RACK1 mediates NLRP3 inflammasome activation during Pasteurella multocida infection.

Pasteurella multocida is a gram-negative bacterium that causes serious diseases in a wide range of animal species. Inflammasomes are intracellular multimolecular protein complexes that play a critical role in host defence against microbial infection. Our previous study showed that bovine P. multocida type A (PmCQ2) infection induces NLRP3 inflammasome activation. However, the exact mechanism underlying PmCQ2-induced NLRP3 inflammasome activation is not clear. Here, we show that NLRP3 inflammasome activation is positively regulated by a scaffold protein called receptor for activated C kinase 1 (RACK1). This study shows that RACK1 expression was downregulated by PmCQ2 infection in primary mouse peritoneal macrophages and mouse tissues, and overexpression of RACK1 prevented PmCQ2-induced cell death and reduced the numbers of adherent and invasive PmCQ2, indicating a modulatory role of RACK1 in the cell death that is induced by P. multocida infection. Next, RACK1 knockdown by siRNA significantly attenuated PmCQ2-induced NLRP3 inflammasome activation, which was accompanied by a reduction in the protein expression of interleukin (IL)-1ß, pro-IL-1ß, caspase-1 and NLRP3 as well as the formation of ASC specks, while RACK1 overexpression by pcDNA3.1-RACK1 plasmid transfection significantly promoted PmCQ2-induced NLRP3 inflammasome activation; these results showed that RACK1 is essential for NLRP3 inflammasome activation. Furthermore, RACK1 knockdown decreased PmCQ2-induced NF-κB activation, but RACK1 overexpression had the opposite effect. In addition, the immunofluorescence staining and immunoprecipitation results showed that RACK1 colocalized with NLRP3 and that NEK7 and interacted with these proteins. However, inhibition of potassium efflux significantly attenuated the RACK1-NLRP3-NEK7 interaction. Our study demonstrated that RACK1 plays an important role in promoting NLRP3 inflammasome activation by regulating NF-κB and promoting NLRP3 inflammasome assembly.

Predictive Nomogram for Hyperprogressive Disease During Anti-PD-1/PD-L1 Treatment in Patients with Advanced Non-Small Cell Lung Cancer.

Introduction: Various studies have reported that anti-PD-1/PD-L1 treatment may lead to the rapid development of tumors called hyperprogressive disease (HPD). A nomogram for HPD prediction in NSCLC patients is urgently needed. Methods: This retrospective cohort study included 176 cases for establishing a model of HPD prediction and 85 cases for validation in advanced NSCLC patients treated with PD-1/PD-L1 inhibitors. HPD was defined as tumor growth rate (TGR, ≥ 2), tumor growth kinetics (TGK, ≥ 2) or time to treatment failure (TTF, ≤ 2 months). Univariate and multivariate logistic regression were used to estimate the specified factors associated with HPD. Then, the nomogram was developed and validated. Results: Anti-PD-1/PD-L1 therapy resulted in a 9.66% (17/176) incidence of HPD in advanced NSCLC. The overall survival (OS) and progression-free survival (PFS) in patients with HPD were significantly shorter than those in patients without HPD (OS: 7.00 vs 12.00 months, P<0.01; PFS: 2.00 vs 5.00 months, P<0.001, respectively). The HPD prediction nomogram included APTT (P<0.01), CD4+ CD25+ CD127-low cells (Treg cells) (P<0.01), the presence of liver metastasis (P<0.05), and more than two metastatic sites (P<0.05). Then, patients were divided into two groups by the "HPD score" calculated by the nomogram. The C-index was 0.845, while the area under the curve (AUC) was 0.830 (sensitivity 75.00%, specificity 91.70%). The calibration plot of HPD probability showed an optimal agreement between the actual observation and prediction by the nomogram. In the validation cohort, the AUC was up to 0.960 (sensitivity 88.70%, specificity 89.80%). Conclusions: The nomogram was constructed with the presence of liver metastasis, more than two metastatic sites, lengthened APTT and a high level of Treg cells, which could be used to predict HPD risk.

The important role of NLRP6 inflammasome in Pasteurella multocida infection.

