Showing papers by "Sai Kiang Lim published in 2019"

Defining mesenchymal stromal cell (MSC)-derived small extracellular vesicles for therapeutic applications

Abstract: Small extracellular vesicles (sEVs) from mesenchymal stromal/stem cells (MSCs) are transiting rapidly towards clinical applications. However, discrepancies and controversies about the biology, functions, and potency of MSC-sEVs have arisen due to several factors: the diversity of MSCs and their preparation; various methods of sEV production and separation; a lack of standardized quality assurance assays; and limited reproducibility of in vitro and in vivo functional assays. To address these issues, members of four societies (SOCRATES, ISEV, ISCT and ISBT) propose specific harmonization criteria for MSC-sEVs to facilitate data sharing and comparison, which should help to advance the field towards clinical applications. Specifically, MSC-sEVs should be defined by quantifiable metrics to identify the cellular origin of the sEVs in a preparation, presence of lipid-membrane vesicles, and the degree of physical and biochemical integrity of the vesicles. For practical purposes, new MSC-sEV preparations might also be measured against a well-characterized MSC-sEV biological reference. The ultimate goal of developing these metrics is to map aspects of MSC-sEV biology and therapeutic potency onto quantifiable features of each preparation.

Proteomic Signature of Mesenchymal Stromal Cell-Derived Small Extracellular Vesicles

Abstract: Small extracellular vesicles (EVs) are 50-200 nm vesicles secreted by most cells. They are considered as mediators of intercellular communication, and EVs from specific cell types, in particular mesenchymal stem/stromal cells (MSCs), offer powerful therapeutic potential, and can provide a novel therapeutic strategy. They appear promising and safe (as EVs are non-self-replicating), and eventually MSC-derived EVs (MSC-EVs) may be developed to standardized, off-the-shelf allogeneic regenerative and immunomodulatory therapeutics. Promising pre-clinical data have been achieved using MSCs from different sources as EV-producing cells. Similarly, a variety EV isolation and characterization methods have been applied. Interestingly, MSC-EVs obtained from different sources and prepared with different methods show in vitro and in vivo therapeutic effects, indicating that isolated EVs share a common potential. Here, well-characterized and controlled, publicly available proteome profiles of MSC-EVs are compared to identify a common MSC-EV protein signature that might be coupled to the MSC-EVs' common therapeutic potential. This protein signature may be helpful in developing MSC-EV quality control platforms required to confirm the identity and test for the purity of potential therapeutic MSC-EVs.

A preliminary investigation of circulating extracellular vesicles and biomarker discovery associated with treatment response in head and neck squamous cell carcinoma

Abstract: There is a paucity of plasma-based biomarkers that prospectively segregate the outcome of patients with head and neck squamous-cell carcinoma (HNSCC) treated with chemoradiation therapy (CRT). Plasma extracellular vesicles (EVs) might be an alternative source for discovery of new specific markers present in patients with HNSCC, which could help to re-direct patients to appropriate curative therapies without delay. In order to identify new markers in plasma compartments, Cholerae toxin B chain (CTB) and Annexin V (AV) were used to isolate EVs from pooled plasma samples from patients with locally advanced HNSCC who responded (CR, n = 6) or presented incomplete response (NR, n = 6) to CRT. The crude plasma and EVs cargo were screened by antibody array. Of the 370 polypeptides detected, 119 proteins were specific to NR patients while 38 were exclusive of the CR subjects. The Gene Set Enrichment Analysis (GSEA) and Search Tool for the Retrieval of Interacting Genes (STRING) database analysis indicated that the content of circulating plasma EVs might have a relevant function for the tumor intercellular communication in the HNSCC patients. This study provides a list of potential markers present in plasma compartments that might contribute to the development of tools for prediction and assessment of CRT response and potentially guide therapeutic decisions in this context.

