Human Prostate Tumor Xenografts Research Articles

Tumor characterization with dynamic contrast enhanced magnetic resonance imaging and biodegradable macromolecular contrast agents in mice.

To investigate the efficacy of polydisulfide-based biodegradable macromolecular contrast agents of different degradability and molecular weight for tumor characterization based on angiogenesis using dynamic contrast enhanced MRI (DCE-MRI). Biodegradable macromolecular MRI contrast agents, Gd-DTPA cystamine copolymers (GDCC) and Gd-DTPA cystine copolymers (GDCP), with molecular weight of 20 and 70 KDa were evaluated for tumor characterization. Gd(DTPA-BMA) and a prototype of macromolecular contrast agent, albumin-(Gd-DTPA), were used as controls. The DCE-MRI studies were performed in nude mice bearing MDA PCa 2b and PC-3 human prostate tumor xenografts. Tumor angiogenic kinetic parameters including endothelium transfer coefficient (K(trans)) and fractional tumor plasma volume (f(PV)) were calculated from the DCE-MRI data using a two-compartment model and compared between the two different tumor models for each contrast agent. There was no significant difference in the f(PV) values between two tumor models estimated with the same agent except for GDCC-70. The K(trans) values in both tumor models decreased with the increase of molecular weight of contrast agents. With the same high molecular weight (70 KDa), GDCC-70 showed a higher K(trans) values than GDCP-70 due to high degradability of the former in both tumor models (p < 0.05). The K(trans) values of MDA PCa 2b tumors were significantly higher than those of PC-3 tumors estimated by Gd(DTPA-BMA), GDCC-20, GDCC-70, GDCP-70, and albumin-(Gd-DTPA) (p < 0.05). The polydisulfide-based biodegradable macromolecular MRI contrast agents are promising in tumor characterization and differentiation with dynamic contrast enhanced MRI.

Synchrotron X‐ray imaging reveals a correlation of tumor copper speciation with Clioquinol's anticancer activity

Tumor development and metastasis depend on angiogenesis that requires certain growth factors, proteases, and the trace element copper (Cu). Recent studies suggest that Cu could be used as a novel target for cancer therapies. Clioquinol (CQ), an antibiotic that is able to form stable complexes with Cu or zinc (Zn), has shown proteasome-inhibitory, androgen receptor-suppressing, apoptosis-inducing, and antitumor activities in human cancer cells and xenografts. The mechanisms underlying the interaction of CQ with cellular Cu, the alteration of the Cu/Zn ratio and the antitumor role of CQ in vivo have not been fully elucidated. We report here that Cu accumulates in tumor tissue and that the Cu/Zn balances in tumor, but not normal, tissue change significantly after the treatment with CQ. Cu speciation analysis showed that the Cu(I) species is predominant in both normal and tumor tissues and that Cu(II) content was significantly increased in tumor, but not normal tissue after CQ treatment. Our findings indicate that CQ can interact with cellular Cu in vivo, dysregulates the Cu/Zn balance and is able to convert Cu(I) to Cu(II) in tumor tissue. This conversion of Cu(I) to Cu(II) may be associated with CQ-induced proteasome inhibition and growth suppression in the human prostate tumor xenografts.

Adenoviral-Mediated Endothelial Precursor Cell Delivery of Soluble CD115 Suppresses Human Prostate Cancer Xenograft Growth in Mice

Prostate cancer tumor growth and neovascularization is promoted by an interplay between migratory tumor stromal cells such as specialized tumor-associated macrophages (TAMs) and circulating endothelial precursor cells (CEPs). As vehicles for tumor therapy, human CEPs are relatively easy to isolate from peripheral blood, are able to proliferate long-term in vitro, are amenable to viral manipulation, and preferentially home to regions of ischemia found in growing tumors. We show here that human peripheral blood CEPs expanded ex vivo migrate to prostate cancer cells in vitro and efficiently home to human prostate tumor xenografts in vivo. Infection of precursors ex vivo with an adenovirus constructed to secrete a soluble form of the colony-stimulating factor-1 receptor CD115 that inhibits macrophage viability and migration in vitro significantly decreases the number of TAMs in xenografts (p < .05), reduces proliferation (p < .01) and vascular density (p < .03), and suppresses the growth of xenografts (p < .03). These data show for the first time that targeting stromal cell processes with cellular therapy has the potential to retard prostate tumor growth.

