P53 Gene Alterations Research Articles

HTLV-I Tax Related Dysfunction of Cell Cycle Regulators and Oncogenesis of Adult T Cell Leukemia

HTLV-I is causually related to the oncogenesis of adult T cell leukemia (ATL). However, the precise mechanism of HTLV-I oncogenesis is unclear. HTLV-I Tax protein functions as an activator of various cellular genes, including IL-2, IL-2 receptor-α, and c-fos through the activation of nuclear transfer factors such as NF-kB and SRF, and also potently activates trascription of viral genes through CREB/ATF sites in the viral LTR. However, Tax activation of HTLV-I infected T cells through the above pathways induces polyclonal proliferation of the cells in vitro; Tax however may function only transiently in the immediate post-infection period following infection in vivo. The long latent period of 60 years from infection to onset of disease suggests other mechanisms for ATL oncogenesis. Recent studies suggest that the malignant transformation of ATL is a multi-hit phenomena, suggesting that discrete genetic events are responsible for ATL oncogenesis. These genetic events could be responsible for the different stages of ATL: smoldering, chronic, lymphoma, and acute type. p16 and p53 genes are important negative regulators of the cell cycle and are often found to be mutated in neoplasms. Recent studies including ours demonstrated a high frequency of alteration of these two genes in primary ATL cells. Furthermore, alteration of the two genes is associated with acute but not chronic type ATL. In addition, p16 gene alteration is linked to the growth rate of ATL cells, suggesting that the alteration of these cell cycle regulatory genes may be related to progression from smoldering or chronic to acute or lymphoma type ATL. Tax may be involved in mutagenesis of these genes through suppression of DNA-β polymerase gene expression during the process from latent period to acute/lymphoma type. Once transformation occurs, activation of the pathway between Tax and the three nuclear transfer factors, NF-kB, SRF, and CREB/ATF, contributes to establish the aggressive manifestations of acute/lymphoma type ATL cells.

Genetic marker analysis in head and neck cancer

Prognostication of head and neck cancer (HNCC) involves molecular identification of residual tumor cells, prediction of recurrence, distant metastases or secondary tumors and prediction of the sensitvity to therapy. Biomarkers of HNCC are mutations of p53, p16 and amplification of Cyclin D and E2F4. One hundred and fifty-two HNCC cases have been evaluated for p53, hMLH1, Cyclin D and p16 gene alterations using PCR-SSCP and Western blot analysis. P53 mutations of HNCC have been found in 37.5% of cases. However, 11% of the cases showed p53 mutations in the normal peritumoral mucosa suggesting "field cancerization" process. Mismatch-repair gene mutations (MMR: hMHL1 and hMSH2) occurred with 17 and 8.6% frequency, respectively, while E2F4 mutations were even more frequent (21.4%) in HNCC. Our data suggest that E2F4 overexpression can be caused by the inactivation of the p16 gene in HNCC, while its mutations are most probably associated to the mutations of the MMR genes. These molecular informations can help to predict the biological potential of HNCC as well as the probability of the development of secondary HNCCs.

Expression and Mutation of p53 in Tumor Effusion Cells of Patients with Ovarian Carcinoma: Response to Cisplatin-Based Chemotherapy

p53 alterations are considered as one of the most important factors responsible for drug resistance in ovarian carcinomas, although the relationship between p53 gene status and response to cisplatin-based chemotherapy in ovarian cancer patients remains unclear. The aim of the study was to evaluate the relationship between p53 protein accumulation, p53 gene mutation and response to cisplatin-based chemotherapy in patients with ovarian carcinoma considering conventional clinicopathological parameters. Tissue sections and corresponding cyst and/or ascitic fluid cells from 79 patients with epithelial ovarian cancer were analyzed immunohistochemically for p53 expression. The PCR-SSCP analysis was performed in 25 cases and the results were compared with immunohistochemical data. It was demonstrated that p53 expression reaching approximately 50% of positive cells in immunostaining was usually associated with PCR-amplified exons showing abnormal migration and suspected for mutation. p53 gene changes were not correlated with histological structure, grade of differentiation or residual tumor after cytoreductive surgery, despite being detected more frequently in III/IV than in II FIGO stages and in patients with residual disease above 2 cm. A significant correlation between p53 accumulation and p53 gene alteration and poor response to cisplatin-based chemotherapy was shown. The overall survival time of patients decreased with an increase in p53 protein expression. A strong p53 expression especially accompanied by p53 changes detectable by PCR-SSCP analysis appears to be a good indicator of the resistance to cisplatin-based chemotherapy. The association between strong p53 overexpression and shorter overall survival time was also revealed.

