Primary Cutaneous Blastoid Mantle Cell Lymphoma-Case Report

Mantle cell lymphoma (MCL) commonly involves extranodal sites, usually as a manifestation of disseminated disease. In rare cases, MCLs may arise as a primary tumor in the skin. Blastoid mantle cell lymphoma (BV-MCL) is a rare variant and has a more aggressive clinical course. The phenotype of BV-MCL is characterized as CD20(+), CD5(+), cyclin D1(+), CD23(-), and CD10(-). Interphase fluorescence in situ hybridization shows a characteristic t(11; 14) fusion pattern. We report a case of a BV-MCL arising in skin as primary cutaneous MCL with the characteristic immunophenotype and translocation.

Mucoepidermoid Carcinoma of the Lung Arising at the Primary Site of A Bronchogenic Cyst: Clinical, Cytogenetic, and Molecular Findings

Primary lung tumors are rare in children, and mucoepidermoid carcinoma (MEC) represents less than 10% of them. Additionally, MEC arising from bronchogenic cysts (BC) is particularly unusual. We describe the clinical and genetic findings on a MEC occurring within a previous location of a BC in an adolescent. This particular association has not been previously reported. The lesion revealed normal karyotype without the typical t(11;19)(q21;p13) translocation. Cyclin D1 overexpression (165-fold increase) was demonstrated by real-time PCR although FISH assessment showed normal hybridization at 11q13. Information on these unusual clinical presentations may present relevant insight on tumorigenesis of infrequent pediatric pulmonary tumors. Pediatr Blood Cancer 2011;56:311-313. (C) 2010 Wiley-Liss, Inc.

Transcriptional changes in the nuc-2A mutant strain of Neurospora crassa cultivated under conditions of phosphate shortage

The molecular mechanism that controls the response to phosphate shortage in Neurospora crassa involves four regulatory genes - nuc-2, preg, pgov, and nuc-1. Phosphate shortage is sensed by the nuc-2 gene, the product of which inhibits the functioning of the PREG-PGOV complex. This allows the translocation of the transcriptional factor NUC-1 into the nucleus, which activates the transcription of phosphate-repressible phosphatases. The nuc-2A mutant strain of N. crassa carries a loss-of-function mutation in the nuc-2 gene, which encodes an ankyrin-like repeat protein. In this study, we identified transcripts that are downregutated in the nuc-2A mutant strain. Functional grouping of these expressed sequence tags allowed the identification of genes that play essential roles in different cellular processes such as transport, transcriptional regulation, signal transduction, metabolism, protein synthesis, protein fate, and development. These results reveal novel aspects of the phosphorus-sensing network in N. crassa. (C) 2009 Elsevier GmbH. All rights reserved.

Long-Term Ethanol Consumption Promotes Hepatic Tumorigenesis but Impairs Normal Hepatocyte Proliferation in Rats

Chronic and excessive alcohol consumption has been related to an increased risk of several cancers, including that of the liver; however, studies in animal models have yet to conclusively determine whether ethanol acts as a tumor promoter in hepatic tumorigenesis. We examined whether prolonged alcohol consumption could act as a hepatic tumor promoter after initiation by diethylnitrosamine (DEN) in a rat model. Male Sprague-Dawley rats were injected with 20 mg DEN/kg body weight 1 wk before introduction of either an ethanol liquid diet or an isoenergic control liquid diet. Hepatic pathological lesions, hepatocyte proliferation, apoptosis, PPAR alpha and PPAR gamma, and plasma insulin-like growth factor 1 IGF-1) levels were assessed after 6 and 10 mo. Mean body and liver weights, plasma IGF-1 concentration, hepatic expressions of proliferating cellular nuclear antigen and Ki-67, and cyclin D1 in ethanol-fed rats were all significantly lower after 10 mo of treatment compared with control rats. In addition, levels of hepatic PPAR gamma protein, not PPAR alpha, were significantly higher in the ethanol-fed rats after prolonged treatment. Although ethanol feeding also resulted in significantly fewer altered hepatic foci, hepatocellular adenoma was detected in ethanol-fed rats at 10 mo, but not in control rats given the same dose of DEN. Together, these results indicate that chronic, excessive ethanol consumption impairs normal hepatocyte proliferation, which is associated with reduced IGF-1 levels, but promotes hepatic carcinogenesis. J. Nutr. 141: 1049-1055, 2011.

