Blood mRNA expression of REV3L and TYMS as potential predictive biomarkers from platinum-based chemotherapy plus pemetrexed in non-small cell lung cancer patients.

PURPOSE: Therapeutic options for cancer patients have increased in the last years, although drugs resistance problem remains unresolved. Genetic background in individual susceptibility to cancer treatment could influence the therapy responses. The aim of this study was to explore the feasibility of using blood 4 genes (AEG-1, BRCA-1, REV3L and TYMS) expression levels as a predictor of the efficacy of pemetrexed therapy in patients with advanced non-small cell lung cancer. METHODS: Sixteen patients from the Medical Oncology Department at "12 de Octubre" Hospital, were included in the study. Total mRNA was isolated from blood samples, and gene expression was analyzed by RT-qPCR. A panel of lung tumor cell lines were used in cell proliferation tests and siRNA-mediated silencing assays. RESULTS: Similarity between blood gene expression levels and protein expression in matched tumor tissue was observed in 54.54% (REV3L) and 81.81% (TYMS) of cases. Gene expression of REV3L and TYMS in blood correlated directly and inversely, respectively, with progression-free survival and overall survival in the patients from our cohort. In tumor cell lines, the knockdown of REV3L conferred resistance to pemetrexed treatment, and the TYMS silencing increased the pemetrexed sensitivity of tumor cells. CONCLUSIONS: The use of peripheral blood samples for expression quantification of interest genes is an affordable method with promising results in the evaluation of response to pemetrexed treatment. Therefore, expression levels of REV3L and TYMS genes might be used as predictive biomarkers in advanced NSCLC patients.

Molecular and Kinetic Analyses of Circulating Tumor Cells as Predictive Markers of Treatment Response in Locally Advanced Rectal Cancer Patients.

Neoadjuvant chemoradiation (NCRT) followed by total mesorectal excision is the standard treatment for locally advanced rectal cancer (LARC). To justify a non-surgical approach, identification of pathologic complete response (pCR) is required. Analysis of circulating tumor cells (CTCs) can be used to evaluate pCR. We hypothesize that monitoring of thymidylate synthase (TYMS) and excision repair protein, RAD23 homolog B (RAD23B), can be used to predict resistance to chemotherapy/radiotherapy. Therefore, the aims of this study were to analyze CTCs from patients with LARC who underwent NCRT plus surgery for expression of TYMS/RAD23B and to evaluate their predictive value. Blood samples from 30 patients were collected prior to NCRT (S1) and prior to surgery (S2). CTCs were isolated and quantified by ISET®, proteins were analyzed by immunocytochemistry, and TYMS mRNA was detected by chromogenic in situ hybridization. CTC counts decreased between S1 and S2 in patients exhibiting pCR (p = 0.02) or partial response (p = 0.01). Regarding protein expression, TYMS was absent in 100% of CTCs from patients with pCR (p = 0.001) yet was expressed in 83% of non-responders at S2 (p < 0.001). Meanwhile, RAD23B was expressed in CTCs from 75% of non-responders at S1 (p = 0.01) and in 100% of non-responders at S2 (p = 0.001). Surprisingly, 100% of non-responders expressed TYMS mRNA at both timepoints (p = 0.001). In addition, TYMS/RAD23B was not detected in the CTCs of patients exhibiting pCR (p = 0.001). We found 83.3% of sensitivity for TYMS mRNA at S1 (p = 0.001) and 100% for TYMS (p = 0.064) and RAD23B (p = 0.01) protein expression at S2. Thus, TYMS mRNA and/or TYMS/RAD23B expression in CTCs, as well as CTC kinetics, have the potential to predict non-response to NCRT and avoid unnecessary radical surgery for LARC patients with pCR.

Autoantibodies against TYMS and PDLIM1 proteins detected as circulatory signatures in Indian breast cancer patients.