Anticancer oncolytic activity of respiratory syncytial virus

Oncolytic virotherapy is an emerging bio-therapeutic platform for cancer treatment, which is based on selective infection/killing of cancer cells by viruses. Herein we identify the human respiratory syncytial virus (RSV) as an oncolytic virus. Using prostate cancer models, we show dramatic enhancement of RSV infectivity in vitro in the androgen-independent, highly metastatic PC-3 human prostate cancer cells compared to the non-tumorigenic RWPE-1 human prostate cells. The oncolytic efficiency of RSV was established in vivo using human prostate tumor xenografts in nude mice. Intra-tumoral and intra-peritoneal injections of RSV led to a significant regression of prostate tumors. Furthermore, enhanced viral burden in PC-3 cells led to selective destruction of PC-3 cancer cells in vitro and in xenograft tumors in vivo due to apoptosis triggered by the down-regulation of NF-κB activity (and the resulting loss of anti-apoptotic function of NF-κB) in RSV-infected PC-3 cells. The intrinsic (mitochondrial) pathway constitutes the major apoptotic pathway; however, the death-receptor-dependent extrinsic pathway, mediated by the paracrine/autocrine action of tumor necrosis factor-α produced from infected cells, also partly contributed to apoptosis. Thus, the oncolytic property of RSV can potentially be exploited to develop targeted therapeutics for the clinical management of prostate tumors.

In vivo targeted killing of prostate tumor cells by a synthetic amoebapore helix 3 peptide modified with two γ-linked glutamate residues at the COOH terminus

We previously designed a pro-cytolytic peptide to target prostate-specific membrane antigen (PSMA)-positive prostate tumor cells. The backbone of the peptide was derived from the cell lytic amoebapore H-3 domain, which becomes completely inactive upon modification by two glutamate residues linked to the ε-amide group of the COOH-terminal lysine through γ-linkages (H-3Glu2). This modified H-3 domain regains its lytic activity against PSMA-positive cells (LNCaP) after the γ-linked glutamate residues are cleaved by PSMA. Our previous in vitro results demonstrate that the modified amoebapore peptide has strong cytolytic activity towards PSMA-positive cells and very little activity towards PSMA-negative cells. In the present study, the in vivo efficacy of this modified peptide was examined in human LNCaP prostate tumor xenografts in nude mice. The results showed significantly decreased tumor size and PSA levels in treated mice as compared to control mice. As well, 5/12 of the treated mice were tumor-free. Peptide distribution studies showed that peptide levels in the prostate tumors maintained a steady concentration for approximately 6 hours. Single-dose toxicity studies showed no toxic effects of the peptide when administered intraperitoneally or intravenously at a dose of 30 mg/kg.

MOLECULAR BASIS OF RADIATION AND CHEMOTHERAPY SENSITIZATION BY ANTI-β2-MICROGLOBULIN ANTIBODY IN HUMAN PROSTATE CANCER CELLS AND TUMOR XENOGRAFTS

You have accessJournal of Urology1 Apr 2009MOLECULAR BASIS OF RADIATION AND CHEMOTHERAPY SENSITIZATION BY ANTI-β2-MICROGLOBULIN ANTIBODY IN HUMAN PROSTATE CANCER CELLS AND TUMOR XENOGRAFTS Yasuhiro Matsuoka, Sajni Josson, Wen-Chin Huang, Fray F Marshall, and Leland Wk Chung Yasuhiro MatsuokaYasuhiro Matsuoka More articles by this author , Sajni JossonSajni Josson More articles by this author , Wen-Chin HuangWen-Chin Huang More articles by this author , Fray F MarshallFray F Marshall More articles by this author , and Leland Wk ChungLeland Wk Chung More articles by this author View All Author Informationhttps://doi.org/10.1016/S0022-5347(09)61453-1AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail "MOLECULAR BASIS OF RADIATION AND CHEMOTHERAPY SENSITIZATION BY ANTI-β2-MICROGLOBULIN ANTIBODY IN HUMAN PROSTATE CANCER CELLS AND TUMOR XENOGRAFTS." The Journal of Urology, 181(4S), pp. 514–515 © 2009 by American Urological AssociationFiguresReferencesRelatedDetails Volume 181Issue 4SApril 2009Page: 514-515 Advertisement Copyright & Permissions© 2009 by American Urological AssociationMetricsAuthor Information Yasuhiro Matsuoka More articles by this author Sajni Josson More articles by this author Wen-Chin Huang More articles by this author Fray F Marshall More articles by this author Leland Wk Chung More articles by this author Expand All Advertisement PDF downloadLoading ...

