RESUMO
OBJECTIVE: In RA, synovial fibroblasts become activated. These cells express fibroblast activation protein (FAP) and contribute to the pathogenesis by producing cytokines, chemokines and proteases. Selective depletion in inflamed joints could therefore constitute a viable treatment option. To this end, we developed and tested a new therapeutic strategy based on the selective destruction of FAP-positive cells by targeted photodynamic therapy (tPDT) using the anti-FAP antibody 28H1 coupled to the photosensitizer IRDye700DX. METHODS: After conjugation of IRDye700DX to 28H1, the immunoreactive binding and specificity of the conjugate were determined. Subsequently, tPDT efficiency was established in vitro using a 3T3 cell line stably transfected with FAP. The biodistribution of [111In]In-DTPA-28H1 with and without IRDye700DX was assessed in healthy C57BL/6N mice and in C57BL/6N mice with antigen-induced arthritis. The potential of FAP-tPDT to induce targeted damage was determined ex vivo by treating knee joints from C57BL/6N mice with antigen-induced arthritis 24 h after injection of the conjugate. Finally, the effect of FAP-tPDT on arthritis development was determined in mice with collagen-induced arthritis. RESULTS: 28H1-700DX was able to efficiently induce FAP-specific cell death in vitro. Accumulation of the anti-FAP antibody in arthritic knee joints was not affected by conjugation with the photosensitizer. Arthritis development was moderately delayed in mice with collagen-induced arthritis after FAP-tPDT. CONCLUSION: Here we demonstrate the feasibility of tPDT to selectively target and kill FAP-positive fibroblasts in vitro and modulate arthritis in vivo using a mouse model of RA. This approach may have therapeutic potential in (refractory) arthritis.
Assuntos
Artrite Experimental/tratamento farmacológico , Fibroblastos/efeitos dos fármacos , Fotoquimioterapia/métodos , Células 3T3/efeitos dos fármacos , Animais , Feminino , Indóis/uso terapêutico , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Compostos de Organossilício/uso terapêuticoRESUMO
OBJECTIVE: Targeting the glucagon-like peptide-1 receptor with radiolabeled exendin is a very promising method to noninvasively determine the ß cell mass in the pancreas, which is needed to unravel the pathophysiology of type 1 and type 2 diabetes. The present study aimed to explore the effects of both hyperglycemia and insulitis on the uptake of exendin in a spontaneous type 1 diabetes mouse model, nonobese diabetic (NOD) mice. METHODS: NOD mice (n = 75, 7-21 weeks old) were injected intravenously with [111In]In-DTPA-exendin-3, and single-photon emission computed tomography (SPECT) images were acquired 1 h pi. The pancreatic accumulation of [111In]In-DTPA-exendin-3 was quantified in vivo using SPECT and by ex vivo counting and correlated to the ß cell mass (BCM). The influence of insulitis and hyperglycemia on the exendin uptake was assessed. RESULTS: The pancreas could be visualized longitudinally using SPECT. A linear correlation was found between the BCM (%) and pancreatic uptake (%ID/g) as measured by ex vivo counting (Pearson r = 0.64, p < 0.001), which was not affected by either insulitis (Pearson r = 0.66, p = 0.83) or hyperglycemia (Pearson r = 0.57, p = 0.51). Biodistribution and ex vivo autoradiography revealed remaining [111In]In-DTPA-exendin-3 uptake in the pancreas despite total ablation of BCM. CONCLUSIONS: Despite hyperglycemia and severe insulitis, we have found a good correlation between BCM and pancreatic exendin uptake, even in a suboptimal model with relatively high background activity.
Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Hiperglicemia/metabolismo , Células Secretoras de Insulina/metabolismo , Peptídeos/metabolismo , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Animais , Autorradiografia , Diabetes Mellitus Tipo 1/diagnóstico por imagem , Modelos Animais de Doenças , Feminino , Imuno-Histoquímica , Radioisótopos de Índio/administração & dosagem , Radioisótopos de Índio/química , Radioisótopos de Índio/metabolismo , Injeções Intravenosas , Camundongos , Camundongos Endogâmicos NOD , Ácido Pentético/administração & dosagem , Ácido Pentético/química , Ácido Pentético/metabolismo , Peptídeos/administração & dosagem , Peptídeos/química , Compostos Radiofarmacêuticos/metabolismo , Distribuição TecidualRESUMO
Complete resection of tumor lesions in advanced stage ovarian cancer patients is of utmost importance, since the extent of residual disease after surgery strongly affects survival. Intraoperative imaging may be useful to improve surgery in these patients. Farletuzumab is a humanized IgG1 antibody that specifically recognizes the folate receptor alpha (FRα). Labeled with a radiolabel and a fluorescent dye, farletuzumab may be used for the intraoperative detection of ovarian cancer lesions. The current aim is to demonstrate the feasibility of FRα-targeted dual-modality imaging using 111In-farletuzumab-IRDye800CW in an intraperitoneal ovarian cancer model. Biodistribution studies were performed 3 days after injection of 3, 10, 30, or 100 µg of 111In-farletuzumab-IRDye800CW in mice with subcutaneous IGROV-1 tumors (5 mice per group). In mice with intraperitoneal IGROV-1 tumors the nonspecific uptake of 111In-farletuzumab-IRDye800CW was determined by coinjecting an excess of unlabeled farletuzumab. MicroSPECT/CT and fluorescence imaging were performed 3 days after injection of 10 µg of 111In-farletuzumab-IRDye800CW. FRα expression in tumors was determined immunohistochemically. Optimal tumor-to-blood-ratios (3.4-3.7) were obtained at protein doses up to 30 µg. Multiple intra-abdominal tumor lesions were clearly visualized by microSPECT/CT, while uptake in normal tissues was limited. Fluorescence imaging was used to visualize and guide resection of superficial tumors. Coinjection of an excess of unlabeled farletuzumab significantly decreased tumor uptake of 111In-farletuzumab-IRDye800CW (69.4 ± 27.6 versus 18.3 ± 2.2% ID/g, p < 0.05). Immunohistochemical analyses demonstrated that the radioactive and fluorescent signal corresponded with FRα-expressing tumor lesions. FRα-targeted SPECT/fluorescence imaging using 111In-farletuzumab-IRDye800CW can be used to detect ovarian cancer in vivo and could be a valuable tool for enhanced intraoperative tumor visualization in patients with intraperitoneal metastases of ovarian cancer.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Receptor 1 de Folato/antagonistas & inibidores , Cuidados Intraoperatórios/métodos , Neoplasias Ovarianas/diagnóstico por imagem , Cirurgia Assistida por Computador/métodos , Animais , Anticorpos Monoclonais Humanizados/química , Benzenossulfonatos/administração & dosagem , Benzenossulfonatos/química , Linhagem Celular Tumoral , Estudos de Viabilidade , Feminino , Corantes Fluorescentes/administração & dosagem , Corantes Fluorescentes/química , Receptor 1 de Folato/imunologia , Humanos , Radioisótopos de Índio/administração & dosagem , Radioisótopos de Índio/química , Indóis/administração & dosagem , Indóis/química , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Imagem Molecular/métodos , Imagem Óptica/métodos , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/cirurgia , Distribuição Tecidual , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A non-invasive imaging method to monitor islet grafts could provide novel and improved insight into the fate of transplanted islets and, potentially, monitor the effect of therapeutic interventions. Therefore, such an imaging method could help improve long-term transplantation outcome. Here, we investigated the use of [ 123 I]IBZM for insulin positive graft volume quantification and longitudinal graft monitoring. SPECT images were acquired 6 weeks after islet transplantation in the calf muscle of rats. For longitudinal graft analysis, rats were monitored by SPECT for 10 weeks. After animals were euthanized, graft containing muscles were dissected for ex vivo analysis and insulin-positive graft volume determination. Six weeks after transplantation, a clear signal was observed in all grafts by SPECT imaging. Moreover, the intensity of the SPECT signal correlated linearly with insulin-positive graft volume, as determined histologically. Longitudinal graft follow-up showed a clear SPECT signal of the transplant from 3 until 10 weeks after transplantation. In this study, we demonstrate for the first time the successful application of a radiotracer, [ 123 I]IBZM, for non-invasive, in vivo graft volume quantification and longitudinal graft monitoring.
Assuntos
Benzamidas , Meios de Contraste , Ilhotas Pancreáticas/diagnóstico por imagem , Extremidade Inferior/diagnóstico por imagem , Pirrolidinas , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Animais , Transplante das Ilhotas Pancreáticas , Período Pós-Operatório , RatosRESUMO
For intraoperative imaging, antibodies labeled with both a radionuclide and a fluorophore may be used to tag the tumor lesion with a radiolabel and a fluorescent signal at high tumor to background ratios. However, labeling antibodies with fluorescent moieties may affect the in vivo behavior of the antibody depending on the dye to antibody substitution ratio. To investigate the optimal substitution ratio for use in dual-modality image-guided surgery, we conjugated three different antibodies, MN-14 (anti-CEACAM5), girentuximab (anti-CAIX), and cetuximab (anti-EGFR), with both diethylene triamine pentaacetic acid (DTPA, for labeling with 111In) and IRdye 800CW at dye to antibody ratios of 0, 1, 1.5, 2, and 3 and assessed in vivo behavior. Biodistribution studies showed that at high dye to antibody ratios, liver uptake of the dual-labeled antibodies increased, whereas tumor uptake decreased. Conversely, very low ratios may not be optimal either because in that case, only a few antibody molecules will be dual-labeled (i.e., contain both a DTPA and an IRDye 800CW moiety), which may complicate interpretation of dual-modality data. The present study shows that, provided that the chelator to antibody ratio is high enough, a dye to antibody ratio in the range of 1 to 1.5 is optimal for antibody-targeted dual-modality imaging applications. However, the optimal configuration is antibody dependent and should be determined for each dual-labeled antibody individually.
