NIH Public Access
`Author Manuscript
`Mol Genet Metab. Author manuscript; available in PMC 2014 December 01.
`Published in final edited form as:
`Mol Genet Metab. 2013 December ; 110(4): 446–453. doi:10.1016/j.ymgme.2013.09.017.
`
`ELEVATED PHENYLACETIC ACID LEVELS DO NOT
`CORRELATE WITH ADVERSE EVENTS IN PATIENTS WITH
`UREA CYCLE DISORDERS OR HEPATIC ENCEPHALOPATHY
`AND CAN BE PREDICTED BASED ON THE PLASMA PAA TO
`PAGN RATIO
`
`M. Mokhtarania, G.A. Diazb, W. Rheadc, S.A. Berryd, U. Lichter-Koneckie, A. Feigenbaumf,
`A. Schulzef, N. Longog, J. Bartleyh, W. Berquisti, R. Gallagherj, W. Smithk, S.E.
`McCandlessl, C. Hardingm, D.C. Rockeyn, J.M. Vierlingo, P. Mantryp, M. Ghabrilq, R.S.
`Brown Jrr, K. Dickinsona, T. Moorsa, C. Norrisa, D. Coakleya, D.A. Milikiens, SC Nagamanit,
`C. LeMonsu, B. Leet, and B.F. Scharschmidta
`a Hyperion Therapeutics, 601 Gateway Blvd., STE 200, South San Francisco, CA 94080, USA
`
`b Icahn School of Medicine at Mount Sinai, Department of Genetics and Genomic Sciences,
`Department of Pediatrics, 1428 Madison Ave., New York, NY 10029, USA
`
`c The Medical College of Wisconsin, 9000 W. Wisconsin Ave., Milwaukee, WI 53226, USA
`
`d University of Minnesota, Minneapolis, 420 Delaware St., SE MMC 75, Minneapolis, MN 55455,
`USA
`
`e Children's National Medical Center, 111 Michigan Ave., NW #1950, Washington, DC 20010,
`USA
`
`f The Hospital for Sick Children and University of Toronto, Division of Clinical and Metabolic
`Genetics,, 555 University Avenue, Toronto, ON, Canada M5G1X8
`
`g The University of Utah, Division of Medical Genetics, 2C412 SOM, 50 North Mario Capecchi
`Drive, Salt Lake City, UT 94132, USA
`
`h Long Beach Memorial Hospital, 2865 Atlantic Avenue, #104, Long Beach, CA 90806, USA
`
`© 2013 Elsevier Inc. All rights reserved.
`Corresponding author: Masoud Mokhtarani, M.D. VP, Clinical Development & Medical Affairs Hyperion Therapeutics, Inc. 601
`Gateway Blvd., Ste. 200 South San Francisco, CA 94080 Phone: 1-650-745-7838 Mobile: 1-650-438-8667 Fax: 1-650-745-3581
`Masoud.mokhtarani@hyperiontx.com.
`Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our
`customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of
`the resulting proof before it is published in its final citable form. Please note that during the production process errors may be
`discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
`Conflict of Interest Statement: D. Coakley, K. Dickinson, M. Mokhtarani, T. Moors, C. Norris and B.F. Scharschmidt are/were
`employees of Hyperion at the time of the study. D. Milikien is an employee of Accudata, which was paid by Hyperion to perform the
`biostatistical analyses.
`None of the other authors have a financial interest in Hyperion, although payments were made by Hyperion to all investigators for
`services related to the clinical trials.
