Non-small cell lung cancer (NSCLC) has a 5-y survival rate of ∼16%, with most deaths associated with uncontrolled metastasis. We screened for stem cell identity-related genes preferentially expressed in a panel of cell lines with high versus low metastatic potential, derived from NSCLC tumors of Kras LA1/+ ;P53 R172HΔG/+ (KP) mice. The Musashi-2 (MSI2) protein, a regulator of mRNA translation, was consistently elevated in metastasis-competent cell lines. MSI2 was overexpressed in 123 human NSCLC tumor specimens versus normal lung, whereas higher expression was associated with disease progression in an independent set of matched normal/primary tumor/lymph node specimens. Depletion of MSI2 in multiple independent metastatic murine and human NSCLC cell lines reduced invasion and metastatic potential, independent of an effect on proliferation. MSI2 depletion significantly induced expression of proteins associated with epithelial identity, including tight junction proteins [claudin 3 (CLDN3), claudin 5 (CLDN5), and claudin 7 (CLDN7)] and down-regulated direct translational targets associated with epithelial-mesenchymal transition, including the TGF-β receptor 1 (TGFβR1), the small mothers against decapentaplegic homolog 3 (SMAD3), and the zinc finger proteins SNAI1 (SNAIL) and SNAI2 (SLUG). Overexpression of TGFβRI reversed the loss of invasion associated with MSI2 depletion, whereas overexpression of CLDN7 inhibited MSI2-dependent invasion. Unexpectedly, MSI2 depletion reduced E-cadherin expression, reflecting a mixed epithelialmesenchymal phenotype. Based on this work, we propose that MSI2 provides essential support for TGFβR1/SMAD3 signaling and contributes to invasive adenocarcinoma of the lung and may serve as a predictive biomarker of NSCLC aggressiveness.on-small cell lung cancer (NSCLC) is the leading cause of cancer-related deaths in the world (1). Approximately 7% of individuals born in the United States in 2013 will ultimately be diagnosed with lung cancer, and 160,000 die from this disease each year (1). The 5-y survival rate for lung cancer is around 16% of diagnosed cases (2). Much of the lethality of lung cancer is due to frequent diagnosis of the malignancy at the metastatic stage, when fundamental changes in tumor biology cause the disease to be refractory to many treatments. A better understanding of the biological processes that promote NSCLC metastasis promises to further improve clinical care of the patients. Kras LA1/+ ;P53 R172HΔG/+ (KP) mice provide a useful and wellvalidated model for the study of NSCLC metastasis. These mice combine a mutant p53 allele (p53 R175HΔG) with an activating KrasG12D allele (Kras LA1 ) (3), leading to development of adenocarcinomas resembling human NSCLC, which are often characterized by mutation of KRAS (∼30%) (4) and loss of TP53 (∼60%) (5). Many of the KP tumors metastasize to sites commonly seen in NSCLC patients (3). These features make the KP murine model a useful tool with which to evaluate factors that underlie NSCLC metastasis. Among the pathways activated...

BackgroundmTORC1 (mammalian target of rapamycin complex 1) activation has been demonstrated in response to endotoxin challenge, but the mechanism and significance are unclear. We investigated the effect of mTORC1 suppression in an animal model of endotoxemia and in a cellular model of endotoxin signaling.Methodology/Principal FindingsMice were treated with the mTORC1 inhibitor rapamycin or vehicle prior to lethal endotoxin challenge. Mortality and cytokine levels were assessed. Cultured macrophage-like cells were challenged with endotoxin with or without inhibitors of various pathways known to be upstream of mTORC1. Activated pathways, including downstream S6K pathway, were assessed by immunoblots. We found that mTORC1-S6K suppression by rapamycin delayed mortality of mice challenged with lethal endotoxin, and was associated with dampened circulating levels of VEGF, IL-1β, IFN-γ and IL-5. Furthermore, in vitro cellular studies demonstrated that LPS (lipopolysaccharide) activation of mTORC1-S6K still occurs in the presence of PI3K-Akt inhibition alone, but can be suppressed by concurrent inhibition of PI3K-Akt and MEK-ERK pathways.Conclusions/SignificanceWe conclude that cellular activation of mTORC1-S6K contributes to cytokine up-regulation and mortality in response to endotoxin, and may occur via multiple pathways.