Assuntos
Anticorpos/metabolismo , Diagnóstico por Imagem/métodos , Cuidados Intraoperatórios/métodos , Neoplasias/metabolismo , Animais , Feminino , Corantes Fluorescentes/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Nus , Coloração e Rotulagem , Distribuição TecidualRESUMO
PURPOSE: Tumor targeted optical imaging using antibodies labeled with near infrared fluorophores is a sensitive imaging modality that might be used during surgery to assure complete removal of malignant tissue. We evaluated the feasibility of dual modality imaging and image guided surgery with the dual labeled anti-carbonic anhydrase IX antibody preparation (111)In-DTPA-G250-IRDye800CW in mice with intraperitoneal clear cell renal cell carcinoma. MATERIALS AND METHODS: BALB/c nu/nu mice with intraperitoneal SK-RC-52 lesions received 10 µg DTPA-G250-IRDye800CW labeled with 15 MBq (111)In or 10 µg of the dual labeled irrelevant control antibody NUH-82 (20 mice each). To evaluate when tumors could be detected, 4 mice per group were imaged weekly during 5 weeks with single photon emission computerized tomography/computerized tomography and the fluorescence imaging followed by ex vivo biodistribution studies. RESULTS: As early as 1 week after tumor cell inoculation single photon emission computerized tomography and fluorescence images showed clear delineation of intraperitoneal clear cell renal cell carcinoma with good concordance between single photon emission computerized tomography/computerized tomography and fluorescence images. The high and specific accumulation of the dual labeled antibody conjugate in tumors was confirmed in the biodistribution studies. Maximum tumor uptake was observed 1 week after inoculation (mean ± SD 58.5% ± 18.7% vs 5.6% ± 2.3% injected dose per gm for DTPA-G250-IRDye800CW vs NUH-82, respectively). High tumor uptake was also observed at other time points. CONCLUSIONS: This study demonstrates the feasibility of dual modality imaging with dual labeled antibody (111)In-DTPA-G250-IRDye800CW in a clear cell renal cell carcinoma model. Results indicate that preoperative and intraoperative detection of carbonic anhydrase IX expressing tumors, positive resection margins and metastasis might be feasible with this approach.
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Anticorpos Monoclonais , Carcinoma de Células Renais/diagnóstico , Diagnóstico por Imagem , Neoplasias Renais/diagnóstico , Neoplasias Experimentais , Imagem Óptica/métodos , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Animais , Linhagem Celular Tumoral , Diagnóstico Diferencial , Feminino , Fluorescência , Camundongos , Camundongos Endogâmicos BALB C , Camundongos NusRESUMO
A new approach in the treatment of clear cell renal carcinoma (ccRCC) is radioimmunotherapy (RIT) using anti-carbonic anhydrase IX (CAIX) antibody G250. To investigate the potential of RIT with lutetium 177 (177Lu)-labeled G250, we conducted a protein dose escalation study and subsequently an RIT study in mice with intraperitoneally growing ccRCC lesions. Mice with intraperitoneal xenografts were injected with 1, 3, 10, 30, or 100 µg of G250 labeled with 10 MBq indium 111 (111In) to determine the optimal protein dose. The optimal protein dose determined with imaging and biodistribution studies was used in a subsequent RIT experiment in three groups of 10 mice with intraperitoneal SK-RC-52 tumors. One group received 13 MBq 177Lu-DOTA-G250, a control group received 13 MBq nonspecific 177Lu-MOPC21, and the second control group was not treated and received 20 MBq 111In-DOTA-G250. The optimal G250 protein dose to target ccRCC in this model was 10 µg G250. Treatment with 13 MBq 177Lu-DOTA-G250 was well tolerated and resulted in significantly prolonged median survival (139 days) compared to controls (49-53 days, p â=â .015), indicating that RIT has potential in this metastatic ccRCC model.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Carcinoma de Células Renais/radioterapia , Imunoconjugados/administração & dosagem , Neoplasias Renais/radioterapia , Lutécio/uso terapêutico , Radioisótopos/uso terapêutico , Compostos Radiofarmacêuticos/administração & dosagem , Animais , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Complexos de Coordenação/uso terapêutico , Relação Dose-Resposta à Radiação , Compostos Heterocíclicos com 1 Anel/uso terapêutico , Xenoenxertos , Humanos , Imunoconjugados/farmacocinética , Imunoconjugados/uso terapêutico , Injeções Intraperitoneais , Neoplasias Renais/patologia , Dose Máxima Tolerável , Camundongos , Transplante de Neoplasias , Neoplasias Experimentais , Radioimunoterapia , Compostos Radiofarmacêuticos/farmacocinética , Compostos Radiofarmacêuticos/uso terapêutico , Distribuição TecidualRESUMO
