The third symposium on treatment-induced neuroendocrine prostate cancer: insights and future directions.

Neuroendocrine prostate cancer (NEPC) is a rare and aggressive subtype of prostate cancer (PCa), emerging from advanced treatments and characterized by loss of androgen receptor (AR) signaling and neuroendocrine features, leading to rapid progression and treatment resistance. The third symposium on treatment-induced NEPC, held from 21 to 23 June 2024, at Harrison Hot Springs Resort, BC, Canada, united leading global researchers and clinicians. Sponsored by the Vancouver Prostate Centre (VPC), Canadian Institute of Health Research, Prostate Cancer Foundation Canada and Pharma Planter Inc, the event focused on the latest NEPC research and innovative treatment strategies. Co-chaired by Drs. Yuzhuo Wang and Martin Gleave, the symposium featured sessions on NEPC's historical context, molecular pathways, epigenetic regulation and the role of the tumor microenvironment and metabolism in its progression. Keynotes from experts like Dr. Himisha Beltran and Dr. Martin Gleave highlighted the complexity of NEPC. The Emerging Talent session showcased new research, pointing to the future of NEPC treatment. The symposium concluded with a consensus on the need for early detection, targeted therapies and personalized medicine to effectively combat NEPC, emphasizing the importance of global collaboration in advancing NEPC understanding and treatment.

Natural compound Alternol exerts a broad anti-cancer spectrum and a superior therapeutic safety index in vivo.

Introduction: Alternol is a natural compound isolated from the fermentation of a mutated fungus. We have demonstrated its potent anti-cancer effect via the accumulation of radical oxygen species (ROS) in prostate cancer cells in vitro and in vivo. In this study, we tested its anti-cancer spectrum in multiple platforms. Methods: We first tested its anti-cancer spectrum using the National Cancer Institute-60 (NCI-60) screening, a protein quantitation-based assay. CellTiter-Glo screening was utilized for ovarian cancer cell lines. Cell cycle distribution was analyzed using flow cytometry. Xenograft models in nude mice were used to assess anti-cancer effect. Healthy mice were tested for the acuate systemic toxicity. Results: Our results showed that Alternol exerted a potent anti-cancer effect on 50 (83%) cancer cell lines with a GI50 less than 5 µM and induced a lethal response in 12 (24%) of those 50 responding cell lines at 10 µM concentration. Consistently, Alternol displayed a similar anti-cancer effect on 14 ovarian cancer cell lines in an ATP quantitation-based assay. Most interestingly, Alternol showed an excellent safety profile with a maximum tolerance dose (MTD) at 665 mg/kg bodyweight in mice. Its therapeutic index was calculated as 13.3 based on the effective tumor-suppressing doses from HeLa and PC-3 cell-derived xenograft models. Conclusion: Taken together, Alternol has a broad anti-cancer spectrum with a safe therapeutic index in vivo.

Natural compound Alternol actives multiple endoplasmic reticulum stress-responding pathways contributing to cell death.

Background: Alternol is a small molecular compound isolated from the fermentation of a mutant fungus obtained from Taxus brevifolia bark. Our previous studies showed that Alternol treatment induced reactive oxygen species (ROS)-dependent immunogenic cell death. This study conducted a comprehensive investigation to explore the mechanisms involved in Alternol-induced immunogenic cell death. Methods: Prostate cancer PC-3, C4-2, and 22RV1 were used in this study. Alternol interaction with heat shock proteins (HSP) was determined using CETSA assay. Alternol-regulated ER stress proteins were assessed with Western blot assay. Extracellular adenosine triphosphate (ATP) was measured using ATPlite Luminescence Assay System. Results: Our results showed that Alternol interacted with multiple cellular chaperone proteins and increased their expression levels, including endoplasmic reticulum (ER) chaperone hypoxia up-regulated 1 (HYOU1) and heat shock protein 90 alpha family class B member 1 (HSP90AB1), as well as cytosolic chaperone heat shock protein family A member 8 (HSPA8). These data represented a potential cause of unfolded protein response (UPR) after Alternol treatment. Further investigation revealed that Alternol treatment triggered ROS-dependent (ER) stress responses via R-like ER kinase (PERK), inositol-requiring enzyme 1α (IRE1α). The double-stranded RNA-dependent protein kinase (PKR) but not activating transcription factor 6 (ATF6) cascades, leading to ATF-3/ATF-4 activation, C/EBP-homologous protein (CHOP) overexpression, and X-box binding protein XBP1 splicing induction. In addition, inhibition of these ER stress responses cascades blunted Alternol-induced extracellular adenosine triphosphate (ATP) release, one of the classical hallmarks of immunogenic cell death. Conclusion: Taken together, our data demonstrate that Alternol treatment triggered multiple ER stress cascades, leading to immunogenic cell death.

