Clinical Kidney Journal, 2020, vol. 13, no. 3, 371–379 doi: 10.1093/ckj/sfz107 Advance Access Publication Date: 26 September 2019 Original Article ORIGINAL ARTICLE The ANTENATAL multicentre study to predict postnatal renal outcome in fetuses with posterior urethral valves: objectives and design Bénédicte Buffin-Meyer1,2, Julie Klein1,2, Loes F.M. van der Zanden3, Elena Levtchenko4, Panogiotis Moulos5, Nadia Lounis6, Françoise Conte-Auriol6, An Hindryckx7, Elke Wühl8, Nicola Persico9,10, Dick Oepkes11, Michiel F. Schreuder 12, Marcin Tkaczyk13, Gema Ariceta14, Magdalena Fossum15, Paloma Parvex16, Wout Feitz17, Henning Olsen18, Giovanni Montini19, Stéphane Decramer1,2,20,21, Joost P. Schanstra1,2 and the ANTENATAL Consortium* 1 Institut National de la Santé et de la Recherche Médicale (INSERM), U1048, Institut of Cardiovascular and Metabolic Disease, Toulouse, France, 2Université Toulouse III Paul-Sabatier, Toulouse, France, 3Department for Health Evidence, Radboud Institute for Health Sciences, Radboud University Medical Center, Nijmegen, The Netherlands, 4Department of Development & Regeneration, KU Leuven, Leuven, Belgium, 5HybridStat Predictive Analytics, Athens, Greece, 6Unité de Recherche Clinique Pédiatrique, Module Plurithématique Pédiatrique du Centre D’Investigation Clinique Toulouse 1436, Hôpital des Enfants, CHU Toulouse, Toulouse, France, 7Department of Obstetrics and Gynaecology, University Hospitals Leuven, Leuven, Belgium, 8Division of Pediatric Nephrology, Center for Pediatrics and Adolescent Medicine, University Hospital Heidelberg, Heidelberg, Germany, 9Department of Clinical Science and Community Health, University of Milan, Milan, Italy, 10Sergio Bonelli Centre for the Prevention of Renal Failure from Fetal to Pediatric Age, Fondazione Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy, 11Department of Prenatal Diagnosis and Therapy, Leiden University Medical Center, Leiden, The Netherlands, 12Department of Pediatric Nephrology, Radboudumc Amalia Children’s Hospital, Radboud Institute for Molecular Life Sciences, Nijmegen, The Netherlands, 13 Department of Pediatrics, Immunology and Nephrology, Polish Mother’s Memorial Hospital Research Institute, Lodz, Poland, 14Servei de Nefrologia Pediátrica Hospital, Universitario Vall d’Hebron, Universitat Autónoma de Barcelona, Barcelona, Spain, 15Section of Pediatric Urology, Department of Highly Specialized Pediatric Surgery and Pediatric Medicine, Karolinska University Hospital and Department of Women’s and Children’s Health, Karolinska Institutet, Stockholm, Sweden, 16Pediatric Nephrology, Unité Romande de Néphrologie Pédiatrique, Hôpitaux Universitaire Genève (HUG), Genève, Switzerland, 17For ERN eUROGEN, Department of Urology, Radboudumc Amalia Children’s Hospital, Radboud University Medical Center, Nijmegen, The Netherlands, 18For ERN eUROGEN, Paediatric Urology, Department of Urology, Aarhus Received: 9.7.2019; Editorial decision: 18.7.2019 C The Author(s) 2019. Published by Oxford University Press on behalf of ERA-EDTA. V This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/ licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com 371 372 | B. Buffin-Meyer et al. University Hospital & Aarhus University, Aarhus, Denmark, 19For ERN ERKNet, Pediatric Nephrology—Centro Sergio Bonelli for the Prevention and Treatment of Urinary Tract Malformations, Fondazione IRCCS Cà Granda, Ospedale Maggiore Policlinico, Milan, Italy, 20Service de Néphrologie Pédiatrique, Hôpital des Enfants, CHU Toulouse, Toulouse, France and 21Centre De Référence des Maladies Rénales Rares du Sud-Ouest (SORARE), Toulouse, France Correspondence and offprint requests to: Joost P. Schanstra; E-mail: joost-peter.schanstra@inserm.fr and Stéphane Decramer; E-mail: decramer.s@chu-toulouse.fr *All members of the ANTENATAL Consortium appear in the Acknowledgements. ABSTRACT Background. Posterior urethral valves (PUV) account for 17% of paediatric end-stage renal disease. A major issue in the management of PUV is prenatal prediction of postnatal renal function. Fetal ultrasound and fetal urine biochemistry are currently employed for this prediction, but clearly lack precision. We previously developed a fetal urine peptide signature that predicted in utero with high precision postnatal renal function in fetuses with PUV. We describe here the objectives and design of the prospective international multicentre ANTENATAL (multicentre