Gen supresor de tumores p53 en neoplasias digestivas

Malignant diseases of the digestive tract cause more than 50% of deaths due to cancer in Chile. There is a high incidence of gastric and gallbladder cancer and an increasing frequency of colorectal cancer. P53 tumor suppressor gene has a great importance in carcinogenesis and its alterations are specially important in digestive tract tumors such as colorectal cancer. There is contradictory evidence about the frequency of p53 gene or protein alterations or their biological significance. There is little information about p53 in Chile and it is mostly limited to immunohistochemical studies. This revision analyzes the frequency of p53 alterations in digestive tract tumors in Chile, using immunohistochemical and molecular biology methods. A special emphasis is given to the prognostic importance of this gene.

Molecular Abnormalities of p53, MDM2, and H-ras in Synovial Sarcoma

Forty-nine cases of synovial sarcoma were evaluated for mutation of the p53 gene, amplification of the MDM2 gene and mutation of the H-ras gene, and for the relation of these factors to overall survival and clinicopathologic parameters. All investigations were carried out on formalin-fixed paraffin-embedded materials. Furthermore, we evaluated the expression of p53 protein, MDM2, and p21WAF1/CIP1 immunohistochemically in these cases, together with an assessment of proliferative activities using monoclonal antibody MIB-1. Nine of the 49 cases (18.4%) had p53 gene alteration detected by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) and direct sequencing. Eleven cases (24%) showed nuclear accumulation of p53 protein in more than 10% of the tumor cells. Among them, only three cases contained gene mutations. There was no correlation between p53 nuclear accumulation and p53 gene alteration. MDM2 gene amplification, as shown by differential PCR, was observed in 19 out of 47 cases (40%). Nineteen out of 49 cases (38.8%) showed immunoreactivity for MDM2. MDM2 gene amplification and the expression of MDM2 protein showed a significant positive relationship (P = 0.0004). Moreover, MDM2 immunoreaction was significantly correlated with nuclear accumulation of p53 protein (P = 0.023). Positive immunoreaction for p21WAF1/CIP1 was observed in 21 out of 48 cases (43.8%). p21WAF1/CIP1 expression was correlated with p53 protein expression. H-ras gene mutations were seen in only three cases (6.1%). All mutations were in codon 12 (one GGC-to-AGC [Gly-to-Ser] mutation and two GGC-to-GAC [Gly-to-Ap] mutations). The gene alteration of p53, MDM2, and H-ras did not affect the patients' prognosis. Although the cases with positive immunoreaction for p53 tended to have a worse prognosis, the difference was not statistically significant (P = 0.13). No correlation was observed between MIB-1 LI and the immunohistochemical expression of p53, MDM2, and p21WAF1/CIP1 or the mutation status of p53 and H-ras. On the other hand, high MIB-1 LI (more than 10) significantly correlated with poor prognosis (P < 0.0001). Our results suggest that p53 gene mutation does not appear to be a major prognostic factor and H-ras mutations are infrequent in synovial sarcoma.

3-methylcholanthrene triggers the differentiation of alveolar tumor cells from canine bronchial basal cells and an altered p53 gene promotes their clonal expansion

3-methylcholanthrene triggers the differentiation of alveolar tumor cells from canine bronchial basal cells and an altered p53 gene promotes their clonal expansion

3-Methylcholanthrene triggers the differentiation of alveolar tumor cells from canine bronchial basal cells and an altered p53 gene promotes their clonal expansion

3-Methylcholanthrene triggers the differentiation of alveolar tumor cells from canine bronchial basal cells and an altered p53 gene promotes their clonal expansion

3-Methylcholanthrene triggers the differentiation of alveolar tumor cells from canine bronchial basal cells and an altered p53 gene promotes their clonal expansion

