Although the incidence of de novo neuroendocrine prostate cancer (PC) is rare, recent data suggest that low expression of prostate-specific membrane antigen (PSMA) is associated with a spectrum of ...neuroendocrine hallmarks and androgen receptor (AR) suppression in PC. Previous clinical reports indicate that PCs with a phenotype similar to neuroendocrine tumors can be more amenable to imaging by
F-FDG than by PSMA-targeting radioligands. In this study, we evaluated the association between neuroendocrine gene signature and
F-FDG uptake-associated genes including glucose transporters (GLUTs) and hexokinases, with the goal of providing a genomic signature to explain the reported
F-FDG avidity of PSMA-suppressed tumors.
Data-mining approaches, cell lines, and patient-derived xenograft models were used to study the levels of 14 members of the
family (encoding GLUT proteins), 4 members of the hexokinase family (genes
-
and
), and PSMA (
gene) after AR inhibition and in correlation with neuroendocrine hallmarks. Also, we characterize a neuroendocrine-like PC (NELPC) subset among a cohort of primary and metastatic PC samples with no neuroendocrine histopathology. We measured glucose uptake in a neuroendocrine-induced in vitro model and a zebrafish model by nonradioactive imaging of glucose uptake using a fluorescent glucose bioprobe, GB2-Cy3.
This work demonstrated that a neuroendocrine gene signature associates with differential expression of genes encoding GLUT and hexokinase proteins. In NELPC, elevated expression of
(encoding glucokinase protein) and decreased expression of
correlated with earlier biochemical recurrence. In tumors treated with AR inhibitors, high expression of
and low expression of
correlated with neuroendocrine histopathology and PSMA gene suppression. GLUT12 suppression and upregulation of glucokinase were observed in neuroendocrine-induced PC cell lines and patient-derived xenograft models. A higher glucose uptake was confirmed in low-PSMA tumors using a GB2-Cy3 probe in a zebrafish model.
A neuroendocrine gene signature in neuroendocrine PC and NELPC associates with a distinct transcriptional profile of GLUTs and hexokinases. PSMA suppression correlates with GLUT12 suppression and glucokinase upregulation. Alteration of
F-FDG uptake-associated genes correlated positively with higher glucose uptake in AR- and PSMA-suppressed tumors. Zebrafish xenograft tumor models are an accurate and efficient preclinical method for monitoring nonradioactive glucose uptake.
Androgen deprivation therapy improves the survival of castration-resistant prostate cancer (CRPC) patients, yet ultimately fails with debilitating side effects. Supraphysiological testosterone ...(SPT)-based therapy produces clinical responses with improved quality of life in a subset of patients. Currently, no information defines a durable response to SPT.
To identify key molecular phenotypes underlying SPT response to improve patient selection and guide combination treatment to achieve a durable response.
A patient-derived xenograft (PDX) preclinical trial was performed with 13 CRPC PDXs to identify molecular features associated with SPT response. Comprehensive intratumoral androgen, tumor growth, and integrated transcriptomic and protein analyses were performed in three PDXs resistant to the newer androgen receptor (AR) pathway inhibitor enzalutamide (ENZ) to define SPT response and resistance.
Testosterone cypionate.
SPT efficacy was evaluated by PDX growth, prostate-specific antigen (PSA) change, and survival. Intratumoral androgens were analyzed using mass spectrometry. Global transcriptome analysis was performed using RNA sequencing, and confirmed by quantitative real-time polymerase chain reaction and immunohistochemistry. Log-rank and Mann-Whitney tests were used for survival and molecular analyses, respectively.
A durable SPT responder was identified, presenting robust repressions of ARv7 and E2F transcriptional outputs, and a DNA damage response (DDR) transcriptomic program that were altogether restored upon SPT resistance in the transient responder. ENZ rechallenge of SPT-relapsed PDXs resulted in PSA decreases but tumor progression.
SPT produces a durable response in AR-pathway inhibitor ENZ CRPC that is associated with sustained suppression of ARv7 and E2F transcriptional outputs, and the DDR transcriptome, highlighting the potential of combination treatments that maintain suppression of these programs to drive a durable response to SPT.
Patients with ENZ-resistant prostate cancer have very limited treatment options. Supraphysiological testosterone presents a prominent option for improved quality of life and a potential durable response in patients with sustained suppression on ARv7/E2F transcriptional outputs and DNA repair program.
Supraphysiological testosterone (SPT) produces clinical and quality-of-life benefits in subsets of castration-resistant prostate cancer patients. Molecular characteristics associated with a response have not been identified yet. This study indicated sustained repressions of ARv7 and E2F transcriptional outputs, and the DNA damage response program as hallmarks of a durable SPT response.