PURPOSE: Breast cancer (BC) is the most common invasive cancer in women worldwide. Autoantibodies (AAbs) to tumor-associated antigens (TAAs) have a great potential for the development of diagnostic biomarkers in cancer. This study was performed to identify AAbs and cognate TAAs that may improve detection of this deadly disease. EXPERIMENTAL DESIGN: Serological proteome analysis of plasma samples of BC patients (N = 30) and healthy controls (N = 30) was performed to identify TAAs. Expressions of selected TAAs were also determined in breast tumor tissues (N = 10) by immunohistochemistry. An independent validation cohort (N = 124) was tested to determine diagnostic accuracy of selected AAbs titer by ELISA. RESULTS: Thymidylate synthase (TYMS) and C-terminal LIM domain protein 1 (PDLIM1) were found to react more specifically with plasma samples of BC patients. Both TAAs were also found to be significantly over expressed (p < 0.001) in breast tumor tissues compared to adjacent normal tissues. TYMS AAbs response was positively correlated (r = 0.778, p < 0.008) with TYMS overexpression in BC tissues. TYMS and PDLIM1 AAbs titers discriminated BC from controls with a sensitivity/specificity of 57.81%/95% and 73.44%/58.33%, respectively. CONCLUSION AND CLINICAL RELEVANCE: High titers of both TYMS and PDLIM1 AAbs were significantly more prevalent in BC cases than controls. Our data recommends further investigations for evaluating their potential for BC detection.

A phase 1 clinical trial of sequential pralatrexate followed by a 48-hour infusion of 5-fluorouracil given every other week in adult patients with solid tumors.

BACKGROUND: Pralatrexate (PDX) is an inhibitor of dihydrofolate reductase that was rationally designed to improve cellular uptake and retention of the drug. Preclinical data have shown synergy with the sequential administration of a dihydrofolate reductase inhibitor followed 24 hours later by 5-fluorouracil (5-FU). METHODS: Twenty-seven patients were enrolled at 1 of 5 PDX dose levels from 75 to 185 mg/m(2) on day 1 followed 24 hours later by 5-FU at a dose of 3000 mg/m(2) /48 hours every 2 weeks with folic acid and vitamin B12 supplementation. Baseline blood was collected for pharmacogenetic analysis of polymorphisms of methylenetetrahydrofolate reductase and thymidylate synthase. RESULTS: Mucositis was the most common dose-limiting toxicity. When the worst toxicities across all cycles were considered, grade 3 to 4 neutropenia, anemia, and thrombocytopenia were found to have occurred in 14.8%, 14.8%, and 0% of patients, respectively. Grade 2 to 3 toxicities included mucositis (66.6%), dehydration (33.3%), fatigue (25.9%), and diarrhea (22.2%). Version 3.0 of the National Cancer Institute Common Toxicity Criteria was used to grade toxicities The median progression-free survival (PFS) was 112 days (range, 28-588 days). Seven patients (26%) had a PFS of >180 days (5 patients with colorectal cancer, 1 patient with pancreatic cancer, and 1 patient with non-small cell lung cancer). Polymorphisms in methylenetetrahydrofolate reductase and thymidylate synthase did not correlate with toxicity. CONCLUSIONS: The recommended dose of PDX was 148 mg/m(2) . A subset of heavily pretreated patients had PFS durations of ≥6 months with this regimen.

Prognostic impact of blood biomarkers TS and DPD in neoadjuvant-treated esophageal cancer patients.

