RESUMO
Dysregulated expression or activation of matrix metalloproteinases (MMPs) is observed in many kinds of life-threatening diseases. Therefore, MMP imaging-for example, with radiolabeled MMP inhibitors (MMPIs)-potentially represents a valuable tool for clinical diagnostics using noninvasive single photon emission computed tomography (SPECT) or positron emission tomography (PET) imaging. Despite numerous preclinical imaging approaches, translation to a clinical setting has not yet been successful. We introduce and oppose three potential radiofluorinated MMP-targeted imaging probes, modified by the introduction of pentamethine cyanine (Cy5) dyes and therefore containing both radio- as well as fluorescent label with respect to their capability to assess MMP activity in vivo by means of scintigraphic (PET) and/or fluorescent (NIRF) imaging. New hybrid MMPI tracer candidates, structurally based on radiofluorinated pyrimidine-2,4,6-triones (barbiturates) from previous approaches, were synthesized by convenient two-step syntheses. In the first step, Cy5 dyes, varying in the number of sulfonate groups (nSO3- = 1, 2, or 4) and bearing an additional "clickable" alkyne moiety, were coupled to the barbiturate MMPI by amide formation. In the second step, the [18F]fluoride radiolabel was introduced into the resulting Cy5 dye conjugates by "radio-click" chemistry. Biodistribution studies of these hybrid tracer candidates were assessed and compared in C57BL/6 mice by PET as well as fluorescence imaging. MMP activity was imaged in a MMP-positive mouse model of irritant contact dermatitis (ICD) by PET and sequential fluorescence reflectance imaging (FRI), respectively. In vivo data were validated by scintillation counting, gelatin zymography, and MMP-histology. Three new potential hybrid MMP imaging probes were prepared, differing essentially in the number of sulfonate groups, introduced by Cy5 dye components. Although the hydrophilicity of these compounds was substantially increased, 10a (nSO3- = 1) and 10b (nSO3- = 2) were still rapidly eliminated via unfavorable hepatobiliary pathways, as observed in earlier approaches. Only 11 (nSO3- = 4) showed delayed in vivo clearance and a shift towards higher renal elimination. In the chosen mouse model of ICD, only 11 (nSO3- = 4) significantly accumulated in the inflamed mouse ear, which could be precisely visualized by means of PET and FRI.
Assuntos
Barbitúricos/química , Barbitúricos/farmacocinética , Corantes Fluorescentes/química , Radioisótopos do Iodo/química , Metaloproteinases da Matriz/metabolismo , Imagem Óptica/métodos , Tomografia por Emissão de Pósitrons/métodos , Animais , Halogenação , Camundongos , Camundongos Endogâmicos C57BL , Traçadores Radioativos , Distribuição TecidualRESUMO
Staphylococcus aureus causes very serious infections of vascular grafts. Knowledge of the molecular mechanisms of this disease is largely lacking because of the absence of representable models. Therefore, the aim of this study was to set up a mouse model of vascular graft infections that closely mimics the human situation. A catheter was inserted into the right carotid artery of mice, which acted as a vascular graft. Mice were infected i.v. using 8 different S. aureus strains, and development of the infection was followed up. Although all strains had varying abilities to form biofilm in vitro and different levels of virulence in mice, they all caused biofilm formation on the grafts. This graft infection was monitored using magnetic resonance imaging (MRI) and 18F-fluordeoxyglucose positron emission tomography (FDG-PET). MRI allowed the quantification of blood flow through the arteries, which was decreased in the catheter after infection. FDG-PET revealed high inflammation levels at the site of the catheter after infection. This model closely resembles the situation in patients, which is characterized by a tight interplay between pathogen and host, and can therefore be used for the testing of novel treatment, diagnosis, and prevention strategies. In addition, combining MRI and PET with microscopic techniques provides an appropriate way to characterize the course of these infections and to precisely analyze biofilm development.
