International researchers headed by a team at Weill Cornell Medicine have identified a new category of nanoparticle messengers—dubbed exomeres—that shuttle molecules such as proteins, fats, and nucleic acids between cells, and could play a role in cancer development or how well patients tolerate chemotherapy. The nanosized exomeres, discovered using a state-of-the-art technique known as asymmetric flow field-flow fractionation (AF4), are produced by both normal and cancer cells, but are distinct from exosomes, the extracellular membrane vesicles that are already known to act as intercellular communicators.

“We found that exomeres are the most predominant particle secreted by cancer cells,” claims senior author David Lyden, M.D., Ph.D., the Stavros S. Niarchos Professor in Pediatric Cardiology, and a scientist in the Sandra and Edward Meyer Cancer Center and the Gale and Ira Drukier Institute for Children's Health at Weill Cornell Medicine. “They are smaller and structurally and functionally distinct from exosomes. Exomeres largely fuse with cells in the bone marrow and liver, where they can alter immune function and metabolism of drugs. The latter finding may explain why many cancer patients are unable to tolerate even small doses of chemotherapy due to toxicity.”

Using the AF4 technology, the team, working with collaborators in the U.S. Portugal, Korea, China, and Spain, was also able to categorize exosomes into two subpopulations, based on vesicle size, which also appeared to have different functions. They report their findings in Nature Cell Biology, in a paper entitled “Identification of Distinct Nanoparticles and Subsets of Extracellular Vesicles by Asymmetric Flow Field-Flow Fractionation.”
Exosomes are nanosized extracellular membrane vesicles that are secreted by most cell types, including cancer cells. “Proteins, genetic material (for example, mRNAs, miRNAs, lncRNAs, DNA), metabolites and lipids are selectively recruited and packaged into exosomes, which horizontally transfer their cargo to recipient cells, thereby acting as vehicles of intercellular communication under both physiological and pathological conditions,” the authors write. Knowledge about exosome biology and function is increasing, but until now the technical challenges associated with separating and characterizing different subpopulations have hampered in-depth studies into their molecular composition.

To address this challenge, the Weill Cornell Medicine team turned to AF4 technology, which is already widely used to characterize nanoparticles and polymers in the pharmaceutical industry, and to study biological macromolecules, protein complexes, and viruses. The researchers used the AF4 technology to fractionate extracellular vesicles from both normal and cancer cells. The results identified two different subpopulations of exosome, which they termed large exosome vesicles (Exo-L, 90–120 nm diameter) and small exosome vesicles (Exo-S, 60–80 nm). The technology also isolated an abundant population of much smaller, nonmembranous nanoparticles—exomeres—which, at about 35 nm diameter, were also much smaller than both the Exo-L and Exo-S nanoparticles.

The technology isolated nanoparticles with diameters corresponding to the Exo-L, Exo-S, and exomere vesicles from more than 20 cell lines, including human melanoma cells. However, each of the three subsets demonstrated very different patterns of biodistribution, “suggesting distinct biological functions,” the authors write.

The exomeres and exosomes had very different biophysical characteristics, such as stiffness and electrical charge, which will ‘likely affect their behavior in the body,” comments lead author Haiying Zhang, Ph.D., an assistant professor of cell and developmental biology in pediatrics at Weill Cornell Medicine. “The more rigid the particle, the easier it is likely taken up by cells, rendering exomeres which are stiffer than exosomes, the more effective messengers of transferring tumor information to recipient cells.”

The two different exosome subpopulations and exomeres also carried distinct types of proteins, nucleic acids, and other components, and exhibited different cellular functions. “Our analyses revealed that exomeres were selectively enriched in proteins involved in metabolism, especially ‘glycolysis’ and ‘mTORC1’ metabolic pathways, suggesting their potential roles in influencing the metabolic program in target organ cells, as well as in proteins associated with coagulation (for example, factors VIII and X) and hypoxia,” the researchers note.  The exomeres were also enriched in key proteins controlling glycan-mediated protein folding control and glycan processing, “suggesting exomere cargo may play a role in modulating glycosylation in distant recipient cells.”

Results from the analyses in addition indicated that each nanoparticle may play distinct roles in how they influence cancer. “Our observation that nanoparticle subtypes have different organ biodistribution patterns suggests that they mediate the pleiotropic effects of cancer,” the authors note.

Exomeres were found to transport metabolic enzymes to the liver, which indicates that they may target the liver and reprogram metabolic function to support tumor progression. Exomeres also carried blood-clotting factors to the liver, whereas the results indicated that Exo-L nanoparticles might promote metastasis to lymph nodes, with Exo-S supporting distant metastasis. “Distinct organ distributions indicate that nanoparticle subsets may be involved in different aspects of tumour progression and metastasis,” the researchers write.

“Cancer is truly a systemic disease that requires multiorgan involvement to progress,” Dr. Lyden comments. “Our finding that tumor cells secrete these three distinct nanoparticles, that then target cells in different organs, reflects this important aspect of the disease.”

The presence of exosomes and exomeres in bodily fluids such as lymphatic fluid could potentially help scientists develop new biomarkers for early detection of cancer or other disorders. “Based on our findings, the next phase will be to measure exosomes and exomeres in plasma samples to help predict organs that may be targeted for metastasis during tumor progression,” Dr. Lyden states. “This will help us better understand the biology of cancer, guide therapeutic decisions, and develop novel therapies.”

“Our identification of exomeres highlights the diversity of EVs [extracellular vesicles] and particles secreted by cells,” the authors conclude. “Elucidating their biogenesis will be essential to unravel their roles in cellular and organ function. Target cells and the functional outcomes exerted by each nanoparticle subset in organs need to be further delineated to advance our understanding of the collective, systemic effects of nanoparticles in the metastasis process. Undoubtedly, these discoveries will open avenues for translational studies of EVs and particles in diagnostic, prognostic and therapeutic applications.”

Cornell University has filed a patent application on the technology described in the paper. Meanwhile, scientists aim to continue studying the vesicles and their cargo, and work to understand their functions at different target organ sites. “Understanding these characteristics may help scientists better understand how exomeres and exosomes help cancers grow and spread to other organs, as well as what role they may play in other diseases,” Dr. Zhang comments.

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