Independent Prostate Cancer Cell Lines Research Articles

Investigating a new C2-symmetric testosterone dimer and its dihydrotestosterone analog: Synthesis, antiproliferative activity on prostate cancer cell lines and interaction with CYP3A4

The synthesis of a 17α-linked C2-symmetric testosterone dimer and its dihydrotestosterone analog is reported. The dimers were synthesized using a short five-step reaction sequence with 28% and 38% overall yield for the testosterone and dihydrotestosterone dimer, respectively. The dimerization reaction was achieved by an olefin metathesis reaction with 2nd generation Hoveyda-Grubbs catalyst. The dimers and their corresponding 17α-allyl precursors were tested for the antiproliferative activity on androgen–dependent (LNCaP) and androgen–independent (PC3) prostate cancer cell lines. The effects on cells were compared with that of the antiandrogen cyproterone acetate (CPA). The results showed that the dimers were active on both cell lines, with an increased activity towards androgen–dependent LNCaP cells. However, the testosterone dimer (11) was fivefold more active than the dihydrotestosterone dimer (15), with an IC50 of 11.7 μM vs. 60.9 μM against LNCaP cells, respectively, and more than threefold more active than the reference drug CPA (IC50 of 40.7 μM). Likewise, studies on the interaction of new compounds with drug-metabolizing cytochrome P450 3A4 (CYP3A4) showed that 11 was a fourfold stronger inhibitor than 15 (IC50 of 3 μM and 12 μM, respectively). This suggests that changes in the chemical structure of sterol moieties and the manner of their linkage could largely affect both the antiproliferative activity of androgen dimers and their crossreactivity with CYP3A4.

Periprostatic adipose tissue promotes prostate cancer resistance to docetaxel by paracrine IGF‐1 upregulation of TUBB2B beta‐tubulin isoform

Growing evidence supports the pivotal role played by periprostatic adipose tissue (PPAT) in prostate cancer (PCa) microenvironment. We investigated whether PPAT can affect response to Docetaxel (DCTX) and the mechanisms associated. Conditioned medium was collected from the in vitro differentiated adipocytes isolated from PPAT which was isolated from PCa patients, during radical prostatectomy. Drug efficacy was studied by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide citotoxicity assay. Culture with CM of human PPAT (AdipoCM) promotes DCTX resistance in two different human prostate cancer cell lines (DU145 and PC3) and upregulated the expression of BCL‐xL, BCL‐2, and TUBB2B. AG1024, a well‐known IGF‐1 receptor inhibitor, counteracts the decreased response to DCTX observed in presence of AdipoCM and decreased TUBB2B expression, suggesting that a paracrine secretion of IGF‐1 by PPAT affect DCTX response of PCa cell. Collectively, our study showed that factors secreted by PPAT elicits DCTX resistance through antiapoptotic proteins and TUBB2B upregulation in androgen independent PCa cell lines. These findings reveal the potential of novel therapeutic strategies targeting adipocyte‐released factors and IGF‐1 axis to overcome DCTX resistance in patients with PCa.

Distinct roles for the RNA-binding protein Staufen1 in prostate cancer

BackgroundProstate cancer is one of the most common malignant cancers with the second highest global rate of mortality in men. During the early stages of disease progression, tumour growth is local and androgen-dependent. Despite treatment, a large percentage of patients develop androgen-independent prostate cancer, which often results in metastases, a leading cause of mortality in these patients. Our previous work on the RNA-binding protein Staufen1 demonstrated its novel role in cancer biology, and in particular rhabdomyosarcoma tumorigenesis. To build upon this work, we have focused on the role of Staufen1 in other forms of cancer and describe here the novel and differential roles of Staufen1 in prostate cancer.MethodsUsing a cell-based approach, three independent prostate cancer cell lines with different characteristics were used to evaluate the expression of Staufen1 in human prostate cancer relative to control prostate cells. The functional impact of Staufen1 on several key oncogenic features of prostate cancer cells including proliferation, apoptosis, migration and invasion were systematically investigated.ResultsWe show that Staufen1 levels are increased in all human prostate cancer cells examined in comparison to normal prostate epithelial cells. Furthermore, Staufen1 differentially regulates growth, migration, and invasion in the various prostate cancer cells assessed. In LNCaP prostate cancer cells, Staufen1 regulates cell proliferation through mTOR activation. Conversely, Staufen1 regulates migration and invasion of the highly invasive, bone metastatic-derived, PC3 prostate cells via the activation of focal adhesion kinase.ConclusionsCollectively, these results show that Staufen1 has a direct impact in prostate cancer development and further demonstrate that its functions vary amongst the prostate cancer cell types. Accordingly, Staufen1 represents a novel target for the development of much-needed therapeutic strategies for prostate cancer.

