Dual Target of EGFR and mTOR Suppresses Triple-Negative Breast Cancer Cell Growth by Regulating the Phosphorylation of mTOR Downstream Proteins.

Objective: To detect the activation of the EGFR and mTOR signaling pathways in the triple negative breast cancer cell line MDA-MB-468 and investigate the inhibitory effect of gefitinib, an epidermal growth factor receptor inhibitor, and everolimus, a target protein inhibitor of rapamycin, on triple negative breast cancer cells. Methods: Triple negative human breast cancer MDA-MB-468 cells were cultured and blank control group, single EGFR inhibitor gefitinib group, single mTOR inhibitor everolimus group, and two drug combination group were set up respectively to detect the effects of single and combined drugs on cell proliferation activity, cell cycle and apoptosis, and the expression of EGFR and mTOR signal pathway proteins in cell lines after single and combined drug intervention was detected again by Western blot. Results: The level of EGFR and p-mTOR protein in triple negative breast cancer was higher than in non triple negative breast cancer (P<0.05). The level of mTOR, S6K1, p-EGFR, p-S6K1 was significantly increased when treated with EGF (0ng/mL, 10ng/mL, 100ng/mL) for 1h, compared to without EGF stimulation (P<0.05). The level of p-EGFR, p-mTOR, p-S6K1 protein increased significantly when the cells were exposed to EGF for 2h, respectively (P<0.05). EGFR inhibitor gefitinib alone and the mTOR inhibitor everolimus alone could significantly inhibit the proliferation of human triple negative breast cancer MDA-MB-468 cells in a dose-dependent manner (P<0.05). The level of p-4EBP1 protein in EGFR and mTOR signal pathway was significantly increased after the intervention of gefitinib alone, everolimus alone, and the combination of two drugs (P<0.05). Conclusion: EGFR and mTOR signaling pathways can be activated in triple negative breast cancer; Both the EGFR inhibitor gefitinib alone and the mTOR inhibitor everolimus alone can significantly inhibit the proliferation of human triple negative breast cancer MDA-MB-468 cells. The combination of the EGFR inhibitor gefitinib and the mTOR inhibitor everolimus may achieve anti-tumor effect similar to that of single drug by reducing the drug dose.

CIABNet: Category imbalance attention block network for the classification of multi-differentiated types of esophageal cancer.

BACKGROUND: Esophageal cancer has become one of the important cancers that seriously threaten human life and health, and its incidence and mortality rate are still among the top malignant tumors. Histopathological image analysis is the gold standard for diagnosing different differentiation types of esophageal cancer. PURPOSE: The grading accuracy and interpretability of the auxiliary diagnostic model for esophageal cancer are seriously affected by small interclass differences, imbalanced data distribution, and poor model interpretability. Therefore, we focused on developing the category imbalance attention block network (CIABNet) model to try to solve the previous problems. METHODS: First, the quantitative metrics and model visualization results are integrated to transfer knowledge from the source domain images to better identify the regions of interest (ROI) in the target domain of esophageal cancer. Second, in order to pay attention to the subtle interclass differences, we propose the concatenate fusion attention block, which can focus on the contextual local feature relationships and the changes of channel attention weights among different regions simultaneously. Third, we proposed a category imbalance attention module, which treats each esophageal cancer differentiation class fairly based on aggregating different intensity information at multiple scales and explores more representative regional features for each class, which effectively mitigates the negative impact of category imbalance. Finally, we use feature map visualization to focus on interpreting whether the ROIs are the same or similar between the model and pathologists, thus better improving the interpretability of the model. RESULTS: The experimental results show that the CIABNet model outperforms other state-of-the-art models, which achieves the most advanced results in classifying the differentiation types of esophageal cancer with an average classification accuracy of 92.24%, an average precision of 93.52%, an average recall of 90.31%, an average F1 value of 91.73%, and an average AUC value of 97.43%. In addition, the CIABNet model has essentially similar or identical to the ROI of pathologists in identifying histopathological images of esophageal cancer. CONCLUSIONS: Our experimental results prove that our proposed computer-aided diagnostic algorithm shows great potential in histopathological images of multi-differentiated types of esophageal cancer.

