Frequent inactivation of PTEN in prostate cancer cell lines and xenografts

Loss of chromosome 10q is a frequently observed genetic defect in prostate cancer. Recently, the PTEN/MMAC1 tumor suppressor gene was identified and mapped to chromosome 10q23.3. We studied PTEN structure and expression in 4 in vitro cell lines and 11 in vivo xenografts derived from six primary and nine metastatic human prostate cancers. DNA samples were allelotyped for eight polymorphic markers within and surrounding the PTEN gene. Additionally, the nine PTEN exons were tested for deletions. In five samples (PC3, PC133, PCEW, PC295, and PC324), homozygous deletions of the PTEN gene or parts of the gene were detected. PC295 contained a small homozygous deletion encompassing PTEN exon 5. In two DNAs (PC82 and PC346), nonsense mutations were found, and in two (LNCaP and PC374), frame-shift mutations were found. Missense mutations were not detected. PTEN mRNA expression was clearly observed in all cell lines and xenografts without large homozygous deletions, showing that PTEN down-regulation is not an important mechanism of PTEN inactivation. The high frequency (60%) of PTEN mutations and deletions indicates a significant role of this tumor suppressor gene in the pathogenesis of prostate cancer.     

p16(INK4a) gene inactivation by deletions, mutations, and hypermethylation is associated with transformed and aggressive variants of non-Hodgkin's lymphomas

The molecular mechanisms underlying the pathogenesis of aggressive lymphomas and the histological transformation of indolent variants are not well known. To determine the role of p16(INK4a) gene alterations in the pathogenesis of non-Hodgkin's lymphomas (NHLs) and the histological progression of indolent variants, we have analyzed the expression, deletions, and mutations of this gene in a series of 112 NHLs. Hypermethylation of the gene was also examined in a subset of tumors with lack of protein expression but without mutations or deletions of the gene. p16(INK4a) gene alterations were detected in 3 out of 64 (5%) indolent lymphomas but in 16 out of 48 (33%) primary or transformed aggressive variants. In the low-grade tumors, p16(INK4a) alterations were detected in 1 (4%) chronic lymphocytic leukemia (hemizygous missense mutation), 1 (6%) follicular lymphoma (homozygous deletion), and 1 (5%) typical mantle cell lymphoma (homozygous deletion). The two later cases followed an aggressive clinical evolution. In the aggressive tumors, p16(INK4a) gene alterations were observed in 2 (29%) Richter's syndromes (2 homozygous deletions), 3 (33%) transformed follicular lymphomas (1 homozygous deletion and 2 nonsense mutations), 3 (43%) blastoid mantle cell lymphomas (2 homozygous and 1 hemizygous deletions), 5 (28%) de novo large-cell lymphomas (1 homozygous deletion and 4 hypermethylations), 2 lymphoblastic lymphomas (2 homozygous deletions), and 1 of 2 anaplastic large cell lymphomas (hypermethylation). Protein expression was lost in all tumors with p16(INK4a) alterations except in the typical chronic lymphocytic leukemia (CLL) with hemizygous point mutation. Sequential samples of the indolent and transformed phase of three cases showed the presence of p16(INK4a) deletions in the Richter's syndrome but not in the CLL component of two cases, whereas in a follicular lymphoma the deletion was present in both the follicular tumor and in the diffuse large-cell lymphoma. In conclusion, these findings indicate that p16(INK4a) gene alterations are a relatively infrequent phenomenon in NHLs. However, deletions, mutations, and hypermethylation of the gene with loss of protein expression are associated with aggressive tumors and they may also participate in the histological progression of indolent lymphomas.     

