Biomedical subjects
Publications and source records attributed to Ramon Parsons.
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In the last 5 years, global gene expression profiling has allowed for the subclassification of the heterogeneous disease of breast cancer into new subgroups with prognostic significance. However, for most subgroups, the nature of the contributions of individual genes to the clinical phenotypes remains largely unknown. In this issue of the JCI, Moyano and colleagues further examine the oncogenic potential of the small heat shock protein alpha-basic-crystallin, commonly expressed in tumors of the basal-like breast cancer subtype associated with poor prognosis, and show that it is an oncogenic protein in the breast.
PTEN, a tumor suppressor gene, is frequently mutated in a variety of human tumors. In mice, monoallelic inactivation of this gene predisposes animals to neoplasia of multiple organs. Interestingly, Pten heterozygous mice develop bilateral hyperplasia of the adrenal medulla. In this report we demonstrate that these neoplasms are hormonally active pheochromocytomas that secrete increased amounts of bioactive catecholamines: norepinephrine and epinephrine. To test a possibility that PTEN might be one of the genes responsible for human sporadic pheochromocytoma, we performed mutation analysis of DNA obtained from tumors of 29 patients. However, direct sequencing of all nine exons of the PTEN gene, including the splice junctions, revealed no mutations. Examination of protein expression by immunohistochemistry using 8 normal adrenals and 11 sporadic pheochromocytomas showed no decrease in the PTEN protein expression in the tumor tissue, but upregulation of insulin-like growth factor II, a peptide implicated in growth of adrenal tissue, was observed in four cases (36%).
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The HIN-1 gene encoding a small, secreted protein is silenced due to methylation in a substantial fraction of breast, prostate, lung, and pancreatic carcinomas, suggesting a potential tumor suppressor function. The receptor of HIN-1 is unknown, but ligand-binding studies indicate the presence of high-affinity cell surface HIN-1 binding on epithelial cells. Here, we report that HIN-1 is a potent inhibitor of anchorage-dependent and anchorage-independent cell growth, cell migration, and invasion. Expression of HIN-1 in synchronized cells inhibits cell cycle reentry and the phosphorylation of the retinoblastoma protein (Rb), whereas in exponentially growing cells, HIN-1 induces apoptosis without apparent cell cycle arrest and effect on Rb phosphorylation. Investigation of multiple signaling pathways revealed that mitogen-induced phosphorylation and activation of AKT are inhibited in HIN-1-expressing cells. In addition, expression of constitutively activate AKT abrogates HIN-1-mediated growth arrest. Taken together, these studies provide further evidence that HIN-1 possesses tumor suppressor functions, and that these activities may be mediated through the AKT signaling pathway.
Many cancers possess elevated levels of PtdIns(3,4,5)P(3), the second messenger that induces activation of the protein kinases PKB/Akt and S6K and thereby stimulates cell proliferation, growth, and survival. The importance of this pathway in tumorigenesis has been highlighted by the finding that PTEN, the lipid phosphatase that breaks down PtdIns(3,4,5)P(3) to PtdIns(4,5)P(2), is frequently mutated in human cancer. Cells lacking PTEN possess elevated levels of PtdIns(3,4,5)P(3), PKB, and S6K activity and heterozygous PTEN(+/-) mice develop a variety of tumors. Knockout of PKBalpha in PTEN-deficient cells reduces aggressive growth and promotes apoptosis, whereas treatment of PTEN(+/-) mice with rapamycin, an inhibitor of the activation of S6K, reduces neoplasia. We explored the importance of PDK1, the protein kinase that activates PKB and S6K, in mediating tumorigenesis caused by the deletion of PTEN. We demonstrate that reducing the expression of PDK1 in PTEN(+/-) mice, markedly protects these animals from developing a wide range of tumors. Our findings provide genetic evidence that PDK1 is a key effector in mediating neoplasia resulting from loss of PTEN and also validate PDK1 as a promising anticancer target for the prevention of tumors that possess elevated PKB and S6K activity.