Chemosensitizing effects of methyl jasmonate on paclitaxel-resistant androgen-dependent and independent prostate cancer cell lines

The taxane-based therapy provides survival benefit in patients with metastatic prostate cancer; however, the average survival is less than 20 months due to the partial taxane-related chemoresistance. Innovative strategies are needed to overcome the chemoresistance for improved patient survival. In this project, paclitaxel-resistance was developed on androgen-independent PC3 and androgen-dependent 22Rv1 and LNCaP human prostate cancer (PCa) cell lines to investigate the efficacy of methyl jasmonate (MeJa), an anti-cancer drug, in overcoming the chemoresistance. The PCa cell lines were maintained in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS) at 37?C under 5% CO2. The cell lines were exposed to the gradually increasing doses of paclitaxel. Since the resistance on LNCaP could not be achieved, the study was continued with 22Rv1 cell line. It was demonstrated that paclitaxel-resistant cell lines overexpress ABCB1. Resistance levels of cells and MeJa activity in all resistant and parental lines were measured using CellTiter-Glo? luminescent assay. Test results were compared with Student?s t-test or analysis of variance (ANOVA). P?0.05 (two-tailed) was considered to be significant. In conclusion, MeJa showed more cytotoxicity on paclitaxel resistant PC3 (PC3-PtxR) cells than resistant 22Rv1 (22Rv1-PtxR) cells. Detection of cytotoxic effects of MeJa in overcoming paclitaxel resistance may contribute to the development of alternative new compounds for the prevention or chemosensitization of resistance to chemotherapeutics such as paclitaxel.

Synthesis, molecular modeling, in vivo study and anticancer activity against prostate cancer of (+) (S)-naproxen derivatives

In this study, (S)-naproxen thiosemicarbazides (3a-d), 1,2,4-triazoles (4a-c), triazole-thioether hybride compounds (5a-p) were synthesized and their structures (3a, 3d, 4a and 5a-p) were confirmed by FT-IR, 1H NMR,13C NMR, HR-Mass spectra and elemental analysis. These compounds are designed to inhibit methionine amino peptidase-2 (MetAP2) enzyme in prostate cancer. These compounds (3d, 5a-p) evaluated against androgen-independent prostate adenocarcinoma (PC-3, DU-145) and androgen-dependent prostate adenocarcinoma (LNCaP) cell lines by using MTS method. Compounds 5a, 5b, 5d and 5e showed 14.2, 5.8, 10.8 and 8.4μM anticancer activity against PC-3cell lines, compounds 5e, 5g and 5n presented anticancer activity against DU-145cell lines 18.8, 12.25 and 10.2μM, and compounds 5g, 5m and 5n exhibited anticancer activity against LNCaP cell lines 12.25, 22.76 and 2.21μM, respectively. Consequently, of these results, compounds 5e and 5n showed the highest activities against androgen dependent and independent prostate cancer cell lines, so these compounds could be potent small molecules against prostate cancer. Furthermore, mitogen-activated protein kinase (MAPK) pathway activation, AKT (protein kinase B) phosphorylation and androgen receptor activation of compound 5n (SGK636) were investigated in LNCaP cells by using Western blot method. Compound 5n (SGK636) was also tested against mRNA expression analysis of the Bax, Bcl-2, Caspase 3, Caspase 9 by using real-time PCR analysis. Compound 5n was given to nude male mice with cancer in comparison to the control group. Compound 5n was found to reverse the malignant phenotype in the nude male mice, whereas the prostate cancer progressed in the control group. Analysis of some blood parameters in the study showed that they were within the normal values with respect to the control. The blood values of the animals treated according to the control group also exhibited compliance with the blood limit values. Molecular docking and dynamics simulation of compound 5n binding to Methionine Aminopeptidase 2 (MetAP2) enzyme rationalized its potential activity. In addition, inhibition assay MetAP2 enzyme of compound 5n was evaluated. Taken together, we suggest compound 5n to be a potential candidate for prostate cancer therapy.