Pasteurella multocida (P. multocida) can cause severe respiratory disease in cattle, resulting in high mortality and morbidity. Inflammasomes are multiprotein complexes in the cytoplasm that recognize pathogens and play an important role in the host defense against microbial infection. In this study, the mechanism of P. multocida-induced NLRP6 inflammasome activation was investigated in vitro and in vivo. Firstly, P. multocida induced severe inflammation with a large number of inflammatory cells infiltrating the lungs of WT and Nlrp6-/- mice. Nlrp6-/- mice were more susceptible to P. multocida infection and they had more bacterial burden in the lungs. Then, the recruitment of macrophages and neutrophils in the lungs was investigated and the results show that the number of immune cells was significantly decreased in Nlrp6-/- mice. Subsequently, NLRP6 was shown to regulate P. multocida-induced inflammatory cytokine secretion including IL-1ß and IL-6 both in vivo and in vitro while TNF-α secretion was not altered. Moreover, NLRP6 was found to mediate caspase-1 activation and ASC oligomerization, resulting in IL-1ß secretion. Furthermore, NLRP6 inflammasome mediated the gene expression of chemokines including CXCL1, CXCL2 and CXCR2 which drive the activation of NLRP3 inflammasomes. Finally, NLRP3 protein expression was detected to be abrogated in P. multocida-infected Nlrp6-/- macrophages, indicating the synergic effect of NLRP6 and NLRP3. Our study demonstrates that NLRP6 inflammasome plays an important role in the host against P. multocida infection and contributes to the development of immune therapeutics against P. multocida.

The Critical Role of Potassium Efflux and Nek7 in Pasteurella multocida-Induced NLRP3 Inflammasome Activation.

Pasteurella multocida is a zoonotic pathogen causing respiratory infection in different animal species such as cattle, sheep, pigs, chickens and humans. Inflammasome is a complex assembled by multiple proteins in the cytoplasm and plays an important role in the host defense against microbial infection. Bovine Pasteurella multocida type A (PmCQ2) infection induces NLRP3 inflammasome activation and IL-1ß secretion, but the mechanism of PmCQ2-induced activation of NLRP3 inflammasome is still unknown. Therefore, the underlying mechanism was investigated in this study. The results showed that potassium efflux mediated PmCQ2-induced IL-1ß secretion and blocking potassium efflux attenuated PmCQ2-induced caspase-1 activation and ASC oligomerization. Furthermore, NIMA-related kinase 7 (Nek7) was also involved in PmCQ2-induced caspase-1 activation and IL-1ß secretion. In addition, PmCQ2 infection promoted Nek7-NLRP3 interaction, which is dependent on potassium efflux. In conclusion, our results indicate the critical role of potassium efflux and Nek7 in Pasteurella multocida-induced NLRP3 inflammasome activation, which provides useful information about Pasteurella multocida-induced host immune response.

The Critical Role of NLRP6 Inflammasome in Streptococcus pneumoniae Infection In Vitro and In Vivo.

Streptococcus pneumoniae (S. pneumoniae) causes severe pulmonary diseases, leading to high morbidity and mortality. It has been reported that inflammasomes such as NLR family pyrin domain containing 3 (NLRP3) and absent in melanoma 2 (AIM2) play an important role in the host defense against S. pneumoniae infection. However, the role of NLRP6 in vivo and in vitro against S. pneumoniae remains unclear. Therefore, we investigated the role of NLRP6 in regulating the S. pneumoniae-induced inflammatory signaling pathway in vitro and the role of NLRP6 in the host defense against S. pneumoniae in vivo by using NLRP6-/- mice. The results showed that the NLRP6 inflammasome regulated the maturation and secretion of IL-1ß, but it did not affect the induction of IL-1ß transcription in S. pneumoniae-infected macrophages. Furthermore, the activation of caspase-1, caspase-11, and gasdermin D (GSDMD) as well as the oligomerization of apoptosis-associated speck-like protein (ASC) were also mediated by NLRP6 in S. pneumoniae-infected macrophages. However, the activation of NLRP6 reduced the expression of NF-κB and ERK signaling pathways in S. pneumoniae-infected macrophages. In vivo study showed that NLRP6-/- mice had a higher survival rate, lower number of bacteria, and milder inflammatory response in the lung compared with wild-type (WT) mice during S. pneumoniae infection, indicating that NLRP6 plays a negative role in the host defense against S. pneumoniae. Furthermore, increased bacterial clearance in NLRP6 deficient mice was modulated by the recruitment of macrophages and neutrophils. Our study provides a new insight on S. pneumoniae-induced activation of NLRP6 and suggests that blocking NLRP6 could be considered as a potential therapeutic strategy to treat S. pneumoniae infection.

