RESUMEN
To enhance uptake of photosensitizers by epithelial tumor cells by targeting these to EGFR, pyropheophorbide derivatives were synthesized that had erlotinib attached to different positions on the macrocycle. Although the addition of erlotinib reduced cellular uptake, several compounds showed prolonged cellular retention and maintained photodynamic efficacy. The aim of this study was to identify whether erlotinib moiety assists in tumor targeting through interaction with EGFR and whether this interaction inhibits EGFR kinase activity. The activity of the conjugates was analyzed in primary cultures of human head and neck tumor cells with high-level expression of EGFR, and in human carcinomas grown as xenografts in mice. Uptake of erlotinib conjugates did not correlate with cellular expression of EGFR and none of the compounds exerted EGFR-inhibitory activity. One derivative with erlotinib at position 3, PS-10, displayed enhanced tumor cell-specific retention in mitochondria/ER and improved PDT efficacy in a subset of tumor cases. Moreover, upon treatment of the conjugates with therapeutic light, EGFR-inhibitory activity was recovered that attenuated EGFR signal-dependent tumor cell proliferation. This finding suggests that tumor cell-specific deposition of erlotinib-pyropheophorbides, followed by light triggered release of EGFR-inhibitory activity, may improve photodynamic therapy by attenuating tumor growth that is dependent on EGFR-derived signals.
Asunto(s)
Fotoquimioterapia , Fármacos Fotosensibilizantes , Animales , Línea Celular Tumoral , Receptores ErbB/metabolismo , Clorhidrato de Erlotinib/farmacología , Clorhidrato de Erlotinib/uso terapéutico , Humanos , Ratones , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/uso terapéutico , Inhibidores de Proteínas Quinasas/farmacología , Transducción de SeñalRESUMEN
Herein we report the syntheses and comparative photophysical, electrochemical, in vitro, and in vivo biological efficacy of 3-(1'-hexyloxy)ethyl-3-devinylpyropheophorbide-cyanine dye (HPPH-CD) and the corresponding indium (In), gallium (Ga), and palladium (Pd) conjugates. The insertion of a heavy metal in the HPPH moiety makes a significant difference in FRET (Förster resonance energy transfer) and electrochemical properties, which correlates with singlet oxygen production [a key cytotoxic agent for photodynamic therapy (PDT)] and long-term in vivo PDT efficacy. Among the metalated analogs, the In(III) HPPH-CD showed the best cancer imaging and PDT efficacy. Interestingly, in contrast to free base HPPH-CD, which requires a significantly higher therapeutic dose (2.5 µmol/kg) than imaging dose (0.3 µmol/kg), the corresponding In(III) HPPH-CD showed excellent imaging and therapeutic potential at a remarkably low dose (0.3 µmol/kg) in BALB/c mice bearing Colon26 tumors. A comparative study of metalated and corresponding nonmetalated conjugates further confirmed that STAT-3 dimerization can be used as a biomarker for determining the level of photoreaction and tumor response.
Asunto(s)
Metales/química , Neoplasias Experimentales/patología , Fotoquimioterapia , Porfirinas/química , Animales , Transferencia Resonante de Energía de Fluorescencia , Ratones , Espectrofotometría UltravioletaRESUMEN
We have previously shown that the (124)I-analog of methyl 3-(1'-m-iodobenzyloxy) ethyl-3-devinyl-pyropheophorbide-a derived as racemic mixture from chlorophyll-a can be used for PET (positron emission tomography)-imaging in animal tumor models. On the other hand, as a non-radioactive analog, it showed excellent fluorescence and photodynamic therapy (PDT) efficacy. Thus, a single agent in a mixture of radioactive ((124)I-) and non-radioactive ((127)I) material can be used for both dual-imaging and PDT of cancer. Before advancing to Phase I human clinical trials, we evaluated the activity of the individual isomers as well as the impact of a chiral center at position-3(1) in directing in vitro/in vivo cellular uptake, intracellular localization, epithelial tumor cell-specific retention, fluorescence/PET imaging, and photosensitizing ability. The results indicate that both isomers (racemates), either as methyl ester or carboxylic acid, were equally effective. However, the methyl ester analogs, due to subcellular deposition into vesicular structures, were preferentially retained. All derivatives containing carboxylic acid at the position-17(2) were noted to be substrate for the ABCG2 (a member of the ATP binding cassette transporters) protein explaining their low retention in lung tumor cells expressing this transporter. The compounds in which the chirality at position-3 has been substituted by a non-chiral functionality showed reduced cellular uptake, retention and lower PDT efficacy in mice bearing murine Colon26 tumors.