High doses of dexamethasone induce increased beta-cell proliferation in pancreatic rat islets

Rafacho A, Cestari TM, Taboga SR, Boschero AC, Bosqueiro JR. High doses of dexamethasone induce increased beta-cell proliferation in pancreatic rat islets. Am J Physiol Endocrinol Metab 296: E681-E689, 2009. First published January 21, 2009; doi:10.1152/ajpendo.90931.2008.-Activation of insulin signaling and cell cycle intermediates is required for adult beta-cell proliferation. Here, we report a model to study beta-cell proliferation in living rats by administering three different doses of dexamethasone (0.1, 0.5, and 1.0 mg/kg ip, DEX 0.1, DEX 0.5, and DEX 1.0, respectively) for 5 days. Insulin sensitivity, insulin secretion, and histomorphometric data were investigated. Western blotting was used to analyze the levels of proteins related to the control of beta-cell growth. DEX 1.0 rats, which present moderate hyperglycemia and marked hyperinsulinemia, exhibited a 5.1-fold increase in beta-cell proliferation and an increase (17%) in beta-cell size, with significant increase in beta-cell mass, compared with control rats. The hyperinsulinemic but euglycemic DEX 0.5 rats also showed a significant 3.6-fold increase in beta-cell proliferation. However, DEX 0.1 rats, which exhibited the lowest degree of insulin resistance, compensate for insulin demand by improving only islet function. Activation of the insulin receptor substrate 2/phosphatidylinositol 3-kinase/serine-threoninekinase/ribosomalprotein S6 kinase pathway, as well as protein retinoblastoma in islets from DEX 1.0 and DEX 0.5, but not in DEX 0.1, rats was also observed. Therefore, increasing doses of dexamethasone induce three different degrees of insulin requirement in living rats, serving as a model to investigate compensatory beta-cell alterations. Augmented beta-cell mass involves beta-cell hyperplasia and, to a lower extent, beta-cell hypertrophy. We suggest that alterations in circulating insulin and, to a lesser extent, glucose levels could be the major stimuli for beta-cell proliferation in the dexamethasone-induced insulin resistance.

Proliferation, apoptosis, and survival in high-level microsatellite instability sporadic colorectal cancer

Sporadic colorectal cancer (CRC) characterized by high-level DNA microsatellite instability (MSI-H) has a favorable prognosis. The reason for this MSI-H survival advantage is not known. The aim of this study was to correlate proliferation, apoptosis, and prognosis in CRC stratified by MSI status. The proliferative index (PI) was measured by immunohistochemical staining with the Ki-67 antibody in a selected series of 100 sporadic colorectal cancers classified according to the level of MSI as 31 MSI-H, 29 MSI-Low (MSI-L), and 40 microsatellite stable (MISS). The Ki-67 index was significantly higher in MSI-H cancers (P < 0.0001) in which the PI was 90.1 1.2% (mean +/- SE) compared with 69.5 +/- 3.1 % and 69.5 +/- 2.3 % in MSI-L and MSS subgroups, respectively. There was a positive linear correlation between the apoptotic index (AI) and PI (r = 0.51; P < 0.001), with MSI-H cancers demonstrating an increased AI:PI ratio indicative of a lower index of cell production. A high PI showed a trend toward predicting improved survival within MSI-H cancers (P = 0.09) but did not predict survival in MSI-L or MSS cancers. The Al was not associated with survival in any MSI subgroup. In conclusion, this is the first study to show that sporadic MSI-H cancers are characterized by a higher AL:PI ratio and increased proliferative activity compared with MSI-L and MSS cancers, and that an elevated PI may confer a survival advantage within the MSI-H subset.

