Presentation on theme: "17.-20. May 2012 Consensus in Pediatrics Moscow Proteomics in Pediatrics Disclosure ES is an employee of mosaiques diagnostics GmbH. Presentation does."— Presentation transcript:
May 2012 Consensus in Pediatrics Moscow Proteomics in Pediatrics Disclosure ES is an employee of mosaiques diagnostics GmbH. Presentation does not include discussion of off-label or investigational use Eric Schiffer 1,2, Jens Drube 3, Lars Pape 3, Jochen HH. Ehrich 3 1 mosaiques diagnostics GmbH, Hannover (Germany), 2 Department of Biomarkers and Systems Medicine, University of Glasgow, Glasgow (UK). 3 Clinic of Pediatric Kidney, Liver and Metabolic Diseases, Hannover Medical School, Hannover, Germany
May 2012 Consensus in Pediatrics Moscow Genomics and Proteomics Genomics approx. 20,000 genes relatively invariant / static Explains vulnerability for a disease static Proteomics approx. 1,000,000 proteins highly dynamic Explains environmental influences dynamic
May 2012 Consensus in Pediatrics Moscow Concentration of analyte Compliance Degree of invasion ProteolysisProximity to disease Tissue Bile Blood Urine Sources of Protein Biomarkers
May 2012 Consensus in Pediatrics Moscow Proteomics Platforms: Pros and Cons 2DE-MSSELDI-TOFLC-MSCE-MS
May 2012 Consensus in Pediatrics Moscow Capillary Electrophoresis coupled to Mass Spectrometry Urine profile SOP QM MIAPE guidelines* *Taylor et al. (2007) Nat. Biotechnol. 25:
May 2012 Consensus in Pediatrics Moscow Low molecular weight urinary proteome profiling
May 2012 Consensus in Pediatrics Moscow Normal Control Amp 2,0 Frequency 28/267 Cardiovascular Disease Amp 3,0 Frequency 22/40 Diabetic Nephropathy Amp 4,3 Frequency 25/34 Bladder Cancer Amp 3,5 Frequency 20/30 Prostate Cancer Amp 2,9 Frequency 49/153 Statistics for Marker ID ID90840 Mass Da CE time22.40 min SequenceMIEGNTKSPLFMGKVVNPTQK Peptidealpha-1-Antitrypsin [aa ] Protein ID: Amp P NC CVD DN BC PC Schiffer et al., Proteomics 2006, 6: Statistical biomarker definition
May 2012 Consensus in Pediatrics Moscow Unblinding Sensitivity and Specificity 94% 89% Clinical Study Design CASE diseased treated (Drug) CONTROL healthy not treated (Placebo) discriminatory pattern compiled pattern discriminatory biomarkers individual analyses
May 2012 Consensus in Pediatrics Moscow DeToni-Debré-Fanconi Syndrome (FS) FS Control CKD
May 2012 Consensus in Pediatrics Moscow FS Control CKD N=11 N=9 N=294 FS: Clinical Validation Blinded: 11 FS 9 Controls AUC=0.86 Open-label:11 FS 294 CKD AUC=0.84 dashed Drube et al. (2009) Nephrol. Dial. Transplant. 24: 2161–2169.
May 2012 Consensus in Pediatrics Moscow Peptide ID Mass (Da) CE-time (min) Median FS (counts) Median HC (counts) maxT (p-value) SequenceProtein VLNLGPITRUromodulin DGPpGRDGQpGHKGCollagen alpha-2 (I) YKRKANDESNEHSOsteopontin PpGEAGKpGEQGVPGDLGCollagen alpha-1 (I) GEpGSpGENGApGQMGPRGCollagen alpha-1 (I) GDDGEAGKPGRpGERGPpGPCollagen alpha-1 (I) GDDGEAGkPGRpGERGPpGPCollagen alpha-1 (I) DGKTGpPGPAGQDGRPGPpG ppG Collagen alpha-1 (I) PpGESGREGAPGAEGSpGRD GSpGAKGDRGETGP Collagen alpha-1 (I) In FS decreased / increased urinary excretion Sequenced marker proteins The marker peptides are fragments derived from osteopontin, uromodulin and collagens
May 2012 Consensus in Pediatrics Moscow FS: Conclusions CE-MS can be used to specifically diagnose FS in pediatric patients CE-MS might be a future tool for the non-invasive diagnosis of FS. Reduced levels of osteopontin and uromodulin might indicate loss of tubular function regardless of underlying cause Fragments of the collagen alpha-1 (I) might hint to change of proteases in collagen degradation as observed in interstitial fibrosis. Biomarkers suggest fibrosis as an early event in the development of renal insufficiency in FS
May 2012 Consensus in Pediatrics Moscow spontaneous remission Surgery ? Prognosis Surgery yes or no Sensitivity94% (21/23) Specificity100% (13/13) Decramer et al. (2006) Nat. Med. 12(4): Ureteropelvic junction obstruction (UPJO)
May 2012 Consensus in Pediatrics Moscow UPJO: Clinical Validation Application of the Decramer et al. pattern to older children Prospective cohort with hydronephrosis enroled at Hannover Diuretic renal scan to identify urodynamically relevant UPJO N=19 1 year of age Sensitivity 83% (5/6) Specificity 92% (12/13) AUC=88%, P< N=8 > 1 year of age Sensitivity 20% (1/5) Specificity 66% (2/3) AUC=57%, P=0.7655
May 2012 Consensus in Pediatrics Moscow UPJO: Cost-effectiveness Marcov process decision tree model Comparison of watchful waiting / imaging to urinary proteomics Incremental gain of $8,000 per QUALY Results insensitive to any included cost parameter Application to 2,000 newborns would save $US 16 Mio. per year while improving quality of life
May 2012 Consensus in Pediatrics Moscow UPJO: Conclusions Confirmation of results reported by Decramer et al. in infants 1 year of age, Urinary proteome analysis predicted obstruction with 83% sensitivity and 92% specificity In older patients sensitivity decreased to 20% and specificity to 66% The proteome pattern established by Decramer and coworkers predicts the need for surgery in infants but not in older children with UPJO Drube et al. (2010) Pediatr. Nephrol. 25:1673–1678..
