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)

Micrometastases of solid tumors are most commonly detected by immunocytochemistry using monoclonal antibodies directed against tissue-specific gene products like cytokeratin-18 (CK-18) and the carcinoembryonic antigen (CEA). While CK-18 is a marker for epithelia in general, CEA is mainly employed in the detection of gastrointestinal and breast carcinomas. To improve the sensitivity and specificity of micrometastasis detection, we planned to establish polymerase chain reaction (PCR) assays for both markers. Here we provide strong evidence for the existence of a CK-18 pseudogene, since specific amplification (i) was readily obtained from healthy bone marrow donors, (ii) did not require reverse transcription of CK-18 mRNA and (iii) was not abolished by RNase treatment. Using a CK-18-specific probe, Southern blot analyses revealed identical-size fragments for both genomic DNA and a CK-18 cDNA after digestion with appropriate restriction enzymes. On the other hand, the amplification of CEA mRNA (i) was never observed in bone marrow samples of healthy donors or patients without solid tumors, (ii) required intact mRNA and the reverse transcriptase reaction, and (iii) could not be obtained after RNase treatment. In reconstitution experiments, single CEA-expressing tumor cells were reliably detected among 2 x 10(7) normal bone marrow cells. We conclude that, due to the presence of pseudogene(s), PCR-based detection systems are not readily suitable for CK-18, while the CEA mRNA amplification should provide a sensitive and specific test for the presence of ectopic, and hence presumed malignant, CEA-expressing cells in body fluids.
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PMID:Diagnosis of micrometastases by the amplification of tissue-specific genes. 754 67

There are few DNA-based studies that detect cancer micrometastases in lymph nodes. We have assayed for the specific detection of carcinoembryonic antigen (CEA)-expressing carcinoma cells in the lymph nodes of patients with gastrointestinal or breast carcinomas. A CEA-specific nested reverse transcriptase (RT)-PCR assay was optimized using limiting dilutions of a CEA-positive cancer cell line mixed with normal lymphocytes. The expression of CEA mRNA was studied in 100 carcinoma tissues, 75 normal mucosal tissues, and 15 lymph nodes from patients with cholelithiasis. Each of 117 lymph nodes from 13 patients with carcinoma was divided into two pieces: one was used for histological examination and the other for RT-PCR, and the results were compared. The sensitivity ratio was one CEA-expressing cancer cell detected in 1 x 10(5) normal lymphocytes. All carcinoma tissues and normal mucosal tissues expressed CEA mRNA, while no amplification was detected in any control lymph nodes. Thirty of 117 lymph nodes were histologically involved by carcinoma cells, and all of these yielded the expected product by RT-PCR. Of the remaining 87 histologically negative nodes, CEA mRNA was detected in 47 lymph nodes by RT-PCR. The positive rate increased from 26% by histological examination to 66% by RT-PCR. The assay by CEA-specific nested RT-PCR is not only sensitive but widely applicable for the detection of cancer micrometastases in lymph nodes. This method may lead to an earlier diagnosis and treatment of patients with subclinical lymph node metastasis.
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PMID:Detection of cancer micrometastases in lymph nodes by reverse transcriptase-polymerase chain reaction. 754 69

Detection of the mRNA of selected genes by reverse transcriptase-polymerase chain reaction (RT-PCR) is a sensitive and powerful tool for detecting cancer cells in bone-marrow or peripheral-blood samples. In this study, we determined whether carcinoembryonic antigen (CEA) mRNA is detectable in the peripheral blood of patients with gastrointestinal or breast cancer. In addition, we studied selected patients undergoing surgical procedures to assess whether tumor manipulation during operation enhances cancer-cell dissemination. Peripheral blood from 55 patients with gastrointestinal or breast cancer and from 22 control cases was analysed for CEA mRNA using RT-PCR. For 15 selected cases undergoing curative surgery for cancer, samples were also obtained during and after surgery. The lower limit of detection was 1 to 10 CEA-positive cells diluted among 1 x 10(7) blood mononuclear cells. The test was positive for 20 of the 55 patients with cancer (36%). None of the 22 control samples were positive. An increase in positivity was observed with increasing stage of disease; however, even some patients with early-stage cancer showed positive results. In addition, CEA mRNA could be detected in the peripheral blood during operation in 3 of 13 patients whose pre-operative CEA mRNA in the peripheral blood had been negative. These findings suggest that, (1) RT-PCR amplification of CEA mRNA is an efficient means of detecting circulating solid cancer cells in the peripheral blood, although long-term clinical studies should be done to evaluate its usefulness; (2) not only breast cancer but also gastrointestinal cancer might be better regarded as a systemic disease even in early stages of carcinoma; and (3) surgical manipulation can provoke cancer-cell dissemination.
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PMID:Molecular detection of circulating solid carcinoma cells in the peripheral blood: the concept of early systemic disease. 898 Jan 76