`ClinicalTrials.gov Identifiers: ClinicalTrials.gov NCT00551200, NCT00947544, NCT00992459, NCT00947297, NCT00999167,
`NCT 01347073
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`i Stanford University, 750 Welch Road, #116, Palo Alto, CA 94305, USA
`
`j Children's Hospital Colorado, 13123 East 16th Avenue, B153, Aurora, CO 80045, USA
`
`k Maine Medical Center, 1577 Congress Street, 2nd Floor, Portland, ME 04102, USA
`
`l Center for Human Genetics, Case Western Reserve University and University Hospitals Case
`Medical Center, 11100 Euclid Avenue Cleveland, OH 44106, USA
`
`m Oregon Health Sciences University, 3181 SW Sam Jackson Park Road, A036/B198, Mailcode
`L103, Portland, OR 97239, USA
`
`n Medical University of South Carolina, Department of Medicine, 96 Jonathan Lucas St. 803CSB
`Charleston, SC 29425, USA
`
`o Baylor College of Medicine, Departments of Medicine and Surgery, 6620 Main St # 1475,
`Houston, TX 77030, USA
`
`p Liver Institute at Methodist Dallas Medical Center, Dallas 1411 N. Beckley Ave., Pavilion III,
`STE 268 Dallas, TX 75203, USA
`
`q Indiana University, Gastroenterology and Hepatology, Regenstrief Health Center, Room 4100
`1050 Wishard Blvd. Indianapolis, IN 46202, USA
`
`r Columbia University Medical Center, Department of Surgery, Room PH 14, 105, 622 West 168th
`St. New York, NY 10032, USA
`
`s Accudata Solutions, Inc., 886 Oak St., Lafayette, CA 94549, USA
`
`t Baylor College of Medicine, One Baylor Plaza, Room R814, Houston, TX, USA
`
`u The National Urea Cycle Disorders Foundation, Pasadena, CA, 75 South Grand Avenue,
`Pasadena, CA, 91105
`
`Abstract
`
`Background—Phenylacetic acid (PAA) is the active moiety in sodium phenylbutyrate (NaPBA)
`and glycerol phenylbutyrate (GPB, HPN-100), both are approved for treatment of urea cycle
`disorders (UCDs) - rare genetic disorders characterized by hyperammonemia. PAA is conjugated
`with glutamine in the liver to form phenylacetyleglutamine (PAGN), which is excreted in urine.
`PAA plasma levels ≥500 μg/dL have been reported to be associated with reversible neurological
`adverse events (AEs) in cancer patients receiving PAA intravenously. Therefore, we have
`investigated the relationship between PAA levels and neurological AEs in patients treated with
`these PAA pro-drugs as well as approaches to identifying patients most likely to experience high
`PAA levels.
`
`Methods—The relationship between nervous system AEs, PAA levels and the ratio of plasma
`PAA to PAGN were examined in 4683 blood samples taken serially from: [1] healthy adults [2],
`UCD patients ≥2 months of age, and [3] patients with cirrhosis and hepatic encephalopathy (HE).
`The plasma ratio of PAA to PAGN was analyzed with respect to its utility in identifying patients
`at risk of high PAA values.
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`Results—Only 0.2% (11) of 4683 samples exceeded 500 ug/ml. There was no relationship
`between neurological AEs and PAA levels in UCD or HE patients, but transient AEs including
`headache and nausea that correlated with PAA levels were observed in healthy adults. Irrespective
`of population, a curvilinear relationship was observed between PAA levels and the plasma
`PAA:PAGN ratio, and a ratio > 2.5 (both in μg/mL) in a random blood draw identified patients at
`risk for PAA levels > 500 μg/ml.
`
`Conclusions—The presence of a relationship between PAA levels and reversible AEs in healthy
`adults but not in UCD or HE patients may reflect intrinsic differences among the populations
`and/or metabolic adaptation with continued dosing. The plasma PAA:PAGN ratio is a functional
`measure of the rate of PAA metabolism and represents a useful dosing biomarker.
`
`Keywords
`BUPHENYL; glycerol phenylbutyrate; HPN-100; neurological adverse events; pharmacokinetics;
`RAVICTI; sodium phenylbutyrate
`
`INTRODUCTION
`
`Glycerol phenylbutyrate, a sodium- and sugar-free phenylbutyrate derivative, and sodium
`phenylbutyrate are approved as ammonia lowering agents in patients with urea cycle
`disorders (UCDs). Both are pro-drugs of phenylacetic acid (PAA), which is formed by beta-
`oxidation from phenylbutyric acid (PBA) delivered either as glycerol phenylbutyrate
`following its intestinal hydrolysis by pancreatic lipases [1] or as sodium phenylbutyrate
`following dissociation in the stomach. PAA is conjugated with glutamine by glutamine-N-
`phenylacetyltransferase, largely in the liver and to a lesser extent in the kidney [2], to form
`phenylacetylglutamine (PAGN), which is excreted in urine, thereby providing an alternate
`pathway to urea for waste nitrogen excretion. In controlled studies population
`pharmacokinetic analyses of sodium phenylbutyrate and glycerol phenylbutyrate, it has been
`shown that the gastrointestinal absorption of PBA is approximately 75% slower when
`delivered as glycerol phenylbutyrate vs. sodium phenylbutyrate and that plasma PAA and
`PAGN levels show less variability during glycerol phenylbutyrate dosing. [3]-[7]. There are
`over 30 reports of the administration of sodium phenylacetate or sodium phenylbutyrate to
`healthy volunteers, patients with UCDs or other metabolic disorders and patients with
`cancer, many of which reported some adverse events (AEs) attributed to PAA
`(Supplemental Table 1) [8]-[36]. These reversible AEs in cancer patients were reported in
`studies involving continuous or intermittent intravenous administration designed to maintain
`high levels of PAA, suggesting that duration of exposure as well as peak PAA levels are
`important [35],[3].