validation of a fetal urine peptidome-based classifier to predict postnatal renal function in posterior urethral valves) study, set up to validate this fetal urine peptide signature. Methods. Participants will be PUV pregnancies enrolled from 2017 to 2021 and followed up until 2023 in >30 European centres endorsed and supported by European reference networks for rare urological disorders (ERN eUROGEN) and rare kidney diseases (ERN ERKNet). The endpoint will be renal/patient survival at 2 years postnatally. Assuming a ¼ 0.05, 1–b ¼ 0.8 and a mean prevalence of severe renal outcome in PUV individuals of 0.35, 400 patients need to be enrolled to validate the previously reported sensitivity and specificity of the peptide signature. Results. In this largest multicentre study of antenatally detected PUV, we anticipate bringing a novel tool to the clinic. Based on urinary peptides and potentially amended in the future with additional omics traits, this tool will be able to precisely quantify postnatal renal survival in PUV pregnancies. The main limitation of the employed approach is the need for specialized equipment. Conclusions. Accurate risk assessment in the prenatal period should strongly improve the management of fetuses with PUV. Keywords: development, kidney disease, obstructive uropathy, prediction, prenatal biomarkers INTRODUCTION In a small-scale proof-of-concept study, we recently identi- fied a fetal urinary peptide signature that predicted in utero the Posterior urethral valves (PUV) are a congenital anomaly asso- postnatal renal function of PUV fetuses with high accuracy [9], ciated with a wide spectrum of outcomes ranging from outperforming routine ultrasound and fetal urine biochemistry. extremely severe phenotypes with prenatal death to live born These results sparked the interest of many obstetricians, children with normal renal function. Exclusively found in paediatric urologists and nephrologists in using this fetal signa- males, PUV is the major cause of severe obstructive nephropa- ture in routine clinical care. On this background, we initiated thy in children. Although a rare disease (1 in 5000–8000 chil- an international investigator-driven prospective large-scale dren [1]), it accounts for 17% of paediatric end-stage renal study, named ANTENATAL (clinicaltrials.gov NCT03116217; disease (ESRD) [2–4]. ‘multicentre vAlidatioN of a fetal urine pepTidome-based The management of PUV is hampered by the absence of classifiEr to predict postnatal reNAl function in posterior tools for adequate antenatal screening of PUV fetuses at a high ureThral vALves’), aiming to validate peptide signature in a risk for developing ESRD. Current clinical practice is based on much larger multicentre setting. The ANTENATAL trial will en- prenatal ultrasound and fetal urine biochemistry, but this is rol 400 PUV pregnancies screened in >30 European centres with clearly insufficient to predict postnatal renal outcome with any involvement of two large European Reference Networks on rare accuracy in PUV patients as shown in several (meta) studies [5– diseases over a period of 4 years and with a patient follow-up 7]. This lack of efficient prognostic methods leads to many until the child’s age of 2 years. The ANTENATAL study will also parents opting for termination of pregnancy (TOP) [6–8], and be an excellent opportunity to evaluate the potential of adding also compromises adequate selection of fetuses that would po- other fetal urinary omics traits to peptide markers for kidney tentially benefit from prenatal intervention (vesicoamniotic disease stratification, including proteins, metabolites and shunting or laser ablation of valves). Hence, new predictive miRNAs, as we suggested in a recent study [10]. In addition, the strategies are necessary to allow optimal diagnostic and thera- ANTENATAL trial will allow exploration of the possibility of peutic management of the disease. extending the omics analysis to amniotic fluid for a less The ANTENATAL study announcement | 373 invasive prediction of postnatal renal function in PUV pregnan- [sodium, b2-microglobulin (b2M)]. Based on accuracy (Figure 1B), cies. The present report introduces an overview, the design and the 12PUV classifier correctly classified 35/38 patients [92% (95% objectives of the ANTENATAL trial. CI 84–100%)] compared with the best ultrasound (multicystic dysplastic kidneys) and biochemical (b2M) parameters, which