we applied an integrated genomic and transcriptomic approach to explore the molecular characteristics of 176 adult patients with MPAL, including 86 patients with T-lymphoid/myeloid MPAL (T/My MPAL-NOS), 42 with Ph+ MPAL, 36 with B-lymphoid/myeloid MPAL (B/My MPAL-NOS), 4 with t(v;11q23), and 8 with MPAL, NOS, rare types. Genetically, T/My MPAL-NOS was similar to B/T MPAL-NOS but differed from Ph+ MPAL and B/My MPAL-NOS. T/My MPAL-NOS exhibited higher CEBPA, DNMT3A, and NOTCH1 mutations. Ph+ MPAL demonstrated higher RUNX1 mutations. B/T MPAL-NOS showed higher NOTCH1 mutations. By integrating next-generation sequencing and RNA sequencing data of 89 MPAL patients, we defined eight molecular subgroups (G1-G8) with distinct mutational and gene expression characteristics. G1 was associated with CEBPA mutations, G2 and G3 with NOTCH1 mutations, G4 with BCL11B rearrangement and FLT3 mutations, G5 and G8 with BCR::ABL1 fusion, G6 with KMT2A rearrangement/KMT2A rearrangement-like features, and G7 with ZNF384 rearrangement/ZNF384 rearrangement-like characteristics. Subsequently, we analyzed single-cell RNA sequencing data from five patients. Groups G1, G2, G3, and G4 exhibited overexpression of hematopoietic stem cell disease-like and common myeloid progenitor disease-like signatures, G5 and G6 had high expression of granulocyte-monocyte progenitor disease-like and monocyte disease-like signatures, and G7 and G8 had common lymphoid progenitor disease-like signatures. Collectively, our findings indicate that integrative genomic and transcriptomic profiling may facilitate more precise diagnosis and develop better treatment options for MPAL.

Background: Although the inactivated SARS-CoV-2 vaccine (CoronaVac) has undergone preclinical tests and clinical trials evaluating its efficacy and safety, few data have been reported in the post-licensure real-world setting. We aimed to assess the safety of the vaccine among healthcare workers. Methods: A self-administered online survey on monitoring adverse reactions post vaccination was conducted among the staff who worked at and were vaccinated in a tertiary hospital in Taizhou, China, from February 24 to 7 March 2021. A total of 1526 subjects responded to the questionnaire when they received an e-mail or an e-poster on WeChat. Results: The incidences of overall adverse reactions after the first and second injections were 15.6% (238/1526) and 14.6% (204/1397), respectively. The most common adverse reaction was localized pain at the injection site, with an incidence of 9.6% and 10.7% after each dose, accounting for 61.8% and 73.0% of adverse reactions, respectively. Fatigue, muscle pain, and headache were the most common systemic adverse reactions. Conclusions: These findings implied that the inactivated CoronaVac vaccine has an acceptable safety profile among healthcare workers due to the low incidence of self-reported adverse reactions. This may boost public confidence in nationwide mass vaccination campaigns.

Cu-0.36wt.%Be-0.46wt.%Co alloy plate with 300 mm in width and 10 mm in thickness prepared by heating-cooling combined mold (HCCM) horizontal continuous casting was cold rolled. Microstructure evolution and mechanical properties of the alloy as well as its deformation mechanism were investigated. The results showed that the as-cast alloy plate had columnar grains along the length direction, good surface quality and elongation of 35%, which was directly large-reduction cold rolled without surface treatment, and the accumulative cold rolling reduction reached 98%. When the reduction was small (20%), numerous dislocations and dislocation cells formed, and the deformation mechanism was dislocation slip. When the reduction was 40%, deformation twins appeared, and interactions between twins and dislocation cells induced strip-like dislocation cells. When the reduction exceeded 60%, shear bands formed and apparent crystal rotation in the micro-region happened. Further increasing the reduction, the amount of the shear bands rose and they interacted with each other, which refined the grains apparently. The tensile strength and hardness increased from 353 MPa and HV 119 of the as-cast alloy to 625 MPa and HV 208 with 95% reduction, respectively, and the elongation reduced from 35% to 7.6%. A process of HCCM horizontal continuous casting-cold rolling can work as a novel compact method to fabricate Cu-Be alloy sheet.

