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PURPOSE: A first of its kind experimental verification of dose-averaged linear energy transfer (LETd) optimized treatment plans for proton therapy has been carried out using a silicon-on-insulator microdosimeter at the Massachusetts General Hospital (MGH), Boston, USA. METHODS AND MATERIALS: Three clinical treatment plans of a typical ependymoma structure set were designed using the standard clinical approach, the proposed protocol approach, and a one-field approach. The plans were then reoptimized to reduce the LETd-weighted dose in the brain stem. All six plans were delivered in a solid water phantom and the experimental yDâ¾ measured. RESULTS: After LETd optimization, a reduction in yDâ¾ was found within the brain stem by an average of 12%, 19%, and 4% for the clinical, protocol, and one-field plans, respectively, while maintaining adequate coverage of the tumor structure. The experimental LETd-weighted doses were in agreement with the treatment planning system calculations and Monte Carlo simulations and reinforced the improvement of the optimization. CONCLUSIONS: This work demonstrates the first experimental verification of the clinical implementation of LETd optimization for patient treatment with proton therapy.
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Neoplasias Encefálicas , Ependimoma , Transferencia Lineal de Energía , Método de Montecarlo , Fantasmas de Imagen , Terapia de Protones , Dosificación Radioterapéutica , Planificación de la Radioterapia Asistida por Computador , Planificación de la Radioterapia Asistida por Computador/métodos , Humanos , Terapia de Protones/métodos , Neoplasias Encefálicas/radioterapia , Neoplasias Encefálicas/diagnóstico por imagen , Ependimoma/radioterapia , Ependimoma/diagnóstico por imagen , Radiometría/métodos , Radiometría/instrumentación , Tronco Encefálico/diagnóstico por imagen , Silicio , Órganos en Riesgo/efectos de la radiaciónRESUMEN
The use of field-specific apertures, routine in scattered or uniform-scanned proton fields, are still a necessity in pencil-beam scanned (PBS) fields to sharpen the penumbral edge at low energies and in high fraction dose application beyond that achievable with small spot size. We describe a model implemented in our clinical pencil-beam algorithm that models the insertion of a shaped aperture, including shapes adapted per energy layer such as may be achieved with a multi-leaf collimator. The model decomposes the spot transport into discrete steps. The first step transport a uniform intensity field of high-resolution sub-pencil-beams at the layer energy through the medium. This transport only considers primary scattering in both the patient and an optional range-shifter. The second step models the aperture areas and edge penumbral transition as a modulation of the uniform intensity. The third step convolves individual steps over the uniform-transported field including the aperture-modified intensities. We also introduce an efficient model based on a Clarkson sector integration for nuclear scattered halo protons. This avoids the explicit modeling of long range halo protons to the detriment of computational efficiency in calculation and optimization. We demonstrate that the aperture effect is primarily due to in-patient and shifter scattering with a small contribution from the apparent beam source position. The model provides insight into the primary physics contributions to the penumbra and the nuclear halo. The model allowed us to fully deploy our PBS capacity at our two-gantry center without which PBS treatments would have been inferior compared to scattered fields with apertures. Finally, Monte Carlo calculations have (nearly) replaced phenomenological pencil-beam models for collimated fields. Phenomenological models do, however, allow exposition of underlying clinical phenomena and closer connection to representative clinical observables.
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Terapia de Protones , Protones , Algoritmos , Humanos , Dosificación Radioterapéutica , Planificación de la Radioterapia Asistida por ComputadorRESUMEN
BACKGROUND: In scanned proton beam therapy systematic deviations in spot size at iso-center can occur as a result of changes in the beam-line optics. There is currently no general guideline of the spot size accuracy required clinically. In this work we quantify treatment plan robustness to systematic spot size variations as a function of spot size and spot spacing, and we suggest guidelines for tolerance levels for spot size variations. METHODS: Through perturbation of spot size in treatment plans for 7 patients and a phantom, we evaluated the dose impact of systematic spot size variations of 5% up to 50%. We investigated the dependence on nominal spot size by studying scenarios with small, medium and large spot sizes for various inter-spot spacings. To come to tolerance levels, we used the Γ passing rate and dose-volume-histograms. RESULTS: Limits on spot size accuracy were extracted for 8 sites, 3 different spot sizes and 3 different inter-spot spacings. While the allowable spot size variation strongly depends on the spot size, the inter-spot spacing turned out to be only of limited influence. CONCLUSIONS: Plan robustness to spot size variations strongly depend on spot size, with small spot plans being much more robust than larger spots plans. Inter-spot spacing did not influence plan robustness. Combining our results with existing literature, we propose limits of ±25%, ±20% and ±10% of the spot width σ, for spots with σ of 2.5, 5.0 and 10â¯mm in proton therapy spot scanning facilities, respectively.