correctly predicted 27/38 [71% (95% CI 57–85%)] and 26/38 ANTENATAL RATIONAL patients [68% (95% CI 54–83%)], respectively. This clearly dem- onstrating that the 12PUV classifier outperformed the conven- Body fluid peptide-based biomarker panels display clear prom- tional approaches. ise for the management of kidney disease because of their im- proved efficacy over single markers and the relative proximity of peptides to the phenotype compared with other omics traits [11, 12]. In the first attempt to show that peptides present in fe- ANTENATAL OBJECTIVES tal urine could serve as biomarkers to predict postnatal renal The primary objective of the ANTENATAL study is to prospec- function in PUV fetuses, we prospectively recruited 66 PUV tively validate the use of the fetal urine peptidome-based 12PUV patients from 26 different French centres. Using 28 PUV fetuses, classifier [9] to predict in utero postnatal renal function in we identified 26 fetal urinary peptides strongly associated with fetuses with PUV in a large international multicentre cohort. postnatal renal survival [9]. Twelve of those were combined in a The secondary objectives are to: (i) evaluate whether adding signature that we called the ‘12PUV classifier’. This signature additional omics traits (miRNAs, proteins and metabolites) to showed excellent performance in predicting the postnatal onset the 12 peptide markers further improves the stratification; of early ESRD in the 38 blindly assessed additional PUV fetuses (ii) explore the predictive biomarker content of amniotic fluid as with a sensitivity of 88% [95% confidence interval (95% CI) a less invasive biological fluid; (iii) compare the predictive effi- 66–98%], a specificity of 95% (95% CI 80–100%) and an area cacy of the 12PUV classifier for renal/patient survival at 2 years under the receiver operator characteristic curve (AUC) of 0.94 postnatally with conventional methods (fetal ultrasound and (95% CI 0.82–0.99) (Figure 1A). The predictive efficacy of the urine biochemistry); (iv) assess whether the accuracy of the 12PUV classifier was compared with routine clinical parame- 12PUV-mediated prediction is modified by intervention (e.g. ters, including ultrasound-measured characteristics of the vesico amniotic shunting or laser cystoscopy ablation of the fetal kidneys (hyperechogenicity, dysplasia, presence of cysts or valves according to local clinical practice); (v) determine the aspect of renal cortex) and amniotic fluid volume (oligohydram- stability and reproducibility of the 12PUV-mediated prediction nios or anhydramnios) as well as fetal urine biochemistry if multiple sequential fetal urine samples of a patient are FIGURE 1: Performance of a panel of 12 fetal urinary peptides (12PUV classifier) in predicting postnatal renal outcome in fetuses with PUV (proof of concept study). (A) Receiver operator characteristic curve for prediction of postnatal renal survival using the 12PUV classifier (38 fetuses, 16 with early ESRD, 22 without ESRD at 2 years postnatally). Adapted from Klein et al. [9] with permission. (B) Predictive accuracy of the 12PUV classifier compared with the clinical parameters including routinely ultrasound-measured characteristics of the fetal kidneys and amniotic fluid volume (oligoamnios or anhydramnios) as well as fetal urine biochem- istry. *P < 0.05, **P < 0.01, ***P < 0.001 versus the 12PUV classifier using the McNemar test for paired proportions. Adapted from Klein et al. [9] with permission. 374 | B. Buffin-Meyer et al. available; and (vi) have a standardized collection of pre- and megabladder in the second ultrasound; (iii) collection of fetal postnatal clinical and demographic data on the outcome of urine taken during the routine management of the disease for PUV pregnancies. biochemistry evaluation; and (iv) signature of a written in- formed consent. Non-inclusion criteria are: (i) persons protected by law and (ii) refusal to participate in the study. The exclusion ANTENATAL IMPLEMENTATION criteria include: (i) patients lost in follow-up, leaving the study or withdrawing consent; (ii) detection of major structural or Design, participants and setting chromosomal anomalies in karyotype; and (iii) lack of PUV con- ANTENATAL is an international multicentre prospective case firmation in the postnatal period or by anatomo-pathological study designed to validate the performance of the 12PUV classi- analysis in case of pregnancy