Characteristics of HPV integration in cervical adenocarcinoma and squamous carcinoma.

PURPOSE: HPV integration usually occurs in HPV-related cancer, and is the main cause of cancer. But the carcinogenic mechanism of HPV integration is unclear. The study aims to provide a theoretical basis for understanding the pathogenesis of cervical adenocarcinoma (AC) and cervical squamous carcinoma (SCC). METHODS: We used HPV capture sequencing to obtain HPV integration sites in AC and SCC, and analyzed cytobands, distribution of genetic and genomic elements, identified integration hotspot genes, clinicopathological parameters, breakpoints of HPV16 and performed pathway analysis. Then we conducted immunohistochemical (IHC) assay to preliminarily verify the expression of most frequently integrated genes in AC, STARD3 and ERBB2. RESULTS: The results revealed that the most frequently observed integrated cytoband was 17q12 in AC and 21p11.2 in SCC, respectively. The breakpoints in both AC and SCC were more tended to occur within gene regions, compared to intergenetic regions. Compared to SCC samples, AC samples had a higher prevalence of genomic elements. In AC, HPV integration has no significantly difference with clinicopathological parameters, but in SCC integration correlated with differentiation (P < 0.05). Breakpoints of HPV in SCC located in LCR more frequently compared to AC, which destroyed the activation of promoter p97. Hotspot genes of HPV integration were STARD3 and ERBB2 in AC, and RNA45S rDNA and MIR3648-1 in SCC, respectively. Meanwhile, we preliminarily proved that the expression of STARD3 and ERBB2, the most frequently integrated genes, would increase after integration. CONCLUSION: These results suggested that HPV may utilize the powerful hosts' promoters to express viral oncogenes and overexpression of viral oncogenes plays a significant role in the carcinogenesis of SCC. In AC, HPV integration may affect hosts' oncogenes, and the dysregulation of oncogenes may primarily contribute to progression of AC.

Detection of DNA Methylation in Gene Loci ASTN1, DLX1, ITGA4, RXFP3, SOX17, and ZNF671 for Diagnosis of Cervical Cancer.

Objective: To evaluate the diagnostic value of DNA methylation detection of multiple gene loci in cervical cancer. Methods: A total of 61 cases requiring cervical biopsy were selected from the outpatient clinic of Maternal and Child Health Hospital of Hubei Province between January 2018 and December 2019. The patients were divided into four groups based on histopathologic diagnosis: cervical cancer (CC) group, high-grade squamous intraepithelial lesion (HSIL) group, low-grade squamous intraepithelial lesion (LSIL) group, and control group. HPV examination, liquid-based cytology examination, and DNA methylation detection at multiple gene sites were performed. The positive rate of DNA methylation, sensitivity, specificity, area under the curve (AUC), and other efficacy indexes were calculated to evaluate the diagnostic value of DNA methylation detection at multiple gene loci in cervical cancer. Results: The positive rates of DNA methylation in CC, HSIL, LSIL, and control groups were 100%, 88%, 83% and 17%, respectively. The ZNF671 gene had the highest positive rate among the cervical lesion group, with rates of 57%, 76%, and 100% in LSIL, HSIL, and CC groups respectively. The combination of DNA methylation detection at multiple gene loci showed the highest diagnostic efficacy for HSIL and cervical cancer, with AUC value of 0.850 (95% CI:0.746-0.954), a Youden index of 0.654, and a sensitivity and specificity of 85% and 85.4%, respectively. The diagnostic efficacy of the combined detection was significantly higher than that of HPV examination and liquid-based cytology examination (P < 0.05). Conclusion: DNA methylation detection at multiple gene loci is highly effective and diagnostic tool for cervical cancer, and has potential application value in clinical practice.

Absence of high-grade cervical intraepithelial neoplasia in conization specimens from patients with colposcopic biopsy-confirmed high-grade cervical intraepithelial neoplasia: Retrospective study of 1695 cases.

