Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Background: Preoperative staging for prostate cancer underestimates the final pathology stage in approximately 40-50% of the cases. Previous work from our institution demonstrated that an enhanced reverse transcriptase polymerase chain reaction (RT-PCR) assay for prostate-specific antigen (PSA) enabled more accurate staging of presumably localized prostate cancer. The goal of the current study is to determine if needle biopsy results when combined with the RT-PCR for PSA assay are a better predictor of final pathology stage. Methods and Results: One hundred sixty-two men with needle biopsy-diagnosed prostate cancer had blood drawn for the RT-PCR for PSA assay before undergoing radical prostatectomy. Polymerase chain reaction primers specific for the PSA gene were run, along with appropriate controls. Tumor was characterized using the TMN staging system: organ confined (pT2a-c), capsular penetration (pT2a-b), seminal vesicle involvement (pT3c). Surgical margins and lymph nodes were also evaluated. Of the 162 patients, the majority had localized disease by digital rectal examination: T2 = 97%, and T3 = 3%. On needle biopsy, 48 cases (30%) had a Gleason score >/=7 and 35 cases (22%) had perineural involvement (PNI). The RT-PCR for PSA assay was positive in 50 patients (31%). Final pathology revealed 39% of patients had pT3 disease; none of the 162 patients had lymph node involvement. Statistical analysis revealed that a Gleason score >/=7 had 81% specificity and 46% sensitivity in predicting pT3 disease (odds ratio 3.6). The presence of PNI on needle biopsy was 89% specific and 38% sensitive in predicting pT3 disease (odds ratio, 4.9). The RT-PCR for PSA assay was 89% specific and 62% sensitive in predicting pT3 disease (odds ratio, 13.0). All 14 cases with both RT-PCR for PSA and PNI positivity had pT3 disease. Logistic regression analysis demonstrated the independent predictive strength of PNI on needle biopsy, Gleason score >/=7, and RT-PCR for PSA positivity for identifying pT3 disease; their combined odds ratio was more than 180. Conclusions: Using the RT-PCR for PSA assay in conjunction with needle biopsy results increases the predictive strength for pT3 disease in patients with presumed organ-confined prostate carcinoma.
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PMID:Enhanced Reverse Transcriptase Polymerase Chain Reaction for Prostate-specific Antigen Combined With Needle Biopsy Results: A Superior Predictor of pT3 Disease. 1046 1

The activity and expression of transgene beta-galactosidase (lacZ) by replication-deficient adenoviral vectors (Ad-lacZ) containing prostate-specific promoters were compared using an in vivo canine model. The prostate tissue-specific promoters were prostate-specific antigen, probasin, and mouse mammary tumor virus long-terminal repeat, which were fused separately to an Escherichia coli lacZ gene. Dogs underwent laparotomy, and adenoviral vectors were delivered by direct intraprostatic injection. At 72 hours postinjection, the prostate and various other organs were harvested to evaluate the degree of prostate expression and dissemination of adenoviral vectors. Expression of lacZ in tissues was determined by 5-bromo-4-chloro-3-indolyl beta-D-galactoside staining, beta-galactosidase assay, and E. coli lacZ reverse transcriptase-polymerase chain reaction (PCR). The presence of adenoviral DNA sequences in canine tissues was determined by PCR using primers specific for the type 5 adenoviral genome. All three of the prostate-specific adenoviruses tested effectively expressed the lacZ gene in the canine prostate, but expression levels were lower than that of the control viral vector AdRSVlacZ following intraprostatic injection. By PCR, adenoviral vector DNA was detected in other organs and tissues, including the bladder and vas deferens. However, reverse transcriptase-PCR analysis revealed that prostate-specific Ad-lacZ vectors only transcribed lacZ mRNA in the prostate and not in nonprostatic tissues. Thus, these novel prostate-specific adenoviral vectors each have equal in vivo expression exclusively in the prostate and may potentially be used for prostate cancer gene therapy.
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PMID:In vivo expression of prostate-specific adenoviral vectors in a canine model. 1050 56