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Terapia de Protones/métodos , Dosis de Radiación , Humanos , Fantasmas de Imagen , Dosificación Radioterapéutica , Planificación de la Radioterapia Asistida por ComputadorRESUMEN
OBJECTIVE: To compare the recurrence rates after complete response to topical treatment with either cidofovir or imiquimod for vulval intraepithelial neoplasia (VIN) 3. DESIGN: A prospective, open, randomised multicentre trial. SETTING: 32 general hospitals located in Wales and England. POPULATION OR SAMPLE: 180 patients were randomised consecutively between 21 October 2009 and 11 January 2013, 89 to cidofoovir (of whom 41 completely responded to treatment) and 91 to imiquimod (of whom 42 completely responded to treatment). METHODS: After 24 weeks of treatment, complete responders were followed up at 6-monthly intervals for 24 months. At each visit, the Common Terminology Criteria for Adverse Events (CTCAE) v3.0 was assessed and any new lesions were biopsied for histology. MAIN OUTCOME MEASURES: Time to histologically confirmed disease recurrence (any grade of VIN). RESULTS: The median length of follow up was 18.4 months. At 18 months, more participants were VIN-free in the cidofovir arm: 94% (95% CI 78.2-98.5) versus 71.6% (95% CI 52.0-84.3) [univariable hazard ratio (HR) 3.46, 95% CI 0.95-12.60, P = 0.059; multivariable HR 3.53, 95% CI 0.96-12.98, P = 0.057). The number of grade 2+ events was similar between treatment arms (imiquimod: 24/42 (57%) versus cidofovir: 27/41 (66%), χ2 = 0.665, P = 0.415), with no grade 4+. CONCLUSIONS: Long-term data indicates a trend towards response being maintained for longer following treatment with cidofovir than with imiquimod, with similar low rates of adverse events for each drug. Adverse event rates indicated acceptable safety of both drugs TWEETABLE ABSTRACT: Long-term follow up in the RT3VIN trial suggests cidofovir may maintain response for longer than imiquimod.
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Antineoplásicos/administración & dosificación , Carcinoma in Situ/tratamiento farmacológico , Cidofovir/administración & dosificación , Imiquimod/administración & dosificación , Recurrencia Local de Neoplasia/epidemiología , Neoplasias de la Vulva/tratamiento farmacológico , Administración Tópica , Antineoplásicos/efectos adversos , Carcinoma in Situ/epidemiología , Carcinoma in Situ/patología , Cidofovir/efectos adversos , Femenino , Humanos , Imiquimod/efectos adversos , Persona de Mediana Edad , Recurrencia Local de Neoplasia/etiología , Recurrencia Local de Neoplasia/patología , Estudios Prospectivos , Resultado del Tratamiento , Neoplasias de la Vulva/epidemiología , Neoplasias de la Vulva/patologíaRESUMEN
We hypothesized that IV busulfan (Bu) dosing could be safely intensified through pharmacokinetic (PK-) dose guidance to minimize the inter-patient variability in systemic exposure (SE) associated with body-sized dosing, and that this should improve outcome of AML/MDS patients undergoing allogeneic stem cell transplantation. To test this hypothesis, we treated 218 patients (median age 50.7 years, male/female 50/50%) with fludarabine 40 mg/m2 once daily x4, each dose followed by IV Bu, randomized to 130 mg/m2 (N=107) or PK-guided to average daily SE, AUC of 6000 µM min (N=111), stratified for remission status and allo-grafting from HLA-matched donors. Toxicity and GvHD rates in the groups were similar; the risk of relapse or treatment-related mortality remained higher in the fixed-dose group throughout the 80-month observation period. Further, PK-guidance yielded safer disease control, leading to improved overall and PFS, most prominently in MDS patients and in AML patients not in remission at allogeneic stem cell transplantation. We conclude that AML/MDS patients receiving pretransplant conditioning treatment with our 4-day regimen may benefit significantly from PK-guided Bu dosing. This could be considered an alternative to fixed-dose delivery since it provides the benefit of precise dose delivery to a predetermined SE without increasing risk(s) of serious toxicity and/or GvHD.