BACKGROUND: Rheumatoid arthritis (RA) is one of the most prevalent and debilitating joint diseases worldwide. RA is characterized by synovial inflammation (synovitis), which is linked to the development of joint destruction. Magnetic resonance imaging and ultrasonography are widely being used to detect the presence and extent of synovitis. However, these techniques do not reveal the activation status of inflammatory cells such as macrophages that play a crucial role in synovitis and express CD64 (Fc gamma receptor (FcγR)I) which is considered as macrophage activation marker. OBJECTIVES: We aimed to investigate CD64 expression and its correlation with pro-inflammatory cytokines and pro-damaging factors in human-derived RA synovium. Furthermore, we aimed to set up a molecular imaging modality using a radiolabeled CD64-specific antibody as a novel imaging tracer that could be used to determine the extent and phenotype of synovitis using optical and nuclear imaging. METHODS: First, we investigated CD64 expression in synovium of early- and late-stage RA patients and studied its correlation with the expression of pro-inflammatory and tissue-damaging factors. Next, we conjugated an anti-CD64 antibody with IRDye 800CW and diethylenetriamine penta-acetic acid (DTPA; used for 111In labeling) and tested its binding on cultured THP1 cells, ex vivo RA synovium explants and its imaging potential in SCID mice implanted with human RA synovium explants obtained from RA patients who underwent total joint replacement. RESULTS: We showed that CD64 is expressed in synovium of early and late-stage RA patients and that FCGR1A/CD64 expression is strongly correlated with factors known to be involved in RA progression. Combined, this makes CD64 a useful marker for imaging the extent and phenotype of synovitis. We reported higher binding of the [111In]In-DTPA-IRDye 800CW anti-CD64 antibody to in vitro cultured THP1 monocytes and ex vivo RA synovium compared to isotype control. In human RA synovial explants implanted in SCID mice, the ratio of uptake of the antibody in synovium over blood was significantly higher when injected with anti-CD64 compared to isotype and injecting an excess of unlabeled antibody significantly reduced the antibody-binding associated signal, both indicating specific receptor binding. CONCLUSION: Taken together, we successfully developed an optical and nuclear imaging modality to detect CD64 in human RA synovium in vivo.
Assuntos
Artrite Reumatoide , Sinovite , Camundongos , Animais , Humanos , Camundongos SCID , Imagem Molecular , Sinovite/diagnóstico por imagem , Artrite Reumatoide/diagnóstico por imagem , Biomarcadores , Anticorpos , Isotipos de Imunoglobulinas , Ácido PentéticoRESUMO
Macrophages play a crucial role in the initiation and progression of rheumatoid arthritis (RA). Liposomes can be used to deliver therapeutics to macrophages by exploiting their phagocytic ability. However, since macrophages serve as the immune system's first responders, it is inadvisable to systemically deplete these cells. By loading the liposomes with the photosensitizer IRDye700DX, we have developed and tested a novel way to perform photodynamic therapy (PDT) on macrophages in inflamed joints. PEGylated liposomes were created using the film method and post-inserted with micelles containing IRDye700DX. For radiolabeling, a chelator was also incorporated. RAW 264.7 cells were incubated with liposomes with or without IRDye700DX and exposed to 689 nm light. Viability was determined using CellTiterGlo. Subsequently, biodistribution and PDT studies were performed on mice with collagen-induced arthritis (CIA). PDT using IRDye700DX-loaded liposomes efficiently induced cell death in vitro, whilst no cell death was observed using the control liposomes. Biodistribution of the two compounds in CIA mice was comparable with excellent correlation of the uptake with macroscopic and microscopic arthritis scores. Treatment with 700DX-loaded liposomes significantly delayed arthritis development. Here we have shown the proof-of-principle of performing PDT in arthritic joints using IRDye700DX-loaded liposomes, allowing locoregional treatment of arthritis.
RESUMO
Treatment of hyperinsulinemic hypoglycemia is challenging. Surgical treatment of insulinomas and focal lesions in congenital hyperinsulinism is invasive and carries major risks of morbidity. Medication to treat nesidioblastosis and diffuse congenital hyperinsulinism has varying efficacy and causes significant side effects. Here, we describe a novel method for therapy of hyperinsulinemic hyperglycemia, highly selectively killing ß-cells by receptor-targeted photodynamic therapy (rtPDT) with exendin-4-IRDye700DX, targeting the glucagon-like peptide 1 receptor (GLP-1R). Methods: A competitive binding assay was performed using Chinese hamster lung (CHL) cells transfected with the GLP-1R. The efficacy and specificity of rtPDT with exendin-4-IRDye700DX were examined in vitro in cells with different levels of GLP-1R expression. Tracer biodistribution was determined in BALB/c nude mice bearing subcutaneous CHL-GLP-1R xenografts. Induction of cellular damage and the effect on tumor growth were analyzed to determine treatment efficacy. Results: Exendin-4-IRDye700DX has a high affinity for the GLP-1R, with a half-maximal inhibitory concentration of 6.3 nM. rtPDT caused significant specific phototoxicity in GLP-1R-positive cells (2.3% ± 0.8% and 2.7% ± 0.3% remaining cell viability in CHL-GLP-1R and INS-1 cells, respectively). The tracer accumulates dose-dependently in GLP-1R-positive tumors. In vivo, rtPDT induces cellular damage in tumors, shown by strong expression of cleaved caspase-3, and leads to a prolonged median survival of the mice (36.5 vs. 22.5 d, respectively; P < 0.05). Conclusion: These data show in vitro as well as in vivo evidence of the potency of rtPDT using exendin-4-IRDye700DX. This approach might in the future provide a new, minimally invasive, highly specific treatment method for hyperinsulinemic hypoglycemia.