Assuntos
Biofilmes/crescimento & desenvolvimento , Infecções Relacionadas a Cateter/diagnóstico por imagem , Modelos Animais de Doenças , Infecções Estafilocócicas/diagnóstico por imagem , Animais , Prótese Vascular/microbiologia , Ensaio de Imunoadsorção Enzimática , Hibridização in Situ Fluorescente , Imageamento por Ressonância Magnética , Camundongos , Microscopia Confocal , Microscopia Eletrônica de Transmissão , Tomografia por Emissão de Pósitrons , Staphylococcus aureusRESUMO
BODIPYs (boron dipyrromethenes) are fluorescent dyes which show high stability and quantum yields. They feature the possibility of selective 18F-fluorination at the boron-core. Attached to a bioactive molecule and labeled with [18F]fluorine, the resulting compounds are promising tracers for multimodal imaging in vivo and can be used for PET and fluorescence imaging. A BODIPY containing a phenyl and a hydroxy substituent on boron was synthesized and characterized. Fluorinated and hydroxy substituted dyes were coupled to an isatin-based caspase inhibitor via cycloaddition and the resulting compounds were evaluated in vitro in caspase inhibition assays. The metabolic stability and the formed metabolites were investigated by incubation with mouse liver microsomes and LC-MS analysis. Subsequently the fluorophores were labeled with [18F]fluorine and an in vivo biodistribution study using dynamic PET was performed.
Assuntos
Compostos de Boro/síntese química , Compostos de Boro/farmacologia , Radioisótopos de Flúor/farmacologia , Animais , Compostos de Boro/farmacocinética , Cromatografia Líquida , Reação de Cicloadição , Radioisótopos de Flúor/farmacocinética , Camundongos , Camundongos Endogâmicos C57BL , Microssomos Hepáticos/efeitos dos fármacos , Imagem Multimodal , Análise Espectral/métodos , Distribuição TecidualRESUMO
PURPOSE: Resistance to bevacizumab (BEV) in glioblastoma is believed to occur via activation of molecular networks including the mTOR/PI3K pathway. Using an MR/PET molecular imaging biomarker approach, we investigated the response to combining BEV with the mTOR/PI3K inhibitor BEZ235. METHODS: Tumours were established by orthotopically implanting U87MG-luc2 cells in mice. Animals were treated with BEZ235 and/or BEV, and imaged using diffusion-weighted-MRI, T2-weighted and T2*-weighted before and after administration of superparamagnetic iron oxide contrast agent. Maps for changes in relaxation rates (ΔR2, ΔR2* and apparent diffusion coefficient) were calculated. Vessel size index and microvessel density index were derived. 3'-Deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT) PET and O-(2-[(18)F]fluoroethyl)-L-tyrosine ([(18)F]FET) PET were further performed and tumour endothelium/proliferation markers assessed by immunohistochemistry. RESULTS: Treatment with BEV resulted in a pronounced decrease in tumour volume (T2-weighted MRI). No additive effect on tumour volume was observed with the BEV/BEZ235 combination compared with BEV monotherapy. The Ki67 proliferation index and [(18)F]FLT uptake studies were used to support the observations. Using ΔR2* and ΔR2 values, respectively, the BEV/BEZ235 combination significantly reduced tumour microvessel volume in comparison to BEV alone. Decreased microvessel density index was further observed in animals treated with the combination, supported by von Willebrand factor (vWF) immunohistochemistry. [(18)F]FET uptake was decreased following treatment with BEV alone, but was not further reduced following treatment with the combination. vWF immunohistochemistry analysis showed that the mean tumour vessel size was increased in all cohorts. CONCLUSION: Assessing MR imaging biomarker parameters together with [(18)F]FET and [(18)F]FLT PET provided information on mechanism of action of the drug combination and clues as to potential clinical responses. Following translation to clinical use, treatment with a BEV/BEZ235 combination could reduce peritumoral oedema obviating the requirement for steroids. The use of hypothesis-driven molecular imaging studies facilitates the preclinical evaluation of drug response. Studies of this kind may more accurately predict the clinical potential of the BEV/BEZ235 combination regimen as a novel therapeutic approach in oncology.
