Introducing fluorescence guided surgery into orthopedic oncology: A systematic review of candidate protein targets for Ewing sarcoma

Orthopaedics, Trauma Surgery and Rehabilitatio

Wacana Kritis Berita Online Kasus Penyadapan Pembicaraan Telepon Elit Indonesia oleh Agen Rahasia Australia

News on online portal has advantages over news on conventional media in constructing reality and affecting the audiences. This research aims to analyze online text news about Indonesian\u27s political elites phone-tapping by Australian intelligence agency in Indonesian and Australian online news portal. This research uses text analyzing technique with interpretative explanation application. Data collection were done by observation on four online news portals, and then continued by data selection in the form of related news text. This news text is acquaired by using search engine in the internet. Collected data then being analyzed using Van Dijk Critical Discourse Analysis to apprehend the social discourses constructed by the online news portals, ideologies behind it, and the impact of those news publication on Indonesia-Australia diplomatic relationship. The results of this research indicates that there is a difference in discourses between Australia and Indonesia\u27s online news portal regarding the phone-tapping of Indonesia\u27s political elites by the Australian intelligence agency. Reality construction which is built by those news portals shows some differences, caused by disparities of press system and communication cultures between the two nations. Besides that, media ideologies also affects that phone-tapping publication. Those news publication in the process inflicts reaction on the people of the two nations, which affects the diplomatic relations between the two country

Ativação de neutrófilos de indivíduos saudáveis e imunocomprometidos por promastigotas de Leishmania spp.

Os neutrófilos são as primeiras células polimorfonucleares do sistema imunitário inato a chegar ao local de infeção, constituindo a primeira linha de defesa contra agentes invasores, como no caso do parasita Leishmania. Este parasita é causador da leishmaniose que tem sido reportada em mais de 98 países e que pode afetar animais e seres humanos. As diferentes espécies do parasita podem causar leishmaniose com diferentes apresentações clinicas, das quais salientamos a leishmaniose visceral e a leishmaniose cutânea. O presente trabalho teve como objetivo analisar a resposta imunitária de células polimórficas nucleares (PMN) de indivíduos com diferentes competências imunitárias quando expostas a parasitas de espécies viscerais (L. infantum) e cutâneas (L. amazonensis, L. shawi e L. guyanensis). A internalização parasitária foi confirmada por microscopia optica. A ativação celular foi avaliada através de técnicas laboratoriais que permitaram analisar os mecanismos oxidativos, nomeadamente a produção de ião superóxido, a exocitose de grânulos ricos em enzimas proteolíticas, como é o caso específico da elastase neutrofilica (NE) e da catepsina G (CatG) e a libertação de armadilhas extracelulares (NET). Verificou-se que as espécies de Leishmania induzem a exocitose de CatG e a libertação de NET, independentemente da competência imunitária dos indivíduos estudados. Contudo, idêntica situação não se verifica no caso do stress oxidativo ou da libertação de NE. Leishmania spp. estimulou o stress oxidativo, com produção do ião superóxido, nos PMN de indivíduos saudáveis enquanto os PMN de indivíduos imunocomprometidos apenas parece ficar ativado na presença de L. infantum e de L. amazonensis apontando para mecanismos de ativação diversos decorrentes quer da espécie do parasita quer do nível imunitário do hospedeiro. Situação similar ocorreu na exocitose de grânulos ricos em NE. L. infantum e L. shawi estimularam a libertação de NE por PMN de indivíduos saudáveis e imunocomprometidos. Porém, os PMN de indivíduos com comprometimento da imunidade também responderam aos parasitas da espécie L. amazonensis enquanto que L. guyanensis unicamente induziu a exocitose de NE nos PMN de indivíduos saudáveis. Estes resultados apontam para a existência de especificidades próprias na ativação destes mecanismos relativamente às espécies de Leishmania e ao estado imunitário do hospedeiro. Estudos complementares são necessários para esclarecer o processo de ativação, bem como identificar os eventuais antigénios parasitários (ou até ilhas CpG) que conduzem a ativação de PMN humanos.Neutrophils are the first polymorphonuclear cells of the innate immune system to reach the infection site, providing the first line of defense against invading pathogens, such as the Leishmania parasite. This parasite causes leishmaniasis that has been reported over 98 countries, affecting both animals and human beings. Different species of the parasite can cause leishmaniasis with diverse clinical presentations, as is the case of visceral leishmaniasis and cutaneous leishmaniasis. The present study aimed to evaluate the immune response of polymorphic nuclear cells (PMN) of individuals with different immune competence when exposed to visceral (L. infantum) and cutaneous parasitic species (L. amazonensis, L. guyanensis and L. shawi). Uptake of parasites by PMN was confirmed by optical microscopy. Laboratory techniques were used to evaluate the PMN activation by assessing oxidative mechanisms, namely the production of ion superoxide exocytosis, exocytosis of granules rich in proteolytic enzymes, as is the specific case of neutrophil elastase (NE) and cathepsin G (CatG) and the release of extracellular traps (NET). It was found that the species of Leishmania induce CatG exocytosis and NET release, regardless of the immune competence of the studied subjects. On the contrary, in the case of oxidative stress or the release of NE some differences were observed between PMN from healthy and immuncompromised individuals. All species of Leishmania induced oxidative stress, with production of superoxide ion in PMN of healthy individuals. However, PMN of immunocompromised individuals just seems to be activated in the presence of L. infantum and L. amazonensis parasites, pointing to the possible existence of more than one activation mechanism in association with the immune competence of the host that can be specifically primed by a particular parasite species. A similar situation was observed in the exocytosis of granules rich in NE. L. infantum and L. shawi stimulated the release of NE by PMN isolated from healthy and immunocompromised individuals. However, PMN from individuals with impaired immunity also appeared to respond to L. amazonensis parasites while L. guyanensis only induced PMN from healthy individuals to exocytosis NE. These findings suggest that the presence of specific features in the activation of these particular mechanisms might be related to the species of Leishmania and also to the immune status of the host. Additional studies are needed to clarify the activation process and identify parasitic antigens (or even CpG islands) that direct cell activation