Few studies have investigated the absence of high-grade cervical intraepithelial neoplasia (CIN) in excised specimens, and sample sizes of these studies were limited. This study retrospectively analyzed clinical characteristics of 1695 patients with CIN 2/3 to determine the incidence rate and relative factors of CIN 1 or less in conization specimens from patients with colposcopic biopsy-confirmed CIN 2/3. The study group comprised 430 cases of CIN 1 or less in conization specimens, and the control group comprised 1142 cases with high-grade CIN lesions in conization specimens. Univariate and multivariate logistic regression models were established to evaluate relative factors. The 1-9 years follow-up data were analyzed to determine the persistence/recurrence rate. Multivariate logistic regression showed that patients aged 18-24 years (OR (95% CI) = 2.224 (1.014, 4.877)); with a negative hrHPV test result (OR (95% CI) = 3.210 (1.627, 6.331)); a cytology test result of normal (OR (95% CI) = 5.184 (3.138, 8.563)), ASC-US (OR (95% CI) = 3.420 (2.102, 5.564)), LSIL (OR (95% CI) = 2.588 (1.475, 4.541)), or ASC-H (OR (95% CI) = 2.434 (1.306, 4.539)); an indication of CIN 2 on biopsy (OR (95% CI) = 2.290 (1.694, 3.096)), and no glandular involvement (OR (95% CI) = 1.616 (1.205, 2.169)) were more likely to have an absence of high-grade dysplasia in conization specimens. There was no difference in the persistence/recurrence rate between the two groups (x2 = 1.55, P = 0.46). An age of 18-24 years, a negative hrHPV test result, a non-HSIL cytology test result, an indication of CIN 2 on biopsy, and no glandular involvement were relative factors for an absence of high-grade dysplasia in conization specimens. For patients with relative factors, especially young women, informed follow-up should be considered.

The Application of DNA Ploidy Analysis in Large-Scale Population Screening for Cervical Cancer.

OBJECTIVE: The objective of this study was to evaluate the application of DNA ploidy analysis in large-scale population screening for cervical cancer. METHODS: From March 2016 to March 2019, eligible subjects were enrolled and recommended to undergo DNA ploidy analysis, the ThinPrep cytology test (TCT), and high-risk human papillomavirus (hrHPV) detection concurrently. Patients with positive results were recommended for colposcopy, and biopsy diagnosis was regarded as the "gold standard." We compared the test efficiencies of the 3 methods and compared the efficiency and accuracy of the TCT in our hospital and the "2-cancer screening" project in Hubei Province during the same period. RESULTS: Among 20,574 women, the positive rates of DNA ploidy analysis, cytology, and hrHPV testing were 4.01%, 4.71%, and 16.28%, respectively. The sensitivities of these methods for screening for grade 2+ cervical intraepithelial neoplasia were 0.70, 0.68, and 0.96, and their specificities were 0.79, 0.82, and 0.45, respectively. On comparing DNA ploidy analysis with the TCT, there was no significant difference in the sensitivity, specificity, positive predictive value, negative predictive value, and missed diagnosis rate. In opportunistic screening and the 2-cancer screening project, the positive rates of cytology were 4.71% and 2.87%, respectively. And the efficiency and accuracy of the TCT in opportunistic screening were higher than in the 2-cancer screening project. CONCLUSION: Therefore, DNA ploidy analysis, which is of low-cost and does not depend on cytopathologists, can replace cytology and be applied in large-scale population screening for cervical cancer.

Stromal Neutrophil Extracellular Trap Density Is an Independent Prognostic Factor for Cervical Cancer Recurrence.

