High Gleason Score Tumors Research Articles

Abstract 2745: Sequential tracking of PD-L1 expression on circulating epithelial tumor cells (CETCs) in men with primary non-metastatic prostate cancer

Abstract Background: Radiotherapy (RT) is an essential component of multimodality therapy in the primary or postoperative treatment of prostate cancer with curative intent by preventing local recurrence and distant metastases, indicating its impact on residual occult tumor, such as circulating tumor cells. Immunogenic changes in cancer cells induced by RT can lead to adaptive up-regulation of PD-L1 expression. As immune checkpoint inhibition plays an increasingly important role in prostate cancer, radiogenic PD-L1 upregulation may be of therapeutic importance. The aim of this study was to determine PD-L1 status on CETCs before and during RT. Methods: Peripheral blood samples were collected from 52 primary non-metastatic prostate cancer patients. CETCs were enumerated and analyzed for PDL-L1 expression prior to (baseline), 3 and 6 weeks after the start of RT using fluorescence scanning microscopy. Results: PD-L1 expression was detectable in CETCs and localized in the membrane and/or cytoplasm. We found a correlation between PD-L1 status in CETCs and aggressiveness of cancer disease prior to RT. The frequency of PD-L1 positive CETCs depended on the grading and Gleason score of the primary tumor. Patients with a high Gleason score and G3 tumors had significantly more PD-L1 positive CETCs compared to patients with a low Gleason Score and G1 tumors. Moreover, patients in stage I/II had significantly less PD-L1 positive CETCs compared to patients in stage III/IV (mean 54.25 vs. mean 69.65; p<0.05). A higher number of PD-L1 positive CETCs were detected in patients who had radical prostatectomy before start of RT than in those who had no surgical intervention (mean 66.24 vs. mean 47.46; p<0.01). Interestingly, the fraction of PD-L1 positive CETCs decreased significantly during RT only in patients with G3 tumors and a high Gleason Score. Conclusion: Our results indicated that PD-L1 in CETCs is closely related to the aggressiveness of the primary tumor and may be a co-factor associated with the progression of prostate cancer. PD-1/PD-L1 blockade combined with RT represents a novel and promising treatment strategy that could contribute to improve the treatment efficacy of prostate cancer. Citation Format: Dorothea Schott, Monika Pizon, Ulrich Pachmann, Katharina Pachmann, Matthias Mäurer. Sequential tracking of PD-L1 expression on circulating epithelial tumor cells (CETCs) in men with primary non-metastatic prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2745.

Abstract 2702: Prostate stromal transcriptome as biomarker of aggressive prostate cancer

Abstract Background: Prostate cancer has a heterogeneous prognosis. Discovery of molecular biomarkers that improve identification of potentially lethal prostate cancer could enhance the ability to determine which patients would benefit most from immediate treatment. Most prognostic biomarkers have focused on those in prostate tumors. However, it is increasingly recognized that the tumor microenvironment contributes to cancer aggressiveness and progression. We therefore examined whole transcriptome expression of the prostate stroma and associations with clinically significant, high-grade prostate cancer. Methods: We performed RNA-sequencing (Illumina TruSeq RNA Exome library preparation) of macrodissected stromal samples (adjacent to and distant from tumor) from 363 radical prostatectomy specimens from Health Professionals Follow-up Study and Physicians' Health Study participants. We performed a differential expression analysis comparing the gene expression and pathways of stroma adjacent to and distant from tumor. We also performed a cluster analysis and computed a single sample gene set enrichment analysis (ssGSEA) score to assess the association of our previously reported 25 stromal signature genes (Tyekucheva et al. 2017) to determine the association with Gleason score. Results: Eighty-eight genes as well as eight Gene Ontology pathways and fourteen Canonical pathways were differentially expressed (FDR<0.05) between tumor adjacent to and distant from tumor. Pathways included cell-substrate adhesion, muscle development, and cardiomyopathy. 30% of men had high-grade (Gleason score 8-10) tumors. We observed higher expression of the individual signature genes in the cluster enriched for high Gleason score tumors, and a higher stromal signature ssGSEA score was associated with high Gleason score (p=0.04). Conclusions: Pathways differentially expressed between stroma adjacent to and distant from tumor suggest known differences in cell types by location or a potential field effect. We observed that the direction of the association of the stromal signature with Gleason score agreed with our previous report. We are currently analyzing the full expression dataset to identify additional stromal genes associated with Gleason score and lethal prostate cancer, and performing multi-plex immunohistochemistry to assess stromal cell-type heterogeneity in regions of tumor and normal prostate across tissue microarrays. Citation Format: Yinglu Zhou, Chaoran Ma, Giuseppe Nicoló Fanelli, Lavinia Stefanizzi, Elizaveta Gazeeva, Lorelei A. Mucci, Massimo Loda, Svitlana Tyekucheva, Kathryn L. Penney. Prostate stromal transcriptome as biomarker of aggressive prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2702.

EXPRESSION OF CD10 IN CARCINOMA PROSTATE AND ITS CORRELATION WITH VARIOUS CLINICOPATHOLOGICAL PARAMETERS.