Integrin αvβ6 as a target for tumor-specific imaging of vulvar squamous cell carcinoma and adjacent premalignant lesions

Surgical removal of vulvar squamous cell carcinoma (VSCC) is associated with significant morbidity and high recurrence rates. This is at least partially related to the limited visual ability to distinguish (pre)malignant from normal vulvar tissue. Illumination of neoplastic tissue based on fluorescent tracers, known as fluorescence-guided surgery (FGS), could help resect involved tissue and decrease ancillary mutilation. To evaluate potential targets for FGS in VSCC, immunohistochemistry was performed on paraffin-embedded premalignant (high grade squamous intraepithelial lesion and differentiated vulvar intraepithelial neoplasia) and VSCC (human papillomavirus (HPV)-dependent and -independent) tissue sections with healthy vulvar skin as controls. Sections were stained for integrin αvβ6, CAIX, CD44v6, EGFR, EpCAM, FRα, MRP1, MUC1 and uPAR. The expression of each marker was quantified using digital image analysis. H-scores were calculated and percentages positive cells, expression pattern, and biomarker localization were assessed. In addition, tumor-to-background ratios were established, which were highest for (pre)malignant vulvar tissues stained for integrin αvβ6. In conclusion, integrin αvβ6 allowed for the most robust discrimination of VSCCs and adjacent premalignant lesions compared to surrounding healthy tissue in immunohistochemically stained tissue sections. The use of an αvβ6 targeted near-infrared fluorescent probe for FGS of vulvar (pre)malignancies should be evaluated in future studies.Drug Delivery Technolog

Endoglin targeting inhibits tumor angiogenesis and metastatic spread in breast cancer

Endoglin, a transforming growth factor-beta co-receptor, is highly expressed on angiogenic endothelial cells in solid tumors. Therefore, targeting endoglin is currently being explored in clinical trials for anti-angiogenic therapy. In this project, the redundancy between endoglin and vascular endothelial growth factor (VEGF) signaling in angiogenesis and the effects of targeting both pathways on breast cancer metastasis were explored. In patient samples, increased endoglin signaling after VEGF inhibition was observed. In vitro TRC105, an endoglin-neutralizing antibody, increased VEGF signaling in endothelial cells. Moreover, combined targeting of the endoglin and VEGF pathway, with the VEGF receptor kinase inhibitor SU5416, increased antiangiogenic effects in vitro and in a zebrafish angiogenesis model. Next, in a mouse model for invasive lobular breast cancer, the effects of TRC105 and SU5416 on tumor growth and metastasis were explored. Although TRC105 and SU5416 decreased tumor vascular density, tumor volume was unaffected. Strikingly, in mice treated with TRC105, or TRC105 and SU5416 combined, a strong inhibition in the number of metastases was seen. Moreover, upon resection of the primary tumor, strong inhibition of metastatic spread by TRC105 was observed in an adjuvant setting. To confirm these data, we assessed the effects of endoglin-Fc (an endoglin ligand trap) on metastasis formation. Similar to treatment with TRC105 in the resection model, endoglin-Fc-expressing tumors showed strong inhibition of distant metastases. These results show, for the first time, that targeting endoglin, either with neutralizing antibodies or a ligand trap, strongly inhibits metastatic spread of breast cancer in vivo.Surgical oncolog