Superiority of fecal carcinoembryonic antigen as diagnosis marker for adenomatous polyposis coli and asymptomatic colorectal cancer.

BACKGROUND: Non-invasive diagnostic tools of adenomatous polyposis coli (APC) and asymptomatic colorectal cancer (CRC) are urgently needed. Although fecal carcinoembryonic antigen (FCEA) has been documented in some studies, the diagnostic potential for the detection of APC and asymptomatic CRC has not been described yet. METHODS: This is a retrospective study. The pre-diagnostic serum carcinoembryonic antigen (SCEA) and fecal occult blood test (FOBT) levels were retrospectively analyzed in 212 patients with intestinal diseases group (IDG) and 224 controls. The levels of FCEA across all the studied groups were measured using electronic chemiluminescence immunoassay (ECLIA), and their sensitivity and specificity were used to evaluate their diagnostic potential. The individual diagnostic accuracy of the three indices, as well as their combined diagnostic potential, was compared using the receiver operating characteristic (ROC) curve and chi-square test. RESULTS: The FCEA had low sensitivity (50%) and high specificity (93.91%) for the diagnosis of IDG, with the area under the curve (AUC) value of 0.781. The AUC of FCEA was higher than that of SCEA for the diagnosis of APC and CRC in the APC, asymptomatic CRC, and APC + CRC-stage I patients. The AUCs of FCEA were 0.708 and 0.691 for the 'double-negative patients' and 'triple-negative patients', respectively. In addition, FCEA could diagnose 45.5% of the 'double-negative' patients, 43.3% of the asymptomatic patients, and 42.9% of the 'triple-negative' patients. The combination of FCEA and FOBT improved the diagnostic value (AUC = 0.916). CONCLUSION: FCEA has been demonstrated to be a favorable diagnostic marker in intestinal diseases, especially in the APC, asymptomatic CRC, and 'double-negative' or 'triple-negative' CRC patients.

Evaluation of 2 Commercially Systems for Detection of Serum Squamous Cell Carcinoma Antigen in Pan Squamous Cell Carcinoma.

BACKGROUND: This study compared the analytical performance of the Elecsys 602 (Roche Diagnostics) system with the I2000 (Abbott laboratories) system for the quantitative measurement of squamous cell carcinoma antigen (SCCA) to assess its role as an indicator in pan squamous cell carcinoma. METHODS: 435 serum samples included pan squamous cell cancer group (n = 318) and healthy subjects (n = 52) and non-squamous cell group (n = 41) and benign diseases group (n = 24) were measured by 2 systems and compared. RESULTS: The within-run precision coefficient of variation (CV) for Abbott and Roche systems were 3.34-4.88% and 0.95 -1.96%, and the total precision CV were 2.89-9.48% and 3.97-5.38%, respectively. Good correlation was showed in Abbott and Roche systems (slopes = 0.749, r = 0.9658). Serum SCCA in the groups of nasopharyngeal carcinomas, lung squamous cell carcinoma, esophageal squamous cell carcinoma, bladder cancer and cervical squamous cell carcinoma under the curve area (AUC) was more than 0.5, while the AUC in the non- nasopharyngeal carcinomas head and neck squamous cell carcinoma was less than 0.5. The AUC of 2 systems was statistically different in lung squamous cell carcinoma and nasopharyngeal carcinomas (P < 0.05). The levels of SCCA of 2 systems were similarities in esophageal squamous cell carcinoma(stage IV vs. stage 0a-II)and bladder cancer(stage I vs. stage Oa)and cervical squamous cell carcinoma(stage IIB-III vs. stage I-IIA), which advanced stage had higher level of SCCA than early stage. But the SCCA levels of 2 systems were inconsistent in bladder cancer (stage II-IV vs. stage Oa in Abbott), head and neck squamous cell carcinoma (stage IV vs. stage Oa-I in the Roche) and lung squamous cell carcinoma (stage III vs. stage I-II in the Roche). (P < 0.05). CONCLUSIONS: 2 systems correlated well in SCCA detection of squamous cell carcinoma, but there were individual differences. Serum SCCA may also contribute to the diagnosis of bladder cancer.

