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Reactive oxygen species (ROS) activate NF-E2-related transcription factor 2 (Nrf2), a key transcriptional regulator driving antioxidant gene expression and protection from oxidant injury. Here, we report that in response to elevation of intracellular ROS above a critical threshold, Nrf2 stimulates expression of transcription Kruppel-like factor 9 (Klf9), resulting in further Klf9-dependent increases in ROS and subsequent cell death. We demonstrated that Klf9 independently causes increased ROS levels in various types of cultured cells and in mouse tissues and is required for pathogenesis of bleomycin-induced pulmonary fibrosis in mice. Mechanistically, Klf9 binds to the promoters and alters the expression of several genes involved in the metabolism of ROS, including suppression of thioredoxin reductase 2, an enzyme participating in ROS clearance. Our data reveal an Nrf2-dependent feedforward regulation of ROS and identify Klf9 as a ubiquitous regulator of oxidative stress and lung injury.
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Regulação da Expressão Gênica , Fatores de Transcrição Kruppel-Like/genética , Fator 2 Relacionado a NF-E2/genética , Estresse Oxidativo , Fibrose Pulmonar/genética , Animais , Sítios de Ligação , Bleomicina , Linhagem Celular Tumoral , Genes Reporter , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Luciferases/genética , Luciferases/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Fator 2 Relacionado a NF-E2/metabolismo , Células NIH 3T3 , Regiões Promotoras Genéticas , Ligação Proteica , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Espécies Reativas de Oxigênio , Transdução de SinaisRESUMO
BACKGROUND: Woodchucks (Marmota monax) are a well-accepted animal model for the investigation of spontaneous hepatocellular carcinoma (HCC). As HCC tumors obtain nutrient blood supply exclusively from the hepatic artery, hepatic artery infusion (HAI) has been applied to HCC. However, there is a scarcity of experimental animal models to standardize drug regimens and examine novel agents. The purpose of this study was to establish an HAI model in woodchucks. MATERIALS AND METHODS: HAI ports were placed in the gastroduodenal artery (GDA) of 11 woodchucks. The ports were infused with either a vehicle (dextrose 5% in water) or an experimental drug, CBL0137, once a week for 3 wk. Technical success rates, anatomical variation, morbidity and mortality, and tumor responses between groups were analyzed. RESULTS: The GDA access was feasible and reproducible in all woodchucks (11/11). The average operation time was 95 ± 20 min with no increase in the levels of liver enzymes detected from either infusate. The most common morbidity of CBL0137 therapy was anorexia after surgery. One woodchuck died due to hemorrhage at the gallbladder removal site from hepatic coagulopathy. Significantly higher CBL0137 concentrations were measured in the liver compared with blood after each HAI. Tumor growth was suppressed after multiple CBL0137 HAI treatments which corresponded to greater T cell infiltration and increased tumor cell apoptosis. CONCLUSIONS: HAI via GDA was a feasible and reproducible approach with low morbidity and mortality in woodchucks. The described techniques serve as a reliable platform for the identification and characterization of therapeutics for HCC.
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Carbazóis/administração & dosagem , Carcinoma Hepatocelular/tratamento farmacológico , Artéria Hepática/cirurgia , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Marmota , Variação Anatômica , Animais , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Artéria Hepática/anatomia & histologia , MasculinoRESUMO
BACKGROUND AND OBJECTIVES: Intraoperative photodynamic therapy (IO-PDT) is typically administered by a handheld light source. This can result in uncontrolled distribution of light irradiance that impacts tissue and tumor response to photodynamic therapy. The objective of this work was to characterize a novel optical surface applicator (OSA) designed to administer controlled light irradiance in IO-PDT. STUDY DESIGN/MATERIALS AND METHODS: An OSA was constructed from a flexible silicone mesh applicator with multiple cylindrically diffusing optical fibers (CDF) placed into channels of the silicone. Light irradiance distribution, at 665 nm, was evaluated on the OSA surface and after passage through solid tissue-mimicking optical phantoms by measurements from a multi-channel dosimetry system. As a proof of concept, the light administration of the OSA was tested in a pilot study by conducting a feasibility and performance test with 665-nm laser light to activate 2-(1'-hexyloxyethyl) pyropheophorbide-a (HPPH) in the thoracic cavity of adult swine. RESULTS: At the OSA surface, the irradiance distribution was non-uniform, ranging from 128 to 346 mW/cm2 . However, in the tissue-mimicking phantoms, beam uniformity improved markedly, with irradiance ranges of 39-153, 33-87, and 12-28 mW/cm2 measured at phantom thicknesses of 3, 5, and 10 mm, respectively. The OSA safely delivered the prescribed light dose to the thoracic cavities of four swine. CONCLUSIONS: The OSA can provide predictable light irradiances for administering a well-defined and potentially effective therapeutic light in IO-PDT. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.