Inhibition of PIM kinase with fractionated radiation and docetaxel in preclinical prostate cancer models.

e17534 Background: PIM kinases are highly expressed in high risk prostate cancer and are associated with aggressiveness and poor prognosis, presenting a therapeutic target. Methods: We studied PIM inhibition (PIMi) in both hormone-dependent (LNCaP) and -independent (DU145, 22Rv1) prostate cancer cell lines and mouse xenografts. We evaluated PIMi effects via Western blotting, immunofluorescence, and RNAseq of multiple relevant pathways, and performed radiosensitization analysis with colony formation assays. We also investigated xenograft tumor growth with PIMi +/- radiation (RT) or docetaxel. Results: Pretreating the cell lines with PIMi prior to RT caused marked changes in H2AX phosphorylation compared to no PIMi + RT, with most significant alteration in DU145. However, in colony formation assays, 22Rv1 showed greatest radiosensitization with colony survival reduced from 23% to 15% with PIMi. 22Rv1 xenografts were grown in castrated mice, which were randomized to 30 Gy in 15 treatments vs no RT over 3 weeks, -/+ PIMi on RT days. RT + PIMi eliminated gross tumor, compared to an average of 78% and 51% of initial tumor volumes remaining with PIMi alone and RT alone, respectively. Immunofluorescence and Western blotting of harvested tumors showed PIMi altered patterns in proteins involved with hypoxic response (COX-2), cell survival (MDM2), and androgen receptor and suggested interactions between RT and PIMi for both MDM2 and androgen receptor but not COX-2. RNAseq analysis of xenografts showed daily PIMi affected multiple pathways involved in metabolism, growth, and DNA repair. Finally, we tested PIMi -/+ docetaxel in castrated mice with hormone-sensitive (LNCaP) or -resistant (22Rv1) xenografts treated with PIMi -/+ docetaxel. With both xenografts, PIMi + docetaxel reduced tumor size best, with 22Rv1 xenografts most dramatically showing 74% volume reduction from baseline with PIMi + docetaxel compared to 50% and 25% reduction in volumes with PIMi alone and docetaxel alone, respectively. Conclusions: PIMi has multiple pathway effects in prostate cancer. PIMi significantly reduces disease burden, particularly in combination with RT or docetaxel. Our data provide rationale for testing PIMi in clinical trials for prostate cancer.

Future Oncotargets: Targeting Overexpressed Conserved Protein Targets in Androgen Independent Prostate Cancer Cell Lines.

Targeting evolutionarily conserved proteins in malignant cells and the adapter proteins involved in signalling that generates from such proteins may play a cardinal role in the selection of anti-cancer drugs. Drugs targeting these proteins could be of importance in developing anti-cancer drugs. We inferred that drugs like loperamide and promethazine that act as antagonists of proteins conserved in cancer cells like voltage-gated Calcium channels (Cav), Calmodulin (CaM) and drug efflux (ABCB1) pump may have the potential to be re-purposed as an anti-cancer agent in Prostate Cancer (PCa). Growth and cytotoxic assays were performed by selecting loperamide and promethazine to target Cav, CaM and drug efflux (ABCB1) pumps to elucidate their effects on androgen-independent PC3 and DU145 PCa cell lines. We show that loperamide and promethazine in doses of 80-100μg/ml exert oncocidal effects when tested in DU145 and PC3 cell lines. Diphenhydramine, which shares its targets with promethazine, except the CaM, failed to exhibit oncocidal effects. Anti-cancer effects can be of significance if structural analogues of loperamide and promethazine that specifically target Cav, CaM and ABCB1 drug efflux pumps can be synthesized, or these two drugs could be re-purposed after human trials in PCa.

Digoxin lowers the incidence of prostate cancer: A nationwide population-based study.

