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2.
Br J Dermatol ; 182(5): 1148-1157, 2020 05.
Article in English | MEDLINE | ID: mdl-31520533

ABSTRACT

BACKGROUND: Guidelines for follow-up of patients with melanoma are based on limited evidence. OBJECTIVES: To guide skin surveillance, we developed a risk prediction model for subsequent primary melanomas, using demographic, phenotypical, histopathological, sun exposure and genomic risk factors. METHODS: Using Cox regression frailty models, we analysed data for 2613 primary melanomas from 1266 patients recruited to the population-based Genes, Environment and Melanoma study in New South Wales, Australia, with a median of 14 years' follow-up via the cancer registry. Discrimination and calibration were assessed. RESULTS: The median time to diagnosis of a subsequent primary melanoma decreased with each new primary melanoma. The final model included 12 risk factors. Harrell's C-statistic was 0·73 [95% confidence interval (CI) 0·68-0·77], 0·65 (95% CI 0·62-0·68) and 0·65 (95% CI 0·61-0·69) for predicting second, third and fourth primary melanomas, respectively. The risk of a subsequent primary melanoma was 4·75 times higher (95% CI 3·87-5·82) for the highest vs. the lowest quintile of the risk score. The mean absolute risk of a subsequent primary melanoma within 5 years was 8·0 ± SD 4.1% after the first primary melanoma, and 46·8 ± 15·0% after the second, but varied substantially by risk score. CONCLUSIONS: The risk of developing a subsequent primary melanoma varies considerably between individuals and is particularly high for those with two or more primary melanomas. The risk prediction model and its associated nomograms enable estimation of the absolute risk of subsequent primary melanoma, on the basis of on an individual's risk factors, and can be used to tailor surveillance intensity, communicate risk and provide patient education. What's already known about this topic? Current guidelines for the frequency and length of follow-up to detect new primary melanomas in patients with one or more previous primary melanomas are based on limited evidence. People with one or more primary melanomas have, on average, a higher risk of developing another primary invasive melanoma, compared with the general population, but an accurate way of estimating individual risk is needed. What does this study add? We provide a comprehensive risk prediction model for subsequent primary melanomas, using data from 1266 participants with melanoma (2613 primary melanomas), over a median 14 years' follow-up. The model includes 12 risk factors comprising demographic, phenotypical, histopathological and genomic factors, and sun exposure. It enables estimation of the absolute risk of subsequent primary melanomas, and can be used to tailor surveillance intensity, communicate individual risk and provide patient education.


Subject(s)
Melanoma , Skin Neoplasms , Australia , Cohort Studies , Humans , Melanoma/epidemiology , Melanoma/etiology , New South Wales/epidemiology , Risk Factors , Skin Neoplasms/epidemiology
3.
J Neurooncol ; 136(3): 613-622, 2018 Feb.
Article in English | MEDLINE | ID: mdl-29168082

ABSTRACT

Brain tumor patients treated with radiotherapy (RT) often develop cognitive dysfunction, and recent studies suggest that the APOE ε-4 allele may influence cognitive outcome. The ε-4 allele is known to promote beta (ß) amyloid deposition in the cortex, and preliminary evidence suggests that RT may be associated with this process. However, it is unknown whether ß-amyloid accumulation contributes to treatment neurotoxicity. In this pilot study, we assessed neuropsychological functions and ß-amyloid retention using 18F-florbetaben (FBB) PET in a subset of brain tumor patients who participated in our study of APOE polymorphisms and cognitive functions. Twenty glioma patients treated with conformal RT ± chemotherapy participated in the study: 6 were APOE ε-4 carriers and 14 were non-ε-4 carriers. Patients completed a neuropsychological re-evaluation (mean time interval = 5 years, SD = 0.83) and brain MRI and FBB PET scans. Wilcoxon signed-rank test comparisons between prior and current neuropsychological assessments showed a significant decline in attention (Brief Test of Attention, p = 0.018), and a near significant decline in verbal learning (Hopkins Verbal learning Test-Learning, p = 0.07). Comparisons by APOE status showed significant differences over time in attention/working memory (WAIS-III digits forward, p = 0.028 and digits backward, p = 0.032), with a decline among APOE ε-4 carriers. There were no significant differences in any of the FBB PET analyses between APOE ε-4 carriers and non-ε-4 carriers. The findings suggest that glioma patients may experience worsening in attention and executive functions several years after treatment, and that the APOE ε-4 allele may modulate cognitive decline, but independent of increased ß-amyloid deposition.


