Phosphoprotein pathway mapping: Akt/Mammalian target of rapamycin activation is negatively associated with childhood rhabdomyosarcoma survival

Mapping of protein signaling networks within tumors can identify new targets for therapy and provide a means to stratify patients for individualized therapy. Despite advances in combination chemotherapy, the overall survival for childhood rhabdomyosarcoma remains ∼60%. A critical goal is to identify functionally important protein signaling defects associated with treatment failure for the 40% nonresponder cohort. Here, we show, by phosphoproteomic network analysis of microdissected tumor cells, that interlinked components of the Akt/mammalian target of rapamycin (mTOR) pathway exhibited increased levels of phosphorylation for tumors of patients with short-term survival. Specimens (n = 59) were obtained from the Children's Oncology Group Intergroup Rhabdomyosarcoma Study (IRS) IV, D9502 and D9803, with 12-year follow-up. High phosphorylation levels were associated with poor overall and poor disease-free survival: Akt Ser473 (overall survival P < 0.001, recurrence-free survival P < 0.0009), 4EBP1 Thr37/46 (overall survival P < 0.0110, recurrence-free survival P < 0.0106), eIF4G Ser1108 (overall survival P < 0.0017, recurrence-free survival P < 0.0072), and p70S6 Thr389 (overall survival P < 0.0085, recurrence-free survival P < 0.0296). Moreover, the findings support an altered interrelationship between the insulin receptor substrate (IRS-1) and Akt/mTOR pathway proteins (P < 0.0027) for tumors from patients with poor survival. The functional significance of this pathway was tested using CCI-779 in a mouse xenograft model. CCI-779 suppressed phosphorylation of mTOR downstream proteins and greatly reduced the growth of two different rhabdomyosarcoma (RD embryonal P = 0.00008; Rh30 alveolar P = 0.0002) cell lines compared with controls. These results suggest that phosphoprotein mapping of the Akt/mTOR pathway should be studied further as a means to select patients to receive mTOR/IRS pathway inhibitors before administration of chemotherapy.

The influence of colour and sound on neuronal activation during visual object naming

This paper investigates how neuronal activation for naming photographs of objects is influenced by the addition of appropriate colour or sound. Behaviourally, both colour and sound are known to facilitate object recognition from visual form. However, previous functional imaging studies have shown inconsistent effects. For example, the addition of appropriate colour has been shown to reduce antero-medial temporal activation whereas the addition of sound has been shown to increase posterior superior temporal activation. Here we compared the effect of adding colour or sound cues in the same experiment. We found that the addition of either the appropriate colour or sound increased activation for naming photographs of objects in bilateral occipital regions and the right anterior fusiform. Moreover, the addition of colour reduced left antero-medial temporal activation but this effect was not observed for the addition of object sound. We propose that activation in bilateral occipital and right fusiform areas precedes the integration of visual form with either its colour or associated sound. In contrast, left antero-medial temporal activation is reduced because object recognition is facilitated after colour and form have been integrated.

Fusiform activation to animals is driven by the process, not the stimulus

Previous studies have found that the lateral posterior fusiform gyri respond more robustly to pictures of animals than pictures of manmade objects and suggested that these regions encode the visual properties characteristic of animals. We suggest that such effects actually reflect processing demands arising when items with similar representations must be finely discriminated. In a positron emission tomography (PET) study of category verification with colored photographs of animals and vehicles, there was robust animal-specific activation in the lateral posterior fusiform gyri when stimuli were categorized at an intermediate level of specificity (e.g., dog or car). However, when the same photographs were categorized at a more specific level (e.g., Labrador or BMW), these regions responded equally strongly to animals and vehicles. We conclude that the lateral posterior fusiform does not encode domain-specific representations of animals or visual properties characteristic of animals. Instead, these regions are strongly activated whenever an item must be discriminated from many close visual or semantic competitors. Apparent category effects arise because, at an intermediate level of specificity, animals have more visual and semantic competitors than do artifacts.

Modeling of protein signaling networks in clinical proteomics

Molecular interactions that underlie pathophysiological states are being elucidated using techniques that profile proteomicend points in cellular systems. Within the field of cancer research, protein interaction networks play pivotal roles in the establishment and maintenance of the hallmarks of malignancy, including cell division, invasion, and migration. Multiple complementary tools enable a multifaceted view of how signal protein pathway alterations contribute to pathophysiological states.One pivotal technique is signal pathway profiling of patient tissue specimens. This microanalysis technology provides a proteomic snapshot at one point in time of cells directly procured from the native context of a tumor micro environment. To study the adaptive patterns of signal pathway events over time, before and after experimental therapy, it is necessary to obtain biopsies from patients before, during, and after therapy. A complementary approach is the profiling of cultured cell lines with and without treatment. Cultured cell models provide the opportunity to study short-term signal changes occurring over minutes to hours. Through this type of system, the effects of particular pharmacological agents may be used to test the effects of signal pathway inhibition or activation on multiple endpoints within a pathway. The complexity of the data generated has necessitated the development of mathematical models for optimal interpretation of interrelated signaling pathways. In combination,clinical proteomic biopsy profiling, tissue culture proteomic profiling, and mathematical modeling synergistically enable a deeper understanding of how protein associations lead to disease states and present new insights into the design of therapeutic regimens.