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Lasers Semicondutores/uso terapêutico , Fotoquimioterapia/instrumentação , Cavidade Torácica/efeitos da radiação , Animais , Humanos , Imagens de Fantasmas , Silicones , SuínosRESUMO
BACKGROUND: Currently delivered light dose (J/cm2) is the principal parameter guiding interstitial photodynamic therapy (I-PDT) of refractory locally advanced cancer. The aim of this study was to investigate the impact of light dose rate (irradiance, mW/cm2) and associated heating on tumour response and cure. METHODS: Finite-element modeling was used to compute intratumoural irradiance and dose to guide Photofrin® I-PDT in locally advanced SCCVII in C3H mice and large VX2 neck tumours in New Zealand White rabbits. Light-induced tissue heating in mice was studied with real-time magnetic resonance thermometry. RESULTS: In the mouse model, cure rates of 70-90% were obtained with I-PDT using 8.4-245 mW/cm2 and ≥45 J/cm2 in 100% of the SCCVII tumour. Increasing irradiance was associated with increase in tissue heating. I-PDT with Photofrin® resulted in significantly (p < 0.05) higher cure rate compared to light delivery alone at same irradiance and light dose. Local control and/or cures of VX2 were obtained using I-PDT with 16.5-398 mW/cm2 and ≥45 J/cm2 in 100% of the tumour. CONCLUSION: In Photofrin®-mediated I-PDT, a selected range of irradiance prompts effective photoreaction with tissue heating in the treatment of locally advanced mouse tumour. These irradiances were translated for effective local control of large VX2 tumours.
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Carcinoma de Células Escamosas/tratamento farmacológico , Éter de Diematoporfirina/uso terapêutico , Neoplasias Experimentais/tratamento farmacológico , Fotoquimioterapia , Fármacos Fotossensibilizantes/uso terapêutico , Animais , Feminino , Temperatura Alta , Camundongos , Camundongos Endogâmicos C3H , Coelhos , TermometriaAssuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Quinolinas , Humanos , Carcinoma Hepatocelular/diagnóstico por imagem , Neoplasias Hepáticas/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Radioisótopos de Gálio , Fluordesoxiglucose F18RESUMO
OBJECTIVE: To describe a detailed step-by-step approach of our technique for robot-assisted intracorporeal 'W'-configuration orthotopic ileal neobladder. PATIENTS AND METHODS: Five patients underwent robot-assisted radical cystectomy (RARC), extended pelvic lymph node dissection and intracorporeal neobladder (ICNB). ICNB was divided into six key steps to facilitate and enable a detailed analysis and auditing of the technique. No conversion to open surgery was required. Timing for each step was noted. All patients had at least 3 months of follow-up. RESULTS: The mean age was 57 years. The mean overall console and diversion times were 357 and 193 min, respectively. None of the patients had any evidence of residual disease after RARC. Four of the five patients had complications; three developed fevers due to urinary tract infections (one required readmission), and one developed myocardial infarction and required coronary angiography and stenting. Looking at the timing for the individual steps, bowel detubularisation and construction of the posterior plate were consistently the longest among the key steps (average 46 min, 13% of the overall operative time), followed by uretero-ileal anastomosis (37 min, 10%), neobladder-urethral anastomosis (23 min, 6%), and identification and fixation of the bowel (26 min, 7%). CONCLUSION: We described our step-by-step technique and initial perioperative outcomes of our first five ICNBs with 'W' configuration.