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Busulfano/administración & dosificación , Monitoreo de Drogas/métodos , Leucemia Mieloide Aguda/terapia , Síndromes Mielodisplásicos/terapia , Acondicionamiento Pretrasplante/métodos , Trasplante Homólogo/métodos , Vidarabina/análogos & derivados , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/toxicidad , Busulfano/farmacocinética , Busulfano/toxicidad , Femenino , Enfermedad Injerto contra Huésped/etiología , Trasplante de Células Madre Hematopoyéticas/métodos , Trasplante de Células Madre Hematopoyéticas/mortalidad , Humanos , Leucemia Mieloide Aguda/mortalidad , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/mortalidad , Recurrencia , Análisis de Supervivencia , Acondicionamiento Pretrasplante/mortalidad , Trasplante Homólogo/mortalidad , Resultado del Tratamiento , Vidarabina/administración & dosificaciónRESUMEN
This study investigates whether 'pencil beam resampling', i.e. iterative selection and weight optimization of randomly placed pencil beams (PBs), reduces optimization time and improves plan quality for multi-criteria optimization in intensity-modulated proton therapy, compared with traditional modes in which PBs are distributed over a regular grid. Resampling consisted of repeatedly performing: (1) random selection of candidate PBs from a very fine grid, (2) inverse multi-criteria optimization, and (3) exclusion of low-weight PBs. The newly selected candidate PBs were added to the PBs in the existing solution, causing the solution to improve with each iteration. Resampling and traditional regular grid planning were implemented into our in-house developed multi-criteria treatment planning system 'Erasmus iCycle'. The system optimizes objectives successively according to their priorities as defined in the so-called 'wish-list'. For five head-and-neck cancer patients and two PB widths (3 and 6 mm sigma at 230 MeV), treatment plans were generated using: (1) resampling, (2) anisotropic regular grids and (3) isotropic regular grids, while using varying sample sizes (resampling) or grid spacings (regular grid). We assessed differences in optimization time (for comparable plan quality) and in plan quality parameters (for comparable optimization time). Resampling reduced optimization time by a factor of 2.8 and 5.6 on average (7.8 and 17.0 at maximum) compared with the use of anisotropic and isotropic grids, respectively. Doses to organs-at-risk were generally reduced when using resampling, with median dose reductions ranging from 0.0 to 3.0 Gy (maximum: 14.3 Gy, relative: 0%-42%) compared with anisotropic grids and from -0.3 to 2.6 Gy (maximum: 11.4 Gy, relative: -4%-19%) compared with isotropic grids. Resampling was especially effective when using thin PBs (3 mm sigma). Resampling plans contained on average fewer PBs, energy layers and protons than anisotropic grid plans and more energy layers and protons than isotropic grid plans. In conclusion, resampling resulted in improved plan quality and in considerable optimization time reduction compared with traditional regular grid planning.
Asunto(s)
Terapia de Protones/métodos , Planificación de la Radioterapia Asistida por Computador/métodos , Radioterapia de Intensidad Modulada/métodos , Anisotropía , Humanos , Órganos en Riesgo/efectos de la radiación , Neoplasias Orofaríngeas/radioterapia , Terapia de Protones/efectos adversos , Radioterapia de Intensidad Modulada/efectos adversosRESUMEN
CONTEXT: The European Association of Urology (EAU) guidelines on urinary incontinence published in March 2012 have been rewritten based on an independent systematic review carried out by the EAU guidelines panel using a sustainable methodology. OBJECTIVE: We present a short version here of the full guidelines on the surgical treatment of patients with urinary incontinence, with the aim of dissemination to a wider audience. EVIDENCE ACQUISITION: Evidence appraisal included a pragmatic review of existing systematic reviews and independent new literature searches based on Population, Intervention, Comparator, Outcome (PICO) questions. The appraisal of papers was carried out by an international panel of experts, who also collaborated in a series of consensus discussions, to develop concise structured evidence summaries and action-based recommendations using a modified Oxford system. EVIDENCE SUMMARY: The full version of the guidance is available online (www.uroweb.org/guidelines/online-guidelines/). The guidance includes algorithms that refer the reader back to the supporting evidence and have greater accessibility in daily clinical practice. Two original meta-analyses were carried out specifically for these guidelines and are included in this report. CONCLUSIONS: These new guidelines present an up-to-date summary of the available evidence, together with clear clinical algorithms and action-based recommendations based on the best available evidence. Where high-level evidence is lacking, they present a consensus of expert panel opinion.
