## 3 Results
Figure 4: Distinct interaction between the testosterone tetrad elicits androgenic signals that drive cell cycle progression, enhanced motility, cancer stemness, and resistance to therapy in patients with PCa. STRING-generated visualization of cancer stemness  
the protein-protein interaction between (**A**) HSD17B2, HSD17B3, SHBG, and SRD5A1, as well as with (**B**) SRD5A2, SRD5A3, HSD3B1, HSD3B2, and CYP17A1. (**C**) Molecular docking showing the direct interaction between HSD17B2 and HSD17B3, SRD5A1 and SHBG (upper left), as well as protein tetrad formation by the HSD17B2/HSD17B3 and SRD5A1/SHBG complexes (lower right). Heatmaps showing the correlation between expression profile of the testosterone tetrad and molecular effectors of (**D**) multidrug resistance, and (**E**) cancer stemness, in the GSE26910 mixed prostate and breast cancer cohort. Columns with similar annotations are collapsed by taking mean inside each group. Rows are centered; unit variance scaling is applied to rows. Both rows and columns are clustered using correlation distance and average linkage. (**F**) Heatmap showing the correlation between expression profile of the testosterone tetrad and ABCC1, ABCB1, ABCG2, ALDH1A1 expression in GSE33455 docetaxel-resistant PCa cell lines dataset. (**G**) Transcriptional profiling heatmap showing the correlation between the differential expression of the testosterone tetrad in human benign prostatic hyperplasia, primary and metastatic PCa samples from the GSE3325 PCa cohort. (**H**) Molecular docking showing the interaction cascade between the testosterone tetrad and molecular effectors or mediators of cell cycle progression, and drug uptake/therapy response. Figure 5: _Cont._  
Compared with PSA, the HSD17B2/HSD17B3/SHBG/SRD5A1 4-Gene Signature Is Capable of Differentiating Recurrent/Nonresponsive from Nonrecurrent/Respusive PCa  
Against the background of the diagnostic and prognostic relevance of PSA in urology practices, our statistical analysis of the TCGA PRAD cohort (n = 497) showed that, compared with selected clinicopathological variables, including PSA, the testosterone tetrad exhibited the largest effect size for prediction of therapy response or disease recurrence (Cohen's \(d\): HSD17B2 = 3.9, HSD17B3 = 7.0, SHBG = 11.9, SRD5A1 = 27.5 vs. PSA = 0.11) (Supplementary Table S1). In concordance, factor inclusion probability analysis using the Bayesian linear regression model showed that, while SRD5A1 (ENSG00000145545.10), SHBG (ENSG00000129214.13), HSD17B3 (ENSG00000130948.8), and HSD17B2 (ENSG00000086696.9) transcended the threshold for inclusion as predictors of recurrence or \"new tumor event after initial treatment\", PSA value and age at diagnosis did not (Figure 6A). In parallel analyses, using K-means clustering, patients from the  
Figure 5: Molecular fine-tuning of components of the 4-gene PCa signature modulate the highly proliferative, metastatic and cancer stem-cell-like (cum disease recurrent) phenotypes of PCa cells. **(A)** Heatmap of the differential expression of SHBG, HSD17B2, TWIST1, SLUG, SNAIL, VIM, TP53, SRD5A1, and GAPDH in the 22 Rv1, DU145, LNCaPAP, MDA-PCa 2b, NCI-H660, PC-3, and VCaP prostate cancer cell lines from the NIH GDC Cancer Cell Line Encyclopedia. **(B)** Line graph showing the effect of SHBG_OE on PC-3 cell proliferation over 8 days. **(C)** Photo-images (upper) and histograms (lower) of the effect of SHBG_OE on PC-3 cell viability on day 0 and day 8. **(D)** Photo-images (upper) and histograms (lower) of the effect of SHBG_OE on PC-3 cell migration or invasion. **(E)** Photo-images and histograms showing the differential effects of SHBG_OE or sgSRD5A1 on the formation of primary or secondary tomycetes by PC-3 or LNCaPAP cells. **(F)** Representative Western blot images of the differential effects of SHBG_OE or sgSRD5A1 on the expression levels of SHBG, SRD5A1, HSD17B2, HSD17B3, KLF4, OCT3/4, ABCC1, and ABCB1 in PC-3 cells. Original figure see Figure S1.