HCCANet: histopathological image grading of colorectal cancer using CNN based on multichannel fusion attention mechanism.

Histopathological image analysis is the gold standard for pathologists to grade colorectal cancers of different differentiation types. However, the diagnosis by pathologists is highly subjective and prone to misdiagnosis. In this study, we constructed a new attention mechanism named MCCBAM based on channel attention mechanism and spatial attention mechanism, and developed a computer-aided diagnosis (CAD) method based on CNN and MCCBAM, called HCCANet. In this study, 630 histopathology images processed with Gaussian filtering denoising were included and gradient-weighted class activation map (Grad-CAM) was used to visualize regions of interest in HCCANet to improve its interpretability. The experimental results show that the proposed HCCANet model outperforms four advanced deep learning (ResNet50, MobileNetV2, Xception, and DenseNet121) and four classical machine learning (KNN, NB, RF, and SVM) techniques, achieved 90.2%, 85%, and 86.7% classification accuracy for colorectal cancers with high, medium, and low differentiation levels, respectively, with an overall accuracy of 87.3% and an average AUC value of 0.9.In addition, the MCCBAM constructed in this study outperforms several commonly used attention mechanisms SAM, SENet, SKNet, Non_Local, CBAM, and BAM on the backbone network. In conclusion, the HCCANet model proposed in this study is feasible for postoperative adjuvant diagnosis and grading of colorectal cancer.

Expression and prognosis analysis of GINS subunits in human breast cancer.

ABSTRACT: GINS subunits, a protein complex composed of GINS1, GINS2, GINS3 and GINS4 in the human genome and the expression level of each GINS subunits plays an important role in different human cancers. As one of the most common malignancies after lung cancer in the world, precise biomarkers for early diagnosis and treatment in breast cancer are important. The purpose of our study was to elucidate the expression and prognostic value of GINS subunits in breast cancer.The purpose of present study was to explore the expression level of GINS subunits in breast cancer patients.In the present study, we investigated the gene alteration, gene expression and potential prognostic value of GINS subunits by using the Gene Expression Profiling Interactive Analysis (GEPIA), UALCAN, cBioPortal, and bc-GenExMiner databases. Then, the GeneMANIA database was used to show the genes that associated with GINS subunits. Furthermore, gene ontology pathway analysis was conducted by using the Metascape database. Finally, immune infiltration analysis in GINS subunits were evaluated using the Tumor Immune Estimation Resource (TIMER) database.Our analyses demonstrated that the expression levels of different GINS subunits were different between breast cancer and normal breast tissues. The expression levels of GINS1, GINS2, and GINS4 were significantly higher in breast cancer tissues than in normal tissues. Survival analysis revealed that increased the expression levels of GINS subunits were associated with poor prognoses in all patients with breast cancer. Gene ontology pathway enrichment analysis of the GINS subunits suggested that GINS subunits involved in pathways including the cell cycle checkpoint, DNA replication and other meaningful signaling pathways.We systemically analyzed the expression, prognostic, clinicopathologic values, and potential functional networks of GINS subunits in breast cancer. Our findings showed that individual GINS subunits could be new potential prognostic biomarkers for breast cancer. However, further verification studies are still needed to prove the clinical value of GINS subunits in breast cancer patients.

Clinical significance of 'double-hit' and 'double-expression' lymphomas.

