An inherited mutation outside the highly conserved DNA-binding domain of the p53 tumor suppressor protein in children and adults with sporadic adrenocortical tumors

Mutations of the p53 tumor suppressor gene are the single most common genetic alterations in human cancers. Recently, a distinct nucleotide substitution was identified in exon 10 of the p53 gene, leading to an Arg337His mutation in 97% of children with adrenocortical tumors from Southern Brazil. In the present study, we investigated the presence of this mutation in a larger series of 55 patients (37 adults and 18 children) with benign and malignant sporadic adrenocortical tumors. None of the patients had family cancer histories that conformed to the criteria for Li-Fraumeni syndrome. Twenty-one asymptomatic close relatives of patients with p53 mutations and 60 normal unrelated individuals were also studied. The missense Arg337His mutation was identified in 19 patients (14 children and 5 adults), and 8 of 11 cases studied had LOH. Among the 19 patients with the Arg337His mutation, only one boy and three adults showed fatal evolution or recurrent metastases. This mutation was also identified in heterozygous state in asymptomatic first-degree relatives of the patients, indicating that Arg337His mutation was inherited in most cases. In contrast, this mutation was not found in 120 alleles of normal unrelated controls. In conclusion, the germ line Arg337His mutation of p53 protein is present at a high frequency (77.7%) in children with benign or malignant sporadic adrenocortical tumors, but it is not restricted to the pediatric group, since 13.5% of adults with adrenocortical tumors also had this mutation. The presence of this mutation was related to unfavorable prognosis in most of the adults, but not in the children with adrenocortical tumors.     

Founder effect for the highly prevalent R337H mutation of tumor suppressor p53 in Brazilian patients with adrenocortical tumors

The incidence of adrenocortical tumors in children from the Southern region of Brazil is higher than in other parts of the world. This fact has been related to the identification of an inherited missense mutation of the p53 (R337H) at high frequency (78-97%) in Brazilian children with adrenocortical tumors. Given the high frequency of this germline mutation in the Brazilian population, it is very likely that the R337H mutation has arisen from a common origin. In this study, we analyzed two highly polymorphic intragenic markers (VNTRp53 and p53CA) in 22 patients (16 children and 6 adults) with adrenocortical tumors carrying the germline R337H mutation and 60 normal individuals using GeneScan Fragment Analysis software. We found six and sixteen different alleles for the VNTRp53 and p53CA polymorphic markers, respectively. Two distinct alleles, both with 122 bp, were found in 56.8% (VNTRp53) and 54.5% (p53CA) of the 44 alleles from patients with adrenocortical tumors associated with the R337H mutation. Differently, these same VNTRp53 and p53CA alleles were found in 18.3% and 14.2% of 120 alleles from normal individuals, respectively (p<0.01, Chi-square test). An identical haplotype for p53 locus was also identified in 95% of the apparently unrelated Brazilian patients with adrenocortical tumors carrying the R337H mutation. In conclusion, we demonstrated a strong evidence of co-segregation between two intragenic polymorphic p53 markers and the germline R337H mutation, indicating that this mutation has originated from a single common ancestral in the great majority of the Brazilian patients with adrenocortical tumors.     

Clinical and molecular aspects of a pediatric metachronous adrenocortical tumor

The occurrence of metachronous adrenocortical carcinoma has rarely been described. We report a case of a child with virilizing adrenocortical metachronous tumors that, despite several metastases, presented long-term survival (15 years). We analyzed in this tumor IGF2, IGF1R and FGFR4 gene expression, and evaluated the presence of p.R337H germline p53 mutation and somatic CTNNB1 mutation. IGF2 gene was over-expressed in both left (Weiss score 5) and right (Weiss 7) adrenocortical tumors. IGF1R expression levels were higher in the right adrenocortical tumor. FGFR4 over-expression was also detected in the right adrenocortical tumor. In addition, this patient harbors the germline p.R337H p53 mutation and loss of heterozygosity (LOH) was detected in the tumors. No somatic CTNNB1 mutations were found in both tumors. In conclusion, we demonstrated in this unusual case the over-expression of growth signaling pathways, which are molecular mechanisms previously related to adrenocortical tumorigenesis. Furthermore, the absence of somatic CTNNB1 mutations, which is a molecular marker of poor prognosis in adults, might be related to the long-term survival of this patient.     

