Knockdown of STIP1 inhibits the invasion of CD133­positive cancer stem­like cells of the osteosarcoma MG63 cell line via the PI3K/Akt and ERK1/2 pathways.

Osteosarcoma is the most common primary malignant tumor of the bone in adolescents and children, with high rates of metastasis and a poor prognosis. Recently, osteosarcoma cancer stem/stem­like cells (CSCs) have been identified as the main cause of recurrence and metastasis. Stress­induced phosphoprotein 1 (STIP1), a co­chaperone that binds to heat shock proteins 70 and 90, is abnormally expressed in several tumor cell lines, and may play an important role in tumor cell migration and invasion. These features indicate that STIP1 may represent a new therapeutic target for osteosarcoma CSCs. However, the role of STIP1 in osteosarcoma CSC migration and invasion remains largely unknown. In the present study, CD133­positive osteosarcoma CSCs were first isolated and cultured by magnetic cell sorting and serum­free medium suspension cell sphere culture, respectively. Knockdown of STIP1 by small interfering RNA significantly was then shown to inhibit the migration and invasion of these cells, possibly due to the regulation of the expression of matrix metalloproteinase (MMP)­2, MMP­9 and tissue inhibitor of metalloproteinase­2. Furthermore, data from the present study suggested that the knockdown of STIP1 decreased the levels of phosphorylated Akt and phosphorylated ERK1/2. In summary, these findings indicate that targeting STIP1 in osteosarcoma may constitute a viable molecular targeted therapy strategy for the inhibition of CSC invasion and migration.

Adjustment Disorder in Female Breast Cancer Patients: Prevalence and Its Accessory Symptoms.

Women diagnosed with breast cancer may have serious psychological problems and will suffer from adjustment disorder (AjD). We investigated the prevalence of AjD in female breast cancer patients who were diagnosed within 1 year and examined the severe life events they experienced, and the most common symptoms of AjD. 342 newly diagnosed (<1 year) female breast cancer patients were recruited from Tongji Hospital and Hubei Cancer Hospital in Hubei, China, from July 2018 to May 2019. The patients completed the self-report questionnaire including demographic characteristics and the scale ADNM-20 for the diagnosis of AjD. SPSS20.0 was used for data analysis. As a result, the prevalence of AjD in breast cancer patients was 38.6%. Patients from rural areas and lacking of exercise were more likely to suffer from AjD (P<0.05). The prevalence of AjD in patients who did not regard breast cancer as the most severe life event was higher than that in patients who took breast cancer as the most severe life event (44.4% vs. 33.9%, P<0.05; OR=1.728, 95% CI=1.072-2.787). The symptom that scored highest was preoccupation (3.15). We found that the prevalence of AjD in women with breast cancer in this study is very high and warrants more attention. Patients from rural areas, lacking of exercise and subject to multiple stressors are more likely to suffer from AjD. The commonest and severe symptom is preoccupation.

Expression of CXCR4 and MMP-2 is associated with poor prognosis in patients with osteosarcoma.

BACKGROUND: Osteosarcoma is a primary malignant tumor with a high tendency to form metastasis and poor prognosis. Consequently, finding effective early indicators of metastases is crucial for identifying and treating high-risk patients. CXCR4 and MMP-2 have been found to strongly correlate with invasion and metastasis of malignant tumors, including osteosarcoma. MATERIALS AND METHODS: Our study evaluated CXCR4 in conjunction with MMP-2 as an important clinicopathological prognostic predictor for metastasis and overall survival of osteosarcoma. 73 patients' clinical data and pathological samples were retrieved for the study. A median time of 36 months follow-up was performed to evaluate for tumor metastasis and patient survival. CXCR4 and MMP-2 proteins in tumor tissues were detected by immunohistochemistry on paraffin-embedded tissue sections. RESULTS: The positive expression rate of CXCR4 and MMP-2 was 68.5% and 54.8% respectively, and of the 45 patients who developed distal metastasis, 33 and 28 patients had positive expression of CXCR4 and MMP-2 respectively. The median metastasis-free survival was 72.00 months in the CXCR4-negative group and 14.00 months in the CXCR4 positive group. Furthermore, median overall survival was 73.77 and 24.00 months in these same two groups. Further, the median metastasis-free survival was 66.51 months in the MMP-2 negative group and 9.00 months in the MMP-2 positive group. The median overall survival was 75.07 and 19.00 months in these same two groups. MMP2 and metastasis remained the significant and independent prognostic factors for metastasis-free survival and overall survival by using the COX regression model adjusted for the multivariate predictors of survival. CONCLUSION: Our results suggest that metastasis and MMP-2 are both independent prognostic indicators for metastasis-free and overall survival of osteosarcoma patients.

