Fig 2: Genetic alterations in SPOP and ERG are synthetic sick.a Distribution of genetic alterations in SPOP and ERG across 333 primary prostate cancers in TCGA database3,8,9. b 3D growth of mouse prostate epithelial organoids derived from C57BL/6 mice over-expressing the indicated SPOP and ERG species (bar represents 20 µm) (n = 3, technical replicates). Representative bright-field pictures and hematoxylin and eosin (H&E) stained sections are shown. c In vivo growth of VCaP xenografts over-expressing the indicated SPOP species in immune-compromised mice (each group, n = 10). d Immunoblot of VCaP cells overexpressing the indicated SPOP species and corresponding quantification of the indicated protein levels depicted as a heatmap. Protein expression changes were normalized to ß-ACTIN and Control cell line, (n = 2). e Tumor growth kinetics of xenografts established from LuCaP-147 PDX (SPOP-Y83C) stably overexpressing ?ERG or Control vector (each group, n = 10). Corresponding immunoblot and quantification are depicted as a heatmap. Protein expression changes were normalized to Vinculin (VCL) and Control cell line. f Dose–response curve of VCaP cells overexpressing the indicated SPOP species and treated with the ETS-inhibitor YK-4-279. All error bars, mean + s.e.m. P-values were determined by one-way ANOVA (b) or two-way ANOVA (c, e, f) with multiple comparisons and adjusted using Benjamini–Hochberg post-test ****P < 0.0001. Molecular weights are indicated in kilodaltons (kDa). Source data are provided as a Source Data File.

Fig 3: Schematic representation of the proposed model for the aversive relationship between mutant SPOP and ERG in prostate cancer.On the left side, SPOP mutant prostate cancer tumorigenesis is depicted. SPOP mutant tumors impair the degradation of substrate proteins such as AR activators (e.g, TRIM24) and ZMIND11, which ultimately triggers the AR pathway and dampens the ERG signaling. In this context, SPOP mutant tumors are more susceptible to ADT therapy. On the right side, TMPRSS2-ERG mutant tumorigenesis is depicted. ERG binds to the promoter of wild-type SPOP, upregulating its protein expression. Consequently, AR activators and ZMYND11 substrate proteins get degraded, leading to the AR pathway downregulation and unleashing the ERG signaling pathway. TMPRSS2-ERG mutant tumors are thus sensitive to high-dose androgen therapy or SPOP inhibition.

Fig 4: SPOP mutant tumors are particularly susceptible to androgen deprivation therapies (ADT).a Progression-free survival of prostate cancer patients derived from the TCGA-cohort. Curves representing TMPRSS2-ERG rearranged, and SPOP-mutant patients are indicated in violet and green, respectively. The area around the curves represents 80% confidence interval. The bar plot in the lower left corner indicates the percentage of SPOP-mutant tumors within all patients diagnosed with prostate cancer (DIAG) and within the individuals who developed a progression of the disease (PROG). P-values for Kaplan–Meier curves were determined using the log-rank test (P = 0.115). b Overall survival of prostate cancer patients derived from the MSK-IMPACT cohort. Curves representing TMPRSS2-ERG rearranged, and SPOP-mutant patients are indicated in violet and green, respectively. The area around the curves represents 80% confidence interval. The bar plot in the lower left corner indicates the percentage of SPOP-mutant tumors within all patients who were diagnosed with prostate cancer (DIAG), within individuals who developed a metastatic progression of the disease (PROG), and within individuals who developed castration-resistant prostate cancer (CRPC). P-values for Kaplan–Meier curves were determined using the log-rank test. (P = 0.247). c Enzalutamide sensitivity of LAPC4 cells overexpressing ΔERG or SPOP mutant species (Y87C, W131G). All error bars, mean + s.e.m. P-values were determined by unpaired, two-tailed Student’s t-test (c), NS, not significant.

Fig 5: Mutant SPOP-induced, androgen receptor signaling, antagonizes ERG activity.a Gene-set enrichment analysis of VCaP cells overexpressing SPOP mutant (SPOP-MTs; SPOP-Y87C, -F102C, -W131G) vs. SPOP-wild type (-WT), based on RNA-seq data. Experiments were performed using three replicates for each condition. Enrichments are determined on custom gene-sets of direct androgen receptor (AR) and ERG target genes (Supplemental Dataset 1). Enrichments and FDR-adjusted P-values are computed with Camera (pre-ranked) b Venn Diagram and heatmap depicting the expression of genes included in the custom gene-set of AR/ERG co-bound genes that are repressed by ERG and induced by DHT in VCaP cells overexpressing SPOP-MTs (SPOP-Y87C, F102C, W131G), SPOP-WT and vector Control. Genes (rows) and samples (columns) were clustered using Euclidean distance. Gene expression values were normalized using variance stabilizing transformation (vst) and subsequently scaled and centered by row prior of clustering. Columns represent the average expression of three replicates for each condition. c Two-dimensional network representing overlaps between the ten most significantly enriched Hallmark and custom gene-sets, identified when comparing SPOP-MTs (SPOP-Y87C, F102C, W131G) to SPOP-wild type (-WT) overexpressing VCaP cells. The thickness of edges is proportional to the significance of the overlap of the connected nodes measured by the Fisher test. Only edges with FDR value <0.05 are shown. The size of nodes is proportional to gene-set enrichment significance and equals to -10 x log10 (FDR). d Heatmap representing gene-set activity stratified according to tumor subtype, derived from TCGA cohort. For each tumor group, the average value of single-sample GSEA scores was considered. Values were scaled and referenced to samples that did not harbor any ETS-fusion (ERG, ETV1, and ETV4) or point mutations in SPOP3.