BACKGROUND: Emerging evidence indicates that the tumor microenvironment influences tumor progression and patient prognosis through various inflammatory cells. Polymorphonuclear neutrophils (PMNs) and their functional structures termed neutrophil extracellular traps (NETs) are prominent constituents of several malignant tumors and affect the tumor microenvironment and cancer evolution. Here, we investigate the prognostic value of PMNs and NETs for recurrence in patients with cervical cancer. METHODS: The study comprised 126 cervical cancer patients who were retrospectively enrolled. CD66b+ neutrophils and myeloperoxidase/citrullinated histone H3 (MPO/H3Cit)-labeled NETs were assessed by immunofluorescence, and the relationships with clinical and histopathologic features and patient outcome were evaluated. RESULTS: The highest density of CD66b+ neutrophils were observed in the stromal compartment (median 55 cells/mm2). Above median densities of stromal CD66b+ neutrophils and NETs were significantly associated with short recurrence-free survival (RFS) (P = 0.041 and P = 0.006, respectively). Multivariate analysis identified high clinical stage (hazard ratio [HR] 6.40; 95% confidence interval [CI] 3.51-11.64; P < 0.001), lymph node metastases (HR 4.69; 95% CI 3.09-9.66; P = 0.006) and high density of NETs (HR 2.66; 95% CI 1.21-5.82; P = 0.015) as independent prognostic factors for short RFS, whereas a high density of CD66b+ neutrophils was not significant. Patients with a high NET density showed worse recurrence status in every stage, but the difference was only significant for stage I (P = 0.042), not stages II, III, or IV (all P > 0.05). Combining stromal NET density and the tumor, nodes, metastasis (TNM) staging system had better prognostic accuracy for cervical cancer than the TNM staging system alone at five and six years respectively (P = 0.010 and P = 0.023). CONCLUSION: Stromal NET density is an independent prognostic factor for RFS in cervical cancer. Combining NETs with the TNM staging system may further improve prognostic stratification.

Asymptomatic and Symptomatic Patients With Non-severe Coronavirus Disease (COVID-19) Have Similar Clinical Features and Virological Courses: A Retrospective Single Center Study.

The current outbreak of coronavirus disease 2019 (COVID-19) has been defined as a pandemic by the World Health Organization. We aimed to evaluate the clinical features and virological course of non-severe COVID-19 patients with or without symptoms who were admitted to a Chinese cabin hospital. In this retrospective single center study, we reviewed 252 laboratory-confirmed COVID-19 patients treated at one temporary cabin hospital in Wuhan, China. Demographic, clinical, serial chest computed tomography (CT), and serial viral test data were compared between asymptomatic and symptomatic patients. The association between clinical features and symptomatic status or patient referral status was analyzed. Among all 252 patients, 74 (29.4%) were asymptomatic and 138 (54.76%) had more than two family members who developed COVID-19. The probability for family clustering was similar between asymptomatic and symptomatic patients (59.70 vs. 61.64%, P = 0.79). Asymptomatic patients and symptomatic patients were equally likely to reach a virus-free state during their stay at the cabin hospital (93.15 vs. 86.44%, P = 0.13). The initial chest CT screening showed that 81 (32.1%) patients had no visible pneumonia, 52 (20.6%) had unilateral pneumonia, and 119 (47.2%) had bilateral pneumonia. Symptomatic patients had a higher chance to have bilateral pneumonia (P < 0.0001) and were less likely to show improvement on the follow-up CT scan (P = 0.0002). In total, 69 (27.4%) patients were referred to the designated hospital and only 23 (9.1%) patients were referred due to the progression of pneumonia. Non-severe COVID-19 patients can transmit the disease regardless of their symptomatic status. It is highly recommended that asymptomatic patients be identified and quarantined to eliminate the transmission of COVID-19.

Expression of HIF-1α is a predictive marker of the efficacy of neoadjuvant chemotherapy for locally advanced cervical cancer.

