Sex-specific transcriptome of the chicken chorioallantoic membrane.

Dimorphism between male and female embryos has been demonstrated in many animal species, including chicken species. Likewise, extraembryonic membranes such as the chorioallantoic membrane (CAM) are likely to exhibit a sex-specific profile. Analysis of the previously published RNA-seq data of the chicken CAM sampled at two incubation times, revealed 783 differentially expressed genes between the CAM of male and female embryos. The expression of some of these genes is sex-dependant only at one or other stage of development, while 415 genes are sex-dependant at both developmental stages. These genes include well-known sex-determining and sex-differentiation genes (DMRT1, HEGM, etc.), and are mainly located on sex chromosomes. This study provides evidence that gene expression of extra-embryonic membranes is differentially regulated between male and female embryos. As such, a better characterisation of associated mechanisms should facilitate the identification of new sex-specific biomarkers.

The chicken chorioallantoic membrane model for isolation of CRISPR/cas9-based HSV-1 mutant expressing tumor suppressor p53.

Oncolytic viruses (OVs) have emerged as a novel cancer treatment modality, which selectively target and kill cancer cells while sparing normal ones. Among them, engineered Herpes simplex virus type 1 (HSV-1) has been proposed as a potential treatment for cancer and was moved to phase III clinical trials. Previous studies showed that design of OV therapy combined with p53 gene therapy increases the anti-cancer activities of OVs. Here, the UL39 gene of the ICP34.5 deleted HSV-1 was manipulated with the insertion of the EGFP-p53 expression cassette utilizing CRISPR/ Cas9 editing approach to enhance oncoselectivity and oncotoxicity capabilities. The ΔUL39/Δγ34.5/HSV1-p53 mutant was isolated using the chorioallantoic membrane (CAM) of fertilized chicken eggs as a complementing membrane to support the growth of the viruses with gene deficiencies. Comparing phenotypic features of ΔUL39/Δγ34.5/HSV1-p53-infected cells with the parent Δγ34.5/HSV-1 in vitro revealed that HSV-1-P53 had cytolytic ability in various cell lines from different origin with different p53 expression rates. Altogether, data presented here illustrate the feasibility of exploiting CAM model as a promising strategy for isolating recombinant viruses such as CRISPR/Cas9 mediated HSV-1-P53 mutant with less virus replication in cell lines due to increased cell mortality induced by exogenous p53.

Human Non-Small Cell Lung Cancer-Chicken Embryo Chorioallantoic Membrane Tumor Models for Experimental Cancer Treatments.

To promote the preclinical development of new treatments for non-small cell lung cancer (NSCLC), we established NSCLC xenograft tumor assays on the chorioallantoic membrane (CAM) of chicken embryos. Five NSCLC cell lines were compared for tumor take rate, tumor growth, and embryo survival. Two of these, A549 and H460 CAM tumors, were histologically characterized and tested for susceptibility to systemic chemotherapy and gene delivery using viral vectors. All cell lines were efficiently engrafted with minimal effect on embryo survival. The A549 cells formed slowly growing tumors, with a relatively uniform distribution of cancer cells and stroma cells, while the H460 cells formed large tumors containing mostly proliferating cancer cells in a bed of vascularized connective tissue. Tumor growth was inhibited via systemic treatment with Pemetrexed and Cisplatin, a chemotherapy combination that is often used to treat patients with advanced NSCLC. Lentiviral and adenoviral vectors expressing firefly luciferase transduced NSCLC tumors in vivo. The adenovirus vector yielded more than 100-fold higher luminescence intensities after a single administration than could be achieved with multiple lentiviral vector deliveries. The adenovirus vector also transduced CAM tissue and organs of developing embryos. Adenovirus delivery to tumors was 100-10,000-fold more efficient than to embryo organs. In conclusion, established human NSCLC-CAM tumor models provide convenient in vivo assays to rapidly evaluate new cancer therapies, particularly cancer gene therapies.

