Karen L Schmeichel

Modeling tissue-specific signaling and organ function in three dimensions

In order to translate the findings from basic cellular research into clinical applications, cell-based models need to recapitulate both the 3D organization and multicellular complexity of an organ but at the same time accommodate systematic experimental intervention. Here we describe a hierarchy of tractable 3D models that range in complexity from organotypic 3D cultures (both monotypic and multicellular) to animal-based recombinations in vivo. Implementation of these physiologically relevant models, illustrated here in the context of human epithelial tissues, has enabled the study of intrinsic cell regulation pathways and also has provided compelling evidence for the role of the stromal compartment in directing epithelial cell function and dysfunction. Furthermore the experimental accessibility afforded by these tissue-specific 3D models has implications for the design and development of cancer therapies.

Structural cues from the tissue microenvironment are essential determinants of the human mammary epithelial cell phenotype

Historically, the study of normal human breastfunction and breast disorders has been significantlyimpaired by limitations inherent to available modelsystems. Recent improvements in human breast epithelial cell lines and three-dimensional(3-D)3 culture systems have contributed tothe development of in vitro model systems thatrecapitulate differentiated epithelial cell phenotypeswith remarkable fidelity. Molecular characterization of these humanbreast cell models has demonstrated that normal breastepithelial cell behavior is determined in part by theprecise interplay that exists between a cell and its surrounding microenvironment. Recent functionalstudies of integrins in a human model system provideevidence to support the idea that the structuralstability afforded by integrin-mediatedcell-extracellular matrix interactions is an important determinantof normal cellular behavior, and that alterations intissue structure can give rise to tumorigenicprogression.

A fly's eye view of tumor progression and metastasis

When developing appropriate models to study tumor progression and metastasis it is essential to balance concerns of tissue fidelity and physiological relevance with the experimental accessibility of the system. Existing models are currently being re-evaluated given an increasing awareness of tumor complexity: in addition to the tumor cells themselves, their neighboring cells and the surrounding stroma are now recognized as important regulators of cancer progression [1]. With these added complexities in mind, two complementary publications have recently made a compelling case for the use of Drosophila melanogaster in understanding tumor progression and the metastatic process [2,3]. n nPrevious studies using Drosophila to model cancer revealed a collection of individual genes [e.g., discs large (dlg), lethal giant larvae (lgl), scribble (scrib), Ras and lats] that when homozygous mutant, resulted in tumor-like over-proliferation in epithelial cell-rich larval imaginal discs (reviewed in [4,5]). The papers in question both suggest that instead of analyzing the effects of cancer-promoting mutations in the context of an entirely mutant animal or tissue, it would be more informative to generate mosaic animals in which cancer-disposing mutations are introduced, either alone or in specific combinations, in only a subset of cells. Such a model would not only shed light on the multi-step nature of tumor progression but would also offer a reasonable approximation of the clonal nature of human cancers.

