carbocyanines and Lung-Neoplasms

Non‑small cell lung cancer (NSCLC) remains an intractable disease, which is primarily due to tumor metastasis and the acquisition of resistance to chemotherapy. Therefore, there is an urgent need for novel therapeutics to overcome these obstacles. It was recently demonstrated that upregulated expression of monoamine oxidase A (MAOA) contributes to the progression of NSCLC. G10, a tumor‑targeting representative conjugate of heptamethine carbocyanine dye and an inhibitor of MAOA, was shown to exert potent cytotoxic effects, comparable to those of doxorubicin, against prostate cancer cell lines, as well as moderate MAOA inhibitory activity. The research described herein aimed to extend our previous study on the antitumor function of G10 in NSCLC in vitro and in vivo, and to elucidate the mechanisms through which G10 exerts its antineoplastic effects. G10 markedly inhibited the proliferation of paclitaxel‑resistant NSCLC cells (H460/PTX) and reduced tumor cell migration and invasion. Gene expression profiling of paclitaxel‑resistant NSCLC cells following treatment with G10 demonstrated that the expression of genes associated with the extracellular matrix was significantly affected, particularly the metastasis‑related genes matrix metallopeptidase (MMP)2, MMP14 and COL6A, which exhibited notably reduced expression. Additionally, the results also demonstrated that MAOA‑related pathways, including AKT and hypoxia‑inducible factor‑1α, were also inhibited by G10 treatment and, subsequently, the downstream molecules of these pathways, such as p21, MMP2 and vascular endothelial growth factor, were also downregulated, highlighting a possible mechanism through which G10 suppresses tumor cell migration, invasion and proliferation. Importantly, in mouse NSCLC xenografts, combined treatment with G10 and paclitaxel resulted in pronounced inhibition of tumor growth. Taken together, the results of the present study highlight the potential of G10 as a novel therapeutic targeting MAOA in paclitaxel‑resistant NSCLC.

Topics: Animals; Antineoplastic Agents; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Drug Repositioning; Drug Resistance, Neoplasm; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Mice; Monoamine Oxidase; Monoamine Oxidase Inhibitors; Paclitaxel; Xenograft Model Antitumor Assays

Abnormally high levels of class I histone deacetylases (HDACs) are associated with triple-negative breast cancer (TNBC) proliferation, malignant transformation, and poor prognosis of patients. Herein, we report a near-infrared imaging probe for TNBC detection via visualizing class I HDACs. Conjugating Cy5.5 to a cyclic depsipeptide inhibitor, we obtained the probe (

Topics: Animals; Biomarkers, Tumor; Carbocyanines; Cell Line, Tumor; Depsipeptides; Epigenesis, Genetic; Female; Fluorometry; Gene Expression Regulation, Neoplastic; Heterografts; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Lung Neoplasms; Mice, Nude; Triple Negative Breast Neoplasms; Vorinostat

In recent years, there has been a progress in the study of glycation reaction which is one the possible reason for multiple metabolic disorders. Glycation is a nonenzymatic reaction between nucleic acids, lipids, and proteins resulting into the formation of early glycation products that may further lead to the accumulation of advanced glycation end products (AGEs). The precipitation of AGEs in various cells, tissues, and organs is one of the factors for the initiation and progression of various metabolic derangements including the cancer. The AGE interaction with its receptor "RAGE" activates the inflammatory pathway; yet, the downregulation of RAGE and its role in these pathways are not clear. We explore the effect of anticancer novel nanoassemblies on AGEs to determine its role in the regulation of the expression of RAGE, NFƙB, TNF-

Topics: A549 Cells; Animals; Benzothiazoles; Blood Glucose; Carbocyanines; Cell Proliferation; Disease Models, Animal; Glycosylation; Humans; Interferon-gamma; Lung Neoplasms; MCF-7 Cells; Mice; Models, Biological; Nanoparticles; NF-kappa B; Oxidative Stress; Rats, Sprague-Dawley; Reactive Oxygen Species; Receptor for Advanced Glycation End Products; Serum Albumin, Human; Spectrophotometry; Trifluridine; Tumor Necrosis Factor-alpha

MHI-148 is a type of heptamethine cyanine dye that can cross the cytoplasmic membrane of lung cancer cells. Here we tested the cytotoxic, in vivo imaging of MHI-148 in lung-cancer nude mice model. Ex vivo imaging was also been measured by testing the major tissue fluorescence intensity. And, the small molecular compound MHI-148 had low cytotoxicity which could be visualized at 1 h post-injection in tumor. From ex vivo fluorescence imaging, the tumor showed the highest uptake of MHI-148 among all the selected organs expect for the time point of 2 h. MHI-148 could be used for effective imaging in lung cancer tissue with good stability and specificity, which suggested that MHI-148 could be an effective tumor clinical imaging agent.

