Journal: Nature Communications

Article Title: Interplay between hypoxia and androgen controls a metabolic switch conferring resistance to androgen/AR-targeted therapy

doi: 10.1038/s41467-018-07411-7

Figure Lengend Snippet: Chronic-ADT in hypoxia induces resistance to targeted disruption of androgen/AR-axis. a Schematic representation of repeated-ADT treatment of LNCaP and LAPC4 cells in hypoxia (1% O 2 ) or normoxia (20% O 2 ). Cells were cultured in charcoal-stripped serum (CSS) media with or without 1 nM synthetic androgen R1881 (R1881 or ADT). Each round/cycle of treatment lasted 48 h, and the survived cells were recovered and re-plated for the same treatment in the following cycle. b Dose-dependent sensitivity of LNCaP clones to enzalutamide. Cells derived from a were cultured with CSS + R1881, and treated by increasing doses of enzalutamide in normoxia or hypoxia for 96 h. Afterwards, cell growth and viability were determined based on viable cells quantitated with SYTO 60 fluorescence staining and imaging. The AdtHs is represented by purple line. All values were mean ± s.d. relative to solvent-treated controls, n = 3. * P

Article Snippet: Androgen-dependent prostate cancer cell lines LNCaP and LAPC4 were purchased from ATCC.

Techniques: Cell Culture, Clone Assay, Derivative Assay, Fluorescence, Staining, Imaging

Journal: Cancer research

Article Title: Loss of FOXO1 cooperates with TMPRSS2-ERG overexpression to promote prostate tumorigenesis and cell invasion

doi: 10.1158/0008-5472.CAN-17-0686

Figure Lengend Snippet: FOXO1 interacts directly with ERG. A, Western blot analysis of endogenous FOXO1 and ERG proteins in VCaP cells immunoprecipitated by indicated antibody or non-specific IgG. B, Western blot analysis of ectopically expressed proteins immunoprecipitated by indicated antibody or non-specific IgG in PC-3 cells transfected with indicated Flag- and Myc-tagged plasmids. C , Schematic depicting a set of GST-ERG recombinant protein constructs. D, Coomassie Blue staining of GST and GST-ERG recombinant proteins purified from bacteria (bottom), and Western blot analysis of FOXO1 proteins in LAPC-4 cells pulled down by GST recombinant proteins (top). Red arrow, non-specific band. * in red, unexpected, but reproducible band shift. * in black, denotes expected fragment size. E, Schematic depicting a set of GST-FOXO1 recombinant protein constructs. F, GelCode Blue staining of GST and GST-FOXO1 recombinant proteins purified from bacteria (bottom), and Western blot analysis of in vitro transcribed and translated ERG proteins pulled down by GST recombinant proteins (top).

Article Snippet: The cell lines VCaP, PC-3, and LAPC-4 were purchased from ATCC (Manassas, VA) and authenticated via STR profiling.

Techniques: Western Blot, Immunoprecipitation, Transfection, Recombinant, Construct, Staining, Purification, Electrophoretic Mobility Shift Assay, In Vitro

Journal: Clinical nutrition experimental

Article Title: Arctigenin inhibits prostate tumor cell growth in vitro and in vivo

doi: 10.1016/j.yclnex.2017.04.001

Figure Lengend Snippet: Inhibition of xenograft tumor growth, blood and tissue levels of arctigenin, and reduced expression of growth factors in SCID mice. Male SCID mice (n=10 per group) were inoculated subcutaneously with 5×10 5 androgen-sensitive LAPC-4 prostate tumor cells. The intervention treatments started either one week after (A, B) or two weeks prior to (C, D) the tumor cell inoculation. Arctigenin was administered daily through oral gavage at dose of 50mg/kg or 100mg/kg bw. Tumor size was measured twice a week using calipers and tumor volume calculated using the formula: length × width × height × 0.5236. Tumor weight was measured at mouse sacrifice. Blood and tumor levels of arctigenin (E) was detected with HPLC coupled with CoulArray electrochemical detector after extraction with ethyl acetate. The concentrations of 8 growth factors in tumor tissues were simultaneously measured using a Human Growth Factor ELISA Strip II kit (F). Data are presented as mean ± SD. Arc: arctigenin. Compared to control, *P

Article Snippet: The androgen-sensitive LNCaP and LAPC-4 human prostate cancer cell lines were purchased from ATCC (Chicago, IL, USA).