Subject(s)
Amyloid/metabolism , Apolipoprotein E4/genetics , Brain Neoplasms/diagnostic imaging , Brain Neoplasms/psychology , Glioma/diagnostic imaging , Glioma/psychology , Adult , Aged , Aniline Compounds , Brain/diagnostic imaging , Brain/metabolism , Brain Neoplasms/genetics , Brain Neoplasms/metabolism , Chemoradiotherapy , Cognition , Cognition Disorders/diagnostic imaging , Cognition Disorders/etiology , Cognition Disorders/genetics , Cognition Disorders/metabolism , Cohort Studies , Female , Glioma/genetics , Glioma/metabolism , Heterozygote , Humans , Magnetic Resonance Imaging , Male , Middle Aged , Neuropsychological Tests , Pilot Projects , Positron-Emission Tomography , Radiopharmaceuticals , Radiotherapy, Conformal , Stilbenes
5.
Br J Dermatol ; 172(4): 1081-9, 2015 Apr.
Article in English | MEDLINE | ID: mdl-25307738

ABSTRACT

BACKGROUND: Melanocytic naevi are an important risk factor for melanoma. Naevi with distinct dermoscopic patterns can differ in size, distribution and host pigmentation characteristics. OBJECTIVES: We examined MC1R and 85 other candidate loci in a cohort of children to test the hypothesis that the development and dermoscopic type of naevi are modulated by genetic variants. METHODS: Buccal DNAs were obtained from a cohort of 353 fifth graders (mean age 10·4 years). Polymorphisms were chosen based on a known or anticipated role in naevi and melanoma. Associations between single-nucleotide polymorphisms (SNPs) and baseline naevus count were determined by multivariate regression adjusting for sex, race/ethnicity and sun sensitivity. Dermoscopic images were available for 853 naevi from 290 children. Associations between SNPs and dermoscopic patterns were determined by polytomous regression. RESULTS: Four SNPs were significantly associated with increasing (IRF4) or decreasing (PARP1, CDK6 and PLA2G6) naevus count in multivariate shrinkage analyses with all SNPs included in the model; IRF4 rs12203952 showed the strongest association with log naevus count (relative risk 1·56, P < 0·001). Using homogeneous naevi as the reference, IRF4 rs12203952 and four other SNPs in TERT, CDKN1B, MTAP and PARP1 were associated with either globular or reticular dermoscopic patterns (P < 0·05). CONCLUSIONS: Our results provide evidence that subsets of naevi defined by dermoscopic patterns differ in their associations with germline genotypes and support the hypothesis that dermoscopically defined subsets of naevi are biologically distinct. These results require confirmation in larger cohorts. If confirmed, these findings will improve the current knowledge of naevogenesis and assist in the identification of individuals with high-risk phenotypes.


Subject(s)
Nevus, Pigmented/genetics , Polymorphism, Single Nucleotide/genetics , Skin Neoplasms/genetics , Alleles , Child , Cyclin-Dependent Kinase 6/genetics , Dermoscopy/methods , Female , Genetic Loci , Genotype , Group VI Phospholipases A2/genetics , Humans , Interferon Regulatory Factors/genetics , Male , Nevus, Pigmented/pathology , Poly (ADP-Ribose) Polymerase-1 , Poly(ADP-ribose) Polymerases/genetics , Prospective Studies , Receptor, Melanocortin, Type 1/genetics , Skin Neoplasms/pathology , Sunlight/adverse effects
6.
Eur J Cancer Prev ; 13(1): 65-70, 2004 Feb.
Article in English | MEDLINE | ID: mdl-15075790

ABSTRACT

Estimation of the relative risk of cancer due to rare germline mutations using population-based epidemiological techniques is challenging, since studies with very large numbers of subjects are required. In this pilot study using a novel study design, we evaluated the role of INK4A mutations in melanoma by comparing patients with multiple primary melanomas to those with single primaries. Patients were ascertained from the Surgery and Dermatology Clinics at Memorial Sloan-Kettering Cancer Center and at the Yale University Pigmented Lesion Clinic. Subjects completed a questionnaire covering risk factors for melanoma and were tested for INK4A mutations. Five (8%) of 65 patients with multiple primaries had a mutation, compared with none of 88 patients with single primaries (P=0.03). Examination of other factors, such as number of nevi on the arms of the patients, fair skin, hair and eye colour, and other phenotypic characteristics associated with the risk of melanoma, demonstrates that these factors exhibit higher prevalence in the multiple primary cases than in the single primaries. These results provide evidence of the utility of the new study design in evaluating the impact of rare but highly penetrant cancer risk factors.


