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Biomedical subjects

I B Weinstein

Publications and source records attributed to I B Weinstein.

At least 19 recordsLinked to original sources
PubMed2006

Hint1 is a haplo-insufficient tumor suppressor in mice.

The HINT1 protein, a member of the histidine triad (HIT) family, is highly conserved in diverse species and ubiquitously expressed in mammalian tissues. However, its precise function in mammalian cells is not known. As a result of its structural similarity to the tumor-suppressor protein FHIT, we used homozygous-deleted Hint1 mice to study its role in tumorigenesis. We discovered that after 2 to 3 years of age the spontaneous tumor incidence in Hint1 -/- mice was significantly greater than that in wild-type Hint1 +/+ mice (P < 0.05). Using a well-established mouse model of 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary carcinogenesis we found a marked and significant (P < 0.05) increase in the incidence of mammary and ovarian tumors in both, Hint1 -/- and +/- mice versus +/+ mice. The Hint1 -/- and +/- mice had similar tumor incidence and similar tumor histologies. Therefore, deletion of Hint1 in mice enhances both spontaneous tumor development and susceptibility to tumor induction by DMBA. In addition, since the Hint1 +/- tumors retained expression of the unmutated wild-type allele, Hint1 is haplo-insufficient with respect to tumor suppression in this model system.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed2001

Effects of H. pylori infection of gastric epithelial cells on cell cycle control.

Chronic infection of the gastric mucosa by the bacterium H. pylori results in an intense inflammatory response which can last for decades. An associated host response is a chronic hyperproliferative state, in which there is increased cell turnover and also increased apoptosis of the gastric epithelial cells. Recent studies have also demonstrated abnormalities in the expression of cell cycle control proteins. This review describes these events, emphasizing recent studies on the effects of H. pylori infection on cell cycle progression and the expression of cell cycle regulatory proteins. The systems that have been studied include in vivo studies in humans and in experimental animals, and in vitro studies in which gastric epithelial cells were co-cultivated with H. pylori. The earliest event following H. pylori's interaction with epithelial cells appears to be growth inhibition and apoptosis. The hyperproliferative response observed in the gastric mucosa is secondary to this initial insult and is associated with increased expression of cyclin D1, the cyclin dependent kinase inhibitor p16ink4a and of p53 and decreased expression of the cyclin dependent kinase inhibitor p27kip1. Dysregulation of the hyperproliferative response may, ultimately, be responsible for the ability of H. pylori to enhance the development of gastric cancer.

Animals↗
PubMed2001

Overexpression of CDC25B overrides radiation-induced G2-M arrest and results in increased apoptosis in esophageal cancer cells.

CDC25B phosphatase plays a key role in controlling G2-M progression by dephosphorylating two inhibitory residues of CDC2 and also has been suggested to have an oncogenic property. In this study, we investigated the effect of CDC25B overexpression on radiation-induced G2-M arrest and radiation sensitivity in esophageal cancer cells. TE8-CDC25B, in which CDC25B was overexpressed under an inducible system, was more radiosensitive than the vector control (TE8-neo) in a clonogenic survival assay. Without radiation, CDC25B overexpression had little effect on cell cycle fractions or growth rate. After 10-Gy radiation, TE8-CDC25B showed decreased G2-M arrest and increased apoptosis, whereas TE8-neo displayed prolonged G2-M arrest and less apoptosis. During this period, there were no differences in the protein amounts of CDC2 and cyclin B1 between the two cell lines. However, more CDC25B expression, which was reduced immediately by radiation, was sustained in TE8-CDC25B than in TE8-neo. Moreover, induction of tyrosine phosphorylation of CDC2 and reduction of CDC2 kinase activity after irradiation was less significant in TE8-CDC25B than in TE8-neo. These results indicate that cancer cells that overexpress CDC25B override G2-M arrest by retaining CDC2 kinase activity and undergo apoptosis after radiation. This may point to an effective approach toward improving radiotherapy outcomes of various cancers.