Assuntos
Hiperinsulinismo Congênito/tratamento farmacológico , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Fotoquimioterapia/métodos , Animais , Linhagem Celular Tumoral , Cricetinae , Cricetulus , Exenatida/metabolismo , Exenatida/uso terapêutico , Feminino , Humanos , Indóis/metabolismo , Indóis/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Nesidioblastose/tratamento farmacológico , Compostos de Organossilício/metabolismo , Compostos de Organossilício/uso terapêutico , RatosRESUMO
Glucagon-like peptide 1 receptor (GLP-1R) imaging with radiolabeled exendin has proven to be a powerful tool to quantify ß-cell mass (BCM) in vivo. As GLP-1R expression is thought to be influenced by glycemic control, we examined the effect of blood glucose (BG) levels on GLP-1R-mediated exendin uptake in both murine and human islets and its implications for BCM quantification. Periods of hyperglycemia significantly reduced exendin uptake in murine and human islets, which was paralleled by a reduction in GLP-1R expression. Detailed mapping of the tracer uptake and insulin and GLP-1R expression conclusively demonstrated that the observed reduction in tracer uptake directly correlates to GLP-1R expression levels. Importantly, the linear correlation between tracer uptake and ß-cell area was maintained in spite of the reduced GLP-1R expression levels. Subsequent normalization of BG levels restored absolute tracer uptake and GLP-1R expression in ß-cells and the observed loss in islet volume was halted. This manuscript emphasizes the potency of nuclear imaging techniques to monitor receptor regulation noninvasively. Our findings have significant implications for clinical practice, indicating that BG levels should be near-normalized for at least 3 weeks prior to GLP-1R agonist treatment or quantitative radiolabeled exendin imaging for BCM analysis.
Assuntos
Glicemia , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Glucose/farmacologia , Ilhotas Pancreáticas/efeitos dos fármacos , Monitorização Fisiológica , Animais , Regulação da Expressão Gênica/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Humanos , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos SCID , Peptídeos/metabolismoRESUMO
The treatment of choice for insulinomas and focal lesions in congenital hyperinsulinism (CHI) is surgery. However, intraoperative detection can be challenging. This challenge could be overcome with intraoperative fluorescence imaging, which provides real-time lesion detection with a high spatial resolution. Here, a novel method for targeted near-infrared (NIR) fluorescence imaging of glucagonlike peptide 1 receptor (GLP-1R)-positive lesions, using the GLP-1 agonist exendin-4 labeled with IRDye 800CW, was examined in vitro and in vivo. Methods: A competitive binding assay was performed using Chinese hamster lung (CHL) cells transfected with GLP-1R. Tracer biodistribution was determined in BALB/c nude mice bearing subcutaneous CHL-GLP-1R xenografts. In vivo NIR fluorescence imaging of CHL-GLP-1R xenografts was performed. Localization of the tracer in the pancreatic islets of BALB/c nude mice was examined using fluorescence microscopy. Laparoscopic imaging was performed to detect the fluorescent signal of the tracer in the pancreas of mini pigs. Results: Exendin-4-IRDye 800CW binds GLP-1R with a half-maximal inhibitory concentration of 3.96 nM. The tracer accumulates in CHL-GLP-1R xenografts. Subcutaneous CHL-GLP-1R xenografts were visualized using in vivo NIR fluorescence imaging. The tracer accumulates specifically in the pancreatic islets of mice, and a clear fluorescent signal was detected in the pancreas of mini pigs. Conclusion: These data provide the first in vivo evidence of the feasibility of targeted fluorescence imaging of GLP-1R-positive lesions. Intraoperative lesion delineation using exendin-4-IRDye 800CW could benefit open as well as laparoscopic surgical procedures for removal of insulinomas and focal lesions in CHI.
