UNIVERSITÀ DI ROMA LA SAPIENZA

CardioNox2: Nox2 and cardiovascular diseases

Cardiovascular diseases (CVDs) are one of the major health issues in Western countries. Cardiovascular risk factors such as diabetes, hypertension and smoking habits contribute to the onset of CVDs and are all associated with an increased oxidative stress. We analyse oxidative stress levels by a method patent by us. This method relies on the identification of a soluble peptide of the NOX (sNOX2-dp), enzyme principally involved in the production of reactive oxygen species. Assessing NOX2 levels in patients with one or more cardiovascular risk factors could help the clinician to stratify the risk of cardiovascular complications.

Patent Status

GRANTED

Priority Number

102017000141849

Priority Date

07/12/2017

License

INTERNATIONAL

Market

Cardiovascular diseases (CVDs) are the most common chronic diseases in industrialized countries. CVDs have a serious impact on quality of life and life expectancy of people and on the health resources. The analysis of Nox2 in patients with cardiovascular risk factors could find a very large market considering the large number of patients in Europe who suffer from these pathologies. In particular, the number of patients suffering from cardiovascular diseases is around 80 million and are responsible for 3.9 million deaths a year. This method may be useful as a support to identify/stratify patients with a predisposition to cardiovascular diseases. Finally, the sNOX2-dp assay’s could provide an immediate approach for the oxidative stress diagnosis.

Problem

To date, oxidative stress is evaluated through indirect methods, expensive and difficult to implement in the clinical field. We have developed, patented and make a prototype for the evaluation of oxidative stress by a method direct and easy to implement. In particular, we have identified a soluble NOX2–derived peptide “sNOX2-dp” that is released in the systemic circulation following the activation of the NOX2 enzyme during an inflammatory process and can be detected by an ELISA method. NOX2 is an isoform of NADPH oxidase, one of the main enzymes able to generating reactive oxygen species (ROS). Increasing ROS production by NOX2 leads to oxidative stress, a condition associated with the onset various diseases such as cardiovascular disease. The application of this method on patients with cardiovascular risk factors will allow an early diagnosis and a stratification of the risk of onset cardiovascular complications in this pathological setting.

Current Technology Limits

Oxidative stress is evaluated through indirect and expensive methodology hard to apply in the clinical setting and unfeasible in all laboratories. We have patented and developed a prototype for the assessment of oxidative stress using a method direct and easy to apply by ELISA procedure. The application of this method can be performed in any type of laboratory as the method is easy to perform and inexpensive.

Killer Application

The main applications of this method are:

Direct measurement of oxidative stress levels
Stratification of risk/cardiovascular complications

Our Technology and Solution

The sNOX2dp ELISA Kit utilizes a microplate-based enzyme immunoassay (ELISA) technique for the semi quantitative measurement of sNOX2-dp in human samples.

The assay is based on:

Immobilization of sNOX2dp contained in standards, positive reference or test samples into 96 well plates;
Addition of the anti-sNox2-dp monoclonal antibody against the amino acidic sequence (224–268) of the extra membrane portion of Nox2;
Detection and quantification of immobilized complex by using an HRP conjugated secondary antibody.

The enzyme activity is measured spectrophotometrically at 450 nm. Optical Density is directly proportional to the amount of sNox2-dp in the unknown samples and can be derived by interpolation from a reference curve generated by serial dilutions of a reference standard. Values were expressed as Arbitrary Units (AU) per microgram of sample. Intra- and Inter- assay are <8% and <15% respectively and accuracy of 107%.

Advantages

The method for the analysis of oxidative stress by the detection of soluble NOX2 derive peptide “sNOX”-dp” was found to be an accurate, specific and reliable method. The application of this method in patients who have one or more cardiovascular risk factors could be of help in the early detection of the development of cardiovascular complications. The stratification of the risk of developing cardiovascular complications is important to reduce the costs and the treatment time. This method identifies specific markers of oxidative stress in patients at high cardiovascular risk.