Mutation analysis of the PTEN/MMAC1 gene in lung cancer

We studied PTEN/MMAC1, a newly discovered candidate tumor suppressor gene at 10q23.3, for mutations in lung cancer. One hundred and thirty-six lung cancer cell line DNAs (66 small cell lung cancers, SCLC, 61 non-small cell lung cancers, NSCLC, four mesotheliomas, five extrapulmonary small cell cancers) were analysed for PTEN/MMAC1 homozygous deletions and five (8%) SCLC lines showed homozygous deletions interrupting the PTEN/MMAC1 gene. Using single stranded conformation polymorphism (SSCP) analysis, we screened the PTEN/MMAC1 open reading frame of 53 lung cancer cell line cDNAs for point mutations and found that 3/35 SCLCs and 3/18 NSCLCs contained homozygous amino acid sequence altering mutations. Northern blot analysis revealed that expression of the PTEN/MMAC1 gene was considerably lower in all the tumor cell lines with point mutations while no expression was detected for cell lines with PTEN/MMAC1 homozygous deletions. Mutation analysis of 22 uncultured, microdissected, primary SCLC tumors and metastases showed two silent mutations, and two apparent homozygous deletions. We also discovered a processed pseudogene (PTEN2) which has 98.5% nt identity to PTEN/MMAC1, that needs to be accounted for in cDNA mutation analysis. Our findings suggest that genetic abnormalities of the PTEN/MMAC1 gene are only involved in a relatively small subset of lung cancers.     

Seasonal rhythm of red pigment concentrating hormone in the crayfish

To evaluate the significance of microsatellite instability (MI) and loss of heterozygosity (LOH) in the development of gastric lymphoma, we examined 33 tissue-samples of 20 primary gastric B-cell lymphomas (6 low-grade lymphomas of mucosa-associated lymphoid tissue [MALT; 10 samples] and 14 diffuse large B-cell lymphomas [23 samples]). MI and LOH were evaluated at 13 microsatellite loci. In MALT lymphoma, four of six cases showed MI at one to two microsatellite loci (average 1.0 per case, 0.8 per sample), whereas in diffuse B-cell lymphoma, all samples showed MI at one to five microsatellite loci (average 2.4 per case, 2.7 per sample) (p < 0.05 and p = 0.0001). MI at the c-myc gene locus was most frequent in both types of gastric lymphomas (3 of 6 and 11 of 14 cases, respectively). Regional heterogeneity of the MI pattern was observed in two of four cases of MALT lymphoma and in four of five cases of diffuse B-cell lymphoma. On the other hand, LOH was observed only in one MALT lymphoma and in three diffuse B-cell lymphomas. Genetic instability may be an important mechanism for the development and progression of gastric lymphoma. Frequent MI at the c-myc locus might reflect an activated state and the importance of this gene in mucosal lymphocytes of chronic gastritis.     

Identification of PTEN/MMAC1 alterations in uncultured melanomas and melanoma cell lines

A novel tumor suppressor gene, PTEN/MMAC1, has been recently shown to be mutated in gliomas, breast, prostate, kidney cancers and melanomas. Loss-of-heterozygosity studies in melanoma have suggested the presence of at least one chromosome 10q locus lost early in tumor progression. In this study, we screened 45 melanoma cell lines and 17 paired uncultured metastatic melanoma and peripheral blood specimens for PTEN/ MMAC1 alterations using PCR-SSCP and direct sequencing. We found nine melanoma cell lines with homozygous deletions (five with intragenic loss) and four cell lines with mutations (one nonsense and one frameshift; two intronic); from among our uncultured melanoma specimens, we found one tumor with a somatic 17 bp duplication in exon 7 leading to a premature stop codon and one tumor with a possible homozygous deletion. Furthermore, we have identified a novel intragenic polymorphism within intron 4 of PTEN/MMAC1. Taken together, these data suggest that PTEN/MMAC1 may be a chromosome 10q tumor suppressor important in melanoma tumor formation or progression.     

