PC Progression Research Articles

Serum CXCL5 level is associated with tumor progression in penile cancer

Chemokine (C-X-C motif) ligand 5 is an important regulator of tumor progression in many cancers, and could serve as potential serum cancer biomarker. Our initial analysis identified CXCL5 as a cancer-related gene highly expressed in PC. Patients with PC exhibited markedly higher preoperative serum CXCL5 levels compared with that in healthy individuals (P<0.001). The area under the curve (AUC) was 0.880 with the sensitivity of 84.0%, and specificity of 80.4% to distinguish PC. Serum CXCL5 levels were also significantly decreased following tumor resection in patients with PC (P=0.001). Preoperative serum CXCL5 level was significantly associated with clinicopathological characteristics including T stage (P=0.001), nodal status (P<0.001), and pelvic lymph node metastasis (P=0.018). Cox regression analysis showed that serum CXCL5 level could serve as an independent prognostic factor for disease-free survival with a HR of 6.363 (95% CI: 2.185–18.531, P=0.001). CXCL5 and its receptor CXCR2 exhibited correlated expression pattern in PC tissues. Differential CXCL5 expression was observed in normal penile tissues, PC cell lines, and their culture supernatants. Furthermore, knockdown of CXCL5 or CXCR2 expression markedly suppressed malignant phenotypes (cell proliferation, clonogenesis, apoptosis escape, migration, and invasion), attenuated STAT3 and AKT signaling, and reduced MMP2/9 secretion in PC cell lines. In conclusion, our findings revealed that serum CXCL5 level might serve as a potential diagnostic and prognostic cancer biomarker for penile cancer. Autocrine CXCL5/CXCR2 signaling might activate multiple downstream oncogenic signaling pathways (STAT3, AKT, MMP2/9) to promote malignant progression of PC, which may warrant further investigation in the future.

HSDL2 Acts as a Promoter in Pancreatic Cancer by Regulating Cell Proliferation and Lipid Metabolism.

BackgroundPancreatic cancer (PC) is a leading cause of cancer mortality worldwide. Hydroxysteroid dehydrogenase like protein 2 (HSDL2) is overexpressed in a variety of malignant tumors and is might be closely related to the development of cancer. It also regulates different metabolism and signaling pathways.PurposeThe purpose of this research was to find HSDL2 expression levels and investigate its underlying molecular mechanism in PC.Patients and MethodsIn the present study, a total of 66 PC samples and 54 normal tissues were used to examine the expression of HSDL2. In order to gain a broader insight into the molecular mechanism of HSDL2 in PC, the HSDL2 siRNA sequences were transfected into PC cell lines (Bxpc-3 and Panc-1), respectively. Cell proliferation was measured by MTT, colony formation assay and EdU assays. Furthermore, the lipid metabolism process was evaluated by triglyceride and phospholipid assay kits, BODIPY 493/503 staining and the expression of several pivotal lipid metabolic enzymes in PC.ResultsIn this study, HSDL2 was highly expressed in PC and connected with shorter overall survival. When HSDL2 was silenced, the cell proliferation was significantly reduced, and the lipid metabolism was further inhibited.ConclusionHigh expression of HSDL2 plays an important role in the progression of PC and might be a potential new biomarker of poor prognosis as well as a therapeutic target in the future.

The RNA methyltransferase NSUN6 suppresses pancreatic cancer development by regulating cell proliferation.

BackgroundPancreatic cancer (PC) is one of the most lethal solid malignancies in the world due to its excessive cell proliferation and aggressive metastatic features. Emerging evidences revealed the importance of posttranscriptional modifications of RNAs in PC progression. However, knowledge about the 5-methylcytosine (m5C) RNA modification in PC is still extremely limited. In this study, we attempted to explore the expression changes and clinical significances of 12 known m5C-related genes among PC patients.MethodsA total of 362 normal and 382 tumor specimens from PC patients were examined for candidate m5C-related gene and protein expression by using quantitative PCR (qPCR) and immunohistochemistry (IHC). The proliferation rate of PC cells was detected by MTS assay. Xenograft mouse models were used to assess the role of NSUN6 in PC tumor formation.FindingsThrough analyzing the four Gene Expression Omnibus (GEO) databases, six m5C-related genes shown significant and consistent alterations were selected for further examination in our 3 independent PC cohorts. Finally, we identified the reduction of NSUN6 as a common feature of all PC sample sets examined. NSUN6 expression correlated with clinicopathologic parameters including T stage, and Ki67+ cell rate. Further assessing the transcriptional profiles of 50 PC tissues, we found biological processes associated with cell proliferation like cell cycle and G2M checkpoint were enriched in NSUN6 lower expression group. Helped by in vitro PC cell lines and in vivo xenograft mouse models, we confirmed the role of NSUN6 in regulating cell proliferation and PC tumor growth. Last but also importantly, we also show the good performance of NSUN6 in evaluating tumor recurrence and survival among PC patients.InterpretationOur data suggested that NSUN6 is an important factor involved in regulating cell proliferation of PC, and highlights the potential of novel m5C-based clinical modalities as a therapeutic approach in PC patients.FundingThis study was supported by the National Natural Science Foundation of China (Grant Nos. 81803014, 81802424, and 81802911).