Topics: Animals; Biological Transport; Carbocyanines; Cell Transformation, Neoplastic; Humans; Indoles; Lung Neoplasms; Mice; Mice, Nude; Optical Imaging

Direct and absolute analysis of microRNAs (miRNAs) in complex media (e.g., human serum) is still a big challenge due to the issues with off-analyte absorption, low sensitivity and specificity. In this work, we have fabricated the erythrocyte membrane-biointerfaced spherical nucleic acids (EMSNAs) for miRNA assay, which not only enables tailor-engineered signal amplification but also exhibits anti-interference property. As a consequence, it is possible to achieve a single-step quantification of miRNAs in complex media without the process of enzymatic amplification, which can vastly simplify the detection procedure. Experimental results reveal that the assay permits ultrasensitive quantification of miR-141, with a limit of detection down to 33.9 aM, and show a high selectivity for discriminating miR-200 family members. More importantly, the assay enables robust miRNA analysis in human serum and can accurately differentiate lung cancer patients and prostate cancer patients from healthy donors. Its performance may satisfy the requirements for direct, rapid, sensitive and specific early diagnosis of cancer, signifying its great potential in clinical diagnostics.

Topics: Animals; Carbocyanines; DNA Probes; Erythrocyte Membrane; Fluorescent Dyes; Gold; Humans; Limit of Detection; Lung Neoplasms; Male; Mice, Inbred BALB C; MicroRNAs; Nanostructures; Nucleic Acid Hybridization; Prostatic Neoplasms; Spectrometry, Fluorescence

Population-based screening of stage 0-I lung cancer is crucial to save lives. In this article, we describe the development of a method for the detection of a CYFRA 21-1-autoantibody complex and CYFRA 21-1 in plasma samples. The CIC/CYFRA 21-1 ratio allows the detection of stage I-IV lung cancer with 76.0% sensitivity and 87.5% specificity.

Topics: Adult; Aged; Animals; Antibodies, Monoclonal, Murine-Derived; Antigen-Antibody Complex; Antigens, Neoplasm; Autoantibodies; Biomarkers, Tumor; Carbocyanines; DNA; Female; Fluorescent Dyes; Goats; Humans; Immunoassay; Immunoglobulin G; Keratin-19; Limit of Detection; Lung Neoplasms; Male; Mice; Middle Aged; Nucleic Acid Hybridization; Sensitivity and Specificity

Lung cancer is the leading cause of cancer-related deaths worldwide, and approximately 85% are diagnosed as non-small-cell lung cancer (NSCLC). However, efficient detection and diagnosis of NSCLC at early stage is still challenging. In this work, we developed a simple, ultrasensitive and high selective strategy for A549 human NSCLC cells detection based on combining the reorganization property of a novel cyanine dye 3,3'-di(3-sulfopropyl)-4,5,4',5'-dibenzo-9-methyl-thiacarbocyanine triethylammonium salt (cy-M) to aptamer S6 G-quadruplex structure having specific affinity to NSCLC cells, which induced a dramatic fluorescence enhancement (~ 10

Topics: A549 Cells; Aptamers, Nucleotide; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Early Detection of Cancer; Fluorescent Dyes; G-Quadruplexes; Humans; Lung Neoplasms; Neoplastic Cells, Circulating; Pleural Effusion; Sensitivity and Specificity

Because the degree of labeling (DOL) of cell-bound antibodies, often required in quantitative fluorescence measurements, is largely unknown, we investigated the effect of labeling with two different fluorophores (AlexaFluor546, AlexaFluor647) in a systematic way using antibody stock solutions with different DOLs. Here, we show that the mean DOL of the cell-bound antibody fraction is lower than that of the stock using single molecule fluorescence measurements. The effect is so pronounced that the mean DOL levels off at approximately two fluorophores/IgG for some antibodies. We developed a method for comparing the average DOL of antibody stocks to that of the isolated, cell-bound fraction based on fluorescence anisotropy measurements confirming the aforementioned conclusions. We created a model in which individual antibody species with different DOLs, present in an antibody stock solution, were assumed to have distinct affinities and quantum yields. The model calculations confirmed that a calibration curve constructed from the anisotropy of antibody stocks can be used for determining the DOL of the bound fraction. The fluorescence intensity of the cell-bound antibody fractions and of the antibody stocks exhibited distinctly different dependence on the DOL. The behavior of the two dyes was systematically different in this respect. Fitting of the model to these data revealed that labeling with each dye affects quantum yield and antibody affinity differentially. These measurements also implied that fluorophores in multiply labeled antibodies exhibit self-quenching and lead to decreased antibody affinity, conclusions directly confirmed by steady-state intensity measurements and competitive binding assays. Although the fluorescence lifetime of antibodies labeled with multiple fluorophores decreased, the magnitude of this change was not sufficient to account for self-quenching indicating that both dynamic and static quenching processes occur involving H-aggregate formation. Our results reveal multiple effects of fluorophore conjugation, which must not be overlooked in quantitative cell biological measurements.