termination or neonatal death. fier for in utero prediction of the postnatal renal outcome of PUV The setup of the ANTENATAL study is reported in Figure 2. fetuses. ANTENATAL anticipates to enrol 400 PUV pregnancies in >30 multidisciplinary prenatal diagnosis centres in nine Ethical aspects European countries (Belgium, France, Germany, Italy, Poland, The ANTENATAL study (clinicaltrials.gov NCT03116217, started Spain, The Netherlands, Switzerland and Sweden). Inclusion on 26 June 2017) was approved by the national ethics committee started in June 2017 and plans to end, under optimal conditions, for the coordinating centre (No. RCB2016-A01914-47, France). in June 2021, while follow-up will continue up to December The study is carried out in accordance with the ethical princi- 2023. Eligible patients are mothers carrying male singleton fetus ples expressed in the Declaration of Helsinki. In particular, with suspected PUV according to the following inclusion appropriate written informed consent for fetal urine sampling criteria: (i) detection of a megabladder associated with urinary and laboratory testing will be obtained from all parents of tract anomalies with or without dysplastic or hyperechogenic fetuses participating in the study, as previously indicated in the parenchyma in a first ultrasound; (ii) confirmation of inclusion criteria. FIGURE 2: ANTENATAL study setup. Four hundred patients recruited in >30 centres will enter the ANTENATAL study with a confirmed presence of a megabladder in the prenatal period. Fetal urine and/or amniotic fluid will be sampled. Fetal urine will be scored with the previously identified peptide-based 12PUV classifier and then compared with renal outcome at 2 years, and renal function at a number of time-points after birth. Fetal urine content in additional omics traits, including proteins, miRNAs and metabolites, will be explored to determine the added value of combining those traits with the peptide-based 12PUV classifier. Amniotic fluid samples will be screened for omics biomarkers of postnatal function in PUV to eventually move the analysis into a less invasive body fluid. Follow-up of all patients will be performed according to local standard care, which can be with or without intervention (vesicoamniotic shunt/laser ablation of valves). The ANTENATAL study announcement | 375 FIGURE 3: ANTENATAL study flow chart. The inclusion period will last for 48 months. The participation of each patient will carry on for 30 months, thereby resulting in a total duration of 78 months for the ANTENATAL trial. Outcomes of TOP or neonatal death, a standardized anatomo-pathological analysis will be proposed to the parents for the determination The primary judgment criterion (endpoint) is renal/patient of the severity of renal lesions (oligomeganephronia, dysplasia, survival at 2 years postnatally, defined by the need for dialysis presence of immature glomeruli and tubules). The final visit or death. The secondary judgment criterion is renal function of includes recording of documentation of surgery, any postnatal the live born at 1 month, 6 months and 1 year postnatally. complications, admissions to the hospital, number of urinary tract infections and medication during the first 2 years of life. Procedure and assessment Four postnatal visits are ideally planned during the 2-year post- natal follow-up, the minimum being one visit. The end of study Study organization. The study organization has been schema- visit equals the 24-month visit (M24), except for TOP or neonatal tized in Figure 3. An in-house-developed electronic case report death, where the last visit will serve as the end of study visit. form (eCRF, Supplementary data) has been established. According to local clinical practice to limit additional burden Biosamples and omics analysis. Details can be found in on the patients, fetal urine, amniotic fluid (if still present in a cer- Supplementary data. tain amount) and postnatal urine samples, as well as pre- and post-clinical data, will be collected. Clinical data can be anony- mously entered in the centralized eCRF at any time during Statistics follow-up. It is anticipated that recruitment will last for 4 years to To reach the primary objective, which is the validation of the fetal reach the 400 patients to be screened. An additional follow-up pe- urine peptidome-based 12PUV classifier to predict in utero postna- riod of at least 2 years will