Long non-coding RNA NORAD contributes to the proliferation, invasion and EMT progression of prostate cancer via the miR-30a-5p/RAB11A/WNT/\u03b2-catenin pathway

BackgroundProstate cancer (PC) is common male cancer with high mortality worldwide. Emerging evidence demonstrated that long noncoding RNAs (lncRNAs) play critical roles in various type of cancers including PC by serving as competing endogenous RNAs (ceRNAs) to modulate microRNAs (miRNAs). LncRNA activated by DNA damage (NORAD) was found to be upregulated in PC cells, while the detailed function and regulatory mechanism of NORAD in PC progression remains largely unclear.MethodsExpression of NORAD in PC tissues and cell lines were detected by real-time quantitative PCR (qRT-PCR). NORAD was respectively overexpressed and knocked down by transfection with pcDNA-NORAD and NORAD siRNA into PC-3 and LNCap cells. Cell proliferation, invasion and apoptosis were determined by using CCK-8, Transwell and Flow cytometry assays, respectively. The target correlations between miR-30-5p and NORAD or RAB11A were confirmed by using dual luciferase reporter assay. Moreover, expression levels of RAB11A, the epithelial-mesenchymal transition (EMT) marker proteins and the Wnt pathway related proteins were measured by Western blotting. Tumor xenograft assay was used to study the effect of NORAD on tumor growth in vivo.ResultsNORAD was upregulated in PC tissues and cells. Overexpression of NORAD promoted cell proliferation, invasion, EMT, and inhibited cell apoptosis; while knockdown of NORAD had the opposite effect. NORAD was found to be functioned as a ceRNA to bind and downregulated miR-30a-5p that was downregulated in PC tumor tissues. Rescue experiments revealed that miR-30a-5p could weaken the NORAD-mediated promoting effects on cell proliferation, invasion and EMT. Furthermore, RAB11A that belongs to a member of RAS oncogene family was verified as a target of miR-30a-5p, and reintroduction of RAB11A attenuated the effects of miR-30a-5p overexpression on cell proliferation, invasion, EMT and apoptosis of PC cells. More importantly, silencing RAB11A partially reversed the promoting effects of NORAD overexpression on cell proliferation, invasion and EMT of PC cells via the WNT/β-catenin pathway. Lastly, tumorigenicity assay in vivo demonstrated that NORAD increased tumor volume and weight via miR-30a-5p /RAB11A pathway.ConclusionOur results indicated a significant role of NORAD in mechanisms associated with PC progression. NORAD promoted cell proliferation, invasion and EMT via the miR-30a-5p/RAB11A/WNT/β-catenin pathway, thus inducing PC tumor growth.

CircDDX42 Accelerates the Development of Pancreatic Cancer via miR-613/ID4/PI3K/AKT Axis.

BackgroundPancreatic cancer (PC) is one of the fatal cancers globally. CircDEAD-box helicase 42 (circDDX42) has been reported to play an oncogenic role in many cancers. The purpose of our study was to explore the relationship between circDDX42 and PC development and the potential mechanism by which circDDX42 modulating the progression of PC.MethodsThe enrichment of circDDX42, miR-613 and inhibitor of DNA binding 4 (ID4) was determined by quantitative real-time polymerase chain reaction (qRT-PCR) in PC tissues and cells. The proliferation, apoptosis and metastasis of PC cells were examined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Western blot, flow cytometry and transwell migration and invasion assays, respectively. The binding sites between miR-613 and circDDX42 or ID4 were predicted by Starbase bioinformatic software, and dual-luciferase reporter assay was conducted to verify the combination between miR-613 and circDDX42 or ID4. Western blot was carried out to detect the abundance of ID4, p-phosphatidylinositol 3-kinase (p-PI3K), PI3K, p-AKT serine/threonine kinase (p-AKT) and AKT in PC cells. The in vivo role of circDDX42 was verified through using murine xenograft model.ResultsThe level of circDDX42 was enhanced in PC tissues and cells compared with that in matching normal tissues and HPDE cells. CircDDX42 promoted the proliferation and metastasis and suppressed the apoptosis of PC cells. CircDDX42 could sponge miR-613, and miR-613 was negatively regulated by circDDX42 in PC cells. MiR-613 suppressed the progression of PC. ID4 was a direct target of miR-613. ID4 was inversely modulated by miR-613 and positively regulated by circDDX42 in PC cells. ID4 played an oncogenic role in the tumorigenesis of PC. CircDDX42/miR-613/ID4 axis regulated the activation of PI3K/AKT pathway in PC cells. ID4 facilitated the progression of PC via activating PI3K/AKT signal pathway. CircDDX42 promoted the tumor growth of PC in vivo.ConclusionCircDDX42 accelerated the proliferation and metastasis while impeded the apoptosis of PC cells via circDDX42/miR-613/ID4/PI3K/AKT axis. This axis might be a promising target for PC therapy.