Alveolar type II cells arising in canine bronchial autografts following exposure to 3-methylcholanthrene (MCA) give rise to carcinomas of varying glandular and squamous growth patterns. To study the role of the tumor suppressor gene p53 in this process, sections from progressive lesions were immunostained for p53 protein; microdissected regions were screened for p53 mutations. Adjacent sections were examined for type II cell expression [cuboid cell shape, large roundish nucleus, cytoplasmic staining for surfactant protein A (SP-A)] and proliferating cell nuclear antigen expression. Evidence for an altered p53 function (nuclear staining, missense mutations) was found in most carcinomas of all histologic types and in all grades of bronchial dysplasia, but not in hyperplastic or normal bronchial epithelium. It was primarily associated with the hyperplastic type II cell populations present in the basal zone of the lesions. In addition, we found SP-A staining in hyperplastic (but not in normal) bronchial basal cells. These data suggest that MCA initiates type II cell differentiation through phenotypic selection (basal cells). Inactivation of the p53 gene promotes the clonal expansion of the type II cells into discernible populations of (squamous or glandular) alveolar tumor cells. This in vivo study is the first to show that p53 is involved in a specific pathway leading to bronchogenic carcinoma.

K-ras gene mutations and loss of heterozygosity at the p53 gene locus relative to histological characteristics of mucin-producing tumors of the pancreas

Mucin-producing tumors (MPTs) of the pancreas accompanied by carcinomas usually include various grades of dysplasia in the ductal epithelium, and invasive areas are histologically similar to those of common invasive ductal carcinomas, suggesting that MPTs provide a good tool to investigate early stages of pancreatic carcinogenesis. Thus, to clarify genetic alterations in the early stage of pancreatic carcinogenesis, we analyzed K-ras gene mutations and loss of heterozygosity (LOH) at the p53 gene locus using 37 cases of MPTs harboring dysplastic epithelium. Further, we conducted an extended, multifocal microdissection analysis focusing on the histological features of ductal epithelium and the distribution of genetic alterations for 3 cases of MPT positive for LOH of the p53 gene to determine the relation to tumor progression. K-ras gene mutations were detected with high frequency in 50% or more cases of the adenomas (14 of 19), borderline tumors (4 of 7), and carcinomas (8 of 11), whereas LOH of the p53 gene was limited to carcinomas (3 of 5 informative cases, 60%) and always accompanied by K-ras gene mutation. Investigation of a total of 126 microdissection sites from 3 cases showed the presence of K-ras gene mutations in mild dysplasia and all (100%) regions of moderate or more marked dysplasia, whereas LOH of the p53 gene showed more gradual tendency to increase with grade from moderate dysplasia. In addition, the multifocal genetic analysis showed K-ras gene mutations to be widely distributed throughout tumors, whereas LOH of the p53 gene was localized to 1 or a few areas. Further, topographically delimited areas with the same histology in the same tumor did not always show the same genetic alteration. In conclusion, we could confirm that both the K-ras and p53 gene alterations occur in the intraductal stage of MPT, and the latter is superimposed on the former during the course of tumor progression. However, the pattern of association of histological features with genetic alteration differs from tumor to tumor. HUM PATHOL 31:795-803.

Allelic imbalance at the 5q14 locus is associated with decreased apoptotic rate in non-small cell lung carcinomas (NSCLCs). Possible synergistic effect with p53 gene alterations on apoptosis

Deletions in the 5q14 region have been described in a variety of neoplasms, such as testicular germ cell tumors, ovarian, gastric and lung cancer. The high frequency of allelic losses observed in this region implies the presence of putative tumor suppressor gene(s) (TSGs). In the present study, we investigated in a series of 56 non-small cell lung carcinomas (NSCLCs) the allelic imbalance (Alm) within the 5q14 region, employing the D5S644 marker, and its relationship with p53 abnormalities, the kinetic parameters [proliferation index (PI) and apoptotic index (AI)] and the ploidy status of the carcinomas. AI at D5S644 was found at a frequency of 51.2%. The rather high percentage of Alm in stage I tumors suggests an early involvement in NSCLC development. LOH at 5q14 was associated with decreased AR in lung tumors insinuating the presence of a putative TSG(s) ( P=0.008). Simultaneous alterations of both p53 and D5S644 locus were the most frequent pattern observed (37.5%). Cases demonstrating this profile also exhibited a marked decrease in AI ( P=0.001). These findings imply a synergistic mechanism of co-operation between different TSGs. However, proliferation activity was dependent only on p53 status, leading to the assumption that the putative TSG(s) present at 5q14 may probably be involved in normal apoptotic procedures. Further studies are needed to identify the candidate gene(s).