Topics: Antibodies, Monoclonal; Antibody Affinity; Binding, Competitive; Breast Neoplasms; Carbocyanines; Female; Fluorescence; Fluorescence Polarization; Humans; Lung Neoplasms; Quinolinium Compounds; Receptor, ErbB-2; Spectrometry, Fluorescence; Tumor Cells, Cultured

The outcome of molecular targeted therapies is restricted by the ambiguous molecular subtypes of nonsmall cell lung cancer (NSCLC), which are difficult to be defined with druggable mutations, and the inevitable emergence of drug-resistance. Here we used the Cu-catalyzed click chemistry to synthesize a chitosan-based self-assembled nanotheranostics (CE7Ns) composed of a near-infrared (NIR) fluorescent photosensitizer Cy7 and molecular targeted drug erlotinib. The well-characterized CE7Ns can release erlotinib and Cy7 fast under acidic condition in the presence of lysozyme, distinguish three molecular subtypes of NSCLC, and specifically bind to the erlotinib-sensitive epidermal growth factor receptor (EGFR)-mutated PC-9 cells. The uptake of CE7Ns is much more in PC-9 cells than in other NSCLC cells, thus generating a notable fluorescence signal in PC-9 cells. Upon NIR irradiation, Cy7 in CE7Ns produces high reactive oxygen species in PC-9 cells. The synergistic effect between erlotinib-targeted therapy and photodynamic therapy significantly up-regulates cancer suppressor p53 and inhibits Survivin, which results in more apoptosis and cell cycle arrest. Upon intravenous administration, the erlotinib-guided CE7Ns significantly accumulate in PC-9-seeded mouse lungs and produce strong fluorescence. Upon NIR irradiation, CE7Ns significantly inhibit the subcutaneously implanted PC-9 tumor growth. This study provides, for the first time, a novel strategy to synthesize a multifunctional theranostic entity to simultaneously distinguish and image druggable mutations and combine targeted therapy with photodynamic therapy to overcome drug resistance.

Topics: Administration, Intravenous; Animals; Antineoplastic Combined Chemotherapy Protocols; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Click Chemistry; Copper; Drug Liberation; Drug Resistance, Neoplasm; Drug Synergism; ErbB Receptors; Erlotinib Hydrochloride; Female; Humans; Infrared Rays; Lung Neoplasms; Mice; Mice, Nude; Microscopy, Confocal; Microscopy, Fluorescence; Molecular Targeted Therapy; Mutation; Photochemotherapy; Theranostic Nanomedicine; Xenograft Model Antitumor Assays

Topics: A549 Cells; Animals; Antineoplastic Agents; Benzimidazoles; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Survival; Drug Screening Assays, Antitumor; Humans; Imidazoles; Kaplan-Meier Estimate; Lung Neoplasms; Membrane Potential, Mitochondrial; Mice; Mice, Inbred C57BL; Microscopy, Fluorescence; Mitochondria; Molecular Conformation; Naphthols; Salts; Structure-Activity Relationship; Transplantation, Heterologous

In the United States, lung cancer is the leading cause of cancer death and ranks second in the number of new cases annually among all types of cancers. Better methods or tools for diagnosing and treating this disease are needed to improve patient outcomes. The delta-opioid receptor (δOR) is reported to be overexpressed in lung cancers and not expressed in normal lung. Thus, we decided to develop a lung cancer-specific imaging agent targeting this receptor. We have previously developed a δOR-targeted fluorescent imaging agent based on a synthetic peptide antagonist (Dmt-Tic) conjugated to a Cy5 fluorescent dye. In this work, we describe the synthesis of Dmt-Tic conjugated to a longer wavelength near-infrared fluorescent (NIRF) dye, Li-cor IR800CW. Binding affinity of Dmt-Tic-IR800 for the δOR was studied using lanthanide time-resolved fluorescence (LTRF) competitive binding assays in cells engineered to overexpress the δOR. In addition, we identified lung cancer cell lines with high and low endogenous expression of the δOR. We confirmed protein expression in these cell lines using confocal fluorescence microscopy imaging and used this technique to estimate the cell-surface receptor number in the endogenously expressing lung cancer cell lines. The selectivity of Dmt-Tic-IR800 for imaging of the δOR in vivo was shown using both engineered cell lines and endogenously expressing lung cancer cells in subcutaneous xenograft models in mice. In conclusion, the δOR-specific fluorescent probe developed in this study displays excellent potential for imaging of lung cancer.