Abstract 1621: Contactin 1 (CNTN1) promotes prostate cancer tumorigenesis in transgenic models

Abstract Background: Prostate cancer (PCa) is one of the most common male-specific malignancies in developed country. The progression of PCa involves complex mechanisms. Contactin 1 (CNTN1) is a neural cell adhesion protein reported to promote lung cancer metastasis and associate with poor prognosis in several cancers. In view of our recent publication reporting CNTN1 as a novel factor promoting PCa metastasis, we have constructed prostate-specific CNTN1 overexpression (CNTN1TG/TG) and PTEN−/− mice to explore CNTN1's role in PC tumorigenesis and progression. Materials and Methods: CNTN1 transgenic C57BL/6 mice was constructed by insertion of a transgene unit into the intron of Rosa26 locus. The CNTN1 transgene (TG) cassette consists of a CAG promoter, a STOP unit flanked by two LoxP sites, followed by CNTN1, which will only be expressed upon removal of the STOP unit by Cre recombinase. The homozygous CNTN1STOP-TG/STOP-TG [without transgene (TG) expression] mice were crossed with prostate-specific Cre mice (PB-Cre4). Prostate expression of CNTN1 was clearly demonstrated only in CNTN1TG/− and CNTN1TG/TG mice but not in CNTN1STOP-TG/STOP-TG animals. In a similar system, PTENflox/flox were crossed with PB-Cre4 to generate prostate-specific PTEN−/- mice. At 21-weeks old when invasive tumors were formed, PTEN−/− mice were surgically castrated and maintained for another 12 weeks to construct the castration-resistant prostate cancer (CRPC) model. Results: At 12-weeks old both CNTN1TG/- and CNTN1TG/TG mice developed hyperplasia in the prostate, evident by an increase in cellularity, nuclear crowding, and mild dysplasia. This supports CNTN1's role in PCa tumorigenesis as prostate epithelial hyperplasia in transgenic mice is a well-regarded precursor of PCa. In line of this observation, immunohistochemistry showed upregulation of proliferation marker Ki67 and AKT phosphorylation at serine 473 (indicative of AKT activation) in the prostate of CNTN1TG/TG compared to control CNTN1STOP-TG/STOP-TG (without Cre) mice. In androgen-sensitive LNCaP cells stably expressing CNTN1, an upregulation of prostate-specific antigen (PSA), a well-regarded AR target, was observed both with and without androgen deprivation. This suggest that CNTN1 likely initiates PCa and contributes to its progression through networks involving AR signaling. The oncogenic role of CNTN1 in association with PTEN deficiency was explored using prostate-specific PTEN−/− mice which recapitulate most clinical features of PCa, including the development of castration-resistant PCa (CRPC). The expression of CNTN1 was undetectable in age-matched C57BL/6 mice but was evident in tumors produced in intact PTEN−/− mice and was further upregulated in CRPC tumors generated in castrated PTEN−/- mice. These observations are consistent with CNTN1-induced AKT activation observed in PCa cells and in CNTN1TG/TG animals. Conclusions: Collectively, these results suggest a role of CNTN1 in promoting PCa tumorigenesis and progression. Evidently, our CNTN1 transgenic mice developed a precancerous lesion of PCa. The oncogenic potential of CNTN1 is further supported by its stimulation of LNCaP cell proliferation. This research provides functional evidence for CNTN1 as an important PCa promoting factor and established a novel transgenic mouse model for PCa; this model may greatly facilitate future PCa research and drug development. Citation Format: Yan Jennifer Gu, Wenjuan Mei, Geoffrey A. Wood, Damu Tang. Contactin 1 (CNTN1) promotes prostate cancer tumorigenesis in transgenic models [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1621.

TUG1 knockdown inhibits the tumorigenesis and progression of prostate cancer by regulating microRNA-496/Wnt/β-catenin pathway.

Our study aimed to further investigate the roles and molecular mechanisms of lncRNA taurine upregulated gene 1 (TUG1) in the development and progression of PC. RT-qPCR assay was carried out to measure expression of TUG1, miR-496, together with β-catenin, cyclin D1 and c-myc. Protein levels of β-catenin, cyclin D1 and c-myc were detected by western blot assay. Cell proliferative ability was assessed by colony formation assay and CCK-8 assay. Cell migratory and invasive capacities were evaluated by Transwell migration and invasion assay. The interaction between miR-496 and TUG1 was explored by bioinformatics analysis, luciferase reporter assay and RNA immunoprecipitation assay. Mouse xenograft experiments were performed to further investigate the roles and molecular basis of TUG1 in the tumorigenesis of PC in vivo. TUG1 was highly expressed in PC tissues and cells (DU145 and PC3). TUG1 knockdown inhibited proliferation, migration and invasion in DU145 and PC3 cells. Moreover, TUG1 suppressed miR-496 expression by direct interaction. TUG1 overexpression abrogated miR-96-mediated antiproliferation, anti-migration and anti-invasion effects in DU145 and PC3 cells. TUG1 knockdown inactivated Wnt/β-catenin signaling pathway by upregulating miR-496 in DU145 and PC3 cells. Additionally, TUG1 knockdown inhibited DU145 cells derived PC xenograft growth by upregulating miR-496 and inactivating Wnt/β-catenin signaling in vivo. TUG1 knockdown suppressed PC cell proliferation, migration and invasion in vitro and curbed PC xenograft growth in vivo by regulating miR-496/Wnt/ β-catenin signaling pathway, deepening our understanding on etiology of PC.