In vitro studies have confirmed that cardiac glycosides can induce apoptosis in both hormone-dependent and -independent prostate cancer (PCa) cell lines. The aim of this study was to investigate the incidence of PCa among patients treated with and without digoxin using a nationwide population-based database in Taiwan. We retrieved data of men aged 30 years or older who were newly diagnosed with heart failure between January 1998 and December 2003 from the National Health Insurance program database in Taiwan. We divided the patients into digoxin users and non-digoxin users. Kaplan-Meier curves and Cox proportional hazard analysis were used to examine the risk of subsequent PCa between the digoxin and non-digoxin groups. The mean ± SD follow-up (years) periods in the digoxin and non-digoxin groups were 8.6 ± 1.78 and 8.3 ± 1.75, respectively. The cumulative incidence of PCa during the follow-up period was 3.5% (147/4233) in the non-digoxin group compared with 3.0% (65/2154) in the digoxin group. The log-rank test revealed that the digoxin group had a similar incidence of PCa to the non-digoxin group (p = 0.18). After adjusting for age, benign prostatic hyperplasia, and comorbidities, Cox proportional hazard regression analysis showed that digoxin was associated with a significantly decreased risk of developing PCa (hazard ratio, 0.74; 95% CI, 0.548-0.993; p = 0.045). Moreover, logistic regression analysis showed that the risk of PCa decreased with a longer duration of digoxin use during the study period compared to those who had never used digoxin (p = 0.043). The cardiac glycoside digoxin had significant effects on reducing the incidence of PCa in a time-dependent manner. Our findings may imply the potential application of cardiac glycosides in the prevention and management of PCa.

Flavonoid Calycopterin Induces Apoptosis in Human Prostate Cancer Cells In-vitro.

Prostate cancer is enumerated as one of the most prevalent cancers in men, with a mortality rate of 18%. Chemotherapy is considered as a common strategy for cancer treatment; however, toxic side effects and drug resistance associated with chemotherapy are major drawbacks with this approach. It is well known that a diet rich in flavonoids can reduce the incidence of many types of cancer in a significant manner, and it was proved that methoxy flavones have greater bioavailability compared to the nonmethylated ones. Calycopterin is a tetramethoxy flavone which was demonstrated to have anti-proliferative effects on colon, gastric, and osteosarcoma cancer cells. Therefore, in the current study, we have evaluated the apoptotic effects of this flavonoid on two prostate cancer cell lines in-vitro. The MTT assay revealed that after 48 h treatment with this flavonoid, cell viability reduced to 50% compared to the control group. However, calycopterin treatment of healthy HUVEC did not cause any significant reduction in cell viability. Moreover, the clonogenic assay demonstrated that after 14 days, colony size and numbers reduced significantly in calycopterin treated cells. In addition, the percentage of the sub-G1 population in calycopterin-treated cells augmented significantly compared to untreated group. Also, calycopterin-treated cells demonstrated shiny condensed nuclei with fragmented DNA indicative of apoptosis. Finally, a significant reduction in the migration ability was observed in both lines subjected to calycopterin after 48 h. To conclude, our results demonstrated the apoptotic and anti-metastatic effects of calycopterin in both hormone-dependent and independent prostate cancer cell lines.

Second-generation testosterone-platinum(II) hybrids for site-specific treatment of androgen receptor positive prostate cancer: Design, synthesis and antiproliferative activity

Prostate cancer is the most diagnosed type of cancer in men in Canada. One out of eight men will be stricken with this disease during the course of his life. It is noteworthy that, at initial diagnoses 80–90% of cancers are androgen dependent. Hence, the androgen receptor is a viable biological target to be considered for drug targeting. We have developed a new generation of testosterone-Pt(II) hybrids for site-specific treatment of hormone-dependent prostate cancer. The hybrid molecules are made from testosterone using an eight-step reaction sequence with about 7% overall yield. They are linked with a stronger tether chain between the testosterone moiety and the Pt(II) moiety in comparison to our first generation hybrids. The new hybrids were tested on hormone-dependent and –independent prostate cancer cell lines. The hybrid 3a presents the best antiproliferative activity and was selective on hormone-dependent prostate cancer with IC50 of 2.2 μM on LNCaP (AR+) in comparison to 13.3 μM on PC3 (AR-) and 8.8 μM on DU145 (AR-) prostate cancer cells. On the same cell lines, CDDP displayed IC50 of 2.1 μM, 0.5 μM and 1.0 μM, respectively. Remarkably, hybrid 3a was inactive on both colon carcinoma (HT-29) and normal human adult keratinocyte cells (HaCat) with an IC50 of >25 μM. This is not the case for CDDP showing IC50 of 1.3 μM and 5.1 μM on HT-29 and HaCat cells, respectively. The potential for selective activity on androgen-receptor positive prostate cancer cells is confirmed with hybrid 3a giving new hope for an efficient and less toxic platinum-based treatment of prostate cancer patients.