Techniques: Inhibition, Expressing, Mouse Assay, High Performance Liquid Chromatography, Enzyme-linked Immunosorbent Assay, Stripping Membranes

Journal: PLoS ONE

Article Title: TIMP-1 Promotes Accumulation of Cancer Associated Fibroblasts and Cancer Progression

doi: 10.1371/journal.pone.0077366

Figure Lengend Snippet: TIMP-1 promotes in vivo growth of prostate cancers. A . Establishment of pooled populations of PC3, 22RV1, and LAPC-4 cells expressing v5 epitope tagged TIMP-1 or transduced with the empty expression vector alone (controls). Secreted v5-tagged TIMP-1 was detected by anti-v5 mAb (Invitrogen). B . TIMP-1 promotes PC3 and 22RV1 prostate cancer growth in vivo . Weights of the tumors derived from PC3 and 22RV1 prostate cancer cells expressing TIMP-1v5 or infected with the empty expression constructs (PC3-control and 22RV1-control) were measured 5 or 10-weeks, respectively, after subcutaneous implantation of the cancer cells into Rag-2/II2rg mice (Taconic). n=6. * p

Article Snippet: Human prostate cancer LAPC-4 cells were obtained from the ATCC Patent Depository.

Techniques: In Vivo, Expressing, Transduction, Plasmid Preparation, Derivative Assay, Infection, Construct, Mouse Assay

Journal: PLoS ONE

Article Title: TIMP-1 Promotes Accumulation of Cancer Associated Fibroblasts and Cancer Progression

doi: 10.1371/journal.pone.0077366

Figure Lengend Snippet: TIMP-1 enhances expression of Snail, MMP-2, and MMP-9 in prostate cancers. Expression levels of several EMT markers, such as Snail (A, E, I, M), Slug (B, F, J, N), MMP-2 (C, G, K, O), and MMP-9 (D, H, L, P) were assessed on the tumor sections derived from PC3/LAPC-4 control cells (A-D and I-L, respectively) and PC3/LAPC-4-TIMP-1 cells (E-H and M-P, respectively). The results show that TIMP-1 enhances expression of Snail, MMP-2, and MMP-9 but not Slug in the prostate cancer tissues. Bar, 50 µm in A, B, E, F, I, J, M, N and 100 µm in C, D, G, H, K, L, O, P.

Article Snippet: Human prostate cancer LAPC-4 cells were obtained from the ATCC Patent Depository.

Techniques: Expressing, Derivative Assay

Journal: Oncotarget

Article Title: Therapy-induced developmental reprogramming of prostate cancer cells and acquired therapy resistance

doi: 10.18632/oncotarget.14850

Figure Lengend Snippet: Developmental reprogramming of PCa Cells to a stem-like intermediate ( A ) Chronic AD of LNCaP cells upregulated genes associated with N/NC stem cells and derivative tissues. LNCaP cells (Left) cultured in androgen-depleted medium for 15-days undergo a morphological transformation (Middle), and overexpress genes associated with N/NC stem cells and derivative tissues shown on heat-mapping (Right). ( B ) Representative phase-contrast images of LNCaP cells over two weeks of culture in STM. ( C ) Representative phase-contrast images of VCaP, LAPC4 or 22Rv1 spheroids after 14-day STM-mediated reprogramming. Scale bars represent 100 μm. ( D ) Proliferation assays of parental LNCaP (P) and reprogrammed stem-like LNCaP cells (SL) showed that the doubling time of LNCaP-SL is significantly greater than parentals. Mean ± standard error; ** P

Article Snippet: Cell culture and xenografts LNCaP, VCaP, 22Rv1 (ATCC, Manassas, VA) and LAPC4 cell line (obtained from Dr. Charles Sawyer), were maintained in phenol red-free ATCC-recommended media.

Techniques: Cell Culture, Transformation Assay

Journal: Hormones & cancer

Article Title: Glucocorticoid Receptor Activity Contributes to Resistance to Androgen-Targeted Therapy in Prostate Cancer

doi: 10.1007/s12672-014-0173-2

Figure Lengend Snippet: Increased regulation of SGK1 by the GR following AR antagonism. A mRNA was collected from LAPC4 and 22Rv1 cells treated for 2 h under various hormonal conditions, and SGK1 gene expression was assessed by qRT-PCR. B Both 22Rv1 and LAPC4 cells were treated for 3 days in either vehicle, R1881 (1 nM), or R1881 + MDV3100 (10 μM)-containing media. Conditions specifying dexamethasone treatment were stimulated with Dex (100 nM) for 1 h prior to chromatin harvest. Targeted qPCR was performed for the region within the SGK1 promoter after ChIP, and y -values represent fold chromatin enrichment (relative to IgG controls for each condition). Error bars represent standard deviation of mean cycle threshold values, and asterisk indicates p

Article Snippet: The LAPC4 human PC cell line was cultured in IMDM (ATCC) supplemented with 10 % FCS, 1 % penicillin/streptomycin, and 1 nM R1881 (Sigma-Aldrich), and VCaP cells lines were cultured in DMEM (Mediatech) supplemented with 10 % FCS and 1 % penicillin/streptomycin.