Lymphangioleiomyomatosis (LAM) is a female-predominant cystic lung disease that can lead to respiratory failure. LAM cells typically have inactivating tuberous sclerosis complex 2 (TSC2) mutations and mammalian target of rapamycin (mTOR) complex (mTORC) 1 activation. Clinical response to the mTORC1 inhibitors has been limited, prompting a search for additional therapy for LAM. In this study, we investigated the impact of TSC2 on the expression of poly (ADP-ribose) polymerase (PARP)-1 that initiates the DNA repair pathway, and tested the efficacy of PARP1 inhibitors in the survival of TSC2-deficient (TSC2(-)) cells. We analyzed publicly available expression arrays of TSC2(-) cells and validated the findings using real-time RT-PCR, immunoblotting, and immunohistochemistry. We examined the impact of rapamycin and Torin 1 on PARP1 expression. We also tested the effect of PARP1 inhibitors, 8-hydroxy-2-methylquinazoline-4-one and 3,4-dihydro-5[4-(1-piperindinyl)butoxy]-1(2H)-isoquinoline, on the survival of TSC2(-) cells. We identified the up-regulation of PARP1 in TSC2(-) cells relative to cells in which wild-type TSC2 has been reintroduced (TSC2-addback [TSC2(+)] cells). The transcript levels of PARP1 in TSC2(-) cells were not affected by rapamycin. PARP1 levels were increased in TSC2(-) cells, xenograft tumors of rat-derived TSC2(-) cells, renal cystadenomas from Tsc2(+/-) mice, and human LAM nodules. RNA interference of mTOR failed to reduce PARP1 levels. Proliferation and survival of TSC2(-) cells was reduced in response to PARP1 inhibitor treatment, more so than TSC2(+) cells. TSC2(-) cells exhibit higher levels of PARP1 relative to TSC2(+) cells in an mTOR-insensitive manner. PARP1 inhibitors selectively suppress the growth and induce apoptosis of TSC2(-) cells from patients with LAM. Targeting PARP1 may be beneficial in the treatment of LAM and other neoplasm with mTORC1 activation.

Factor X deficiency is a rare bleeding disorder that can be associated with life-threatening bleeding events. Factor X deficiency can either be inherited or acquired. Acquired cases of factor X deficiency can be seen in patients with plasma cell dyscrasias as well as amyloidosis. Coagulopathy, with clinically relevant bleeding events, although rare, is not an unusual phenomenon for patients with systemic amyloidosis. However, clinically relevant bleeding in patients with symptomatic multiple myeloma, without associated amyloidosis, has not been reported in literature before. We present a rare case of multiple myeloma without concomitant amyloidosis that presented with life-threatening bleeding from acquired deficiency of factor X and responded remarkably to treatment for underlying multiple myeloma. This case not only highlights the diagnostic workup required in patients with factor X deficiency but also provides the principles of management of acquired coagulopathy in plasma cell dyscrasias.

We present a Trefftz-type finite element method on meshes consisting of curvilinear polygons. Local basis functions are computed using integral equation techniques that allow for the efficient and accurate evaluation of quantities needed in the formation of local stiffness matrices. To define our local finite element spaces in the presence of curved edges, we must also properly define what it means for a function defined on a curved edge to be "polynomial" of a given degree on that edge. We consider two natural choices, before settling on the one that yields the inclusion of complete polynomial spaces in our local finite element spaces, and discuss how to work with these edge polynomial spaces in practice. An interpolation operator is introduced for the resulting finite elements, and we prove that it provides optimal order convergence for interpolation error under reasonable assumptions. We provide a description of the integral equation approach used for the examples in this paper, which was recently developed precisely with these applications in mind. A few numerical examples illustrate this optimal order convergence of the finite element solution on some families meshes in which every element has at least one curved edge. We also demonstrate that it is possible to exploit the approximation power of locally singular functions that may exist in our finite element spaces in order to achieve optimal order convergence without the typical adaptive refinement toward singular points.