Expression of DSG1 and DSC1 are prognostic markers in anal carcinoma patients

Background:

Our purpose was to investigate if dysregulation of cell adhesion molecules could be linked to prognosis in squamous cell carcinomas (SCCs) of the anal region.

Methods:

Protein expression of desmoglein-1 (DSG1), desmocollin-1 (DSC1) and E-cadherin was studied by immunohistochemistry in a cohort of 53 anal carcinoma patients treated by radiation alone or combined with 5-fluorouracil and mitomycin C.

Results:

Univariate analyses identified, among others, negative membranous DSG1 staining (P=0.009), negative cytoplasmic DSC1 staining (P=0.012) and negative DSG1 (membranous)+negative DSC1 (cytoplasmic) staining (P=0.004) to be associated with improved cancer-specific survival (CSS). On multivariate analyses positive DSG1 (membranous)+DSC1 (cytoplasmic) staining (HR 6.95, P=0.044), large tumour size and lymph node metastases (HR 6.44, P=0.004) and radiation without chemotherapy (HR 6.73 P=0.004) were associated with worse CSS. On univariate analysis, improved disease-free survival was associated with negative membranous staining of DSG1 (P=0.047), and negative DSG1 (membranous)+negative DSC1 (cytoplasmic) staining (P=0.025), among others.

Conclusion:

Membrane negativity for DSG1 and cytoplasmic negativity for DSC1 are favourable markers for CSS in SCCs of the anal region.     

Early expression of the fractalkine receptor CX3CR1 in pancreatic carcinogenesis

Background:

In pancreatic ductal adenocarcinoma (PDAC), fractalkine receptor CX3CR1 contributes to perineural invasion (PNI). We investigated whether CX3CR1 expression occurs early in PDAC and correlates with tumour features other than PNI.

Methods:

We studied CX3CR1 and CX3CL1 expression by immunohistochemistry in 104 human PDAC and coexisting Pancreatic Intraepithelial Neoplasia (PanIN), and in PdxCre/LSL-Kras^G12D mouse model of PDAC. CX3CR1 expression in vitro was studied by a spheroid model, and in vivo by syngenic mouse graft of tumour cells.

Results:

In total, 56 (53.9%) PDAC expressed CX3CR1, 70 (67.3%) CX3CL1, and 45 (43.3%) both. CX3CR1 expression was independently associated with tumour glandular differentiation (P=0.005) and PNI (P=0.01). Pancreatic Intraepithelial Neoplasias were more frequently CX3CR1+ (80.3%, P<0.001) and CX3CL1+ (86.8%, P=0.002) than matched cancers. The survival of PDAC patients was better in those with CX3CR1+ tumour (P=0.05). Mouse PanINs were also CX3CR1⁺ and -CL1⁺. In vitro, cytokines significantly increased CX3CL1 but not CX3CR1 expression. Differently, CX3CR1 was upregulated in tumour spheroids, and in vivo only in well-differentiated tumours.

Conclusion:

Tumour differentiation, rather than inflammatory signalling, modulates CX3CR1 expression in PanINs and PDAC. CX3CR1 expression pattern suggests its early involvement in PDAC progression, outlining a potential target for interfering with the PanIN transition to invasive cancer.     

Expression and prognostic value of transcription factor PROX1 in colorectal cancer

Background:

PROX1 is a specific target of the β-catenin/TCF pathway in the intestinal epithelium. It acts as a regulator of progression from a benign to a highly dysplastic phenotype in colorectal tumours. However, the clinical significance of PROX1 expression is not known.

Methods:

We studied the prognostic value of immunohistochemical expression of PROX1 in a series of 517 patients with colorectal cancer (CRC).

Results:

The majority of the tumour samples expressed PROX1 (91%, 471 out of 517). High PROX1 expression was associated with a poor grade of tumour differentiation (P<0.0001). In the subgroup of patients with colon cancer, high PROX1 expression was associated with unfavourable colorectal cancer-specific survival (CCSS) as compared with low PROX1 expression (CCSS 47% vs 62% P=0.045; RR 1.47). The association between high PROX1 and poor outcome was further strengthened in female colon cancer patients (CCSS 38% vs 63% P=0.007; RR 2.02). Nonetheless, in multivariate survival analysis PROX1 expression was not retained as an independent prognostic factor.