To investigate the accuracy and safety of pedicle screw placement assisted by orthopedic robot and C-arm fluoroscopy.The clinical data of 36 patients with spinal diseases underwent surgical treatment from January 2019 to August 2020 was retrospectively analyzed. Among them, 18 cases were implanted pedicle screws assisted by orthopaedic robot(observation group), including 12 males and 6 females, aged from 16 to 61 years with an average of (38.44±3.60) years;there were 1 case of adolescent scoliosis, 1 case of spinal tuberculosis, 7 cases of lumbar spondylolisthesis, 4 cases of thoracic fracture and 5 cases of lumbar fracture. Another 18 cases were implanted pedicle screws assisted by C-arm fluoroscopy(control group), including 10 males and 8 females, aged from 18 to 58 years with an average of (43.22±2.53) years;there were 1 case of adolescent scoliosis, 6 cases of lumbar spondylolisthesis, 6 cases of thoracic fracture and 5 cases of lumbar fracture. The intraoperative fluoroscopy times, nail placement time and postoperative complications were recorded in two groups. CT scan was performed after operation. The Gertzbein-Robbins standard was used to evaluate the accuracy of pedicle screw placement which was calculated.The number of intraoperative fluoroscopy in observation group was(6.89±0.20) times, which was significantly higher than that in control group(14.00±0.18)times(Orthopaedic robot assisted pedicle screw placement has the advantages of less fluoroscopy times, shorter screw placement time and higher accuracy, which can further improve the surgical safety and has a broad application prospect in the orthopaedic.

To explore the clinical application of Orthopedic Tianji Robot in the treatment of thoracolumbar fractures with minimally invasive percutaneous pedicle screw internal fixation.The clinical data of 46 patients with thoracolumbar fractures treated by minimally invasive percutaneous pedicle screw internal fixation from June 2018 to January 2020 in Gansu Provincial People's Hospital was retrospectively analyzed. According to the different aided equipments, the patients were divided into a C-arm X-ray plus robot assisted nail placement group (observation group) and simple C-arm X-ray assisted nail placement group (control group). Twenty-two patients in observation group, including 12males and 10 females, aged from 24 to 61 years old, with a mean of(40.23±12.19) years, 3 cases were TAll patients were followed up for 5 to 12 months with an average of (7.07±0.83) months. There was no significant differences in intraoperative blood loss, operation time and hospitalization days between two groups (P0.05). The number of intraoperative fluoroscopy was (5.62±0.51) times in observation group and (12.54±0.52) in control group, the difference between two groups was statistically significant (P0.05). The VAS, the percentage of the height of the injured vertebra anterior edge and the Cobb angle of the injured vertebral plane at each time point after operation were improved(P0.05), but the difference between two groups was not statistically significant (P0.05). One case of postoperative incision infection occurred in the control group, which was cured after active dressing changes. There were no serious complications such as screw loosening and breakage in all patients, and there was no statistically significant difference in the incidence of complications between two groups(P0.05). The postoperative nail placement accuracy ratesof the observation group and the control group were 98.4% (126/128) and 90.3% (121/134), respectively, and the difference was statistically significant (P0.05).In the surgical treatment of thoracolumbar fractures, compared with simple C-arm fluoroscopy, the pedicle screw placement assisted by Orthopedic Tianji Robot overcomes the shortcomings of surgeon's physiological tremor, fatigue and multiple intraoperative fluoroscopy, improves the accuracy of screw placement, and is a more accurate and safer way of pedicle screw placement for thoracolumbar fractures.

Study design Retrospective case–control study. Objectives Proximal junctional kyphosis (PJK) is a postoperative complication involving the proximal segments which is commonly seen in patients with degenerative spine diseases (DSD). The purpose of the present study was to identify predictive factors for postoperative PJK in elderly patients with DSD. Methods We reviewed elderly patients with DSD who underwent thoracolumbar fusion involving no less than 3 levels. Patients who developed PJK were propensity score-matched with patients with DSD who received the same procedure but did not develop PJK. Demographic characteristics, sagittal vertical axis (SVA), computed tomography (CT) value (Hounsfield unit), and paraspinal muscle parameters were compared between PJK and non-PJK groups. Results Eighty-three PJK and non-PJK patients were selected by propensity score matching for age, sex, history of smoking, body mass index, number of fused segments, and upper instrumented vertebra (UIV) location. SVA showed no significant difference between the two groups. In PJK group, fatty infiltration (FI) in erector spinae and multifidus was significantly greater, while the relative cross-sectional area (rCSA) of erector spinae was significantly smaller than that in non-PJK group. CT value was significantly lower in PJK group. Lower erector spinae rCSA and CT value of the UIV, higher erector spinae FI and multifidus FI were identified as predictors of postoperative PJK. Conclusions PJK is a common complication in older patients with DSD. Paraspinal muscle degeneration and low bone mineral density of the UIV are predictors of PJK. Protective measures targeting paraspinal muscles and the UIV may help prevent postoperative PJK.