Apoptosis↗
PubMed2001

Protein kinase G activates the JNK1 pathway via phosphorylation of MEKK1.

We recently obtained evidence that treatment of human colon cancer cells with exisulind (sulindac sulfone) and related compounds induces apoptosis by activation of protein kinase G (PKG) and c-Jun kinase (JNK1). The present study further explores this mechanism. We demonstrate that in NIH3T3 cells a constitutively active mutant of PKG causes a dose-dependent activation of JNK1 and thereby transactivates c-Jun and stimulates transcription from the AP-1 enhancer element. The activation of JNK1 and the transactivation of c-Jun by this mutant of PKG were inhibited by a dominant negative MEKK1. In vitro assays showed that a purified PKG directly phosphorylated the N-terminal domain of MEKK1. PKG also directly phosphorylated a full-length MEKK1, and this was associated with enhanced MEKK1 phosphorylation. Thus, it appears that PKG activates JNK1 through a novel PKG-MEKK1-SEK1-JNK1 pathway, by directly phosphorylating and activating MEKK1.

3T3 Cells↗
PubMed2001

Antiproliferative effects of S-allylmercaptocysteine on colon cancer cells when tested alone or in combination with sulindac sulfide.

Epidemiological studies link increased garlic (Allium sativum) consumption with a reduced incidence of colon cancer in various human populations. Experimental carcinogenesis studies in animal models and in cell culture systems indicate that several allium-derived compounds exhibit inhibitory effects and that the underlying mechanisms may involve both the initiation and promotion phases of carcinogenesis. To provide a better understanding of the effects of allium derivatives on the prevention of colon cancer, we examined two water-soluble derivatives of garlic, S-allylcysteine (SAC) and S-allylmercaptocysteine (SAMC), for their effects on proliferation and cell cycle progression in two human colon cancer cell lines, SW-480 and HT-29. For comparison, we included the compound sulindac sulfide (SS), because sulindac compounds are well-established colon cancer chemopreventive agents. We found that SAMC, but not SAC, inhibited the growth of both cell lines at doses similar to that of SS. SAMC also induced apoptosis, and this was associated with an increase in caspase3-like activity. These affects of SAMC were accompanied by induction of jun kinase activity and a marked increase in endogenous levels of reduced glutathione. Although SS caused inhibition of cell cycle progression from G1 to S, SAMC inhibited progression at G2-M, and a fraction of the SW-480 and HT-29 cells were specifically arrested in mitosis. Coadministration of SS with SAMC enhanced the growth inhibitory and apoptotic effects of SS. These findings suggest that SAMC may be useful in colon cancer prevention when used alone or in combination with SS or other chemopreventive agents.

Antineoplastic Agents↗
PubMed2001

Allelic exclusion and differentiation by protein kinase C-mediated signals in immature thymocytes.

Pre-T cell receptor (preTCR)-derived signals mediate the transition of thymocytes from the CD4(-) CD8(-) double-negative (DN) to CD4(+) CD8(+) double-positive stage of T lymphocyte development. This progression, termed beta-selection, is limited to thymocytes that have generated a functional TCR-beta chain able to associate with pTalpha to form the preTCR complex. Formation of the preTCR complex not only induces differentiation, survival, and proliferation of DN thymocytes; it also inhibits further TCR-beta gene rearrangement through an ill-defined process known as allelic exclusion. The signaling pathways controlling this critical developmental checkpoint have not been characterized. Here we demonstrate that formation of the preTCR complex leads to the activation of protein kinase C (PKC), and that activation of PKC is necessary for the differentiation and expansion of DN thymocytes. Importantly, we also show that allelic exclusion at the TCR-beta gene loci is enforced by PKC-mediated signals. These results define PKC as a central mediator of both differentiation and allelic exclusion during thymocyte development.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Safety and efficacy of exisulind for treatment of recurrent prostate cancer after radical prostatectomy.

