Benign Prostate Tissue Research Articles

Role of SELENBP1 and SELENOF in prostate cancer bioenergetics

The contribution of selenium and selenoproteins in prostate cancer etiology remains elusive, potentially due to insufficient information regarding the biochemical pathways in which they are involved. There are twenty-five human selenocysteine-containing proteins or selenoproteins as well as a smaller class of selenium-containing proteins that do not include selenocysteine, and their cancer-associated aberrations, both genetic and functional, have evoked special interest, although their contribution to the metabolic reprogramming of prostate cancers remains has not been extensively studied. While benign prostate tissue exhibits a glycolytic phenotype, neoplastic events restore the truncated tricarboxylic acid cycle and enhance oxidative phosphorylation. Two selenium-containing proteins, selenium binding protein 1 and selenoprotein F, affect prostate cancer phenotypes by modulating tumor cell metabolic profiles with significant effects on mitochondrial biology, including oxidative phosphorylation and ATP synthesis. One of the pathways affected by both proteins is the activation of adenosine monophosphate kinase and its downstream signaling with concomitant induction of glycolysis. This review focuses on highlighting the role of these two proteins in modulating the bioenergetic profile of prostate cancer and in maintaining the metabolic plasticity of these cells rendering growth advantage and possible therapeutic resistance.

TMEM158 expression is negatively regulated by AR signaling and associated with favorite survival outcomes in prostate cancers.

Membrane protein TMEM158 was initially reported as a Ras-induced gene during senescence and has been implicated as either an oncogenic factor or tumor suppressor, depending on tumor types. It is unknown if TMEM158 expression is altered in prostate cancers. Multiple public gene expression datasets from RNA-seq and cDNA microarray assays were utilized to analyze candidate gene expression profiles. TMEM158 protein expression was assessed using an immunohistochemistry approach on a tissue section array from benign and malignant prostate tissues. Comparisons of gene expression profiles were conducted using the bioinformatics software R package. COX regression-based screening identified the membrane protein TMEM158 gene as negatively associated with disease-specific and progression-free survival in prostate cancer patients. Gene expression at the mRNA and protein levels revealed that TMEM158 expression was significantly reduced in malignant tissues compared to benign compartments. Meanwhile, TMEM158 downregulation was strongly correlated with advanced clinicopathological features, including late-stage diseases, lymph node invasion, higher PSA levels, residual tumors after surgery, and adverse Gleason scores. In castration-resistant prostate cancers, TMEM158 expression was negatively correlated with AR signaling activity but positively correlated with neuroendocrinal progression index. Consistently, in cell culture models, androgen treatment reduced TMEM158 expression, while androgen deprivation led to upregulation of TMEM158 expression. Correlation analysis showed a tight correlation of TMEM158 expression with the level of R-Ras gene expression, which was also significantly downregulated in prostate cancers. Tumor immune infiltration profiling analysis discovered a strong association of TMEM158 expression with NK cell and Mast cell enrichment. The membrane protein TMEM158 is significantly downregulated in prostate cancer and is tightly associated with disease progression, anti-tumor immune infiltration, and patient survival outcome.

Optical anisotropy composition of benign and malignant prostate tissues revealed by Mueller-matrix imaging.

A Mueller matrix imaging approach is employed to disclose the three-dimensional composition framework of optical anisotropy within cancerous biotissues. Visualized by the Mueller matrix technique spatial architecture of optical anisotropy of tissues is characterised by high-order statistical moments. Thus, quantitative analysis of the spatial distribution of optical anisotropy, such as linear and circular birefringence and dichroism, is revealed by using high-order statistical moments, enabling definitively discriminate prostate adenoma and carcinoma. The developed approach provides greater (>90%) accuracy of diagnostic achieved by using either the 3-rd or 4-th order statistical moments of the linear anisotropy parameters. Noticeable difference is observed between prostate adenoma and carcinoma tissue samples in terms of the extinction coefficient and the degree of depolarisation. Juxtaposition to other optical diagnostic modalities demonstrates the greater accuracy of the approach described herein, paving the way for its wider application in cancer diagnosis and tissue characterization.

Overexpression of ELL-associated factor 2 suppresses invasion, migration, and angiogenesis in colorectal cancer.

BACKGROUNDThe androgen responsive gene, ELL-associated factor 2 (EAF2), expressed in benign prostate tissues, has been shown to play an important role in tumor suppression in a variety of malignant tumors. In addition, some scholars found that EAF2 frameshift mutations are associated with intratumor heterogeneity in colorectal cancer (CRC) and inactivation of EAF2 in microsatellite instability-high CRC. However, the molecular mechanism by which EAF2 is involved in CRC invasion and metastasis remains unclear.AIMTo determine the clinical value of expression of EAF2 protein in CRC, and to study the effects of EAF2 on the invasion, migration, and angiogenesis of CRC cells in vitro.METHODSIn this study, we collected colorectal adenocarcinoma and corresponding adjacent tissues to investigate the clinical expression of EAF2 protein in patients with advanced CRC. Subsequently, we investigated the effect of EAF2 on the invasion, migration, and angiogenesis of CRC cells in vitro using plasmid transfection.RESULTSEAF2 protein was lowly expressed in cancer tissues of patients with advanced CRC. Kaplan-Meier survival analysis showed that the survival rate of the high EAF2 level group was higher than that of the low EAF2 level group.CONCLUSIONOur results demonstrated that EAF2, as a tumor suppressor, may inhibit the invasion, metastasis, and angiogenesis of CRC cells by regulating the signal transducer and activator of transcription 3/transforming growth factor-β1 crosstalk pathway, and play a cancer suppressive and protective role in the occurrence and development of CRC. Our findings are of great significance to provide a new idea and theoretical basis for the targeted diagnosis and treatment of CRC.