BACKGROUND/AIM: The prognostic value of TS (thymidylate synthase) and DPD (dihydropyrimidine dehydrogenase) RNA expression in the blood of patients with esophageal cancer is not known. The aim of the present study was to evaluate the significance of these molecular alterations in the blood as a prognostic marker for patients with neoadjuvant-treated esophageal cancer. PATIENTS AND METHODS: A total of 29 patients with locally advanced esophageal cancer (cT3-T4, Nx, M0) were enrolled in this prospective study. All patients received neoadjuvant chemoradiation followed by a transthoracic resection (curative transthoracic en bloc esophagectomy, RO). Peripheral blood samples were drawn before initiation of therapy. The analysis was performed using quantitative real-time-polymerase chain reaction (RT-PCR). The histomorphological regressions grading after neoadjuvant therapy was defined as follows: major response (MaR)=less than 10% vital tumor tissue, minor response (MiR)=more than 10% vital tumor tissue. RESULTS: Nineteen out of 29 patients (65.5%) had a MiR and 10 (34.5%) had a MaR. The median survival of patients was 2.08 years (range=0.15-4.53). Among the tested genes, the RNA expression of TS was significantly associated with prognosis of patients. Patients with TS expression above 0.78 had a median survival of 1.1 years (range=0.21-3.96) compared to 2.6 years (range=0.15 to 4.53) in patients with TS expression lower than 0.78 (p=0.031, log rank test). There was no association between clinical variables (e.g., tumor stage, gender, age, etc.) and the RNA expression of TS in the serum. CONCLUSION: The RNA expression of TS in the blood is a potential prognostic marker in patients with neoadjuvant-treated esophageal cancer. The significance of these molecular alterations as non-invasive prognostic marker for esophageal cancer should be evaluated in prospective studies.

Evaluation of antigen detection tests, microscopy, and polymerase chain reaction for diagnosis of malaria in peripheral blood in asymptomatic pregnant women in Nanoro, Burkina Faso.

Rapid diagnostics tests (RDTs) detect malaria specific antigen(s) in the circulation, even when parasites are sequestered in the placenta and not visible by microscopy. However, research on their diagnostic accuracy during pregnancy is limited. Pregnant women (n = 418) were screened for malaria during routine antenatal care by using two RDTs that detect histidine-rich protein 2 (HRP2) or Plasmodium lactate dehydrogenase, and enzyme-linked immunosorbent assays with antibodies that detect dihydrofolate reductase-thymidylate synthase or heme-detoxification protein, and compared with real-time polymerase chain reaction (RT-PCR) and microscopy for evaluation of their diagnostic accuracy. Prevalence of malaria infection was high (53% by PCR). The RT-PCR and the HRP2 RDT detected most cases of malaria during pregnancy, whereas microscopy, the Plasmodium lactate dehydrogenase RDT, and enzyme-linked immunosorbent assays for dihydrofolate reductase-thymidylate synthase and heme-detoxification protein antibodies did not detect several low-density infections. Therefore, the HRP2 RDT could be a useful tool in high-transmission areas for diagnosis of malaria in asymptomatic pregnant women.

Antigen persistence of rapid diagnostic tests in pregnant women in Nanoro, Burkina Faso, and the implications for the diagnosis of malaria in pregnancy.

OBJECTIVES: To evaluate persistence of several Plasmodium antigens in pregnant women after treatment and compare diagnostics during treatment follow-up. METHODS: Thirty-two pregnant women (N = 32) with confirmed malaria infection by a histidine-rich protein 2 (HRP2)-based rapid diagnostic test (RDT) and microscopy were followed for 28 days after artemisinin-based combination therapy (ACT). A Plasmodium lactate dehydrogenase (pLDH)-based RDT and two ELISAs based on the detection of dihydrofolate reductase-thymidylate synthase (DHFR-TS) and haeme detoxification protein (HDP) were compared with each other and to RT-PCR at each visit. RESULTS: The mean visit number (95% confidence interval) on which the HRP2-based RDT was still positive after treatment was 3.4 (2.7-4.1) visits with some patients still positive at day 28. This is significantly later than the pLDH-based RDT [0.84 (0.55-1.1)], microscopy (median 1, range 1-3), DHFR-TS-ELISA [1.7 (1.1-2.3)] and RT-PCR (median 2, range 1-5) (P < 0.05), but not significantly later than HDP-ELISA [2.1 (1.6-2.7)]. Lower gravidity and higher parasite density at day 0 resulted in significantly longer positive results with most tests (P < 0.05). CONCLUSIONS: HRP2 can persist up to 28 days after ACT treatment; therefore, this test is not suitable for treatment follow-up in pregnant women and can generate problems when using this test during intermittent preventive treatment (IPTp). DHFR-TS is less persistent than HRP2, making it a potentially interesting target for diagnosis.