Topics: Animals; Binding, Competitive; Carbocyanines; Cell Line, Tumor; Dipeptides; Female; Fluorescent Dyes; Humans; Lung; Lung Neoplasms; Mice; Mice, Nude; Optical Imaging; Receptors, Opioid, delta; Tetrahydroisoquinolines

Despite the increasing lung cancer-associated death rate, its therapy has been constrained by impasse of early diagnosis. To apply non-invasive imaging for potential cancer diagnosis system, we screened human lung adenocarcinoma-specific peptides using the phage display technique. For in vivo phage-displayed peptide screening, M13 phage library displaying 2.9 × 10(9) random peptides was injected through tail vein to lung adenocarcinoma cell-derived xenograft mouse model. Through four rounds of biopanning, a specific peptide sequence (CAKATCPAC) was screened out with the highest frequency and was named as Pep-1, and it was analyzed for its targeting ability as an imaging probe by in vitro competitive assay to test its cell-binding ability, immunohistochemical detection in the tumor tissue, and in vivo NIR fluorescent optical imaging. The specificity of Pep-1 toward lung cancer was ensured by in vivo imaging using xenograft animals of various cancer types. The results suggest that Pep-1 is a promising diagnostic lead molecule for rapid and accurate detection of human lung adenocarcinoma. In addition, it was found that the targeting ability was much enhanced by ionizing radiation in both cell-derived and patient-derived lung adenocarcinoma xenografts, suggesting the possibility of applying Pep-1 for prognostic diagnosis after radiotherapy. Taken together, this study suggests that Pep-1 possesses a specific-targeting ability for human lung adenocarcinoma and that this peptide could be directly used as a clinically applicable imaging probe.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Amino Acid Sequence; Animals; Bacteriophage M13; Carbocyanines; Cell Line, Tumor; Drug Delivery Systems; Early Diagnosis; Fluorescent Dyes; Gamma Rays; Heterografts; Humans; Injections, Subcutaneous; Lung Neoplasms; Male; Mice; Mice, Nude; Molecular Probes; Optical Imaging; Peptide Fragments; Peptide Library

To enhance the diagnosis of non-small cell lung cancer (NSCLC), we prepared a dual-modal probe Cy5.5-Tf-Gd-DTPA. Gd-DTPA and near-infrared (NIR) dyes were conjugated to holo-Transferrin (Tf) sequentially, the result of ICP-AES and UV showed 25 Gd ions and 1 Cy5.5 could be loaded per protein, respectively. The calculated longitudinal relaxivity R1 of Cy5.5-Tf-DTPA-Gd was 4.21 mM-1S-1 per Gd while that of Magnevist (Gd-DTPA) was only 4.02 mM-1S-1. Confocal laser scanning microscopy and immunohistochemical analyses revealed that the Cy5.5-Tf-DTPA-Gd was localized and accumulated in cytoplasmic vesicles; the cell toxicity assay showed no apparent toxicity. MR and NIR imaging of mice with subcutaneous H1299 xenografte tumors following intravenous injection of Cy5.5-Tf-DTPA-Gd revealed a strong positive contrast of the tumors, which caused a longer lasting enhancement of the MRI signal and fluorescence signal. Taken together, these studies indicate that Cy5.5-Tf-DTPA-Gd could be a good agent for MR/NIRF dual mode applications to detect both tumor in situ and its metastasis.

