Dehm, Scott M

Abstract The roles of nucleoplasm-residing nucleoporins (NUP) in solid tumors, including prostate cancer, remain unknown. In this study, we reveal the clinical significance and mechanistic role of the off-pore NUP, soluble POM121 (sPOM121), as a crucial transcriptional regulator that enhances the aggressiveness of metastatic prostate cancer. Using orthogonal methodologies in human samples, sPOM121 was identified as the predominantly expressed nucleoplasmic NUP in prostate cancer. Unbiased proteomic and epigenomic studies demonstrate that sPOM121, through its C-terminus, interacts with the chromatin remodeler SMARCA5 at gene promoter sites and localizes at nuclear condensates, reprogramming gene expression. Indeed, sPOM121 regulates a distinct oncogenic gene network, including β-catenin, leading to prostate cancer progression and immune evasion. Importantly, targeting the sPOM121/β-catenin axis in patient-derived preclinical and syngeneic mouse models halts prostate cancer aggressiveness and enhances antitumor immunity. Taken together, these findings reveal previously unknown actionable reprogramming functions of off-pore NUPs in solid tumors. Significance: This study uncovers how oncogenic signaling programs are transcriptionally heightened by the NUP sPOM121 in metastatic prostate cancer. Localization of sPOM121 at active transcriptional nuclear condensates propels disease progression and immune evasion, offering novel anticancer therapeutic opportunities.

Abstract Purpose: Germline alterations in homologous recombination repair (gHRR) genes affect the pathogenesis, treatment options, and survival of patients with cancer. However, distinct gHRR gene alterations may differentially affect treatment response and oncogenic signaling. In this study, we interrogated genomic and transcriptomic data and assessed clinical outcomes of patients with gHRR mutations across four BRCA-associated cancers (breast, ovarian, pancreatic, and prostate cancers) to identify therapeutic vulnerabilities. Experimental Design: We assessed 24,309 patients undergoing matched tumor/normal next-generation DNA and RNA sequencing. Annotated gHRR gene variants [germline BRCA1, germline BRCA2, germline PALB2, germline ATM (gATM), and germline CHEK2] were analyzed. HRs were used to assess survival outcomes comparing germline versus sporadic groups. Somatic alterations and their frequencies were compared across gHRR-altered groups. Differential gene expression and gene set enrichment analysis were used to compare transcriptomic profiles. Results: Somatic TP53 mutations were depleted in gATM carriers (P < 0.05) across all four BRCA-associated cancers by up to 2.5-fold. Tumors with germline BRCA1/2 mutations were associated with improved survival in patients with ovarian cancer and had consistent enrichment of TP53 mutations in all four cancers. gATM mutations displayed elevated p53 transcriptional activity in all four cancers, with significance reached in breast and prostate cancers (P < 0.01). In breast, ovarian, and prostate cancers, gATM tumors demonstrated significantly increased inflammatory pathways (P < 0.001). Finally, using gene dependency data, we found that cell lines that were highly dependent on ATM were co-dependent on canonical p53 function. Conclusions: gATM-associated cancers seem to require intact p53 activity and this synthetic essentiality may be used to guide targeted therapies that perturb canonical TP53 function.