Roles of thymidylate synthase and dihydropyrimidine dehydrogenase expression in blood as predictors of response to multimodal therapy in esophageal cancer.

BACKGROUND: Thymidylate synthetase (TS) and dihydropyrimidine dehydrogenase (DPD) RNA expression in peripheral blood was examined as a noninvasive molecular predictor of response to neoadjuvant radiochemotherapy in patients with locally advanced cancer of the esophagus. METHODS: Blood samples were drawn from 29 patients with esophageal cancer (10 squamous cell carcinomas and 19 adenocarciomas) before neoadjuvant radiochemotherapy. After extraction of cellular tumor RNA from blood samples, quantitative expression analysis of TS and DPD was performed with quantitative real-time reverse-transcription polymerase chain reaction. RESULTS: Twenty of 29 (68%) of patients had a minor histopathologic response, and 9 of 29 (32%) had a major response to neadjuvant radiochemotherapy. RNA expression in the blood of patients was detectable for TS in 86%, for DPD in 97%, and in 100% for ß-actin. No significant associations were detected between TS and DPD expression levels and clinical variables of the patients. A high expression level for TS was associated with a minor response to neoadjuvant treatment (P = .046), while there was no significant association between DPD and response to therapy. Combined analysis of TS and DPD expression increased the specificity for the prediction of response to 100%. No major responder to therapy had high expression levels for both genes in their peripheral blood. CONCLUSION: Quantitation of TS and DPD in peripheral blood may be a highly specific analysis to identify a subset of patients who do not respond to neoadjuvant radiochemotherapy and may therefore prevent expensive, noneffective, and potentially harmful therapies in a substantial number of patients with esophageal cancer.

A predictive factor for the response to S-1 plus cisplatin in gastric cancer.

AIM: To prove that the protein expression level of thymidylate synthase is a predictive factor for the response to S-1/cisplatin (CDDP) chemotherapy in gastric cancer. METHODS: We measured the protein expression levels of thymidylate synthase (TS), dihydropyrimidine dehydrogenase (DPD), and orotate phosphoribosyltransferase (OPRT) in advanced gastric cancer. Before S-1/CDDP chemotherapy, tumor specimens from primary sites were obtained by endoscopic biopsy and analyzed by enzyme-linked immunosorbent assay. The chemotherapeutic effects on the primary sites were evaluated by endoscopic biopsy performed more than once after S-1/CDDP chemotherapy. The effects are a predictive factor for the response to S-1/CDDP chemotherapy in patients with advanced gastric cancer, as evaluated by endoscopic biopsy over time. RESULTS: The protein expression level of TS was significantly higher (P < 0.05) in the tumor than in the normal tissue, and significantly lower (P < 0.05) in the responders than in the non-responders. We were able to evaluate the correlation between changes in the protein expression levels of TS, DPD and OPRT and chemotherapeutic responses in 7 patients by assessing tumor tissues more than twice. In the responders, the protein expression level of TS was < 40 ng/mg protein. However, there were significant increases in the protein expression levels of TS (P < 0.01) and DPD (P < 0.05) after chemotherapy in 3 patients. In these cases, the patient assessment changed from "responder" to "non-responder". In the non-responders, the protein expression level of TS was > 40 ng/mg protein. CONCLUSION: We have confirmed that the protein expression level of TS is a predictive factor for the response to S-1/CDDP chemotherapy in patients with advanced gastric cancer.

Biomarkers for response to neoadjuvant chemoradiation for rectal cancer.

