PTEN mutations do not cause nuclear beta-catenin accumulation in endometrial carcinomas

PTEN: and beta-catenin mutations constitute the predominant genetic alterations in endometrioid carcinomas of the endometrium. PTEN encodes a dual-specificity phosphatase with lipid phosphatase and protein tyrosine phosphatase activities that regulate both apoptosis and interactions with the extracellular matrix. Recent studies have associated PTEN mutations with tumorigenesis of prostate carcinoma via the Wnt signaling pathway, leading to nuclear beta-catenin accumulation. To elucidate the potential interaction of PTEN and beta-catenin in endometrial cancer, we performed mutation analyses of the entire PTEN gene and of exon 3 of the beta-catenin gene that is most frequently targeted by mutations. A total of 82 endometrial carcinomas comprising 62 type I endometrioid carcinomas and 20 type II high-grade carcinomas were investigated. In addition in a subset of 22 carcinomas, the intracellular beta-catenin distribution was analyzed by immunohistochemistry. Overall, 20 (24.4%) of 82 tumors revealed mutations in the PTEN gene, and 16 (19.5%) of 82, in the beta-catenin gene. Six tumors (7.3%) showed mutations in both the PTEN and beta-catenin gene. Mutations were mainly detected in endometrioid carcinomas of the endometrium. As expected, a striking nuclear accumulation of beta-catenin could be shown in tumors with beta-catenin mutations. In the vast majority of tumors with PTEN mutations, a regular staining pattern of the cytoplasmic and membranous compartments was found. We therefore conclude that, in contrast to prostate cancer, mutations in the PTEN gene seem not to affect cellular distribution of the beta-catenin protein in endometrial carcinomas.     

Aberrant expression of beta-catenin and mutation of exon 3 of the beta-catenin gene in renal and urothelial carcinomas

The present study attempted to clarify the significance of aberrant expression of beta-catenin protein and mutation of exon 3 of the beta-catenin gene in renal and urothelial carcinogenesis. beta-Catenin expression was examined immunohistochemically and mutation of the beta-catenin gene was analyzed by polymerase chain reaction-single strand conformation polymorphism (SSCP) and direct sequencing. beta-Catenin immunoreactivity was observed at the cell membrane in all 30 renal cell carcinomas (RCC) examined, and no RCC showed a mobility-shifted SSCP band. Of 46 transitional cell carcinomas (TCC) examined, there was reduced expression of beta-catenin, as compared with its expression in non-cancerous transitional epithelium, in 22 cases (48%) and beta-catenin accumulation in the nucleus in five cases (11%). Of four renal pelvis TCC examined, point mutation of exon 3 of the beta-catenin gene at codon 45 resulting in amino acid substitution (Ser to Phe) was detected in one (25%). The incidence of reduced expression of beta-catenin correlated significantly with the growth pattern (superficial type vs invasive type) of TCC (P < 0.05). These data indicate that: (1) aberrant beta-catenin expression may be at least partly involved in urothelial carcinogenesis, but less significantly so in renal carcinogenesis, and (2) it may be associated with the progression of TCC showing invasive growth.     

Renal medullary carcinomas: histopathologic phenotype associated with diverse genotypes

Chromosomal abnormalities and gene mutations have become major determinants in the classification of kidney carcinomas. Most renal medullary carcinomas develop in patients with hereditary sickle cell disease, but sporadic cases unassociated with sickle cell disease have also been described, for which underlying genetic abnormality is unknown. We evaluated 3 patients with renal medullary carcinoma (1 patient with sickle cell disease and 2 patients without sickle cell disease) for germ line and somatic mutations in genes commonly involved in pathogenesis of renal carcinomas using denaturing high-performance liquid chromatography and direct sequencing. Chromosomal abnormalities were studied by the conventional cytogenetic and SNP arrays analysis. Expression of hypoxia-inducible factor 1α was examined using immunohistochemistry. Two new mutations in the gene for fumarate hydratase were identified in 1 case of medullary renal carcinoma without sickle cell disease: a germ line mutation in exon 6 (R233H) and an acquired (somatic) mutation in exon 8 (P374S). No fumarate hydratase mutations were identified in the other 2 patients. The second sporadic case of renal medullary carcinoma harbored double somatic mutations in von Hippel-Lindau gene, and renal medullary carcinoma in the patient with sickle cell disease showed von Hippel-Lindau gene promoter methylation (epigenetic silencing). No consistent pattern of chromosomal abnormalities was found between 2 cases tested. All 3 cases showed increased hypoxia-inducible factor 1α expression. Medullary renal carcinomas from patients with or without sickle cell disease show involvement of genes important in hypoxia-induced signaling pathways. Generalized cellular hypoxia (in sickle cell disease) or pseudohypoxia (in tumors with fumarate hydratase and von Hippel-Lindau mutations or epigenetic silencing) may act alone or in concert at the level of medullary tubular epithelium to promote development of this rare type of renal carcinoma, which could then be genetically reclassified as either fumarate hydratase–associated renal carcinomas or high-grade clear cell renal cell carcinomas.     