The Irs2 branch of the insulin/insulin-like growth factor signaling cascade activates the phosphatidylinositol 3-kinase --> Akt --> Foxo1 cascade in many tissues, including hepatocytes and pancreatic beta-cells. The 3'-lipid phosphatase Pten ordinarily attenuates this cascade; however, its influence on beta-cell growth or function is unknown. To determine whether decreased Pten expression could restore beta-cell function and prevent diabetes in Irs2(-/-) mice, we generated wild type or Irs2 knock-out mice that were haploinsufficient for Pten (Irs2(-/-)::Pten(+/-)). Irs2(-/-) mice develop diabetes by 3 months of age as beta-cell mass declined progressively until insulin production was lost. Pten insufficiency increased peripheral insulin sensitivity in wild type and Irs2(-/-) mice and increased Akt and Foxo1 phosphorylation in the islets. Glucose tolerance improved in the Pten(+/-) mice, although beta-cell mass and circulating insulin levels decreased. Compared with Irs2(-/-) mice, the Irs2(-/-)::Pten(+/-) mice displayed nearly normal glucose tolerance and survived without diabetes, because normal but small islets produced sufficient insulin until the mice died of lymphoproliferative disease at 12 months age. Thus, steps to enhance phosphatidylinositol 3-kinase signaling can promote beta-cell growth, function, and survival without the Irs2 branch of the insulin/insulin-like growth factor signaling cascade.
CHK1 is an essential kinase involved in the regulation of the cell cycle progression and preservation of genomic integrity. Inhibition of CHK1 leads to the accumulation of double-stranded DNA breaks. Loss of PTEN impairs CHK1-mediated checkpoint activation due to cytoplasmic sequestration of ubiquitinated CHK1. Here, we provide evidence that another consequence of reduced CHK1 function in PTEN deficient cells is the accumulation of double-stranded DNA breaks. Moreover, we show that the site of CHK1 ubiquitination (K274) is near the site of AKT phosphorylation (S280). Overall, these data demonstrate that lack of PTEN generates DNA damage due to inappropriate inactivation of CHK1.DNA damage due to the loss of PTEN is likely to stimulate tumor development.
PBAF and BAF are two chromatin-remodeling complexes of the SWI/SNF family essential for mammalian transcription and development. Although these complexes share eight identical subunits, only PBAF can facilitate transcriptional activation by nuclear receptors in vitro. Here we show that these complexes have selectivity in mediating transcription of different interferon-responsive genes. The selectivity by PBAF requires a novel subunit, BAF200, but not the previously described PBAF-specificity subunit, BAF180 (Polybromo). Our study provides in vivo evidence that PBAF and BAF regulate expression of distinct genes, and suggests that BAF200 plays a key role in PBAF function.
To understand how a signaling molecule's activities are regulated, we need insight into the processes controlling the dynamic balance between its synthesis and degradation. For the Ins(1,3,4,5,6)P5 signal, this information is woefully inadequate. For example, the only known cytosolic enzyme with the capacity to degrade Ins(1,3,4,5,6)P5 is the tumour-suppressor PTEN [J.J. Caffrey, T. Darden, M.R. Wenk, S.B. Shears, FEBS Lett. 499 (2001) 6 ], but the biological relevance has been questioned by others [E.A. Orchiston, D. Bennett, N.R. Leslie, R.G. Clarke, L. Winward, C.P. Downes, S.T. Safrany, J. Biol. Chem. 279 (2004) 1116 ]. The current study emphasizes the role of physiological levels of PTEN in Ins(1,3,4,5,6)P5 homeostasis. We employed two cell models. First, we used a human U87MG glioblastoma PTEN-null cell line that hosts an ecdysone-inducible PTEN expression system. Second, the human H1299 bronchial cell line, in which PTEN is hypomorphic due to promoter methylation, has been stably transfected with physiologically relevant levels of PTEN. In both models, a novel consequence of PTEN expression was to increase Ins(1,3,4,5,6)P5 pool size by 30-40% (p<0.01); this response was wortmannin-insensitive and, therefore, independent of the PtdIns 3-kinase pathway. In U87MG cells, induction of the G129R catalytically inactive PTEN mutant did not affect Ins(1,3,4,5,6)P(5) levels. PTEN induction did not alter the expression of enzymes participating in Ins(1,3,4,5,6)P5 synthesis. Another effect of PTEN expression in U87MG cells was to decrease InsP6 levels by 13% (p<0.02). The InsP6-phosphatase, MIPP, may be responsible for the latter effect; we show that recombinant human MIPP dephosphorylates InsP6 to D/L-Ins(1,2,4,5,6)P5, levels of which increased 60% (p<0.05) following PTEN expression in U87MG cells. Overall, our data add higher inositol phosphates to the list of important cellular regulators [Y. Huang, R.P. Wernyj, D.D. Norton, P. Precht, M.C. Seminario, R.L. Wange, Oncogene, 24 (2005) 3819 ] the levels of which are modulated by expression of the highly pleiotropic PTEN protein.