Conclusion:

High PROX1 expression is associated with a poor grade of tumour differentiation, and, in colon cancer patients, also with less favourable patient outcome. Our results strengthen the previous preclinical observations that PROX1 has a role in tumour progression in CRC.     

Haemoptysis as a prognostic factor in lung adenocarcinoma after curative resection

Background:

Haemoptysis is a common symptom of lung cancer. Its prognostic role and mechanisms are still poorly understood.

Methods:

We retrospectively reviewed 666 consecutive patients with primary lung adenocarcinoma who underwent complete resection. The prognostic value of haemoptysis with respect to overall survival (OS), disease-specific survival (DSS) and disease-free survival (DFS) was analysed. To further explore the possible mechanisms of haemoptysis, we evaluated vascular endothelial growth factor (VEGF) expression, tumour necrosis, vascular invasion and extratumoural microvessel density (MVD) in 112 randomly selected patients.

Results:

Haemoptysis predicted poor OS, DSS and DFS in operable lung adenocarcinoma (all P<0.001). In addition, haemoptysis was associated with high white blood cell (WBC) count (P=0.032), high fibrinogen (Fib; P<0.001), high tumour greatest dimension (P<0.001), severe vascular invasion (P=0.002) and central tumour location (P<0.001). We obtained no statistically significant differences of VEGF expression, tumour necrosis and extratumoural MVD in haemoptysis and non-haemoptysis groups.

Conclusion:

Our study demonstrates that haemoptysis predicts poor OS, DSS and DFS in lung adenocarcinoma after curative resection. Vascular invasion rather than angiogenesis or tumour necrosis could be the most important mechanism of haemoptysis in lung adenocarcinoma.     

Does thyroid-stimulating hormone influence the prognosis of patients with endometrial cancer? A multicentre trial

Background:

Thyroid function has been suggested to interfere with tumour biology and prognosis in different cancers. The present study was performed to investigate the impact of pre-therapeutic serum thyroid-stimulating hormone (TSH) levels on the prognosis of patients with endometrial cancer.

Methods:

Pre-therapeutic serum TSH was investigated in 199 patients with endometrial cancer. After stratification in TSH risk groups, univariate and multivariable survival analyses were performed.

Results:

Elevated TSH was independently associated with poor disease-specific survival in univariate/multivariable survival analyses (P=0.01 and P=0.03, respectively).

Conclusion:

Thyroid-stimulating hormone may serve as a novel and independent prognostic parameter for disease-specific survival in patients with endometrial cancer.     

Clinical prognostic value of combined analysis of Aldh1, Survivin,and EpCAM expression in colorectal cancer

Background:

Tumour aggressiveness might be related to the degree of main cancer hallmark acquirement of tumour cells, reflected by expression levels of specific biomarkers. We investigated the expression of Aldh1, Survivin, and EpCAM, together reflecting main cancer hallmarks, in relation to clinical outcome of colorectal cancer (CRC) patients.

Methods:

Immunohistochemistry was performed using a tumour tissue microarray of TNM (Tumour, Node, Metastasis)-stage I–IV CRC tissues. Single-marker expression or their combination was assessed for associations with the clinical outcome of CRC patients (N=309).

Results:

Increased expression of Aldh1 or Survivin, or decreased expression of EpCAM was each associated with poor clinical outcome, and was therefore identified as clinically unfavourable expression. Analyses of the combination of all three markers showed worse clinical outcome, specifically in colon cancer patients, with an increasing number of markers showing unfavourable expression. Hazard ratios ranged up to 8.3 for overall survival (P<0.001), 36.6 for disease-specific survival (P<0.001), and 27.1 for distant recurrence-free survival (P<0.001).

