Biomedical subjects

Raheela Ashfaq

Publications and source records attributed to Raheela Ashfaq.

At least 37 records · Page 2Linked to original sources

PubMed2005

Loss of FHIT expression in breast cancer is correlated with poor prognostic markers.

OBJECTIVE: The fragile histidine triad (FHIT) gene is a putative tumor suppressor gene that is thought to be involved in the carcinogenesis of breast cancer. Loss of FHIT expression has been observed in up to 72% of breast cancers and has been associated with increased p53, a high proliferation index, and increased tumor size and grade. However, loss of FHIT expression has not been investigated in association with apoptosis and cyclooxygenase-2 (COX-2) expression in breast cancer. Furthermore, expression of FHIT in primary breast tumors and their metastatic axillary lymph nodes has also not been previously described. The purpose of this study was to evaluate the expression of FHIT, COX-2, bcl-2, and p53 in primary breast tumor tissue; correlate their expression with known clinical and pathologic markers; and in cases when tissue was available, evaluate the expression of FHIT and COX-2 in the corresponding metastatic axillary lymph node in the same patient. METHODS: Primary breast tumor specimens from 80 patients were examined for the presence of FHIT, COX-2, bcl-2, and p53 expression by immunohistochemistry using standard methods. When tissue was available, the expression of FHIT and COX-2 was also evaluated in the corresponding metastatic axillary lymph node specimen. RESULTS: FHIT expression in primary breast tumors was 56%. There was a significant correlation between FHIT expression in primary breast tumor and bcl-2 expression (P = 0.017). We also observed a significant inverse correlation between FHIT expression in primary breast tumor tissue and p53 expression (P = 0.023) in lymph node-negative cases. A significant inverse correlation between FHIT expression in the primary tumor and Ki-67 (P = 0.009) was also observed in lymph node-negative cases. FHIT expression in primary tumors correlated with FHIT expression in the metastatic lymph node (52.5%; P = 0.001). FHIT expression in primary tumors did not correlate with COX-2 expression. CONCLUSION: Our results suggest that loss of FHIT expression in breast cancer is associated with poor prognostic features. Furthermore, loss of FHIT expression is also seen in metastatic axillary lymph node. The prognostic and predictive value of these findings needs to be further evaluated in larger trials with longer follow-up.

Acid Anhydride Hydrolases↗

PubMed2005

A subset of pancreatic adenocarcinomas demonstrates coamplification of topoisomerase IIalpha and HER2/neu: use of immunolabeling and multicolor FISH for potential patient screening andtreatment.

We sought to identify the frequency of amplification of the topoisomerase IIalpha gene (TOP2A) in pancreatic cancer and determine the usefulness of TOP2A immunolabeling in screening for TOP2A and human epidermal growth factor receptor (HER)2/neu amplification. We examined 55 pancreatic adenocarcinoma specimens for TOP2A immunolabeling and identified TOP2A protein expression in all specimens with a nuclear labeling index (NLI; positive nuclei/total nuclei x 100) of 5% to 80%. Normal pancreatic ductal epithelium, proposed to give rise to pancreatic adenocarcinoma, did not demonstrate detectable TOP2A expression. In a subset of specimens selected for fluorescence in situ hybridization analysis of TOP2A and HER2/neu amplification using a recently developed multicolor probe, 7 of 8 lesions with an NLI of 25% or more demonstrated TOP2A amplification, in contrast with 2 of 14 lesions with a TOP2A NLI of less than 25%. In 8 of 9 TOP2A-amplified cases, coamplification of HER2/neu was present, suggesting a potential relationship between TOP2A and HER2/neu in pancreatic adenocarcinoma. We propose that TOP2A immunolabeling be used in conjunction with a newly developed multicolor probe to screen patients with pancreatic adenocarcinoma to determine the best potential therapeutic modalities, such as TOP2A inhibitors, trastuzumab, or both.

Adenocarcinoma↗

PubMed2004

An epidermal growth factor receptor intron 1 polymorphism mediates response to epidermal growth factor receptor inhibitors.

