Fluorescent molecular rotors as sensors for the detection of thymidine phosphorylase.

Three new fluorescent molecular rotors were synthesized with the aim of using them as sensors to dose thymidine phosphorylase, one of the target enzymes of 5-fluorouracil, a potent chemotherapic drug largely used in the treatment of many solid tumors, that acts by hindering the metabolism of pyrimidines. 5-Fluorouracil has a very narrowtherapeutic window, in fact, its optimal dosage is strictly related to the level of its target enzymes that vary significantly among patients, and it would be of the utmost importance to have an easy and fast method to detect and quantify them. The three molecular rotors developed as TP sensors differ in the length of the alkylic spacer joining the ligand unit, a thymine moiety, and the fluorescent molecular rotor, a [4-(1-dimethylamino)phenyl]-pyridinium bromide. Their ability to trigger an optical signal upon the interaction with thymidine phosphorylase was investigated by fluorescent measurements.

Thymidine phosphorylase promotes metastasis and serves as a marker of poor prognosis in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) ranks as one of the most common and lethal malignancies worldwide. A better understanding of the mechanism responsible for HCC metastasis will be helpful for the treatment of HCC patients. Thymidine phosphorylase (TP), a key enzyme that catalyzes the conversion of thymidine to thymine and deoxyribose-1-phosphate, was demonstrated to promote the invasion and metastasis of HCC in our study. Clinical retrospective analysis revealed that metastatic HCC tumor tissues have higher TP expression, and TP expression was significantly correlated with matrix metalloproteinase (MMP) 2 and 9 expression. Survival analysis revealed that TP expression was negatively correlated with the prognosis of HCC patients. Moreover, in vitro cell experiments confirmed that TP could promote the migration and invasion of HCC cells. In addition, MMP2 and MMP9 were activated by TP overexpression. Overall, this study suggests that TP promotes metastasis and may serve as a marker of poor prognosis in HCC. Thus, TP is a potential target for the treatment of HCC.

[Morphological assessment of predictive factors in adenocarcinoma of the colon].

Immunohistochemical investigation of 15 different markers in colon adenocarcinoma was carried out. Prognostic significance showed chemokine receptor CXCR4 and Ki-67. Predictive significance was revealed for thymidylate synthase (TS) and thymidine phosphorylase (TP).

Metronomic doxifluridine chemotherapy combined with the anti-angiogenic agent TNP-470 inhibits the growth of human uterine carcinosarcoma xenografts.

Uterine carcinosarcoma is a highly aggressive gynecological neoplasm that responds poorly to conventional chemotherapy and radiotherapy. Metronomic chemotherapy is accepted as a new approach for cancer treatment, and its underlying mechanism seems to involve the suppression of angiogenesis. However, the efficacy of metronomic and anti-angiogenic therapies against uterine carcinosarcoma is unknown. The anti-angiogenic effect of doxifluridine was assessed in vitro using human umbilical vein endothelial cells (HUVEC) co-cultured with FU-MMT-1 human uterine carcinosarcoma cells. The antitumor and anti-angiogenic effects of metronomic doxifluridine (delivered via oral gavage) in combination with TNP-470 were evaluated in vivo. Tumor vascularity was assessed by contrast-enhanced color Doppler ultrasound, laser Doppler and microvessel density staining. Doxifluridine suppressed tube formation of HUVEC and vascular endothelial growth factor production by FU-MMT-1 cells. Metronomic doxifluridine alone significantly suppressed tumor growth compared with the untreated (control) group, while metronomic doxifluridine in combination with TNP-470 significantly inhibited tumor growth compared with each treatment alone. A significant reduction of intratumoral vascularity was observed in FU-MMT-1 xenografts following treatment with metronomic doxifluridine in combination with TNP-470, as compared with each treatment alone. Intestinal bleeding was only observed when the maximum tolerated dose of doxifluridine was administered in combination with TNP-470. Metronomic doxifluridine chemotherapy in combination with TNP-470 might be effective for uterine carcinosarcoma without marked toxicity, possibly acting via its potent anti-angiogenic effects. Clinical studies are needed to evaluate the safety and efficacy of this treatment in humans.

[Ki-67 expression, thymidine phosphorylase and PTEN in intraepithelial cervical carcinoma].