Conclusions:

Our data identified combined expression levels of Aldh1, Survivin, and EpCAM as strong independent prognostic factors, with high hazard ratios, for survival and tumour recurrence in colon cancer patients, and therefore reflect tumour aggressiveness.     

High levels of carbonic anhydrase IX in tumour tissue and plasma are biomarkers of poor prognostic in patients with non-small cell lung cancer

Background:

Carbonic anhydrase IX (CAIX) is an enzyme upregulated by hypoxia during tumour development and progression. This study was conducted to assess if the expression of CAIX in tumour tissue and/or plasma can be a prognostic factor in patients with non-small cell lung cancer (NSCLC).

Methods:

Tissue microarrays containing 555 NSCLC tissue samples were generated for quantification of CAIX expression. The plasma level of CAIX was determined by ELISA in 209 of these NSCLC patients and in 58 healthy individuals. The CAIX tissue immunostaining and plasma levels were correlated with clinicopathological factors and patient outcome.

Results:

CAIX tissue overexpression correlated with shorter overall survival (OS) (P=0.05) and disease-specific survival (DSS) of patients (P=0.002). The CAIX plasma level was significantly higher in patients with NSCLC than in healthy individuals (P<0.001). A high level of CAIX in the plasma of patients was associated with shorter OS (P<0.001) and DSS (P<0.001), mostly in early stage I+II NSCLC. Multivariate Cox analyses revealed that high CAIX tissue expression (P=0.002) was a factor of poor prognosis in patients with resectable NSCLC. In addition, a high CAIX plasma level was an independent variable predicting poor OS (P<0.001) in patients with NSCLC.

Conclusion:

High expression of CAIX in tumour tissue is a predictor of worse survival, and a high CAIX plasma level is an independent prognostic biomarker in patients with NSCLC, in particular in early-stage I+II carcinomas.     

Associations between hepatocyte growth factor, c-Met, and basic fibroblast growth factor and survival in endometrial cancer patients

Background:

Hepatocyte growth factor (HGF), c-Met, and basic fibroblast growth factor (bFGF) are molecular markers that contribute to angiogenesis and proliferation in numerous cancers. We assessed the prognostic significance of these factors in tumour and stroma of endometrial cancer (EC) patients (n=211).

Methods:

Immunohistochemistry (IHC) was used to detect tumour and stromal protein expression of the biomarkers. Associations between expression and clinicopathological factors were assessed using Chi-square tests. Kaplan–Meier curves, log-rank tests, and Cox regression were used to summarise associations between biomarker expression and overall survival (OS) and recurrence-free survival (RFS).

Results:

Tumour bFGF was significantly associated with high-grade endometrioid and clear cell histology (P<0.001), advanced stage (P=0.008), positive lymph-node involvement (P=0.002), poor OS (log-rank test, P=0.009), and poor RFS (P<0.001). In multivariable analyses, cases with HGF-positive, stromal bFGF-positive tumours had a lower risk of death compared with cases with HGF-positive, stromal bFGF-negative tumours (hazard ratio (HR): 0.14, 95% CI: 0.03, 0.60). Cases with HGF-positive, bFGF-positive tumours had a higher risk of recurrence compared with cases with negative expression of both markers (HR: 9.88, 95% CI: 2.63, 37.16).

Conclusion:

These IHC data show that tumour and stromal bFGF expression have opposite associations with survival outcomes in EC patients. If confirmed in larger studies, tumour-derived bFGF could be an attractive target in EC therapy.     

The chemokine, CXCL12, is an independent predictor of poor survival in ovarian cancer

Background:

The chemokine CXCL12 and its cognate receptor, CXCR4, have been implicated in numerous tumour types where expression promotes tumour growth, angiogenesis, metastasis and suppresses tumour immunity.

Methods:

Using a tissue microarray of 289 primary ovarian cancers coupled to a comprehensive database of clinicopathological variables, the expression of CXCL12 and CXCR4 was assessed by immunohistochemistry and its impact in terms of survival and clinicopathological variables was determined.