This study tested the hypothesis that the number of CA single sequence repeat (CA-SSR) in the intron 1 of the epidermal growth factor receptor (egfr) gene, which affects transcription efficiency of the gene, is associated with the response to EGFR inhibitors. To this end, we determined the number of CA dinucleotides in the intron 1 of the egfr gene in a panel of 12 head and neck cancer cell lines that lack egfr gene amplification and measured the expression of EGFR (mRNA and protein), as well as response to EGFR inhibition. Cells with lower number of CA dinucleotides in the CA-SSR had higher expression of the EGFR gene and protein and were more sensitive to the inhibitory effects of erlotinib, a small molecule inhibitor of the EGFR tyrosine-kinase. Phenotypic modification by silencing EGFR mRNA expression in a susceptible cell line induced resistance to the drug. The number of CA dinucleotide was equivalent in genomic and tumor DNA obtained from 30 patients with head and neck cancer. In a clinical study in colorectal cancer, subjects with lower number of CA dinucleotide frequently developed skin toxicity, a feature that is related to the antitumor activity of this class of drugs. These results suggest that polymorphic variations in the intron 1 of the egfr gene is associated with response to EGFR inhibitors and may provide an explanation as to why the development of skin toxicity is associated with a favorable outcome in patients treated with these agents.

Cell Line, Tumor↗

PubMed2004

Met proto-oncogene and insulin-like growth factor binding protein 3 overexpression correlates with metastatic ability in well-differentiated pancreatic endocrine neoplasms.

Pancreatic endocrine neoplasms are neoplastic proliferations of islet cells or islet cell precursors and are capable of secreting a variety of synthetic products, including insulin, glucagon, gastrin, and vasoactive intestinal peptide. The biological behavior of pancreatic endocrine neoplasms is often unpredictable, and there are few reliable histopathologic criteria reliably correlating with metastatic ability. We have used the Affymetrix U133 GeneChip set (HG_U133 A and B; Affymetrix; Santa Clara, CA) representing approximately 33,000 characterized transcripts to examine global gene expression profiles from well-differentiated nonmetastatic (n=5) and metastatic (n=7) pancreatic endocrine neoplasms to determine molecular markers that predict disease progression. Microarray hybridization data were normalized using the GeneLogic GeneExpress Software System to identify differentially up- and down-regulated genes in metastatic versus nonmetastatic pancreatic endocrine neoplasms. Using a 3-fold change in gene expression as a threshold, we have identified 65 overexpressed and 57 underexpressed genes in metastatic pancreatic endocrine neoplasms as compared with nonmetastatic pancreatic endocrine neoplasms. Several classes of genes, including growth factors and growth factor-related molecules (IGFBP1, IGFBP3, and MET), developmental factors (TBX3 and MEIS2), cytoskeletal factors (beta 1 tubulin and ACTN2), cholesterol homeostasis mediators (LRP5, SLC27A2, and RXRG), intracellular signaling pathway mediators (DYRK1A, PKIB, and AK2), methyltransferases (MGMT and GAMT), and DNA repair and regulatory molecules (CHEK1 and ZNF198), were identified as differentially over- or underexpressed via this method. Immunohistochemical validation of microarray data were performed for two overexpressed genes, namely, the met proto-oncogene (MET) and insulin-like growth factor binding protein 3 (IGFBP3) with tissue microarrays of nonmetastatic (n=24) and metastatic (n=15) pancreatic endocrine neoplasms. Increased expression of IGFBP3 was confirmed in metastatic versus nonmetastatic pancreatic endocrine neoplasms (12 of 15, 80% versus 10 of 24, 42%), as well as in lymph node (6 of 7, 86%) and liver (9 of 9, 100%) metastases. Similarly, overexpression of MET was confirmed in metastatic versus nonmetastatic pancreatic endocrine neoplasms (5 of 15, 33% versus 4 of 24, 17%), as well as in lymph node metastases (4 of 7, 57%) and liver metastases (5 of 9, 56%). The majority of genes that demonstrated altered expression has not been previously identified as differentially expressed in metastatic pancreatic endocrine neoplasm lesions and may therefore represent newly identified molecules in the progression of these lesions.

