The International Journal of Biological Markers

Many studies have evaluated the accuracy of EGFR mutation status in blood against that in tumor tissues as the reference. We conducted this systematic review and meta-analysis to assess whether blood can be used as a substitute for tumor tissue in detecting EGFR mutations.

Investigations that provided data on EGFR mutation status in blood were searched in the databases of Medline, Embase, Ovid Technologies and Web of Science. The detect efficiency of EGFR mutations in paired blood and tissues was compared using a random-effects model of meta-analysis. Pooled sensitivity and specificity and diagnostic accuracy were calculated by receiver operating characteristic curve.

A total of 19 studies with 2,922 individuals were involved in this meta-analysis. The pooled results showed the positive detection rate of EGFR mutations in lung cancer tissues was remarkably higher than that of paired blood samples (odds ratio [OR] = 1.47, p<0.001). The pooled sensitivity and specificity of blood were 0.65 and 0.91, respectively, and the area under the receiver operating characteristic curve was 0.89.

Although blood had a better specificity for detecting EGFR mutations, the absence of blood positivity should not necessarily be construed as confirmed negativity. Patients with negative results for blood should decidedly undergo further biopsies to ascertain EGFR mutations.

This study aimed to elucidate the concordance between human epidermal growth factor receptor 2 (HER2) immunohistochemistry (IHC) and in situ hybridization (ISH) and the diagnostic accuracy of HER2 IHC in non-small cell lung cancer (NSCLC) through a meta-analysis and diagnostic test accuracy review.

Seven eligible studies and 1,217 patients with NSCLC were included in the review. The concordance between HER2 IHC and ISH was analyzed. To confirm the diagnostic accuracy of HER2 IHC, the sensitivity and specificity were analyzed and the area under the curve (AUC) in the summary receiver operating characteristic (SROC) curve was calculated.

The concordance rate between HER2 IHC and ISH was 0.795 (95% confidence interval [CI] 0.534-0.929). In the HER2 IHC-negative (score 0/1+) subgroup, the concordance rate was 0.975 (95% CI 0.854-0.996). The concordance rates of the HER2 IHC score 2+ and 3+ subgroups were 0.091 (95% CI 0.039-0.197) and 0.665 (95% CI 0.446-0.830), respectively. In diagnostic test accuracy review, the pooled sensitivity and specificity were 0.67 (95% CI 0.54-0.78) and 0.89 (95% CI 0.87-0.91), respectively. The AUC in the SROC curve was 0.891 and the diagnostic odds ratio was 16.99 (95% CI 5.08-56.76).

HER2 IHC was largely in agreement with ISH in cases of HER2 IHC score 0/1+. Because the concordance rates of HER2 IHC score 2/3+ cases were lower than that of HER2 IHC score 0/1+ cases, further studies for detailed analysis criteria for HER2 IHC score 2+ or 3+ are required.

Recently, pro-gastrin-releasing peptide (pro-GRP) became available as an alternative sensitive, specific and reliable tumor marker for patients with small cell lung cancer (SCLC), both in limited (LD) and diffuse disease (DD).

We retrospectively analyzed pro-GRP, neuron-specific enolase (NSE) and CEA in patients with SCLC and non-small cell lung cancer (NSCLC). Serum pro-GRP level was measured with electrochemiluminescence at our laboratory (cutoff 77.8 pg/mL). Continuous variables were analyzed with the Mann-Whitney test, contingency data with Fisher’s exact test. Receiver operator characteristic (ROC) curve analysis was performed to identify threshold values to set the highest sensitivity (Sn) and specificity (Sp) values.

A total of 65 patients were studied (49 men, median age 67 years, range 27-79). Thirty-seven patients had SCLC (29 DD, 8 LD) and 28 advanced NSCLC. Median pro-GRP level was 919 pg/mL (range 22-147,350) in SCLC and 32 pg/mL (range 10-119.2) in NSCLC (p<0.0001). NSE was 4.38-fold higher in SCLC patients (p = 0.0005); CEA did not reveal significant differences between groups. Pro-GRP Sn and Sp were 86.4% and 96.4%, respectively. With ROC curve analysis, a cutoff value of 329.3 pg/mL showed a Sn of 75.8% and Sp of 87.5% in discriminating DD from LD. Pro-GRP was not influenced by either liver metastases or renal impairment.

Pro-GRP is sensitive for SCLC diagnosis. Since high marker levels are related to high disease burden, pro-GRP may have a negative prognostic significance. Follow-up studies are required to define its role in clinical practice in monitoring responses to treatment and early relapses.

