RESUMEN
Transcutaneous stimulation of the auricular branch of the vagus nerve (tVNS) has been proposed as a treatment for a spectrum of physical and psychological disorders. One of the proposed working mechanisms of tVNS is a modulatory effect on the locus coeruleus - noradrenaline (LC-NA) network. We tested this hypothesis in humans in a series of three studies: one focusing on high trait worriers, and two in healthy populations. In all three studies, we tested whether tVNS increases resting pupil diameter - as an index of LC-NA network activity. Additionally, we tested whether tVNS affects task performance and task-related pupil dilation during an Attentional Blink task. We found no evidence that tVNS increases pupil diameter or task-related pupil dilation in any of the tasks. No consistent effects of tVNS on performance on the attentional blink task were found. Overall, the results of these studies indicate that tVNS does not affect these behavioral and physiological indices of noradrenergic activity.
Asunto(s)
Pupila , Estimulación Eléctrica Transcutánea del Nervio , Estimulación del Nervio Vago , Humanos , Norepinefrina , Pupila/fisiología , Nervio VagoRESUMEN
Fear overgeneralization is thought to be one of the cardinal processes underlying anxiety disorders, and a determinant of the onset, maintenance and recurrence of these disorders. Animal studies have shown that stimulating the vagus nerve (VNS) affects neuronal pathways implicated in pattern separation and completion, suggesting it may reduce the generalization of a fear memory to novel situations. In a one-day study, 58 healthy students were subjected to a fear conditioning, fear generalization, and fear extinction paradigm. Participants were randomly assigned to receive either transcutaneous auricular VNS (tVNS; final Nâ¯=â¯29) or sham stimulation (final Nâ¯=â¯29) during the generalization and extinction phases. tVNS did not affect fear generalization, as reflected by US expectancy ratings and fear potentiated startle responses. However, participants who received tVNS reported lower US expectancy ratings to the CS+ during the extinction phase, possibly reflecting a stronger declarative extinction of fear. No effects of tVNS on fear potentiated startle responses during extinction were found. The pattern of findings regarding extinction of declarative fear suggest a facilitating effect of tVNS.
Asunto(s)
Condicionamiento Clásico/fisiología , Extinción Psicológica/fisiología , Miedo/fisiología , Generalización Psicológica/fisiología , Estimulación Eléctrica Transcutánea del Nervio , Nervio Vago/fisiología , Adolescente , Adulto , Femenino , Frecuencia Cardíaca/fisiología , Humanos , Masculino , Reflejo de Sobresalto/fisiología , Adulto JovenRESUMEN
Worrying is a central component of anxiety disorders. We tested whether non-invasive vagus nerve stimulation reduces negative thought intrusions in high worriers. Worry was assessed with a Breathing Focus Task, which consists of a pre-worry period, a worry induction, and a post-worry period. Ninety-seven high worriers were randomly allocated to receive transcutaneous electrical stimulation of the auricular branch of the vagus nerve at the concha (tVNS), or of the earlobe (sham stimulation) throughout the lab session. Participants who received tVNS reported significantly fewer negative thought intrusions during the pre-worry period, but the effects of tVNS after the worry induction were mixed. An exploratory analysis indicated that participants in the tVNS condition were more likely to report negative thought intrusions shortly after the worry induction, but became less likely to do so as the post-worry period went on. No effects of tVNS on RMSSD were observed. These findings provide preliminary indications that tVNS may decrease the occurrence of worrisome thoughts.
Asunto(s)
Trastornos de Ansiedad/psicología , Trastornos de Ansiedad/terapia , Pensamiento/fisiología , Estimulación Eléctrica Transcutánea del Nervio/métodos , Estimulación del Nervio Vago/métodos , Adulto , Trastornos de Ansiedad/fisiopatología , Femenino , Humanos , Masculino , Nervio Vago/fisiología , Adulto JovenRESUMEN
Extinction memories are fragile and their formation has been proposed to partially rely on vagus nerve activity. We tested whether stimulating the auricular branch of the vagus (transcutaneous VNS; tVNS) accelerates extinction and reduces spontaneous recovery of fear. Forty-two healthy students participated in a 3-day fear conditioning study, where we tested fear acquisition (day 1), fear extinction (day 2) and the retention of the extinction memory (day 3). During extinction, participants were randomly allocated to receive tVNS or sham stimulation concurrently with each CS presentation. During the acquisition and retention phases, all participants received sham stimulation. Indexes of fear included US-expectancy, startle blink EMG and skin conductance responses. Results showed successful acquisition and extinction of fear in all measures. tVNS facilitated the extinction of declarative fear (US expectancy ratings), but did not promote a stronger retention of the declarative extinction memory. No clear effects of tVNS on extinction and retention of extinction were found for the psychophysiological indexes. The present findings provide tentative indications that tVNS could be a promising tool to improve fear extinction and call for larger scale studies to replicate these effects.