Platinum-based, arterial infusion chemotherapy as a neoadjuvant chemotherapy (NACT) followed by hysterectomy may be efficient for the treatment of locally advanced cervical cancer and improve prognosis. It is important to predict whether the NACT would be effective before it is launched. Hypoxia inducible factor-1α (HIF-1α) is the master transcriptional regulator of the cellular response to altered oxygen concentration. HIF-1α protein expression is elevated in numerous human malignancies, contributes to poor disease outcome, and has been reported to induce tumorigenesis and chemoresistance. In the present study, patients with International Federation of Gynecology and Obstetrics stage IIB-IIIB cervical cancer (n=59) between 2008 and 2014 were assessed for HIF-1α expression by immunohistochemistry. Tumor samples were obtained by biopsy before any treatment. A double-path chemotherapy regimen, paclitaxel (intravenous) plus cisplatin (intra-arterial injection into the uterine region), was used as NACT. The patients were then separated into two groups according to NACT response: One group comprised patients with NACT, for whom the response to treatment was efficient resulting in complete/partial remission of the tumor (CR + PR group; n=52), the other group contained patients with NACT, for whom the result of the treatment was a stable/progressive disease (SD + PD group; n=7). HIF-1α expression was tested in paraffin-embedded sections using immunohistochemistry. HIF-1α expression was significantly higher in the SD + PD group compared with the CR + PR group (P=0.029). The overall survival time was significantly longer in the CR + PR group compared with the SD + PD group (P<0.001). When the patients were divided into two groups based on HIF-1α expression levels. Low (weighted score ≤4, n=39) and high (weighted score ≥6, n=20) expression level groups; the low HIF-1α expression group was significantly more susceptible to NACT treatment (P=0.025). Cox hazard analysis revealed that a high level of HIF-1α expression and lymph node metastases were significant independent predictors of poor overall survival (P=0.025, HR=6.354; P=0.020, HR=6.909, respectively). These results indicated that the expression of HIF-1α may be able to predict the efficiency of NACT and may be considered an independent prognostic factor for stage IIB-IIIB cervical cancer.

Prospective observational study of the efficacy of mixed methylene blue compound injection for treatment of vulvar non-neoplastic epithelial disorders.

OBJECTIVE: To investigate the safety and efficacy of mixed methylene blue (MB) compound injection for treatment of vulvar non-neoplastic epithelial disorders (NNEDs). METHODS: A prospective observational study among 118 women with vulvar NNEDs treated with intradermal injection of mixed MB compound at a hospital in Wuhan, China, between 2013 and 2016. Itching score, skin hypopigmentation area percentage (SHAP), and recurrence were assessed by interview and physical examination before and after treatment. Adverse effects were recorded. RESULTS: Before treatment, mean ± SD itching score was 7.78 ± 1.59. It decreased rapidly immediately after treatment and remained low thereafter (1.82 ± 2.31, 1.69 ± 2.39, 1.97 ± 2.73, 2.05 ± 2.72, and 2.19 ± 2.86 at 1, 3, 6, 12, and 24 months, respectively). Before treatment, mean ± SD SHAP was 28.01% ± 18.28%. SHAP decreased gradually and remained stably low 6 months later (26.28% ± 17.95%, 21.19% ± 18.42%, 19.19% ± 18.67%, 18.68% ± 18.91%, and 18.65% ± 19.20% at 1, 3, 6, 12, and 24 months, respectively). The recurrence rate in 2 years was 21.2% (25/118) with no major complications. CONCLUSIONS: Intradermal injection of mixed MB compound was found to be an effective and safe treatment for vulvar NNEDs. ClinicalTrials.gov: NCT03200808.

Semaphorin 4C Promotes Macrophage Recruitment and Angiogenesis in Breast Cancer.

