Differential Expression of the KLK2 and KLK3 Genes in Peripheral Blood and Tissues Samples of Iranian Patients with Prostate Cancer (original) (raw)
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Genetics and Molecular Biology, 2006
We used the multiplex semi-quantitative reverse-transcriptase PCR (RT-PCR) to investigate kallikrein 2 and 3 (KLK2 and KLK3) mRNA levels in prostate tissue from 42 prostate cancer patients, 33 of whom were also assessed for peripheral blood KLK2 expression by qualitative semi-nested RT-PCR. We found that KLK2 was an important tissue biomarker for distinguishing between prostate cancer patients and those with benign prostatic hyperplasia, particularly when KLK2 expression was > 60% of that of the β2-microglobulin constitutive gene. Patients with an average relative expression value ≥ 0.6 (cutoff value) had an eleven-fold higher chance of having prostate cancer. When one or two tissues samples were evaluated for KLK2 expression using the cutoff value the estimated chance of having prostate cancer was increased by seven times for one positive sample and 45 times for two positive samples. There was no significant correlation between KLK3 gene expression and prostate cancer diagnosis. Logistic regression for blood and tissue KLK2 expression successfully detected 92% of the prostate cancer cases. The detection of KLK2 in blood showed a sensitivity of 59% and a specificity of 82%. This study indicates that the KLK2 gene may be a useful molecular marker for the diagnosis of prostate cancer and that analysis of KLK2 expression in blood and tissues could provide a novel approach for the clinical investigation of this type of cancer.
The Prostate, 2003
BACKGROUND. Many members of the human kallikrein gene family are differentially expressed in cancer and a few have potential as diagnostic/prognostic markers. KLK14 is a newly discovered human kallikrein gene that is mainly expressed in the central nervous system and endocrine tissues. Since KLK14 was found to be regulated by steroid hormones in prostate cancer cell lines, we hypothesized that it will be differentially expressed in prostate cancer tissues compared to their normal counterparts. METHODS. Matched prostate tissue samples from the cancerous and non-cancerous parts of the same prostates were obtained from 100 patients who underwent radical prostatectomy. Quantitative analysis of KLK14 expression levels were performed by real-time RT-PCR using SYBR Green I dye on the LightCycler TM system. Associations with clinico-pathological parameters were analyzed. RESULTS. KLK14 overexpression in the cancerous compared to non-cancerous tissue was found in 74% of patients (P < 0.001). Mean level of expression was 154 arbitrary units (Au) in cancerous tissues and 14.2 Au in the non-cancerous tissues. The ratio of the cancerous to noncancerous KLK14 expression values was higher in patients with late stage (stage III) compared to stage II (P ¼ 0.002), and in grade 3 compared to grade 1/2 tumors (P ¼ 0.001). A statistically significant increase was also observed in patients with higher in Gleason score (>6) compared to Gleason score ¼ 6 tumors (P ¼ 0.027). No correlation was found between KLK14 tissue expression levels and serum prostate-specific antigen. CONCLUSIONS. KLK14 expression is significantly higher in cancerous compared to noncancerous prostatic tissue. The up-regulation of the KLK14 gene in advanced and more aggressive tumors may indicate a possible role for the hK14 protein in tumor spread and opens the possibility of hK14 being a candidate new marker for prostate cancer diagnosis and prognosis.
The Prostate, 2008
BACKGROUND. The kallikrein-related (KLK) serine protease, prostate specific antigen is the current marker for prostate cancer (PCa). Other members of the KLK family are also emerging as potential adjunct biomarkers for this disease. Our aim was to identify and characterize novel KLK-related genes with potential as PCa bio-markers. METHODS. Low stringency DNA screening was coupled with amplification techniques to identify novel sequences. Transcripts were examined by Northern blot, RT-PCR, and in situ hybridization analysis and in silico bioinformatics approaches. Protein characterization was performed by Western blot and confocal microscopy analysis. Gene regulation studies were performed by quantitative PCR and promoter reporter assays. RESULTS. We identified a novel kallikrein-related mRNA designated KRIP1 (kallikreinrelated, expressed in prostate 1) which, together with the recently reported CKLK1 and KLK31P transcripts, is transcribed from KLKP1; a gene evolved from, and located within, the KLK locus. Significantly, in contrast to these other non-coding KLKP1 transcripts, the KRIP1 mRNA generates an $18 kDa intracellular protein-the first non-serine protease identified from the KLK locus. KRIP1 mRNA is abundant only in normal prostate and is restricted to cells of epithelial origin in normal and diseased glands. Ligand binding of the androgen receptor increases transcription from the KLKP1 gene. Consistently, KRIP1 mRNA levels are lower in PCa samples compared to benign prostatic hyperplasia. CONCLUSIONS. Transcription from KLKP1 is reduced as cells de-differentiate on the pathway to malignancy. KLKP1/KRIP1 has potential as a marker of both PCa progression and recent evolutionary events within the KLK locus.