PURPOSE: We evaluated the safety and efficacy of exisulind for delaying disease progression in men with increasing prostate specific antigen (PSA) after radical prostatectomy. MATERIALS AND METHODS: A total of 96 men with increasing PSA after radical prostatectomy were randomized to receive placebo (49) or 250 mg. exisulind twice daily (47) for 12 months. The primary efficacy parameter was the difference in change from baseline PSA between the placebo and exisulind groups. The PSA doubling time was also evaluated before and during study. A subgroup analysis classified patients based on the risk of developing metastatic disease. RESULTS: Compared with placebo, exisulind significantly suppressed the increase in PSA in all patients (p = 0.017). The results were also statistically significant in men at high risk for metastasis (p = 0.0003) and those who could not be classified according to risk (p = 0.0009). In addition, median PSA doubling time was lengthened in high risk patients on exisulind (2.12 month increase) compared with those on placebo (3.37 month decrease, p = 0.048). Exisulind was well tolerated. CONCLUSIONS: Exisulind inhibited the increase in PSA overall and prolonged PSA doubling time in high risk patients compared with placebo. These results suggest that Exisulind has the potential to extend the time from biochemical recurrence to the need for androgen deprivation therapy. Exisulind was well tolerated in this patient population. Our results support further study of Exisulind in the treatment of patients with prostate cancer.

Aged↗
PubMed2001

Effects of epigallocatechin-3-gallate on growth, epidermal growth factor receptor signaling pathways, gene expression, and chemosensitivity in human head and neck squamous cell carcinoma cell lines.

The antitumor effects of the green tea compound epigallocatechin-3-gallate (EGCG) have not been studied in detail previously in head and neck squamous cell carcinoma (HNSCC) cells. Overexpression of the epidermal growth factor receptor (EGFR) occurs frequently in HNSCC, which is an adverse prognostic factor. Therefore, we examined in detail the molecular effects of EGCG on two human HNSCC cell lines, YCU-N861 and YCU-H891, focusing on the EGFR signaling pathway. The 70% lethal dose (IC(70)) of EGCG for both cell lines was 10 microg/ml. Treatment with EGCG increased the proportion of cells in the G(1) phase of the cell cycle and induced apoptosis. In cells treated with EGCG, there was a decrease in the cyclin D1 protein, an increase in the p21(Cip1) and p27(Kip1) proteins, and a reduction in the hyperphosphorylated form of pRB, changes that may account for the arrest in G(1). EGCG also caused a decrease in the Bcl-2 and Bcl-X(L) proteins, an increase in the Bax protein, and activation of caspase 9, suggesting that EGCG induces apoptosis via a mitochondrial pathway. Treatment with EGCG inhibited phosphorylation of the EGFR, signal transducer and activator of transcription3 (Stat3), and extracellular regulated kinase (ERK) proteins and also inhibited basal and transforming growth factor-alpha-stimulated c-fos and cyclin D1 promoter activity. EGCG at 0.1 microg/ml (a concentration found in serum after oral administration) markedly enhanced the growth-inhibitory effects of 5-fluorouracil. Taken together, these findings provide insights into molecular mechanisms of growth inhibition by EGCG and suggest that this naturally occurring compound may be useful, when used alone or in combination with other agents, in the chemoprevention and/or treatment of HNSCC.

Antineoplastic Agents↗
PubMed2000

Cloning and characterization of DIP1, a novel protein that is related to the Id family of proteins.