Semaphorins are a large family of evolutionarily conserved morphogenetic molecules that are associated with repelling axonal guidance. Intriguingly, recent researches indicate that semaphorins are involved in cancer progression. Semaphorin 4C (SEMA4C) has long been considered a neuronal migration gene, but we detected that it is also highly expressed in many malignant human cancers. During an investigation of subcutaneous tumor models, we found that SEMA4C expression promoted tumor growth and progression. We discovered that SEMA4C was involved in maintaining tumor cell self-renewal, likely by regulating the p53 pathway. Inhibiting the expression of endogenous SEMA4C in tumor cells impaired growth and induced senescence and cell-cycle arrest in the G2-phase. In addition, we found that SEMA4C induced the production of angiogenin and colony-stimulating factor-1 (CSF-1) in tumor cells by activating the NF-κB pathway in a plexinB2-dependent manner. In conclusion, SEMA4C expression in breast cancer cells promotes cancer cell proliferation, macrophage recruitment, and angiogenesis. Thus, inhibition of SEMA4C activity may be a novel therapeutic strategy for human breast cancer. IMPLICATIONS: In breast cancer, therapeutic targeting of the SEMA4C pathway may prevent tumor growth, angiogenesis, metastasis, and progression.

Prevalence and Determinants of High-risk HPV Infection among 11549 Women from an Opportunistic Screening in Hubei Province.

High-risk human papillomavirus (hrHPV) infection plays an important role in the development of cervical intraepithelial neoplasia and cervical cancer. A total of 11 549 women were enrolled from the Maternal and Child Health Hospital of Hubei Province. Each participant accepted hrHPV testing and completed a self-administered questionnaire about basic information and potential risk factors. The univariable and multivariable logistic regression model was used to explore the associations between variants and hrHPV infection. Our results showed that hrHPV prevalence was 16.09% in Hubei Province, among which, hrHPV was more likely to be positive in women aged 51 years or above (OR=1.65, 95% CI: 1.28-2.14), and in women who had symptoms of bleeding after intercourse (OR=1.32, 95% CI:1.17-1.50), had first sexual intercourse at the age of 18 years or below (OR=1.33, 95% CI:1.07-1.64), had at least three male sexual partners (OR=2.50, 95% CI:2.07-3.03), and who had been diagnosed with sexually transmitted infections (OR=1.50, 95% CI:1.12-2.03). Married women (OR=0.66, 95% CI: 0.55-0.78) and women who frequently used condoms (OR=0.75, 95% CI:0.67-0.84) had a relatively lower hrHPV prevalence. This study confirms that hrHPV infection was associated with age, marital status, symptoms of intercourse bleeding, history of sexually transmitted infections, and sex-related behaviors. Above all, this study provides a baseline database prior to obtaining vaccinations for dynamic tracking of the changes in hrHPV prevalence.

Isoform expression patterns of EPHA10 protein mediate breast cancer progression by regulating the E-Cadherin and ß-catenin complex.

Overexpression of EPHA10 protein was reported in concomitance with clinical severity of breast cancer. In this study, we annotate overexpression of EPHA10 protein with changes of isoform expression as EphA10s (EPHA10 isoform 2) and EphA10 (EPHA10 isoform 3). In the process of malignant transformation, secretory protein EphA10s is in low expression, and pseudo-kinase EphA10 is overexpressed and cytoplasmically enriched. Down-regulated EphA10s blunts stabilization of membrane-associate ß-catenin via the interaction with ephrin A5. Cytoplasmic EphA10 maintains phosphorylation of E-cadherin. Restoring isoform expression pattern by up-regulated EphA10s and down-regulated cytoplasmic EphA10 inhibits cell invasion and lymph node metastasis by strengthening the stability of the complex of E-cadherin and ß-catenin in membrane. Taken together, we defined the novel interaction via expression patterns of EphA10s and EphA10 that promote malignant transformation of breast cancer, and demonstrated the potential benefit in clinical usage.

Tumor-associated Lymphatic Endothelial Cells Promote Lymphatic Metastasis By Highly Expressing and Secreting SEMA4C.