Chemokine receptor 7 contributes to T- and B-cell filtering in ageing bladder, cystitis and bladder cancer.

BACKGROUND: Research has suggested significant correlations among ageing, immune microenvironment, inflammation and tumours. However, the relationships among ageing, immune microenvironment, cystitis and bladder urothelial carcinoma (BLCA) in the bladder have rarely been reported. METHODS: Bladder single-cell and transcriptomic data from young and old mice were used for immune landscape analysis. Transcriptome, single-cell and The Cancer Genome Atlas Program datasets of BLCA and interstitial cystitis/bladder pain syndrome (IC/BPS) were used to analyse immune cell infiltration and molecular expression. Bladder tissues from mice, IC/BPS and BLCA were collected to validate the results. RESULTS: Eight types of immune cells (macrophages, B-cells, dendritic cells, T-cells, monocytes, natural killer cells, γδ T-cells and ILC2) were identified in the bladder of mice. Aged mice bladder tissues had a significantly higher number of T-cells, γδ T-cells, ILC2 and B-cells than those in the young group (P < 0.05). Three types of T-cells (NK T-cells, γδ T-cells and naïve T-cells) and three types of B-cells (follicular B-cells, plasma and memory B-cells) were identified in aged mice bladder. Chemokine receptor 7 (CCR7) is highly expressed in aged bladder, IC/BPS and BLCA (P < 0.05). CCR7 is likely to be involved in T- and B-cell infiltration in aged bladder, IC/BPS and BLCA. Interestingly, the high CCR7 expression on BLCA cell membranes was a prognostic protective factor. CONCLUSIONS: In this study, we characterised the expression profiles of immune cells in bladder tissues of aged and young mice and demonstrated that CCR7-mediated T- and B-cell filtration contributes to the development of bladder ageing, IC/BPS and BLCA.

Expressions of glucose transporter genes are diversely attenuated and significantly associated with prostate cancer progression.

Prostate cancer is a health-threaten disease in men worldwide, however, lacking is the reliable biomarkers for patient management. Aberrant metabolic events including glucose metabolism are involved in prostate cancer progression. To examine the involvement of glucose metabolic pathways in prostate cancer, we analyzed the expression profiles of glucose transporter family genes using multiple RNA-seq datasets. Our results showed that three SLC2A family genes (SLC2A4/5/9) were significantly downregulated in primary prostate cancers compared to their benign compartments. These down-regulated expressions were inversely correlated with their gene promoter methylation and genome abnormalities. Among these three SLC2A genes, only SLC2A4 showed a significantly reverse correlation with all clinicopathological parameters, including TNM stage, disease relapse, Gleason score, disease-specific survival, and progression-free interval. In addition, the expression levels of these three genes were strongly correlated with anti-cancer immune cell filtration in primary prostate cancers. In a group of patients with early-onset prostate cancers, SLC2A4 also showed a strong negative correlation with multiple clinicopathological parameters, such as tumor mutation burden, biochemical relapse, pre-surgical PSA levels, and Gleason score but a positive correlation with progression-free interval after surgery. In metastatic castration-resistant prostate cancers (CRPC), SLC2A9 gene expression but not SLC2A4 or SLC2A5 genes showed a significant correlation with androgen receptor (AR) activity score and neuroendocrinal (NE) activity score. Meanwhile, SLC2A2/9/13 expression was significantly elevated in CRPC tumors with neuroendocrinal features compared to those without NE features. On the other hand, SLC2A10 and SlC2A12 gene expression were significantly reduced in NEPC tumors compared to CRPC tumors. Consistently, SLC2A10/12 expression levels were significantly reduced in castrated animals carrying the LuCaP35 xenograft models. Survival outcome analysis revealed that SLC2A4 expression in primary tumors is a favorable prognostic factor and SLC2A6 is a worse prognostic factor for disease-specific survival and progression-free survival in prostate cancer patients. In conclusion, our results suggest that SLC2A4/6 expressions are strong prognostic factors for prostate cancer progression and survival. The significance of SLC2A2/9/13 over-expression during NEPC progression needs more investigation.

