Clinically relevant humanized mouse models of metastatic prostate cancer facilitate therapeutic evaluation.

There is tremendous need for improved prostate cancer (PCa) models. The mouse prostate is anatomically and developmentally different from the human prostate and does not spontaneously form tumors. Genetically engineered mouse models lack the heterogeneity of human cancer and rarely establish metastatic growth. Human xenografts are an alternative but must rely on an immunocompromised host. Therefore, we generated PCa murine xenograft models with an intact human immune system (huNOG and huNOG-EXL mice) to test whether humanizing tumor-immune interactions would improve modeling of metastatic PCa and the impact of androgen receptor-targeted and immunotherapies. These mice maintain multiple human immune cell lineages, including functional human T-cells and myeloid cells. Implications: To our knowledge, results illustrate the first model of human PCa that has an intact human immune system, metastasizes to clinically relevant locations, responds appropriately to standard-of-care hormonal therapies, and can model both an immunosuppressive and checkpoint-inhibition responsive immune microenvironment.

ERBB3 Overexpression is Enriched in Diverse Patient Populations with Castration-sensitive Prostate Cancer and is Associated with a Unique AR Activity Signature.

PURPOSE: Despite successful clinical management of castration-sensitive prostate cancer (CSPC), the 5-year survival rate for men with castration-resistant prostate cancer is only 32%. Combination treatment strategies to prevent disease recurrence are increasing, albeit in biomarker-unselected patients. Identifying a biomarker in CSPC to stratify patients who will progress on standard-of-care therapy could guide therapeutic strategies. EXPERIMENTAL DESIGN: Targeted deep sequencing was performed for the University of Illinois (UI) cohort (n = 30), and immunostaining was performed on a patient tissue microarray (n = 149). Bioinformatic analyses identified pathways associated with biomarker overexpression (OE) in the UI cohort, consolidated RNA sequencing samples accessed from Database of Genotypes and Phenotypes (n = 664), and GSE209954 (n = 68). Neutralizing antibody patritumab and ectopic HER3 OE were utilized for functional mechanistic experiments. RESULTS: We identified ERBB3 OE in diverse patient populations with CSPC, where it was associated with advanced disease at diagnosis. Bioinformatic analyses showed a positive correlation between ERBB3 expression and the androgen response pathway despite low dihydrotestosterone and stable expression of androgen receptor (AR) transcript in Black/African American men. At the protein level, HER3 expression was negatively correlated with intraprostatic androgen in Black/African American men. Mechanistically, HER3 promoted enzalutamide resistance in prostate cancer cell line models and HER3-targeted therapy resensitized therapy-resistant prostate cancer cell lines to enzalutamide. CONCLUSIONS: In diverse patient populations with CSPC, ERBB3 OE was associated with high AR signaling despite low intraprostatic androgen. Mechanistic studies demonstrated a direct link between HER3 and enzalutamide resistance. ERBB3 OE as a biomarker could thus stratify patients for intensification of therapy in castration-sensitive disease, including targeting HER3 directly to improve sensitivity to AR-targeted therapies.

The Impact of Inherited Genetic Variation on DNA Methylation in Prostate Cancer and Benign Tissues of African American and European American Men.

BACKGROUND: American men of African ancestry (AA) have higher prostate cancer incidence and mortality rates compared with American men of European ancestry (EA). Differences in genetic susceptibility mechanisms may contribute to this disparity. METHODS: To gain insights into the regulatory mechanisms of prostate cancer susceptibility variants, we tested the association between SNPs and DNA methylation (DNAm) at nearby CpG sites across the genome in benign and cancer prostate tissue from 74 AA and 74 EA men. Genome-wide SNP data (from benign tissue) and DNAm were generated using Illumina arrays. RESULTS: Among AA men, we identified 6,298 and 2,641 cis-methylation QTLs (meQTL; FDR of 0.05) in benign and tumor tissue, respectively, with 6,960 and 1,700 detected in EA men. We leveraged genome-wide association study (GWAS) summary statistics to identify previously reported prostate cancer GWAS signals likely to share a common causal variant with a detected meQTL. We identified nine GWAS-meQTL pairs with strong evidence of colocalization (four in EA benign, three in EA tumor, two in AA benign, and three in AA tumor). Among these colocalized GWAS-meQTL pairs, we identified colocalizing expression quantitative trait loci (eQTL) impacting four eGenes with known roles in tumorigenesis. CONCLUSIONS: These findings highlight epigenetic regulatory mechanisms by which prostate cancer-risk SNPs can modify local DNAm and/or gene expression in prostate tissue. IMPACT: Overall, our findings showed general consistency in the meQTL landscape of AA and EA men, but meQTLs often differ by tissue type (normal vs. cancer). Ancestry-based linkage disequilibrium differences and lack of AA representation in GWAS decrease statistical power to detect colocalization for some regions.