The expression of CD26 and CD40 ligand is mutually exclusive in human T-cell non-Hodgkin's lymphomas/leukemias

CD26 and CD40 ligand (CD40L) are surface molecules on human activated T lymphocytes that play a critical role in the regulation of lymphopoiesis. Both molecules are expressed on a restricted fraction of human T-cell non-Hodgkin's lymphomas (NHL)/leukemias; however, little is known about their functional and/or clinical significance in these disorders. In this study, the pattern of expression of CD40L was compared with that of the CD26 molecule. A series of 67 human T-cell NHL/leukemias and a panel of leukemia/lymphoma T-cell lines were evaluated by immunohistochemistry, flow cytometry, and RNA studies. The overall frequency of CD26+ and CD40L+ samples was rather similar (25/67 [37%] v 18/67 [27%]). However, the majority of CD26-expressing cases clustered in the lymphoblastic lymphomas (LBL)/T-acute lymphoblastic leukemias (ALL; 12/23) and CD30+ anaplastic large-cell (ALC) lymphomas (5/8), whereas CD40L+ lymphomas included a large fraction of mycosis fungoides (11/21 [52%]). CD26 and CD40L coexpression was found only in 2 myocosis fungoides cases and 1 small lymphocytic lymphoma. Thus, the expression of the two antigens was mutually exclusive in almost all T-cell lymphomas/leukemias. Accordingly, lymphoma cell lines expressed either one of the molecules or the relative amounts of CD26 and CD40L were inversely proportional. In contrast, reactive T lymphocytes from patients with non-neoplastic T-cell expansions and in vitro activated CD3+ or CD4+ normal T cells were found to coexpress CD40L and CD26. Results of a multivariate analysis showed that the expression of CD26 in T-cell LBL/ALL patients was associated to a worse outcome in terms of survival, as compared with patients with CD26- tumors (P < or = .0001). Based on our results, it can be concluded that, (1) as opposed to activated or reactive normal T cells, the expression of CD26 and of CD40L is mutually exclusive in human T-cell lymphomas/leukemias; (2) expression of CD26 is restricted to aggressive pathologic entities, such as T-cell LBL/ALL and T-cell CD30+ ALC lymphomas, whereas CD40L is expressed on slow progressing diseases such as mycosis fungoides; and (3) within the T-cell LBL/ALL group of tumors, CD26 may identify a subset of poor prognosis patients.     

Deletion analysis of the p16 tumor suppressor gene in gastrointestinal mucosa-associated lymphoid tissue lymphomas

BACKGROUND & AIMS: The molecular mechanisms responsible for initiation and progression of gastrointestinal mucosa-associated lymphoid tissue (MALT) lymphomas are largely unknown. The aim of this study was to analyze the p16 tumor suppressor gene in MALT lymphomas of the stomach and colon. METHODS: Tumor samples were obtained from 28 patients with low-grade (n = 12) and high-grade (n = 14) gastric MALT lymphomas and from 2 patients with colonic MALT lymphomas. DNA was extracted from microdissected areas with at least 80% tumor cells. To detect homozygous p16 deletions, a semiquantitative polymerase chain reaction assay was used, whereby either p16 exon 1 or exon 2 was coamplified with an unrelated sequence as internal control. RESULTS: Homozygous p16 deletions were found in 2 of 14 (14%) cases with high-grade gastric MALT lymphomas. Both patients had Helicobacter pylori-associated gastritis; however, DNA extracted from areas of gastritis showed a normal p16 complement. No deletion was found in any of the low-grade gastric or the colonic MALT lymphoma specimens. CONCLUSIONS: In a subset of gastric MALT lymphomas, homozygous p16 deletions are acquired and may contribute to the transformation from a low-grade to a high-grade malignancy.     

Disruption of the MMAC1/PTEN gene by deletion or mutation is a frequent event in malignant melanoma

The MMAC1/PTEN gene, located at 10q23.3, is a candidate tumor suppressor commonly mutated in glioma. We have studied the pattern of deletion, mutation, and expression of MMAC1/PTEN in 35 unrelated melanoma cell lines. Nine (26%) of the cell lines showed partial or complete homozygous deletion of the MMAC1/PTEN gene, and another six (17%) harbored a mutation in combination with loss of the second allele. Mutations could also be demonstrated in uncultured tumor specimens from which the cell lines had been established, and cell lines derived from two different metastases from one individual carried the same missense mutation. Collectively, these findings suggest that disruption of MMAC1/PTEN by allelic loss or mutation may contribute to the pathogenesis or neoplastic evolution in a large proportion of malignant melanomas.     

