Investigation on cell proliferation with a new antibody against thymidine kinase 1.
01 Jan 2001-Analytical Cellular Pathology (Hindawi Publishing Corporation)-Vol. 23, Iss: 1, pp 11-19
TL;DR: A polyclonal anti‐TK1 antibody against a synthetic peptide from the C‐terminus of human TK1 is developed and demonstrated, demonstrating the exclusive location of TK 1 in the cytoplasm of cells.
Abstract: The cytosolic thymidine kinase 1 (TK1) is one of the enzymes involved in DNA replication. Based on biochemical studies, TK1 is activated at late G1 of cell cycle, and its activity correlates with the cell proliferation. We have developed a polyclonal anti‐TK1 antibody against a synthetic peptide from the C‐terminus of human TK1. Using this antibody, here we demonstrate the exclusive location of TK1 in the cytoplasm of cells. Cell cycle dependent TK1 expression was studied by simultaneous fluorescence staining for TK1 and bromodeoxyuridine, by using elutriated cells, and by quantitation of the amount TK1 in relation to the cellular DNA content. TK1, which was strongly expressed in the cells in S+G2 period, raised at late G1 and decreased during mitosis. The amount of TK1 increased three folds from late G1 to G2. TK1 positive cells were demonstrated in areas of proliferation activity of various normal and malignant tissues. The new anti‐TK1 antibody works in archival specimens and is a specific marker of cell proliferation.
Citations
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TL;DR: The potential clinical applications of proliferative indices are discussed, including their use as prognostic indicators and predictors of response to systemic therapy.
Abstract: Various methods are available for the measurement of proliferation rates in tumours, including mitotic counts, estimation of the fraction of cells in S-phase of the cell cycle and immunohistochemistry of proliferation-associated antigens. The evidence, advantages and disadvantages for each of these methods along with other novel approaches is reviewed in relation to breast cancer. The potential clinical applications of proliferative indices are discussed, including their use as prognostic indicators and predictors of response to systemic therapy.
193 citations
Cites background from "Investigation on cell proliferation..."
...Wang and colleagues [78] have developed a polyclonal anti-TK1 antibody and demonstrated cell cycle-dependent expression of the enzyme....
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TL;DR: Chicken egg yolk immunoglobulins generated to a synthetic 31-amino acid peptide from the C-terminal of human HeLa thymidine kinase 1 (TK1) enzyme are suggested to be potentially useful for serological and immunohistochemical detection of TK1 as an early prognosis and for monitoring patients undergoing treatment.
74 citations
Journal Article•
TL;DR: The results indicate that the S-TK1 concentration is higher in patients developing distant and/or loco-regional recurrence 3 months post-surgery.
Abstract: Background The prognostic value of the concentration of serum thymidine kinase 1 (S-TK1) with regard to recurrence in low risk breast cancer patients, 3 months after surgery was evaluated. Patients and methods The concentration of S-TK1 in serum was determined in 120 breast cancer patients at the time of surgery and in 67 patients 3 months after surgery, by anti-TK1 chicken IgY antibody, using a dot-blot immuno-assay. The S-TK1 concentration was compared with the serological activity of thymidine kinase (STK) and of carbohydrate antigen (CA 15-3). Results A statistically significant trend (unadjusted) was found for recurrence (distant or loco-regional) in patients with a higher S-TK1 concentration, as compared with patients with a lower S-TK1 concentration. A multivariate analysis gave the same results. The hazard rate ratio for developing distant and/or loco-regional recurrence in patients with a higher S-TK1 concentration was about six to seven times higher than in patients with a lower S-TK1 concentration. Conclusion Our results indicate that the S-TK1 concentration is higher in patients developing distant and/or loco-regional recurrence 3 months post-surgery.
72 citations
TL;DR: The status of TK1 for cancer monitoring and its use as a proliferation marker are summarized and a comprehensive overview about the association of Tk-1 with various entities is given.
70 citations
Cites background from "Investigation on cell proliferation..."