Gel-based chemical cross-linking analysis of 20S proteasome subunit-subunit interactions in breast cancer.

The ubiquitin-proteasome system plays a pivotal role in breast tumorigenesis by controlling transcription factors, thus promoting cell cycle growth, and degradation of tumor suppressor proteins. However, breast cancer patients have failed to benefit from proteasome inhibitor treatment partially due to proteasome heterogeneity, which is poorly understood in malignant breast neoplasm. Chemical crosslinking is an increasingly important tool for mapping protein three-dimensional structures and proteinprotein interactions. In the present study, two cross-linkers, bis (sulfosuccinimidyl) suberate (BS(3)) and its water-insoluble analog disuccinimidyl suberate (DSS), were used to map the subunit-subunit interactions in 20S proteasome core particle (CP) from MDA-MB-231 cells. Different types of gel electrophoresis technologies were used. In combination with chemical cross-linking and mass spectrometry, we applied these gel electrophoresis technologies to the study of the noncovalent interactions among 20S proteasome subunits. Firstly, the CP subunit isoforms were profiled. Subsequently, using native/SDSPAGE, it was observed that 0.5 mmol/L BS(3) was a relatively optimal cross-linking concentration for CP subunit-subunit interaction study. 2-DE analysis of the cross-linked CP revealed that α1 might preinteract with α2, and α3 might pre-interact with α4. Moreover, there were different subtypes of α1α2 and α3α4 due to proteasome heterogeneity. There was no significant difference in cross-linking pattern for CP subunits between BS(3) and DSS. Taken together, the gel-based characterization in combination with chemical cross-linking could serve as a tool for the study of subunit interactions within a multi-subunit protein complex. The heterogeneity of 20S proteasome subunit observed in breast cancer cells may provide some key information for proteasome inhibition strategy.

MiR-21 mediates the radiation resistance of glioblastoma cells by regulating PDCD4 and hMSH2.

The purpose of this study was to investigate the molecular mechanism by which miR-21 and its target genes mediate radiation resistance of glioblastoma cells. Real-time PCR was employed to detect miR-21 expression in normal brain tissues, glioblastoma tissues and glioblastoma cell lines (A172, T98G and U87MG). T98G cells were transfected with anti-miR-21 oligonucleotides, or plasmids containing PDCD4 or hMSH2 (PDCD4-pcDNA3 and hMSH2-pcDNA3). The survival curve was obtained to investigate the sensitivity of T98G cells to radiation. Cell apoptosis was measured by using the Caspase-3/7 kit and cell cycle by flow cytometry. Western blotting was performed to detect the expression of hMSH2 and PDCD4 in miR-21-inhibiting T98G cells. The results showed that miR-21 expression in glioblastoma cells and tissues was conversely associated with the radiation sensitivity. Over-expression of miR-21 resulted in radiation resistance, while knockdown of miR-21 led to higher sensitivity of glioblastma cells to radiation. After miR-21 knockdown, the apoptosis of T98G cells was significantly increased and the G(2) phase arrest was more significant. In addition, miR-21 knockdown increased the expression of endogenous PDCD4 and hMSH2, which contributed to the apoptosis and G(2) arrest of T98G cells. The findings suggested that miR-21 may mediate the resistance of glioblastoma cells against radiation via its target genes PDCD4 and hMSH2. MiR-21 and its target genes may be used as potential molecular targets for clinical radiotherapy sensitization in the future.