Characterization of a targeted nanoparticle functionalized with a urea-based inhibitor of prostate-specific membrane antigen (PSMA)

Polymeric nanoparticles represent a form of targeted therapy due to their ability to passively accumulate within the tumor interstitium via the enhanced permeability and retention (EPR) effect. We used a combined approach to decorate the surface of a nanoparticle with a urea-based small-molecule peptidomimetic inhibitor of prostate specific membrane antigen (PSMA). PSMA is expressed by normal and malignant prostate epithelial cells and by the neovasculature of almost all solid tumors. This strategy takes advantage of both the avidity of the functionalized nanoparticle for binding to PSMA and the ability of the nanoparticle to be retained for longer periods of time in the tumor due to enhanced leakage via EPR into the tumor interstitium. As an initial step to introducing the targeting moiety, the amino terminus of the small-molecule PSMA inhibitor was conjugated to PEG (Mn 3400) bearing an activated carboxyl group to obtain a PEGylated inhibitor. Studies undertaken using a radiolabeled PSMA-substrate based assay established that the PEGylated inhibitor had an IC50 value similar to the uncomplexed inhibitor. Subsequently, nanoparticles loaded with docetaxel were formulated using a mixture of poly(lactide-β-ethylene glycol-β-lactide) and PSMA-inhibitor bound α-amino-ω-hydroxy terminated poly(ethylene glycol-β-ε-caprolactone). In vitro studies using these nanoparticles demonstrated selective cytotoxicity against PSMA-producing cells. Binding of fluorescently labeled PSMA-targeted particles to PSMA-producing cells has also been directly observed using fluorescence microscopy and observed in secondary fashion using a PSMA substrate based enzyme inhibition assay. Ongoing in vivo studies address the localization, activity and toxicity of these targeted nanoparticles against PSMA-producing human prostate tumor xenografts.

Choline Autoradiography of Human Prostate Cancer Xenograft: Effect of Castration

The purpose of this study was to investigate the effects of castration and tracer uptake time interval on the level of radiolabeled choline accumulation in murine-implanted human prostate tumor xenografts using quantitative autoradiography. We implanted androgen-dependent (CWR22) and androgen-independent (PC3) human prostate cancer cells in castrated (n = 9) and noncastrated (n = 9) athymic male mice and allowed tumors to grow to 1 cm3. The mice were euthanized at 5, 10, and 20 minutes after injection of 5 microCi [14C]-choline. Mice were prepared for quantitative autoradiography with density light units of viable tumor sections converted to units of radioactivity (nCi/mm2) using calibration. Two-group comparisons were performed using a two-tailed Student t-test with unequal variance and with a significance probability level of less than .05. Two-group comparisons between the means of the tracer uptake level for each tumor type at each of three time points for each of two host types showed that (1) the level of tracer localization in the two tumor types was affected little in relation to the host type and (2) PC3 tumor uptake level tended to increase slowly with time only in the noncastrated host, whereas this was not observed in the castrated host or with CWR22 tumor in either host type. The uptake time interval and castration do not appear to significantly affect the level of radiolabeled choline uptake by the human prostate cancer xenograft.

A 19F-NMR approach using reporter molecule pairs to assess beta-galactosidase in human xenograft tumors in vivo.