Cell-based tracers as Trojan horses for image-guided surgery

Surgeons rely almost completely on their own vision and palpation to recognize affected tissues during surgery. Consequently, they are often unable to distinguish between different cells and tissue types. This makes accurate and complete resection cumbersome. Targeted image-guided surgery (IGS) provides a solution by enabling real-time tissue recognition. Most current targeting agents (tracers) consist of antibodies or peptides equipped with a radiolabel for Positron Emission Tomography (PET) and Single Photon Emission Computed Tomography (SPECT), magnetic resonance imaging (MRI) labels, or a near-infrared fluorescent (NIRF) dye. These tracers are preoperatively administered to patients, home in on targeted cells or tissues, and are visualized in the operating room via dedicated imaging systems. Instead of using these ‘passive’ tracers, there are other, more ‘active’ approaches of probe delivery conceivable by using living cells (macrophages/monocytes, neutrophils, T cells, mesenchymal stromal cells), cell(-derived) fragments (platelets, extracellular vesicles (exosomes)), and microorganisms (bacteria, viruses) or, alternatively, ‘humanized’ nanoparticles. Compared with current tracers, these active contrast agents might be more efficient for the specific targeting of tumors or other pathological tissues (e.g., atherosclerotic plaques). This review provides an overview of the arsenal of possibilities applicable for the concept of cell-based tracers for IGS

Preclinical evaluation of EpCAM-binding designed ankyrin repeat proteins (DARPins) as targeting moieties for bimodal near-infrared fluorescence and photoacoustic imaging of cancer

PurposeFluorescence-guided surgery (FGS) can play a key role in improving radical resection rates by assisting surgeons to gain adequate visualization of malignant tissue intraoperatively. Designed ankyrin repeat proteins (DARPins) possess optimal pharmacokinetic and other properties for in vivo imaging. This study aims to evaluate the preclinical potential of epithelial cell adhesion molecule (EpCAM)-binding DARPins as targeting moieties for near-infrared fluorescence (NIRF) and photoacoustic (PA) imaging of cancer.MethodsEpCAM-binding DARPins Ac2, Ec4.1, and non-binding control DARPin Off7 were conjugated to IRDye 800CW and their binding efficacy was evaluated on EpCAM-positive HT-29 and EpCAM-negative COLO-320 human colon cancer cell lines. Thereafter, NIRF and PA imaging of all three conjugates were performed in HT-29_luc2 tumor-bearing mice. At 24 h post-injection, tumors and organs were resected and tracer biodistributions were analyzed.ResultsAc2-800CW and Ec4.1-800CW specifically bound to HT-29 cells, but not to COLO-320 cells. Next, 6 nmol and 24 h were established as the optimal in vivo dose and imaging time point for both DARPin tracers. At 24 h post-injection, mean tumor-to-background ratios of 2.60 & PLUSMN; 0.3 and 3.1 & PLUSMN; 0.3 were observed for Ac2-800CW and Ec4.1-800CW, respectively, allowing clear tumor delineation using the clinical Artemis NIRF imager. Biodistribution analyses in non-neoplastic tissue solely showed high fluorescence signal in the liver and kidney, which reflects the clearance of the DARPin tracers.ConclusionOur encouraging results show that EpCAM-binding DARPins are a promising class of targeting moieties for pan-carcinoma targeting, providing clear tumor delineation at 24 h post-injection. The work described provides the preclinical foundation for DARPin-based bimodal NIRF/PA imaging of cancer.Vascular SurgerySurgical oncolog

Glycan-based near-infrared fluorescent (NIRF) imaging of gastrointestinal tumors: a preclinical proof-of-conceptIn vivostudy

