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Biomedical subjects

W B Isaacs

Publications and source records attributed to W B Isaacs.

At least 19 recordsLinked to original sources
PubMed2001

Cyclooxygenase-2 is up-regulated in proliferative inflammatory atrophy of the prostate, but not in prostate carcinoma.

Cyclooxygenase-2 (COX-2) is the inducible isoform of the rate-limiting enzymes that convert arachidonic acid to proinflammatory prostaglandins as well as a primary target for nonsteroidal anti-inflammatory drugs. Accumulating evidence suggests that up-regulation of COX-2 is associated with carcinogenesis in multiple organ systems including the large bowel, lung, breast, and prostate. In this report, we examine the expression of COX-2 protein and mRNA in prostate tissue containing various lesions and in prostate cancer cell lines. In the cell lines, LNCaP, DU145, PC-3, and TSU, COX-2 protein expression was undetectable under basal conditions but could be induced transiently by phorbol ester treatment in PC-3 and TSU cells, but not in DU145 and LNCaP cells. Immunohistochemical analysis of 144 human prostate cancer cases suggested that, in contrast to several previous reports, there was no consistent overexpression of COX-2 in established prostate cancer or high-grade prostatic intraepithelial neoplasia, as compared with adjacent normal prostate tissue. Positive staining was seen only in scattered cells (<1%) in both tumor and normal tissue regions but was much more consistently observed in areas of proliferative inflammatory atrophy, lesions that have been implicated in prostatic carcinogenesis. Staining was also seen at times in macrophages. Western blotting and quantitative RT-PCR analyses confirmed these patterns of expression. These results suggest that if nonsteroidal anti-inflammatory drugs are indeed chemopreventive and/or chemotherapeutic for prostate cancer, their effects are likely to be mediated by modulating COX-2 activity in non-PCa cells (either inflammatory cells or atrophic epithelial cells) or by affecting a COX-2-independent pathway.

Atrophy↗
PubMed2001

Linkage and association of CYP17 gene in hereditary and sporadic prostate cancer.

Androgens are essential for prostate development, growth and maintenance and the association between androgen levels and prostate cancer is well established. Since the CYP17 gene encodes the enzyme cytochrome P450c17alpha, which mediates 17alpha-hydroxylase and 17,20-lyase activities in the androgen biosynthesis pathway, sequence variations in the gene and association with increased risk to prostate cancer has been studied. In particular, several groups have studied the association between a polymorphism in the 5' promoter region and prostate cancer using a population-based association approach. However, the results from these studies were inconclusive. To further study this polymorphism and its possible role in hereditary prostate cancer (HPC), we performed a genetic linkage analysis and family-based association analysis in 159 families, each of which contains at least 3 first-degree relatives with prostate cancer. In addition, we performed a population-based association analysis to compare the risk of this polymorphism to hereditary and sporadic prostate cancer in 159 HPC probands, 249 sporadic prostate cancer patients and 211 unaffected control subjects. Evidence for linkage at the CYP17 gene region was found in the total 159 HPC families (LOD = 1.3, p = 0.01, at marker D10S222). However, family-based association tests did not provide evidence for overtransmission of either allele of the CYP17 polymorphism to affected individuals in the HPC families. The allele and genotype frequencies of the polymorphism were not statistically different among the HPC probands, sporadic cases and unaffected control subjects. In conclusion, our results suggest that the CYP17 gene or other genes in the region may increase the susceptibility to prostate cancer in men; however, the polymorphism in the 5' promoter region has a minor role if any in increasing prostate cancer susceptibility in our study sample.

Adult↗
PubMed2001

Methylation and mutational analysis of p27(kip1) in prostate carcinoma.