The androgen receptor (AR) pathway regulates key cell survival programs in prostate epithelium. The AR represents a near-universal driver and therapeutic vulnerability in metastatic prostate cancer, ...and targeting AR has a remarkable therapeutic index. Though most approaches directed toward AR focus on inhibiting AR signaling, laboratory and now clinical data have shown that high dose, supraphysiological androgen treatment (SPA) results in growth repression and improved outcomes in subsets of prostate cancer patients. A better understanding of the mechanisms contributing to SPA response and resistance could help guide patient selection and combination therapies to improve efficacy. To characterize SPA signaling, we integrated metrics of gene expression changes induced by SPA together with cistrome data and protein-interactomes. These analyses indicated that the Dimerization partner, RB-like, E2F and Multi-vulval class B (DREAM) complex mediates growth repression and downregulation of E2F targets in response to SPA. Notably, prostate cancers with complete genomic loss of RB1 responded to SPA treatment whereas loss of DREAM complex components such as RBL1/2 promoted resistance. Overexpression of MYC resulted in complete resistance to SPA and attenuated the SPA/AR-mediated repression of E2F target genes. These findings support a model of SPA-mediated growth repression that relies on the negative regulation of MYC by AR leading to repression of E2F1 signaling via the DREAM complex. The integrity of MYC signaling and DREAM complex assembly may consequently serve as determinants of SPA responses and as pathways mediating SPA resistance.
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
. 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.
Prostate cancer (PC) is driven by androgen receptor (AR) activity, a master regulator of prostate development and homeostasis. Frontline therapies for metastatic PC deprive the AR of the activating ...ligands testosterone (T) and dihydrotestosterone (DHT) by limiting their biosynthesis or blocking AR binding. Notably, AR signaling is dichotomous, inducing growth at lower activity levels, while suppressing growth at higher levels. Recent clinical studies have exploited this effect by administration of supraphysiological concentrations of T, resulting in clinical responses and improvements in quality of life. However, the use of T as a therapeutic agent in oncology is limited by poor drug-like properties as well as rapid and variable metabolism. Here, we investigated the antitumor effects of selective AR modulators (SARMs), which are small-molecule nonsteroidal AR agonists developed to treat muscle wasting and cachexia. Several orally administered SARMs activated the AR program in PC models. AR cistromes regulated by steroidal androgens and SARMs were superimposable. Coregulatory proteins including HOXB13 and GRHL2 comprised AR complexes assembled by both androgens and SARMs. At bioavailable concentrations, SARMs repressed MYC oncoprotein expression and inhibited the growth of castration-sensitive and castration-resistant PC in vitro and in vivo. These results support further clinical investigation of SARMs for treating advanced PC.
The incidence of prostate cancer is directly linked to age, but age-associated changes that facilitate prostate cancer development and progression are poorly understood. This study investigated ...age-related changes in the prostate microenvironment for their influence on prostate cancer behavior. Prostate cancer cells implanted orthotopically into the prostate demonstrated accelerated tumor growth in aged compared with young mice. Metastatic lesions following intravenous injection were also more numerous in aged mice. Tumors from young and aged mice showed no significant differences concerning their proliferation index, apoptosis, or angiogenesis. However, analysis of tumor-infiltrating immune cells by IHC and RNA sequencing (RNA-seq) revealed elevated numbers of macrophages in prostates from aged mice, which are quickly polarized towards a phenotype resembling protumorigenic tumor-associated macrophages upon tumor cell engraftment. Older patients with prostate cancer (>60 years old) in The Cancer Genome Atlas Prostate Adenocarcinoma (TCGA-PRAD) dataset displayed higher expression of macrophage markers (CD163 and VSIG4) which associated with higher rates of biochemical relapse. Remodeling of the collagenous extracellular matrix (ECM) was associated with prostate cancer growth and invasion in the aged microenvironment. Moreover, the collagen matrix extracted from aged mice enhanced the invasiveness and proliferation of prostate cancer cells
. Together, these results demonstrate that the aged prostatic microenvironment can regulate the growth and metastasis of malignant prostate cells, highlighting the role of resident macrophages and their polarization towards a protumorigenic phenotype, along with remodeling of the ECM. IMPLICATIONS: These findings demonstrate the importance of age-associated tumor microenvironment alterations in regulating key aspects of prostate cancer progression.