Techniques: Expressing, Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Chromatin Immunoprecipitation, Standard Deviation

Journal: Hormones & cancer

Article Title: Glucocorticoid Receptor Activity Contributes to Resistance to Androgen-Targeted Therapy in Prostate Cancer

doi: 10.1007/s12672-014-0173-2

Figure Lengend Snippet: GR expression in prostate cancer following androgen signaling inhibition. A Western blot of GR and AR expression for PC cell lines PC3, DU145, LNCaP, LAPC4, CWR-22Rv1, and VCaP. B NRC31 ( GR ) mRNA expression ( above ) and protein expression ( below ) in PC cell lines after long-term ( > 30 days) treatment with AR antagonist MDV3100. Error bars represent standard error of the mean, and asterisk indicates p

Article Snippet: The LAPC4 human PC cell line was cultured in IMDM (ATCC) supplemented with 10 % FCS, 1 % penicillin/streptomycin, and 1 nM R1881 (Sigma-Aldrich), and VCaP cells lines were cultured in DMEM (Mediatech) supplemented with 10 % FCS and 1 % penicillin/streptomycin.

Techniques: Expressing, Inhibition, Western Blot

Journal: Hormones & cancer

Article Title: Glucocorticoid Receptor Activity Contributes to Resistance to Androgen-Targeted Therapy in Prostate Cancer

doi: 10.1007/s12672-014-0173-2

Figure Lengend Snippet: GR expression/activation levels following AR inhibitor therapy. Hormonal therapies utilized included: R1881 (1 nM), a synthetic androgen; dexamethasone (Dex, 100 nM), a GR agonist; and MDV3100 (MDV, 10 μM), an AR antagonist. A 22Rv1 and LAPC4 GR mRNA expression analysis by qRT-PCR after 3 days of treatment; normalized to GAPDH and depicted as fold change relative to vehicle. B Western blot analysis of 22Rv1 and LAPC4 GR and AR protein expression after 3 and 7 days of treatment. All conditions were treated with Dex and R1881. Asterisk denotes AR splice variants within 22Rv1 cells. C 22Rv1 and LAPC4 cells were treated for 14 days under various conditions

Article Snippet: The LAPC4 human PC cell line was cultured in IMDM (ATCC) supplemented with 10 % FCS, 1 % penicillin/streptomycin, and 1 nM R1881 (Sigma-Aldrich), and VCaP cells lines were cultured in DMEM (Mediatech) supplemented with 10 % FCS and 1 % penicillin/streptomycin.

Techniques: Expressing, Activation Assay, Quantitative RT-PCR, Western Blot

Journal: Hormones & cancer

Article Title: Glucocorticoid Receptor Activity Contributes to Resistance to Androgen-Targeted Therapy in Prostate Cancer

doi: 10.1007/s12672-014-0173-2

Figure Lengend Snippet: Increased regulation of PSA by the GR following AR antagonism. A mRNA was collected from LAPC4 and 22Rv1 cells treated for 2 h under various hormonal conditions, and PSA gene expression was assessed by qRT-PCR. B Both 22Rv1 and LAPC4 cells were treated for 3 days in either vehicle, R1881 (1 nM), or R1881 + MDV3100 (10 μM) containing media. Conditions specifying dexamethasone treatment were stimulated with Dex (100 nM) for 1 h prior to chromatin harvest. Targeted qPCR was performed for the PSA promoter after ChIP, and y -values represent fold chromatin enrichment (relative to IgG controls for each condition). Error bars represent standard deviation of mean cycle threshold values, and asterisk indicates p

Article Snippet: The LAPC4 human PC cell line was cultured in IMDM (ATCC) supplemented with 10 % FCS, 1 % penicillin/streptomycin, and 1 nM R1881 (Sigma-Aldrich), and VCaP cells lines were cultured in DMEM (Mediatech) supplemented with 10 % FCS and 1 % penicillin/streptomycin.

Techniques: Expressing, Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Chromatin Immunoprecipitation, Standard Deviation

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: MEIS1 and MEIS2 Expression and Prostate Cancer Progression: A Role For HOXB13 Binding Partners in Metastatic Disease

doi: 10.1158/1078-0432.CCR-17-3673

Figure Lengend Snippet: Functional Characterization of Total MEIS Knockdown LAPC-4 Cells (A) Assessment of MEIS1 and MEIS2 knockdown in LAPC-4 cell using RT-PCR (top) and Western blot analysis (bottom). Two shRNA constructs against MEIS1 and two shRNA constructs against MEIS2 were utilized to control for off-target effects. To determine MEIS1 and MEIS2 protein levels after shRNA depletion, western blot analyses was conducted using a mouse monoclonal targeting both MEIS1 and MEIS2 (Pan-MEIS). (B) Xenograft growth of MEIS1 and MEIS2 double knockdown LAPC-4 cells injected subcutaneously on the flanks of hormonally-intact male nude mice compared to non-silencing controls. Differences in tumor volumes at days 25, 27 and 29 post-injection were statistically significant (Control N=8; double knockdown N=8; * p -value

Article Snippet: Cell authentication of LAPC-4 cells was confirmed via DDC Medical services (Fairfield, OH).

Techniques: Functional Assay, Reverse Transcription Polymerase Chain Reaction, Western Blot, shRNA, Construct, Injection, Mouse Assay