Cytological examination of peritoneal washes is a useful predictor of peritoneal recurrence in gastric carcinoma patients. In the present study, even more sensitive detection of free cancer cells could be achieved through amplification of carcinoembryonic antigen (CEA) mRNA by means of the reverse transcriptase-polymerase chain reaction (RT-PCR). CEA was first confirmed to be present in all the gastric cancer cell lines examined, irrespective of the differentiation degree, and absent in blood and mesothelium, indicating the specificity of this approach for detection of carcinoma cells in peritoneal lavage fluid. In sensitivity tests, CEA RT-PCR proved to be capable of detecting 10 carcinoma cells per sample. Peritoneal washes of 15 of 48 gastric carcinoma patients, including all 10 patients with positive cytology results, proved positive for CEA mRNA. None of the 5 patients with benign disease was positive. Moreover, a close association with the depth of cancer invasion was established. The results indicate that the assay is more sensitive for detection of free carcinoma cells in the peritoneal cavity than conventional cytology. This is the first study to suggest the feasibility of the RT-PCR method for prediction of peritoneal recurrence in gastric cancer patients.
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PMID:Detection of carcinoembryonic antigen-expressing free tumor cells in peritoneal washes from patients with gastric carcinoma by polymerase chain reaction. 931 Jan 42

Circulating tumour cells play a central role in the metastatic process, but little is known about the relationship between this cellular subpopulation and the development of secondary disease. This study was aimed at assessing the presence of colonic cells in peripheral blood of patients with colorectal cancer in different evolutionary stages, by means of reverse transcriptase polymerase chain reaction (RT-PCR) targeted to carcinoembryonic antigen (CEA) mRNA. In vitro sensitivity was established in a recovery experiment by preparing serial colorectal cancer cell dilutions. Thereafter, 95 colorectal cancer patients and a control group including healthy subjects (n=11), patients with other gastrointestinal neoplasms (n=11) or inflammatory bowel disease (n=9) were analysed. Specific cDNA primers for CEA transcripts were used to apply RT-PCR to peripheral blood samples. Tumour cells were detected down to five cells per 10 ml blood, thus indicating a sensitivity limit of approximately one tumour cell per 10(7) white blood cells. CEA mRNA expression was detected in 39 out of 95 colorectal cancer patients (41.1%), there being a significant correlation with the presence of distant metastases at inclusion. None of the healthy volunteers and only 1 of 11 patients (9.1%) with other gastrointestinal neoplasms had detectable CEA mRNA in peripheral blood. By contrast, CEA mRNA was detected in five of the nine patients (55.6%) with inflammatory bowel disease. These results confirm that it is feasible to amplify CEA mRNA in the peripheral blood, its presence being almost certainly derived from circulating malignant cells in colorectal cancer patients. However, CEA mRNA detectable in blood of patients with inflammatory bowel disease suggests the presence of circulating non-neoplastic colonic epithelial cells.
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PMID:Detection of colonic cells in peripheral blood of colorectal cancer patients by means of reverse transcriptase and polymerase chain reaction. 982 81

