Clinical validation of the novel fluorescent glomerular filtration rate tracer agent relmapirazin (MB-102)

Abstract

The fluorescent compound relmapirazin has been rationally designed for use in point-of-care measurement of glomerular filtration rate (GFR), with attributes including negligible protein binding, negligible metabolites in vivo, negligible tubular secretion, and excellent chemical and photo stability. Twenty-four nonclinical assays were performed in accordance with FDA requirements yielding negligible toxicology concerns. Here, a clinical study was performed to validate relmapirazin as a GFR tracer in patients by comparison to iohexol. This was evaluated in 120 adults at three clinical sites with eGFR values ranging from normal to Stage 4 chronic kidney disease. Relmapirazin and iohexol were administered intravenously in consecutive boluses to each subject and serial blood samples obtained over the subsequent 12 hours. Plasma concentrations were measured and the corresponding plasma GFR for each agent was determined using a standard two-compartment pharmacokinetic assessment. Urine from each subject was collected for the entire 12-hour study period to measure the amount of administered dose appearing in the urine. A near perfect linear regression correlation was observed between the GFRs measured by these two tracers (r2=0.99). Bland-Altman analysis confirmed agreement between these two measures of GFR (limits of agreement -7.0 to +5.6 mL/min; mean of -0.7 mL/min). The GFR determined by relmapirazin was independent of GFR stratification by chronic kidney disease stage, and importantly by race. The percent of the administered relmapirazin dose recovered in the urine was greater than or equal to that of iohexol with no reported severe adverse events. Thus, relmapirazin may be used as a GFR tracer agent in humans. The fluorescent compound relmapirazin has been rationally designed for use in point-of-care measurement of glomerular filtration rate (GFR), with attributes including negligible protein binding, negligible metabolites in vivo, negligible tubular secretion, and excellent chemical and photo stability. Twenty-four nonclinical assays were performed in accordance with FDA requirements yielding negligible toxicology concerns. Here, a clinical study was performed to validate relmapirazin as a GFR tracer in patients by comparison to iohexol. This was evaluated in 120 adults at three clinical sites with eGFR values ranging from normal to Stage 4 chronic kidney disease. Relmapirazin and iohexol were administered intravenously in consecutive boluses to each subject and serial blood samples obtained over the subsequent 12 hours. Plasma concentrations were measured and the corresponding plasma GFR for each agent was determined using a standard two-compartment pharmacokinetic assessment. Urine from each subject was collected for the entire 12-hour study period to measure the amount of administered dose appearing in the urine. A near perfect linear regression correlation was observed between the GFRs measured by these two tracers (r2=0.99). Bland-Altman analysis confirmed agreement between these two measures of GFR (limits of agreement -7.0 to +5.6 mL/min; mean of -0.7 mL/min). The GFR determined by relmapirazin was independent of GFR stratification by chronic kidney disease stage, and importantly by race. The percent of the administered relmapirazin dose recovered in the urine was greater than or equal to that of iohexol with no reported severe adverse events. Thus, relmapirazin may be used as a GFR tracer agent in humans. Lay SummaryCurrent estimation of kidney function, specifically glomerular filtration rate (GFR), in the clinical setting most commonly employs a single time point measurement of endogenous serum creatinine with empirical equations resulting in an estimated GFR, which is known to often be inaccurate for an individual subject. We performed a clinical study comparing GFR determined from the acknowledged standard of iohexol plasma clearance to GFR determined using a novel fluorescent compound relmapirazin (MB-102), which was designed specifically to be a marker of glomerular filtration. A near perfect correlation was observed between the plasma GFR of these 2 agents across a wide range of individual GFR. Thus, the novel fluorescent agent relmapirazin is a GFR tracer agent in humans. Current estimation of kidney function, specifically glomerular filtration rate (GFR), in the clinical setting most commonly employs a single time point measurement of endogenous serum creatinine with empirical equations resulting in an estimated GFR, which is known to often be inaccurate for an individual subject. We performed a clinical study comparing GFR determined from the acknowledged standard of iohexol plasma clearance to GFR determined using a novel fluorescent compound relmapirazin (MB-102), which was designed specifically to be a marker of glomerular filtration. A near perfect correlation was observed between the plasma GFR of these 2 agents across a wide range of individual