Locally advanced rectal cancer (LARC) is currently treated with neoadjuvant chemoradiation. Although approximately 45% of patients respond to neoadjuvant therapy with T-level downstaging, there is no effective method of predicting which patients will respond. Molecular biomarkers have been investigated for their ability to predict outcome in LARC treated with neoadjuvant chemotherapy and radiation. A literature search using PubMed resulted in the initial assessment of 1,204 articles. Articles addressing the ability of a biomarker to predict outcome for LARC treated with neoadjuvant chemotherapy and radiation were included. Six biomarkers met the criteria for review: p53, epidermal growth factor receptor (EGFR), thymidylate synthase, Ki-67, p21, and bcl-2/bax. On the basis of composite data, p53 is unlikely to have utility as a predictor of response. Epidermal growth factor receptor has shown promise as a predictor when quantitatively evaluated in pretreatment biopsies or when EGFR polymorphisms are evaluated in germline DNA. Thymidylate synthase, when evaluated for polymorphisms in germline DNA, is promising as a predictive biomarker. Ki-67 and bcl-2 are not useful in predicting outcome. p21 needs to be further evaluated to determine its usefulness in predicting outcome. Bax requires more investigation to determine its usefulness. Epidermal growth factor receptor, thymidylate synthase, and p21 should be evaluated in larger prospective clinical trials for their ability to guide preoperative therapy choices in LARC.

Free circulating mRNA in plasma from breast cancer patients and clinical outcome.

We studied by real-time PCR cyclin D1 and thymidylate synthase (TS) mRNA in plasma as possible markers of clinical outcome in breast cancer. We observed poor outcome in patients with presence of cyclin D1 mRNA in good-prognosis groups, such as negative vascular invasion. Presence of both markers was associated with non-response to treatment after relapse. In patients treated with tamoxifen, a trend to significant relation between poor outcome and cyclin D1 mRNA was found. Cyclin D1 mRNA in plasma could identify patients with poor overall survival in good-prognosis groups and patients non-responsive to tamoxifen.

Arsenic trioxide/ascorbic acid therapy in patients with refractory metastatic colorectal carcinoma: a clinical experience.

Arsenic trioxide (As2O3) has demonstrated effectiveness in treating acute promyelocytic leukemia (APL). Therefore the FDA has approved it to treat APL. In patients with refractory metastatic colorectal carcinoma (CRC), we assessed the efficacy and toxicity of As2O3/AA (ascorbic acid) as the outcome of this trial. Five patients with refractory metastatic CRC who failed all previous standard chemotherapy were enrolled in this study. They were treated with 0.25 mg/kg body weight/day As2O3 and 1000 mg/day of ascorbic acid for 5 days a week for 5 weeks. Each treatment cycle extended for 7 weeks with 5 weeks of treatment and 2 weeks of rest. All the patients developed moderate to severe toxic side effects to arsenic trioxide/AA therapy and therefore the study was discontinued. No CR (complete remission) or PR (partial remission) was observed. CT scans demonstrated stable or progressive disease. Three of the five patients died within 2 to 5 months after cessation of the therapy. None of the deaths could be related to this clinical trial. Two years of follow-up study showed that two patients were alive with stable disease. Under the current treatment regimen all patients developed moderate to severe side effects with no clinically measurable activity. As an alternate, efforts may be made to reduce the dose and arsenic trioxide may be combined with other standard regimen in reversing the chemo resistance.

Thymidylate synthase messenger RNA expression in plasma from patients with colon cancer: prognostic potential.

PURPOSE: Thymidylate synthase (TS), a critical target in fluorouracil-based chemotherapy, is a prognostic marker in colon carcinomas and a predictor of response to treatment. Tumor RNA has been detected in plasma from cancer patients and is associated with poor prognosis. This is the first study to examine extracellular TS mRNA in plasma from patients with colon carcinoma, and its possible relation with TS promoter enhancer region (TSER) polymorphism. EXPERIMENTAL DESIGN: TS expression was measured in plasma from 88 patients and 26 controls, and in a tumor subgroup of this series by quantitative PCR. Genotyping for TSER polymorphism was done in 60 patients. Clinicopathologic variables were correlated with these molecular changes. RESULTS: TS mRNA was detected in plasma in 47% of patients, showing significant differences from healthy controls. Patients with TS mRNA in plasma had higher levels of TS in tumor tissue than patients without. The presence of TS mRNA was associated with lymph node metastases and more advanced stages. Polymorphism TSER 3/3 was found in 38% of cases, and was significantly correlated with high amounts of TS mRNA in plasma. CONCLUSIONS: Our results suggest that TS mRNA in plasma originated from tumors, it may indicate poor prognosis and might help to classify tumors in Dukes' stages B and C. The TSER genotype may influence TS mRNA expression in plasma.