Point mutation and homozygous deletion of PTEN/MMAC1 in primary bladder cancers

A new tumor suppressor gene PTEN/MMAC1 was recently isolated at chromosome 10q23 and found to be inactivated by point mutation or homozygous deletion in glioma, prostate and breast cancer. PTEN/MMAC1 was also identified as the gene predisposing to Cowden disease, an autosomal dominant cancer predisposition syndrome associated with an increased risk of breast, skin and thyroid tumors and occasional cases of other cancers including bladder and renal cell carcinoma. We screened 345 urinary tract cancers by microsatellite analysis and found chromosome 10q to be deleted in 65 of 285 (23%) bladder and 15 of 60 (25%) renal cell cancers. We then screened the entire PTEN/MMAC1 coding region for mutation in 25 bladder and 15 renal cell primary tumors with deletion of chromosome 10q. Two somatic point mutations, a frameshift and a splicing variant, were found in the panel of bladder tumors while no mutation was observed in the renal cell carcinomas. To screen for homozygous deletion, we isolated two polymorphic microsatellite repeats from genomic BAC clones containing the PTEN/MMAC1 gene. Using these new informative markers, we identified apparent retention at the gene locus indicative of homozygous deletion of PTEN/MMAC1 in four of 65 bladder and 0 of 15 renal cell tumors with LOH through chromosome 10q. Identification of the second inactivation event in six bladder tumors with LOH of 10q implies that the PTEN/MMAC1 gene is occasionally involved in bladder tumorigenesis. However, the low frequency of biallelic inactivation suggests that either PTEN/MMAC1 is inactivated by other mechanisms or it is not the only target of chromosome 10q deletion in primary bladder and renal cell cancer.     

Analysis of PTEN and the 10q23 region in primary prostate carcinomas

Deletions involving chromosome 10q23 occur frequently in prostatic carcinomas. Recently, a novel tumour suppressor gene, PTEN, mapping to this interval, has been identified. Mutation or deletion of PTEN has been observed in a proportion of prostate cancer cell lines; however, primary prostate carcinomas have not been studied. We have investigated the involvement of PTEN in primary prostatic adenocarcinomas using a panel of 51 matched normal and prostate tumour DNAs. We first determined the proportion of tumours with allele loss at loci in 10q23 which span the region containing the PTEN gene. Our results show that LOH involving 10q23 is common in primary prostate carcinomas. Twenty-five of 51 (49%) tumours showed loss of heterozygosity (LOH) over the region spanning the PTEN locus. We next directly analysed the PTEN gene for mutations of the coding region using single strand conformation polymorphism (SSCP) and sequence analyses. Of those tumours with LOH, only a single tumour was found to carry a missense mutation in PTEN. No mutations in PTEN were identified in tumours without LOH. Our results suggest either that mutation of PTEN is a late event in prostate tumorigenesis, or that another tumour suppressor gene important in prostate cancer may lie close to PTEN in 10q23.     

Review of alterations of the cyclin-dependent kinase inhibitor INK4 family genes p15, p16, p18 and p19 in human leukemia-lymphoma cells