be required to obtain endpoint data for tal renal function in fetuses with PUV, we need to confirm its the last included patients. This period is paralleled by fetal uri- both sensitivity (88%) and specificity (95%). The 12PUV classifier nary peptidome analysis and initiation of the analysis of addi- comprises a scoring scheme, which includes positive and nega- tional omics traits in both fetal urine and amniotic fluid. tive predictive values (PPV and NPV, respectively). Power and sample size calculation for this specific setting and the validation Routine monitoring. Prenatal visits (Table 1) aim to obtain a set of both PPV and NPV have been addressed by Steinberg et al., de- of ultrasound-measured parameters of kidneys and urinary veloped especially for this type of diagnostic tests [13]. Hence, tract of PUV fetuses. A minimum of one prenatal visit is re- according to Steinberg’s method, focusing on the test’s specificity quired and optimally data from four prenatal visits are without compromising sensitivity, using a range of prevalence of expected. This will provide information on the antenatal evolu- ESRD in PUV individuals from 0.24–0.45 (mean: 0.35) [14, 15] and tion of the disease and will allow comparison of the efficacy of assuming a ¼ 0.05 (significance level) and 1  b ¼ 0.8 (power), we the omics-based prognosis tools with the routine clinical calculated that the sample size estimation to reach the primary parameters. Postnatal visits (Table 1) are scheduled to perform objective is 400 patients to be enrolled. anthropometric examination, postnatal biochemistry, and kid- ney and urinary tract ultrasound. The first postnatal visit fur- ther aims to confirm the presence of PUV (or confirm the correct DISCUSSION removal of the valves in case of prenatal laser intervention); it It is essential to confirm that any developed diagnostic or prog- will also serve to record the outcome of the pregnancy. In case nostic method also predicts well in, and thus is generalizable to, Table 1. Synopsis of clinical data obtained in the ANTENATAL study 376 Prenatal visits Postnatal visits | Prenatal Prenatal Prenatal Postnatal Postnatal Postnatal Data Screening Inclusion 14–23 WAa,b 23–32 WAa,b 32–41 WAa,b Birth–M1a,g M5–M7a,g M11–M13a,g M23–M25a,g Ultrasound of kidneys and urinary tract Morphologic datac X X X X X Doppler analysisd X X X Amniotic fluid volume X X X X Keyhole sign X Cervical length X X X X B. Buffin-Meyer et al. Confirmation of PUV X Outcome of pregnancy miscarriage, perinatal X mortality, neonatal mortality or survival Anthropometric examinatione X X X X Ultrasound of kidneys and urinary tract morphologic datac X X X X Documentation of any postnatal surgery X Documentation of any postnatal complica- X tionsincluding start of dialysis Admissions to hospital during first two year- X snumber and length Number of acute renal tract infections during X first two years lower urinary tract infection or ascending infection Medication during first two years X Fetopathology/anatomopathology if TOPf or X neonatal death a Range accepted. b Weeks of amenorrhea. c Kidney size and volume, absence or presence of renal hyperechogenicity, corticomedullary differentiation or renal cysts, kidney cortex thickness, pelvis and ureter maximum diameter if pelvic or ureter dilatation is present, respectively, bladder volume and wall thickness. d Renal artery pulsatility index PI) and peak systolic velocity PSV) on color and pulsed wave doppler. e Weight, length, head circumference, systolic and diastolic blood pressure. f Termination of pregnancy. g Months. The ANTENATAL study announcement | 377 ‘similar but different’ individuals outside the setting in which maternal plasma or urine even if such body fluids are easier to the method was developed [16]. The ANTENATAL trial is collect than amniotic fluid [18]. Furthermore, it appears that the expected to validate the previously identified 12PUV classifier risk of miscarriage and stillbirth in women undergoing amnio- [9] in the first and the largest prospective cohort focusing on centesis is mostly increased in high-risk pregnancies [20, 21]. fetuses with PUV. Here, validation will be simultaneously tem- Announcement of anomalies such as PUV is a traumatic and poral and geographical, since some new patients will be emotive event for families anticipating an unsecured long-term recruited in the same institutions (France) as in the develop- health