BACKGROUND: 'Double-hit' lymphoma (DHL) and 'double-expression' lymphoma (DEL) involving gene rearrangement and protein expression of MYC and BCL2/BCL6 have recently become the most commonly used terms to describe the poor prognostic types of diffuse large B-cell lymphoma (DLBCL). However, the clinical and pathological spectra of these rare entities have not been well defined. The aim of this study was to determine the frequency of DHL and DEL in DLBCL and their prognostic impacts in the era of cyclophosphamide, doxorubicin, vincristine and prednisone plus rituximab therapy. METHODS: The data and tissues from 98 patients diagnosed with DLBCL were used in this study. Formalin-fixed and paraffin-embedded tissues were constructed for immunohistochemistry (IHC) and interphase fluorescene in situ hybridisation (FISH) analysis for MYC, BCL6 and BCL2 rearrangements. RESULTS: There were 14 cases (14.29%, 14/98) and 34 cases (34.70%, 34/98) of lymphomas with the DHL and DEL of MYC and BCL2/BCL6, respectively. DLBCL patients with MYC/BCL2 rearrangements more frequently had bone marrow involvement (p=0.002), and their tumours were commonly of the germinal centre B cell (GCB) subtype. Patients with MYC+/BCL2 +coexpression were more often assessed using the International Prognostic Index (IPI), (Performance Status) PS score and bone marrow involvement. Patients with MYC+/BCL6 +coexpression were associated with their IPI values, Eastern Cooperative Oncology Group scores and occurrence of B symptoms. Their lymphomas were more often of the non-GCB subtype (p=0.010). Multivariate analysis showed that IPI, bone marrow involvement, rearrangements of BCL2, BCL6 and MYC, expression concurrent with rearrangements and coexpression were all significantly associated with overall survival and progression-free survival, with the exception of MYC+/BCL6 +coexpression. CONCLUSIONS: MYC/BCL2 DHL, MYC/BCL6 DHL and MYC/BCL2 DEL are an aggressive B cell lymphoma and patients have a poor prognosis. IPI and bone marrow involvement were independent prognostic factors for DHL and DEL. BCL2, BCL6 and MYC rearrangements translocation, expression concurrent rearrangements and coexpression were were independent prognostic factors for survival. POTENTIAL IMPLICATIONS: The present study analysed the genomic alterations and protein expression levels of MYC, BCL2 and BCL6 using FISH and IHC in Chinese patients with DLBCL.We assessed the frequency, clinicopathological features and the prognostic impacts of DHL and DEL in a cohort of de novo DLBCL patients and evaluated the role of each genetic translocation separately and in combination in order to provide reliable conclusions and practical recommendations for diagnostic workups and prognostic predictions.

Clinicopathologic characteristics of non-small cell lung cancer patients with seldom mutation types of epidermal growth factor receptor and treatment analysis / 中国肿瘤临床

Objective:To compare clinicopathologic parameters of uncommon mutations of epidermal growth factor receptor (EGFR) and preliminary therapeutic effects of EGFR-TKI treatment in patients with non-small cell lung cancer. Methods:We collected clinico-pathological data from 29 patients with non-small cell lung cancer who carry uncommon mutations of EGFR, which were pathological-ly confirmed in the Tumor Hospital Affiliated to Xinjiang Medical University, from January 2012 to April 2016. Then we analyzed the re-lationship between the clinicopathologic characteristics of uncommon mutations and therapeutic effects of EGFR-TKIs. Results:Among the 29 cases of patients with uncommon mutations, the most common distant metastasis organs were ipsilateral/contralateral lung tissue, bone, brain, liver, and adrenal gland;the most common metastatic lymph nodes were hilar lymph node, supraclavicular/subclavian lymph node, neck-root lymph node, and mediastinal lymph node. In seldom mutations, 16 cases of single mutation were found:5 cases of L861Q, 5 cases of G719X, 4 cases of 20ins, and 2 cases of S768I. By contrast, 11 cases of double mutations were found:4 cases of S768I and 20ins, 1 case of double mutation of L858R and S768I, 1 case of double mutation of 19Del and T790M, 2 cases of double mutations of L861Q and G719X, 1 case of 19Del and S768I, 1 case of 20ins and G719X, and 1 case of T790M and G719X. Moreover, 2 cases of triple mutation were found:1 case of L858R, L861Q, and G719X;1 case of S768I, 20ins, and G719X. The objective response rate (ORR) of the first-line EGFR-TKI therapy was 43.75%, the disease control rate (DCR) was 50%, and the median progression-free survival (mPFS) was 5.5 months. Furthermore, the ORR of the second-line EGFR-TKI therapy was 28.57%, the DCR was 42.85%, and the mPFS was 4 months. Moreover, the ORR of the third-line EGFR-TKI therapy was 33.33%, the DCR was 50.00%, and the mPFS was 2.67 months. Conclusion:Great individual differences were found on EGFR uncommon mutations for effective rate and sur-vival time of EGFR-TKI treatment;in general, ORR and mPFS of EGFR seldom mutations were lower than classical mutations and partly higher than wild types. The first-line therapeutic effects of EGFR-TKI therapy was slightly better than the second-line or third-line thera-peutic effects;however, no significant statistical difference was observed .

