ABSTRACT
The P-N bond in oligonucleotide P3' --> N5' phosphoramidates (5'-amino-DNA) is known to be chemoselectively cleaved under mild acidic conditions. We prepared homopyrimidine oligonucleotides containing 5'-amino-5'-deoxythymidine (5'-amino-DNA thymine monomer) or its conformationally locked congener, 5'-amino-2',4'-BNA thymine monomer, at midpoint of the sequence. The effect of triplex formation with homopurineohomopyrimidine dsDNA targets on acid-mediated hydrolysis of the P3' --> N5' phosphoramidate linkage was evaluated. Very interestingly, it was found that the triplex formation significantly accelerates the P-N bond cleavage.
Subject(s)
Oligodeoxyribonucleotides/chemical synthesis , Amides/chemical synthesis , Amides/chemistry , Base Sequence , Bridged-Ring Compounds/chemical synthesis , Bridged-Ring Compounds/chemistry , Hydrogen-Ion Concentration , Hydrolysis , Molecular Structure , Oligodeoxyribonucleotides/chemistry , Phosphoric Acids/chemical synthesis , Phosphoric Acids/chemistry , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
BACKGROUND: Although ascites cytology is important for therapeutic strategies, it is difficult to distinguish cancer infiltration from reactive mesothelial proliferation in some patients. In this study, the authors applied CDX2 immunocytochemistry to improve diagnostic accuracy. METHODS: The authors examined the distribution of CDX2 expression in carcinoma specimens using paraffin-embedded tissues from various organs from 549 cancer patients. CDX2 immunostaining was applied to the 116 ascites specimens. RESULTS: CDX2 expression was detected in a restricted range of cancers, with the vast majority of them originating from the gastrointestinal tract and pancreas. When applied to ascites specimens, no positive reactions were detected in any of the 81 cytology-negative and molecular genetic analysis-negative specimens. By contrast, 28 of 35 specimens diagnosed as suspicious for malignancy or malignancy showed a positive reaction. Furthermore, the authors found that a nuclear-positive reaction was easily evaluated, even with a high level of background staining, and single cancer cells in 10(6) normal cells could be detected. CONCLUSION: Results suggest that CDX2 is a specific and sensitive marker to detect gastrointestinal and pancreatic malignancies in ascites cytology.
Subject(s)
Adenocarcinoma/diagnosis , Ascites/pathology , Biomarkers, Tumor/analysis , Gastrointestinal Neoplasms/diagnosis , Homeodomain Proteins/analysis , Immunohistochemistry , Pancreatic Neoplasms/diagnosis , Adenocarcinoma/chemistry , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Ascites/genetics , Biomarkers, Tumor/genetics , CDX2 Transcription Factor , Carcinoembryonic Antigen/analysis , Carcinoembryonic Antigen/genetics , Cytodiagnosis/methods , Gastrointestinal Neoplasms/chemistry , Gastrointestinal Neoplasms/genetics , Gastrointestinal Neoplasms/pathology , Humans , Pancreatic Neoplasms/chemistry , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , RNA, Messenger/analysis , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction , Sensitivity and SpecificityABSTRACT
Tamoxifen (TAM) causes cancer in rat liver and human endometrium, whereas the carcinogenicity of its chlorinated analogue toremifene (TOR) has not been observed. To elucidate the genotoxicity of TOR, the capability of forming DNA adducts by TOR was examined in the leukocytes of patients treated with TOR. Leukocytes were collected from 27 breast cancer patients (57.7 +/- 11.4 years old) taking TOR (40 mg/day for 25 patients, 80 mg/day for one patient, and 120 mg/day for one patient; average duration, approximately 12 months) and 20 untreated breast cancer patients (58.2 +/- 12.3 years old). The DNA extracted was analyzed by (32)P-postlabeling/high-performance liquid chromatography. No DNA adducts were detected in the leukocytes of either TOR-treated or nontreated patients. Our results contrast to the previous observation detecting TAM-DNA adducts in several patients treated with TAM, indicating that TOR is less genotoxic to humans.