Downregulation of miR-182-5p inhibits the proliferation and invasion of triple-negative breast cancer cells through regulating TLR4/NF-κB pathway activity by targeting FBXW7.

BACKGROUND: To investigate the role of miR-182-5p in the proliferation and invasion of triple-negative breast cancer (TNBC) cells, as well as the underlying mechanism. METHODS: qRT-PCR was used to detect the level of miR-182-5p in tissues and cells. CCK-8 assay, flow cytometry, and Transwell assay were performed to detect cell proliferation, apoptosis rate, and invasion, respectively. Pearson correlation analysis was used to determine the correlation between miR-182-5p and its predicted target gene FBXW7, and the target of miR-182-5p was identified by dual-luciferase reporter gene assay. ELISA assay was used to examine the levels of cytokines in the culture supernatant of tumor cells. Western blot analysis was used to detect the expressions of FBXW7, TLR4, and NF-κB) pathways in tumor cells. RESULTS: In MDA-MB-231 and BT-549 cells, downregulation of miR-182-5p significantly inhibited cell proliferation and invasion and promoted tumor cell apoptosis. Pearson correlation analysis showed that miR-182-5p had a negative correlation with FBXW7. Dual-luciferase reporter gene assay showed that miR-182-5p could directly target FBXW7. Further studies showed that FBXW7 overexpression significantly inhibited cell proliferation and invasion and increased the apoptosis rate. Downregulation of miR-182-5p significantly reduced the levels of TNF-α, IL-1 ß, IL-6, and IL-18 in the culture supernatant, and decreased the activity of TLR4/NF-κB pathway in tumor cells, while downregulation of FBXW7 significantly inhibited the effect of miR-182-5p on tumor cells. CONCLUSIONS: Downregulation of miR-182-5p inhibits TLR4/NF-κB pathway activity by increasing FBXW7 expression, thereby suppressing the proliferation and invasion of TNBC cells.

Apatinib sensitizes chemoresistant NSCLC cells to doxetaxel via regulating autophagy and enhances the therapeutic efficacy in advanced and refractory/recurrent NSCLC.

The prognosis of advanced non­small cell lung cancer (NSCLC) is poor; therefore, identifying novel treatment strategies for patients with NSCLC is important. The present study aimed to investigate the efficacy of apatinib plus docetaxel vs. docetaxel alone, as well as their effects on regulating autophagy markers in patients with advanced NSCLC. Furthermore, it was evaluated whether apatinib sensitized chemoresistant NSCLC cells to docetaxel via regulating autophagy. A total of 39 patients with advanced NSCLC were consecutively enrolled and treated with apatinib plus docetaxel (n=19) or docetaxel alone (n=20) for four treatment cycles. The treatment response, adverse events and expression levels of autophagy markers [(light chain 3 α (LC3A) and Beclin­1] were evaluated in tumor samples, which were obtained via biopsy, before treatment and after 2­cycle treatment. In addition, in a mechanistic in vitro experiment, apatinib, docetaxel, the autophagy activator rapamycin and the autophagy inhibitor 3­methyladenine (3­MA) were used to treat docetaxel­resistant A549 (A549/DTX) cells alone or in various combinations. The expression levels of LC3A, Beclin­1, poly (ADP) ribose polymerase (PARP) and phosphorylated (p)­AKT were detected via western blotting, while the cell apoptosis rate was detected with an Annexin V/PI assay. The overall remission rate (37 vs. 10%; P=0.047) and disease control rate (84 vs. 45%; P=0.011) were increased in the Apatinib plus docetaxel group compared with the Docetaxel group. Most of the adverse events were mild and tolerable, and there was no difference between the two groups except for total hypertension and hand­foot syndrome, which were higher in the Apatinib plus docetaxel group). Compared with the levels prior to treatment, Beclin­1 and LC3A remained unchanged post­treatment in the Apatinib plus docetaxel group, while they were increased in the Docetaxel group. Docetaxel increased LC3A, Beclin­1 and p­AKT expression levels, PARP cleavage and the cell apoptosis rate in A549/DTX cells, and rapamycin further enhanced, while 3­MA reduced these effects of docetaxel. Moreover, apatinib repressed LC3A, Beclin­1, p­AKT expression levels and promoted the cell apoptosis rate in A549/DTX cells and docetaxel­treated A549/DTX cells. In conclusion, apatinib synergize the effect of docetaxel in treating patients with advanced NSCLC and chemoresistant NSCLC cells via inhibiting autophagy.