Single-cell RNA sequencing of human prostate basal epithelial cells reveals zone-specific cellular populations and gene expression signatures.

Despite evidence of genetic signatures in normal tissue correlating with disease risk, prospectively identifying genetic drivers and cell types that underlie subsequent pathologies has historically been challenging. The human prostate is an ideal model to investigate this phenomenon because it is anatomically segregated into three glandular zones (central, peripheral, and transition) that develop differential pathologies: prostate cancer in the peripheral zone (PZ) and benign prostatic hyperplasia (BPH) in the transition zone (TZ), with the central zone (CZ) rarely developing disease. More specifically, prostatic basal cells have been implicated in differentiation and proliferation during prostate development and regeneration; however, the contribution of zonal variation and the critical role of basal cells in prostatic disease etiology are not well understood. Using single-cell RNA sequencing of primary prostate epithelial cultures, we elucidated organ-specific, zone-specific, and cluster-specific gene expression differences in basal cells isolated from human prostate and seminal vesicle (SV). Aggregated analysis identified ten distinct basal clusters by Uniform Manifold Approximation and Projection. Organ specificity compared gene expression in SV with the prostate. As expected, SV cells were distinct from prostate cells by clustering, gene expression, and pathway analysis. For prostate zone specificity, we identified two CZ-specific clusters, while the TZ and PZ populations clustered together. Despite these similarities, differential gene expression was identified between PZ and TZ samples that correlated with gene expression profiles in prostate cancer and BPH, respectively. Zone-specific profiles and cell type-specific markers were validated using immunostaining and bioinformatic analyses of publicly available RNA-seq datasets. Understanding the baseline differences at the organ, zonal, and cellular level provides important insight into the potential drivers of prostatic disease and guides the investigation of novel preventive or curative treatments. Importantly, this study identifies multiple prostate basal cell populations and cell type-specific gene signatures within prostate basal epithelial cells that have potential critical roles in driving prostatic diseases. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Glucocorticoid Receptor (GR) Activation Is Associated with Increased cAMP/PKA Signaling in Castration-Resistant Prostate Cancer.

In castration-resistant prostate cancer (CRPC), increased glucocorticoid receptor (GR) expression and ensuing transcriptional activity have been proposed as an oncogenic "bypass" mechanism in response to androgen receptor (AR) signaling inhibition (ARSi). Here, we report that GR transcriptional activity acquired following ARSi is associated with the upregulation of cyclic adenosine monophosphate (cAMP)-associated gene expression pathways in both model systems and metastatic prostate cancer patient samples. In the context of ARSi, the expression of GR-mediated genes encoding cAMP signaling pathway-associated proteins can be inhibited by treatment with selective GR modulators (SGRMs). For example, in the context of ARSi, we found that GR activation resulted in upregulation of protein kinase inhibitor beta (PKIB) mRNA and protein levels, leading to nuclear accumulation of the cAMP-dependent protein kinase A catalytic subunit (PKA-c). Increased PKA-c, in turn, is associated with increased cAMP response element-binding protein phosphorylation and activity. Furthermore, enzalutamide and SGRM combination therapy in mice bearing CRPC xenografts delayed CRPC progression compared with enzalutamide therapy alone, and reduced tumor PKIB mRNA expression. Supporting the clinical importance of GR/PKA signaling activation in CRPC, we found a significant enrichment of both cAMP pathway signaling-associated gene expression and high NR3C1 (GR) activity in patient-derived xenograft models and metastatic human CRPC samples. These findings suggest a novel mechanism linking CRPC-induced GR transcriptional activity with increased cAMP signaling in AR-antagonized CRPC. Furthermore, our findings suggest that GR-specific modulation in addition to AR antagonism may delay GR+ CRPC time to recurrence, at least in part, by inhibiting tumor cAMP/PKA pathways.