Reduction to homozygosity involving p53 in esophageal cancers demonstrated by the polymerase chain reaction.

Loss of heterozygosity affecting chromosome 17p has been detected at high frequencies in a variety of human tumors, including cancers of the colon, breast, lung, and brain. One presumed target of these losses is p53, a tumor suppressor gene located on 17p. To our knowledge, loss of heterozygosity has not yet been reported at any locus, including p53, in human esophageal cancer. Moreover, current methods of detecting loss of heterozygosity depend on the availability of large amounts of high molecular weight DNA, making the study of small biopsy specimens or paraffin-embedded tissues problematic. We examined 52 primary human esophageal neoplasms for loss of heterozygosity affecting the p53 gene by using the polymerase chain reaction. Loss of one allele was detected in 52% of informative cases and was more common in squamous carcinomas than in adenocarcinomas. Southern blot analysis was used to confirm polymerase chain reaction-derived data. The identification of allelic loss in approximately half of the tumors analyzed supports the hypothesis that inactivation of p53 is involved in the pathogenesis of esophageal cancer.     

Association Between Recurrence of Sporadic Colorectal Cancer,High Level of Microsatellite Instability,and Loss of Heterozygosity at Chromosome 18q

PURPOSE: Microsatellite instability and loss of heterozygosity of chromosomes 18q, 8p, and 4p are genetic alterations commonly found in colorectal cancer. We investigated whether these genetic markers allow for the stratification of patients with Stage II to III colorectal cancer into groups with different recurrence risks, and with different prognoses.METHODS: Tumors of 113 patients were evaluated for loss of heterozygosity of chromosomes 18q, 8p, and 4p and for microsatellite instability by use of six microsatellite markers. Genetic alterations involving each of these genetic markers were examined for association with disease recurrences and survival.RESULTS: Loss of heterozygosity of chromosomes 18q, informative in 96 percent of cases, in Stage III tumors was associated with higher risk of overall recurrence (P < 0.001), local recurrence (P < 0.001), distant metastases (P < 0.001), decreased overall survival (P = 0.002), and disease-free survival (P < 0.001). The recurrence rates and survival rates among patients with Stage II colorectal cancer were independent of loss of heterozygosity of chromosome 18q. Stage III and loss of heterozygosity of chromosome 8p also were associated with a higher risk of recurrences when these factors were considered individually. In multivariate analysis, only loss of heterozygosity of chromosome 18q was independently associated with risk of recurrences (P < 0.001) and with disease-free survival (P = 0.001). No correlation was observed between microsatellite instability and recurrence rates. However, microsatellite instability was associated with improved overall survival (P = 0.04) and with a longer disease-free interval (P = 0.002). Only in five cases (16.7 percent) was it possible to perform resection of recurrences; two of these patients had microsatellite instability tumor. In no cases was it possible to resect recurrence of tumors with loss of heterozygosity of chromosome 18q.CONCLUSIONS: Loss of heterozygosity of chromosome 18q is an informative genetic marker, which in resected Stage III colorectal cancer can be used to predict recurrences and survival. Microsatellite instability identified cases that, even in the case of recurrence, have a more favorable prognosis.Supported by a grant from Lega Italiana per la Lotta Contro i Tumori, Sezione di Parma.     

Genotyping of adrenocortical tumors: very frequent deletions of the MEN1 locus in 11q13 and of a 1-centimorgan region in 2p16