BACKGROUND: We have previously identified 12p12-13 as a region of frequent genetic loss in prostate carcinoma. A candidate tumor suppressor gene at this locus is the cyclin dependent kinase inhibitor p27(kip1), which has been implicated as a marker of aggressive prostate carcinoma. Herein, we examine metastatic prostate tumors, xenografts, and cell lines for gene inactivation via mutational inactivation or promoter hypermethylation. METHODS: Mutation analysis was performed on metastatic prostate tumors of 18 patients, eight prostate carcinoma cell lines, and 18 xenografts by PCR amplification of the entire open reading frame of p27(kip1). PCR products were sequenced directly using internal primers. Methylation analysis was performed on four cell lines and nine xenografts using direct sequencing of cloned PCR products of bisulfite treated DNA. Presence of a CpG was consistent with methylation of that cytosine in the original sample. RESULTS: With the exception of the previously reported homozygous deletion, no additional mutations were identified. Methylated CpG residues were identified in three xenografts (LuCAP23, LuCAP35, and PC82) and the methylated residues clustered at six sites; the cytosines 69, 149, 191, 286, 349, and 487 base pairs 5' of the ATG start codon. However, no sample demonstrated promotor methylation in all sequenced clones and the number of methylated base pairs ranged from seven to three, not the level usually associated with gene silencing. CONCLUSIONS: Mutational inactivation of p27(kip1) is a rare event in metastatic prostate carcinoma. While CpG methylation does occur, it is an infrequent event and does not appear to be the mechanism of p27(kip1) down regulation in prostate carcinoma.

Cell Cycle Proteins↗
PubMed2001

A novel human cell culture model for the study of familial prostate cancer.

Research into molecular and genetic mechanisms underlying familial prostate cancer would be greatly advanced by in vitro models of prostate tumor cells representing primary tumors. We have successfully established an immortalized human prostate epithelial cell culture derived from primary tumors of familial prostate cancer patients with telomerase. The actively proliferating early-passaged 957E cells were transduced through infection with a retrovirus expressing the human telomerase catalytic subunit, human telomerase reverse transcriptase (hTERT). A high level of telomerase activity was detected in 957E/hTERT cells, but not in 957E cells. 957E/hTERT cells are currently growing well at passage 40, whereas 957E cells senesced at passage 5. 957E/hTERT cells exhibit epithelial morphology. Expression of an androgen-regulated prostate specific homeobox gene NKX3.1 and an epithelial cell-specific cytokeratin 8, but not prostate specific antigen or androgen receptor, was detected in 957E/hTERT cells. Prostatic stem cell antigen and p16 were also expressed in this line. 957E/hTERT cells showed growth inhibition when exposed to retinoic acid and transforming growth factor beta1, potent inhibitors of prostate epithelial cell growth. Chromosome analysis showed that the 957E/hTERT cell line (passage 10) was near diploid human male (XY), with most chromosome counts in the 44-46 range. However, there was random loss of chromosomes 8, 13, X, Y, and alteration in chromosome 4q. The late passage 957E/hTERT cell line (passage 32) was karyologically similar to the early passage 957E/hTERT cell line (passage 10) and also had the same alteration of 4q observed in the early passage 957E/hTERT cell line (passage 10) as well as a trisomy of chromosome 20. The well-characterized human cancer lines derived from such patients will be useful for the identification and characterization of prostate cancer susceptibility genes. This is the first documented case of an established human prostate cancer cell line from primary tumor of a familial prostate cancer patient.

Adenocarcinoma↗
PubMed2001

Linkage and association studies of prostate cancer susceptibility: evidence for linkage at 8p22-23.