Gene therapy has emerged as a promising strategy for treatment of various diseases. However, widespread implementation is hampered by difficulties in assessing the success of transfection in the target tissue and the longevity of gene expression. Thus, there is increasing interest in the development of non-invasive in vivo reporter techniques to assay gene expression. We recently demonstrated the ability to detect beta-galactosidase activity in stably transfected human prostate tumor xenografts in mice in vivo using 19F NMR. We now extend the studies to human MCF7 breast cancer cells growing as xenografts in nude mice. Moreover, by using two spectrally resolved reporters (o-fluoro-p-nitrophenyl-beta-D-galactopyranoside and an isomer), two tumors could be interrogated simultaneously revealing lacZ transgene activity in a stably transfected tumor versus no activity in a wild-type tumor. Most significantly, hydrolytic activity observed by 19F NMR corresponded to differential activity in lacZ-expressing tumors.

Improved synthesis of histone deacetylase inhibitors (HDIs) (MS-275 and CI-994) and inhibitory effects of HDIs alone or in combination with RAMBAs or retinoids on growth of human LNCaP prostate cancer cells and tumor xenografts

We have developed new, simple, and efficient procedures for the synthesis of two promising histone deacetylase inhibitors (HDIs), CI-994, (N-(2-aminophenyl)-4-acetylaminobenzamide), and MS-275 (N-(2-aminophenyl)4-[N-(pyridine-3-yl-methoxycarbonyl)aminomethyl]benzamide) from commercially available acetamidobenzoic acid and 3-(hydroxymethyl)pyridine, respectively. The procedures provide CI-994 and MS-275 in 80% and 72% overall yields, respectively. We found that the combination of four HDIs (CI-994, MS-275, SAHA, and TSA) with retinoids all-trans-retinoic acid (ATRA) or 13-cis-retinoic acid (13-CRA) or our atypical retinoic acid metabolism blocking agents (RAMBAs) 1 (VN/14-1) or 2 (VN/66-1) produced synergistic anti-neoplastic activity on human LNCaP prostate cancer cells. The combination of 2 and SAHA induced G1 and G2/M cell cycle arrest and a decrease in the S phase in LNCaP cells. 2+SAHA treatment effectively down-regulated cyclin D1 and cdk4, and up-regulated pro-differentiation markers cytokeratins 8/18 and pro-apoptotic Bad and Bax. Following subcutaneous administration, 2, SAHA or 2+SAHA were well tolerated and caused significant suppression/regression of tumor growth compared with control. These results demonstrate that compound 2 and its combination with SAHA are potentially useful agents that warrant further preclinical development for treatment of prostate cancer.

44. Phase I Study of Ad5-TRAIL in Men with Clinically Organ Confined Prostate Cancer

AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA 4870 We have developed a recombinant adenovirus that encodes human TNF-related apoptosis-inducing ligand (TRAIL; Ad5-TRAIL), which is a potent inducer of tumor cell apoptosis but is not cytotoxic to normal cells. In vitro infection of human prostate tumor cells with Ad5-TRAIL resulted in the rapid transcription and translation of the transferred TRAIL gene into functional TRAIL protein that, when expressed on the cell surface, induced apoptotic death in TRAIL-sensitive prostate tumor cell targets but not normal prostate epithelial cells. Furthermore, in vivo preclinical studies have revealed Ad5-TRAIL can inhibit or slow the outgrowth of human prostate tumor xenografts in SCID mice. Thus, the primary objective of the study is to determine the local and systemic toxicity associated with intraprostatic injection of Ad5-TRAIL delivered in a collagen matrix in men with locally confined prostate cancer, and determine the maximum tolerated dose from four different dose levels. Secondary study objectives will examine the distribution of Ad5-TRAIL within the human prostate after transrectal injection, to determine the effect of transrectal injection of Ad5-TRAIL on existent prostate cancer, and to determine the effect of Ad5-TRAIL-induced apoptosis on immune activation to prostate antigens. Prostate cancer patients (T1c, T2a, T2b and N0, M0) scheduled to undergo radical prostatectomy (RRP) within 10 d following study entry were chosen. The patients had 4 intraprostatic injections of Ad5-TRAIL, 2 per prostate lobe. One week later, endorectal MRI was performed prior to RRP to rule out abscess. Laboratory assessment was performed to assess systemic infection. RRP was then performed if there was no adverse reaction. Operation difficulty was compared to non-injected control patients. Standard histologic evaluation and Gleason grading was performed. Evaluation of the inflammatory infiltrates in Ad5-TRAIL-treated prostates was assessed and compared to control prostate biopsies. To date (11/06), three patients have received the initial vector concentration. There were no immediate or delayed allergic reactions. None of the patients showed abscess on MRI. Operatively, there was no indication of increased inflammation (p=.495), difficulty of the operation, length of stay (p=0.724), or total estimated blood loss. There was no significant increased morbidity post-operatively. All patients had negative margins. Pathologically, each patient had an increase in staging from clinically, and viral vector uptake was shown through visualization of the collagen matrix in the specimen. Ad-5 TRAIL in vivo is known to induce apoptosis in prostate cancer. The initial dose level shows intraprostatic injection is well-tolerated and remains in the prostate after injection. Further increases in dosing will reveal the maximum tolerated dose, and likelihood of increased apoptosis in prostatectomy specimens.