SOX2 expression in prostate cancer drives resistance to nuclear hormone receptor signaling inhibition through the WEE1/CDK1 signaling axis.

The development of androgen receptor signaling inhibitor (ARSI) drug resistance in prostate cancer (PC) remains therapeutically challenging. Our group has described the role of sex determining region Y-box 2 (SOX2) overexpression in ARSI-resistant PC. Continuing this work, we report that NR3C1, the gene encoding glucocorticoid receptor (GR), is a novel SOX2 target in PC, positively regulating its expression. Similar to ARSI treatment, SOX2-positive PC cells are insensitive to GR signaling inhibition using a GR modulating therapy. To understand SOX2-mediated nuclear hormone receptor signaling inhibitor (NHRSI) insensitivity, we performed RNA-seq in SOX2-positive and -negative PC cells following NHRSI treatment. RNA-seq prioritized differentially regulated genes mediating the cell cycle, including G2 checkpoint WEE1 Kinase (WEE1) and cyclin-dependent kinase 1 (CDK1). Additionally, WEE1 and CDK1 were differentially expressed in PC patient tumors dichotomized by high vs low SOX2 gene expression. Importantly, pharmacological targeting of WEE1 (WEE1i) in combination with an ARSI or GR modulator re-sensitizes SOX2-positive PC cells to nuclear hormone receptor signaling inhibition in vitro, and WEE1i combined with ARSI significantly slowed tumor growth in vivo. Collectively, our data suggest SOX2 predicts NHRSI resistance, and simultaneously indicates the addition of WEE1i to improve therapeutic efficacy of NHRSIs in SOX2-positive PC.

Regulation of Prostate Androgens by Megalin and 25-hydroxyvitamin D Status: Mechanism for High Prostate Androgens in African American Men.

Vitamin D deficiency is associated with an increased risk of prostate cancer mortality and is hypothesized to contribute to prostate cancer aggressiveness and disparities in African American populations. The prostate epithelium was recently shown to express megalin, an endocytic receptor that internalizes circulating globulin-bound hormones, which suggests regulation of intracellular prostate hormone levels. This contrasts with passive diffusion of hormones that is posited by the free hormone hypothesis. Here, we demonstrate that megalin imports testosterone bound to sex hormone-binding globulin into prostate cells. Prostatic loss of Lrp2 (megalin) in a mouse model resulted in reduced prostate testosterone and dihydrotestosterone levels. Megalin expression was regulated and suppressed by 25-hydroxyvitamin D (25D) in cell lines, patient-derived prostate epithelial cells, and prostate tissue explants. In patients, the relationships between hormones support this regulatory mechanism, as prostatic DHT levels are higher in African American men and are inversely correlated with serum 25D status. Megalin levels are reduced in localized prostate cancer by Gleason grade. Our findings suggest that the free hormone hypothesis should be revisited for testosterone and highlight the impact of vitamin D deficiency on prostate androgen levels, which is a known driver of prostate cancer. Thus, we revealed a mechanistic link between vitamin D and prostate cancer disparities observed in African Americans. Significance: These findings link vitamin D deficiency and the megalin protein to increased levels of prostate androgens, which may underpin the disparity in lethal prostate cancer in African America men.

The ß-Secretase 1 Enzyme as a Novel Therapeutic Target for Prostate Cancer.