WALNUTS for POWER: A Protocol for the Polyphenols, Omega-3 Fatty Acids, Weight Loss, and EneRgy Randomized Controlled Trial

Abstract Objectives Prostate cancer (PC) is the most common non-skin cancer in men and the second leading cause of cancer death. Omega-3 fatty acids and polyphenols have bioactive properties that may alter tumor biology and reduce PC progression. Walnuts (Juglans regia L.) are one of the best sources of plant-based omega-3 s (2.5 g per oz) and polyphenols (total polyphenols 462 mg per oz). We propose to determine the effect of walnuts added to a typical diet on markers of PC progression in men scheduled for radical prostatectomy (RP). The primary objective of the study is to determine the effect of walnut intake on Ki67 expression in prostatic tissue. Secondary objectives are to determine the effect of walnuts on prostatic tissue oxidative stress and inflammation. Exploratory objectives include measuring the effect of walnuts on the gut microbiome, insulin/IGF signaling and gene expression. Methods 50 men with PC scheduled for RP are recruited from the Durham Veterans Medical Center and Cedars-Sinai Medical Center. Men are randomized to receive 2 oz. of walnuts added to a usual diet (intervention) or control (usual diet) using a 1:1 allocation during the 3–10-week pre-RP period. An experienced dietitian provides counseling and recipes to men in the intervention arm and monitor dietary compliance. RP tissue is stained and scored for Ki67 by an experienced pathologist. Tumors are processed for RNA extraction and RNA-sequenced at the UCLA genomics core. Fasting blood are collected at baseline and pre-RP visits, spun, separated into serum, plasma, and red blood cells and stored at −80°C. Serum are batch-analyzed for insulin, IGF-1, IGFBP-3, and IL-6. Stool samples for microbiota analyses are collected at baseline and pre-RP visits and undergo 16 s rRNA gene sequencing and statistical analyses to identify microbiome community content. Between-arm differences during the study are assessed by t-test for continuous variables and chi-square test for categorical variables. The study will be completed in 3 years. To date, 7 men have been recruited. Results N/A Conclusions If shown to be effective in improving PC outcomes in men diagnosed with early stage PC, this modest dietary change could provide a much needed feasible, low-cost, easily accessible, non-toxic option that could be sustained over the long-term. Funding Sources California Walnut Commission.

The role of response as predictor of improved outcome in advanced pancreatic cancer (APC) patients (pts) treated with first-line Gemcitabine plus Nab-paclitaxel (GemNab).

e16758 Background: GemNab is one of the first-line standard treatment (tx) of APC. To date, no predictive factors, both clinical and molecular, of benefit from this regimen exist. Two retrospective studies showed that early tumor shrinkage (ETS) can predict an improved outcome in APC pts receiving a first-line tx with FOLFIRINOX or GemNab. However, data regarding GemNab, limited to a small population of only 57 pts, seem to not confirm the association of ETS with a better outcome. Hence, we retrospectively analysed an homogeneous population of APC treated with first-line GemNab at our Institution, investigating the impact of several clinical factors, including response and ETS. Methods: APC pts receiving a first-line tx with GemNab were included in the analysis. The association of RECIST response and ETS with PFS and OS was evaluated. The following variables were collected: gender; age ( &gt; vs ≤ 55 years and ≥ vs &lt; 70 years ); baseline ECOG PS; Ca 19.9 baseline level (≥ vs &lt; 200); anamnesis of diabetes; site of primary tumor (head/uncinate process vs body/tail); locally A vs metastatic (m) PC; synchronous vs metachronous; number of m sites (1 vs &gt; 1); m sites (liver, peritoneum, lung, nodes); number of tx lines (1 vs &gt; 1). Univariate and multivariate analyses for PFS and OS were performed. Results: A total of 184 APC pts receiving first-line GemNab at our Institution from February 2014 to May 2019 were included in the analysis. RR and ETS were assessed in 174 and 168 pts, respectively. RR was 30%, disease control rate (DCR) 63% and ETS was 24%. Responders had a significant better PFS (12.5 vs 5.7 months, p &lt; 0.0001) and OS (25.1 vs 12.1 months, p &lt; 0.0001). ETS was significantly associated with improved PFS (12.3 vs 6.2 months, p &lt; 0.0001) and OS (24.0 vs 12.8 months, p &lt; 0.0001). At the multivariate analysis a significant association with survival parameters was confirmed for RECIST response, but not for ETS. At the multivariate analysis, also metachronous disease and number of tx lines &gt; 1 were independently associated with better OS. Conclusions: Despite its retrospective nature, this is one of the largest series of APC pts treated with first-line GemNab investigating the role of RECIST response and ETS in predicting outcome. On the basis of our results, RECIST response may be considered a positive prognostic factor, whereas ETS does not. In conclusion, achieving tumor shrinkage, not necessarily early, significantly delays PC progression and prolongs survival in pts treated with first-line GemNab.