Expression of Ki-67, thymidine phosphorylase (TP) and PTEN were assessed in various grades of cervical intraepithelial neoplasia (CIN) in order to evaluate their potentials of predicting the gravity of possible damage to the epithelium as well as pro- or regression of CIN. Ki-67 and TP levels were shown to correlate directly with CIN grade. It was suggested that a small number of cases of Ki-67 and TP expression absence (15%), exclusively in CIN3 samples, be due to imminent progression to invasive cancer. Both separately and in combination, Ki-67 and TP expression indices should be regarded as having a potential as markers for cervical carcinoma diagnosis, grade and clinical course.

Thymidine phosphorylase expression and benefit from capecitabine in patients with advanced breast cancer.

BACKGROUND AND AIM: Capecitabine is an orally bioavailable prodrug that is converted to 5-fluorouracil through several enzymatic steps, the last of which is mediated by thymidine phosphorylase (TP). TP has been reported to be expressed at higher levels in cancer tissue compared with normal counterpart. The present study aimed at evaluating the potential relationship between TP expression and benefit from capecitabine in patients with metastatic breast cancer (BC). METHODS: Immunohistochemistry for TP and other biological markers was carried out on paraffin-embedded cancer tissues of 61 patients with BC treated with at least three cycles of capecitabine as single agent for metastatic disease. All patients had received capecitabine 1000 mg/m(2) b.i.d. days 1-14 every 21 days. The following variables were analyzed as potential determinants of benefit from capecitabine: TP expression, estrogen receptor (ER) and progesterone receptor status, human epidermal growth factor receptor-2 (HER-2) status, MIB-1 expression, performance status at the beginning of capecitabine treatment, stage at diagnosis, grade, presence of visceral metastases at the beginning of capecitabine treatment, and previous chemotherapy. RESULTS: Overall, median time to progression (TTP) was 6.5 months (range 1.4-33). On multivariate analysis, ER status [hazard ratio (HR) for progression = 0.31; 95% confidence interval (CI) = 0.15-0.64; P = 0.002], presence of visceral metastases at the beginning of capecitabine treatment (HR = 2.30; 95% CI = 1.21-4.39; P = 0.01), and capecitabine as first- or second-line treatment (HR = 2.28; 95% CI = 1.21-4.32; P = 0.01) independently predicted TTP. TP was highly expressed in 34 of 61 cases (55.7%). In the subgroup of patients with TP-expressing tumor, TTP was significantly longer in patients who received anthracyclines and taxanes before capecitabine (median TTP 7.5 versus 3.3 months, P = 0.01, log-rank test). Similarly, patients with a TP-positive tumor showed a longer TTP if they received taxanes before capecitabine than patients with TP-positive tumor who did not receive this treatment (7.3 versus 3.4 months, P = 0.03). CONCLUSIONS: These data provide further evidence that TP expression in BC could represent a biomarker of sensitivity to capecitabine treatment. Prospective studies with translational approach are desirable to confirm the predictive and prognostic role of TP.

The effect of neoadjuvant therapy on the 5-fluorouracil metabolic and relative enzymes of oral squamous cell carcinoma.

To assess the effect of neoadjuvant therapy using tegafur/uracil (UFT) and radiation therapy on the 5-fluorouracil (5-FU) metabolic and relative enzymes, thymidylate synthase (TS), dihydropyrimidine dehydrogenase (DPD), orotate phosphoribosyl transferase (OPRT) and thymidine phosphorylase (TP) in oral squamous cell carcinoma (OSCC), we examined the mRNA expression and immunohistochemical staining status of these enzymes using 17 surgical specimens. Seven patients did not receive any neoadjuvant therapy and 10 were treated with UFT and local irradiation therapy. Our result showed that the mRNA expression of these enzymes in neoadjuvant group was not significantly different from that of non-treated group using real-time quantitative PCR. To confirm the protein expression, we also carried out immunohistological staining of TS and DPD two key enzymes in the 5-FU metabolism, using the same specimens. Immunohistological staining status did not correspond to the results of mRNA analysis completely, though no significant difference between the groups was observed. Furthermore, no significant relationship between the UFT administration period and mRNA expression of the 5-FU metabolic enzymes was observed in neoadjuvant therapy group and also the distribution of the enzyme mRNA expression levels was similar to that of non-treated group. The results suggested that the neoadjuvant therapy of OSCC might not affect the expression status of 5-FU metabolic and relative enzymes in surgical tumor samples and the tumor tissues might serve as a useful specimen source to analyze the expression status of the 5-FU metabolic and relative enzymes and to determine the prospective efficiency of 5-FU-based adjuvant chemotherapy.