Influence of combined methylenetetrahydrofolate reductase (MTHFR) and thymidylate synthase enhancer region (TSER) polymorphisms to plasma homocysteine levels in Korean patients with recurrent spontaneous abortion.

OBJECTIVES: Methylenetetrahydrofolate reductase (MTHFR) mutations known to be associated with hyperhomocysteinemia may be a risk factor for recurrent spontaneous abortion. Recently 28-bp tandem repeat polymorphism in thymidylate synthase enhancer region (TSER) was reported to affect plasma homocysteine level. We investigated the association between plasma homocysteine level and MTHFR and TSER genotypes. METHODS: Plasma homocysteine level was measured by fluorescent polarizing immunoassay. MTHFR mutations (C677T and A1298C) were identified by PCR-restriction fragment length polymorphism assay. TSER mutation was analyzed by PCR method. RESULTS: Average homocysteine level was significantly higher in MTHFR 677TT genotype (9.80+/-3.87 micromol/L) than in MTHFR 677CT (7.04+/-1.99 micromol/L) in MTHFR 677CC genotype (8.14+/-1.74 micromol/L) in Korean patients with unexplained recurrent spontaneous abortion (p=0.0143). While MTHFR 1298AA showed the highest level, plasma homocysteine levels were not significantly different among MTHFR 1298AA (8.42+/-2.65 micromol/L), 1298AC (6.98+/-2.44 micromol/L) and 1298CC (6.09+/-0.32 micromol/L) (p=0.2058). There was no significant difference among TSER genotypes (2R2R, 8.61+/-1.68 micromol/L; 2R3R, 7.84+/-2.16 micromol/L; 3R3R, 8.05+/-2.81 micromol/L; p=0.9319). Among the combined genotypes of MTHFR C677T and TSER, 677TT-3R3R genotype had the highest homocysteine level (11.47+/-4.66 micromol/L). 1298AA-3R3R had the highest level (8.54+/-3.05 micromol/L) among the combined genotypes of MTHFR A1298C and TSER. CONCLUSION: Although there was no significant difference found among combined genotypes, 3R3R showed elevated homocysteine levels in MTHFR 677TT and 1298AA in Korean patients with unexplained recurrent spontaneous abortion. Thus TSER polymorphism may be a genetic determinant of plasma homocysteine level in Korean patients as well as MTHFR C677T polymorphism.

Recurrence and 5-FU sensitivity of stage III/Dukes' C colorectal cancer with occult neoplastic cells in lymph node sinuses.

In 72 patients without occult neoplastic cells (ONCs) in their lymph node sinuses, the 5-year relapse-free survival (RFS) rate and overall survival (OS) rate were 71.3% and 69.2%, respectively. In 33 patients with ONCs, the 5-year RFS rate and OS rate were 33.9% and 31.3%, respectively. There was a marked difference of survival between the two groups (p=0.0001 and p=0.0003). The metastatic lymph nodes of the 33 ONC-positive patients had high and low levels of thymidilate synthase (TS) expression in 38.1% (8/21) and 61.9% (13/21) of the recurrence group (n=21), respectively, while high and low levels of dihydropyrimidine dehydrogenase (DPD) expression were found in 38.1% (8/21) and 61.9% (13/21), respectively. In the non-recurrence group (n=12), high and low levels of TS or DPD expression were detected in 58.3% (7/12) and 41.7% (5/12) versus 16.7% (2/12) and 83.3% (10/12), respectively. Patients with high TS and low DPD expression accounted for 9.5% (2/21) of the recurrence group and 50.0% (6/12) of the non-recurrence group (p<0.01). These results suggest that ONCs are clearly associated with the 5-year RFS and OS rates. Unlike the non-recurrence group, the recurrence group of ONC-positive patients with Dukes' C colorectal cancer is unlikely to respond well to treatment with 5-FU plus LV and require combination chemotherapy based on CPT-11 and/or L-OHP.