PURPOSE: Lymphatic vessels are mainly regarded as passive conduits for the dissemination of cancer cells. In this study, we investigate whether and how the tumor-associated lymphatic vessels may play an active role in tumor metastasis. EXPERIMENTAL DESIGN: In situ laser capture microdissection of lymphatic vessels followed by cDNA microarray analysis was used to determine the expression profiling of lymphatic endothelial cells (LEC). Gene expression levels and activity of signaling pathways were measured by real-time RT-PCR, ELISA, or immunoblotting. Lymphangiogenesis was assessed by IHC. Lymph node metastasis was measured using fluorescence imaging. The effects of SEMA4C on lymphangiogenesis in vitro were evaluated using migration assay and tube-formation assay of LECs. RESULTS: Tumor-associated LECs are molecularly and functionally different from their normal counterparts. In addition to expressing high levels of membrane-bound SEMA4C, tumor-associated LECs also produced soluble SEMA4C (sSEMA4C). Increased serum sSEMA4C was detected in patients with breast cancer and cervical cancer. Patients with metastasis had much higher levels of serum sSEMA4C. sSEMA4C promoted lymphangiogenesis by activating PlexinB2-ERBB2 signaling in LECs, and promoted the proliferation and migration of tumor cells by activating PlexinB2-MET signaling, thus promoting lymphatic metastasis. Although the SEMA4C signaling pathways differ between LECs and tumor cells, RHOA activation was necessary for the effects of SEMA4C in both types of cells. CONCLUSIONS: Tumor-associated LECs produce sSEMA4C to promote lymphatic metastasis of tumors. Our results suggest that SEMA4C and RHOA might be potential therapeutic targets, and that higher serum sSEMA4C could be a marker for breast cancer and cervical cancer. Clin Cancer Res; 23(1); 214-24. ©2016 AACR.

Smad4 is required for the development of cardiac and skeletal muscle in zebrafish.

Transforming growth factor-beta (TGF-beta) regulates cellular functions and plays key roles in development and carcinogenesis. Smad4 is the central intracellular mediator of TGF-beta signaling and plays crucial roles in tissue regeneration, cell differentiation, embryonic development, regulation of the immune system and tumor progression. To clarify the role of smad4 in development, we examined both the pattern of smad4 expression in zebrafish embryos and the effect of smad4 suppression on embryonic development using smad4-specific antisense morpholino-oligonucleotides. We show that smad4 is expressed in zebrafish embryos at all developmental stages examined and that embryonic knockdown of smad4 results in pericardial edema, decreased heartbeat and defects in the trunk structure. Additionally, these phenotypes were associated with abnormal expression of the two heart-chamber markers, cmlc2 and vmhc, as well as abnormal expression of three makers of myogenic terminal differentiation, mylz2, smyhc1 and mck. Furthermore, a notable increase in apoptosis was apparent in the smad4 knockdown embryos, while no obvious reduction in cell proliferation was observed. Collectively, these data suggest that smad4 plays an important role in heart and skeletal muscle development.

Inhibition of STAT3 activity re-activates anti-tumor immunity but fails to restore the immunogenicity of tumor cells in a B-cell lymphoma model.

A large number of patients with advanced lymphoma become refractory or relapse after initial treatment due to the persistence of minimal residual disease. Ideal immunotherapy strategy for eradicating the minimal residual disease of lymphoma and preventing the tendency to relapse need to be developed. Here, we use a mice model mimicked the disease entities of aggressive B-cell lymphoma dynamically to analyze the host anti-lymphoma immunity during the progression of lymphoma. We have shown that STAT3 activity was gradually enhanced in host immune effector cells with the progression of lymphoma. Inhibition of the STAT3 activity with a small molecule inhibitor was able to effectively enhance the function of both host innate and adaptive immunity, and thereby delayed the progression of lymphoma. Despite the therapeutic benefits were achieved by using of the STAT3 inhibitor, disrupting of STAT3 pathway did not prevent the eventual development of lymphoma due to the presence of point mutation of ß2M, which controls immune recognition by T cells. Our findings highlight the complexity of the mechanism of immune evasion; therefore a detailed analysis of genes involved in the immune recognition process should be essential before an elegant immunotherapy strategy could be conducted.