Enrichment of “Cribriform” morphologies (intraductal and cribriform adenocarcinoma) and genomic alterations in radiorecurrent prostate cancer

AbstractLocally relapsed prostate cancer (PCa) after radiation therapy (RT) is associated with substantial morbidity and mortality. Morphological and molecular consequences that may contribute to RT resistance and local recurrence remain poorly understood. Locally recurrent PCa tissue from 53 patients with clinically localized PCa who failed with primary RT and subsequently underwent salvage radical prostatectomy (RP) was analyzed for tumor focality, clinicopathological, molecular, and genomic characteristics. Targeted next-generation sequencing with full exon coverage of 1,425 cancer-related genes was performed on 10 representative radiorecurrent PCas exhibiting no RT effect with matched adjacent benign prostate tissue. At RP, 37 (70%) of PCas had no RT effect with the following characteristics: grade group (GG) ≥ 3 (70%), unifocal tumor (75%), extraprostatic disease (78%), lymph node metastasis (8%), and “cribriform” morphologies (84%) [cribriform PCa (78%) or intraductal carcinoma (IDC-P) (61%)] at a median percentage of approximately 80% of tumor volume. In the setting of multifocal tumors (25%) at RP, the cribriform morphologies were restricted to index tumors. Of 32 patients with available pre-RT biopsy information, 16 had GG1 PCa, none had cribriform morphologies at baseline but 81% demonstrated cribriform morphologies at RP. Notable alterations detected in the sequenced tumors included: defects in DNA damage response and repair (DDR) genes (70%) (TP53, BRCA2, PALB2, ATR, POLQ), PTEN loss (50%), loss of 8p (80%), and gain of MYC (70%). The median tumor mutational burden was 4.18 mutations/Mb with a range of 2.16 to 31.86. Our findings suggest that most radiorecurrent PCas are enriched in cribriform morphologies with potentially targetable genomic alterations. Understanding this phenotypic and genotypic diversity of radiorecurrent PCa is critically important to facilitate optimal patient management.

ZFP36 Inhibits Tumor Progression of Human Prostate Cancer by Targeting CDK6 and Oxidative Stress.

Background The expression of ZFP36 in previous study was reduced in prostate cancer (PCa) tissues as compared to benign prostate tissues, indicating the potential of ZFP36 as an auxiliary marker for PCa. Further evaluation was conducted in clinical samples for in vitro and in vivo experiments, to prove the potential possibility that ZFP36 dysregulation participated in the malignant phenotype of PCa, to determine its potential mechanism for tumor regulation, and to provide a new theoretical basis for gene therapy of PCa. Methods First, the expression of ZFP36 in prostate tissue and PCa tissue was explored, and the relationship between ZFP36 and clinical features of PCa patients was illustrated. Subsequently, the impact of ZFP36 on the biology of PCa cells and relevant downstream pathways of ZFP36's biological impact on PCa were elucidated. Finally, whether oxidative stress mediated the regulation of ZFP36 in PCa was verified by the determination of oxidative stress-related indicators and bioinformatics analysis. Results The downregulation of ZFP36 in PCa tissue had a positive correlation with high Gleason scores, advanced pathological stage, and biochemical recurrence. ZFP36 was identified as an independent prognostic factor for PCa patients' BCR-free survival (P = 0.022) by survival analysis. Following a subsequent experiment of function gain and loss, ZFP36 inhibited the proliferation, invasion, and migration in DU145 and 22RV1 cells and inhibits tumor growth in the mouse model. Additionally, high-throughput sequencing screened out CDK6 as the downstream target gene of ZFP36. Western blot/Q-PCR demonstrated that overexpression of ZFP36 could reduce the expression of CDK6 at both cellular and animal levels, and the dual-luciferase experiment and RIP experiment proved that CDK6 was the downstream target of ZFP36, indicating that CDK6 was a downstream target of ZFP36, which mediated tumor cell growth by blocking cell cycle at the G1 stage. Furthermore, ZFP36 inhibited oxidative stress in PCa cells. Conclusions In PCa, ZFP36 might be a tumor suppressor that regulated growth, invasion, and migration of PCa cells. The lately discovered ZFP36-CDK6 axis demonstrated the molecular mechanism of PCa progression to a certain extent which might act as a new possible therapeutic target of PCa therapy.

Prostate Cancer in Male-to-Female Transgender Individuals: Histopathologic Findings and Association With Gender-affirming Hormonal Therapy.

Male-to-female (MtF) transgender individuals are at risk for prostate cancer, although guidelines for screening and management in this population are not well established. We describe a series of 9 MtF transgender patients who underwent prostate tissue sampling and highlight histopathologic features and challenges related to pathologic interpretation of prostate tissue in this patient population. Seven of 9 total patients were diagnosed with prostate cancer and all had elevated prostate-specific antigen at the time of diagnosis. Three of the 7 patients diagnosed with prostate cancer had received different types of hormone therapy for gender affirmation before the diagnosis of prostate cancer, and in all 3 of these patients, there was histologic evidence of hormone therapy effect in both benign prostate tissue and/or the adenocarcinoma. The 2 patients with benign prostate tissue underwent transurethral resection for lower urinary tract symptoms and were previously on hormone therapy for gender affirmation. Both of these specimens showed diffuse glandular atrophy and basal cell hyperplasia, indicative of hormone therapy effect on benign prostatic tissue. In the patients diagnosed with prostate cancer, a spectrum of grades was observed, ranging from Grade Group 1 to Grade Group 5. Four patients underwent radical prostatectomy, with 2 cases showing extraprostatic extension and Grade Group 5 prostatic adenocarcinoma, and 2 showing Grade Group 2 prostatic adenocarcinoma. Three of the 4 patients who underwent radical prostatectomy had received gender-affirming hormone therapy before surgery, and all 3 of these specimens showed hormone therapy effect in non-neoplastic prostate tissue and focal hormone therapy effect in prostatic adenocarcinoma. The presence of areas of viable carcinoma without hormone therapy effect enabled the assignment of a Gleason score and Grade Group in these 3 cases. Hormone therapy administered for gender identity affirmation induces histopathologic changes to both benign prostate tissue (nonkeratinizing squamous metaplasia, diffuse atrophy, basal cell hyperplasia, and stromal dominance with decreased numbers of glands) and prostatic adenocarcinoma (nuclear pyknosis, atrophy, cytoplasmic vacuolization, and architectural patterns that would qualify for Gleason 4 and 5 in the absence of hormone therapy effect) that have been traditionally seen in cis-male prostate cancer patients receiving hormone therapy. In the absence of hormone therapy, the morphology of prostatic adenocarcinoma in transgender patients shows classic morphologic features similar to those seen in cis-male patients not on hormone therapy. Prostate cancer with hormone therapy effect may not only be histologically quite subtle and may be overlooked if not suspected, but also should not be assigned a Gleason score because the Gleason score would substantially overstate its biologic potential. Therefore, similar to cis-male patients who have received androgen deprivation therapy for prostate cancer, transgender patients on hormone therapy for gender affirmation may be at risk for both underrecognition and over-grading of prostate cancer, particularly if the pathologist is not aware of the clinical history.