Antitumor activity of bevacizumab in combination with capecitabine and oxaliplatin in human colorectal cancer xenograft models.

To understand the mechanisms of the effects of combination treatments, we established animal models showing antitumor activity of bevacizumab as a monotherapy and in combination with capecitabine or capecitabine and oxaliplatin and measured thymidine phosphorylase (TP) and vascular endothelial growth factor (VEGF) levels. Tumor-inoculated nude mice were treated with bevacizumab, capecitabine, and oxaliplatin, alone or in combination, after tumor growth was confirmed and volume and microvessel density (MVD) in tumors were evaluated. Levels of TP and VEGF in the tumor were examined by ELISA. Bevacizumab showed significant antitumor activity as a monotherapy in three xenograft models (COL-16-JCK, COLO 205 and CXF280). The MVD in tumor tissues treated with bevacizumab was lower than that of the control. Antitumor activity of bevacizumab in combination with capecitabine was significantly higher than that of each agent alone (COL-16-JCK, COLO 205). Furthermore, the antitumor activity of bevacizumab in combination with capecitabine + oxaliplatin was significantly superior to that of capecitabine + oxaliplatin (COL-16-JCK). TP and VEGF levels were not increased by bevacizumab or capecitabine, respectively, suggesting there are other potentially efficacious mechanisms involved. In the present study we established human colorectal cancer xenograft models which reflect the efficacy of clinical combination therapies, capecitabine + bevacizumab and capecitabine + oxaliplatin + bevacizumab. We will further investigate the mechanisms of the combination therapies using these models.

Prognostic significance of periodic acid-Schiff-positive patterns in clear cell renal cell carcinoma.

BACKGROUND: The ability of aggressive tumors to form nonendothelial tumor cell-lined microvascular channels is known as "vasculogenic mimicry" (VM). VM channels are revealed as periodic acid-Schiff (PAS)-positive patterns, and in some tumors their presence predicts clinical outcomes. OBJECTIVE: We aimed to study VM channels in clear cell renal cell carcinoma (cRCC) tumors and explore their prognostic significance and relationship to other suggested prognostic factors such as thymidine phosphorylase (TP) and vascular endothelial growth factor (VEGF) expression. METHODS: We retrospectively studied 45 patients who had undergone radical nephrectomy for clinically confined cRCC (stage T2-T3NOMO) at the Russian Cancer Research Center. The tumor sections were reviewed for disease stage, nuclear grade, perirenal fat invasion, and lymph node involvement, and we performed immunohistochemical staining for VEGF and TP expression, and PAS staining. Disease-free survival probabilities were determined by Kaplan-Meier estimates and prognostic factors were evaluated by univariate analysis. RESULTS: PAS-positive patterns observed in the cRCC tumor included back-to-back closed loops, networks, arcs, and parallel patterns. There was a significant decrease in disease-free survival among patients with PAS-positive networks (p = 0.005), but not among patients with other PAS-positive patterns. TP expression was also a significant predictor of disease-free survival (p = 0.035), but this factor did not correlate with the presence of PAS-positive networks. Notably, in our small sample, the six patients whose tumors were positive for both factors had the highest risk of cancer recurrence. CONCLUSIONS: The presence of PAS-positive networks is an independent and relevant prognostic parameter for disease-free survival in patients with cRCC. Our data suggest that the combination of PAS-positive networks and TP expression may identify patients with the highest risk of cancer recurrence.

Prognostic significance of thymidylate synthase, dihydropyrimidine dehydrogenase and thymidine phosphorylase protein expression in colorectal cancer patients treated with or without 5-fluorouracil-based chemotherapy.