The potential effect of leptin co-administration on photodynamic damage using quail chorioallantoic membrane model.

BACKGROUND: The chorioallantoic membrane (CAM) of the Japanese quail is an excellent model for studying photodynamic therapy (PDT) due to its rich vascularization. PDT is used not only in oncological treatment but also in infectious diseases, or psoriasis, where it yields significant advantages. This treatment also has its limitations, such as burning, itching, erythema, redness, swelling, and delayed wound healing. The aim of this study was to analyse the potentially protective properties of the tissue hormone leptin during PDT. METHODS: Japanese quail embryos incubated ex ovo were used in this experiment. On the 9th day of embryonic development, leptin (5 µg) and photosensitiser hypericin (79 µM) were topically applied, followed by irradiation. The effect of leptin co-administration was evaluated from CAM images and histological structure analysis, histological samples, and qPCR, where the expression of genes involved in angiogenesis, apoptosis, and oxidative stress was monitored. RESULTS: We observed vascular damage in all experimental groups, the highest damage was found after the application of hypericin without leptin coadministration. Histological analysis confirmed the protective effect of leptin. qPCR analysis presented differences in FREK gene expression, but also in genes involved in oxidative stress like SOD, NRF-1, NRF-2, and GPX7. The application of leptin significantly reduced the expression of apoptosis regulatory proteins CASP3, cytochrome C, and APAF1. CONCLUSIONS: Our results in the CAM model suggest a possible protective effect of leptin to prevent PDT damage and aid in the subsequent regeneration of target tissues after antimicrobial PDT.

How to Xenograft Cancer Cells on the Chorioallantoic Membrane of a Fertilized Hen's Egg and Its Visualization by PET/CT and MRI.

The chorioallantoic membrane (CAM) of fertilized hen's eggs represents a unique and alternative model for cancer research. The CAM model provides an optimal platform for xenografting cancer cell lines and studying essential key factors. Tumor size and growth as well as angiogenesis can be investigated to evaluate the response of therapies and strategies against cancer. Preclinical imaging represented by magnetic resonance imaging and positron emission tomography/computed tomography can generate detailed anatomical and functional information and reveal excellent metabolic sensitivity. In the following, a guideline is introduced in order to find a simplified entrance to the CAM model in combination with modern preclinical imaging techniques. Finally, the presented procedures are additionally completed by histological studies in the form of hematoxylin and eosin as well as immunohistochemical staining.

Use of Chicken Embryos as an Angiogenesis Model for Central Nervous System Malignant Tumor Research.

AIM: To demonstrate the usability of chicken chorioallantoic membrane (CAM) as an angiogenesis model for the development and treatment of malignant tumors of the central nervous system. MATERIAL AND METHODS: A fresh tumor tissue piece taken from Glioblastoma patients, a malignant tumor of the central nervous system, was transferred to the CAM of chicken embryos and left to incubate in the incubator and their development was monitored. After examining the results of the study macroscopically, CAM tissue samples were evaluated both histochemically and immunohistochemically in terms of angiogenic factors VEGF (Vascular Endothelial Growth Factor), bFGF (basic Fibroblast Growth Factor) and PDGF (Platelet Derived Growth Factor). RESULTS: According to histochemical findings obtained from our study when compared with control embryos, blood vessels, fibroblast count and inflammatory infiltration were observed more in the tumor transplanted groups, especially in the tumordeveloping CAM region. There was also intense pleomorphism and marked hypercellularity in the cells. In our immunohistochemical findings, it was determined that bFGF, PDGF, VEGF staining intensities were higher in tumor transplanted groups compared to control groups, and this elevation was more pronounced in the tumor-developing region. CONCLUSION: As a result, it has been shown that the chicken embryo CAM model may be a suitable in vivo model for cancer angiogenesis studies. The protocol we created in this study will be a source for projects related to the use of therapeutic agents in cancer angiogenesis.

