As an application of near-infrared spectroscopy, pulse oximetry is widely used to non-invasively measure the arterial oxygen saturation (SpO2). To improve the maternal and fetal outcome during delivery, transabdominal fetal pulse oximetry can be used to measure the fetal SpO2. The layered tissue structure above the fetus, however, is complex and thick. In order to understand the feasibility of transabdominal pulse oximetry, we simulated light propagation through the maternal abdomen. For realistic geometry, we segmented a magnetic resonance imaging (MRI) scan of a pregnant women to create a 3D anatomical model, from which a 3D tetrahedron mesh was generated. Using this mesh, we then simulated photon propagation for 5 wavelengths and a grid of 70 sources and detectors (35 each) on the maternal abdomen above the fetal head with NIRFAST. Finally, depth sensitives were examined with Jacobian (J) and flat field. For a fetal head at ~4 cm depth, we found the normalized J at this depth is ~0.1-1% for source-detector distances within 9-10 cm. We also observed that at the same depth, the normalized flat field sensitivity is ~5-10%, which is 1-2 orders of magnitude higher than the normalized J. These results indicate that enough light can reach the fetus when considering ~9 cm source detector distances.
Point-of-care (POC) measured glomerular filtration rate (GFR) has been a goal of nephrologists for the last 30 years. To this end, a fluorescent GFR tracer agent, relmapirazin, with appropriate photophysical properties for transdermal detection has been rationally designed. Twenty-four standard nonclinical assays to evaluate toxicity as required by the FDA have been performed. Fluorescent detection instrumentation to acquire and process the emission signal from the agent through the skin has been developed. The pilot clinical study objectives were to: • Demonstrate relmapirazin is a GFR agent in humans by comparison to a known non-fluorescent standard agent, • Establish that the GFR as measured by the transdermal fluorescence excretion rate matches the standard labor-intensive non-POC plasma GFR. This combination product was evaluated on 120 subjects covering three clinical sites. Subjects were enrolled spanning normal to very impaired renal function, and for all six skin colors on the Fitzpatrick Skin Scale. Relmapirazin and the standard agent were intravenously administered in consecutive boluses. Prior to dosing, the transdermal sensor was placed on the chest of each subject and fluorescent readings were initiated. The plasma-derived GFR measured from relmapirazin matched the plasma-derived GFR measured from the standard agent. An algorithm was developed to convert the transdermal fluorescence measurement directly into a GFR, applicable to the entire GFR range and for all skin colors. No serious nor significant adverse events were reported. Clinically amenable point-of-care measured GFR has been demonstrated for subjects with a range of GFR values and for all skin colors.
The intestinal mucosal barrier prevents macromolecules and pathogens from entering the circulatory stream in a healthy gut. Tight junctions in this barrier are compromised in many intestinal disorders including inflammatory bowel diseases (such as Crohn’s and ulcerative colitis), celiac disease, graft vs host disease, environmental enteropathy, and enteric dysfunction. Dual sugar absorption tests are a standard method for measuring gastrointestinal integrity in humans. The larger molecular weight sugar, lactulose, is minimally absorbed through a healthy gut. The smaller molecular weight sugar, mannitol or rhamnose, in contrast, is readily absorbed through both a healthy and inflamed gut. Thus, a higher ratio of lactulose to mannitol or rhamnose reflects increased intestinal permeability. However, several issues prevent the widespread use of the dual sugar assay including requirements for lengthy urine collection, transport of specimens under conditions free from contamination and bacterial growth, analysis by sophisticated laboratory equipment, and the associated lengthy turnaround time. In a recent publication, the feasibility of employing a dual fluorescent tracer agent assay to mimic the dual sugar absorption test without the need for specimen collection was reported. This dual fluorophore assay for GI permeability was demonstrated in an indomethacin-induced bowel disease model in rats. However, one of the fluorophores was not entirely biocompatible. Herein is reported a dual fluorophore system that is totally biocompatible, with emphasis on the transdermal detection of the fluorophores, thus enabling the use of the technology for noninvasive point-of-care gastrointestinal permeability determination.