`
`The AEs reportedly associated with high levels of PAA have most commonly included
`nausea, headache, emesis, fatigue, weakness, lethargy, somnolence, dizziness, slurred
`speech, memory loss, confusion, and disorientation [35], [36]. Except for the symptoms of
`Kussmaul respiration, metabolic acidosis, cerebral edema, and coma associated with a fatal
`overdose of sodium phenylacetate/sodium benzoate (AMMONUL®)[13], the symptoms
`were rapidly reversible with reduced dosing or interruption of dosing.
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`Based on a detailed analysis of the timing of the AEs in relation to blood PAA
`concentrations, Simell calculated the safe upper PAA concentration limit to be 3.5 mmol/L,
`equivalent to 476 μg/mL [22], and Thibault reported that AEs were associated with PAA
`levels ranging from 499–1285 μg/mL [35], [36] .
`
`Sodium phenylbutyrate (BUPHENYL®) has been used for over three decades in the
`treatment of UCDs. Despite the fact that the AEs reportedly associated with elevated plasma
`PAA levels can mimic those associated with hyperammonemia, little is known regarding the
`relationship between PAA levels and AEs in UCD patients. The clinical trials of glycerol
`phenylbutyrate (RAVICTI®, HPN-100), which included over 100 UCD patients, 80 of
`whom underwent comparative study of sodium phenylbutyrate and glycerol phenylbutyrate
`[3] - [6] (the largest prospectively studied group of patients with this rare disorder), 193
`patients with advanced cirrhosis complicated by hepatic encephalopathy (HE) [37], and
`more than 90 healthy adult subjects have afforded a unique dataset and opportunity to
`systematically examine the relationship between PAA levels and AEs and to explore
`biomarkers indicative of patients most likely to experience elevated PAA levels.
`
`METHODS
`
`Clinical Studies (Table 1)
`
`Data from a thorough QTc study in healthy adults, five clinical studies in UCD patients and
`an open label safety and dose escalation study as well as a randomized, double-blinded
`controlled phase 2 study of patients with decompensated cirrhosis complicated by HE
`formed the basis for these analyses.
`
`UCD Patients
`
`Eighty UCD patients completed 4 short-term (10 to 28 days) cross-over studies of sodium
`phenylbutyrate vs. glycerol phenylbutyrate (Table 1). The short-term UCD study population
`included 26 pediatric patients ages ≥2 mos through 17 years who received a mean (range)
`dose of 8 (1-19) g/day of glycerol phenylbutyrate or an equivalent dose of sodium
`phenylbutyrate and 54 adults patients ages 18 years or older who received a mean (range)
`dose of 13 (2-34) g/day of glycerol phenylbutyrate or an equivalent dose of sodium
`phenylbutyrate [3] [4] [5][6]. In addition, data from 100 UCD patients enrolled in 12-month
`glycerol phenylbutyrate treatment protocols including 49 children and 51 adults were
`analyzed in relation to PAA levels over time and the occurrence of the symptoms reported in
`cancer patients by Thibault [35][36][4] [5][6] during 12 months treatment.
`
`Patients with Cirrhosis and HE
`
`Data from a 4-week safety and dose escalation study and a multicenter, randomized placebo-
`controlled study of 178 patients with cirrhosis and hepatic encephalopathy who received
`13.2 g/day of glycerol phenylbutyrate (N=90) or placebo (N=88) for 16 weeks were
`analyzed [37], [38] (Table 1). Patients were monitored for safety and frequent PK samples
`were taken over the course of the study.
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`Healthy Adults
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`Page 5
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`A total of 98 healthy adults (mean age of 28; 53 male 45 female) participated in a blinded,
`randomized, cross over study to assess effects of glycerol phenylbutyrate and its metabolites
`on QTc and other ECG parameters (Table 1). In this protocol 12 subjects received 29.7 g/
`day, 4 subjects 39.6 g/day of glycerol phenylbutyrate and 68 subjects received placebo,
`moxifloxacin as the positive control and glycerol phenylbutyrate at doses of 13.2 g/day and
`19.8 g/day administered three times daily for 3 days.