RNA-seq analysis of the active chick embryo chorioallantoic membrane reveals genes that encode proteins assigned to ion transport and innate immunity.

The chicken chorioallantoic membrane (CAM) is an extraembryonic membrane that is vital for the embryo. It undergoes profound cell differentiation between 11 and 15 days of embryonic incubation (EID), which corresponds to the acquisition of its physiological functions. To gain insight into the functional genes that accompany these biological changes, RNA sequencing of the CAM at EID11 and EID15 was performed. Results showed that CAM maturation coincides with the overexpression of 4225 genes, including many genes encoding proteins involved in mineral metabolism, innate immunity, homeostasis, angiogenesis, reproduction, and regulation of hypoxia. Of these genes, some exhibit variability in expression depending on the chicken breed (broiler versus layer breeds). Besides the interest of these results for the poultry sector, the identification of new functional gene candidates opens additional research avenues in the field of developmental biology.

Development of in vitro and in vivo tools to evaluate the antiangiogenic potential of melatonin to neutralize the angiogenic effects of VEGF and breast cancer cells: CAM assay and 3D endothelial cell spheroids.

Melatonin is a molecule with different antitumor actions in breast cancer and has been described as an inhibitor of vascular endothelial growth factor (VEGF). Despite the recognition of the key role exerted by VEGF in tumor angiogenesis, limitations arise when developing models to test new antiangiogenic molecules. Thus, the aim of this study was to develop rapid, economic, high capacity and easy handling angiogenesis assays to test the antiangiogenic effects of melatonin and demonstrate its most effective dose to neutralize and interfere with the angiogenic sprouting effect induced by VEGF and MCF-7. To perform this, 3D endothelial cell (HUVEC) spheroids and a chicken embryo chorioallantoic membrane (CAM) assay were used. The results showed that VEGF and MCF-7 were able to stimulate the sprouting of the new vessels in 3D endothelial spheroids and the CAM assay, and that melatonin had an inhibitory effect on angiogenesis. Specifically, as the 1 mM pharmacological dose was the only effective dose able to inhibit the formation of ramifications around the alginate in the CAM assay model, this inhibition was shown to occur in a dose-dependent manner. Taken together, these techniques represent novel tools for the development of antiangiogenic molecules such as melatonin, with possible implications for the therapy of breast cancer.

Chick chorioallantoic membrane: a valuable 3D in vivo model for screening nanoformulations for tumor antiangiogenic therapeutics.

Drug discovery is an extensive process. From identifying lead compounds to approval for clinical application, it goes through a sequence of labor-intensive in vitro, in vivo preclinical screening and clinical trials. Among thousands of drugs screened only a few get approval for clinical trials. Furthermore, these approved drugs are often discontinued due to systemic toxicity and comorbidity at clinically administered dosages. To overcome these limitations, nanoformulations have emerged as the most sought-after strategy to safely and effectively deliver drugs within tumors at therapeutic concentrations. Most importantly, the employment of suitably variable preclinical models is considered highly critical for the therapeutic evaluation of candidate drugs or their formulations. A review of literature from the past 10 years on antiangiogenic nanoformulations shows the employment of limited types of preclinical models mainly the 2-dimensional (2D) monolayer cell culture and murine models as the mainstay for drug uptake, toxicity and efficiency studies. To top it all, murine models are highly expensive, time-consuming and require expertise in handling them. The current review highlights the utilization of the age-old chicken chorioallantoic membrane (CAM), a well-defined angiogenic model in the investigation of antiangiogenic compounds and nanoformulations in an economic framework. For practical applicability, we have evaluated the CAM model to demonstrate the screening of antiangiogenic compounds and that tumor cells can be implanted onto developing CAM for growing xenografts by recruiting host endothelial and other cellular components. In addition, the exploitation of CAM tumor xenograft models for the evaluation of nanoparticle distribution has also been reinforced by demonstrating that intravenously administered iron oxide nanoparticles (IONPs) passively accumulate and exhibit intracellular as well as extracellular compartment accumulation in highly vascular xenografts. Finally, the ethical considerations, benefits, and drawbacks, of using CAM as an experimental model for testing potential therapeutics are also highlighted.