Small cell carcinoma of the endometrium: report of a case with analysis of Wnt/beta-catenin pathway

Small cell carcinoma of the endometrium (SCCE) is extremely rare. Previous reports indicate that SCCE frequently shows systemic spread and has a poor prognosis. Beta-catenin has been shown to be a key downstream effector of the Wnt signaling pathway, which regulates cell growth and survival. Decreased membranous expression of beta-catenin in cancers correlates with poor prognosis and is associated with dissemination of tumor cells and the formation of metastases. Recently, some different investigators demonstrated aberrant beta-catenin accumulation in neuroendocrine tumors arising in different organs, suggesting a role for the Wnt/beta-catenin signaling pathway during neuroendocrine tumorigenesis. Here, we report a new case of SCCE associated with peritoneal spreading and aggressive course; the patient died one month after surgery. This study also aimed at assessing the involvement of the Wnt signaling pathway in this rare neuroendocrine tumor. Interestingly, both intense nuclear beta-catenin accumulation and cyclin D1 immunoreactivity were restricted to carcinoma cells invading lymphatic vessels. However, mutation analysis failed to demonstrate any mutation in exon 3 of the beta-catenin gene or exon 15 of the APC gene in the present case. Although the mechanism of nuclear accumulation of beta-catenin is still unknown, the heterotopic nuclear localization of beta-catenin may play a role in the tumor invasion process and, subsequently, may be associated with the aggressive behavior of SCCE.     

Nuclear Accumulation of Mutated β-Catenin in Hepatocellular Carcinoma Is Associated with Increased Cell Proliferation

Inappropriate activation of the Wnt pathway resulting from beta-catenin gene alterations has recently been implicated in the development of hepatocellular carcinoma (HCC). To explore the in vivo effects of mutated beta-catenin, HCC specimens from 32 patients carrying one or several tumors were screened for somatic mutations in exon 3 of the beta-catenin gene, and the expression and subcellular localization of beta-catenin was studied by immunohistochemistry. Missense mutations or interstitial deletions in beta-catenin exon 3 were detected in 12 of 35 (34%) HCC samples. After immunostaining, most tumors exhibited increased membranous and/or cytoplasmic expression of beta-catenin compared with adjacent nontumoral liver. Strong nuclear accumulation of beta-catenin was observed either focally or uniformly in 15 of 35 (43%) tumor specimens, but not in cirrhotic nodules or dysplastic liver cells in adjacent liver. Aberrant nuclear expression of beta-catenin was significantly associated with the presence of mutations in the beta-catenin gene (P < 0.005). Moreover, nuclear beta-catenin staining correlated significantly with increased Ki-67 proliferative index in tumor (P < 0.001) and seemed to be associated with poor outcome in patients with HCC. In conclusion, our data indicate that activation of the Wnt/beta-catenin pathway in HCC results mainly from somatic mutations in the beta-catenin gene and may promote tumor progression by stimulating tumor cell proliferation.     

Solid-pseudopapillary tumors of the pancreas are genetically distinct from pancreatic ductal adenocarcinomas and almost always harbor beta-catenin mutations

Solid-pseudopapillary tumors (SPTs) are unusual pancreatic neoplasms of low malignant potential that most frequently affect young women. Genetic events contributing to the development of SPTs are unknown. Whereas the more common ductal adenocarcinomas of the pancreas essentially never harbor beta-catenin or APC gene mutations, we have recently identified alterations of the APC/beta-catenin pathway in other nonductal pancreatic neoplasms including pancreatoblastomas and acinar cell carcinomas. We analyzed a series of 20 SPTs for somatic alterations of the APC/beta-catenin pathway using immunohistochemistry for beta-catenin protein accumulation, direct DNA sequencing of beta-catenin exon 3, and direct DNA sequencing of the mutation cluster region in exon 15 of the APC gene in those SPTs that did not harbor beta-catenin mutations. Immunohistochemical labeling for cyclin D1 was performed to evaluate the overexpression of this cell-cycle protein as one of the putative downstream effectors of beta-catenin dysregulation. In addition, we analyzed the SPTs for genetic alterations commonly found in pancreatic ductal adenocarcinomas, including mutations in the K-ras oncogene and p53 and DPC4 tumor suppressor genes, using direct DNA sequencing of K-ras and immunostaining for p53 and Dpc4. Almost all SPTs harbored alterations in the APC/beta-catenin pathway. Nuclear accumulation of beta-catenin protein was present in 95% (19 of 20), and activating beta-catenin oncogene mutations were identified in 90% (18 of 20) of the SPTs. Seventy-four percent (14 of 19) showed overexpression of cyclin D1, ranging from 10 to 70% of tumor nuclei. In contrast, no K-ras mutations were present in any of the 20 SPTs, and Dpc4 expression was intact in all 16 SPTs for which immunohistochemical labeling was successful. Overexpression of p53 was limited to only 3 of 19 (15.8%) SPTs. These results emphasize the two distinct, divergent genetic pathways of neoplastic progression in pancreatic ductal and nonductal neoplasms.     

