Citation Details

Journal Article

PC-3M-luc-C6, PC-3M-luc, IVIS, Bioware, Prostate cancer, Bioluminescence, Animals; Cell Line, Tumor; Endocytosis; Fluorescent Dyes/chemistry/*diagnostic use; Glucosamine/chemistry/*diagnostic use; Humans; Kinetics; Male; Mice; Mice, Nude; Molecular Imaging/*methods; Neoplasms/*diagnosis/pathology; Optical Devices; Prostatic Neoplasms/diagnosis/pathology; Spectroscopy, Near-Infrared; Tissue Distribution; *Xenograft Model Antitumor Assays

PURPOSE: Near-infrared fluorescence (NIRF) imaging is an attractive technique for studying diseases at the molecular level in vivo. Glucose transporters are often used as targets for in vivo imaging of tumors. The efficiency of a tumor-seeking fluorescent probe can be enhanced by attaching one or more glucosamine (GlcN) moieties. This study was designed to evaluate the use of previously developed GlcN-linked NIRF probes for in vitro and in vivo optical imaging of cancer. PROCEDURES: Cellular uptake of the probes (1 muM) was investigated in monolayer cultures of luciferase-expressing PC3 (PC3-luc) cells. The prostate tumors were established as subcutaneous xenografts using PC3-luc cells in nude mice. The biodistributions and tumor-targeting specificities of cypate (cyp), cypate-D: -(+)-glucosamine (cyp-GlcN), and D: -(+)-gluosamine-cypate-D: -(+)-gluosamine (cyp-2GlcN) were studied. The tumor, muscle, and major organs were collected for ex vivo optical imaging. RESULTS: The tumor cell uptake of the probe containing two glucosamine residues, cyp-2GlcN, was significantly higher than the uptake of both the probe with one glucosamine residue, cyp-GlcN, and the probe without glucosamine, cyp only. Similarly, in in vivo experiments, cyp-2GlcN demonstrated higher maximum fluorescence intensity and longer residence lifetime in tumors than cyp-GlcN or cyp. The ex vivo biodistribution analysis revealed that tumor uptake of cyp-2GlcN and cyp-GlcN was four- and twofold higher than that of cyp at 24 h post-injection, respectively. CONCLUSION: Both cyp-GlcN and cyp-2GlcN NIRF probes exhibited good tumor-targeting properties in prostate cancer cell cultures and live mice. The cyp-2GlcN probe showed the highest uptake with good retention characteristics in vivo. The uptake of cyp-2GlcN and cyp-GlcN is likely mediated by glucosamine-recognizing transporters. The uptake mechanism is being explored further for developing cypate-glucosamine-based probes for in vivo imaging.

Journal Article

PC-3M-luc-C6, PC-3M-luc, IVIS, Bioware, Prostate cancer, Bioluminescence, Animals; Chromatography, Liquid; Enzyme-Linked Immunosorbent Assay; Ephrins/genetics/*metabolism; Humans; Mass Spectrometry/*methods; Mice; Receptor, EphA1/genetics/*metabolism

The Eph receptor tyrosine kinase family includes many members, which are often expressed together in various combinations and can promiscuously interact with multiple ephrin ligands, generating intricate networks of intracellular signals that control physiological and pathological processes. Knowing the entire repertoire of Eph receptors and ephrins expressed in a biological sample is important when studying their biological roles. Moreover, given the correlation between Eph receptor/ephrin expression and cancer pathogenesis, their expression patterns could serve important diagnostic and prognostic purposes. However, profiling Eph receptor and ephrin expression has been challenging. Here we describe a novel and straightforward approach to catalog the Eph receptors present in cultured cells and tissues. By measuring the binding of ephrin Fc fusion proteins to Eph receptors in ELISA and pull-down assays, we determined that a mixture of four ephrins is suitable for isolating both EphA and EphB receptors in a single pull-down. We then used mass spectrometry to identify the Eph receptors present in the pull-downs and estimate their relative levels. This approach was validated in cultured human cancer cell lines, human tumor xenograft tissue grown in mice, and mouse brain tissue. The new mass spectrometry approach we have developed represents a useful tool for the identification of the spectrum of Eph receptors present in a biological sample and could also be extended to profiling ephrin expression.