BAF180 encoding a subunit of the human SWI/SNF chromatin remodeling complex maps to 3p21, in a region where frequent allele loss has been detected in lung cancer. BAF180 can be mutated and has tumor suppressing properties in breast cancer. In addition, another member of this complex, hSNF5/INI1, is a known tumor suppressor gene (TSG) for malignant rhabdoid and childhood central nervous system tumors. Thus, BAF180 is a strong candidate TSG for lung cancer. The objective of this study was to determine whether BAF180 mRNA or protein expression was inactivated or abnormal in lung cancers to prompt detailed DNA promoter methylation or mutational analyses. In 30 non-small-cell and 26 small-cell lung cancer cell lines, most of which had 3p21 allele loss, BAF180 mRNA and protein expression were evaluated by RT-PCR using three sets of primers and Western blotting using two anti-BAF180 antibodies. In all cases, BAF180 was expressed and no abnormal size BAF180 protein was detected. Finally, we found no amino-acid sequence coding mutations in five non-small-cell and five small-cell lung cancer cell lines, while we did find a new splicing isoform of BAF180 (AY281068). We conclude that abnormalities of BAF180 are not frequently involved in the pathogenesis of lung cancer.
Deregulation of the phosphatidylinositol 3-kinase (PI3K) pathway either through loss of PTEN or mutation of the catalytic subunit alpha of PI3K (PIK3CA) occurs frequently in human cancer. We identified PIK3CA mutations in 26% of 342 human breast tumor samples and cell lines at about equal frequency in tumor stages I to IV. To investigate the relationship between PTEN and PIK3CA, we generated a cohort of tumors that had lost PTEN expression and compared it with a matched control set that had retained PTEN. A highly significant association between PIK3CA mutations and retention of PTEN protein expression was observed. In addition, PIK3CA mutations were associated with expression of estrogen and progesterone receptors (ER/PR), lymph node metastasis, and ERBB2 overexpression. The fact that PIK3CA mutations and PTEN loss are nearly mutually exclusive implies that deregulated phosphatidylinositol-3,4,5-triphosphate (PIP(3)) is critical for tumorigenesis in a significant fraction of breast cancers and that loss of PIP(3) homeostasis by abrogation of either PIK3CA or PTEN relieves selective pressure for targeting of the other gene. The correlation of PIK3CA mutation to ER/PR-positive tumors and PTEN loss to ER/PR-negative tumors argues for disparate branches of tumor evolution. Furthermore, the association between ERBB2 overexpression and PIK3CA mutation implies that more than one input activating the PI3K/AKT pathway may be required to overcome intact PTEN. Thus, mutation of PIK3CA is frequent, occurs early in carcinoma development, and has prognostic and therapeutic implications.
Loss of chromosome arm 18q is a common event in human pancreatic, colon, and breast cancers and is often interpreted as representing loss of one or more tumor-suppressor genes. In this article, we describe two novel biallelic deletions at chromosome band 18q21.1 in a recently characterized human breast cancer cell line, HCC-1428. One lesion deletes a fragment of approximately 300 kb between SMAD4 and DCC that encodes no known genes. The second lesion is an in-frame SMAD4 deletion (amino acids 49-51) that affects the level of SMAD4 protein but not the SMAD4 message. This change accelerates 26S proteasome-mediated degradation of both endogenous and exogenous mutant SMAD4. Examination of normal DNA from the same patient demonstrated that both lesions are somatic and associated with loss of both normal alleles. These data support the concept that two independent tumor-suppressor loci exist at chromosome segment 18q21.1, one at SMAD4 and the other potentially at an enhancer of DCC or an unrelated novel gene.
