ABSTRACT
BACKGROUND: Tobacco control is important for cancer patient health, but delivering effective low-dose CT (LDCT) screening and tobacco cessation is more difficult in underserved and patients from racial and ethnic minority groups. At City of Hope (COH), we have developed strategies to overcome barriers to the delivery of LDCT and tobacco cessation. METHODS: We performed a needs assessment. New tobacco control program services were implemented focusing on patients from racial and ethnic minority groups. Innovations included Whole Person Care with motivational counseling, placing clinician and nurse champions at points of care, training module and leadership newsletters, and a patient-centric personalized medicine Personalized Pathways to Success (PPS) program. RESULTS: Emphasis on patients from racial and ethnic minority groups was implemented by training cessation personnel and lung cancer control champions. LDCT increased. Tobacco use assessment increased and abstinence was 27.2%. The PPS pilot program achieved 47% engagement in cessation, with self-reported abstinence at 3 months of 38%, with both results slightly higher in patients from racial and ethnic minority groups than in Caucasian patients. CONCLUSIONS: Tobacco cessation barrier-focused innovations can result in increased lung cancer screening and tobacco cessation reach and effectiveness, especially among patients from racial and ethnic minority groups. The PPS program is promising as a personalized medicine patient-centric approach to cessation and lung cancer screening.
ABSTRACT
An iodine-mediated hydration reaction of alkynes serves as a green alternative to metal-catalyzed procedures. Previous work has shown that this method works well with terminal alkynes on keto-functionalized scaffolds, including 1,3-dicarbonyls and their heteroatom analogues. It was hypothesized that the reaction proceeds through a 5-exo-dig neighboring group participation (NGP) cyclization and an α-iodo intermediate. The work described herein probes the existence of the intermediate through NMR investigations and explores the scope of the hydration process with internal alkynes. The NMR experiments confirm the existence of the α-iodo intermediate, and methodology studies demonstrate that alkyl-capped, asymmetric, internal alkynes undergo a regiospecific hydration, also via the 5-exo-dig NGP pathway.
ABSTRACT
mda-7/IL-24 (HGMW-approved symbol IL24) is a tumor suppressor gene whose expression is lost during tumor progression. Gene transfer using adenoviral mda-7/IL-24 (Ad-mda7) exhibits minimal toxicity on normal cells while inducing potent apoptosis in a variety of cancer cell lines. Ad-mda7-transduced cells express high levels of MDA-7 protein intracellularly and also secrete a soluble form of MDA-7 protein. In this study, we sought to determine whether the intracellular or secreted MDA-7 protein was responsible for anti-tumor activity in H1299 lung tumor cells. Ad-mda7 transduction of lung tumor cells increased expression of stress-related proteins, including BiP, GADD34, PP2A, caspases 7 and 12, and XBP-1, consistent with activation of the UPR pathway, a key sensor of endoplasmic reticulum (ER)-mediated stress. Blocking secretion of MDA-7 did not inhibit apoptosis, demonstrating that intracellular MDA-7 was responsible for cytotoxicity. Consistent with this result, when applied directly to lung cancer cells, soluble MDA-7 protein exhibited minimal cytotoxic effect. We then generated mda-7 expression constructs using vectors that target the expressed protein to various subcellular compartments, including cytoplasm, nucleus, and ER. Only full-length and ER-targeted MDA-7 elicited cell death in tumor cells. Thus in lung cancer cells, Ad-mda7 activates the UPR stress pathway and induces apoptosis via intracellular MDA-7 expression in the secretory pathway.
Subject(s)
Carcinoma, Non-Small-Cell Lung/genetics , Cytokines/genetics , Genetic Therapy/methods , Interleukins/genetics , Lung Neoplasms/genetics , Adenoviridae/genetics , Antigens, Differentiation , Apoptosis , Blotting, Western , Brefeldin A/pharmacology , Caspase 12 , Caspase 7 , Caspases/metabolism , Cell Cycle Proteins , Cell Line , Cell Line, Tumor , Cell Separation , Cell Survival , Disease Progression , Dose-Response Relationship, Drug , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum Chaperone BiP , Flow Cytometry , Gene Transfer Techniques , Genes, Tumor Suppressor , Genetic Vectors , Heat-Shock Proteins/metabolism , Humans , Microscopy, Fluorescence , Models, Genetic , Molecular Chaperones/metabolism , Oligonucleotide Array Sequence Analysis , Phosphoprotein Phosphatases/metabolism , Plasmids/metabolism , Protein Folding , Protein Phosphatase 1 , Protein Phosphatase 2 , Proteins/metabolism , Tunicamycin/pharmacologyABSTRACT
The melanoma differentiation-associated gene 7 (mda-7), also called interleukin (IL)-24, suppresses the growth of some cancers in vitro and in vivo as a result of the ectopic expression of its protein. However, the function of the secreted form of the protein in cancer has not been previously studied. The purpose of this study was to determine the antiangiogenic function of a secreted form of the MDA-7/IL-24 protein (sMDA-7/IL-24). In vitro, sMDA-7/IL-24 inhibited both endothelial cell differentiation and migration of endothelial cells induced by vascular endothelial growth factor and basic fibroblast growth factor. The sMDA-7/IL-24-mediated inhibitory effect was 10-50 times more potent than endostatin, IFN-gamma, and IFN-inducible protein 10 in vitro. Furthermore, the inhibitory effect was not mediated by IFN or IFN-inducible protein 10. IL-22 