The aim of this study was to elucidate the clinicopathological significance of enhancer of zeste homolog 2 (EZH2) expression in gliomas, including its incidence and prognostic role. In addition, we investigated the concordance between immunohistochemistry and polymerase chain reaction for determining the presence or absence of EZH2 in these tumors.

The current meta-analysis included 1,049 gliomas with various WHO tumor grades from 12 eligible studies, which were analyzed for positivity of EZH2 and correlation between EZH2 expression and prognosis. Subgroup analyses were performed based on detection methods and WHO tumor grades.

We found the estimated positive rate of EZH2 in gliomas to be 0.663 (95% confidence interval [95% CI], 0.549-0.761). There was no difference between immunohistochemistry and polymerase chain reaction in determination of EZH2 positivity (0.706, 95% CI, 0.539-0.831 vs. 0.673, 95% CI, 0.472-0.825). The positive rate of EZH2 increased by increasing WHO tumor grade. EZH2 expression was significantly correlated with worse overall and progression-free survival (hazard ratio [HR] = 2.436, 95% CI, 1.350-4.393, and HR = 4.071, 95% CI, 1.325-12.508, respectively). The overall concordance rate between immunohistochemistry and polymerase chain reaction was 0.885 (95% CI, 0.300-0.993).

EZH2 positivity was significantly correlated with WHO tumor grade and worse prognosis in gliomas.

We aimed to evaluate the genetic variation of poly(ADP-ribose) polymerase-1 (PARP-1) in the development of gliomas among Chinese individuals.

Patients with a confirmed diagnosis of glioma and healthy individuals with no clinical symptoms of glioma were enrolled at Liaocheng People’s Hospital, China. Genetic polymorphisms were studied in plasma samples by polymerase chain reaction-restriction fragment length polymorphism assay. Cytokine levels were measured routinely in serum samples by sandwich ELISA technique.

A total of 120 Chinese patients with gliomas and 120 healthy Chinese individuals were included. We found that patients with the GG genotype (odds ratio [OR] 2.53, 95% confidence interval [CI] 1.46-4.38, p<0.001) and carriers of the G allele (OR 11.5, 95% CI 6.31-21.3, p<0.0001) were at high risk of developing glioma. A del/ins polymorphism of the NF-κB1 gene (OR 4.27, 95% CI 2.43-7.50, p<0.001) was also found to be associated with glioma. In addition, significantly increased cytokine levels were observed in patients with glioma (p<0.05).

Our findings showed that PARP-1 polymorphisms are involved in the development of glioma in Chinese individuals. Also serum cytokine levels can be considered among the potential risk factors for developing glioma.

Laryngeal squamous cell carcinoma (LSCC) has a poor prognosis due to recurrence and metastasis. IQ-domain GTPase-activating protein 1 (IQGAP1), a scaffold protein, plays an important role in tumorigenesis and malignant development. In this study, we aimed to explore the role of IQGAP1 in LSCC.

Expression of IQGAP1 in human LSCC specimens was assessed by immunohistochemistry. We also evaluated the roles of IQGAP1 in cell proliferation, migration and invasion and epithelial-to-mesenchymal transition (EMT) in Hep-2 cells.

The expression of IQGAP1 protein was significantly up-regulated in LSCC tissues compared with normal laryngeal tissues (p = 0.002). Furthermore, the knockdown of IQGAP1 in Hep-2 cells inhibited cell growth, migration and invasion. Moreover, we found that IQGAP1 silencing reversed EMT.

These results show for the first time that IQGAP1 is up-regulated in LSCC tissues and plays an important role in LSCC cell proliferation and invasiveness, which indicates that IQGAP1 could work as an oncogene and may serve as a promising molecular target for treatment of LSCC.

The most important adverse prognostic factor for laryngeal squamous cell carcinoma (LSCC) is the presence of cervical lymph node metastases. The supraglottic area of the larynx is richly supplied with lymphatics, and 25%-75% of supraglottic carcinomas metastasize in neck lymph nodes. Cortactin is a multidomain protein related to actin cytoskeleton regulation, podosome and lamellipodia formation, integrin signaling, axon guidance and extracellular matrix degradation. Cortactin is involved in metastasis formation because of its role in cell mobility. The present study focused mainly on the role of cortactin and phosphorylated cortactin (residues tyr⁴²¹ and tyr⁴⁶⁶) expression and subcellular localization in primary supraglottic LSCCs and their cervical lymph node metastases.