Early systemic spread of occult tumor cells that may develop into founders of incurable distant metastasis has been identified in prostate cancer patients by reverse transcription-PCR (RT-PCR) amplification of prostate-specific antigen (PSA) mRNA. Nevertheless, the introduction of this new staging tool into the clinical setting has been hampered by the disparate and contradictory data on the sensitivity and specificity of RT-PCR methods reported recently. We used PSA RT-PCR to examine the influence of analytical variables such as priming and enzyme of reverse transcriptase reaction, temperature and time of primer annealing, primer extension and denaturation, as well as the concentrations of magnesium chloride, Taq polymerase, deoxynucleotide triphosphate, primers and BSA on the amplification process. By systematically varying these chemical and physical components, we could demonstrate a significant increase in amplification yield and in stringency of primer annealing. This may explain the wide variety of published findings on molecular staging of prostate cancer, which currently impedes the clinical introduction of PSA RT-PCR assays in prostate cancer. Methodological analyses are needed for standardization and quality assurance to achieve reproducible molecular methods that can be used in clinical practice.
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PMID:Analytical variables of reverse transcription-polymerase chain reaction-based detection of disseminated prostate cancer cells. 1091 19

Trypsinogen is a serine proteinase produced mainly by the pancreas, but it has recently been found to be expressed also in several cancers such as ovarian and colon cancer and in vascular endothelial cells. In this study, we found that trypsinogen-1 and -2 are present at high concentrations (median levels, 0.4 and 0.5 mg/L, respectively) in human seminal fluid and purified them to homogeneity by immunoaffinity and anion exchange chromatography. Purified trypsinogen isoenzymes displayed a M(r) of 25 to 28 kd in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Most of the trypsinogen-1 purified from seminal fluid was enzymatically active whereas trypsinogen-2 occurred as the proform, which could be activated by enteropeptidase in vitro. Immunohistochemically, trypsinogen protein was detected in the human prostate, urethra, utriculus, ejaculatory duct, seminal vesicles, deferent duct, epididymal glands, and testis. Expression of trypsinogen mRNA in the same organs was demonstrated by in situ hybridization. Trypsinogen mRNA was also detected in the prostate and seminal vesicles by reverse transcriptase-polymerase chain reaction and Northern blotting. Isolated trypsin was shown to activate the proenzyme form of prostate-specific antigen. These results suggest that trypsinogen isoenzymes found in seminal fluid are produced locally in the male genital tract and that they may play a physiological role in the semen.
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PMID:Expression and characterization of trypsinogen produced in the human male genital tract. 1110 74

Circulating prostate cells can be detected in peripheral blood of patients with clinically localized or advanced prostate carcinoma. Traditionally, nested reverse transcriptase-polymerase chain reaction (RT-PCR) is used for this as a sensitive, but qualitative only, detection system. We developed a quantitative real-time RT-PCR method for measuring prostate-specific antigen (PSA) mRNA in peripheral blood of prostate cancer patients. A quantitative assay was developed using an external standard reference curve generated with RNA from the human prostate cell line LNCaP. Basal blood samples were collected from 44 patients without evidence of distant metastases and from 30 healthy controls. In 29 patients surgically treated with radical prostatectomy, the measurement of PSA mRNA was performed in blood samples collected before, at the end and 6 days after surgery. In 14 patients treated with radiotherapy, the measurements were repeated at 3-month intervals to evaluate time-related changes during therapy. The measurements were also performed for one year at 3-month intervals in one patient treated with anti-androgen therapy. We found detectable PSA mRNA in 14/44 (32%) basal blood samples. A wide range of values were observed in these patients, ranging from 0.5 to 1724 pg of total LNCaP RNA/ml blood. In patients undergoing radical prostatectomy, circulating PSA mRNA was detectable in eight patients in basal samples, and in seven of them also in blood specimens collected at the end of surgery, showing an increase in only two patients. In blood samples collected 6 days later, PSA mRNA was dramatically reduced in all patients, but still present in seven of them. In four patients, whose basal samples were negative, PSA mRNA was detectable in samples collected at the end of surgery and three of them were negative after 6 days. In patients who did not receive surgical treatment, a rapid decrease in PSA mRNA was demonstrated in five patients treated with radiotherapy and in one patient undergoing androgen deprivation. No detectable PSA mRNA was found in healthy controls. The levels of PSA mRNA in peripheral blood from patients with prostate carcinoma can be easily measured by this sensitive, quantitative and reliable procedure. This assay is a promising tool for the detection and follow-up of these patients.
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PMID:Real-Time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) for the measurement of prostate-specific antigen mRNA in the peripheral blood of patients with prostate carcinoma using the taqman detection system. 1143 86