outcome for their future newborn. In addition, the lim- ment sample, but at a later point in time, while other patients ited value of ultrasound-based analysis and fetal urinary bio- will be enrolled from eight additional countries. On 1 July 2019, chemistry in predicting postnatal ESRD led in the past to the eCRF contained 55 patient entries from 15 different centres. unnecessary fetal interventions, TOPs, sometimes contradicted Since PUV is a rare disorder, international collaboration by fetopathology, or continuation of pregnancies resulting in is necessary to obtain data from the patients needed for this early ESRD [5–8]. The introduction of an ANTENATAL-validated study. The European Reference Networks (ERNs) were estab- prognostic panel of fetal urinary peptides in the prenatal care lished to tackle rare conditions by bringing together healthcare of patients with PUV can guide clinicians in the counselling providers across Europe. ANTENATAL is an international study process by delivering unbiased and unambiguous prognostic in- that was initiated before the start of the ERNs, but after their formation. This will improve disease management in many acceptance, we approached both the centres participating in ways, impacting both affected patients and their family by: the European Rare Kidney Diseases Reference Network (ERN (i) reducing stress of parents related to the uncertainty of well- ERKNet, www.erknet.org; 31 July 2019, date last accessed) and being of their unborn infant; (ii) alleviating the enormous psy- the centres participating in the Rare Urogenital Diseases chological burden of parents associated with their decision Network (ERN eUROGEN, http://eurogen-ern.eu/; 31 July 2019, to continue or terminate pregnancy; (iii) potentially avoiding date last accessed). This led to additional centres including unnecessary interventions, stressful for the mother and risky patients for ANTENATAL, and endorsement of the ANTENATAL even if performed by fetoscopic surgery; (iv) potentially avoiding study by those two ERNs. unnecessary pregnancy termination; and (v) anticipating renal Collaboration was also established with the Radboudumc replacement therapy or palliative care of newborns if continua- Aetiologic research into Genetic and Occupational/ tion of the pregnancy is decided in fetuses predicted in the Environmental Risk Factors for Anomalies in Children high-risk stratum. (AGORA) data and biobank [17]. AGORA currently contains data of 500 PUV patients. To further enrich this resource, we asked the ANTENATAL centres to collect DNA samples of SUPPLEMENTARY DATA children and their parents and parental questionnaires on a Supplementary data are available at ckj online. range of non-genetic risk factors for AGORA. This collabora- tion allows the AGORA researchers to include more patients, and allows us to also evaluate the added value of genomic ACKNOWLEDGEMENTS biomarkers. We would like to thank Nadège Algans (Direction de la With 88% sensitivity and 95% specificity shown in the proof- of-concept study [9], we anticipate that the 12PUV classifier will Recherche, du Développement et de l’Innovation (DRDI), display superiority over conventional methods in predicting CHU de Toulouse, Toulouse, France) for her excellent orga- postnatal renal outcome. Beyond its best predictive accuracy, nizational assistance and the help of all the other DRDI- this fetal urine peptide-based tool will have the double benefit members who participate in the clinical trial. of being objective and quantifiable. Indeed, the measurement of fetal urine peptides is not subjected to personal interpretation, as can be the case for sonographic imaging. In addition, the Antenatal Consortium Investigators by recent and fast evolution of techniques for the analysis of Country the molecular content of biological fluids opens the possibility Belgium: An Hindryckx, Luc De Catte, UZ Leuven, Obstetrics of studying additional molecular levels, including miRNAs, and Gynaecology, Herestraat 49, 3000 Leuven, Belgium. proteins and metabolites. As demonstrated recently [10], the France: Christophe Vayssieres, Agnès Sartor, Marion combination of fetal urinary peptide and metabolite markers Groussolles, Christelle Plard, Paul Guerby, Laure Connan extracted from the same sample generated a clear added value and Mathieu Morin, CHU Toulouse 31059 Toulouse, France; with respect to metabolites alone for the