Protective effect of leaf extract of Abutilon indicum on DNA damage and peripheral blood lymphocytes in combating the oxidative stress.

The current research explores in vitro antioxidant characteristics, radiation-induced DNA damage protection and quenching effects of the oxidative stress by the ethanolic leaf extract of Abutilon indicum (EEAI) on human peripheral blood lymphocytes (PBLs). PBLs were incubated with various concentrations of EEAI accompanied by pre- and post-treatment with hydrogen peroxide. Cell viability was investigated by MTT assay. In addition, quenching of free radicals were measured in vitro using DPPH, superoxide anion, hydrogen peroxide, reducing power and nitric oxide radical scavenging assays. These activities were compared with ascorbic acid as standard antioxidants. Furthermore, inhibition of UV radiation-induced strand break formation in plasmid pBR322 DNA and anti-Fenton reactions in calf thymus DNA was evaluated. Cytotoxic effects of hydrogen peroxide on PBLs were significantly reduced with EEAI pre-treatment compared to post-treatment in a dose-dependent manner comparable with similar cytoprotective effects of ascorbic acid (p > 0.05). EEAI has shown strong antioxidant effects in the scavenging of DPPH, superoxide anion, hydrogen peroxide, and nitric oxide. EEAI also has a strong protective effect of UV-induced plasmid pBR322 DNA cleavage and Fenton-induced DNA damage. Overall, the results revealed that Abutilon indicum has a cytoprotective, potent antioxidant and DNA protective effect that provide pharmacological credence to justify its overall biological activity. Furthermore, future studies to identifying bioactive molecules and its molecular mechanisms responsible for promising therapeutic applications in the rescue of disease-induced cellular oxidative damage.

Target Inhibition of CBP Induced Cell Senescence in BCR-ABL- T315I Mutant Chronic Myeloid Leukemia.

The treatment of chronic myeloid leukemia (CML) with BCR-ABL tyrosine kinase inhibitors (TKIs), such as imatinib, has yielded clinical success. However, the direct targeting of BCR-ABL does not eradicate CML cells expressing mutant BCR-ABL, especially the T315I mutation in BCR-ABL. Moreover, increasing mutations were identified in BCR-ABL domain, resulting in TKIs resistance recently. It is necessary to find BCR-ABL-independent target for treating CML patients with various mutations, including T315I mutation in BCR-ABL. The dichotomous behavior of CREB binding protein (CBP) and E1A protein (p300), recruited by ß-catenin associated with self-renewal and differentiation, have been identified in hematopoietic stem cells, respectively. In this study, CBP was aberrantly expressed in CML cells on the basis of Oncomine dataset. The ß-catenin bound with much more CBP than p300 in CML cells. Down-regulation of CBP inhibited cell proliferation capacity and increased the binding of ß-catenin to p300, thus promoting cell differentiation and p53-dependent cell senescence in CML cells with either wild type or T315I mutant BCR-ABL in vitro and in vivo models. These demonstrate CBP blockage can be developed for the treatment of CML independent of BCR-ABL mutation status including T315I.

Atom Probe Tomography Characterization of Dopant Distributions in Si FinFET: Challenges and Solutions.

Atom probe tomography (APT) has emerged as an important tool in characterizing three-dimensional semiconductor devices. However, the complex structure and hybrid nature of a semiconductor device can pose serious challenges to the accurate measurement of dopants. In particular, local magnification and trajectory aberration observed when analyzing hybrid materials with different evaporation fields can cause severe distortions in reconstructed geometry and uncertainty in local chemistry measurement. To address these challenges, this study systematically investigates the effect of APT sampling directions on the measurement of n-type dopants P and As in an Si fin field-effect transistor (FinFET). We demonstrate that the APT samples made with their Z-axis perpendicular to the center axis of the fin are effective to minimize the negative effects that result from evaporation field differences between the Si fin and SiO2 on reconstruction and achieve improved measurement of dopant distributions. In addition, new insights have been gained regarding the distribution of ion-implanted P and As in the Si FinFET.

