• cancer imaging
• kallikreins
• metastasis
• protease inhibitors
• serine proteases

• Male
• Humans
• Female
• Serine Proteinase Inhibitors/therapeutic use
• Amyloid beta-Peptides/therapeutic use
• Prostate/pathology
• Amyloid beta-Protein Precursor/pharmacology
• Amyloid beta-Protein Precursor/therapeutic use
• Breast Neoplasms/drug therapy
• Breast Neoplasms/genetics
• Breast Neoplasms/pathology
• Ovarian Neoplasms/drug therapy
• Ovarian Neoplasms/genetics
• Kallikreins/genetics

• 20-0238 Worldwide Cancer Research
• Worldwide Cancer Research

The kallikrein (KLK) family of genes consists of 15 members, many of which are highly expressed in number of cancers compared to their normal parent tissues. KLK7 was initially characterized as an enzyme implicated in the degradation of intercellular cohesive structures in the stratum corneum of stratified squamous epithelia, preceding desquamation in the skin. It catalyzes the degradation of desmosomes in the outermost layer of skin and permits cell shedding to take place at the skin surface. Overexpression of KLK7 in tumor cells has been reported to significantly enhance the invasive potential in intracranial malignancies and ovarian cancer cells. Thus, KLK7 could contribute to the degradation of extracellular matrices in oral squamous cell carcinoma (OSCC) tissues, promoting invasion of neoplastic cells locally and facilitating metastasis to regional lymph nodes. The objectives of the present study were to compare the expression of KLK 7 in normal subjects and patients with OSCC, to correlate the expression of KLK 7 with respect to the clinical staging of OSCC and to evaluate the expression of KLK 7with respect to different histopathological grades of OSCC.

Thirty cases of OSCC were staged clinically and graded histopathologically. The immunohistochemical method was used to detect the expression of KLK 7 in OSCC. The scores obtained were documented and compared statistically.

KLK 7 immunoreactivity was noticed in all cases of OSCC. A statistically significant difference was observed in immunoreactivity of KLK 7 between the normal and OSCC (P = 0.0001*) and in different histopathological grades (P = 0.0001*) and in different clinical stages (P = 0.0127*) of OSCC using Kruskal–Wallis analysis of variance test.

The KLK 7 immunoexpression histopathologically increased from low grade to high grade and clinically from Stage 1 to Stage 4 in OSCC. Hence, increased expression of KLK 7 may be related to poor prognosis in patients with OSCC.

• Immunohistochemistry
• Kallikrein 7
• oral squamous cell carcinoma
• prognosis

Tumor‐associated macrophages (TAMs) have important roles in the growth, angiogenesis and progression of various tumors. Although we have demonstrated the association of an increased number of infiltrating CD204⁺ TAMs with poor prognosis in esophageal squamous cell carcinomas (ESCCs), the roles of TAMs in ESCC remain unclear. Here, to study the effects of TAMs on the tumor microenvironment of ESCCs, we established a co‐culture assay using a human ESCC cell line and TAM‐like peripheral blood monocyte‐derived macrophages and performed a cDNA microarray analysis between monocultured and co‐cultured ESCC cell lines. Our qRT‐PCR confirmed that in the co‐cultured ESCC cell lines, CYP1A1, DHRS3, ANXA10, KLK6 and CYP1B1 mRNA were highly up‐regulated; AMTN and IGFL1 mRNA were down‐regulated. We observed that the high expression of a calcium‐dependent phospholipid‐binding protein ANXA10 was closely associated with the depth of invasion and high numbers of infiltrating CD68⁺ and CD204⁺ TAMs and poor disease‐free survival (P = 0.0216). We also found ANXA10 promoted the cell growth of ESCC cell lines via the phosphorylation of Akt and Erk1/2 pathways in vitro. These results suggest that ANXA10 induced by the interaction with TAMs in the tumor microenvironment is associated with cell growth and poor prognosis in human ESCC tissues.

• ANXA10
• cell growth
• esophageal squamous cell carcinoma
• tumor microenvironment
• tumor-associated macrophage

Kallikrein (KLK) proteases are hormone-like signaling molecules with critical functions in different cancers. This study investigated the expression of KLK6 in gastric cancer and its potential role in the growth, migration, and invasion of gastric cancer cells.