Cdc25B activity is regulated by 14-3-3

In the G2 phase cell cycle checkpoint arrest, the cdc25-dependent activation of cyclin B/cdc2, a critical step in regulating entry into mitosis, is blocked. Studies in yeast have demonstrated that the inhibition of cdc25 function involves 14-3-3 binding to cdc25, In humans, two cdc25 isoforms have roles in G2/M progression, cdc25B and cdc25C, both bind 14-3-3, Abrogating 14-3-3 binding to cdc25C attenuates the G2 checkpoint arrest, but the contribution of 14-3-3 binding to the regulation of cdc25B function is unknown. Here we demonstrate that high level over-expression of cdc25B in G2 checkpoint arrested cells can activate cyclin B/cdc2 and overcome the checkpoint arrest. Mutation of the major 14-3-3 binding site, S323, or removal of the N-terminal regulatory domain are strong activating mutations, increasing the efficiency with which the mutant forms of cdc25B not only overcome the arrest, but also initiate aberrant mitosis, We also demonstrate that 14-3-3 binding to the S323 site on cdc25B blocks access of the substrate cyclin/cdks to the catalytic site of the enzyme, thereby directly inhibiting the activity of cdc25B, This provides direct mechanistic evidence that 14-3-3 binding to cdc25B can regulate its activity, thereby controlling progression into mitosis.

Up-regulation of p21(WAF1)/(CIP1) by histone deacetylase inhibitors reduces their cytotoxicity

Histone deacetylase inhibitors show promise as chemotherapeutic agents and have been demonstrated to block proliferation in a wide range of tumor cell lines. Much of this antiproliferative effect has been ascribed to the up-regulated expression of the cyclin-dependent kinase inhibitor p21(WAF1/CIP1). In this article, we report that p21 expression was up-regulated by relatively low doses of the histone deacetylase inhibitor azelaic bishydroxamic acid (ABHA) and correlated with a proliferative arrest. Higher doses of ABHA were cytotoxic. Cells that did not up-regulate p21 expression were hypersensitive to killing by ABHA and died via apoptosis, whereas up-regulation of p21 correlated with reduced sensitivity and a block in the apoptotic mechanism, and these cells seemed to die by necrosis. Using isogenic p21(+/+) and p21(-/-) cell lines and direct inhibition of caspase activity, we demonstrate that the reduced sensitivity to killing by ABHA is a consequence of inhibition of apoptosis by up-regulated p21 expression. These data indicate the enormous potential of therapeutic strategies that bypass the cytoprotective effect of p21 and act on the same molecular targets as the histone deacetylase inhibitors.

MC1R genotype modifies risk of melanoma in families segregating CDKN2A mutations

Mutations in the exons of the cyclin-dependent kinase inhibitor gene CDKN2A are melanoma-predisposition alleles which have high penetrance, although they have low population frequencies. In contrast, variants of the melanocortin-1 receptor gene, MC1R, confer much lower melanoma risk but are common in European populations. Fifteen Australian CDKN2A mutation-carrying melanoma pedigrees were assessed for MC1R genotype, to test for possible modifier effects on melanoma risk. A CDKN2A mutation in the presence of a homozygous consensus MC1R genotype had a raw penetrance of 50%, with a mean age at onset of 58.1 years. When an MC1R variant allele was also present, the raw penetrance of the CDKN2A mutation increased to 84%, with a mean age at onset of 37.8 years (P=0.1). The presence of a CDKN2A mutation gave a hazard ratio of 13.35, and the hazard ratio of 3.72 for MC1R variant alleles was also significant. The impact of MC1R variants on risk of melanoma was mediated largely through the action of three common alleles, Arg151Cys, Arg160Trp, and Asp294His, that have previously been associated with red hair, fair skin, and skin sensitivity to ultraviolet light.

