(1) Department of Surgical, Medical, Molecular Pathology and Emergency Medicine, University of Pisa, Pisa, Italy
(2) Tissue Regeneration Department, University of Twente, Enschede, The Netherlands
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Understanding cancer biology is a major challenge of this century. The recent insight about carcinogenesis mechanisms, including the role exerted by the tumour microenvironment and cancer stem cells in chemoresistance, relapse and metastases, has made it self-evident that only new cancer models, with increased predictability, will allow the development of efficient therapies. The aims of this critical review are to briefly summarise and discuss the key aspects in the development of three-dimensional biomimetic tumour models. In this review, tissue engineering (TE) retains a valuable and highly exploitable potential. Tissue-engineered tumour models can account for a number of advantages, such as reproducibility, tailourable complexities (e.g., cell types, size, chemistry, architecture, mechanical properties, bioresorption and diffusion gradients) and ethical sustainability, making them suitable tools not only for mimicking normal tissue regeneration, but also, and most interestingly, for cancer development and resistance to therapies. Finally, we will focus upon interesting studies recently reported in the published literature about cancer TE, grouping their findings by tumour type, in order to give a snapshot picture of the current achievements to those cancer scientists, who are wishing to approach the field of TE. A special focus was given to pancreas, breast and prostate tumours.
There are marked intent affinities indicating TE as a suitable discipline to model cancer tissues. This is a topic of current efforts by several research groups worldwide, although, to date, well-defined guidelines have not been outlined yet, but rather preliminary individual studies have been reported.
Despite our body develops and evolves since the very first embryological events in a three-dimensional (3D) environment, nowadays we are still studying the processes at the base of developmental biology with a two-dimensional (2D) technology, i.e., with traditional
The concept of cancer TE is very recent, but holds great promise; indeed, convergences of objectives and methodologies between both disciplines have been highlighted and discussed elsewhere[10,11,12]. In 2006, at the dawn of cancer tissue engineering (TE) studies, the TE community pointed out their next-generation guidelines, underlining the necessity of complex biomimetic models, nicely correlating stem cell differentiation on TE scaffolds with developmental biology. To achieve the formation of mature functional substitutes
In this critical review, we aim at collecting and discussing with educational intent, the key aspects involved in the design of new biomimetic cancer models, with a special focus on the role, potential and actual—so far—played by TE. Finally, the ultimate purpose of this critical review is to stimulate a propulsive interaction between cancer scientists and tissue engineers, to respond, via a highly multidisciplinary approach, to still unmet therapeutic needs.
In this review, the authors have referenced some of their own studies. The protocols of these studies have been approved by the relevant ethics committees associated to the institution in which they were performed.
The search for cancer models has started in the second half of the last century and it is still in progress (Figure 1A–B). Traditional
Schematic picture of (A) a tumour and (B) tumour models, with their main characteristics. Some important aspects were identified and qualitatively scored according to the findings of the published literature and to our personal experience. They include model-inherent features, such as model type and reproducibility, and some model biomimetic capabilities.
2D, two-dimensional; 3D, three-dimensional.
The most widely used 3D (complex) model of cancer biology is typically an animal model (Fig. 1B).
Recently, microfluidics circuits have been developed to make a further step towards 3D cultures in cancer. Yet, when macroscopically relevant dimensions (higher than 1 mm) are achieved, nutrient diffusions and cell survival remain problematic. To solve these challenges, microfluidic well systems, with the capacity of controlling nutrient perfusion, have been developed and used alone or in combination with hydrogels.
Different from xenograft, spheroids and gel embedding, TE models can potentially offer all the fundamental achievements to cancer studies obtained so far for the regeneration of normal tissues as follows high standardisation of assays, multiple cell-type interaction, tailourable architecture allowing spontaneous 3D cell disposition and ECM synthesis, mechanical properties matching those of the tissue and tuneable diffusion profiles, thus appearing, in the end, as potentially elective models for the regeneration of 3D tumours (Figure 1B)[1,3,10,11,12,23].
A TE model of cancer should be a bottom-up 3D reconstruction of the tissue, using selected cells (CSCs or tumour cell mixtures), derived from primary cultures or from tissues, thus retracing the schematic diagram shown in Figure 2. For each tumour type, suitable scaffold architecture should be identified, ideally which is able to match the topographic and mechanical aspects of the native tissues[1,3,10,11,12,23].
Flowsheet of TE cancer models. This example is rendered with images of a study related to human pancreatic ductal adencocarcinoma (hPDAC) performed in our laboratories. Consequentially, single images/image groups show the following: light micrograph of hPDAC morphology (haematoxylin and eosin stain); light micrograph of isolated primary hPDAC cells (PP244); scanning electron microscopy (SEM) micrograph of scaffold inner structures (3D fiber deposition via Bioplotter®, sponge via emulsion and freeze-drying, microfibers via electrospinning); hPDAC cell/scaffold constructs under different viability assays (colour gradients of the scaffold surfaces highlight spatial localisation of viable cells) and light micrograph of a spongy construct (haematoxylin and eosin stain) showing presence of 3D hPDAC cell clusters within the scaffold pores, whose morphology mimics that of native PDAC (see the tumour biopsy micrograph).