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Incontinencia Urinaria/cirugía , Algoritmos , Femenino , Humanos , Masculino , Procedimientos Quirúrgicos Urológicos/normasRESUMEN
CONTEXT: The previous European Association of Urology (EAU) guidelines on urinary incontinence comprised a summary of sections of the 2009 International Consultation on Incontinence. A decision was made in 2010 to rewrite these guidelines based on an independent systematic review carried out by the EAU guidelines panel, using a sustainable methodology. OBJECTIVE: We present a short version of the full guidelines on assessment, diagnosis, and nonsurgical treatment of urinary incontinence, with the aim of increasing their dissemination. EVIDENCE ACQUISITION: Evidence appraisal included a pragmatic review of existing systematic reviews and independent new literature searches, based on Population, Intervention, Comparator, Outcome questions. Appraisal of papers was carried out by an international panel of experts, who also collaborated on a series of consensus discussions, to develop concise structured evidence summaries and action-based recommendations using a modified Oxford system. EVIDENCE SUMMARY: The full version of the guidelines is available online (http://www.uroweb.org/guidelines/online-guidelines/). The guidelines include algorithms that refer the reader back to the supporting evidence, and they are more immediately useable in daily clinical practice. CONCLUSIONS: These new guidelines present an up-to-date summary of the available evidence, together with clear clinical algorithms and action-based recommendations based on the best available evidence. Where such evidence does not exist, they present a consensus of expert opinion.
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Incontinencia Urinaria/terapia , Algoritmos , Humanos , Incontinencia Urinaria/diagnóstico , Incontinencia Urinaria/tratamiento farmacológicoRESUMEN
We investigated the administration of i.v. BU combined with melphalan (Mel) in patients with ALL undergoing allogeneic hematopoietic SCT. Forty-seven patients with a median age of 33 years (range 20-61) received a matched sibling (n=27) or matched unrelated donor transplant (n=20) for ALL in first CR (n=26), second CR (n=13), or with more advanced disease (n=8). BU was infused daily for 4 days, either at a fixed dose of 130 mg/m² (5 patients) or using pharmacokinetic (PK) dose adjustment (42 patients), to target an average daily area-under-the-curve (AUC) of 5000 µmol/min, determined by a test dose of i.v. BU at 32 mg/m². This was followed by a rest day, then two daily doses of Mel at 70 mg/m². Stem cells were infused on the following day. The 2-year OS, PFS and non-relapse mortality (NRM) rates were 35% (95% confidence interval (CI), 23-51%), 31% (95% CI, 21-48%) and 37% (95% CI, 23-50%), respectively. Acute NRM at 100 days was favorable at 12% (95% CI, 5-24%); however, the 2-year NRM was significantly higher for patients older than 40 years, 58% vs 20%, mainly due to GVHD.
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Antineoplásicos Alquilantes/uso terapéutico , Busulfano/uso terapéutico , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Melfalán/uso terapéutico , Agonistas Mieloablativos/uso terapéutico , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Acondicionamiento Pretrasplante , Adulto , Factores de Edad , Antineoplásicos Alquilantes/administración & dosificación , Antineoplásicos Alquilantes/efectos adversos , Busulfano/administración & dosificación , Busulfano/efectos adversos , Quimioterapia Combinada/efectos adversos , Femenino , Estudios de Seguimiento , Enfermedad Injerto contra Huésped/epidemiología , Enfermedad Injerto contra Huésped/etiología , Enfermedad Injerto contra Huésped/prevención & control , Humanos , Incidencia , Infusiones Intravenosas , Masculino , Melfalán/efectos adversos , Persona de Mediana Edad , Agonistas Mieloablativos/administración & dosificación , Agonistas Mieloablativos/efectos adversos , Leucemia-Linfoma Linfoblástico de Células Precursoras/prevención & control , Prevención Secundaria , Análisis de Supervivencia , Texas , Acondicionamiento Pretrasplante/efectos adversos , Trasplante Homólogo , Adulto JovenRESUMEN
We present a data acquisition system to perform on-the-fly background subtraction and lower-level discrimination compression of streaming x-ray photon correlation spectroscopy data from a fast charge-coupled device (CCD) area detector. The system is built using a commercial frame grabber with an on-board field-programmable gate array. The system is capable of continuously processing at least 60 CCD frames per second each consisting of 1024 × 1024 16-bit pixels with â² 15,000 photon hits per frame at a maximum compression factor of ≈95%.