Aged↗

PubMed2004

Receptor for the globular heads of C1q (gC1q-R, p33, hyaluronan-binding protein) is preferentially expressed by adenocarcinoma cells.

Combinatorial Ig libraries with phage display allow in vitro generation of human Ig fragments without the need to maintain hybridomas in ongoing cell culture or to select circulating Ig from human serum. Identifying tumor-associated antigens on the surface of intact tumor cells, as opposed to purified proteins, presents a challenge due to the difficulty of preserving complex 3-D epitopic sites on the cell surface, the variable expression of antigens on different malignant cell types and the stereotactic interference of closely associated proteins on the intact membrane surface limiting accessibility to antigenic sites. A combinatorial Ig library of 10(10) clones was generated from the cDNA of PBMCs derived from patients with breast adenocarcinoma. Following subtractive panning, the library was enriched for Ig (Fab fragment) binding to intact adenocarcinoma cells and the resultant Fabs were screened against a cDNA expression library, itself generated from breast cancer cells. Using this approach, we isolated clones from the cDNA library expressing gC1q-R, a glycoprotein comprising the major structure of C1, the first component of the complement system. gC1q-R is a 33 kDa glycoprotein expressed not only on the cell surface but also intracellularly, with motifs that target it to mitochondria and complete homology with HABP and human HeLa cell protein p32, which is copurified with pre-mRNA SF2. Sequencing of the gene encoding tumor-associated gC1q-R did not reveal any consistent tumor-specific mutations. However, histochemical staining with anti-gC1q-R MAb demonstrated marked differential expression of gC1q-R in thyroid, colon, pancreatic, gastric, esophageal and lung adenocarcinomas compared to their nonmalignant histologic counterparts. In contrast, differential expression was not seen in endometrial, renal and prostate carcinomas. Despite high expression in breast carcinoma, gC1q-R was also expressed in nonmalignant breast tissue. Although the precise relation of gC1q-R to carcinogenesis remains unclear, our finding of tumor overexpression and the known multivalent binding of gC1q-R to not only C1q itself but also a variety of circulating plasma proteins as well as its involvement in cell-to-cell interactions suggest that gC1q-R may have a role in tumor metastases and potentially serve in molecule-specific targeting of malignant cells.

Adenocarcinoma↗

PubMed2004

Gene expression profiling identifies markers of ampullary adenocarcinoma.

Ampullary adenocarcinoma is an aggressive cancer with a poor prognosis. Without surgical resection, ampullary adenocarcinomas can be difficult to distinguish from ampullary adenomas. The aim of this study was to identify differentially expressed genes in ampullary adenocarcinoma in order to identify candidate markers of the disease. The Affymetrix Human Genome U133 GeneChip set (HG-U133A and HG-U133B) was used to obtain gene expression profiles of 5 ampullary adenocarcinomas and 10 normal duodenal samples. Using fold change analysis we identified 235 fragments expressed at least fivefold higher in ampullary cancers than in normal duodenum. The expression profiles of eight candidate overexpressed genes (osteopontin, mesothelin, tissue inhibitor of metalloproteinases 1, mucin-1, mucin-5, fascin, heat shock protein 47, fibronectin 1) were confirmed by immunohistochemistry or in situ hybridization on tissue microarrays (TMA) containing 54 ampullary adenocarcinomas. One of these genes, osteopontin, was expressed 27-fold higher levels in ampullary adenocarcinomas compared to normal duodenum by genechip analysis. We therefore determined serum osteopontin levels in patients with ampullary neoplasms, patients with other periampullary diseases and in normal controls. Mean preoperative serum osteopontin levels as measured by competitive ELISA were 906 +/- 268 ng/ml in patients with ampullary cancer, 867 +/- 160 ng/ml in patients with an ampullary adenoma, 327.1 +/- 195.6 ng/ml in patients with nonmalignant periampullary diseases and 204 +/- 65 ng/ml in age-matched healthy controls (p < 0.001). Measurement of markers of ampullary cancer such as osteopontin may aid in the early detection and differential diagnosis of patients with periampullary lesions.

Adenocarcinoma↗

PubMed2004

HER-2 gene amplification can be acquired as breast cancer progresses.