Optomizing Transfection Efficiency of Cervical Cancer Cells Transfected by Cationic Liposomes LipofectamineTM2000.

BACKGROUND: Currently, cationic liposome has become the commonly used vehicles for gene transfection. Furthermore, one of the most significant steps in microRNAs expression studies is transferring microRNAs into cell cultures successfully. In this study we aim to approach the feasibility of transfection of cervical cancer cell lines mediated by liposome and to obtain the optimized transfection condition for cervical cancer cell lines. MATERIALS AND METHODS: Lipofectamine(TM)2000 as the carrier, miR-101 mimic was transfected into Hela cells and Siha cells. Using green fluorescent protein as reporter gene, to set different groups according to cell seeding density, the amount of miRNA , miRNA and the proportion of Liposomes, Whether to add serum into medium to study their impact on the liposomal transfection efficiency. Finally, MTT assay was used to analyze the relative minimal cell toxicity of liposome reagents. RESULTS: The seeding density of Hela cell line and Siha are 1.5 x 10(4) (per well of 24 well plates), miRNA amount is 1ul of both, the ratio of miRNA and liposome is 1:0.5 of Hela cell line; 1:0.7 of Siha cell line respectively, after 24 hours we can get the highest transfection efficiency. Compared with serum medium, only Siha cells cultured with serum-free medium obtained higher transfection efficiency before transfection (P<0.01).MTT assay showed that according to the above conditions which has the lowest cytotoxicity. CONCLUSIONS: The method of Liposome to transfected is a suitable way and it can be an efficient reagent for miRNA delivery for Hela cells and Siha cells in vitro. It may serve as a reference for the further research or application.

Syntheses and characterization of nearly monodispersed, size-tunable silver nanoparticles over a wide size range of 7-200 nm by tannic acid reduction.

Nearly monodispersed spherical silver nanoparticles (Ag NPs) were synthesized by using tannic acid (TA) as both reductant and stabilizer in a 30 °C water bath. The size of the as-prepared Ag NPs could be tuned in a range of 7-66 nm by changing the molar ratio of TA to silver nitrate and pH of the reaction solutions. UV-vis spectra, TEM observations, and temporal evolution of the monomer concentrations for the reactions carried out at different experimental conditions showed that the improved size distribution and size tunability of the Ag NPs were mainly attributed to the use of TA, which could promote the balance of nucleation and growth processes of the NPs effectively. The size of the Ag NPs was extendable up to 200 nm in one-pot fashion by the multi-injection approach. The size-dependent surface-enhanced Raman scattering (SERS) activity of the as-prepared Ag NPs was evaluated, and the NPs with size around 100 nm were identified to show a maximum enhanced factor of 3.6 × 10(5). Moreover, the as-prepared TA-coated Ag NPs presented excellent colloidal stability compared to the conventional citrate-coated ones.