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Erik Sahai

Publications and source records attributed to Erik Sahai.

17 recordsLinked to original sources
PubMed2026

Plasma signals of lung tumor promotion for molecular cancer prevention.

Predicting lung cancer risk would enhance prevention trials. Although the Canakinumab Anti-inflammatory Thrombosis Outcome Study (CANTOS) trial demonstrated reduced lung cancer incidence with interleukin (IL)-1β inhibition, the high number needed to treat (NNT) to prevent lung cancer limits its use in unselected populations. Using machine learning, we identified a 14-protein plasma signature predicting lung cancer more than 5 years before diagnosis. The signature, validated across eight cohorts, was elevated in current smokers and individuals exposed to particulate matter (PM) and linked to lung myeloid and alveolar cells. In epidermal growth factor receptor (EGFR)-driven lung adenocarcinoma, diverse epithelial lineages converged on a keratin8+/claudin4+ alveolar transitional state (KAC), whose transcriptional programs correlated with signature emergence. Components of the signature were induced by PM, oncogenic EGFR, or IL-1β, whereas IL-1β inhibition restrained PM-driven KAC expansion and early tumorigenesis. In CANTOS, the signature identified individuals who seemed to benefit more from anti-IL-1β therapy, lowering the NNT threshold and nominating circulating signals of tumor promotion for prevention.

Humans↗
PubMed2006

Smurf1 regulates tumor cell plasticity and motility through degradation of RhoA leading to localized inhibition of contractility.

Rho GTPases participate in various cellular processes, including normal and tumor cell migration. It has been reported that RhoA is targeted for degradation at the leading edge of migrating cells by the E3 ubiquitin ligase Smurf1, and that this is required for the formation of protrusions. We report that Smurf1-dependent RhoA degradation in tumor cells results in the down-regulation of Rho kinase (ROCK) activity and myosin light chain 2 (MLC2) phosphorylation at the cell periphery. The localized inhibition of contractile forces is necessary for the formation of lamellipodia and for tumor cell motility in 2D tissue culture assays. In 3D invasion assays, and in in vivo tumor cell migration, the inhibition of Smurf1 induces a mesenchymal-amoeboid-like transition that is associated with a more invasive phenotype. Our results suggest that Smurf1 is a pivotal regulator of tumor cell movement through its regulation of RhoA signaling.

Animals↗
PubMed2006

ROCK- and myosin-dependent matrix deformation enables protease-independent tumor-cell invasion in vivo.

Tumor cells invading three-dimensional matrices need to remodel the extracellular matrix (ECM) in their path. Many studies have focused on the role of extracellular proteases; however, cells with amoeboid or rounded morphologies are able to invade even when these enzymes are inhibited. Here, we describe the mechanism by which cells move through a dense ECM without proteolysis. Amoeboid tumor cells generate sufficient actomyosin force to deform collagen fibers and are able to push through the ECM. Force generation is elevated in metastatic MTLn3E cells, and this correlates with increased invasion and altered myosin light chain (MLC) organization. In metastatic cells, MLC is organized perpendicularly to the direction of movement behind the invading edge. Both the organization of MLC and force generation are dependent upon ROCK function. We demonstrate that ROCK regulates the phosphorylation of MLC just behind the invading margin of the cell. Imaging of live tumors shows that MLC is organized in a similar ROCK-dependent fashion in vivo and that inhibition of ROCK but not matrix-metalloproteases reduces cancer cell motility in vivo.

Animals↗
PubMed2006

Epidermal growth factor receptor overexpression results in increased tumor cell motility in vivo coordinately with enhanced intravasation and metastasis.

Although overexpression of the epidermal growth factor receptor (EGFR; ErbB1) has been correlated with poor prognosis in breast and other cancers, clinical trials of ErbB1 inhibitors have shown limited efficacy in inhibiting tumor proliferation. To evaluate other possible roles of ErbB1 in tumor malignancy besides proliferation, we have developed a series of tools for analysis of intravasation. Overexpression of ErbB1 in MTLn3 mammary adenocarcinoma cells results in increased intravasation and lung metastasis from tumors formed by injection of cells in the mammary fat pad. However, increased ErbB1 expression has no effect on primary tumor growth and lung seeding efficiency of cells injected i.v. Chemotactic responses to low concentrations of EGF in vitro and cell motility in vivo in the primary tumor measured using intravital imaging are significantly increased by ErbB1 overexpression. The increased cell motility is restricted to ErbB1-overexpressing cells in tumors containing mixtures of cells expressing different ErbB1 levels, arguing for a cell-autonomous effect of increased ErbB1 expression rather than alteration of the tumor microenvironment. In summary, we propose that ErbB1 overexpression makes more significant contributions to intravasation than growth in some tumors and present a novel model for studying ErbB1 contributions to tumor metastasis via chemotaxis and intravasation.

Adenocarcinoma↗
PubMed2006

Purification of TAT-C3 exoenzyme.