Highly efficient cationic hydroxyethylated cholesterol-based nanoparticle-mediated gene transfer in vivo and in vitro in prostate carcinoma PC-3 cells

Optimal gene therapy for tumors must deliver DNA to tumor cells with high efficiency and minimal toxicity. It has been reported that in non-viral gene delivery, the hydroxyethyl group at the amino terminal in cationic lipid was important for high transfection efficiency. Therefore, in this study, we developed new cationic nanoparticles ( NP-OH) composed of cholesteryl-3β-carboxyamidoethylene- N-hydroxyethylamine and Tween 80, and optimized in vitro and in vivo transfections for potential use as a non-viral DNA vector into human prostate tumor PC-3 cells and xenografts. In vitro transfection resulted in efficient DNA transfer when positive-charged nanoplex was prepared in the presence of sodium chloride (NaCl). In in vivo transfection, negative-charged nanoplex formed in water strongly induced the gene expression compared with positive-charged nanoplex when directly transfected into xenografts. These transfection efficiencies in vitro and in vivo were comparable to each commercial product. Furthermore, NP-OH nanoplexes displayed no induction of tumor necrosis factor (TNF)-α when administered by intravenous injection. The results of the experiments provided optimal conditions to form NP-OH nanoplex for gene delivery in vitro and in vivo. NP-OH is a potential non-viral DNA vector for the local treatment of tumor and in vitro.

Modulation of liver X receptor signaling as novel therapy for prostate cancer

Liver X receptors (LXRs) are important regulators of cholesterol, fatty acid, and glucose homeostasis. LXR agonists are effective for treatment of murine models of atherosclerosis, diabetes, and Alzheimer's disease. Recently we observed that LXR agonists suppressed proliferation of prostate and breast cancer cells in vitro and treatment of mice with the LXR agonist T0901317 suppressed the growth of prostate tumor xenografts. LXR agonists appear to cause G1 cell cycle arrest in cells by reducing expression of Skp2 and inducing the accumulation of p27(Kip). T0901317 induced expression of ATP-binding cassette transporter A1 (ABCA1) and delayed the progression of androgen-dependent human prostate tumor xenografts towards androgen-independency in mice. Phytosterols, the plant equivalent of mammalian cholesterol, have recently been shown to be agonists for LXRs. beta-Sitosterol and campesterol, the two most common phytosterols, suppressed proliferation of prostate and breast cancer cells. The anticancer activity of phytosterols may be due to LXR signaling. This review examines the potential use of LXR signaling as a therapeutic target in prostate and other cancers.

Knock-down of Bcl-2 by antisense oligodeoxynucleotides induces radiosensitization and inhibition of angiogenesis in human PC-3 prostate tumor xenografts