`
`Adverse Event Mapping
`
`All treatment emergent adverse events (AEs) coded as to Body System as Nervous System
`Disorders using the Medical Dictionary for Regulatory Activities (MedDRA) in subjects
`enrolled in these studies were included in the analyses. For UCD patients, the specific
`toxicities reported by Thibault [35], [36], including nausea, headache, emesis, fatigue,
`weakness, lethargy, somnolence, dizziness, slurred speech, memory loss, confusion, and
`disorientation, exacerbation of neuropathy, pedal edema, hearing loss, abnormal taste,
`arrhythmia, rash, Kussmaul respiration, metabolic acidosis, increased anion gap, tachypnea,
`abdominal discomfort, cerebral edema, and obtundation or coma, were mapped to the
`MedDRA preferred terms in the clinical trial databases.
`
`Analysis of AEs in Relation to PAA Levels
`
`Analyses were based on (a) 2126 samples from 98 healthy adults, (b) 1281 blood PAA and
`PAGN values derived from 80 UCD patients during the short term-switchover studies who
`received both sodium phenylbutyrate and glycerol phenylbutyrate, and (c) 428 samples from
`90 patients with cirrhosis and HE who received glycerol phenylbutyrate. Because plasma
`PAA levels were not always available at the time the patient was experiencing an AE, the
`following rules were applied to associate an AE to a known PAA level. For healthy subjects,
`maximum PAA values recorded after the first dose but within 24 hours of the last dose and
`the incidence of neurological AEs (yes/no) were summarized by dosing period; for periods
`where subjects received placebo or moxifloxacin, the PAA levels were set to 0. For UCD
`patients, maximum PAA values (Cmax) recorded during each dosing period and the
`incidence of neurological AEs were summarized by treatment (glycerol phenylbutyrate or
`sodium phenylbutyrate). For HE patients, each AE was attributed to the PAA result that was
`closest in time to the AE.
`
`The contribution of a 20 μg/mL increase in PAA levels to the probability of a neurological
`AE regardless of relationship to the study drug was examined using Generalized Estimating
`Equations [39]. For healthy subjects, data were summarized for each dose group. Since
`UCD patients received a range of doses, data were summarized for patients receiving a dose
`greater or less than the median dose (equivalent to 11.7 g/day). For HE patients,
`neurological AEs were examined both in relation to blinded treatment group assignment; i.e.
`glycerol phenylbutyrate or placebo, as well as in relation to PAA levels among patients
`treated with glycerol phenylbutyrate.
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`Analysis of PAA in Relation to Plasma PAA:PAGN Ratio
`
`GEE were used to model the predictive value of plasma PAA:PAGN ratio in identifying
`patients at risk of a high plasma PAA level as defined to have a PAA level equal or greater
`than 400 μg/mL or 500 μg/mL during 24 hours of dosing . Plasma PAA:PAGN ratios were
`grouped into binary categorical range of less than 2.5 or greater than 2.5. The repeated
`measures categorical outcome was modeled using GEE with a logit link function, ratio
`category as the independent variable, and the individual subject ID as the repeated measures
`factor. Confidence intervals for the predicted probabilities were computed by bootstrap
`estimation of 1000 re-samplings of the original data, as detailed in Davison and Hinkley
`[40].
`
`RESULTS
`
`UCD patients (Table 2, Figure 1)
`
`Common AEs reported by at least 10% of patients during short-term treatment with either
`drug included diarrhea, flatulence, and headache. Neurological AEs reported by more than 1
`UCD patient included headache, dizziness and dysgeusia. The mean (SD) PAA Cmax was
`similar in patients who reported at least one neurological AE, as compared with those who
`did not (50.8 (34.5) μg/mL vs 51.5 (49.23) μg/mL respectively). There was no statistically
`significant relationship in UCD patients between the presence or absence of neurological
`AEs and PAA levels during either glycerol phenylbutyrate or sodium phenylbutyrate
`treatment. The odds ratio of a neurological AE occurring for each 20 μg/mL increase in
`PAA levels for the two drugs combined, controlling for dose level, was 0.929, very close to
`1 indicating that increasing levels of PAA were not associated with an increase in
`neurological AEs in these studies. There was no difference in the frequency of the PAA-
`associated AEs reported in cancer patients by Thibault [35], [36] in adult vs. pediatric UCD
`patients in the short-term controlled studies, despite the generally higher PAA levels in
`pediatric patients (Supplemental Table 2).