Subject(s)
Cyclin-Dependent Kinase Inhibitor p16/genetics , Melanoma/genetics , Neoplasms, Multiple Primary/genetics , Skin Neoplasms/genetics , Adolescent , Adult , Case-Control Studies , Chromatography, Liquid , Female , Germ-Line Mutation/genetics , Humans , Male , Melanoma/epidemiology , Middle Aged , Neoplasms, Multiple Primary/epidemiology , Nevus/genetics , Nucleic Acid Denaturation/genetics , Odds Ratio , Pilot Projects , Polymerase Chain Reaction , Prevalence , Risk Factors , Skin Neoplasms/epidemiology , Surveys and Questionnaires
8.
Environ Mol Mutagen ; 38(2-3): 223-6, 2001.
Article in English | MEDLINE | ID: mdl-11746758

ABSTRACT

The etiology of soft tissue sarcoma is poorly understood. Exposure to environmental chemicals may play a role, but the data are not clear. We compared a group of soft tissue sarcoma patients with healthy controls to determine whether the mutagen sensitivity assay, a simple chromosome aberration assay using the radiomimetic bleomycin, might be useful to identify patients at risk for soft tissue sarcoma. Patients with a diagnosis of soft tissue sarcoma at Memorial Sloan-Kettering's outpatient clinic signed informed consent and donated 30 ml of blood. Controls were selected from the general population of Connecticut by random digit dialing. Unrepaired DNA damage was assessed for 100 metaphase spreads for each individual, with the number of breaks in chromatids being counted as breaks per cell (b/c). The 20 cases with soft tissue sarcoma had 1.03 mean b/c and the controls had 0.88 b/c (P = 0.16). Patients with soft tissue sarcoma were 5.7 times more likely to be mutagen sensitive than controls (P = 0.01), as determined after dividing subjects into sensitive or not sensitive groups based on the median b/c among controls. As mutagen sensitivity has been shown to be associated with a number of cancers and appears to reflect genetic susceptibility, this assay may be an appropriate biomarker for radiation sensitivity or it may be a marker of susceptibility to soft tissue sarcoma. Larger studies should be undertaken to assess these possibilities.


Subject(s)
Bleomycin/toxicity , DNA Damage/drug effects , Mutagens/toxicity , Sarcoma/genetics , Adult , Aged , DNA Damage/genetics , Female , Genetic Predisposition to Disease , Humans , Male , Middle Aged , Mutagenicity Tests , Radiation Tolerance , Risk , Sarcoma/chemically induced
9.
J Mol Diagn ; 3(4): 158-63, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11687599

ABSTRACT

The incidence of melanoma is increasing rapidly in western countries. Genetic predisposition in familial and in some sporadic melanomas has been associated with the presence of INK4A gene mutations. To better define the risk for developing sporadic melanoma based on genetic and environmental interactions, large groups of cases need to be studied. Mutational analysis of genes lacking hot spots for sequence variations is time consuming and expensive. In this study we present the application of denaturing high performance liquid chromatography (DHPLC) for screening of mutations. Exons 1alpha, 2, and 3 were amplified from 129 samples and 13 known mutants, yielding 347 products that were examined at different temperatures. Forty-two of these amplicons showed a distinct non-wild-type profile on the chromatogram. Independent sequencing analysis confirmed 16 different nucleotide variations in Leu32Pro; Ile49Thr; 88 del G; Gln50Arg; Arg24Pro; Met53Ile; Met53Thr; Arg58stop; Pro81Leu; Asp84Ala; Arg80stop; Gly101Trp; Val106Val; Ala148Thr; and in positions (-2) in intron 1 (C --> T); and in the 3' UTR, nucleotide 500 (C --> G). No false negatives or false positives were obtained by DHPLC in samples with mutations or polymorphisms. We conclude that the DHPLC is a fast, sensitive, cost-efficient, and reliable method for the scanning of INK4A somatic or germline mutations and polymorphisms of large number of samples.


Subject(s)
Chromatography, High Pressure Liquid/methods , Cyclin-Dependent Kinase Inhibitor p16/genetics , Epithelial Cells/cytology , Nucleic Acid Denaturation/genetics , Sequence Analysis, DNA/methods , Buffers , Exons , Humans , Melanoma/blood , Mouth Mucosa , Mutation , Temperature , Time Factors
10.
Hum Pathol ; 32(5): 537-44, 2001 May.
Article in English | MEDLINE | ID: mdl-11381373

