Evidence is mounting for exosomes playing a central role in many aspects of cancer biology. As researchers start to unravel the secrets of these elusive little vesicles, this is opening a host of exciting new opportunities to tackle cancer – from non-invasive diagnostics to innovative new therapeutics.
Exosomes are thought to be a specific class of vesicle that arise from multivesicular bodies (MVBs) and are released from cells. Around 50 to 150 nanometres in size, these tiny membrane-wrapped packages contain a variety of cargo – such as small molecules, proteins or nucleic acids.
However, with no markers available to distinguish them from other vesicles, exosomes are difficult to define – and their nomenclature is currently a subject of hot debate.
“I quite like the term exosomes because that’s what I think I’m studying – but I can’t prove it!” says Professor Aled Clayton of the Division of Cancer and Genetics at the University of Cardiff, UK.
“The umbrella term that is probably more correct is extracellular vesicles, which also includes larger plasma membrane-derived vesicles that are structurally and physically quite different.”
As well as what to call them, the role of exosomes in healthy systems is also unclear.
“We think they may facilitate a few things, one is potentially to remove unwanted proteins that may have misfolded, and another is to transfer information between cells,” says Dr Richard Kelwick, a Research Associate at Imperial College London, UK.
Exosomes are integral in cancer biology
However, there is an increasing number of studies pointing the finger at exosomes in many aspects of cancer development, including hypoxia driven epithelial-mesenchymal transition (EMT), cancer stemness, angiogenesis, metastasis and modulating the immune response.
“Just think of everything that tumors need to do – these vesicles are in some fashion involved,” says Clayton.
Exosomes may even help explain why particular cancer types show a preference for metastatic spread to specific organs.
“Some papers suggest that these vesicles might be a way for tumor cells to prime other organs – so they are like seeds, eliciting some change that then allows tumor cells to metastasize there,” explains Clayton.
They may also help cells to develop drug resistance, for example by packaging the drug into exosomes and throwing it out of the cell.
“We think they can even share these properties with neighboring cells, with a bystander cell that may be drug susceptible receiving vesicles from a drug resistant cell – and by that interaction becoming drug resistant,” says Clayton.
Liquid biopsies for early detection and personalized treatment
With the evidence building for exosome involvement in many different facets of cancer, this is opening a wealth of new opportunities for potential clinical applications.
At the forefront of this gold rush is using cancer exosomes as a source of readily accessible biomarkers for diagnostics.
“While it can sometimes be difficult to access tumors, it can be very easy to access exosomes as they’re found in pretty much all bodily fluids like blood, saliva or urine,” says Kelwick.
As exosomes find their way into the bloodstream at an early stage of a tumor’s development, they offer an exciting window into the body that could help detect cancers very early on.
“For example, my part in a wider collaborative project to develop a new screening programme to detect early-stage lung cancer is to isolate exosomes from blood samples and look at the micro RNAs,” says Kelwick.
It may even be possible to use them as tests to monitor tumors for changes that indicate when they are becoming dangerous.
“By serially tracking people as they age and identifying the point at which a tumor is beginning to show signs of becoming aggressive, which is when you need to treat it,” explains Clayton.
Exosome biomarkers could also provide insights to enable personalized treatment based on a what’s driving a patient’s own cancer, or as a companion diagnostic to figure out if their treatment is working in real-time.
Exosomes as cancer therapeutics
Looking even further ahead into the future another exciting possibility is to use exosomes to treat cancer, exploiting their ability to get inside cells.
“You could load the vesicles with a drug and then you have a system that is protecting it, which is being disseminated around the body, can target a particular cell or tissue, and can get through the plasma membrane and into the cell in an extremely efficient fashion,” explains Clayton.
There is also the added potential to design more complex therapeutics by loading combinations of different treatment modalities into the exosomes – for example, small molecules, proteins, or nucleic acids.
“This multimodal approach seems to be the way we’re heading for diseases that are complex and when one target isn’t enough, which is especially true for cancer where resistance can occur,” enthuses Kelwick.
The challenges of studying exosomes
If we are to exploit the array of these new opportunities, we will first need to improve our understanding of their fundamental biology – by isolating them and studying their components.
The standard technique for isolating exosomes is ultracentrifugation. However, there can be problems due to contamination with other proteins – for example, from the blood – or damage to the exosomes due to high centrifugal forces.
“Microfluidics provides a promising alternative. These tend to be silicon or plastic chips with carefully designed channels that help you to separate out exosomes based on their physical and electrophysical characteristics,” says Kelwick.
Once isolated, nanoparticle tracking analysis is useful to get a sense of the populations of exosomes within a sample. Researchers then use many different downstream techniques to characterize the exosomes’ surface markers or contents, including flow cytometry, mass spectrometry or next-generation sequencing.
However, biologists admit they are struggling to extract these tiny exosomes and quantify their molecules – particularly from the blood that is full of contaminating factors.
“I think we’ve reached a point where we do need help from other disciplines, maybe chemists or engineers or physicists, who can help us devise new methods which are very sensitive to enable us to measure these very small things in the context of a very complex background,” says Clayton.
Studying their normal biology using cell culture is also challenging as it is often impractical due the sheer volume of flasks needed to generate sufficient exosomes – and using 2D cell culture often isn’t ideal for understanding complex processes. To address this, some companies are developing new ways of culturing cells specifically for exosome production.
“I think that the best I’ve seen is hollow fiber cell culture, which is a kind of a mix between 2D and 3D culture. It allows you to have a very high density in a small volume enabling you to get very concentrated exosomes,” says Kelwick.
Exosomes in the clinic
Looking into the future, exosomes may play a key part in cancer diagnosis and treatment in patients – from several different perspectives.
“I think exosome diagnostics will be normal for anyone who is suspected of having cancer or is under treatment. And perhaps one day we may be able to engineer personalized exosomes – designed to treat a patient’s own specific cancer,” suggests Kelwick.
Potential clinical applications for tackling metastasis are also exciting, but currently hard to scope.
“As we don’t currently have any anti-metastatic drugs, this could open a whole new avenue of controlling the disease so that people could live with tumors and prevent them from spreading and causing lethal disease,” says Clayton.
Small packages with big potential
These long overlooked, tiny vesicles are still proving elusive – with many challenges remaining with how to capture and study them, and even around what to call them. But they are certainly worth pursuing as they have the potential to transform the way we diagnose and treat cancer in the future.
“I would say they are absolutely fundamental to our understanding of biology – you’d have to be an idiot to ignore or dismiss them just because it’s difficult,” says Clayton.