Expression of the proto-oncogene Bcl-2 is associated with tumor progression. Bcl-2's broad expression in tumors, coupled with its role in resistance to chemotherapy and radiation therapy-induced apoptosis, makes it a rational target for anticancer therapy. Antisense Bcl-2 oligodeoxynucleotide (ODN) reagents have been shown to be effective in reducing Bcl-2 expression in a number of systems. We investigated whether treating human prostate cancer cells with antisense Bcl-2 ODN (G3139, oblimersen sodium, Genasense) before irradiation would render them more susceptible to radiation effects. Two prostate cancer cell lines expressing Bcl-2 at different levels (PC-3-Bcl-2 and PC-3-Neo) were subjected to antisense Bcl-2 ODN, reverse control (CTL), or mock treatment. Antisense Bcl-2 ODN alone produced no cytotoxic effects and was associated with G(1) cell cycle arrest. The combination of antisense Bcl-2 ODN with irradiation sensitized both cell lines to the killing effects of radiation. Both PC-3-Bcl-2 and PC-3-Neo xenografts in mice treated with the combination of antisense Bcl-2 ODN and irradiation were more than three times smaller by volume compared with xenografts in mice treated with reverse CTL alone, antisense Bcl-2 ODN alone, irradiation alone, or reverse CTL plus radiotherapy (P = 0.0001). Specifically, PC-3-Bcl-2 xenograft tumors treated with antisense Bcl-2 ODN and irradiation had increased rates of apoptosis and decreased rates of angiogenesis and proliferation. PC-3-Neo xenograft tumors had decreased proliferation only. This is the first study which shows that therapy directed at Bcl-2 affects tumor vasculature. Together, these findings warrant further study of this novel combination of Bcl-2 reduction and radiation therapy, as well as Bcl-2 reduction and angiogenic therapy.

Prostate cancer chemoprevention by silibinin: Bench to bedside

Prostate cancer (PCA) is the most invasive malignancy and second leading cause of cancer deaths in American males. One approach to reduce PCA incidence, growth and metastasis is prevention and intervention targeted towards mitogenic and survival signaling and cell-cycle regulation. This approach is based on the rationale that overexpression of receptor tyrosine kinases (RTKs) and/or non-receptor tyrosine kinases leads to persistent autocrine stimulation of malignant cells for deregulated cell-cycle progression and uncontrolled growth. PCA progression has also been associated with transition from a paracrine to an autocrine relationship between receptors and growth ligands as this malignancy progresses to an advanced androgen-independent aggressive stage. Together, these studies suggest that targeting RTK-mediated signaling pathways along with cell-cycle regulators could be a practical and translational approach for PCA prevention and intervention. Here, we provide evidence that a naturally occurring nontoxic flavanoid, silibinin, targets the epidermal growth factor receptor (EGFR), insulin-like growth factor-1 receptor (IGF-1R) and NF-kappaB (nuclear factor-kappa B) pathways in PCA. Furthermore, it modulates cell-cycle regulators, including cyclin-dependent kinases (CDKs), Cip/Kip and cyclins for its anticancer efficacy against PCA. Silibinin inhibits growth of PCA cells from human, mouse, and rat origins, and also suppresses human prostate tumor xenograft growth in nude mice. Silibinin also inhibits PCA growth in the transgenic adenocarcinoma of mouse prostate (TRAMP) mouse model. Now, silibinin has been entered into phase I/II clinical trials in human PCA patients where preliminary observations were suggestive of its further study in a larger base of the patient population.

Effective Growth Arrest of Human Colon Cancer in Mice, Using Rat Sodium Iodide Symporter and Radioiodine Therapy

We have demonstrated that the rat sodium iodide symporter (rNIS) and 131I can effectively induce growth arrest of human prostate tumor xenografts [Mitrofanova, E., Unfer, R., Vahanian, N., Daniels, W., Roberson, E., Seregina, T., Seth, P., and Link, C. (2004). Rat sodium iodide symporter (rNIS) for radioiodide therapy of cancer. Clin. Cancer Res. 10, 6969-6976]. In that study the average size of tumors established in athymic nude mice was 200 +/- 50 mm3 when treated. Testing under more rigorous and extreme in vitro conditions will better evaluate the ability of an anticancer approach to induce tumor regression or killing capacity in preclinical studies. In this work the ability of the rNIS and 131I system to inhibit the growth of relatively large (about 800 mm3 when treated with 131I) and rapidly growing colon tumors in an animal model was examined. in vitro experiments demonstrated that transduction of human colon cancer cells with Ad-rNIS resulted in a 100- to 150-fold increase in 125I uptake compared with nontransduced cells. Western blot analysis revealed robust expression of rNIS protein in cells 72-120 hr posttransduction with Ad-rNIS. Immunocytochemical analysis demonstrated that intracellular localization of rNIS-specific staining was observed mainly in plasma membranes of cells. in vitro studies revealed an immediate inhibition of growth of rapidly expanding tumors after radioiodine injection in the rNIS and 131I treatment group of mice. Twenty-seven percent of experimental mice survived more than 30 days (p = 0.019), whereas control groups had only 7% survival over 30 days. This is the first report demonstrating that rat NIS and 131I can effectively induce growth arrest of relatively large tumors in an animal model.