ABSTRACT

We undertook the present study to examine alterations affecting the RB pathway in the G1 checkpoint and to determine their potential clinical significance in children affected with nonfamilial retinoblastoma. Using immunohistochemistry, patterns of expression of pRB, p16/INK4A, and E2F1 were analyzed in tissue from a cohort of 86 well-characterized patients with nonfamilial retinoblastoma diagnosed at the "Instituto Nacional de Pediatria" in Mexico City. The relationship of these phenotypes to proliferative index was assessed by analysis of Ki67 antigen expression. pRB expression was found in 11 (13%) cases. Using a hypophosphorylated specific pRB antibody, we observed low levels of underphosphorylated pRB expression in only 1 of 9 evaluable positive cases. These data suggest that the detected pRB products were hyperphosphorylated and thus had decreased functional activity. Increased p16 nuclear expression was found in only 6 tumors. No tumors showed deletions or mobility shifts of the INK4A gene. Undetectable pRB levels were significantly associated with undetectable p16 expression (odds ratio, 10.8; 95% confidence interval, 1.4-81.3; P =.03). All tumors showed nuclear immunoreactivities for E2F1 and Ki67. Increased Ki67 proliferative index was associated with increased staining for E2F1 (r =.44; P =.008) and increasing clinical stage (P =.03). Among children with unilateral disease, the mean Ki67 proliferative index was significantly higher in children with advanced clinical disease (stages 3 and 4) (mean 81.25; SD 6.78) than in those with earlier stage disease (mean 69.50; SD 9.45) (P = 0.001). Among children with bilateral disease, however, the mean proliferative index was not significantly higher for children with advanced clinical stage. When examining all cases together, there was a significant trend toward increasing proliferative index with increasing clinical stage (P =.03). In unilateral tumors, we also found that presence of detectable pRB was associated with a lower percentage of cells expressing E2F1 (46.7% v 70.8%) (P = 0.05), whereas there was no association between presence of pRB and E2F1 among bilateral tumors. We have found that expression of some of the cell cycle markers examined varies according to laterality, suggesting underlying differences in the capacity for cell cycle regulation between these 2 forms of the disease. Differences in capacities for cell cycle regulation may account for some differences in clinical behavior. Thus, the inclusion of molecular markers may become useful adjuncts to clinicopathological staging and subsequent determination of therapy.


Subject(s)
Carrier Proteins , Cell Cycle Proteins/analysis , DNA-Binding Proteins , Retinal Neoplasms/chemistry , Retinoblastoma/chemistry , Age Factors , Cell Division , Cell Nucleus/chemistry , Child , Cyclin-Dependent Kinase Inhibitor p16/analysis , Cyclin-Dependent Kinase Inhibitor p16/genetics , DNA Mutational Analysis , E2F Transcription Factors , E2F1 Transcription Factor , Female , Gene Deletion , Humans , Ki-67 Antigen/analysis , Male , Neoplasm Staging , Optic Nerve/pathology , Phenotype , Phosphorylation , Polymorphism, Single-Stranded Conformational , Retinal Neoplasms/mortality , Retinal Neoplasms/pathology , Retinoblastoma/mortality , Retinoblastoma/pathology , Retinoblastoma Protein/analysis , Retinoblastoma Protein/metabolism , Retinoblastoma-Binding Protein 1 , Reverse Transcriptase Polymerase Chain Reaction , Transcription Factor DP1 , Transcription Factors/analysis
11.
Int J Cancer ; 95(4): 223-7, 2001 Jul 20.
Article in English | MEDLINE | ID: mdl-11400114

ABSTRACT

During the metaphase-anaphase transition, the spindle checkpoint prevents segregation of chromosomes if the spindle assembly is perturbed. Critical components of this checkpoint are the MAD and BUB families of proteins, which prevent the proteolysis of Pds1 and B cyclins, producing mitotic arrest. In the present study, we first intended to resolve the role of the hsMAD2 gene in human cancer by determining the potential presence of hsMAD2 mutations in 44 primary bladder tumors, 42 soft-tissue sarcomas and 10 hepatocellular carcinomas. The entire coding region of the hsMAD2 gene was analyzed using PCR-SSCP and sequencing. One of the bladder tumor samples showed a point mutation consisting of a transition, ATC-->GTC (Ile-->Val) in codon 190 of hsMAD2. However, no differences were found in the mitotic arrest between cells transfected with mutant and wild-type MAD2 cDNA. We also identified mobility shifts in hsMAD2 in both normal and tumor DNA in 3 bladder tumors, 3 soft-tissue sarcomas and 1 hepatocellular carcinoma, consistent with a polymorphism at codon 143, CCA-->CCG (Pro-->Pro). Another polymorphism was identified in a hepatocellular carcinoma case at codon 22, GAG-->GAA (Glu-->Glu). In addition, a subgroup of 67 primary tumors was analyzed by Southern blot hybridization. No deletion or visible re-arrangements were detected by comparing tumor and normal DNA band signals. Two other important components of the spindle mitotic checkpoint, hBUB1 and hBUB3, were also screened for mutations: hBUB1 in 43 bladder tumors and 9 bladder cell lines and hBUB3 only in the cell lines. Two polymorphisms were found in hBUB1 at positions 144, CAG-->CAA (Gln-->Gln) in 1 primary tumor and 1 bladder cell line, and 913 (ATC-->ATT, Ile-->Ile) in 1 primary tumor. We did not find sequence alterations in hBUB3. These results suggest that mutations of the hsMAD2, hBUB1 and hBUB3 genes are very rare in bladder tumors and that hsMAD2 alterations are also infrequent in soft-tissue sarcomas and hepatocellular carcinomas.