Asunto(s)
Carcinoma de Células Escamosas/radioterapia , Clorofila/análogos & derivados , Neoplasias del Colon/radioterapia , Neoplasias Pulmonares/radioterapia , Fármacos Fotosensibilizantes/farmacología , Animales , Transporte Biológico , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Carcinoma de Células Escamosas/ultraestructura , Línea Celular Tumoral , Clorofila/síntesis química , Clorofila/química , Clorofila/farmacología , Clorofila A , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Neoplasias del Colon/ultraestructura , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Radioisótopos de Yodo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/ultraestructura , Ratones , Ratones Endogámicos BALB C , Imagen Molecular/métodos , Trasplante de Neoplasias , Especificidad de Órganos , Fotoquimioterapia/métodos , Fármacos Fotosensibilizantes/síntesis química , Spirulina/química , Estereoisomerismo , Carga Tumoral/efectos de los fármacosRESUMEN
The latent transcription factor Stat3 is activated by gp130, the common receptor for the interleukin (IL)-6 cytokine family and other growth factor and cytokine receptors. Ligand-induced dimerization of gp130 leads to activation of the Stat1, Stat3 and Shp2-Ras-Erk signaling pathways. Here we assess genetically the contribution of exaggerated Stat3 activation to the phenotype of gp130 (Y757F/Y757F) mice, in which a knock-in mutation disrupts the negative feedback mechanism on gp130-dependent Stat signaling. Compared to gp130 (Y757F/Y757F) mice, reduced Stat3 activation in gp130 (Y757F/Y757F) Stat3(+/-) mice increased their lifespan, prevented splenomegaly, normalized exaggerated hepatic acute-phase response and lymphocyte trafficking, and suppressed the growth of spontaneously arising gastric adenomas in young mice. These lesions share histological features of gastric polyps in aging mice with monoallelic null mutations in Smad4, which encodes the common transducer for transforming growth factor (TGF)-beta signaling. Indeed, hyperactivation of Stat3 desensitizes gp130 (Y757F/Y757F) cells to the cytostatic effect of TGF-beta through transcriptional induction of inhibitory Smad7, thereby providing a novel link for cross-talk between Stat and Smad signaling in gastric homeostasis.
Asunto(s)
Adenoma/metabolismo , Receptor gp130 de Citocinas/metabolismo , Factor de Transcripción STAT3/metabolismo , Transducción de Señal/fisiología , Neoplasias Gástricas/metabolismo , Activación Transcripcional/fisiología , Factor de Crecimiento Transformador beta/metabolismo , Adenoma/fisiopatología , Animales , Receptor gp130 de Citocinas/genética , Técnicas Histológicas , Immunoblotting , Ratones , Ratones Mutantes , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína smad7/metabolismo , Neoplasias Gástricas/fisiopatologíaRESUMEN
Pheophorbide-based photosensitizers have demonstrated tumor cell-specific retention. The lead compound 3-[1'-hexyloxyethyl]-2-devinylpyropheophorbide-a (HPPH) in a clinical trial for photodynamic therapy of head and neck cancer lesions indicated a complete response in 80% of patients. The question arises whether the partial response in 20% of patients is due to inefficient retention of photosensitizers by tumor cells and, if so, can the photosensitizer preference of individual cancer cases be identified prior to photodynamic therapy. This study determined the specificity of head and neck cancer cells and tumor tissues for the uptake and retention of diffusible pheophorbides differing in peripheral groups on the macrocycle that contribute to cellular binding. The relationship between photosensitizer level and light-mediated photoreaction was characterized to identify markers for predicting the effectiveness of photodynamic therapy in situ. The experimental models were stromal and epithelial cells isolated from head and neck tumor samples and integrated into monotypic tissue cultures, reconstituted three-dimensional co-cultures, and xenografts. Tumor cell-specific photosensitizer retention patterns were identified, and a procedure was developed to allow the diagnostic evaluation of HPPH binding by tumor cells in individual cancer cases. The findings of this study may assist in designing conditions for photosensitizer application and photodynamic therapy of head and neck cancer lesions optimized for each patient's case.