Asunto(s)
Condicionamiento Psicológico/fisiología , Extinción Psicológica/fisiología , Miedo/fisiología , Memoria/fisiología , Estimulación Eléctrica Transcutánea del Nervio , Estimulación del Nervio Vago , Adulto , Femenino , Respuesta Galvánica de la Piel/fisiología , Humanos , Masculino , Reflejo de Sobresalto/fisiología , Adulto JovenRESUMEN
BACKGROUND: The expression of side-population (SP) cells and their relation to tumour-initiating cells (T-ICs) have been insufficiently studied in breast cancer (BC). We therefore evaluated primary cell cultures derived from patients and a panel of human BC cell lines with luminal- or basal-molecular signatures for the presence of SP and BC stem cell markers. METHODS: The SPs from luminal-type BC were analysed for BC T-IC characteristics, including human epidermal growth factor receptor 2 (HER2), ERalpha, IGFBP7 expression and their ability to initiate tumours in non-obese diabetic severe combined immunodeficiency (NOD/SCID) mice. Pharmacological modulators were used to assess the effects of HER2 signalling and breast cancer-resistance protein (BCRP) expression on SPs. RESULTS: The SP was more prevalent in the luminal subtype of BC compared with the basal subtype. HER2 expression was significantly correlated with the occurrence of an SP (r(2)=0.75, P=0.0003). Disappearance of SP in the presence of Ko143, a specific inhibitor of the ATP-binding cassette transporter BCRP, suggests that BCRP is the predominant transporter expressed in this population. The SP also decreased in the presence of HER2 signalling inhibitors AG825 or trastuzumab, strengthening the notion that HER2 contributed to the SP phenotype, likely through downstream AKT signalling. The SP cells from luminal-type MCF-7 cells with enforced expression of HER2, and primary cells with luminal-like properties from a BC patient, displayed enrichment in cells capable of repopulating tumours in NOD/SCID mice. Engraftment of SP cells was inhibited by pretreatment with AG825 or by in vivo treatment with trastuzumab. INTERPRETATION: Our findings indicate an important role of HER2 in regulating SP and hence T-ICs in BC, which may account for the poor responsiveness of HER2-positive BCs to chemotherapy, as well as their aggressiveness.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Proteínas de Neoplasias/metabolismo , Células Madre Neoplásicas/patología , Receptor ErbB-2/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/antagonistas & inhibidores , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales Humanizados , Western Blotting , Resistencia a Antineoplásicos , Femenino , Citometría de Flujo , Humanos , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/antagonistas & inhibidores , Ratones , Ratones Endogámicos NOD , Ratones Desnudos , Ratones SCID , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor ErbB-2/antagonistas & inhibidores , Transducción de Señal , Trastuzumab , Células Tumorales CultivadasRESUMEN
Telomeres and telomerase are targets for anticancer drug development and specific inhibitors are currently under clinical investigation. However, it has been reported that standard cytotoxic agents can affect telomere length and telomerase activity suggesting that they also have of a role in drug resistance. in this study, telomere lengths and telomerase activity as well as drug efflux pump expression, glutathione (GSH) levels and polyadenosine-ribose polymerase (PARP) cleavage were assessed in a panel of human tumor cell lines made resistant to vindesine, gemcitabine and cisplatin. these included two lung cancer cell lines resistant to vindesine (LXFL 529L/Vind, LXFA 526L/Vind), a renal cancer cell line (RXF944L/Gem) and an ovarian cancer cell line (AG6000) resistant to gemcitabine, and one resistant to cisplatin (ADDP). The resistant clones were compared to their parental lines and evaluated for cross resistance to other cytotoxic agents. Several drug specific resistance patterns were found, and various complex patterns of cross resistance emerged from some cell lines, but these mechanisms of resistance could not be related to drug efflux pump expression, GSH levels or pARp cleavage. However, all displayed changes in telomerase activity and/or telomere length. Our studies present evidence that telomere maintenance should be taken into consideration in efforts not only to overcome drug resistance, but also to optimize the use of telomere-based therapeutics.