Recent studies have demonstrated the association of APP and Aß with cancer, suggesting that BACE1 may play an important role in carcinogenesis. In the present study, we assessed BACE1's usefulness as a therapeutic target in prostate cancer (PCa). BACE1 expression was observed in human PCa tissue samples, patient-derived xenografts (PDX), human PCa xenograft tissue in nude mice, and transgenic adenocarcinoma of the mouse prostate (TRAMP) tissues by immunohistochemistry (IHC) analysis. Additionally, the downstream product of BACE1 activity, i.e., Aß1-42 expression, was also observed in these PCa tissues by IHC as well as by PET imaging in TRAMP mice. Furthermore, BACE1 gene expression and activity was confirmed in several established PCa cell lines (LNCaP, C4-2B-enzalutamide sensitive [S], C4-2B-enzalutamide resistant [R], 22Rv1-S, 22Rv1-R, PC3, DU145, and TRAMP-C1) by real-time PCR and fluorometric assay, respectively. Treatment with a pharmacological inhibitor of BACE1 (MK-8931) strongly reduced the proliferation of PCa cells in in vitro and in vivo models, analyzed by multiple assays (MTT, clonogenic, and trypan blue exclusion assays and IHC). Cell cycle analyses revealed an increase in the sub-G1 population and a significant modulation in other cell cycle stages (G1/S/G2/M) following MK-8931 treatment. Most importantly, in vivo administration of MK-8931 intraperitoneal (30 mg/kg) strongly inhibited TRAMP-C1 allograft growth in immunocompetent C57BL/6 mice (approximately 81% decrease, p = 0.019). Furthermore, analysis of tumor tissue using the prostate cancer-specific pathway array revealed the alteration of several genes involved in PCa growth and progression including Forkhead O1 (FOXO1). All together, these findings suggest BACE1 as a novel therapeutic target in advanced PCa.

Ectopic JAK-STAT activation enables the transition to a stem-like and multilineage state conferring AR-targeted therapy resistance.

Emerging evidence indicates that various cancers can gain resistance to targeted therapies by acquiring lineage plasticity. Although various genomic and transcriptomic aberrations correlate with lineage plasticity, the molecular mechanisms enabling the acquisition of lineage plasticity have not been fully elucidated. We reveal that Janus kinase (JAK)-signal transducer and activator of transcription (STAT) signaling is a crucial executor in promoting lineage plasticity-driven androgen receptor (AR)-targeted therapy resistance in prostate cancer. Importantly, ectopic JAK-STAT activation is specifically required for the resistance of stem-like subclones expressing multilineage transcriptional programs but not subclones exclusively expressing the neuroendocrine-like lineage program. Both genetic and pharmaceutical inhibition of JAK-STAT signaling resensitizes resistant tumors to AR-targeted therapy. Together, these results suggest that JAK-STAT are compelling therapeutic targets for overcoming lineage plasticity-driven AR-targeted therapy resistance.

SOX2 mediates metabolic reprogramming of prostate cancer cells.

New strategies are needed to predict and overcome metastatic progression and therapy resistance in prostate cancer. One potential clinical target is the stem cell transcription factor SOX2, which has a critical role in prostate development and cancer. We thus investigated the impact of SOX2 expression on patient outcomes and its function within prostate cancer cells. Analyses of SOX2 expression among a case-control cohort of 1028 annotated tumor specimens demonstrated that SOX2 expression confers a more rapid time to metastasis and decreased patient survival after biochemical recurrence. SOX2 ChIP-Seq analyses revealed SOX2-binding sites within prostate cancer cells which differ significantly from canonical embryonic SOX2 gene targets, and prostate-specific SOX2 gene targets are associated with multiple oncogenic pathways. Interestingly, phenotypic and gene expression analyses after CRISPR-mediated deletion of SOX2 in castration-resistant prostate cancer cells, as well as ectopic SOX2 expression in androgen-sensitive prostate cancer cells, demonstrated that SOX2 promotes changes in multiple metabolic pathways and metabolites. SOX2 expression in prostate cancer cell lines confers increased glycolysis and glycolytic capacity, as well as increased basal and maximal oxidative respiration and increased spare respiratory capacity. Further, SOX2 expression was associated with increased quantities of mitochondria, and metabolomic analyses revealed SOX2-associated changes in the metabolism of purines, pyrimidines, amino acids and sugars, and the pentose phosphate pathway. Analyses of SOX2 gene targets with central functions metabolism (CERK, ECHS1, HS6SDT1, LPCAT4, PFKP, SLC16A3, SLC46A1, and TST) document significant expression correlation with SOX2 among RNA-Seq datasets derived from patient tumors and metastases. These data support a key role for SOX2 in metabolic reprogramming of prostate cancer cells and reveal new mechanisms to understand how SOX2 enables metastatic progression, lineage plasticity, and therapy resistance. Further, our data suggest clinical opportunities to exploit SOX2 as a biomarker for staging and imaging, as well as a potential pharmacologic target.