Non-metastatic prostate cancer: rationale for conservative treatment and impact on disease-related morbidity and mortality in the elderly

Purpose: To determine the rationale for not offering local treatment to prostate cancer patients with non-metastatic disease at diagnosis who later died of prostate cancer and to document local and systemic complications caused by disease progression.Material and Methods: In this population-based, retrospective study we reviewed the medical records of all patients who died of prostate cancer in 2009–2014 in Vestfold County (Vestfold Mortality Study), who were non-metastatic at diagnosis and who had received no local treatment to the prostate (n = 117).Results: A review of patient records demonstrated that the chronological age of 75 years or older was the main rationale for not offering local treatment to the prostate (37%, n = 43). No consideration was given to the functional status and patient health. These elderly patients stood for almost one-fifth of the total PC mortality in Vestfold County. In addition to dying from PC, 86% of patients developed local complications attributable to PC progression. Observation of strict limits for local treatment with regard to tumor characteristics contributed further to the underuse of local treatment.Conclusions: Our study demonstrated systematic undertreatment of elderly patients with aggressive, non-metastatic PC with regard to local treatment based on chronological age alone. The patients in this study died of prostate cancer and the majority experienced significant morbidity caused by local tumor growth.

Interferon gamma inhibits CXCL8–CXCR2 axis mediated tumor-associated macrophages tumor trafficking and enhances anti-PD1 efficacy in pancreatic cancer

BackgroundPancreatic cancer (PC) is a common malignancy of the digestive system and is characterized by poor prognosis and early metastasis. Tumor immune escape plays an important role in PC progression....

MLPH Accelerates the Epithelial-Mesenchymal Transition in Prostate Cancer.

IntroductionProstate cancer (PC) is the second greatest cause of cancer deaths globally. PC presents a poor prognosis once it metastasizes. There is considerable proof of vital epithelial–mesenchymal transition (EMT) functionality in PC metastasis. Previous studies revealed that melanophilin (MLPH) is associated with PC; however, its role in PC remains poorly understood.MethodsBioinformatics analyses were performed. The cellular responses to MLPH knockdown were examined in HCC cell lines via wound healing assay, migration and invasion assay, Western blotting.ResultsAnalysis of the PROGgeneV2 database revealed that high MLPH expression might indicate poor overall survival. MLPH knockdown reduced PC cell migration, proliferation, and invasion. MLPH downregulation in vivo resulted in a lower growth rate and fewer metastatic nodules in lung tissues. Furthermore, MLPH knockdown recovered downregulated expression of the mesenchymal marker N-cadherin and the epithelial marker E-cadherin following a decrease in β-catenin.ConclusionThese results indicate that progression of PC is stimulated via MLPH-dependent initiation of the EMT.

MiR-129-5p suppresses proliferation, migration, and induces apoptosis in pancreatic cancer cells by targeting PBX3.

Pancreatic cancer (PC) is the seventh most frequent cause of cancer-related deaths worldwide with a high mortality. MicroRNAs (miRNAs) act as important regulators for the development of PC and participate in the progression of PC. miR-129-5p was reported to regulate the progression of tumors, such as thyroid cancer and gastric cancer. However, the function of miR-129-5p in PC is still unclear. In this study, the down-regulation of miR-129-5p was detected in PC tissues and PC cells. miR-129-5p was overexpressed or knocked down in AsPC-1 and BxPC-3 cells. The results showed that miR-129-5p overexpression suppressed proliferation, migration and invasion, and induced apoptosis of PC cells, whereas miR-129-5p knockdown showed opposite effects. In addition, we found that pre-B-cell leukemia homeobox 3 (PBX3) overexpression promoted proliferation, migration and invasion, but reduced apoptosis of PC cells. PBX3 was identified as a target of miR-129-5p by informatics analysis and dual luciferase reporter assay. Finally, our results indicated that miR-129-5p suppressed cell proliferation and migration by targeting PBX3. This study demonstrated that miR-129-5p could function as a tumor suppressor in the progression and development of PC by targeting PBX3, providing a reliable prognostic factor and a new therapeutic strategy for PC.

Cross-talk among AFAP1-AS1, ACVR1 and microRNA-384 regulates the stemness of pancreatic cancer cells and tumorigenicity in nude mice

BackgroundPancreatic cancer (PC) represents one of the most aggressive forms of cancer. The role of long non-coding RNAs (lncRNAs) has been highlighted in various malignancies including PC. The aim of the present study was to investigate the effects associated with actin filament-associated protein 1 antisense RNA 1 (AFAP1-AS1) on the progression of PC and the underlying mechanism.MethodsMicroarray-based gene expression profiling of PC was performed to identify PC-related lncRNAs, after which the expression of AFAP1-AS1 and cancer stem cell (CSC) markers in PC tissues and cells were determined accordingly. The potential microRNA-384 (miR-384) capable of binding to AFAP1-AS1, in addition to its ability to regulate activin receptor A type I (ACVR1) were analyzed. In order to investigate the effect of the AFAP1-AS1/miR-384/ACVR1 axis on self-renewal ability, tumorigenicity, invasion, migration and stemness of PC cells, shRNA-AFAP1-AS1, miR-384 mimic and inhibitor were cloned into cells.ResultsHigh expression of AFAP1-AS1 and ACVR1 with low expression of miR-384 were detected in PC tissues. ACVR1 was determined to be down-regulated when miR-384 was overexpressed, while the inhibition of AFAP1-AS1 decreased its ability to binding competitively to miR-384, resulting in the down-regulation of ACVR1 and enhancing miR-384 expression, ultimately inhibiting the progression of PC. The knockdown of AFAP1-AS1 or overexpression of miR-384 was confirmed to impair PC cell self-renewal ability, tumorigenicity, invasion, migration and stemness.ConclusionsTaken together, AFAP1-AS1 functions as an endogenous RNA by competitively binding to miR-384 to regulate ACVR1, thus conferring inhibitory effects on PC cell stemness and tumorigenicity.

