Slide 1: NETs: Casting a new light on sepsis management
NETs: Casting a new light on sepsis management
Slide 2: Bienvenue!
Bienvenue! Professor Djillali Annane
Slide 3: Neutrophil Extracellular Traps (NETs)
Neutrophil Extracellular Traps (NETs) NETs: • are produced by ejecting chromosomal material out of the cell • catch and kill bacteria and viruses • can sterilize blood in minutes • first reported in 20041 • now the subject of > 5000 publications 1. Brinkmann V, Reichard U, Goosmann C, Fauler B, Uhle
Slide 4: 1 in 5 deaths worldwide are associated with sepsis
1 in 5 deaths worldwide are associated with sepsis It’s the number 1... Almost 50 million cases resulting in Cause of death in Over 40% of 11 million deaths hospitals survivors suffer from long-term physical Over 40% of cases Cause for hospital or psychological are children under readmissions effec
Slide 5: Volition's Mission
Volition’s Mission Develop a low-cost, easy-to-use, rapid diagnostic test to save lives and improve outcomes for patients worldwide. We are here to present Nu.Q® NETs H3.1 assay, a novel, clinically relevant biomarker which has the potential to change the management of patients with sepsis. Property
Slide 6: Executive Summary: consolidated conclusions
Executive Summary: consolidated conclusions Results from three independent studies totalling over 3,000 patients These findings are consistent across cohorts1-3 An elevated H3.1 level reflects a dysregulated immune response and is associated with: • a risk of increased mortality • an increased risk
Slide 7: Speakers
Speakers Dr. Andrew Retter Terry Kelly, PhD Dr. Caroline Neumann Clinical Lead in Critical Chief Innovation Officer, Senior Consultant in Intensive Care Medicine, ECMO and VolitionRx, Care Medicine, Thrombosis USA Jena University Hospital, Germany Chief Medical Officer at VolitionRx, UK Property of
Slide 8: Why is H3.1 key: the biology & scientific rationale
Why is H3.1 key: the biology & scientific rationale Dr. Andrew Retter, MBBS, MRCP, FRCPath (Haem), DICM, FFICM Clinical Lead in Intensive Care, ECMO and Thrombosis Confidential - Property of Volition© 2024 Slide 8
Slide 9: Conflicts of interest to declare
Conflicts of interest to declare • Employee and shareholder of VolitionRX Limited Property of Volition© 2024 Slide 9
Slide 10: Nucleosomes and histones:
Nucleosomes and histones: Property of Volition© 2024 Slide 10
Slide 11: Nucleosomes and histones:
Nucleosomes and histones: Key message: The H3.1 assay can detect nucleosomes using chemiluminescence technology and provide a result within 15 minutes The lower limit of quantification is 20ng/ml The upper limit of quantification is 20,000ng/ml Property of Volition© 2024 Slide 11
Slide 12: H3.1 as a Damage-Associated Molecular Protein
H3.1 as a Damage-Associated Molecular Protein Silk et al, Cell Death & Disease, 2017 http://dx.doi.org/10.1038/cddis.2017.52 Property of Volition© 2024 Slide 12
Slide 13: H3.1 sits as a triumvirate of innate immunity, inflammation and coagulation
H3.1 sits as a triumvirate of innate immunity, inflammation and coagulation. The majority of extracellular pathology is due to the indiscriminate binding of anionic components of the circulation and vasculature. Property of Volition© 2024 Slide 13
Slide 14: Extracellular Histones and Organ Injury
Extracellular Histones and Organ Injury Silk et al, Cell Death & Disease, 2017 http://dx.doi.org/10.1038/cddis.2017.52 Property of Volition© 2024 Slide 14
Slide 15: Bacteria Alveolus Platelet plug
Bacteria Alveolus Active TF TF rich microparticles Monocyte H3.1 Pathogen PRR Elastase, Cathepsin G, Coagulation PSGL1 Myeloperoxidase, complement are enhanced Monocyte Reactive oxygen species P-selectin Platelet plug Property of Volition© 2024 Slide 15
Slide 16: The role of H3.1 in NETosis
The role of H3.1 in NETosis: …proving we are measuring what we say we are measuring! Terry Kelly, PhD., Chief Innovation Officer, Volition Confidential - Property of Volition© 2024 Slide 16
Slide 17: NETs Contain Proteins and Trap Pathogens
NETs Contain Proteins and Trap Pathogens A neutrophil granulocyte (yellow) has ejected a NET (green) to capture bacteria (purple). A red blood cell (orange) is also trapped in the NET. Stained scanning electron microscope image by Volker Brinkmann. © Volker Brinkmann / Max Planck Institute for Infec
Slide 18: Nu.Q Antibodies Label NETs in Isolated Neutrophils
Nu.Q® Antibodies Label NETs in Isolated Neutrophils H3.1 DNA Membranes Merge Nucleosomes DNA Membranes Merge Property of Volition© 2024 Slide 18
Slide 19: H3.1 Nucleosome Levels Increase With NETosis