Using human cyclin D1 as the "bait" in a yeast two-hybrid system, together with a HL60 cDNA library, we identified a novel human nuclear protein designated DIP1. This protein is expressed in a variety of cell types, and in fibroblasts its level remains constant throughout the cell cycle. However, the level of this protein increases severalfold during the differentiation of HL60 cells. The DIP1 protein can be phosphorylated in vitro by a cellular kinase and this activity reaches its maximum in extracts obtained from cells in the G1 phase of the cell cycle. DIP1 contains a helix-loop-helix motif but lacks an adjacent basic DNA-binding domain, thus resembling the Id family of proteins. The dip1 gene is located on human chromosome 16p11.2-12, a locus that is amplified in several types of human cancer. These results suggest that DIP1 may be involved in the control of gene expression and differentiation, but its precise function remains to be determined.

Amino Acid Sequence↗
PubMed2000

Herbimycin A induces G1 arrest through accumulation of p27(Kip1) in cyclin D1-overexpressing fibroblasts.

The ansamycin antibiotic herbimycin A is a potent tyrosine kinase inhibitor and reduces the growth rate of various types of mammalian cells. When quiescent Rat6 fibroblast cells were treated with herbimycin A, serum-induced expression of cyclin D1 was inhibited, and this was associated with inhibition of G1 phase progression. However, herbimycin A also inhibited serum-induced G1 progression in derivatives of the Rat6 fibroblast cell line that stably overexpress a human cyclin D1 cDNA (R6ccnD1#4 cells), without affecting the expression levels of G1 cyclins. We found that herbimycin A prevented serum-induced downregulation of the cyclin-dependent kinase inhibitor p27(Kip1), thereby leading to inactivation of the protein kinase activity of CDK2. These results suggest that herbimycin A inhibits a tyrosine kinase(s) that plays a role in degradation of the p27(Kop1) protein.

Animals↗
PubMed2000

Multiple functions of p27(Kip1) and its alterations in tumor cells: a review.

Cyclin-dependent kinases (CDKs), together with cyclins, their regulatory subunits, govern cell-cycle progression in eukaryotic cells. p27(Kip1) is a member of a family of CDK inhibitors (CDIs) that bind to cyclin/CDK complexes and arrest cell division. There is considerable evidence that p27(Kip1) plays an important role in multiple fundamental cellular processes, including cell proliferation, cell differentiation, and apoptosis. Moreover, p27(Kip1) is a putative tumor-suppressor gene that appears to play a critical role in the pathogenesis of several human malignancies and its reduced expression has been shown to correlate with poor prognosis in cancer patients. This study reviews current information on the functions of p27(Kip1), its abnormalities found in human tumors, and the possible clinical implications of these findings with respect to the management of cancer patients.

Animals↗
PubMed2000

Overexpression of protein kinase C-beta1 isoenzyme suppresses indomethacin-induced apoptosis in gastric epithelial cells.

BACKGROUND & AIMS: We have previously reported that nonsteroidal anti-inflammatory drugs (NSAIDs) could induce apoptosis of gastric epithelial cells both in vivo and in vitro. This study investigated the role of protein kinase C (PKC) isoforms in the regulation of NSAID-induced apoptosis. METHODS: Protein levels of 12 PKC isoforms in AGS cells, in the presence or absence of indomethacin, were determined by Western blot. The effect of PKC-beta1 overexpression by transfection with its complementary DNA (cDNA) on indomethacin-induced apoptosis and apoptosis-related genes, including p53, p21(waf1/cip1), and c-myc, was further investigated. RESULTS: Treatment with indomethacin decreased the abundance of PKC-beta1 and increased that of PKC-beta2, eta, and epsilon, but did not alter the expression of PKC alpha, gamma, zeta, delta, iota, and micro. Overexpression of PKC-beta1 attenuated the apoptotic response of AGS cells to indomethacin, associated with overexpression of p21(waf1/cip1) in both messenger RNA and protein levels. Inhibition of PKC-beta1-mediated overexpression of p21(waf1/cip1) by its antisense cDNA partially reduced the antiapoptotic effect of PKC-beta1. CONCLUSIONS: Indomethacin-induced apoptosis in gastric cancer cells is partly mediated by differential regulation of PKC isoform expression. Enhanced expression of exogenous PKC-beta1 protects against indomethacin-induced apoptosis through up-regulation of p21(waf1/cip1).