FTY720, a fungus metabolite, inhibits in vivo growth of androgen‐independent prostate cancer

FTY720, a derivative of fungus, has demonstrated dramatic anticancer effect in several malignancies recently. Our study evaluates the therapeutic potential of FTY720 in the treatment of androgen-independent prostate cancer using a human prostate cancer xenograft in nude mice. CWR22R, an androgen-independent human prostate tumor xenograft was inoculated into castrated nude mice and the animals were administrated with either normal saline or FTY720 (10 mg/kg) through intraperitoneal (i.p.) injection for 20 days. Body weight and tumor volume were recorded every 2 days, and serum prostate specific antigen (PSA) levels were also measured before and after the treatment. The effect of FTY720 on tumor cell proliferation was examined using antibodies against PCNA and Ki-67 by immunohistochemical staining, MTT assay and colony forming assay, whereas apoptotic effect of FTY720 was evaluated by TUNEL assay and immunostaining using antibodies against cleaved caspase 3 and Bcl-2. In addition, the potential inhibitory effect of FTY720 on prostate cancer angiogenesis and metastasis was investigated by immunostaining of CD31, VEGF, E-cadherin and beta-catenin. Our results showed that FTY720 treatment led to suppression of CWR22R tumor growth without causing any detectable side effects in nude mice. The FTY720-induced tumor suppression was correlated with decreased serum PSA level as well as reduced proliferation rate, suppression of angiogenic factors, and restoration of E-cadherin and beta-catenin expression. In addition, the FTY720-treated tumors showed increased apoptosis rate demonstrated by increased TUNEL- and cleaved caspase 3-positive cells, and decreased Bcl-2 expression. Our results suggest a potential novel agent in the suppression of androgen-independent prostate cancer.

Targeting Tomoregulin for Radioimmunotherapy of Prostate Cancer

Radiotherapy is an effective approach for the treatment of local prostate cancer. However, once prostate cancer metastasizes, radiotherapy cannot be used due to the distribution of multiple metastases to lymph nodes and bones. In contrast, radioimmunotherapy should still be efficacious in metastatic prostate cancer as radioisotopes are brought to tumor cells by targeting antibodies. Here we identify and validate a prostate-expressed molecule, tomoregulin, as a target for radioimmunotherapy of prostate cancer. Tomoregulin is a transmembrane protein selectively expressed in the brain, prostate, and prostate cancer, but not expressed in other normal tissues. Immunohistochemical studies of tomoregulin protein in clinical samples show its location in the luminal epithelium of normal prostate, benign prostatic hyperplasia, and prostatic intraepithelial neoplasia. More importantly, the tomoregulin protein is expressed in primary prostate tumors and in their lymph node and bone metastases. The nature of tomoregulin as a transmembrane protein and its tissue-specific expression make tomoregulin an attractive target for radioimmunotherapy, in which tomoregulin-specific antibodies will deliver a radioisotope to prostate tumor cells and metastases. Indeed, biodistribution studies using a prostate tumor xenograft model showed that the (111)In-labeled anti-tomoregulin antibody 2H8 specifically recognizes tomoregulin protein in vivo, leading to a strong tumor-specific accumulation of the antibody. In efficacy studies, a single i.p. dose of 150 microCi (163 microg) (90)Y-labeled 2H8 substantially inhibits the growth rate of established LNCaP human prostate tumor xenograft in nude mice but produces no overt toxicity despite cross-reactivity of 2H8 with mouse tomoregulin. Our data clearly validate tomoregulin as a target for radioimmunotherapy of prostate cancer.