BACKGROUND: Low tumour expression levels of thymidylate synthase (TS), dihydropyrimidine dehydrogenase (DPD) and thymidine phosphorylase (TP) have been linked with improved outcome for colorectal cancer (CRC) patients treated with 5-fluorouracil (5-FU). It is unclear whether this occurs because such tumours have better prognosis or they are more sensitive to 5-FU treatment. PATIENTS AND METHODS: Associations between TS, DPD and TP levels, determined by tissue microarrays and immunohistochemistry, and survival was evaluated in 945 CRC patients according to treatment status. RESULTS: Low TS and DPD expression associated with worse prognosis in stage II [hazard ratio (HR) = 1.69, 95% confidence interval (CI) (1.09-2.63) and HR = 1.92 (95% CI 1.23-2.94), respectively] and stage III CRC patients treated by surgery alone [HR = 1.39 (95% CI 0.92-2.13) and HR = 1.49 (95% CI 1.02-2.17), respectively]. Low TS, DPD and TP associated with trends for better outcome in stage III patients treated with 5-FU [HR = 0.81 (95% CI 0.49-1.33), HR = 0.70 (95% CI 0.42-1.15) and HR = 0.66 (95% CI 0.39-1.12), respectively]. CONCLUSION: Low TS and DPD expression are prognostic for worse outcome in CRC patients treated by surgery alone, whereas low TS, DPD and TP expression are prognostic for better outcome in patients treated with 5-FU chemotherapy. These results provide indirect evidence that low TS, DPD and TP protein expression are predictive of good response to 5-FU chemotherapy.

Thymidine phosphorylase expression is associated with time to progression in patients receiving low-dose, docetaxel-modulated capecitabine for metastatic breast cancer.

BACKGROUND: Preclinical data have indicated a synergistic interaction between docetaxel and capecitabine by means of taxane-induced up-regulation of thymidine phosphorylase (TP). On the basis of such premises, we conducted a phase II trial to determine the activity and tolerability of weekly docetaxel plus capecitabine in patients with metastatic breast cancer (MBC). Furthermore, we explored the relationship between TP tumor expression and benefit from this regimen. PATIENTS AND METHODS: Patients received docetaxel 36 mg/m(2) i.v. on days 1, 8, and 15 and capecitabine orally 625 mg/m(2) b.i.d. from days 8 to 21. Cycles were repeated every 4 weeks. In the correlative study, we evaluated the TP expression by immunohistochemistry and the TP messenger RNA expression by real-time RT-PCR in the primary tumor. RESULTS: Forty-seven women were enrolled. In the intention-to-treat analysis, objective responses were achieved in 24 patients (51%). Fourteen additional patients (30%) had stable disease. The median time to progression (TTP) was 6 months (range 1-44 months). Median survival was 17 months (range 1-48 months). Overall, the treatment was well tolerated. The most common clinical adverse events (all grades) were alopecia (55%), nail changes (53%), fatigue/asthenia (51%), nausea/vomiting (51%), neutropenia (49%), and neuropathy (49%). A significantly higher TTP was observed in patients with TP-positive tumors (log-rank test, P = 0.009). Interestingly, a subgroup analysis confirmed this TTP benefit in patients with TP-positive tumors obtaining a tumor response (log-rank test, P = 0.03), whereas the statistical significance was lost in nonresponders (log-rank test, P = 0.3). CONCLUSIONS: This study indicates that a regimen with low doses of capecitabine plus weekly docetaxel is active against MBC. The correlative analysis provides preliminary evidence that TP expression may be a predictive marker for therapeutic benefit.

Thymidylate synthase, dihydropyrimidine dehydrogenase, ERCC1, and thymidine phosphorylase gene expression in primary and metastatic gastrointestinal adenocarcinoma tissue in patients treated on a phase I trial of oxaliplatin and capecitabine.

