Values of multiparametric and biparametric MRI in diagnosing clinically significant prostate cancer: a multivariate analysis.

BACKGROUND: To investigate the value of semi-quantitative and quantitative parameters (PI-RADS score, T2WI score, ADC, Ktrans, and Kep) based on multiparametric MRI (mpMRI) or biparametric MRI (bpMRI) combined with prostate specific antigen density (PSAD) in detecting clinically significant prostate cancer (csPCa). METHODS: A total of 561 patients (276 with csPCa; 285 with non-csPCa) with biopsy-confirmed prostate diseases who underwent preoperative mpMRI were included. Prostate volume was measured for calculation of PSAD. Prostate index lesions were scored on a five-point scale on T2WI images (T2WI score) and mpMRI images (PI-RADS score) according to the PI-RADS v2.1 scoring standard. DWI and DCE-MRI images were processed to measure the quantitative parameters of the index lesion, including ADC, Kep, and Ktrans values. The predictors of csPCa were screened by logistics regression analysis. Predictive models of bpMRI and mpMRI were established. ROC curves were used to evaluate the efficacy of parameters and the model in diagnosing csPCa. RESULTS: The independent diagnostic accuracy of PSA density, PI-RADS score, T2WI score, ADCrec, Ktrans, and Kep for csPCa were 80.2%, 89.5%, 88.3%, 84.6%, 58.5% and 61.6%, respectively. The diagnostic accuracy of bpMRI T2WI score and ADC value combined with PSAD was higher than that of PI-RADS score. The combination of mpMRI PI­RADS score, ADC value with PSAD had the highest diagnostic accuracy. CONCLUSIONS: PI-RADS score according to the PI-RADS v2.1 scoring standard was the most accurate independent diagnostic index. The predictive value of bpMRI model for csPCa was slightly lower than that of mpMRI model, but higher than that of PI-RADS score.

The pH and mercury ion stimuli-responsive supramolecular assemblies of cucurbit[7]uril and Hoechst 33342.

In the present work, we have investigated the effect of pH and mercury ions on the host-guest complex formed between cucurbit[7]uril (Q[7]) and Hoechst 33342 (H33342). 1H NMR, UV-vis and fluorescence spectroscopy revealed that acid/base stimulation could change the binding stoichiometry between Q[7] and H33342. The results suggest that two complexation equilibria (1:1 and 2:1) may exist between H33342 and Q[7] at pH 2.0 and 4.5, respectively. However, a 1:1 host-guest complex was formed between H33342 and Q[7] at pH 7.0 and 10.0. Q[7] shows differential affinities for the protonated and neutral forms of H33342 dye. Moreover, the switching between H33342∙2H+@2Q[7](1:2) at pH 4.5 and H33342∙H+@Q[7](1:1) at pH 7.0 was reversible. Furthermore, as a metal stimulus, Hg2+ ions could push (i) Q[7] from the piperazine ring to the benzimidazole position with a 1:1 guest-host ratio and (ii) a second Q[7] onto the ethyl position with a 1:2 guest-host stoichiometry. This stimulus response system will have potential applications in the field of molecular switch design.

CRISPR-Assisted Multiplex Base Editing System in Pseudomonas putida KT2440.