H3.1 Nucleosome Levels Increase With NETosis Zukas et al, Journal of Thrombosis & Hematology, 2024 https://doi.org/10.1016/j.jtha.2024.05.028 JTH Commentary, Sept 2024 https://doi.org/10.1016/j.jtha.2024.06.016 Property of Volition© 2024 Slide 19
Slide 20: Kinetic Information
Kinetic Information H3.1 is not impacted by height, weight, age, sex1 H3.1 is not impacted by the circadian rhythm2 1. German Data Set, data on file; 2. RHU Records Data Set, data on file Property of Volition© 2024 Slide 20
Slide 21: cfDNA Profiles Vary Across Disease and Cell Death Mechanisms
cfDNA Profiles Vary Across Disease and Cell Death Mechanisms Apoptosis Length of Plasma Derived DNA Cancer Sepsis NETs Circulating nucleosome levels increase as they are released faster than they can be removed Data on file Property of Volition© 2024 Slide 21
Slide 22: Size Distribution Distinguishes Sepsis from Cancer
Size Distribution Distinguishes Sepsis from Cancer & DNA Methylation Patterns Identify Cell of Origin cfDNA from NHL sample primarily Length of Plasma Derived DNA derived from B-cell (B-Cell Lymphoma) Cancer INV001 – Prostate cancer INV002 – Sepsis INV006- NHL Sepsis INV007 – NHL cfDNA from sepsis s
Slide 23: cfDNA Derived From Neutrophils is Enriched in Sepsis Patients
cfDNA Derived From Neutrophils is Enriched in Sepsis Patients % neutrophils Healthy Individuals ICU Non-sepsis Sepsis 0% 32% 75% Fragment length (log scale) Data on file Property of Volition© 2024 Slide 23
Slide 24: Clinical Data
Clinical Data Dr. Andrew Retter Confidential - Property of Volition© 2024 Slide 24
Slide 25: KOL meeting: Sept 2024
KOL meeting: Sept 2024 Chaired by Professor Djillali Annane, Professor of Medicine at University Paris Saclay- UVSQ. • Professor Derek Angus, Professor and Chair of the Critical Care Medicine Department at the University of Pittsburgh. (Partial attendance; virtual) • Professor Michael Bauer, Profess
Slide 26: Studies at Centers of Excellence: >3000 patients
Studies at Centers of Excellence: >3000 patients Study Country Description Cohort Size SISPCT Germany Retrospective analysis of 971 intensive care patients prospectively collected cohort Multiple timepoints Amsterdam Netherlands Retrospective analysis of 1,713 intensive care patients UMC prospective
Slide 27: H3.1 only correlated weakly with the neutrophil count
H3.1 only correlated weakly with the neutrophil count 1. German Data Set, data on file; 2. RHU Records Data Set, data on file Property of Volition© 2024 Slide 27
Slide 28: H3.1 only weakly correlated with the neutrophil count
H3.1 only weakly correlated with the neutrophil count This is critical as H3.1 is giving us NEW information! You don’t just have to measure an FBC. 1. Amsterdam UMC Data Set, data on file Property of Volition© 2024 Slide 28
Slide 29: H3.1 predicts mortality
H3.1 predicts mortality Property of Volition© 2024 Confidential Slide 29
Slide 30: Initial H3.1 readings above 10,000 predicted mortality within 14 days
Initial H3.1 readings above 10,000 predicted mortality within 14 days ag os s g se t c shoc f st se e e se s s Overall survivors not plotted a s o eath a s German Data Set, data on file All this work is based on admission H3.1 Property of Volition© 2024 Slide 30
Slide 31: Mortality risk for different levels of H3.1 on admission
Mortality risk for different levels of H3.1 on admission German Data Set, data on file All this work is based on admission H3.1 Property of Volition© 2024 Slide 31
Slide 32: Kaplan-Meier plot of survival based on initial H3.1
Kaplan-Meier plot of survival based on initial H3.1 – high > 1,143.3 ng/mL Cox H3.1_first 7.080e-05 1.000e+00 1.195e-05 5.923 3.16e-09 *** --- German Data Set, data on file Property of Volition© 2024 Slide 32
Slide 33: Clinical utility of Nu.Q in septic Acute Kidney Injury (AKI)
Clinical utility of Nu.Q® in septic Acute Kidney Injury (AKI): Data from SISPCT Dr. Caroline Neumann, DESA, EDIC, Infectious Diseases Specialist (DGI) Senior Intensive Care Consultant Confidential - Property of Volition© 2024 Slide 33
Slide 34: A Unified Theory of Sepsis-Induced AKI
A Unified Theory of Sepsis-Induced AKI Adapted from Gomez H, Ince C, De Backer D, Pickkers P, Payen D, Hotchkiss J, Kellum JA. A unified theory of sepsis-induced acute kidney injury: inflammation, microcirculatory dysfunction, bioenergetics, and the tubular cell adaptation to injury. Shock. 2014 Jan
Slide 35: A Unified Theory of Sepsis-Induced AKI
A Unified Theory of Sepsis-Induced AKI Gomez H, Ince C, De Backer D, Pickkers P, Payen D, Hotchkiss J, Kellum JA. A unified theory of sepsis-induced acute kidney injury: inflammation, microcirculatory dysfunction, bioenergetics, and the tubular cell adaptation to injury. Shock. 2014 Jan;41(1):3-11.