prediction of progres- Elizabeth Simon and Jean Breaud, CHU Lenval 6200 Nice, sion of PUV disease. France; Anne-Hélène Saliou, Loic de Parscau, Nadine Jay and In addition to fetal urine, omics analysis in amniotic fluid dis- Isabelle Germouty, CHRU Brest 29609 Brest, France; plays promise for the clinic, since it has emerged as a reliable source of biomarkers for fetal diseases [18, 19]. In this context, we Gwenaelle le Bouar and Amelie Ryckewaert, CHU Rennes recently identified a novel amniotic fluid peptide signature that 35000 Rennes, France; Marie-Christine Manca-Pellissier and predicts postnatal renal function in bilateral congenital anoma- Thierry Merrot, CHU de la Timone 13385 Marseille, France; lies of kidney and urinary tract, including but not limited to PUV, Helene Laurichesse, D. Gallot and Lucie Bessenay, CHU thereby potentially strongly improving the prenatal diagnostic Estaing 63100 Clermont-Ferrand, France; Laurent Bidat and workup of the disease (clinicaltrials.gov NCT02675686, 178 Philippe Boize, CH René-Dubos 95303 Pontoise, France; fetuses, unpublished results). Hence, the expansion of biomarker Norbert Winer, Emma Allain-Launey and Claudine le research to amniotic fluid in the ANTENATAL project opens the Vaillant, CHU de Nantes 44093 Nantes, France; Fabienne possibility of predicting postnatal renal function in fetuses with Prieur and Marie-Pierre Lavocat, CHU Hôpitaux de St- PUV using a less invasive sample. Of note, the omics analysis of Etienne 42055 Saint-Etienne, France; Frederic Coatleven, Eric markers of PUV severity can most likely not be extended to Debromez, Jérôme Harembat and Brigitte Llanas, CHU 378 | B. Buffin-Meyer et al. Pellegrin 33078 Bordeaux, France; Romain Favre, Raphael Netherlands; Michiel Schreuder, Pediatric Nephrology, Moog and Ariane Zaloszyc, SIHCUS-CMCO 67300 Radboudumc Amalia Children’s Hospital, P.O. Box 9101, Schiltigheim and Hôpital de Hautepierre 67200, Strasbourg, 6500 HB Nijmegen, The Netherlands; Frank Vandenbussche, France; Jérôme Massardier and Delphine Demede, Hôpital Obstetrics and Gynecology, Radboudumc Amalia Children’s Femme Mère Enfant 69677 Lyon, France; Franck Perrotin Hospital, P.O. Box 9101, 6500 HB Nijmegen. and Sylvie Cloarec, CHRU de Tours and Hôpital Clocheville Poland: Marcin Tkaczyk and Małgorzata Stan  czyk, Pediatrics 37044, Tours, France; Valérie Vequeau-Goua and Immunology and Nephrology, Polish Mothers Memorial Emmanuelle Descombes, CHU Poitiers 86021 Poitiers, Hospital Research Institute, 281/289 Rzgowska Str., 93-338 France; Pierre Boulot, Denis Morin, Florent Fuchs and Julie , Poland; Krzysztof Szaflik, Justyna Wojtera and Łódz Tenenbaum, CHU Montpellier 34295 Montpellier, France; Waldemar Krzeszowski, Gynecology, Fertility and Fetal Yves Ville, Thomas Blanc and Laurence Heidet, AP-HP, hôpi- Therapy, Polish Mothers Memorial Hospital Research tal Necker 75015 Paris, France; Anne Paris, Hôpital Bagatelle Institute, 281/289 Rzgowska Str., 93-338 Łódz , Poland; 33400 Talence, France; Eric Dobremez and Marie-Françoise Tomasz Talar and Barbara Pawłowska, Neonatology, Polish Froute, CHU Pellegrin 33078 Bordeaux, France; Jean Gondry, Mothers Memorial Hospital Research Institute, 281/289 Charles Muszynski, Elodie Haraux, Fabienne Lobelle and Rzgowska Str., 93-338 Łódz , Poland; Katarzyna Fortecka- Julien Chevreau, CHU Amiens 80480 Amiens, France; Piestrzeniewicz, Intensive Care and Congenital Jonathan Rosenblatt, Véronique Baudoin and Georges Malformations of Neonate and Infant, Polish Mothers Deschenes, AP-HP, Robert-Debré 75019 Paris, France; Memorial Hospital Research Institute, 281/289 Rzgowska Virginie Guigue, Florence Amblard and Guylhène Bourdat- Str., 93-338 Łódz, Poland; Dariusz Olejniczak, Pediatric Michel, CHU Grenoble 38043 Grenoble, France. Surgery and Urology, Polish Mothers Memorial Hospital Germany: Elke Wühl and Franz Schaefer, Pediatrics Research Institute, 281/289 Rzgowska Str., 93-338 Łódz , Department, Heidelberg University Hospital Im Poland. Neuenheimer Feld 430 60120 Heidelberg, Germany; Michael Spain: Gema Ariceta, Pediatric Nephrology, Hospital Elsässer, Gynecology & Obstetrics, Heidelberg University Universitari Vall d’Hebron, Pg/Vall d’Hebron 119-129, 08035 Hospital Im Neuenheimer Feld 430 60120 Heidelberg, Barcelona, Spain; Elena Carreras, Obstetrics, Hospital Germany. Universitari Vall d’Hebron, Pg/Vall d’Hebron 119-129, 08035 Italy: Nicola Persico and Federica