Topics: Animals; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Survival; Fluorescence; Fluorescent Dyes; Gadolinium; Gene Expression Regulation, Neoplastic; Humans; Ions; Lung Neoplasms; Magnetic Resonance Imaging; Magnetics; Male; Mice; Mice, Inbred BALB C; Neoplasm Metastasis; Neoplasm Transplantation; Spectroscopy, Near-Infrared; Transferrin; Ultraviolet Rays

Conventional chemotherapeutics nonselectively kill all rapidly dividing cells, which produces numerous side effects. To address this challenge, we report the discovery of functional polyesters that are capable of delivering siRNA drugs selectively to lung cancer cells and not to normal lung cells. Selective polyplex nanoparticles (NPs) were identified by high-throughput library screening on a unique pair of matched cancer/normal cell lines obtained from a single patient. Selective NPs promoted rapid endocytosis into HCC4017 cancer cells, but were arrested at the membrane of HBEC30-KT normal cells during the initial transfection period. When injected into tumor xenografts in mice, cancer-selective NPs were retained in tumors for over 1 wk, whereas nonselective NPs were cleared within hours. This translated to improved siRNA-mediated cancer cell apoptosis and significant suppression of tumor growth. Selective NPs were also able to mediate gene silencing in xenograft and orthotopic tumors via i.v. injection or aerosol inhalation, respectively. Importantly, this work highlights that different cells respond differentially to the same drug carrier, an important factor that should be considered in the design and evaluation of all NP carriers. Because no targeting ligands are required, these functional polyester NPs provide an exciting alternative approach for selective drug delivery to tumor cells that may improve efficacy and reduce adverse side effects of cancer therapies.

Topics: Animals; Apoptosis; Carbocyanines; Cell Line, Tumor; Cell Proliferation; Combinatorial Chemistry Techniques; Endocytosis; Gene Silencing; Gene Transfer Techniques; Humans; Lung Neoplasms; Mice; Nanoparticles; Polyesters; RNA, Small Interfering; Ubiquitin; Xenograft Model Antitumor Assays

Direct pulmonary delivery of anti-cancer agents is viewed as an effective way of treating lung cancer. Here, we fabricated inhalable nanoparticles made of human serum albumin (HSA) conjugated with doxorubicin and octyl aldehyde and adsorbed with apoptotic TRAIL protein (TRAIL/Dox HSA-NP). The octyl aldehyde and doxorubicin endowed HSA with significant hydrophobicity that facilitated self-assembly. TRAIL/Dox HSA-NP was found to have excellent particle size (~340nm), morphology, dispersability, and aerosolization properties. TRAIL/Dox HSA-NP displayed synergistic cytotoxicity and apoptotic activity in H226 lung cancer cells vs. HSA-NP containing TRAIL or Dox alone. TRAIL/Dox HSA-NP was well deposited in the mouse lungs using an aerosolizer, and TRAIL and Dox-HSA were found to be gradually released over 3days. The anti-tumor efficacy of pulmonary administered TRAIL/Dox HSA-NP was evaluated in BALB/c nu/nu mice bearing H226 cell-induced metastatic tumors. It was found that the tumors of H226-implanted mice treated with TRAIL/Dox HSA-NP were remarkably smaller and lighter than those of mice treated with TRAIL or Dox HSA-NP alone (337.5±7.5; 678.2±51.5; and 598.9±24.8mg, respectively). Importantly, this improved anti-tumor efficacy was found to be due to the synergistic apoptotic effects of Dox and TRAIL. In the authors' opinion, TRAIL/Dox HSA-NP offers a potential inhalable anti-lung cancer drug delivery system. Furthermore, the synergism displayed by combined use of Dox and TRAIL could be used to markedly reduce doxorubicin doses and minimize its side effects.

Topics: Administration, Inhalation; Adsorption; Albumins; Aldehydes; Animals; Antineoplastic Agents; Apoptosis; Carbocyanines; Cell Line, Tumor; Doxorubicin; Drug Carriers; Drug Resistance, Neoplasm; Humans; Lung; Lung Neoplasms; Male; Mice, Inbred BALB C; Mice, Inbred ICR; Nanoparticles; TNF-Related Apoptosis-Inducing Ligand

Cellular viability is usually determined by measuring the capacity of cells to exclude vital dyes such as 4',6-diamidino-2-phenylindole (DAPI), or by assessing nuclear morphology with chromatinophilic plasma membrane-permeant dyes, such as Hoechst 33342. However, a fraction of cells that exclude DAPI or exhibit normal nuclear morphology have already lost mitochondrial functions and/or manifest massive activation of apoptotic caspases, and hence are irremediably committed to death. Here, we developed a protocol for the simultaneous detection of plasma membrane integrity (based on DAPI) or nuclear morphology (based on Hoechst 33342), mitochondrial functions (based on the mitochondrial transmembrane potential probe DiOC6(3)) and caspase activation (based on YO-PRO®-3, which can enter cells exclusively upon the caspase-mediated activation of pannexin 1 channels). This method, which allows for the precise quantification of dead, dying and healthy cells, can be implemented on epifluorescence microscopy or flow cytometry platforms and is compatible with a robotized, high-throughput workflow.