Abstract Purpose: Around 10% to 15% of prostate cancers harbor recurrent aberrations in the Forkhead Box A1 gene, FOXA1, whereby the alteration type and the effect on the forkhead (FKH) domain affect protein function. We developed a FOXA1 classification system to inform clinical management. Experimental Design: A total of 5,014 prostate cancer samples were examined using whole-exome and -transcriptome sequencing from the Caris Life Sciences database. We denoted class 1 FOXA1 alterations as missense and in-frame insertions/deletions with subclasses oriented with respect to the FKH domain. These were in the first part of the FKH domain [class 1A: amino acids (AA) 168–246], within the Wing2 region of FKH (class 1B: AA 247–269), or outside FKH (class 1C: AA 1–167, 270+). Two hotspot missense mutations at R219 were denoted class 2. Class 3 included predicted truncating mutations with subclasses partitioned based on the FKH domain (class 3A: AA 1–269 and class 3B: AA 270+). Class 4 represented FOXA1 amplifications. Real-world overall survival and therapy outcomes were determined from insurance claims. Results: FOXA1 alterations did not influence survival when considered in aggregate but had distinct prognostic effects when stratified by class. In primary prostate samples, class 1A alterations were associated with overall improved survival (HR, 0.57; P = 0.03); a similar trend was seen in metastatic biopsies with class 1B (HR, 0.84; P = 0.09). Conversely, in primary specimens, class 1C exhibited worse survival upon second-generation androgen receptor signaling inhibitor treatment (HR, 1.93; P < 0.001). Class 2 mutations (R219C/S) were enriched in neuroendocrine prostate cancers and were associated with overall poor survival (HR, 2.05; P < 0.001) and worse outcomes to first-line androgen-deprivation therapies (HR, 2.5; P < 0.001). Class 3A alterations indicated improved survival (HR, 0.70; P = 0.01), whereas class 3B alterations portended poor outcomes (HR, 1.50; P < 0.001). Amplifications (class 4) indicated poor outcomes in metastatic samples (HR, 1.48; P = 0.02). Molecularly, different FOXA1 alteration classes harbored distinct mutational and immunologic features as well as unique transcriptional programs. Finally, relative to European Americans, African Americans had increased class 1C alterations, whereas Asian/Pacific Islander patients had increased class 1B alterations. Conclusions: FOXA1 alterations should not be interpreted in aggregate, as different classes are associated with divergent molecular features and clinical outcomes. Our revised classification schema facilitates clinical decision-making for patients with prostate cancer and uncovers important racial differences.

Abstract B7-H3 (CD276) is a transmembrane glycoprotein of the B7 immune checkpoint superfamily that has emerged as a promising therapeutic target. To better understand the applicability of B7-H3–directed therapies, we analyzed 156,791 samples comprising 50 cancer types to interrogate the clinical, genomic, transcriptomic, and immunologic correlates of B7-H3 mRNA expression. DNA (592-gene/whole-exome) and RNA (whole-transcriptome) sequencing was performed from samples submitted to Caris Life Sciences. B7-H3 high versus low expression was based on top and bottom quartiles for each cancer type. Patients’ overall survival was determined from insurance claims data. Pathway analysis was performed using gene set enrichment analyses. Immune cell fractions were inferred using quanTIseq. B7-H3 is expressed across several human malignancies including prostate, pancreatic, ovarian, and lung cancers. High B7-H3 expression is associated with differences in overall survival, possibly indicating a prognostic role of B7-H3 for some cancers. When examining molecular features across all cancer types, we did not identify recurrent associations between B7-H3 expression and genetic alterations in TP53, RB1, and KRAS. However, we find consistent enrichment of epithelial-to-mesenchymal transition, Wnt, TGFβ, and Notch signaling pathways. In addition, tumors with high B7-H3 expression are associated with greater proportions of M1 macrophages, but lower fractions of CD8+ T cells. We have begun to define the genomic, transcriptomic, clinical, and immunologic features associated with B7-H3 expression in 50 cancer types. We report novel clinical and molecular features of B7-H3–high tumors which may inform how current B7-H3 therapeutics should be deployed and prioritized. Significance: B7-H3–targeting therapeutics have shown promising results in initial clinical trials. In this pan-cancer analysis of B7-H3 mRNA expression, we found that B7-H3 exhibits robust expression in many common cancer types. These results may inform further development of B7-H3–targeting therapeutics and may guide clinical decisions for patients with limited treatment options.