Amplification and overexpression of the HER-2 oncogene in breast cancer is felt to be stable over the course of disease and concordant between primary tumor and metastases. Therefore, patients with HER-2-negative primary tumors rarely will receive anti-Her-2 antibody (trastuzumab, Herceptin) therapy. A very sensitive blood test was used to capture circulating tumor cells (CTCs) and evaluate their HER-2 gene status by fluorescence in situ hybridization. The HER-2 status of the primary tumor and corresponding CTCs in 31 patients showed 97% agreement, with no false positives. In 10 patients with HER-2-positive tumors, the HER-2/chromosome enumerator probe 17 ratio in each tumor was about twice that of the corresponding CTCs (mean 6.64 +/- 2.72 vs. 2.8 +/- 0.6). Hence, the ratio of the CTCs is a reliable surrogate marker for the expected high ratio in the primary tumor. Her-2 protein expression of 10 CTCs was sufficient to make a definitive diagnosis of the HER-2 gene status of the whole population of CTCs in 19 patients with recurrent breast cancer. Nine of 24 breast cancer patients whose primary tumor was HER-2-negative each acquired HER-2 gene amplification in their CTCs during cancer progression, i.e., 37.5% (95% confidence interval of 18.8-59.4%). Four of the 9 patients were treated with Herceptin-containing therapy. One had a complete response and 2 had a partial response.

Antibodies, Monoclonal↗

PubMed2004

Reprocessing unsatisfactory ThinPrep Papanicolaou test specimens increases sample adequacy and detection of significant cervicovaginal lesions.

BACKGROUND: The objective of the current study was to determine the effect of reprocessing bloody ThinPrep (TP) samples using a glacial acetic acid technique on the unsatisfactory rate. METHODS: During a 12-month study period, all TP gynecologic samples received by the Cytology Laboratory from inpatient, outpatient, and community-based clinics at the University of Texas Southwestern Medical Center and the Parkland Health and Hospital System in Dallas, Texas, were enrolled prospectively into the study. The initial TP slides were evaluated for specimen adequacy based on the 2001 Bethesda System. Any TP sample that contained abnormal cells, by definition, was not considered unsatisfactory. The criteria for reprocessing included scant cellularity in a background of abundant blood. Biopsy correlations for all abnormal cytologic diagnoses were established between TP smears and follow-up biopsies. RESULTS: During the course of the 1-year study period, a total volume of 57,296 TP samples were evaluated by the study laboratory. Prior to reprocessing, the laboratory unsatisfactory rate was 8.32% (4767 of 57,296 TP samples). After reprocessing those samples that were compromised by blood (2593 of 4767 TP samples), the unsatisfactory rate was reduced to 5.47% (3134 of 57,296 TP samples), with an overall reduction by 34.25% (2.85 of 8.32 TP samples). Of 2593 samples that were reprocessed, 62.98% (1633 of 2593 TP samples) changed from inadequate to adequate for the purposes of evaluation. Of those 1633 adequate samples, 1509 samples (92.41%) were negative for intraepithelial lesion or malignancy, 72 samples (4.40%) were atypical squamous cells of unknown significance, 5 samples (0.31%) were atypical glandular cells of undetermined significance, 36 samples (2.20%) were low-grade squamous intraepithelial lesions, 10 samples (0.61%) were high-grade squamous intraepithelial lesions, and 1 sample (0.06%) was adenocarcinoma. CONCLUSIONS: The authors conclude that the reprocessing of unsatisfactory ThinPrep cervicovaginal cytology samples decrease the unsatisfactory rate considerably, with an increase in the detection of significant abnormal cervicovaginal lesions.

Female↗

PubMed2004

Survivin expression is associated with features of biologically aggressive prostate carcinoma.