The Clostridium botulinum C3 exoenzyme has been an invaluable tool for the study of the biological functions of Rho GTPases. The C3 enzyme selectively catalyzes the ADP-ribosylation, and consequent inactivation, of RhoA, RhoB, and RhoC of the Rho GTPase protein family. Through the experimental use of C3, it has been possible to determine the contributions made by these signaling proteins to processes including the regulation of cell morphology, cell cycle progression, and gene transcription. Unlike bacterial toxins that have some means to attach to and/or enter cells, C3 does not have an element that facilitates efficient entry. As a result, numerous methods have been used to effectively deliver C3 into cells. One approach has been to engineer a recombinant C3 with an HIV TAT leader sequence that permits transduction of the protein across the plasma membrane. In this chapter, the purification and characterization of the recombinant TAT-C3 protein is described.

ADP Ribose Transferases↗
PubMed2006

Tumor cell migration in three dimensions.

In almost all physiological and pathological situations, cells migrate through three-dimensional environments, yet most studies of cell motility have used two-dimensional substrates. It is clear that two-dimensional substrates do not mimic the in vivo environment accurately, and recent work using three-dimensional environments has revealed many different mechanisms of cell migration (Abbott, 2003; Sahai and Marshall, 2003; Wolf et al., 2003). This chapter will describe methods for generating three-dimensional matrices suitable for studying cell motility, methods for imaging the morphology of motile cells in situ, and methods for quantifying cell migration through three-dimensional environments.

Amides↗
PubMed2005

Macrophages promote the invasion of breast carcinoma cells via a colony-stimulating factor-1/epidermal growth factor paracrine loop.

Previous studies have shown that macrophages and tumor cells are comigratory in mammary tumors and that these cell types are mutually dependent for invasion. Here we show that macrophages and tumor cells are necessary and sufficient for comigration and invasion into collagen I and that this process involves a paracrine loop. Macrophages express epidermal growth factor (EGF), which promotes the formation of elongated protrusions and cell invasion by carcinoma cells. Colony stimulating factor 1 (CSF-1) produced by carcinoma cells promotes the expression of EGF by macrophages. In addition, EGF promotes the expression of CSF-1 by carcinoma cells thereby generating a positive feedback loop. Disruption of this loop by blockade of either EGF receptor or CSF-1 receptor signaling is sufficient to inhibit both macrophage and tumor cell migration and invasion.

Animals↗
PubMed2005

Simultaneous imaging of GFP, CFP and collagen in tumors in vivo using multiphoton microscopy.

BACKGROUND: The development of multiphoton laser scanning microscopy has greatly facilitated the imaging of living tissues. However, the use of genetically encoded fluorescent proteins to distinguish different cell types in living animals has not been described at single cell resolution using multiphoton microscopy. RESULTS: Here we describe a method for the simultaneous imaging, by multiphoton microscopy, of Green Fluorescent Protein, Cyan Fluorescent Protein and collagen in vivo in living tumors. This novel method enables: 1) the simultaneous visualization of overall cell shape and sub-cellular structures such as the plasma membrane or proteins of interest in cells inside living animals, 2) direct comparison of the behavior of single cells from different cell lines in the same microenvironment in vivo. CONCLUSION: Using this multi-fluor, multiphoton technique, we demonstrate that motility and metastatic differences between carcinoma cells of differing metastatic potential can be imaged in the same animal simultaneously at sub-cellular resolution.

Animals↗
PubMed2005

Mechanisms of cancer cell invasion.

The movement of cancer cells into tissue surrounding the tumour and the vasculature is the first step in the spread of metastatic cancers. Recent advances in imaging, the use of 3D model systems and the application of microarray technologies have yielded new insights into these processes. This work has challenged our views about what causes cancer cells to become motile in the first place, and has demonstrated that cancer cells can move in many different ways.

Animals↗
PubMed2005

Tumor cells caught in the act of invading: their strategy for enhanced cell motility.

Invasion of neighboring extracellular matrix tissue, the lymphatic system and blood vessels is a key element of tumor cell metastasis in many epithelial tumors. Understanding the cell motility pathways that contribute to invasion can provide new approaches and targets for anticancer therapy. The recent convergence of technologies for expression profiling and intravital imaging has revealed the identities of some of the genes that contribute to motility and chemotaxis of cancer cells in tumors. In particular, the genes encoding a minimum motility machine are coordinately upregulated in tumor cells collected by an in vivo invasion assay. These results support a "tumor microenvironment invasion model" and provide new target opportunities for cancer therapy.

Animals↗
PubMed2004

Conditional ROCK activation in vivo induces tumor cell dissemination and angiogenesis.

