A Simple Technique for Direct Immobilization of Target Enzymes from Cell Lysates Based on the SpyTag/SpyCatcher Spontaneous Reaction.

Many of the current methods for enzyme purification and immobilization suffer from several drawbacks, such as requiring tedious multistep procedures or long preparation, and being environmentally unfriendly, due to the chemicals and conditions involved. Thus, a simple technique for direct purification and immobilization of target enzymes from cell lysates was proposed. The elastin-like polypeptides (ELPs)-SpyCatcher chimera could mediate the formation of silica carriers within seconds and the target enzymes were then covalently immobilized on silica carriers via SpyCatcher/SpyTag spontaneous reaction. These tailor-made carriers were easily prepared, with precisely controlled morphology and size, as well as none-consuming surface modification needed, which could specifically immobilize the SpyTag-fused target enzymes from the cell lysate without pre-purification.

Construction and validation of a prognostic model for stage IIIC endometrial cancer patients after surgery.

BACKGROUND: To explore the most predictive lymph node (LN) scheme for stage IIIC endometrial cancer (EC) patients after hysterectomy and develop a scheme-based nomogram. METHODS: Data from 2626 stage IIIC EC patients, diagnosed between 2010 and 2014, were extracted from the Surveillance, Epidemiology, and End Results (SEER) registry. The predictive ability of four LN schemes was assessed using C-index and Akaike information criterion (AIC). A nomogram based on the most predictive LN scheme was constructed and validated. The comparison of the predictive ability between nomogram and FIGO stage was conducted using the area under the receiver operating characteristic curve (AUC) and decision curve analysis (DCA). RESULTS: FIGO stage (stage IIIC1/stage IIIC2) was not an independent risk factor for OS in stage IIIC EC patients (P = 0.672) and log odds of positive lymph nodes (LODDS) had the best predictive ability (C-index: 0.742; AIC: 8228.95). A nomogram based on LODDS was constructed and validated, which had a decent C-index of 0.742 (0.723-0.762). The nomogram showed a better predictive ability than that of the FIGO staging system. CONCLUSION: FIGO IIIC1/FIGO IIIC2 could not differentiate the prognosis for stage IIIC EC patients. We developed and validated a nomogram based on LODDS to predict OS for post-operative patients with stage IIIC EC.

[Relationship between the radiosensitivity of hepatic carcinoma cells and their survivin expression levels].

OBJECTIVE: To investigate the relationship between the radiosensitivity of hepatic carcinoma cells and their survivin expression levels. METHODS: Hepatic carcinoma cell lines HepG2 and SMMC-7721 were irradiated with various doses of 60Co gamma-rays. The cell survival rate, expression of survivin, cell cycle profile and activity of caspase-3 were respectively detected by clonogenic assay, immunocytochemistry, flow cytometry and chromatometry. RESULTS: The surviving fraction at 2Gy (SF2) of HepG2 and SMMC-7721 were 0.43+/-0.01 vs 0.70+/-0.02, and HepG2 had higher radiosensitivity than SMMC-7721. gamma-rays radiation up-regulated the expression of survivin. SMMC-7721 had a significantly higher expression of survivin than HepG2 (t = 2.81-5.20, P < 0.05). The activity of caspase-3 was more powerful in HepG2 than in SMMC-7721 (t = 6.05-6.72, P < 0.01). CONCLUSION: Survivin may play a critical role in mediating radiation resistance in SMMC-7721 through its up-regulation and caspase-3 dependent manner.

Detecting normal cell radiosensitivity via assay of DNA damage in lymphocytes for individualizing radiotherapy in head and neck cancer patients.

PURPOSE: The purpose of this study was to determine whether the distribution of radiosensitivities in normal tissues of head and neck cancer patients, measured using a DNA damage assay on lymphocytes, is likely to provide sufficient discrimination to enable reliable identification of patients with abnormal sensitivities. MATERIAL AND METHODS: Radiosensitivity was assessed in 307 lymphocyte samples from unselected head and neck cancer patients and was quantified as the initial number of DNA double-strand breaks (dsb) induced per Gray and per DNA unit (200 Mbp). RESULTS: The existence of an inter-individual variation in the radiosensitivity parameter is described by the range (0.41--9.38 dsb/Gy/DNA unit) of the values found. We detected 37 patients who developed severe skin reactions during radiotherapy treatment and we compared their radiosensitivity values with the remaining patients treated. Radiosensitivity values of >7.20 dsb/Gy/DNA unit should theoretically correspond to highly radiosensitive patients. CONCLUSIONS: Our results suggest that initial DNA damage measured on lymphocytes offers an approach to predict the acute response of human normal tissues prior to radiotherapy. .

Correlation between DNA repair capacity in lymphocytes and acute side effects to skin during radiotherapy in nasopharyngeal cancer patients.

PURPOSE: Repair of radiation-induced DNA damage plays a critical role for both the susceptibility of patients to side effects after radiotherapy and their subsequent cancer risk. The study objective was to evaluate whether DNA repair data determined in vitro are correlated with the occurrence of acute side effects during radiotherapy. EXPERIMENTAL DESIGN: Nasopharyngeal cancer patients receiving radiation therapy were recruited in a prospective epidemiologic study. As an indicator for clinical radiosensitivity, adverse reactions of the skin were recorded. Cryopreserved lymphocytes from 100 study participants were gamma-irradiated with 5 Gy in vitro and analyzed using the alkaline comet assay. Reproducibility of the assay was determined by repeated analysis (n = 22) of cells from a healthy donor. A coefficient of variation of 0.24 was calculated. RESULTS: The various parameters determined to characterize the individual DNA repair capacity showed large differences between patients. Twenty-one patients were identified with considerably enhanced DNA damage induction, and 19 patients exhibited severely reduced DNA repair capacity after 15 and 30 minutes. Eight patients were considered as clinically radiosensitive, indicated by moist desquamation of the skin after a total radiation dose of 70 Gy. CONCLUSIONS: Using the alkaline comet assay as described here, nasopharyngeal cancer patients were identified showing abnormal cellular radiation effects, but this repair deficiency corresponded only at a very limited extent to the acute radiation sensitivity of the skin.

[Inhibition effects of c-erbB-2 and c-raf-1 antisense oligodeoxynucleotides combined transfection on the human ovarian carcinoma transplanted subcutaneously in nude mice].

OBJECTIVE: To investigate the inhibition effects of c-erbB-2 and c-raf-1 antisense oligodeoxynucleotides (ASODN) combined transfection on the human ovarian epithelial cancer transplanted subcutaneously in nude mice. METHODS: There were 7 groups: normal control group, c-erbB-2 sense observed group, c-raf-1 sense observed group, c-erbB-2 antisense observed group, c-raf-1 antisense observed group, whole dose combined group, half dose combined group. Human ovarian epithelial cancer cells SKOV3 were treated by different oligodeoxynucleotides, then transplanted subcutaneously in nude mice, respectively. The changes of tumor volume were observed and the tumor growth inhibitory rate was calculated. RESULTS: There was no difference between sense observed group and normal control group. There was a larger growth inhibitory rate in whole -dose combined group and half -dose combined group, the first time that can be detected was 13.7 days and 15.2 days, and the maximum tumor growth inhibitory rates were 61.1% and 71.3%, respectively. CONCLUSIONS: The results suggested that ASODN combined transfection can inhibit the tumorigenesis of ovarian epithelial cancer cells in nude mice, it may be a more useful gene therapy for the ovarian epithelial carcinoma.