BACKGROUND: Survivin counteracts cell death and controls mitotic progression. The objectives of the current study were to compare the expression patterns of survivin in normal prostate, primary prostate carcinoma, and lymph node tissues involved with prostate carcinoma and to determine whether the expression of survivin is associated with prostate carcinoma characteristics and progression. METHODS: Immunohistochemical staining for survivin and for transforming growth factor beta1 (TGF-beta1) and its receptors (types I and II; TGF-betaR1 and TGF-betaR2, respectively) was carried out on archival specimens from 114 consecutive patients who underwent radical prostatectomy (median follow-up, 64.8 months). Punch biopsies of the index carcinoma and normal tissue from each specimen were sectioned onto a single slide and stained. The authors also evaluated the expression of survivin in normal and malignant lymph node tissue from eight patients. RESULTS: Survivin was expressed in 41 of 114 normal prostate specimens (36%) from prostates that contained carcinoma, in 81 of 114 primary prostate carcinoma specimens (71%), in 3 of 8 normal lymphoid specimens (38%), and in 7 of 8 prostate carcinoma lymphoid specimens (88%). Survivin expression was associated with higher final Gleason sum (P = 0.001), loss of TGF-betaR1 and TGF-betaR2 expression (P = 0.041 and P = 0.008, respectively), and an increased risk of biochemical progression on univariate analysis (P = 0.0441). Among patients who had disease progression, survivin was expressed more commonly in those who had tumors with features of aggressive behavior. CONCLUSIONS: The expression of survivin gradually increased from normal prostate tissue, to low-grade primary carcinoma, to high-grade primary carcinoma and was highest in lymph node metastases. Survivin expression was associated further with alteration of the TGF-beta pathway and with overall and aggressive biochemical progression after radical prostatectomy.

Adult↗

PubMed2004

P16 as a molecular biomarker of cervical adenocarcinoma.

OBJECTIVE: Cervical adenocarcinomas are increasing in incidence each year. The aim of this study was to identify a molecular biomarker to improve early detection. STUDY DESIGN: Fifty-five in situ and invasive cervical adenocarcinomas were compared with 5 normal endocervical controls by immunohistochemical analysis of p16, p21, p27, cyclin D1, cyclin E, p53, and Ki-67. Expression was scored from 0 to 8 by using an automated imaging system. Western blotting and polymerase chain reaction-based human papillomavirus (HPV) testing were performed on 16 of the invasive cases having fresh-frozen tissue. RESULTS: P16 exhibited a higher mean expression score for in situ (7.4; P<.0001) and invasive cervical adenocarcinoma (6.6; P<.0001) versus controls (2.0). A cutoff p16 expression score of 5 had a sensitivity of 94.5% and a specificity of 100%. Western blotting confirmed p16 protein expression. Fourteen (88%) of 16 invasive cervical adenocarcinomas were HPV-positive. CONCLUSION: P16 is a putative molecular biomarker of cervical adenocarcinoma. Overexpression appears to primarily reflect HPV-induced cell cycle dysregulation.

Adenocarcinoma↗

PubMed2004

Patient and duct selection for nipple duct lavage.

BACKGROUND: Nipple ductal lavage (NDL) is a new minimally invasive procedure with the potential to help identify women who could benefit from breast cancer risk intervention. NDL is currently encouraged for women with fluid-producing ducts and a 5-year Gail risk > or =1.7%. The purpose of this study was to evaluate the atypia rate by NDL in fluid-producing ducts compared with non-fluid-producing ducts and the atypia rate in high-risk verses low-risk patients to determine if current recommendations are supported. METHODS: Fifty-nine women were studied with NDL. The 226 ducts lavaged included all fluid-producing ducts (n = 136) and any dry ducts we could cannulate (n = 90). Breast cancer risk was calculated using mathematic models. RESULTS: There were 26 (44%) women with a 5-year Gail risk > or =1.7% and 33 (56%) with a 5-year Gail risk <1.7%. Cytologic atypia was diagnosed in 20 of 59 (34%) of patients. The atypia rate was similar for women with a 5-year Gail risk > or =1.7% (9 of 26 or 35%) compared with lower-risk women (11 of 33 or 33%, P = 1.0) and for fluid-producing ducts (26 of 136 or 19%) compared with dry ducts (14 of 90 or 15%, P = 0.61). No significant differences were found when the atypia was categorized as mild versus marked. Of note, the insufficient sample rate was higher for dry ducts (33%) compared with fluid-producing duct (22%, P = 0.07). CONCLUSIONS: If NDL results are found to correlate with breast cancer incidence, it will be important to apply the test in a way that maximizes sensitivity for the detection of atypia in a screened population. We were unable to identify patient or duct characteristics that predict NDL atypia rates.