Purpose Aberrantly expressed glycans in cancer are of particular interest for tumor targeting. This proof-of-conceptin vivostudy aims to validate the use of aberrant Lewis glycans as target for antibody-based, real-time imaging of gastrointestinal cancers. Procedures Immunohistochemical (IHC) staining with monoclonal antibody FG88.2, targeting Lewis(a/c/x), was performed on gastrointestinal tumors and their healthy counterparts. Then, FG88.2 and its chimeric human/mouse variant CH88.2 were conjugated with near-infrared fluorescent (NIRF) IRDye 800CW for real-time imaging. Specific binding was evaluatedin vitroon human gastrointestinal cancer cell lines with cell-based plate assays, flow cytometry, and immune-fluorescence microscopy. Subsequently, mice bearing human colon and pancreatic subcutaneous tumors were imagedin vivoafter intravenous administration of 1 nmol (150 mu g) CH88.2-800CW with the clinical Artemis NIRF imaging system using the Pearl Trilogy small animal imager as reference. One week post-injection of the tracer, tumors and organs were resected and tracer uptake was analyzedex vivo. Results IHC analysis showed strong FG88.2 staining on colonic, gastric, and pancreatic tumors, while staining on their normal tissue counterparts was limited. Next, human cancer cell lines HT-29 (colon) and BxPC-3 and PANC-1 (both pancreatic) were identified as respectively high, moderate, and low Lewis(a/c/x)-expressing. Using the clinical NIRF camera system for tumor-bearing mice, a mean tumor-to-background ratio (TBR) of 2.2 +/- 0.3 (Pearl: 3.1 +/- 0.8) was observed in the HT-29 tumors and a TBR of 1.8 +/- 0.3 (Pearl: 1.9 +/- 0.5) was achieved in the moderate expression BxPC-3 model. In both models, tumors could be adequately localized and delineated by NIRF for up to 1 week.Ex vivoanalysis confirmed full tumor penetration of the tracer and low fluorescence signals in other organs. Conclusions Using a novel chimeric Lewis(a/c/x)-targeting tracer in combination with a clinical NIRF imager, we demonstrate the potential of targeting Lewis glycans for fluorescence-guided surgery of gastrointestinal tumors.Surgical oncolog

Novel Molecular Targets for Tumor-Specific Imaging of Epithelial Ovarian Cancer Metastases

In epithelial ovarian cancer (EOC), the strongest prognostic factor is the completeness of surgery. Intraoperative molecular imaging that targets cell-surface proteins on tumor cells may guide surgeons to detect metastases otherwise not visible to the naked eye. Previously, we identified 29% more metastatic lesions during cytoreductive surgery using OTL-38, a fluorescent tracer targeting folate receptor-α (FRα). Unfortunately, eleven out of thirteen fluorescent lymph nodes were tumor negative. The current study evaluates the suitability of five biomarkers (EGFR, VEGF-A, L1CAM, integrin αvβ6 and EpCAM) as alternative targets for molecular imaging of EOC metastases and included FRα as a reference. Immunohistochemistry was performed on paraffin-embedded tissue sections of primary ovarian tumors, omental, peritoneal and lymph node metastases from 84 EOC patients. Tumor-negative tissue specimens from these patients were included as controls. EGFR, VEGF-A and L1CAM were highly expressed in tumor-negative tissue, whereas αvβ6 showed heterogeneous expression in metastases. The expression of EpCAM was most comparable to FRα in metastatic lesions and completely absent in the lymph nodes that were false-positively illuminated with OTL-38 in our previous study. Hence, EpCAM seems to be a promising novel target for intraoperative imaging and may contribute to a more reliable detection of true metastatic EOC lesions.Surgical oncolog

Introducing fluorescence-guided surgery for pediatric Ewing, osteo-, and Rhabdomyosarcomas: a literature review

Sarcomas are a rare heterogeneous group of malignant neoplasms of mesenchymal origin which represent approximately 13% of all cancers in pediatric patients. The most prevalent pediatric bone sarcomas are osteosarcoma (OS) and Ewing sarcoma (ES). Rhabdomyosarcoma (RMS) is the most frequently occurring pediatric soft tissue sarcoma. The median age of OS and ES is approximately 17 years, so this disease is also commonly seen in adults while non-pleiomorphic RMS is rare in the adult population. The mainstay of all treatment regimens is multimodal treatment containing chemotherapy, surgical resection, and sometimes (neo)adjuvant radiotherapy. A clear resection margin improves both local control and overall survival and should be the goal during surgery with a curative intent. Real-time intraoperative fluorescence-guided imaging could facilitate complete resections by visualizing tumor tissue during surgery. This review evaluates whether non-targeted and targeted fluorescence-guided surgery (FGS) could be beneficial for pediatric OS, ES, and RMS patients. Necessities for clinical implementation, current literature, and the positive as well as negative aspects of non-targeted FGS using the NIR dye Indocyanine Green (ICG) were evaluated. In addition, we provide an overview of targets that could potentially be used for FGS in OS, ES, and RMS. Then, due to the time- and cost-efficient translational perspective, we elaborate on the use of antibody-based tracers as well as their disadvantages and alternatives. Finally, we conclude with recommendations for the experiments needed before FGS can be implemented for pediatric OS, ES, and RMS patients.Experimentele farmacotherapi