Glycolytic potential enhanced by blockade of pyruvate influx into mitochondria sensitizes prostate cancer to detection and radiotherapy.

Objective:This study aimed to evaluate the effects of mitochondrial pyruvate carrier (MPC) blockade on the sensitivity of detection and radiotherapy of prostate cancer (PCa).Methods:We investigated glycolysis reprogramming and MPC changes in patients with PCa by using metabolic profiling, RNA-Seq, and tissue microarrays. Transient blockade of pyruvate influx into mitochondria was observed in cellular studies to detect its different effects on prostate carcinoma cells and benign prostate cells. Xenograft mouse models were injected with an MPC inhibitor to evaluate the sensitivity of 18F-fluorodeoxyglucose positron emission tomography with computed tomography and radiotherapy of PCa. Furthermore, the molecular mechanism of this different effect of transient blockage towards benign prostate cells and prostate cancer cells was studied in vitro.Results:MPC was elevated in PCa tissue compared with benign prostate tissue, but decreased during cancer progression. The transient blockade increased PCa cell proliferation while decreasing benign prostate cell proliferation, thus increasing the sensitivity of PCa cells to 18F-PET/CT (SUVavg, P = 0.016; SUVmax, P = 0.03) and radiotherapy (P < 0.01). This differential effect of MPC on PCa and benign prostate cells was dependent on regulation by a VDAC1-MPC-mitochondrial homeostasis-glycolysis pathway.Conclusions:Blockade of pyruvate influx into mitochondria increased glycolysis levels in PCa but not in non-carcinoma prostate tissue. This transient blockage sensitized PCa to both detection and radiotherapy, thus indicating that glycolytic potential is a novel mechanism underlying PCa progression. The change in the mitochondrial pyruvate influx caused by transient MPC blockade provides a critical target for PCa diagnosis and treatment.

Abstract 3632: Differential DNA methylation in the benign and cancerous prostate tissue of African American and European American men

Abstract In this study, we compare patterns of differential DNA methylation (DNAm) distinguishing tumor and benign prostate tissue in European American (EA) and African American (AA) men to improve our understanding of biological mechanisms that may underlie prostate cancer (PCa) disparities. In the United States, AA men are more likely to be diagnosed with and to die of PCa compared with EA men. Changes in DNAm accompany tumor development and progression, and differences in DNAm across ancestry groups may be an important lens through which to study PCa disparities. We collected paired tumor and histologically benign formalin-fixed paraffin-embedded tissue blocks from 151 (76 AA, 75 EA) PCa patients undergoing prostatectomy at the University of Chicago. DNA was extracted and genome-wide DNAm was measured at ~850,000 CpG sites using the Illumina MethylationEpic Array. After quality control, 682,694 CpGs remained for analysis. We conducted differential methylation analyses, adjusting for batch, age, and individual and identified 29,236 and 38,035 tumor-associated CpGs in AA and EA respectively (p&amp;lt;5x10-8 and Δβ&amp;gt;0.3) with 25,263 CpGs common to both sets. There was a depletion of differentially methylated CpGs in CpG islands and promoters in both groups, but depletion was stronger in AA. Tumor-associated CpGs within islands and in promoters were more likely to be hypermethylated for both groups, though this was stronger in EA. We identified 2,392 genes with differentially methylated promoters common to both ancestry groups; 223 and 1,045 were unique to AA and EA respectively. Shared gene regions included GSTP1, APC, RARB, and GRASP, which have been previously reported. We assigned CpG sites to genes and used Gene Set Enrichment Analysis to identify differentially methylated pathways. The identified pathways include several previously associated with cancer development including the epithelial mesenchymal transition. We also identified ancestry-specific pathways including the early and late estrogen response pathways in AA. Examining the predictive ability of our differentially methylated CpGs, we found that relatively few CpGs (~10) were sufficient to distinguish tumor and benign tissue in AA and EA men (AUC&amp;gt;0.9). Scores constructed from differentially methylated CpGs in EA were able to predict tumor vs benign in AA (and vice versa) with high accuracy. Finally, we examined the association between ancestry and DNAm in both tumor and benign tissue. Here, 89 and 423 ancestry-associated CpGs were identified in tumor and benign respectively. Overall, we find that differential methylation patterns distinguishing tumor and benign tissue are generally similar for EA and AA men. However, differenced in tumor-associated DNAm as well as the presence of ancestry-associated CpGs provide a rich area for future studies exploring the impact of these differences on cancer development in AA and EA men. Citation Format: Meytal B. Chernoff, Marc Gillard, Kathryn Demanelis, Dayana Delgado, Anthony Williams, Donald Vander Griend, Brandon L. Pierce. Differential DNA methylation in the benign and cancerous prostate tissue of African American and European American men [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3632.