The cyclin-dependent kinase inhibitors known as p15, p16, p18 and p19 have been suggested as candidates for tumor suppressor genes. The main genetic alterations are deletions (bi- or monoallelic) or 5' CpG island methylation of p15 and p16; very few cases or cell lines had p18 or p19 deletions or hypermethylation. Hypermethylation and homozygous deletions of tumor suppressor genes establish a new paradigm of inactivation by lack of expression, in contrast to the previously identified tumor suppressors which are predominantly inactivated by point mutations followed by loss of the wild-type allele. Here, the literature data on alterations of this gene family in more than 4700 primary cases of leukemia or lymphoma and some 320 continuous leukemia-lymphoma cell lines are summarized. Among hematopoietic malignancies, the highest frequencies of p15del and p16del were seen in acute lymphoblastic leukemia (ALL) (>30%) with striking rates in T-ALL (>50%), but also high rates in B cell precursor (BCP)-ALL (>20%); the rates of deletions in chronic lymphoid leukemia (CLL), multiple myeloma, acute and chronic myeloid leukemia (AML and CML), and myelodysplastic syndromes (MDS) were rather low, only some B cell and T cell lymphomas showed increased frequencies. Results are quite different with regard to the second mode of inactivation, hypermethylation of the promoter region. Here, p15 is most often inactivated, at particularly high frequencies in the disorders lacking any p15/p16 deletions: 40-80% p15met in AML, MDS and multiple myeloma. Also p15met rates in BCP- and T-ALL cases were high (c. 40%). There is controversy concerning the prognostic impact of p15 and p16 aberrations with some studies describing a significant correlation between inactivation of these genes and poor prognosis, while most others did not detect any prognostic relevance, at least in pediatric ALL; there may be a worse prognosis for adults with B or T cell lymphomas. Despite the small number of cases studied, paired sequential analyses suggested that disease progression is associated with loss of p15/p16 activity in a certain percentage of adult patients. p15del/p16del and p15met/p16met were also detected in the large panel of leukemia-lymphoma cell lines studied. In general, the results in cell lines reproduce the data seen in primary cells with the important difference that the rates of p15/p16 inactivation are clearly higher in the cultured cells compared with the freshly explanted cells. Retrovirus- or electroporation-mediated ectopic gene transfer of p16 wild-type into p16-deficient cell lines led to growth inhibition, arrest in G1 (without apoptosis) and occasionally to differentiation, suggesting that the malignant phenotype of p16-/- cell lines can, at least partially, be reversed by restoring p16 gene expression. A striking inverse correlation between the absence of p16 (due to deletion) and presence of wild-type retinoblastoma gene was observed in cell lines confirming a common growth suppressor pathway; no comparable relationship of p16 inactivation with p53 was detected. Paired analysis of cell lines and corresponding primary cell material showed that in all instances tested both populations carried the same gene configuration of p15 and p16. Thus, p15del or p16del did not occur during establishment of the cell lines or during prolonged culture. It is likely that p15 or p16 deletions already acquired in vivo provide a dramatic growth advantage for the immortalization process in vitro, thus increasing the success rate for cell line establishment which is commonly extremely difficult. In conclusion, the present review suggests an involvement of the p15 and p16 tumor suppressor genes in leukemo- and lymphomagenesis. Future studies will determine their exact role in the development and progression of hematopoietic neoplasms. These genes may represent interesting targets for new therapeutic strategies.     

The t(11;18)(q21;q21) chromosome translocation is a frequent and specific aberration in low-grade but not high-grade malignant non-Hodgkin's lymphomas of the mucosa-associated lymphoid tissue (MALT-) type

Primary extranodal malignant non-Hodgkin's lymphoma arising from the mucosa-associated lymphoid tissue (MALT-type lymphoma) represents a subtype of B-cell lymphoid malignancies with distinct clinicopathological features and is often associated with a favorable prognosis. Unlike the situation in nodal non-Hodgkin's lymphoma of B-cell lineage, few data are still available concerning the chromosomal constitution of MALT-type lymphomas. Until now, cytogenetic data from 29 low-grade MALT lymphomas with karyotypic alterations have been reported from different institutions, and virtually no data were available for high-grade MALT-type lymphomas. We have analyzed the cytogenetics of 44 MALT lymphomas arising in the stomach, parotid gland, thyroid gland, lung, breast, and conjunctiva. Clonal chromosome aberrations have been detected in 13 of 20 (65%) low-grade and 20 of 24 (83%) high-grade tumors. More than half of the low-grade lymphomas with abnormal karyotypes (7 of 13 cases, 53%) displayed clonal t(11;18)(q21;q21), thus specifically associating this translocation with MALT-type lymphomas for the first time in a larger series. In contrast, t(11;18) was not found in a single case of 20 high-grade MALT-type lymphomas with abnormal karyotypes, nor were translocations t(14;18) or t(3;14), characterizing about 10-35% of primary nodal large cell lymphomas. Instead, these lymphomas were associated with t(8;14)(q24;q32) in three cases, frequent deletions in the long arm of chromosome 6, and partial or whole gains of chromosomes 3, 7, 17, 18, and 21.     