No evidence of a role for activating CDK2 mutations in melanoma

Inactivation of p16(INK4a) and/or activation of cyclin-dependent kinase-4 (CDK4) are strongly associated with both susceptibility and progression in melanoma. Activating CDK4 mutations prevent the binding and inhibition of CDK4 by p16(INK4a). A second, more indirect role for CDK4 is in late G(1), where It may sequester the inhibitors p27(KIP1) or p21(CIP1) away from CDK2, and in doing so upregulate the CDK2 activity necessary for cells to proceed completely through G(1) into S phase. As the pivotal residues around the most predominant R24C activating CDK4 mutation are invariant between CDK2 and CDK4, we speculated that the pivotal arginine (position 22 in CDK2), or a nearby residue, may be mutated in some melanomas, resulting in the diminution of its binding and inhibition by p27(KIP1) or p21(CIP1). However, except for a silent polymorphism, we detected no variants within this region of the CDK2 gene in 60 melanoma cell lines. Thus, if CDK2 activity is dysregulated in melanoma it is likely to occur by a means other than mutations causing loss of direct inhibition. We also examined the expression of the CDK2 gene in melanoma cell lines, to assess its possible co-regulation with the gene for the melanocyte-lineage antigen pmel17, which maps less than 1 kb away in head to head orientation with CDK2 and may be transcribed off the same bidirectional promoter. However, expression of the genes is not co-regulated. (C) 2001 Lippincott Williams & Wilkins.

Proteomic analysis reveals that 14-3-3 sigma in downregulated in human breast cancer cells

The class of molecular chaperones known as 14-3-3 is involved in the control of cellular growth by virtue of its apparent regulation of various signaling pathways, including the Raf/mitogen-activated protein kinase pathway. In breast cancer cells, the sigma form of 14-3-3 has been shown to interact with cyclin-dependent kinases and to control the rate of entry into mitosis. To test for a direct role for 14-3-3 in breast epithelial cell neoplasia, me have quantitated 14-3-3 protein levels using a proteomic approach based on two-dimensional electrophoresis and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF). We show here that 14-3-3 sigma protein is strongly down-regulated in the prototypic breast cancer cell lines MCF-7 and MDA-MB-231 and in primary breast carcinomas as compared with normal breast epithelial cells. In contrast, levels of the alpha, beta, delta, or zeta isoforms of 14-3-3 mere the same in both normal and transformed cells. The data support the idea that 14-3-3 sigma is involved in the neoplastic transition of breast epithelial cells by virtue of its role as a tumor suppressor; as such, it may constitute a robust marker with clinical efficacy for this pathology.

Mechanism of mitosis-specific activation of MEK1

Activation of cyclin B-Cdc2 is an absolute requirement for entry into mitosis, but other protein kinase pathways that also have mitotic functions are activated during G(2)/M progression. The MAPK cascade has well established roles in entry and exit from mitosis in Xenopus, but relatively little is known about the regulation and function of this pathway in mammalian mitosis. Here we report a detailed analysis of the activity of all components of the Ras/Raf/MEK/ERK pathway in HeLa cells during normal G(2)/M. The focus of this pathway is the dramatic activation of an endomembrane-associated MEK1 without the corresponding activation of the MEK substrate ERK. This is because of the uncoupling of MEK1 activation from ERK activation. The mechanism of this uncoupling involves the cyclin B-Cdc2-dependent proteolytic cleavage of the N-terminal ERK-binding domain of MEK1 and the phosphorylation of Thr(286). These results demonstrate that cyclin B-Cdc2 activity regulates signaling through the MAPK pathway in mitosis.