Multiple lines of evidence have implicated the short arm of chromosome 8 as harboring genes important in prostate carcinogenesis. Although most of this evidence comes from the identification of frequent somatic alterations of 8p loci in prostate cancer cells (e.g., loss of heterozygosity), studies have also suggested a role for 8p genes in mediation of inherited susceptibility to prostate cancer. To further examine this latter possibility, we performed linkage analyses, in 159 pedigrees affected by hereditary prostate cancer (HPC), using 24 markers on the short arm of chromosome 8. In the complete set of families, evidence for prostate cancer linkage was found at 8p22-23, with a peak HLOD of 1.84 (P=.004), and an estimate of the proportion of families linked (alpha) of 0.14, at D8S1130. In the 79 families with average age at diagnosis >65 years, an allele-sharing LOD score of 2.64 (P=.0005) was observed, and six markers spanning a distance of 10 cM had LOD scores >2.0. Interestingly, the small number of Ashkenazi Jewish pedigrees (n=11) analyzed in this study contributed disproportionately to this linkage. Mutation screening in HPC probands and association analyses in case subjects (a group that includes HPC probands and unrelated case subjects) and unaffected control subjects were carried out for the putative prostate cancer-susceptibility gene, PG1, previously localized to the 8p22-23 region. No statistical differences in the allele, genotype, or haplotype frequencies of the SNPs or other sequence variants in the PG1 gene were observed between case and control subjects. However, case subjects demonstrated a trend toward higher homozygous rates of less-frequent alleles in all three PG1 SNPs, and overtransmission of a PG1 variant to case subjects was observed. In summary, these results provide evidence for the existence of a prostate cancer-susceptibility gene at 8p22-23. Evaluation of the PG1 gene and other candidate genes in this area appears warranted.

Age of Onset↗
PubMed2001

Human prostate cancer and benign prostatic hyperplasia: molecular dissection by gene expression profiling.

Critical aspects of the biology and molecular basis for prostate malignancy remain poorly understood. To reveal fundamental differences between benign and malignant growth of prostate cells, we performed gene expression profiling of primary human prostate cancer and benign prostatic hyperplasia (BPH) using cDNA microarrays consisting of 6500 human genes. Frozen prostate specimens were processed to facilitate extraction of RNA from regions of tissue enriched in either benign or malignant epithelial cell growth within a given specimen. Gene expression in each of the 16 prostate cancer and nine BPH specimens was compared with a common reference to generate normalized measures for each gene across all of the samples. Using an analysis of complete pairwise comparisons of expression profiles among all of the samples, we observed clearly discernable patterns of overall gene expression that differentiated prostate cancer from BPH. Further analysis of the data identified 210 genes with statistically significant differences in expression between prostate cancer and BPH. These genes include many not recognized previously as differentially expressed in prostate cancer and BPH, including hepsin, which codes for a transmembrane serine protease. This study reveals for the first time that significant and widespread differences in gene expression patterns exist between benign and malignant growth of the prostate gland. Gene expression analysis of prostate tissues should help to disclose the molecular mechanisms underlying prostate malignant growth and identify molecular markers for diagnostic, prognostic, and therapeutic use.

Gene Expression Profiling↗
PubMed2001

Evaluation of linkage and association of HPC2/ELAC2 in patients with familial or sporadic prostate cancer.

To investigate the relationship between HPC2/ELAC2 and prostate cancer risk, we performed the following analyses: (1) a linkage study of six markers in and around the HPC2/ELAC2 gene at 17p11 in 159 pedigrees with hereditary prostate cancer (HPC); (2) a mutation-screening analysis of all coding exons of the gene in 93 probands with HPC; (3) family-based and population-based association study of common HPC2/ELAC2 missense variants in 159 probands with HPC, 249 patients with sporadic prostate cancer, and 222 unaffected male control subjects. No evidence for linkage was found in the total sample, nor in any subset of pedigrees based on characteristics that included age at onset, number of affected members, male-to-male disease transmission, or race. Furthermore, only the two previously reported missense changes (Ser217Leu and Ala541Thr) were identified by mutational analysis of all HPC2/ELAC exons in 93 probands with HPC. In association analyses, family-based tests did not reveal excess transmission of the Leu217 and/or Thr541 alleles to affected offspring, and population-based tests failed to reveal any statistically significant difference in the allele frequencies of the two polymorphisms between patients with prostate cancer and control subjects. The results of this study lead us to reject the three alternative hypotheses of (1) a highly penetrant, major prostate cancer-susceptibility gene at 17p11, (2) the allelic variants Leu217 or Thr541 of HPC2/ELAC2 as high-penetrance mutations, and (3) the variants Leu217 or Thr541 as low-penetrance, risk-modifying alleles. However, we did observe a trend of higher Leu217 homozygous carrier rates in patients than in control subjects. Considering the impact of genetic heterogeneity, phenocopies, and incomplete penetrance on the linkage and association studies of prostate cancer and on the power to detect linkage and association in our study sample, our results cannot rule out the possibility of a highly penetrant prostate cancer gene at this locus that only segregates in a small number of pedigrees. Nor can we rule out a prostate cancer-modifier gene that confers a lower-than-reported risk. Additional larger studies are needed to more fully evaluate the role of this gene in prostate cancer risk.