Free cancer cells exfoliated from cancer-invaded serosa contribute to peritoneal dissemination, the most frequent pattern of recurrence in patients with gastric and ovarian cancers. This study was designed to evaluate the prognostic significance of free cancer cells in peritoneal washes detected using the reverse transcriptase-polymerase chain reaction (RT-PCR) and cytology. RT-PCR analysis with primers specific for the carcinoembryonic antigen (CEA) gene was found to be more sensitive than cytology for detection of free tumor cells in the peritoneal washes, collected at laparotomy from 199 gastric carcinoma patients, with higher detection rates for each of the T-categories in the TNM classification. Six patients with synchronous and 5 with recurrent peritoneal dissemination were found among 25 advanced cancer patients with positive PCR and negative cytology results. Positive PCR results were significantly associated with poor survival of curatively resected advanced gastric carcinoma patients (P < 0.001). A rapid method for detecting CEA mRNA using the LightCycler and the dsDNA binding dye SYBR green I was also developed. The results obtained using this technique were essentially the same as those obtained using the conventional RT-PCR method. Furthermore, RT-PCR analysis with primers specific for MUC1 epithelial mucin were performed on peritoneal washes from patients with ovarian cancer. Peritoneal washes from 21 of 25 ovarian carcinoma patients, including all 17 with positive cytology results, were positive for MUC1 mRNA, again indicating a higher sensitivity using this method than conventional cytology. Highly sensitive and rapid detection of free cancer cells in peritoneal washes, most reliably by RT-PCR, is a powerful technique to predict peritoneal dissemination in patients with gastric and ovarian cancers.
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PMID:Molecular diagnostic detection of free cancer cells in the peritoneal cavity of patients with gastrointestinal and gynecologic malignancies. 1035 56

We analyzed the peripheral blood of patients with gastrointestinal tract cancer at different stages to assess the presence of carcinoembryonic antigen (CEA) mRNA by reverse transcriptase-polymerase chain reaction (RT-PCR), which we used as an indicator for micrometastatic malignant cells. A total of 35 gastric, 24 colorectal, 4 esophageal and 4 biliary tract cancer patients and nine normal healthy subjects were studied. No CEA mRNA was detected in the nine normal healthy volunteers. CEA mRNA was detected in 100% (10/10) of metastatic, 33.3% (3/9) of early gastric cancer (EGC), and 18.8% (3/16) resectable gastric cancer patients, respectively. In colorectal cancer, 55.6% (5/9) of metastatic cancers were positive for CEA mRNA, and 26.7% (4/15) Duke stage B/C showed positive. One patient with stage III gastric cancer who was negative CEA mRNA initially and turned positive during follow-up, developed multiple bone metastasis one month later. Another stage III patient, who was positive for CEA mRNA, preoperatively revealed early relapse in two months. These results suggest that the identification of circulating tumor cells using RT-PCR for the detection of CEA mRNA is feasible and this analysis may be a promising tool for early detection of micrometastatic circulating malignant cells in patients with gastrointestinal tract cancer.
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PMID:Detection of tumor cell contamination in peripheral blood by RT-PCR in gastrointestinal cancer patients. 1064 39

To determine whether the tumor cell contamination of peripheral blood stem cells influences clinical impacts on high-dose chemotherapy in patients with metastatic breast cancer, we analyzed carcinoembryonic antigen (CEA) mRNA in the apheresis products by nested RT-PCR (reverse transcriptase-polymerase chain reaction). A total of 38 metastatic breast cancer patients and ten normal healthy subjects as a negative control were included. Twenty out of 38 (51.3%) apheresis products from patients with metastatic breast cancer were positive for CEA mRNA. CEA mRNA was noted in 54.8% (17/31) of patients mobilized with chemotherapy plus G-CSF and 42.8% (3/7) of patients with G-CSF alone. There was no significant difference in age, estrogen receptor, menopausal status, mobilization method, disease free interval, or number of metastasis sites (1 vs > or = 2) between positive and negative groups. The presence of CEA mRNA in apheresis products did not influence the time to progression and overall survival in both groups. However, both the univariate and the multivariate analysis disclosed that the number of metastasis was associated with survival significantly. We suggest that the tumor cell contamination does not predict poor treatment outcome in patients with metastatic breast cancer.
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PMID:Clinical impacts of tumor cell contamination of hematopoietic stem cell products in metastatic breast cancer patients undergoing autologous peripheral blood stem cell transplantation: multicenter trial. 1130 43