GFR. Thus, the novel fluorescent agent relmapirazin is a GFR tracer agent in humans. The current clinical standard determination of glomerular filtration rate (GFR), which employs empirical equations that estimate GFR (eGFR), is based on a single time point serum creatinine (SCr) value. It is well-known that a change in SCr concentration lags a kidney insult or injury by 24 to 48 hours, approximately one-half of kidney function may be lost before the SCr concentration reaches an abnormal level, and the SCr concentration is affected by attributes other than kidney function (such as muscle mass and diet).1Endre Z.H. Pickering J.W. Walker R.J. Clearance and beyond: the complementary roles of GFR measurement and injury biomarkers in acute kidney injury (AKI).Am J Physiol Renal Physiol. 2011; 301: F697-F707Crossref PubMed Scopus (125) Google Scholar, 2Speeckaert M.M. Seegmiller J. Glorieux G. et al.Measured glomerular filtration rate: the query for a workable golden standard technique.J Pers Med. 2021; 11: 949Crossref PubMed Scopus (16) Google Scholar, 3Zitta S. Schrabmair W. Reibnegger G. et al.Glomerular filtration rate (GFR) determination via individual kinetics of the inulin-like polyfructosan sinistrin versus creatinine-based population-derived regression formulae.BMC Nephrol. 2013; 14: 159Crossref PubMed Google Scholar Furthermore, the eGFR equations are ensemble averages and hence may be unpredictive for a specific individual.4Shafi T. Zhu X. Lirette S.T. et al.Quantifying individual-level inaccuracy in glomerular filtration rate estimation: a cross-sectional study.Ann Intern Med. 2022; 175: 1073-1082Crossref PubMed Scopus (36) Google Scholar Fluorescent detection of an exogenous GFR tracer agent at the point of care would be a direct measurement of GFR and has been pursued by several groups.5Chinen L.K. Galen K.P. Kuan K.T. et al.Fluorescence-enhanced europium-diethylenetriaminepentaacetic (DTPA)-monoamide complexes for the assessment of renal function.J Med Chem. 2008; 51: 957-962Crossref PubMed Scopus (34) Google Scholar, 6Schock-Kusch D. Sadick M. Henninger N. et al.Transcutaneous measurement of glomerular filtration rate using FITC-sinistrin in rats.Nephrol Dial Transplant. 2009; 24: 2997-3001Crossref PubMed Scopus (111) Google Scholar, 7Yu W. Sandoval R.M. Molitoris B.A. Rapid determination of renal filtration function using an optical ratiometric imaging approach.Am J Physiol Renal Physiol. 2007; 292: F1873-F1880Crossref PubMed Scopus (90) Google Scholar, 8Rizk D.V. Meier D. Sandoval R.M. et al.A novel method for rapid bedside measurement of GFR.J Am Soc Nephrol. 2018; 29: 1609-1613Crossref PubMed Scopus (47) Google Scholar, 9Huang J. Weinfurter S. Daniele C. et al.Zwitterionic near infrared fluorescent agents for noninvasive real-time transcutaneous assessment of kidney function.Chem Sci. 2017; 8: 2652-2660Crossref PubMed Scopus (25) Google Scholar However, to date, no agents nor measurement procedures have been approved by a regulatory authority. The aim of this study was to evaluate the use of the fluorescent compound relmapirazin as a new GFR tracer agent in humans by comparison to the often used and widely accepted GFR tracer agent iohexol as the reference standard. Relmapirazin (International Nonproprietary Names [INN] designation), also known as MB-102, is a novel fluorescent tracer agent engineered for real-time point-of-care GFR determination. The chemical name is 3,6-diamino-2,5-bis{N-[(1R)-1-carboxy-2-hydroxyethyl]carbamoyl}pyrazine. The chemical structure is shown in Supplementary Figure S1. Characteristics and properties of this molecule needed for a GFR tracer agent (such as hydrophilicity, negligible protein binding, negligible metabolism, negligible photobleaching, negligible tubular secretion, excellent solubility, and excellent stability), as well as its photophysical properties, have been previously published.10Rajagopalan R. Neumann W.L. Poreddy A.R. et al.Hydrophilic pyrazine dyes as exogenous fluorescent tracer agents for real-time point-of-care measurement of glomerular filtration rate.J Med Chem. 2011; 54: 5048-5058Crossref PubMed Scopus (45) Google Scholar, 11Shieh J.-J. Riley I.R. Rogers T.E. et al.Characterization of MB-102, a new fluorescent tracer agent for point-of-care renal function monitoring.J Pharm Sci. 2020; 109: 1191-1198Abstract Full Text Full Text PDF PubMed Scopus (14) Google Scholar, 12Shieh J.-J. Riley I.R. Rogers T.E. Dorshow R.B. Novel chiral high-performance liquid chromatographic (HPLC) determination of MB-102, a new fluorescent tracer agent enabling the measurement of point-of-care glomerular filtration rate.Anal Lett. 2023; 56: 781-788Crossref Scopus (0) Google Scholar Relmapirazin has been evaluated in 24 formal toxicology, safety, and pharmacology in vitro and in vivo nonclinical studies necessary for advancement to human clinical studies.13Bugaj J.E. Dorshow R.B. Pre-clinical toxicity evaluation of MB-102, a novel fluorescent tracer agent for real-time measurement of glomerular filtration rate.Regul Toxicol Pharmacol. 