Assuntos
Bevacizumab/farmacologia , Glioblastoma/patologia , Imidazóis/farmacologia , Imageamento por Ressonância Magnética , Inibidores de Fosfoinositídeo-3 Quinase , Tomografia por Emissão de Pósitrons , Quinolinas/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Transporte Biológico/efeitos dos fármacos , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Interações Medicamentosas , Feminino , Glioblastoma/irrigação sanguínea , Glioblastoma/diagnóstico por imagem , Glioblastoma/metabolismo , Humanos , Camundongos , Microvasos/efeitos dos fármacos , Microvasos/patologia , Microvasos/fisiopatologia , Imagem Multimodal , Inibidores de Proteínas Quinases/farmacologia , Carga Tumoral/efeitos dos fármacos , Tirosina/análogos & derivados , Tirosina/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
INTRODUCTION: Dysregulated activity of matrix metalloproteinases (MMPs) drives a variety of pathophysiological conditions. Non-invasive imaging of MMP activity in vivo promises diagnostic and prognostic value. However, current targeting strategies by small molecules are typically limited with respect to the bioavailability of the labeled MMP binders in vivo. To this end, we here introduce and compare three chemical modifications of a recently developed barbiturate-based radiotracer with respect to bioavailability and potential to image MMP activity in vivo. METHODS: Barbiturate-based MMP inhibitors with an identical targeting unit but varying hydrophilicity were synthesized, labeled with technetium-99m, and evaluated in vitro and in vivo. Biodistribution and radiotracer elimination were determined in C57/BL6 mice by serial SPECT imaging. MMP activity was imaged in a MMP-positive subcutaneous xenograft model of human K1 papillary thyroid tumors. In vivo data were validated by scintillation counting, autoradiography, and MMP immunohistochemistry. RESULTS: We prepared three new 99mTc-labeled MMP inhibitors, bearing either a glycine ([99mTc]MEA39), lysine ([99mTc]MEA61), or the ligand HYNIC with the ionic co-ligand TPPTS ([99mTc]MEA223) yielding gradually increasing hydrophilicity. [99mTc]MEA39 and [99mTc]MEA61 were rapidly eliminated via hepatobiliary pathways. In contrast, [99mTc]MEA223 showed delayed in vivo clearance and primary renal elimination. In a thyroid tumor xenograft model, only [99mTc]MEA223 exhibited a high tumor-to-blood ratio that could easily be delineated in SPECT images. CONCLUSION: Introduction of HYNIC/TPPTS into the barbiturate lead structure ([99mTc]MEA223) results in delayed renal elimination and allows non-invasive MMP imaging with high signal-to-noise ratios in a papillary thyroid tumor xenograft model.
Assuntos
Inibidores de Metaloproteinases de Matriz , Neoplasias da Glândula Tireoide , Animais , Barbitúricos , Disponibilidade Biológica , Humanos , Ligantes , Metaloproteinases da Matriz/metabolismo , Camundongos , Tecnécio/química , Distribuição Tecidual , Tomografia Computadorizada de Emissão de Fóton Único/métodosRESUMO
κ-Opioid receptors (KORs) play a predominant role in pain alleviation, itching skin diseases, depression and neurodegenerative disorders such as multiple sclerosis. Therefore, imaging of KOR by a fluorinated PET tracer was envisaged. Two strategies were followed to introduce a F atom into the very potent class of cis,trans-configured perhydroquinoxalines. Whereas the synthesis of fluoroethyltriazole 2 has already been reported, fluoropyrrolidines 14 (1-[2-(3,4-dichlorophenyl)acetyl]-8-[(R)-3-fluoropyrrolidin-1-yl]-perhydroquinoxalines) were prepared by SN2 substitution of a cyclic sulfuric acid derivative with hydroxypyrrolidine and subsequent transformation of the OH moiety into a F substituent. Fluoropyrrolidines 14 showed similar low-nanomolar KOR affinity and selectivity to the corresponding pyrrolidines, but the corresponding alcohols were slightly less active. In the cAMP and ß-arrestin assay, 14b (proton at the 4-position) exhibited similar KOR agonistic activity as U-50,488. The fluoro derivatives 14b and 14c (CO2CH3 at the 4-position) revealed KOR-mediated anti-inflammatory activity as CD11c and the IFN-γ production were reduced significantly in mouse and human dendritic cells. Compounds 14b and 14-c also displayed anti-inflammatory and immunomodulatory activity in mouse and human T cells. The PET tracer [18F]-2 was prepared by 1,3-dipolar cycloaddition. In vivo, [18F]-2 did not label KOR due to very fast elimination kinetics. Nucleophilic substitution of a mesylate precursor provided [18F]-14c. Unfortunately, defluorination of [18F]-14c occurred inâ vivo, which was analyzed in detail by inâ vitro studies.