Adult↗

PubMed2004

Risk stratification for bladder tumor recurrence, stage and grade by urinary nuclear matrix protein 22 and cytology.

PURPOSE: To test the hypothesis that voided urinary levels of nuclear matrix protein 22 (NMP22) would add to the predictive ability of urine cytology in the diagnosis, staging and grading of bladder transitional cell carcinoma (TCC), and to evaluate the diagnostic performance of different NMP22 cut-points. MATERIALS: NMP22 level and barbotage cytology were evaluated in voided urine specimens collected before cystoscopy from 302 subjects with a history of TCC, 32 subjects with benign urologic pathologies, and 10 healthy volunteers. RESULTS: 180 patients (52%) had bladder TCC. Higher levels of NMP22 and positive cytology were independently associated with an increased risk of TCC, invasive stage, and high grade. The NMP22 value with equal sensitivity and specificity for prediction of bladder cancer was 6.5U/ml; for prediction of grade 3 TCC it was 13.5U/ml; and for prediction of invasive tumor stage it was 17.4U/ml. The NMP22 cut-point of 6.5U/ml outperformed the 10U/ml cut-point in all pathologic stages and grades. The diagnostic sensitivity of the cytology and NMP22 combined was superior across all pathologic stages and grades to that of either marker alone. NMP22 and cytology stratified patients into groups with significantly different risk for TCC presence, invasive stage, and high grade. CONCLUSIONS: 6.5U/ml is a robust NMP22 cut-point for bladder cancer surveillance. The diagnostic sensitivities of the combined NMP22 and cytology for TCC presence, stage, and grade were significantly higher than those of single marker alone. The combination of urine cytology and NMP22 could be used to tailor the frequency of cystoscopic follow up.

Adult↗

PubMed2004

Analysis of novel tumor markers in pancreatic and biliary carcinomas using tissue microarrays.

Using global gene expression analyses, multiple novel tumor markers overexpressed in infiltrating ductal adenocarcinomas of the pancreas have recently been identified. However, the expression of these markers in morphologically similar adenocarcinomas of the biliary tree has not been investigated. The purpose of the present study was 3-fold. First, we used 8 markers that have been shown to be overexpressed in whole tissue sections of pancreatic adenocarcinomas to validate tissue microarrays (TMAs) created from a series of pancreatic adenocarcinomas (n=68). The labeling patterns of 6 epithelial markers (fascin, mucin 4, 14-3-3sigma, prostate stem cell antigen, topoisomerase IIalpha, and cdc2/p34) were concordant with previously published studies on whole tissue sections, yet required far fewer slides and reagents. Mesothelin, an epithelial marker, and heat shock protein 47, a marker of peritumoral desmoplasia, showed lower levels of expression in the TMAs when compared with whole tissue sections. Second, we examined the previously unknown expression of the same 8 novel tumor proteins in cancers of the biliary tree by using TMAs created from a series of intrahepatic cholangiocarcinomas, gallbladder adenocarcinomas, and adenocarcinomas of the distal common bile duct (n=38). Each of the 8 markers was overexpressed in the biliary cancers, ranging from 14% demonstrating at least focal labeling with prostate stem cell antigen to 100% labeling with cdc2/p34. Most of the markers showed lower frequencies of expression in the biliary tract carcinomas in comparison to the pancreatic adenocarcinomas. In addition, expression patterns varied with location in the biliary system (intrahepatic versus gallbladder versus distal common bile duct). These differences were statistically significant (P<0.05) for mesothelin, mucin 4, and heat shock protein 47. Finally, the expression of selected markers in neoplastic progression of gallbladder cancer was examined. Two markers, fascin and mesothelin, showed up-regulation of expression with transition from carcinoma in situ to invasive adenocarcinoma, implicating a role for these markers in neoplastic progression. The results of this study indicate that TMA technology provides valid and cost-effective means to screen large numbers of novel tumor markers, even in tumors such as pancreatic and biliary adenocarcinomas that characteristically have abundant desmoplastic stroma. In addition, novel tumor markers of pancreatic adenocarcinomas show similar, yet not identical, expression patterns in biliary carcinomas. Therefore, these markers are potentially useful in developing diagnostic tests and treatment paradigms for tumors involving the biliary system.