Age of Onset↗
PubMed2001

Protection against 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine cytotoxicity and DNA adduct formation in human prostate by glutathione S-transferase P1.

The prostate has been identified as a target for 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-induced carcinogenesis. Humans are exposed to PhIP through ingestion of well-done cooked meats, and there is evidence from epidemiological studies that implicates red meat consumption in prostate carcinogenesis. The alpha and pi class isoforms of glutathione S-transferases (GSTs) have been shown to inhibit adduction of activated PhIP metabolites to DNA in cell-free systems. In humans, silencing of GST pi(GSTP1) through CpG island hypermethylation is found in nearly all prostate carcinomas and is believed to be an early event in prostate carcinogenesis. We hypothesized that suppressed GSTP1 expression in prostate cells would increase their vulnerability to cytotoxicity and DNA adduct formation mediated by activated PhIP metabolites. To test this hypothesis, the human prostate adenocarcinoma cell line, LNCaP, which contains a silenced GSTP1 gene, was genetically modified to constitutively express high levels of GSTP1. Both LNCaP and LNCaP-GSTP1 cells exposed to N-OH-PhIP, but not parent PhIP, for 24 h showed a dose-dependent decrease in cell viability. GSTP1-overexpressing cells had LC50s 30-40% higher than cells transfected with the vector alone. PhIP-DNA adducts isolated from LNCaP-derived cells and primary human prostate tissue cultures exposed to N-OH-PhIP were analyzed by liquid chromatography/electrospray ionization mass spectrometry. Primary cultures of human prostate tissue and LNCaP-GSTP1 cells had approximately 50% lower adduct levels than parental LNCaP and vector control cells. Bioactivation assays using LNCaP cytosols showed that enzymatic activation of N-OH-PhIP to a DNA binding species was dependent on ATP and could be inhibited by recombinant human GSTP1 in the presence of glutathione. This evidence confirms that N-OH-PhIP can be bioactivated to a DNA binding species in human prostate and human prostate-derived cells. These observations provide the basis for using LNCaP and LNCaP-GSTP1 cells as a model system for studying the role of this enzyme in protection against N-OH-PhIP induced DNA damage in prostate carcinogenesis. Loss of GSTP1 expression in human prostate may, therefore, enhance its susceptibility to carcinogenic insult by compounds such as N-OH-PhIP. Conversely, induction of GSTs in early-stage prostate carcinogenesis may be a useful protective strategy.

Adenocarcinoma↗
PubMed2001

Multiple antibodies to titin immunoreact with AHNAK and localize to the mitotic spindle machinery.