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Quinoxalinas/farmacologia , Compostos Radiofarmacêuticos/farmacologia , Receptores Opioides kappa/agonistas , Animais , Anti-Inflamatórios não Esteroides/síntese química , Anti-Inflamatórios não Esteroides/química , Encéfalo/metabolismo , Células Cultivadas , Citocinas/análise , Radioisótopos de Flúor , Cobaias , Halogenação , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Tomografia por Emissão de Pósitrons , Quinoxalinas/síntese química , Quinoxalinas/química , Compostos Radiofarmacêuticos/síntese química , Compostos Radiofarmacêuticos/química , Receptores Opioides kappa/metabolismoRESUMO
Dysregulated levels of activated matrix metalloproteinases (MMPs) are linked to different pathologies, such as cancer, atherosclerosis, neuroinflammation, and arthritis. Therefore, imaging of MMPs with positron-emission tomography (PET) represents a powerful tool for the diagnosis of MMP-associated diseases. Moreover, to distinguish between the distinct functions and roles of individual MMPs in particular pathophysiological processes, their specific imaging must be realized with radiolabeled tracers, such as fluorine-18-labeled MMP inhibitors (MMPIs). Therefore, fluorinated dibenzofuransulfonamide-based MMPIs showing excellent inhibition of MMP-12 and selectivity for MMP-12 over other MMPs were prepared. MMP-12 is a key enzyme in diseases such as chronic obstructive pulmonary disease (COPD) and atherosclerosis. Because of their promising in vitro properties, three candidates (4, 9, and 19) were selected from this library, and radiofluorinated analogues ([18F]4, [18F]9, and [18F]19) were successfully synthesized. Initial in vitro serum stability and in vivo biodistribution studies of the radiolabeled MMPIs with PET demonstrated their potential benefit for preferable MMP-12 imaging.
Assuntos
Desenho de Fármacos , Metaloproteinase 12 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz/síntese química , Inibidores de Metaloproteinases de Matriz/farmacologia , Tomografia por Emissão de Pósitrons/métodos , Animais , Técnicas de Química Sintética , Humanos , Masculino , Inibidores de Metaloproteinases de Matriz/química , Inibidores de Metaloproteinases de Matriz/farmacocinética , Camundongos , Camundongos Endogâmicos C57BL , Traçadores Radioativos , Radioquímica , Distribuição TecidualRESUMO
BACKGROUND: Disturbance of intestinal wound closure leads to insufficient anastomotic healing and is associated with considerable morbidity following colorectal resections. Matrix metalloproteinases (MMPs) play a crucial role in regulation of wound closure. Here fluorescence endoscopy was evaluated for assessment of MMP-2/-9 expression during failed intestinal anastomotic healing. METHODS: Distal colonic anastomoses were performed as a model for disturbed healing in 36 Balb/c mice. Healing was evaluated endoscopically, macroscopically, and histologically after 1, 3 and 5 days. For detection of MMP-2/-9 expression fluorescence endoscopy (FE) was used following i.v.-administration of a Cy5.5-labeled MMP-2/-9 specific tracer. FE was complemented by quantification of the fluorescence signal using the MS-FX PRO Optical Imaging System. An overall leakage score was calculated and correlated with the results of FE. RESULTS: With increasing incidence of anastomotic leakage from POD1 (17%) to POD5 (83%) the uptake of the MMP tracer gradually increased (signal-to-noise ratio (SNR), POD1: 17.91 ± 1.251 vs. POD3: 30.56 ± 3.03 vs. POD5: 44.8 ± 4.473, P<0.0001). Mice with defective anastomotic healing showed significantly higher uptake compared to non-defective (SNR: 37.37± 3.63 vs. 26.16± 3.635, P = 0.0369). White light endoscopy and FE allowed evaluation of anastomotic healing and visualization of mucosal MMPs in vivo. Using FE based detection of MMPs in the anastomosis, an overall positive predictive value of 71.4% and negative predictive value of 66.6% was calculated for detection of anastomotic leakage. CONCLUSION: During disturbed anastomotic healing increased expression of MMP-2/-9 was observed in the anastomotic tissue. Fluorescence endoscopy for detection of MMP-2/-9 during the healing process might be a promising tool for early identification of anastomotic leakage.