MicroRNA-7-5p induces cell growth inhibition, cell cycle arrest and apoptosis by targeting PAK2 in non-small cell lung cancer.

MicroRNAs (miR) are known to be critical regulators in tumor progression. miR-7-5p was reported to be involved in several cancers, including glioblastoma, cervical cancer, and melanoma, but its prognostic value and biological function in non-small-cell lung cancer (NSCLC) remain unclear. In this study, using quantitative real-time PCR analysis, we found that miR-7-5p was significantly downregulated in NSCLC tissues and cell lines. Lower miR-7-5p expression was associated with tumor-node-metastasis stage and tumor size by chi-squared test. Deceased miR-7-5p expression was associated with a worse prognosis in patients with NSCLC using Kaplan-Meier survival analysis and multivariate Cox regression analysis. Experiments in NSCLC cell lines (A549 and H1299) demonstrated that upregulation of miR-7-5p significantly suppressed cell proliferation, but induced cell cycle G0/G1 phase arrest and apoptosis using Cell Counting Kit-8, colony formation, and flow cytometry analysis. Through loss-of-function assays, we further demonstrated that downregulation of miR-7-5p promoted cell proliferation and cell cycle G1/S transition, but decreased cell apoptosis in SPC-A1 cells. Furthermore, P21-activated kinase 2 (PAK2) was predicted and confirmed as a direct target gene of miR-7-5p in NSCLC cells by luciferase reporter assay. In addition, we found PAK2 overexpression could partially reverse the effects of miR-7-5p on cell proliferation, cell cycle distribution, and apoptosis. We thus concluded that lower expression of miR-7-5p was associated with poor prognosis and NSCLC progression by directly targeting PAK2.

SphK1 functions downstream of IGF-1 to modulate IGF-1-induced EMT, migration and paclitaxel resistance of A549 cells: A preliminary in vitro study.

Insulin-like growth factor-1 (IGF-1) -induced epithelial-mesenchymal transition (EMT) plays a key role in the metastasis and drug resistance of non-small cell lung cancer (NSCLC). Sphingosine kinase-1 (SphK1) is also involved in EMT of NSCLC. However, the interaction between SphK1 and IGF-1 in the EMT of NSCLC is largely unknown. To clarify this issue, we examined the involvement of SphK1 in IGF-1-induced EMT using human lung cancer cell line A549, and its paclitaxel-resistant subline. Cell viability was evaluated by cell counting kit-8 assay; Migratory ability was examined using scratch wound healing test; Protein expression levels of SphK1, vimentin, fibronectin, N-cadherin and E-cadherin were detected by western blot analysis, respectively. The results showed that, IGF-1 treatment of A549 cells stimulated the expression of SphK1, the activation of ERK and AKT, the cell migration, and the expression of EMT hallmark proteins, while inhibition of SphK1 by its specific inhibitor SKI-II suppressed all the above changes and increased the sensitivity of A549 cells to paclitaxel. Our data demonstrate that SphK1 acts as a downstream effector of IGF-1 and plays a critical role in IGF-1-induced EMT, cell migration and paclitaxel resistance of A549 cells, suggesting that SphK1 might be a potential therapeutic target for NSCLC.

Serum hepatitis B viral (HBV) DNA is a predictive biomarker for survival in non-small cell lung cancer patients with chronic HBV infection.