Aim: Adenocarcinoma of the prostate is the second most common cause of cancer. Loss of CD10 is a common early event in human prostate cancer and is seen in lower Gleason Score malignancies while increased and altered expression is seen in high Gleason Score tumors, lymph node and bone metastasis. Materials and methods: The present study was a prospective observational study conducted on seventy-five patients suspected of having prostate cancer. Immunohistochemical profile was assessed for PSA, AMACR and CD10 immunostain. The intensity of CD10 expression and pattern of CD10 staining of tumor cells was evaluated. Results: The patients were in age group of 50-90 years with a mean age of 70.97 ± 9.51 years. As the grade group/gleason score increased the number of cases showing negative expression decreased and the pattern of expression changed from membranous to cytoplasmic to both types of expression. As the serum PSA levels increased the intensity of expression changed from focally positive to diffusely positive. The pattern of expression also changed from membranous to cytoplasmic to both ( membranous + cytoplasmic ) types of expression with increase in PSA levels. Conclusion: By immunohistochemical analysis we can identify CD10 positive tumors, which may warrant more aggressive initial therapy. A number of drugs against CD10 are available based on which potential targeted therapies could be formulated.

Prospective comparison of 68Ga-RM2 PET/CT and 68Ga-PSMA-617 PET/CT for initial staging of prostate cancer.

21 Background: PSMA ( Prostate Specific Membrane Antigen) and GRP-R ( Gastrin Releasing Peptide Receptor) are overexpressed on prostatic cancer cells and can be targeted with a PSMA inhibitor such as 68Ga-PSMA-617 (Liu 2018) or a GRP-R antagonist such as 68Ga-RM2 (Touijer 2019). We aimed to compare 68Ga-PSMA-617 PET/CT and 68Ga-RM2 PET/CT at initial staging of prostate cancer. Methods: We prospectively enrolled 10 treatment-naïve patients with biopsy-confirmed prostate cancer. Fifteen lesions were found (two with Gleason score (GS) 6; seven GS 7(3+4), two GS 7(4+3) and four GS ≥ 8). 68Ga-PSMA-617 PET/CT and 68Ga-RM2 PET/CT were randomly performed before prostatectomy with imaging at 1 and 2 hours after injection (2MBq/kg). Manual (MS) and automatic (AS) segmentation were performed on each PET imaging and then compared to histology (standard-of- truth). Prostatectomy samples were analyzed by an experimented pathologist who identified and staged each lesion according to GS. Each prostatectomy slice available was scanned for comparison between PET and histologic segmentation. Uptake intensity (SUVmax) and signal-to-noise ratio (SNR) relative to non-lesional prostate gland parenchyma were analyzed. Per-lesion analysis was performed on final GS (prostatectomy). Results: Using AS, 68Ga-RM2 PET/CT showed better sensitivity (Se) than 68Ga-PSMA-617 PET/CT: (0.85 vs 0.75) and better specificity (Sp) (0.89 vs 0.60). MS found similar Se (0.94 vs 0.93) but better Sp (0.89 vs 0.80) with 68Ga-RM2 PET/CT. SNR of 68Ga-RM2 PET/CT was high whatever the GS was; while 68Ga-PSMA-617 PET/CT exhibited higher SNR in high GS (p = 0.044). Finally, in low GS, SNR of 68Ga-RM2 PET/CT was significantly higher than 68Ga-PSMA-617 PET/CT (p = 0.004). Conclusions: GRP-R-targeting and PSMA-targeting have been previously compared in patients with biochemically recurrence (Minamimoto 2016) but not at primary staging. Our initial results, in this indication, showed great diagnostic performance of 68Ga-RM2 PET/CT compared to 68Ga-PSMA-617 PET/CT to characterize low Gleason score tumors. 68Ga-PSMA-617 PET/CT is promising for initial staging of high Gleason score tumors. These results need to be confirmed in a larger study.

Abstract 334: Proteostasis by HSP70/STUB1 complex regulates androgen receptor variants expression and confers resistance to enzalutamide and abiraterone

Abstract INTRODUCTION AND OBJECTIVES: Proteostasis is a complementary process by which cells control the protein biosynthesis, folding, trafficking and degradation. Evidence suggests that proteomic instability, such as protein misfolding and aggregation plays pivotal roles in cancer cell survival and progression. AR-V7 lacking the ligand binding domain confers resistance to enzalutamide and abiraterone in prostate cancer, targeting AR-V7 is an logical strategy to overcome the resistance. The mechanisms of AR-V7 proteostasis haven’t been fully studied so far. The objectives of this project are to investigate the roles of proteostasis in regulating AR-V7 expression and sensitivity to enzalutamide and abiraterone treatment. METHODS: Expression of HSP70 and STUB1 was determined by qRT-PCR and western blot. Expression of HSP70 and STUB1 was downregulated using specific siRNA. HSP70/STUB1 and AR-V7 interaction was determined by co-immunoprecipitation and dual immunofluorescence. The gene regulating mechanisms underlying the HSP70 inhibition in drug resistant prostate cancer cells was determined by RNA sequencing analyses. The effects of HSP70 inhibition on enzalutamide sensitivity were examined in vitro and in vivo. The correlation between HSP70 and AR-V7 in high Gleason score prostate tumors was determined by qRT-PCR. RESULTS: In the present study, we analyzed enzalutamide and abiraterone resistant prostate cancer cells and found ubiquitin mediated proteolysis pathway was suppressed, and E3 ubiquitin ligases STUB1 is downregulated in enzalutamide and abiraterone resistant prostate cancer cells. STUB1 binds to AR-V7, degrades AR-V7 expression and suppresses its activity. HSP70, the STUB1 binding protein, also binds to AR-V7 and enhances AR-V7 transcriptional activity. Mechanistically, STUB1 disassociates HSP70 from AR-V7 binding and increases AR-V7 degradation. Targeting HSP70 by siRNA or small molecular inhibitors (Apoptozole and Ver155008) significantly suppressed prostate cancer growth and improved enzalutamide and abiraterone treatment through AR-V7 inhibition in vitro and in vivo. Additionally, HSP70 expression is upregulated in mCRPC tumors and correlates with AR-V7 levels in high Gleason score and metastatic prostate tumor specimens. CONCLUSION: Enzalutamide and abiraterone treatment induces the imbalance of AR-V7 proteostasis through the ubiquitin-proteolysis alteration. STUB1/HSP70 complex controls AR and AR variants proteostasis. Targeting HSP70 could be a valuable strategy to overcome the next generation anti-androgen resistance and improve their therapy. Citation Format: Chengfei Liu, Wei Lou, Joy C. Yang, Liangren Liu, Cameron M. Armstrong, Alan P. Lombard, Ruining Zhao, Onika DV Noel, Clifford G. Tepper, Hong-Wu Chen, Marc Dall’Era, Marc Dall’Era, Christopher P. Evans, Allen C. Gao. Proteostasis by HSP70/STUB1 complex regulates androgen receptor variants expression and confers resistance to enzalutamide and abiraterone [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 334.