BACKGROUND: Over-expression of thymidylate synthase (TS) and dihydropyrimidine dehydrogenase (DPD) in tumor tissue is associated with insensitivity to 5-fluorouracil (5-FU). Over-expression of ERCC1 correlates with insensitivity to oxaliplatin (OX) therapy, while high thymidine phosphorylase (TP) levels predict for increased sensitivity to capecitabine (Xel). METHODS: Biopsies of metastatic tumor were taken before OX (130 mg/m2 day 1) given with Xel (1200-3000 mg/m2 in two divided doses days 1-5 and 8-12) every 3-weeks. Micro-dissected metastatic and primary tumors were analyzed for relative gene expression by real-time quantitative polymerase chain reaction. The clinical protocol prospectively identified the molecular targets of interest that would be tested. Endpoints for the molecular analyses were correlation of median, first and third quartiles for relative gene expression of each target with response, time to treatment failure (TTF), and survival. RESULTS: Among 91 patients participating in this trial; 97% had colorectal cancer. The median number of prior chemotherapy regimens was 2, and most had prior 5-FU and irinotecan. In paired samples, median mRNA levels were significantly higher in metastatic versus primary tumor (-fold): TS (1.9), DPD (3.8), ERCC1 (2.1) and TP (1.6). A strong positive correlation was noted between DPD and TP mRNA levels in both primary (r = 0.693, p < 0.0005) and metastatic tissue (r = 0.697, p < 0.00001). There was an association between TS gene expression and responsive and stable disease: patients whose intratumoral TS mRNA levels were above the median value had significantly greater risk of early disease progression (43% vs 17%), but this did not translate into a significant difference in TTF. ERCC1 gene expression above the third quartile was associated with a shorter TTF (median 85 vs 162 days, p = 0.046). Patients whose TS mRNA levels in metastatic tumor tissue were below the median had a longer overall survival (median 417 vs 294 days, p = 0.042). CONCLUSION: Target gene expression in primary tumor was significantly lower than that in paired metastatic tissue. High ERCC1 mRNA levels in metastatic tumor was associated with a shorter TTF. Lower expression of TS mRNA correlated with a lower chance of early PD with XelOX therapy and improved overall survival.

A phase II study of capecitabine and cisplatin (XP) as first-line chemotherapy in patients with advanced esophageal squamous cell carcinoma.

PURPOSE: The combination of 5-fluorouracil (5-FU) and cisplatin (FP) remains the mostly used regimen for metastatic esophageal squamous carcinoma. This phase II study assessed the efficacy and safety of capecitabine/cisplatin (XP) as a first-line chemotherapy in a homogenous cohort of patients with metastatic or recurrent esophageal squamous cell carcinoma. MATERIALS AND METHODS: Patients received 60 mg/m(2) of cisplatin intravenously (IV) on day 1 and capecitabine 1,250 mg/m(2)/dose orally twice a day on days 1-14. Treatment cycles were repeated every 3 weeks until the documented disease progression, unacceptable toxicity, or patient's refusal. Immunohistochemical studies against thymidylate synthase (TS) and thymidine phosphorylase (TP) were performed to seek predictive markers for treatment response. RESULTS: Between December 2003 and March 2006, 45 patients entered the study. All patients had histologically proven squamous cell carcinoma of the esophagus. The overall response rate (ORR) was 57.8% (95% CI, 43.3-72.2) with 0 CR and 26 PRs. The median duration of response in responders was 4.6 months (1.0-15.6 months). With a median follow-up duration of 25.7 months (10.8-42.6 months), the median time to progression was 4.7 months (95% CI, 2.5-7.0) and the median survival time was 11.2 months (95% CI, 8.5-13.9). Common grade 3 or 4 non-hematological adverse events were anorexia (18/191, 9.4%), fatigue (9/191, 4.7%), constipation (6/191, 3.1%), hand-foot syndrome (6/191, 3.1%) and diarrhea (4/191, 2.1%). The most common grade 3 or 4 hematological adverse events were neutropenia (33/191, 17.3%), followed by leucopenia (11/191, 5.8%), anemia (2/191, 1.0%) and thrombocytopenia (1/191, 0.5%). There was no treatment-related death. Neither TS nor TP showed predictive value for treatment response. CONCLUSION: The XP regimen demonstrated a promising antitumor activity in metastatic esophageal squamous cell carcinoma, which may potentially replace the FP regimen.

Prognostic significance of numeric aberrations of genes for thymidylate synthase, thymidine phosphorylase and dihydrofolate reductase in colorectal cancer.