Purpose: To study the association between pretreatment serum hepatitis B viral (HBV) DNA copy numbers and clinical outcome of non-small cell lung cancer (NSCLC) patients with chronic HBV infection. Patients and methods: We retrospectively evaluated the medical records of NSCLC HBV (+) patients between January 2008 and December 2010. The HBV DNA copy numbers and other prognostic factors including albumin (ALB), C-reactive protein (CRP), platelet, neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), and Glasgow Prognostic Score (GPS) were obtained before any antitumor treatment. Kaplan-Meier curves and the log-rank test were used to calculate prognostic significance. A multivariable Cox proportional hazard regression was modeled to analyze the independent prognostic factors for NSCLC HBV (+) patients. All independent prognostic factors in the Cox multivariable analysis were used to build a nomogram. The predictive accuracy of HBV DNA, TNM stage and nomogram was evaluated with the concordance index (C-index) and time-dependent receiver operating characteristics (ROC) curves, and simultaneously compared with traditional TNM staging system respectively. Results: A total of 188 patients were recruited in this study; the median age was 56 years, and the median overall survival (OS) was 34 months. Cox multivariate analysis results showed independent factors for OS including TNM stage (P=0.028), treatment (P=0.002), HBV DNA (P<0.001), and GPS (P=0.026). The nomogram model for survival was built based on four prognostic factors. The C-index for HBV DNA was 0.67, 0.69 for TNM stage, and 0.76 for the nomogram model. There was no statistical difference between HBV DNA and TNM stage (P=0.48). However, the C-index values of nomogram model were statistically higher both than HBV DNA (0.76 vs 0.67, P<0.001), and TNM stage (0.76 vs 0.69, P<0.001). Conclusion: Pretreatment serum HBV DNA copy numbers can act as a prognostic marker of survival for NSCLC patients with chronic HBV infection.

Study on the degradation rule of calcitonin in vitro in patients with medullary thyroid carcinoma.

BACKGROUND: It was found that calcitonin in vitro can degrade rapidly whether in refrigeration or at room temperature, which increases the difficulty of accurate detecting of calcitonin greatly. This study was designed to solve this problem. METHODS: Serum samples with increased calcitonin from patients with medullary thyroid carcinoma were collected and divided into refrigeration and room temperature group. Analyze the degradation rule of calcitonin in the two groups according to the time of blood draw (T0), the time of sample detection (Tx), and the concentration (y) measured at Tx. RESULTS: The degradation rate of calcitonin was significantly different between the two storage conditions. The degradation has plateau stage and its concentration is associated with the original concentration. The degradation rule of the two groups can be expressed by the formula: y = -aln(x) + b. In the formula, b/a are constants associated with storage condition and gender. "x" is the time interval between blood draw and detection, that is, x = Tx - T0. CONCLUSIONS: The significance of the presence of immunoreactive isoforms and fragments of calcitonin with different stability in patients with medullary thyroid carcinoma is unclear. To make the inspection report more accurate and reliable, maybe the inspection personnel should report both the instrumental and corrected result, and also indicate the correction formula on the inspection report.

Loss of MED12 Induces Tumor Dormancy in Human Epithelial Ovarian Cancer via Downregulation of EGFR.

A high rate of disease relapse makes epithelial ovarian cancer (EOC) the leading cause of death among all gynecologic malignancies. These relapses are often due to tumor dormancy. Here we identify the RNA polymerase II transcriptional mediator subunit 12 (MED12) as an important molecular regulator of tumor dormancy. MED12 knockout (KO) induced dormancy of EOC cells in vitro and in vivo, and microarray analysis showed that MED12 KO decreased expression of EGFR. Restoration of EGFR expression in MED12 KO cells restored proliferation. Additionally, MED12 bound to the promoter of EGFR, and correlation studies showed that MED12 expression positively correlated with EGFR expression in EOC patient samples. Clinical data demonstrated that chemotherapy-resistant patients expressed lower levels of MED12 compared with responsive patients. Overall, our data show that MED12 plays an important role in regulating dormancy of EOC through regulation of EGFR.Significance: MED12 is identified as a novel, important regulator of tumor dormancy in human ovarian cancer. Cancer Res; 78(13); 3532-43. ©2018 AACR.

Simple quantitative PCR analysis for allelic Pten loss in tumor progression.

We describe a simple method to accurately detect and quantify both Pten mutation and allele-specific loss using allele-specific PCR analysis. Our approach used a heterozygous genomic DNA with one wild-type and one mutant Pten allele as a reference at a single concentration to calculate the percent ratio of the wild-type Pten gene for the detection of allele-specific gene loss. With a standard curve, ratios from PCR data were used to quantitate the wild-type Pten allele copy number loss in tumor specimens. We demonstrate the utility of our approach to calculate allele-specific Pten loss during tumor progression and show that our approach generates quantitative data that are comparable to those obtained from digital droplet PCR. As a method to detect both mutation and allele-specific gene loss, our approach is less subject to the variability of sample amount that are often very limited in clinical analysis. Since conventional PCR is easy to be carried out, our method simplifies the workflow in any laboratory and would provide significant advantages for simplicity to quantify allele-specific gene loss.