Long noncoding RNA SNHG20 promotes prostate cancer progression via upregulating DDX17.

IntroductionAccumulating evidence has revealed the critical roles of long noncoding RNAs (lncRNAs) in various cancers. LncRNA SNHG20 has been shown to be a cancer-associated lncRNA in several cancers with diverse mechanisms. However, the clinical references, biological roles, and mechanisms of action of SNHG20 in prostate cancer (PCa) are still unclear.Material and methodsThe expression of SNHG20 in PCa tissues and cell lines was detected by RT-qPCR. The correlations between SNHG20 expression and clinicopathological features were analyzed by χ2 test. The roles of SNHG20 in PCa cell proliferation and migration were detected by CCK-8, EdU incorporation, and transwell assays. The regulatory mechanisms of SNHG20 on DDX17 were detected by dual luciferase reporter assay, RT-qPCR, and western blot.ResultsSNHG20 is highly expressed in PCa tissues and cell lines. High expression of SNHG20 is positively correlated with high Gleason score and advanced tumor stage. Functional experiments revealed that overexpression of SNHG20 promotes PCa cell proliferation and migration. SNHG20 knockdown represses PCa cell proliferation and migration. Mechanistically, SNHG20 was verified to act as a competing endogenous RNA (ceRNA) to upregulate DDX17. DDX17 is also highly expressed and has oncogenic roles in PCa. Furthermore, the expression of DDX17 is significantly positively correlated with that of SNHG20 in PCa tissues. Depletion of DDX17 reverses the oncogenic roles of SNHG20 in PCa.ConclusionsThese data showed that SNHG20 promotes PCa cell proliferation and migration via acting as a ceRNA to upregulate DDX17. This study also suggested that SNHG20 may be a potential novel therapeutic target for PCa.

Study the Expression of CD10 in Prostate Carcinoma and its Correlation with Various Clinicopathological Parameters.

Background and Objective:Adenocarcinoma of the prostate is the second most common cause of cancer. The loss of CD10 is a common early event in human prostate cancer and is seen in lower Gleason Score malignancies while increased and altered expression is seen in high Gleason Score tumors, lymph nodes and bone metastasis. Material and Methods:This was a prospective observational study conducted on 75 patients suspected to have prostate cancer. Immunohistochemical profile was assessed for PSA, AMACR and CD10 immunostaining. The intensity of CD10 expression and pattern of CD10 staining of tumor cells was evaluated.Results:The patients were in age group of 50-90 years with a mean age of 70.97 ± 9.51 years. As the Grade Group/Gleason Score increased, the number of cases showing negative expression decreased and the pattern of expression changed from membranous to cytoplasmic to both types of expression. As the serum PSA levels increased the intensity of expression changed from focally positive to diffusely positive. The pattern of expression also changed from membranous to cytoplasmic to both (membranous + cytoplasmic) types of expression with an increase in PSA levels.Conclusion:By immunohistochemical analysis we can identify CD10 positive tumors, which may warrant more aggressive initial therapy. A number of drugs against CD10 are available based on which potential targeted therapies could be formulated.

Abstract CT016: A first in man phase I/II adjuvant dendritic cell vaccine study in high-risk prostate cancer patients following radical surgery reduce the incidence of biochemical relapse