Natural course of pineal cysts\u2014a radiographic study

BackgroundPineal cysts (PCs) are a benign lesion of the pineal gland that have been known to the medical community for a long time. With a prevalence rate of approximately 1% in the general population, PC is often a reason for medical counseling. The natural course of PC morphology has not been well described. In this study, we present a longitudinal magnetic resonance imaging (MRI) study of patients with PCs, with special focus on those who showed an increase or decrease in PC size.MethodsWe enrolled all patients with a PC who were referred to our department between January 2000 and January 2018. Each patient underwent a clinical examination, and the patient’s age, sex, and presenting signs and symptoms were noted. MRI was performed during periodic examinations, and a clinical and radiological course was reassessed.ResultsIn total, 133 patients (99 women, 34 men) were enrolled. The mean maximum diameter was 12.7 ± 5.2 mm (range 7–35 mm). PCs increased in size during the follow-up in seven patients (5.3%) and decreased in size in 10 (7.5%). The remaining cysts (n = 116, 87.2%) were stable over the follow-up period. Analyzing patients according to cyst size change, we found a significant difference in the mean age between the PC progression group and PC regression group (p = 0.01). The mean size of the PCs at the time of diagnosis did not differ significantly between the two groups (p = 0.81). We diagnosed two cases of pineal apoplexy.ConclusionWe found that PCs are a dynamic structure that may change in size during the patient’s lifetime. Patients with an increase in PC size were significantly younger than patients with a decrease in size. Therefore, PC growth in the first, second, and third decennium is normal and does not justify medical intervention. Surgery is indicated in cases of hydrocephalus and Parinaud’s syndrome or in atypical cysts when neoplasia is suspected. The size of a PC does not predict PC behavior in terms of a future increase or decrease in size.

Abstract A050: Integrated system-level analyses define an androgen receptor variant oncogenic network for advanced prostate cancer

Abstract Unbiased high-throughput approaches provide a global view of the massive genetic and transcriptomic changes that occur in and potentially drive cancer. Such approaches permit the discovery of novel networks that can serve as prognostic markers and therapeutic targets. We performed a systems-biology approach to identify genes linked to the androgen receptor (AR) variant AR-V7 in castration-resistant prostate cancer (CRPC). AR variants are C-terminal truncated, constitutively active, and play key transcriptional regulatory roles resulting in treatment resistance and disease progression. These AR variants lack the C-terminal ligand binding domain, which is the target of all approved drugs against AR, and consequently there is considerable interest in novel therapeutic strategies that are effective against AR variant-expressing prostate tumors. Directly targeting AR variants is challenging, and thus an attractive alternative approach is to identify downstream AR-V7 hub genes that not only drive cancer progression but also feedback to enhance AR-V7 activity. Thus, our goals were to: 1) define clinically relevant gene sets or modules in human PC progression using unbiased gene expression meta-analysis of PC patient samples by Weighted Gene-Co-expression Network Analysis (WGCNA); 2) map the AR-V7 functional interactome using a unique high-throughput synthetic genetic array (SGA) screening in the yeast S. pombe; and 3) identify AR-V7 regulated genes in PC cells. Integration of these approaches resulted in the identification of seven genes that are upregulated in human PC (as determined by WGCNA), functionally interact with AR-V7 (as determined by SGA), and are regulated by AR-V7. This gene set encoded select proteins previously linked to PC, such as members of the cyclin and topoisomerase families, but also proteins with no prior association with AR-V7 or PC. The seven genes comprised a signature that correlated with patient Gleason score and predicted disease free-survival and overall survival in large independent PC patient cohorts. Individual depletion of the expression of these genes decreased ligand-independent AR transcriptional activity concomitant with reduced CRPC cell proliferation. Because CRPC cells were highly dependent on these inter-related genes, combination therapy targeting two of the seven genes led to potent inhibition of CRPC cell proliferation. In contrast, the combination of these agents had little to no effect on nontumorigenic prostate epithelial cells or AR-null PC cells. In summary, we utilized an unbiased and novel gene discovery strategy that combined sophisticated bioinformatics and experimental approaches. This multifaceted approach resulted in the identification of disease-relevant genes that were regulated by AR-V7 and that reciprocally enhanced AR-V7 oncogenic activity. This novel gene set has a prognostic value and provides a rationale for combinatorial CRPC therapy. Citation Format: Fiorella Magani, Eric Bray, Maria Julia Martinez, Valeria Copello, Ning Zhao, Stephanie Peacock, David Wiley, Gennaro D’urso, Kerry L. Burnstein. Integrated system-level analyses define an androgen receptor variant oncogenic network for advanced prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A050.