To identify chromosomal regions that may contain loci for tumor suppressor genes involved in adrenocortical tumor development, a panel of 60 tumors (39 carcinomas and 21 adenomas) were screened for loss of heterozygosity. Although the vast majority of loss of heterozygosity (LOH) were detected in the carcinomas and involved chromosomes 2, 4, 11, and 18, only few were found in the adenomas. Therefore, 2 loci that harbor the familial cancer syndromes Carney complex in 2p16 and the multiple endocrine neoplasia type 1 gene in 11q13 were further studied in 27 (13 carcinomas and 14 adenomas) of the 60 tumors. Detailed analysis of the 2p16 region mapped a minimal area of overlapping deletions to a 1-centimorgan region, which is separate from the Carney complex locus. LOH for a microsatellite marker (PYGM), very close to the MEN1 gene, was detected in all 8 informative carcinomas (100%) and in 2 of 14 adenomas. Of the 27 cases analyzed in detail, 13 cases (11 carcinomas and 2 adenomas) showed LOH on chromosome 11 and was therefore selected for MEN1 gene mutation analysis. In 6 cases a common polymorphism (Asp418Asp) was found, but no mutation was detected. In conclusion, our data indicate the existence of tumor suppressor genes at multiple chromosomal locations, whose inactivations are involved in the development of adrenocortical carcinomas. Loss of genetic material from 2p16 was strongly associated with the malignant phenotype, as it was seen in almost all carcinomas but not in any of the adenomas. LOH in 11q13 also occurred frequently in the carcinomas, but was not associated with a MEN1 mutation, suggesting the involvement of a different tumor suppressor gene on this chromosome.     

Allelic loss of chromosomes 16q and 10q in human prostate cancer.

Recent advances in understanding the molecular genetics of common adult tumors have indicated that multiple genetic alterations including the activation of oncogenes and the inactivation of tumor suppressor genes are important in the pathogenesis of these tumors. Loss of heterozygosity is a hallmark of tumor suppressor gene inactivation and has been used to identify chromosomal regions that contain these genes. We have examined allelic loss in the most common tumor in men, prostate cancer. Twenty-eight prostate cancer specimens have been examined for loss of heterozygosity at 11 different chromosomal arms including 3p, 7q, 9q, 10p, 10q, 11p, 13q, 16p, 16q, 17p, and 18q. Fifty-four percent (13/24) of clinically localized tumors and 4 of 4 metastatic tumors showed loss of heterozygosity on at least one chromosome. Chromosomes 16q and 10q exhibited the highest frequency of loss of heterozygosity with 30% of tumors showing loss at these chromosomes. These data demonstrate that allelic loss is a common event in prostate cancer and suggest that chromosomes 16q and 10q may contain the sites of tumor suppressor genes important in the pathogenesis of human prostate cancer.     

Loss of heterozygosity on chromosomes 3, 13, and 17 in small-cell carcinoma and on chromosome 3 in adenocarcinoma of the lung.

By a molecular genetic approach using polymorphic DNA markers that detect allelic deletion of specific chromosomal regions, we analyzed for possible loss of chromosomal heterozygosity in five different histological types of lung cancers obtained from 47 patients. In small-cell carcinomas, the incidence of allelic deletions at three different chromosomal loci was extremely high; loss of heterozygosity was detected on chromosomes 3p in 7 of 7 patients (100%), 13q in 10 of 11 patients (91%), and 17p in 5 of 5 patients (100%). The deletions at these loci in small-cell carcinomas were observed even in the tumors without any clinical evidence of metastasis. Furthermore, loss of heterozygosity on chromosomes 3p and 13q occurred prior to NMYC amplification and chromosome 11p deletion. Loss of heterozygosity on chromosome 3p was also detected with high frequency in adenocarcinomas [5 of 6 patients (83%)]. Heterozygosity of chromosomes 13q and 17p was lost in 10 of 31 patients (32%) and in 3 of 12 patients (25%), respectively, of lung cancers other than small-cell carcinomas. These results indicate that recessive genetic changes involving sequences on chromosomes 3p, 13q, and 17p may play important roles in the genesis of small-cell carcinoma, and those on chromosome 3p may play an important role in the genesis of adenocarcinoma.     

Mitotic recombination of chromosome 17 in astrocytomas.