RESUMEN
Dental caries and periodontal disease are the most common oral conditions experienced by adults today. The treatment of these diseases by the dental team can only be performed when patients attend dental practices. There is recognition that the preventive measures patients perform at home between dental visits is of vital importance in the control of these diseases. Water fluoridation and fluoridated toothpastes have made enormous progress into the prevention of dental caries worldwide. However, prevention of periodontal disease is yet to enjoy the same success. A number of toothpastes have been developed for the prevention and control of periodontal disease. One such toothpaste - containing triclosan/copolymer - has been thoroughly researched. The literature pertaining to the efficacy, mode of action and safety of triclosan/copolymer toothpaste has been reviewed. A MEDLINE search identified 198 articles dated from 1989 to 2008. The findings of this body of research are discussed and conclusions regarding the efficacy of triclosan/copolymer toothpaste in the home-care management of periodontal disease are presented.
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Antiinfecciosos Locales/uso terapéutico , Enfermedades Periodontales/prevención & control , Pastas de Dientes/uso terapéutico , Triclosán/uso terapéutico , Adulto , Antiinfecciosos Locales/administración & dosificación , Preparaciones de Acción Retardada , Placa Dental/prevención & control , Gingivitis/prevención & control , Humanos , Maleatos , Higiene Bucal , Periodontitis/prevención & control , Vehículos Farmacéuticos , Polietilenos , Seguridad , Pastas de Dientes/administración & dosificación , Resultado del Tratamiento , Triclosán/administración & dosificaciónRESUMEN
Bone loss is a feature of both periodontitis and osteoporosis, and periodontal destruction may be influenced by systemic bone loss. This study evaluated the association between periodontal disease and bone mineral density (BMD) in a cohort of 1347 (137 edentulous) older men followed for an average of 2.7 years. Participants were recruited from the Osteoporotic Fractures in Men Study. Random half-mouth dental measures included clinical attachment loss (CAL), pocket depth (PD), calculus, plaque, and bleeding. BMD was measured at the hip, spine, and whole-body, by dual-energy x-ray absorptiometry, and at the heel by ultrasound. After adjustment for age, smoking, race, education, body mass index, and calculus, there was no association between number of teeth, periodontitis, periodontal disease progression, and either BMD or annualized rate of BMD change. We found little evidence of an association between periodontitis and skeletal BMD among older men.
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Densidad Ósea , Osteoporosis/fisiopatología , Periodontitis/fisiopatología , Pérdida de Diente/fisiopatología , Absorciometría de Fotón , Anciano , Calcáneo/diagnóstico por imagen , Estudios Transversales , Progresión de la Enfermedad , Cadera/diagnóstico por imagen , Humanos , Masculino , Estudios Prospectivos , UltrasonografíaRESUMEN
Signal transducer and activator of transcription-3 (STAT3) is constitutively activated in a variety of cancer types, including malignant gliomas. STAT3 is activated by phosphorylation of a tyrosine residue, after which it dimerizes and translocates into the nucleus. There it regulates the expression of several genes responsible for proliferation and survival at the transcriptional level. A selective inhibitor of STAT3 phosphorylation, AG490, has been shown to inhibit growth and induce apoptosis in some cancer cell types. However, although AG490 routinely shows in vitro anticancer activity, it has not consistently demonstrated an in vivo anticancer effect in animal models. Here, we have tested WP1066, a novel inhibitor structurally related to AG490 but significantly more potent and active, against human malignant glioma U87-MG and U373-MG cells in vitro and in vivo. IC(50) values for WP1066 were 5.6 muM in U87-MG cells and 3.7 muM in U373-MG cells, which represents 18-fold and eightfold increases in potency, respectively, over that of AG490. WP1066 activated Bax, suppressed the expression of c-myc, Bcl-X(L) and Mcl-1, and induced apoptosis. Systemic intraperitoneal administration of WP1066 in mice significantly (P<0.001) inhibited the growth of subcutaneous malignant glioma xenografts during the 30-day follow-up period. Immunohistochemical analysis of the excised tumors revealed that phosphorylated STAT3 levels in the WP1066 treatment group remained inhibited at 3 weeks after the final WP1066 injection, whereas tumors from the control group expressed high levels of phosphorylated STAT3. We conclude that WP1066 holds promise as a therapeutic agent against malignant gliomas.