Asunto(s)
Antineoplásicos/farmacología , Resistencia a Antineoplásicos , Telomerasa/metabolismo , Telómero/genética , Western Blotting , Ensayo de Unidades Formadoras de Colonias , Femenino , Glutatión/metabolismo , Humanos , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/metabolismoRESUMEN
Telomerase and telomeres are attractive targets for anticancer therapy. This is supported by the fact that the majority of human cancers express the enzyme telomerase which is essential to maintain their telomere length and thus, to ensure indefinite cell proliferation--a hallmark of cancer. Tumours have relatively shorter telomeres compared to normal cell types, opening the possibility that human cancers may be considerably more susceptible to killing by agents that inhibit telomere replication than normal cells. Advances in the understanding of the regulation of telomerase activity and the telomere structure, as well as the identification of telomerase and telomere associated binding proteins have opened new avenues for therapeutic intervention. Here, we review telomere and telomerase biology and the various approaches which have been developed to inhibit the telomere/telomerase complex over the past decade. They include inhibitors of the enzyme catalytic subunit and RNA component, agents that target telomeres, telomerase vaccines and drugs targeting binding proteins. The emerging role of telomerase in cancer stem cells and the implications for cancer therapy are also discussed.
Asunto(s)
Vacunas contra el Cáncer , Inhibidores Enzimáticos , Neoplasias/tratamiento farmacológico , Telomerasa/antagonistas & inhibidores , Telómero/efectos de los fármacos , Animales , Vacunas contra el Cáncer/química , Vacunas contra el Cáncer/farmacología , Vacunas contra el Cáncer/uso terapéutico , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Humanos , Modelos Biológicos , Estructura Molecular , Neoplasias/enzimología , Neoplasias/patologíaRESUMEN
The pentacyclic acridinium methosulfate salt RHPS4 induces the 3'single-stranded guanine-rich telomeric overhang to fold into a G-quadruplex structure. Stabilisation of the latter is incompatible with an attachment of telomerase to the telomere and thus G-quadruplex ligands can effectively inhibit both the catalytic and capping functions of telomerase. In this study, we examined mechanisms underlying telomere uncapping by RHPS4 in uterus carcinoma cells (UXF1138L) with short telomeres and compared the susceptibility of bulk and clonogenic cancer cells to the G-quadruplex ligand. We show that treatment of UXF1138L cells with RHPS4 leads to the displacement of the telomerase catalytic subunit (hTERT) from the nucleus, induction of telomere-initiated DNA-damage signalling and chromosome fusions. We further report that RHPS4 is more potent against cancer cells that grow as colonies in soft agar than cells growing as monolayers. Human cord blood and HEK293T embryonic kidney cell colony forming units, however, were more resistant to RHPS4. RHPS4-treated UXF1138L xenografts had a decreased clonogenicity, showed loss of nuclear hTERT expression and an induction of mitotic abnormalities compared with controls. Although single-agent RHPS4 had limited in vivo efficacy, a combination of RHPS4 with the mitotic spindle poison Taxol caused tumour remissions and further enhancement of telomere dysfunction.
Asunto(s)
Acridinas/farmacología , Antineoplásicos/farmacología , Inhibidores Enzimáticos/farmacología , Células Madre Neoplásicas/efectos de los fármacos , Telomerasa/antagonistas & inhibidores , Telómero/efectos de los fármacos , Animales , Proliferación Celular/efectos de los fármacos , Sinergismo Farmacológico , Femenino , Humanos , Ratones , Paclitaxel/farmacologíaRESUMEN
The pyrrolobenzodiazepine monomer DRH-417 is a member of the anthramycin group of anti-tumor antibiotics that bind covalently to the N2 of guanine within the minor groove of DNA. DRH-417 emerged from the EORTC-Drug Discovery Committee and NCI 60 cell line in vitro screening programs as a potent antiproliferative agent with differential sensitivity towards certain cancer types such as melanoma, breast and renal cell carcinoma (mean IC(50) = 3 nM). DRH-417 was therefore tested for in vivo activity. The maximum tolerated dose (MTD) was established as 0.5 mg/kg given i.p. Marked anti-tumor activity was seen in two human renal cell cancers, one breast cancer and a murine colon tumor model (p<0.01). A selective HPLC (LC/MS) analytical method was developed and plasma pharmacokinetics determined. At a dose of 0.5 mg kg(-1), the plasma AUC was 540 nM h (197.1 ng h ml(-1)) and the peak plasma concentration (171 nM [62.4 ng ml(-1)]) occurred at 30 min., reaching doses levels well above those needed for in vitro antiproliferative activity. Genomic profiling of in vivo sensitive tumors revealed that the latter have an activated insulin-like growth factor signaling pathway.