Abstract 5856: Differential expresion of miR-9 and their putative targets in prostate cancer

Abstract Background: Deregulation of microRNA (miRNA) plays an important role in the growth, invasion and migration of prostate cancer cells. miR-9 has been found overexpressed in prostate cancer (Pca) compared to benign prostate tissue (BPT) and proposed as an oncomiR with a potential as a therapeutic target. Little is known about the regulatory mechanisms of miR-9 in PCa and opposing roles in supporting or suppressing tumor development and metastasis has been reported in different tumor tissues. We aimed to explore the miR-9 expression and their putative targets in BPT and PCa tissue through in situ Hybridization (ISH) and bioinformatics analysis of miRNA and gene expression databases. Design: A prostate tissue microarray was constructed with representative areas of BPT and PCa. The expression of miR-9-5p was evaluated through a miRNAscope 2.5 LS Red ISH, it was scored using a semiquantitative analysis of the average number of dots per cell; mean score values were compared between BPT and PCa tissues using Mann-Whitney test, a p value less than 0.05 was considered statistically significant. We look for possible mir-9 targets in the database of experimentally validated microRNA-target (miRTarBase), and we use the AACR Project GENIE data base to analyze the behavior of the putative miR-9 targets in PCa vs BPT and explored their expression in the tumor tissues of the Cancer Genome Atlas (TCGA) through the bioinformatics tools of the GEPIA2 web server. Results: The analysis of miR-9 expression in 294 prostate tissue cores (145 BPT and 149 PCa) showed an overexpression of this miRNA in PCa (1.60, 95% CI:1.51-1.69) compared to BPT (0.86, 95% CI: 0.77-0.94), (p less than 0.0001). We identified 526 mir-9 target genes in the miRTarBase validated with at least one experimental assay, and selected the 24 genes with the strongest evidence of interaction with miR-9. The gene targets with higher expression level in PCa were CDH1, RAB34, AP3B1, CCNG1, SRF, TGFB1, ID2, FOXO3 y CCND1 (Log2 (TPM+1) &amp;gt; 4). Only 4 genes showed a statistically significant difference in their expression when BPT and PCa tissue data from the TCGA were compared. CDH1 was overexpressed in the PCa datasets and BCL6, RAB34 and NTRK3 were downregulated in cancer compared to normal tissues. Most of the miR-9 gene targets did not show alterations at the DNA sequence level according to the analysis of the AACR Project GENIE data using the cBioPortal, except for CDH1, BCL6, CCND1 and PRDM1, which showed truncating mutations, amplifications, deep deletion and splice mutations in some of the cancer datasets. These results suggest that mir-9 may be regulating tumor initiation, progression, and metastasis processes through the Neurotrophic Receptor Tyrosine Kinase 3 (NTRK3), which has been found altered in breast carcinomas and other cancers, or the downregulation of Cadherin 1 (CDH1), whose loss of function is thought to contribute to cancer progression by increasing proliferation, invasion, and metastasis. Citation Format: Ines Benedetti, Lia Barrios, Juan Rebollo. Differential expresion of miR-9 and their putative targets in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5856.

Abstract 5718: Rab34 is downregulated in human prostate cancer tissue

Abstract Background: Rab34 encodes a protein belonging to the Rab family of proteins, which are small GTPases important in regulating signal transduction and cellular processes. A potential role of Rab34 in cancer has been suggested, but few studies have reported its function in human epithelial cancers. It was reported that prostate cancer (PCa) patients were more prone to biochemical recurrence when Rab34 was downregulated in PCa tissue compared with benign prostate tissue (BPT), being negatively associated with miR-148, a miRNA that increased the growth in cell lines and was up-regulated in PCa samples. Rab34 is a putative target gene for miR-9. We aimed to explore the Rab34 and miR-9 expression in PCa using a bioinformatic analysis, however, the mere presence of miRNA-binding sites is insufficient for predicting target regulation, and therefore, we also analyzed Rab34 expression in BPT and PCa tissues. Design: The expression of miR9-5p and Rab34 mRNA was explored in the tumor tissues of the Cancer Genome Atlas (TCGA) through the bioinformatics tools of the dbDEMC2 and GEPIA2 web servers, and additional evidence of miR-9/Rab34 interaction was sought in the miRTarBase of experimentally validated microRNA-target interactions. Two pathologists evaluated Rab34 immunohistochemical expression in localized PCa and BPT in a prostate tissue microarray. For each spot an H-score was obtained by microscopic assessment of epithelial cells with positive staining. Differences in Rab34 expression were found by comparing H-Scores between BPT and PCa tissues using Mann-Whitney test, a p value less than 0.05 was considered statistically significant. Results: The analysis of miR-9 and Rab34 expression in the TCGA showed that Rab34 has a diverse expression in tumor tissues, being downregulated in colon, rectum and PCa, and upregulated in lymphoma, leukemia and pancreatic adenocarcinoma. On the other hand, miR-9 was upregulated in PCa and downregulated in pancreatic adenocarcinoma. There were no experimental evidence supporting the miR-9/Rab34 interaction in PCa tissues from the miRTarBase. However, most of the breast cancer datasets showed a negative correlation in the expression of both genes. Rab34 expression at protein level was assessed in 123 tissue cores representing PCa, and 86 tissue cores representing BPT. In cores with PCa mean Rab34 expression was significantly lower than the mean Rab34 expression in cores with BPT (p less than 0.0001). There was a lower mean staining score in PCa (87.3, 95% CI: 73.78 - 100.9), compared to BPT (156.3, 95% CI: 144 - 168.5). Conclusions: Our study shows that Rab34 is downregulated in PCa tissue. These findings are in accordance to a previous report that associated its lower expression to worse prognosis and tumor recurrence. The downregulation of Rab34 in PCa tissue might be related to overexpression of its putative microRNA, miR-9. Additional studies to detect the in situ expression of miR-9 in PCa are in progress to validate these results. Citation Format: Ines Benedetti, Lia Barrios, Juan Rebollo. Rab34 is downregulated in human prostate cancer tissue [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5718.

