Biomarkers and Outcome Predictors of Pediatric Nephrotic Syndrome: A Genetic, Transcriptomic, and Secretome Multiomics Study

Biomarkers and Outcome Predictors of Pediatric Nephrotic Syndrome: A Genetic, Transcriptomic, and Secretome Multiomics Study

Idiopathic Nephrotic Syndrome is a rare disease of the kidneys, which typically affects children. For most affected children there is the need of a prolonged treatment with drugs reducing the activity of the immune system, also resulting in many side effects. Those patients, who do not respond to treatment, are at risk of kidney damage and of dialysis or kidney transplantation. It is currently impossible to predict the response to treatment, leading to unnecessary therapies with side effects as well as unclear prognosis in the affected children. The response of the idiopathic nephrotic syndrome to medications acting on the immune system explains its important role in the occurrence of the disease.

With this study we aim to obtain predictors of the response to treatment right at the beginning of the disease, to adapt the therapy avoiding needless side effects. This will be done evaluating the blood and urine of affected children using state of the art molecular characterisation. We will evaluate the genetic predisposition, the cell trait changes and the presence of molecules in blood and urine that may affect the interaction between the immune system and the kidneys. We expect that the findings will improve treatment of children with idiopathic nephrotic syndrome and reduce the number of children suffering from unnecessary drugs related side effects.

The PRECISE study (Biomarkers and Outcome Predictors of Pediatric Nephrotic Syndrome) is an innovative, multicenter research project focused on advancing our understanding of pediatric nephrotic syndrome (NS) and improving treatment outcomes for affected children across Europe. This project, which involves multiple European institutions, centers around discovering predictive biomarkers using advanced genetic, transcriptomic, and secretome multiomics methods. Nephrotic syndrome, a complex kidney disease that primarily impacts children, can lead to significant health challenges and often necessitates prolonged treatment with immunosuppressive drugs that carry side effects.

Overview of Nephrotic Syndrome

Nephrotic syndrome is characterized by symptoms such as proteinuria (excessive protein in the urine), hypoalbuminemia (low blood albumin levels), edema (swelling), and hyperlipidemia (high blood lipids). Pediatric nephrotic syndrome, known for its unpredictable nature, is categorized as Idiopathic Nephrotic Syndrome (INS) and affects around 2-7 children per 100,000 each year. While some children respond well to steroid treatment (steroid-sensitive), a significant proportion may become steroid-resistant, risking severe kidney damage that may lead to dialysis or even kidney transplant.

Why This Study is Important

Currently, it is challenging to predict which children will respond to treatment, how their disease will progress, or which children may experience repeated relapses. The standard treatment with immunosuppressive drugs, although effective for some, can bring numerous side effects, especially with prolonged use. Therefore, one of the most pressing unmet needs in pediatric nephrology is to develop early predictive biomarkers for individualized treatment plans, ultimately reducing unnecessary drug exposure and improving long-term outcomes.

The PRECISE study is designed to fill this critical gap by identifying genetic and molecular markers that can help clinicians:

1. Predict Treatment Response - Identify children who may or may not respond to steroid therapy.
2. Anticipate Disease Progression - Understand which patients are more likely to experience relapses or progress toward kidney failure.
3. Personalize Therapy - Tailor treatments based on each patient's unique biological profile to minimize side effects and maximize effectiveness.

Objectives of the Study

The primary goal of the PRECISE study is to create a predictive model for differentiating children with Steroid-Resistant Nephrotic Syndrome (SRNS) or Steroid-Dependent Nephrotic Syndrome (SDNS) from those with Infrequent Relapsing Nephrotic Syndrome (IRNS) right at the onset of the disease. This model will integrate various data sources, including omics, genetic, and clinical information, to produce a reliable "biomarker signature" that can guide therapeutic decisions from the start.

The study has several secondary objectives, including:

1. Genetic and Epigenetic Analysis - Identifying the genetic and epigenetic factors that contribute to nephrotic syndrome's pathogenesis.
2. Immune System Profiling - Understanding how adaptive immune system alterations may influence disease onset and progression.
3. Molecular Characterization - Using advanced proteomics to identify distinct protein patterns in serum and urine that differ between disease subtypes.
4. Healthy Controls Comparison - Analyzing the molecular characteristics of healthy children to enhance the accuracy of disease-specific biomarkers.