Asunto(s)
Antibióticos Antineoplásicos/farmacología , Benzodiazepinas/farmacología , Pirroles/farmacología , Animales , Antramicina/farmacología , Antibióticos Antineoplásicos/análisis , Antibióticos Antineoplásicos/farmacocinética , Benzodiazepinas/análisis , Benzodiazepinas/uso terapéutico , Proliferación Celular/efectos de los fármacos , Cromatografía Líquida de Alta Presión , Perfilación de la Expresión Génica , Humanos , Espectrometría de Masas , Ratones , Trasplante de Neoplasias , Neoplasias Experimentales/tratamiento farmacológico , Neoplasias Experimentales/metabolismo , Pirroles/análisis , Pirroles/uso terapéutico , Trasplante HeterólogoRESUMEN
Tumour hypoxia confers poor prognosis in a wide range of solid tumours, due to an increased malignancy, increased likelihood of metastasis and treatment resistance. Poorly oxygenated tumours are resistant to both radiation therapy and chemotherapy. However, although the link between radiation therapy and hypoxia is well established in a range of clinical studies, evidence of its influence on chemotherapy response is lacking. In this study, a panel of human tumour-derived xenografts that have been characterised previously for in vivo response to a large series of anti-cancer agents, and have been found to show chemosensitivities that correlate strongly with the parent tumour, were used to address this issue. Immunohistochemistry was carried out on formalin-fixed, paraffin-embedded sections of xenograft samples to detect expression of the intrinsic hypoxia marker Glut-1 and adducts of the bioreductive hypoxia marker pimonidazole. Glut-1 scores correlated significantly with T/C values for CCNU sensitivity (r = 0.439, P = 0.036, n = 23) and showed a borderline significant correlation with dacarbazine T/C (r = 0.405, P = 0.076, n = 20). However, there was no correlation between both Glut-1 and pimonidazole scores and T/C obtained for the bioreductive drug mitomycin C. The use of human tumour-derived xenografts offers a potentially useful way of using archival material to determine the influence of hypoxia and other tumour-microenvironmental factors on chemosensitivity without the direct use of human subjects.
Asunto(s)
Antineoplásicos Alquilantes/farmacología , Proteínas de Transporte de Monosacáridos/fisiología , Neoplasias Experimentales/tratamiento farmacológico , Animales , Biomarcadores , Hipoxia de la Célula , Chaperón BiP del Retículo Endoplásmico , Transportador de Glucosa de Tipo 1 , Proteínas de Choque Térmico/fisiología , Humanos , Ratones , Chaperonas Moleculares/fisiología , Trasplante de Neoplasias , Neoplasias Experimentales/metabolismo , Trasplante HeterólogoRESUMEN
Pluripotent cells can be grown in clonogenic assays. The tumour stem-cell fraction, which accounts for <0.4% of the total cells, and which is considered the most relevant cell type in the development of metastases and recurrences, is able to divide and to form colonies in a semisolid matrix (agar or methylcellulose). Major applications of the tumour clonogenic assay (TCA) are chemosensitivity testing of tumours and xenografts, and for assessments within drug discovery programmes. Of critical relevance for the usefulness of the TCA is whether it can predict sensitivity or resistance towards clinically used agents. When we compared the response of human tumours established as xenografts in nude mice in the TCA in vitro to that of the clinical response, 62% of the comparisons for drug sensitivity, and 92% of the comparisons for drug resistance were correct. The same percentage of true/false observations was found when tumours were tested after serial passage in nude mice in the TCA in vitro and their response compared to in vivo activity in corresponding xenografts (60% and 90%, respectively). The highest correct predictive values were, however, found when the clinical response of tumours was compared to their explants established in the nude mouse and treated in vivo. Of 80 comparisons performed, we observed a correct prediction for tumour resistance in 97% and for tumour sensitivity in 90%. In our opinion, the TCA with established human tumour xenografts has an important role in current drug discovery strategies. We therefore included the TCA as secondary assay in our approach to anticancer drug discovery and found that a number of novel agents were active; these are now in advanced preclinical development or clinical trials. Thus, the tumour clonogenic assay has proven predictive value in the chemosensitivity testing of standard and experimental anticancer drugs.