Subject(s)
Calcium-Binding Proteins/genetics , Cell Cycle Proteins/genetics , DNA, Neoplasm/genetics , Mutation , Protein Kinases/genetics , Proteins/genetics , Calcium-Binding Proteins/metabolism , Carcinoma, Hepatocellular/genetics , Cell Cycle Proteins/metabolism , DNA Mutational Analysis , Gene Deletion , Humans , Liver Neoplasms/genetics , Mad2 Proteins , Poly-ADP-Ribose Binding Proteins , Polymorphism, Genetic , Protein Kinases/metabolism , Protein Serine-Threonine Kinases , Proteins/metabolism , Repressor Proteins , Sarcoma/genetics , Soft Tissue Neoplasms/genetics , Tumor Cells, Cultured , Urinary Bladder Neoplasms/genetics
12.
Cancer Genet Cytogenet ; 125(2): 131-8, 2001 Mar.
Article in English | MEDLINE | ID: mdl-11369056

ABSTRACT

The INK4A and INK4B loci are located at 9p21 and have been implicated in the tumorigenesis of various human malignancies. The INK4A gene encodes two cell cycle regulators, p16(INK4A) and ARF, while INK4B encodes p15(INK4B). Previously, we have shown that the p16(INK4) tumor suppressor was not mutated or deleted in primary breast carcinomas. However, primary and metastatic breast carcinomas exhibited a relative hypomethylation of p16(INK4A), which is associated with expression, compared to normal breast tissue. The present study was conducted to determine if inactivation of p15(INK4B) and INK4A exon 1beta (ARF) are common events in breast carcinoma. Mutational analysis was performed by PCR-SSCP, and mRNA expression was evaluated by RT-PCR. Methylation-specific PCR was used to determine the methylation status of the p15(INK4B) promoter. Our results demonstrate that the p15(INK4B) gene was altered in 3 (21%) of the 14 breast cell lines; one had a silent mutation and two had homozygous deletion of the gene. None of the cell lines showed methylation of p15(INK4B). Two (14%) cell lines had homozygous deletion of INK4A exon 1beta. All normal and malignant breast tissue samples were wild-type and non-methylated for p15(INK4B) and wild-type for exon 1beta. Our results show that these structurally and functionally related genes are not invariably affected together, and the most frequently observed alteration at the INK4A and INK4B loci in breast carcinoma appears to be p16(INK4A) hypomethylation.


Subject(s)
Breast Neoplasms/genetics , Carcinoma/genetics , Carrier Proteins/genetics , Cell Cycle Proteins , Chromosomes, Human, Pair 9/genetics , DNA Methylation , Gene Expression Regulation, Neoplastic , Genes, Tumor Suppressor , Genes, p16 , Tumor Suppressor Proteins , Blotting, Southern , Breast/metabolism , Breast Neoplasms/pathology , Carcinoma/pathology , Carrier Proteins/biosynthesis , CpG Islands , Cyclin-Dependent Kinase Inhibitor p15 , Cyclin-Dependent Kinase Inhibitor p16/biosynthesis , DNA Mutational Analysis , DNA, Neoplasm/genetics , Exons/genetics , Female , Gene Expression Profiling , Gene Silencing , Humans , Neoplasm Proteins/biosynthesis , Neoplasm Proteins/genetics , Polymorphism, Single-Stranded Conformational , RNA, Messenger/biosynthesis , RNA, Neoplasm/biosynthesis , Reverse Transcriptase Polymerase Chain Reaction , Sequence Deletion , Tumor Cells, Cultured , Viral Proteins/biosynthesis , Viral Proteins/genetics
13.
Clin Cancer Res ; 6(10): 4010-6, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11051250

ABSTRACT

Epidemiological studies have shown that the use of barrier methods of contraception is associated with a decreased incidence of papilloma virus infection and reduced risk of having a child with retinoblastoma. Thirty-nine primary retinoblastomas were analyzed for the presence of papilloma virus sequences. Tumor tissue sections were also used to assess the expression of the retinoblastoma protein and proliferative index. Papilloma sequences were detected in 14 of 39 (36%) tumors. Tumors in which viral sequences were detected were associated with a lower proliferative index (68% versus 78%; P = 0.015). Children with tumors containing viral sequences had a lower risk of extraocular disease (odds ratio, 9.0; 95% confidence interval, 1.6-49; P = 0.008) and a lower birth weight (2.9 versus 3.5 kg; P = 0.030). Based on these data, it is our hypothesis that papilloma viruses may play a role in the development of sporadic retinoblastoma. Detection of papilloma virus sequences and retinoblastoma protein in certain primary lesions suggests an alternative mechanism of tumor development for sporadic retinoblastoma.