Single-cell RNA-Seq identifies factors necessary for the regenerative phenotype of prostate luminal epithelial progenitors.

Benign prostate hyperplasia and prostate cancer are common diseases that involve the overgrowth of prostatic tissue. Although their pathologies and symptoms differ, both diseases show aberrant activation of prostate progenitor cell phenotypes in a tissue that should be relatively quiescent. This phenomenon prompts a need to better define the normal prostate progenitor cell phenotype and pursue the discovery of causal networks that could yield druggable targets to combat hyperplastic prostate diseases. We used single-cell (sc) RNA-Seq analysis to confirm the identity of a luminal progenitor cell population in both the hormonally intact and castrated mouse prostate. Using marker genes from our scRNA-Seq analysis, we identified factors necessary for the regeneration phenotype of prostate organoids derived from mice and humans in vitro. These data outline potential factors necessary for prostate regeneration and utilization of scRNA-Seq approaches for the identification of pharmacologic strategies targeting critical cell populations that drive prostate disease.

The role of the androgen receptor in prostate development and benign prostatic hyperplasia: A review.

Benign prostatic hyperplasia (BPH) is a benign enlargement of the prostate in which incidence increases linearly with age, beginning at about 50 years old. BPH is a significant source of morbidity in aging men by causing lower urinary tract symptoms and acute urinary retention. Unfortunately, the etiology of BPH incidence and progression is not clear. This review highlights the role of the androgen receptor (AR) in prostate development and the evidence for its involvement in BPH. The AR is essential for normal prostate development, and individuals with defective AR signaling, such as after castration, do not experience prostate enlargement with age. Furthermore, decreasing dihydrotestosterone availability through therapeutic targeting with 5α-reductase inhibitors diminishes AR activity and results in reduced prostate size and symptoms in some BPH patients. While there is some evidence that AR expression is elevated in certain cellular compartments, how exactly AR is involved in BPH progression has yet to be elucidated. It is possible that AR signaling within stromal cells alters intercellular signaling and a "reawakening" of the embryonic mesenchyme, loss of epithelial AR leads to changes in paracrine signaling interactions, and/or chronic inflammation aids in stromal or epithelial proliferation evident in BPH. Unfortunately, a subset of patients fails to respond to current medical approaches, forcing surgical treatment even though age or associated co-morbidities make surgery less attractive. Fundamentally, new therapeutic approaches to treat BPH are not currently forthcoming, so a more complete molecular understanding of BPH etiology is necessary to identify new treatment options.

Imaging Fibroblast Activation Protein Alpha Improves Diagnosis of Metastatic Prostate Cancer with Positron Emission Tomography.

PURPOSE: Metastatic castration-resistant prostate cancer (mCRPC) is a lethal, heterogeneous disease with few therapeutic strategies that significantly prolong survival. Innovative therapies for mCRPC are needed; however, the development of new therapies relies on accurate imaging to assess metastasis and monitor response. Standard imaging modalities for prostate cancer require improvement and there remains a need for selective and sensitive imaging probes that can be widely used in patients with mCRPC. EXPERIMENTAL DESIGN: We evaluated the transmembrane protease fibroblast activation protein alpha (FAP) as a targetable cell surface antigen for mCRPC. Genomic and IHC analyses were performed to investigate FAP expression in prostate cancer. Our FAP-targeted antibody imaging probe, [89Zr]Zr-B12 IgG, was evaluated by PET/CT imaging in preclinical prostate cancer models. RESULTS: Analysis of patient data documented FAP overexpression in metastatic disease across tumor subtypes. PET imaging with [89Zr]Zr-B12 IgG demonstrated high tumor uptake and long-term retention of the probe in the preclinical models examined. FAP-positive stroma tumor uptake of [89Zr]Zr-B12 IgG was 5-fold higher than the isotype control with mean %ID/cc of 34.13 ± 1.99 versus 6.12 ± 2.03 (n = 3/group; P = 0.0006) at 72 hours. Ex vivo biodistribution corroborated these results documenting rapid blood clearance by 24 hours and high tumor uptake of [89Zr]Zr-B12 IgG by 72 hours. CONCLUSIONS: Our study reveals FAP as a target for imaging the tumor microenvironment of prostate cancer. Validation of [89Zr]Zr-B12 IgG as a selective imaging probe for FAP-expressing tumors presents a new approach for noninvasive PET/CT imaging of mCRPC.