Asunto(s)
Antineoplásicos/uso terapéutico , Neoplasias/tratamiento farmacológico , Ensayo de Tumor de Célula Madre/métodos , Animales , División Celular , Diseño de Fármacos , Células Madre Hematopoyéticas/efectos de los fármacos , Humanos , Técnicas In Vitro , Ratones , Ratones Desnudos , Metástasis de la Neoplasia/patología , Trasplante de Neoplasias , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
The in vitro antiproliferative or stimulatory activity of an aqueous mistletoe extract (AME) with a defined content of bioactive mistletoe lectin (ML) was investigated in 6 human tumor cell lines, including two melanomas and leiomyosarcomas, each of which had previously been reported to show evidence of growth stimulation if treated with low concentrations of isolated ML. The effects of AME were compared to that of the standard cytotoxic agent adriamycin (ADR) using the well established propidium iodide and sulforhodamin B proliferation assays. The AME concentrations used ranged from 0.5 pg to 5 ng (0.82 fMol-85 pM) bioactive ML/ml in melanoma (HT-144, SK-MEL-28) and leiomyosarcoma (SK-MLS-1, S-UT-1B) cell lines and from 0.1-100 ng ML/ml (1.7 pM-1.7 nM) in MCF-7 breast cancer and SW620 colon carcinoma cell lines, respectively. The influence of AME on cell growth was determined at various time-points from 24 hours to 6 days of exposure. We found a time- and cell line-dependent inhibition of tumor cell growth, but no reproducible stimulation of tumor cell proliferation. Inhibitory concentrations 50% (IC50) for e.g. the SK-MEL-28 melanoma cell line, decreased from 4.1 ng ML/ml at 24 hours to 0.16 ng ML/ml at 72 hours and 0.18 ng ML/ml at 5 days. Our data clearly demonstrate that, by applying scientifically valid methods and procedures, the standardized AME did not stimulate tumor cell proliferation but showed time- and concentration-dependent antiproliferative effects.
Asunto(s)
Muérdago/química , Preparaciones de Plantas/farmacología , Proteínas de Plantas , Toxinas Biológicas/farmacología , División Celular/efectos de los fármacos , Línea Celular Tumoral , Inhibidores de Crecimiento/farmacología , Humanos , Concentración 50 Inhibidora , Leiomiosarcoma/tratamiento farmacológico , Leiomiosarcoma/patología , Melanoma/tratamiento farmacológico , Melanoma/patología , Proteínas Inactivadoras de Ribosomas Tipo 2 , Estimulación Química , Agua/químicaRESUMEN
The breast cancer-associated T2A10 clone was originally isolated from a cDNA library enriched for tumour messenger ribonucleic acids. Our survey of 125 microarrayed primary tumour tissues using affinity purified polyclonal antibodies has revealed that corresponding protein is overexpressed in invasive breast cancer and is weakly expressed in kidney and prostate tumours. Now known as RNF11, the gene encodes a RING-H2 domain and a PY motif, both of which mediate protein-protein interactions. In particular, the PPPPY sequence of RNF11 PY motif is identical to that of Smad7, which has been shown to bind to WW domains of Smurf2, an E3 ubiquitin ligase that mediates the ubiquitination and degradation of the TGFbeta receptor complex. Using various mutants of RNF11 in GST pulldown and immunoprecipitation assays, we found that RNF11 interacts with Smurf2 through the PY motif, leading to ubiquitination of both proteins. Smurf2 plays an active role in the repression of TGFbeta signalling, and our data indicate that overexpression of RNF11, through its interaction with Smurf2, can restore TGFbeta responsiveness in transfected cells.
Asunto(s)
Neoplasias de la Mama/genética , Proteínas Portadoras/biosíntesis , Regulación Neoplásica de la Expresión Génica , Neoplasias de la Mama/patología , Proteínas de Unión al ADN , Humanos , Inmunohistoquímica , Invasividad Neoplásica , Análisis de Secuencia por Matrices de Oligonucleótidos , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/fisiología , Transducción de Señal , Transfección , Células Tumorales Cultivadas , Ubiquitina-Proteína Ligasas/farmacología , Dedos de ZincRESUMEN
Vesicular phospholipid gels (VPG), i.e. highly concentrated liposomal dispersions, are suitable for entrapping substances such as anticancer drugs with particular high encapsulation efficiencies (EE). We prepared different formulations of VPG with 30% (w/w) lipid containing 5-fluorouracil (5-FU) by high pressure homogenization and analysed their EE and drug release. Using mixtures of hydrogenated soy phosphatidylcholine and cholesterol with molar ratios ranging from 55/45 to 75/25, a decreasing amount of cholesterol correlated with an increasing EE, which is probably due to a reduced amount of smaller vesicles and number of lamellae. Using a 5-FU solution of pH 8.6 for VPG preparation, an EE of approximately 40% was found after redispersion of the gel to a liposomal dispersion and separation of free drug from liposomal drug by size exclusion chromatography. The reduced EE for preparations with lower pH values was attributed to a fast initial drug release due to the increased drug lipophilicity below the pK(a) value of 8. After redispersion of a VPG of pH 8.0, an initially faster release of about a third of the entrapped drug was found during the first 20 min, followed by stable entrapment over many hours. The rapid initial release may be due to the portion of liposomes smaller than 40 nm in diameter, determined by photon correlation spectroscopy. Cryo electron microscopic pictures show a lentil-like shape of these small liposomes. The membrane defects on the edges are probably the reason for the very high initial drug release rate. The half-life time of the release of 5-FU from intact FU-VPG at both pH 7.4 and 8.0 was found to be in the order of 4-5 h and the kinetics are typical for matrix-controlled drug diffusion. The in vitro data of 5-FU loaded VPG suggest their applicability as implants with controlled release properties or, after redispersion, as intravenously injected liposomal formulations.