The immunohistochemical expression of cortactin, p-Y466-cortactin and p-Y421-cortactin was assessed in 38 primary supraglottic LSCCs and 10 lymph node metastases. The statistical approach included bootstrapping analysis.

Despite a significantly higher expression of cortactin in carcinoma cells than in adjacent normal laryngeal mucosa, no associations emerged between prognosis and the expression of cortactin or its isoforms in supraglottic LSCC. Statistical analysis found cortactin expression higher in less-differentiated LSCCs (p = 0.03). A significant direct correlation was found between cortactin and p-Y466-cortactin levels (p = 0.031), and between p-Y466-cortactin and p-Y421-cortactin levels (p = 0.001).

Cortactin expression in carcinoma cells and its known involvement in the EGFR pathway suggest a role for this protein as a target for LSCC therapy. Further prospective studies are needed to investigate the potential of cortactin, p-Y466-cortactin and p-Y421-cortactin expression as markers of response to treatment (particularly EGFR-directed agents) in LSCC.

This study aimed to uncover potential microRNA (miRNA) targets in head and neck squamous cell carcinoma (HNSCC) using an ensemble method which combined 3 different methods: Pearson’s correlation coefficient (PCC), Lasso and a causal inference method (i.e., intervention calculus when the directed acyclic graph (DAG) is absent [IDA]), based on Borda count election.

The Borda count election method was used to integrate the top 100 predicted targets of each miRNA generated by individual methods. Afterwards, to validate the performance ability of our method, we checked the TarBase v6.0, miRecords v2013, miRWalk v2.0 and miRTarBase v4.5 databases to validate predictions for miRNAs. Pathway enrichment analysis of target genes in the top 1,000 miRNA–messenger RNA (mRNA) interactions was conducted to focus on significant KEGG pathways. Finally, we extracted target genes based on occurrence frequency ≥3.

Based on an absolute value of PCC >0.7, we found 33 miRNAs and 288 mRNAs for further analysis. We extracted 10 target genes with predicted frequencies not less than 3. The target gene MYO5C possessed the highest frequency, which was predicted by 7 different miRNAs. Significantly, a total of 8 pathways were identified; the pathways of cytokine–cytokine receptor interaction and chemokine signaling pathway were the most significant.

We successfully predicted target genes and pathways for HNSCC relying on miRNA expression data, mRNA expression profile, an ensemble method and pathway information. Our results may offer new information for the diagnosis and estimation of the prognosis of HNSCC.

Bladder cancer (BC) represents the most common neoplasm of the urinary tract. Although cystoscopy and urine cytology represent the gold standard methods to monitor BC, both procedures have limitations. Therefore, the identification of reliable biomarkers for early and noninvasive detection of BC is urgently required.

In this study, we analyzed nicotinamide N-methyltransferase (NNMT) expression in urine samples from 55 BC patients and 107 controls, using real-time polymerase chain reaction (PCR). Receiver operating characteristic (ROC) analysis was used to identify the best cutoff value to discriminate BC patients from healthy donors, and to evaluate the diagnostic accuracy of a urine-based NNMT test.

The results demonstrated that urinary NNMT expression was significantly (p<0.05) higher in BC patients. Moreover, a significant (p<0.05) inverse correlation was found between NNMT expression and histological grade. The ROC analysis revealed that a ΔCq of 13.3 was the best cutoff value, since it was associated with the highest combination of sensitivity and specificity. Moreover, the area under the curve (AUC) value was 0.913 (p<0.05), indicating the excellent diagnostic accuracy of a urine-based NNMT test.

Our data indicate that NNMT is a promising biomarker that could be used to support the early and noninvasive diagnosis of BC.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, characterized by advanced disease stage and poor prognosis. Moreover, due to the lack of therapeutic markers, TNBC patients can’t benefit fully from currently available targeted therapies.

To fully understand the molecular basis of TNBC, we used gene set enrichment analysis (GSEA) to screen out the most altered functional module in TNBC, from publicly available microarray data and studied the association of the candidate gene with TNBC development.

We found that the proteasome was significantly activated in TNBC. As compared with other breast cancer subtypes and normal tissue, proteasome subunit beta 5 (PSMB5), the key regulator of proteasome function, was overexpressed in TNBC tissue and predictive of poor prognosis. Moreover, we also found that PSMB5 knockdown induced TNBC apoptosis and significantly enhanced cancer cell sensitivity to the chemotherapeutic agents bortezomib and paclitaxel.

Our results suggest a potential role for PSMB5 as a biomarker and therapeutic target for TNBC.