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Cistectomia , Íleo/cirurgia , Procedimentos Cirúrgicos Robóticos , Derivação Urinária , Cistectomia/métodos , Feminino , Guias como Assunto , Humanos , Excisão de Linfonodo , Masculino , Pessoa de Meia-Idade , Cirurgia Assistida por Computador , Resultado do TratamentoRESUMO
BACKGROUND: The liver has unique anatomy in that most blood flow to normal hepatocytes is derived from the portal venous system, whereas liver tumors obtain their nutrient blood supply exclusively from the hepatic artery. The focused arterial delivery of anticancer agents to liver tumors has been performed for decades; however, preclinical models to standardize drug regimens and examine novel agents have been lacking. The purpose of this study was to establish preclinical hepatic artery infusion (HAI) models in a mouse and to evaluate the safety and delivery capability of the models. MATERIAL AND METHODS: C57BL/6 and BALB/c mice were used to develop models of HAI via the hepatic artery (HA), superior pancreaticoduodenal artery (SPDA), or lienogastric artery (LGA). Success rates, distribution of perfusion, and associated morbidity and mortality were analyzed between groups. RESULTS: All three models were feasible and reproducible in mice, and there was no statistical difference on body weight change between models. The HA model had a 13.3% mortality from acute liver failure, and the SPDA model demonstrated duodenal and pancreatic toxicity. SPDA and LGA routes had the highest success rates (96.7% and 91.4%, respectively) with low mortality, better drug delivery, and preserved physiologic liver function compared with the HA model. CONCLUSIONS: The optimal route of HAI was mouse breed specific; SPDA access in BALB/c mice, and the LGA access in C57BL/6 mice. The described techniques serve as a reproducible platform for the identification and characterization of therapeutics for diverse metastatic liver tumors.
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Antineoplásicos/administração & dosagem , Neoplasias do Colo/patologia , Fluoruracila/administração & dosagem , Artéria Hepática , Neoplasias Hepáticas Experimentais/secundário , Neoplasias Hepáticas/secundário , Animais , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Feminino , Fluoruracila/uso terapêutico , Infusões Intra-Arteriais , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Reprodutibilidade dos Testes , Resultado do TratamentoRESUMO
BACKGROUND: Renin-angiotensin system (RAS) activation increases angiotensin II production stimulating profibrotic factors, especially in the setting of chronic kidney disease. Nephrogenic systemic fibrosis (NSF) has been associated with gadolinium (Gd) exposure and renal failure. RAS involvement in NSF is unclear compared to transforming growth factor beta and Smad. RenTag mice were chosen to investigate the role of RAS in NSF-like dermal fibrosis because they demonstrated dermal fibrosis at birth, perturbations of RAS in subcutaneous tissue, and renal failure within 4 weeks of age. METHODS: Wild-type and RenTag mice were injected weekly with a supratherapeutic dose of intravenous gadodiamide (3.0 mmol/kg body weight) and killed at 12 weeks of age for skin and kidney histology. RESULTS: RenTag mice had elevated BUN levels, pitted kidneys, and glomerular damage. RenTag mice skin revealed an increased density of fibroblasts, no mucopolysaccharide deposits, and increased collagen fibril density regardless of Gd exposure. Skin and kidney histopathology of wild-type mice were normal regardless of Gd exposure. CD34 positivity was higher in RenTag compared to wild-type. CONCLUSIONS: Since RenTag dermal lesions remained unchanged after gadolinium exposure in the setting of renal failure, this animal model suggests perturbations of subcutaneous RAS may be involved in Gd-naïve dermal fibrosis.
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Modelos Animais de Doenças , Rim/patologia , Dermopatia Fibrosante Nefrogênica , Sistema Renina-Angiotensina , Animais , Camundongos Transgênicos , Dermopatia Fibrosante Nefrogênica/patologiaRESUMO
We show here that fundamental aspects of antitumor immunity in mice are significantly influenced by ambient housing temperature. Standard housing temperature for laboratory mice in research facilities is mandated to be between 20-26 °C; however, these subthermoneutral temperatures cause mild chronic cold stress, activating thermogenesis to maintain normal body temperature. When stress is alleviated by housing at thermoneutral ambient temperature (30-31 °C), we observe a striking reduction in tumor formation, growth rate and metastasis. This improved control of tumor growth is dependent upon the adaptive immune system. We observe significantly increased numbers of antigen-specific CD8(+) T lymphocytes and CD8(+) T cells with an activated phenotype in the tumor microenvironment at thermoneutrality. At the same time there is a significant reduction in numbers of immunosuppressive MDSCs and regulatory T lymphocytes. Notably, in temperature preference studies, tumor-bearing mice select a higher ambient temperature than non-tumor-bearing mice, suggesting that tumor-bearing mice experience a greater degree of cold-stress. Overall, our data raise the hypothesis that suppression of antitumor immunity is an outcome of cold stress-induced thermogenesis. Therefore, the common approach of studying immunity against tumors in mice housed only at standard room temperature may be limiting our understanding of the full potential of the antitumor immune response.