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Antimetabolitos Antineoplásicos/química , Fluorouracilo/química , Fosfatidilcolinas/química , Colesterol/química , Cromatografía Líquida de Alta Presión , Microscopía por Crioelectrón , Composición de Medicamentos/métodos , Geles , Concentración de Iones de Hidrógeno , Bombas de Infusión Implantables , Liposomas , Tamaño de la Partícula , Solubilidad , Factores de TiempoRESUMEN
PURPOSE: The in vivo pharmacokinetics (PK), biodistribution and antitumor activity of a new liposomal formulation of gemcitabine (GemLip) were compared to the conventional (clinical) formulation of gemcitabine (GemConv). METHODS: Gemcitabine was entrapped in a vesicular phospholipid gel (VPG) consisting of densely packed liposomes. Redispersed VPG containing GemLip consisted of 33% liposomally entrapped and 67% free gemcitabine. The in vivo efficacies of GemLip and GemConv were compared using the subcutaneously growing human soft tissue sarcoma SXF 1301 and the orthotopically growing human bladder cancer BXF 1299T. PK and biodistribution were evaluated using radiolabeled drug and lipid in SXF 1301 tumor-bearing nude mice. RESULTS: GemLip was highly active in SXF 1301 at a gemcitabine dose of 6-9 mg/kg (days 1, 8 and 15; dose near the MTD). In the 6-mg/kg groups, complete tumor remissions were observed in seven of eight mice. Equimolar doses of GemConv resulted in only moderate tumor growth inhibition. Even at equitoxic doses (360 mg/kg given on days 1, 8 and 15, or 120 mg/kg on days 1, 5 and 8) GemConv was less active than GemLip. Furthermore, GemLip was active in the orthotopically growing BXF 1299T bladder cancer model at 6 mg/kg and prevented distant organ metastasis. In the PK study, GemLip achieved a 35-fold higher plasma AUC (1680 mg x h/ml) than GemConv (47.6 mg x h/ml). The serum half-lives were 0.15 h for free gemcitabine and 13.3 h for liposomal gemcitabine (6 mg/kg each i.v.). Moreover, gemcitabine levels in tumors were fourfold higher following injection of GemLip than following injection of GemConv. CONCLUSIONS: GemLip is a highly effective gemcitabine delivery system which results in superior gemcitabine pharmacodynamics and PK than GemConv. The enhanced in vivo efficacy might be explained by sustained release and passive tumor targeting.