Subject(s)
Carrier Proteins , Cell Cycle Proteins , DNA-Binding Proteins , Papillomaviridae/metabolism , Retinoblastoma/etiology , Retinoblastoma/virology , Age of Onset , Blotting, Southern , Cell Division , Child, Preschool , E2F Transcription Factors , HeLa Cells , Humans , Immunohistochemistry , Infant , Polymerase Chain Reaction , Retinoblastoma/pathology , Retinoblastoma Protein/biosynthesis , Retinoblastoma-Binding Protein 1 , Transcription Factor DP1 , Transcription Factors/biosynthesis
14.
Oncol Rep ; 7(2): 421-5, 2000.
Article in English | MEDLINE | ID: mdl-10671696

ABSTRACT

Mutations in proto-oncogenes and tumor suppressor genes have been associated with tumor development and/or progression in many neoplasms. It has been reported that parathyroid tumors have deletions affecting the retinoblastoma gene (RB), and overexpression of cyclin D1 (Cyc D1). The aim of the present study was to evaluate the alterations in the components of the pRB pathway in parathyroid adenomas and parathyroid aggressive tumors, including patterns of expression of pRB, Cyc D1, and p16/INK4A. Paired normal and tumor DNA from 6 parathyroid adenomas and 5 aggressive tumors were analyzed for loss of heterozygosity (LOH) at the RB locus. The expression of pRB, Cyc D1 and p16 was studied in 4 adenomas and 5 aggressive tumors. RB LOH was found in 1 of 6 adenomas, and in 1 of 2 informative aggressive tumors. Immunohistochemical analysis revealed undetectable pRB in 4 of 5 aggressive tumors and presence of pRB in all adenomas. Conversely, Cyc D1 expression was found in 3 of 4 aggressive tumors, but was undetectable in the adenomas. Expression of p16 was identified only in one aggressive tumor. Thus, alterations in the pRB pathway seem to prevail in the aggressive form of parathyroid neoplasms. Our results warrant further investigation of these cell cycle regulators in order to determine their potential role as tumor markers in parathyroid tumors.


Subject(s)
Biomarkers, Tumor , Cell Cycle/genetics , Cyclin D1/biosynthesis , Gene Deletion , Genes, Retinoblastoma , Parathyroid Neoplasms , Adult , Aged , Cyclin-Dependent Kinase Inhibitor p16/metabolism , Female , Humans , Male , Middle Aged , Parathyroid Neoplasms/genetics , Parathyroid Neoplasms/metabolism , Parathyroid Neoplasms/pathology , Retinoblastoma Protein/metabolism , Signal Transduction/genetics
15.
Am J Pathol ; 155(6): 1855-60, 1999 Dec.
Article in English | MEDLINE | ID: mdl-10595915

ABSTRACT

The INK4A gene, a candidate tumor suppressor gene located on chromosome 9p21, encodes two protein products, p16 and p19(ARF). p16 is a negative cell cycle regulator capable of arresting cells in the G1 phase by inhibiting cyclin-dependent kinases 4 (Cdk4) and 6 (Cdk6), thus preventing pRB phosphorylation. p19(ARF) prevents Mdm2-mediated neutralization of p53. Loss of INK4A is a frequent molecular alteration involved in the genesis of several neoplasms, including tumors of neuroectodermal origin. This study investigated the frequency of INK4A gene alterations in a series of malignant peripheral nerve sheath tumors (MPNSTs) and neurofibromas (NFs). INK4A gene and the p19(ARF)-specific exon 1beta were studied in 11 MPNST samples from 8 patients and 7 neurofibromas. Presence of INK4A deletions was assessed by Southern blotting hybridization and by a multiplex polymerase chain reaction (mPCR). INK4A point mutations were examined by single-strand conformation polymorphism (SSCP) and sequencing. The p16 promoter methylation status was determined by PCR amplification of bisulfite-treated DNA. Homozygous deletions of exon 2, thus affecting both p16 and p19(ARF), were identified in MPNSTs from 4 of 8 patients. Deletions, mutations, or silencing by methylation were not identified in the neurofibromas analyzed. Based on our results, we conclude that INK4A deletions are frequent events in MPNSTs and may participate in tumor progression. Silencing of p16 by methylation, which occurs often in several tumor types, is uncommon in MPNSTs.


Subject(s)
Cyclin-Dependent Kinases/antagonists & inhibitors , Gene Deletion , Genes, Tumor Suppressor , Genes, p16 , Nerve Sheath Neoplasms/genetics , Neurofibroma/genetics , Proteins/genetics , Blotting, Southern , Cell Cycle , DNA Methylation , DNA, Neoplasm/analysis , Exons , Humans , Nerve Sheath Neoplasms/pathology , Neurofibroma/pathology , Point Mutation , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Tumor Suppressor Protein p14ARF
16.
Am J Pathol ; 155(1): 105-13, 1999 Jul.
Article in English | MEDLINE | ID: mdl-10393843

ABSTRACT

The INK4A and the INK4B genes map to chromosome 9p21, an area frequently deleted in bladder neoplasms. In addition to the p16 protein, the INK4A encodes for a second product, termed p19(ARF). We analyzed tissues from 121 patients with initial Ta and T1 tumors. Deletions of the INK4A gene were observed in 17 of 121 (14.1%) cases. Point mutations were identified in 2 of 64 (3.1%) tumors. The INK4A-exon 1beta and the INK4B gene were codeleted with INK4A in all of the homozygously deleted cases analyzed. The p16 promoter underwent de novo methylation in 7 of 47 (14.9%) evaluable cases. The p16-positive phenotype was observed in 18 of 56 (32%) evaluable cases. p16 negative phenotype correlated with deletion and methylation status. A statistically significant association between INK4A homozygous deletions and tumor size was observed (P = 0.003). Patients bearing tumors with INK4A homozygous deletions had a lower recurrence-free survival (P = 0.040) than those with wild type INK4A. In conclusion, deletions and methylation of the INK4A gene occur frequently in superficial bladder tumors. However, only those deletions that affect both the p16 and the p19(ARF), deregulating both the pRb and p53 pathways, correlated with clinicopathological parameters of worse prognosis.