Results:

Patients whose tumours expressed high levels of CXCL12 had significantly poorer survival (P=0.026) than patients whose tumours failed to produce this chemokine. Lack of CXCL12 expression within tumours was associated with a 51-month survival advantage for patients when compared with patients whose tumours expressed high levels of CXCL12. FIGO stage, adjuvant chemotherapy and the absence of macroscopic disease after surgery were all shown to predict prognosis independently of each other in this cohort of patients. CXCL12 was independently predictive of prognosis on multivariate analysis (P=0.016). There was no correlation between CXCL12 and any clinicopathological variable.

Conclusion:

The chemokine CXCL12 is an independent predictor of poor survival in ovarian cancer. High expression of CXCL12 was seen in only 20% of the tumours, suggesting a role for anti-CXCL12/CXCR4 therapy in the management of these patients.     

Aurora kinase A (AURKA) expression in colorectal cancer liver metastasis is associated with poor prognosis

Background:

Five-year survival after resection of colorectal cancer liver metastasis (CRLCM) is <30%. We recently found that aurora kinase A (AURKA) drives 20q gain-associated tumour progression and is associated with disease recurrence. This study evaluates the prognostic value of AURKA expression in CRCLM of patients who underwent liver resection.

Methods:

Tissue microarrays (TMAs) were generated using formalin-fixed paraffin-embedded CRCLM and matched primary tumour from a multi-institutional cohort of patients with CRCLM who underwent liver resection between 1990 and 2010. Tissue microarrays were stained for AURKA using immunohistochemistry, and a hazard rate ratio (HRR) for the association between overall survival (OS) and nuclear AURKA expression in CRCLM was calculated. Results were validated by 500-fold cross-validation.

Results:

The expression of AURKA was evaluated in CRCLM of 343 patients. High AURKA expression was associated with poor OS (HRR 1.55, P<0.01), with a cross-validated average HRR of 1.57 (P=0.02). Average HRR was adjusted for the established prognostic clinicopathological variables in a multivariate analysis (average HRR 1.66; P=0.02). The expression of AURKA in CRCLM was correlated to its expression in corresponding primary tumour (P<0.01).

Conclusion:

The expression of AURKA protein is a molecular biomarker with prognostic value for patients with CRCLM, independent of established clinicopathological variables.     

The prognostic impact of a combined carbonic anhydrase IX and Ki67 signature in oral squamous cell carcinoma

Background:

Tumour hypoxia is associated with impaired apoptosis, resistance to therapy and poor prognosis. We previously reported that high stromal expression of the endogenous marker of hypoxia, carbonic anhydrase IX (CAIX), is associated with significantly reduced survival in oral squamous cell carcinoma (OSCC). In addition to hypoxia, CAIX expression is regulated by proliferation-associated signalling. We hypothesised that incorporating Ki67, a proliferation marker, into our existing CAIX-based stratification of OSCC would identify patients with the least favourable prognosis.

Methods:

Surgically resected tumours from 60 OSCC patients were analysed for CAIX, Ki67 and BAX expression using fluorescence immunohistochemistry and automated quantitative analysis (AQUA).

Results:

In patients expressing high stromal CAIX (sCAIX), stratification by tumour Ki67 expression revealed significantly distinct survival outcomes (P=0.005). In our OSCC cohort, below-median Ki67 and top-quartile sCAIX expression (Ki67^losCAIX^hi) were associated with significantly worse disease-specific survival in univariate (HR 7.2 (2.5–20.4), P=0.001) and multivariate (HR 4.2 (1.4–12.8), P=0.011) analyses. Hypoxia is associated with decreased BAX expression; the Ki67^losCAIX^hi group was more strongly associated with low BAX expression than high sCAIX alone.

Conclusion:

These data suggest that combined analysis of tumour Ki67 and sCAIX expression may provide a more clinically relevant assessment of tumour hypoxia in OSCC.     

CXCR7 expression is associated with disease-free and disease-specific survival in cervical cancer patients

Background:

The CXC chemokine receptor (CXCR)7 is involved in tumour development and metastases formation. The aim of the present study was to determine protein expression of CXCR7, its putative co-receptors epidermal growth factor receptor (EGFR) and CXCR4, its predominant ligand CXCL12, their co-dependency and their association with survival in cervical cancer patients.