May 2012 Consensus in Pediatrics Moscow Vesicoureteral reflux (VUR) High grade VUR (grade IV or V) is a risk factor for renal scarring, impaired renal function and arterial hypertension. Voiding cystourethrography (VCUG) is the gold standard for detecting the severity of VUR. High grade VUR is present in the minority of children with urinary tract infection (UTI), thus exposing the majority to invasive diagnostics which have no surgical consequence. We therefore aimed at establishing a non-invasive test to identify children with high grade VUR.
May 2012 Consensus in Pediatrics Moscow High grade VUR (grade IV or V) Case-control study to establish specific urinary proteome pattern by CE-MS 18 cases with primary VUR grade IV or V 19 controls without VUR after UTI. VURControls CE Time (min) Mass (Da) Drube et al. (2012) Pediatrics 129(2):e
May 2012 Consensus in Pediatrics Moscow VUR: Blinded Clinical Validation The tests accuracy was independent of age, gender and grade of VUR in the contra-lateral kidney. The odds ratio of suffering from VUR grade IV or V when tested positive was 28 (95%CI: 4.5 to 176) Sensitivity 1-Specificity Sensitivity 88% (15/17) Specificity 79% (15/19) AUC=80%, P<
May 2012 Consensus in Pediatrics Moscow VUR: Sequencing of Markers Peptide ID Mass (Da) CE-Time (min) VURUTISequenceProtein DRGEpGPpGPA Collagen alpha-1(I) ( ) DGGGSPKGDVDP Sodium/potassium- transporting ATPase, subunit gamma (7-18) DFDDFNLED CD99 antigen-like protein 2 (26-34) KGEAGLpGApGSPG Q Collagen alpha-1(XIX) ( ) NDGAKGDAGApGAp GSQGApG Collagen alpha-1(I) ( ) In VUR decreased / increased urinary excretion
May 2012 Consensus in Pediatrics Moscow VUR: Pathophysiology of Markers CD99 is strongly expressed in normal urinary mucosa, which is known to show histological changes in animal models for VUR Collagen alpha-1 (I) fragments suggest alterations of extra cellular matrix turnover Na/K-transporting ATPase plays a key role in the active transportation of Na + and K + across basolateral membranes of nephron epithelial cells Biomarkers might indicate early onset of reflux nephropathy
May 2012 Consensus in Pediatrics Moscow Proteomics in Pediatrics: Conclusion Proteomic patterns tolerate instability and inconsistency of individual biomarkers CE-MS platform allows biomarker discovery and validation as a pattern in large patient cohorts Proteomic biomarkers generate novel hypothesis for potential pathological processes during disease development CE-MS enables transfer of urinary proteome analysis from bench side to bed side
May 2012 Consensus in Pediatrics Moscow 1 Coronary Artery Disease 2 Diabetic Nephropathy 3 Bladder cancer 4 Prostate cancer Furthermore: Proteomics in adults
May 2012 Consensus in Pediatrics Moscow CE-time Mass CAD negativeCAD positive Δ CAD Score Irbesartan 300 mg/day Placebo P= Delles et al. (2010) J. Hypertension 28: Furthermore: Treatment Monitoring Long-term treatment effects. Patients with type 2 diabetes irbesartan 300 mg once daily (n=11) or placebo (n=11) At baseline and after 2 years of Decrease in CADScore toward healthier
May 2012 Consensus in Pediatrics Moscow The Physician Gerrit Dou Leiden ( ) Acknowledgment Mosaiques: Harald Mischak Mohammed Dakna Igor Golovko Annika Dorban Justyna Siwy Jochen Metzger Petra Zürbig Partners: Markus J. Kemper (Hamburg, Germany) Thomas Neuhaus (Zurich, Switzerland) Ralf Lichtinghagen (Hannover, Germany) Esther Lau (Hannover, Germany) Benno Ure (Hannover, Germany) Sylvia Glüer (Hannover, Germany) Martin Kirschstein (Celle, Hannover) Stéphane Decramer (Toulouse, France) Jean-Loup Bascands (Toulouse, France) Joost P. Schanstra (Toulouse, France) Claus Petersen (Hannover, Germany)
May 2012 Consensus in Pediatrics Moscow Implementation of Proteomics The number of publications on proteomics has increased tremendously, however, the implementation of urinary proteomics into routine nephrology diagnostics is awaiting further progress. Combined efforts should focus on implementing clinical proteomics after identification of disease specific proteome patterns by providing guidance for further analysis of samples from biobanks, and providing guidance for clinical study design concerning intervention studies. Feedback mechanisms to evaluating cost- effectiveness and clinical adoption are urgently needed to allow a timely introduction of proteomics into every day clinical care.