`
`A total of 100 UCD patients enrolled in 12-month studies of glycerol phenylbutyrate
`received a mean (SD) total dose of 11.01 (5.970) g/day (range: 0.8–34.3 g). Overall common
`AEs reported in at least 10% of UCD patients during long-term treatment included vomiting,
`upper respiratory tract infection, nausea, nasopharyngitis, diarrhea, headache,
`hyperammonemia, decreased appetite, cough, fatigue, dizziness, and oropharyngeal pain.
`There was no increase either in plasma PAA levels (Supplemental Figure 1) or the rate of
`AEs over time. Just as in the short-term studies there was no difference between pediatric
`and adult patients in the frequency of the PAA-associated AEs reported in cancer patients by
`Thibault (Supplemental Table 2).
`
`Patients with cirrhosis and HE (Table 2, Figure 1)
`
`Of 88 patients randomized to placebo, 48.9% reported a neurological AE as compared to
`40.9% of 90 patients randomized to glycerol phenylbutyrate. Of the 428 PAA data points
`from patients randomized to glycerol phenylbutyrate, 46 were in patients who reported a
`neurological AE and 382 in patients who did not. The mean (SD) PAA value closest to
`occurrence of an AE was 61.4 (75.3) μg/mL while the mean PAA value not temporally
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`associated with an AE was 36.4 (55.6) μg/mL (p=0.77) (Figure 2). Similar to UCD patients,
`there was no increase in the odds of experiencing a neurological AE with each 20 μg/mL
`increment in PAA levels in cirrhosis patients (odds ratio 1.086; p=0.172) indicating that at
`the dose of 13.2 g/day the odds of experiencing a neurological AE did not increase with an
`increase in PAA level.
`
`Healthy subjects
`
`Common AEs in ≥ 10% of healthy volunteers included headache, nausea, and dizziness.
`Neurological AEs increased in frequency with increasing dose, ranging from 26.5% for 13.2
`g/day to 91.7% for 29.7 g/day. Among those who reported a neurological AE, PAA values
`were higher for the 19.8 g/day, 29.7 g/day, and 39.6 g/day dosing periods than for the 13.2
`g/day dosing period (Figure 2, Table 2). PAA levels increased as the dose of glycerol
`phenylbutyrate increased. In the case of the 13.2 g/day dose group, the difference was
`statistically significant (73.3 vs. 41.6, p <0.001) (Table 2). Logistic regression analysis
`indicated that each increment in PAA of 20 μg/mL was associated with increasing odds of
`experiencing a neurological AE (odds ratio = 1.75; p = 0.006). Individual AEs reported by
`healthy adults were generally transient and typically began within 36 hours of dosing and
`generally resolved with continued dosing, as depicted in Supplemental Figure 2.
`
`Plasma PAA:PAGN ratio as a Predictor of Elevated PAA Levels
`
`PAA levels showed considerable variation over a 24-hr period in all patients regardless of
`the dose, drug and population (Figure 3). Unlike PAA, the ratio of PAA:PAGN was
`comparatively constant over 24 hours (data not shown). A curvilinear relationship was
`observed between PAA and PAA:PAGN in all populations, with a sharp upward inflexion
`beginning with PAA concentrations approaching 200 μg/ml and a PAA:PAGN of
`approximately 2.5 or greater (Figure 4). Only 11 of a total of 4683 samples exceeded the
`500 ug/ml threshold level reported by Thibault to be associated with occurrence of
`neurological AEs in cancer patients. The estimated probabilities of correctly detecting a ratio
`≥2.0 based on a single plasma sample taken at any time between the fasting morning sample
`(0 hr time point) and early evening (12 hr time point) remained relatively constant (77% to
`84%), indicating that the timing of blood draw did not have an impact on the ratio of
`PAA:PAGN in plasma regardless of the PAA concentration. Patients with a ratio ≥2.5 had
`significantly higher PAA levels than those with a ratio ≤2.5 (p<0.0001) and PAA:PAGN
`ratios ≥2.5 had an approximately 20 times higher probability of being associated with PAA
`levels > 400 μg/ml (0.8% vs. 19.1%) or 500 μg/ml (0.3% vs. 8.4%) (Table 3).