Assuntos
Benzenossulfonatos/química , Exenatida/química , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Indóis/química , Imagem Óptica/métodos , Animais , Transporte Biológico , Células CHO , Cricetulus , Exenatida/metabolismo , Exenatida/farmacocinética , Feminino , Camundongos , Camundongos Nus , Pâncreas/metabolismo , Suínos , Distribuição TecidualRESUMO
PURPOSE: Intraoperative image guidance may aid in clinical decision-making during surgical treatment of colorectal cancer. We developed the dual-labeled carcinoembryonic antigen-targeting tracer, [111In]In-DTPA-SGM-101, for pre- and intraoperative imaging of colorectal cancer. Subsequently, we investigated the tracer in preclinical biodistribution and multimodal image-guided surgery studies, and assessed the clinical feasibility on patient-derived colorectal cancer samples, paving the way for rapid clinical translation. EXPERIMENTAL DESIGN: SGM-101 was conjugated with p-isothiocyanatobenzyl-diethylenetriaminepentaacetic acid (DTPA) and labeled with Indium-111 (111In). The biodistribution of 3, 10, 30, and 100 µg [111In]In-DTPA-SGM-101 was assessed in a dose escalation study in BALB/c nude mice with subcutaneous LS174T human colonic tumors, followed by a study to determine the optimal timepoint for imaging. Mice with intraperitoneal LS174T tumors underwent micro-SPECT/CT imaging and fluorescence image-guided resection. In a final translational experiment, we incubated freshly resected human tumor specimens with the tracer and assessed the tumor-to-adjacent tissue ratio of both signals. RESULTS: The optimal protein dose of [111In]In-DTPA-SGM-101 was 30 µg (tumor-to-blood ratio, 5.8 ± 1.1) and the optimal timepoint for imaging was 72 hours after injection (tumor-to-blood ratio, 5.1 ± 1.0). In mice with intraperitoneal tumors, [111In]In-DTPA-SGM-101 enabled preoperative SPECT/CT imaging and fluorescence image-guided resection. After incubation of human tumor samples, overall fluorescence and radiosignal intensities were higher in tumor areas compared with adjacent nontumor tissue (P < 0.001). CONCLUSIONS: [111In]In-DTPA-SGM-101 showed specific accumulation in colorectal tumors, and enabled micro-SPECT/CT imaging and fluorescence image-guided tumor resection. Thus, [111In]In-DTPA-SGM-101 could be a valuable tool for preoperative SPECT/CT imaging and intraoperative radio-guided localization and fluorescence image-guided resection of colorectal cancer.
Assuntos
Anticorpos Monoclonais/farmacologia , Antígeno Carcinoembrionário/isolamento & purificação , Neoplasias Colorretais/cirurgia , Cirurgia Assistida por Computador/métodos , Animais , Anticorpos Monoclonais/química , Antígeno Carcinoembrionário/genética , Linhagem Celular Tumoral , Neoplasias Colorretais/diagnóstico por imagem , Neoplasias Colorretais/patologia , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/isolamento & purificação , Xenoenxertos , Humanos , Radioisótopos de Índio/farmacologia , Camundongos , Imagem Óptica/métodos , Tomografia Computadorizada com Tomografia Computadorizada de Emissão de Fóton Único , Distribuição Tecidual/efeitos da radiaçãoRESUMO
Image-guided surgery can aid in achieving complete tumor resection. The development and assessment of tumor-targeted imaging probes for near-infrared fluorescence image-guided surgery relies mainly on preclinical models, but the translation to clinical use remains challenging. In the current study, we introduce and evaluate the application of a dual-labelled tumor-targeting antibody for ex vivo incubation of freshly resected human tumor specimens and assessed the tumor-to-adjacent tissue ratio of the detectable signals. Immediately after surgical resection, peritoneal tumors of colorectal origin were placed in cold medium. Subsequently, tumors were incubated with 111In-DOTA-hMN-14-IRDye800CW, an anti-carcinoembryonic antigen (CEA) antibody with a fluorescent and radioactive label. Tumors were then washed, fixed, and analyzed for the presence and location of tumor cells, CEA expression, fluorescence, and radioactivity. Twenty-six of 29 tumor samples obtained from 10 patients contained malignant cells. Overall, fluorescence intensity was higher in tumor areas compared to adjacent non-tumor tissue parts (p < 0.001). The average fluorescence tumor-to-background ratio was 11.8 ± 9.1:1. A similar ratio was found in the autoradiographic analyses. Incubation with a non-specific control antibody confirmed that tumor targeting of our tracer was CEA-specific. Our results demonstrate the feasibility of this tracer for multimodal image-guided surgery. Furthermore, this ex vivo incubation method may help to bridge the gap between preclinical research and clinical application of new agents for radioactive, near infrared fluorescence or multimodal imaging studies.