MET expression in sporadic renal cell carcinomas

Although germline mutations of met proto-oncogene on human chromosome 7q31-34 have been known as useful molecular markers of hereditary papillary renal cell carcinoma (RCC), the expression of MET, a product of met proto-oncogene, has not been fully studied in sporadic RCC, along with its clinical significance. We investigated the expression of MET by immunohistochemistry in 182 cases of renal neoplasm encompassing 145 RCC, 25 urothelial carcinomas of renal pelvis, and 12 oncocytomas. MET was diffusely and strongly expressed in 90% of papillary RCC, all collecting duct carcinomas, and 92% of urothelial carcinomas of renal pelvis. On the contrary, clear cell RCC, chromophobe RCC, and oncocytomas were negative or focally positive for MET expression. In clear cell RCC, MET expression was positively correlated with high nuclear grade, presence of infiltrative growth, tumoral necrosis, papillary architecture, sarcomatoid component, tumoral involvement of the renal pelvis or ureter, involvement of the calyx, and lymphatic invasion. In conclusion, diffuse and strong expression of MET in papillary RCC and collecting duct carcinoma might be helpful in discriminating from the other subtypes of RCC with tubular or papillary growth. In case of MET expression observed in clear cell RCC, it might correlate with those clinicopathological parameters implying aggressive behavior.     

Beta-catenin dysregulation in thyroid neoplasms: down-regulation, aberrant nuclear expression, and CTNNB1 exon 3 mutations are markers for aggressive tumor phenotypes and poor prognosis

beta-catenin has a role in cell adhesion and Wnt signaling. It is mutated or otherwise dysregulated in a variety of human cancers. In this study we assess beta-catenin alteration in 145 thyroid tumors samples from 127 patients. beta-catenin was localized using immunofluorescence and mutational analysis was performed by single-strand conformational polymorphism. Membrane beta-catenin expression was decreased in eight of 12 (66%) adenomas and in all 115 carcinomas (P: < 0.0001). Among carcinomas, reduced membrane beta-catenin was associated with progressive loss of tumor differentiation (P: < 0.0001). CTNNB1 exon 3 mutations and nuclear beta-catenin localization were restricted to poorly differentiated [7 of 28 (25%) and 6 of 28 cases (21.4%), respectively] or undifferentiated carcinomas [19 of 29 (65.5%) and 14 of 29 (48.3%) cases, respectively]. Poorly differentiated tumors always featured mutations involving Ser and Thr residues and were characterized by Thr to Ile amino acid substitutions (P: = 0.0283). The association between CTNNB1 exon 3 mutations and aberrant nuclear immunoreactivity (P: = 0.0020) is consistent with Wnt activation because of stabilizing beta-catenin mutations. Low membrane beta-catenin expression as well as its nuclear localization or CTNNB1 exon 3 mutations are significantly associated with poor prognosis, independent of conventional prognostic indicators for thyroid cancer but not of tumor differentiation. Analysis of beta-catenin dysregulation may be useful to objectively subtype thyroid neoplasms and more accurately predict outcomes.     

Aberrant nuclear localization and gene mutation of beta-catenin in low-grade adenocarcinoma of fetal lung type: up-regulation of the Wnt signaling pathway may be a common denominator for the development of tumors that form morules.