Asunto(s)
Neoplasias de Cabeza y Cuello , Fotoquimioterapia , Clorofila/uso terapéutico , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Humanos , Fármacos Fotosensibilizantes/metabolismo , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/uso terapéuticoRESUMEN
This study investigated the impact of anionic and cationic substituents of the pyropheophorbide-based photosensitizers (PS) on uptake and retention by tumor epithelial cells and photodynamic therapy (PDT). A series of PSs were generated that bear carboxylic acid functionalities, alkyl amines with variable length of carbon units or as a quaternary ammonium salt introduced at position 172 of 3-(1'-hexyloxy)ethyl-3-devinylpyropheophorbide-a (HPPH). The nature of the functionalities in the macrocycle made a significant difference in overall lipophilicity (log D values at pH 7.4), and in binding to and retention by human and murine tumor cells. Depending on the presence of functional groups, the PSs showed a change in cellular uptake from diffusion to endocytosis and in the preference for subcellular localization to mitochondria/ER or lysosomes. Two and more carboxylic groups drastically reduced uptake by all cell types. In contrast, PSs with amine and quaternary amine salt showed higher cellular binding, uptake and in vitro PDT efficacy than HPPH. The enhanced cellular uptake of the cationic PSs was accompanied by a loss of tumor cell specificity and contributed to severe systemic toxicity in tumor-bearing mice intravenously injected with the PS and subjected to investigate their therapeutic potential.
Asunto(s)
Fotoquimioterapia , Fármacos Fotosensibilizantes , Animales , Transporte Biológico , Lisosomas/metabolismo , Ratones , Fármacos Fotosensibilizantes/metabolismo , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/uso terapéuticoRESUMEN
To investigate the impact of mono- and di-ß-galactose moieties in tumor uptake and photodynamic therapy (PDT) efficacy, HPPH [3-(1'-hexyloxy)ethyl-3-devinylpyropheophorobide-a], the meso pyropheophorbide-a [3-ethyl-3-devinyl-pyropheophorbide-a], and the corresponding 20-benzoic acid analogs were used as starting materials. Reaction of the intermediates containing one or two carboxylic acid functionalities with 1-aminogalactose afforded the desired 172- or 20(4')- mono- and 172, 20(4')-di galactose conjugated photosensitizers (PSs) with and without a carboxylic acid group. The overall lipophilicity caused by the presence of galactose in combination with either an ethyl or (1'-hexyloxy)ethyl side chain at position-3 of the macrocycle made a significant difference in in vitro uptake by tumor cells and photoreaction upon light exposure. Interestingly, among the PSs investigated, compared to HPPH 1 the carbohydrate conjugates 2 and 11 in which ß-galactose moieties are conjugated at positions 172 and 20(4') of meso-pyro pheophorbide-a showed similar in vitro efficacy in FaDu cell lines, but in SCID mice bearing FaDu tumors (head & neck) Ps 11 gave significantly improved long-term tumor cure.
RESUMEN
BACKGROUND: Constitutive activation of signal transducer and activator of transcription-3 (STAT3) was detected in blasts from approximately 50% of patients with acute myeloid leukemia (AML) and was correlated with an adverse outcome. In vitro treatment of AML blasts with arsenic trioxide (ATO) down-regulated STAT3 activity within 6 hours associated with a reduced viability within 48 hours. METHODS: A phase 1 clinical trial to evaluate the biologically effective dose and/or the maximally tolerated dose (MTD) of ATO in vivo in conjunction with high-dose cytarabine (Hidac) and idarubicin (Ida) in patients with AML aged <60 years was conducted. Data were compared with 117 historic AML patients who had received treatment with Hidac/Ida. RESULTS: In total, 61 patients were enrolled onto 11 different dose levels (from 0.01 to 0.65 mg/kg ideal body weight). The MTD was 0.5 mg/kg. Compared with historic controls, patients who received ATO/Hidac/Ida, although they had similar pretreatment characteristics, had better overall survival (P = .039). CONCLUSIONS: ATO priming may have improved the outcome of patients aged <60 years with AML who received Hidac/Ida. The current data suggested that ATO may enhance the effect of chemotherapy. The authors concluded that further studies of this novel combination are warranted.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Arsenicales/administración & dosificación , Citarabina/administración & dosificación , Idarrubicina/administración & dosificación , Leucemia Mieloide Aguda/tratamiento farmacológico , Óxidos/administración & dosificación , Factor de Transcripción STAT3/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Trióxido de Arsénico , Arsenicales/efectos adversos , Citarabina/efectos adversos , Regulación hacia Abajo , Femenino , Humanos , Idarrubicina/efectos adversos , Masculino , Persona de Mediana Edad , Óxidos/efectos adversosRESUMEN
A variety of cytokines play a role in the response to an inflammatory stimulus. The interleukin-6 (IL-6)-type cytokines are released in response to tissue injury or an inflammatory stimulus. They act locally and systemically to generate a variety of physiologic responses, principal among them is the acute phase response. The IL-6 type cytokines demonstrate pleiotropy and redundancy of actions. This is made possible by the distinctive characteristics of the IL-6 receptor complex, which contains an ubiquitous subunit that is shared by most IL-6-type cytokines, as well as a cytokine-specific subunit.