RESUMO
BACKGROUND: In colorectal cancer, survival of patients is drastically reduced when complete resection is hampered by involvement of critical structures. Targeted photodynamic therapy (tPDT) is a local and targeted therapy which could play a role in eradicating residual tumor cells after incomplete resection. Since carcinoembryonic antigen (CEA; CEACAM5) is abundantly overexpressed in colorectal cancer, it is a potential target for tPDT of colorectal cancer. METHODS: To address the potential of CEA-targeted PDT, we compared colorectal cancer cell lines with different CEA-expression levels (SW-48, SW-480, SW-620, SW-1222, WiDr, HT-29, DLD-1, LS174T, and LoVo) under identical experimental conditions. We evaluated the susceptibility to tPDT by varying radiant exposure and concentration of our antibody conjugate (DTPA-hMN-14-IRDye700DX). Finally, we assessed the efficacy of tPDT in vivo in 18 mice (BALB/cAnNRj-Foxn1nu/nu) with subcutaneously xenografted LoVo tumors. RESULTS: In vitro, the treatment effect of tPDT varied per cell line and was dependent on both radiant exposure and antibody concentration. Under standardized conditions (94.5 J/cm2 and 0.5 µg/µL antibody conjugate concentration), the effect of tPDT was higher in cells with higher CEA availability: SW-1222, LS174T, LoVo, and SW-48 (22.8%, 52.8%, 49.9%, and 51.9% reduction of viable cells, respectively) compared to cells with lower CEA availability. Compared to control groups (light or antibody conjugate only), tumor growth rate was reduced in mice with s.c. LoVo tumors receiving tPDT. CONCLUSION: Our findings suggest cells (and tumors) have different levels of susceptibility for tPDT even though they all express CEA. Furthermore, tPDT can effectively reduce tumor growth in vivo.
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BACKGROUND: Image-guided surgery may improve surgical outcome for colorectal cancer patients. Here, we evaluated the feasibility of a pretargeting strategy for multimodal imaging in colorectal cancer using an anti-carcinoembryonic antigen (CEA) x anti-histamine-succinyl-glycine (HSG) bispecific antibody (TF2) in conjunction with the dual-labeled diHSG peptide (RDC018), using both a fluorophore for near-infrared fluorescence imaging and a chelator for radiolabeling. METHODS: Nude mice with subcutaneous (s.c) CEA-expressing LS174T human colonic tumors and CEA-negative control tumors were injected with TF2. After 16 h, different doses of 111In-labeled IMP-288 (non-fluorescent) or its fluorescent derivative RDC018 were administered to compare biodistributions. MicroSPECT/CT and near-infrared fluorescence imaging were performed 2 and 24 h after injection. Next, the biodistribution of the dual-labeled humanized anti-CEA IgG antibody [111In]In-DTPA-hMN-14-IRDye800CW (direct targeting) was compared with the biodistribution of 111In-RDC018 in mice with TF2-pretargeted tumors, using fluorescence imaging and gamma counting. Lastly, mice with intraperitoneal LS174T tumors underwent near-infrared fluorescence image-guided resection combined with pre- and post-resection microSPECT/CT imaging. RESULTS: 111In-RDC018 showed specific tumor targeting in pretargeted CEA-positive tumors (21.9 ± 4.5 and 10.0 ± 4.7% injected activity per gram (mean ± SD %IA/g), at 2 and 24 hours post-injection (p.i.), respectively) and a biodistribution similar to 111In-IMP288. Both fluorescence and microSPECT/CT images confirmed preferential tumor accumulation. At post mortem dissection, intraperitoneal tumors were successfully identified and removed using pretargeting with TF2 and 111In-RDC018. CONCLUSION: A pretargeted approach for multimodal image-guided resection of colorectal cancer in a preclinical xenograft model is feasible, enables preoperative SPECT/CT, and might facilitate intraoperative fluorescence imaging.
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BACKGROUND: Combining modalities using dual-labeled antibodies may allow preoperative and intraoperative tumor localization and could be used in image-guided surgery to improve complete tumor resection. Trastuzumab is a monoclonal antibody against the human epidermal growth factor-2 (HER2) receptor and dual-labeled trastuzumab with both a fluorophore (IRDye800CW) and a radioactive label (111In) can be used for multimodal imaging of HER2-positive breast cancer. The aim of this study was to demonstrate the feasibility of HER2-targeted multimodal imaging using [111In]In-DTPA-trastuzumab-IRDye800CW in an orthotopic breast cancer model. METHODS: Trastuzumab was conjugated with p-isothiocyanatobenzyl (ITC)-diethylenetriaminepentaacetic acid (DTPA) and IRDye800CW-NHS ester and subsequently labeled with 111In. In a dose escalation study, the biodistribution of 10, 30, and 100 µg [111In]In-DTPA-trastuzumab-IRDye800CW was determined 48 h after injection in BALB/c nude mice with orthotopic high HER2-expressing tumors. Also, a biodistribution study was performed in a low HER2-expressing breast cancer model. In addition, multimodal image-guided surgery was performed in each group. Autoradiography, fluorescence microscopy, and immunohistochemically stained slices of the tumors were compared for co-localization of tumor tissue, HER2 expression, fluorescence, and radiosignal. RESULTS: Based on the biodistribution data, a 30 µg dose of dual-labeled trastuzumab (tumor-to-blood ratio 13 ± 2) was chosen for all subsequent studies. [111In]In-DTPA-trastuzumab-IRDye800CW specifically accumulated in orthotopic HER2-positive BT474 tumors (101 ± 7 %IA/g), whereas uptake in orthotopic low HER2-expressing MCF7 tumor was significantly lower (1.2 ± 0.2 %IA/g, p = 0.007). BT474 tumors could clearly be visualized with both micro-SPECT/CT, fluorescence imaging and subsequently, image-guided resection was performed. Immunohistochemical analyses of BT474 tumors demonstrated correspondence in fluorescence, radiosignal, and high HER2 expression. CONCLUSIONS: Dual-labeled trastuzumab showed specific accumulation in orthotopic HER2-positive BT474 breast tumors with micro-SPECT/CT and fluorescence imaging and enabled image-guided tumor resection. In the clinical setting, [111In]In-DTPA-trastuzumab-IRDye800CW could be valuable for preoperative detection of (metastatic) tumors by SPECT/CT imaging, and intraoperative localization by using a gamma probe and fluorescence image-guided surgery to improve radical resection of tumor tissue in patients with HER2-positive tumors.