Adenocarcinoma↗

PubMed2004

Understanding the mechanisms of FHIT inactivation in cervical cancer for biomarker development.

OBJECTIVE: Loss of the fragile histidine triad (Fhit) protein has been documented in cervical cancer and dysplasia. The goal of this study was to confirm the utility of homozygous deletions, aberrant methylation, and immunohistochemical evaluations of FHIT as functionally relevant determinants of FHIT expression. METHODS: We studied matched DNA, RNA, and protein from nine early-passage cervical cancer cell lines. DNA markers spanning FHIT were used to examine the extent of homozygous deletions for each cell line. 5 CpG island methylation of FHIT was investigated by methylation-specific polymerase chain reaction (PCR) assays. FHIT transcripts were characterized by reverse transcriptase (RT)-PCR. Western blot analysis and immunohistochemistry were performed to characterize Fhit protein expression. RESULTS: Homozygous deletions were found in six of nine cervical cancer cell lines, but only one had homozygous deletions involving an exon. All nine lines had both methylated and unmethylated alleles according to methylation-specific PCR. Loss of wild-type FHIT transcripts were found in five of nine lines. By western blot analysis, Fhit protein expression was lost in five of nine lines, producing an exact correlation with RT-PCR results. Immunohistochemical staining was concordant with Fhit protein expression by western blotting in eight of nine cell lines. CONCLUSION: A perfect correlation was found between FHIT mRNA expression and western blot analysis. Assays for Fhit protein expression and large FHIT homozygous deletions are representative biomarkers of Fhit expression. By contrast, the aberrant methylation assay is not concordant with FHIT gene expression, and we suggest caution in its use as a functionally relevant biomarker for cervical cancer.

Acid Anhydride Hydrolases↗

PubMed2004

FHIT mRNA and protein expression in hepatocellular carcinoma.

Loss of fragile histidine triad (FHIT) gene expression is seen in approximately 50% of hepatocellular carcinomas in China. However, little information is available on FHIT expression in hepatocellular carcinoma in the United States, where carcinogen exposure is generally lower. Investigations of FHIT mRNA in hepatocellular neoplasms and paired non-neoplastic tissues demonstrated normal-sized FHIT transcripts in all non-neoplastic tissues and in all neoplasms including 11 hepatocellular carcinomas, two fibrolamellar carcinomas, and four benign proliferative lesions. In addition, all but one malignant and all benign neoplasms showed aberrant smaller transcripts. The smaller aberrant transcripts were overexpressed in 6/11 hepatocellular carcinomas and 1/2 fibrolamellar carcinomas. An additional 79 hepatocellular carcinomas, 12 fibrolamellar carcinomas and 15 hepatic adenomas were examined for FHIT expression by immunohistochemistry. No loss of immunostaining was seen in 67/79 (85%) of hepatocellular carcinomas, while a moderate or marked decrease was seen in 12/79 (15%). Fibrolamellar carcinomas and hepatic adenomas showed no loss of FHIT expression. In conclusion, hepatocellular carcinomas retained expression of normal FHIT mRNA transcripts, but also showed universal expression of smaller sized aberrant transcripts and commonly overexpressed these aberrant transcripts. Loss of FHIT protein expression is relatively uncommon in this cohort from the United States, where exposure to hepatic carcinogens is generally low.

Acid Anhydride Hydrolases↗

PubMed2004

MIB1 labeling index as an indicator of chemoresponse in carcinoma of the breast.