Subject(s)
Gene Deletion , Genes, p16/genetics , Neoplasm Recurrence, Local/genetics , Urinary Bladder Neoplasms/genetics , Exons/genetics , Gene Frequency , Homozygote , Humans , Methylation , Mutation/genetics , Phenotype , Prognosis , Promoter Regions, Genetic/physiology , Proteins/genetics , Survival Analysis , Tumor Suppressor Protein p14ARF , Urinary Bladder Neoplasms/pathology
17.
Int J Oncol ; 15(1): 17-24, 1999 Jul.
Article in English | MEDLINE | ID: mdl-10375589

ABSTRACT

The INK4A and INK4B genes map to 9p21, with the INK4A gene encoding two products, p16 and p19ARF. Many neoplasms in which INK4A and INK4B genes are altered show deletions involving both genes. Mice carrying a targeted Ink4a deletion develop tumors at an early age. In the present study we examined the genetic alterations affecting the remaining Ink4a allele and the Ink4b gene in tumors arising in heterozygous Ink4a mice. We identified deletion of the remaining Ink4a allele in 7 of 18 (39%) tumors. We also observed deletion of the exon 1beta in 3 cases, one of them presenting this deletion as a unique alteration. In conclusion, the deletion of the remaining Ink4a allele was the alteration most frequently observed, representing the inactivation of two proteins capable of arresting the cell cycle through different pathways that involve the tumor suppressors pRB and p53.


Subject(s)
Cyclin-Dependent Kinase Inhibitor p16/physiology , Genes, p16 , Neoplasms, Experimental/genetics , Proteins/physiology , Animals , Cell Cycle/genetics , Cyclin-Dependent Kinase Inhibitor p16/deficiency , Cyclin-Dependent Kinase Inhibitor p16/genetics , DNA, Neoplasm/genetics , Gene Deletion , Gene Targeting , Genetic Predisposition to Disease , Heterozygote , Mice , Mice, Knockout , Neoplasms, Experimental/chemically induced , Neoplasms, Experimental/pathology , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Proteins/genetics , Retinoblastoma Protein/physiology , Tumor Suppressor Protein p14ARF
18.
J Natl Cancer Inst ; 91(10): 874-81, 1999 May 19.
Article in English | MEDLINE | ID: mdl-10340908

ABSTRACT

BACKGROUND: We sought to identify and characterize potential alterations in E2F-1, a transcription factor that binds to the retinoblastoma protein (pRB), in bladder neoplasms and to elucidate a possible role for E2F-1 as an oncogene or a tumor suppressor gene. METHODS: Tumor samples from 133 evaluable patients with bladder cancer were analyzed for E2F-1 gene mutations by use of polymerase chain reaction-single-strand conformational polymorphism (PCR-SSCP) analysis and DNA sequencing. In addition, tumors were studied for E2F-1 and pRB protein expression by use of immunohistochemistry. Results from the above analyses were correlated with clinicopathologic parameters and outcome. All P values are two-sided. RESULTS: A polymorphism, consisting of a nucleotide change at amino acid codon 393 in exon 7 (GGC-->AGC [Gly-->Ser]), was identified in seven of 133 case patients, being present in both tumor and corresponding normal tissues. No bandshifts were identified in the nuclear-localization or DNA-binding domains on PCR-SSCP analysis. On immunohistochemical analysis, E2F-1 nuclear reactivity was observed in less than 5% of the cells from 53 tumors and in 5%-75% of the cells from the remaining 80 tumors. The pattern of E2F-1 protein expression was not altered in relation to the identified polymorphism. pRB nuclear reactivity greater than 20% (of tumor cells stained) was present in 66% of the samples. E2F-1 nuclear reactivity correlated inversely with the percentage of cells showing pRB reactivity (Kendall tau(b) = -0.18; P = .019). On multivariate analysis, patients with lower E2F-1 reactivity had statistically significantly increased risks of progression to metastases (P = .001) and death (P = .02). CONCLUSIONS: E2F-1 alterations occur at the phenotypic level, rather than at the genotypic level, in bladder cancer. The adverse outcome for patients whose tumors exhibit low E2F-1 nuclear expression suggests a possible tumor suppressor role for E2F-1 in bladder cancer.