Abstract Prostate cancer patients diagnosed with high Gleason score (≥ 8) and large tumors (≥T2c) are considered high-risk patients and >50% will develop an early biochemical relapse following radical surgery. Presently, there is no curative therapy available for patients once when biochemical relapse occur. Based on encouraging clinical results from 6 relapsed prostate cancer patients treated under hospital exemption with dendritic cell (DC) vaccines we started an adjuvant, first in man, phase I/II study using autologous, monocyte derived DCs targeting autologous tumor antigens from primary tumor, combined with hTERT and survivin. Twenty patients were included in the study. All patients had pathological stage pT2-pT3b and Gleason score 7b-10. Following surgery prostate specific antigen (PSA) was < 0,2 µg/L in all patients. Fifteen patients have received 3-days DCs generated according to our standard protocol with a maturation cocktail composed of GM-CSF, IL-4, TNFα, IL1β and PgE2. Five patients were treated with a TLR 7/8 ligand containing maturation cocktail, resulting in DCs with a polarized release of IL-12p70 and no or low IL-10. The patients received 4 weekly vaccinations, then DTH vaccine at week 8 and thereafter-monthly vaccinations the first year, and every third month the second and third year. All patients except of two have completed vaccination. Of the 15 first patients using the standard DCs, three patients developed PSA relapse during vaccination (PSA measured twice >0,5 μg/L with minimum 4 weeks interval) and two additional patients relapsed after completed 3 years of vaccination. From the last 5 patients given the new type of DCs none have experienced PSA relapse. Seventy-five percent of the patients remain without biochemical relapse with a mean observation time of 47,5 (range 29-82) months. We confirm that the study is feasible and safe. Immune responses in the patients are under investigation. Altogether, our clinical results are promising and the use of adjuvant DC vaccines might become a new approach to prevent biochemical relapse in high-risk prostate cancer patients following radical surgery. Citation Format: Anne Merete Tryggestad, Iris Bigalke, Karol Axcrona, Bjørn Brennhovd, T Kirsti Hønnåshagen, Lisbeth Skoge, Lena Tjeldhorn, Lene Mowinckel, Stein Sæbøe-Larsen, Jens A L Jørgensen, Grete S Andreassen, Gjertrud Skorstad, Dag Josefsen, Ulrika Axcrona, Steinar Aamadal, Wolfgang Lilleby, Svein Dueland, Gunnar Kvalheim. A first in man phase I/II adjuvant dendritic cell vaccine study in high-risk prostate cancer patients following radical surgery reduce the incidence of biochemical relapse [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT016.

CDC6 mRNA Expression Is Associated with the Aggressiveness of Prostate Cancer.

BackgroundCell division cycle 6 (CDC6) is an essential regulator of DNA replication and plays important roles in the activation and maintenance of the checkpoint mechanisms in the cell cycle. CDC6 has been associated with oncogenic activities in human cancers; however, the clinical significance of CDC6 in prostate cancer (PCa) remains unclear. Therefore, we investigated whether the CDC6 mRNA expression level is a diagnostic and prognostic marker in PCa.MethodsThe study subjects included 121 PCa patients and 66 age-matched benign prostatic hyperplasia (BPH) patients. CDC6 expression was evaluated using real-time polymerase chain reaction and immunohistochemical (IH) staining, and then compared according to the clinicopathological characteristics of PCa.ResultsCDC6 mRNA expression was significantly higher in PCa tissues than in BPH control tissues (P = 0.005). In addition, CDC6 expression was significantly higher in patients with elevated prostate-specific antigen (PSA) levels (> 20 ng/mL), a high Gleason score, and advanced stage than in those with low PSA levels, a low Gleason score, and earlier stage, respectively. Multivariate logistic regression analysis showed that high expression of CDC6 was significantly associated with advanced stage (≥ T3b) (odds ratio [OR], 3.005; confidence interval [CI], 1.212–7.450; P = 0.018) and metastasis (OR, 4.192; CI, 1.079–16.286; P = 0.038). Intense IH staining for CDC6 was significantly associated with a high Gleason score and advanced tumor stage including lymph node metastasis stage (linear-by-linear association, P = 0.044 and P = 0.003, respectively).ConclusionCDC6 expression is associated with aggressive clinicopathological characteristics in PCa. CDC6 may be a potential diagnostic and prognostic marker in PCa patients.

The oncogenic role of the spliced somatostatin receptor sst5TMD4 variant in prostate cancer.

sst5TMD4, a splice variant of the sst5 gene, is overexpressed and associated with aggressiveness in various endocrine-related tumors, but its presence, functional role, and mechanisms of actions in prostate cancer (PCa)-the most common cancer type in males-is completely unexplored. In this study, formalin-fixed, paraffin-embedded prostate pieces from patients with localized PCa, which included tumoral and nontumoral adjacent regions (n = 45), fresh biopsies from patients with high-risk PCa (n = 52), and healthy fresh prostates from cystoprostatectomies (n = 14) were examined. In addition, PCa cell lines and xenograft models were used to determine the presence and functional role of sst5TMD4. Results demonstrated that sst5TMD4 is overexpressed (mRNA/protein) in PCa samples, and this is especially drastic in metastatic and/or high Gleason score tumor samples. Remarkably, sst5TMD4 expression was associated with an altered frequency of 2 single-nucleotide polymorphisms: rs197055 and rs12599155. In addition, PCa cell lines and xenograft models were used to demonstrate that sst5TMD4 overexpression increases cell proliferation and migration in PCa cells and induces larger tumors in nude mice, whereas its silencing decreased proliferation and migration. Remarkably, sst5TMD4 overexpression activated multiple intracellular pathways (ERK/JNK, MYC/MAX, WNT, retinoblastoma), altered oncogenes and tumor suppressor gene expression, and disrupted the normal response to somatostatin analogs in PCa cells. Altogether, we demonstrate that sst5TMD4 is overexpressed in PCa, especially in those patients with a worse prognosis, and plays an important pathophysiologic role in PCa, which suggesting its potential as a biomarker and/or therapeutic target.-Hormaechea-Agulla, D., Jiménez-Vacas, J. M., Gómez-Gómez, E., L.-López, F., Carrasco-Valiente, J., Valero-Rosa, J., Moreno, M. M., Sánchez-Sánchez, R., Ortega-Salas, R., Gracia-Navarro, F., Culler, M. D., Ibáñez-Costa, A., Gahete, M. D., Requena, M. J., Castaño, J. P., Luque, R. M. The oncogenic role of the spliced somatostatin receptor sst5TMD4 variant in prostate cancer.