Neuroendocrine (NE) differentiation in metastatic castration-resistant prostate cancer (mCRPC) is an increasingly common clinical feature arising from cellular plasticity. We recently characterized ...two mCRPC phenotypes with NE features: androgen receptor (AR)-positive NE-positive amphicrine prostate cancer (AMPC) and AR-negative small cell or neuroendocrine prostate cancer (SCNPC). Here, we interrogated the regulation of RE1-silencing transcription factor (REST), a transcriptional repressor of neuronal genes, and elucidated molecular programs driving AMPC and SCNPC biology. Analysis of prostate cancer cell lines, mCRPC specimens, and LuCaP patient-derived xenograft models detected alternative splicing of REST to REST4 and attenuated REST repressor activity in AMPC and SCNPC. The REST locus was also hypermethylated and REST expression was reduced in SCNPC. While serine/arginine repetitive matrix protein 4 (SRRM4) was previously implicated in alternative splicing of REST in mCRPC, we detected SRRM3 expression in REST4-positive, SRRM4-negative AMPC, and SCNPC. In CRPC cell lines, SRRM3 induced alternative splicing of REST to REST4 and exacerbated the expression of REST-repressed genes. Furthermore, SRRM3 and SRRM4 expression defined molecular subsets of AMPC and SCNPC across species and tumor types. Two AMPC phenotypes and three SCNPC phenotypes were characterized, denoted either by REST attenuation and ASCL1 activity or by progressive activation of neuronal transcription factor programs, respectively. These results nominate SRRM3 as the principal REST splicing factor expressed in early NE differentiation and provide a framework to molecularly classify diverse NE phenotypes in mCRPC. SIGNIFICANCE: This study identifies SRRM3 as a key inducer of cellular plasticity in prostate cancer with neuroendocrine features and delineates distinct neuroendocrine phenotypes to inform therapeutic development and precision medicine applications.
The androgen receptor (AR) is a driver of cellular differentiation and prostate cancer development. An extensive body of work has linked these normal and aberrant cellular processes to mRNA ...transcription; however, the extent to which AR regulates posttranscriptional gene regulation remains unknown. Here, we demonstrate that AR uses the translation machinery to shape the cellular proteome. We show that AR is a negative regulator of protein synthesis and identify an unexpected relationship between AR and the process of translation initiation in vivo. This is mediated through direct transcriptional control of the translation inhibitor 4EBP1. We demonstrate that lowering AR abundance increases the assembly of the eIF4F translation initiation complex, which drives enhanced tumor cell proliferation. Furthermore, we uncover a network of pro-proliferation mRNAs characterized by a guanine-rich cis-regulatory element that is particularly sensitive to eIF4F hyperactivity. Using both genetic and pharmacologic methods, we demonstrate that dissociation of the eIF4F complex reverses the proliferation program, resulting in decreased tumor growth and improved survival in preclinical models. Our findings reveal a druggable nexus that functionally links the processes of mRNA transcription and translation initiation in an emerging class of lethal AR-deficient prostate cancer.
Prostate cancer-associated stroma (CAS) plays an active role in malignant transformation, tumor progression, and metastasis. Molecular analyses of CAS have demonstrated significant changes in gene ...expression; however, conflicting evidence exists on whether genomic alterations in benign cells comprising the tumor microenvironment (TME) underlie gene expression changes and oncogenic phenotypes. This study evaluates the nuclear and mitochondrial DNA integrity of prostate carcinoma cells, CAS, matched benign epithelium and benign epithelium-associated stroma by whole-genome copy-number analyses, targeted sequencing of TP53, and FISH. Array comparative genomic hybridization (aCGH) of CAS revealed a copy-neutral diploid genome with only rare and small somatic copy-number aberrations (SCNA). In contrast, several expected recurrent SCNAs were evident in the adjacent prostate carcinoma cells, including gains at 3q, 7p, and 8q, and losses at 8p and 10q. No somatic TP53 mutations were observed in CAS. Mitochondrial DNA (mtDNA) extracted from carcinoma cells and stroma identified 23 somatic mtDNA mutations in neoplastic epithelial cells, but only one mutation in stroma. Finally, genomic analyses identified no SCNAs, LOH, or copy-neutral LOH in cultured cancer-associated fibroblasts, which are known to promote prostate cancer progression in vivo
The gene expression changes observed in prostate cancer-adjacent stroma and the attendant contribution of the stroma to the development and progression of prostate cancer are not due to frequent or recurrent genomic alterations in the TME.
We show that the VEGF receptor neuropilin-2 (NRP2) is associated with high-grade, PTEN-null prostate cancer and that its expression in tumor cells is induced by PTEN loss as a consequence of c-Jun ...activation. VEGF/NRP2 signaling represses insulin-like growth factor-1 receptor (IGF-IR) expression and signaling, and the mechanism involves Bmi-1-mediated transcriptional repression of the IGF-IR. This mechanism has significant functional and therapeutic implications that were evaluated. IGF-IR expression positively correlates with PTEN and inversely correlates with NRP2 in prostate tumors. NRP2 is a robust biomarker for predicting response to IGF-IR therapy because prostate carcinomas that express NRP2 exhibit low levels of IGF-IR. Conversely, targeting NRP2 is only modestly effective because NRP2 inhibition induces compensatory IGF-IR signaling. Inhibition of both NRP2 and IGF-IR, however, completely blocks tumor growth in vivo.
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