Objective The objective of this open-label, randomized study was to compare dose-dense paclitaxel plus carboplatin (PCdd) with dose-dense epirubicin and cyclophosphamide followed by paclitaxel (ECdd-P) as an adjuvant chemotherapy for early triple-negative breast cancer (TNBC). Methods We included Chinese patients with high recurrence risk TNBC who underwent primary breast cancer surgery. They were randomly assigned to receive PCdd [paclitaxel 150 mg/m2 on d 1 and carboplatin, the area under the curve, (AUC)=3 on d 2] or ECdd-P (epirubicin 80 mg/m2 divided in 2 d and cyclophosphamide 600 mg/m2 on d 1 for 4 cycles followed by paclitaxel 175 mg/m2 on d 1 for 4 cycles) every 2 weeks with granulocyte colony-stimulating factor (G-CSF) support. The primary endpoint was 3-year disease-free survival (DFS); the secondary endpoints were overall survival (OS) and safety. Results The intent-to-treat population included 143 patients (70 in the PCdd arm and 73 in the ECdd-P arm). Compared with the ECdd-P arm, the PCdd arm had significantly higher 3-year DFS [93.9% vs. 79.1%; hazard ratio (HR)=0.310; 95% confidence interval (95% CI), 0.137-0.704; log-rank, P=0.005] and OS (98.5% vs. 92.9%; HR=0.142; 95% CI, 0.060-0.825; log-rank, P=0.028). Worse neutropenia (grade 3/4) was found in the ECdd-P than the PCdd arm (47.9% vs. 21.4%, P=0.001). Conclusions PCdd was superior to ECdd-P as an adjuvant chemotherapy for early TNBC with respect to improving the 3-year DFS and OS. PCdd also yielded lower hematological toxicity. Thus, PCdd might be a preferred regimen for early TNBC patients with a high recurrence risk.

Aim: This study aimed to develop an effective risk predictor for patients with stage II and III colorectal cancer (CRC). Materials & methods: The prognostic value of p-mTOR (Ser2448) levels was analyzed using Kaplan–Meier survival analysis and Cox regression analysis. Results: The levels of p-mTOR were increased in CRC specimens and significantly correlated with poor prognosis in patients with stage II and III CRC. Notably, the p-mTOR level was an independent poor prognostic factor for disease-free survival and overall survival in stage II CRC. Conclusion: Aberrant mTOR activation was significantly associated with the risk of recurrence or death in patients with stage II and III CRC, thus this activated proteins that may serve as a potential biomarker for high-risk CRC.