Subject(s)
Antineoplastic Agents, Hormonal/toxicity , Breast Neoplasms/drug therapy , Breast Neoplasms/metabolism , DNA Adducts/analysis , Leukocytes/chemistry , Toremifene/toxicity , Adult , Aged , Aged, 80 and over , Antineoplastic Agents, Hormonal/therapeutic use , Chromatography, High Pressure Liquid , DNA, Neoplasm/chemistry , DNA, Neoplasm/drug effects , DNA, Neoplasm/isolation & purification , Female , Humans , Middle Aged , Toremifene/therapeutic useABSTRACT
BACKGROUND: Despite advances in diagnostic imaging techniques, the differentiation between pancreatic cancer and focal pancreatitis remains difficult. This study evaluated the effectiveness of EUS-guided FNA in the differential diagnosis between pancreatic cancer and focal pancreatitis, with particular reference to detection of the K-ras point mutation. METHODS: The study included 62 consecutive patients with pancreatic ductal cancer and 15 patients with focal pancreatitis demonstrated as a pancreatic mass lesion by EUS. RESULTS: Sensitivity, specificity, overall accuracy, positive predictive value, and negative predictive value of cytopathologic diagnosis were 82%, 100%, 86%, 100%, and 58%, respectively. Sensitivity, specificity, overall accuracy, positive predictive value, and negative predictive value of histopathologic diagnosis were 44%, 100%, 55%, 100%, and 32%, respectively. The K-ras point mutation was found in 74% of pancreatic cancers and 0% of focal pancreatitis lesions. No complication of EUS-guided FNA was observed. CONCLUSIONS: EUS-guided FNA is useful for the differential diagnosis of pancreatic mass lesions caused by pancreatic cancer and focal pancreatitis. Analysis for the K-ras point mutation in specimens obtained by EUS-guided FNA may enhance diagnostic accuracy in indeterminate cases.
Subject(s)
Biopsy, Fine-Needle , Endosonography , Pancreatic Neoplasms/diagnostic imaging , Pancreatic Neoplasms/pathology , Pancreatitis/diagnostic imaging , Pancreatitis/pathology , Adult , Aged , Diagnosis, Differential , Female , Genes, ras/genetics , Humans , Male , Middle Aged , Pancreatic Neoplasms/genetics , Pancreatitis/genetics , Point Mutation/genetics , Predictive Value of Tests , Reproducibility of Results , Retrospective StudiesABSTRACT
Tamoxifen (TAM), a widely used antiestrogen for breast cancer therapy and chemoprevention, increases the incidence of endometrial cancer in women. The formation of DNA adducts induced by tamoxifen may initiate endometrial cancer. To evaluate the genotoxic risk of TAM, the formation of DNA adducts in leukocytes was examined. Blood samples were collected from 47 breast cancer patients (61.7 +/- 12.5 years) taking TAM (20 mg/day; average duration until sampling, approximately 37 months) and 20 untreated patients (58.2 +/- 12.3 years), and their leukocyte DNA was analyzed by 32P-postlabeling/HPLC analysis. This assay resolves synthetic standards, trans- and cis-diastereoisomers of alpha-(N2-deoxyguanosinyl)tamoxifen 3'-monophosphate (dG3'P-N2-TAM), alpha-(N2-deoxyguanosinyl)-N-desmethyltamoxifen 3'-monophosphate (dG3'P-N2-N-dMeTAM), and alpha-(N2-deoxyguanosinyl)tamoxifen N-oxide 3'-monophosphate', and is capable of determining TAM adducts quantitatively. The detection limit of this assay is 0.6 adducts/10(9) nucleotides. trans-dG3'P-N2-TAM (fr-2; one of the two trans-isomers) was detected in six of 47 breast cancer patients treated with TAM. Among them, trans-dG(3'P-N2-N-dMeTAM (fr-2) was also detected in two patients. The total amounts of TAM-DNA adducts in the positive patients were 2.6 +/- 3.0 adducts/10(9) nucleotides. No adducts were detected in the controls. The presence of TAM-DNA adducts in the leukocyte DNA samples was confirmed using several 32P-postlabeling/HPLC systems.