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Linfócitos T CD8-Positivos/imunologia , Abrigo para Animais/normas , Imunoterapia/métodos , Neoplasias/imunologia , Estresse Fisiológico/imunologia , Temperatura , Análise de Variância , Animais , Contagem de Células Sanguíneas , Linhagem Celular Tumoral , Feminino , Imuno-Histoquímica , Modelos Lineares , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Y15 or inhibitor 14 (1,2,4,5-benzenetetramine tetrahydrochloride) is a potent and specific inhibitor of focal adhesion kinase that inhibits its autophosphorylation activity, decreases the viability of cancer cells, and blocks tumor growth. In this preclinical study, we analyzed the pharmacokinetics of Y15 in mice plasma, its metabolic stability in mouse and human liver microsomes and toxicity in mice. The pharmacokinetics study in mice demonstrated that, following intraperitoneal administration at 30 mg/kg dose, Y15 was very rapidly absorbed in mice, reaching maximum plasma concentration in 4.8 min. Y15 rapidly metabolized in mouse and human liver microsomes with half-life t1/2 of 6.9 and 11.6 min, respectively. The maximal tolerated dose of single-dose administration of Y15 by oral administration was 200 mg/kg, and the multiple maximum tolerated dose of Y15 was 100 mg/kg by PO during 7 day study. Y15 did not cause any mortality or statistically significant differences in the body weight at 30 mg/kg by IP during 28-day study, and at 100 mg/kg by PO during the 7-day study. There were no clinical chemical, hematological, or histopathological changes in different mice organs at 30 mg/kg by IP during 28 days and at 100 mg/kg dose by PO during 7 days. Thus, this is the first preclinical toxicity, pharmacokinetics, and metabolic stability study of Y15 inhibitor. Further development of Y15 will provide a basis for new therapeutic and future clinical studies.
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Compostos de Anilina/farmacocinética , Compostos de Anilina/toxicidade , Antineoplásicos/farmacocinética , Antineoplásicos/toxicidade , Quinase 1 de Adesão Focal/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacocinética , Inibidores de Proteínas Quinases/toxicidade , Administração Oral , Compostos de Anilina/administração & dosagem , Compostos de Anilina/sangue , Animais , Antineoplásicos/administração & dosagem , Biotransformação , Estabilidade de Medicamentos , Feminino , Quinase 1 de Adesão Focal/metabolismo , Meia-Vida , Humanos , Injeções Intraperitoneais , Masculino , Dose Máxima Tolerável , Camundongos , Microssomos Hepáticos/metabolismo , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/sangue , Medição de Risco , Testes de ToxicidadeRESUMO
INTRODUCTION: Robot-assisted surgery (RAS) has been integrated into the surgical armamentarium and generated wide-spread interest among practicing, non-robotic surgeons (NRS). While methods for training novice non-robotic surgeons have emerged, the effectiveness of these training programs has endured minimal scrutiny. This study aims to establish effectiveness of the RAST training program. MATERIALS AND METHODS: A formal RAST program was established at Roswell Park Cancer Institute (RPCI) in 2008. From July 2010 to October 2012, 43 non-robotic surgeons participated in the program. The 1 to 4 week program included the validated fundamentals skills of robotic surgery (FSRS) curriculum, hands-on bedside trouble-shooting training, case observation with an expert robotic surgeon, hands on surgical training (HoST) procedure modules, da Vinci robotic surgical hands-on experience and finally a compulsory animal laboratory utilizing the da Vinci. As part of our training and credentialing quality assurance program, all participants were prospectively evaluated employing a survey. This survey aimed to evaluate the enduring impact of the RAST through time-sensitive interventions that allowed participants to reacclimatize themselves to their prospective practice as independently performing surgeons. RESULTS: The survey responses received from the participating NRS were collected over 27 months, with a response rate of 84%. The average follow up period post-RAST completion was 6 months (2-19). Overall, participants felt that the FSRS curriculum (81%), bedside trouble shooting (7%), and animal laboratory (53%) were beneficial program features that enabled NRS to become adequately acquainted with the basic principles of RAS. Approximately 5 weeks after RAST program completion, 64% of surgeons performed robot-assisted surgery. The two most commonly performed procedures were robot-assisted radical prostatectomy and gastrointestinal surgeries where eight surgeons performed independently while 12 performed procedures under the supervision of an expert robotic surgeon. The overall conversion rate to open was reported to be 1.3%. CONCLUSIONS: A dedicated surgical training program focused on learning key steps of RAS enabled most participants to successfully incorporate and maintain their RAS skills in clinical practice.