eMelanoBase: An online locus-specific variant database for familial melanoma

A proportion of melanoma,prone individuals in both familial and non,familial contexts has been shown to carry inactivating mutations in either CDKN2A or, rarely, CDK4. CDKN2A is a complex locus that encodes two unrelated proteins from alternately spliced transcripts that are read in different frames. The alpha transcript (exons 1a, 2, and 3) produces the p16INK4A cyclin-dependent kinase inhibitor, while the beta transcript (exons 1beta and 2) is translated as p14ARF, a stabilizing factor of p53 levels through binding to MDM2. Mutations in exon 2 can impair both polypeptides and insertions and deletions in exons 1alpha, 1beta, and 2, which can theoretically generate p16INK4A,p14ARF fusion proteins. No online database currently takes into account all the consequences of these genotypes, a situation compounded by some problematic previous annotations of CDKN2A related sequences and descriptions of their mutations. As an initiative of the international Melanoma Genetics Consortium, we have therefore established a database of germline variants observed in all loci implicated in familial melanoma susceptibility. Such a comprehensive, publicly accessible database is an essential foundation for research on melanoma susceptibility and its clinical application. Our database serves two types of data as defined by HUGO. The core dataset includes the nucleotide variants on the genomic and transcript levels, amino acid variants, and citation. The ancillary dataset includes keyword description of events at the transcription and translation levels and epidemiological data. The application that handles users' queries was designed in the model,view. controller architecture and was implemented in Java. The object-relational database schema was deduced using functional dependency analysis. We hereby present our first functional prototype of eMelanoBase. The service is accessible via the URL www.wmi.usyd.e, du.au:8080/melanoma.html.

14-3-3 acts as an intramolecular bridge to regulate cdc25B localization and activity

One of the major regulators of mitosis in somatic cells is cdc25B. cdc25B is tightly regulated at multiple levels. The final activation step involves the regulated binding of 14-3-3 proteins. Previous studies have demonstrated that Ser-323 is a primary 14-3-3 binding site in cdc25B, which influences its activity and cellular localization. 14-3-3 binding to this site appeared to interact with the N-terminal domain of cdc25B to regulate its activity. The presence of consensus 14-3-3 binding sites in the N-terminal domain suggested that the interaction is through direct binding of the 14-3-3 dimer to sites in the N-terminal domain. We have identified Ser-151 and Ser-230 in the N-terminal domain as functional 14-3-3 binding sites utilized by cdc25B in vivo. These low affinity sites cooperate to bind the 14-3-3 dimer bound to the high affinity Ser-323 site, thus forming an intramolecular bridge that constrains cdc25B structure to prevent access of the catalytic site. Loss of 14-3-3 binding to either N-terminal site relaxes cdc25B structure sufficiently to permit access to the catalytic site, and the nuclear export sequence located in the N-terminal domain. Mutation of the Ser-323 site was functionally equivalent to the mutation of all three sites, resulting in the complete loss of 14-3-3 binding, increased access of the catalytic site, and access to nuclear localization sequence.

Differential expression of CDC25 phosphatases splice variants in human breast cancer cells

Background: CDC25 phosphatases control cell cycle progression by activating cyclin dependent kinases. The three CDC25 isoforms encoding genes are submitted to alternative splicing events which generate at least two variants for CDC25A and five for both CDC25B and CDC25C. An over-expression of CDC25 was reported in several types of cancer, including breast cancer, and is often associated with a poor prognosis. Nevertheless, most of the previous studies did not address the expression of CDC25 splice variants. Here, we evaluated CDC25 spliced transcripts expression in anti-cancerous drug-sensitive and resistant breast cancer cell lines in order to identify potential breast cancer biomarkers. Methods: CDC25 splice variants mRNA levels were evaluated by semi-quantitative RT-PCR and by an original real-time RT-PCR assay. Results: CDC25 spliced transcripts are differentially expres-sed in the breast cancer cell lines studied. An up-regulation of CDC25A2 variant and an increase of the CDC25C5/C1 ratio are associated to the multidrug-resistance in VCREMS and DOXOR breast cancer cells, compared to their sensitive counterpart cell line MCF-7. Additionally, CDC25B2 tran-script is exclusively over-expressed in VCREMS resistant cells and could therefore be involved in the development of certain type of drug resistance. Conclusions: CDC25 splice variants could represent interesting potential breast cancer prognostic biomarkers.