A prototype medical device for monitoring kidney function by transdermal measurement of the clearance rate of the exogenous fluorescent tracer agent MB-102 (administered intravenously) was developed. Verification of the device with an in vitro protocol is described. The expected renal clearance of the agent was mimicked by preparing a dilution series of MB-102 in the presence of a scattering agent. The slope of a linear fit to the logarithm of fluorescence intensity as a function of dilution step agreed with predictions within 5%, a level of accuracy that would be adequate in assessment of GFR to prevent misdiagnosis of kidney disease. Transdermal measurement was validated using a rat model. A two-compartment pharmacokinetic dependence was observed, with equilibration of the fluorescent agent between the vascular space into which it was injected and the extracellular space into which it subsequently diffused. The best observed signal-to-noise ratios were about 150, allowing determination of the renal clearance time with 5% precision using a 10-min fitting window. Based on the verification and validation methods for transdermal fluorescence detection described herein, the instrument has been approved by the FDA for a first-in-human clinical study, and a first transdermal clearance curve in a human is presented herein.
The fluorescent tracer agent 3,6-diamino-2,5-bisN-[(1R)-1-carboxy-2-hydroxyethyl]carbamoylpyrazine, designated MB-102, is cleared from the body solely by the kidneys. A prototype noninvasive fluorescence detection device has been developed for monitoring transdermal fluorescence after bolus intravenous injection of MB-102 in order to measure kidney function. A mathematical model of the detected fluorescence signal was created for evaluation of observed variations in agent kinetics across body locations and for analysis of candidate instrument geometries. The model comprises pharmacokinetics of agent distribution within body compartments, local diffusion of the agent within the skin, Monte Carlo photon transport through tissue, and ray tracing of the instrument optics. Data from eight human subjects with normal renal function and a range of skin colors shows good agreement with simulated data. Body site dependence of equilibration kinetics was explored using the model to find the local vasculature-to-interstitial diffusion time constant, blood volume fraction, and interstitial volume fraction. Finally, candidate instrument geometries were evaluated using the model. While an increase in source-detector separation was found to increase sensitivity to tissue optical properties, it reduced the relative intensity of the background signal with minimal effect on the measured equilibration kinetics.
A prototype medical device for trans-cutaneous monitoring of kidney function has been developed, validated, and used in a clinical trial on 16 healthy subjects having a wide range of skin color types. The fluorescent tracer agent MB-102 was administered intravenously as a bolus that was varied between 0.5 and 4 μmole/kg subject weight. The tracer agent was tracked as a function of time in plasma by blood sampling and trans-cutaneously at four body sites (sternum, forehead, arm, and side) simultaneously. Excitation was performed with a very low level of amplitude-modulated LED light at 450 nm (<50 μW/cm2), and fluorescence emission was synchronously detected at 570 nm. With adjustment of detection gain between subjects, no skin color dependence was observed of the signal-to-noise ratio (SNR) of the transcutaneous measurements. The primary source of measurement noise appeared to be subject motion, likely due to variations in blood content at the skin measurement site. A typical two-compartment pharmacokinetic dependence was observed with equilibration of the fluorescent agent between the vascular space into which it was injected and the extracellular space into which it subsequently diffused. Variation of this equilibration time was observed across body sites, with the sternum providing the shortest and most consistent equilibration. After equilibration, the terminal fluorescence time dependence at the sternum site was found to be highly correlated with tracer agent concentration time dependence sampled from the blood plasma.
The fluorescent tracer agent 2,5-bis[N-(1-carboxy-2-hydroxy)]carbamoyl-3,6-diaminopyrazine, designated MB-102, has been developed with properties and attributes necessary for use as a direct measure of glomerular filtration rate (GFR). Comparison to known standard exogenous GFR agents in animal models has demonstrated an excellent correlation. A clinical trial to demonstrate this same correlation in humans is in progress. This clinical trial is the first in a series of trials necessary to obtain regulatory clearance from the FDA. We report herein the comparison of plasma pharmacokinetics between MB-102 and the known standard exogenous GFR agent Iohexol in healthy subjects with normal renal function. Post simultaneous administration of both agents, blood samples over a period of 12 hours were collected from each subject to assess pharmacokinetic parameters including GFR. Urine samples were collected over this same period to assess percent injected dose recovered in the urine. Results indicate MB-102 is a GFR agent in humans from the comparison to the standard agent.