2015; 72: 26-38Crossref PubMed Google Scholar, 14Dorshow R.B. Bugaj J.E. Next tier in vitro and in vivo nonclinical studies further elucidating the safety and toxicity profile of MB-102, a novel fluorescent tracer agent for measurement of glomerular filtration rate.Regul Toxicol Pharmacol. 2019; 107104417Crossref PubMed Scopus (14) Google Scholar, 15Bugaj J.E. Dorshow R.B. Absence of developmental or reproductive toxicity in rats for MB-102, a fluorescent tracer agent for point-of-care measurement of glomerular filtration rate.Regul Toxicol Pharmacol. 2022; 131105158Crossref PubMed Scopus (3) Google Scholar, 16Bugaj J.E. Dorshow R.B. Evaluation of developmental and reproductive toxicity in rabbits for MB-102, a fluorescent tracer agent designed for real-time measurement of glomerular filtration rate.Int J Toxicol. 2022; 41: 380-388Crossref PubMed Scopus (1) Google Scholar No toxicological effects nor safety concerns were reported. The drug product (drug dosing solution) is a formulation of the drug substance in phosphate-buffered saline. No other excipients are present. No change in photophysical properties nor chemical properties are observed on formulation with respect to the drug substance. pH = 7.3 and hence is intravenously administered at physiological conditions. The comparator and reference GFR tracer agent for the clinical study, chosen with agreement by the U.S. Food and Drug Administration, is the X-ray contrast agent iohexol. Iohexol is a U.S. Food and Drug Administration–approved iodinated contrast media used to enhance X-ray or computerized tomography imaging tests. Iohexol administered intravenously in small quantities has a history of off-label use as a GFR tracer agent.17Delanaye P. Ebert N. Melsom T. et al.Iohexol plasma clearance for measuring glomerular filtration rate in clinical practice and research: a review. Part 1: how to measure glomerular filtration rate with iohexol.Clin Kidney J. 2016; 9: 682-689Crossref PubMed Scopus (170) Google Scholar,18Delanaye P. Melsom T. Ebert N. et al.Why to measure glomerular filtration rate with iohexol? Part 2: how to measure glomerular filtration rate with iohexol.Clin Kidney J. 2016; 9: 700-704Crossref PubMed Scopus (105) Google Scholar The specific brand and concentration used in this clinical study was Omnipaque 300 (manufactured by GE Healthcare). Iohexol was provided as a sterile, pyrogen-free, colorless to pale-yellow solution. The iohexol concentration used as the reference standard in this trial was 647 mg/ml. In formulation, each milliliter of iohexol solution contains 1.21 mg of tromethamine and 0.1 mg edetate calcium disodium. This was a pilot phase II clinical study authorized under an investigation device exemption due to the regulatory pathway of the eventual combination product. It was a prospective, open study conducted at 3 clinical sites: the Washington University clinical research unit, the Saint Louis University clinical research unit, and the Orlando Clinical Research Center. The study followed International Council for Harmonization (ICH) guidelines for Good Clinical Practice (ICH R2, 2016), the Code of Federal Regulations Title 21 parts 812, 11, 50, 54, and 56 and International Organization for Standardization 14155: 2011(E) Clinical Investigation of Medical Devices for Human Subjects–Good Clinical Practice, as well as the ethical principles defined by the World Medical Association Declaration of Helsinki, and the requirements of national device and data protection laws and other regulatory to in the study was obtained from each the study was This was a study of relmapirazin for kidney The study in was a cross-sectional analysis of with iohexol as the this was a pilot study, no formal of was However, the were to have a of that the GFR range from normal to 4 chronic kidney disease and for one-half of the to have to and one-half with to of were in this A of with normal kidney function 2 of 120 to of with 3 and 4 of to to have of with to and with to were to the Kidney was used in et Kidney A new to estimate glomerular filtration Intern Med. 2009; Intern Med. PubMed Scopus Google Scholar greater than or equal to normal or and kidney function, and use of were or to history of or to use of drug two of the dosing history of or 3 or or or the U.S. Food and Drug the range to to of and adults than of were from the A of the and is provided on the clinical under provided at the to and and was to be of the dosing safety and adverse were 3 of to the study for safety The 2 agents were administered to each subject Relmapirazin was administered by over followed by a over this iohexol was administered to each subject by over followed by a over The subject dose of relmapirazin was of of iohexol. blood samples were collected and at and were and used in the pharmacokinetic in to 48 hours, with blood at and Urine was collected and urine was collected to the blood for most 48 for the in blood samples were and the plasma was for to a that the agent concentration measured the concentration of the 2 agents at each time were as to the of the iohexol concentration versus time analysis relmapirazin concentration versus time The determination of iohexol concentration in plasma was performed with the of protein of an of the resulting in a for liquid mass and analysis of liquid mass The samples of iohexol in human plasma from were as of the for this et comparison of iohexol Med. 2018; 2: PubMed Google G. S. C. et of determination of the glomerular filtration marker iohexol by as by Med. 2019; PubMed Scopus Google Scholar The measurement of the concentration of relmapirazin in plasma was and J.