Opportunities and challenges for use of tumor spheroids as models to test drug delivery and efficacy

Department of Periodontics & Oral Medicine, University of Michigan School of Dentistry, The University of Michigan, Ann Arbor, MI 48109-2099, United States.
Journal of Controlled Release (Impact Factor: 7.71). 05/2012; 164(2). DOI: 10.1016/j.jconrel.2012.04.045
Source: PubMed


Multicellular spheroids are three dimensional in vitro microscale tissue analogs. The current article examines the suitability of spheroids as an in vitro platform for testing drug delivery systems. Spheroids model critical physiologic parameters present in vivo, including complex multicellular architecture, barriers to mass transport, and extracellular matrix deposition. Relative to two-dimensional cultures, spheroids also provide better target cells for drug testing and are appropriate in vitro models for studies of drug penetration. Key challenges associated with creation of uniformly sized spheroids, spheroids with small number of cells and co-culture spheroids are emphasized in the article. Moreover, the assay techniques required for the characterization of drug delivery and efficacy in spheroids and the challenges associated with such studies are discussed. Examples for the use of spheroids in drug delivery and testing are also emphasized. By addressing these challenges with possible solutions, multicellular spheroids are becoming an increasingly useful in vitro tool for drug screening and delivery to pathological tissues and organs.

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    • "In the context of tumor cell cultures, most bioreactor-based approaches have introduced agitation techniques and microfluidic platforms [17e19], generating hydrodynamic conditions in the form of convectional fluid flow around cells and tissues. However, resulting superficial flows in those systems are of limited effectiveness to address internal transport limitations [20], which in turn critically affect cell behavior and function as well as drug penetration [21]. Bioreactor devices applying direct perfusion were shown to provide uniform cell distribution [22], allowing the development and maintenance of uniformly viable large tissues for prolonged culture times [22] [23]. "
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    ABSTRACT: Anticancer compound screening on 2D cell cultures poorly predicts "in vivo" performance, while conventional 3D culture systems are usually characterized by limited cell proliferation, failing to produce tissue-like-structures (TLS) suitable for drug testing. We addressed engineering of TLS by culturing cancer cells in porous scaffolds under perfusion flow. Colorectal cancer (CRC) HT-29 cells were cultured in 2D, on collagen sponges in static conditions or in perfused bioreactors, or injected subcutaneously in immunodeficient mice. Perfused 3D (p3D) cultures resulted in significantly higher (p < 0.0001) cell proliferation than static 3D (s3D) cultures and yielded more homogeneous TLS, with morphology and phenotypes similar to xenografts. Transcriptome analysis revealed a high correlation between xenografts and p3D cultures, particularly for gene clusters regulating apoptotic processes and response to hypoxia. Treatment with 5-Fluorouracil (5-FU), a frequently used but often clinically ineffective chemotherapy drug, induced apoptosis, down-regulation of anti-apoptotic genes (BCL-2, TRAF1, and c-FLIP) and decreased cell numbers in 2D, but only "nucleolar stress" in p3D and xenografts. Conversely, BCL-2 inhibitor ABT-199 induced cytotoxic effects in p3D but not in 2D cultures. Our findings advocate the importance of perfusion flow in 3D cultures of tumor cells to efficiently mimic functional features observed "in vivo" and to test anticancer compounds. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Biomaterials 09/2015; 62. DOI:10.1016/j.biomaterials.2015.05.037 · 8.56 Impact Factor
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    • "Often, promising results obtained from 2D cannot be translated similarly into in vivo settings (Goodman et al., 2008). Whereas cells on 2D are exposed to a uniform environment with sufficient oxygen and nutrients, cells in solid tumors are exposed to gradients of critical chemical and biological signals (Mehta et al., 2012), which can exert both stimulatory and inhibitory effects on tumor progression (Mehta et al., 2012). Intriguingly, certain tumor cells from cancer patients are intrinsically resistant to a broad spectrum of chemotherapeutic drugs without any previous exposure to those cytotoxic agents (Sanchez et al., 2009; Zhu et al., 2005, 2012). "
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    ABSTRACT: Cancer occurs when cells acquire genomic instability and inflammation, produce abnormal levels of epigenetic factors/proteins and tumor suppressors, reprogram the energy metabolism and evade immune destruction, leading to the disruption of cell cycle/normal growth. An early event in carcinogenesis is loss of polarity and detachment from the natural basement membrane, allowing cells to form distinct three-dimensional (3D) structures that interact with each other and with the surrounding microenvironment. Although valuable information has been accumulated from traditional in vitro studies in which cells are grown on flat and hard plastic surfaces (2D culture), this culture condition does not reflect the essential features of tumor tissues. Further, fundamental understanding of cancer metastasis cannot be obtained readily from 2D studies because they lack the complex and dynamic cell-cell communications and cell-matrix interactions that occur during cancer metastasis. These shortcomings, along with lack of spatial depth and cell connectivity, limit the applicability of 2D cultures to accurate testing of pharmacologically active compounds, free or sequestered in nanoparticles. To recapitulate features of native tumor microenvironments, various biomimetic 3D tumor models have been developed to incorporate cancer and stromal cells, relevant matrix components, and biochemical and biophysical cues, into one spatially and temporally integrated system. In this article, we review recent advances in creating 3D tumor models employing tissue engineering principles. We then evaluate the utilities of these novel models for the testing of anticancer drugs and their delivery systems. We highlight the profound differences in responses from 3D in vitro tumors and conventional monolayer cultures. Overall, strategic integration of biological principles and engineering approaches will both improve understanding of tumor progression and invasion and support discovery of more personalized first line treatments for cancer patients.
    Biotechnology Advances 11/2014; 32(7). DOI:10.1016/j.biotechadv.2014.07.009 · 9.02 Impact Factor
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    • "Second, single cells and monolayers are not biologically relevant models, especially for studying complex biological processes [14]. Altogether, a more appropriate approach would consist of performing SECM measurements on arrays of 3D cell aggregates such as spheroids, which are acknowledged as closer models to the in vivo situation [15]. "
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    ABSTRACT: While scanning electrochemical microscopy (SECM) is a powerful technique for non-invasive analysis of cells, SECM-based assays remain scarce and have been mainly limited so far to single cells, which is mostly due to the absence of suitable platform for experimentation on 3D cellular aggregates or microtissues. Here, we report stamping of a Petri dish with a microwell array for large-scale production of microtissues followed by their in situ analysis using SECM. The platform is realized by hot embossing arrays of microwells (200 μm depth; 400 μm diameter) in commercially available Petri dishes, using a PDMS stamp. Microtissues form spontaneously in the microwells, which is demonstrated here using various cell lines (e.g., HeLa, C2C12, HepG2 and MCF-7). Next, the respiratory activity of live HeLa microtissues is assessed by monitoring the oxygen reduction current in constant height mode and at various distances above the platform surface. Typically, at a 40 μm distance from the microtissue, a 30% decrease in the oxygen reduction current is measured, while above 250 μm, no influence of the presence of the microtissues is detected. After exposure to a model drug (50% ethanol), no such changes in oxygen concentration are found at any height in solution, which reflects that microtissues are not viable anymore. This is furthermore confirmed using conventional live/dead fluorescent stains. This live/dead assay demonstrates the capability of the proposed approach combining SECM and microtissue arrays formed in a stamped Petri dish for conducting cellular assays in a non-invasive way on 3D cellular models.
    PLoS ONE 04/2014; 9(4):e93618. DOI:10.1371/journal.pone.0093618 · 3.23 Impact Factor
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