Microsatellite instability and mutation of mitochondrial and nuclear DNA in gastric carcinoma

Background & Aims: Microsatellite instability (MSI) in mitochondrial DNA (mtDNA) is observed in some colorectal carcinomas. We attempted to determine if mitochondrial MSI (mtMSI) and mutations occur in gastric carcinomas and if the mtMSI phenotype underlies specific clinicopathologic profiles. Methods: Sixty-two gastric carcinomas (34 intestinal and 28 diffuse types) were investigated. Coding mutations in 8 different mitochondrial genes, mtMSI in a noncoding (C)n tract, and p53 gene mutations were examined by polymerase chain reaction–single-strand conformation polymorphism (PCR-SSCP) analysis. MSI in nuclear DNA (nMSI) and loss of the p53 gene were examined using microsatellite markers. Results: Ten of 62 (16%) carcinomas showed the mtMSI phenotype. Mitochondrial gene mutation was detected in 5 carcinomas, 4 of which also showed the mtMSI phenotype. There was a positive correlation between mtMSI and nMSI status. In intestinal carcinomas, mtMSI, nMSI, and p53 gene alterations were frequently detected from early to advanced stages. In diffuse carcinomas, both kinds of MSI were found in only advanced (subserosal or serosal invasion) carcinomas. Six of 7 carcinomas with the nMSI phenotype and all 5 carcinomas with mitochondrial coding mutations had a considerable intestinal-type tumor cell component. Conclusions: Mitochondrial gene mutations, which are associated with the mtMSI phenotype, may play a specific role in the tumorigenesis of intestinal-type gastric carcinomas. GASTROENTEROLOGY 2000;118:835-841

P53 gene mutations occurring in spontaneous benign and malignant mammary tumors of the dog.

Sixty-three cases of benign and malignant canine mammary tumors were analyzed to define the alteration of exons 5-8 for the p53 tumor suppressor gene using polymerase chain reaction direct sequence analysis with paraffin-embedded tissues. Four missense mutations were found in 38 benign mammary tumors (11%), and five missense (one tumor had two missense mutations) and one nonsense mutations were found in 25 mammary carcinomas (20%). These data suggest that the p53 gene alterations might be initiated at an early stage of canine mammary carcinogenesis and p53 mutations might be associated with malignancy. However, there was no evidence of any relationship between the p53 alterations and the histologic types of tumors or breeds of dogs.

Amplification and Overexpression of mdm2 Gene in Ependymomas

Ependymomas rarely show p53 gene alteration, and the tumorigenic mechanism of ependymomas still remains to be elucidated. We investigated the amplification and overexpression of mdm2 gene, whose product (MDM2) is considered to be one of the major cellular regulators of p53-mediated growth control, in 26 specimens of ependymomas obtained from 20 patients. The majority of the ependymomatous samples (96%) showed at least focal immunopositivity for MDM2; however, only 8% of the samples were immunopositive for p53. mdm2 gene amplification was detected in 35% of the samples by differential polymerase chain reaction, all of which overexpressed MDM2. These results suggest that the amplification and/or overexpression of mdm2 may be one of the major molecular events occurring in the tumorigenesis of ependymomas.