In this study, we compared protein levels of KLK6, vascular endothelial growth factor (VEGF), and matrix metallopeptidase (MMP) 9 in normal gastric epithelial and gastric cancer cell lines by western blot. Fluorescence-activated cell sorting was employed to sort 2 clones of SGC-7901 cells with distinct KLK6 expression, namely, KLK6-high (KLK6^high) and KLK6-low (KLK6^low), which were then expanded. Lastly, immunohistochemical analysis was performed to investigate KLK6 expression in gastric cancer patients.

The expression levels of KLK6, VEGF, and MMP 9, were significantly higher in the gastric cancer cell lines SGC-7901, BGC-823, MKN-28, and MGC-803 than in the normal gastric epithelial cell line GES-1. Compared to KLK6^low cells, KLK6^high cells showed enhanced viability, colony-forming ability, migration, and invasion potential in vitro. Importantly, immunohistochemical analysis of a human gastric cancer tissue cohort revealed that the staining for KLK6, VEGF, and MMP9 was markedly stronger in the cancerous tissues than in the adjacent normal tissues. KLK6 expression also correlated with that of VEGF and MMP9 expression, as well as several key clinicopathological parameters.

Together, these results suggest an important role for KLK6 in human gastric cancer progression.

• Gastric cancer
• KLK6 protein, human
• MMP9 protein, human
• VEGF

• X-ray crystallography
• cell invasion
• directed evolution
• enzyme inhibition
• protease inhibitor
• protein engineering
• proteolysis
• selective binding
• yeast surface display

• ELISA
• KLK6
• immunocapture
• kallikrein 6
• mass spectrometry
• α1-antitrypsin

• Animals
• Antibodies, Monoclonal/immunology
• Biomarkers/blood
• Body Fluids/chemistry
• Enzyme-Linked Immunosorbent Assay
• Female
• Humans
• Kallikreins/blood
• Kallikreins/immunology
• Mice
• Mice, Inbred BALB C
• Ovarian Neoplasms/blood
• Ovarian Neoplasms/diagnosis
• Ovarian Neoplasms/immunology
• alpha 1-Antitrypsin/blood
• alpha 1-Antitrypsin/immunology

• Amino Acids/chemistry
• Amino Acids/genetics
• Computer Simulation
• Genes, Neoplasm/genetics
• Genomics/methods
• Humans
• Hydrophobic and Hydrophilic Interactions
• Models, Theoretical
• Neoplasms/classification
• Neoplasms/diagnosis
• Neoplasms/genetics
• ROC Curve
• Sequence Analysis, DNA/methods

• EOC
• TH
• epithelial ovarian cancer
• genetic and epigenetic alterations
• molecular targeted treatment
• tumoural heterogeneity

• Antineoplastic Agents/therapeutic use
• BRCA1 Protein/genetics
• BRCA1 Protein/metabolism
• Carcinoma, Ovarian Epithelial
• Chromosome Aberrations
• Drug Resistance, Neoplasm/genetics
• Epigenesis, Genetic
• Female
• Genetic Heterogeneity
• Humans
• Molecular Targeted Therapy
• Neoplasm Grading
• Neoplasm Proteins/genetics
• Neoplasm Proteins/metabolism
• Neoplasm Staging
• Neoplasms, Glandular and Epithelial/diagnosis
• Neoplasms, Glandular and Epithelial/drug therapy
• Neoplasms, Glandular and Epithelial/genetics
• Neoplasms, Glandular and Epithelial/pathology
• Ovarian Neoplasms/diagnosis
• Ovarian Neoplasms/drug therapy
• Ovarian Neoplasms/genetics
• Ovarian Neoplasms/pathology
• Poly(ADP-ribose) Polymerase Inhibitors/therapeutic use
• Poly(ADP-ribose) Polymerases/genetics
• Poly(ADP-ribose) Polymerases/metabolism
• Prognosis

• Cancer
• Carcinogenesis
• Function
• KLK
• Kallikrein-related peptidase
• Protease