2D, two-dimensional; 3D, three-dimensional.
The current state-of-the-art about the development of
Tumour cell/biomaterial models for different cancer types.
Due to its inauspicious prognosis, pancreatic ductal adenocarcinoma (PDAC) is the object of persistent studies. The development of an
The 3D models have been developed to study metastasis initiation and development, with the use of cellular aggregates or spheroids, and microfluidic devices[22,23]. Considering the relevance of breast and prostate cancer mortality due to their metastasis to bone, 3D models derived from TE know-how, have been developed to study metastatic events of these cancer types to bone engineered tissue. Cancer cell angiogenic signalling was regulated by integrin and correlated with enhanced production of interleukin-8 (IL-8). Further control over tumour angiogenesis was influenced by oxygen availability in 3D tumour culture models, with increased levels of IL-8 secretion in normoxia and of vascular endothelial growth factor in hypoxic culture conditions. Similarly, porous biomaterials containing inorganic phases like hydroxyapatite (HA) were used to create initial models of breast metastasis into bones and revealed a role of HA crystal size in tumour cell adhesion and proliferation[29,30].
Basic 3D systems have shown that breast and prostate cancer cells, among others, are indeed more resistant to chemotherapies than when cultured on 2D substrates, thus justifying the continued development of advanced
There are marked intent affinities indicating TE as a suitable discipline to model cancer tissues. This is a topic of current efforts by several research groups worldwide, although, to date, well-defined guidelines have not been outlined yet, but rather preliminary individual studies have been reported. Recent studies have reinforced the theoretical hypothesis that tissue-engineered cancer constructs can mimic the tumour microenvironment because of their three-dimensionality and their multi-parametric tailourability. The interactions between tumour cells and different biomaterials seem to play a key role in tumour biomimetics to be finely exploited in the very near future.
2D, two-dimensional; 3D, three-dimensional; CSC, cancer stem cells; ECM, extracellular matrix; HA, hydroxyapatite; hPDAC, human PDAC; IL-8, interleukin-8; PDAC, pancreatic ductal adenocarcinoma.
Authors wish to acknowledge Dr. Niccola Funel and all members of the Anatomical Pathology Unit of Cisanello Hospital (AOUP, Pisa, Italy) for experimental and theoretical support on pancreas cancer.
All authors contributed to the conception, design, and preparation of the manuscript, as well as read and approved the final manuscript.
All authors abide by the Association for Medical Ethics (AME) ethical rules of disclosure.
Tumour cell/biomaterial models for different cancer types.
|Tumour type||Model||Biomaterials||Cell line; species||Main results||Year||Ref. #|
|Pancreas||TE||PVA + gelatine||PP244; human||Good growth and viability||2008|
|TE||PGA-TMC + gelatine||isolated CSCs (CD24+, CD44+); human||Expression of cancer markers and cancer morphology||2013|
|Gel||Fibronectin-gelatine||K643f, NIH3T3; murine||More biomimetic drug delivery and ECM||2013|
|Spheroids||Methylcellulose||Panc-01, Capan-1 ASPC-1, BxPC-3; human||Improved chemoresistance with respect to 2D||2013|
|Breast||TE||Chitosan||MCF-7; human||3D growth conferred drug resistance||2005|
|TE||PLA, PLGA||MCF-7; human||Tissue-like structure and drug resistance||2005|
|TE||PLG + HA||MDA-MB231; human||HA improved cell adhesion||2010|
|TE||PLG + HA||MDA-MB231; human||Good proliferation||2011|
|Prostate||TE||PCL-TCP||PC3, LNCaP; human||Increased invasion potential||2010|
|Gel||PEG-Gln/PEG-MMP-Ly||LNCaP; human||Upregulated expression of MMPs, steroidogenic enzymes, and prostate specific antigen||2012|
|Oral||TE||PLG||LLC, MCF-7, U87; human||Tumour-similar ECM and hypoxic condition in 3D model||2007|
|Colorectal||Gel||lrECM/matrigel||CACO-2, COLO-206F, DLD-1, HT-29 SW-480 COLO-205; human||Different morphology from metastasis and primary cells||2013|
|Lymphoma||TE||PS||Z138, HBL2; human||Higher growth in 3D||2013|
|Lung||Spheroids||AlgiMatrix™||NSCLC cell lines (H460, A549, H1650, H1650 stem cells); human||Higher resistance to anticancer drugs than 2D (increased IC50 values of drug and reduced cleaved caspase-3 expression)||2013|
|Ewing Sarcoma||TE||Electrospun PCL||TC-71; human||Tumour biomimetics of morphology, growth kinetics and protein expression profile||2013|
2D, two-dimensional; 3D, three-dimensional; CSC, cancer stem cell; ECM, extracellular matrix; HA, hydroxyapatite; MMPs, matrix metalloproteinases; PCL-TCP, polycaprolactone-tricalcium phosphate; PCL, polycaprolactone; PEG, polyethylene glycol; PGA-TMC, poly(glycolide-co-trimethylene carbonate); PLGA, poly lactic-co-glycolic acid; PLG, poly(lactide-co-glycolide); PVA, poly(vinyl alcohol); PS, polystyrene; TE, tissue engineered.