Background/Aims: The purpose of this study was to probe the clinico-pathological significance and the underlying mechanism of miR-30d-5p expression in non-small cell lung cancer (NSCLC). Methods: We initially examined the level of miR-30d-5p expression in NSCLC and non-cancer tissues using RT-qPCR. Then, a series of validation analyses including a meta-analysis of data from microarray chips in Gene Expression Omnibus (GEO), data mining of the cancer genome atlas (TCGA) and an integrated meta-analysis incorporating GEO microarray chips, TCGA data, in-house RT-qPCR and literature studies were performed to examine the clinico-pathological value of miR-30d-5p expression in NSCLC. In vitro experiments were further conducted to investigate the impact of miR-30d-5p on NSCLC cell growth. The molecular mechanism by which miR-30d-5p regulates the pathogenesis of NSCLC was probed through a bioinformatics analysis of its target genes. Moreover, dual luciferase reporter assay was conducted to verify the targeting regulatory relationship between miR-30d-5p and CCNE2. Results: Based on results from RT-qPCR, GEO meta-analysis, TCGA data mining and the integrated meta-analysis incorporating GEO microarray chips, TCGA data, in-house RT-qPCR and literature studies, miR-30d-5p expression was decreased in NSCLC tissues, and patients with NSCLC who presented with lower miR-30d-5p expression tended to display an advanced clinical progression. Significant pathways including the Mucin type O-glycan biosynthesis pathway, cell cycle pathway and cysteine and methionine metabolism pathway (all P< 0.05) revealed potential roles of the target genes of miR-30d-5p in the oncogenesis of NSCLC. Results from in vitro experiments indicated that miR-30d-5p could attenuate proliferation and viability of NSCLC cells. Among the 12 identified hub genes, nine genes including E2F3, CCNE2, SKP2, CDK6, TFDP1, LDHA, GOT2, DNMT3B and ST6GALNAC1 were validated by Pearson’s correlation test and the human protein atlas (HPA) database as targets of miR-30d-5p with higher probability. Specifically, dual luciferase reporter assay confirmed that CCNE2 was directly targeted by miR-30d-5p. Conclusion: In summary, miR-30d-5p expression is decreased in NSCLC, and it might play the role as tumor suppressor in NSCLC by regulating target genes.

Background The MET gene has been recognized as a potential important therapeutic target in non–small-cell lung cancer (NSCLC). We sought to investigate the MET exon 14 mutations in a cohort of Chinese patients with NSCLC. Methods We tested 461 NSCLCs for MET exon 14 mutations by sequencing whole exon 14 and its flanking introns. The protein expression was determined by immunohistochemical analysis. Results In this study, we identified MET exon 14 mutations in 9 (2.0%) of 461 NSCLCs. Of these 9 mutations, 7 (77.8%) were located in the splice sites of MET exon 14, with MET overexpression in 6. One point mutation c.3010C>T (p.Arg1004Ter) was nonsense mutation with no MET expression. One insertion mutation was within exon 14 of MET with MET overexpression. MET protein localization in tumor cells with MET exon 14 mutations was different between mutation types. Three point mutations that disrupted the splice donor site of intron 14 were membranous staining, whereas the other mutations were cytoplasmic staining. Patients with MET exon 14 splice site mutations were significantly older. The incidence of MET exon 14 mutations in sarcomatoid carcinoma was significantly higher than in other histologic types (P = .034). Conclusion Distinct MET protein localization is associated with MET exon 14 mutation types in patients with NSCLC. Different MET exon 14 mutation types were identified in a subset of Chinese patients with NSCLC who could possibly benefit from MET targeted therapy.

Objective Long non-coding RNA (lncRNA) HOXC-AS3 has been characterized as a cancer-related lncRNA in many types of cancer, while its role in colorectal cancer (CRC) is unknown. Methods The expression of HOXC-AS3 and TGF-β2 were detected by RT-qPCR. Overexpression assays were performed to explore the interaction between HOXC-AS3 and TGF-β2. A follow-up study was performed to explore the prognostic value of HOXC-AS3 for CRC. The direct interaction between HOXC-AS3 and miR-1269 was assessed with RNA-RNA pulldown assay. Transwell assays were performed to determine the role of HOXC-AS3 and TGF-β2 in regulating CRC cell invasion and migration. Results HOXC-AS3 was significantly downregulated in CRC tissues, while TGF-β2 was significantly upregulated in CRC tissues compared to that in adjacent non-cancer tissues of CRC patients. The follow-up study showed that low expression levels of HOXC-AS3 in CRC tissues were closely correlated with poor survival. Correlation analysis showed that HOXC-AS3 and TGF-β2 were inversely correlated across CRC tissues but not non-cancer tissues. Overexpression of HOXC-AS3 in the two cell lines resulted in downregulation of TGF-β2, while the expression of HOXC-AS3 was not affected by TGF-β2. Transwell migration and invasion assay showed that overexpression of TGF-β2 increased cell invasion and migration, while overexpression of HOXC-AS3 decreased cell migration and invasion. In addition, overexpression of HOXC-AS3 attenuated the effects of overexpression of TGF-β2. MiR-1269 increased the expression of TGF-β2. HOXC-AS3 directly interacted with miR-1269 in CRC cells. Conclusions Upregulation of HOXC-AS3 inhibited TGF-β2-induced colorectal cancer (CRC) cell migration and invasion possibly by sponging miR-1269.