Asunto(s)
Antimetabolitos Antineoplásicos/farmacología , Antimetabolitos Antineoplásicos/farmacocinética , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacología , Desoxicitidina/farmacocinética , Paclitaxel/análogos & derivados , Sarcoma/tratamiento farmacológico , Taxoides , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Animales , Antimetabolitos Antineoplásicos/administración & dosificación , Antineoplásicos Fitogénicos/administración & dosificación , Antineoplásicos Fitogénicos/farmacocinética , Antineoplásicos Fitogénicos/farmacología , Desoxicitidina/administración & dosificación , Docetaxel , Geles , Semivida , Humanos , Liposomas , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Paclitaxel/administración & dosificación , Paclitaxel/farmacocinética , Paclitaxel/farmacología , Fosfolípidos , Distribución Tisular , Trasplante Heterólogo , Vindesina/administración & dosificación , Vindesina/farmacocinética , Vindesina/farmacología , GemcitabinaRESUMEN
A new class of simple synthetic antimitotic compounds based on 2-aroylindoles was discovered. (5-Methoxy-1H-2-indolyl)-phenylmethanone (1) as well as analogous 3-fluorophenyl- (36) and 3-methoxyphenyl (3) derivatives displayed high cytotoxicity of IC(50) = 20 to 75 nM against the human HeLa/KB cervical, SK-OV-3 ovarian, and U373 astrocytoma carcinoma cell lines. The inhibition of proliferation correlated with the arrest in the G2/M phase of the cell cycle. In in vitro assays with tubulin isolated from bovine brain, in general antiproliferative activity correlated with inhibition of tubulin polymerization. Thus, the antimitotic activity of 2-aroylindoles is explained by interference with the mitotic spindle apparatus and destabilization of microtubules. In contrast to colchicine, vincristine, nocodazole, or taxol, 1 did not significantly affect the GTPase activity of beta-tubulin. Interestingly, selected compounds inhibited angiogenesis in the chorioallantoic membrane (CAM) assay. In xenograft experiments, 1 was highly active after oral administration at 200 mg/kg against the human amelanocytic melanoma MEXF 989 in athymic nude mice. We conclude, that 2-aroylindoles constitute an interesting new class of antitubulin agents with the potential to be clinically developed for cancer treatment.
Asunto(s)
Antineoplásicos/síntesis química , Indoles/síntesis química , Tubulina (Proteína)/química , Alantoides/irrigación sanguínea , Inhibidores de la Angiogénesis/síntesis química , Inhibidores de la Angiogénesis/química , Inhibidores de la Angiogénesis/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Biopolímeros , Bovinos , Corion/irrigación sanguínea , Ensayos de Selección de Medicamentos Antitumorales , Fase G2/efectos de los fármacos , GTP Fosfohidrolasas/química , Humanos , Técnicas In Vitro , Indoles/química , Indoles/farmacología , Melanoma/tratamiento farmacológico , Ratones , Ratones Desnudos , Mitosis/efectos de los fármacos , Relación Estructura-Actividad , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
Pharmacogenetic analysis of polymorphisms in drug metabolizing enzymes is currently generating considerable interest as a means of individualizing patient therapy. Recent studies have suggested that patients that are homozygous for a polymorphic variant (a C to T transition at position 609 of the cDNA sequence) of the enzyme NAD(P)H:quinone oxidoreductase (NQO1) may be resistant to Mitomycin C (MMC). Genotyping of a panel of 54 human tumor xenografts by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP), classified tumors as wild type (40/54), heterozygotes (11/54), and homozygous mutants (3/54). Previously, 37 of these tumors had been characterized in terms of their response to MMC in vivo, and in this study, a further nine tumor xenografts have been characterized in terms of their response to MMC. No correlation could be found between the NQO1 polymorphic status of xenografts and their response to MMC in vivo. In terms of genotype/phenotype relationships, NQO1 activity in tumors genotyped as wild type, heterozygotes, and homozygous mutants were 311.1 +/- 421.9 (N = 40), 76.9 +/- 109.5 (N = 11), and 0.2 +/- 0.17 (N = 3) nmol/min/mg, respectively. Genotyping of patients may provide a useful initial step in identifying patients who are unlikely to benefit from quinone-based chemotherapy. In the case of MMC, however, the work presented here demonstrates that genotyping of individuals with respect to NQO1 is unlikely to be beneficial in terms of predicting tumor responses to MMC.