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Educação Médica Continuada/métodos , Laparoscopia/educação , Robótica/educação , Animais , Atitude do Pessoal de Saúde , Competência Clínica , Simulação por Computador , Humanos , Resolução de ProblemasRESUMO
Purpose: We tested a recently developed short peptide radioligand for PET imaging of hepatocellular carcinoma (HCC) by targeting an oncoprotein, extra-domain B fibronectin (EDB-FN) in the tumor microenvironment. Methods: The radioligand consists of a small linear peptide ZD2 with 68Ga-NOTA chelator, and specifically binds to EDB-FN. PET images were acquired dynamically for 1 hour after intravenously (i.v.) injecting 37 MBq (1.0 mCi) of the radioligand into the woodchuck model of naturally occurring HCC. Woodchuck HCC originated from chronic viral hepatitis infection, which recapitulates the corresponding human primary liver cancer. The animals were euthanized post-imaging for tissue collection and validation. Results: For ZD2 avid liver tumors, the radioligand accumulation plateaued a few minutes after injection, while the liver background uptake stabilized 20 min post-injection. The status of EDB-FN in woodchuck HCC was confirmed by histology and validated by PCR and western blocking. Conclusion: We have showed the viability of using the ZD2 short peptide radioligand targeting EDB-FN in liver tumor tissue for PET imaging of HCC, which can potentially impact the clinical care for HCC patients.
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Photobac is a near infrared photosensitizer (PS) derived from naturally occurring bacteriochlorophyll- a, with a potential for treating a variety of cancer types (U87, F98 and C6 tumor cells in vitro). The main objective of the studies presented herein was to evaluate the efficacy, toxicity and pharmacokinetic profile of Photobac in animals (mice, rats and dogs) and submit these results to the United States Food and Drug Administration (US FDA) for its approval to initiate Phase I human clinical trials of glioblastoma, a deadly cancer disease with no long term cure. The photodynamic therapy (PDT) efficacy of Photobac was evaluated in mice subcutaneously implanted with U87 tumors, and in rats bearing C6 tumors implanted in brain. In both tumor types, the Photobac-PDT was quite effective. The long-term cure in rats was monitored by magnetic resonance imaging (MRI) and histopathology analysis. A detailed pharmacology, pharmacokinetics and toxicokinetic study of Photobac was investigated in both non-GLP and GLP facilities at variable doses following the US FDA parameters. Safety Pharmacology studies suggest that there is no phototoxicity, cerebral or retinal toxicity with Photobac. No metabolites of Photobac were observed following incubation in rat, dog, mini-pig and human hepatocytes. Based on current biological data, Photobac-IND received the approval for Phase-I human clinical trials to treat Glioblastoma (brain cancer), which is currently underway at our institute. Photobac has also received an orphan drug status from the US FDA, because of its potential for treating Glioblastoma as no effective treatment is currently available for this deadly disease.
