Tumor metastasis: molecular insights and evolving paradigms.

Metastases represent the end products of a multistep cell-biological process termed the invasion-metastasis cascade, which involves dissemination of cancer cells to anatomically distant organ sites and their subsequent adaptation to foreign tissue microenvironments. Each of these events is driven by the acquisition of genetic and/or epigenetic alterations within tumor cells and the co-option of nonneoplastic stromal cells, which together endow incipient metastatic cells with traits needed to generate macroscopic metastases. Recent advances provide provocative insights into these cell-biological and molecular changes, which have implications regarding the steps of the invasion-metastasis cascade that appear amenable to therapeutic targeting.

A pleiotropically acting microRNA, miR-31, inhibits breast cancer metastasis.

MicroRNAs are well suited to regulate tumor metastasis because of their capacity to coordinately repress numerous target genes, thereby potentially enabling their intervention at multiple steps of the invasion-metastasis cascade. We identify a microRNA exemplifying these attributes, miR-31, whose expression correlates inversely with metastasis in human breast cancer patients. Overexpression of miR-31 in otherwise-aggressive breast tumor cells suppresses metastasis. We deploy a stable microRNA sponge strategy to inhibit miR-31 in vivo; this allows otherwise-nonaggressive breast cancer cells to metastasize. These phenotypes do not involve confounding influences on primary tumor development and are specifically attributable to miR-31-mediated inhibition of several steps of metastasis, including local invasion, extravasation or initial survival at a distant site, and metastatic colonization. Such pleiotropy is achieved via coordinate repression of a cohort of metastasis-promoting genes, including RhoA. Indeed, RhoA re-expression partially reverses miR-31-imposed metastasis suppression. These findings indicate that miR-31 uses multiple mechanisms to oppose metastasis.

Activation of miR-31 function in already-established metastases elicits metastatic regression.

Distant metastases, rather than the primary tumors from which these lesions arise, are responsible for >90% of carcinoma-associated mortality. Many patients already harbor disseminated tumor cells in their bloodstream, bone marrow, and distant organs when they initially present with cancer. Hence, truly effective anti-metastatic therapeutics must impair the proliferation and survival of already-established metastases. Here, we assess the therapeutic potential of acutely expressing the microRNA miR-31 in already-formed breast cancer metastases. Activation of miR-31 in established metastases elicits metastatic regression and prolongs survival. Remarkably, even brief induction of miR-31 in macroscopic pulmonary metastases diminishes metastatic burden. In contrast, acute miR-31 expression fails to affect primary mammary tumor growth. miR-31 triggers metastatic regression in the lungs by eliciting cell cycle arrest and apoptosis; these responses occur specifically in metastases and can be explained by miR-31-mediated suppression of integrin-α5, radixin, and RhoA. Indeed, concomitant re-expression of these three proteins renders already-seeded pulmonary metastases refractory to miR-31-conferred regression. Upon miR-31 activation, Akt-dependent signaling is attenuated and the proapoptotic molecule Bim is induced; these effects occur in a metastasis-specific manner in pulmonary lesions and are abrogated by concurrent re-expression of integrin-α5, radixin, and RhoA. Collectively, these findings raise the possibility that intervention strategies centered on restoring miR-31 function may prove clinically useful for combating metastatic disease.

Concomitant suppression of three target genes can explain the impact of a microRNA on metastasis.

It remains unclear whether a microRNA (miRNA) affects a given phenotype via concomitant down-regulation of its entire repertoire of targets or instead by suppression of only a modest subset of effectors. We demonstrate that inhibition of breast cancer metastasis by miR-31-a miRNA predicted to modulate >200 mRNAs-can be entirely explained by miR-31's pleiotropic regulation of three targets. Thus, concurrent re-expression of integrin-alpha5, radixin, and RhoA abrogates miR-31-imposed metastasis suppression. These effectors influence distinct steps of the metastatic process. Our findings have implications concerning the importance of pleiotropy for the biological actions of miRNAs and provide mechanistic insights into metastasis.

Roles for microRNAs in the regulation of cell adhesion molecules.