receptor mediated the antiangiogenic activity of sMDA-7/IL-24. Administration of a blocking antibody to IL-22 receptor in conjunction with sMDA-7/IL-24 led to abrogation of inhibition of endothelial differentiation. sMDA-7/IL-24 inhibited vascular endothelial growth factor-induced angiogenesis as evidenced by reduced vascularization and hemoglobin content in in vivo Matrigel plug assays. In vivo, the growth of human lung tumor cells was significantly inhibited, and vascularization was reduced when the cells were mixed with 293 cells stably expressing sMDA-7/IL-24. Systemic administration of sMDA-7/IL-24 inhibited lung tumor growth in a mouse xenograft model. Associated with tumor growth inhibition was decreased tumor microvessel density and hemoglobin content, indicating the presence of antiangiogenic activity. These data demonstrate that sMDA-7/IL-24 is a novel and potent antiangiogenic effector and support the development of MDA-7/IL-24-based therapeutics.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Interleukins/pharmacology , Receptors, Interleukin/physiology , Animals , Cell Differentiation/drug effects , Cell Division/drug effects , Cell Movement/drug effects , Chemokine CXCL10 , Chemokines, CXC/physiology , Collagen/pharmacology , DNA-Binding Proteins/metabolism , Endostatins , Endothelium, Vascular/cytology , Female , Genes, Tumor Suppressor , Humans , Interferon-gamma/physiology , Interleukins/therapeutic use , Mice , Mice, Inbred BALB C , Neoplasm Transplantation , Neoplasms, Experimental/drug therapy , Peptide Fragments/pharmacology , STAT3 Transcription Factor , Trans-Activators/metabolism , Transplantation, Heterologous , Tumor Cells, CulturedABSTRACT
mda-7 is a novel tumor suppressor with cytokine properties. Adenoviral mda-7 (Ad-mda7) induces apoptosis and cell death selectively in tumor cells. The molecular mechanisms underlying the anti-tumor activity of Ad-mda7 in breast and lung cancer lines were investigated. Microarray analyses implicated both the beta-catenin and the PI3K signaling pathways. Ad-mda7 treatment increased protein expression from tumor suppressor genes, including E-cadherin, APC, GSK-3beta, and PTEN, and decreased expression of proto-oncogenes involved in beta-catenin and PI3K signaling. Ad-mda7 caused a redistribution of cellular beta-catenin from the nucleus to the plasma membrane, resulting in reduced TCF/LEF transcriptional activity, and upregulated the E-cadherin-beta-catenin adhesion complex in a tumor cell-specific manner. Expression of the PI3K pathway members (p85 PI3K, FAK, ILK-1, Akt, and PLC-gamma) was downregulated and expression of the PI3K antagonist PTEN was increased. Consistent with this result, pharmacological inhibition of PI3K by wortmannin did not abrogate killing by Ad-mda7. Killing of breast cancer cells by Ad-mda7 required both MAPK and MEK1/2 signaling pathways, whereas these pathways were not essential for MDA-7-mediated killing in lung cancer cells. Thus, in breast and lung tumor cells MDA-7 protein expression modulates cell-cell adhesion and intracellular signaling via coordinate regulation of the beta-catenin and PI3K pathways.
Subject(s)
Breast Neoplasms/metabolism , Cytoskeletal Proteins/metabolism , Interleukins/metabolism , Lung Neoplasms/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Signal Transduction , Trans-Activators/metabolism , Adenoviridae/genetics , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Cadherins/metabolism , Cell Adhesion , Cell Death , Cell Line, Tumor , Cell Movement , Cytoskeletal Proteins/antagonists & inhibitors , Gene Expression Regulation, Neoplastic , Genes, Tumor Suppressor , Humans , Interleukins/genetics , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Oligonucleotide Array Sequence Analysis , Phosphoinositide-3 Kinase Inhibitors , Protein Binding , Proto-Oncogenes/genetics , Trans-Activators/antagonists & inhibitors , beta CateninABSTRACT
The melanoma differentiation association gene-7 (mda-7) is a novel tumor suppressor gene of which the protein expression decreases to nearly undetectable levels in metastatic melanoma. In contrast, expression of inducible nitric oxide synthase (iNOS) is increased in advanced stages of melanoma, and iNOS expression has been proposed as a potential prognostic marker in this disease. Thus, expression of these molecules in the same tumor appears to exhibit reciprocal characteristics. We hypothesize that the relative ratios of these melanoma progression molecules may define either tumor progression or tumor suppression in human melanoma. The first goal of this study was to determine whether MDA-7 expression in melanoma negatively correlates with iNOS expression. The second goal was to determine whether iNOS expression could be regulated by MDA-7 expression in melanoma cells. Expression of MDA-7 and iNOS proteins were evaluated by immunohistochemistry in a total of 81 tumor samples: 38 primary melanomas and 43 metastatic melanomas. Evaluation of these melanoma patient samples showed that expression of MDA-7 and iNOS exhibits a significant negative correlation (P = 0.03). In vitro studies revealed that iNOS expression in melanoma cell lines is lost in a dose-dependent fashion after treatment with an adenoviral vector encoding the mda-7 gene (Ad-mda7) or with rhMDA-7 protein, demonstrating that MDA-7 down-regulates iNOS expression. Furthermore, we demonstrate that the STAT-3-modulated expression of IFN regulatory factors 1 and 2 is regulated by MDA-7, which may alter iNOS gene expression. These studies demonstrate that expression of the MDA-7 tumor suppressor can negatively regulate iNOS expression in malignant melanoma cell lines.