Abstract 4013: Functional heterogeneity of cancer-associated stromal subtypes in the prostate cancer microenvironment

Abstract The stromal tumor microenvironment plays a central role in the development, progression and therapy resistance of prostate cancer (PCa), the second leading cause of cancer-related death in males in western nations. Functional and single cell RNA sequencing (scRNA-seq) studies unequivocally demonstrate the coexistence of functionally and spatially distinct fibroblast and mural cell states within the tumor microenvironment of various non-prostate tumors. Stromal cellular heterogeneity in PCa however remains poorly understood. This study therefore aimed to (1) comprehensively analyze stromal cell states within the PCa microenvironment and (2) to identify and characterize onco-supportive fibromuscular entities in respect to their potential clinical exploitation. Thus, we performed bulk tissue scRNA-seq on biopsy cores from benign and cancerous prostate tissue of 4 different patients. Analyses presented here focused on the fibroblast and mural cell clusters, which each delineated further into 2 subclusters. One fibroblast subcluster showed strong similarity to published signatures of inflammatory cancer-associated fibroblasts (iCAF). A closely related but distinct fibroblast subcluster exhibited hallmarks of benign interstitial fibroblasts with enrichment of GO terms such as antigen processing and presentation. Based on expression signatures and spatial localization via immunohistochemistry, the 2 mural cell subclusters (MCAM+) were identified as vascular smooth muscle cells (vSMCs) and dedifferentiated prostate SMCs/myofibroblasts. Notably, the gene signature of this latter subcluster correlated with increasing Gleason score and decreased disease-specific survival. On the basis of scRNA-seq data from this and independent datasets, panels of subcluster-specific markers were selected for validation studies via FACS, multiplex immunohistochemistry and quantitative real time PCR. Thus far, immunohistochemical studies validated not only the in vivo presence of the 4 distinct cellular states but also revealed their distinct spatial localization. We will present molecular and functional data from these ongoing analyses in (i) immortalized fibroblast and smooth muscle cell lines, (ii) heterogeneous ex vivo outgrowth cultures of primary PCa fibroblasts/myofibroblasts and (iii) primary cells from single cell suspensions of PCa biopsies enzymatically digested using an in-house approach optimized for maximal recovery of fibromuscular cells. These studies aim to evaluate the specificity, robustness and suitability of the chosen markers to delineate distinct tumor-relevant stromal cell subtypes enabling their functional contribution to PCa pathophysiology and clinical relevance to be assessed in future experiments. Citation Format: Elisabeth Damisch, Elena Brunner, Sieghart Sopper, Isabel Heidegger, Andreas Pircher, Georgios Fotakis, Zlatko Trajanoski, Sandra Theresa Deichsler, Georg Schäfer, Natalie Sampson. Functional heterogeneity of cancer-associated stromal subtypes in the prostate cancer microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4013.

Abstract 2372: Targeting a metabolic compensatory mechanism for heat shock factor 1 inhibition in prostate cancer

Abstract While inhibition of androgen receptor signaling has greatly improved the outcome for many prostate cancer patients (PCa), there are limited therapeutic interventions for advanced and metastatic PCa. Heat shock factor 1 (HSF1) is activated by cellular stress and regulates transcription, proteome stability, and oxidative stress. HSF1 is elevated in aggressive cancers from many tissue types and has been directly implicated in driving cancer progression. We found that HSF1 predicts survival for metastatic castration resistant prostate cancer patients, who are unable to be treated by inhibition of the androgen receptor signaling. In comparison to benign prostate tissue, HSF1 had increased protein levels in primary PCa, castration resistant PCa, and small cell neuroendocrine carcinoma the most aggressive histologic type of PCa. Knockdown of HSF1 decreases cellular growth in PCa cell lines. In collaboration with the laboratory of Dr. Dennis Thiele, we identified a direct HSF inhibitor that targets nuclear HSF1 for degradation. Prostate cancer cells experience cellular senescence when treated with the HSF1 inhibitor. To better understand how HSF1 regulates PCa cellular growth, we investigated metabolic changes in response to the selective HSF1 inhibitor. We found that a rate limiting enzyme in the transsulfuration pathway was reproducibly altered after HSF1 knockdown and pharmacological inhibition. This is the first linkage of HSF1 and the transsulfuration pathway in cancer. The transsulfuration pathway is important for production of cellular energy in the form of pyruvate, so overactive HSF1 likely increases cellular growth through increased pyruvate production. The transsulfuration pathways also produces the antioxidant hydrogen sulfide. The increased production of antioxidants likely aid cancer cells in responding to the accumulation of reactive oxygen species that form in cancer cells from oncogene activation, decreased blood supply, and tumor hypoxia. We have identified novel HSF1 binding sites in the gene encoding this transsulfuration pathway enzyme, which regulate levels of gene expression. Small cell neuroendocrine carcinoma cells treated with both the selective HSF1 inhibitor and the transsulfuration pathway inhibitor have reduced growth and increased cellular death by caspase 3 cleavage. Targeting the transsulfuration pathway in combination with HSF1 inhibition may delay recurrence in PCa patients by killing tumor cells instead of solely inducing cellular senescence. Since HSF1 is increased in many forms of cancer, these data may provide novel therapeutic targets for aggressive cancer from many tissue types. Citation Format: J. Spencer Hauck, Xia Gao, William Butler, Lingfan Xu, Jiaoti Huang. Targeting a metabolic compensatory mechanism for heat shock factor 1 inhibition in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2372.