PO-150 Modulation of the expression of casein kinase i isoform alpha in the initiation and progression of prostate cancer

IntroductionProstate cancer (PC) is the first cause of fatality related to the reproductive system in men. One out of seven men encounter this pathology once in their lifetime. Prostate Specific Antigen along with digital rectal exam are the first line of screening for prostate pathologies but numerous studies showed that these techniques lack specificity and in sometimes validity for screening. From this notion comes the need to find new biomarkers and molecular players that help get an early detection and effective treatment of PC. Casein Kinase 1 Alpha (CSNK1A1) is an enzyme involved in multiple cellular processes such as the regulation of the oncogenic Wnt/beta-catenin signalling pathway. Its implication in carcinogenesis and cancer progression is still unclear. In this study, we aim to establish a CSNK1A1 expression profile in BPH and in differently advanced PC grades and to investigate the localization of the enzyme compared to beta catenin in the different samples.Material and methodsFormalin-fixed paraffin-embedded human prostatic tissues were collected as follows: 85 BPH, 64 PC and 3 controls. Gene expression was assessed by quantitative RT-PCR of cellular transcripts. Protein localization was monitored by tissue immunostaining for CSNK1A1 and Beta-Catenin. Further in-vitro validation of the results is to be made on normal prostate epithelial cell line and PC cell lines using RT-PCR and western blotting.Results and discussionsWe were able to identify a continuous increase in the CSNK1A1 expression between controls, BPH and PC. These results are being validated in vitro. Preliminary immunostaining results showed membranous beta catenin in BPH and that seemed to be more heterogeneous in PC sections. A correlation with CSNK1A1 localization and amount is still to be achieved. Our results suggest a role of CSNK1A1 in the pathogenesis of BPH and PC by working on the proliferation induction axis, and malignant behaviour of cells. Our staining results suggest a minor role of beta catenin and Wnt pathway in these axes from which emerges the need to deeply investigate the pathways in which CSNK1A1 is implicated in BPH and PC initiation and progression.ConclusionOur results suggest an increase in CSNK1A1 in BPH and PC, which flags this protein as a potential marker in the progression of PC. More investigation is needed to verify whether this protein is involved in cancer initiation or in the direct inhibition of the oncogenic Wnt/beta-catenin signalling pathway and other pathways.

PO-316 Genome-wide gene expression analysis of a murine model of prostate cancer cell progression: towards identification of high-potential therapeutic targets

IntroductionProstate cancer (PC) is the most frequently diagnosed cancer among men and the second leading cause of male cancer-related deaths worldwide. The poor prognosis of PC is largely due to late diagnosis of the disease when it has progressed to advanced stages marked by androgen-independence. We construe that these direly needed advances are limited by our poor understanding of early events in the progression of PC and that would thus represent ideal targets for early intervention. To begin to fill this void, we interrogated molecular ‘oncophenotypes’ that embody the transition of PC from an androgen-dependent (AD) to –independent (AI) state.Material and methodsWe have previously established AD and AI murine PC cell lines, PLum-AD and PLum-AI, respectively, which recapitulate primary and progressive PC morphologically and genetically. We statistically surveyed global gene expression in these cell lines by microarray analysis. Differential profiles were functionally interrogated by pathways, gene set enrichment and topological gene-gene network analyses – features built in the commercially available software Ingenuity Pathways Analysis.Results and discussionsGene expression analysis of PLum-AD and PLum-AI transcriptomes (n=3 each), revealed 723 differentially expressed genes (392 upregulated and 331 downregulated) in PLum-AI compared with PLum-AD cells. Gene set analysis demonstrated enrichment of biological functions and pathways in PLum-AI cells that are central to tumour aggressiveness including epithelial-to-mesenchymal transition (EMT), cell migration, and cell invasion. Further analysis demonstrated that the p38 mitogen activated protein kinase (MAPK) was predicted to be significantly activated in the PLum-AI cells. Conversely, gene sets that previously showed to be associated with favourable response to the p38 inhibitor SB203580 were attenuated in the PLum-AI cells, suggesting that these aggressive cells may be therapeutically vulnerable to p38 inhibition. Network analysis also alluded to activation of other signalling networks particularly those associated with enhanced EMT, inflammation and immune function/response.ConclusionOur pilot genome-wide gene expression analysis of a murine model of PC progression identified molecular oncophenotypes (e.g. inflammation and EMT) that denote this progression and that comprise high-potential targets (e.g. p38) for early treatment of PC. Efforts are underway to functionally assess the potential roles of these genes as therapeutic targets for PC progression.

