Vascular endothelial growth factor (VEGF) expression is a prognostic factor for radiotherapy outcome in advanced carcinoma of the cervix

The aim of the study was to evaluate VEGF expression in tumour biopsies as a prognostic factor for radiotherapy outcome in advanced carcinoma of the cervix. A retrospective study was carried out on 100 patients. Pre-treatment tumour VEGF expression was examined immunohistochemically in formalin-fixed, paraffin-embedded biopsies using a widely available commercial antibody. A semi-quantitative analysis was made using a scoring system of 0, 1, 2, and 3, for increasing intensity of staining. High VEGF expression was associated with a poor prognosis. A univariate log rank analysis found a significant relationship with overall survival (P = 0.0008) and metastasis-free survival (P = 0.0062), but not local control (P = 0.23). There was no correlation between VEGF expression and disease stage, tumour differentiation, patient age, or tumour radiosensitivity (SF2). In a Cox multivariate analysis of survival VEGF expression was the most significant independent prognostic factor (P = 0.001). After allowing for VEGF only SF2 was a significant prognostic factor (P = 0.003). In conclusion, immunohistochemical analysis of VEGF expression is a highly significant and independent prognostic indicator of overall and metastasis-free survival for patients treated with radiotherapy for advanced carcinoma of the cervix. It is also a rapid and easy method that could be used in the clinical setting, to identify patients at high risk of failure with conventional radiotherapy who may benefit from novel approaches or chemoradiotherapy. © 2000 Cancer Research Campaig

The Prognostic Effect of VEGF Expression in Squamous Cell Carcinoma of the Cervix Treated with Radiation Therapy Alone

We investigated the relationship between vascular endothelial growth factor (VEGF) expression and clinical outcome in squamous cell carcinoma of the cervix treated with radiotherapy alone. The immunohistochemical study was performed for forty-two paraffin embedded specimens with anti-VEGF mouse monoclonal antibody. Staining was defined as positive for VEGF when more than 10% of the tumor cells were stained from 500 cells counted. Positive VEGF expression was observed in twenty-one among forty-two patients. VEGF expression according to stage (p=0.101), lymph node status (p=0.621), parametrial invasion (p=0.268), and age (p=0.5) revealed no significant difference. But the VEGF expression was significantly higher in tumors larger than 4 cm (p=0.031). Five year survival rates according to VEGF expression status were 89% for VEGF negative group and 47% for VEGF positive group (p=0.02). FIGO stage (p=0.007), tumor size (p=0.025) and the duration of external beam radiation therapy (p=0.006) were also significant prognostic factors for overall survival. We suggest that VEGF expression may be a prognotic factor of the cervix cancer patients treated with radiation therapy alone

No relationship between thymidine phosphorylase (TP, PD-ECGF) expression and hypoxia in carcinoma of the cervix

The expression of hypoxia-regulated genes promotes an aggressive tumour phenotype and is associated with an adverse cancer treatment outcome. Thymidine phosphorylase (TP) levels increase under hypoxia, but the protein has not been studied in association with hypoxia in human tumours. An investigation was made, therefore, of the relationship of tumour TP with hypoxia, the expression of other hypoxia-associated markers and clinical outcome. This retrospective study was carried out in patients with locally advanced cervical carcinoma who underwent radiotherapy. Protein expression was evaluated with immunohistochemistry. Hypoxia was measured using microelectrodes and the level of pimonidazole binding. There was no relationship of TP expression with tumour pO2 (r=−0.091, P=0.59, n=87) or pimonidazole binding (r=0.13, P=0.45, n=38). There was no relationship between TP and HIF-1α, but there was a weak borderline significant relationship with HIF-2α expression. There were weak but significant correlations of TP with the expression of VEGF, CA IX and Glut-1. In 119 patients, the presence of TP expression predicted for disease-specific (P=0.032) and metastasis-free (P=0.050) survival. The results suggest that TP is not a surrogate marker of hypoxia, but is linked to the expression of hypoxia-associated genes and has weak prognostic power

Carbonic anhydrase IX promotes tumor growth and necrosis in vivo and inhibition enhances anti-VEGF therapy.

PURPOSE: Bevacizumab, an anti-VEGFA antibody, inhibits the developing vasculature of tumors, but resistance is common. Antiangiogenic therapy induces hypoxia and we observed increased expression of hypoxia-regulated genes, including carbonic anhydrase IX (CAIX), in response to bevacizumab treatment in xenografts. CAIX expression correlates with poor prognosis in most tumor types and with worse outcome in bevacizumab-treated patients with metastatic colorectal cancer, malignant astrocytoma, and recurrent malignant glioma. EXPERIMENTAL DESIGN: We knocked down CAIX expression by short hairpin RNA in a colon cancer (HT29) and a glioblastoma (U87) cell line which have high hypoxic induction of CAIX and overexpressed CAIX in HCT116 cells which has low CAIX. We investigated the effect on growth rate in three-dimensional (3D) culture and in vivo, and examined the effect of CAIX knockdown in combination with bevacizumab. RESULTS: CAIX expression was associated with increased growth rate in spheroids and in vivo. Surprisingly, CAIX expression was associated with increased necrosis and apoptosis in vivo and in vitro. We found that acidity inhibits CAIX activity over the pH range found in tumors (pK = 6.84), and this may be the mechanism whereby excess acid self-limits the build-up of extracellular acid. Expression of another hypoxia inducible CA isoform, CAXII, was upregulated in 3D but not two-dimensional culture in response to CAIX knockdown. CAIX knockdown enhanced the effect of bevacizumab treatment, reducing tumor growth rate in vivo. CONCLUSION: This work provides evidence that inhibition of the hypoxic adaptation to antiangiogenic therapy enhances bevacizumab treatment and highlights the value of developing small molecules or antibodies which inhibit CAIX for combination therapy