The method has a low analytical cost and provides rapid results.

The method has a wide application because it is based on an ELISA test

Roadmap

The validation and production of a commercial kit for the detection of the sNOX2-dp will require additional investments such as:

Validation of the method by clinical trial on a large population with cardiovascular risk factors;
Identification of a company that will be involved in the production and distribution of the sNOX2-dp ELISA KIT.

Review the Technology

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Team

Simona Bartimoccia

Research fellow at Università di Roma - La Sapienza

Research fellow at the Clinical Internal, Anaesthesiological and Cardiovascular Sciences Sapienza University of Rome. She earned hes PhD in “CARDIO-THORACO-VASCULAR PHYSIOPATHOLOGY AND IMAGING” at the Sapienza University of Rome. Her experience concerns the study of oxidative stress and platelet activation pathways in several pathological setting associated to atherosclerosis.

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Vittoria Cammisotto

Reseacher at l'Università di Roma - La Sapienza

Researcher at the Clinical Internal, Anaesthesiological and Cardiovascular Sciences, Sapienza University of Rome. She earned his PhD in “CARDIO-THORACO-VASCULAR PHYSIOPATHOLOGY AND IMAGING” at the Sapienza University of Rome. She specialises in “Clinical Pharmacology and Toxicology” and her interest and experience concern the study of platelet pathophysiology. This specifically deals with the pathways of platelet activation, from oxidative stress mediated by the activation of NOX2 up to the most recent studies on the role of Lipopolysaccharide and the functional modulation of such pathways through pharmacological or nutraceutical approaches.

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Cristina Nocella

Assistant Professor at Università di Roma - La Sapienza

Assistant Professor at the “Sapienza” University of Rome. She holds a Biological Sciences degree at the “Sapienza” University of Rome where she also completed a PhD in Immunological, Haematological and Rheumatological Sciences. Her scientific interest and experience concern the study of mechanisms, which play a role in the onset of cardiovascular disease. Specifically, she study the pathways of platelet activation, from oxidative stress-mediated by the activation of NOX2 up to the most recent studies about on platelet activation mediated by various promoting factors such as LPS. Finally, her interest is about the functional modulation of these pathways through pharmacological or nutraceutical approaches.

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Francesco Violi

Professor Emeritus at Sapienza University of Rome

Professor Emeritus at Sapienza University of Rome. He studies the mechanisms of platelet activation and oxidative stress in clinical models of atherosclerosis. He works on human models of genetic deletion inherent in gp91phox, catalytic "core" of the NADPH oxidase demonstrating its involvement in platelet thrombus formation. He conducted and analysed the risk of cardiovascular events associated with the use of drugs and the relationship between atherosclerotic risk factors and cardiovascular events in several pathological setting.

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Pasquale Pignatelli

Full Professor at the Clinical Internal Anaesthesiological and Cardiovascular Sciences, Sapienza University of Rome

Full Professor at the Clinical Internal Anaesthesiological and Cardiovascular Sciences, Sapienza University of Rome. He studies problems relating to atherothrombotic pathology and the mechanisms involved such as oxidative stress, endothelial dysfunction and platelet activation. He is director of the UOC of internal medicine and prevention of atherosclerosis which is divided into various ambulatories capable of designing personalized programs for the prevention of atherosclerosis and thrombosis.

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Roberto Carnevale

Associate Professor at the Department of Medical-Surgical Sciences and Biotechnologies, Sapienza University of Rome

Associate Professor at the Department of Medical-Surgical Sciences and Biotechnologies, Sapienza University of Rome. He earned his PhD in Human Pathology at the “Sapienza” University of Rome. His scientific interest and experience have been completely committed to and involved in the pathophysiology of atherosclerosis. Most of his studies focused on research direct and non-invasive biomarkers for the diagnosis of oxidative stress.

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