BACKGROUND: Most human cancer cells have structural aberrations of chromosomal regions leading to loss or gain of gene specific alleles. This study aimed to assess the range of gene copies per nucleus of thymidylate synthase (TYMS), thymidine phosphorylase (TP) and dihydrofolate reductase (DHFR) in colorectal cancer, and to evaluate its prognostic significance following adjuvant chemotherapy, since these enzymes are closely related to efficacy of 5-fluorouracil (5FU). PATIENTS AND METHODS: Consecutive patients (n = 314), who were completely resected for colorectal cancer stages II-IV and adjuvantly treated with 5-FU were retrospectively evaluated. Paraffin embedded tumor specimens were assessed for gene copies per nucleus of TYMS, TP and DHFR by fluorescence in situ hybridisation (FISH) using specific peptide nucleic acid probes. Outcome according to gene copies per nucleus above and below the median were compared. Also TYMS expression, assessed by immunohistochemistry, was associated with TYMS copies per nucleus. RESULTS: The number of gene copies per nucleus were 1.7 (0.7-2.8), 1.8 (0.9-3.1) and 1.8 (1.1-2.7) median (range) for TYMS, TP and DHFR, respectively. TYMS expression was directly associated with TYMS genes per nucleus (p = 0.05). Cox multivariate analysis, adjusted for the prognostic impact of disease stage, vascular tumor invasion, and bowel obstruction at resection, revealed that high TYMS gene copy number was associated with significantly higher risk of recurrence (HR = 1.6; 95%CI 1.1-2.2; p = 0.02) and death (HR = 1.6; 95%CI 1.1-2.3; p = 0.01). No significant differences in outcome appeared according to TP and DHFR gene ratios. CONCLUSION: Aberration of TYMS gene is of significance to expression of TYMS, which may influence the biology and 5-FU sensitivity of colorectal cancer. This may be utilized in the allocation of patients for treatment approaches and for decision on follow-up programs.

Thymidine phosphorylase and dihydropyrimidine dehydrogenase are predictive factors of therapeutic efficacy of capecitabine monotherapy for breast cancer-preliminary results.

Capecitabine monotherapy was administered for 25 patients with advanced or recurrent breast cancer, and the clinical therapeutic efficacy and its relationship to expression of 5-fluorouracil-related enzymes (i. e., thymidine phosphorylase (TP), thymidylate synthase (TS) and dihydropyrimidine dehydrogenase (DPD)) were investigated. The expressions of TP, TS and DPD were determined by immunohistochemical staining techniques and rated using a scoring system of 1~4. The expression score for TP/DPD showed a statistically significant correlation with the clinical response, whereas the expression score for TP/TS also showed a correlation but it was not statistically significant. The number of patients was small, but the results revealed the potential of application of the TP/DPD expression score as a factor for predicting the efficacy of the drug in individual patients.

Neoadjuvant docetaxel and capecitabine and the use of thymidine phosphorylase as a predictive biomarker in breast cancer.

PURPOSE: Thymidine phosphorylase (TP) induction by docetaxel is a proposed mechanism for the observed preclinical synergy of docetaxel and capecitabine (DC). We evaluated whether TP protein expression is increased by docetaxel and correlates with pathologic complete response (pCR) in breast cancer patients. EXPERIMENTAL DESIGN: Women with stage II to III breast cancer were given four cycles of neoadjuvant docetaxel 36 mg/m(2) i.v. over 30 min on days 1, 8, and 15 and capecitabine 2,000 mg/d, in two divided doses, on days 5 to 21 of a 28-day cycle. Radiology-directed biopsies of the breast tumors were done at baseline and 5 days after the first dose of docetaxel to evaluate TP expression. Following DC therapy, patients had core breast biopsies, and if residual disease was present, received four cycles of standard dose-dense doxorubin and cyclophosphamide (AC). RESULTS: The pCR rate was 26.9% (95% confidence interval, 11.6-47.8). Up-regulation of TP expression was not observed by either quantitative immunofluorescence (QIF) or immunohistochemistry. Radiology-directed core biopsy after neoadjuvant chemotherapy accurately predicted pathologic response in 88% (95% confidence interval, 69.8-97.6) of the cases. Neither level of TP expression nor TP up-regulation correlated with pCR. Significant toxicity resulted in therapy discontinuation in 3 of 26 patients. CONCLUSIONS: DC chemotherapy exhibited a similar pCR rate compared with standard taxane regimens, with increased toxicity. TP expression was not up-regulated after docetaxel and did not correlate with therapeutic response. Core breast biopsy after neoadjuvant chemotherapy accurately predicted pathologic response.

Correlations between cyclooxygenase-2 expression and angiogenic factors in primary tumors and liver metastases in colorectal cancer.