Maintenance of appropriate cell adhesion is crucial for normal cellular and organismal homeostasis. Certain microRNAs have recently been found capable of regulating molecules that oversee the fundamental cell biological events that drive cellular adhesion. It is now apparent that microRNAs play crucial roles in the great majority of biochemical pathways that contribute to normal cell adhesion. In this Commentary, we describe the latest advances within this still-emerging field, and highlight connections between the deregulation of microRNAs that affect cell-adhesion-associated molecules and the pathogenesis of several human diseases. Current evidence suggests that the ability of certain microRNAs--notably miR-17, miR-29, miR-31, miR-124 and miR-200--to pleiotropically regulate multiple molecular components of the cell adhesion machinery endows these microRNAs with the capacity to function as key modulators of adhesion-associated processes. This, in turn, holds important implications for our understanding of both the basic biology of cell adhesion and the etiology of multiple pathological conditions.

Roles of microRNAs and other non-coding RNAs in breast cancer metastasis.

Despite the fact that metastases are responsible for the overwhelming majority of human cancer deaths, our comprehension of the molecular events that drive metastatic progression remains woefully incomplete. Excitingly, the recent appreciation that various species of non-coding RNAs­including microRNAs­play pivotal roles in dictating the malignant behaviors of breast carcinoma cells promises to afford new insights into the molecular circuitry that determines metastatic propensity. Here, I summarize our current knowledge regarding these still-emerging functions for non-coding RNAs in the pathogenesis of breast cancer metastasis, with an emphasis placed upon the roles played by microRNAs in these processes. Additionally, I discuss the potential translational opportunities afforded by these research findings for the diagnosis and treatment of human breast tumors. When assessed collectively, it is apparent that although this field of research is still in its infancy, comprehension of the biological actions of microRNAs and other non-coding RNAs will hold important consequences for our understanding of the etiology of metastatic disease, as well as its clinical management and treatment.

Assaying microRNA loss-of-function phenotypes in mammalian cells: emerging tools and their potential therapeutic utility.

MicroRNAs are small, non-coding RNAs that are increasingly appreciated to play critical roles in the modulation of gene expression. In mammalian cells, our knowledge regarding the full impact of microRNAs on cellular behavior remains fragmentary. This has been due, in significant part, to the limited availability of experimental tools for studying microRNA loss-of-function phenotypes. Recently, several strategies for achieving this goal have been developed. Here, we discuss these methodologies for inhibiting specific microRNAs in mammalian cells both in vitro and in vivo, compare and contrast the strengths and weaknesses of these approaches, and speculate regarding the future impact of these antagonists on the treatment of human diseases such as cancer. These emerging techniques enable the attenuation of microRNA function in a manner that is quite sequence-specific, relatively long-lasting and increasingly cost-effective. As such, some of these advances hold great promise in terms of their eventual utility as therapeutic agents.

Endogenous anticancer mechanisms: metastasis.

Metastases, rather than the primary tumors from which these malignant growths are spawned, are culpable for greater than 90 % of human cancer-associated mortality. Metastases arise through the completion of a series of cell-biological events - collectively termed "the invasion-metastasis cascade" - which involve the dissemination of tumor cells to distant organ sites and their subsequent adaptation to these foreign microenvironments. Importantly, a number of endogenous mechanisms exist that serve to prevent metastatic progression. These safeguards must be overcome by incipient metastatic tumor cells in order for them to generate detectable metastases. Here, I highlight four endogenous mechanisms that protect against the development of metastatic disease in breast carcinomas. I discuss how the expression of these genes are dampened during malignant progression, the downstream responses they orchestrate, and clinical opportunities to therapeutically target these mechanisms. Indeed, one potentially effective strategy for the remediation of metastatic disease involves the reactivation of endogenous anti-metastasis mechanisms. Therefore, knowledge regarding endogenous anti-metastasis mechanisms may both further our comprehension of the basic etiology of metastasis and also guide the treatment of human tumors.

Metastasis suppression: a role of the Dice(r).

Recent studies have implicated the microRNA biogenesis enzyme Dicer as a suppressor of breast carcinoma metastasis and elucidated upstream signaling pathways that control Dicer levels.

miR-31: a crucial overseer of tumor metastasis and other emerging roles.

MicroRNAs constitute a family of pleiotropically acting short regulatory RNAs. Increasingly, specific microRNAs have been implicated as key modulators of a variety of normal physiologic processes; moreover, the aberrant activity of certain microRNAs has been linked to the pathogenesis of multiple diseases. The microRNA miR-31 has been identified as a crucial overseer of several normal and diseased phenotypes. Here, we describe current knowledge regarding the functions of miR-31, with an emphasis placed upon the role of this microRNA in neoplastic development and tumor metastasis. Additionally, we highlight a number of recent reports concerning the contributions of miR-31 to other pathological states, the role of this microRNA in normal physiology, and the upstream mechanisms by which miR-31 expression levels are regulated. Assessed collectively, existing evidence suggests that miR-31 concomitantly regulates a number of essential signaling pathways in mammalian cells. For these reasons, further elucidation of the biological actions of miR-31 may prove significant for the prognosis and remediation of various pathological states.