Hypoxia-regulated carbonic anhydrase IX expression is associated with poor survival in patients with invasive breast cancer.

Tumour hypoxia is a microenvironmental factor related to poor response to radiation, chemotherapy, genetic instability, selection for resistance to apoptosis, and increased risk of invasion and metastasis. Hypoxia-regulated carbonic anhydrase IX (CA IX) has been studied in various tumour sites and its expression has been correlated with the clinical outcome. The purpose of this study was to investigate the correlation of CA IX expression with outcome in patients with invasive breast cancer. We conducted a retrospective study examining the effects of carbonic anhydrase IX (CA IX) on survival in patients with breast cancer. To facilitate the screening of multiple tissue blocks from each patient, tissue microarrays were prepared containing between two and five representative samples of tumour per patient. Immunohistochemistry was used to examine expression of CA IX in patients with breast cancer. The study includes a cohort of 144 unselected patients with early invasive breast cancer who underwent surgery, and had CA IX expression and follow-up data available for analysis. At the time of analysis, there were 28 deaths and median follow-up of 48 months with 96% of patients having at least 2 years of follow-up. CA IX was negative for 107 patients (17 deaths) and positive for 37 patients (11 deaths). Kaplan-Meier survival curves show that survival was superior in the CA IX-negative group with a 2-year survival of 97% for negatives and 83% for positives (log-rank test P=0.01). Allowing for potential prognostic variables in a Cox regression analysis, CA IX remained a significant independent predictor of survival (P=0.035). This study showed in both univariate and multivariate analysis that survival is significantly inferior in patients with tumour expressing CA IX. Prospective studies are underway to investigate this correlation in clinical trial setting

Online accounts of gene expression profiling in early-stage breast cancer: interpreting genomic testing for chemotherapy decision-making

Background Genomic techniques are being developed within oncology and beginning to be experienced within routine cancer care. Little is known about how these tools feature in patients’ experiences of treatment decision making. Objective This research explores the ways in which women interpret and discuss gene expression profiling for breast cancer treatment decision making, as articulated within online accounts. Design This study used a qualitative approach to analyse written exchanges focusing on gene expression profiling in the UK (Oncotype DX test). Accounts are taken from online forums hosted by two UK cancer charity websites, comprising 132 discussion threads from a total of seven forums. Authors qualitatively analysed the data and developed key themes drawing on existing literature from medical sociology. Findings Women used online spaces to share and discuss results of gene expression profiling. Women interpreted results in the context of indirect experience of cancer treatment, and sociocultural depictions of cancer and chemotherapy. Users largely represented the test positively, emphasizing its ability to “personalize” treatment pathways, though many also pointed to inherent uncertainties with regards the possibility of cancer recurrence. Discussion and Conclusions We highlight the complex contexts in which genomic techniques are experienced, with these shaped by personal biographies, online environments and pervasive cultural narratives of cancer and its treatment. We highlight tensions between the claims of genomic testing to aid treatment decision making and patient reflections on the capability of these techniques to resolve uncertainties surrounding treatment decisions

Hypoxia leads to significant changes in alternative splicing and elevated expression of CLK splice factor kinases in PC3 prostate cancer cells