Subject(s)
Interleukins/genetics , Melanoma/enzymology , Melanoma/genetics , Nitric Oxide Synthase/genetics , Skin Neoplasms/enzymology , Skin Neoplasms/genetics , DNA-Binding Proteins/analysis , DNA-Binding Proteins/genetics , Disease Progression , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Neoplastic , Genes, Tumor Suppressor , Humans , Immunohistochemistry , Interferon Regulatory Factor-1 , Interleukins/analysis , Melanoma/pathology , Neoplasm Metastasis , Nitric Oxide Synthase Type II , Phosphoproteins/analysis , Phosphoproteins/genetics , Skin Neoplasms/pathology , Transcription Factors/analysis , Transcription Factors/genetics , Tumor Cells, CulturedABSTRACT
BACKGROUND: Melanoma is an aggressive tumor with a propensity to rapidly metastasize. The PTEN gene encodes a phosphatase with an unusual dual specificity for proteins and lipids. Mutations of PTEN have been found in various human cancers, including glioblastoma, prostate, breast, lung, and melanoma. Here we investigate in vitro the effects of blocking PI3K signaling using adenoviral-delivered PTEN (Ad-PTEN) in cell lines derived from both early- and late-stage melanoma. MATERIALS AND METHODS: Ad-PTEN transduced melanoma cell lines or normal cells were assayed for cell death, apoptosis, gene expression, invasion and migration, and regulation of angiogenesis. RESULTS: The PTEN locus from RGP and metastatic melanoma cell lines was sequenced; no coding region mutations were found. Adenoviral transfer of PTEN into melanoma cells containing wild-type PTEN alleles led to tumor-specific apoptosis and growth inhibition, with coordinate inhibition of AKT phosphorylation. Ad-PTEN suppressed cell migration by metastatic melanoma cells with concomitant increase in the level of cell surface E-cadherin. Immunohistochemical and confocal analyses localized PTEN to the cytoplasm and demonstrated enrichment at the cell membrane. Ad-PTEN inhibited angiogenesis as demonstrated by the tube formation assay using human vascular endothelial cells. CONCLUSIONS: These studies indicate that Ad-PTEN can inhibit tumor cells via multiple mechanisms and has pro-apoptotic, anti-metastatic, and anti-angiogenic properties. Thus, PI3K blockade via Ad-PTEN may be a promising approach for the treatment of early- and late-stage melanoma, even in tumors that do not harbor PTEN mutations.
Subject(s)
Apoptosis/physiology , Melanoma/metabolism , Phosphoinositide-3 Kinase Inhibitors , Phosphoric Monoester Hydrolases/metabolism , Tumor Suppressor Proteins/metabolism , Adenoviridae , Cell Cycle/physiology , Cell Differentiation/physiology , Endothelium/physiopathology , Gene Transfer Techniques , Genetic Therapy , Genetic Vectors , Humans , Melanoma/pathology , Melanoma/therapy , Neovascularization, Pathologic/physiopathology , PTEN Phosphohydrolase , Phosphatidylinositol 3-Kinases/metabolism , Phosphoric Monoester Hydrolases/genetics , Transgenes , Tumor Suppressor Proteins/geneticsABSTRACT
Adenoviral-mediated overexpression of the melanoma differentiation-associated gene 7 (Ad-mda7) induces apoptosis in a wide range of cancer cells, although themechanism is not well understood. We report that Ad-mda7 induces andactivates the double-stranded RNA-dependent protein kinase (PKR), which leads to phosphorylation of the alpha subunit of eukaryotic translation initiation factor 2 (eIF-2alpha) and the induction of apoptosis in lung cancer cells. Treatment with 2-aminopurine (2-AP), a serine/threonine kinase inhibitor, inhibits PKR activation, eIF2alpha phosphorylation, and apoptosis induction by Ad-mda7. Additionally, PKR null but not wild-type fibroblasts are resistant to Ad-mda7-induced apoptosis. These results suggest that the activation of PKR and its downstream targets may be a critical pathway for Ad-mda7-mediated apoptosis.