MEIS-mediated suppression of human prostate cancer growth and metastasis through HOXB13-dependent regulation of proteoglycans.

The molecular roles of HOX transcriptional activity in human prostate epithelial cells remain unclear, impeding the implementation of new treatment strategies for cancer prevention and therapy. MEIS proteins are transcription factors that bind and direct HOX protein activity. MEIS proteins are putative tumor suppressors that are frequently silenced in aggressive forms of prostate cancer. Here we show that MEIS1 expression is sufficient to decrease proliferation and metastasis of prostate cancer cells in vitro and in vivo murine xenograft models. HOXB13 deletion demonstrates that the tumor-suppressive activity of MEIS1 is dependent on HOXB13. Integration of ChIP-seq and RNA-seq data revealed direct and HOXB13-dependent regulation of proteoglycans including decorin (DCN) as a mechanism of MEIS1-driven tumor suppression. These results define and underscore the importance of MEIS1-HOXB13 transcriptional regulation in suppressing prostate cancer progression and provide a mechanistic framework for the investigation of HOXB13 mutants and oncogenic cofactors when MEIS1/2 are silenced.

Decisions regarding the treatment of patients with early-stage prostate cancer are often based on the risk that the cancer could grow and spread quickly. However, it is not always straightforward to predict how the cancer will behave. Studies from 2017 and 2018 found that samples of less aggressive prostate cancer have higher levels of a group of proteins called MEIS proteins. MEIS proteins help control the production of numerous other proteins, which could affect the behavior of prostate cancer cells in many ways. VanOpstall et al. ­ including some of the researchers that performed the 2017 and 2018 studies ­ have investigated how MEIS proteins affect prostate cancer. When prostate cancer cells are implanted into mice, they result in tumors. VanOpstall et al. found that tumors that produced MEIS proteins grew more slowly. Next, MEIS proteins were extracted from the prostate cancer cells and were found to interact with another protein called HOXB13, which regulates the activity of numerous genes. When the cells were genetically modified to prevent HOXB13 being produced, the protective effect of MEIS proteins was lost. MEIS proteins work with HOXB13 to regulate the production of several other proteins, in particular a protein called Decorin that can suppress tumors. When MEIS proteins and HOXB13 are present, the cell produces more Decorin and the tumors grow more slowly and are less likely to spread. VanOpstall et al. found that blocking Decorin production rendered MEIS proteins less able to slow the spread of prostate cancer. These results suggest that MEIS proteins and HOXB13 are needed to stop tumors from growing and spreading, and some of this ability is by prompting production of Decorin. This study explains how MEIS proteins can reduce prostate cancer growth, providing greater confidence in using them to determine whether aggressive treatment is needed. A greater understanding of this pathway for tumor suppression could also provide an opportunity for developing anti-cancer drugs.

Genetic Determinants of Reduced Arsenic Metabolism Efficiency in the 10q24.32 Region Are Associated With Reduced AS3MT Expression in Multiple Human Tissue Types.

Approximately 140 million people worldwide are exposed to inorganic arsenic through contaminated drinking water. Chronic exposure increases risk for cancers as well as cardiovascular, respiratory, and neurologic diseases. Arsenic metabolism involves the AS3MT (arsenic methyltransferase) gene, and arsenic metabolism efficiency (AME, measured as relative concentrations of arsenic metabolites in urine) varies among individuals. Inherited genetic variation in the 10q24.32 region, containing AS3MT, influences AME, but the mechanisms remain unclear. To better understand these mechanisms, we use tissue-specific expression data from GTEx (Genotype-tissue Expression project) to identify cis-eQTLs (expression quantitative trait loci) for AS3MT and other nearby genes. We combined these data with results from a genome-wide association study of AME using "colocalization analysis," to determine if 10q24.32 SNPs (single nucleotide polymorphisms) that affect AME also affect expression of AS3MT or nearby genes. These analyses identified cis-eQTLs for AS3MT in 38 tissue types. Colocalization results suggest that the casual variant represented by AME lead SNP rs4919690 impacts expression of AS3MT in 13 tissue types (> 80% probability). Our results suggest this causal SNP also regulates/coregulates expression of nearby genes: BORCS7 (43 tissues), NT5C2 (2 tissues), CYP17A1-AS1 (1 tissue), and RP11-724N1.1 (1 tissue). The rs4919690 allele associated with decreased AME is associated with decreased expression of AS3MT (and other coregulated genes). Our study provides a potential biological mechanism for the association between 10q24.32 variation and AME and suggests that the causal variant, represented by rs4919690, may impact AME (as measured in urine) through its effects on arsenic metabolism occurring in multiple tissue types.