Aberrant expression of multiple glycolytic enzyme genes is significantly associated with disease progression and survival outcomes in prostate cancers.

Prostate cancer is the leading cause of cancer death after lung cancer in men. Recent studies showed that aberrant metabolic pathways are involved in prostate cancer development and progression. In this study, we performed a systemic analysis of glycolytic enzyme gene expression using the TCGA-PRAD RNAseq dataset. Our analysis revealed that among 25 genes, only four genes (HK2/GPI/PFKL/PGAM5) were significantly upregulated while nine genes (HK1/GCK/PFKM/PFKP/ALDOC/PGK1/PGAM1/ENO2/PKM) were downregulated in primary prostate cancer tissues compared to benign compartments. Among these 13 altered genes, four genes (ENO2/ALDOC/GPI/GCK) exhibited strong diagnostic potential in distinguishing malignant and benign tissues. Meanwhile, GPI expression exerted as a prognostic factor of progression-free and disease-specific survival. PFKL and PGAM5 gene expressions were associated with AR signaling scores in castration-resistant patients, and AR-targeted therapy suppressed their expression. In LuCap35 xenograft tumors, PFKL and PGAM5 expression was significantly reduced after animal castration, confirming the AR dependency. Conversely, GCK/PKLR genes were significantly associated with neuroendocrinal progression, representing two novel neuroendocrinal biomarkers for prostate cancer. In conclusion, our results suggest that GPI expression is a strong prognostic factor for prostate cancer progression and survival while GCK/PKLR are two novel biomarkers of prostate cancer progression to neuroendocrinal status.

Red blood cell membrane-coated functionalized Au nanocage as a biomimetic platform for improved MicroRNA delivery in hepatocellular carcinoma.

Dysregulation of microRNAs (miRNAs) expression is closely related to cancers and managing miRNA expression holds great promise for cancer therapy. However, their wide clinical application has been hampered by their poor stability, short half-life and non-specific biodistribution in vivo. Herein, a novel biomimetic platform designated as RHAuNCs-miRNA for improved miRNA delivery was prepared through wrapping miRNA-loaded functionalized Au nanocages (AuNCs) with red blood cell (RBC) membrane. RHAuNCs-miRNA not only successfully loaded miRNAs but also effectively protected them from enzymatic degradation. With good stability, RHAuNCs-miRNA had the characteristics of photothermal conversion and sustained release. Cellular uptake of RHAuNCs-miRNA by SMMC-7721 cells was in a time-dependent manner via clathrin- and caveolin-mediated endocytosis. The uptake of RHAuNCs-miRNAs was affected by cell types and improved by mild near infrared (NIR) laser irradiation. More importantly, RHAuNCs-miRNA exhibited a prolonged circulation time without the occurrence of accelerated blood clearance (ABC) in vivo, resulting in efficient delivery to tumor tissues. This study may demonstrate the great potential of RHAuNCs-miRNA for improved miRNAs delivery.

Jumonji domain-containing protein RIOX2 is overexpressed and associated with worse survival outcomes in prostate cancers.