The salient histopathologic features of low-grade adenocarcinoma of the fetal lung type (L-FLAC)/well-differentiated fetal adenocarcinoma (WDFA) include complex glandular structures and morules with biotin-rich optically clear nuclei. Interestingly, these characteristic features are shared by the cribriform-morular variant of papillary thyroid carcinoma, whose morphology is identical to that of familial adenomatous polyposis (FAP)-associated thyroid carcinoma. Furthermore, the single reported case of lung cancer associated with FAP was L-FLAC/WDFA. These observations lead us to hypothesize that up-regulation of the Wnt signaling pathway underlies the development of L-FLAC/WDFA. To verify this hypothesis, 11 cases of L-FLAC/WDFA, including the one FAP-associated case, eight cases of high-grade adenocarcinoma of the fetal lung type (H-FLAC), 24 cases of conventional pulmonary adenocarcinoma (CAC), and 13 fetal lungs were immunostained for beta-catenin. All cases of L-FLAC/WDFA showed predominantly aberrant nuclear/cytoplasmic expression, especially in budding glands and morules, whereas six of eight cases (75%) of H-FLAC and all but one case (96%) of CAC showed predominantly membranous expression. Fetal lungs showed nuclear/cytoplasmic expression restricted to the distal branching airway epithelium. Mutational analysis of exon 3 of the beta-catenin gene in five sporadic cases of L-FLAC/WDFA showed a point mutation at codon 34 and codon 37 in two cases, respectively. The present study indicates that up-regulating disturbances in the Wnt signaling pathway, including mutation of the beta-catenin gene, underlie tumorigenesis of L-FLAC/WDFA. The expression pattern of beta-catenin in L-FLAC/WDFA resembles that of the developing fetal lung airway. With the expression pattern of beta-catenin as a marker, most cases of H-FLAC as well as CAC appear to have different oncogenic pathways from cases of L-FLAC/WDFA. The present study together with other available data also suggests that abnormal up-regulation of the Wnt signaling pathway may be a common denominator for the development of tumors with morular formation from a variety of anatomic sites.     

青少年肾细胞癌的临床病理及分子遗传学研究

目的 探讨青少年肾细胞癌的临床病理特征、遗传学改变、鉴别诊断及预后.方法 对46例青少年肾细胞癌进行光镜观察及免疫组织化学染色,随访并复习相关文献.对46例肿瘤进行von Hippel-Lindau(VHL)基因区域杂合性缺失(LOH)及VHL基因突变筛查.结果 共诊断19例Xp11.2易位/TFE3基因融合相关性肾癌(Xp11 RCC)、9例透明细胞癌、17例乳头状肾细胞癌(PRCC)和1例不能分类肾细胞癌.19例Xp11 RCC均TFE3阳性,而TFEB阴性.8例肿瘤具有巢状和乳头状结构形态类似t(X;17)ASPL-TFE3型肾癌,6例肿瘤组织学类似t(X;1)PRCC-TFE3型肾癌,4例肿瘤形态像透明细胞癌,1例肿瘤组织学形态文献中未被检索到,表现为细胞核呈毛玻璃样,核仁不明显,可见核沟,肿瘤间质见大量黏液.LOH及VHL突变检测结果显示,仅1例透明细胞癌和1例2型PRCC存在LOH,并且该2型PRCC的VHL基因的一个剪切位点存在胚系突变,553+5 G→C.其余45例均未检测出VHL突变.统计学分析表明TFE3阳性肾细胞癌比TFE3阴性肾细胞癌更倾向于高病理分期(pT3/pT4),并且预后较差(P=0.035).结论 青少年肾细胞癌表现出不同的组织学形态以及分子遗传学背景.其中Xp11 RCC为最常见的肾癌亚型.TFE3阳性肾细胞癌的预后要差于TFE3阴性肾细胞癌.     

Mutations of the p53 gene in carcinomas of the urinary system

Deletion of p53, which is an anti-oncogene located on chromosome 17p, was reported to be present at a high incidence in tumor cells of colorectal carcinoma, as well as osteosarcoma of the familial cancer syndrome. Mutations of the p53 gene were investigated in 59 surgical specimens of primary carcinomas of the urinary system from 57 patients, using the polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis. The PCR products were sequenced using the dideoxy chain termination method or the DNA sequencer. The tumors examined were 20 transitional cell carcinomas (TCC) and 39 renal cell carcinomas (RCC). Mutations of the p53 gene were detected in 20.0% (4/20) of TCC and were present in 16.7% (1/6) of the tumors invading the muscular layer. In two patients with simultaneous double bladder TCC, the mutations were found only in the larger tumors. In RCC, mutations were detected in 7.7% (3/39) of patients. No significant correlation between the presence of the mutation and the clinicopathologic parameters was found in RCC except that the three tumors with p53 gene mutations were clear cell carcinomas. These results suggest that p53 gene mutations play a possible role in both carcinogenesis and progression of TCC, but the p53 gene mutations may not be significant in development of RCC.     

