Biomedical subjects

Pamela J Maxwell

Publications and source records attributed to Pamela J Maxwell.

4 recordsLinked to original sources

PubMed2003

Identification of 5-fluorouracil-inducible target genes using cDNA microarray profiling.

The fluoropyrimidine 5-Fluorouracil (5-FU) is widely used in the treatment of cancer. To identify novel downstream mediators of tumor cell response to 5-FU, we used DNA microarray technology to identify genes that are transcriptionally activated by 5-FU treatment in the MCF-7 breast cancer cell line. Of 2400 genes analyzed, 619 were up-regulated by >3-fold. Highly up-regulated genes (>6-fold) with signal intensities of >3000 were analyzed by Northern blot. Genes that were consistently found to be up-regulated were spermine/spermidine acetyl transferase (SSAT), annexin II, thymosin-beta-10, chaperonin-10, and MAT-8. Treatment of MCF-7 cells with the antifolate tomudex and DNA-damaging agent oxaliplatin also resulted in up-regulation of each of these targets. The 5-FU-induced activation of MAT-8, thymosin-beta-10, and chaperonin-10 was abrogated by inactivation of p53 in MCF-7 cells, whereas induction of SSAT and annexin II was significantly reduced in the absence of p53. Moreover, each of these genes contained more than one potential p53-binding site, suggesting that p53 may play an important regulatory role in 5-FU-induced expression of these genes. In addition, we found that basal expression levels of SSAT, annexin II, thymosin beta-10, and chaperonin-10 were increased (by approximately 2-3-fold), and MAT-8 expression dramatically increased (by approximately 10-fold) in a 5-FU-resistant colorectal cancer cell line (H630-R10) compared with the parental H630 cell line, suggesting these genes may be useful biomarkers of resistance. These results demonstrate the potential of DNA microarrays to identify novel genes involved in mediating the response of tumor cells to chemotherapy.

Antimetabolites, Antineoplastic↗

PubMed2003

The interaction of thymidylate synthase expression with p53-regulated signaling pathways in tumor cells.

Thymidylate synthase (TS) is a chemotherapeutic target for the fluoropyrimidine 5-fluorouracil (5-FU) and antifolate tomudex (TDX). Using the MCF-7 breast cancer line, we have developed a cell line with inducible TS expression termed M7TS90. Inducible TS expression in this line resulted in a moderate (approximately 3-fold) increase in 5-FU 50% inhibitory concentration at 72 hours (IC-50(72 h)) dose and a dramatic (approximately 24-fold) increase in the IC-50(72 h) dose of TDX, but did not affect chemosensitivity to cisplatin, oxaliplatin, irinotecan, and paclitaxel. In the absence of drug treatment, inducible TS expression had no effect on expression of the p53 tumor suppressor gene. However, TS induction abrogated p53, p21, Fas, and Bak induction in response to TDX, but not 5-FU. Similarly, downregulation of Bcl-2 was reversed by inducible TS expression in TDX, but not 5-FU-treated cells. Our results indicate that inducible TS expression in M7TS90 cells modulates p53 and p53 target gene expression in response to TDX, but not 5-FU.

Antimetabolites, Antineoplastic↗

PubMed2002

The role of thymidylate synthase induction in modulating p53-regulated gene expression in response to 5-fluorouracil and antifolates.

Thymidylate synthase (TS) is a critical target for chemotherapeutic agents such as 5-fluorouracil (5-FU) and antifolates such as tomudex (TDX),multitargeted antifolate, and ZD9331. Using the MCF-7 breast cancer line, we have developed p53 wild-type (M7TS90) and null (M7TS90-E6) isogenic lines with inducible TS expression (approximately 6-fold induction compared with control after 48 h). In the M7TS90 line, inducible TS expression resulted in a moderate approximately 3-fold increase in 5-FU IC-50(72 h) dose and a dramatic >20-fold increase in the IC-50(72 h) doses of TDX, multitargeted antifolate, and ZD9331. S-phase cell cycle arrest and apoptosis induced by the antifolates were abrogated by TS induction. In contrast, cell cycle arrest and apoptosis induced by 5-FU was unaffected by TS expression levels. Inactivation of p53 significantly increased resistance to 5-FU and the antifolates with IC-50(72 h) doses for 5-FU and TDX of >100 and >10 microM, respectively, in the M7TS90-E6 cell line. Furthermore, p53 inactivation completely abrogated the cell cycle arrest and apoptosis induced by 5-FU. The antifolates induced S-phase arrest in the p53 null cell line; however, the induction of apoptosis by these agents was significantly reduced compared with p53 wild-type cells. Both inducible TS expression and the addition of exogenous thymidine (10 microM) blocked p53 and p21 induction by the antifolates but not by 5-FU in the M7TS90 cell line. Similarly, inducible TS expression and exogenous thymidine abrogated antifolate but not 5-FU-mediated up-regulation of Fas/CD95 in M7TS90 cells. Our results indicate that in M7TS90 cells, inducible TS expression modulates p53 and p53 target gene expression in response to TS-targeted antifolate therapies but not to 5-FU.

Breast Neoplasms↗

PubMedDate not supplied

5-Fluorouracil: identification of novel downstream mediators of tumour response.

BACKGROUND: 5-Fluorouracil (5-FU) is routinely used in the treatment of gastrointestinal, breast and head and neck cancers. A major limitation to the use of this drug is acquired or inherent resistance. MATERIALS AND METHODS: To examine the downstream molecular signals activated in response to 5-FU, we used DNA microarray technology to examine global transcriptional changes in 5-FU-treated MCF-7 breast cancer cells. RESULTS: We identified several novel 5-FU-inducible target genes that have not previously been linked to 5-FU response, including spermine/spermidine acetyl transferase (SSAT) and annexin II. Treatment of MCF-7 cells with the antifolate tomudex (TDX) and the DNA damaging agent oxaliplatin also caused up-regulation of each target gene. Inactivation of wild-type p53 abrogated the 5-FU-mediated induction of SSAT and annexin II. Inducible expression of thymidylate synthase completely abrogated TDX-, but not 5-FU-mediated induction of each gene. Furthermore, basal expression of SSAT and annexin II was elevated in cells resistant to 5-FU. CONCLUSION: These data demonstrate the potential of microarray analysis to identify novel genes associated with response or resistance to chemotherapeutic agents.

Acetyltransferases↗