Slide 36: The role of immune cells in early injury and repair from AKI
The role of immune cells in early injury and repair from AKI Volition Veterinary Diagnostics© 2019 Property of Volition© 2023 Confidential 2024 Slide 36
Slide 37: Role of Immune Cells in AKI and Repair
Role of Immune Cells in AKI and Repair Lee SA, Noel S, Sadasivam M, Hamad ARA, Rabb H. Role of Immune Cells in Acute Kidney Injury and Repair. Nephron. 2017;137(4):282-286. doi: 10.1159/000477181 Property of Volition© 2024 Slide 37
Slide 38: Neutrophil Extracellular Traps in the Establishment and Progression of Renal Disease
Neutrophil Extracellular Traps in the Establishment and Progression of Renal Disease Salazar-Gonzalez H, Zepeda-Hernandez A, Melo Z, Saavedra-Mayorga DE, Echavarria R. Neutrophil Extracellular Traps in the Establishment and Progression of Renal Diseases. Medicina (Kaunas). 2019 Aug 2;55(8):431. doi:
Slide 39: How to assess damage and failing repair?
How to assess damage and failing repair? Volition Veterinary Diagnostics© 2019 Property of Volition© 2023 Confidential 2024 Slide 39
Slide 40: Observational study of nucleosome levels in septic acute kidney injury
Observational study of the clinical utility of the nucleosome levels in septic acute kidney injury: improved classification and targeted intervention in AKI? Objectives and methods: • secondary analysis of patients recruited for the SISPCT trial [1] • evaluation of levels of circulating H3.1 nucleos
Slide 41: Study Design
n=1089 Study Design patients included in SISPCT n=208 *G : GFR ≥ 9 l *G2: GFR 60-89 ml/min exclusion due to n=881 pre-existing RRT *G3a: GFR 45-59 ml/min *G3b: GFR 30-44 ml/min eligible for analysis *G4: GFR 15-29 ml/min *G5: GFR<15 ml/min, but without RRT n=567 n=67 (7.6%) n=28 (3.2%) n=219 (24.8%)
Slide 42: Measurement of H3.1 using Nu.Q H3.1 Assay
Measurement of H3.1 using Nu.Q® H3.1 Assay H3.1 nucleosome levels were analyzed at admission and serially in frozen citrate plasma samples D0 D2 D7 D-1 randomization D28 and H3.1 screening D90 H3.1 H3.1 Mortality, LOS ICU AKI/no AKI and hospital, RRT dependence... German Data Set, data on file Prope
Slide 43: SISPCT sub-cohort baseline characteristics
SISPCT sub-cohort baseline characteristics Variable AKI 0 (n=567) AKI 1 (n=67) AKI 2 (n=28) AKI 3 (n=219) Male sex, No. (%) 365 (64%) 46 (69%) 18 (64%) 143 (65%) Age, med. (SD), y 67 (56, 75) 71 (60, 77) 66 (56, 74) 70 (60, 76) SOFA 9 (7, 11) 9 (8, 11) 8 (7, 10) 11 (9, 15) Scr 24 h before study 97 (
Slide 44: H3.1 levels in patients with preexisting impaired kidney function without evolving AKI
H3.1 levels in patients with preexisting impaired kidney function without evolving AKI (p<0.001) G : GFR≥ 9 l G2: GFR 60-89 ml/min G3a: GFR 45-59ml/min G3b: GFR 30-44ml/min G4: GFR 15-29ml/min G5: GFR <15ml/min H3.1 levels in patients with preexisting impaired kidney function without evolving AKI (p
Slide 45: H3.1 Nucleosome levels over time
H3.1 Nucleosome levels over time AKI stage D0 (Nu.Q® Level) D2 (Nu.Q® Level) D7 (Nu.Q® Level) p-value 0 484 (216-1127) 518 (249-974) 492 (229-969) 0.785 1 577 (266-1881) 790 (393-1319) 608 (234-820) 0.084 2 518 (319-1917) 809 (477-1477) 658 (356-2864) 0.574 3 1151 (509-3797) 1169 (611-2881) 885 (438
Slide 46: Kaplan-Meier: Quintiles of Creatinine first for AKI Stage