Oral squamous cell carcinomas (OSCC) induced in F344 rats by 4-nitroquinoline-1-oxide (4-NQO) demonstrate considerable phenotypic similarity to human oral cancers. Gene expression studies (microarray and PCR) were coupled with methylation analysis of selected genes to identify molecular markers of carcinogenesis in this model and potential biochemical and molecular targets for oral cancer chemoprevention. Microarray analysis of 11 pairs of OSCC and site-matched phenotypically normal oral tissues from 4-NQO-treated rats identified more than 3500 differentially expressed genes; 1735 genes were up-regulated in rat OSCC versus non-malignant tissues, while 1803 genes were down-regulated. In addition to several genes involved in normal digestion, genes demonstrating the largest fold increases in expression in 4-NQO-induced OSCC include three lipocalins (VEGP1, VEGP2, LCN2) and three chemokines (CCL, CXCL2, CXCL3); both classes are potentially druggable targets for oral cancer chemoprevention and/or therapy. Down-regulated genes in 4-NQO-induced OSCC include numerous keratins and keratin-associated proteins, suggesting that alterations in keratin expression profiles may provide a useful biomarker of oral cancer in F344 rats treated with 4-NQO. Confirming and extending our previous results, PTGS2 (cyclooxygenase-2) and several cyclooxygenase-related genes were significantly up-regulated in 4-NQO-induced oral cancers; up-regulation of PTGS2 was associated with promoter hypomethylation. Rat OSCC also demonstrated increased methylation of the first exon of APC2; the increased methylation was correlated with down-regulation of this tumor suppressor gene. Overexpression of pro-inflammatory chemokines, hypomethylation of PTGS2, and hypermethylation of APC2 may be causally linked to the etiology of oral cancer in this model.

• 4-Nitroquinoline-1-oxide
• Animals
• Carcinoma, Squamous Cell/chemically induced
• Carcinoma, Squamous Cell/genetics
• Carcinoma, Squamous Cell/metabolism
• Chemokines/genetics
• Chemokines/metabolism
• Cyclooxygenase 1/genetics
• Cyclooxygenase 1/metabolism
• Cyclooxygenase 2/genetics
• Cyclooxygenase 2/metabolism
• DNA Methylation
• Gene Expression Regulation, Neoplastic
• Genes, Tumor Suppressor
• Keratins/genetics
• Keratins/metabolism
• Lipocalins/genetics
• Lipocalins/metabolism
• Male
• Membrane Proteins/genetics
• Membrane Proteins/metabolism
• Mouth Neoplasms/chemically induced
• Mouth Neoplasms/genetics
• Mouth Neoplasms/metabolism
• Oligonucleotide Array Sequence Analysis
• Promoter Regions, Genetic
• Rats, Inbred F344
• Tongue/pathology
• Transcriptome

Early detection of ovarian cancer remains a challenge due to widespread metastases and a lack of biomarkers for early-stage disease. This study was conducted to identify relevant biomarkers for both laparoscopic and serum diagnostics in ovarian cancer.

Bioinformatics analysis and expression screening in ovarian cancer cell lines were employed. Selected biomarkers were further validated in bio-specimens of diverse cancer types and ovarian cancer subtypes. For non-invasive detection, biomarker proteins were evaluated in serum samples from ovarian cancer patients.

Two kallikrein (KLK) serine protease family members (KLK6 and KLK7) were found to be significantly overexpressed relative to normal controls in most of the ovarian cancer cell lines examined. Overexpression of KLK6 and KLK7 mRNA was specific to ovarian cancer, in particular to serous and papillary serous subtypes. In situ hybridization and histopathology further confirmed significantly elevated levels of KLK6 and KLK7 mRNA and proteins in tissue epithelium and a lack of expression in neighboring stroma. Lastly, KLK6 and KLK7 protein levels were significantly elevated in serum samples from serous and papillary serous subtypes in the early stages of ovarian cancer, and therefore could potentially decrease the high “false negative” rates found in the same patients with the common ovarian cancer biomarkers human epididymis protein 4 (HE4) and cancer antigen 125 (CA-125).

KLK6 and KLK7 mRNA and protein overexpression is directly associated with early-stage ovarian tumors and can be measured in patient tissue and serum samples. Assays based on KLK6 and KLK7 expression may provide specific and sensitive information for early detection of ovarian cancer.

The online version of this article (doi:10.1186/s13048-014-0109-z) contains supplementary material, which is available to authorized users.

We measured the impact of changing KLK6 expression levels on the pathological grade of gliomas and on proliferation rate, cell cycle progression, and apoptosis in the U251 glioblastoma cell line.

The expression of KLK6 in 35 brain glioma tissues and adjacent noncancerous tissues was measured using real-time quantitative polymerase chain reaction (PCR) and the relationship between KLK6 expression and pathological grades was analysed.