...The presence of TK1 positive cells could be demonstrated in both, normal and malignant tissue [11]....
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TL;DR: Determination of thymidine kinase helps to monitor the follow-up of solid tumours and haematological malignancies as well as indicating the efficacy of adjuvant and palliative chemotherapy.
Abstract: Thymidine kinase 1 (TK 1-fetal) is a cell cycle-dependent marker that increases dramatically during the S-phase of the cell cycle. In this review, the authors discuss serum levels of thymidine kinase in a variety of neoplasias. Determination of thymidine kinase helps to monitor the follow-up of solid tumours and haematological malignancies as well as indicating the efficacy of adjuvant and palliative chemotherapy. Elevated levels of thymidine kinase must always be interpreted together with a detailed knowledge of the patient's condition because nonspecific elevations of serum levels (inflammatory and autoimmune diseases) must be excluded.
70 citations
Cites methods from "Investigation on cell proliferation..."
...Although immunochemical methods have been reported, a highly sensitive but convenient approach for routine measurement of serum TK1 in haematological malignancies has not been forthcoming [16-19] ....
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References
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TL;DR: Examination of all 49 breast tumors revealed a 61% frequency of deletion at or near the p53 locus, however, the presence of allelic deletion did not correlate with overexpression of the protein.
Abstract: Overexpression of an activated form of the p53 protein may be involved in neoplastic transformation. We found widespread overexpression of p53 by immunohistochemical staining in 11 (22%) of 49 primary invasive human breast cancers. Northern blot analysis showed that this overexpression was not due to an increase in the steady-state level of p53 mRNA. The p53 gene was directly sequenced in 7 of these tumors with elevated levels of the protein and, in each case, a mutation that altered the coding sequence for p53 was found in a highly conserved region of the gene. Whereas 4 of these tumors contained only a mutant p53 allele, the other 3 tumors exhibited coding sequences from both a mutant and a wild-type allele. p53 mutations have previously been correlated with allelic loss of part of chromosome 17p that contains the p53 locus. Examination of all 49 breast tumors revealed a 61% frequency of deletion at or near the p53 locus. However, the presence of allelic deletion did not correlate with overexpression of the protein. Six tumors that were deleted but did not express high levels of the protein were sequenced and all retained a wild-type p53 allele. In this series of human breast cancers, overexpression of the p53 protein, not allelic loss on chromosome 17p, was always associated with mutation of the p53 gene.
280 citations
"Investigation on cell proliferation..." refers background in this paper
...Recently, also cell cycle related gene expression such as the expression of the p53 tumor suppressor gene has been the subject of studies relating to the prognosis of malignancies [4,16,30]....
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Journal Article•
TL;DR: Cyclin D1 is mainly overexpressed in the well differentiated and lobular types of invasive breast cancer and is strongly associated with estrogen receptor positivity and it is negatively correlated with the proliferation marker mitoses count and with the differentiation markers nuclear area and nuclear volume.
Abstract: Cyclin D1 overexpression, detected by standard immunohistochemistry, was correlated with other prognostic variables and its prognostic value was evaluated in a group of 148 invasive breast cancers with long-term follow-up. Overexpression of cyclin D1 (59% of cases) was negatively correlated (chi 2 test) with histological grade (P = 0.0001), mean nuclear area (P = 0.004), mean nuclear volume (P = 0.02), and mitotic activity (P = 0.03) and positively correlated with estrogen receptor (P = 0.0001). There was a strong correlation between cyclin D1 overexpression and histological type (P = 0.0001). Positive cyclin D1 staining was seen in 11 of 13 tubular carcinomas, 3 of 3 mucinous carcinomas, 4 of 4 invasive cribriform carcinomas, and 17 of 20 lobular carcinomas. Of 102 ductal cancers, 52 were positive, and all 6 medullary carcinomas were negative. There were no significant correlations with lymph node status, tumor size, or DNA ploidy. In survival analysis, cyclin D1 overexpression did not provide significant univariate or multivariate prognostic value. In conclusion, cyclin D1 is mainly overexpressed in the well differentiated and lobular types of invasive breast cancer and is strongly associated with estrogen receptor positivity. It is negatively correlated with the proliferation marker mitoses count and with the differentiation markers nuclear area and nuclear volume. However, cyclin D1 overexpression does not seem to have prognostic value in invasive breast cancer when no adjuvant treatment is given.
