Anti-CD44 monoclonal antibody, clone LN92

Emerging evidence has reported that dysregulation of microRNAs (miRNAs) participated in the development of diverse types of cancers. Our initial microarray-based analysis identified differentially expressed NEK2 related to breast cancer and predicted the regulatory microRNA-128-3p (miR-128-3p). Herein, this study aimed to characterize the tumour-suppressive role of miR-128-3p in regulating the biological characteristics of breast cancer stem cells (BCSCs). CD44(& xff0b;)CD24(-/low)cells were selected for subsequent experiments. After verification of the target relationship between miR-128-3p and NEK2, the relationship among miR-128-3p, NEK2 and BCSCs was further investigated with the involvement of the Wnt signalling pathway. The regulatory effects of miR-128-3p on proliferation, migration, invasion and self-renewal in vitro as well as tumorigenicity in vivo of BCSCs were examined via gain- and loss-of-function approaches. Highly expressed NEK2 was found in breast cancer based on GSE61304 expression profile. Breast cancer stem cells and breast cancer cells showed a down-regulation of miR-128-3p. Overexpression of miR-128-3p was found to inhibit proliferation, migration, invasion, self-renewal in vitro and tumorigenicity in vivo of BCSCs, which was further validated to be achieved through inhibition of Wnt signalling pathway by down-regulating NEK2. In summary, this study indicates that miR-128-3p inhibits the stem-like cell features of BCSCs via inhibition of the Wnt signalling pathway by down-regulating NEK2, which provides a new target for breast cancer treatment.

De novo expressed CD44 isoforms containing exon-v6 are frequently associated with gastric cancer (GC) aggressiveness, and may predict chemotherapy response in vitro. Whether exon-v6 itself is responsible for conferring these properties to CD44v6-containing isoforms remains to be elucidated. CRISPR/Cas9 and Phosphorodiamidate Morpholino oligomers (PMOs) were used to induce specific exon-v6 skipping, maintaining the CD44 reading frame, in two GC cell lines endogenously expressing CD44v6. Cisplatin and 5-fluorouracil treatment response, and self-renewal ability was compared between CRISPR/Cas9-edited, CD44v6 knockdown and mock cells. We obtained homozygous genome-edited cell lines with exon-v6 deletion. Edited cells transcribed CD44v isoforms presenting in frame v5-v7 splicing, mimicking exon-v6 skipping. Results showed that removing specifically exon-v6 sensitizes cells to cisplatin and impairs cells' self-renewal ability, similarly to CD44v6 knockdown. In parallel, we also tested a clinically feasible approach for transient exon-v6 skipping with a PMO-based strategy. We demonstrate that exon-v6 specific removal from CD44v isoforms increases cell sensitivity to cisplatin and impairs GC cells self-renewal. We trust that a PMO approach designed towards CD44v6 overexpressing GC cells may be a suitable approach to sensitize tumor cells for conventional therapy.