Profiling plasma extracellular vesicle by pluronic block‐copolymer based enrichment method unveils features associated with breast cancer aggression, metastasis and invasion

Abstract

Extracellular vesicle (EV)-based liquid biopsies have been proposed to be a readily obtainable biological substrate recently for both profiling and diagnostics purposes. Development of a fast and reliable preparation protocol to enrich such small particles could accelerate the discovery of informative, disease-related biomarkers. Though multiple EV enrichment protocols are available, in terms of efficiency, reproducibility and simplicity, precipitation-based methods are most amenable to studies with large numbers of subjects. However, the selectivity of the precipitation becomes critical. Here, we present a simple plasma EV enrichment protocol based on pluronic block copolymer. The enriched plasma EV was able to be verified by multiple platforms. Our results showed that the particles enriched from plasma by the copolymer were EV size vesicles with membrane structure; proteomic profiling showed that EV-related proteins were significantly enriched, while high-abundant plasma proteins were significantly reduced in comparison to other precipitation-based enrichment methods. Next-generation sequencing confirmed the existence of various RNA species that have been observed in EVs from previous studies. Small RNA sequencing showed enriched species compared to the corresponding plasma. Moreover, plasma EVs enriched from 20 advanced breast cancer patients and 20 age-matched non-cancer controls were profiled by semi-quantitative mass spectrometry. Protein features were further screened by EV proteomic profiles generated from four breast cancer cell lines, and then selected in cross-validation models. A total of 60 protein features that highly contributed in model prediction were identified. Interestingly, a large portion of these features were associated with breast cancer aggression, metastasis as well as invasion, consistent with the advanced clinical stage of the patients. In summary, we have developed a plasma EV enrichment method with improved precipitation selectivity and it might be suitable for larger-scale discovery studies.

KIAA0100 Modulates Cancer Cell Aggression Behavior of MDA-MB-231 through Microtubule and Heat Shock Proteins

“…MVBs or even EVs might be one of its potential destinations. This is consistent with our previous finding that showed KIAA0100 was detected in EVs of all three breast cancer cell line EVs as well as in a plasma EV fraction from breast cancer patients [ 13 ].…”

“…The development and progression of breast cancer are involved in a complex process including interaction of many genetic, epigenetic and environmental factors at different stages of cancer cells, such as proliferation/growth, anchorage/re-attachment, adhesion/aggregation, anoikis resistance, and metastasis/invasion. KIAA0100 has been found to be elevated in breast cancer tissues from multiple studies [ 4 , 8 , 9 , 10 ] as well as in malignant transformed benign breast cells [ 5 ]; in addition, our previous study also showed that KIAA0100 protein was found in all three different breast cancer cell lines including MCF7, T47D and MDA-MB-231 as well as their respective extracellular vesicles (EVs) preparations [ 13 ], suggesting this protein might be related to breast cancer aggression.…”

“…Meanwhile, multiple bioinformatics tools have been adopted to predict the functions of KIAA0100, and realize that it might be an anti-apoptotic factor related to carcinogenesis or progression [ 2 , 12 ]. Interestingly, our recent study showed that KIAA0100 was elevated in the extracellular vesicles (EVs) fraction in the plasma from breast cancer patients compared to non-cancer controls [ 13 ], suggesting KIAA0100 may be linked to EV pathway. However, the molecular and cellular functions that KIAA0100 plays and how it contributes to cancer development, especially in breast cancer cells, remain elusive.…”

Cancer Nanomedicine: A New Era of Successful Targeted Therapy

“…The accumulation of doxorubicin in tumors was more than free doxorubicin with normal distribution of polymer in normal tissues [85,86]. The mechanism of action of micelles is changing the structure of the membrane, decreasing membrane fluidization, and inhibiting function and expression of efflux transporters, such as P-gp and MRPs [87][88][89][90]. These subsequently sensitize resistant cancer cell to the chemotherapeutic agents [91], increasing the proapoptotic [92], decreasing the levels of glutathione (GSH) and glutathione-S-transferase (GST) activity, inhibiting the mitochondrial respiratory chain [92], decreasing oxygen consumption [93], and decreasing both mitochondrial membrane potentials [94], and the production of reactive oxygen species and release of cytochrome C in MDR cells are additional mechanisms [95,96].…”

Pluronic-F127/Platelet Microvesicles nanocomplex delivers stem cells in high doses to the bone marrow and confers post-irradiation survival

“…The plasma extracellular vesicles (EVs), of which the pMVs are also a major component, were particularly found to bind with pluronics. In their study, Zhong et al were able to selectively enrich the EVs from the plasma by using Pluronic-F68 (PF68) 19 . We through our study deduced that the pMVs were able to bind Pluronic-F127 (PF127 or poloxamer407) more efficiently than PF68.…”