The correlation between IGF-II and Bcl-2 expression in colorectal adenocarcinoma.

Our aim for this study was to investigate the correlation and clinical significance between the expression of IGF-II and Bcl-2 in colorectal adenocarcinoma, especially in terms of the metastasis of colorectal adenocarcinoma. Sixty paraffin embedded samples of colorectal adenocarcinoma were selected, and fifteen normal colorectal tissues were used as controls. IGF-II mRNA was detected using in situ hybridization, and the expression of Bcl-2 along with the proliferating cell nuclear antigen (PCNA) protein was detected through immunohistochemistry. The TUNEL assay was used to detect apoptosis. Specimens with a positive cell ratio less than 30% were defined as negative. The levels of IGF-II mRNA and the Bcl-2 protein were significantly higher in colorectal adenocarcinoma (39.64 ± 7.38% and 30.74 ± 7.22%, respectively) than in normal colorectal tissues (22.56 ± 4.21% and 12.17 ± 1.94%, respectively) (P < 0.01). The levels were related to Dukes' stage and lymph node metastases, but were unrelated to patient age, gender, tumor site, tumor size, and tumor differentiation. Also, a negative correlation was observed between IGF-II mRNA and Bcl-2 protein (P < 0.05) during Dukes' stages. In addition, a positive correlation between IGF-II mRNA and PCNA or apoptosis, as well as a negative correlation between Bcl-2 and apoptosis were observed (P < 0.01). There was no correlation between Bcl-2 and PCNA (P > 0.05). The patients detected as IGF-II mRNA (+) and Bcl-2 (-) showed the worst prognosis. The expression of IGF-II and Bcl-2 was correlated with the clinical manifestation of colorectal adenocarcinoma; thus, the assessment of both IGF-II and Bcl-2's status will provide important information regarding the diagnosis and prognosis of colorectal adenocarcinoma.

[Potentials of DNA-PKcs, Ku80, and ATM in enhancing radiosensitivity of cervical carcinoma cells].

BACKGROUND & OBJECTIVE: DNA double strand break (DSB) is the lethal damage of cells after irradiation. DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and ataxia-telangiectasia mutated (ATM) are the most important repair proteins of DSB. Cervical carcinoma is mainly treated by radiotherapy; however, the tumor cells display different radiosensitivity. This study tried to explore the correlations of DNA-PKcs, Ku80, and ATM expression to radiosensitivity of cervical cancer cells, and to probe their potentials in enhancing the radiosensitivity of cervical carcinoma. METHODS: The expression of DNA-PKcs, Ku80, and ATM in 41 specimens of cervical carcinoma was detected by immunohistochemistry. Their expression in 8 tumor cell lines (including 4 cervical carcinoma cell lines) was measured by Western blot; the survival fraction at 2 Gy (SF2) and alpha value were measured by colony formation test; the correlations of protein expression to SF2 and alpha values were analyzed by Pearson linear correlation analysis. The small hairpin RNA (shRNA) targeting DNA-PKcs and a competitive DNA-PKcs inhibitor LY294002 were used to inhibit DNA-PKcs expression and activity in cervical carcinoma cell line HeLa. The SF2 and alpha values of HeLa cells after X-ray irradiation were measured by colony formation test; cell apoptosis was analyzed by flow cytometry. RESULTS: The positive rates of Ku80, DNA-PKcs, and ATM in the 41 specimens of cervical carcinoma were 70.73%, 68.29%, and 19.51%, respectively. The expression of DNA-PKcs was positively related to the SF2 values of the 8 tumor cell lines (r = 0.72, P = 0.04); the expression of Ku80 and ATM had no correlation to SF2 and alpha values. The SF2 value was lower in DNA-PKcs shRNA-transfected HeLa cells than in control HeLa cells (0.37 vs. 0.53, P < 0.05). When treated with 50 mumol/L LY294002 for 1 h, the apoptosis rate of HeLa cells had no significant change ( P > 0.05); when irradiated by 6 Gy X-ray for 48 h and 72 h, the apoptosis rate was significantly higher in LY294002-pretreated HeLa cells than in control HeLa cells (t = 3.25, P = 0.03; t = 3.01, P = 0.04). CONCLUSIONS: DNA-PKcs protein is highly expressed in cervical carcinoma, and its expression level could prognosticate the radiosensitivity of tumor cells. Inhibiting DNA-PKcs expression or activity may sensitize HeLa cells to X-ray.