Abstract 226: Verification of prostate cancer genomics biomarker candidates at protein level using PRISM-SRM

Abstract Mass spectrometry (MS) based targeted proteomics such as selected reaction monitoring (SRM) provides an antibody-independent strategy for sensitive, specific and multiplexed verification of genomics biomarker candidates at the protein level. In order to identify a panel of proteins with the potential to discriminate between aggressive and indolent forms of prostate cancer and predict prostate cancer progression, we have selected 52 protein candidates from existing prostate cancer genomics data sets and validated cancer drivers, and performed quantitative proteomics analysis in tumor and control tissue samples using the highly sensitive PRISM (high-pressure, high-resolution separations coupled with intelligent selection and multiplexing)-SRM approach. PRISM-SRM assays have been developed for the 52 prostate cancer biomarker candidates including: prostate cancer prognosis associated genes, prostate cancer associated genes that were up-regulated in transcriptomics studies, and other cancer-related genes (including the ERG or ETV1 isoforms). Two sets of tissue samples were analyzed using PRISM-SRM with heavy isotope-labeled synthetic peptides as internal standards: 1) 10 high Gleason-score (7-9) primary prostate tumors and 10 benign prostatic hyperplasia (BPH) tissues (OCT-embedded specimens); and 2) 10 primary tumors from patients showing metastatic progression, 10 primary tumors from patients who showed biochemical recurrence (BCR), and 10 primary tumors from patients with no BCR or metastatic progression after more than ten years of follow-up after radical prostatectomy (FFPE whole mount prostate specimens). Overall, PRISM-SRM analyses of all the patient tissue samples enabled the detection of 48 out of 52 biomarker candidates, suggesting extremely low level of expression of the remaining 6 genes (HXC6, OSTP, TWST1, and ERG8); in comparison regular LC-SRM can only detect 21 of these candidates at the protein level. In the 10 high Gleason-score tumors and 10 BPH controls, 13 proteins were found differentially abundant with P<0.05. In the 10X10X10 FFPE sample analysis, there were three proteins discriminating between “metastatic progression” and “no progression” tumors, one protein discriminated between BCR and “no progression” tumors, and four proteins discriminated between metastatic progression and BCR tumors (P<0.05). These promising biomarker candidates will be further evaluated, individually and in panels, in independent, larger cohort for their potential prognostic applications. In summary, PRISM-SRM provides a highly sensitive method for quantification and rapid screening of multiple potential biomarker candidates at the protein level. This approach holds great potential for rapidly translating genomics-based discovery candidates into protein-based biomarkers. Citation Format: Hui Wang, Yuqian Gao, Athena Schepmoes, Gyorgy Petrovics, Jennifer Cullen, Thomas Fillmore, Tujin Shi, Wei-Jun Qian, Richard Smith, Brandi Weaver, Robin Leach, Ian Thompson, Sudhir Srivastava, Jacob Kagan, Albert Dobi, Karin Rodland, Shiv Srivastava, Tao Liu. Verification of prostate cancer genomics biomarker candidates at protein level using PRISM-SRM [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 226. doi:10.1158/1538-7445.AM2017-226

Epigenetic signature of Gleason score and prostate cancer recurrence after radical prostatectomy.

BackgroundIdentifying the subset of patients with clinically localized prostate cancer (PCa) at the highest risk of recurrence remains challenging, and better prognostic markers are needed. Gleason score is the best predictor of PCa aggressiveness and prognosis. In the present study, we generated an epigenetic signature based on high versus low Gleason score tumors and evaluated its ability to predict recurrence after radical prostatectomy.MethodsGenome-wide DNA methylation data from The Cancer Genome Atlas (TCGA; no. of patients = 333) and the elastic net method were used to generate an epigenetic signature by contrasting patients with high (8–10) versus low (≤6) Gleason score tumors. The signature was then tested in a cohort of 523 patients with clinically localized disease who had radical prostatectomy. Samples taken from the primary tumor were used for DNA methylation and mRNA expression profiling. Patients were followed for PCa recurrence on average for 8 years after diagnosis.ResultsThe epigenetic signature includes 52 differentially methylated CpG sites. In the testing cohort, the signature was associated with poorer recurrence-free survival (hazard ratio per 25 % increase = 1.78; 95 % confidence interval 1.48, 2.16). The signature significantly improved the area under the curve (AUC) for PCa recurrence compared to clinical-pathological parameters alone, particularly among patients diagnosed with Gleason score 7 tumors (0.64 vs. 0.76, P = 1.34E−4). Results were comparable for patients with Gleason 3 + 4 and those with 4 + 3 tumors. Gene Set Enrichment Analysis showed that higher levels of the signature were associated with increased expression of genes related to cell cycle proliferation and decreased expression of androgen-responsive genes.ConclusionsThis report shows evidence that DNA methylation patterns measured in prostate tumor cells are predictive of PCa aggressiveness. The epigenetic signature may have clinical utility to improve prognostication particularly in patients with intermediate Gleason score 7 tumors.Electronic supplementary materialThe online version of this article (doi:10.1186/s13148-016-0260-z) contains supplementary material, which is available to authorized users.