Journal Article

PC-3M-luc-C6, PC-3M-luc, IVIS, Bioware, Prostate cancer, Bioluminescence

Prostate cancer remains the second leading cause of cancer deaths among American men. Early diagnosis increases survival rate in patients; however, treatments for advanced disease are limited to hormone ablation techniques and palliative care. Thus, new methods of treatment are necessary for inhibiting prostate cancer disease progression. Here, we have shown that miRNA-29b (miR-29b) expression was lower in prostate cancer cells (PC3 and LNCaP) as compared with immortalized prostate epithelial cells. Between these two prostate cancer cell lines, metastatic prostate cancer PC3 cells displayed lower expression of miR-29b. We also observed a significant downregulation of miR-29b expression in human prostate cancer tissues as compared with patient-matched nontumor tissues. PC3 cells ectopically expressing miR-29b inhibited wound healing, invasiveness, and failed to colonize in the lungs and liver of severe combined immunodeficient mice after intravenous injection, while PC3 cells expressing a control miRNA displayed metastasis. Epithelial cell marker E-cadherin expression was enhanced miR-29b transfected in prostate cancer cells as compared with cells expressing control miRNA. On the other hand, N-cadherin, Twist, and Snail expression was downregulated in PC3 cells expressing miR-29b. Together these results suggested that miR-29b acts as an antimetastatic miRNA for prostate cancer cells at multiple steps in a metastatic cascade. Therefore, miR-29b could be a potentially new attractive target for therapeutic intervention in prostate cancer.

Journal Article

PC-3M-luc-C6, PC-3M-luc, IVIS, Bioware, Prostate cancer, Bioluminescence

Frequent side effects of radical treatment modalities and the availability of novel diagnostics have raised the interest in focal therapies for localized prostate cancer. To improve the selectivity and therapeutic efficacy of such therapies, we developed a minimally invasive procedure, based on a novel polymeric photosensitizer prodrug sensitive to urokinase-like plasminogen activator (uPA). The compound is inactive in its prodrug form and accumulates passively at the tumor site by the enhanced permeability and retention effect. There, the prodrug is selectively converted to its photoactive form by uPA which is over-expressed by prostate cancer cells. Irradiation of the activated photosensitizer exerts a tumor-selective phototoxic effect. The prodrug alone (8 microM) showed no toxic effect on PC-3 cells, but upon irradiation the cell viability was reduced by 90%. In vivo, after systemic administration of the prodrug, PC-3 xenografts became selectively fluorescent. This is indicative of the prodrug accumulation in the tumor and selective local enzymatic activation. Qualitative analysis of the activated compound confirmed that the enzymatic cleavage occurred selectively in the tumor, with only trace amounts in the neighboring skin or muscle. Subsequent photodynamic therapy studies demonstrated complete tumor eradication of animals treated with light (150 J/cm2 at 665 nm) 16 hours after the injection of the prodrug (7.5 mg/kg). These promising results evidence the excellent selectivity of our prodrug with the potential to be used for both, imaging and therapy of localized prostate cancer.

Journal Article

LL/2-luc-M38, LL/2-luc, Lewis Lung Carcinoma, IVIS, Adenocarcinoma/drug therapy/genetics/*immunology/metabolism/pathology; Administration, Intranasal; Adult; Aged; Animals; Antibodies, Neutralizing/administration & dosage/*therapeutic use; Antigens, CD/immunology; Female; Forkhead Transcription Factors/genetics/*immunology/metabolism; Gene Expression Regulation, Neoplastic/drug effects/*immunology; Humans; Immunologic Surveillance; Interferon-gamma/biosynthesis/immunology; Interleukin-17/immunology/metabolism; Interleukin-23/immunology/metabolism; Lung/drug effects/*immunology/metabolism/pathology; Lung Neoplasms/drug therapy/genetics/*immunology/metabolism/pathology; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Middle Aged; T-Box Domain Proteins/deficiency/*genetics/immunology; T-Lymphocytes, Regulatory/immunology

Lung cancer is the leading cause of cancer deaths worldwide. The cytokine interleukin-17A supports tumour vascularization and growth, however, its role in lung cancer is unknown. Here we show, in the lungs of patients with lung adenocarcinoma, an increase in interleukin-17A that is inversely correlated with the expression of T-bet and correlated with the T regulatory cell transcription factor Foxp3. Local targeting of interleukin-17A in experimental lung adenocarcinoma results in a reduction in tumour load, local expansion of interferon-gamma-producing CD4(+) T cells and a reduction in lung CD4(+)CD25(+)Foxp3(+) regulatory T cells. T-bet((-/-)) mice have a significantly higher tumour load compared with wild-type mice. This is associated with the local upregulation of interleukin-23 and induction of interleukin-17A/interleukin-17R-expressing T cells infiltrating the tumour. Local anti-interleukin-17A antibody treatment partially improves the survival of T-bet((-/-)) mice. These results suggest that local anti-interleukin-17A antibody therapy could be considered for the treatment of lung tumours.