Chang-Yao Hsieh

Interleukin-6 promotes cervical tumor growth by VEGF-dependent angiogenesis via a STAT3 pathway.

Interleukin-6 (IL-6) has received particular attention in the pathogenesis of cervical cancer, although the underlying mechanism remains elusive. This study revealed that IL-6 promotes in vivo tumor growth of human cervical cancer C33A cells, but does not substantially alter their in vitro growth kinetics. The in vivo angiogenic assays showed that IL-6 increases angiogenic activity in human cervical cancer cells, an effect that is specifically associated with upregulation of vascular endothelial growth factor (VEGF). Also, using anti-VEGF antibody to block VEGF function significantly inhibited IL-6-mediated angiogenesis and tumor growth in nude mice, strongly supporting the critical role of VEGF in the IL-6-mediated cervical tumorigenesis. Accordingly, the signaling pathway downstream of IL-6/IL-6R responsible for the regulation of VEGF was investigated. Notably, pharmacological inhibition of PI3-K or MAPK failed to inhibit IL-6-mediated transcriptional upregulation of VEGF. Meanwhile, blocking STAT3 pathway with dominant-negative mutant STAT3D effectively abolished IL-6-induced VEGF mRNA. In transient transfections, a luciferase reporter construct containing the full-length 1.5-kb VEGF promoter or a 1.2-kb fragment lacking the known hypoxic-response element also exhibited the same degree of response to IL-6. Additionally, transient transfection of STAT3D downregulated the 1.2-kb VEGF promoter luciferase reporter stimulated by IL-6. Based on the above phenomenon combined with the concomitant increased tumor expression of IL-6 and VEGF in cervical cancer tissues, we conclude that IL-6 may promote cervical tumorigenesis by activating VEGF-mediated angiogenesis via a STAT3 pathway.

Genetic polymorphisms of p53 and GSTP1,but not NAT2,are associated with susceptibility to squamous-cell carcinoma of the esophagus.

The interaction of genetic and environmental factors can determine an individuals susceptibility to various cancers. We present a hospital‐based case‐control study, which included 90 patients of esophageal squamous‐cell carcinoma (ESCC) and 254 healthy people in Taiwan, to investigate the effects of genetic polymorphisms of p53, GSTP1 and NAT2 on the risk of ESCC. Polymorphisms of p53, NAT2 and GSTP1 were determined by PCR‐RFLP. The codon 72 p53 Pro allele was more frequently found in ESCC patients [odds ratio (OR) 1.86, 95% confidence interval (CI) 1.04–3.35 for Arg/Pro genotype and OR 2.56, 95% CI 1.29–5.08 for Pro/Pro genotype]. In cigarette smokers, the frequency of GSTP1 Ile/Ile genotype was higher in ESCC patients (OR 2.8, 95% CI 1.4–5.7). Among alcohol drinkers, borderline significance was also found for GSTP1 Ile/Ile genotype (OR 2.0, 95% CI 0.9–4.4). Results were not similar for the NAT2 genetic polymorphism. Using logistic analyses, we found that individuals with p53 Pro/Pro genotype had a significantly higher risk of developing ESCC than those with Arg/Arg genotype (OR 2.3, 95% CI 1.1–5.1), after adjusting for other significant environmental risk factors. This result remained similar (OR 2.2, 95% CI 1.0–4.8 for p53 Pro/Pro vs. Arg/Arg), even after further adjustment for NAT2 and GSTP1 polymorphisms. The codon 72 p53 Pro/Pro genotype in the general population and GSTP1 Ile/Ile in cigarette smokers may predict a higher risk of developing ESCC. Int. J. Cancer 89:458–464, 2000.

Vascular endothelial growth factor and prognosis of cervical carcinoma

Objective To evaluate vascular endothelial growth factor (VEGF) as a marker for predicting lymph node metastasis and an independent prognostic factor of early-stage cervical carcinoma. Methods One hundred thirty-five women with stage IB–IIA cervical carcinoma had radical abdominal hysterectomies and pelvic lymph node dissections. Intratumoral cytosol VEGF concentrations were assayed with enzyme immunoassay. Histopathologic items and cytosol VEGF-influencing clinical outcomes were compared. Results Twenty-two women (16.3%) who had disease recurrence had higher levels of cytosol VEGF (1020 versus 112 pg/mg protein, P < .001) than those without recurrence. Using a cutoff value of 400 pg/mg protein resulted in best sensitivity of 75%, best specificity of 70%, positive predictive value of 41%, and negative predictive value of 92%. Only overexpressed cytosol VEGF (hazard ratio 6.44, P < .001) was an independent prognostic factor of disease-free survival. The overexpressed cytosol VEGF (hazard ratio 4.50, P = .021) and positive lymphovascular emboli (hazard ratio 4.11, P = .045) were independent prognostic factor of overall survival. Conclusion Cytosol VEGF might be a biomarker for the status of pelvic lymph nodes in early-stage cervical carcinoma and an independent prognostic indicator of its outcome.