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Neoplasias Encefálicas , Glioblastoma , Fotoquimioterapia , Ratos , Cães , Animais , Camundongos , Humanos , Suínos , Bacterioclorofilas/uso terapêutico , Glioblastoma/patologia , Fotoquimioterapia/métodos , Bacterioclorofila A/uso terapêutico , Porco Miniatura , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/patologia , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Modelos AnimaisRESUMO
Although melanoma accounts for only 5.3% of skin cancer, it results in >75% of skin-cancer-related deaths. To avoid disfiguring surgeries on the head and neck associated with surgical excision, there is a clear unmet need for other strategies to selectively remove cutaneous melanoma lesions. Mohs surgery is the current treatment for cutaneous melanoma lesions and squamous and basal cell carcinoma. While Mohs surgery is an effective way to remove melanomas in situ, normal tissue is also excised to achieve histologically negative margins. This paper describes a novel combination therapy of nonthermal plasma (NTP) which emits a multitude of reactive oxygen species (ROS) and the injection of a pharmaceutical agent. We have shown that the effects of NTP are augmented by the DNA-damaging prodrug, tirapazamine (TPZ), which becomes a free radical only in conditions of hypoxemia, which is often enhanced in the tumor microenvironment. In this study, we demonstrate the efficacy of the combination therapy through experiments with B16-F10 and 1205 Lu metastatic melanoma cells both in vitro and in vivo. We also show the safety parameters of the therapy with no significant effects of the therapy when applied to porcine skin. We show the need for the intratumor delivery of TPZ in combination with the surface treatment of NTP and present a model of a medical device to deliver this combination therapy. The importance of functional gap junctions is indicated as a mechanism to promote the therapeutic effect. Collectively, the data support a novel therapeutic combination to treat melanoma and the development of a medical device to deliver the treatment in situ.
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Melanoma , Neoplasias Cutâneas , Suínos , Animais , Melanoma/tratamento farmacológico , Neoplasias Cutâneas/tratamento farmacológico , Tirapazamina/farmacologia , Terapia Combinada , Microambiente Tumoral , Melanoma Maligno CutâneoRESUMO
BACKGROUND: Cardioprotective effects of N-acetyl-ser-asp-lys-pro (Ac-SDKP) have been reported in preclinical models of myocardial remodeling. However, the rapid degradation of this endogenous peptide in vivo limits its clinical use. METHOD: To prolong its bioavailability, Ac-SDKP was encapsulated by phosphocholine lipid bilayers (liposomes) similar to mammalian cell membranes. The physical properties of the liposome structures were assessed by dynamic light scattering and scanning electron microscopy. The uptake of Ac-SDKP by RAW 264.7 macrophages and human and murine primary cardiac fibroblasts was confirmed by fluorescence microscopy and flow cytometry. Spectrum computerized tomography and competitive enzyme-linked immunoassays were performed to measure the ex vivo cardiac biodistribution of Ac-SDKP. The biological effects of this novel synthetic compound were examined in cultured macrophages and cardiac fibroblasts and in a murine model of acute myocardial infarction induced by permanent coronary artery ligation. RESULTS: A liposome formulation resulted in the greater uptake of Ac-SDKP than the naked peptide by cultured RAW 264.7 macrophages and cardiac fibroblasts. Liposome-delivered Ac-SDKP decreased fibroinflammatory genes in cultured cardiac fibroblasts co-treated with TGF-ß1 and macrophages stimulated with LPS. Serial tissue and serum immunoassays showed the high bioavailability of Ac-SDKP in mouse myocardium and in circulation. Liposome-delivered Ac-SDKP improved cardiac function and reduced myocardial fibroinflammatory responses in mice with acute myocardial infarction. CONCLUSION: Encapsulation of Ac-SDKP in a cell membrane-like phospholipid bilayer enhances its plasma and tissue bioavailability and offers cardioprotection against ischemic myocardial injury. Future clinical trials can use this novel approach to test small protective endogenous peptides in myocardial remodeling.