Asunto(s)
Interleucina-6/fisiología , Heridas y Lesiones/inmunología , Cuidados Críticos , Cirugía General , Humanos , Interleucina-6/química , Interleucina-6/metabolismo , Receptores de Interleucina-6/química , Receptores de Interleucina-6/metabolismo , Sepsis/inmunologíaRESUMEN
A variety of cytokines play a role in the inflammatory response. Interleukin-6 (IL-6)-type cytokines are released in response to tissue injury or an inflammatory stimulus, and act locally and systemically to generate a variety of physiologic responses. Interleukin-6 concentrations are elevated after surgery, trauma, and critical illness. The magnitude of IL-6 elevation correlates with the extent of tissue trauma/injury severity. Furthermore, there is an association between IL-6 elevation and adverse outcome. Interleukin-6 levels can also be used to stratify patients for therapeutic intervention.
Asunto(s)
Enfermedad Crítica , Interleucina-6/fisiología , Procedimientos Quirúrgicos Operativos , Heridas y Lesiones/sangre , Humanos , Inflamación/sangre , Inflamación/etiología , Procedimientos Quirúrgicos Operativos/efectos adversos , Heridas y Lesiones/complicacionesRESUMEN
To investigate the importance of the chirality and precise structure at position 3(1') of pyropheophorbide-a for tumor cell specificity and photodynamic therapy (PDT), a series of photosensitizers (PSs) was synthesized: (a) with and without chirality at position 3(1'), (b) alkyl ether chain with a variable number of chiral centers, (c) hexyl ether versus thioether side chain, and (d) methyl ester versus carboxylic acid group at position 172. The cellular uptake and specificity were defined in human lung and head/neck cancer cells. PSs without a chiral center and with an alkyl chain or thioether functionalities showed limited uptake and PDT efficacy. Replacing the methyl group at the chiral center with a propyl group or introducing an additional chiral center improved cellular retention and tumor cell specificity. Replacing the carboxylic acid with methyl ester at position 172 lowered cellular uptake and PDT efficacy. A direct correlation between the PS uptake in vitro and in vivo was identified.
Asunto(s)
Clorofila/análogos & derivados , Fármacos Fotosensibilizantes/metabolismo , Animales , Clorofila/química , Clorofila/metabolismo , Clorofila/uso terapéutico , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Neoplasias de Cabeza y Cuello/patología , Humanos , Luz , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Ratones , Ratones SCID , Microscopía Fluorescente , Fotoquimioterapia , Fármacos Fotosensibilizantes/síntesis química , Fármacos Fotosensibilizantes/uso terapéutico , Solubilidad , Estereoisomerismo , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
Erlotinib was covalently linked to 3-(1'-hexyloxy)ethyl-3-devinylpyropheophorbide-a (HPPH) and structurally related chlorins and bacteriochlorins at different positions of the tetrapyrrole ring. The functional consequence of each modification was determined by quantifying the uptake and subcellular deposition of the erlotinib conjugates, cellular response to therapeutic light treatment in tissue cultures, and in eliminating of corresponding tumors grown as a xenograft in SCID mice. The experimental human cancer models the established cell lines UMUC3 (bladder), FaDu (hypopharynx), and primary cultures of head and neck tumor cells. The effectiveness of the compounds was compared to that of HPPH. Furthermore, specific functional contribution of the carboxylic acid side group at position 172 and the chiral methyl group at 3(1') to the overall activity of the chimeric compounds was assessed. Among the conjugates investigated, the PS 10 was identified as the most effective candidate for achieving tumor cell-specific accumulation and yielding improved long-term tumor control.