BACKGROUND: Angiogenesis is required for growth and metastasis of colorectal cancer (CRC), and several positive regulators of tumor angiogenesis have been identified. Cyclooxygenase-2 (COX-2), known to be elevated in several human cancers, regulates angiogenesis by inducing angiogenic factors. The aim of this study was to clarify the levels and evaluate the relationships of COX-2, vascular endothelial growth factor A and C, thymidine phosphorylase (TP) and microvascular density (MVD) in paired tissue specimens between primary CRC and corresponding metastatic liver cancer. METHODS: Tissue samples from pairs of primary tumors and corresponding metastatic liver tumors from 44 patients with CRC were immunohistochemically evaluated for COX-2, VEGF-A, VEGF-C, TP and MVD. RESULTS: The primary and corresponding metastatic liver tumors tended to show concordant immunoreactivity for COX-2 (P = 0.005, rs = 0.428), VEGF-A (P = 0.039, rs = 0.314), TP (P = 0.005, rs = 0.422) and MVD (P = 0.046, rs = 0.304) by Spearman rank test. The rate of COX-2 immunoreactivity was higher in liver metastases than in primary tumors (P = 0.002), while the rate of VEGF-A was higher in primary tumors than in liver metastases (P = 0.0004). The incidence of TP immunoreactivity and the level of MVD did not differ between primary and metastatic liver tumors (P = 0.247; P = 0.229). Significant correlations were found between COX-2 immunoreactivity and VEGF-A immunoreactivity in metastatic liver tumors (P = 0.033) as well as in primary tumors (P = 0.008). CONCLUSION: The positive correlations between COX-2, VEGF-A, TP and MVD in primary CRC and liver metastasis as demonstrated here will help to predict the angiogenic activity of liver metastasis by analyzing primary tumors, allowing for individualized cancer treatment options.

Thymidine phosphorylase expression as a prognostic marker for predicting recurrence in primary superficial bladder cancer.

No prognostic marker for predicting recurrence in primary superficial bladder cancer has been established. The aim of this study was to investigate the expression levels of thymidine phosphorylase (TP) by enzyme-linked immunosorbent assay (ELISA) and the immunohistological localization of TP in primary superficial bladder cancer tissue to assess its association with tumor recurrence and stage progression as well as the pathological parameters of the tumor. TP expression in cancer tissues from 77 patients was measured by sandwich-type ELISA. Clinicopathological factors and the clinical prognosis were examined in relation to the expression levels of TP. To clarify the clinicopathological implication of TP localization in primary superficial bladder cancer tissue, the immunohistochemically determined TP expression was assessed for histological grades 1-3. The TP expression in primary superficial bladder cancer significantly increased with the histological grade and stage. The TP expression in patients with a shift to invasive cancer was significantly higher than in those without invasive cancer. Patients with low TP expression had a significant longer postoperative tumor-free period than those with high TP expression (P=0.011). High TP expression was an independent prognostic factor for tumor recurrence (P=0.0441). Strong immunoreactivity for TP was observed in the cytoplasms of tumor cells and vascular endothelial cells. This study suggests that elevated TP expression may be a prognostic marker for predicting recurrence in primary superficial bladder cancer, and that high TP expression may be associated with the marked proliferation of tumor cells and increased vascular endothelial cells, showing strong immunoreactivity for TP.

Apoptosis induced by preoperative oral 5'-DFUR administration in gastric adenocarcinoma and its mechanism of action.