We report the clinical evolution of a prostate cancer, metastasizing to lungs and bones, recurring locally, and escaping from anti-androgen therapy. Key event of biological progression of the patient's tumor was the coincidence of allelic imbalance accumulation and of bone metastases occurrence. The recurrent tumor was established as the transplantable xenograft PAC120 in nude mice, where it grew locally. PAC120 displayed the same immunophenotype of the original tumor (positive for keratin, vimentin, prostatic acid phosphatase, and Leu-7) and expressed human HOXB9, HOXA4, HER-2/neu, and prostate-specific antigen genes, as detected by reverse transcriptase-polymerase chain reaction. It formed lung micrometastases detected by mRNA expression of human genes. Cytogenetic analysis demonstrated numerous alterations reflecting the tumor evolution. PAC120 was still hormone-dependent; its growth was strongly inhibited by the new gonadotropin-releasing hormone antagonist FE 200486 but weakly by gonadotropin-releasing hormone superagonist D-Trp(6)-luteinizing-hormone releasing hormone (decapeptyl). Tumor growth inhibition induced by anti-hormone therapy was linked to the hormone deprivation degree, more important and more stable with FE 200486 than with D-Trp(6)-luteinizing-hormone releasing hormone. Surgical castration of mice led to tumor regressions but did not prevent late recurrences. Transition to hormone-independent tumors was frequently associated with a mucoid differentiation or with a neuroendocrine-like pattern. Independent variations of mRNA expression of HER-2/neu and prostate-specific antigen were observed in hormone-independent tumors whereas HOXB9 gene expression was constant. In conclusion, PAC120 xenograft, a new model of hormone-dependent prostate cancer retained the progression potential of the original tumor, opening the opportunity to study the hormone dependence escape mechanism.
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PMID:Clinical and experimental progression of a new model of human prostate cancer and therapeutic approach. 1148 33

Prostate androgen regulated transcript 1 (PART-1), is a gene predominantly expressed in the prostate gland and is regulated by androgens in human prostate cancer cell lines. Here, we report additional characteristics of PART-1 tissue expression and hormonal regulation and study its expression profile in human normal and matched prostate cancer tissues. Since PART-1 shows similarity to prostate-specific antigen (PSA) in prostate specificity and regulation, we hypothesized that it may be implicated in prostate carcinogenesis or may be a potential new biomarker. We used reverse transcriptase polymerase chain reaction (RT-PCR) to further characterize PART-1 tissue expression and hormonal regulation in the LNCaP prostate cancer cell line. RT-PCR analysis revealed that PART-1 is expressed not only in the prostate and salivary gland, but also in other tissues, including the thymus and placenta. In addition to androgen stimulation, PART-1 is also up-regulated by progestins, oestrogens and glucocorticoids. We further studied the expression of PART-1 in 27 paired (from the same patient) cancerous and non-cancerous prostatic tissues, with qualitative and quantitative RT-PCR (LightCycler technology), in order to examine whether PART-1 is overexpressed or underexpressed in cancer. Our results indicated that PART-1 is more frequently overexpressed in the cancerous prostatic tissue. We conclude that this gene is overexpressed in prostate cancer and may represent a novel prostate cancer tumour marker.
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PMID:Expression and regulation of prostate androgen regulated transcript-1 (PART-1) and identification of differential expression in prostatic cancer. 1148 71