Sema4d is required for the development of the hindbrain boundary and skeletal muscle in zebrafish.

Semaphorin4d (SEMA4D), also known as CD100, an oligodendrocyte secreted R-Ras GTPase-activating protein (GAP), affecting axonal growth is involved in a range of processes including cell adhesion, motility, angiogenesis, immune responses and tumour progression. However, its actual physiological mechanisms and its role in development remain unclear. This study has focused on the role of sema4d in the development and expression patterns in zebrafish embryos and the effect of its suppression on development using sema4d-specific antisense morpholino-oligonucleotides. In this study the knockdown of sema4d, expressed at all developmental stages, lead to defects in the hindbrain and trunk structure of zebrafish embryos. In addition, these phenotypes appeared to be associated with the abnormal expression of three hindbrain rhombomere boundary markers, wnt1, epha4a and foxb1.2, and two myogenic regulatory factors, myod and myog. Further, a notable increase of cell apoptosis appeared in the sema4d knockdown embryos, while no obvious reduction in cell proliferation was observed. Collectively, these data suggest that sema4d plays an important role in the development of the hindbrain and skeletal muscle.

[Expression and clinical significance of Sema4C in esophageal cancer, gastric cancer and rectal cancer].

OBJECTIVE: To explore the expression and clinical significance of signal protein Sema4C in esophageal cancer, gastric cancer and rectal cancer. METHODS: Fifty esophageal cancer, 75 gastric cancer, 50 rectal cancer and 20 corresponding normal mucous membrane specimens, collected during the period of January 2008 to December 2010, were detected with streptavidin-peroxidase immunohistochemistry to detect the expression levels of Sema4C. And the relationships of the Sema4C expression with clinicopathological data was analyzed. RESULTS: The expression levels of Sema4C in three kinds of cancers were significantly higher than the corresponding normal mucous membranes (80.0% (n = 40) vs 20.0% (n = 4), 77.3% (n = 58) vs 25.0% (n = 5), 80.0% (n = 40) vs 15.0% (n = 3), all P = 0.000). Furthermore, the percentage of Sema4C positive cells was significantly higher in carcinoma nests of tumors with lymphatic metastasis than those without (90.3% (n = 28) vs 63.2% (n = 12), 85.0% (n = 51) vs 46.7% (n = 7), 92.0% (n = 23) vs 68.0% (n = 17), P = 0.049, 0.005, 0.034). However, no significant correlations were found between the Sema4C expression with gender, age, location of tumors, types of cancer cells, cell differentiation, tumor size, depth of invasion or tumor stage (all P > 0.05). CONCLUSION: There is a high expression of Sema4C in esophageal cancer, gastric cancer and rectal cancer. And it is strongly correlated with lymphatic metastasis. Thus Sema4C may play critical roles in the invasion and lymphatic metastasis of esophageal cancer, gastric cancer and rectal cancer.

Immune safety of a novel oncolytic mutant M1 after administration in vivo.

The aim of this study was to evaluate the safety and efficiency of a novel, oncolytic adenovirus mutant M1 administered in conjunction with immunosuppressive agents. Animal models were established by administering purified M1 either intravenously or retroperitoneally. At different time points, blood samples were taken from the mice for testing of liver and renal function. Microscopic examination of the liver was performed to observe pathological changes. Immunohistochemical analyses were used to evaluate the expression of the adenovirus in the liver. Lymphocyte recruitment to the liver and the activation of adenovirus specific T cells were also analyzed. No signs of general toxicity were observed, but transient increases in ALT and Scr were observed following the administration of M1. Microscopic examination revealed a mild inflammatory response in the liver. Compared to intravenous injection, higher expression levels of adenoviral proteins were observed after retroperitoneal injection. Combined treatment with cyclosporine A resolved the liver and kidney dysfunction and increased the concentration of the adenovirus in the liver. The use of the novel oncolytic adenovirus mutant M1 in vivo is safe, and the combined administration of M1 with immunosuppressive agents was able to enhance the effectiveness and safety profile of M1.

