Introduction

Cancer metastasis (distraction of cancer cells from the main tumor site to distant organs) is the most common cancer death worldwide. But why tumors can spread so easily, and how did it get that way? This was one of the biggest mysteries of oncology. 2D cell cultures and animal models have done us, but they can't fully represent how cancer cells interact with the microenvironment as they spread. Therefore, 3D printing technology lets scientists print intricate, 3D micro-models of tumor metastases. These models are more accurate and predictable, and provide new insights into the biology of tumors and a way to accelerate therapy. The following article is a review of the promise, application, and difficulties of 3D printed tumor metastasis models and where it will lead in the world of cancer research.

Figure 1. A 3D bioprinted ovarian cancer model. (Datta P, et al.; 2020)

The Tumor Metastasis Process: A Twisted Process

Among the stages of metastasis, in which the tumor cells permeate the body, flow into the bloodstream or lymphatics, and colonize distant organs. In all stages, cancer cells and the surrounding microenvironment (ECM), stromal cells, immune cells and endothelial cells) must interact complexly. And second, the microenvironment at the secondary site (the lung, liver, bone) controls how cancer cells live and generate secondary tumors.

To understand this fluid, multifactorial dynamic, researchers need novel models that are not limited to in vitro 2D cultures, where the cell-to-cell dynamic and 3-dimensional nature of tumors are too difficult to study. 3D printing is one possibility: models of metastases could be made in a very similar way to what we can expect from the human in vivo world.

Metastasis Modelling 3D Printer Technology for 3D Printing

Known as additive manufacturing, 3D printing constructs objects by layering components (biocompatible hydrogels, cells, ECM parts). It can be used to create the precise geometry, cellular makeup and microstructure of the printed model that emulates the complexity of metastatic tumors.

1. Bioinks and Materials

The choice of bioinks is important to re-create the metastatic tumor microenvironment. For cell encapsulation, hydrogels – which can imitate tissues' suppleness and wettability – are common. These hydrogels can be blended with other ingredients including collagen, fibrin and hyaluronic acid to create the ECM that helps with metastasis through adhesion, migration and invasion of cells.

So can bioinks made of cancer cells, stromal cells (fibroblasts and endothelial cells), and immune cells, which form an amorphous microenvironment that resembles the in vivo tumor. The ECM proteins and signalling molecules that allow cancer cells to interact with the stroma must be added specifically to models of metastases.

2. Modelling Tumor Metastases Printing Techniques for Metastasis Tumors

For tumor metastasis micro-models, 3D bioprinting methods are applied:

Extrusion-based Bioprinting: Here you layer-by-layer deposition of cell-rich bioinks through a nozzle. It's also used for creating larger models with more cells and higher cell densities where you can include a range of cell types like tumor cells, endothelial cells, and immune cells. In a way, extrusion bioprinting also can build strong models, which is useful for modeling the stiffness of tissue in metastasis.

Inkjet Bioprinting: Inkjet printing layers layer using bioink droplets. It is ideal for high-resolution models and allows the deposition of various materials with high control. It's a common procedure when you need very detailed cell or biomaterial patterning – for example, to recreate the vascularisation and stromal structure of metastasis.

Stereolithography (SLA): SLA is a process of polymerizing photo-crosslinkable material using light, which results in very fine and intricate geometries.

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Uses of 3D Printed Tumor Metastasis Sculptures

1. Studying Metastatic Mechanisms

The molecular and cellular processes governing metastasis is one of the major uses of 3D printed tumor metastasis models. These models can be employed to study how cancer cells are in contact with the ECM, how they invade tissues, and how they can spread and colonise other distant organs. Imagining the basement membrane, for instance, or introducing gradients of ECM stiffness, researchers could examine the effect of physics on cancer cell migration.

These models could also be applied to study the contribution of immune cells and stromal tissues to metastasis. Co-culturing cancer cells with immune cells (macrophages, T cells, etc.) could, for example, give clues as to how the immune system supports or blocks metastatic activity.

2. Drug Testing and Screening

3D printed metastasis models are a great way to test drugs especially for metastasis therapies. Traditionally used 2D cell cultures don't work well to find therapies that stop the spread of disease, because they lack a realistic replica of the tumor microenvironment. On the contrary, 3D-printed models enable drugs to be evaluated in a more natural setting and give better data on how well they are blocking tumor invasion, migration and dissemination.

They could also be used to test combination treatments for each stage of metastasis, including anti-mobility drugs, inhibitors of new blood vessel formation (anti-angiogenic drugs) or immune checkpoint inhibitors that boost the body's ability to fight off the metastatic cells.

3. Personalized Medicine

3D printed tumor models, created from patient biopsies, are a path to personalised medicine, and allow oncologists to trial therapies on simulations that are more faithful to a patient's tumor.

Challenges

Although 3D printing is promising for the study of cancer, it's also been quite difficult to build tumor metastasis micro-models due to some hurdles.

• Model Complexity: Tumor metastasis is so intricate and there are too many steps and interactions to model all the complexities in a single model. Current models are moving very fast, but usually they lack such characteristics as vascular meshwork or the capability to mimic distant organ-specific metastases (eg, the colonisation of cancer cells in bone or brain).
• Reproducibility: 3D bioprinting is still in its infancy and reproducibility in model design is hard to guarantee. The bioink formulation, print conditions, and cell culture might be different for different models and this makes comparing study results hard.
• Vascularization: One of the biggest limitations with metastasis modeling is that it is not possible to build completely functioning vasculature on a printed structure. Cancer cells require arteries for blood to get nutrients and to go far, so modelling a blood flow is essential for modeling metastasis properly.

Future Directions and Conclusion

The future for tumor metastasis studies with 3D printing appears promising and the development of bioinks, print quality and model complexity will only make things more interesting. Across the vascular networks, scientists are trying to integrate them even more fully in more advanced models that better capture what cancer cells face during metastasis. And the integration of AI and machine learning might be used to better parse the rich data produced by these models, to support drug discovery and personalised care.

To sum up, 3D printed tumor metastasis models are the future of cancer. They provide a better, more reproducible and more morally sound substitute for conventional models, which allows us to understand metastatic activity better and will lead to better treatments. Increasingly, as technology develops, these models will become essential to the biology of cancer, and thus to patient outcomes.

Reference

1. Datta P, et al.; 3D bioprinting for reconstituting the cancer microenvironment. NPJ Precis Oncol. 2020, 4:18.