Asparaginyl endopeptidase promotes proliferation and invasiveness of prostate cancer cells via PI3K/AKT signaling pathway

Recurrence and metastasis are the major lethal causes of prostate cancer. It is urgent to find out the mechanisms and key factors governing prostate cancer progression and metastasis for developing new therapeutic strategies. Asparaginyl endopeptidase (AEP) overexpression has been found in a number of solid tumors. In prostate cancer, AEP has also been shown to exhibit a vesicular staining pattern and significantly associated with advanced tumor stage, high Gleason score, perineural invasion, and larger tumor. Here, we found that AEP was differentially expressed in prostate cancer cells with higher expression in 22RV1 cells and lower expression in PC-3 cells. AEP knockdown in 22RV1 cells significantly inhibited cell proliferation and invasion abilities while overexpression of AEP in PC-3 cells prompted cell proliferation and invasion abilities. Meanwhile, AEP knockdown upregulated cell apoptosis and vice versa. Further, we firstly identified that AEP promotes activation of the PI3K-AKT signaling pathway in prostate cancer cells. Taken together, our results suggest that AEP may be an attractive target for prostate cancer therapy.

Abstract 6: Metabolomics of Gleason score in prostate tumor tissue and serum

Abstract Background: Gleason score is currently the best clinical predictor of prostate cancer specific-mortality. Identifying biomarkers associated with Gleason score can improve prediction and advance the understanding of the biology of aggressive disease. Using gene expression data, we previously determined that metabolism pathways, including pyrimidine, propanoate, and beta-alanine, were differentially expressed in high and low grade tumors. The goal of this study is to identify metabolites that differ in low and high Gleason score tumors and in blood from these patients. Methods: Metabolic profiling was performed on Gleason score 6 (n = 53) and Gleason score ≥8 (n = 32) radical prostatectomy tumor tissue specimens, and on serum samples from the time of diagnosis (n = 30 Gleason score 6 and n = 19 Gleason score ≥8) from the DF/HCC Prostate Cancer SPORE Cohort. Samples were prepared for analysis by Metabolon, Inc. with their standard solvent extraction method to recover small molecules. Metabolites were identified and quantified by gas and liquid chromatography and mass spectrometry. Missing values were calculated using k-nearest neighbor imputation, and sample values were normalized across batches. The metabolite levels were compared across Gleason score 6 and Gleason score ≥8 tumors and serum using a Wilcoxon test. Results: In the tumor tissue, 207 metabolites were identified. Of these, 30 were present in significantly different amounts in high and low Gleason score tumors, including previously reported citrate (p = 0.002) and spermine (p = 0.003). In a pathway analysis, the propanoate pathway was significantly different in high and low Gleason (p = 0.03), but not the pyrimidine or beta-alanine pathways (p>0.05). In serum, 15 individual metabolites had significantly different levels, including uridine (p = 0.008) and N-acetylserine (p = 0.0001). Conclusions: Metabolite levels do differ in high and low Gleason score prostate tumor tissue. The novel serum results suggest that metabolites in blood may serve as biomarkers of tumor differentiation. Our ongoing research will build prediction models, and will determine if the blood metabolites can indicate the presence of high-Gleason score tumor not detected at biopsy and could serve as biomarkers for monitoring disease progression in active surveillance patients. Citation Format: Kathryn Penney, Svitlana Tyekucheva, Massimo Loda. Metabolomics of Gleason score in prostate tumor tissue and serum. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 6.

Abstract 2235: Clinical results of a Phase I/II trial of adjuvant therapeutic vaccination in high risk resected prostate cancer patients using autologous dendritic cells loaded with mRNA from primary prostate cancer tissue, hTERT and survivin

Abstract Prostate cancer patients diagnosed with high Gleason score (≥ 8) and large tumors (≥T2c) are considered high-risk patients and >50% will develop an early biochemical relapse. Presently, there is no curative therapy available for patients with biochemical relapse. Based on these findings we initiated in January 2011 a Phase I/II dendritic cell (DC) vaccine study. Patients included have pathological stage pT2 - pT3b, Gleason score 7b-10, pN0, pN+ or pNx and postoperative PSA < 0.2 μg/L. Following surgery autologous tumor cell lines were established from each patient using an in-house culturing method. mRNA from the tumor cell line was produced and used for DC vaccination in combination with mRNA hTERT and mRNA Survivin. DCs were differentiated from enriched monocytes, cultured for 2 days with IL4 and GM-CSF and matured with Jonuleit-maturation cocktail for 24 hours. The matured DCs were transfected separately with the 3 different mRNAs and then frozen and stored until use. The vaccination regimen includes one vaccine per week for four weeks, followed by monthly “vaccine boost” during the first year, then every 3 months the second and third year. Recently, a novel 3 days DC protocol using a TLR7/8-agonist maturation cocktail has been implemented at our department. Based on encouraging clinical results with this type of DCs in compassionate use patients with different types of tumors, we decided to change our DCs protocol to the new generation DCs. Of the 20 patients included in this trial 15 patients have been given the standard fast DCs and 5 have been vaccinated with the new type of DCs. 8 Patients given standard DC has completed 3 years of vaccination and 4 has completed 2 years of vaccination. 3 of 15 patient given standard DC has experienced PSA relapse during vaccination. None of the patients given the new type of DC has so far experienced raise in PSA levels. To our knowledge this is the first adjuvant DC vaccine study in high risk prostate cancer and we conclude that the study is feasible, safe and utmost promising. Extensive immune monitoring is ongoing taking advantage of the established autologous tumor cell lines from all patients. Citation Format: Anne Merete Aa. Tryggestad, Karol Axcrona, Iris Bigalke, Else M. Inderberg-Suso, Gjertrud Skorstad, Ulrika Axcrona, Steinar Aamdal, Gustav Gaudernack, Dolores Schendel, Wolfgang Lilleby, Svein Dueland, Gunnar Kvalheim. Clinical results of a Phase I/II trial of adjuvant therapeutic vaccination in high risk resected prostate cancer patients using autologous dendritic cells loaded with mRNA from primary prostate cancer tissue, hTERT and survivin. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2235.