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed2000

Molecular epidemiology: recent advances and future directions.

In 1982 we proposed the concept and a framework for implementing molecular cancer epidemiology. Here, we review progress during the past 17 years in validating and applying this approach to cancer prevention. There have been major advances, notably in the understanding of environment-susceptibility interactions in human cancer. However, a review of major findings to date reveals several urgent research needs to keep pace with the rapid evolution in knowledge of mechanisms in carcinogenesis. Although much valuable progress continues to be made in the study of carcinogens that cause direct DNA damage and are mutagenic, exogenous and endogenous carcinogens can also act by altering gene expression, cell proliferation and differentiation. The mechanisms include aberrant DNA methylation, oxidative damage, effects on metabolism of nitrogen oxide and nitrites, activation of receptors and transcription factors, cyclins and other cell cycle proteins. Sensitive, validated biomarkers are needed to detect these mechanisms in small numbers of cells, tissues or fluids. There is also increasing recognition that individual risk from carcinogen exposure varies as a function of both inherited and acquired factors. Recent advances in genomics, microassay technologies and informatics hold promise for rapid identification of polymorphic variants or changes in expression of genes influencing both response and susceptibility to carcinogens. Another emerging area of molecular epidemiology concerns the role of nutrition and specific dietary factors (including studies on antioxidants, energy metabolism, insulin and various growth factors) and the modulating effect of genetic polymorphisms. Finally, molecular epidemiology has enormous potential in cancer prevention through the early identification of 'at risk' populations and the rapid assessment of intervention efficacy. Its success in fully reaching this potential will depend on the application of validated biomarkers, with adherence to sound epidemiologic and ethical principles.

Carcinogens↗
PubMed2000

Disorders in cell circuitry during multistage carcinogenesis: the role of homeostasis.

The multistage process of carcinogenesis involves the progressive acquisition of mutations, and epigenetic abnormalities in the expression, of multiple genes that have highly diverse functions. An important group of these genes are involved in cell cycle control. Thus, cyclin D1 is frequently overexpressed in a varety of human cancers. Cylin D1 plays a critical role in carcinogenesis because (i) overexpression enhances cell transformation and tumorigenesis, and enhances the amplification of other genes, and (ii) an antisense cyclin D1 cDNA reverts the malignant phenotype of carcinoma cells. Therefore, cyclin D1 may be a useful biomarker in molecular epidemiology studies, and inhibitors of its function may be useful in both cancer chemoprevention and therapy. We discovered a paradoxical increase in the cell cycle inhibitors protein p27(Kip1) in a subset of human cancers, and obtained evidence for homeostatic feedback loops between cyclins D1 or E and p27(Kip1). Furthermore, derivatives of HT29 colon cancer cells with increased levels of p27(Kip1) showed increased sensitivity to induction of differentiation. This may explain why decreased p27(Kip1) in a subset of human cancers is associated with a high grade (poorly differentiated) histology and poor prognosis. Agents that increase cellular levels of p27(Kip1) may, therefore, also be useful in cancer therapy. Using an antisense Rb oligonucleotide we obtained evidence that the paradoxical increase in pRb often seen in human colon cancers protects these cells from growth inhibition and apopotosis. On the basis of these, and other findings, we hypothesize that homeostatic feedback mechanisms play a critical role in multistage carcinogenesis. Furthermore, because of their bizarre circuitry, cancer cells suffer from 'gene addiction' and 'gene hypersensitivity' disorders that might be exploited in both cancer prevention and chemotherapy.

Cell Cycle↗
PubMed2000

Chronic Helicobacter pylori infection induces an apoptosis-resistant phenotype associated with decreased expression of p27(kip1).