Assuntos
Anastomose Cirúrgica/efeitos adversos , Fístula Anastomótica/diagnóstico por imagem , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Cicatrização , Administração Intravenosa , Fístula Anastomótica/patologia , Animais , Carbocianinas/administração & dosagem , Colo/diagnóstico por imagem , Colo/patologia , Colo/cirurgia , Colonoscopia/métodos , Modelos Animais de Doenças , Estudos de Viabilidade , Fluorescência , Corantes Fluorescentes/administração & dosagem , Humanos , Mucosa Intestinal/diagnóstico por imagem , Mucosa Intestinal/patologia , Mucosa Intestinal/cirurgia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Coloração e Rotulagem/métodosRESUMO
Background: To facilitate onsite decision-making during endoscopy, both accurate detection and in vivo characterization of preneoplasia are prerequisites. However, no endoscopy technique is available that meets both demands satisfactorily. We evaluated endothelin-receptor A (ETAR)-guided fluorescence endoscopy (FE) in vivo and fluorescence reflectance imaging (FRI) ex vivo for detection and characterization of early dysplastic colitis-associated colonic lesions. Methods: Colorectal cancerogenesis was investigated in the inflammatory driven AOM-DSS model and spontaneous adenoma development in ApcMin mice. A Cy5.5-labeled nonpeptidic ETAR-specific imaging probe was injected intravenously to assess tumor development in vivo by white light endoscopy (WLE) and FE. Ex vivo tumors were evaluated by FRI, histological examination, and western blot analysis. In addition, tissue samples from patients with colitis-associated malignant and nonmalignant mucosal alterations were analyzed. Specificity experiments were performed using an unspecific Cy3.5-glycine tracer. Results: Overall, 62 adenomas were observed. FE was able to detect and quantify ETAR expression targeting the ETAR-specific photoprobe. A significantly higher fluorescent contrast was detected in colonic adenomas compared to adjacent nonmalignant mucosa by FE (64.3 ± 7.9 vs. 56.6. ± 7.0; P < 0.001). These results were confirmed by FRI examination, immunochemistry, and western blot analysis. Additionally, ETAR expression in samples from human patients with colitis-associated cancer was highly elevated compared to nonmalignant alterations. Specificity experiments indicated a high binding-specificity of the applied ETAR photoprobe (1.4 ± 0.3 vs. 2.5 ± 0.7; P < 0.001). Conclusions: We introduced ETAR guided FE in mice for successful in vivo detection and characterization of colorectal neoplasia on a molecular level.