MiR-221-5p regulates proliferation and migration in human prostate cancer cells and reduces tumor growth in vivo

BackgroundDespite latest advances in prostate cancer (PCa) therapy, PCa remains the third-leading cause of cancer-related death in European men. Dysregulation of microRNAs (miRNAs), small non-coding RNA molecules with gene expression regulatory function, has been reported in all types of epithelial and haematological cancers. In particular, miR-221-5p alterations have been reported in PCa.MethodsmiRNA expression data was retrieved from a comprehensive publicly available dataset of 218 PCa patients (GSE21036) and miR-221-5p expression levels were analysed. The functional role of miR-221-5p was characterised in androgen- dependent and androgen- independent PCa cell line models (C4–2 and PC-3M-Pro4 cells) by miR-221-5p overexpression and knock-down experiments. The metastatic potential of highly aggressive PC-3M-Pro4 cells overexpressing miR-221-5p was determined by studying extravasation in a zebrafish model. Finally, the effect of miR-221-5p overexpression on the growth of PC-3M-Pro4luc2 cells in vivo was studied by orthotopic implantation in male Balb/cByJ nude mice and assessment of tumor growth.ResultsAnalysis of microRNA expression dataset for human primary and metastatic PCa samples and control normal adjacent benign prostate revealed miR-221-5p to be significantly downregulated in PCa compared to normal prostate tissue and in metastasis compared to primary PCa. Our in vitro data suggest that miR-221-5p overexpression reduced PCa cell proliferation and colony formation. Furthermore, miR-221-5p overexpression dramatically reduced migration of PCa cells, which was associated with differential expression of selected EMT markers. The functional changes of miR-221-5p overexpression were reversible by the loss of miR-221-5p levels, indicating that the tumor suppressive effects were specific to miR-221-5p. Additionally, miR-221-5p overexpression significantly reduced PC-3M-Pro4 cell extravasation and metastasis formation in a zebrafish model and decreased tumor burden in an orthotopic mouse model of PCa.ConclusionsTogether these data strongly support a tumor suppressive role of miR-221-5p in the context of PCa and its potential as therapeutic target.

Novel monoclonal antibody therapeutics for metastatic castration resistant prostate cancer.

e14222 Background: Therapeutic potential of innate immunity comprising Natural killer cell based targets are beginning to unravel the complexity of immune responses. NK cells recognize and induce cytotoxicity of wide range of target cells, such as, tumor cells without prior antigen sensitization. In this study, we have studied Lectin-like transcript 1 (LLT1), a member of the C-type lectin super family, is expressed on target cells and various immune cells. LLT1 isoform 1, is known to interact with CD161, a critical receptor on NK cells. CD161 is expressed on most of human NK cells, NK-T cells, γδ T cells and so on. Tumors exploit the CD161- LLT1 interaction to evade host defense mechanism (“DO NOT KILL” signal); indicating LLT1 as an attractive immunotherapeutic strategy. Methods: Prostate cancer cell lines and other tumor cell lines were used to evaluate novel anti LLT1 antibodies for therapeutic potential - IFNγ production assays and tumor cell death assays were carried out. In vivo efficacy of these antibodies were established using PC3 xenograft in humanized mouse model (HuNOG-EXL). Results: Human androgen independent prostate cancer cell line, PC3 was studied for LLT1 expression and interactions with immune cells, to understand role of LLT1 in metastatic castration resistant prostate cancer (mCRPC). Overexpression of LLT1 on tumor cells was influenced by cytokines and various TLRs. Inhibition of CD161-LLT1 interaction with novel anti LLT1 antibodies leads to IFNγ production and consequent NK cell mediated cytotoxicity – hall mark of anti-tumor responses. Disruption of LLT1 - CD161 innate immunity axis with anti LLT1 antibody releases the break on NK cell cytotoxicity and hence, established a new therapeutic option. PC3 xenograft on HuNOG mouse revealed in vivo efficacy of LLT1 antibody. Significant tumor growth reduction was observed with specific anti LLT1 antibodies alone and in combination with check point antibodies. Thus, synergistic tumor growth reduction was established by combinatorial application of anti LLT1 antibody and PD1/PDL1 axis inhibitors. Conclusions: PC3 xenograft study and other results point to therapeutic opportunities in metastatic castration resistant prostate cancer, a disease condition which needs improved patient outcomes. The ligation of CD161/LLT1 will serve as a new immuno-oncology pair regulating innate and adaptive immune responses; novel human antibodies against LLT1 described here will bring therapeutic benefit to patients in need.