Rossi, Obstetrics and Barcelona, Spain; Silvia Arevalo and Carlota Rodo, Fetal Gynecology, Fondazione IRCCS Cà Granda, Ospedale Medicine, Hospital Universitari Vall d’Hebron, Pg/Vall d’ Maggiore Policlinico, Milan, Italy; Gianantonio Manzoni, Hebron 119-129, 08035 Barcelona, Spain; Alexandra Navarro, Pediatric Urology—Centro Sergio Bonelli for the Prevention Pathology, Hospital Universitari Vall d’Hebron, Pg/Vall d’ and Treatment of Urinary Tract Malformations, Fondazione Hebron 119-129, 08035 Barcelona, Spain. IRCCS Cà Granda, Ospedale Maggiore Policlinico, Milan, Sweden: Magdalena Fossum, Section of Pediatric Urology, Italy; Erika A. De Marco, Pediatric Urology—Centro Sergio Department of Highly Specialized Pediatric Surgery and Bonelli for the Prevention and Treatment of Urinary Tract Pediatric Medicine, Karolinska University Hospital and Dept. Malformations, Fondazione IRCCS Cà Granda, Ospedale of Women’s and Children’s Health, Karolinska Institutet, Maggiore Policlinico, Milan, Italy; Giovanni Montini, SE 171 76 Stockholm, Sweden; Gillian Barker, Section of Pediatric Nephrology—Centro Sergio Bonelli for the Pediatric Urology, Department of Highly Specialized Prevention and Treatment of Urinary Tract Malformations, Pediatric Surgery and Pediatric Medicine, Karolinska Fondazione IRCCS Cà Granda, Ospedale Maggiore University Hospital SE 171 76 Stockholm, and Akademiska Policlinico, Milan, Italy; Valentina Capone, Pediatric Children’s hospital, Uppsala, Sweden; Peter Lindgren, Nephrology—Centro Sergio Bonelli for the Prevention and Department of Obstetrics and Gynecology, Karolinska Treatment of Urinary Tract Malformations, Fondazione University Hospital, SE 141 86 Stockholm, Sweden. IRCCS Cà Granda, Ospedale Maggiore Policlinico, Milan, Switzerland: Paloma Parvex and Hassib Chehade, Pediatric Italy; Leonardo Caforio, Medical and Surgical Neonatology, Nephrology Unité Romande de Néphrologie Pédiatrique, Unit of Medical and Surgical Fetal Therapy, Bambino Gesù Hôpitaux universitaire Genève (HUG), 6 rue Willy-Donzé, Children’s Hospital—Rome, Piazza S. Onofrio 4, 00165, CH-1211 Genève, Switzerland; Jacques Birraux, Pediatric Rome, Italy; Antonio Zaccara, Pediatric Urology Therapy, Urology, Centre Universitaire Romand de Chirurgie Bambino Gesù Children’s Hospital—Rome, Piazza S. Onofrio Pédiatrique Hôpitaux Universitaire Genève (HUG), 6 rue 4, 00165, Rome, Italy; Michele Innocenzi, Pediatric Urology, Willy-Donzé, CH-1211 Genève, Switzerland; Gianmaria Bambino Gesù Children’s Hospital—Rome, Piazza S. Onofrio Pellegrinelli, Gynéco-Obstétrique, Hôpitaux Universitaire 4, 00165, Rome, Italy; Pietro Bagolan and Nicola Capozza, Genève (HUG), 6 rue Willy-Donzé, CH-1211 Genève, Bambino Gesù Children’s Hospital—Rome, Piazza S. Onofrio Switzerland. 4, 00165, Rome, Italy; Marco Castagnetti and Mariangela Mancini, Urology Department, Azienda Ospedaliera di Padova, Via Giustiniani, 2-35128—Padova, Italy. FUNDING The Netherlands: Dick Oepkes and Phebe Adama van The study is partially made possible by support of the Scheltema, Department of Prenatal Diagnosis and Therapy, ‘Programme Hospitalier de Recherche Clinique and ‘La Leiden University Medical Center, Albinusdreef 2, 2333 ZA Foundation de la Recherche Médicale (grant number Leiden, The Netherlands; Wout Feitz and Barbara DEQ20170336759, France). M.T. was supported by the Polish Kortmann, Pediatric Urology, Radboudumc Amalia Mothers Memorial Hospital Research Institute (internal Children’s Hospital, P.O. Box 9101, 6500 HB Nijmegen, The grant number 2016/IV/54-GW. LvdZ is supported by a Kolff The ANTENATAL study announcement | 379 grant from the Dutch Kidney Foundation (13OKJ36) and a 10. Buffin-Meyer B, Klein J, Breuil B et al. Combination of the fetal ZonMW-VENI grant from the Netherlands Organisation for urinary metabolome and peptidome for the prediction of Scientific Research (91618036). This project has been sup- postnatal renal outcome in fetuses with PUV. J Proteomics ported by ERN ERKNet and ERN eUROGEN, which are partly 2018; 184: 1–9 co-funded by the European Union within the framework of 11. Mischak H, Delles C, Vlahou A, Vanholder R. Proteomic bio- the Third Health Programme ‘ERN-2016–Framework markers in kidney disease: issues in development and im- Partnership Agreement 2017–2021’. plementation. Nat Rev Nephrol 2015; 11: 221–232 12. Decramer S, Wittke S, Mischak H et al. 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