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Infarto do Miocárdio , Fosfolipídeos , Humanos , Camundongos , Animais , Fosfolipídeos/metabolismo , Lipossomos/metabolismo , Distribuição Tecidual , Colágeno/metabolismo , Miocárdio/metabolismo , Fibrose , Infarto do Miocárdio/metabolismo , Mamíferos/metabolismoRESUMO
Mice are the most common animal used to study disease, but there are real concerns about the reproducibility of many of these experiments. This review discusses how several different sources of chronic stress can directly impact experimental outcomes. Mandated housing conditions induce an underappreciated level of chronic stress but are not usually considered or reported as part of the experimental design. Since chronic stress plays a critical role in the development and progression of many somatic diseases including cancer, obesity, and auto-immune diseases, this baseline stress can directly affect outcomes of such experiments. To study the role of stress in both physical and psychiatric diseases, there has been a proliferation of protocols for imposing chronic stress on mice. For somatic diseases, biomarkers can be used to compare the models with the disease in patients, but to evaluate the validity of psychiatric models, behavioral tests are carried out to assess changes in behavior and these tests may themselves cause an underappreciated degree of additional stress. Therefore, it is important for animal welfare to reduce baseline stress and select the most humane protocols for inducing and assessing chronic stress to obtain the most reliable outcomes.
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BACKGROUND: Epidermal growth factor receptor (EGFR) inhibitors, including cetuximab and panitumumab, are valuable therapeutics for colorectal cancer (CRC), but resistance to these inhibitors is common. The reason for such resistance is not well understood, which hampers development of better therapeutic strategies. Although activating mutations in KRAS, BRAF and PIK3CA are considered major drivers of CRC resistance to EGFR inhibitors, therapeutic targeting of these drug resistance drivers has not produced substantial clinical benefit. METHODS: We exploited cell lines and mouse tumor models (cell line xenografts and patient derived xenografts) for experiments of genetic and pharmacologic depletion of EGFR and/or its family member HER2, including EGFR mutants, inhibition of EGFR ligand shedding, and biochemical analysis of signaling proteins, to delineate the mechanism of CRC resistance to EGFR inhibitors and to assess the therapeutic activity of PEPDG278D, which is a recombinant human protein that induces the degradation of both EGFR and HER2. RESULTS: The sensitivity of CRC cells to cetuximab and panitumumab correlates with the ability of these drugs to induce EGFR downregulation. PEPDG278D strongly inhibits oncogenic signaling and growth of CRC cells by causing profound depletion of EGFR and HER2, regardless of activating mutations of KRAS, BRAF and PIK3CA. siRNA knockdown of EGFR or HER2 also inhibits CRC cells resistant to EGFR inhibitors. Tumors harboring mutated KRAS, BRAF and/or PIK3CA also overexpress EGFR ligands, further suggesting that EGFR signaling remains important to the tumors. While excessive tumor-generated high-affinity EGFR ligands block target engagement by PEPDG278D, aderbasib, an inhibitor of ADAM10 and ADAM17, enables PEPDG278D to exert strong antitumor activity by inhibiting ligand shedding. Moreover, adding fluorouracil, which is commonly used in CRC treatment, to the combination of PEPDG278D and aderbasib further enhances tumor inhibition. CONCLUSIONS: Our study shows that CRC resistance to EGFR inhibitors results primarily from the inability of the inhibitors to downregulate their target and that a PEPDG278D-based combination treatment overcomes the resistance.
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Neoplasias Colorretais , Proteínas Proto-Oncogênicas B-raf , Animais , Cetuximab/farmacologia , Cetuximab/uso terapêutico , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/metabolismo , Humanos , Ligantes , Camundongos , Panitumumabe/farmacologia , Panitumumabe/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Receptores Proteína Tirosina Quinases/metabolismoRESUMO
Interstitial photodynamic therapy (I-PDT) is a promising therapy considered for patients with locally advanced cancer. In I-PDT, laser fibers are inserted into the tumor for effective illumination and activation of the photosensitizer in a large tumor. The intratumoral light irradiance and fluence are critical parameters that affect the response to I-PDT. In vivo animal models are required to conduct light dose studies, to define optimal irradiance and fluence for I-PDT. Here we describe two animal models with locally advanced tumors that can be used to evaluate the response to I-PDT. One model is the C3H mouse bearing large subcutaneous SCCVII carcinoma (400-600 mm3). Using this murine model, multiple light regimens with one or two optical fibers with cylindrical diffuser ends (cylindrical diffuser fiber, CDF) can be used to study tumor response to I-PDT. However, tissue heating may occur when 630 nm therapeutic light is delivered through CDF at an intensity ≥60 mW/cm and energy ≥100 J/cm. These thermal effects can impact tumor response while treating locally advanced mice tumors. Magnetic resonance imaging and thermometry can be used to study these thermal effects. A larger animal model, New Zealand White rabbit with VX2 carcinoma (~5000 mm3) implanted in either the sternomastoid (neck implantation model) or the biceps femoris muscle (thigh implantation model), can be used to study I-PDT with image-based pretreatment planning using computed tomography. In the VX2 model, the light delivery can include the use of multiple laser fibers to test light dosimetry and delivery that are relevant for clinical use of I-PDT.