Silencing Wnt2B by siRNA interference inhibits metastasis and enhances chemotherapy sensitivity in ovarian cancer.

OBJECTIVE: Wnt2B overexpression is thought to be involved in tumor progression through the activation of the canonical Wingless and INT-1 signaling pathway. However, the mechanism of Wnt2B signaling in oncogenesis is unknown. In this study, we investigated whether silencing Wnt2B expression could inhibit the invasiveness of ovarian cancer cells and reduce drug resistance. METHODS/MATERIALS: Four ovarian carcinoma cell lines, SKOV3, OV2008, A2780, and C13K, were used. Protein levels were studied by Western blotting. The colony formation ability and invasive ability were determined through colony formation assay and the Matrigel transwell assay, respectively. Cell viability was determined by the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay, whereas apoptosis was assessed using flow cytometry analysis. RESULTS: Among the 4 ovarian carcinoma cell lines, the A2780 cells and C13K cells expressed Wnt2B, and these 2 cell lines were used for analyzing the mechanism of Wnt2B. The down-regulation of Wnt2B inhibited cell colony formation and invasiveness. Enhanced paclitaxel or cisplatin sensitivity was observed in A2780 cells or C13K cells treated with Wnt2B siRNA, respectively. In the presence of Wnt2B siRNA treatment, the caspase-9/B-cell lymphoma 2 (BCL2)/B-cell lymphoma-xL (BCL-xL) pathway and the epithelial-mesenchymal transition/phosphorylated protein kinase B pathway were inhibited. CONCLUSION: These data suggest that Wnt2B indeed plays an important role in ovarian cancer metastasis and drug resistance. This study may provide a new therapeutic target for and a better understanding of ovarian cancer therapy.

Mesenchymal stem cells as carriers and amplifiers in CRAd delivery to tumors.

BACKGROUND: Mesenchymal stem cells (MSCs) have been considered to be the attractive vehicles for delivering therapeutic agents toward various tumor diseases. This study was to explore the distribution pattern, kinetic delivery of adenovirus, and therapeutic efficacy of the MSC loading of E1A mutant conditionally replicative adenovirus Adv-Stat3(-) which selectively replicated and expressed high levels of anti-sense Stat3 complementary DNA in breast cancer and melanoma cells. METHODS: We assessed the release ability of conditionally replicative adenovirus (CRAd) from MSC using crystal violet staining, TCID(50) assay, and quantitative PCR. In vitro killing competence of MSCs carrying Adv-Stat3(-) toward breast cancer and melanoma was performed using co-culture system of transwell plates. We examined tumor tropism of MSC by Prussian blue staining and immunofluorescence. In vivo killing competence of MSCs carrying Adv-Stat3(-) toward breast tumor was analyzed by comparison of tumor volumes and survival periods. RESULTS: Adv-Stat3(-) amplified in MSCs and were released 4 days after infection. MSCs carrying Adv-Stat3(-) caused viral amplification, depletion of Stat3 and its downstream proteins, and led to significant apoptosis in breast cancer and melanoma cell lines. In vivo experiments confirmed the preferential localization of MSCs in the tumor periphery 24 hours after tail vein injection, and this localization was mainly detected in the tumor parenchyma after 72 hours. Intravenous injection of MSCs carrying Adv-Stat3(-) suppressed the Stat3 pathway, down-regulated Ki67 expression, and recruited CD11b-positive cells in the local tumor, inhibiting tumor growth and increasing the survival of tumor-bearing mice. CONCLUSIONS: These results indicate that MSCs migrate to the tumor site in a time-dependent manner and could be an effective platform for the targeted delivery of CRAd and the amplification of tumor killing effects.