Background: Histone demethylase RIOX2 was cloned as a c-Myc downstream gene involved in cell proliferation and has been implicated as an oncogenic factor in multiple tumor types. Its expression profiles and correlation with disease progression in prostate cancers are unknown. Methods: Transcriptomic profiles of Jumanji domain-containing protein genes were assessed using multiple public expression datasets generated from RNA-seq and cDNA microarray assays. RIOX2 protein expression was assessed using an immunohistochemistry approach on a tissue section array from benign and malignant prostate tissues. Gene expression profiles were analyzed using the bioinformatics software R package. Western blot assay examined androgen stimulation on RIOX2 protein expression in LNCaP cells. Results: Among 35 Jumanji domain-containing protein genes, 12 genes were significantly upregulated in prostate cancers compared to benign compartments. COX regression analysis identified that the ribosomal oxygenase 2 (RIOX2) gene was the only one significantly associated with disease-specific survival outcomes in prostate cancer patients. RIOX2 upregulation was confirmed at the protein levels using immunohistochemical assays on prostate cancer tissue sections. Meanwhile, RIOX2 upregulation was associated with clinicopathological features, including late-stage diseases, adverse Gleason scores, TP53 gene mutation, and disease-free status. In castration-resistant prostate cancers (CRPC), RIOX2 expression was positively correlated with AR signaling index but negatively correlated with the neuroendocrinal progression index. However, androgen treatment had no significant stimulatory effect on RIOX2 expression, indicating a parallel but not a causative effect of androgen signaling on RIOX2 gene expression. Further analysis discovered that RIOX2 expression was tightly correlated with its promoter hypomethylation and MYC gene expression, consistent with the notion that RIOX2 was a c-Myc target gene. Conclusion: The Jumanji domain-containing protein RIOX2 was significantly overexpressed in prostate cancer, possibly due to c-Myc upregulation. RIOX2 upregulation was identified as an independent prognostic factor for disease-specific survival.

Scaffold protein MAPK8IP2 expression is a robust prognostic factor in prostate cancer associated with AR signaling activity.

Mitogen-activated protein kinase-8-interacting protein 2 (MAPK8IP2) is a scaffold protein that modulates MAPK signal cascades. Although MAPK pathways were heavily implicated in prostate cancer progression, the regulation of MAPK8IP2 expression in prostate cancer is not yet reported. We assessed MAPK8IP2 gene expression in prostate cancer related to disease progression and patient survival outcomes. MAPK8IP2 expression was analyzed using multiple genome-wide gene expression datasets derived from The Cancer Genome Atlas (TCGA) RNA-sequence project and complementary DNA (cDNA) microarrays. Multivariable Cox regressions and log-rank tests were used to analyze the overall survival outcome and progression-free interval. MAPK8IP2 protein expression was evaluated using the immunohistochemistry approach. The quantitative PCR and Western blot methods analyzed androgen-stimulated MAPK8IP2 expression in LNCaP cells. In primary prostate cancer tissues, MAPK8IP2 mRNA expression levels were significantly higher than those in the case-matched benign prostatic tissues. Increased MAPK8IP2 expression was strongly correlated with late tumor stages, lymph node invasion, residual tumors after surgery, higher Gleason scores, and preoperational serum prostate-specific antigen (PSA) levels. MAPK8IP2 upregulation was significantly associated with worse overall survival outcomes and progression-free intervals. In castration-resistant prostate cancers, MAPK8IP2 expression strongly correlated with androgen receptor (AR) signaling activity. In cell culture-based experiments, MAPK8IP2 expression was stimulated by androgens in AR-positive prostate cancer cells. However, MAPK8IP2 expression was blocked by AR antagonists only in androgen-sensitive LNCaP but not castration-resistant C4-2B and 22RV1 cells. These results indicate that MAPK8IP2 is a robust prognostic factor and therapeutic biomarker for prostate cancer. The potential role of MAPK8IP2 in the castration-resistant progression is under further investigation.