RGD-Coated Polymer Nanoworms for Enriching Cancer Stem Cells

SIMPLE SUMMARY: Cancer stem cells are a small portion of tumor cells, and it is also hard to keep their stem features during isolation and culture, thus making the study of them very difficult. However, they play a critical role in tumor development, drug resistance, and metastasis. Therefore, an effective method to isolate them and maintain their stem feature is required. In this study, we explored a new method with our synthesized thermo-responsive polymer coated with triple peptides RGD to meet the requirement. We demonstrate that, with this system, we can promote cancer stem cell growth and sustain their stem nature. Furthermore, the new system does not need enzyme treatment to dissociate cells. This method can thus be widely used to isolate and culture cancer stem cells from primary tumor or cancer cell lines to facilitate the study of identifying therapeutic targets and screening drugs sensitive to cancer stem cells. ABSTRACT: Cancer stem cells (CSCs) are primarily responsible for tumour drug resistance and metastasis; thus, targeting CSCs can be a promising approach to stop cancer recurrence. However, CSCs are small in numbers and readily differentiate into matured cancer cells, making the study of their biological features, including therapeutic targets, difficult. The use of three-dimensional (3D) culture systems to enrich CSCs has some limitations, including low sphere forming efficiency, enzymatic digestion that may damage surface proteins, and more importantly no means to sustain the stem properties. A responsive 3D polymer extracellular matrix (ECM) system coated with RGD was used to enrich CSCs, sustain stemness and avoid enzymatic dissociation. RGD was used as a targeting motif and a ligand to bind integrin receptors. We found that the system was able to increase sphere forming efficiency, promote the growth of spheric cells, and maintain stemness-associated properties compared to the current 3D culture. We showed that continuous culture for three generations of colon tumour spheroid led to the stem marker CD24 gradually increasing. Furthermore, the new system could enhance the cancer cell sphere forming ability for the difficult triple negative breast cancer cells, MBA-MD-231. The key stem gene expression for colon cancer also increased with the new system. Further studies indicated that the concentration of RGD, especially at high doses, could inhibit stemness. Taken together, our data demonstrate that our RGD-based ECM system can facilitate the enrichment of CSCs and now allow for the investigation of new therapeutic approaches for colorectal cancer or other cancers.