-J. Riley I.R. Dorshow R.B. Clinical analysis and of MB-102, a novel tracer in human 2018; Google Scholar the plasma was with phosphate-buffered and This solution was an liquid with The was at the and for and The of was with linear to these the entire concentration range of the clinical The was to urine of The of urine at each was and were for subsequent analysis of agent The amount of agent recovered was from the urine and to the agent amount for each subject. The for determination of each agent concentration in urine the procedures in with the of the due to the agent concentration greater in the urine samples than in the plasma versus time data for each subject were for pharmacokinetic A standard with and was chosen as the pharmacokinetic for The the under the and the clearance hence with other pharmacokinetic data were used in this analysis time for the 12-hour study and time for the study regression analysis and was used to evaluate the of the resulting GFRs between the 2 The Bland-Altman method comparing a new measurement to an measurement was also correlation was also In the used in eGFR J. et a J Kidney Full Text Full Text PDF PubMed Scopus Google Scholar of and of that by no than and from the reference were also comparing GFR to GFR. A of were for A and and are in for This the eGFR from the as well as mass and race. with eGFR of 3 and 4 were on than in for are in Supplementary of study A = = (0) mass (34) (16) estimated glomerular filtration are mean or A are with eGFR are with eGFR in a new estimated glomerular filtration are mean or A are with eGFR are with eGFR Relmapirazin concentration was measured for plasma Figure 2 relmapirazin plasma concentrations with time for with kidney function ranging from normal to from the of the pharmacokinetic analysis the that was observed in subject of kidney function The of agent is in the at the time that is of the is between these 2 pharmacokinetic the phase by the in these is of glomerular filtration A rapid of the agent from the is from with GFR, a of the agent from the is from with GFR. Relmapirazin pharmacokinetic were also from the pharmacokinetic for each subject. The and values for several pharmacokinetic are in Supplementary Iohexol concentration as the reference comparator was measured for plasma The iohexol plasma concentrations with time for the as in Figure 2 function ranging from normal to are in Supplementary Figure data also the of an GFR tracer agent P. Ebert N. Melsom T. et al.Iohexol plasma clearance for measuring glomerular filtration rate in clinical practice and research: a review. Part 1: how to measure glomerular filtration rate with iohexol.Clin Kidney J. 2016; 9: 682-689Crossref PubMed Scopus (170) Google Scholar GFR was from the agent concentration versus time data using standard pharmacokinetic analysis each a GFR was from the relmapirazin as the and from the iohexol as the reference data a range of measured GFRs from normal kidney function to that of with 4 The of these 2 GFR values for each subject is in Supplementary The GFR from the relmapirazin plasma is the GFR from the iohexol plasma in Figure for data from across The linear regression analysis with at a of with of with and for the of and the on the an of with no change in the A Bland-Altman analysis of this data in Figure 4 agreement between these 2 measures of GFR. The and are and -7.0 The are across the GFR and no is The mean is and The linear regression analysis was performed on the data by A of was which is the commonly accepted versus kidney and values and this are the as for the entire data in 2 and in Figure of linear regression analysis between and of iohexol of Figure of iohexol of Figure glomerular filtration GFR of iohexol of Figure in a new GFR, glomerular filtration The linear regression analysis was also performed on the data by race. were and and values for each are the no was data are in 2 also and in Supplementary Figure The GFR data in Figure 3 and in Supplementary were also as and the of relmapirazin GFR that by no than or from the iohexol GFR = = and = are in with the mean and the This method also an agreement between the GFR and the of between and and are of that by no than and in a new and are of that by no than and Furthermore, a of correlation a of the 120 in this study, were due to urine the study, urine collected over 12 hours, and urine collected over 48 hours. Figure the of of administered dose in urine between the relmapirazin and iohexol as a function of