Subtypes of renal cell carcinoma with defined genomic alterations: diagnostic and prognostic significance

The World Health Organization (WHO) 2016 classification of renal neoplasia defines renal cell carcinomas (RCCs) with genomic alterations: (1) succinate dehydrogenase deficient (SDH) RCC, (2) hereditary leiomyomatosis and renal cell carcinoma (HLRCC) syndrome-associated RCC and (3) the MiT family translocation carcinoma (Xp11 translocation carcinoma and t(6;11) carcinoma). Genomic alterations also define two syndromes associated with RCC that have a varied histology; tuberous sclerosis complex and Birt-Hogg-Dube. This review will examine the WHO entities and the two aforementioned syndromes, and discuss a genomic alteration (TCEB1 mutation) that appears to define a subset of clear cell RCCs that histomorphologically overlap with clear cell tubulopapillary RCC. The focus will be on diagnostic considerations from the pathologic perspective, and include discussion of clinical features, prevalence, prognosis, common and uncommon immunohistochemistical stains and molecular testing.     

beta-Catenin expression pattern, beta-catenin gene mutations, and microsatellite instability in endometrioid ovarian carcinomas and synchronous endometrial carcinomas.

beta-Catenin gene mutations and microsatellite instability (MI) have been reported in endometrioid ovarian carcinomas. In colon but not endometrial cancer, beta-catenin gene mutations are associated with a replication error phenotype and MI. In this study the authors investigate whether beta-catenin mutations and MI are two independent oncogenic pathways in endometrioid ovarian carcinomas. They also evaluate the usefulness of these molecular markers in determining the primary origin of simultaneous tumors in the ovary and endometrium. This study was performed on 26 patients diagnosed with primary endometrioid ovarian carcinoma, five of whom also had pathologically diagnosed primary synchronous endometrioid endometrial carcinoma. Immunohistochemical and molecular analyses indicated that there were 25 primary ovarian tumors with four primary synchronous endometrial cancers and one ovarian metastasis of a primary endometrial carcinoma. All studies were performed on formalin-fixed, paraffin-embedded tissue samples. The beta-catenin expression pattern (nuclear vs. membranous) was analyzed immunohistochemically. Mutations in exon 3 of the beta-catenin gene were studied by polymerase chain reaction, single-strand conformational polymorphism, and direct sequencing. MI status was established by studying BAT-26 and BAT-25 mononucleotide repeats. In the group with 21 single ovarian tumors, 18 (85%) had beta-catenin nuclear expression, eight (38%) had beta-catenin gene mutations (always associated with beta-catenin nuclear expression), and four (19%) had MI. Only one case (5%) had both beta-catenin gene mutations and MI. The mutations affected one of the serine/threonine residues targeted for phosphorylation by glycogen synthase kinase-3beta or adjacent residues. At codon 32, a GAC-to-TAC (D32Y) change was found; at codon 33, two TCT-to-TGT (S33C) changes were found; at codon 37, three TCT-to-TTT (S37F) changes and one TCT-to-TGT (S37C) change were found; and, lastly, one ACC-to-GCC change at codon 41 (T41A) was detected. Four of the 25 endometrioid ovarian carcinomas (16%) had an associated synchronous endometrial carcinoma. There was a higher percentage of beta-catenin mutations (n = 3, 75%) in synchronous ovarian carcinomas than in single ones, although with a similar percentage of MI (n = 1, 25%). beta-catenin mutations were S37C in two cases and D32G in one. One of the four endometrial carcinomas showed an S33C beta-catenin mutation, and two carcinomas had MI. None of the four tumors had both beta-catenin gene mutation and MI. beta-catenin gene mutations were always associated with a nuclear beta-catenin expression pattern, whereas MI was associated with a membranous pattern. In one patient both the ovarian and the endometrial carcinomas had beta-catenin gene mutations, in another patient both tumors showed MI, whereas in the remaining two patients the ovarian carcinomas showed beta-catenin gene mutations and the endometrial carcinomas showed MI. To summarize, the results of this study suggest that beta-catenin mutations and MI could represent two independent pathways in endometrioid ovarian carcinomas because they occur simultaneously very infrequently (in 5% of these cases). beta-catenin mutations are always associated with a nuclear beta-catenin expression pattern, whereas cases with a replication error -plus phenotype showed no abnormal beta-catenin subcellular localization. The study of the beta-catenin expression pattern, beta-catenin mutations, and MI, together with conventional clinicopathologic findings, could aid in distinguishing between the metastatic or independent origin of simultaneous endometrioid ovarian and endometrial carcinomas. Tumors with identical immunohistochemical and molecular features should therefore be considered to have a common origin.