CXCR3 antagonist AMG487 ameliorates experimental autoimmune prostatitis by diminishing Th1 cell differentiation and inhibiting macrophage M1 phenotypic activation.

Chronic prostatitis and chronic pelvic pain syndrome (CP/CPPS) is an inflammatory immune disease that is characterized by infiltrating inflammatory cells in the prostate and pelvic or by perineal pain. Receptor CXCR3modulates immune and inflammatory responses; however, the effects of CXCR3 antagonist AMG487 in the context of CP/CPPS are unknown. Therefore, we investigated the effect of AMG487 in experimental autoimmune prostatitis (EAP) mice and explored the potential functional mechanisms. The EAP model was induced by intradermally injecting a mixture of prostate antigensand complete Freund's adjuvant on Days 0 and 28. To evaluate the effect of AMG487 on EAP mice, treatment with AMG487 and vehicle solution was conducted for the indicated period. Then, procedures were performed, including behavioral test, to evaluate the pain response to stimulation before the mice were killed and a histological assessment to evaluate the inflammation afterthe mice were killed. Immunofluorescence, flow cytometry, and Western blot assay were used to analyze the functional phenotype and regulation mechanism of AMG487 on T helper type 1 (Th1) cells and macrophages. We found high expression of CXCR3 in human benign prostate tissues with inflammation and EAP mice. The elevated CXCR3 in prostate tissues correlates with the severity of inflammation. CXCR3 antagonist AMG487 treatment ameliorated the inflammatory changes and the pelvic pain of EAP mice. AMG487 inhibits Th1 cell differentiation through the IL-12/STAT4pathwayand inhibits pro-inflammatory M1 macrophages through the lipopolysaccharide/NF-κB p65signaling. AMG487 could inhibit the secretion of inflammatory mediators in EAP mice. CXCR3 antagonist AMG487 could ameliorate the inflammatory changes and the pelvic pain of EAP mice by diminishing Th1 cell differentiation and inhibiting macrophage M1 phenotypic activation. Thus, the results imply that AMG487 has the potential as an effective therapeutic agent in the prevention and treatment of EAP.

Molecular correlates of high B7-H3-expressing metastatic castrate-resistant prostate cancers (mCRPC) via exome, transcriptome, and epigenome analyses.

5045 Background: mCRPC is a lethal condition with limited effective treatment options. B7-H3, a transmembrane protein of the B7 checkpoint superfamily is overexpressed in prostate cancer (PC) and is associated with poor prognosis. While several novel approaches target B7-H3 in prostate cancer, lack of knowledge about the molecular features and regulatory mechanisms of B7-H3 expression in mCRPC prevents the optimal design of these interventions. We aimed to characterize B7-H3 in mCRPC with the purpose to reveal the roles of B7-H3 in mCRPC pathogenesis, and stratify the patient population optimal for B7-H3–targeted therapies. Methods: We conducted bioinformatic interrogations at genome-scale on whole-exome and whole-transcriptome sequencing (WES, WTS) data from SU2C/PCF (n = 209, mCRPC), SUWC (n = 101, mCRPC) and GTEx (n = 246, benign prostate tissue). We developed and utilized novel machine learning (ML) algorithms to examine the association of B7-H3 with other key PC pathways. We examined the genetic and epigenetic regulation of B7-H3 by CHIP-seq. We also performed single-cell mRNA sequencing (scRNA-seq) analysis on one patient before and after treatment with the AR inhibitor, enzalutamide. We utilized both Gene Set Enrichment Analysis (GSEA) and our ML approaches to identify enriched signaling pathways in high B7-H3–expressing mCRPC. Results: Expression of B7-H3 was significantly elevated in mCRPC compared to benign prostate tissues, and was distinctive from other B7 family members. High B7-H3 expression was significantly associated with ERG fusions, AR-V7 variant, and increased FOXA1 and androgen receptor (AR) mRNA expression. At gene-level analysis, our ML algorithm suggested that B7-H3 shared similar gene networks as AR and its co-regulators, HOXB13, and FOXA1. CHIP-seq analysis revealed that the distal enhancers of B7-H3 in mCRPC were hyperactive (based on enhanced H3K27Ac marks) relative to primary tumors. Notably, these B7-H3 enhancers were bound by HOXB13, FOXA1 and AR in mCRPC samples. Our scRNA-seq analysis of one patient resistant to enzalutamide showed increased number of B7-H3-expressing cells. Based on integrated GSEA and ML analyses on WTS data, B7-H3 was positively associated with TGF-beta signaling, an enzalutamide resistance pathway. Conclusions: In mCRPC, we found B7-H3 overexpression as compared to other B7 family members. The association of B7-H3-high expression and certain genomic alterations may stratify mCRPC patients for B7-H3–targeted therapy. Further, B7-H3 exhibited regulation by AR, HOXB13 and FOXA1, at the B7-H3 enhancers. The activating epigenetic modification at the distal enhancers represents another potential marker to examine B7-H3 expression in tumors. Altogether, our study indicates that B7-H3 is a critical target in mCRPC patients, including those resistant to enzalutamide.