The KLK6 expression in U251 cells was silenced by a specific siRNA, and the effects on proliferation, the cell cycle, and apoptosis were compared to wild type cells. Expression of KLK6 was downregulated in gliomas relative to matched noncancerous tissue. There was no obvious relationship between patient sex, pathological grade, or tumour classification and the expression of KLK6. In the U251 cell line, cell proliferation was enhanced and the fractions of cells in the G2 and S phases were increased by siRNA-mediated KLK6 silencing.

Expression of KLK6 inhibits tumour growth. Decreased KLK6 expression may be a possible risk factor for glioma.

• KLK6
• U251
• gliomas
• kallikrien
• siRNA

AIM: To analyze the expression of kallikrein gene 10 (KLK10) in gastric cancer and to determine whether KLK10 has independent prognostic value in gastric cancer.

METHODS: We studied KLK10 expression in 80 histologically confirmed gastric cancer samples using real-time quantitative reverse transcription-PCR and hK10 expression using immunohistochemistry. Correlations with clinicopathological variables (lymph node metastasis, depth of invasion and histology) and with outcomes (disease-free survival and overall survival) during a median follow-up period of 31 mo were assessed. Gastric cancer tissues were then classified as KLK10 positive or negative.

RESULTS: KLK10 was found to be highly expressed in 57/80 (70%) of gastric cancer samples, while its expression was very low in normal gastric tissues. Positive relationships between KLK10 expression and lymph node metastasis (P = 0.048), depth of invasion (P = 0.034) and histology (P = 0.015) were observed. Univariate survival analysis revealed that gastric cancer patients with positive KLK10 expression had an increased risk for relapse/metastasis and death (P = 0.005 and 0.002, respectively). Cox multivariate analysis indicated that KLK10 was an independent prognostic indicator of disease-free survival and overall survival in patients with gastric cancer.

CONCLUSION: KLK10 expression is an independent biomarker of unfavorable prognosis in patients with gastric cancer.

• Kallikrein gene 10
• Gastric cancer
• Survival analysis
• Prognostic biomarkers

• Adult
• Aged
• Biomarkers, Tumor/analysis
• Biomarkers, Tumor/genetics
• Chi-Square Distribution
• Disease-Free Survival
• Female
• Humans
• Immunohistochemistry
• Kallikreins/analysis
• Kallikreins/genetics
• Kaplan-Meier Estimate
• Lymphatic Metastasis
• Male
• Middle Aged
• Multivariate Analysis
• Neoplasm Invasiveness
• Predictive Value of Tests
• Proportional Hazards Models
• RNA, Messenger/analysis
• Real-Time Polymerase Chain Reaction
• Reverse Transcriptase Polymerase Chain Reaction
• Risk Factors
• Stomach Neoplasms/enzymology
• Stomach Neoplasms/genetics
• Stomach Neoplasms/mortality
• Stomach Neoplasms/pathology
• Stomach Neoplasms/therapy
• Time Factors
• Treatment Outcome
• Up-Regulation

Human kallikrein gene 6 (KLK6) is a member of the human kallikrein gene family (Kallikreins, KLKs). Human kallikrein-related peptidase 6 (hK6) is a trypsin-like serine protease encoded by the KLK6, has been reported to be highly expressed in several cancers including gastric cancer. In this study, we investigated the the correlation of hK6 expression with clinicopathological characteristics, tumor recurrence and prognosis in advanced gastric carcinoma after curative resection.

We retrospectively analyzed the clinical data of 129 cases advanced gastric cancer after curative gastrectomy. The expression of hK6 in advanced gastric cancer tissues compared to adjacent noncancerous tissues were examined, and the relationship between hK6 expression and clinicopathological characteristics was evaluated. In additional, these patients were followed up to investigate the relationship between hK6 expression and the survival time.

The positive rate of hK6 expression was significantly higher in advanced gastric cancer tissue, than that in adjacent noncancerous and gastric ulcer tissues (36.5%, 33.3%, respectively, P < 0.001). There was a close relationship between hK6 expression and TNM stage (P = 0.005), vascular invasion (P = 0.037) and perineural invasion (P = 0.035). Furthermore, patients with hK6 positive showed significantly higher recurrence and poorer prognosis than those with hK6 negative. Multivariate analysis showed that hK6 expression was a significant independent factor for tumor recurrence and overall survival.

hK6 is overexpressed in advanced gastric cancer tissues. Its clinical utility may be used as an unfavorable indicator in predicting tumor recurrence and prognosis for advanced gastric cancer after operation. This study also suggests that hK6 might be a potential therapeutic target for gastric cancer.