Cabazitaxel and silibinin co-encapsulated cationic liposomes for CD44 targeted delivery: A new insight into nanomedicine based combinational chemotherapy for prostate cancer

Cancer stem cells (CSCs) are the promising targets for cancer chemotherapy that cannot be eliminated by conventional chemotherapy. In this study cationic liposomes of cabazitaxel (CBX) and silibinin (SIL) were prepared with an aim to kill cancer cells and CSCs for prostate cancer. CBX act as cancer cell inhibitor and SIL as CSC inhibitor. Hyaluronic acid (HA), an endogenous anionic polysaccharide was coated on cationic liposomes for targeting CD44 receptors over expressed on CSCs. Liposomes were prepared by ethanol injection method with particle size below 100 nm and entrapment efficiency of more than 90% at 10% w/w drug loading. Liposomes were characterized by dynamic light scattering, transmission electron microscopy, 1H nuclear magnetic resonance and scanning electron microscopy-energy dispersive x-ray spectroscopy. Liposomes were evaluated for their anticancer action in androgen independent human prostate cancer cell lines (PC-3 and DU-145). HA coated liposomes showed potential cytotoxicity over other groups with low IC50, significantly inhibited cell migration and induced apoptosis. Synergistic cytotoxic effect was also observed with HA coated liposomes that resulted in colony formation inhibition and G2/M phase arrest. Proficient cytotoxicity against CD44+ cells (14.87 ± 0.41% in PC-3 and 33.95 ± 0.68% in DU-145 cells) indicated the efficiency of HA coated liposomes towards CSC targeting. Hence, the outcome of this combinational therapy with CD44 targeting indicates the suitability of HA coated CBX and SIL co-loaded liposomes as a potential approach for eradicating prostate cancer and herein might provide a insight for future studies.

Abstract 1516: BAT-1 up-regulation decreases cell invasion and migration in vitro

Abstract Prostate cancer is the most common cancer in men in the United States and is the third cause of cancer related deaths in men. In Puerto Rico, prostate cancer is the most common type of cancer and the leading cause of cancer death in men. Approximately one out of seven men will be diagnosed with prostate cancer in his lifetime. The current available biomarkers are unable to predict malignant outcomes such as recurrence. Thus, there is a critical demand for the development of innovative diagnostic and prognostic tools for the management of prostate cancer. In the current study, we evaluated the biological role of BAT-1 in prostate cancer. Preliminary data from patients who had prostate cancer recurrence identified that HLA-B associated transcript 1 (BAT-1) was down-regulated in patients with prostate cancer recurrence when compared with non-recurrent patients. We up-regulated BAT-1 in androgen independent PC3 and androgen dependent 22RV1 prostate cancer cell lines using a BAT-1 recombinant protein. In vitro assays were performed to measure proliferation, migration and invasion. Proliferation assays using MTS showed no significant change in BAT-1 recombinant protein concentrations ranging from 0.25-2.0μg/mL at 12, 24 and 48hrs for PC3 and 22RV1 cells. Wound healing assay showed that cells treated with BAT-1 recombinant protein at 0.25μg/mL and 2.0μg/mL significantly decreased migration at 6, 12hrs and 24hrs in PC3 cells when compared to control. Boyden assay showed that cells treated with BAT-1 recombinant protein at 0.25μg/mL and 2.0μg/mL significantly decreased invasion in PC3 and 22RV1 cells at 24hrs when compared to control. Our results showed that BAT-1 up-regulation decreased migration and cell motility. Cells did not show a significant change in proliferation. These results were expected due to previous data showing that BAT-1 down-regulation increased cell migration and invasion, suggesting that BAT-1 expression promotes aggressiveness in prostate cancer recurrence. A possible mechanism of action for BAT1 is the modulation of cell migration and invasion. Citation Format: Aileen M. Garcia-Vargas, Maria Sanchez, Magaly Martinez-Ferrer. BAT-1 up-regulation decreases cell invasion and migration in vitro [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 1516.