MP45-06 CLINICAL SIGNIFICANCE OF α2,3-SIALYLATED PROSTATE-SPECIFIC ANTIGEN DENSITY AND MRI FOR HIGH-GRADE PROSTATE CANCER IN BIOPSY-NAÏVE MEN WITH ELEVATED PSA LEVEL

You have accessJournal of UrologyCME1 May 2022MP45-06 CLINICAL SIGNIFICANCE OF α2,3-SIALYLATED PROSTATE-SPECIFIC ANTIGEN DENSITY AND MRI FOR HIGH-GRADE PROSTATE CANCER IN BIOPSY-NAÏVE MEN WITH ELEVATED PSA LEVEL Tohru Yoneyama, Shingo Hatakeyama, Mihoko Sutoh Yoneyama, Yuki Tobisawa, Takuma Narita, Hirotake Kodama, Masaki Momota, Hiroyuki Ito, Shintaro Narita, Fumiyasu Tsushima, Koji Mitsuzuka, Takahiro Yoneyama, Yasuhiro Hashimoto, Wilhelmina Duivenvoorden, Jehonathan H. Pinthus, Shingo Kakeda, Akihiro Ito, Norihiko Tsuchiya, Tomonori Habuchi, and Chikara Ohyama Tohru YoneyamaTohru Yoneyama More articles by this author , Shingo HatakeyamaShingo Hatakeyama More articles by this author , Mihoko Sutoh YoneyamaMihoko Sutoh Yoneyama More articles by this author , Yuki TobisawaYuki Tobisawa More articles by this author , Takuma NaritaTakuma Narita More articles by this author , Hirotake KodamaHirotake Kodama More articles by this author , Masaki MomotaMasaki Momota More articles by this author , Hiroyuki ItoHiroyuki Ito More articles by this author , Shintaro NaritaShintaro Narita More articles by this author , Fumiyasu TsushimaFumiyasu Tsushima More articles by this author , Koji MitsuzukaKoji Mitsuzuka More articles by this author , Takahiro YoneyamaTakahiro Yoneyama More articles by this author , Yasuhiro HashimotoYasuhiro Hashimoto More articles by this author , Wilhelmina DuivenvoordenWilhelmina Duivenvoorden More articles by this author , Jehonathan H. PinthusJehonathan H. Pinthus More articles by this author , Shingo KakedaShingo Kakeda More articles by this author , Akihiro ItoAkihiro Ito More articles by this author , Norihiko TsuchiyaNorihiko Tsuchiya More articles by this author , Tomonori HabuchiTomonori Habuchi More articles by this author , and Chikara OhyamaChikara Ohyama More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002611.06AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: To reduce unnecessary biopsies, biomarker discriminating high-grade prostate cancer (HGPC) greater than grade group 2 are needed. The presence of α2,3-sialylated prostate-specific antigen (S2,3PSA) in sera with prostate cancer (PC) patients is a potential marker of their aggressiveness. The objective was to characterized the origin of S2,3PSA in prostate tissue, and studied the clinical performance of %S2,3PSA density (%S2,3PSAD) alone and in combination with MRI for the detection of high-grade PC (HGPC) in men with elevated PSA. METHODS: %S2,3PSA of benign and cancer tissue was measured in 71 radical prostatectomy specimens. The diagnostic performance of %S2,3PSA density (%S2,3PSAD) alone (primary outcome) and %S2,3PSAD combined with Prostate Imaging Reporting and Data System (PI-RADS) scores (secondary outcome) was evaluated by retrospective systematic biopsy cohort (n = 654) and prospective MRI-targeted biopsy cohort (n = 79) respectively, analyzed using ROC and DCA analysis. RESULTS: %S2,3PSA was significantly higher in Gleason pattern 4 and 5 tissues compared to benign prostate tissue. In the retrospective cohort, the AUC of %S2,3PSAD for detecting HGPC was superior to total PSA (tPSA) or PSA density (PSAD) (0.7704 vs. 0.5431 and 0.7140, all p <0.001). In the prospective cohort, %S2,3PSAD was superior to tPSA, PSAD, and PI-RADS (AUC: 0.8194 vs. 0.5058, 0.7177 and 0.7661, all p <0.05) and biopsy avoidance rate of %S2,3PSAD+PI-RADS+tPSA+digital rectal examination (DRE) was superior to PI-RADS+tPSA+DRE (24% vs. 13.9%) at 30% risk threshold. CONCLUSIONS: The diagnostic performance of %S2,3PSAD was superior to conventional strategies but comparable to MRI. Source of Funding: This work was supported by the Japan Society for the Promotion of Science (JSPS) KAKENHI (grant nos. 15K15579 and 15H02563) and also supported by Sakurai Memorial Medical Research Foundation © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e764 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Tohru Yoneyama More articles by this author Shingo Hatakeyama More articles by this author Mihoko Sutoh Yoneyama More articles by this author Yuki Tobisawa More articles by this author Takuma Narita More articles by this author Hirotake Kodama More articles by this author Masaki Momota More articles by this author Hiroyuki Ito More articles by this author Shintaro Narita More articles by this author Fumiyasu Tsushima More articles by this author Koji Mitsuzuka More articles by this author Takahiro Yoneyama More articles by this author Yasuhiro Hashimoto More articles by this author Wilhelmina Duivenvoorden More articles by this author Jehonathan H. Pinthus More articles by this author Shingo Kakeda More articles by this author Akihiro Ito More articles by this author Norihiko Tsuchiya More articles by this author Tomonori Habuchi More articles by this author Chikara Ohyama More articles by this author Expand All Advertisement PDF DownloadLoading ...

Automated quantitative high-throughput multiplex immunofluorescence pipeline to evaluate OXPHOS defects in formalin-fixed human prostate tissue

Advances in multiplex immunofluorescence (mIF) and digital image analysis has enabled simultaneous assessment of protein defects in electron transport chain components. However, current manual methodology is time consuming and labour intensive. Therefore, we developed an automated high-throughput mIF workflow for quantitative single-cell level assessment of formalin fixed paraffin embedded tissue (FFPE), leveraging tyramide signal amplification on a Ventana Ultra platform coupled with automated multispectral imaging on a Vectra 3 platform. Utilising this protocol, we assessed the mitochondrial oxidative phosphorylation (OXPHOS) protein alterations in a cohort of benign and malignant prostate samples. Mitochondrial OXPHOS plays a critical role in cell metabolism, and OXPHOS perturbation is implicated in carcinogenesis. Marked inter-patient, intra-patient and spatial cellular heterogeneity in OXPHOS protein abundance was observed. We noted frequent Complex IV loss in benign prostate tissue and Complex I loss in age matched prostate cancer tissues. Malignant regions within prostate cancer samples more frequently contained cells with low Complex I & IV and high mitochondrial mass in comparison to benign–adjacent regions. This methodology can now be applied more widely to study the frequency and distribution of OXPHOS alterations in formalin-fixed tissues, and their impact on long-term clinical outcomes.