Pten-/- cells display a partially defective checkpoint in response to ionizing radiation (IR). The checkpoint defect was traced to the ability of AKT to phosphorylate CHK1 at serine 280, since a nonphosphorylated mutant of CHK1 (S280A) complemented the checkpoint defect and restored CDC25A degradation. CHK1 phosphorylation at serine 280 led to covalent binding of 1 to 2 molecules of ubiquitin and cytoplasmic CHK1 localization. Primary breast carcinomas lacking PTEN expression and having elevated AKT phosphorylation had increased cytoplasmic CHK1 and displayed aneuploidy (p <0.005). We conclude that loss of PTEN and subsequent activation of AKT impair CHK1 through phosphorylation, ubiquitination, and reduced nuclear localization to promote genomic instability in tumor cells.
Despite expression of the high-affinity IL-2R, CD4(+)CD25(+) regulatory T cells (Tregs) are hypoproliferative upon IL-2R stimulation in vitro. However the mechanisms by which CD4(+)CD25(+) T cells respond to IL-2 signals are undefined. In this report, we examine the cellular and molecular responses of CD4(+)CD25(+) Tregs to IL-2. IL-2R stimulation results in a G(1) cell cycle arrest, cellular enlargement and increased cellular survival of CD4(+)CD25(+) T cells. We find a distinct pattern of IL-2R signaling in which the Janus kinase/STAT pathway remains intact, whereas IL-2 does not activate downstream targets of phosphatidylinositol 3-kinase. Negative regulation of phosphatidylinositol 3-kinase signaling and IL-2-mediated proliferation of CD4(+)CD25(+) T cells is inversely associated with expression of the phosphatase and tensin homologue deleted on chromosome 10, PTEN.
The PI-3 kinase pathway is a major driving force for human cancer. One common way of stimulating the PI-3 kinase pathway occurs through inactivation of the PTEN tumor suppressor. The mechanisms of PTEN inactivation include mutation, epigenetic silencing and post-translational modification. Improved insight into the regulation of PTEN is leading to a richer understanding of the contribution of PTEN and the PI-3 kinase pathway to human tumors. Understanding the pathology of PI-3 kinase signaling in tumors improves knowledge of cancer etiology and provides novel therapeutic targets.
Cancer is the second leading cause of death in New York City, with nearly 15,000 deaths each year. The urban setting of New York City provides ready access to large and diverse populations for whom racial/ethnic disparities in cancer risk and outcomes can be examined. A new cohort study was undertaken with several aims: (1) to provide a database and biorepository for studies of cancer etiology and pathogenesis, including host genetics; (2) to differentiate risk factors that contribute to racial/ethnic disparities in cancer risk, prevention, control, incidence, mortality, and survival; (3) to provide timely data on cancer risk and preventive behaviors that can be used to mobilize and then evaluate public health programs. Scientists from multiple institutions contributed to protocol design and implementation. Study instruments included demographics, personal and family history of cancer, risk and prevention efforts. End points include linkage with registries and medical record reviews. Using venue-based sampling with quotas, 18,187 adults aged 30 years or older were recruited over a year to undergo a baseline questionnaire, venipuncture, and contact information. The sample was 39% male, 37% older than 50 years, 58% white, 20% African American, 18% Hispanic, and 9% Asian. In terms of family history of cancer, 21% reported mother, 21% reported father, and 5.9% reported both parents with cancer; 8.5% reported any sibling with cancer. At baseline, 1,231 participants reported prior cancer. Showing the feasibility of constructing a cohort based in New York City, plans proceed for additional recruitment and analyses on the salient questions about cancer.
The PTEN tumour suppressor gene is mutated frequently in many malignancies and its importance in the development of cancer is probably underestimated. As the primary phosphatase of phosphatidylinositol (3,4,5)-trisphosphate, PTEN has a central role in reigning in the phosphoinositide 3-kinase (PI 3-kinase) network to control cellular homeostasis. Cells that lack PTEN are unable to regulate the PtdIns 3-kinase programme, which stimulates a variety of cellular phenotypes that favour oncogenesis. As well as the well-known role as tumour suppressor, recent studies show that PTEN is involved in the regulation of several basic cellular functions, such as cell migration, cell size, contractility of cardiac myocytes and chemotaxis. Here, we review the roles of PTEN in normal cellular functions and disease development.