Topics: Amino Acid Chloromethyl Ketones; Apoptosis; Benzimidazoles; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Caspases; Cell Line, Tumor; Cell Membrane Permeability; Cell Survival; Connexins; Drug Synergism; Flow Cytometry; Fluorescent Dyes; High-Throughput Screening Assays; Humans; Indoles; Lung Neoplasms; Membrane Potential, Mitochondrial; Microscopy, Fluorescence; Nerve Tissue Proteins; Organoplatinum Compounds; Oxaliplatin; Robotics; Staining and Labeling; Staurosporine; Workflow

DNA methylation is the most frequently studied epigenetic modification that is strongly involved in genomic stability and cellular plasticity. Aberrant changes in DNA methylation status are ubiquitous in human cancer and the detection of these changes can be informative for cancer diagnosis. Herein, we reported a facile quantum dot-based (QD-based) fluorescence resonance energy transfer (FRET) technique for the detection of DNA methylation. The method relies on methylation-sensitive restriction enzymes for the differential digestion of genomic DNA based on its methylation status. Digested DNA is then subjected to PCR amplification for the incorporation of Alexa Fluor-647 (A647) fluorophores. DNA methylation levels can be detected qualitatively through gel analysis and quantitatively by the signal amplification from QDs to A647 during FRET. Furthermore, the methylation levels of three tumor suppressor genes, PCDHGB6, HOXA9 and RASSF1A, in 20 lung adenocarcinoma and 20 corresponding adjacent nontumorous tissue (NT) samples were measured to verify the feasibility of the QD-based FRET method and a high sensitivity for cancer detection (up to 90%) was achieved. Our QD-based FRET method is a convenient, continuous and high-throughput method, and is expected to be an alternative for detecting DNA methylation as a biomarker for certain human cancers.

Topics: Adenocarcinoma; Carbocyanines; Cell Line, Tumor; DNA Methylation; DNA, Neoplasm; Fluorescence Resonance Energy Transfer; Genes, Tumor Suppressor; Humans; Lung Neoplasms; Quantum Dots

A near-infrared fluorescent probe (Trp-Cy) for endogenous ozone is presented, which exhibited a large stokes shift about 140 nm and a rapid fluorescence response to ozone with high selectivity and sensitivity.

Topics: Animals; Carbocyanines; Cell Line, Tumor; Fluorescent Dyes; Half-Life; Humans; Lung Neoplasms; Macrophages; Magnetic Resonance Spectroscopy; Mice; Microscopy, Confocal; Molecular Imaging; Ozone; Reactive Oxygen Species; Sensitivity and Specificity; Spectroscopy, Near-Infrared; Tetradecanoylphorbol Acetate; Tryptophan

The putative presence of the erythropoietin receptor (EpoR) on human cancer cells has given rise to controversial discussion about the use of recombinant human erythropoietin (rhuEpo) for treatment of patients with chemotherapy-induced anemia. In vivo analysis of the EpoR status in tumors could help in elucidating the role of erythropoietin in cancer. Thus, the aim of this study was to develop a targeted EpoR probe for the investigation of EpoR expression in human lung cancer xenografts by fluorescence-mediated tomography.. Epo-Cy5.5 was generated by coupling Cy5.5 to rhuEpo. In vitro binding assays were performed using the EpoR-positive non-small cell lung cancer (NSCLC) cell lines A549 (lower EpoR expression) and H838 (higher EpoR expression), the EpoR-negative cell line H2030, and EpoR/EGFP-overexpressing HeLa cells. In vivo specificity of Epo-Cy5.5 was confirmed by competition analyses using micro-CT/fluorescence-mediated tomography fusion imaging. Biodistribution was analyzed over 50 h after injection. Binding of Epo-Cy5.5 was validated on tumor cryosections.. After intravenous injection, the probe was rapidly cleared from the circulation. An accumulation was observed in liver and kidneys, with a maximum at 7 h after injection followed by a decline, indicating renal excretion. Almost constant accumulation of Epo-Cy5.5 was found in bone marrow and tumors, indicating specific receptor binding. The probe allowed the discrimination between H838 with higher EpoR expression (89.54 ± 15.91 nM at 25 h) and A549 tumors with lower EpoR expression (60.45 ± 14.59 nM at 25 h, P < 0.05). Tumor accumulation of Epo-Cy5.5 could be significantly reduced by adding unlabeled rhuEpo (P < 0.05 at 4, 7, and 24 h). In vitro validation confirmed specific binding of Epo-Cy5.5 to the tumor cells, and this binding correlated with the EpoR expression level. Binding was also observed on endothelial cells. Vessel density and Epo-Cy5.5 binding on endothelial cells were comparable.. Epo-Cy5.5 allows the longitudinal analysis of EpoR expression in tumors and thereby can investigate the influence of erythropoietin on EpoR expression, tumor growth, and angiogenesis.