[Expression of thymidine phosphorylase in cancer].

OBJECTIVE: To study the thymidine phosphorylase (TP) expression in different types of cancer and its correlation with tumor microvessel density (MVD). METHODS: The expression of TP and MVD was detected by immunohistochemistry method. In a series of 251 cancer patients there were 48 patients with gastric cancer, 53 with colorectal cancer, 47 with breast cancer, 56 with cervical cancer, 47 with lung cancer. Normal gastric (n = 25), colorectal (n = 25), cervical (n = 17) and lung (n = 25) tissues around the cancer were also examined. RESULTS: The TP expression rate was 64.6% in gastric cancer, 67.9% in colorectal cancer, 80.9% in breast cancer, 82.1% in cervical cancer, and 63.8% in lung cancer, which was significantly higher than that in normal tissues (P = 0.0000). TP expression was positively correlated with MVD in gastric, colorectal, breast, and cervical cancers. The correlation was not statistically significant in lung cancer. CONCLUSION: This study indicates that TP overexpression in cancer may be associated with tumor angiogenesis.

[First-line Xeloda (Capecitabine) treatment for advanced and recurrent colorectal cancer].

OBJECTIVE: To evaluate the efficacy and safety of capecitabine as first-line therapy in patients with advanced and recurrent colorectal cancer. METHODS: From December 2000 to November 2001, sixty patients with advanced and recurrent colorectal cancer received first-line capecitabine treatment given at a dose of 1250 mg/m(2) twice daily, on days 1 - 14 every 21 days. At least 2 cycles were administered. RESULTS: The overall response rate was 23.3% with 14 PR, 24 SD (40.0%) and 15 PD. The median survival time was 14.7 months. The survival rate was 63.9% at 12-months and 33.4% at 24-months. Grade III-IV adverse effects were diarrhea in 4 patients (6.6%), anemia in 2 (3.3%) and hand-foot syndrome (HFS) in 1 (1.7%); Grade I-II adverse effects were hyperpigmentation in 20 (33.3%), HFS in 18 (30.0%) and diarrhea in 10 (16.7%). CONCLUSION: Capecitabine is an efficacious and better-tolerated alternative treatment for the patients with advanced and recurrent colorectal cancer.

[Result of phase II clinical trial of herceptin in advanced Chinese breast cancer patients].

OBJECTIVE: To observe the clinical efficacy and adverse effects of herceptin for advanced Chinese breast cancer patients. METHODS: Thirty-one pathologically proved advanced breast cancer women were treated by herceptin. In the first week, a loading dose 4 mg/kg was administered by intravenous infusion and from the second week, a routine dose of 2 mg/kg was given every week for at least 3 months. RESULTS: There were 2 CR, 6 PR, 7 SD, and 16 PD among 31 patients after treatment by herceptin, the response rate being 25.8%. In factors influencing the prognosis, age and general condition were factors favoring the results, and pathological type, site of metastasis, grade of her-2 over expression and prior treatment were irrelevant to the results. The adverse effects were mild but different from those of the common anticancer drugs. CONCLUSION: Herceptin is effective and well tolerated by the Chinese breast cancer patients.