Helicobacter pylori infection is associated with the development of gastric cancer. In short-term coculture with AGS gastric cells, H. pylori inhibits cell cycle progression and induces dose-dependent apoptosis. Based on the concept that an imbalance between proliferation and apoptosis may contribute to the emergence of gastric cancer, we chronically exposed AGS cells to H. pylori as a model of chronic exposure in humans. The AGS derivatives selected by this process were stably resistant not only to H. pylori-induced apoptosis but also to apoptosis induced by other enteric bacteria and by several toxic agents including radiation and cancer chemotherapy. Like the parental AGS cells, the derivatives underwent G(1)/S-phase cell cycle inhibition in response to H. pylori. The AGS derivatives displayed a marked decrease in cellular levels of the cell cycle control protein p27(kip1). We found a similar decrease in epithelial cell p27(kip1) expression in gastric biopsy specimens from H. pylori-infected patients. These findings are consistent with observations that link decreases in the p27(kip1) level to increased susceptibility to cancer in mice with p27(kip1) deleted and to a poor prognosis of gastric cancer in humans. This is the first demonstration that bacterial infection can lead to apoptosis resistance and to cross-resistance to other inducers of apoptosis such as bacteria, chemotherapeutic agents, and radiation. The development of apoptosis resistance and downmodulation of p27(kip1) may contribute to the increased risk for gastric cancer observed in humans chronically exposed to H. pylori.

Apoptosis↗
PubMed2000

Different protein kinase C isoforms determine growth factor specificity in neuronal cells.

Although mitogenic and differentiating factors often activate a number of common signaling pathways, the mechanisms leading to their distinct cellular outcomes have not been elucidated. In a previous report, we demonstrated that mitogen-activated protein (MAP) kinase (ERK) activation by the neurogenic agents fibroblast growth factor (FGF) and nerve growth factor is dependent on protein kinase Cdelta (PKCdelta), whereas MAP kinase activation in response to the mitogen epidermal growth factor (EGF) is independent of PKCdelta in rat hippocampal (H19-7) and pheochromocytoma (PC12) cells. We now show that EGF activates MAP kinase through a PKCzeta-dependent pathway involving phosphatidylinositol 3-kinase and PDK1 in H19-7 cells. PKCzeta, like PKCdelta, acts upstream of MEK, and PKCzeta can potentiate Raf-1 activation by EGF. Inhibition of PKCzeta also blocks EGF-induced DNA synthesis as monitored by bromodeoxyuridine incorporation in H19-7 cells. Finally, in embryonic rat brain hippocampal cell cultures, inhibitors of PKCzeta or PKCdelta suppress MAP kinase activation by EGF or FGF, respectively, indicating that these factors activate distinct signaling pathways in primary as well as immortalized neural cells. Taken together, these results implicate different PKC isoforms as determinants of growth factor signaling specificity within the same cell. Furthermore, these data provide a mechanism whereby different growth factors can differentially activate a common signaling intermediate and thereby generate biological diversity.

3-Phosphoinositide-Dependent Protein Kinases↗
PubMed2000

The protein kinase C beta-specific inhibitor LY379196 blocks TPA-induced monocytic differentiation of HL60 cells the protein kinase C beta-specific inhibitor LY379196 blocks TPA-induced monocytic differentiation of HL60 cells.

The ability of the promyelocytic leukemia HL60 cell line to differentiate in response to various stimuli has provided a widely used model of differentiation. The phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), acting via its cellular receptor protein kinase C(PKC), induces these cells to acquire a monocytic phenotype. We set out to identify the specific isoform of the multigene PKC family that is involved in this differentiation event. To do so, we utilized a highly specific PKCbeta inhibitor, LY379196. We found that LY379196 could prevent the growth arrest, cellular adherence, and changes in several marker proteins that occur after the addition of TPA to HL60 cells and that these effects were not simply due to nonspecific cytotoxicity. Thus, the present studies provide strong evidence that the beta isoform of PKC plays a critical role in TPA-induced HL60 monocytic differentiation.

Blotting, Western↗