Concurrent suppression of integrin alpha5, radixin, and RhoA phenocopies the effects of miR-31 on metastasis.

miR-31 inhibits breast cancer metastasis via the pleiotropic suppression of a cohort of prometastatic target genes that include integrin alpha(5) (ITGA5), radixin (RDX), and RhoA. We previously showed that the concomitant overexpression of ITGA5, RDX, and RhoA was capable of overriding the antimetastatic effects of ectopically expressed miR-31 in vivo. However, these prior studies failed to investigate whether the combined suppression of the endogenous mRNAs encoding these three proteins recapitulated the in vivo consequences of miR-31 expression on metastasis. We show here that short hairpin RNA-mediated concurrent downregulation of ITGA5, RDX, and RhoA is sufficient to phenocopy the full spectrum of described influences of miR-31 on metastasis in vivo, including the effects of this microRNA (miRNA) on local invasion, early post-intravasation events, and metastatic colonization. These findings provide mechanistic insights into the metastatic process and have implications about the importance of pleiotropy for the biological actions of miRNAs.

miR-9, a MYC/MYCN-activated microRNA, regulates E-cadherin and cancer metastasis.

MicroRNAs (miRNAs) are increasingly implicated in regulating the malignant progression of cancer. Here we show that miR-9, which is upregulated in breast cancer cells, directly targets CDH1, the E-cadherin-encoding messenger RNA, leading to increased cell motility and invasiveness. miR-9-mediated E-cadherin downregulation results in the activation of beta-catenin signalling, which contributes to upregulated expression of the gene encoding vascular endothelial growth factor (VEGF); this leads, in turn, to increased tumour angiogenesis. Overexpression of miR-9 in otherwise non-metastatic breast tumour cells enables these cells to form pulmonary micrometastases in mice. Conversely, inhibiting miR-9 by using a 'miRNA sponge' in highly malignant cells inhibits metastasis formation. Expression of miR-9 is activated by MYC and MYCN, both of which directly bind to the mir-9-3 locus. Significantly, in human cancers, miR-9 levels correlate with MYCN amplification, tumour grade and metastatic status. These findings uncover a regulatory and signalling pathway involving a metastasis-promoting miRNA that is predicted to directly target expression of the key metastasis-suppressing protein E-cadherin.

MicroRNAs: Crucial multi-tasking components in the complex circuitry of tumor metastasis.

Distant metastases are the underlying cause of patient mortality in an overwhelming majority of human carcinomas. Certain microRNAs have recently been found capable of regulating the process of tumor metastasis. In this review, we highlight advances within this rapidly emerging field, endeavor to connect known microRNA pathways with recent conceptual advances in the larger field of metastasis research, and speculate regarding the future utility of microRNAs in the diagnosis and treatment of human cancers. Assessed collectively, current evidence suggests that the pleiotropic activities of microRNAs endow them with the capacity to function as crucial, yet previously unappreciated, nodes within already-identified metastasis regulatory circuitry. This has important implications for our understanding of the pathogenesis of high-grade malignancies.

Novel transcriptional activities of vitamin E: inhibition of cholesterol biosynthesis.

Vitamin E is a dietary lipid that is essential for vertebrate health and fertility. The biological activity of vitamin E is thought to reflect its ability to quench oxygen- and carbon-based free radicals and thus to protect the organism from oxidative damage. However, recent reports suggest that vitamin E may also display other biological activities. Here, to examine possible mechanisms that may underlie such nonclassical activities of vitamin E, we investigated the possibility that it functions as a specific modulator of gene expression. We show that treatment of cultured hepatocytes with (RRR)-alpha-tocopherol alters the expression of multiple genes and that these effects are distinct from those elicited by another antioxidant. Genes modulated by vitamin E include those that encode key enzymes in the cholesterol biosynthetic pathway. Correspondingly, vitamin E caused a pronounced inhibition of de novo cholesterol biosynthesis. The transcriptional activities of vitamin E were mediated by attenuating the post-translational processing of the transcription factor SREBP-2 that, in turn, led to a decreased transcriptional activity of sterol-responsive elements in the promoters of target genes. These observations indicate that vitamin E possesses novel transcriptional activities that affect fundamental biological processes. Cross talk between tocopherol levels and cholesterol status may be an important facet of the biological activities of vitamin E.