© 2018 The Author(s). Background: Mounting evidence suggests that one of the ways that cells adapt to hypoxia is through alternative splicing. The aim of this study was firstly to examine the effect of hypoxia on the alternative splicing of cancer associated genes using the prostate cancer cell line PC3 as a model. Secondly, the effect of hypoxia on the expression of several regulators of splicing was examined. Methods: PC3 cells were grown in 1% oxygen in a hypoxic chamber for 48 h, RNA extracted and sent for high throughput PCR analysis at the RNomics platform at the University of Sherbrooke, Canada. Genes whose exon inclusion rate PSI (ψ) changed significantly were identified, and their altered exon inclusion rates verified by RT-PCR in three cell lines. The expression of splice factors and splice factor kinases in response to hypoxia was examined by qPCR and western blotting. The splice factor kinase CLK1 was inhibited with the benzothiazole TG003. Results: In PC3 cells the exon inclusion rate PSI (ψ) was seen to change by >25% in 12 cancer-associated genes; MBP, APAF1, PUF60, SYNE2, CDC42BPA, FGFR10P, BTN2A2, UTRN, RAP1GDS1, PTPN13, TTC23 and CASP9 (caspase 9). The expression of the splice factors SRSF1, SRSF2, SRSF3, SAM68, HuR, hnRNPA1, and of the splice factor kinases SRPK1 and CLK1 increased significantly in hypoxia. We also observed that the splice factor kinase CLK3, but not CLK2 and CLK4, was also induced in hypoxic DU145 prostate, HT29 colon and MCF7 breast cancer cell lines. Lastly, we show that the inhibition of CLK1 in PC3 cells with the benzothiazole TG003 increased expression of the anti-apoptotic isoform caspase 9b. Conclusions: Significant changes in alternative splicing of cancer associated genes occur in prostate cancer cells in hypoxic conditions. The expression of several splice factors and splice factor kinases increases during hypoxia, in particular the Cdc-like splice factor kinases CLK1 and CLK3. We suggest that in hypoxia the elevated expression of these regulators of splicing helps cells adapt through alternative splicing of key cancer-associated genes. We suggest that the CLK splice factor kinases could be targeted in cancers in which hypoxia contributes to resistance to therapy

Necrosis related HIF-1α expression predicts prognosis in patients with endometrioid endometrial carcinoma

<p>Abstract</p> <p>Background</p> <p>Hypoxia inducible factor 1α (HIF-1α) plays an essential role in the adaptive response of cells to hypoxia and is associated with aggressive tumour behaviour. We have shown p27^kip1, which is generally reduced in endometrial cancer, to be re-expressed in hypoxic regions. This possibly contributes to survival of cancer cells. The aim of this study was to evaluate the prognostic value of HIF-1α and p27^kipexpression in patients with endometrioid endometrial cancer.</p> <p>Methods</p> <p>Expression levels of HIF-1α, CAIX, Glut-1, and p27^kip1were analyzed by immunohistochemistry. Percentage of positive cells, staining pattern (perinecrotic, diffuse, or mixed) and presence of necrosis were noted.</p> <p>Results</p> <p>Necrosis was correlated with shortened disease free survival (DFS) (p <it>= </it>0.008) and overall survival (OS) (p <it>= </it>0.045). For DFS, perinecrotic HIF-1α expression was also prognostic (p <it>= </it>0.044). Moreover, high p27^kip1expression was an additional prognostic factor for these patients with perinecrotic HIF-1α expression. In multivariate Cox regression, perinecrotic HIF-expression emerged as an independent prognostic factor. Perinecrotic HIF-1α expression was significantly associated with CAIX and Glut-1 expression, pointing towards functional HIF-1.</p> <p>Conclusions</p> <p>In patients with endometrioid endometrial cancer, necrosis and necrosis-related expression of HIF-1α are important prognostic factors. More aggressive adjuvant treatment might be necessary to improve the outcome of patients with these characteristics.</p

Differential Impact of EGFR-Targeted Therapies on Hypoxia Responses: Implications for Treatment Sensitivity in Triple-Negative Metastatic Breast Cancer

In solid tumors, such as breast cancer, cells are exposed to hypoxia. Cancer cells adapt their metabolism by activating hypoxia-inducible factors (HIFs) that promote the transcription of genes involved in processes such as cell survival, drug resistance and metastasis. HIF-1 is also induced in an oxygen-independent manner through the activation of epidermal growth factor receptor tyrosine kinase (EGFR-TK). Triple-negative breast cancer (TNBC) is a subtype of invasive breast cancer characterized by negative expression of hormonal and HER2 receptors, and this subtype generally overexpresses EGFR. Sensitivity to three EGFR inhibitors (cetuximab, gefitinib and lapatinib, an HER2/EGFR-TK inhibitor) was evaluated in a metastatic TNBC cell model (MDA-MB-231), and the impact of these drugs on the activity and stability of HIF was assessed.MDA-MB-231 cells were genetically modified to stably express an enhanced green fluorescent protein (EGFP) induced by hypoxia; the Ca9-GFP cell model reports HIF activity, whereas GFP-P564 reports HIF stability. The reporter signal was monitored by flow cytometry. HIF-1 DNA-binding activity, cell migration and viability were also evaluated in response to EGFR inhibitors. Cell fluorescence signals strongly increased under hypoxic conditions (> 30-fold). Cetuximab and lapatinib did not affect the signal induced by hypoxia, whereas gefitinib sharply reduced its intensity in both cell models and also diminished HIF-1 alpha levels and HIF-1 DNA-binding activity in MDA-MB-231 cells. This gefitinib feature was associated with its ability to inhibit MDA-MB-231 cell migration and to induce cell mortality in a dose-dependent manner. Cetuximab and lapatinib had no effect on cell migration or cell viability.Resistance to cetuximab and lapatinib and sensitivity to gefitinib were associated with their ability to modulate HIF activity and stability. In conclusion, downregulation of HIF-1 through EGFR signaling seems to be required for the induction of a positive response to EGFR-targeted therapies in TNBC