The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/8558403578787206

Despite the widespread use of conventional and contemporary methods to detect ovarian cancer development, ovarian cancer remains a common and commonly fatal gynecological malignancy. The identification and validation of early detection biomarkers highly specific to ovarian cancer, which would permit development of minimally invasive screening methods for detecting early onset of the disease, are urgently needed. Current practices for early detection of ovarian cancer include transvaginal ultrasonography, biomarker analysis, or a combination of both. In this paper we review recent research on novel and robust biomarkers for early detection of ovarian cancer and provide specific details on their contributions to tumorigenesis. Promising biomarkers for early detection of ovarian cancer include KLK6/7, GSTT1, PRSS8, FOLR1, ALDH1, and miRNAs.

Human kallikrein-related peptidase 6 (KLK6) has been implicated in various types of cancer and in neurodegenerative and demyelinating diseases including multiple sclerosis. Further, anti-KLK6 antibodies attenuated disease manifestations in the mouse model of multiple sclerosis. Availability of specific antibodies against KLK6 is fundamental to the development of improved diagnostic and/or immunotherapeutic applications. Here, we exploited the enhanced immunogenicity of mammalian proteins in avian species to generate a polyclonal antibody against KLK6.

Chicken were immunized with recombinant KLK6 and antibodies Y (IgYs) were purified from egg yolk with a simple procedure and evaluated for KLK6 detection by ELISA and Western blot using recombinant proteins and human cell lysates and supernatants. The anti-KLK6 Y polyclonal exhibited high affinity for KLK6 with a detection limit of 30 fmol. On the other hand, the widely used rabbit polyclonal antibody that was raised against the same recombinant KLK6 had a detection limit of 300 fmol. Moreover, the IgYs did not display any crossreactivity with recombinant KLKs or endogenous KLKs and other cellular proteins.

Based on its high specificity and sensitivity the developed anti-KLK6 IgY is expected to aid the development of improved diagnostic tools for the detection of KLK6 in biological and clinical samples.

• fish
• copy-number
• chromosomal instability
• structural rearrangement
• kallikrein locus
• ovarian carcinoma

• Chromosomes, Human, Pair 19
• Female
• Gene Dosage
• Genetic Heterogeneity
• Genomic Instability
• Humans
• Immunohistochemistry
• Kallikreins/genetics
• Ovarian Neoplasms/genetics

• R01 CA120197 NCI NIH HHS
• R01 CA120197-01 NCI NIH HHS
• R01 CA120197-02 NCI NIH HHS
• R01 CA120197-03 NCI NIH HHS

The human kallikrein-related peptidase family consists of 15 genes. Twelve of these genes are overexpressed in ovarian cancer and may represent potential markers for diagnosis, prognosis, and/or response to treatment. The aim of this study was to determine the prognostic significance of kallikrein-related peptidase 6 (KLK6) and kallikrein-related peptidase 13 (KLK13) in epithelial ovarian cancer by quantifying gene expression levels with tumour pathology and patient survival data.

Total RNA was isolated from 106 patients diagnosed with primary ovarian cancer, as well as 8 normal ovary controls. Samples were analysed by quantitative real-time PCR for KLK6 and KLK13 expression. Correlation between kallikrein gene expression and clinical characteristics was evaluated with the χ²-test. Survival analysis was performed using Kaplan–Meier and Cox proportional hazards regression models.

Expression levels of both KLK6 and KLK13 mRNA were significantly increased in invasive cancers relative to normal ovaries (P=0.002 and 0.039 respectively). High KLK6 and KLK13 expression was an indicator of poor prognosis, with patients having a shorter recurrence-free survival (P=0.002 and 0.027 respectively). High KLK6 expression was also significantly associated with lower overall survival (P=0.011). When subjected to multivariate analysis, patients with either high KLK6 or KLK13 were 3- and 2.2-fold, respectively, more likely to have a recurrence than patients with low kallikrein expression.

These data show increased mRNA expression of KLK6 and KLK13 in ovarian cancer compared to normal ovarian tissues. High KLK6 or KLK13 expression in primary ovarian tumours can significantly predict prognosis in terms of recurrence-free survival and overall survival. In all, this study shows KLK6 and KLK13 as potential biomarkers and may be therapeutic targets for treatment of ovarian cancer.