Kaplan-Meier: Quintiles of Creatinine first for AKI Stage 4 at 28 days G ou u t le Ra ge: to c ol o o t o of coho t u t le Ra ge: to 9 c ol u t le Ra ge: 9 to c ol u t le Ra ge: to c ol u t le Ra ge: to 9 c ol a s German Data Set, data on file Property of Volition© 2024 Slide 46
Slide 47: Kaplan-Meier: Quintiles of H3.1 First for AKI Stage
Kaplan-Meier: Quintiles of H3.1 First for AKI Stage 4 at 28 days G ou u t le Ra ge: to 9 h o o t o of coho t u t le Ra ge: to h u t le Ra ge: to h u t le H3.1 Ra ge: to h First u t le Ra ge: to h a s German Data Set, data on file Property of Volition© 2024 Slide 47
Slide 48: Clinical Model of H3.1 Dose + Platelets + Urine24hr for AnyRRT28
Clinical Model of H3.1 Dose + Platelets + Urine24hr for AnyRRT28 G ou co e o o t o of coho t 9 at s co e at s co e at s co e at s a s German Data Set, data on file Property of Volition© 2024 Nucleosomics pipeline version 4.0.6 Slide 48
Slide 49: Kidney Failure: summary
Kidney Failure: summary • Sepsis-induced AKI involves complex pathophysiology, with NETs playing a crucial role. • Nu.Q® H3.1, as a marker of NETosis, shows promise as a biomarker in this context. • H3.1 levels correlate with AKI severity and show distinct temporal patterns, particularly in severe A
Slide 50: Respiratory Failure
Respiratory Failure Dr. Andrew Retter Confidential - Property of Volition© 2024 Slide 50
Slide 51: Restoring discarded porcine lungs by ex vivo removal of NETs
Restoring discarded porcine lungs by ex vivo removal of neutrophil extracellular traps. Mittendorfer, Margareta et al. The Journal of Heart and Lung Transplantation, Volume 0, Issue 0. https://doi.org/10.1016/j.healun.2024.07.007 Property of Volition© 2024 Slide 51
Slide 52: Restoring discarded porcine lungs by ex vivo removal of NETs
Restoring discarded porcine lungs by ex vivo removal of neutrophil extracellular traps. Mittendorfer, Margareta et al. The Journal of Heart and Lung Transplantation, Volume 0, Issue 0. https://doi.org/10.1016/j.healun.2024.07.007 Property of Volition© 2024 Slide 52
Slide 53: Restoring discarded porcine lungs by ex vivo removal of NETs
Restoring discarded porcine lungs by ex vivo removal of neutrophil extracellular traps. Mittendorfer, Margareta et al. The Journal of Heart and Lung Transplantation, Volume 0, Issue 0. https://doi.org/10.1016/j.healun.2024.07.007 Property of Volition© 2024 Slide 53
Slide 54: Restoring discarded porcine lungs by ex vivo removal of NETs
Restoring discarded porcine lungs by ex vivo removal of neutrophil extracellular traps. Mittendorfer, Margareta et al. The Journal of Heart and Lung Transplantation, Volume 0, Issue 0. https://doi.org/10.1016/j.healun.2024.07.007 Property of Volition© 2024 Slide 54
Slide 55: Restoring discarded porcine lungs by ex vivo removal of NETs
Restoring discarded porcine lungs by ex vivo removal of neutrophil extracellular traps. Mittendorfer, Margareta et al. The Journal of Heart and Lung Transplantation, Volume 0, Issue 0. https://doi.org/10.1016/j.healun.2024.07.007 Property of Volition© 2024 Slide 55
Slide 56: Our human data is consistent with this animal model of ARDS
Our human data is consistent with this animal model of ARDS Respiratory parameters at time of admission and ARDs Nu.Q® H3.1 Nu.Q® H3.1 Statistical levels levels Sepsis Septic Shock No ARDs 138 70 63 285.7 0.0017 647.8 (14.2%) (15.8%) (12.1%) Mild ARDs 201 102 96 0.0044 396.3 646.7 (20.7%) (23%) (18.