Prostate Cancer and Prostatic Diseases, 2003
Human kallikreins 6, 10 and 13 (hK6, hK10 and hK13) are expressed by many normal, mainly glandular tissues, including prostatic epithelium. Some kallikreins may function as tumor suppressors or are downregulated during cancer progression. The aim of this study was to evaluate the immunoexpression of these kallikreins in benign and malignant prostatic tissues and correlate their expression with prostate cancer (PC) prognosis. Included in the study were 25 cases of nonmalignant prostate and 179 cases of PC. Among them, 122 PC cases were immunostained for hK6, 94 for hK10 and 113 for hK13, respectively. The follow-up period for a subset of 68 patients who had undergone radical prostatectomy (RP) was 1-58 months (mean=13.4 +/- 1.7 and median=8.0 months). A cutoff value of 0.2 microg/l of serum PSA was established as a biochemical recurrence threshold. Follow-up information was available for 26/55 RP cases stained for hK6, 14/32 cases stained for hK10 and 25/59 cases stained for hK13. Gleason score (GS) 7 carcinomas were stratified as 7a and 7b, according to the primary grade. PC with GS 2-7a were histologically categorized as low malignant (LM) and PC with GS 7b-10 as high malignant (HM). The immunohistochemical method of streptavidin-biotin-peroxidase using monoclonal and polyclonal antibodies was performed. In the benign prostate and in prostatic intraepithelial neoplasia, a cytoplasmic immunostaining of varying intensity was evident. In PC, the immunoexpression of all kallikreins was decreased: 102/122 cases (84%) were positive for hK6, 73/94 (78%) for hK10 and 97/113 (86%) for hK13, respectively. A statistically significant difference in expression was found, in comparison to nonmalignant prostates (P=0.029, 0.009 and 0.045, respectively). Also, a positive correlation was observed between the immunoexpression of these three kallikreins. Concerning the histological grade, HM-PC expressed all three kallikreins with a slightly higher percentage than LM-PC: 79 vs 88% for hK6, 76 vs 79% for hK10 and 76 vs 92% for hK13. These differences were statistically significant only in the case of hK13 (P=0.024). Serum PSA did not correlate with kallikrein immunoexpression in PC. Furthermore, there was no significant correlation between kallikrein expression and pathological stage or recurrence, in the cases of RP. All three kallikreins are expressed in the nonmalignant and malignant prostate, with cancer tissues demonstrating slightly lower expression. Expression levels did not correlate with aggressiveness and they do not seem to have value for prostate cancer prognosis.
Prostate-specific antigen (PSA, hK3) is a diagnostic marker for prostatic cancer but lacks the specificity to sufficiently distinguish between prostatic cancer and benign prostatic hyperplasia (BPH). Human glandular kallikrein 2 (hK2) has been proposed as a potential diagnostic marker for prostate cancer that could complement the current PSA test. Recently we demonstrated that proPSA is present in prostate cancer sera. This study examines the expression of prohK2 in prostate cells and its presence in human sera. Western blot analysis was used to assess prohK2 expression in the human carcinoma cell line, LNCaP. A highly specific and sensitive dual monoclonal immunoassay for prohK2 was developed and used to assess the presence of prohK2 in human sera. prohK2 was detected in the spent media of LNCaP cells. Furthermore, prohK2 was present at immunodetectable concentrations in human sera, and its concentration was increased in prostatic cancer and BPH. These results indicate for the first time that prohK2 is secreted by human prostate cells and is a major component of uncomplexed (free) hK2 in human sera. In addition, prohK2 in human sera is associated with prostate disease and thus may be a useful marker for prostatic cancer and BPH.
Kallikrein gene family as biomarkers for recurrent prostate cancer
Croatian Medical Journal, 2020
AimTo assess kallikrein (KLK) expression in recurrent and non-recurrent prostate tumors and adjacent healthy prostate tissues.MethodsThe expression levels of 15 KLK genes in 34 recurrent and 36 non-recurrent prostate cancer samples and 19 adjacent healthy prostate tissue samples was assessed with quantitative reverse-transcription polymerase chain reaction. The samples were obtained from Baylor College of Medicine, Houston, TX, USA between 2013 and 2016 .ResultsCompared with controls, prostate cancer samples showed a strong decrease in KLK1, KLK4, KLK9, and KLK14. Recurrent samples were negative for KLK1, KLK2, and KLK14 but demonstrated higher levels of KLK3, KLK4, and KLK9 than controls. Other KLKs were not significantly expressed.ConclusionThis study for the first time showed a difference in the expression levels of the KLK gene family in recurrent prostate cancer. KLKs could be used as recurrence markers for prostate cancer.