Laparoscopic surgery for treatment of colorectal cancer has been suggested to enhance tumor dissemination. Recently, molecular techniques have been developed to detect micrometastatic disease in patients with solid tumors, with a higher accuracy than cytologic or immunohistochemical approaches. This study was undertaken to investigate the potential harmful effects of laparoscopic-assisted colectomy on neoplastic cell mobilization in patients with resectable colorectal cancer. Fifty patients with nonmetastatic colorectal cancer were randomly assigned to laparoscopic-assisted (LAC, n = 26) or open (OC, n = 24) colectomy. Peripheral venous blood samples were obtained preoperatively, immediately after tumor removal, and 24 hours later. In 10 patients from each treatment group, portal blood and peritoneal fluid samples were also obtained before and after resection. Neoplastic cells were detected by means of reverse transcriptase-polymerase chain reaction targeted to carcinoembryonic antigen (CEA) transcription. CEA mRNA was detected in peripheral venous blood samples from 35 of 50 colorectal cancer patients preoperatively. Among those 15 baseline-negative patients, four experienced conversion 24 hours after tumor resection (2 [33%] of 6 in the LAC group vs. 2 [22%] of 9 in the OC group; NS). At that time point, clearance of CEA mRNA expression was observed in 14 of the 35 baseline-positive patients (9 [45%] of 20 in the LAC group vs. 5 [33%] of 15 in the OC group; NS). In addition, only one patient in the LAC group with baseline-negative CEA mRNA expression experienced portal blood conversion after tumor removal, although his peripheral blood level remained negative. Finally, baseline peritoneal fluid CEA mRNA expression was never detected, but one patient in each group became positive postoperatively. These results confirm that preoperative and perioperative mobilization of neoplastic cells is a frequent occurrence in patients with colorectal cancer, but the surgical approach (LAC vs. OC) does not seem to be a determining factor.
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PMID:Laparoscopic-assisted vs. open colectomy for colorectal cancer: influence on neoplastic cell mobilization. 1130 50

Detection of breast cancer micrometastases based on specific genetic markers may provide useful information to justify appropriate therapeutic strategies. We examined the presence of a carcinoembryonic antigen (CEA) messenger RNA(mRNA) in the peripheral blood of 32 patients with varying stages of breast cancers by means of the reverse transcriptase-polymerase chain reaction (RT-PCR) assay prior to and after the curative operation. CEA mRNA were detected in the peripheral blood samples from 12 (38%) out of 32 breast cancer patients prior to surgery. Among 12 CEA mRNA-positive patients prior to surgery, 4 (33.3%) relapsed from breast cancer within 2 years after surgery. Moreover, CEA mRNA was detected in the peripheral blood samples obtained prior to surgery in 3 out of 11 patients (27.2%) with a stage I disease. One out of three of these patients had a relapse in lung. There were four patterns of CEA mRNA expression, ( +, + ), (+, -), (-, + ), and (-, -) in the pre- and post-operative blood samples. In 12 CEA mRNA-positive patients submitted to surgical resection of the primary tumor, persistence of CEA mRNA expression was observed in five patients (+, +) within a month after surgical treatment. Three out of these 5 patients (60%) relapsed from breast cancer within 2 years after surgery. In 7 other patients (+, -), CEA mRNA expression was not detected within a month after tumor removal, and recurrence occurred in 1 out of the 7 patients (14%) within 2 years after surgery. In 19 patients, CEA mRNA expression was not detected in pre- or post-operative blood samples (-, -). There was a patient whose blood sample was negative for CEA mRNA before the operation, but changed to show a positive result after surgery (-,+). No recurrence was found in 20 of CEA mRNA-negative patients prior to surgery (-, +), (-, -). This study suggested that the presence of CEA mRNA expression in preoperative peripheral blood sample represent the progression of the disease, especially the risk of hematogenous metastasis in the patients in spite of their clinical stage, and the presence of CEA mRNA in the postoperative blood sample may represent the evidence of a residual disease. Thus consideration might be given for adding combined multi-modal therapy.
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PMID:Clinical significance of circulating cancer cells in peripheral blood detected by reverse transcriptase-polymerase chain reaction in patients with breast cancer. 1131 30


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