Pseudomonas putida (P. putida) KT2440 is a paradigmatic environmental-bacterium that possesses significant potential in synthetic biology, metabolic engineering and biodegradation applications. However, most genome editing methods of P. putida KT2440 depend on heterologous repair proteins and the provision of donor DNA templates, which is laborious and inefficient. In this report, an efficient cytosine base editing system was established by using cytidine deaminase (APOBEC1), enhanced specificity Cas9 nickase (eSpCas9ppD10A) and the uracil DNA glycosylase inhibitor (UGI). This constructed base editor converts C-G into T-A in the absence of DNA strands breaks and donor DNA templates. By introducing a premature stop codon in target spacers, we successfully applied this system for gene inactivation with an efficiency of 25-100% in various Pseudomonas species, including P. putida KT2440, P. aeruginosa PAO1, P. fluorescens Pf-5 and P. entomophila L48. We engineered an eSpCas9ppD10A-NG variant with a NG protospacer adjacent motif to expand base editing candidate sites. By modifying the APOBEC1 domain, we successfully narrowed the editable window to increase gene inactivation efficiency in cytidine-rich spacers. Additionally, multiplex base editing in double and triple loci was achieved with mutation efficiencies of 90-100% and 25-35%, respectively. Taken together, the establishment of a fast, convenient and universal base editing system will accelerate the pace of future research undertaken with P. putida KT2440 and other Pseudomonas species.

Inflammation promotes oral squamous carcinoma immune evasion via induced programmed death ligand-1 surface expression.

The association between inflammation and cancer provides a new target for tumor biotherapy. The inflammatory cells and molecules within the tumor microenvironment have decisive dual roles in antitumor immunity and immune evasion. In the present study, phytohemagglutinin (PHA) was used to stimulate peripheral blood mononuclear cells (PBMCs) to simulate the tumor inflammatory microenvironment. The effect of immune cells and inflammatory cytokines on the surface expression of programmed cell death-1 ligand 1 (PD-L1) and tumor immune evasion was investigated using flow cytometry (FCM) and an in vivo xenotransplantation model. Based on the data, PHA-activated, but not resting, immune cells were able to promote the surface expression of PD-L1 in Tca8113 oral squamous carcinoma cells via the secretion of inflammatory cytokines, but not by cell-cell contact. The majority of the inflammatory cytokines had no significant effect on the proliferation, cell cycle progression and apoptosis of the Tca8113 cells, although they each induced the expression of PD-L1 in a dose-dependent manner. In total, 99% of the Tca8113 cells expressed PD-L1 following treatment with the supernatant of PHA-stimulated PBMCs. The PHA-supernatant pretreated Tca8113 cells unusually induced Tca8113 antigen-specific CD8+ T cell apoptosis in vitro and the evasion of antigen-specific T cell attraction in a nude mouse tumor-bearing model. These results indicate a new mechanism for the promotion of tumor immune evasion by the tumor inflammatory microenvironment.

[Inflammatory factors promote the expression of PD-L1 in Tca8113].

OBJECTIVE: To investigate the mechanism of immunologic escape in the tumor microenvironment, study the expression of programmed death 1 ligand-1 (PD-L1) in Tca8113 with treatment of inflammatory factors. METHODS: The expression of PD-L1 treated with inflammatory factors (IL-1beta, IL-2, IL-6, TNF-alpha, IFN-gamma) was detected by flow cytometry (FCM). RESULTS: The expression of PD-L1 in Tca8113 was up-regulated conspicuously with treatment of inflammatory factors (P < 0.05), including: IL-1beta, IL-6, TNF-alpha, IFN-y. And the factors played the role in synergistic effects, most significantly in the groups of IL-1beta + IFN-gamma, TNF-alpha + IFN-gamma and IL-1beta + IL-6 + IFN-gamma + TNF-alpha. But the influence of IL-2 on the expression of PD-L1 was not significant (P > 0.05). CONCLUSION: With the up-regulated expression of PD-L1, the tumor would be escaped more easily from the immunoreactions, while there were an abundance of inflammatory factors in the tumor microenvironment.

[Study on prohibition of high mobility group chromosomal protein N2 against human oral squamous cell carcinoma in vitro].