Absence of p16, p21 and p53 gene alterations in hepatocellular carcinomas induced by N-nitrosodiethylamine or a choline-deficient l-amino acid-defined diet in rats

To clarify the involvement of tumor suppressor genes in exogenous and endogenous liver carcinogenesis, alterations of p16, p21 and p53 in hepatocellular carcinomas (HCCs) induced by N-nitrosodiethylamine (DEN) and a choline deficient l-amino acid-defined (CDAA) diet in rats were investigated. Male Fischer 344 rats received DEN at 6-week of age followed by partial hepatectomy (PH), with colchicine to induce cell cycle disturbance, and a selection pressure regimen. Sacrifice was after 42 weeks. Other animals continuously received a CDAA diet for 75 weeks and were then killed. Eleven and 15 HCCs were obtained, respectively. Total RNA was extracted from and cDNA was synthesized with reverse transcriptase to allow investigation of mutations in p16, p21 and p53 by polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) analysis. Expression of p16 and p21 mRNA was also analyzed by reverse transcription (RT)-PCR. The results showed no mutations or deletions of p16, p21 and p53 in any of the HCCs induced by DEN or CDAA. Loss or decrease of p16 and p21 expression were also not found, suggesting that p16, p21 and p53 alteration may not be necessary for either exogenous or endogenous liver carcinogenesis in rats.

Neoplastic cell apoptosis in nude mice transplants with nasopharyngeal carcinoma cell lines

Objective: To observe the morphological features of neoplastic cell apoptosis developed in nude mice transplants with nasopharyngeal carcinoma (NPC) cell lines, CNE-1 and CNE-2, and to investigate the roles of p53, bcl-2 and bax playing in the process of apoptosis. Methods: CNE-1 and CNE-2 cell lines were inoculated and passed in nude mice for 3 generations. The cell apoptosis was detected on H & E and TUNEL staining slides. The expression of p53, bcl-2 and bax were detected by using immunohistochemistry. p53 gene alteration was assayed in cell lines and transplants by PCR-SSCP. Results: A considerable number of neoplastic cells underwent apoptosis in CNE-1 and CNE-2 transplant tissues. The “shrinkage necrosis” and apoptotic bodies were the main appearances of apoptosis. The p53 alteration was detected in exon 8 by PCR-SSCP and p53 protein accumulation observed in the cell smears and nude mice transplant tissue sections. All the transplant tissue sections of 3 passages showed bcl-2 negativity and bax overexpression. Conclusion: The neoplastic cells of CNE-1 and CNE-2 transplants underwent death mainly taking the way of apoptosis. The “shrinkage necrosis” and apoptotic bodies were the main morphological features of apoptosis seen in those transplants. The apoptosis in CNE-1 or CNE-2 nude mice transplant is highly probable through a p53-independent and bax-mediated pathway.

The clinicopathologic significance of the p16 gene expression in primary lung cancer.

To explore the relationship between p16 gene expression and oncogenesis , and development of human lung cancer. Immunohistochemistry was employed to detect the expression of p16 protein in 114 paraffin-embedded primary lung cancer tissues. Loss of p16 INK4 gene exon 2 (E2) was analyzed using PCR technique in 62 cancer samples with p16 protein-negative stain and 14 paracancerous lung tissues. The positive rates of p16 protein expression in squamous cell carcinoma , adenocarcinoma and small cell lung cancer (SCLC) were 43. 3 % , 48. 2 % and 27. 0 % respectively , and no significant difference was found among them ( P > 0. 05) . The expression level of p16 protein was closely correlated with histological grade and lymph node metastasis of lung cancer. The deletion of p16 E2 was 45. 2 % in NSCLC and closely related to lymph node metastasis and histological grade ( P < 0. 05) . No deletion was observed in 37 cases of SCLC and 14 paracancerous lung tissues. Alteration of p16 gene and protein is closely related to the histological grade and lymph node metastasis of NSCLC. It may play an important role in the oncogenesis , development and lymph node metastasis of NSCLC.

Spontaneous apoptosis in ovarian carcinomas: a positive association with p53 gene mutation is dependent on growth fraction.