Assuntos
Adenoma/diagnóstico , Colite/complicações , Neoplasias do Colo/diagnóstico , Colonoscopia/métodos , Mucosa Intestinal/metabolismo , Imagem Óptica/métodos , Receptor de Endotelina A/metabolismo , Adenoma/etiologia , Adenoma/metabolismo , Animais , Colite/induzido quimicamente , Colite/fisiopatologia , Neoplasias do Colo/etiologia , Neoplasias do Colo/metabolismo , Feminino , Corantes Fluorescentes/metabolismo , Humanos , Mucosa Intestinal/patologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
The tumor microenvironment is highly heterogeneous. For gliomas, the tumor-associated inflammatory response is pivotal to support growth and invasion. Factors of glioma growth, inflammation, and invasion, such as the translocator protein (TSPO) and matrix metalloproteinases (MMP), may serve as specific imaging biomarkers of the glioma microenvironment. In this study, noninvasive imaging by PET with [18F]DPA-714 (TSPO) and [18F]BR-351 (MMP) was used for the assessment of localization and quantification of the expression of TSPO and MMP. Imaging was performed in addition to established clinical imaging biomarker of active tumor volume ([18F]FET) in conjunction with MRI. We hypothesized that each imaging biomarker revealed distinct areas of the heterogeneous glioma tissue in a mouse model of human glioma. Tracers were found to be increased 1.4- to 1.7-fold, with [18F]FET showing the biggest volume as depicted by a thresholding-based, volumes of interest analysis. Tumor areas, which could not be detected by a single tracer and/or MRI parameter alone, were measured. Specific compartments of [18F]DPA-714 (14%) and [18F]BR-351 (11%) volumes along the tumor rim could be identified. [18F]DPA-714 (TSPO) and [18F]BR-351 (MMP) matched with histology. Glioma-associated microglia/macrophages (GAM) were identified as TSPO and MMP sources. Multitracer and multimodal molecular imaging approaches may allow us to gain important insights into glioma-associated inflammation (GAM, MMP). Moreover, this noninvasive technique enables characterization of the glioma microenvironment with respect to the disease-driving cellular compartments at the various disease stages. Cancer Res; 77(8); 1831-41. ©2017 AACR.
Assuntos
Neoplasias Encefálicas/diagnóstico por imagem , Glioma/diagnóstico por imagem , Animais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Feminino , Radioisótopos de Flúor , Glioma/metabolismo , Glioma/patologia , Metaloproteinases da Matriz/metabolismo , Camundongos , Camundongos Nus , Microglia/patologia , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos , Receptores de GABA/metabolismo , Microambiente TumoralRESUMO
Advanced stage colorectal cancer (CRC) is still associated with limited prognosis. For preclinical evaluation of novel therapeutic approaches, murine models with orthotopic tumor growth and distant metastases are required. However, these models usually require surgical procedures possibly influencing tumor immunogenicity and development. The aim of this study was to establish a minimal-invasive endoscopy-based murine orthotopic model of metastatic CRC. During colonoscopy of CD-1 nude and non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, implantation of Caco-2 and HT-29 CRC cells was performed subcutaneously (s.c.) or orthotopic into the colonic submucosa. White light endoscopy (WLE) and fluorescence endoscopy (FE) were applied for tumor detection in vivo. Ex vivo, resected tumors were examined by fluorescence reflectance imaging (FRI), histology, gelatin zymography and immunohistochemistry. In CD-1 nude mice, marked tumor growth was observed within 14 days after subcutaneous implantation while submucosal implantation failed to induce CRC after 17 weeks. In contrast, in NOD/SCID mice submucosal injection of HT-29 cells resulted in pronounced tumor growth 12 days post injectionem. Subsequently, rapid tumor expansion occurred, occupying the entire colonic circumference. Importantly, post mortem histological analyses confirmed liver metastases in 28.6 % and peritoneal metastases in 14.3 % of all mice. FRI and gelatin zymography did not detect a significantly increased matrix metalloproteinases (MMPs) expression in s.c. implanted tumors while MMP-tracer uptake was significantly enhanced in orthotopic implanted tumors. Neither s.c. nor orthotopic Caco-2 cell implantation resulted in tumor development. We successfully established an endoscopy-based model of metastatic CRC in immunodeficient mice.