Syntaxin 6-mediated exosome secretion regulates enzalutamide resistance in prostate cancer.

Prostate cancer (PCa) deaths are typically the result of metastatic castration-resistant PCa (mCRPC). Recently, enzalutamide (Enz), an oral androgen receptor inhibitor, was approved for treating patients with mCRPC. Invariably, all PCa patients eventually develop resistance against Enz. Therefore, novel strategies aimed at overcoming Enz resistance are needed to improve the survival of PCa patients. The role of exosomes in drug resistance has not been fully elucidated in PCa. Therefore, we set out to better understand the exosome's role in the mechanism underlying Enz-resistant PCa. Results showed that Enz-resistant PCa cells (C4-2B, CWR-R1, and LNCaP) secreted significantly higher amounts of exosomes (2-4 folds) compared to Enz-sensitive counterparts. Inhibition of exosome biogenesis in resistant cells by GW4869 and dimethyl amiloride strongly decreased their cell viability. Mechanistic studies revealed upregulation of syntaxin 6 as well as its increased colocalization with CD63 in Enz-resistant PCa cells compared to Enz-sensitive cells. Syntaxin 6 knockdown by specific small interfering RNAs in Enz-resistant PCa cells (C4-2B and CWR-R1) resulted in reduced cell number and increased cell death in the presence of Enz. Furthermore, syntaxin 6 knockdown significantly reduced the exosome secretion in both Enz-resistant C4-2B and CWR-R1 cells. The Cancer Genome Atlas analysis showed increased syntaxin 6 expressions associated with higher Gleason score and decreased progression-free survival in PCa patients. Importantly, IHC analysis showed higher syntaxin 6 expression in cancer tissues from Enz-treated patients compared to Enz naïve patients. Overall, syntaxin 6 plays an important role in the secretion of exosomes and increased survival of Enz-resistant PCa cells.

PEG10 Promoter-Driven Expression of Reporter Genes Enables Molecular Imaging of Lethal Prostate Cancer.

The retrotransposon-derived paternally expressed gene 10 (PEG10) protein is ordinarily expressed at high levels in the placenta. Recently, it was discovered that PEG10 isoforms promote the progression of prostate cancer to a highly lethal androgen receptor (AR)-negative phenotype. The presence of PEG10 in other subtypes of prostate cancer has not been explored and a utility for PEG10 overexpression has not been developed. Here, we found that in addition to AR-null disease, PEG10 was also expressed in prostate cancer with constitutively active AR-splice variants. A molecular genetic imaging strategy for noninvasive imaging of AR-splice variant prostate cancer was developed by utilizing the cancer specificity of the PEG10 promoter to drive the expression of reporter genes. Plasmid insertion of a PEG10 promoter sequence optimized for enhanced output upstream of a reporter gene allowed detection of prostate cancer by near-infrared and positron emission tomography imaging after systemic administration of the plasmid in vivo. PEG10 expressing subcutaneous xenograft and intratibial tumor models were imaged by both modalities using this molecular genetic imaging strategy. This study demonstrates a preclinical proof-of-concept that the PEG10 promoter is a powerful and specific tool that can be utilized for noninvasive detection of aggressive prostate cancer subtypes. SIGNIFICANCE: PEG10 is expressed by prostate cancer with constitutively active AR-splice variants that can be exploited for noninvasive molecular imaging of this aggressive prostate cancer subytpe.