Longitudinal bioluminescence imaging to monitor breast tumor growth and treatment response using the chick chorioallantoic membrane model.

The development of successful treatment regimens for breast cancer requires strong pre-clinical data generated in physiologically relevant pre-clinical models. Here we report the development of the chick embryo chorioallantoic membrane (CAM) model to study tumor growth and angiogenesis using breast cancer cell lines. MDA-MB-231 and MCF7 tumor cell lines were engrafted onto the chick embryo CAM to study tumor growth and treatment response. Tumor growth was evaluated through bioluminescence imaging and a significant increase in tumor size and vascularization was found over a 9-day period. We then evaluated the impact of anti-angiogenic drugs, axitinib and bevacizumab, on tumor growth and angiogenesis. Drug treatment significantly reduced tumor vascularization and size. Overall, our findings demonstrate that the chick embryo CAM is a clinically relevant model to monitor therapeutic response in breast cancer and can be used as a platform for drug screening to evaluate not only gross changes in tumor burden but physiological processes such as angiogenesis.

[The hen embryo: An alternative preclinical model in cancer]. / L'embryon de poule - Un modèle préclinique alternatif en cancérologie.

For therapeutic purposes, the development of new anti-cancer drugs requires their evaluation in terms of activity, cytotoxicity and pharmacokinetics. The candidate drugs are tested in vitro on cell lines and primary cells isolated from patients, and in vivo, often, using xenografts in immuno-compromised mice. In recent years, administrative constraints have become increasingly stringent and the 3R rule (reduce, refine, replace) requires the elaboration of alternative models capable to replace mouse models or at least to limit their use. Among them, xenograft on chick embryo chorioallantoic membrane (CAM assay) seems particularly efficient. It makes it possible to monitor and quantify tumor growth and tumor-associated parameters such as neoangiogenesis, invasion and migration. It allows the screening of drugs effective both on tumor cells and their microenvironment. Finally, the model seems adapted to the development of personalized medicine to which current research in cancerology is tending. In this context, this review focuses on the technique itself and its advantages.

Title: L'embryon de poule - Un modèle préclinique alternatif en cancérologie. Abstract: Le développement de drogues anti-cancéreuses à visée thérapeutique nécessite leur évaluation. Ces drogues candidates sont généralement testées in vitro, sur des lignées cellulaires ou sur des cellules isolées à partir de patients, et, in vivo, dans des modèles de xénogreffe chez la souris immunodéprimée. Depuis quelques années, les contraintes réglementaires (règle des 3R : réduire, raffiner, remplacer) imposent de mettre en place des modèles alternatifs qui se substituent aux modèles murins ou, au moins, en limitent l'utilisation. Parmi les modèles alternatifs proposés, la greffe sur membrane chorio-allantoïdienne d'embryon de poule semble performante. Elle permet de suivre et de quantifier la croissance tumorale et d'autres paramètres associés, comme la néo-angiogenèse, l'invasion et la migration tumorales. Elle permet aussi le criblage de drogues. Ce modèle semble également adapté à la médecine personnalisée en cancérologie. Nous présentons dans cette revue la technique et ses avantages.

The Comparative Experimental Study of Sodium and Magnesium Dichloroacetate Effects on Pediatric PBT24 and SF8628 Cell Glioblastoma Tumors Using a Chicken Embryo Chorioallantoic Membrane Model and on Cells In Vitro.