Circulating extracellular vesicle-associated TGFβ3 modulates response to cytotoxic therapy in head and neck squamous cell carcinoma

Abstract: Management of locally advanced head and neck squamous cell carcinoma (HNSCC) requires a multi-prong approach comprising surgery, radiation and/or chemotherapy, yet outcomes are limited. This is largely due to a paucity of biomarkers that can predict response to specific treatment modalities. Here, we evaluated TGFβ3 protein levels in extracellular vesicles (EVs) released by HNSCC cells as a predictor for response to chemoradiation therapy (CRT). To this end, specific EV-fractions were isolated from cell lines or HNSCC patient plasma, and TGFβ3 protein was quantified. In patients treated with CRT, TGFβ3 levels were found to be significantly higher in plasma EV-fractions or non-responders compared with responders. High levels of TGFβ3 levels in Annexin V-EVs were associated with the worst progression-free survival. In vitro experiments demonstrated that TGFβ3 silencing sensitized HNSCC cells to cytotoxic therapies, and this phenotype could be rescued by treatment with exogenous. In addition, specific EV-fractions shed by cisplatin-resistant cells were sufficient to transfer the resistant phenotype to sensitive cells through activation of TGFβ-signaling pathway. Therefore, our data show that TGFβ3 transmitted through EV plays a significant role in response to cytotoxic therapy, which can be exploited as a potential biomarker for CRT response in HNSCC patients treated with curative intent.

Membrane lipid binding molecules for the isolation of bona fide extracellular vesicle types and associated biomarkers in liquid biopsy

Abstract: Cancer exacts a heavy socioeconomic cost. Earlier detection and treatment are likely to mitigate this cost. Unfortunately, conventional tissue biopsy, the gold standard in cancer diagnosis cannot fulfill the goal of earlier detection. While liquid biopsy is a promising alternative to tissue biopsy, it has its challenges and limitations. A major challenge is the isolation of bona fide lipid membrane vesicles from biological fluids. In this review, we presented a new perspective of isolating different types of extracellular vesicles (EVs) by their affinity for membrane lipid binding ligands for liquid biopsy. EVs are lipid membrane particles naturally released by almost all cells and are found in almost all biological fluids suitable for liquid biopsy. They carry materials from the secreting cells that could affect the biology of the recipient cells and could thus inform on the state and progress of the disease. However, isolating bona fide EVs is a technical challenge as biological fluids have a complex composition and contain particles or aggregates that are physically similar to EVs. Here we review the use of membrane lipidbinding ligands to isolate different bona fide EV subtypes, and to circumvent the problem of co-isolating physically similar non-EV complexes in current EV isolation protocols. We will discuss the advantages of this technique and its potential for accelerated biomarker discovery and validation through examples of pre-clinical studies. We propose that isolating EV subtypes is a technically viable and robust strategy to overcome the current bottleneck of isolating EVs for liquid biopsy. Page 2 of 9 Lai et al . J Cancer Metastasis Treat 2019;5:65 I http://dx.doi.org/10.20517/2394-4722.2019.011

High expression of MLANA in the plasma of patients with head and neck squamous cell carcinoma as a predictor of tumor progression

Abstract: BACKGROUND There is a paucity of plasma-based biomarkers that predict outcome in patients with head and neck squamous cell carcinoma (HNSCC) treated with chemoradiation therapy (CRT). Here, we evaluate the prognostic potential of plasma Melanoma-Antigen Recognized by T-cells 1 (MLANA) in this setting. METHODS MLANA expression in HNSCC lines were evaluated by reverse transcription polymerase chain reaction, whereas plasma levels were quantified using ELISA in 48 patients with locally advanced HNSCC undergoing a phase 2 trial with CRT. RESULTS MLANA is expressed at variable levels in a panel of HNSCC lines. In plasma, levels were elevated in patients with tumor relapse compared to those without (P < .004); 73.9% of the patients expressing high plasma MLANA levels progressed with recurrent disease (P = .020). Multivariate analysis showed that plasma MLANA levels and tumor resectability were independent prognostic factors for progression free survival. CONCLUSION Plasma MLANA expression appears to be an effective noninvasive biomarker for outcomes in patients treated with CRT, and could potentially guide therapeutic decisions in this context.