Assuntos
Carcinoma , Segunda Neoplasia Primária , Fotoquimioterapia , Animais , Humanos , Camundongos , Camundongos Endogâmicos C3H , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/uso terapêutico , CoelhosRESUMO
Increased red cell distribution width (RDW), which measures erythrocyte mean corpuscular volume (MCV) variability (anisocytosis), has been linked to early mortality in many diseases and in older adults through unknown mechanisms. Hypoxic stress has been proposed as a potential mechanism. However, experimental models to investigate the link between increased RDW and reduced survival are lacking. Here, we show that lifelong hypobaric hypoxia (~10% O2) increased erythrocyte numbers, hemoglobin, and RDW, while reducing longevity in male mice. Compound heterozygous knockout (hKO) mutations in succinate dehydrogenase (Sdh; mitochondrial complex II) genes Sdhb, Sdhc, and Sdhd reduced Sdh subunit protein levels, reduced RDW, and increased healthy life span compared with WT mice in chronic hypoxia. RDW-SD, a direct measure of MCV variability, and the SD of MCV showed the most statistically significant reductions in Sdh hKO mice. Tissue metabolomic profiling of 147 common metabolites showed the largest increase in succinate with elevated succinate/fumarate and succinate/oxoglutarate (2-ketoglutarate) ratios in Sdh hKO mice. These results demonstrate that mitochondrial complex II level is an underlying determinant of both RDW and healthy life span in hypoxia and suggest that therapeutic targeting of Sdh might reduce high RDW-associated clinical mortality in hypoxic diseases.
Assuntos
Índices de Eritrócitos , Succinato Desidrogenase , Animais , Hipóxia , Longevidade , Masculino , Camundongos , Succinato Desidrogenase/genética , Succinato Desidrogenase/metabolismo , SuccinatosRESUMO
BACKGROUND AND AIMS: Radiolabeled short peptide ligands targeting prostate-specific membrane antigen (PSMA) were developed initially for imaging and treatment of prostate cancers. While many nonprostate solid tumors including hepatocellular carcinoma (HCC) express little PSMA, their neovasculature expresses a high level of PSMA, which is avid for Gallium-68-labeled PSMA-targeting radio-ligand (68Ga-PSMA-11) for positron emission tomography (PET). However, the lack of a spontaneous animal model of tumor-associated vascular PSMA overexpression has hindered the development and assessment of PSMA-targeting radioligands for imaging and therapy of the nonprostatic cancers. We identified detectable indigenous PSMA expression on tumor neovascular endothelia in a naturally occurring woodchuck model of HCC. METHODS: Molecular docking was performed with 3 bait PSMA ligands and compared between human and woodchuck PSMA. Initially, PET images were acquired dynamically after intravenously injecting 37 MBq (1.0 mCi) of 68Ga-PSMA-11 into woodchuck models of HCC. Subsequently, 10-minute static PET scans were conducted for other animals 1-hour after injection due to HCC and liver background uptake stabilization at 30-45 minutes after injection. Liver tissue samples were harvested after imaging, fresh-frozen for quantitative reverse transcription polymerase chain reaction and western blot for validation, or fixed for histology for correlation. RESULTS: Our preclinical studies confirmed the initial clinical findings of 68Ga-PSMA-11 uptake in HCC. The agents (ligands and antibodies) developed against human PSMA were found to be reactive against the woodchuck PSMA. CONCLUSION: This animal model offers a unique opportunity for investigating the biogenesis of tumor-associated vascular PSMA, its functional role(s), and potentials for future treatment strategies targeting tumor vascular PSMA using already developed PSMA-targeting agents.