216 Background: PCa harbors recurrent FOXA1 alterations, yet distinct alteration types may drive divergent consequences. Here, we established several classes of FOXA1 alterations and evaluated their clinical outcomes, molecular correlates, and immunologic landscape in a large cohort of primary and metastatic PCa samples from Caris Life Sciences. Methods: We interrogated the Caris DNA (592-gene/whole exome) and RNA (whole transcriptome) NGS databases. FOXA1 alterations were annotated based on amino acid (a.a.) location and prior functional studies (Table). Real-world overall survival (OS) was determined from insurance claims data using Kaplan-Meier estimates. Gene set enrichment analyses (GSEA) was used to estimate signaling pathway activity. Immune-cell fractions were inferred from bulk RNA-seq using quanTIseq. All comparisons are made relative to FOXA1 WT cases. Results: The overall prevalence of FOXA1 alterations was 16%, with no enrichment in metastatic PCa samples (15.5%). Class 2 mutations were enriched in histologic NEPC, were associated with a classical NEPC mRNA signature, and showed poor survival with ADT (HR 5.6, 95% CI: 2.5–12.5, p<0.001). Conversely, metastatic PCa patients with Class 1B mutations had numerically improved survival with ADT (HR 0.76, 95% CI: 0.50–1.14, p=0.18). All FOXA1 alterations (except Class 2) resulted in increased FOXA1 transcripts, with 1B and 1C having the greatest effects (q<0.0001). Class 0 and 3 alterations had 2–5 fold higher frequency of TMB-high and MSI-high status (q<0.05). Conversely, Class 1B exhibited lower TMB and markedly fewer TP53 mutations (35.7% vs 20.3%, q<0.001) and TMPRSS2-ERG fusions (34.5% vs 4.6%, q<0.001). Class 1B was depleted of AR transcriptional signatures (q=0.01) but enriched of MYC signaling in GSEA analysis (FDR<0.001), and showed reduced expression of B7-H3. On immune-cell deconvolution analysis, Class 1B showed reduced dendritic cells, but increased B cells and neutrophils (DC: 1.96% vs 0.01%, B cell: 4.27% vs 5.29%, neutrophils: 5.05% vs 8.00%, each q<0.001). Conclusions: Different FOXA1 alterations exhibit divergent molecular and clinical features, and should not be interpreted in aggregate. In particular, Class 1B mutations are associated with a unique molecular and immunological landscape with potentially better outcomes to ADT, while Class 2 mutations are associated with NEPC phenotype with inferior ADT sensitivity. [Table: see text]

Abstract Purpose: Men with metastatic castration-resistant prostate cancer (mCRPC) frequently develop resistance to androgen receptor signaling inhibitor (ARSI) treatment; therefore, new therapies are needed. Trophoblastic cell-surface antigen (TROP-2) is a transmembrane protein identified in prostate cancer and overexpressed in multiple malignancies. TROP-2 is a therapeutic target for antibody–drug conjugates (ADC). Experimental Design: TROP-2 gene (TACSTD2) expression and markers of treatment resistance from prostate biopsies were analyzed using data from four previously curated cohorts of mCRPC (n = 634) and the PROMOTE study (dbGaP accession phs001141.v1.p1, n = 88). EPCAM or TROP-2–positive circulating tumor cells (CTC) were captured from peripheral blood for comparison of protein (n = 15) and gene expression signatures of treatment resistance (n = 40). We assessed the efficacy of TROP-2–targeting agents in a mouse xenograft model generated from prostate cancer cell lines. Results: We demonstrated that TACSTD2 is expressed in mCRPC from luminal and basal tumors but at lower levels in patients with neuroendocrine prostate cancer. Patients previously treated with ARSI showed no significant difference in TACSTD2 expression, whereas patients with detectable AR-V7 expression showed increased expression. We observed that TROP-2 can serve as a cell surface target for isolating CTCs, which may serve as a predictive biomarker for ADCs. We also demonstrated that prostate cancer cell line xenografts can be targeted specifically by labeled anti–TROP-2 agents in vivo. Conclusions: These results support further studies on TROP-2 as a therapeutic and diagnostic target for mCRPC.

Abstract We identified resistance mechanisms to abiraterone acetate/prednisone (AA/P) in patients with metastatic castration-resistant prostate cancer (mCRPC) in the Prostate Cancer Medically Optimized Genome-Enhanced Therapy (PROMOTE) study. We analyzed whole-exome sequencing (WES) and RNA-sequencing data from 83 patients with metastatic biopsies before (V1) and after 12 weeks of AA/P treatment (V2). Resistance was determined by time to treatment change (TTTC). At V2, 18 and 11 of 58 patients had either short-term (median 3.6 months; range 1.4–4.5) or long-term (median 29 months; range 23.5–41.7) responses, respectively. Nonresponders had low expression of TGFBR3 and increased activation of the Wnt pathway, cell cycle, upregulation of AR variants, both pre- and posttreatment, with further deletion of AR inhibitor CDK11B posttreatment. Deletion of androgen processing genes, HSD17B11, CYP19A1 were observed in nonresponders posttreatment. Genes involved in cell cycle, DNA repair, Wnt-signaling, and Aurora kinase pathways were differentially expressed between the responder and non-responder at V2. Activation of Wnt signaling in nonresponder and deactivation of MYC or its target genes in responders was detected via SCN loss, somatic mutations, and transcriptomics. Upregulation of genes in the AURKA pathway are consistent with the activation of MYC regulated genes in nonresponders. Several genes in the AKT1 axis had increased mutation rate in nonresponders. We also found evidence of resistance via PDCD1 overexpression in responders. Implications: Finally, we identified candidates drugs to reverse AA/P resistance: topoisomerase inhibitors and drugs targeting the cell cycle via the MYC/AURKA/AURKB/TOP2A and/or PI3K_AKT_MTOR pathways.