Preoperative erectile function and the pathologic features of prostate cancer

Purpose We evaluated whether preoperative erectile function is associated with pathologic features in the patients who underwent radical prostatectomy (RP).Materials and Methods We reviewed medical records of 1,743 men who underwent RP from November 2003 through May 2012. Of these, 50 patients who had prior hormone therapy and 272 patients who had lacking data of International Index of Erectile Function-5 (IIEF-5) were excluded. Men whose IIEF-5 was in the lower 25 percentile were assigned as Low Erectile Function group and the others were assigned as Control group. We compared pathologic features using univariable and multivariable logistic regression analysis between two groups.Results A total of 1,421 patients were included in the analysis. Patients’ age was 65.8 ± 6.7 years and prostate-specific antigen (PSA) was 12.8±16.1 ng/mL. Median and low 25 percentile of IIEF-5 were 14 and 8, respectively. Low Erectile Function group (IIEF-5<8) had higher risk to have high Gleason score (≥7(4+3), odds ratio (OR) 1.642, p<0.001) and large tumor volume (≥5 mL, OR 1.292, p=0.042). Even after adjusting age, year of surgery, body mass index, Charlson comorbidity index, PSA, clinical stage and biopsy Gleason score, Low Erectile Function group still had higher risk of high Gleason score (OR 1.910, p<0.001) and large tumor volume (OR 1.390, p=0.04) by multivariable logistic regressions.Conclusions Lower erectile function before RP was associated with higher Gleason’s score and larger tumor volume in final pathology. Thus, erectile function could be a surrogate barometer for prostate cancer aggressiveness.

Immunohistological analysis of ABCD3 expression in Caucasian and African American prostate tumors.

In a previously published study, we showed that expression of the ABCD3 gene increased with increasing metastatic potential in a panel of prostate cancer cell lines derived from African American and Caucasian American men. Given importance of identifying biomarker(s) that can distinguish indolent versus aggressive prostate tumors, we conducted an immunohistochemical analysis of ABCD3 expression Caucasian and African American prostate tumors. ABCD3 expression in each patient population was compared with clinicopathologic characteristics, Gleason score, and age. ABCD3 expression increased with increasing Gleason score (P = 0.0094), age (P = 0.0014), and pathology grade (P = 0.0007) in Caucasian patients. Interestingly, in the AA patients, ABCD3 expression highly increased to the same degree in both low and high Gleason score tumors. Similarly, ABCD3 expression was elevated to the same degree in BPH derived from AA. Our findings demonstrate that increased ABCD3 expression correlates with Gleason Score in CA prostate tumors. However, in AA prostate tumors, ABCD3 expression was higher and was sustained in both low Gleason and high Gleason AA tumors. While the functional role of ABCD3 in prostate cancer is not completely elucidated, this gene warrants further study as a potential biomarker for aggressive prostate.

Influence of peripheral whole-blood microRNA-7 and microRNA-221 high expression levels on the acquisition of castration-resistant prostate cancer: evidences from in vitro and in vivo studies.

Prostate cancer (PC) is the more frequently diagnosed neoplasia in men in developed countries. The evolution of PC to castration-resistant prostate cancer (CRPC) represents real problems of clinical management, in consequence to the limited therapeutic options. MicroRNAs (miRNAs) are small noncoding RNAs that play an important role in gene expression and function regulation. The increased evidence that miRNAs are involved in cancer development and progression has made them potential biomarkers for cancer diagnosis, prognosis, and aggressiveness. Our purpose was to identify a miRNA expression profile associated with the development of CRPC. We firstly observed a miRNA expression profile differentially expressed between the castration-resistant (CR) PC3 cell line and the hormone-sensitive LnCaP cell line, where miR-7, miR-221, and miR-222 were upregulated in PC3 (11.3-fold increase, P = 0.012; 11.3-fold increase, P = 0.002; 8.6-fold increase, P = 0.002, respectively). We also observed that the trend of miR-1233 expression levels was higher in PC3 (3.7-fold increase, P = 0.057). These miRNAs differentially expressed in vitro were studied in a peripheral whole-blood samples from PC patients. We observed that patients presenting an early CR acquisition (≤ 20 months) had higher expression levels of miR-7 and miR-221 (P = 0.034 and P = 0.036, respectively). Furthermore, we found that patients diagnosed with high-Gleason score tumors and presenting simultaneous higher miR-7 expression levels have a significant reduce time to CR compared with patients who present lower miR-7 expression levels (11 vs. 51 months, log-rank test P = 0.004). We also found that patients diagnosed with high-Gleason score tumors and higher expression levels of miR-221 have an early CRPC compared to patients with lower miR-221 expression levels (10 vs. 46 months, log-rank test P = 0.012). We observed a significantly lower overall survival in patients with higher peripheral whole-blood expression levels of miR-7 (28 vs. 116 months, log-rank test P = 0.001). Our results suggest that miR-7 and miR-221 peripheral whole-blood expression levels can be potential predictive biomarkers of CRPC development.

