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Background: Kallikrein-related peptidases (KLKs) are a subgroup of serine proteases located on chromosome 19q13.3. Most KLKs have been extensively studied as potential biomarkers for several carcinomas and other diseases. KLK5 was originally identified from a keratinocyte library, and its enzyme was purified from the stratum corneum of human skin. KLK5 was shown to be differentially expressed in a variety of endocrine tumors, although it is not as yet examined widely in colorectal cancer (CRC). Methods: In this study, we quantitatively assessed the mRNA expression status of KLK5 in 197 colorectal tissues from 133 patients (70 cancerous and their paired normal colonic mucosa for 64 of them, as well as 63 colorectal adenomas) by quantitative real-time PCR (qPCR) using TaqMan probes. Statistical analysis evaluated the results. Results: It was shown that KLK5 expression is reduced following the histologically non-cancerous-adenoma sequence (p<0.001), whereas it is increased during the sequence adenoma-carcinoma (p<0.001). Furthermore, KLK5 positive expression is associated with positive nodal status (p=0.022), advanced tumor stage (p=0.038) and high histological grade (p=0.033). Cox univariate analysis revealed that KLK5 positive expression is associated with disease-free survival (DFS) (p=0.028) and overall survival (OS) of patients (p=0.048). Kaplan-Meyer survival models showed that patients with positive KLK5 expression have lower DFS (p=0.009) and OS (p=0.019). Receiver operating characteristic (ROC) analysis demonstrated for first time that KLK5 expression had significant discriminatory values between cancer and adenoma tissues (area under the curve [AUC] 0.77; 95% confidence interval [CI]=0.69–0.85, p=0.03). Conclusions: KLK5 mRNA expression may be useful for the differentiation of CRC from colorectal adenoma and represents a potential unfavorable prognostic biomarker for CRC. [ABSTRACT FROM AUTHOR]

Apoptosis is a type of programmed cell death involved in many crucial biological processes. It represents the basic mechanism for the action of chemotherapeutic agents, such as doxorubicin and carboplatin. Both are able to cause cell death through the induction of apoptosis in the human leukemic cell line HL-60. We investigated the possible alterations in the expression of apoptosis-related genes, including the novel BCL2L12 gene, which was recently cloned in our group. The kinetics of apoptosis induction and cell toxicity was investigated by DNA laddering and by the MTT method, respectively. Total RNA was extracted and cDNA was prepared by reverse transcription. BCL2 , BAX , FAS , caspase-9, caspase-3 and BCL2L12 were amplified by PCR. Overexpression of FAS , BCL2L12 and caspase-3 was observed after treatment of HL-60 cells for 3 or 6 h with carboplatin, while their expression was decreased after a 12-h treatment, demonstrating that these genes may take part in the early stages of apoptosis. Overexpression of the same genes was also observed after 6 h of treatment with doxorubicin (concomitantly with DNA laddering). In the case of carboplatin-induced apoptosis we detected down-regulation of BAX , BCL2 and caspase-9, whereas in the case of doxorubicin, BAX and BCL2 remained at control levels and caspase-9 was increased. [ABSTRACT FROM AUTHOR]

The anti-apoptotic molecule BCL2 delays cell-cycle entry from quiescence. We have recently cloned a new member of the BCL2 family of apoptosis-related genes, BCL2L12. In the present study, the expression of BCL2 and BCL2L12 genes during cisplatin-induced apoptosis in HL-60 leukemic cells was investigated. The kinetics of apoptosis induction and cell toxicity were evaluated by DNA laddering and the MTT method, respectively. BCL2 and BCL2L12 expression was analyzed by RT-PCR using gene-specific primers. The ratio of apoptotic cells increased with increasing concentrations of cisplatin and exposure time of cell culture to the drug. Gradual, time-dependent downregulation of BCL2 gene was observed during cisplatin treatment. Up-regulation of BCL2L12 was observed 3 h (no DNA fragmentation) and 6 h (initiation of the DNA fragmentation) after treatment with cisplatin, followed by a decrease of its expression after 12-h continuous treatment with the drug. It is known that the main anti-carcinogenic effect of cisplatin is due to the induction of cell apoptosis. Present results indicate that downregulation of the BCL2 and upregulation of the BCL2L12 gene may be the underlying mechanisms. [ABSTRACT FROM AUTHOR]

The article discusses the 2nd International Symposium on Kallikreins and Kallikrein-Related Peptidases Symposium. The symposium was held by the Hellenic Anticancer Institute and organized by president Litsa Talieri and Judith Clements, chair of the scientific advisory board. Research on structural features and functional roles of kallikrein was presented by 135 participants. Clements was awarded for her contributions to the identification of the expanded tissue kallikrein gene locus.

