Browsing by Author "Gazdar, Adi F."

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    Aberrant promoter hypermethylation of multiple genes in gallbladder carcinoma and chronic cholecystitis
    (American Association Cancer Research, 2004) Takahashi, Takao; Shivapurkar, Narayan; Riquelme Sánchez, Erick Marcelo; Shigematsu, Hisayuki; Reddy, Jyotsna; Suzuki, Makoto; Miyajima, Kuniharu; Zhou, Xian; Bekele, B. Nebiyou; Gazdar, Adi F.; Wistuba Oyarzún, Ignacio
    Purpose: Aberrant methylation of 5' gene promoter regions is an epigenetic phenomenon that is a major mechanism for silencing of tumor suppressor genes in many cancer types. There is limited information about the molecular changes involved in the pathogenesis of gallbladder carcinoma (GBC), including methylation status. Experimental Design: We investigated the aberrant promoter methylation profile of 24 known or suspected tumor suppressor genes in 50 GBCs and compared those results with the findings in 25 chronic cholecystitis (CC) specimens without cancer. The methylation-specific polymerase chain reaction and combined restriction analysis methods were used to detect methylation, and the results were confirmed by sequencing of cloned polymerase chain reaction products. Results: In GBC, gene methylation frequencies varied from 0% to 80%. Ten genes demonstrated relatively high frequencies of aberrant methylation: SHP1 (80%), 3-OST-2 (72%), CDH13 (44%), P-15(INK4B) (44%), CDH1 (38%), RUNX3 (32%), APC (30%), RIZ1 (26%), P16(INK4A) (24%), and HPP1 (20 %). Eight genes (P73, RARbeta2, SOCS-1, DAPK, DcR2, DcR1, HIN1, and CHFR) showed low frequencies (2-14%) of methylation, and no methylation of the remaining six genes (TIMP-3, P57, RASSF1A, CRBP1, SYK, and NORE1) was detected. In CC, methylation was detected for seven genes: SHP1 (88 %), P15(INK4B) (28 %), 3-OST-2 (12%), CDH1 (12 %), CDH13 (8 %), DcR2 (4 %), and P16(INK4A) (4%) Significantly higher frequencies of methylation in GBC compared with CC were detected for eight genes (3-OST-2, CDH13, CDH1, RUNX3, APC, RIZ1, P16(INK4A), and HPP1). Of those, four genes showed frequent methylation (>30%) in GBCs. The mean methylation index, an expression of the amount of methylated genes by case, was significantly higher in GBC (0.196 +/- 0.013) compared with CC (0.065 +/- 0.008; P < 0.001). Conclusions: Our study constitutes the most comprehensive methylation profile report available in GBC and demonstrates that this neoplasm has a distinct pattern of abnormal gene methylation. Whereas gallbladders from healthy individual were not available, our finding of methylation in CC cases without cancer suggests that this phenomenon represents an early event in the pathogenesis of GBC.
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    Fragile histidine triad gene abnormalities in the pathogenesis of gallbladder carcinoma
    (2002) Wistuba Oyarzún, Ignacio; Ashfaq, Raheela; Maitra, Anirban; Álvarez Pando, Héctor Andrés; Riquelme Sánchez, Erick Marcelo; Gazdar, Adi F.
    There is limited information about the molecular changes involved in the pathogenesis of gallbladder carcinoma (GBC). Our recent allelotyping analyses have indicated that chromosome 3p loss of heterozygosity (LOH), including the fragile histidine triad (FHIT) candidate tumor-suppressor gene locus at 3p14.2, is frequently detected in this neoplasm. To investigate the role of the FHIT abnormalities in the multistage sequential development of GBC, 33 formalin-fixed paraffin-embedded invasive GBC specimens and 76 accompanying histologically normal (n = 43) and dysplastic (n = 33) epithelia were examined by nostaining for expression of Fhit protein. Allele loss at the FHIT gene locus (3p14.2) was studied in all GBCs and in a subset of accompanying gallbladder epithelia by polymerase chain reaction-based LOH analysis, using three 3p14.2 microsatellite markers. In addition, histologically normal epithelium from chronic cholecystitis (n = 19) and dysplasia (it = 13) from gallbladder specimens without cancer were examined for immunostaining and LOH. There was a progressive increase in both the frequency of loss of Fhit expression and LOH at FHIT with increasing severity of histopathological changes. FHIT abnormalities were occasionally demonstrated in histologically normal gallbladder epithelium. Dysplastic foci demonstrated frequent reduction or absence of Fhit immunostaining (38 to 55%) and FHIT allelic loss (33 to 46%). In invasive tumors, these abnormalities were even higher, with 79% reduction or absence of Fhit immunostaining and 76% FHIT allele loss. A high correlation (70%) was observed between Fhit immunostaining abnormalities and allele loss in GBC specimens (P < 0.05). Although a high frequency of FHIT locus breakpoints; were detected in both invasive and dysplastic gallbladder specimens, no intronic homozygous deletions on FHIT were detected in GBCs. FHIT gene abnormalities are nearly universal in GBC and these changes are detected early in the sequential development of this neoplasm. Our findings indicate that the FHIT gene is one of the chromosome 3p putative tumor suppressor genes involved in the pathogenesis of this highly malignant neoplasm.
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    Tp53 Abnormalities Are Frequent and Early Events in the Sequential Pathogenesis of Gallbladder Carcinoma
    (2005) Moreno, M.; Pimentel González, Eduardo Fernando; Gazdar, Adi F.; Wistuba Oyarzun, Ignacio; Miquel Poblete Juan Francisco
    Background: Gallbladder carcinoma (GBC) is a frequent neoplasm in Hispanic and native American populations. GBC is preceded by gallstones, chronic cholecystitis and dysplastic changes of the gallbladder epithelium. The knowledge of the molecular events involved in its pathogenesis is scarce. Aims: We investigated the role of TP53 inactivation in the sequential pathogenesis of GBC. Methods: Invasive tumor-, dysplastic- and histologically normal GB ep ithelial-cells were obtained from archival formalin fixed tissues from GBC and GB from gallstone pa tients without GBC. Normal GB epithelia from 5 non-gallstone specimens were also studied. DNA extracted was examined for loss of heterozygosity (LOH) using 2 microsatellite markers and for TP53 mutations at exons 5 to 8. Results: GBCs demonstrated a high frequency of LOH (81%) and mutation (67%), and both abnormalities indicating gene inactivation were detected in 52%. Similar frequency of TP53 Abnormalities are frequent and early events in the sequential pathogenesis of gallbladder carcinomaivation (38%) were detected in their accompanying normal and dysplastic epithelia. Noteworthy, one third of normal and dysplastic epithelia obtained from GBs of gallstone patients without GBC demonstrated either TP53 allele loss or mutation, but gene inactivation was less frequent (11%). Most mutations affected exons 5 and 7, and they were more frequently missense point mutations. The same TP53 mutation was de tected in only a subset (27%) of comparisons between non-malignant epithelia adjacent to GBCs, indicating that TP53 mutation occurs independently at several epithelial foci. Conclusions: These findings indicate that TP53 abnormalities are early and frequent events in the pathogenesis of GBC, starting from chronic cholecystitis.