Methods:

CXC chemokine receptor 7, EGFR, CXCR4 and CXCL12 expression were determined immunohistochemically in 103 paraffin-embedded, cervical cancers. Subsequently, associations with patient characteristics were assessed and survival analyses were performed.

Results:

CXC chemokine receptor 7 was expressed by 43% of tumour specimens, in a large majority of cases together with either EGFR or CXCR4 (double positive), or both (triple positive). The CXCR7 expression was associated with tumour size (P=0.013), lymph node metastasis (P=0.001) and EGFR expression (P=0.009). CXC chemokine receptor 7 was independently associated with disease-free survival (hazard ratio (HR)=4.3, 95% confidence intervals (CI) 1.7–11.0, P=0.002), and strongly associated with disease-specific survival (HR=3.9, 95% CI 1.5–10.2, P=0.005).

Conclusion:

CXC chemokine receptor 7 expression predicts poor disease-free and disease-specific survival in cervical cancer patients, and might be a promising new therapeutic marker. In a large majority of cases, CXCR7 is co-expressed with CXCR4 and/or EGFR, supporting the hypothesis that these receptors assist in CXCR7 signal transduction.     

Quantitative assessment of miR34a as an independent prognostic marker in breast cancer

Background:

Aberrant expression of microRNAs (miRNAs) is associated with cancer progression, initiation and metastasis. MiR34a is a miRNA that has been previously described as a tumour suppressor. Herein, we assess the expression of miR34a in three independent breast cancer cohorts using a quantitative in situ hybridisation assay (qISH) and determined its association with disease-specific death in breast cancer.

Methods:

The qISH method was applied to three independent primary breast cancer cohorts (Cohort 1 with 461, Cohort 2 with 279 and Cohort 3 with 795 patients) using 5′ and 3′ double DIG-labelled LNA-modified probe against miR34a using the protocol described previously. Level of expression measured as automated quantitative analysis (AQUA) score for miR34a was determined for each patient and assessed for association with risk of disease-specific death. An optimal cutpoint was determined using the X-tile software for disease-specific survival in Cohort 1 and this cutpoint was then applied to the other two cohorts after median normalisation of AQUA scores.

Results:

Loss of miR34a is associated with poor outcome in three independent breast cancer cohorts (uncorrected log-rank P=0.0188 for Cohort 1, log-rank P=0.0024 for Cohort 2 and log-rank P=0.0455 for Cohort 3). In all three cohorts, loss of miR34a is able to stratify patients with poor disease-specific survival among node-negative patients, but not in node-positive population. Multivariate Cox proportional hazards analysis in Cohort 1 (P=0.0381) and Cohort 2 (P=0.0468) revealed that loss of miR34a is associated with poor outcome, independent of age, node status, receptor status and tumour size.

Conclusion:

Loss of the tumour suppressor, miR34a, identifies a subgroup of breast cancer patients with poor disease-specific survival. This study is consistent with the well-established preclinical observations for miR34a as a tumour suppressor and suggests that miR34a may have future value as a biomarker in breast cancer.     

Urinary TIMP-1 and MMP-2 levels detect the presence of pancreatic malignancies

Background:

A majority of patients with pancreatic malignancies, including both pancreatic ductal adenocarcinoma (PDAC) and pancreatic neuroendocrine tumours (pNETs), present with advanced disease due to a lack of specific symptoms and current diagnostic limitations, making this disease extremely difficult to detect. Our goal was to determine whether urinary matrix metalloproteases (uMMPs) and/or their endogenous inhibitors, urinary tissue inhibitor of metalloproteases (uTIMPs), could be detected in the urine of patients with pancreatic malignancies and whether they may serve as independent predictors of disease status.

Methods:

Retrospective analyses of urine samples (n=139) from PDAC and pNET patients as well as age- and sex-matched controls were conducted. Urinary MMP-2 and uTIMP-1 levels were determined using ELISA and zymography. Biomarker expression in tumour and normal pancreatic tissues was analysed via immunohistochemistry (IHC).