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Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Glioma/tratamiento farmacológico , Glioma/patología , Piridinas/farmacología , Factor de Transcripción STAT3/antagonistas & inhibidores , Transducción de Señal/efectos de los fármacos , Tirfostinos/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Humanos , Piridinas/química , Factor de Transcripción STAT3/fisiología , Transducción de Señal/fisiología , Tirfostinos/químicaRESUMEN
BACKGROUND: No longitudinal studies have tracked cognitive performance through the menopausal transition and thus the impact of the transition on cognition, independent of aging, is not known. The authors hypothesized that a decline in cognitive functioning occurs as women progress through the menopausal transition, independent of age, educational level, family income, ethnicity, and baseline self-perceived health. METHOD: The authors began a population-based, longitudinal study in January 1996 with yearly follow-up interviews. This report includes follow-up through November 2001. The authors randomly selected African American and white women from a census of two contiguous Chicago communities. After screening for eligibility (age 42 to 52 years, premenopausal or early perimenopausal, no exogenous hormone use in the past 3 months, and no hysterectomy), 868 agreed to participate. Women who became pregnant, had a hysterectomy, or began using hormones were censored from that time onward. This study reports on 803 women for whom cognitive assessments were available. The authors assessed working memory (Digit Span Backward) and perceptual speed (Symbol Digit Modalities Test). RESULTS: Contrary to the hypothesis, the authors found small but significant increases over time during the premenopausal and perimenopausal phases. This trend was not accounted for by chronological age, education, family income, ethnicity, or baseline self-perceived health. CONCLUSIONS: Transition through menopause is not accompanied by a decline in working memory and perceptual speed.
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Cognición/fisiología , Menopausia/psicología , Adulto , Envejecimiento/psicología , Población Negra/psicología , Chicago/epidemiología , Estudios de Cohortes , Escolaridad , Femenino , Estudios de Seguimiento , Humanos , Renta , Estudios Longitudinales , Memoria/fisiología , Persona de Mediana Edad , Posmenopausia/psicología , Factores Socioeconómicos , Población Blanca/psicologíaRESUMEN
Although many estrogen receptor-positive breast cancers initially respond to antihormones, responses are commonly incomplete with resistance ultimately emerging. Delineation of signaling mechanisms underlying these phenomena would allow development of therapies to improve antihormone response and compromise resistance. This in vitro investigation in MCF-7 breast cancer cells examines whether epidermal growth factor receptor (EGFR) signaling limits antiproliferative and proapoptotic activity of antihormones and ultimately supports development of resistance. It addresses whether the anti-EGFR agent gefitinib (ZD1839/Iressa; TKI: 1 mum) combined with the antihormones 4-hydroxytamoxifen (TAM: 0.1 mum) or fulvestrant (Faslodex; 0.1 mum) enhances growth inhibition and prevents resistance. TAM significantly suppressed MCF-7 growth over wk 2-5, reducing proliferation detected by immunocytochemistry and fluorescence-activated cell sorter cell cycle analysis. A modest apoptotic increase was observed by fluorescence-activated cell sorter and fluorescence microscopy, with incomplete bcl-2 suppression. EGFR induction occurred during TAM response, as measured by immunocytochemistry and Western blotting, with EGFR-positive, highly proliferative resistant growth subsequently emerging. Although TKI alone was ineffective on growth, TAM plus TKI cotreatment exhibited superior antigrowth activity vs. TAM, with no viable cells by wk 12. Cotreatment was more effective in inhibiting proliferation, promoting apoptosis, and eliminating bcl-2. Cotreatment blocked EGFR induction, markedly depleted ERK1/2 MAPK and protein kinase B phosphorylation, and prevented emergence of EGFR-positive resistance. Faslodex plus TKI cotreatment was also a superior antitumor strategy. Thus, increased EGFR evolves during treatment with antihormones, limiting their efficacy and promoting resistance. Gefitinib addition to antihormonal therapy could prove more effective in treating estrogen receptor-positive breast cancer and may combat development of resistance.
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Antineoplásicos Hormonales/farmacología , Neoplasias de la Mama/patología , Neoplasias de la Mama/fisiopatología , Factor de Crecimiento Epidérmico/antagonistas & inhibidores , Estradiol/análogos & derivados , Estradiol/farmacología , Antagonistas de Estrógenos/farmacología , Proteínas Serina-Treonina Quinasas , Quinazolinas/farmacología , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacología , Apoptosis/efectos de los fármacos , División Celular/efectos de los fármacos , Línea Celular Tumoral , Combinación de Medicamentos , Resistencia a Medicamentos/efectos de los fármacos , Sinergismo Farmacológico , Receptores ErbB/metabolismo , Receptores ErbB/fisiología , Femenino , Fulvestrant , Gefitinib , Humanos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , Transducción de Señal/efectos de los fármacosRESUMEN
A validated method is described for the simultaneous analysis of PGE2, 11-, 12-, and 5-HETEs from cultured cells using HPLC negative electrospray ionization tandem mass spectrometry (LC/MS/MS). This method permits quantification of selected individual arachidonic acid metabolites from cell extracts without derivatization, multiple purification steps, or lengthy separation times required by traditional GC-MS- or HPLC-UV -based methods. Accuracy assessments of values calculated using this method showed deviations from nominal values were < or =15%. An average relative deviation of 7% of mean calculated values was observed for values taken on separate days. The lower limit of detection for all metabolites was 1.3 pg. The method was used to quantify arachidonic acid metabolites present in various cancer cell lines after incubation with arachidonic acid and the selective cyclooxygenase-2 inhibitor celecoxib. Results showed that the presence of celecoxib in lung cancer A549 cells reduced production of both PGE2 and 11-HETE in a concentration-dependent manner.