MP53-01 PERFORMANCE OF MULTI-PARAMETRIC MRI FOR HIGH GRADE PROSTATE CANCER: CORRELATION WITH WHOLE-MOUNT PATHOLOGY

INTRODUCTION AND OBJECTIVES: Higher Gleason score (GS) is associated with increased level of suspicion on multi-parametric magnetic resonance imaging (mp-MRI) of the prostate, but some tumors still evade detection by MRI. The objective of this study was to characterize the performance of mp-MRI for detection of high GS tumors and identify risk factors for missed tumor detection. METHODS: A retrospective study was performed of 122 consecutive men who underwent mp-MRI prior to radical prostatectomy from 10/2010-2/2013. Clinical, MRI, and pathologic features were obtained. A genitourinary radiologist and pathologist collectively reviewed each case and matched the MRI to whole-mount pathology lesions. Index tumor was defined as the highest GS tumor; in cases of multifocal tumors of identical GS, the largest tumor was considered the index. Chi-square or Fisher’s exact analysis was performed for categorical and t-test for continuous variables. RESULTS: 180 regions of interest were identified by mp-MRI and 283 tumors were histologically confirmed among 122 men. Among the 57 tumors determined to be GS 4+3, 15/57 tumors (26%) were missed by mp-MRI in 11/50 men (22%), corresponding to a sensitivity of 74%. Index tumor status, tumor size, number of whole mount levels involved, and presence of extra-capsular extension were all associated with tumor detection (p<0.05) (table). 37/44 (84%) of index tumors were correctly identified by mp-MRI. Missed tumors were: less likely to be index tumors (47% vs 88% of MRI-pathology concordant tumors); likely to be smaller (1.24 cm 0.78 cm SD vs 2.41 2.06 SD), involving fewer whole-mount levels; and less likely to have extra-capsular extension (13% vs 55%). Of missed non-index tumors, 4/8 (50%) of the corresponding index tumors were detected by mp-MRI. Of the missed index tumors, no corresponding non-index tumors were detected; review of these slides demonstrated tumor infiltrating benign glands (3/7), small volume tumor within large overall prostate volume (3/7), or abundant foamy gland variant pathology (2/7) as possible reasons for missed tumor detection. CONCLUSIONS: High Gleason score tumors evaded mp-MRI detection in 26% of cases. Smaller, non-index, organ-confined tumors were more likely to be missed; there may be a histologic basis for nonvisualization of tumors by mp-MRI. Table Characteristics of tumors missed versus detected by mp-MRI.

MP53-02 IMPACT OF MULTIFOCALITY ON PROSTATE CANCER DETECTION BY MULTI-PARAMETRIC MRI

INTRODUCTION AND OBJECTIVES: Higher Gleason score (GS) is associated with increased level of suspicion on multi-parametric magnetic resonance imaging (mp-MRI) of the prostate, but some tumors still evade detection by MRI. The objective of this study was to characterize the performance of mp-MRI for detection of high GS tumors and identify risk factors for missed tumor detection. METHODS: A retrospective study was performed of 122 consecutive men who underwent mp-MRI prior to radical prostatectomy from 10/2010-2/2013. Clinical, MRI, and pathologic features were obtained. A genitourinary radiologist and pathologist collectively reviewed each case and matched the MRI to whole-mount pathology lesions. Index tumor was defined as the highest GS tumor; in cases of multifocal tumors of identical GS, the largest tumor was considered the index. Chi-square or Fisher’s exact analysis was performed for categorical and t-test for continuous variables. RESULTS: 180 regions of interest were identified by mp-MRI and 283 tumors were histologically confirmed among 122 men. Among the 57 tumors determined to be GS 4+3, 15/57 tumors (26%) were missed by mp-MRI in 11/50 men (22%), corresponding to a sensitivity of 74%. Index tumor status, tumor size, number of whole mount levels involved, and presence of extra-capsular extension were all associated with tumor detection (p<0.05) (table). 37/44 (84%) of index tumors were correctly identified by mp-MRI. Missed tumors were: less likely to be index tumors (47% vs 88% of MRI-pathology concordant tumors); likely to be smaller (1.24 cm 0.78 cm SD vs 2.41 2.06 SD), involving fewer whole-mount levels; and less likely to have extra-capsular extension (13% vs 55%). Of missed non-index tumors, 4/8 (50%) of the corresponding index tumors were detected by mp-MRI. Of the missed index tumors, no corresponding non-index tumors were detected; review of these slides demonstrated tumor infiltrating benign glands (3/7), small volume tumor within large overall prostate volume (3/7), or abundant foamy gland variant pathology (2/7) as possible reasons for missed tumor detection. CONCLUSIONS: High Gleason score tumors evaded mp-MRI detection in 26% of cases. Smaller, non-index, organ-confined tumors were more likely to be missed; there may be a histologic basis for nonvisualization of tumors by mp-MRI. Table Characteristics of tumors missed versus detected by mp-MRI.