Serous ovarian carcinoma patients with high alpha-folate receptor had reducing survival and cytotoxic chemo-response

The alpha‐folate receptor (α‐FR) is highly‐expressed in various non‐mucinous tumors of epithelial origin, including ovarian carcinoma. The aim of this study was to investigate the relationship between alpha‐folate receptor (α‐FR) and the clinico‐pathologic features and outcomes of serous ovarian carcinoma patients and the possible mechanism of α‐FR to chemo‐resistance. Therefore, semi‐quantitative reverse‐transcription polymerase chain reactions for α‐FR expression were performed in the 91 specimens of serous ovarian carcinomas. The expression of α‐FR in each ovarian cancer tissue specimen was defined as the ratio of density of α‐FR to density of glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH). In vitro apoptotic experiments were tested in the original OVCAR‐3 tumor cells and various OVCAR‐3 α‐FR‐transfectants. Patients with an increased α‐FR expression level had poorer responses to chemotherapy (per α‐FR expression level increase: odds ratio (OR): 8.97 (95% confidence interval (CI): 1.40–57.36), p = 0.021). An increased α‐FR expression level was an independently poor prognostic factor for disease free interval (DFI) (per α‐FR expression level increase: hazard ratio (HR): 2.45 (95% CI: 1.16–5.18), p = 0.02) and had a negative impact on overall survival (OS) of these serous ovarian cancer patients (per α‐FR expression level increase: HR: 3.6 (95% CI: 0.93–13.29), p = 0.03) by multivariate analyses. α‐FR inhibited cytotoxic drug‐induced apoptosis in our in vitro apoptotic assays. α‐FR could induce chemo‐resistance via regulating the expression of apoptosis‐related molecules, Bcl‐2 and Bax. Therefore, α‐FR can be a potential biomarker for the prediction of chemotherapeutic responses and clinical prognosis. It also could be the target of ovarian cancer treatment.

Metronomic Chemotherapy Enhances Antitumor Effects of Cancer Vaccine by Depleting Regulatory T Lymphocytes and Inhibiting Tumor Angiogenesis

Although cancer vaccines are emerging as innovative methods for cancer treatment, these alone have limited potential for treating measurable tumor burden. Thus, the importance of identifying anticancer strategies with greater potency is necessary. The chimeric DNA vaccine CTGF/E7 (connective tissue growth factor linked to the tumor antigen human papillomavirus 16 E7) generates potent E7-specific immunity and antitumor effects. We tested immune-modulating doses of chemotherapy in combination with the CTGF/E7 DNA vaccine to treat existing tumors in mice. Metronomic low doses of paclitaxel, not the maximal tolerable dose, are synergistic with the antigen-specific DNA vaccine. Paclitaxel, given in metronomic sequence with the CTGF/E7 DNA vaccine enhanced the vaccines potential to delay tumor growth and decreased metastatic tumors in vivo better than the CTGF/E7 DNA vaccine alone. The two possible mechanisms of metronomic paclitaxel chemotherapy are the depletion of regulatory T cells and the inhibition of tumor angiogenesis rather than direct cancer cell cytolytic effects. Results indicate that combination treatment of metronomic chemotherapy and antigen-specific DNA vaccine can induce more potent antigen-specific immune responses and antitumor effects. This provides an immunologic basis for further testing in cancer patients.

Arsenic trioxide prevents radiation-enhanced tumor invasiveness and inhibits matrix metalloproteinase-9 through downregulation of nuclear factor kappaB.

Arsenic trioxide (ATO) has been implicated as a promising anticancer agent by inhibiting cell growth and inducing apoptosis in certain types of cancer cells. This study explored the antimetastasis property of arsenic, drew potential link between arsenic use and radiotherapy, and uncovered the specific mechanisms underlying these remarkable responses. Using gelatin invasion assay and intravasation assay, we report the novel finding that low-dose ATO (1 μM) reduced the intrinsic migration ability of HeLa cells and significantly inhibited radiation-promoted tumor invasive potential of CaSki cells without inducing apoptotic cell death. Using the murine Lewis lung carcinoma model, the control animals and ATO treatment animals (1 mg/kg i.p., twice weekly) displayed similar in vivo growth kinetics, whereas the radiation (30 Gy in one fraction) and concurrent treatment groups showed considerable growth inhibition. Importantly, although concurrent treatment did not enhance the effectiveness of radiation therapy to the primary tumor, further examination of the lungs revealed that all animals succumbed to radiation-accelerated lung metastases could be effectively treated by combination of ATO and radiation. Radiation-induced matrix metalloproteinase-9 (MMP-9) expression was significantly inhibited by ATO using sequential analysis of the expression of MMPs in xenografts. Supporting this observation, ATO directly downregulates radiation-induced MMP-9 mRNA expression by inhibiting nuclear factor κB activity in human cervical cancer cells. In sum, concurrent arsenic–radiation therapy not only achieves local tumor control but also inhibits distant metastasis. Experimental results of this study highlight a novel strategy in cancer treatment.