Subject(s)
Carcinoma/genetics , Carcinoma/pathology , Carrier Proteins , Cell Cycle Proteins/genetics , DNA-Binding Proteins/genetics , Mutation , Transcription Factors/genetics , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/pathology , E2F Transcription Factors , E2F1 Transcription Factor , Genotype , Glycine/genetics , Humans , Immunohistochemistry , Lymphatic Metastasis , Multivariate Analysis , Neoplasm Invasiveness , Neoplasm Staging , Phenotype , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Prognosis , Retinoblastoma-Binding Protein 1 , Serine/genetics , Transcription Factor DP1
19.
J Natl Cancer Inst ; 91(1): 73-9, 1999 Jan 06.
Article in English | MEDLINE | ID: mdl-9890173

ABSTRACT

BACKGROUND: The INK4A and INK4B genes map to chromosome 9p21, with the INK4A gene encoding two protein products, p16 and pl9ARF. Alterations of the INK4A and INK4B genes occur frequently in certain primary malignant neoplasms. This study was undertaken to evaluate the frequency of INK4A and INK4B gene alterations in a cohort of adult soft tissue sarcomas. METHODS: The status of the INK4A and INK4B genes was determined in 46 soft tissue sarcomas by use of the following methods: Southern blotting, polymerase chain reaction (PCR), single-strand conformation polymorphism analysis, comparative multiplex PCR, and a methylation assay focusing on the p16 promoter. Associations between alterations of the INK4A and INK4B genes and clinicopathologic variables, as well as with p53 and pRB (retinoblastoma protein) status, were evaluated by use of the two-tailed Fisher's exact test. Disease-specific survival was evaluated by use of the Kaplan-Meier method and the logrank test. Proportional hazards analysis was used to obtain estimates of relative risks. All P values are two-sided. RESULTS: Homozygous and hemizygous deletions, but no point mutations, were observed in these two genes. The overall frequency of gene alteration (deletion or rearrangement) was approximately 15% for the INK4A and INK4B genes, with changes restricted to high-grade sarcomas. Statistically significant associations were observed between INK4A/INK4B deletions (P = .036) or alterations (P = .005) and poor survival. Alteration of the INK4A and INK4B genes was the only statistically significant predictor for poor survival when controlling for tumor grade and size (P = .03). CONCLUSION/IMPLICATIONS: Coincident homozygous deletion of the INK4A and INK4B genes occurs frequently in adult soft tissue sarcomas. Loss of p16 and pl9ARF function in primary tumors, although not equivalent to alterations in p53 and pRB function, appears to be associated with cancers that have an aggressive biologic behavior.


Subject(s)
Carrier Proteins/genetics , Cell Cycle Proteins , Chromosomes, Human, Pair 9/genetics , Cyclin-Dependent Kinase Inhibitor p16 , DNA, Neoplasm/genetics , Genes, p16 , Neoplasm Proteins/genetics , Sarcoma/genetics , Soft Tissue Neoplasms/genetics , Tumor Suppressor Proteins , Adult , Animals , Antibodies, Monoclonal/immunology , Cohort Studies , Cyclin-Dependent Kinase Inhibitor p15 , DNA Methylation , Genes, Retinoblastoma , Genes, p53 , Humans , Life Tables , Loss of Heterozygosity , Mice , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Prognosis , Sarcoma/mortality , Soft Tissue Neoplasms/mortality
20.
Int J Oncol ; 12(3): 583-8, 1998 Mar.
Article in English | MEDLINE | ID: mdl-9472096

ABSTRACT

p16 and p15 are representative members of cyclin-dependent kinase inhibitors. Because the selective expression of p57KIP2 in liver, and because p16CDKN2/MTS1/INK4A has been found altered in many primary tumors, we undertook the present study to determine the presence of alterations in these genes in a group of hepatocellular carcinomas (HCC). Seventeen tumor and normal DNA pairs were analyzed by Southern blot, PCR-SSCP and DNA sequencing. Microsatellite markers surrounding the area of the p16 gene was also used. Southern blot analysis did not show allelic losses of the p16 or p57KIP2 genes. In 4 cases, an extra band was observed when hybridizing with the specific p16 cDNA. Overall, 4/17 (24%) cases presented microsatellite alterations at the 9p21-24 region. These results suggest that deletions or point mutations in these genes are not frequent if present at all in HCC, but reveals the existence of microsatellite alterations at the 9p21-24 region in HCC.


Subject(s)
Carcinoma, Hepatocellular/genetics , Cyclin-Dependent Kinase Inhibitor p16/genetics , Enzyme Inhibitors , Liver Neoplasms/genetics , Nuclear Proteins/genetics , Point Mutation , Amino Acid Substitution , Base Sequence , Blotting, Southern , Cyclin-Dependent Kinase Inhibitor p57 , Humans , Liver/metabolism , Microsatellite Repeats , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Reference Values
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