The efficacy of therapy with targeted cytotoxic luteinizing hormone-releasing hormone (LHRH) analog AN-207 consisting of superactive doxorubicin derivative AN-201 linked to carrier [D-Lys(6)]LH-RH was evaluated in vivo in nude mice bearing xenografts of MDA-PCa-2b prostate cancer line. AN-207 was administered intravenously (i.v.) at 200 nmol/kg on day 1 and at 150 nmol/kg on day 14. After 4 weeks of treatment with AN-207, tumor growth was inhibited as shown by a 63% (P<0.01) decrease in tumor volume and a 55% (P<0.05) reduction in tumor weight, compared with controls. None of the animals died after administration of AN-207 at the total dose of 350 nmol/kg, and at the end of the experiment the body weights of mice given AN-207 did not differ significantly from controls. A single injection of cytotoxic radical AN-201 at 200 nmol/kg resulted in 43% mortality. In the surviving mice, AN-201 caused a 50% inhibition in tumor volume and a 27% reduction in tumor weight, which were non-significant, as compared to the controls. After 4 weeks, serum prostate-specific antigen concentrations in mice treated with AN-207 were 65% lower than those in controls (P<0.05), while in animals given AN-201 the reduction in serum prostate-specific antigen was only 40% (NS). The expression of mRNA for LHRH receptors was detected by reverse transcriptase polymerase chain reaction (RT-PCR) in MDA-PCa-2b tumors. The present study indicates that chemotherapy targeted to LHRH receptors on tumors inhibits growth of MDA-PCa-2B prostate cancers representative of human carcinoma disseminated to the bone and progressing despite androgen withdrawal.
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PMID:Inhibition of in vivo proliferation of MDA-PCa-2b human prostate cancer by a targeted cytotoxic analog of luteinizing hormone-releasing hormone AN-207. 1179 Apr 54

Uroguanylin, a peptide hormone highly expressed in the gastrointestinal tract, is implicated in the regulation of epithelial salt and water transport processes. Since little is known about a possible role of uroguanylin in the reproductive system, we investigated for the first time the occurrence of this peptide in the human prostate using specimens of benign prostatic hyperplasia. Northern blot analyses detected a single uroguanylin transcript of approximately 600 bp in prostate RNA. The uroguanylin expression was further investigated by reverse transcriptase polymerase chain reaction of prostate RNA with uroguanylin-specific primers. Sequencing of the fragments obtained indicated the presence of a uroguanylin molecule with a sequence identical to its intestinal counterpart. Furthermore, in situ hybridization and immunohistochemistry revealed that uroguanylin mRNA and peptide are confined to epithelial cells of the prostate glands. Comparison with the distribution pattern of immunoreactivity for prostate-specific antigen (PSA) showed a high degree of colocalization of uroguanylin- and PSA-immunoreactive cells. In addition, by western blotting techniques we detected the presence of high molecular weight uroguanylin-immunoreactive material in prostatic fluid. In conclusion, our study indicates that the human prostate glands synthesize and secrete (pro-)uroguanylin. We hypothesize that this hormone may play a novel role in the male reproductive tract.
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PMID:Occurrence and localization of uroguanylin in the aging human prostate. 1254 7

In recent years, the mRNA for prostate-specific antigen (PSA) has been investigated as a potential marker for molecular staging of prostate cancer. We report a simple, rapid, and sensitive assay protocol for the quantification of PSA mRNA in peripheral blood by using reverse transcriptase polymerase chain reaction (RT-PCR) and a chemiluminometric hybridization assay. A recombinant RNA internal standard (IS) that has the same size and primer binding sites as the PSA mRNA is included in the RT-PCR mixture. Total RNA from the sample is coextracted with a constant amount of IS RNA and subjected to RT-PCR. Amplified sequences are labeled with biotin during PCR by using a biotinylated upstream primer. The products are heat-denatured and hybridized with oligonucleotide-specific probes (for PSA and IS) that are immobilized in microtiter wells. Immobilization of oligonucleotide probes is achieved by adsorption of their conjugates with bovine serum albumin. The hybrids are measured using alkaline phosphatase-labeled streptavidin and a dioxetane chemiluminogenic substrate. The ratio of the luminescence values obtained for the PSA mRNA and the RNA IS is a linear function of the initial amount of PSA mRNA present in the sample prior to RT-PCR amplification. The linear range extended from 50 to 500,000 PSA mRNA copies, and the overall reproducibility of the assay, including RT-PCR and hybridization, ranged from 11.5 to 14.2%. Samples containing total RNA from PSA-expressing LNCaP cells give luminescence ratios that are linearly related to the number of cells in the range of 0.04-400 cells. The method was applied to PSA mRNA determination in peripheral blood of healthy individuals, patients with benign prostate hyperplasia, patients with prostate cancer, and patients with other types of localized cancer.
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PMID:Determination of prostate specific antigen mRNA in peripheral blood by reverse transcriptase polymerase chain reaction and a simple chemiluminometric hybridization assay in a high-throughput format. 1257 64


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