the measured GFR for the with urine over 12 hours. The standard of the mean is also The values in Figure are the of in each The values are in Supplementary normal kidney function and for in and 2 the of agents in the is 2 to hours, hence a 12-hour to of the relmapirazin administered dose was recovered in the 12 of this The of these agents is 4 for with patients 3 and for with 4 is in 12 for these for with 3 and for with The data that the recovered administered dose amount in the urine for relmapirazin that of iohexol in these kidney function in urine collected over a time period of were chosen to the range of kidney function from normal to 4 Relmapirazin dose was greater than or equal to iohexol dose in The relmapirazin dose for these was to iohexol of Supplementary Figure of in urine between the relmapirazin and iohexol for each of these The individual and values are in Supplementary Relmapirazin was rationally designed with the necessary properties to be a fluorescent GFR tracer R. Neumann W.L. Poreddy A.R. et al.Hydrophilic pyrazine dyes as exogenous fluorescent tracer agents for real-time point-of-care measurement of glomerular filtration rate.J Med Chem. 2011; 54: 5048-5058Crossref PubMed Scopus (45) Google J.-J. Riley I.R. Rogers T.E. et al.Characterization of MB-102, a new fluorescent tracer agent for point-of-care renal function monitoring.J Pharm Sci. 2020; 109: 1191-1198Abstract Full Text Full Text PDF PubMed Scopus (14) Google Scholar Relmapirazin concentration versus time data the observed for kidney tracer The time to between these 2 to be between and 2 on subject which is also to small molecule kidney tracer P. Ebert N. Melsom T. et al.Iohexol plasma clearance for measuring glomerular filtration rate in clinical practice and research: a review. Part 1: how to measure glomerular filtration rate with iohexol.Clin Kidney J. 2016; 9: 682-689Crossref PubMed Scopus (170) Google Scholar The comparison of the plasma GFR, to the reference plasma GFR, excellent by the regression analysis with = an perfect between the 2 agent GFR The in 3 the excellent agreement between the as the correlation that relmapirazin is a kidney tracer agent in humans was observed in the mass with the that the of of the administered dose of relmapirazin in urine that of iohexol for each kidney function Relmapirazin was to be the in this No severe adverse were and adverse were or in in Supplementary were no clinical or Relmapirazin has no with or commonly used or The glomerular filtration in the of Figure 2 no over the 12-hour data In this most 3 with a of on the study dosing of by the with 3 and 4 in the study is in Supplementary relmapirazin is by and of this study be the measured GFR data the range the is are than in the study is than as with were well in the study C. et glomerular filtration rate determination using and the for measurement 2021; Full Text Full Text PDF PubMed Scopus Google Scholar and a was used to from each subject. the reference GFR iohexol has its as a GFR tracer specifically that is to have an clearance P. Ebert N. Melsom T. et al.Iohexol plasma clearance for measuring glomerular filtration rate in clinical practice and research: a review. Part 1: how to measure glomerular filtration rate with iohexol.Clin Kidney J. 2016; 9: 682-689Crossref PubMed Scopus (170) Google Scholar However, in clinical study, the of of the urine from the of a clearance determination for relmapirazin at this of in the is point of care real-time direct measurement of Z.H. Pickering J.W. Walker R.J. Clearance and beyond: the complementary roles of GFR measurement and injury biomarkers in acute kidney injury (AKI).Am J Physiol Renal Physiol. 2011; 301: F697-F707Crossref PubMed Scopus (125) Google Scholar that are direct measures of GFR are to point of serial blood of known GFR as or of GFR is on or for determination of plasma followed by pharmacokinetic using the measured agent concentration as a function of This is also of the GFR measurement using a fluorescent which for GFR D.V. Meier D. Sandoval R.M. et al.A novel method for rapid bedside measurement of GFR.J Am Soc Nephrol. 2018; 29: 1609-1613Crossref PubMed Scopus (47) Google Scholar We to the fluorescent of relmapirazin for point-of-care detection in a phase clinical study In an in vivo of renal and excellent correlation between and relmapirazin as well as excellent correlation with real-time assessment of drug is and of for patients renal R.B. et detection of and correlation to renal in vivo J. 2023; PubMed Scopus (0) Google Scholar In phase II pilot data relmapirazin is a novel GFR tracer agent in humans. studies are needed and to relmapirazin and in the and in acute GFR is is a the the and a of is an and of is the of clinical and a of is a and of data in the are the and the The data enabling of Supplementary an are from the corresponding on We the W. at Washington University in J. at Louis and at the Orlando Clinical Research in and The to M. of for In D. for a of the for this was provided by

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