In this study, pyruvate dehydrogenase kinase-1 inhibition with dichloroacetate (DCA) was explored as an alternative cancer therapy. The study's aim was to compare the effectiveness of NaDCA and MgDCA on pediatric glioblastoma PBT24 and SF8628 tumors and cells. The treatment effects were evaluated on xenografts growth on a chicken embryo chorioallantoic membrane. The PCNA, EZH2, p53, survivin expression in tumor, and the SLC12A2, SLC12A5, SLC5A8, CDH1, and CDH2 expression in cells were studied. The tumor groups were: control, cells treated with 10 mM and 5 mM of NaDCA, and 5 mM and 2.5 mM of MgDCA. The cells were also treated with 3 mM DCA. Both the 10 mM DCA preparations significantly reduced PBT24 and SF8624 tumor invasion rates, while 5 mM NaDCA reduced it only in the SF8628 tumors. The 5 mM MgDCA inhibited tumor-associated neoangiogenesis in PBT24; both doses of NaDCA inhibited tumor-associated neoangiogenesis in SF8628. The 10 mM DCA inhibited the expression of markers tested in PBT24 and SF8628 tumors, but the 5 mM DCA affect on their expression depended on the cation. The DCA treatment did not affect the SLC12A2, SLC12A5, and SLC5A8 expression in cells but increased CDH1 expression in SF8628. The tumor response to DCA at different doses indicated that a contrast between NaDCA and MgDCA effectiveness reflects the differences in the tested cells' biologies.

The Japanese quail chorioallantoic membrane as a model to study an amphiphilic gradient copoly(2-oxazoline)s- based drug delivery system for photodynamic diagnosis and therapy research.

Amphiphilic gradient copoly(2-oxazoline)s are widely researched in the field of drug delivery. They could be used as a transport system for hydrophobic drugs such as hypericin (HYP). We prepared six gradient copolymers (EtOx)-grad-(ROPhOx) by living cationic ring-opening polymerization of a hydrophilic comonomer 2-ethyl-2-oxazoline (EtOx) and a hydrophobic comonomer 2-(4-alkyloxyphenyl)-2-oxazoline (ROPhOx), with different composition ratio (88:12 and 85:15) and three different alkyl chain lengths of alkyl (R) substituents. As an experimental model, Japanese quail chorioallantoic membrane (CAM) was used. The effect of nanoparticles loaded with HYP was evaluated by the changes of fluorescence intensity during photodynamic diagnosis (PDD) monitored under 405 nm LED light before administration, and 0,1,3 and 24 h after topical administration. The effectiveness of photodynamic therapy (PDT) (405 nm, 285 mW/cm2) applied 1h after the administration of HYP-loaded nanoparticles was evaluated using vascular damage score and histological sections. Molecular analysis was done by measuring angiogenesis-related gene expression by qPCR. The application of nanoparticles unloaded or loaded with HYP proved to be biocompatible, non-toxic, and undamaging to the CAM tissue, while they successfully altered the HYP fluorescence. We observed a possible anti-angiogenic potential of prepared nanoparticles, which could present an advantage for PDT used for tumour treatment.

Deep Learning-Based Image Analysis for the Quantification of Tumor-Induced Angiogenesis in the 3D In Vivo Tumor Model-Establishment and Addition to Laser Speckle Contrast Imaging (LSCI).

(1) Background: angiogenesis plays an important role in the growth and metastasis of tumors. We established the CAM assay application, an image analysis software of the IKOSA platform by KML Vision, for the quantification of blood vessels with the in ovo chorioallantoic membrane (CAM) model. We added this proprietary deep learning algorithm to the already established laser speckle contrast imaging (LSCI). (2) Methods: angiosarcoma cell line tumors were grafted onto the CAM. Angiogenesis was measured at the beginning and at the end of tumor growth with both measurement methods. The CAM assay application was trained to enable the recognition of in ovo CAM vessels. Histological stains of the tissue were performed and gluconate, an anti-angiogenic substance, was applied to the tumors. (3) Results: the angiosarcoma cells formed tumors on the CAM that appeared to stay vital and proliferated. An increase in perfusion was observed using both methods. The CAM assay application was successfully established in the in ovo CAM model and anti-angiogenic effects of gluconate were observed. (4) Conclusions: the CAM assay application appears to be a useful method for the quantification of angiogenesis in the CAM model and gluconate could be a potential treatment of angiosarcomas. Both aspects should be evaluated in further research.