Morphine induces apoptosis of human endothelial cells through nitric oxide and reactive oxygen species pathways

Morphine has been widely used for pain management. Other than analgesia, it has effects on vascular endothelial cells, including angiogenesis and apoptosis. An in vitro model of human umbilical vein endothelial cells (HUVECs) was made to investigate the effects and comprehensive mechanisms of morphine on vascular endothelial cells. Morphine enhanced apoptosis of HUVECs, increased intracellular reactive oxygen species (ROS), and reduced mitochondrial membrane potentials (MMPs). It also induced the release of NO and activated NF-kappaB in HUVECs. Naloxone, the opioid receptor antagonist, could reverse cell apoptosis and ROS generation, NO production, and MMP loss. Expression levels of Bak and Bax, and the activation of caspases 3 and 7 in HUVECs significantly increased when treated with morphine. Inhibition of NO production by NO synthase inhibitor reduced morphine-induced apoptosis. Morphine could induce apoptosis of HUVECs through both the NO and ROS pathways. Thus, inhibiting NO or ROS may be a potential target in blocking morphine-induced apoptosis of endothelial cells.

Fusion Protein Vaccine by Domains of Bacterial Exotoxin Linked with a Tumor Antigen Generates Potent Immunologic Responses and Antitumor Effects

Antigen-specific immunotherapy represents an attractive approach for cancer treatment because of the capacity to eradicate systemic tumors at multiple sites in the body while retaining the requisite specificity to discriminate between neoplastic and nonneoplastic cells. It has been shown that certain domains of bacterial exotoxins facilitate translocation from extracellular and vesicular compartments into the cytoplasm. This feature provides an opportunity to enhance class I and/or II presentation of exogenous antigen to T lymphocytes. We investigated previously whether the translocation domain (domain II) of Pseudomonas aeruginosa exotoxin A with a model tumor antigen, human papillomavirus type 16 E7, in the context of a DNA vaccine could enhance vaccine potency. We then attempted to determine whether this chimeric molecule could also generate strong antigen-specific immunologic responses and enhance the potency of cancer vaccine in the protein format. Our results show that vaccination with the PE(DeltaIII)-E7-KDEL3 fusion protein enhances MHC class I and II presentation of E7, leading to dramatic increases in the number of E7-specific CD8+ and CD4+ T-cell precursors and markedly raised titers of E7-specific antibodies. Furthermore, the PE(DeltaIII)-E7-KDEL3 protein generates potent antitumor effects against s.c. E7-expressing tumors and preestablished E7-expressing metastatic lung tumors. Further, mice immunized with PE(DeltaIII)-E7-KDEL3 protein vaccine also retained long-term immunologic responses and antitumor effects. Our results indicate that retrograde-fusion protein via the delivery domains of exotoxins with an antigen greatly enhances in vivo antigen-specific immunologic responses and represents a novel strategy to improve cancer vaccine potency.

Vascular endothelial growth factor in cervical carcinoma

OBJECTIVE To quantitate vascular endothelial growth factor of cervical carcinoma and elucidate its clinical correlation. METHODS Intratumoral protein levels of vascular endothelial growth factor were measured in 104 cervical cancer patients and in 30 cervical tissue specimens of benign gynecologic diseases as controls. The concentrations were correlated with clinical and pathologic characteristics. RESULTS The median concentrations of vascular endothelial growth factor in cervical cancer tissues were higher than those in benign cervical tissues (180.0 versus 0.0 pg/mg of protein, P < .001). Tumors larger than 4 cm (1030.0 versus 118.0 pg/mg of protein, P < .001) and with deep stromal invasion (364.0 versus 111.0 pg/mg of protein, P = .016) had higher levels than those smaller than 4 cm or with superficial stromal invasion. Higher levels were also found in tumors with lymphovascular emboli (568.0 versus 118.0 pg/mg of protein, P = .006), parametrial invasion (582.0 versus 117.0 pg/mg of protein, P = .04), and pelvic lymph node metastasis (759.5 versus 121.0 pg/mg of protein, P = .002) than in those without. The protein levels of vascular endothelial growth factor correlated positively with tumor sizes (r = 0.340, P < .001). Tumors with overexpressed VEGF were larger (3.35 +/- 1.17 versus 2.13 +/- 1.28 cm, P < .001) and had higher incidence of deep stromal invasion (20 of 57 versus 6 of 47, P = .009), lymphovascular emboli (15 of 33 versus 11 of 71, P = .011), parametrial invasion (15 of 32 versus 11 of 72, P = .002), and lymph node metastasis (10 of 20 versus 16 of 84, P = .004). CONCLUSION Intratumoral protein level of vascular endothelial growth factor in cervical cancer tissue correlates well with local tumor progression and tumor metastasis. Vascular endothelial growth factor might be a marker for evaluating disease severity.

Clinical application of intratumoral blood flow study in patients with endometrial carcinoma

The objective of this study was to evaluate the correlation between intratumoral blood flow as assessed by color Doppler ultrasound with stage, tumor grade, depth of invasion, and lymph node metastasis in endometrial carcinoma and determine its clinical usefulness.