Changes in cell survival contribute to tumour development, influence tumour biology and its response to chemotherapy. p53 gene alterations should negatively affect apoptosis by impaired p53-dependent apoptotic response. We looked for associations between spontaneous apoptosis, p53 gene mutation, p53 protein accumulation, growth fraction, bcl-2 expression and histological parameters in 64 ovarian, four tubal and three peritoneal carcinomas. Apoptotic cells were detected with the TUNEL method. p53 gene variants were detected by the single-strand conformation polymorphism and were sequenced directly. P53, Ki-67 and bcl-2 protein expressions were detected immunohistochemically. A weighed multiple logistic regression model was applied. Apoptotic index (Al) ranged 0.02–0.18 (mean 0.11); proliferation index (PI) ranged 3–90% (mean 54%). p53 gene mutations were present in 51, p53 protein accumulation in 46, and diffuse bcl-2 expression in 29 of 71 tumours. The AI was positively associated with the presence of p53 gene mutation (P = 0.011). However, the PI included into the analysis did positively influence the AI (P = 0.02) and diminished the association with p53 gene mutation (P = 0.082). The AI was negatively associated with good histological differentiation (P = 0.0006), the serous tumour type (P = 0.002), and diffuse bcl-2 expression (P = 0.025). Strong bcl-2 expression was associated with endometrioid tumour type (P = 0.002). FIGO stage and p53 protein accumulation were the only parameters that influenced overall survival time. Thus, our results suggest that histological tumour type and grade are major determinants of spontaneous apoptosis in ovarian carcinomas;p53 alterations do not adversely but rather positively affect spontaneous apoptosis by increasing growth fraction. This, in turn, suggests p53-independency of spontaneous apoptosis in ovarian carcinomas. © 2000 Cancer Research Campaign

The role of p16-cyclin d/CDK-pRb pathway in the tumorigenesis of endometrioid-type endometrial carcinoma.

We analysed p16 gene alteration and p16, cyclin-dependent kinase 4 (CDK4), CDK6, cyclin D1, cyclin D2, cyclin D3 and retinoblastoma protein (pRb) expression in ten normal endometriums (PE), 18 endometrial hyperplasias (EH) and 35 endometrial cancers (EC). Two of ten PE (20%), nine of 18 EH (50.0%) and 29 of 35 EC (82.9%) exhibited p16 nuclear staining. p16 expression was significantly higher in EC than EH (P = 0.0119). In the six p16 (–) EC, one was considered to have reduced gene dosage consistent with possible homozygous deletion of the CDKN2 gene and three had methylation in 5′CpG island in the promoter region of the p16 gene, whereas none showed such reduced gene dosage and four had methylation in the nine p16 (–) EH. Strong CDK4 staining was observed in 12 of 35 EC (34.3%) and one of 18 EH (5.6%). The strong expression of CDK4 was higher in EC than in EH (P = 0.0399). The expression of CDK4 was higher in EH than PE (P = 0.0054). The abnormalities of p16-cyclin D/CDK-pRb pathway were detected in 18 of 35 EC (51.4%). In conclusion, the expression of p16 and CDK4 may be an early event in the neoplastic transformation of endometrial cancer. © 2000 Cancer Research Campaign

P16 gene alterations in locally advanced squamous cell carcinoma of head and neck

p16 gene alterations in locally advanced squamous cell carcinoma of head and neck

Analysis of mdm2 and p53 gene alterations in glioblastomas and its correlation with clinical factors.

Malignant gliomas are the most frequent primary brain tumors. Recent studies defined several genetic markers, which might characterize molecular-biological subsets of glioblastomas with probably prognostic implications. To elucidate the involvement of murine-double-minute (mdm)2 gene amplifications and mutations of the tumor suppressor gene p53 in the tumorigenesis of malignant gliomas we analyzed a series of 75 glioblastomas. The p53 mutations occur in one-third of glioblastomas, mdm2 amplifications were found in 13% of cases. Our analysis revealed a hot spot in the p53 gene locus in codon 156, the same point mutation was detected in 4 tumor samples. None of the mdm2 amplified tumors had p53 mutations, supporting the hypothesis, that mdm2 amplifications are alternative mechanisms for p53 inactivation. Patients with p53 mutated tumors were significantly younger characterized by a mean age of 44 years. Additionally association with longer overall survival could be detected for this subgroup of patients. In our study, survival estimation revealed a significant correlation of mdm2 gene amplification with shorter survival time, and support the hypothesis, that mdm2 oncogene activation appears to occur late in tumor progression and may be characteristic as negative prognostic marker.