Fig 2: Cell proliferation assay in response to enzalutamide treatment combined with SNRNP200, SRRM1 and SRSF3 silencing. Proliferation rate of 22Rv1 (a) and LNCaP (b) cell line was measured after 24 h of SNRNP200-, SRRM1- and SRSF3-silencing in the presence (DHT) or absence (no DHT) of 5a-dihydrotestosterone with or without enzalutamide (ENZA; n = 4). Results were expressed as percentage referred to scramble vehicle-treated control with DHT (mean ± SEM). Asterisks (* p < 0.05; ** p < 0.01), dash (#p < 0.05) and dollar sign ($p < 0.05) indicate statistically significant differences compared to DHT of scramble, DHT of each condition and DHT+ENZA of scramble, respectively.

Fig 3: Functional consequences of SNRNP200, SRRM1 and SRSF3 silencing in prostate-derived cell lines. a) Comparison of SNRNP200, SRRM1 and SRSF3 expression levels between a non-tumor prostate cell line (PNT2) and PCa cell lines LNCaP, 22Rv1, DU145 and PC-3 (n = 5). mRNA levels were determined by qPCR and adjusted by normalization factor calculated from ACTB and GAPDH expression levels. b) Validation by qPCR of SNRNP200, SRRM1 and SRSF3 silencing (si-SNRNP200, si-SRRM1 and si-SRSF3, respectively). mRNA levels were determined by qPCR and adjusted by normalization factor calculated from ACTB and GAPDH expression levels. Data were represented as percent of scramble cells (mean ± SEM). c) Proliferation rate of LNCaP (upper panel), 22Rv1 (middlepanel) and DU145 (bottom panel) cell lines after 24-, 48- and 72 h of SNRNP200-, SRRM1- and SRSF3-silencing (n = 4). d) Effect of SNRNP200-, SRRM1- and SRSF3-silencing on the migration rate of DU145 cell line was determined by wound-healing assay (12 h; n = 3). Representative images are depicted in right panel. Data were represented as percent of scramble cells (mean ± SEM). Asterisks (* p < 0.05; ** p < 0.01; *** p < 0.001) indicate statistically significant differences between groups.

Fig 4: Immunohistochemical analysis of SNRNP200,SRRM1 and SRSF3 in prostate cancer (PCa) samples. a) Comparison of SNRNP200,SRRM1 (b) and SRSF3 (c) protein levels by immunohistochemistry (IHC) between a representative set of PCa samples (n = 47), prostatic intraepithelial neoplasia (PIN; n = 6) and benign prostatic hyperplasia (BPH; n = 7). Association of protein levels with clinically significant PCa (SigPCa; defined as Gleason score higher than 7) and the presence of metastasis at diagnosis (central panel and right panel, respectively). Representative images of BPH, PIN, PCa with Gleason score = 6 and SigPCa stained with SNRNP200 (400× magnification), SRRM1 (400× magnification) and SRSF3 (200× magnification) antibodies are showed below a, b and c panels, respectively. Scale bar indicates 100 µm. Data are expressed as mean ± SEM of IHC staining scaled from low [1] to high [3] intensity. Asterisks (* p < 0.05; ** p < 0.01; *** p < 0.001) indicate statistically significant differences between groups.

Fig 5: Expression of SNRNP200, SRRM1 and SRSF3 in a highly aggressive cohort of prostate cancer (PCa) samples. (a) SNRNP200, SRRM1 and SRSF3 expression levels in a battery of highly-aggressive PCa samples, with or without the presence of metastasis (n = 42). represent the mean ± SEM of mRNA expression levels. b-f) Correlation between SNRNP200, SRRM1 and SRSF3 expression levels and Gleason score (b), expression levels of AR-v7 (c), AR (d) MKI67 (e) and KLK3 (f). mRNA levels were determined by a microfluidic-based qPCR array and adjusted by normalization factor calculated from ACTB and GAPDH expression levels. Asterisk (* p < 0.05) indicates statistically significant differences between groups.