Asunto(s)
Antibióticos Antineoplásicos/uso terapéutico , Mitomicina/uso terapéutico , NADH NADPH Oxidorreductasas/genética , Neoplasias Experimentales/tratamiento farmacológico , Animales , Modelos Animales de Enfermedad , FMN Reductasa , Genotipo , Humanos , Ratones , Ratones Desnudos , NADH NADPH Oxidorreductasas/metabolismo , Trasplante de Neoplasias , Neoplasias Experimentales/enzimología , Polimorfismo Genético , Trasplante Heterólogo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The in vitro antiproliferative activity of an aqueous mistletoe extract (AME) with a defined content of bioactive mistletoe lectin (ML) was tested in 25 human tumor cell lines, including 20 solid and 5 hematological malignancies and 47 human tumor xenografts. The antiproliferative activity of AME was compared to that of the standard cytotoxic agent doxorubicin (CAS 23214-92-8, adriamycin, ADR) using the sulforhodamin B, propidium iodide and soft agar colony forming assays, respectively. AME was highly cytotoxic in solid human tumors with mean IC70 values in the range of 0.17-1 ng ML/ml (2.8-17 pmol bioactive ML). On a molar basis, AME was 3 to 4 logs more potent than ADR and showed differential cytotoxicity towards tumors of the breast, small cell and non-small cell lung, prostate and renal cell cancers. AME was also highly active in hematological malignancies with steep dose response curves resulting in mean IC70 values of 0.12 ng ML/ml (2 pmol). The acute lymphoblastic leukemia cell line HL-60 was the most sensitive, the histiocytic lymphoma cell line U937 the most resistant hematological malignancy. It is important to stress that AME did not induce a biologically relevant increase of cell proliferation in any of the tumor cell lines tested. Our data suggest that AME has in vitro antitumor profiles similar to those of classical anticancer agents. Clear dose-response relationships were found in all of the performed experiments and interesting differential cytotoxicity patterns were observed. Experiments with sensitive tumor types identified in these in vitro studies are currently ongoing in order to demonstrate the anticancer activity of AME in different animal tumor models.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Muérdago/química , Plantas Medicinales , Animales , Antibióticos Antineoplásicos/farmacología , División Celular/efectos de los fármacos , Doxorrubicina/farmacología , Eritrocitos/inmunología , Humanos , Sustancias Intercalantes/farmacología , Trasplante de Neoplasias , Extractos Vegetales/farmacología , Propidio/farmacología , Ovinos/inmunología , Células Tumorales Cultivadas , Ensayo de Tumor de Célula MadreRESUMEN
Mistletoe extracts have been used for decades for non-specific stimulation of the immune system in cancer therapy. Mistletoe lectins (ML) have been identified as the active principle with cytotoxic and immunomodulatory potencies. In the present in vivo experiments, the anticancer effects of an aqueous mistletoe extract (AME) were investigated in different subcutaneously growing syngeneic murine tumors such as Renca renal cell carcinoma, C8 colon 38 carcinoma, F9 testicular carcinoma, B16 melanoma and Lewis lung carcinoma. The animals used were immunocompetent mice of different strains (C57BL/6, BALB/c), depending on the type of tumor tested. After tumor transplantation, the mice were treated with AME at dose levels corresponding to 0, 0.3, 3, 30 or 300 ng ML/kg/d by the i.p. or s.c. route for a maximum of 4 consecutive weeks. The tumor volume was determined by serial caliper measurements and expressed relative to controls. Significant tumor growth inhibition was observed with the Renca , C8 colon 38 and F9 testicular carcinomas at 30 and 300 ng ML/kg/d. These findings were confirmed in independent repeat experiments. No inhibitory effects were seen with the Lewis lung carcinoma and B16 melanoma under the conditions described above. In conclusion, AME showed in vivo anticancer activity in different transplantable syngeneic murine tumor models following repeated parenteral treatment. In view of the low dose levels used, the effects are most likely due to the immunostimulatory rather than to the cytotoxic potencies of AME.
Asunto(s)
Muérdago/química , Muérdago/uso terapéutico , Neoplasias/tratamiento farmacológico , Fitoterapia , Extractos Vegetales/farmacología , Plantas Medicinales , Animales , Relación Dosis-Respuesta a Droga , Femenino , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Neoplasias Experimentales , Factores de Tiempo , Células Tumorales CultivadasRESUMEN
The complex insulin-like growth factor network of ligands, receptors and binding proteins has been shown to be disturbed in breast cancer. In addition to defects in proteins controlling cell cycle checkpoints, this type of aberrations could affect tumor growth and survival thereby influencing both tumor aggressiveness and potential response to treatments. We have previously identified the T1A12/mac25 protein, which is identical to the IGFBP-rP1, as a differentially expressed gene product in breast cancer cells compared with normal cells. Here we compare the expression of IGFBP-rP1 in 106 tumor samples with known status of cell cycle aberrations and other clinicopathological data. This was done using a tumor tissue section array system that allows for simultaneous immunohistochemical staining of all samples in parallel. Cytoplasmic staining of variable intensity was observed in most tumors, 15% lacked IGFBP-rP1 staining completely, 20% had weak staining, 32% intermediate and 33% showed strong staining. Low IGFBP-rP1 was associated with high cyclin E protein content, retinoblastoma protein (pRb) inactivation, low bcl-2 protein, poorly differentiated tumors and higher stage. There was a significantly impaired prognosis for patients with low IGFBP-rP1 protein tumors. Interestingly, IGFBP-rP1 showed an inverse association with proliferation (Ki-67%) in estrogen receptor negative tumors as well as in cyclin E high tumors suggesting a separate cell cycle regulatory function for IGFBP-rP1 independent of interaction with the estrogen receptor or the pRb pathway.