Asunto(s)
Clorhidrato de Erlotinib/química , Fármacos Fotosensibilizantes/síntesis química , Porfirinas/química , Animales , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Técnicas de Cocultivo , Femenino , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Neoplasias de Cabeza y Cuello/mortalidad , Neoplasias de Cabeza y Cuello/patología , Humanos , Ratones , Ratones SCID , Fotoquimioterapia , Fármacos Fotosensibilizantes/química , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/uso terapéutico , Estereoisomerismo , Relación Estructura-Actividad , Tasa de SupervivenciaRESUMEN
We present initial results obtained during the course of a Phase I clinical trial of 2-1[hexyloxyethyl]-2-devinylpyropheophorbide-a (HPPH)-mediated photo-dynamic therapy (PDT) in a head and neck cancer patient. We quantified blood flow, oxygenation and HPPH drug photobleaching before and after therapeutic light treatment by utilizing fast, non-invasive diffuse optical methods. Our results showed that HPPH-PDT induced significant drug photobleaching, and reduction in blood flow and oxygenation suggesting significant vascular and cellular reaction. These changes were accompanied by cross-linking of the signal transducer and activator of transcription 3 (STAT3), a molecular measure for the oxidative photoreaction. These preliminary results suggest diffuse optical spectroscopies permit non-invasive monitoring of PDT in clinical settings of head and neck cancer patients.
Asunto(s)
Clorofila/análogos & derivados , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Hemodinámica/fisiología , Fotoblanqueo , Fotoquimioterapia , Clorofila/uso terapéutico , Reactivos de Enlaces Cruzados/farmacología , Tecnología de Fibra Óptica , Humanos , Masculino , Fotoblanqueo/efectos de los fármacos , Factor de Transcripción STAT3/metabolismoRESUMEN
The contribution of acute phase plasma proteins to host immune responses remains poorly characterized. To better understand the role of the acute phase reactant and major hemoglobin-binding protein haptoglobin (Hp) on the function of immune cells, we generated Hp-deficient C57BL/6J mice. These mice exhibit stunted development of lymphoid organs associated with lower counts of mature T and B cells in the blood and secondary lymphoid compartments. Moreover, these mice show markedly reduced adaptive immune responses as represented by reduced accumulation of IgG antibody after immunization with adjuvant and nominal antigen, abrogation of Th1-dominated delayed-type hypersensitivity reaction, loss of mitogenic responses mounted by T cells, and reduced T cell responses conveyed by APCs. Collectively, these defects are in agreement with the observations that Hp-deficient mice are not capable of generating a recall response or deterring a Salmonella infection as well as failing to generate tumor antigen-specific responses. The administration of Hp to lymphocytes in tissue culture partially ameliorates these functional defects, lending further support to our contention that the acute phase response protein Hp has the ability to regulate immune cell responses and host immunity. The phenotype of Hp-deficient mice suggests a major regulatory activity for Hp in supporting proliferation and functional differentiation of B and T cells as part of homeostasis and in response to antigen stimulation.
Asunto(s)
Haptoglobinas/inmunología , Inmunidad/inmunología , Reacción de Fase Aguda/inmunología , Traslado Adoptivo , Animales , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Epítopos , Regulación de la Expresión Génica/efectos de los fármacos , Haptoglobinas/deficiencia , Haptoglobinas/genética , Haptoglobinas/metabolismo , Hipersensibilidad Tardía/inmunología , Inmunidad/efectos de los fármacos , Inmunización , Lipopolisacáridos/farmacología , Activación de Linfocitos/efectos de los fármacos , Tejido Linfoide/embriología , Ratones , Ratones Endogámicos C57BL , Mitógenos/farmacología , Ovalbúmina , Fenotipo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunologíaRESUMEN
This study determined in primary cultures of human lung cancer cells the cell specificity of chlorin-based photosensitizers. Epithelial cells (ECs) preferentially retained 3-[1-hexyloxyethyl]-2-devinylpyropheophorbide-a (HPPH) and related structural variants. Tumor-associated fibroblasts (Fb) differ from EC by a higher efflux rate of HPPH. Immunoblot analyses indicated dimerization of STAT3 as a reliable biomarker of the photoreaction. Compared to mitochondria/ER-localized photoreaction by HPPH, the photoreaction by lysosomally targeted HPPH-lactose showed a trend toward lower STAT3 cross-linking. Lethal consequence of the photoreaction differed between EC and Fb with the latter cells being more resistant. A survey of lung tumor cases indicated a large quantitative range by which EC retains HPPH. The specificity of HPPH retention defined in vitro could be confirmed in vivo in selected cases grown as xenografts. HPPH retention as a function of the tetrapyrrole structure was evaluated by altering side groups on the porphyrin macrocycle. The presence or absence of a carboxylic acid at position 172 proved to be critical. A benzyl group at position 20 enhanced retention in a subset of cancer cells with low HPPH binding. This study indicated experimental tools that are potentially effective in defining the photosensitizer preference and application for individual patient's cancer lesions.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Clorofila/análogos & derivados , Neoplasias Pulmonares/metabolismo , Fármacos Fotosensibilizantes/farmacología , Animales , Carcinoma de Pulmón de Células no Pequeñas/patología , Clorofila/metabolismo , Clorofila/farmacología , Femenino , Humanos , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones SCID , Fotoquimioterapia , Fármacos Fotosensibilizantes/metabolismo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Tuberous sclerosis complex (TSC) is a tumor predisposition syndrome with significant renal cystic and solid tumor disease. While the most common renal tumor in TSC, the angiomyolipoma, exhibits a loss of heterozygosity associated with disease, we have discovered that the renal cystic epithelium is composed of type A intercalated cells that have an intact Tsc gene that have been induced to exhibit Tsc-mutant disease phenotype. This mechanism appears to be different than that for ADPKD. The murine models described here closely resemble the human disease and both appear to be mTORC1 inhibitor responsive. The induction signaling driving cystogenesis may be mediated by extracellular vesicle trafficking.
Asunto(s)
Enfermedades Renales Quísticas/patología , Esclerosis Tuberosa/patología , Animales , Modelos Animales de Enfermedad , Vesículas Extracelulares/genética , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/patología , Femenino , Enfermedades Renales Quísticas/genética , Enfermedades Renales Quísticas/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Ratones Noqueados , Esclerosis Tuberosa/genética , Esclerosis Tuberosa/metabolismo , Proteína 1 del Complejo de la Esclerosis Tuberosa/genética , Proteína 1 del Complejo de la Esclerosis Tuberosa/metabolismo , Proteína 2 del Complejo de la Esclerosis Tuberosa/genética , Proteína 2 del Complejo de la Esclerosis Tuberosa/metabolismoRESUMEN
Interleukin-31 (IL-31), an IL-6 cytokine family member, is proposed to play a role in animal models of airway hyperreactivity. It is produced by activated T cells and signals via a heterodimeric receptor complex composed of IL-31Ralpha and OSMRbeta. Only low levels of IL-31Ralpha expression have been demonstrated in pulmonary epithelial cell lines, however, and little is known about the ability to regulate its expression and signaling. Therefore, primary cultures of human bronchial and alveolar epithelial cells, pulmonary fibroblasts, pulmonary macrophages, and established lines of immortalized bronchial epithelial cells (HBE) and alveolar carcinoma cells (A549) were analyzed by RT-PCR, immunoblotting, and thymidine incorporation. Distinct, cell type-specific regulation of IL-31Ralpha expression was detected. Transforming growth factor-beta (TGF-beta) enhanced IL-31Ralpha mRNA expression in primary cultures and established lines of epithelial cells, but not in macrophages. In contrast, interferon-gamma (IFN-gamma) induced IL-31Ralpha mRNA expression in macrophages. IL-31Ralpha protein expression was below detection threshold in primary epithelial cell cultures but was detectable in A549 cells and increased with TGF-beta treatment. In HBE and A549 cells, TGF-beta pretreatment increased IL-31-mediated Stat3 and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation. In A549 cells, TGF-beta magnified IL-31-dependent suppression of proliferation. The data suggest that increased IL-31Ralpha expression correlates with an enhanced response to IL-31.