Allelic combinations at seven loci on human chromosome 17 defined by restriction fragment length polymorphisms were determined in tumor and normal tissues from 35 patients with gliomas. Loss of constitutional heterozygosity at one or more of these loci was observed in 8 of the 24 tumors displaying astrocytic differentiation and in the single primitive neuroectodermal tumor examined. The astrocytomas showing these losses included examples of each adult malignancy grade of the disease, including glioblastoma (malignancy grade IV), and seven of them demonstrated concurrent maintenance of heterozygosity for at least one chromosome 17 locus. Determination of allele dosage together with the genotypic data indicated that the tumor chromosomes 17 were derived by mitotic recombination in 7 of the 9 cases with shared homozygosity of the region 17p11.2-pter in all cases. In contrast, tumors of oligodendrocytic, ependymal, or mixed cellular differentiation did not exhibit loss of alleles at any of the loci examined. These data suggest that the somatic attainment of homozygosity for loci on chromosome 17p is frequently associated with the oncogenesis of central nervous system tumors, particularly those showing solely astrocytic differentiation, and that mitotic recombination mapping is a useful approach towards the subregional localization of a locus whose rearrangement is involved in this disease.     

Loss of heterozygosity on the short arm of chromosome 17 is associated with high proliferative capacity and DNA aneuploidy in primary human breast cancer.

Loss of heterozygosity (LOH) on the short arm of chromosome 17 (17p) was found in 27 of 52 (52%) previously untreated primary breast cancers. There was a significant correlation between this 17p allelic loss and two parameters associated with aggressive tumor behavior: high cellular proliferative fraction and DNA aneuploidy. These correlations with high cellular proliferative fraction and DNA aneuploidy were not found in tumors with LOH at nine other chromosome locations. The p53 gene, a putative tumor suppressor gene located at 17p13, was examined for aberrations to determine whether it is the target for the 17p LOH in breast cancer. Unlike other types of human cancer, there were no homozygous deletions or rearrangements of the p53 gene, and only 2 of 13 (15%) were mutated in the conserved region where mutational "hot spots" have been previously located. Therefore, we hypothesize that, in breast cancer, either loss or inactivation of gene(s) on chromosome 17p other than the p53 gene or a different mechanism of p53 gene inactivation may be responsible for the observed high labeling index and DNA aneuploidy associated with LOH at 17p.     

Evaluation of PCNA, p53, K-ras and LOH in endocrine pancreas tumors

BACKGROUND/AIMS: The object of this study was to evaluate the characters of the endocrine pancreas tumors including proliferative activity, p53 mutation, K-ras mutation and microsatellite instability. METHODOLOGY: The 13 endocrine tumors of the pancreas were enrolled in this study. There were 8 hypervascular tumors and 4 normo- or hypovascular tumors. All cases were immunohistochemically characterized in paraffin sections for the presence of proliferating cell nuclear antigen and p53 protein. Mutation in K-ras at codon 12 was detected by the Mutant-allele-specific amplification system. Microsatellite instability was examined by using frozen tissues in the 2 cases. RESULTS: Proliferating cell nuclear antigen labeling index range was 0.00-0.62 (0.26 +/- 0.23). p53 was positive in 4/13 tumors. K-ras codon 12 mutation was not detected in any tumors. PCNA LI was significantly lower in hypervascular tumors (0.16 +/- 0.20) than normo- or hypovascular tumors (0.44 +/- 0.17) (P < 0.05). PCNA LI was significantly lower in the p53-positive tumors (0.48 +/- 0.17) than the p53-negative tumors (0.17 +/- 0.18) (P < 0.05). K-ras codon 12 mutation was not detected in any tumors. Loss of heterozygosity in 3p was detected in 1 tumor. CONCLUSIONS: Hypervascular endocrine pancreas tumors have low proliferative activity. p53 mutation influences proliferation as the late event of tumor progression.     

