Ve NEPC transdifferentiation. These Clemizole In Vivo studies could result in valuable insights towards identifying novel therapeutic targets for the remedy from the disease progression from CRPC to NEPC. Primarily based on our previous study [26], we discovered the loss-of-function research as excellent for blocking TBX2, and therefore, we applied this approach for the present study. Further, the repressive function of TBX2 within this study is in line with previous reports which have focused on TBX2 repression of its effectors [42,44,45]. Although the TBX2 protein consists of each the activation and repression domains, TBX2 has predominantly been reported to function as a transcriptional repressor [44]. The TBX2 DN mutant construct contains the T-box DNA-binding domain but lacks the carboxy-terminal residues vital for transcriptional repression [50]–thereby generating it a perfect approach to specifically investigate transcriptional repression. Furthermore, previous reports such as our study had identified that TBX2DN functions in congruence with all the RNA interference approach and upregulates p21, a known TBX2 Momelotinib Autophagy target [26,45,50]. Lastly, though the present study was focused around the role of exosomal miR-200c3p in advertising the NEPC phenotype in between neighboring PCa cells, in our orthotopic mouse model of PCa metastasis, we observed improved expression of miR-200c-3p in the human TBX2DN PCa xenografts that display abrogated metastatic capacity for the lymph nodes (compared with Neo controls) (Figure 3E). This opens up an intriguing query if TBX2/miR-200c-3p/SOX2/N-MYC signaling could potentially drive metastatic manifestation at the secondary websites via exosomal transfer. The insights offered by these investigations could offer additional clues in to the NEPC transdifferentiation puzzle especially in lieu of our earlier report that delineated the role of TBX2 in many facets of PCa progression including distant metastasis [26]. As research on the clinical challenges posed by potent ADTs is garnering increasing recognition, the emphasis on discovering crucial drivers of t-NEPC/NEPC transdifferentiation is gaining momentum, and the list of crucial drivers keeps growing [63]. The objective of these research such as ours is to boost PCa therapy by means of advancing our understanding from the molecular effectors/signaling pathways that orchestrate t-NEPC/NEPC transdifferentiation as a mechanism of acquired therapeutic resistance. 5. Conclusions Our study has identified a novel mechanism wherein TBX2 drives NEPC transdifferentiation via miR-200c-3p/SOX2/N-MYC signaling. Additional, our investigations point to good correlations amongst TBX2 and SOX2/N-MYC expression in human PCa patient samples. Our findings may perhaps pave the way for the development of novel and helpful therapeutic methods against the progression from CRPC to NEPC by way of targeting the TBX2/miR-200c-3p/SOX2/N-MYC axis.Cancers 2021, 13,15 ofSupplementary Materials: The following data is accessible on-line at https://www.mdpi.com/ article/10.3390/cancers13195020/s1, Figure S1: Larger extracellular vesicles [such as apoptotic bodies (ABs), microvesicles (MVs)] or soluble aspects (SFs) did not influence the expression of neuroendocrine markers in LNCaP cells, Figure S2: Larger extracellular vesicles [such as apoptotic bodies (ABs), microvesicles (MVs)] or soluble components (SFs) didn’t influence the expression of neuroendocrine markers in 22Rv1 cells, Figure S3: Magnified image of Figure 2C, Figure S4: Densitometric analysis of the Western blot image.