Biomedical subjects

Paul J Goodfellow

Publications and source records attributed to Paul J Goodfellow.

24 records · Page 2Linked to original sources

PubMed2003

A panel of repeat markers for detection of microsatellite instability in murine tumors.

A substantial fraction of human malignancies lack functional DNA mismatch repair (MMR), accumulate mutations at high frequency, and exhibit microsatellite instability (MSI). In order to distinguish between MMR-competent and MMR-deficient malignancies, a consensus panel of microsatellite repeats has been adopted for MSI analysis of human tumors. There is no reference panel of repeats for MSI typing of murine malignancies. In this study, we present six new microsatellite repeat markers for MSI typing of mouse tumors. Analysis of polymerase chain reaction (PCR)-amplified tumor DNA from MMR-deficient and -proficient mice on denaturing polyacrylamide gels revealed that the new panel of microsatellites was more sensitive in detecting MSI than six commonly used CA(n) repeats. Using the new set of microsatellite markers, we demonstrated the presence of MSI in endometrial carcinoma and cancer precursors from diethylstilbestrol (DES)-treated mice, pointing to a possible role for loss of MMR in hormonally promoted endometrial tumorigenesis.

Adaptor Proteins, Signal Transducing↗

PubMed2003

Expression profiling of mouse endometrial cancers microdissected from ethanol-fixed, paraffin-embedded tissues.

Expression-profiling studies have helped define genetic changes associated with carcinogenesis. Determining which alterations in gene expression are causally associated with cancer and which result from the general dysregulation in gene expression that is characteristic of malignancies remains a problem. Transcriptional profiling of early lesions (small cancers or precancers) holds promise for identifying biologically important changes in gene expression. There are, however, technical barriers to the study of small tumors. The total number of cells available for analysis is limiting. It is also often difficult to distinguish cancer cells from normal proliferating cells in frozen sections that are typically used as a source of RNA. Here we describe an ethanol fixation and paraffin-embedding protocol that preserves tissue architecture and cellular morphology of the mouse endometrium, and allows for the recovery of high-quality RNA from microdissected cells. We performed GeneChip expression profiling using RNA from 800 to 4400 cells microdissected from ethanol-fixed, paraffin-embedded uteri. Endometrial adenocarcinomas exhibited changes in the levels of a number of messages known to be abnormally expressed in cancer, and differential expression of additional transcripts not previously implicated in carcinogenesis. We confirmed increased Amd1 expression in RNAs from mouse endometrial carcinomas that were hybridized to GeneChips and validated overexpression of this transcript in additional tumors.

Animals↗

PubMed2003

Antisense transcripts at the EMX2 locus in human and mouse.

The homeodomain transcription factor EMX2 is critical for central nervous system and urogenital development. In addition, EMX2 maps to a region of allelic deletion corresponding to a putative endometrial tumor suppressor at 10q26. We now report another polyadenylated transcript that is transcribed on the strand opposite to EMX2 and overlaps with the EMX2 transcript. This transcript was designated EMX2OS (OS, opposite strand), and an orthologous transcript present at the murine Emx2 locus was designated Emx2os. Alternative splicing to generate transcripts with varying 5' sequences was detected in the human but not the mouse. Neither ortholog contains a significant open reading frame, nor is primary sequence conserved between the two species. The sense and antisense transcripts display coordinate expression in that EMX2 and EMX2OS are abundant in normal postmenopausal endometrium, reduced in premenopausal endometrium, and absent or reduced in a majority of primary endometrial tumors. EMX2, EMX2OS, Emx2, and Emx2os are abundant in the uterine endometrium, with sense and antisense transcripts exhibiting identical expression patterns. Conservation of functional human and murine EMX2 antisense genes, of overlap between the sense and the antisense transcripts, and of identical cellular expression patterns suggests a biological function for EMX2OS, presumably to regulate EMX2.

Animals↗

PubMed2003

Frequent HOXA11 and THBS2 promoter methylation, and a methylator phenotype in endometrial adenocarcinoma.