Chromosome instability contributes to loss of heterozygosity in mice lacking p53

The p53 tumor suppressor protein participates in multiple cellular processes including cell cycle checkpoints and programmed cell death. In cell lines, loss of p53 function is associated with increased genetic instability including aneuploidy, gene amplification, and point mutation. Although similar genetic instability often accompanies the progression of malignancy in tumors, its role in tumor initiation in normal cells is not clear. To study whether or not loss of p53 leads to genetic instability in normal cells in vivo, we have examined mechanisms of loss of heterozygosity (LOH) at the Aprt (adenine phosphoribsyltransferase) and flanking loci in normal fibroblasts and T lymphocytes of p53-deficient mice. Somatic cell variants that arose in vivo as a consequence of genetic or epigenetic alterations abolishing Aprt function were selected and expanded in vitro by virtue of their resistance to 2,6-diaminopurine (DAP). We observed that p53 null mice produced about three times as many DAP-resistant fibroblast colonies than wild-type mice, but the frequency of DAP-resistant T lymphocyte colonies was not significantly changed. Mitotic recombination, but not point mutation, partly accounted for the increase in the frequency of DAP-resistant fibroblasts. Most significantly, chromosome loss/duplication and interstitial deletion, which were extremely rare events in the wild-type mice, represented a significant proportion of LOH events in both fibroblasts and T lymphocytes of p53 null mice. Also, increased interstitial deletion was observed in fibroblasts of p53 heterozygous mice. These data suggest that increased genetic variation, including chromosome instability, starts at the initiation stage of tumorigenesis when functional p53 is absent or reduced.     

Constant denaturant gel electrophoresis as a rapid screening technique for p53 mutations.

At present, mutation of the p53 gene appears to be the most common genetic alteration found in human cancers. These mutations can occur within many different regions of the gene. We have developed a modification of denaturing gradient gel electrophoresis termed "constant denaturant gel electrophoresis" (CDGE), which provides a rapid and sensitive method to screen the four conserved regions within the p53 gene where the majority of p53 mutations have been reported. The sensitivity of CDGE was first tested with known p53 mutations in all four conserved regions. The CDGE technique was then used to screen 32 breast carcinomas that had been analyzed by immunohistochemical methods for altered p53 protein levels and whose DNA had already been shown to have loss of heterozygosity for a chromosome 17p marker. By immunostaining techniques, only 6 of the 32 tumors had elevated p53 expression. However, CDGE detected p53 mutations in 11 of the 32 tumors. DNA sequence analysis was performed to determine the nucleotide positions of these mutations in all 11 samples. Loss of heterozygosity for the pYNZ22 or p144D6 markers did not associate with either the loss of heterozygosity at the p53 locus or the mutations detected by CDGE. We conclude that CDGE is a rapid and effective technique to screen for p53 mutations.     

Loss of heterozygosity on chromosome 17p predicts neoplastic progression in Barrett's esophagus

BACKGROUND AND AIM: Endoscopic surveillance for adenocarcinoma in patients with Barrett's esophagus is costly, with one cancer detected every 48-441 patient years of follow up. Genetic abnormalities, including loss of heterozygosity at sites of tumor suppressor genes, have been detected in malignant and premalignant Barrett's esophagus. The aim of this prospective study was to determine if loss of heterozygosity analysis could identify patients with Barrett's esophagus at greatest risk of adenocarcinoma, for whom endoscopic surveillance is most appropriate. METHODS: Loss of heterozygosity analysis was performed on endoscopic biopsies from 48 patients as part of a Barrett's surveillance program using 14 microsatellite markers shown previously to detect loss of heterozygosity in more than 30% of esophageal adenocarcinomas. Patients were followed up endoscopically for a median of 5 years. RESULTS: Loss of heterozygosity was detected in nine patients. Three patients with loss of heterozygosity on chromosome 5q or 9p did not progress beyond metaplasia. Loss of heterozygosity at 17p11.1-p13 was detected in six patients, all of whom demonstrated dysplasia and/or carcinoma during follow up (four low-grade dysplasia, one high-grade dysplasia and one adenocarcinoma). CONCLUSION: Loss of heterozygosity at 17p11.1-p13 on chromosome 17p identifies patients with Barrett's esophagus at risk of neoplastic progression and can supplement histology in determining the frequency of endoscopy during surveillance.     

Loss of distinct regions on the short arm of chromosome 17 associated with tumorigenesis of human astrocytomas.