Recently, the large filamentous striated-muscle protein titin has been observed in non-muscle cells, and, in one instance, has been proposed to have a nuclear function as a chromosomal component contributing to structure and elasticity. In this study, we sought to further characterize the presumptive nuclear isoform of titin. Immunofluorescence microscopy with multiple titin-specific monoclonal antibodies shows localization to the nucleus in interphase cells and to the spindle machinery in mitotic cells in all cell types examined; localization to condensed chromosomes is not observed. An abundant 700-kDa phosphoprotein is the predominant species immunoprecipitated with these antibodies. Sequencing of peptide fragments of the immunopurified protein reveals identity to AHNAK, a nuclear phosphoprotein, an identification that was confirmed by Western blot analysis with antibodies to AHNAK and peptide fragmentation patterns. Sequence comparison suggests similarities between the repetitive heptad phi+/-phiP+/-phi+/- motif in AHNAK and the PEVK region of titin, potentially explaining the cross-reactivity observed between AHNAK antibodies and titin antibodies. Interestingly, although some AHNAK antibodies stain interphase nuclei, no evidence of mitotic spindle localization is seen, suggesting that the identity of the protein at the latter location is more closely related to titin than AHNAK. This concept is further supported by observations that cell lines not expressing AHNAK have similar antititin antibody localization to the mitotic spindle. We conclude that (1) multiple titin antibodies, particularly those recognizing the PEVK region, cross-react with AHNAK, and (2) the mitotic spindle staining observed with antititin antibodies is most likely due to the association of titin or a titin-like molecule with this structure.

Amino Acid Motifs↗
PubMed2001

Linkage of prostate cancer susceptibility loci to chromosome 1.

Three prostate cancer susceptibility genes have been reported to be linked to different regions on chromosome 1: HPC1 at 1q24-25, PCAP at 1q42-43, and CAPB at 1p36. Replication studies analyzing each of these regions have yielded inconsistent results. To evaluate linkage across this chromosome systematically, we performed multipoint linkage analyses with 50 microsatellite markers spanning chromosome 1 in 159 hereditary prostate cancer families (HPC), including 79 families analyzed in the original report describing HPC1 linkage. The highest lod scores for the complete dataset of 159 families were observed at 1q24-25 at which the parametric lod score assuming heterogeneity (hlod) was 2.54 (P=0.0006) with an allele sharing lod of 2.34 (P=0.001) at marker D1S413, although only weak evidence was observed in the 80 families not previously analyzed for this region (hlod=0.44, P=0.14, and allele sharing lod=0.67, P=0.08). In the complete data set, the evidence for linkage across this region was very broad, with allele sharing lod scores greater than 0.5 extending approximately 100 cM from 1p13 to 1q32, possibly indicating the presence of multiple susceptibility genes. Elsewhere on chromosome 1, some evidence of linkage was observed at 1q42-43, with a peak allele sharing lod of 0.56 (P=0.11) and hlod of 0.24 (P=0.25) at D1S235. For analysis of the CAPB locus at 1p36, we focused on six HPC families in our collection with a history of primary brain cancer; four of these families had positive linkage results at 1p36, with a peak allele sharing lod of 0.61 (P=0.09) and hlod of 0.39 (P=0.16) at D1S407 in all six families. These results are consistent with the heterogeneous nature of hereditary prostate cancer, and the existence of multiple loci on chromosome 1 for this disease.

Chromosome Mapping↗
PubMed2001

Evidence for a prostate cancer linkage to chromosome 20 in 159 hereditary prostate cancer families.

Prostate cancer is the most common malignancy diagnosed in men in the US. Genetic susceptibility to prostate cancer has been well documented. A region at chromosome 20q13 (HPC20) has been reported to be linked to a prostate cancer susceptibility gene. To confirm this finding, we genotyped 16 markers spanning approximately 95 cM on chromosome 20 in 159 hereditary prostate cancer (HPC) families. Positive (but not statistically significant) linkage scores were observed from 20pter to 20q11, with the highest non-parametric linkage (NPL) score for the complete dataset of 1.02 (P=0.15) being observed at D20S195 at 20q11. Evidence for linkage from parametric analyses with a dominant or a recessive model was weak. Interestingly, consistent with the original findings of linkage to 20 g higher linkage scores were observed in the subsets of families with a later age at diagnosis (> or =65 years; n=80, NPL=1.94, P=0.029 at D20S186), fewer than five affected family members (n=69, NPL=1.74, P=0.037 at D20S889), or without male-to-male disease transmission (n=60, NPL=1.01, P=0.15 at D20S117). The region with positive linkage scores spanned approximately 60 cM from 20pter to 20q11 in these subsets of families. Our results are consistent with a prostate cancer susceptibility locus on chromosome 20.