Centrosome cycle studies reveal promising candidates for anti-cancer drug design

Centrosomes are complex cellular substructures that are dynamically regulated by a series of biochemical and morphological changes that parallel the progression of the cell cycle. In addition to organizing cytoplasmic microtubule arrays during interphase, centrosomes direct formation and positioning of the bipolar mitotic spindle and thereby enable equal partitioning of chromosomal material during mitosis. Because impaired centrosomal function during cell division has the potential to cause chromosomal missegregation, and thus lead to genetic instability, the centrosome has become an important focus for cancer research. In fact, many tumor cells, including those from the breast, exhibit excessive numbers of centrosomes and/or centrosomes with aberrant morphologies. A collection of recent papers, some of which challenge prevailing dogmas, provide new insight into the biology of the metazoan centrosome cycle (i.e., duplication, maturation and separation) and elucidate novel regulatory pathways that are reasonable targets for oncogenic therapy development. n nGiven the correlation between centrosome number and tumor progression, molecules regulating centrosomal duplication at the G1/S transition have been under considerable scrutiny. Fisk and Winey recently used a culture-based assay of centrosome duplication to query the relevance of mMps-1p, the mouse orthologue of the Saccharomyces cerevisiae MPS1p protein kinase, a protein known to regulate spindle pole body replication [1]. While overexpression of wild-type mMps-1p promotes excessive duplication events, a kinase-inactive form of mMps-1p exerts a dominant negative effect antagonizing centrosome replication. Furthermore, mMps-1p protein stability, and thus function, is regulated via phosphorylation through cyclin-dependent kinase 2 (CDK2), a known mediator, and possible coordinator, of DNA replication and centrosomal duplication. Matsumoto and Maller have demonstrated that, in the context of S-phase-arrested Xenopus extracts, it is calcium/calmodulin-dependent kinase (CaMKII), and not CDK2, that actually initiates centrosomal duplication [2]. CaMKII is likely triggered in response to the periodic calcium oscillations that occur at the G1/S transition and are required for centrosome duplication. Finally, a provocative study by Khodjakov et al. challenges the concept that centrosome replication is an obligate semi-conservative process [3]. They show that new centrosomes form de novo in S phase-arrested cells in which centrosomes have been removed by laser ablation. This finding not only raises important questions regarding centrosome duplication checkpoints but also suggests that tumor-associated centrosome amplification could conceivably be attributed to de novo synthesis events. n nAurora serine/threonine kinases have received recent attention as critical regulators of the cell and centrosome cycle. One family member, called Aurora A, is over-expressed in many tumors and is found in the 20q13 breast cancer amplicon. In cultured cells, over-expression of Aurora A results in centrosome amplification and aneuploidy. Recent results indicate that Aurora A is an important determinant of centrosome maturation. Hannak et al. employed an RNAi strategy to query the role of Caenorhabditis elegans Aurora A (or AIR-1) during the first embryonic cell division [4]. AIR-1 depletion resulted in the incomplete separation of centrosomes, an event likely due to the impaired accumulation of γ-tubulin and other pericentriolar components required for centrosome maturation. Aurora A depletion in Drosophila also induces centrosomal maturation and spindle defects. For example, Giet and colleagues showed that in both neuroblasts and syncytial embryos, mutations in Aurora A lead to a decrease in the length of astral microtubules and to a disorganization of centrosomes [5]. Aurora A-depleted cells also displayed mitotic spindles with gross metaphase defects. Moreover, reduced Aurora A function correlated with aberrant centrosomal recruitment of the D-TACC/MSPS complex, which plays a role in the regulation of astral microtubules of the spindle. Likewise, in an independent study of Drosophila sensory organ precursor cells, Berdnik and Knoblich showed that mutations in Aurora A impair centrosomal accumulation of both centrosomin and γ-tubulin and give rise to spindle defects [6]. n nThe genetic models described above argue strongly for a role for Aurora A in centrosome maturation and suggest that perturbed Aurora A function has severe consequences on spindle function during subsequent mitotic progression. A recent study by Meraldi et al. in cultured cells has suggested that Aurora A over-expression induces centrosomal amplification, not by modulating duplication machinery, but by promoting aberrant mitosis and cytokinesis [7]. Aurora A-induced malfunctions lead to an abundance of multinucleated cells and a seeming amplification of what are actually normally duplicated centrosomes. This assessment is further supported by the fact that centrosome amplification is exacerbated in cells defective in the mitotic checkpoint protein p53. Collectively, these studies demonstrate the importance of Aurora A as a target for cancer study. In this regard, a recent structural analysis, by Cheetham and colleagues, of Aurora A highlights a number of features unique to this kinase [8], and in doing so provides potentially useful information for the development of specific Aurora A modulators. n nOther centrosomal constituents of recent notice are centrosomal Nek2-associated protein 1(C-Nap1) and the Rho-dependent protein kinase, p160ROCK, both of which appear to be involved in centrosome organization via their contributions to intercentriolar linkages. C-Nap1 localizes preferentially to the ends of parental centrioles and has been implicated in cell-cycle-regulated cohesion of centrosomes following duplication. Mayor and colleagues recently showed that C-Nap1 is phosphorylated at the onset of mitosis. This phosphorylation drives dissociation of C-Nap1 from the centrioles, and may thereby contribute to centrosome separation leading to bipolar spindle formation [9]. p160ROCK, which was originally identified from a panel of centrosome-directed monoclonal antibodies, plays a role in intercentriolar positioning. Chevrier et al. have shown that within a single centrosome, p160ROCK localizes to the maternal centriole and the intercentriolar linker [10]. p160ROCK-specific inhibitors induced centriole separation in G1 cells and caused accelerated movement of the maternal centriole to the midplate after mitosis. Thus, p160ROCK is an important regulator of centrosome organization and positioning throughout the cell cycle. n nClearly, irregularities in centrosome duplication, maturation and/or separation have potential to wreak havoc on the fidelity of mitotic events. Presumably, these centrosomal defects contribute to tumor cell aneuploidy, which is observed in many human cancers. Given the abundance of centrosome cycle regulators, it is likely that studies such as those described above will contribute to the development of important cancer diagnostics and/or therapies.

91st Annual Meeting of the American Association for Cancer Research

The 91st Annual Meeting of the American Association for Cancer Research (AACR) was held at the Moscone Convention Center in San Francisco, California, USA, April 1-5, 2000. The comprehensive and multidisciplinary nature of the AACR meeting was conveyed through a collection of concurrent symposia, minisymposia and poster sessions. These standard meeting forums were further supplemented by additional educational workshops, interactive meet-the-expert sessions at the beginning of each day and panel discussions approaching more general interest topics, such as the relationship of media, science and consumers. Although it was generally impossible to attend all of the sessions on a particular topic, many topics were repeated in different formats throughout the meeting to allow for attendees to get a reasonable sampling of the current trends in each field. n nA number of scientists were honoured at the meeting for their outstanding contributions to cancer research. Charles Sherr (St Judes Childrens Research Hospital, Memphis, TN, USA) was awarded the Pezcoller International Cancer Research Award for his work on the mechanisms of cell growth control and neoplastic transformation. The Bruce F Cain Memorial Award was given to Axel Ullrich (Max-Planck Institute for Biochemistry, Martinsried, Germany) who has successfully translated his pioneering work on tyrosine kinase receptors, such as HER2/neu, into actual treatment strategies. Edison T Liu (National Cancer Institute, Bethesda, MD, USA) was also recognized for his work in establishing a correlation between HER2/neu overexpression and those breast cancers that have an unfavourable prognosis and high probability of responding to doxorubicin therapy. Finally, the prestigious GHA Clowes Memorial Award was presented to Elizabeth Blackburn (University of California, San Francisco, CA, USA) for her pioneering work in the discovery of telomerase and its potential role in cancer. n nHerein we outline a few of the many provocative studies discussed at the meeting. Although some of the topics discussed below are specific to the breast, others addressing global mechanisms of tumour progression are also considered because they may be appropriate paradigms for understanding and treating breast cancer in the future.