OBJECTIVE: Take human oral squamous cell carcinoma Tca8113 as experimental model, and study the anti oral squamous cell carcinoma activity of high mobility group chromosomal protein N2 (HMGN2) molecule. METHODS: Train a large number of recombinant human HMGN2 expression vector Escherichia coli BL21. HMGN2 was expressed under isopropyl-1-thio-beta-galactopyranoside (IPTG) induction and purified by B-PER GST Fusion Protein Purification Kit. A variety of concentrations HMGN2 were added to cell culture medium, cells were tested by MTT, Hoechst 33342 fluorescence staining, flow cytometry assay and Western-blot. RESULTS: MTT results proved that HMGN2 could significantly inhibit human oral squamous cell carcinoma Tca8113 growth. Hoechst 33342 fluorescence staining, flow cytometry assay test and Western-blot proved HMGN2 could make Tca8113 cells morphological change, make Tca8113 cells block in S period of cell cycle and strongly promote Tca8113 cells to apoptosis. CONCLUSION: HMGN2 can promote apoptosis of oral squamous cell carcinoma cells.

Interferon-γ-induced PD-L1 surface expression on human oral squamous carcinoma via PKD2 signal pathway.

Many cells located in the tumor microenvironment function to protect or promote the ability of tumor cells to escape immune destruction. Previous studies have shown that programmed death ligand-1 (PD-L1), a ligand of the B7 superfamily, is expressed on a series of human tumors and can inhibit anti-tumor immune responses. Interferon-γ (IFN-γ), a cytokine produced and secreted by inflammatory cells in the tumor microenvironment, is a main stimulator of PD-L1 expression in tumor cells. Making clear the mechanism of IFN-γ induced the expression of PD-L1 on tumor cells that is benefit to find a way to inhibit the function of PD-L1 and improve cancer cell-reactive immune responses. Herein, we have identified protein kinase D isoform 2 (PKD2) as an important regulator of PD-L1 expression on human oral squamous carcinoma cells induced by IFN-γ. IFN-γ induced the expression of PD-L1 and PKD2 in human oral squamous carcinoma Tca8113 in both time and dose dependent manner. The expression of PD-L1 was decreased significantly after PKD2 knockdown with shRNA/siRNA interference or PKD chemical inhibitor following induction with IFN-γ. The apoptosis of CD8(+) T cell which is induced by tumor cells via PD-1/PD-L1 pathway was significantly decreased, as a result, the anti-tumor effects of tumor antigen specific T cell were increased in vivo. Together, these data combined with our previous results, indicate PKD2 as an important target candidate for tumor biotherapy. Inhibition of PKD2 activation not only inhibits PD-L1 expression and promotes an anti-tumor effect, but also decreases drug resistance in chemotherapy.

PKD2 mediates multi-drug resistance in breast cancer cells through modulation of P-glycoprotein expression.

Multi-drug resistance (MDR) represents a major obstacle for chemotherapeutic treatment of a wide variety of human cancers. Increased expression of drug efflux pumps, such as the P-glycoprotein (P-gp) have been linked to development of MDR. Herein, we have identified protein kinase D isoform 2 (PKD2) as an important regulator of MDR and P-gp expression in paclitaxel-treated breast cancer cell lines. PKD2 was expressed with the highest phosphorylated activation status in the MDA-MB-231 cell line. MDA-MB-231 cells were also found to exhibit the highest level of resistance to an array of chemotherapeutic drugs. To further characterize the relationship between PKD2 activation and MDR, we next focused on the effects of the chemotherapeutic agent paclitaxel in MDA-MB-231 cells. Treatment with paclitaxel was shown to induce both PKD2 phosphorylation and P-gp expression in a time-dependent manner. Importantly, shRNA-mediated knockdown of PKD2 in MDA-MB-231 cells resulted in a significant decrease in resistance to paclitaxel, evident as significant decreases in both the IC(50) value and the resistance index (RI). Concurrent with the decrease in drug resistance, paclitaxel-induced expression of P-gp was also significantly reduced in PKD knockdown cells. These results indicate that PKD2 is required for paclitaxel-induced MDR and expression of P-gp. Therefore, modulation of PKD2 activity represents an attractive therapeutic strategy for improvement of the clinical effectiveness of chemotherapy.