Progression of tumors to invasive and metastatic forms requires that tumor cells undergo dramatic morphologic changes, a process regulated by Rho GTPases. Elevated expression of RhoA and RhoC, as well as the Rho effector proteins ROCK I and ROCK II, are commonly observed in human cancers and are often associated with more invasive and metastatic phenotypes. To examine how ROCK contributes to the progression of solid tumors, we established a conditionally activated form of ROCK II by fusing the kinase domain to the estrogen receptor hormone-binding domain (ROCK:ER). ROCK:ER-expressing colon carcinoma cells grown as tumors in immunocompromised nude mice organized into discrete clusters surrounding blood vessels. However, ROCK:ER activation resulted in the aggressive dissemination of tumor cells into the surrounding stroma, indicating that increased ROCK signaling is sufficient to promote invasion from solid tumors. In addition, tumors in which ROCK:ER was activated were more highly vascularized, indicating that ROCK contributes to tumor angiogenesis. ROCK:ER activation resulted in changes to epithelial morphology and organization that facilitated motility in vitro, likely by inducing the redistribution of proteins such as ezrin, as well as adherens junction and extracellular matrix-binding proteins. These results suggest that ROCK inhibitors would be useful antimetastatic and antiangiogenic chemotherapeutic agents in tumors associated with elevated RhoA, RhoC, ROCK I, or ROCK II expression.

Animals↗
PubMed2004

Identification and testing of a gene expression signature of invasive carcinoma cells within primary mammary tumors.

We subjected cells collected using an in vivo invasion assay to cDNA microarray analysis to identify the gene expression profile of invasive carcinoma cells in primary mammary tumors. Expression of genes involved in cell division, survival, and cell motility were most dramatically changed in invasive cells indicating a population that is neither dividing nor apoptotic but intensely motile. In particular, the genes coding for the minimum motility machine that regulates beta-actin polymerization at the leading edge and, therefore, the motility and chemotaxis of carcinoma cells, were dramatically up-regulated. However, ZBP1, which restricts the localization of beta-actin, the substrate for the minimum motility machine, was down-regulated. This pattern of expression implicated ZBP1 as a suppressor of invasion. Reexpression of ZBP1 in metastatic cells with otherwise low levels of ZBP1 reestablished normal patterns of beta-actin mRNA targeting and suppressed chemotaxis and invasion in primary tumors. ZBP1 reexpression also inhibited metastasis from tumors. These experiments support the involvement in metastasis of the pathways identified in invasive cells, which are regulated by ZBP1.

Actins↗
PubMed2003

ERK-MAPK signaling coordinately regulates activity of Rac1 and RhoA for tumor cell motility.

We describe two signaling events downstream of ERK-MAP kinase contributing to cell motility in colon carcinoma cells. The Fos family member Fra-1 is expressed in an ERK-dependent manner. Silencing of Fra-1 expression with short interfering RNAs leads to losses of cell polarization, motility, and invasiveness in vitro. These effects of ablating Fra-1 are a consequence of activation of a RhoA-ROCK pathway by beta1-integrin, leading to an increase in the amount of stress fibers and stabilization of focal adhesions. We propose that Fra-1 promotes cell motility by inactivating beta1-integrin and keeping RhoA activity low. This depression of RhoA activity is necessary to permit a second ERK-dependent signaling event via uPAR, the receptor for urokinase-type plasminogen activator, to activate Rac and to drive motility through polarized lamellipodia extension.

Cell Movement↗
PubMed2003

Differing modes of tumour cell invasion have distinct requirements for Rho/ROCK signalling and extracellular proteolysis.

Rho family GTPases regulate the cytoskeleton and cell migration and are frequently overexpressed in tumours. Here, we identify two modes of tumour-cell motility in 3D matrices that involve different usage of Rho signalling. Rho signalling through ROCK promotes a rounded bleb-associated mode of motility that does not require pericellular proteolysis. This form of motility requires ezrin, which is localized in the direction of cell movement. In contrast, elongated cell motility is associated with Rac-dependent F-actin-rich protrusions and does not require Rho, ROCK or ezrin function. Combined blockade of extracellular proteases and ROCK negates the ability of tumour cells to switch between modes of motility and synergises to prevent tumour cell invasion.

Cell Culture Techniques↗
PubMed2002

ROCK and Dia have opposing effects on adherens junctions downstream of Rho.

Adherens junctions (AJs) are crucial for maintaining the integrity of epithelial tissues and are often disrupted during tumour progression. Rho family proteins have been shown to regulate adherens junctions. We find that activation of the effector kinase ROCK and acto-myosin contraction disrupts AJs downstream of Rho. In contrast, signalling through the Rho effector Dia1 is required to ensure a dynamically stable interface between cells and the maintenance of adherens junction complexes. The ability of Dia1 to regulate the actin network is crucial for the localization of adherens junction components to the cell periphery.

Actins↗
PubMed2002

RHO-GTPases and cancer.

The RAS oncogenes were identified almost 20 years ago. Since then, we have learnt that they are members of a large family of small GTPases that bind GTP and hydrolyse it to GDP. This is then exchanged for GTP and the cycle is repeated. The switching between these two states regulates a wide range of cellular processes. A branch of the RAS family--the RHO proteins--is also involved in cancer, but what is the role of these proteins and would they make good therapeutic targets?

Animals↗