The objective of this study was to examine the prognostic significance of carbonic anhydrase IX (CAIX), an endogenous marker for tumor hypoxia; the cellular tumor antigen p53; and the apoptosis regulator Bcl-2, in triple-negative breast cancer (TNBC) patients.

Immunohistochemically determined expression of CAIX, p53, Bcl-2 and proliferation factor Ki-67, analyzed in 64 paraffin-embedded TNBC tissue samples, was used to assess their relation to clinicopathological variables and prognostic implications for overall survival (OS).

Bcl-2 expression was negatively correlated with histological grade of tumor, while expression of p53 was positively correlated with the same clinical variable (p = 0.036 and p = 0.033, respectively). The p53 expression was also positively correlated with tumor size (p = 0.010). Survival analysis showed that patients with high Bcl-2 expression (above cutoff value determined by receiver operator characteristic [ROC] curve analysis) had shorter OS (p = 0.020). The same was observed for patients with tumors larger than 5 cm (p = 0.034) or positive lymph nodes (p = 0.004). Among all 3 examined markers, multivariate analysis showed that only Bcl-2 expression was a strong independent prognostic indicator for decreased OS (hazard ratio [HR] = 15.16, 95% confidence interval [95% CI], 2.881-79.727, p = 0.001).

Elevated expression of Bcl-2 was an independent prognostic factor for poorer OS in TNBC and as such a significant marker for tumor aggressiveness.

Several studies have suggested that there are single nucleotide polymorphisms (SNPs) that can be considered potential biomarkers in the prognosis and therapeutic response of cancer patients. The present study investigated the association between ERCC1 rs3212986 and XRCC3 rs861539 polymorphisms and clinical toxicities induced by chemoradiotherapy (CRT) in cervical cancer.

This hospital-based retrospective cohort study included 260 patients with cervical cancer, FIGO stages Ib2-IVa, who underwent CRT (cisplatin). Genetic polymorphisms analysis was performed by allelic discrimination with real-time polymerase chain reaction (RT-PCR).

Our results indicated a link between ERCC1 rs3212986 and the onset of late gastrointestinal toxicity (p = 0.038). Furthermore, using a recessive model (AA vs. CC/CA), we found that patients carrying AA homozygous genotype presented a fourfold increased risk of developing late gastrointestinal toxicity when compared with patients with the C allele (odds ratio = 3.727, 95% confidence interval, 1.199-11.588; p = 0.017). No association was found regarding the XRCC3 rs861539 polymorphism and any clinical toxicity event.

This is the first study evaluating the relationship between these polymorphisms and clinical toxicities in cervical cancer patients submitted to CRT with cisplatin. These results may contribute toward a better understanding of the influence of genetic polymorphisms in genes associated with DNA repair in the clinical response to CRT of patients with cervical cancer.

In this study we investigated the function of the non-catalytic region of tyrosine kinase adaptor protein 2 (NCK2) and its correlation with ITGB1 and ITGB4 integrins in driving ovarian cancer (OvCa) aggressiveness. We also evaluated whether NCK2 may influence prognosis in OvCa patients.

Nanofluidic technology was used to analyze expression of NCK2 in 332 OvCa patients. To evaluate mRNA expression of NCK2, integrins and VEGFA in OvCa cell lines, qRT-PCR was performed. Stable NCK2 overexpression was obtained in OVCAR3. qRT-PCR and Western blot were performed to evaluate expression changes of VEGFA, vimentin, ITGB1, ITGB4, MMP2 and MMP9 under normoxia and hypoxia conditions. Coimmunoprecipitation (Co-IP) was performed in the A2780 cell line to study the interaction between NCK2 and proteins of interest. To investigate whether NCK2 can influence anchorage-independent growth, a soft agar assay was completed. Transwell invasion assay was performed on stable-transfected OVCAR-3 cell lines.

Nanofluidic data showed NCK2 can play an important role as a factor promoting tumor aggressiveness and survival in OvCa. This role was also linked to the behaviors of ITGB1 and ITGB4. Moreover, in cells overexpressing NCK2, the expression of vimentin, MMP2, MMP9, VEGFA and ITGB1, but not of ITGB4 was induced by hypoxia. Co-IP showed that NCK2 can directly bind ITGB1, but not VEGFA. NCK2 may be involved in mediating cell-extracellular matrix interactions in OvCa cells by influencing tumor aggressiveness.

This study provides evidence of a possible role of NCK2 as biomarker of OvCa progression.