Assuntos
Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Endoscopia/métodos , Neoplasias Hepáticas/secundário , Cirurgia Assistida por Computador/métodos , Animais , Neoplasias Colorretais/diagnóstico por imagem , Neoplasias Colorretais/cirurgia , Células HT29 , Humanos , Processamento de Imagem Assistida por Computador/métodos , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/cirurgia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Nus , Camundongos SCID , Transplante de NeoplasiasRESUMO
BACKGROUND: Patients with ulcerative colitis are at increased risk for colorectal cancer and endoscopic surveillance is mandatory. Matrix metalloproteinases (MMPs)-2 and -9 activities are increased in malignant colonic mucosa. The aim of the study was to evaluate molecular imaging of MMP-2/-9 by fluorescence endoscopy (FE) for early tumor detection. METHODS: Colorectal cancer in mice (n = 28) was induced by azoxymethane and dextran sodium sulfate. Twenty-four hours after intravenous injection of a nonpeptidic, Cy5.5-labeled MMP-selective tracer, tumor development was assessed in vivo by white light endoscopy and FE. Topical administration of the tracer was also investigated (after 5 minutes and 24 hours). Colonic tumors were evaluated ex vivo by fluorescence reflectance imaging, immunohistochemistry, Western blot analysis, and zymography. RESULTS: Imaging of MMP-2/-9 expression by FE achieved a significantly higher contrast of the fluorescence signal in colonic adenomas compared with the adjacent nonmalignant mucosa (P < 0.001). Fluorescence reflectance imaging detected a significantly higher tracer uptake in adenoma compared with healthy mucosa (P < 0.001) and revealed a tumor size-dependent increase of tracer uptake (P < 0.01). Topical tracer administration did not facilitate tumor detection. Immunohistochemistry, Western blot analysis, and zymography indicated higher levels of MMP-2 and -9 in high-grade dysplasia and pT1 tumors ex vivo. CONCLUSIONS: MMP-2/-9 expression was significantly increased in colorectal neoplasia. FE allows direct visualization of a prognostic parameter (here MMP-2/-9) on a molecular level and may improve the characterization of colorectal lesions and the adenoma detection rate in the future.
Assuntos
Neoplasias Colorretais/diagnóstico , Endoscopia Gastrointestinal , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Imagem Óptica/métodos , Animais , Azoximetano/toxicidade , Western Blotting , Carbocianinas/metabolismo , Carcinógenos/toxicidade , Neoplasias Colorretais/induzido quimicamente , Neoplasias Colorretais/enzimologia , Sulfato de Dextrana/toxicidade , Feminino , Técnicas Imunoenzimáticas , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
UNLABELLED: Addition of temozolomide (TMZ) to radiation therapy is the standard treatment for patients with glioblastoma (GBM). However, there is uncertainty regarding the effectiveness of TMZ. Considering the rapid evolution of the disease, methods to assess TMZ efficacy early during treatment would be of great benefit. Our aim was to monitor early effects of TMZ in a mouse model of GBM using positron emission tomography (PET) with 3'-deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT). METHODS: Human glioma cells sensitive to TMZ (Gli36dEGFR-1) were treated with sub-lethal doses of TMZ to obtain cells with lower sensitivity to TMZ (Gli36dEGFR-2), as measured by growth and clonogenic assays. Gli36dEGFR-1 and Gli36dEGFR-2 cells were subcutaneously (s.c.) or intracranially (i.c.) xenografted into nude mice. Mice were treated for 7 days with daily injection of 25 or 50 mg/kg TMZ. Treatment efficacy was measured using [(18)F]FLT-PET before treatment and after 2 days. Computed Tomography (CT) or Magnetic Resonance Imaging (MRI) were used to determine tumor volumes before treatment and after 7 days. RESULTS: A significant difference was observed between TMZ and DMSO treated tumors in terms of variations of [(18)F]FLT T/B ratio as soon as day 2 in the i.c. as well as in the s.c. mouse model. Variations of [(18)F]FLT T/B uptake ratio between days 0 and 2 correlated with variations of tumor size between days 0 and 7 (s.c. model: n(tumor)â=â17 in n(mice)â=â11, P<0.01; i.c. model: n(tumor/mice)â=â9, P<0.01). CONCLUSIONS: Our results indicate that [(18)F]FLT-PET may be useful for an early evaluation of the response of GBM to TMZ chemotherapy in patients with glioma.