The aim of this work was to establish measurement conditions under which endogenous skin fluorescence ("auto-fluorescence")
is relatively invariant, so that changes in exogenous agents can be accurately determined. Fluorescence
emission was measured on the volar forearm of 36 subjects, chosen to be equally representative of all 6 Fitzpatrick skin
types. All subjects were exposed to approximately 40 minutes of optical excitation at 450 and 500 nm with 4 irradiances
between 0.3 and 9 mW/cm2. Both non-optically-induced (e.g. tissue settling and fluctuation) and optically-induced
variations were observed in the measured fluorescence and mechanisms explaining these effects are proposed. The
optically-induced auto-fluorescence decay was independent of skin type when excited at 450 nm, but significantly
dependent on skin type when excited at 500 nm. Further, the extent of decay over time was linearly related to irradiance
at 500 nm, but at 450 nm was non-linear, with the extent of decay rolling off between 2 and 9 mW/cm2. In order to
maintain the auto-fluorescence signal within 95% of its original value over a 30 minute period, the excitation at 450 nm
would need to be limited to 1.5 mW/cm2, while excitation at 500 nm should be limited to 5 mW/cm2.
Accurate measurement of glomerular filtration rate (GFR) at the bedside is highly desirable in order to assess renal
function in real-time, which is currently an unmet clinical need. In our pursuit to develop exogenous fluorescent tracers
as GFR markers, various hydrophilic derivatives of 3,6-diaminopyrazine-2,5-dicarboxylic acid with varying molecular
weights and absorption/emission characteristics were synthesized. These include polyhydroxyalkyl based small
molecules and poly(ethylene glycol) (PEG) substituted moderate molecular weight compounds, which were further sub-grouped
into analogs having blue excitation with green emission, and relatively longer wavelength analogs having green
excitation with orange emission. Lead compounds were identified in each of the four classes on the basis of structure-
activity relationship studies, which included in vitro plasma protein binding, in vivo urine recovery of administered dose,
and in vivo optical monitoring. The in vivo optical monitoring experiments with lead candidates have been correlated
with plasma pharmacokinetic (PK) data for measurement of clearance and hence GFR. Renal clearance of these
compounds, occurring exclusively via glomerular filtration, was established by probenecid blocking experiments. The
renal clearance property of all these advanced candidates was superior to that of the iothalamate, which is currently an
accepted standard for the measurement of GFR.
Rapid assessment of glomerular filtration rate (GFR), which measures the amount of plasma filtered through the
kidney within a given time, would greatly facilitate monitoring of renal function for patients at the bedside in the clinic.
In our pursuit to develop exogenous fluorescent tracers for real-time monitoring of renal function by optical methods, N-alkylated
aminopyrazine dyes and their hydrophilic conjugates based on poly (ethylene glycol) (PEG) were synthesized
via reductive amination as the key step. Photophysical properties indicated a bathochromic shift on the order of 50 nm in
both absorption and emission compared to naked aminopyrazines which could be very useful in enhancing both tissue
penetration as well as easier detection methods. Structure-activity relationship (SAR) and pharmacokinetic (PK) studies,
and the correlation of in vivo optical data with plasma PK for measurement of clearance (and hence GFR) are focus of
the current investigation.
The ability to continuously monitor renal function via the glomerular filtration rate (GFR) in the clinic is currently an
unmet medical need. To address this need we have developed a new series of hydrophilic fluorescent probes designed
to clear via glomerular filtration for use as real time optical monitoring agents at the bedside. The ideal molecule should
be freely filtered via the glomerular filtration barrier and be neither reabsorbed nor secreted by the renal tubule. In
addition, we have hypothesized that a low volume of distribution into the interstitial space could also be advantageous.
Our primary molecular design strategy employs a very small pyrazine-based fluorophore as the core unit. Modular
chemistry for functionalizing these systems for optimal pharmacokinetics (PK) and photophysical properties have been
developed. Structure-activity relationship (SAR) and pharmacokinetic (PK) studies involving hydrophilic pyrazine
analogues incorporating polyethylene glycol (PEG), carbohydrate, amino acid and peptide functionality have been a
focus of this work. Secondary design strategies for minimizing distribution into the interstitium while maintaining
glomerular filtration include enhancing molecular volume through PEG substitution. In vivo optical monitoring
experiments with advanced candidates have been correlated with plasma PK for measurement of clearance and hence
GFR.
A series of experiments were performed to assess the feasibility of measuring total hemoglobin (HbT) non-invasively using light in the visible to near infrared (NIR) range. The experiments included spectroscopic measurements of both in vitro isolated tissue components and in vivo human tissue (73 subjects) in a clinical setting. Several different methods of estimating HbT, all by optical non-invasive means, were tested and compared for accuracy relative to standard invasive methods. By combining time-resolved optical absorbances at 3 wavelengths, 750, 965, and 1320 nm, in a fashion that is highly analogous to pulse oximetry, HbT was predicted with a standard error of 1.1 g/dL.
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