The dynamics of cellular automata on 2-manifolds is affected by topology

In this paper, we demonstrate that the distribution of Wolfram classes within a cellular automata rule space in the triangular tessellation is not consistent across different topological general. Using a statistical mechanics approach, cellular automata dynamical classes were approximated for cellular automata defined on genus-0, genus-1 and genus-2 2-manifolds. A distribution-free equality test for empirical distributions was applied to identify cases in which Wolfram classes were distributed differently across topologies. This result implies that global structure and local dynamics contribute to the long term evolution of cellular automata.

On the effect of topology on cellular automata rule spaces

This thesis presents an empirical study of the effects of topology on cellular automata rule spaces. The classical definition of a cellular automaton is restricted to that of a regular lattice, often with periodic boundary conditions. This definition is extended to allow for arbitrary topologies. The dynamics of cellular automata within the triangular tessellation were analysed when transformed to 2-manifolds of topological genus 0, genus 1 and genus 2. Cellular automata dynamics were analysed from a statistical mechanics perspective. The sample sizes required to obtain accurate entropy calculations were determined by an entropy error analysis which observed the error in the computed entropy against increasing sample sizes. Each cellular automata rule space was sampled repeatedly and the selected cellular automata were simulated over many thousands of trials for each topology. This resulted in an entropy distribution for each rule space. The computed entropy distributions are indicative of the cellular automata dynamical class distribution. Through the comparison of these dynamical class distributions using the E-statistic, it was identified that such topological changes cause these distributions to alter. This is a significant result which implies that both global structure and local dynamics play a important role in defining long term behaviour of cellular automata.

Nilotinib and MEK inhibitors induce synthetic lethality through paradoxical activation of RAF in drug-resistant chronic myeloid leukemia

We show that imatinib, nilotinib, and dasatinib possess weak off-target activity against RAF and, therefore, drive paradoxical activation of BRAF and CRAF in a RAS-dependent manner. Critically, because RAS is activated by BCR-ABL, in drug-resistant chronic myeloid leukemia (CML) cells, RAS activity persists in the presence of these drugs, driving paradoxical activation of BRAF, CRAF, MEK, and ERK, and leading to an unexpected dependency on the pathway. Consequently, nilotinib synergizes with MEK inhibitors to kill drug-resistant CML cells and block tumor growth in mice. Thus, we show that imatinib, nilotinib, and dasatinib drive paradoxical RAF/MEK/ERK pathway activation and have uncovered a synthetic lethal interaction that can be used to kill drug-resistant CML cells in vitro and in vivo.

Activation of forkhead box O transcription factors by oncogenic BRAF promotes p21cip1-dependent senescence

Oncogene-induced senescence (OIS) is a potent tumor-suppressive mechanism that is thought to come at the cost of aging. The Forkhead box O (FOXO) transcription factors are regulators of life span and tumor suppression. However, whether and how FOXOs function in OIS have been unclear. Here, we show a role for FOXO4 in mediating senescence by the human BRAFV600E oncogene, which arises commonly in melanoma. BRAFV600E signaling through mitogen-activated protein kinase/extracellular signal-regulated kinase kinase resulted in increased reactive oxygen species levels and c-Jun NH 2 terminal kinase-mediated activation of FOXO4 via its phosphorylation on Thr223, Ser226, Thr447, and Thr451. BRAFV600E-induced FOXO4 phosphorylation resulted in p21cip1-mediated cell senescence independent of p16 ink4a or p27kip1. Importantly, melanocyte-specific activation of BRAFV600E in vivo resulted in the formation of skin nevi expressing Thr223/Ser226-phosphorylated FOXO4 and elevated p21cip1. Together, these findings support a model in which FOXOs mediate a trade-off between cancer and aging.

Activation of toxic chemicals by cytochrome P450 enzymes : regio- and stereoselective oxidation of aflatoxin B1

Cytochrome P450 (P450) enzymes are involved in the oxidations of numerous steroids, eicosanoids, alkaloids, and other endogenous substrates. These enzymes are also the major ones involved in the oxidation of potential toxicants and carcinogens such as those encountered among pollutants, solvents, and pesticides, as well as many natural products. A proper understanding of the basic mechanisms by which the P450 enzymes oxidize such compounds is important in developing rational strategies for the evaluation of the risks of these compounds.