Results:

Multivariable logistic regression analyses indicated that, when controlling for age and sex, uMMP-2 (P<0.0001) and uTIMP-1 (P<0.0001) but not uMMP-9, were significant independent predictors for distinguishing between PDAC patients and healthy controls. Our data also indicated that uMMP-2 was an independent predictor of the presence of pNET. In addition, uTIMP-1 levels could differentiate the two cancer groups, PDAC and pNET, respectively. Immunohistochemistry analysis confirmed that MMP-2 and TIMP-1 protein expression is significantly upregulated in PDAC tissue compared with the normal pancreas.

Conclusions:

Taken together, our results suggest that the detection of uMMP-2 and uTIMP-1 may have diagnostic value in the detection of pancreatic malignancies and that uTIMP-1 may be useful in distinguishing between pancreatic adenocarcinoma and neuroendocrine tumours.     

Stromal infiltration of CD8 T cells is associated with improved clinical outcome in HPV-positive oropharyngeal squamous carcinoma

Background:

Patients with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) have a better prognosis than those with HPV-negative tumours. There is interest in de-escalating their treatment but strategies are needed for risk stratification to identify subsets with a poor prognosis. This study investigated tumour-infiltrating lymphocytes (TILs) in relation to HPV tumour status and patient survival.

Methods:

Biopsies from 218 patients diagnosed with OPSCC between 2002 and 2011, who underwent chemo/radiotherapy were analysed for HPV by PCR, in-situ hybridisation and p16 immunohistochemistry (IHC). One hundred and thirty-nine samples with concordant HPV detection were analysed for CD3, CD4, CD8 and FoxP3 expression in tumour and stromal regions using multiplexIHC and multispectral image analysis. Labelling of smooth muscle actin (SMA) identified activated stroma.

Results:

Human papillomavirus-positive compared with HPV-negative OPSCC had higher infiltration in both tumour and stromal areas of CD4 and CD8 T cells but not FoxP3 T regulatory cells. Only CD3+CD8+ stromal and not tumour area infiltration was associated with increased survival (P=0.02). There was significantly higher SMA expression in HPV-positive compared with -negative tumours, which did not correlate with survival.

Conclusions:

Studies of TILs for risk stratification in OPSCC should assess stromal infiltration.     

hERG1 channels drive tumour malignancy and may serve as prognostic factor in pancreatic ductal adenocarcinoma

Background:

hERG1 channels are aberrantly expressed in human cancers. The expression, functional role and clinical significance of hERG1 channels in pancreatic ductal adenocarcinoma (PDAC) is lacking.

Methods:

hERG1 expression was tested in PDAC primary samples assembled as tissue microarray by immunohistochemistry using an anti-hERG1 monoclonal antibody (α-hERG1-MoAb). The functional role of hERG1 was studied in PDAC cell lines and primary cultures. ERG1 expression during PDAC progression was studied in Pdx-1-Cre,LSL-Kras^G12D/+,LSL-Trp53^R175H/+ transgenic (KPC) mice. ERG1 expression in vivo was determined by optical imaging using Alexa-680-labelled α-hERG1-MoAb.

Results:

(i) hERG1 was expressed at high levels in 59% of primary PDAC; (ii) hERG1 blockade decreased PDAC cell growth and migration; (iii) hERG1 was physically and functionally linked to the Epidermal Growth Factor-Receptor pathway; (iv) in transgenic mice, ERG1 was expressed in PanIN lesions, reaching high expression levels in PDAC; (v) PDAC patients whose primary tumour showed high hERG1 expression had a worse prognosis; (vi) the α-hERG1-MoAb could detect PDAC in vivo.

Conclusions:

hERG1 regulates PDAC malignancy and its expression, once validated in a larger cohort also comprising of late-stage, non-surgically resected cases, may be exploited for diagnostic and prognostic purposes in PDAC either ex vivo or in vivo.