Recurrence and 5-FU sensitivity of stage II/Dukes' B colorectal cancer with occult neoplastic cells in lymph node sinuses.

In 103 patients without occult neoplastic cells (ONCs) in the lymph node sinuses, the 5-year relapse-free survival (RFS) rate and overall survival (OS) rate were 90.2% and 91.8%, respectively. In 21 patients with ONCs, the 5-year RFS and OS rates were 34.9% and 62.3%, respectively. There were marked differences of survival between the two groups (p=0.0000 and p=0.0003). In the primary tumors of the 21 ONC-positive patients, high and low TS levels were found in 46.2% (6/13) and 53.8% (7/13) of the recurrence group (n=13), respectively. High and low DPD levels were found in 23.1% (3/13) and 76.9% (10/13), respectively. In the non-recurrence group (n=8), high and low TS or DPD levels were found in 75.0% (6/8) and 25.0% (2/8) versus 12.5% (1/8) and 87.5% (7/8), respectively. The percentage of patients with high TS and low DPD levels was 23.1% (3/13) in the recurrence group and 62.5% (5/8) in the non-recurrence group (p=0.07). These results suggest that the presence of ONCs had a clear association with the 5-year RFS and OS rates. The recurrence group of ONC-positive patients with stage II/Dukes' B colorectal cancer was unlikely to be highly responsive to 5-FU-based treatment, thus requiring multi-combination chemotherapy using CPT-11 and/or L-OHP.

The thymidylate synthase tandem repeat polymorphism is not associated with homocysteine concentrations in healthy young subjects.

Thymidylate synthase (TYMS) and 5,10-methylenetetrahydrofolate reductase (MTHFR) may compete for their common cofactor 5,10-methylenetetrahyhdrofolate (5,10-meTHF). Limiting 5,10-meTHF results in elevated homocysteine, especially in individuals homozygous for the T allele of the MTHFR C677T polymorphism. The TYMS gene has a tandem repeat polymorphism (two repeats or three repeats, designated 2R or 3R, respectively), which may also affect homocysteine concentrations. The 3R allele is subject to increased translational efficiency in vitro and the 3R3R genotype is associated with both decreased serum folate and elevated plasma homocysteine (tHcy) in a population of Singapore Chinese. We assessed the relationship between TYMS genotype and key biochemical and genetic variables in a random sample of 392 healthy young Northwestern European men and women. The tHcy concentrations for 3R3R homozygotes (median 8.5 micromol/l) did not differ significantly from those for 2R2R homozygotes (median 8.7 micromol/l) or 2R3R heterozygotes (median 9.3 micromol/l) in the population as a whole (P=0.43), or in subsets of subjects with low serum folate (P=0.60) or the MTHFR 677TT genotype (P=0.90). Furthermore, there was no trend toward elevated tHcy in 3R3R homozygotes. Similarly, the TYMS tandem repeat polymorphism was not associated with serum folate concentrations. Our findings indicate that the TYMS 3R3R genotype is not a determinant of homocysteine in this sample of healthy young Caucasian adults from Northern Ireland.

Human colorectal malignancy and oral UFT.

Thymidylate synthase (EC 2.1.1.45) and thymidine kinase (EC 2.71.21) are key enzymes involved in de novo and salvage pathways for pyrimidine nucleotide synthesis. Both enzyme activities are increased in rapidly proliferating normal, fetal and neoplastic tissues. In a previous study, the activities of thymidylate synthase and thymidine kinase were relatively predominant in the poorly-and well-differentiated types of a gastric cancer. In the present study of patients with colorectal cancer, the serum thymidine kinase activities are elevated in cases at a clinically late stage, and in cases with a recurrence and a distant metastasis associated with abundant blood supply, i.e. metastasis to the liver, lung and bone. Well-differentiated colorectal cancer shows higher thymidine kinase activity than moderately-well-differentiated type as was previously shown in gastric cancer patients. Furthermore, neoadjuvant chemotherapy using the 5-fluorouracil derivative UFT demonstrates a stronger suppression of increased activities of thymidylate synthase in the tumorous tissues than in the non-tumorous mucosa.