Topics: Animals; Bone Marrow; Carbocyanines; Cell Line, Tumor; Cell Transformation, Neoplastic; Endothelial Cells; Erythropoietin; Female; Gene Expression Regulation, Neoplastic; Humans; Infrared Rays; Lung Neoplasms; Mice; Molecular Imaging; Molecular Probes; Receptors, Erythropoietin; Substrate Specificity

Although oestrogen-related receptor alpha (ERRalpha) is primarily thought to regulate energy homeostasis, it also serves as a prognostic marker for cancer. The aim of this study was to investigate any connection between ERRalpha activity and cell population growth.. XCT-790, an ERRa specific inverse agonist, was employed to suppress ERRa activity in human non-small cell lung cancer cells (NSCLC) A549. Gene expressions were detected using quantitative real-time PCR and Western blot analysis. Mitochondrial mass, membrane potential and reactive oxygen species (ROS) production were measured by staining with Mitotracker green, JC-1 and CM-H(2)DCFDA dyes respectively. Rate of progression through the tricarboxylic acid (TCA) cycle was analysed by measuring activities of citrate synthase and succinate dehydrogenase. Cell cycle analysis was performed by using flow cytometry.. We found that XCT-790 treatment reduced mitochondrial mass but enhanced mitochondrial ROS production by increasing rate through the TCA cycle, elevating mitochondrial membrane potential (DeltaPsi(m)) and down-regulating expression of superoxide dismutase. It was further demonstrated that XCT-790-induced ROS modulated p53 and Rb signalling pathways and suppressed cell replication.. ERRalpha affects cell cycle mechanisms through modulating mitochondrial mass and function. Dysregulation of this essential pathway leads to elevation in mitochondrial ROS production, which in turn modulates activities of tumour suppressors, resulting in cell cycle arrest.

Topics: Benzimidazoles; Carbocyanines; Carcinoma, Non-Small-Cell Lung; Cell Cycle; Cell Line, Tumor; Dose-Response Relationship, Drug; ERRalpha Estrogen-Related Receptor; Fluorescent Dyes; Humans; Lung; Lung Neoplasms; Membrane Potential, Mitochondrial; Membrane Potentials; Mitochondria; Nitriles; Reactive Oxygen Species; Receptors, Estrogen; Signal Transduction; Superoxide Dismutase; Thiazoles; Time Factors; Tumor Suppressor Protein p53

Monitoring and targeting protein kinases is widely accepted as a promising approach for disease diagnosis and drug discovery. For this purpose, the authors have developed an original type of peptide array as a high-throughput screening assay for quantitatively evaluating kinase activity. A volume of 2 nL of peptide solution was spotted onto a formyl group-modified glass slide by using an arrayer, which was designed for use with protein chip technology. The phosphorylation was recognized by fluorescence-label antibody and detected with an automatic microarray scanner widely used in DNA chip technology. The system needs low sample volume, provides a high-density peptide array, and supplies high reproducibility. It provided enough sensitivity for inhibitor screening, even though a relatively low concentration of purified kinase was employed. The assay also proved useful for the detection of intracellular kinase activity as well as for the measurement of the fluctuations of intracellular protein kinase activity with drug stimulation. Thus, this peptide array would be applicable for kinase-targeted diagnosis, cell-based drug screening, and signal pathway investigation.

Topics: Animals; Antibodies; Breast Neoplasms; Carbocyanines; Cell Extracts; Cell Line, Tumor; CHO Cells; Cricetinae; Cricetulus; Female; Fluorescent Dyes; Humans; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Peptides; Phosphorylation; Protein Array Analysis; Protein Kinases; Reproducibility of Results; Sensitivity and Specificity

The effect of dye localization and dye distribution on the antineoplastic behavior of photosensitizers was investigated with a homologous series of trimethine thiacarbocyanine dyes in L1210 leukemia and A549 lung carcinoma cells. These dyes were synthesized with N-alkyl groups of different sizes (ethyl to octadecyl) to vary their lipophilic properties without compromising their photophysics. While dyes with smaller N-alkyl groups (ethyl to decyl) were already cytotoxic in the dark, longer chain cyanines exhibited antineoplastic activity only after exposure to light. Results from this study indicate that the switch from dark cytotoxicity to phototoxicity occurred when dyes, due to a decrease in cationic character with increasing size of alkyl substituents, were no longer able to cross the plasma membrane. Dark cytotoxicity decreased with increasing size of N-alkyl groups and was cell-line independent. On the other hand, photodynamic damage varied by several orders of magnitude depending on the cell line and the length of the alkyl substituents. The most effective photosensitizer was the dioctadecyl dye which achieved a 4- to 5-log reduction of leukemia cells, although it had very modest triplet and singlet oxygen quantum yields of 0.008 and 0.006, respectively. This study also showed that photobiological performance can be improved greatly by optimizing dye binding properties via structural modifications.