Allelic Variants of KLK2 Gene Predict Presence of Prostate Cancer at Biopsy
Journal of Cancer Diagnosis, 2016
Objective: Several single nucleotide polymorphisms associated with prostate cancer risk have been reported in recent years. We evaluated polymorphisms in the human glandular kallikrein 2 (KLK2) genes because the protein product of this gene is known to be increased in prostate cancer. Materials and methods: Blood samples were collected from sixty patients who underwent prostate biopsy sectioning, and from their genomic DNA the SNPs in KLK2 gene were investigated by direct DNA sequencing. Another 138 archived prostate tissue sections were also evaluated using the TaqMan SNP genotyping assay. Results: Eighteen known SNPs were identified in the KLK2 gene. The SNPs were located in introns, coding exons and untranslated regions of the gene. Further analysis showed that two of the SNPs were associated with prostate disease. The T/T allele of rs198977 was significantly predictive of the presence of prostate cancer at biopsy and was also associated with high tumour grade. The A/A allele of rs2664155 was also significantly associated with the presence of benign hyperplasia at biopsy. Conclusion: Our results support previous reports of association of the rs198977 SNP with prostate cancer risk and also indicated a link with the disease phenotype. However, the second SNP (rs2664155) was more associated with benign hyperplasia than prostate cancer risk. The method of TaqMan SNP genotyping could be clinically useful in genetic screening and risk stratification of patients for prostate diseases.
We present a multiplexed and internally calibrated quantitative reverse transcription-PCR (QRT-PCR) assay to detect human glandular kallikrein 2 (hK2) and prostate-specific antigen (PSA) transcripts in blood samples from healthy subjects and prostate cancer (PC) patients. The assay detected 50 copies of hK2 and PSA mRNA, and 1 PSA-and 10 hK2-expressing LNCaP cells in the presence of 2.5 ؋ 10 6 PSA-and hK2negative cells. In PC patients, 20 of 25 and 19 of 25 gave detectable PSA and hK2 mRNAs, respectively. Number of hK2 mRNA copies was significantly higher than that of PSA mRNA copies in patients with biochemically progressive (P ؍ 0.02) PC, and with locally advanced and metastasized (P ؍ 0.004) PC. Patients with rapidly progressive and hormone refractory PC gave detectable hK2 mRNA only in 2 of 8 and PSA mRNA in 3 of 8 patients. Neither PSA nor hK2 mRNAs were detected in 16 healthy subjects. PSA and hK2 discriminated PC patients with biochemically progressive and advanced disease from the controls and from the aggressive distant metastatic disease. The assay provides a reliable quantification of the number of hK2 and PSA mRNA copies, allows to discriminate PC cases from healthy subjects, and offers a tool for further studies on molecular staging of PC.
Urology, 1997
Objectives. We describe the expression of a potentially new tumor marker, human glandular kallikrein 2 (hK2), that may be useful as an adjunct to prostate-specific antigen (PSA) in the diagnosis and monitoring of prostate cancer. Methods. We evaluated 257 radical prostatectomy specimens removed at the Mayo Clinic with pathologic Stage T2 adenocarcinoma to compare the cytoplasmic expression of hK2, PSA, and prostatic acid phosphatase (PAP) in benign tissue, high-grade prostatic intraepithelial neoplasia (PIN), and adenocarcinoma. Two monoclonal antibodies, hK2-A523 and hK2-G586, specific for hK2 were used, as well as antibodies against PSA (PSM-773) and PAP (polyclonal). Results. Intense epithelial cytoplasmic immunoreactivity was observed in every case for hK2-A523, hK2-G586, PSA, and PAP (100% of cases, respectively). The intensity and extent of hK2 expression for both antibodies were greater in cancer than high-grade PIN; furthermore, high-grade PIN was greater than benign epithelium. Cases of Gleason primary grade 4 and 5 cancer showed hK2 staining in almost every cell, whereas there was greater heterogeneity of staining in lower grades of cancer. In marked contrast to hK2, PSA and PAP immunoreactivity was most intense in benign epithelium and stained to a lesser extent in PIN and carcinoma. The number of immunoreactive cells for hK2 and PSA was not predictive of cancer recurrence. Conclusions. hK2 was expressed in every cancer, and the expression incrementally increased from benign epithelium to high-grade PIN and adenocarcinoma. PSA and PAP displayed inverse immunoreactivity compared with hK2. The expression of hK2 and PSA was not predictive of cancer recurrence in patients with Stage T2 carcinoma. Expression of hK2 indicates that this kallikrein antigen is both prostate localized and tumor associated. Tissue expression of hK2 appears to be regulated independently of PSA and PAP. Further studies are needed to determine whether tissue immunoreactjvity of hK2 will prove clinically useful in the diagnosis and monitoring of prostate cancer. UROLOGY 49: 857-862, 1997. © 1997, Elsevier Science Inc. All rights reserved. p rostate-specific antigen (PSA) (human glandular kallikrein 3 [hK3]) is the most important tumor marker for prostate cancer. It is useful for diagnosis and for monitoring progression but is limited by modest levels of sensitivity and speci-