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The changes in ß-cell mass (BCM) during the development and progression of diabetes could potentially be measured by radionuclide imaging using radiolabeled exendin. In this study, we investigated the potential of 111In-diethylenetriaminepentaacetic acid-exendin-3 (111In-exendin) in a rat model that closely mimics the development of type 1 diabetes (T1D) in humans: BioBreeding diabetes-prone (BBDP) rats. BBDP rats of 4-18 weeks of age were injected intravenously with 111In-exendin, and single-photon emission computed tomography (SPECT) images were acquired. The accumulation of the radiotracer was measured as well as the BCM and grade of insulitis by histology. 111In-exendin accumulated specifically in the islets, resulting in a linear correlation with the BCM (%) (Pearson r = 0.89, P < 0.0001, and r = 0.64 for SPECT). Insulitis did not have an influence on this correlation. These results indicate that 111In-exendin is a promising tracer to determine the BCM during the development of T1D, irrespective of the degree of insulitis.
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Radioisótopos de Índio/análise , Células Secretoras de Insulina/metabolismo , Peptídeos/análise , Animais , Diabetes Mellitus Tipo 1/metabolismo , Feminino , Humanos , Ratos , Tomografia Computadorizada de Emissão de Fóton ÚnicoRESUMO
Head and neck squamous cell carcinoma (HNSCC) is an often highly invasive tumor, infiltrating functionally important tissue areas. Achieving complete tumor resection and preserving functionally relevant tissue structures depends on precise identification of tumor-free resection margins during surgery. Fluorescence-guided surgery (FGS), by intraoperative detection of tumor cells using a fluorescent tracer, may guide surgical excision and identify tumor-positive resection margins. Using a literature survey on potential surface molecules followed by immunohistochemical validation, we identified CD44 variant 6 (CD44v6) as a constitutively expressed antigen in the invasion zone of HNSCC lesions. The monoclonal anti-CD44v6 antibody BIWA was labeled with both a near-infrared fluorescent dye (IRDye800CW) and a radioactive label (Indium-111) and dual-modality imaging was applied in a locally invasive tumor mouse model. BIWA accurately detected human HNSCC xenografts in mice with a tumor uptake of 54 ± 11% ID/g and invasion regions with an accuracy of 94%. When dissected under clinical-like conditions, tumor remnants approximately 0.7 mm in diameter consisting of a few thousand cells were identified by fluorescence imaging, resulting in reliable dissection of invasive microregions. These data indicate that CD44v6 is a suitable target for reliable near-infrared detection and FGS of invasive HNSCC lesions in vivo.
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Antígenos de Neoplasias/uso terapêutico , Receptores de Hialuronatos/uso terapêutico , Imagem Óptica/métodos , Carcinoma de Células Escamosas de Cabeça e Pescoço/cirurgia , Animais , Corantes Fluorescentes , Xenoenxertos , Humanos , Radioisótopos de Índio , Período Intraoperatório , Camundongos , Invasividade Neoplásica , Carcinoma de Células Escamosas de Cabeça e Pescoço/diagnóstico por imagemRESUMO
Pancreatic islet transplantation is a promising therapy for patients with type 1 diabetes. However, the duration of long-term graft survival is limited due to inflammatory as well as non-inflammatory processes and routine clinical tests are not suitable to monitor islet survival. 111In-exendin-SPECT (single photon emission computed tomography) is a promising method to non-invasively image islets after transplantation and has the potential to help improve the clinical outcome. Whether 111In-exendin-SPECT allows detecting small differences in beta-cell mass (BCM) and measuring the actual volume of islets that were successfully engrafted has yet to be demonstrated. Here, we evaluated the performance of 111In-exendin-SPECT using an intramuscular islet transplantation model in C3H mice. In vivo imaging of animals transplanted with 50, 100, 200, 400 and 800 islets revealed an excellent linear correlation between SPECT quantification of 111In-exendin uptake and insulin-positive area of islet transplants, demonstrating that 111In-exendin-SPECT specifically and accurately measures BCM. The high sensitivity of the method allowed measuring small differences in graft volumes, including grafts that contained less than 50 islets. The presented method is reliable, convenient and holds great potential for non-invasive monitoring of BCM after islet transplantation in humans.