Proteomic Analysis of Chicken Chorioallantoic Membrane (CAM) during Embryonic Development Provides Functional Insight.

In oviparous animals, the egg contains all resources required for embryonic development. The chorioallantoic membrane (CAM) is a placenta-like structure produced by the embryo for acid-base balance, respiration, and calcium solubilization from the eggshell for bone mineralization. The CAM is a valuable in vivo model in cancer research for development of drug delivery systems and has been used to study tissue grafts, tumor metastasis, toxicology, angiogenesis, and assessment of bacterial invasion. However, the protein constituents involved in different CAM functions are poorly understood. Therefore, we have characterized the CAM proteome at two stages of development (ED12 and ED19) and assessed the contribution of the embryonic blood serum (EBS) proteome to identify CAM-unique proteins. LC/MS/MS-based proteomics allowed the identification of 1470, 1445, and 791 proteins in CAM (ED12), CAM (ED19), and EBS, respectively. In total, 1796 unique proteins were identified. Of these, 175 (ED12), 177 (ED19), and 105 (EBS) were specific to these stages/compartments. This study attributed specific CAM protein constituents to functions such as calcium ion transport, gas exchange, vasculature development, and chemical protection against invading pathogens. Defining the complex nature of the CAM proteome provides a crucial basis to expand its biomedical applications for pharmaceutical and cancer research.

Human Lung Tissue Implanted on the Chick Chorioallantoic Membrane as a Novel In Vivo Model of Idiopathic Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal lung disease with no curative pharmacological treatment. Current preclinical models fail to accurately reproduce human pathophysiology and are therefore poor predictors of clinical outcomes. Here, we investigated whether the chick embryo chorioallantoic membrane (CAM) assay supports the implantation of xenografts derived from IPF lung tissue and primary IPF lung fibroblasts and can be used to evaluate the efficacy of antifibrotic drugs. We demonstrate that IPF xenografts maintain their integrity and are perfused with chick embryo blood. Size measurements indicate that the xenografts amplify on the CAM, and Ki67 and pro-collagen type I immunohistochemical staining highlight the presence of proliferative and functional cells in the xenografts. Moreover, the IPF phenotype and immune microenvironment of lung tissues are retained when cultivated on the CAM and the fibroblast xenografts mimic invasive IPF fibroblastic foci. Daily treatments of the xenografts with nintedanib and PBI-4050 significantly reduce their size, fibrosis-associated gene expression, and collagen deposition. Similar effects are found with GLPG1205 and fenofibric acid, two drugs that target the immune microenvironment. Our CAM-IPF model represents the first in vivo model of IPF that uses human lung tissue. This rapid and cost-effective assay could become a valuable tool for predicting the efficacy of antifibrotic drug candidates for IPF.

The 3D in vivo chorioallantoic membrane model and its role in breast cancer research.

PURPOSE: We aimed to evaluate the role of the chorioallantoic membrane model (CAM) in breast cancer research. METHODS: The following is an overview of the use of the CAM in the field of breast cancer research based on a PubMed literature query. RESULTS: The CAM is a 3D in vivo model that can be used for the analysis of tumor growth, biology and angiogenesis of primary tumor tissue or tumor cell lines. The CAM model has been used in breast cancer research for drug testing, migration assays and the evaluation of vascularization, amongst others. The CAM model is a valuable method that offers a better imitation of the physiological phenomena compared to 2D or 3D in vitro models. CONCLUSION: The CAM model has primarily and successfully been utilized for the assessment of the tumor biology of established breast cancer cell lines. Further, the CAM model is a promising method to analyze patient derived primary tumor material and could be used as a "patient-specific 3D-tumor-therapy-model" for the cost-efficient evaluation of anti-cancer drugs to find the optimal treatment for breast cancer patients.