In this study, we determined the extent of variation in proliferative markers and hormone receptor status in breast carcinoma between core biopsies and subsequent resections, and determined the impact of clinical and histologic parameters on such variation. We performed a paired comparison of biomarkers in 87 core biopsies and subsequent resections of breast carcinomas in patients with and without preoperative chemotherapy. The markers included estrogen receptor, progesterone receptor, Her2/neu, DNA ploidy (diploidy versus nondiploidy), DNA index (difference of > or = 0.5), and MIB1 labeling index (<15% vs. >15%). The tumor biomarkers were evaluated with standard IHC and scored by automated cellular imaging systems. The number of patients showing prominent changes were as follows: ploidy, 12; DI, 15; ER, 6; PR, 15; MIB1, 17; and Her2/neu, 15. Seventeen of 87 patients sustained a significant change in MIB1 index (above or below 15%). The patients who received chemotherapy had a larger proportion with a change in MIB1 status (P < 0.05). DI showed a similar trend, although it was not statistically significant. Of the patients with MIB1 values higher in biopsy specimens, a majority showed values to decrease below 15%. MIB1 index reduction by greater than 25% was seen in 8 of 16 (50%) cases with chemotherapy, compared with only 3 of 32 (9%) without chemotherapy. MIB1 and DNA index values postchemotherapy can be a useful measure of chemoresponse. Our study underscores the need to perform prognostic markers on both biopsy and subsequent resections, especially in the setting of preoperative chemotherapy.

Biopsy, Fine-Needle↗

PubMed2004

Claudin 4 protein expression in primary and metastatic pancreatic cancer: support for use as a therapeutic target.

We performed a comprehensive immunohistochemical evaluation of claudin 4 protein expression in paraffin-embedded tissue samples from 72 patients with primary infiltrating pancreatic cancer, 38 patients with metastatic pancreatic cancer, and a panel of normal control tissue samples from various organs. In 11 samples of primary infiltrating pancreatic cancer, foci of pancreatic intraepithelial neoplasia (PanIN) were present and also were analyzed for claudin 4 protein expression. Intense positive claudin 4 immunolabeling was noted within virtually all primary (71/72 [99%]) and metastatic (49/49 [100%]) pancreatic cancer tissue samples analyzed and in 10 of 11 samples of PanIN. In all cases, immunolabeling was noted in a membranous distribution. Claudin 4 protein also was detectable in normal breast, prostate, bladder, and gastrointestinal mucosa, although expression was substantially less intense than that seen in pancreatic cancer tissue samples. Our findings support the use of claudin 4 as a target for novel therapeutics or radioimaging of infiltrating pancreatic cancer. Furthermore, the finding of claudin 4 overexpression within pancreatic intraepithelial neoplasia, the precursor lesion of pancreatic cancer, suggests a potential benefit of imaging claudin 4 before the development of an invasive carcinoma.

Antibodies, Monoclonal↗

PubMed2004

BRCAPAP: feasibility of clinical BRCA testing on liquid-based cervical cytology: implications for biomarker development.

OBJECTIVE: The study was designed to test the feasibility that lower genital tract cytology is a compatible medium for robust germ line genetic analyses. METHOD: BRCA1 and/or BRCA2 gene mutational analysis was done on DNA isolated from liquid-based cervical or vaginal cytology taken from 17 consenting women (age 29-65 years) who previously had genetic counseling followed by BRACAnalysis (Myriad Genetics, Salt Lake City, UT) blood analyses. Eleven women had known mutations in either BRCA1 or BRCA2 (cases) and six had no identified mutations (controls) on entry into the study. Anonymized cytology samples were sent to Myriad Genetics with a request for testing that was limited to the degree of genomic testing previously done on the blood samples. RESULTS: One cervicovaginal specimen from a test-positive woman had inadequate cellular content that precluded gene sequencing and therefore was excluded from this analysis. For the 16 women with adequate cytologic specimens, there was 100% concordance for BRCA mutation test results between blood and genital tract cytology (kappa = 1.0; 95% confidence interval, 0.51-1.0). CONCLUSION: We have shown the feasibility of using liquid-based genital tract cytology as an alternative biospecimen to blood for germ line genetic analysis using a clinical approved assay. It needs to be emphasized that any type of testing for BRCA1 or BRCA2 mutation genotype should only be done in the setting of pretest and posttest counseling.

Adult↗