Asunto(s)
Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/análisis , Dinoprostona/análisis , Ácidos Hidroxieicosatetraenoicos/análisis , Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/metabolismo , Animales , Ácido Araquidónico/metabolismo , Celecoxib , Cromatografía Líquida de Alta Presión/métodos , Ciclooxigenasa 2 , Dinoprostona/metabolismo , Humanos , Ácidos Hidroxieicosatetraenoicos/metabolismo , Isoenzimas/antagonistas & inhibidores , Leucemia/metabolismo , Lipooxigenasa/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas de la Membrana , Prostaglandina-Endoperóxido Sintasas/metabolismo , Pirazoles , Ratas , Espectrometría de Masa por Ionización de Electrospray/métodos , Sulfonamidas/metabolismo , Sulfonamidas/farmacología , Células Tumorales Cultivadas/citología , Células Tumorales Cultivadas/metabolismoRESUMEN
UCN-01 is a naturally derived anticancer agent isolated in the culture broth of actinomyces streptomyces. We have developed a sensitive high-performance liquid chromatographic method for the determination of UCN-01 in human plasma. UCN-01 was isolated from human plasma after intravenous administration, by using 100% ice-cold acetonitrile liquid-liquid phase extraction. Liquid chromatographic separation was achieved by isocratic elution on a phenyl analytical column. The mobile phase consisted of acetonitrile-0.5 M ammonium acetate (45:55) with 0.2% triethylamine added as a modifier. The UCN-01 peak was identified from other peaks using fluorescence excitation energy and emission energy wavelengths of 310 and 410 nm, respectively. Retention time for UCN-01 was 4.2 +/- 0.5 min. The UCN-01 peak was baseline resolved, with nearest peak at 2.6 min distance. No interfering peaks were observed at the retention time of UCN-01. Peak area amounts from extracted samples were proportional over the dynamic concentration range used: 0.2 to 30 microg/ml. Mean recoveries of UCN-01 at concentrations of 0.5 and 25 microg/ml were 89 and 90.2%, respectively. Relative standard deviations for UCN-01 calibration standards ranged from 1.89 to 2.31%, with relative errors ranging from 0.3 to 11.6%. Assay precision for UCN-01 based on quality control samples of 0.50 microg/ml was +/- 4.86% with an accuracy of +/-5.7%. For drug extracted from plasma the lowest limit of detection was 0.1 microg/ml, with the lowest limit of quantitation being 0.2 microg/ml. This method is suitable for routine analysis of UCN-01 in human plasma at concentration from 0.2 to 30 microg/ml.
Asunto(s)
Alcaloides/sangre , Antineoplásicos/sangre , Cromatografía Líquida de Alta Presión/métodos , Calibración , Humanos , Estándares de Referencia , Sensibilidad y Especificidad , Espectrometría de Fluorescencia , Estaurosporina/análogos & derivadosRESUMEN
Karenitecin is a novel, highly lipophilic camptothecin derivative with potent anticancer potential. We have developed a sensitive high-performance liquid chromatographic method for the determination of karenitecin concentration in human plasma and urine. Karenitecin was isolated from human plasma and urine using solid-phase extraction. Separation was achieved by gradient elution, using a water and acetonitrile mobile phase, on an ODS analytical column. Karenitecin was detected using fluorescence detection at excitation and emission wavelengths of 370 and 490 nm, respectively. Retention time for karenitecin was 16.2 +/- 0.5 min and 8.0 +/- 0.2 min for camptothecin, the internal standard. The karenitecin peak was baseline resolved, with the nearest peak at 3.1 min distance. Using normal volunteer plasma and urine from multiple individuals, as well as samples from the 50 patients analyzed to date, no interfering peaks were detected. Inter- and intra-day coefficients of variance were <4.4 and 7.1% for plasma and <4.9 and 11.6% for urine. Assay precision, based on an extracted karenitecin standard plasma sample of 2.5 ng/ml, was +4.46% with a mean accuracy of 92.4%. For extracted karenitecin standard urine samples of 2.5 ng/ml assay precision was +2.35% with a mean accuracy of 99.5%. The mean recovery of karenitecin, at plasma concentrations of 1.0 and 50 ng/ml, was 81.9 and 87.8% respectively. In urine, at concentrations of 1.5 and 50 ng/ml, the mean recoveries were 90.3 and 78.4% respectively. The lower limit of detection (LLD) for karenitecin was 0.5 ng/ml in plasma and 1.0 ng/ml in urine. The lower limit of quantification (LLQ) for karenitecin was 1 ng/ml and 1.5 ng/ml for plasma and urine, respectively. Stability studies indicate that when frozen at -70 degrees C, karenitecin is stable in human plasma for up to 3 months and in human urine for up to 1 month. This method is useful for the quantification of karenitecin in plasma and urine samples for clinical pharmacology studies in patients receiving this agent in clinical trials.