Asunto(s)
Bronquios/citología , Células Epiteliales/metabolismo , Fibroblastos/metabolismo , Regulación de la Expresión Génica , Macrófagos Alveolares/metabolismo , Alveolos Pulmonares/citología , Receptores de Interleucina/metabolismo , Bronquios/metabolismo , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Medios de Cultivo Condicionados , ADN/biosíntesis , Dexametasona/farmacología , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Immunoblotting , Macrófagos Alveolares/citología , Macrófagos Alveolares/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de Interleucina/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/efectos de los fármacos , Factor de Crecimiento Transformador beta/farmacologíaRESUMEN
We have demonstrated that constitutive signal transducer and activator of transcription (STAT) 3 activity, observed in approximately 50% of acute myeloid leukemia (AML) cases, is associated with adverse treatment outcome. Constitutive STAT3 activation may result from the expression of oncogenic protein tyrosine kinases or from autocrine stimulation by hematopoietic growth factors. These causes are generally neither necessary nor sufficient for leukemogenesis; additional transforming events or growth stimulatory processes are needed. Here we review the literature addressing epigenetic regulation as a mechanism controlling STAT3 signaling in AML. A better understanding of mechanisms of dysregulation of STAT signaling pathways may serve as a basis for designing novel therapeutic strategies that target these pathways in leukemia cells.
Asunto(s)
Epigénesis Genética , Leucemia Mieloide Aguda/genética , Factor de Transcripción STAT3/genética , Humanos , Leucemia Mieloide Aguda/metabolismo , Metilación , Factor de Transcripción STAT3/metabolismo , Transducción de SeñalRESUMEN
BACKGROUND: Hepatocellular carcinomas (HCC) associated with inflammation that undergo radiofrequency ablation (RFA) appear to have poorer local control rates. Little is known of how mediators of inflammation influence HCC cellular thermotolerance, which in part is mediated by heat shock protein 70 (HSP70). This study determines how inflammatory mediators affect cellular thermotolerance and provides insight into how associated inflammation may impact HCC RFA local control rates. METHODS: HepG2 cell lines were cultured in control medium (CM) or CM containing conditioned medium of endotoxin-activated macrophage (CMM). Serial dilutions of CMM established microenvironments approximating low, medium, and high CMM. All groups underwent a heat shock challenge (HSC) at 45 degrees C for 10 min. Western blot, Northern blot, densitometric analysis, along with thymidine and clonogenic assays determined how inflammation influenced multiple biological endpoints. RESULTS: Cells cultured in low CMM expressed significantly more HSP70 RNA and protein compared with control cells after HSC. The cells also had a higher proliferative and survival rate after HSC compared with control cells. Medium CMM cultured cells had no significant difference in HSP70 RNA and protein production or proliferation and survival rates after HSC, compared with CM cultured cells. AT high CMM, the inhibitory effects of inflammatory mediators prevailed and all of the measured endpoints were significantly less compared with CM cultured cells. CONCLUSIONS: This study demonstrates that inflammation can alter the responsiveness of HCC cells to a HSC in a dose-dependent manner. This study supports the clinical observation that HCC associated with chronic inflammation have worse RFA local control rates.
Asunto(s)
Carcinoma Hepatocelular/complicaciones , Calor , Mediadores de Inflamación/metabolismo , Inflamación/complicaciones , Neoplasias Hepáticas/complicaciones , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/terapia , Ablación por Catéter , Línea Celular Tumoral , Citocinas/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Humanos , Inflamación/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/terapiaRESUMEN
PURPOSE: Photodynamic therapy (PDT) depends on the delivery of a photosensitizer to the target tissue that, under light exposure, produces singlet oxygen and other reactive oxygen species, which in turn cause the death of the treated cell. This study establishes a quantitative marker for the photoreaction that will predict the outcome of PDT. EXPERIMENTAL DESIGN: Cells in tissue culture, murine s.c. tumors, and endobronchial carcinomas in patients were treated with PDT, and the noncleavable cross-linking of the latent signal transducer and activator of transcription 3 (STAT3) was determined. RESULTS: Murine and human cancer cell lines reacted to PDT by an immediate covalent cross-linking of STAT3 to homodimeric and other complexes. The magnitude of this effect was strictly a function of the PDT reaction that is determined by the photosensitizer concentration and light dose. The cross-link reaction of STAT3 was proportional to the subsequent cytotoxic outcome of PDT. An equivalent photoreaction as detected in vitro occurred in tumors treated in situ with PDT. The light dose-dependent STAT3 cross-linking indicated the relative effectiveness of PDT as a function of the distance of the tissue to the treating laser light source. Absence of cross-links correlated with treatment failure. CONCLUSIONS: The data suggest that the relative amount of cross-linked STAT3 predicts the probability for beneficial outcome, whereas absence of cross-links predicts treatment failure. Determination of STAT3 cross-links after PDT might be clinically useful for early assessment of PDT response.