Cytotoxic Action of Palladium-Based Compound on Prostate Stem Cells, Primary Prostate Epithelial Cells, Prostate Epithelial Cells, and Prostate Cell Lines.

Objective Prostate cancer is one of the most common types of cancer found to occur in males and is ranked as the second-highest cause of cancer-associated deaths among male patients. In this study, we have shown the influence of a new palladium-based anticancer agent in contrast to the six distinct prostate cancer lines and the primary cultures. Methods In this study, we have used six distinct prostate cell lines, that is, PNT2-C2, LNCaP, BPH-1, PC-3, PNT1A, and P4E6. The MTP and ATP assay were performed to evaluate the growth of the cell and the flow cytometry to investigate the status of the cell cycle. The antigrowth effect of the palladium complex was evaluated against different cell lines at three time zones 24 h, 48 h, and 72 h. [PdCl(terpy)] (capsule)-2H2O is synthesized by direct encapsulation of equimolar amounts of capsule ions into [Pd (terpy) Cl] Cl-2H2O. Results A comparative analysis was done on 25 mM etoposide and 12 mM cisplatin, cytotoxic agents. The lowest IC50 value at 72 hours was 0.128 mM for BPH-1 cell lines with 0.139 mM, whereas PNT2-C2 cells were found to be most resistant with IC50 values of 0.829 mM. The antigrowth effect of palladium complex on cell lines was measured using the MTS assay at 24, 48, and 72 hours. BPH-1, PNT2-C2, and PNT1A either possess normal tissues or have benign prostatic hyperplasia tissues whereas P4E6, PC-3, and LNCaP cell lines possess malignant origin. The Pd complex exhibited significant cytotoxic action in stem cells when compared against etoposide. An antigrowth effect was reported for Pd complex at lower concentration, but it was more cytotoxic than etoposide with significant cytotoxicity (P=0.001). Conclusion The palladium complex experienced a substantial antigrowth influence over most of the prostate tumor cell lines and the primary cultures, eventually, leading to the implementation of this Pd complex in the treating procedure of metastatic prostate cancer, which is tremendously resistant to the traditional treatment.

Race Differences in Telomere Length in Benign Prostate Biopsies and Subsequent Risk of Prostate Cancer.

Telomere shortening is linked to aging and may be associated with increased risk for cancer. Most cancer studies have used telomere length in leukocytes rather than in the target tissue of cancer origin. A case-control study of 524 case-control pairs with a benign prostate biopsy nested within a historical cohort of 10,478 men was conducted to determine whether premalignant prostate telomere length (assessed using a modified qRT-PCR) is associated with prostate cancer risk. Telomere lengths in benign prostate biopsies of cases versus controls were similar (1.46 ± 0.38 vs. 1.45 ± 0.42; P = 0.49). African American (AA) men had significantly shorter telomeres compared with White men (1.51 ± 0.38 vs. 1.63 ± 0.39; P < 0.0001). In race-stratified analyses, increasing telomere length was more strongly associated with prostate cancer risk in White men, wherein those with telomere length in the highest quartile had 1.9-fold greater adjusted risk of prostate cancer compared with men with prostate telomere lengths in the lowest quartile [OR = 1.90; 95% confidence interval (CI) = 1.08-3.36]. Men in the highest telomere length quartile also had a greater risk of aggressive prostate cancer compared with men with telomere lengths in the lowest quartile (OR = 2.78; 95% CI = 1.25-6.19). White men have longer telomeres in benign prostate tissue compared with AA men, and those with the longest telomeres may be at increased risk for prostate cancer, particularly the more aggressive form of the disease. Race-specific telomere length measures may be an early biomarker of aggressive prostate cancer.

Tenascin C regulates cancer cell glycolysis and tumor progression in prostate cancer.

Tenascin C is a potential biomarker of cancer-associated fibroblasts and has been significantly associated with poor prognosis in patients with prostate cancer. However, the effects of Tenascin C in prostate cancer cell glycolysis largely remain unclear. Thus, this study aimed to investigate the Tenascin C expression in prostate cancer and its correlation to glycolysis-related protein and gene expression, clinicopathological parameters, and survival of patients. We performed immunohistochemical staining for Tenascin C in 141 cases of primary prostate cancer. Based on public data sets, we explored the association of Tenascin C with angiogenesis-related genes, M2 macrophage-related gene, androgen receptor levels, PI3K/AKT/NF-κB pathway genes, and glycolytic enzyme expression. The glucose uptake, lactate production, and glycolytic enzyme levels were detected by glycolysis assay and western blotting. Our results showed that Tenascin C expression is upregulated in prostate cancer tissues compared with benign prostatic hyperplasia tissues. High Tenascin C expression in prostate cancer cells was positively associated with lymph node metastasis, advanced clinical stage, the expression of CD105, CD206, and androgen receptor levels. The Kaplan-Meier curves showed a significant association of Tenascin C expression with the patient's overall survival. Tenascin C expression was positively associated with PI3K p85, pAKT-ser308, and NF-κB p65 protein expression in prostate cancer samples. Moreover, siRNA-mediated knockdown of Tenascin C expression inhibited cell glucose uptake, lactate production, and glycolytic-enzyme expression in prostate cancer cells invitro. Together, our findings suggest that Tenascin C is a prognostic marker for patients with prostate cancer and that its effects might be mediated via regulation of the glycolysis process of prostate cancer cells.