Study Design and Methodology

The PRECISE study is an open, multicenter, observational study. It will enroll a total of 310 children diagnosed with INS and an additional control group of 40 healthy children. The study is structured as follows:

1. Discovery Phase - This phase will involve 110 treatment-naive children with INS who will be followed for 12 months. During this period, biological samples will be collected at several key points, such as at the time of diagnosis, during remission, and in case of relapse. These samples will undergo extensive molecular profiling, including genetic, proteomic, and transcriptomic analyses, which will enable the identification of key biomarkers.
2. Validation Phase - In this phase, an independent cohort of 200 INS patients (prevalent cases) will be studied to confirm the reliability of biomarkers identified in the discovery phase.

Key Elements of the Study Protocol

Inclusion Criteria for Participants

For the main study group, children must be aged between 1 and 18 years and have a clinical diagnosis of idiopathic nephrotic syndrome. They should not have undergone any previous treatment for INS and must meet specific criteria for disease markers, such as proteinuria and hypoalbuminemia. A validation cohort includes children with similar characteristics but who have already been treated, while a control group comprises healthy children with no proteinuria and minor congenital kidney conditions.

Data Collection and Sampling

At each assessment visit, clinical data will be gathered, including physical exam results, medical history, vital signs, and laboratory values. Biological samples (blood and urine) will also be collected and stored at centralized biobanking facilities, with samples being analyzed for a wide range of molecular markers. The study uses high-level biostatistics and bioinformatics to interpret the complex data, aiming to provide a multidimensional view of each participant's health profile.

Analysis and Expected Outcomes

Using advanced statistical techniques and machine learning algorithms, researchers will analyze differences in genetic, immune, and molecular markers among the different patient groups. The primary endpoint is to develop molecular signatures predictive of disease progression, which can differentiate between SRNS, SDNS, and IRNS groups. Secondary endpoints include:

• Genetic Predispositions - Assessing variants in immune- and kidney-related genes that may be associated with disease progression.
• Immune Alterations - Evaluating specific immune cell types and their behaviors, which may be linked to the risk of relapse or resistance to treatment.
• Proteomic Profiles - Identifying protein patterns in blood and urine that are unique to the disease and could be used to classify disease stages and predict outcomes.

Impact and Implications

The PRECISE study represents a major step forward in pediatric nephrology by leveraging cutting-edge scientific methods to address the pressing challenges faced by children with nephrotic syndrome. By identifying biomarkers that predict treatment response and disease progression, this study can help tailor treatments to each child's needs, reducing the risk of adverse side effects and improving overall outcomes.

Consortium and Collaborating Institutions

The project is led by the Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico in Milan, Italy, under the guidance of Principal Investigator Prof. Giovanni Montini. It brings together several renowned research centers and universities across Europe, each contributing unique expertise in pediatric nephrology, genetics, and molecular biology. Key partners include Heidelberg University, University Hospital of Cologne, Medical University of Gdansk, Istanbul University, and Vilnius University, along with patient advocacy organizations such as NephCEurope and various national nephrology groups.

Through this consortium, the study integrates expertise and resources from across Europe, ensuring a robust and comprehensive approach to this challenging area of pediatric medicine.

Summary

In summary, the PRECISE study is a pioneering effort to improve the diagnosis, prognosis, and treatment of pediatric nephrotic syndrome through the identification of precise molecular biomarkers. This study not only seeks to benefit children affected by nephrotic syndrome today but also lays the groundwork for future research in rare kidney diseases. By aligning clinical care with the latest in genetic and molecular science, the PRECISE study aims to provide new hope to families affected by this chronic and often debilitating condition.

Inclusion Criteria:

• Clinical diagnosis of idiopathic nephrotic syndrome (INS) with nephrotic range proteinuria (uPr/uCr ratio > 2 mg/mg).
• Hypoalbuminemia with serum albumin < 3.0 g/dL.
• Presence of edema.
• No prior treatment for idiopathic nephrotic syndrome.
• Age between 1 and 18 years at the time of enrollment.
• igned informed consent by a parent or legal guardian.

Exclusion Criteria:

• Diagnosis of congenital or infantile nephrotic syndrome (age < 1 year).
• Diagnosis of secondary nephrotic syndrome.
• Presence of glomerulonephritis, autoimmune diseases, or vasculitis.
• Lack of signed informed consent by a parent or legal guardian.
• Previous treatment with prednisone or prednisolone for nephrotic syndrome.