儿童侵袭性成熟B细胞淋巴瘤的临床病理学及分子遗传学特点

目的 分析总结中国儿童各类型侵袭性成熟B细胞淋巴瘤的临床病理学及分子遗传学特点,为其诊断的标准化提供依据.方法 收集97例儿童侵袭性成熟B细胞淋巴瘤石蜡包埋组织标本,包括伯基特淋巴瘤(BL)81例、弥漫大B细胞淋巴瘤(DLBCL)8例、介于BL和DLBCL间的不能分类的B细胞淋巴瘤(BL/DLBCL)8例,利用免疫组织化学技术和间期荧光原位杂交(FISH)技术检测其免疫表型和分子遗传学特征.结果 BL的bcl-2和MUM1的阳性率分别为3%(2/66)和17%(12/71),DLBCL分别为50%(4/8)和63%(5/8),BL/DLBCL分别为50%(4/8)和63%(5/8).BL、DLBCL和BL/DLBCL的Ki-67平均值分别为(93±4.4)%、(83±14.3)%和(80±11.5)%.BL、DLBCL和BL/DLBCL的c-myc基因易位的比例分别为98%(79/81)、38%(3/8)和50%(4/8).38%(3/8)的DLBCL和25%(2/8)的BL/DLBCL存在bcl-6基因的多拷贝,BL与DLBCL之间、BL与BL/DLBCL之间bcl-2、MUM1和Ki-67平均值的差异及c-myc基因易位和bcl-6基因多拷贝的差异均有统计学意义(均P＜0.05).结论 儿童侵袭性成熟B细胞淋巴瘤的诊断和分型需要综合分析形态学、免疫表型和分子遗传学特征.儿童BL/DLBCL可能是DLBCL的一个亚型.CD10+、bcl-6+、bcl-2-、Ki-67＞90%、伴有IGH/c-myc重排、不伴有bcl-2和bcl-6重排时,支持BL的诊断;bcl-2+、Ki-67为50%～90%,同时伴有bcl-6基因的多拷贝时,支持DLBCL或BL/DLBCL的诊断.     

Biased VH family usage in primary gastric B cell lymphoma of mucosa-associated lymphoid tissue-type and the selected V beta expression of T cells infiltrating into the lymphoma cells

Seven cases of primary gastric low-grade B cell lymphoma of mucosa-associated lymphoid tissue (MALT) type, two cases of high-grade B cell lymphoma with a low-grade component and three cases of pure high-grade lymphoma were selected for the current study. The Ig VH gene use of lymphoma cells and the V beta repertoires of infiltrating T cells were investigated. The VH gene analysis showed multiple VH family usage in 12 cases, but the MALT-type lymphoma cell usage was found to be biased for the families that have a low number of VH genes (VHIV and V). Another analysis of lymphoma-infiltrating T cells showed restricted expressions of the V beta repertoire in all seven low-grade cases and three high-grade cases. In those 10 cases, a considerable number of CD4-positive T cells infiltrated into lymphoma cells and RAG-1 was also prominently expressed. Based on these findings, it was thus assumed that the normal counterpart of gastric B cell lymphoma of MALT type is different from the conventional B cell lymphoma, and the restricted expression of V beta repertoires is therefore considered to be a characteristic finding in low-grade B cell lymphomas of MALT type as well as in a proportion of high-grade lymphomas (the so called 'high-grade lymphoma of MALT type').     