Currently available colon cancer (CC) markers lack sensitivity and specificity. Kallikrein-related peptidases (KLKs) present a new class of biomarkers under investigation for diverse diseases, including cancer. KLKs are co-expressed in various tissues participating in proteolytic cascades. KLK7 in human tumours facilitates metastasis by degrading components of the extracellular matrix. KLK14 promotes tumourigenesis by activating proteinase-activated receptors. In the present study we examined the concomitant expression of KLK7 and KLK14 in245 colonic tissue specimens from 175 patients; 70 were pairs of cancerous-normal tissues, 31 were cancerous tissues and 74 were colonic adenomas. We used quantitative real-time PCR and proved that both genes are up-regulated in CC at the mRNA level. Receiver-operating characteristic (ROC) analysis of our results showed that both genes have discriminatory value between CC and adenoma tissues, with KLK14 obtaining greater distinguishing power (area under the curve [AUC]=0.708 for KLK14; AUC=0.669 for KLK7). Current work showed that the two genes are fairly co-expressed in all three types of colon tissues examined (normal rs=0.667, p<0.001, adenomas rs=0.373, p=0.001, carcinomas rs=0.478, p<0.001). KLK14 is associated with shorter disease-free survival (DFS) and overall survival (OS) of patients (p=0.003, p=0.016 respectively), whereas KLK7only with shorter DFS (p=0.004). KLK7 and KLK14 gene expression can be regarded as markers of poor prognosis for CC patients with discriminating power between CC and adenoma patients.

Background: Kallikrein-related peptidases (KLKs) have been proposed as potential cancer biomarkers. Contradictions in literature led us to clarify the role of KLK5 as a breast cancer predictor, as well as its association with KLK7 expression., Patients and Methods: Semi-quantitative RT-PCR detected KLKs 5 and 7 in 80 breast tissues, 74 neoplastic and 6 normal. Steroid hormone receptors were quantified in all samples. Associations between KLK5 status and clinicopathological variables, as well as disease-free survival (DFS) and overall survival (OS) of patients were analyzed., Results: Forty tumor tissues showed high KLK5 expression, which was significantly associated with estrogen receptor status. Significant co-expression of KLKs 5 and 7 was observed in the same cancer samples (p=0.02). Increased KLK5 expression was a statistically significant independent prognostic factor for DFS (p=0.009 univariate analysis and p=0.028 multivariate analysis) and OS of patients (p=0.014, univariate analysis)., Conclusion: Increased KLK5 expression can contribute to the prognosis of DFS and OS of breast cancer patients. KLKs 5 and 7 are co-expressed in breast cancer.

Human tissue kallikrein-related peptidases are a family of 15 secreted serine proteases, located at chromosome 19q13.4. Most of them have been reported to be potential biomarkers for several carcinomas and other diseases. Human tissue kallikrein-related peptidase 7 (KLK7) has been purified from human stratum corneum and resembles a chymotryptic endopeptidase originally called stratum corneum chymotryptic enzyme (SCCE). In this study, we examined for the first time, the prognostic value of KLK7 mRNA expression, using a semi-quantitative RT-PCR method, in 105 colorectal cancer tissues for 54 of which, paired normal colonic mucosa were available. Furthermore, we analysed the expression of KLK7 in 10 adenomas, in 18 biopsies of inflamed colon mucosa, as well as in 22 human cancer cell lines of various origin, four of them being of colon. A defined number of colon cancer samples were also examined by immunohistochemistry. KLK7 expression was higher in cancerous than in normal tissues. Less differentiated tumors of more advanced stage showed higher KLK7 expression. Follow-up analysis revealed that KLK7 was significantly associated with shorter overall survival (OS) and disease-free survival (DFS). In addition, selected colon cancer samples highly expressing KLK7 gene, showed intense immunohistochemical staining for KLK7, enhancing RT-PCR results. Present data suggest that KLK7 gene is up-regulated in colon cancer and its expression predicts poor prognosis for colon cancer patients.