Purpose/objective(s) To investigate the timing and pattern of recurrence in stage IIIA-N2 non-small cell lung cancer (NSCLC) patients after complete resection and adjuvant chemotherapy. Materials/methods The study cohort included pathologically confirmed stage IIIA-N2 NSCLC patients treated with complete resection and adjuvant chemotherapy from 2003 to 2015 in our single institution. Recurrence was categorized as local-regional recurrence (LRR), distant metastasis (DM) and both LRR and DM. The risk distribution was assessed by using clinical and pathological factors. The hazard rate function and competing risk analysis were used to evaluate the recurrence dynamics. The Gray's test was employed to estimate the cumulative recurrence rates and compare the differences between groups. Results Among 854 patients, 61.9% had multiple station N2 involvement. The 1, 3 and 5-year cumulative incidence rates of recurrence was 16.85%, 50.74% and 67.37%, respectively. Of the 510 patients who experienced recurrence, 95 (18.6%) experienced LRR, 285 (55.9%) experienced DM, whereas 130 (25.5%) had both LRR and DM. The hazard rate function for overall recurrence revealed a continuous increase between 0-18 months after surgery, consistent high level during 8-48 months and marked decline thereafter. And the DM displayed a hazard rate curve similar to that of overall recurrence. However, a double-peaked pattern of hazard rate was present in LRR and both LRR and DM. What's more, the peak recurrence frequency of DM differed by organs. A comparison of clinical and pathological factors revealed that patients with lower pT stage, single pN2 station and postoperative radiotherapy had a lower recurrence risk but similar pattern of recurrence. Conclusion The recurrence risk of IIIA-N2 NSCLC after complete resection and adjuvant chemotherapy increased to a high level in 18 months and maintained continuously high till 48 months after surgery, which hinted necessity of intensive follow-up during this period of time. This follow-up strategy implied an individualized surveillance for N2 disease which was different from clinical routine, and should be verified in further studies.

Objective To explore the protective effect and possible mechanism of Astragalus immortal prescription (AIP) from hepatic oxidative stress in mice irradiated by X ray. Methods Six to eight weeks old Kunming mice were randomly divided into normal control group, irradiation alone group, positive control group, and low, medium and high dose groups of AIP (10 rats in each group), physiological saline, thymus peptide and different concentration decocta of AIP were given respectively. After continuous gavage for 10 days, the whole body was irradiated with X ray at the dose of 8 Gy at a time, and then continued lavaging for 3 days, with weighing the mice and observing the diet, water intake and general condition. After killing the mice, the liver was take and the liver index was calculated; the morphological changes of the liver tissues were observed and the expressions of superoxide dismutase (SOD), malondiadehyde (MDA), glutathione peroxidase (GSH-Px) and nuclear factor erythroid 2-related factor 2 (Nrf2) in the liver tissues were detected. Results Compared with the normal control group, the weight of mice in model group decreased obviously (P

Microbial communities in subsurface soil are specialized for their environment, which is distinct from that of the surface communities. However, little is known about the microbial communities (bacteria and fungi) that exist in the deeper soil horizons. Vertical changes in microbial alpha-diversity (Chao1 and Shannon indices) and community composition were investigated at four soil depths (0-10, 10-20, 20-40, and 40-60 cm) in a natural secondary forest of Betula albosinensis by high-throughput sequencing of the 16S and internal transcribed spacer rDNA regions. The numbers of operational taxonomic units (OTUs), and the Chao1 and Shannon indices decreased in the deeper soil layers. Each soil layer contained both mutual and specific OTUs. In the 40-60 cm soil layer, 175 and 235 specific bacterial and fungal OTUs were identified, respectively. Acidobacteria was the most dominant bacterial group in all four soil layers, but reached its maximum at 40-60 cm (62.88%). In particular, the 40-60 cm soil layer typically showed the highest abundance of the fungal genus Inocybe (47.46%). The Chao1 and Shannon indices were significantly correlated with the soil organic carbon content. Redundancy analysis indicated that the bacterial communities were closely correlated with soil organic carbon content (P = 0.001). Collectively, these results indicate that soil nutrients alter the microbial diversity and relative abundance and affect the microbial composition.