Abstract 4062: Novel synthetic peptide targets CD206 receptor promoting the repolarization of prostate cancer (PCa) TAMs and increase the efficacy of conventional prostate cancer therapeutic

Abstract Background: Tumor associated M-ϕs (TAMs) have a complex etiology as a result of soluble factors secreted by prostate tumor cells, creating an immunosuppressive micro-environment (M-E) for the progression of late stage PC that has metastasized to the bone. The result is an M2-like polarization that expresses a number of M2-TAM specific markers, including CD206. Therapeutic targeting of TAMs has proven to be a challenge, due partly to a lack of understanding of the function of this biomarker and the bi-directional effects of TAMs on tumor cells. Our objective is to characterize the CD206 receptor in PC patients, and to test the potential of a novel engineered CD206 targeted peptide (T-P) to reverse the immune-suppressive M-Env in late stage PC. Methods: The TGCA Prostate Dataset was used for patient data and R software was used for informatics analysis. Small-angle X-ray scattering (SAXS) was used to generate the NMR structure of CD206. Peptide binding to the naïve CD206 receptor was determined in silico using PIPER, and was validated by Surface Plasmon Resonance (SPR). In vivo efficacy of CD206 T-P was determined in mice inoculated with PC-3 intratibially. Mice were treated with peptide alone, daily or twice a week at 10, 20, and 50mg/Kg, as a monotherapy or in combination with 20mg/Kg Docetaxel (DTX) over a 21day period. CD86 (M1) and CD206/CD163 (M2-TAM) markers and pro-inflammatory cytokines were determined by FACS. Bone osteolysis was determined by IHC of the entire bone, and bone turnover was determined by IHC analysis of RANKL. Results: TCGA data showed that over-expression of CD206 in Human PC is significantly associated with poor survival. In silico analyses showed the T-P exhibited strong affinity towards CD206 and was validated by SPR (Kd =3.57 µM). In bone-metastatic xenograft models of CRPC (PC-3), the flux differential of relative fluorescence in IVIS imaging showed peptide alone (at 20mg/kg qD sQ) decreased fluorescence by 64.36% compared with the untreated group. FACS analyses of M1/M2 markers CD86/CD206 in mice treated with peptide showed a 33.87 fold decrease in CD206 (M2-TAM) expression and a 7.6 fold increase in CD86 (M1-TAM) expression, compared with only a 4.4 fold decrease in CD206 expression and 0.44 fold decrease in CD86 expression in DTX treated mice. Stem cell marker CD133 showed a 4.7 fold decrease expression compared to the untreated group further suggesting that TAM repolarization reversed the immunosuppressive M-E. X-ray of tumors in bone suggests a reduction of osteolysis and was confirmed by IHC staining and analysis of whole bone showing fibrous and inflammatory remodeling patterns of tumor regression in the peptide only group. Conclusion: These results suggest that the novel engineered CD206 T-P repolarizes M2-TAM phenotype to M1-TAM which reverses the immunosuppressive cancer M-E in late stage PC. Citation Format: Ahmad B. Salam, Benjamin Adu-Addai, Huixian Lin, Anghesom Ghebremedhin, Jason White, Ruksana Amin, Balasubramanyam Karanam, George Martin, Charles Garvin, Henry Lopez, Jesse M. Jaynes, Clayton Yates. Novel synthetic peptide targets CD206 receptor promoting the repolarization of prostate cancer (PCa) TAMs and increase the efficacy of conventional prostate cancer therapeutic [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 4062.

Abstract 5085: CXCR2 signaling in pancreatic cancer induces a tumor supporting inflammatory phenotype of the cancer-associated fibroblasts

Abstract Pancreatic cancer (PC) remains one of the deadliest types of solid malignancies. Making progress in dealing with PC starts by understanding the complex interaction of cellular components in the tumor microenvironment and identifying the crucial checkpoints for tumor progression and metastasis. Desmoplasia and inflammation are two major hallmarks of PC. Cancer-associated fibroblasts (CAFs) are known for inducing fibrosis in several pathologies of the pancreas including PC. Recent reports have described that CAFs can assume the fibrosis-inducing phenotype, when close or have a direct contact with tumor cells, or a tumor-promoting inflammatory phenotype, through distant paracrine signaling. Known for their role in recruiting granulocytes and inducing angiogenesis, CXC chemokines (1-3 and 5-8) contain a conserved motif of Glutamic acid-Leucine-Arginine (ELR) and signal through CXCR1/2 chemokine receptors. In PC, ELR+ chemokines produced by malignant cells and stromal cells contribute to tumor growth, angiogenesis, and tumor-supporting immunosuppression. The deletion of CXCR2 in the genetically engineered PC mouse model (KC) promoted fibrosis and tumor metastasis. Taken together, we hypothesize that ELR+ chemokines signaling through CXCR2 may trigger the inflammatory phenotype of CAFs in PC that promotes tumor growth and progression. Utilizing unidirectional coculture studies, incubating CAFs in conditioned media of PC cells inhibited their growth, decreased the expression of the fibrotic CAFs-associated markers such as αSMA and collagen I, and increased the expression of immunosuppressive cytokines and tumor-promoting chemokines including IL-4, IL-10, IL-13, and CXCL7. Treating CAFs with exogenous ELR+ chemokines (CXCL1 and CXCL8) exhibited a similar phenotype. Furthermore, treating CAFs with exogenous ELR+ chemokines or PC cells conditioned media in the presence of pharmacological inhibitors of CXCR2 rescued this phenotype. In summary, we believe that ELR+ chemokine signaling through CXCR2 induces the inflammatory phenotype of CAFs that promotes immunosuppression, tumor growth, and tumor progression of PC. Citation Format: Mohammad Awaji, Michelle Varney, Abhilasha Purohit, Surinder Batra, Rakesh Singh. CXCR2 signaling in pancreatic cancer induces a tumor supporting inflammatory phenotype of the cancer-associated fibroblasts [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 5085.