Background: In this study, the possible relation of the expression pattern of arginine methyltransferase 1 and colon cancer progression is investigated., Materials and Methods: Colon cancer samples as well as normal colon samples were used to define the arginine methyltransferase 1 expression by RT-PCR. The results were associated with clinical and histological parameters of the tissues., Results: In colon cancer tissue, only PRMT1 variants v1 and v2 were often expressed. Statistical significance for the clinicopathological parameters examined was found only for PRMT1 variant v2. PRMT1 v2 expression was associated with nodal status and tumour grade. PRMT1 v2 expression analysis in 25 pairs of cancerous/non-cancerous colon tissue showed higher or equal expression in cancer versus normal tissue. In 18 inflamed colon tissues examined for PRMT1 expression and compared with the expression of 90 colon cancer tissue samples, statistical significance was found only for variants v1 and v2. A higher percentage of PRMT1 v2 expression was observed in older patients., Conclusion: From the present preliminary results, it can be said that PRMT1 variant v2 can probably be regarded as a marker of unfavourable prognosis in colon cancer patients.

Several of the novel kallikrein-related peptidases (tissue kallikreins; KLKs) are emerging new serum and/or tissue biomarkers for prostate cancer (CaP) diagnosis, prognosis and monitoring. In the present research approach, our objective was to investigate the possible alterations in the mRNA expression levels of KLK5 and KLK11 genes in prostate cancer cells PC3 as a response to treatment with mitoxantrone, etoposide, doxorubicin and carboplatin. Viability was assessed with the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay after cell treatment with either mitoxantrone (2 microM), etoposide (20 microM), doxorubicin (1 microM), or carboplatin (15 microM), for 24, 48 and 72 hours. Additionally, trypan blue staining revealed that in PC3 cells all drugs displayed almost the same limited necrotic effects which appeared mainly at 72 hours of treatment. PC3 prostate cancer cells showed a concentration- and time-dependent increased cytotoxicity to the drugs under study which was mainly due to reduction of cell proliferation efficiency. Distinct modulations of KLK5 and KLK11 genes, at the mRNA level, were observed, supporting a drug-dependent cell response. Our experimental data demonstrate that the molecular profile mainly of KLK5 gene may serve as a new potential molecular biomarker predicting treatment response in CaP cells.

Background: The human tissue kallikrein gene family (KLK1 to KLK15) encodes a group of 15 serine proteases (hK1 to hK15), several of which have been implicated in cancer-related processes., Methods: We established a specific quantitative reverse transcription-PCR assay for full-length KLK7 mRNA that excluded amplification of the exon 2 deletion splice variant (the latter does not encode a functional protease), and evaluated full-length KLK7 mRNA expression [normalized to human glucose-6-phosphate dehydrogenase (h-G6PDH)] in tumor tissue specimens from 155 breast cancer patients., Results: High KLK7 mRNA expression (continuous) was significantly associated with a better patient outcome according to both univariate (P = 0.005) and multivariate (P = 0.046) Cox survival analysis. Separation of patients by optimized dichotomization revealed a significantly better prognosis for patients with high KLK7 mRNA status (n = 89) compared with patients with low KLK7 mRNA status (n = 66) [univariate hazard ratio (HR) = 0.45 (P = 0.001); multivariate HR = 0.50 (P = 0.005)]. In the subgroup of patients not receiving adjuvant treatment (n = 69), KLK7 mRNA status was a significant prognosticator [univariate HR = 0.29 (P = 0.002); multivariate HR = 0.40 (P = 0.034)]. This subgroup was least influenced by postoperative treatment and thus best showed the impact of KLK7 expression on the natural course of breast cancer disease., Conclusion: Expression of full-length KLK7 mRNA may represent a new prognostic marker in breast cancer disease.