Novel C-17-Heteroaryl Steroidal CYP17 Inhibitors/Antiandrogens: Synthesis, in Vitro Biological Activity, Pharmacokinetics, and Antitumor Activity in the LAPC4 Human Prostate Cancer Xenograft Model

New chemical entities, steroidal C-17 benzoazoles (5, 6, 9 and 10) and pyrazines (14 and 15) were rationally designed and synthesized. The key reaction for synthesis of the benzoazoles involved the nucleophilic vinylic "addition-elimination" substitution reaction of 3beta-acetoxy-17-chloro-16-formylandrosta-5,16-diene (2) and benzoazole nucleophiles, while that for synthesis of pyrazines involved palladium-catalyzed cross-coupling reaction of 17-iodoandrosta-5,16-dien-3beta-ol (13) with tributylstannyl diazines. Some of the compounds were shown to be potent inhibitors of human CYP17 enzyme as well as potent antagonist of both wild type and mutant androgen receptors (AR). The most potent CYP17 inhibitors were 3beta-hydroxy-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (5, code named VN/124-1), 3beta-hydroxy-17-(5(1)-pyrimidyl)androsta-5,16-diene (15) and 17-(1H-benzimidazole-1-yl)androsta-4,16-dien-3-one (6), with IC(50) values of 300, 500 and 915 nM, respectively. Compounds 5, 6, 14 and 15 were effective at preventing binding of (3)H-R1881 (methyltrienolone, a stable synthetic androgen) to both the mutant LNCaP AR and the wild-type AR, but with a 2.2- to 5-fold higher binding efficiency to the latter. Compounds 5 and 6 were also shown to be potent pure AR antagonists. The cell growth studies showed that 5 and 6 inhibit the growth of DHT-stimulated LNCaP and LAPC4 prostate cancer cells with IC(50) values in the low micromolar range (i.e., <10 microM). Their inhibitory potencies were comparable to that of casodex but remarkably superior to that of flutamide. The pharmacokinetics of compounds 5 and 6 in mice were investigated. Following s.c. administration of 50 mg/kg of 5 and 6, peak plasma levels of 16.82 and 5.15 ng/mL, respectively, occurred after 30 to 60 min, both compounds were cleared rapidly from plasma (terminal half-lives of 44.17 and 39.93 min, respectively), and neither was detectable at 8 h. Remarkably, compound 5 was rapidly converted into a metabolite tentatively identified as 17-(1H-benzimidazol-1-yl)androsta-3-one. When tested in vivo, 5 proved to be very effective at inhibiting the growth of androgen-dependent LAPC4 human prostate tumor xenograft, while 6 was ineffective. Compound 5 (50 mg/kg/twice daily) resulted in a 93.8% reduction (P = 0.00065) in the mean final tumor volume compared with controls, and it was also significantly more effective than castration. To our knowledge, this is the first example of an antihormonal agent (an inhibitor of androgen synthesis (CYP17 inhibitor)/antiandrogen) that is significantly more effective than castration in suppression of androgen-dependent prostate tumor growth. In view of these impressive anticancer properties, compound 5 is a strong candidate for development for the treatment of human prostate cancer.

Adenovirus-mediated FLT1-targeted proapoptotic gene therapy of human prostate cancer

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/Apo2L) is of particular interest in the development of prostate carcinoma therapeutics as it preferentially induces apoptosis of tumor cells. To employ adenoviral vectors for highly efficient and specific TRAIL gene transfer into cancer cells could overcome some potential problems for recombinant TRAIL. The vascular endothelial growth factor receptor FLT-1 is involved in regulation of angiogenesis and tumor growth, invasion, and metastasis of prostate carcinoma. FLT-1 expression is observed in both tumor endothelial cells and prostate cancer cells. We developed an adenoviral vector encoding the TRAIL gene under control of the FLT1 promoter (AdFlt-TRAIL), which produced endothelial and prostate cancer cell death. The combination of ionizing radiation and adenovirus-driven TRAIL expression overcame human prostate cancer cell resistance to TRAIL. Furthermore, in vivo administration of AdFlt-TRAIL at the site of tumor growth in combination with radiation treatment produced significant suppression of the growth of DU145 human prostate tumor xenografts in athymic nude mice. Our results suggest that specific TRAIL delivery employing the FLT1 promoter can effectively inhibit tumor growth and demonstrate the advantage of combination radiotherapy and gene therapy for the treatment of prostate cancer.