Probing the prostate tumour microenvironment I: impact of glucose deprivation on a cell model of prostate cancer progression.

In the developed world, prostate cancer is the most common cancer diagnosis in men. Although prostate cancer initially presents as a non life-threatening disease, 90% of patients will develop castration resistant prostate cancer (CRPC), which preludes distant metastasis and is largely accountable for prostate cancer associated deaths. This is because as yet, there are no viable molecular therapeutic targets for effective treatment of CRPC. It is now widely accepted that cancer cells can alter their metabolic profile during the course of tumourgenesis and metastasis such that they are able to survive in oxygen and nutrient-poor environments. This work was aimed towards gaining greater mechanistic understanding of how such stresses in the tumour microenvironment impact on both androgen sensitive (LNCaP) and androgen independent (LNCaP-abl and LNCaP-abl-Hof) prostate cancer cell lines. Here we have applied technically robust and reproducible label-free liquid chromatography mass spectrometry analysis for comprehensive proteomic profiling of prostate cancer cell lines under nutrient deficient (low glucose) conditions. This led to the identification of approximately 4,000 proteins - one of the largest protein datasets for prostate cancer cell lines established to date. The biological and clinical significance of proteins showing a significant change in expression as result of low glucose conditions was established. Novel, intuitive workflows were subsequently implemented to ensure the verification of selected proteins of interest in a robust, reproducible and high throughput manner. Overall, these data suggest that this strategy supports identification of protein biomarkers of prostate cancer progression and potential therapeutic targets for CRPC.

Probing the prostate tumour microenvironment II: Impact of hypoxia on a cell model of prostate cancer progression

Approximately one in six men are diagnosed with Prostate Cancer every year in the Western world. Although it can be well managed and non-life threatening in the early stages, over time many patients cease to respond to treatment and develop castrate resistant prostate cancer (CRPC). CRPC represents a clinically challenging and lethal form of prostate cancer. Progression of CRPC is, in part, driven by the ability of cancer cells to alter their metabolic profile during the course of tumourgenesis and metastasis so that they can survive in oxygen and nutrient-poor environments and even withstand treatment. This work was carried out as a continuation of a study aimed towards gaining greater mechanistic understanding of how conditions within the tumour microenvironment impact on both androgen sensitive (LNCaP) and androgen independent (LNCaP-abl and LNCaP-abl-Hof) prostate cancer cell lines. Here we have applied technically robust and reproducible label-free liquid chromatography mass spectrometry analysis for comprehensive proteomic profiling of prostate cancer cell lines under hypoxic conditions. This led to the identification of over 4,000 proteins – one of the largest protein datasets for prostate cancer cell lines established to date. The biological and clinical significance of proteins showing a significant change in expression as result of hypoxic conditions was established. Novel, intuitive workflows were subsequently implemented to enable robust, reproducible and high throughput verification of selected proteins of interest. Overall, these data suggest that this strategy supports identification of protein biomarkers of prostate cancer progression and potential therapeutic targets for CRPC.

Investigating the role of caveolin-2 in prostate cancer cell line

Prostate cancer is a worldwide problem. While the role of caveolin-1 has been extensively studied, little is known about the role of caveolin-2 (CAV2) in prostate cancer. Up-regulation of CAV2 in androgen independent PC3 cells compared to normal prostate cell line and androgen dependent prostate cancer cell lines has been observed. Recent studies suggest that up-regulation of CAV2 plays an important role in androgen independent prostate cancer. This study investigates whether CAV2 is important in mediating the aggressive phenotypes seen in androgen independent prostate cancer cells. The androgen independent prostate cancer cell line, PC3 was used that has been shown to express CAV2, and CAV2 knock down was performed using siRNA system. Changes to cell number, migration and invasion were assessed after knocking down CAV2. Our results showed that down-regulating CAV2 resulted in reduced cell numbers, migration and invasion in PC3 cells. This preliminary study suggests that CAV2 may act to promote malignant behavior in an androgen independent prostate cancer cell line. Further studies are required to fully elucidate the role of CAV2 in androgen independent prostate cancer.