PURPOSE: This study was designed to determine whether there is a methylator phenotype in stage I and II endometrioid endometrial adenocarcinoma, and if so, whether methylation correlates with recurrence. EXPERIMENTAL DESIGN: Bisulfite-converted DNAs from 24 stage I and II primary cancers (12 recurrent and 12 nonrecurrent), and 5 endometrial cancer cell lines were analyzed for methylation in the promoter regions of seven genes. A methylation index (MeI) was calculated for each tumor. Frequent HOXA11 and THBS2 methylation prompted analysis of case-matched bloods and 25 additional nonrecurrent primary cancers. Statistical analysis included Fisher's exact and Student t tests. RESULTS: Rates of methylation in the initial tumor series were as follows: HOXA11, 70.8%; THBS2, 62.5%; MLH1, 33.3%; CTNNB1, 16.7%; VDR, 4.2%; CDKN2A, 4.2%; and THBS1, 0%. There was no difference in the MeI of recurrent and nonrecurrent cases. However, cell lines had higher mean MeI. High rates of HOXA11 and THBS2 methylation were confirmed in the additional nonrecurrent tumors. None of the 24 case-matched bloods had HOXA11 methylation, whereas three blood DNAs showed THBS2 methylation. There was a statistically significant difference in the rate of HOXA11 methylation in recurrent and nonrecurrent tumors (P = 0.0167). CONCLUSIONS: Endometrial adenocarcinomas have a methylator phenotype. No correlation between MeI and clinicopathologic variables in early stage tumors was observed. High rates of methylation were found in the HOXA11 and THBS2 promoter regions. HOXA11 promoter methylation was significantly more frequent in recurrent than nonrecurrent cases. HOXA11 methylation in early stage endometrial cancer is associated with poor outcome.

Adenocarcinoma↗

PubMed2002

MSI in endometrial carcinoma: absence of MLH1 promoter methylation is associated with increased familial risk for cancers.

Loss of DNA mismatch repair occurs in a variety of malignancies and is associated with genome-wide instability of microsatellite repeats, a molecular phenotype referred to as microsatellite instability (MSI). MSI is a consistent feature of colorectal and endometrial tumors from patients with hereditary non-polyposis colorectal cancer (HNPCC). Sporadic colorectal and endometrial cancers that exhibit MSI frequently have methylation of the MLH1 promoter. We undertook a detailed family and medical history study to compare family cancer risk for women with MSI-positive and -negative endometrial cancers. The MLH1 promoter methylation status was determined for all cancers. Family histories were developed for 80 probands (40 with MSI-positive and 40 with MSI-negative tumors). The numbers of reported cancers in first- and second-degree relatives of the 2 groups were similar. There was a modest increase in familial cancer clustering for MSI-positive probands. When MSI-positive tumors were subclassified according to MLH1 promoter methylation, a clear association between methylation status and familial cancer risk was evident. Women with MSI-positive endometrial cancers in which the MLH1 promoter was unmethylated had a 7-fold relative risk (RR) of demonstrating familial clustering of cancers [RR 7.07 (95% confidence interval 2.29-21.81)]. The women with MSI-positive, MLH1-unmethylated tumors were significantly younger than the rest of the study population (56.1 years vs. 65.4, p < or = 0.01). Age of onset and tumor MSI not associated with MLH1 promoter methylation may point to women with a genetic susceptibility to malignancies.

Adaptor Proteins, Signal Transducing↗

PubMed2002

Ribosomal DNA methylation in patients with endometrial carcinoma: an independent prognostic marker.

BACKGROUND: Surgery is curative for the majority of patients with endometrial carcinoma: Consequently, the use of adjuvant therapy has been controversial. More effective methods to identify patients who are at risk for recurrence and who would benefit from adjuvant therapy are needed. The objective of this study was to investigate the prognostic significance of ribosomal DNA (rDNA) methylation in patients with endometrial carcinoma. METHODS: rDNA methylation was assessed in 215 endometrial tumors by Southern blot analysis of HpaII-digested DNA using a probe corresponding to the 5.8 and 28S ribosomal subunits. Tumor rDNA methylation status was correlated with patient outcome. RESULTS: The majority of tumors demonstrated high levels of rDNA methylation (rDNA-high; 74%). Both disease free survival and overall survival were significantly worse for patients with low-level rDNA methylation (rDNA-low; P < 0.0001). African-American patients were more likely to have rDNA-low tumors than Caucasian patients (P = 0.002). Among the subpopulation of patients with endometrial carcinoma for whom the use of adjuvant therapy is most controversial (148 women with Stage I-II endometrioid tumors), survival was significantly worse for the patients with rDNA-low tumors (P < 0.0001); and, using multivariate analyses, tumor rDNA level was the only significant prognostic factor for both disease free survival and overall survival (hazard ratios, 11.0 and 26.3, respectively; P < 0.01 for both). CONCLUSIONS: rDNA methylation is an independent prognostic indicator for patients with endometrial carcinoma that may serve to identify women with early-stage disease at who are at high risk for recurrence. Racial differences in DNA methylation may explain the historically observed disparity in survival between African-American patients and Caucasian patients.

Adenocarcinoma, Clear Cell↗