Aged↗
PubMed2001

GSTP1 CpG island hypermethylation is responsible for the absence of GSTP1 expression in human prostate cancer cells.

GSTP1 CpG island hypermethylation is the most common somatic genome alteration described for human prostate cancer (PCA); lack of GSTP1 expression is characteristic of human PCA cells in vivo. We report here that loss of GSTP1 function may have been selected during the pathogenesis of human PCA. Using a variety of techniques to detect GSTP1 CpG island DNA hypermethylation in PCA DNA, we found only hypermethylated GSTP1 alleles in each PCA cell in all but two PCA cases studied. In these two cases, CpG island hypermethylation was present at only one of two GSTP1 alleles in PCA DNA. In one of the cases, DNA hypermethylation at one GSTP1 allele and deletion of the other GSTP1 allele were evident. In the other case, an unmethylated GSTP1 allele was detected, accompanied by abundant GSTP1 expression. GSTP1 CpG island DNA hypermethylation was responsible for lack of GSTP1 expression by LNCaP PCA cells: treatment of the cells with 5-azacytidine (5-aza-C), an inhibitor of DNA methyltransferases, reversed the GSTP1 promoter DNA hypermethylation, activated GSTP1 transcription, and restored GSTP1 expression. GSTP1 promoter activity, assessed via transfection of GSTP1 promoter-CAT reporter constructs in LNCaP cells, was inhibited by SssI-catalyzed CpG dinucleotide methylation. Remarkably, although selection for loss of GSTP1 function may be inferred for human PCA, GSTP1 did not act like a tumor suppressor gene, as LNCaP cells expressing GSTP1, either after 5-aza-C treatment or as a consequence of transfection with GSTP1 cDNA, grew well in vitro and in vivo. Perhaps, GSTP1 inactivation may render prostatic cells susceptible to additional genome alterations, caused by electrophilic or oxidant carcinogens, that provide a selective growth advantage.

Alleles↗
PubMed2000

Detection and analysis of beta-catenin mutations in prostate cancer.

BACKGROUND: E-cadherin and alpha-catenin are components of adherens junctions which mediate calcium-dependent, cell-cell adhesion in a homotypic manner. Both these molecules have been defined as useful tumor markers as their altered expression correlates with increased tumor aggressiveness and dedifferentiation. More recently, alterations of a third component of adherens junctions, beta-catenin, have been observed to play a role in several human cancers. Dysregulation of beta-catenin, either by direct mutation or by defects in interacting pathways/regulators, can result in its cytoplasmic accumulation and nuclear translocation. In the nucleus, beta-catenin forms a transcriptional complex capable of upregulating target genes, many of which encode proliferative factors. Given its oncogenic activity and connection to human cancer, we examined the beta-catenin gene and its expression in prostate cancer. METHODS: By single-stranded conformational polymorphism (SSCP) and DNA sequencing analyses, we screened exon 3 of beta-catenin from a panel of 81 primary tumors obtained at radical prostatectomy, 22 lymph node metastases from untreated patients, and a unique set of 61 metastatic tissues from 19 patients who died of hormone-refractory disease. RESULTS: We found putative activating mutations (missense and deletion) at a rate of 5% (7/138). One patient had the same 72 base pair deletion in each of nine separate metastases examined, indicating that this change was associated with a clonal population of metastatic cells. CONCLUSIONS: Immunohistological staining of mutation-positive tumors demonstrated beta-catenin accumulation and nuclear localization in a heterogeneous fashion. Consistent with this in vivo finding, our in vitro analyses demonstrate that certain mutations can result in increased beta-catenin nuclear activity in prostate cancer cell lines. These data implicate the beta-catenin signaling pathway in the development of a subset of prostate cancers.