Bifunctional conjugates with potent inhibitory activity towards cyclooxygenase and histone deacetylase

We herein disclose a series of compounds with potent inhibitory activities towards histone deacetylases (HDAC) and cyclooxygenases (COX). These compounds potently inhibited the growth of cancer cell lines consistent with their anti-COX and anti-HDAC activities. While compound 2b showed comparable level of COX-2 selectivity as celecoxib, compound 11b outperformed indomethacin in terms of selectivity towards COX-2 relative to COX-1. An important observation with our lead compounds (2b, 8, 11b, and 17b) is their enhanced cytotoxicity towards androgen dependent prostate cancer cell line (LNCaP) relative to androgen independent prostate cancer cell line (DU-145). Interestingly, compounds 2b and 17b arrested the cell cycle progression of LNCaP in the S-phase, while compound 8 showed a G0/G1 arrest, similar to SAHA. Relative to SAHA, these compounds displayed tumor-selective cytotoxicity as they have low anti-proliferative activity towards healthy cells (VERO); an attribute that makes them attractive candidates for drug development.

Synthesis of novel C2-symmetric testosterone dimers and evaluation of antiproliferative activity on androgen-dependent and -independent prostate cancer cell lines

A series of 7α-linked testosterone dimers were made and tested for biological activity on both androgen-dependent (LNCaP) and androgen-independent (DU-145 and PC3) prostate cancer cell lines. The synthesis proceeds through the formation of a trans-4-(17β-acetoxy-4-androsten-3-one-7α-yl)-but-2-enoic acid 4-hydroxy-alkyl ester intermediate of various length (7a–d) followed by the final dimerization step. The dimers showed interesting biological activity in comparison to the ω-hydroxyalkyl ester intermediates 7a–d. The most active dimer 8a (n=1) showed IC50 of 3.8, 1.4 and 1.8μM, respectively on LNCaP, DU-145 and PC3 cancer cell lines. On these cell lines, this dimer is about 12, 70 and 47 times more powerful than cyproterone acetate (CPA) the reference antiandrogen. Furthermore, dimers 8b–d (n=2, 3, 4) were less active than 8a but showed selective activity on androgen-dependent LNCaP prostate cancer cells. This indicates possible application for the treatment of androgen-dependent prostate cancer.

Abstract 1360: Genistein-AR antagonist conjugate potently induce growth arrest in both androgen dependent and independent prostate cancer cell lines

Abstract The ability of prostate cancer to transition from androgen dependent- to androgen independent state (castration resistant), during androgen deprivation therapy, requires that new chemotherapeutic agents be introduced that will retain activity in the two prostate cancer types. Though described as androgen independent, castration resistant prostate cancer (CRPC) still relies heavily on androgen receptor (AR) signaling for survival. Enzalutamide, an AR antagonist, is probably that most effective drug used in androgen deprivation therapy for CRPC and AR dependent prostate cancer. Despite its clinical success, there is the fear of resistant to enzalutamide due to mutation in the ligand binding domain of AR. This poses a challenge that requires developing new agents devoid of resistance, in the event of mutation. One of the approaches being proposed is to develop new agents that can selectively induce AR degradation, an approach that was successfully used to develop the selective estrogen receptor degrader, fulvestrant. In our study, we incorporate genistein, a soy isoflavone known to perturb multiple biochemical pathways relevant for cancer survival, into an AR binding template to give AG-1-33. AG-1-33 was evaluated for growth inhibitory activity in LNCaP (AR dependent) and DU-145 (AR independent) prostate cancer cell lines. This compound retained its AR antagonist activity and also showed low micromolar anti-proliferation activity in the two prostate cancer cell lines. We also evaluated the effect of AG-1-33 on AR expression, as well as its ability to act as a selective AR degrader in LNCaP. Citation Format: Idris O. Raji, Alex George, Omer Kucuk, Adegboyega K. Oyelere. Genistein-AR antagonist conjugate potently induce growth arrest in both androgen dependent and independent prostate cancer cell lines. [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 1360.