Topics: Animals; Antineoplastic Agents; Carbocyanines; Fatty Acids, Unsaturated; Fluorescent Dyes; Humans; Leukemia L1210; Lung Neoplasms; Mice; Photochemistry; Structure-Activity Relationship; Tumor Cells, Cultured

The search for improved photosensitizers for laser phototherapy of malignancies has led to the examination of a new group of carbocyanine dyes as effective fluorochromes. In this study, four carbocyanine dyes with different absorption maxima of 483 nm [DiOC6(3)], 545.5 nm (DiIC5(3)], 556.6 nm [DiSC5(3)], and 651.0 nm [DiSC3(5)] were tested in vitro. The kinetics of uptake and toxicity of these four dyes were assessed for P3 human squamous cell carcinoma, HT29 colon carcinoma, M26 melanoma, and TE671 fibrosarcoma cell lines at 15, 30, 45, 60, and 180 minutes after exposure with each dye. After sensitization with DiOC6(3), the P3 and M26 cell lines were also tested for phototherapy by treatment with 488-nm light from an argon laser. The results showed that these four carbocyanine dyes had rapid and significant uptake by the carcinoma cell lines with no toxicity at concentrations < 0.1 micrograms/mL. Nontoxic DiOC6(3) levels in sensitized tumor cells after laser phototherapy resulted in approximately 85% inhibition of P3 and approximately 95% inhibition of M26 cell lines by MTT assays. The results suggest that these carbocyanine dyes can be used for tumor photosensitization and wavelength-matched laser photodynamic therapy. Further in vivo studies will be necessary to define the clinical potential of carbocyanine dyes as tumor-targeting agents for phototherapy of cancer.

Topics: Adenocarcinoma; Argon; Benzothiazoles; Carbocyanines; Carcinoma, Squamous Cell; Cell Survival; Colonic Neoplasms; Fibrosarcoma; Fluorescent Dyes; Humans; Laser Therapy; Lung Neoplasms; Medulloblastoma; Melanoma; Neoplasms; Photochemotherapy; Tetrazolium Salts; Tumor Cells, Cultured

Certain positively charged, lipophilic dyes have been noted by various authors to localize selectively in the mitochondria of carcinoma cells. Oseroff et al. (Proc. Natl. Acad. Sci. USA, 83:9729-9733, 1986) studied 10 carcinoma-specific mitochondrial photosensitizers and judged N,N'-bis(2-ethyl-1,3-dioxolane)kryptocyanine (EDKC) to be the most effective in selective carcinoma cell photolysis, a system where light-absorbing molecules accumulate only in carcinoma cells and on illumination initiate a reaction that kills or damages those cells. The present study duplicated the published EDKC retention result for the normal monkey kidney epithelial cell line CV-1. A series of nontumorigenic and tumorigenic human bronchial epithelial and human pleural mesothelial cells were assayed for EDKC uptake and retention, with the intent of using selective carcinoma cell photolysis to isolate nontumorigenic revertants of the tumorigenic lung cell lines. In addition, the uptake and retention of the fluorescent, mitochondria- and carcinoma-specific dye rhodamine-123 were surveyed in a series of hybrids between tumorigenic and nontumorigenic human bronchial epithelial cells. The half-life of dye retention ranged from 6 to 12 h in all the bronchial epithelial and mesothelial cells studied, with little or no dye selectivity for tumorigenic cells. When EDKC-retaining bronchial epithelial cells were illuminated with red light, significant reductions in short term viability and colony-forming efficiency were seen, which became more pronounced as light and dye doses were increased. However, these effects did not correlate with tumorigenicity within the cell series. The method, therefore, does not appear generally useful for the selection of nontumorigenic variants of human bronchial epithelial or pleural mesothelial cancers of the lung.

Topics: Carbocyanines; Coloring Agents; Humans; Kinetics; Light; Lung Neoplasms; Photolysis; Quinolines; Radiation-Sensitizing Agents; Rubidium; Tumor Cells, Cultured