Nurses achieve quality with pre-assessment clinics

Twelve patients with diagnosis of B-cell non-Hodgkin's lymphoma/leukemia and del[7q] were studied for their clinical, cytogenetic, and molecular characteristics. Eleven patients were classified as small cell lymphoma whereas one had a diffuse large cell lymphoma. Lymphoplasmacytic features were observed in six out of eleven small cell lymphomas. Morphologically and immunologically these small cell lymphomas could be classified as chronic lymphocytic leukemia (typical or atypical; 4 cases), marginal zone lymphoma (splenic lymphoma with villous lymphocytes; 1 case), mantle cell lymphoma (2 cases), or nonspecified, non-Hodgkin's lymphoma (4 cases). Eleven of twelve patients presented with peripheral blood and bone marrow involvement. Two of twelve cases showed del[7q] as the sole anomaly. Two different types of deletions were present: ten cases had del(7)(q21q31) and two cases had del(7)(q31q34). Cases that could be molecularly investigated did not show any involvement of BCL2, BCL3, or BCL6, and only one case had BCL1 rearrangement. The data indicate that del(7q) is associated with a subset of mature small B-cell lymphoproliferative disorders of which some but not all show lymphoplasmatic features.     

Genotypic analysis of tumor suppressor genes PTEN/MMAC1 and p53 in head and neck squamous cell carcinomas

OBJECTIVES: Tumor suppressor gene mutations in both p53 and PTEN/MMAC1 genomic DNA have been detected in many types of cancer. The purpose of this study was to investigate the presence and importance of PTEN/MMAC1 mutations in squamous cell carcinomas. METHODS: Exons of each gene were amplified after polymerase chain reaction (PCR) using genomic DNA derived from cell lines of squamous cell carcinoma of the head and neck (SCCHN) and snap-frozen biopsy specimens from primary established head and neck tumors. The amplified and purified DNA was then sequenced directly. RESULT: As anticipated, point mutations of the p53 gene were found in 80% of cell lines examined. A single base mutation in codon 151 was found in six of 10 cell lines studied. PTEN/MMAC1 gene mutations were found in neither the cell lines tested nor the tumor biopsy samples. CONCLUSION: This study, as well as a large volume of data, confirms that mutations of the p53 gene are frequent events in head and neck cancer cell lines. Although PTEN/MMAC1 gene mutations have been found in a variety of carcinomas, this gene was not found to be mutated in SCCHN cell lines or in primary squamous cell carcinomas of the head and neck. This information is useful for further studies of mutations in these cell lines.     

Frequent deletion and 5' CpG island methylation of the p16 gene in primary malignant lymphoma of the brain

A total of 10 primary malignant lymphomas of the brain were examined for deletion, mutation, and 5' CpG island methylation of the p16 gene, which is a candidate tumor suppressor gene with CDK-inhibitory function. In Southern blot analysis, p16 gene deletion was suggested in nine cases, homozygously (five cases) or hemizygously (four cases). In the remaining one case, p16 gene deletion was not suggested. Although single-strand conformation polymorphism and nucleotide analyses suggested no mutations of the p16 gene in these cases, methylation analyses revealed 5' CpG island methylation in three cases, of which two were those with presumed hemizygous deletion and one was that without deletion in Southern blot analysis. Thus, p16 gene abnormality was detected in all 10 of the brain lymphomas examined, and in 8 of them, actual p16 gene inactivation was suggested by their homozygous deletion (5 cases) or 5' CpG island methylation (3 cases). These findings suggest that p16 gene abnormality and inactivation are closely related to carcinogenesis in primary malignant lymphoma of the brain. The p15 gene, another candidate tumor suppressor gene located in the vicinity of the p16 gene, to which it shows structural and functional similarity, was also presumed to be deleted similarly in most cases. Its methylation was seen in one case, the case without the methylated p16 gene.