Patterns of Tumor Vasculogenesis on a Chick Embryo Chorioallantoic Membrane In Vivo Model of Breast Cancer.

BACKGROUND/AIM: Understanding tumor vasculogenesis is a cornerstone for the inhibition of tumor progression. This study aimed to generate an in vivo breast cancer environment to analyze the patterns of tumor vasculogenesis. MATERIALS AND METHODS: Human mesenchymal stem cells (hMSC) and breast cancer MCF-7 cells (MCF-7) were seeded onto a chorioallantoic membrane (CAM) and, after a 7-day incubation, we performed a morphological and immunohistochemical analysis of CAM. RESULTS: hMSC and MCF-7 activated vasculogenesis and hematopoiesis on CAM. They stimulated the development of cord/capillary-like structures (CLS), formed by endothelial-like cells and hematopoietic cells. CLS presented a polygonal pattern, evolving towards a clearly visible plexus. Immunohistochemically, CLS were CD105+/AC133+/Oct3/4+, and the intensity was weak-moderate in the endothelial-like cells (inconstant) and weak in the hematopoietic cells. CONCLUSION: Tumor and embryonic vasculogenesis share a common paradigm, while CD105, AC133, and Oct3/4 were found to play a role in establishing the vasculogenic and hematopoietic stage.

Applications of the Chick Chorioallantoic Membrane as an Alternative Model for Cancer Studies.

A variety of in vivo experimental models have been established for the studies of human cancer using both cancer cell lines and patient-derived xenografts (PDXs). In order to meet the aspiration of precision medicine, the in vivomurine models have been widely adopted. However, common constraints such as high cost, long duration of experiments, and low engraftment efficiency remained to be resolved. The chick embryo chorioallantoic membrane (CAM) is an alternative model to overcome some of these limitations. Here, we provide an overview of the applications of the chick CAM model in the study of oncology. The CAM model has shown significant retention of tumor heterogeneity alongside increased xenograft take rates in several PDX studies. Various imaging techniques and data analysis have been applied to study tumor metastasis, angiogenesis, and therapeutic response to novel agents. Lastly, to practically illustrate the feasibility of utilizing the CAM model, we summarize the general protocol used in a case study utilizing an ovarian cancer PDX.

The Chick Embryo Chorioallantoic Membrane Model: A Research Approach for Ex Vivo and In Vivo Experiments.

BACKGROUND: The chick chorioallantoic membrane (CAM) model has attracted a great deal of interest in pharmaceutical and biological research as an alternative or complimentary in vivo assay to animal models. Traditionally, CAM assay has been widely used to perform some toxicological studies, specifically to evaluate the skin, ocular and embryo toxicity of new drugs and formulations, and to perform angiogenesis studies. Due to the possibility to generate the tumors onto the CAM, this model has also become an excellent strategy to evaluate the metastatic potential of different tumours and to test the efficacy of novel anticancer therapies in vivo. Moreover, in the recent years, its use has considerably grown in other research areas, including the evaluation of new anti-infective agents, the development of biodistribution studies and in tissue engineering research. OBJECTIVE: This manuscript provides a critical overview of the use of CAM model in pharmaceutical and biological research, especially to test the toxicity of new drugs and formulations and the biodistribution and the efficacy of novel anticancer and antiinfective therapies, analyzing its advantages and disadvantages in comparison to animal models. CONCLUSION: The chick chorioallantoic membrane model shows a great utility in several research areas, such as cancer, toxicology, biodistribution studies and anti-infective therapies. In fact, it has become an intermediate stage between in vitro experiments and animal studies, and, in the case of toxicological studies (skin and ocular toxicity), it has even replaced the animal models.