AIM: To study the apoptosis induced by preoperative oral 5'-DFUR administration in gastric adenocarcinoma and its mechanism of action. METHODS: Sixty gastric cancer patients were divided randomly into three groups (20 each group) before operation: group one:5'-DFUR oral administration at the dose of 800-1200 mg/d for 3 - 5 d, group two: 500 mg 5-FU + 200 mg/d CF by venous drip for 3 - 5 d,group three (control group). One or two days after chemotherapy, the patients were operated. Fas/FasL,PD-ECGF and PCNA were examined by immunohistochemistry and apoptotic tumor cells were detected by in situ TUNEL method. Fifty-four patients received gastrectomy, including 12 palliative resections and 42 radical resections. Six patients were excluded. Finally 18 cases in 5'-DFUR group, 16 cases in CF+5-FU group, and 20 cases in control group were analyzed. RESULTS: There was no significant difference in patient mean age, gender, white blood cell count, haematoglobin (HB),thromboplastin, perioperative complication incidence, radical or palliation resection, invasion depth (T), lymphonode involvement (N),metastasis (M) and TNM staging among the three groups. However,the PCNA index (PI) in 5'-DFUR group (40.51+/-12.62) and 5-FU+CF group (41.12+/-15.26) was significantly lower than that in control group (58.33+/-15.69) (F=9.083, P=0.000). The apoptotic index (AI) in 5'-DFUR group (14.39+/-9.49) and 5-FU+CF group (14.11+/-9.68)was significantly higher than that in control group (6.88+/-7.37) (F=4.409, P=0.017).The expression rates of Fas and FasL in group one and group three were 66.7% (12/18) and 50% (9/18), 43.8% (7/16) and 81.3% (13/16), 45.0% (9/20) and 85% (17/20), respectively. The expression rate of FasL in 5'-DFUR group was significantly lower than that in the other two groups (chi2=6.708, P=0.035). Meanwhile, the expression rate of PD-ECGF was significantly lower in 5'-DFUR group (4/18,28.6%) than in CF+5-FU group(9/16,56.3%)and control group (13/20,65.0%) (chi2=7.542, P=0.023). The frequency of Fas expression was significantly correlated with palliative or radical resection (chi2=7.651, P=0.006), invasion depth (chi2=8.927, P=0.003), lymphatic spread (chi2=4.488, P=0.034) and UICC stages (chi2=8.063, P=0.045) respectively. By the end of March 2005,45 patients were followed up. The 0.5-, 1-, 2-, 3-year survival rates were 96%,73%,60%,48%, respectively, which were related with T, N, M and Fas expression, but not with PD-ECGF and FasL expression. CONCLUSION: Preoperative oral 5'-DFUR administration may induce apoptosis of gastric carcinoma cells and decrease tumor cell proliferation index,but cannot improve the prognosis of patients with gastric cancer.Down-regulation of FasL and PD-ECGF expression mediated by 5'-DFUR may be one of its anti-cancer mechanisms.Fas expression correlates with the progression of gastric carcinoma and may be an effective prognostic factor.

Evaluation of tumor angiogenesis and thymidine phosphorylase tissue expression in patients with endometrial cancer.

The formation of new blood vessels in endometrial cancer tissue is a main process, which leads to tumor progression, and is connected with tumor expansion and invasiveness. The aim of the study was evaluation of thymidine phosphorylase protein (TP) expression in human endometrial cancer cells by immunohistochemistry and comparison obtained data with intensity of angiogenesis process and clinicopathological factors as FIGO stage of disease and histopathologic grade. Endometrial cancer specimens were obtained from 55 postmenopausal patients (aged 52 to 74 years) underwent total abdominal hysterectomy with bilateral salpingo-oophorectomy. None of patients received preoperative pelvic irradiation. Histopathological typing and grading of the endometrial tumors (G-1, G-2, G-3) as well as myometrial invasion (<1/2, >1/2) were assessed using standard criteria, on hematoxylin-eosin sections. At the surgery, FIGO clinical stage of disease was determined. Thymidine phosphorylase overexpression was observed in 23 of 55 (41.8%) cases of endometrial cancer. Although we found no statistically significant differences in TP expression between histopathologic grades, particular FIGO stages showed a significant trend of increase TP tumor overexpression. Thymidine phosphorylase overexpression cases demonstrate higher intensity of angiogenesis in comparison to negative samples and results are statistically significant for t-test (p<0.0001). The most intensive new blood vessel formation was observed in G-2 of tumor differentiation grade (p=0.013 for ANOVA test) Mean angiogenic points density (APD) values in cases of G-1 histopathologic grade reached 135.7; values of G-2 and G-3 grades reached 213.8 and 162.8, respectively. Mean intensity of angiogenesis in the first FIGO stage of disease reached 160.0 APD, in stage II 205.6 APD, and in the third 286.9, respectively. Angiogenesis was more intensive in cases of advanced tumors - analysis of variance (ANOVA) confirmed statistically significant differences in APD values between FIGO stage groups (p=0.0007). In conclusion, thymidine phosphorylase expression correlates with increased microvessel density in endometrial cancer. The intensity of angiogenesis process increases according to FIGO stage of disease, which is connected with progressing of cancer disease. Thymidine phosphorylase can play an important role in endometrial cancer progression and could offer additional information about advance of disease.