Abstract Purpose: Androgen receptor (AR) variant AR-V7 is a ligand-independent transcription factor that promotes prostate cancer resistance to AR-targeted therapies. Accordingly, efforts are under way to develop strategies for monitoring and inhibiting AR-V7 in castration-resistant prostate cancer (CRPC). The purpose of this study was to understand whether other AR variants may be coexpressed with AR-V7 and promote resistance to AR-targeted therapies. Experimental Design: We utilized complementary short- and long-read sequencing of intact AR mRNA isoforms to characterize AR expression in CRPC models. Coexpression of AR-V7 and AR-V9 mRNA in CRPC metastases and circulating tumor cells was assessed by RNA-seq and RT-PCR, respectively. Expression of AR-V9 protein in CRPC models was evaluated with polyclonal antisera. Multivariate analysis was performed to test whether AR variant mRNA expression in metastatic tissues was associated with a 12-week progression-free survival endpoint in a prospective clinical trial of 78 CRPC-stage patients initiating therapy with the androgen synthesis inhibitor, abiraterone acetate. Results: AR-V9 was frequently coexpressed with AR-V7. Both AR variant species were found to share a common 3′ terminal cryptic exon, which rendered AR-V9 susceptible to experimental manipulations that were previously thought to target AR-V7 uniquely. AR-V9 promoted ligand-independent growth of prostate cancer cells. High AR-V9 mRNA expression in CRPC metastases was predictive of primary resistance to abiraterone acetate (HR = 4.0; 95% confidence interval, 1.31–12.2; P = 0.02). Conclusions: AR-V9 may be an important component of therapeutic resistance in CRPC. Clin Cancer Res; 23(16); 4704–15. ©2017 AACR.

Recent therapeutic advances for managing advanced prostate cancer include the successful targeting of the androgen-AR axis with several new drugs in castrate resistant prostate cancer including abiraterone acetate and enzalutamide (MDV3100). This translational progress from “bench to bed-side” has resulted in an enlarging repertoire of novel and traditional drug choices now available for use in advanced prostate cancer therapeutics, which has had a positive clinical impact in prolonging longevity and quality of life of advanced prostate cancer patients. In order to further the clinical utility of these drugs, development of predictive biomarkers guiding individual therapeutic choices remains an ongoing challenge. This paper will summarize the potential in developing predictive biomarkers based on the pathophysiology of the androgen-AR axis in tumor tissue from patients with advanced prostate cancer as well as inherited variation in the patient’s genome. Specific examples of rational clinical trial designs incorporating potential predictive biomarkers from these pathways will illustrate several aspects of pharmacogenetic and pharmacogenomic predictive biomarker development in advanced prostate cancer therapeutics.

Abstract ETS variant 1 (ETV1), also known as ETS-related protein 81, is overexpressed in prostate tumors, but whether and how this transcription factor affects tumorigenesis has remained elusive. Here, we show that ETV1 is primarily overexpressed in the most aggressive human prostate tumors. Transgenic ETV1 mice developed prostatic intraepithelial neoplasia as well as hyperplasia/neoplasia in seminal vesicles. Moreover, ETV1 cooperated with the androgen receptor (AR) to bind to the prostate-specific antigen enhancer and stimulate gene transcription. Consistent with its ability to physically interact with AR, ETV1 rendered an ETV1 binding site–driven reporter androgen inducible, and, on the other hand, ETV1 superinduced transcription from an AR binding site on androgen stimulation. In conclusion, our study substantiates that ETV1 overexpression is an underlying cause in the development of prostate and possibly also seminal vesicle cancer. Its interaction with and activation of AR provides a molecular mechanism on how ETV1 exerts its deleterious function. Thus, inhibiting ETV1 or blocking its interaction with AR may represent novel strategies in prostate cancer therapy. [Cancer Res 2009;69(20):8102–10]