Astrocytomas, including glioblastoma multiforme, represent the most frequent and deadly primary neoplasms of the human nervous system. Despite a number of previous cytogenetic and oncogene studies primarily focusing on malignant astrocytomas, the primary mechanism of tumor initiation has remained obscure. The loss or inactivation of "tumor suppressor" genes are thought to play a fundamental role in the development of many human cancers. Thus, we have analyzed astrocytomas of various histological malignancy grades with polymorphic DNA markers to search for specific chromosomal deletions potentially pointing to loci containing tumor suppressor genes. Loss of constitutional heterozygosity indicating chromosomal loss or deletions was most frequently seen for markers on the short arm of chromosome 17 in 50% of the informative tumors (5 of 10 informative cases) and, to a lesser extent, for markers on chromosomes 1 and 10. Deletions on chromosome 17p were seen in both low-grade and high-grade malignant astrocytomas, suggesting that this chromosome may contain a tumor suppressor gene associated with the early events in tumorigenesis. The common region of deletions on the short arm of chromosome 17 is, therefore, clearly distinct from the gene causing von Recklinghausen neurofibromatosis (NF1), a tumor syndrome associated with glial tumors that maps to the long arm of chromosome 17. The search for progressively smaller deletions on chromosome 17p in astrocytomas may be the way to clone and characterize this locus, thus leading to insights into normal and abnormal growth and differentiation of glial cells.     

Chromosome 17p deletions and p53 gene mutations associated with the formation of malignant neurofibrosarcomas in von Recklinghausen neurofibromatosis.

von Recklinghausen neurofibromatosis (NF1) is a common hereditary disorder characterized by neural crest-derived tumors, particularly benign neurofibromas whose malignant transformation to neurofibrosarcomas can be fatal. The NF1 gene has been mapped to a small region of chromosome 17q, but neither the nature of the primary defect nor the mechanisms involved in tumor progression are understood. We have tested whether NF1 might be caused by the inactivation of a tumor suppressor gene on 17q, analogous to that on chromosome 22 in NF2, by searching for deletions of chromosome 17 in NF1-derived tumor specimens. Both neurofibrosarcomas from patients with "atypical" NF and 5 of 6 neurofibrosarcomas from NF1 patients displayed loss of alleles for polymorphic DNA markers on chromosome 17. However, the common region of deletion was on 17p and did not include the NF1 region of 17q. Since no loss of markers on chromosome 17 was observed in any of 30 benign tumors from NF1 patients, the 17p deletions seen in neurofibrosarcomas are probably associated with tumor progression and/or malignancy. This region contains a candidate gene for tumor progression, p53, which has recently been implicated in the progression of a broad array of human cancers. In a preliminary search for p53 aberrations by direct sequencing of polymerase chain reaction-amplified DNA from 7 neurofibrosarcomas, 2 tumors that contained point mutations in exon 4 of the p53 gene were found, suggesting a role for this gene in at least some neurofibrosarcomas. Thus the formation of malignant neurofibrosarcomas may result from several independent genetic events including mutation of the NF1 gene, whose mechanism of tumorigenesis remains uncertain, and subsequent loss of a "tumor suppressor" gene on 17p, most likely p53.     

Comparative genomic hybridization analysis of adrenocortical tumors

Comparative genomic hybridization (CGH) is a molecular cytogenetic technique that allows the entire genome of a tumor to be surveyed for gains and losses of DNA copy sequences. A limited number of studies reporting the use of this technique in adult adrenocortical tumors have yielded conflicting results. In this study we performed CGH analysis on 13 malignant, 18 benign, and 1 tumor of indeterminate malignant potential with the aim of identifying genetic loci consistently implicated in the development and progression of adrenocortical tumors. Tissue samples from 32 patients with histologically proven adrenocortical tumors were available for CGH analysis. CGH changes were seen in all cancers, 11 of 18 (61%) adenomas, and the 1 tumor of indeterminate malignant potential. Of the adrenal cancers, the most common gains were seen on chromosomes 5 (46%), 12 (38%), 19 (31%), and 4 (31%). Losses were most frequently seen at 1p (62%), 17p (54%), 22 (38%), 2q (31%), and 11q (31%). Of the benign adenomas, the most common change was gain of 4q (22%). Mann-Whitney analysis showed a highly significant difference between the cancer group (mean changes, 7.6) and the adenoma group (mean changes, 1.1) for the number of observed CGH changes (P < 0.01). Logistic regression analysis showed that the number of CGH changes was highly predictive of tumor type (P < 0.01). This study has identified several chromosomal loci implicated in adrenocortical tumorigenesis. Activation of a protooncogene(s) on chromosome 4 may be an early event, with progression from adenoma to carcinoma involving activation of oncogenes on chromosomes 5 and 12 and inactivation of tumor suppressor genes on chromosome arms 1p and 17p.     