Asunto(s)
Antineoplásicos/sangre , Camptotecina/análogos & derivados , Camptotecina/sangre , Cromatografía Líquida de Alta Presión/métodos , Inhibidores Enzimáticos/sangre , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Espectrometría de FluorescenciaRESUMEN
Programmable fusogenic vesicles (PFV) are liposomes composed of non-bilayer lipid components stabilized by the inclusion of an exchangeable poly(ethylene glycol) (PEG)-lipid conjugate. Vesicle destabilization by loss of the PEG-lipid results in recovery of the inherent fusogenic character. As a result, PFV can be designed to display a long circulation lifetime after i.v. administration, high accumulation at disease sites and full bioavailability of an encapsulated compound. In the present study, we investigated the potential application of PFV as carriers for intracellular delivery of antisense oligodeoxynucleotides (ODN). Antisense phosphorothioate ODN were encapsulated into PFV containing dioleoylphosphatidylethanolamine, cholesterol, dioleyldimethylammonium chloride and PEG-ceramides with different carbon chain length (C(8), C(14) and C(20)). In vitro fluorescent microscopy and flow cytometry analysis demonstrated that PFV containing PEG-ceramide C(14) provided enhanced intracellular delivery of FITC-labelled antisense ODN compared to PFV displaying faster or slower rates of destabilization (containing PEG-ceramide C(8) or C(20), respectively). Therapeutic efficacy of PFV-encapsulated antisense ODN against two proto-oncogenes, c-myc and bcl-2, was examined in various cell lines. At antisense concentrations of 0.5 microM, no significant downregulation of c-myc mRNA levels was observed in HEK293, B16 and MCA207 cells. However, treatment of 518A2 melanoma cells with PFV-encapsulated antisense targeting bcl-2 at concentrations of 0.5 microM and 1.0 microM resulted in reduced bcl-2 mRNA level by about 20% and 25% after 48 h incubation. Free antisense ODN did not affect bcl-2 mRNA expression at the concentrations used in this study and encapsulated control antisense (reverse polarity) led to a non-specific increase in mRNA levels. Our results suggest that PFV carriers displaying appropriate rates of destabilization have the potential to act as intracellular delivery vehicles and may improve the bioavailability and potency of antisense oligonucleotides.
Asunto(s)
Células/efectos de los fármacos , Liposomas , Oligonucleótidos Antisentido/farmacología , Animales , Línea Celular , Células/metabolismo , Medio de Cultivo Libre de Suero , Regulación hacia Abajo , Fluoresceína-5-Isotiocianato , Colorantes Fluorescentes , Genes bcl-2 , Genes myc , Técnicas Genéticas , Humanos , Ratones , Oligonucleótidos Antisentido/administración & dosificación , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales CultivadasRESUMEN
PSI-BLAST is an iterative program to search a database for proteins with distant similarity to a query sequence. We investigated over a dozen modifications to the methods used in PSI-BLAST, with the goal of improving accuracy in finding true positive matches. To evaluate performance we used a set of 103 queries for which the true positives in yeast had been annotated by human experts, and a popular measure of retrieval accuracy (ROC) that can be normalized to take on values between 0 (worst) and 1 (best). The modifications we consider novel improve the ROC score from 0.758 +/- 0.005 to 0.895 +/- 0.003. This does not include the benefits from four modifications we included in the 'baseline' version, even though they were not implemented in PSI-BLAST version 2.0. The improvement in accuracy was confirmed on a small second test set. This test involved analyzing three protein families with curated lists of true positives from the non-redundant protein database. The modification that accounts for the majority of the improvement is the use, for each database sequence, of a position-specific scoring system tuned to that sequence's amino acid composition. The use of composition-based statistics is particularly beneficial for large-scale automated applications of PSI-BLAST.