Slide 57: Results from other Clinical Studies
Results from other Clinical Studies Dr. Andrew Retter Property of Volition© 2024 Slide 57
Slide 58: Studies at Centers of Excellence: >3000 patients
Studies at Centers of Excellence: >3000 patients Study Country Description Cohort Size SISPCT Germany Retrospective analysis of 971 intensive care patients prospectively collected cohort Multiple timepoints Amsterdam Netherlands Retrospective analysis of 1,713 intensive care patients UMC prospective
Slide 59: Relationship with mortality
Relationship with mortality Amsterdam UMC Data Set, data on file Confidential - Property of Volition© 2024 Slide 59
Slide 60: Relationship with mortality
Relationship with mortality Amsterdam UMC Data Set, data on file Confidential - Property of Volition© 2024 Slide 60
Slide 61: Relationship with AKI
Relationship with AKI Amsterdam UMC Data Set, data on file Confidential - Property of Volition© 2024 Slide 61
Slide 62: Total number of organ failures
Total number of organ failures Amsterdam UMC Data Set, data on file Confidential - Property of Volition© 2024 Slide 62
Slide 63: Total number of organ failures
Total number of organ failures Amsterdam UMC Data Set, data on file Confidential - Property of Volition© 2024 Slide 63
Slide 64: H3.1 levels and outcome: higher initial level in patients who will die
H3.1 levels and outcome: Higher initial level of H3.1- nuclesomes in patient who will die (7 days follow-up) p=0.007 Alive Dead n= 374 34 Median 846 2106,8 (ng/ml) Records Data Set, data on file Confidential - Property of Volition© 2024 Slide 64
Slide 65: Higher level of H3.1 in septic shock group
Higher level of H3.1 in septic shock group p=0.001 RECORDS Septic Sepsis Shock n= 183 233 Median 727 1029 (ng/ml) Sepsis Septic Shock Records Data Set, data on file Confidential - Property of Volition© 2024 Slide 65
Slide 66: H3.1 levels at admission higher in patients who will need RRT
H3.1 levels at admission is higher in patient who will need RRT at Day 1 and/or Day 2 No Continu Disconti DAY 1 dialysis ous nuous n= 375 18 11 Median (ng/ml) 847,1 3460 2424,4 Records Data Set, data on file Confidential - Property of Volition© 2024 Slide 66
Slide 67: The risk to need any RRT increases with the level of nucleosomes at Day 0
The risk to need any RRT increases with the level of nucleosomes at Day 0 56 patients had at least one (any) RRT within 7 Days ➔ The distributions are significantly ≠ between categories (p = 0,002) Records Data Set, data on file Confidential - Property of Volition© 2024 Slide 67
Slide 68: Higher H3.1-nucleosomes in patients who required respiratory support
Higher level of H3.1-nucleosomes in patient who required respiratory support (at admission) p<0.001 p=0.009 No assisted Assisted No invasive Invasive ventilation ventilation ventilation ventilation n= 181 235 n= 29 206 Median Median 673,4 1132,4 528,6 1261,9 (ng/ml) (ng/ml) Records Data Set, data on
Slide 69: Concluding Remarks
Concluding Remarks Professor Djillali Annane Confidential - Property of Volition© 2024 Slide 69
Slide 70: Studies at Centers of Excellence: >3000 patients
Studies at Centers of Excellence: >3000 patients Study Country Description Cohort Size SISPCT Germany Retrospective analysis of 971 intensive care patients prospectively collected cohort Multiple timepoints Amsterdam Netherlands Retrospective analysis of 1,713 intensive care patients UMC prospective
Slide 71: Executive Summary: consolidated conclusions
Executive Summary: consolidated conclusions Results from three independent studies totalling over 3,000 patients These findings are consistent across cohorts1-3 An elevated H3.1 level reflects a dysregulated immune response and is associated with: • a risk of increased mortality • an increased risk
Slide 72: Question & Answer Session
Question & Answer Session Confidential - Property of Volition© 2024 Slide 72