Inactivation of the p16 tumor suppressor gene in adrenocortical tumors.

The mechanisms of adrenocortical tumorigenesis are still unknown. Evidence that the majority of adrenocortical tumors are monoclonal in origin suggests that a progressive accumulation of genetic aberrations, due to activation of protooncogenes and/or inactivation of tumor suppressor genes, leads to abnormal cell proliferation through a multistep process. Inactivation of the p16 tumor suppressor gene (p16INK4A), which encodes the cell cycle protein p16, was investigated in a series of 14 adrenocortical tumors. Using 11 polymorphic microsatellite markers spanning the short arm of chromosome 9, we demonstrated that three of seven adrenocortical carcinomas and one of seven adrenocortical adenomas had loss of heterozygosity (LOH) within chromosome 9p21, the region containing p16NK4A. Immunohistochemistry showed the absence of p16 nuclear staining in all adrenocortical tumors with LOH within 9p21, and positive staining in all remaining tumors without LOH. In conclusion, LOH within 9p21 associated with lack of p16 expression occurs in a considerable proportion of adrenocortical malignant tumors, but is rare in adenomas. Inactivation of p16INK4A may contribute to the deregulation of cell proliferation in this neoplastic disease.     

Loss of heterozygosity on chromosomes 17 and 18 in breast carcinoma: two additional regions identified.

The loss of heterozygosity (LOH) at specific regions of the human genome in tumor DNA is recognized as evidence for a tumor-suppressor gene located within the corresponding region of the homologous chromosome. Restriction fragment length polymorphism analysis of a panel of primary human breast tumor DNAs has led to the identification of two additional regions on chromosomes 17q and 18q that frequently are affected by LOH. Deletions of each of these regions have a significant correlation with clinical parameters that are associated with aggressive breast carcinomas. Previous restriction fragment length polymorphism analysis of this panel of tumors has uncovered several other frequently occurring mutations. LOH on chromosome 18q frequently occurs in tumors with concomitant LOH of loci on chromosomes 17p and 11p. Similarly, tumors having LOH on 17q also have LOH on chromosomes 1p and 3p. This suggests that certain combinations of mutations may collaborate in the development and malignant progression of breast carcinomas.     

Allelic alterations in pancreatic endocrine tumors identified by genome-wide single nucleotide polymorphism analysis

Pancreatic endocrine tumors (PETs) are uncommon and the genetic alterations in these indolent tumors are not well characterized. Chromosomal imbalances are frequent in tumors but PETs have not been studied by high-density single nucleotide polymorphism (SNP) array. We used genome-wide high-density SNP array analysis to detect copy number alterations using matched tumor and non-neoplastic tissue samples from 15 patients with PETs. In our study, whole or partial loss of chromosomes 1, 3, 11, 22 was present in 40, 47, 53, 40% of tumors respectively, and gain of chromosomes 5, 7, 12, 14, 17, and 20 was present in 47, 60, 47, 53, 53, and 47% of tumors respectively. One tumor had loss of heterozygosity of chromosome 3 and another of chromosome 22 without copy number alterations, suggesting uniparental disomy due to non-disjunction and deletion or to chromosomal recombination. Chromosomal aberrations of the autosomal chromosomes were correlated with chromosomal loss or gain of other chromosomes (r>0.5, P<0.5). About 60% of PETs had high allelic imbalances (AI) defined by more than four chromosomal aberrations, and 40% of tumors had low AI. The PETs with high AI were larger: the mean tumor size with high AI was 5.4 +/- 3.1 cm compared with 2.3 +/- 1.3 cm for low AI (P = 0.03). Our study shows that genome-wide allelotyping is a powerful new tool for the analysis of AI in PETs.