Animals↗
PubMed2000

GSTP1 CpG island DNA hypermethylation in hepatocellular carcinomas.

Glutathione S-transferases, enzymes that defend cells against damage mediated by oxidant and electrophilic carcinogens, may be critical determinants of cancer pathogenesis. We report here that the pathogenesis of hepatocellular carcinoma (HCC), one of the most common cancers in the world, frequently involves an accumulation of somatic DNA methylation changes at GSTP1, the gene encoding the pi-class glutathione S-transferase. For our study, Hep3B HCC cells and a cohort of 20 HCC tissue specimens were subjected to analysis for GSTP1 expression and for somatic GSTP1 alterations. GSTP1 DNA hypermethylation in HCC DNA was assessed by Southern blot analysis, via a polymerase chain reaction (PCR) assay, and by using a genomic sequencing approach. Hep3B HCC cells failed to express GSTP1 mRNA or GSTP1 polypeptides. Similarly, HCC cells in 19 of 20 HCC cases were devoid of GSTP1 polypeptides. By Southern blot analysis, DNA from Hep3B HCC cells displayed abnormal GSTP1 hypermethylation. Treatment of Hep3B HCC cells in vitro with the DNA methyltransferase inhibitor 5-aza-deoxycytidine both reversed GSTP1 DNA hypermethylation and restored GSTP1 expression. Using a PCR assay, somatic GSTP1 DNA hypermethylation was also detected in HCC DNA from 17 of 20 HCC cases. Genomic sequencing analyses, undertaken to map 5-methyldeoxycytidine nucleotides located at the GSTP1 transcriptional regulatory region, frequently detected somatic DNA hypermethylation near the gene promoter in HCC DNA. The data indicate that GSTP1 DNA hypermethylation changes appear frequently in human HCC. In addition, the data raise the possibility that somatic GSTP1 inactivation, via hypermethylation, may contribute to the pathogenesis of HCC.

Adult↗
PubMed2000

Loss of heterozygosity at 12P12-13 in primary and metastatic prostate adenocarcinoma.

PURPOSE: Our laboratory has recently identified a 1 to 2 Mb homozygous deletion at 12p12-13 in a prostate cancer specimen and determined that the p27/kip1 gene lies within the deletion. While immunohistochemical analysis has implicated p27/kip1 in prostate carcinoma, no previous studies had identified genetic abnormalities at this locus. Here, we examined primary and metastatic prostate tumors to determine if allelic loss occurs at this locus in localized disease and if it increases the risk of metastatic, high stage or high-grade disease. MATERIALS AND METHODS: DNA was extracted from prostate tumors and normal tissue of 99 patients. 60 tumors were primary, 20 were metastatic pelvic lymph nodes, and 19 were distant metastases. Multiple metastases were analyzed from 11 of 19 patients with metastatic disease. Polymorphic markers spanning our region of interest were PCR amplified from tumor and normal DNA. PCR products were then scored for allelic loss. RESULTS: Loss of heterozygosity (LOH) was identified in 14/60 (23%) primary tumors, 6/20 (30%) lymph node metastasis, and 9/19 (47%) distant metastases. The difference between primary and distant metastatic disease was statistically significant (p = 0.045, Fisher's exact test). The pattern of LOH was identical in all metastatic sites obtained from individual patients, indicating that genetic loss occurred prior to metastasis. Subset analysis of the 60 primary tumors demonstrated no association between LOH and adverse pathological feature [nodal involvement, seminal vesicle invasion, margin positivity, high Gleason score (7-10)]. CONCLUSIONS: Demonstrating that 12p12-13 LOH is a prominent feature of primary prostate tumors and that multiple metastatic foci have an identical LOH pattern, provides evidence that gene inactivation in this region occurs prior to metastasis. In addition, the strong association between LOH and distant metastasis raises the possibility that mutational inactivation of a gene at 12p12-13, possibly p27/kip1, plays a pivotal role in the development of metastatic disease.

Adenocarcinoma↗