207 citations
"Investigation on cell proliferation..." refers background in this paper
...We have developed a polyclonal anti-TK1 antibody against a synthetic peptide from the C-terminus of human TK1....
[...]
TL;DR: A new method of analysis utilizing a monoclonal antibody to bromodeoxyuridine (BrdUrd) has been developed which makes it possible in most cases to accurately determine phase fractions without resorting to mathematical models.
Abstract: The flow cytometric measurement of DNA distributions of cells has many applications in biomedical research. Phase fractions estimated (calculated) from such distributions are used to study the growth characteristics of various types of cells, particularly when the cells have been exposed to perturbing agents such as chemotherapeutic drugs. For more than 10 years many methods for resolving DNA distributions into the three cell subpopulations (G1, S and G2 + M) have been reported in the literature. A new method of analysis utilizing a monoclonal antibody to bromodeoxyuridine (BrdUrd) has been developed (Gratzner, 1982; Dolbeare et al., 1983) which makes it possible in most cases to accurately determine phase fractions without resorting to mathematical models. The procedure involves the incorporation of BrdUrd by growing (DNA synthesizing) S phase cells, labelling the BrdUrd with a fluorescent monoclonal antibody, and the bivariate measurement of the antibody and of total DNA content, the latter through propidium-iodide staining. The resulting bivariate distributions clearly and simply resolve the three subpopulations. This paper describes the method and illustrates its use in the analysis of various fractions of elutriated exponentially growing Chinese hamster ovary (CHO) cells.
206 citations
"Investigation on cell proliferation..." refers methods in this paper
...In clinical investigations, mitotic count, 3H-thymidine labeling index, bromodeoxyuridine (BrdU) incorporation, expression of Ki-67, proliferating cell nuclear antigen, and cyclins, as well as measurement of S-phase by means of DNA flow cytometry have been used in the assessment of cell proliferation [5,8,17,22,33,36,37]....
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TL;DR: The ability of semiquantitative PCNA grading to allow for intra-tumoural variation suggests it may have advantages over absolute counting, which is prone to sampling error when tumour heterogeneity is a major factor.
Abstract: A new monoclonal antibody to proliferating cell nuclear antigen (PCNA), PC10, which can be used on routinely processed tissue, was applied to 93 cases of gastric carcinoma. Significant intra-tumoural variation in staining occurred. In addition to a PCNA index (percentage of positive cells per 1000 tumour cells), a semiquantitative PCNA grading system was devised, based on estimates of less than or more than 50% of positive tumour cells in whole sections. Neither PCNA index nor PCNA grade showed any correlation with established histological variables, tumour stage, or the presence of lymph node metastases. No significant correlation was observed between PCNA index and S + G2M phase fraction measured by flow cytometric analysis. To analyse survival tumours with PCNA indices above and below the median level (41%) were compared. Those with a higher index tended to have a worse prognosis, but when PCNA grade was considered, it was found to have definite independent prognostic value, tumours of low grade surviving better than those of high grade. The ability of semiquantitative PCNA grading to allow for intra-tumoural variation suggests it may have advantages over absolute counting, which is prone to sampling error when tumour heterogeneity is a major factor. The prognostic value of PC10 staining in gastric carcinoma is therefore promising.
189 citations
"Investigation on cell proliferation..." refers methods in this paper
...In clinical investigations, mitotic count, 3H-thymidine labeling index, bromodeoxyuridine (BrdU) incorporation, expression of Ki-67, proliferating cell nuclear antigen, and cyclins, as well as measurement of S-phase by means of DNA flow cytometry have been used in the assessment of cell proliferation [5,8,17,22,33,36,37]....
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