`
`DISCUSSION
`
`No relationship was observed among UCD patients between PAA levels and either
`neurological AEs, or the specific AEs reported by Thibault, during treatment with either
`glycerol phenylbutyrate or sodium phenylbutyrate. This is supported by (a) the absence of a
`relationship during short term treatment in UCD patients, in which the odds ratio for the
`likelihood of a neurological AE for every 20 μg/mL increase in PAA levels was 0.929, (b)
`the absence of a difference in the frequency of AEs similar to those reported in cancer
`patients by Thibault between pediatric and adult UCD patients during short or long-term
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`treatment, despite generally higher PAA levels in pediatric patients, and (c) the absence of
`any change in either PAA levels or the pattern of AEs during 12 months of dosing.
`Similarly, no statistical relationship was noted between PAA levels and neurological AEs
`among HE patients treated with 13.2g/day of glycerol phenylbutyrate for 16 weeks, as there
`was no difference in neurological AEs between the glycerol phenylbutyrate and placebo
`treatment arms, nor was there a relationship between PAA levels and the occurrence of
`neurological AEs.
`
`Among the healthy adult volunteers, a relationship was observed between PAA levels and
`the occurrence of any neurological AE (e.g. headache, dizziness, vomiting and nausea).
`These AEs were generally mild, started early in the dosing period, and disappeared with
`continued dosing. The theoretical risk of PAA toxicity is expected to be similar for sodium
`phenylbutyrate or glycerol phenylbutyrate, as both drugs convert to PAA upon absorption.
`The AEs reported by healthy volunteers in these studies receiving glycerol phenylbutyrate
`are generally consistent with prior reports involving administration of sodium
`phenylbutyrate. The mechanism for these AEs is unknown, although interference with brain
`biochemical function has been suggested [41]. These differences between populations may
`be attributable either to metabolic differences between UCD and HE patients, who exhibit
`pathological nitrogen retention and high glutamine levels, as compared with healthy adults,
`and/or metabolic adaptation that may occur with continued exposure to PAA in chronically
`treated patients. Consistent with adaptation are the findings that AEs tended to disappear
`with continued dosing in healthy adults and that the UCD patients enrolled in these studies
`had been treated with sodium phenylbutyrate for an average of more than 9 years.
`
`While most human tissues are capable of beta-oxidation and, hence, conversion of
`phenylbutyrate to PAA [42], enzymatic conversion of PAA to PAGN occurs primarily in the
`liver [2]. This may explain why conversion of PAA to PAGN appears to be a rate-limiting
`step in the metabolism of PAA prodrugs and why PAA metabolism may be compromised
`when liver function is poor, when availability of the precursor glutamine may be limited as
`in healthy subjects, and/or when the capacity of the enzymatic conversion may be limited as
`in very young children [7]. Regardless of the reason, decreases in the rate of PAGN
`formation are associated with an increased ratio of PAA to PAGN in plasma. It is interesting
`in this regard that the upward inflexion in PAA values assessed as a function of the
`PAA:PAGN ratio occurs at a concentration similar to the estimated Km of this reaction
`based on population PK modeling, which is approximately 190 μM as previously described
`by Monteleone et al [7].
`
`In clinical practice, interpretation of an individual PAA value is compromised by the fact
`that concentrations vary considerably over the course of the day due to the relatively short
`half-lives of PBA and PAA. For example among the clinical trials comprising the present
`analyses, plasma PAA fluctuation index varied from 843% - 3931%; and fasting and
`maximal PAA levels in HE patients ranged from 0 - 1.3 μg/mL and 248 - 532 μg/mL,
`respectively. As compared with measurement of PAA alone, measurement of the
`PAA:PAGN ratio appears to be a useful proxy for the efficiency with which an individual
`patient converts PAA to PAGN, and a predictor of patients at risk of having an elevated
`PAA level. The PAA:PAGN ratio also has an important clinical advantage in that it remains
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`comparatively constant over the day and, therefore, is more readily interpretable in a random
`blood draw.
`
`These analyses have several limitations. First, although pharmacokinetic and safety data
`were derived from controlled prospective studies, the analyses of the frequency of the
`specific AEs reported by Thibault et al. [35], [36] were done as post hoc analysis. Second,
`PAA levels were not always available at precisely the time of occurrence of neurological
`AEs, though a conservative approach was taken in these analyses by utilizing the highest
`recorded PAA for that dosing period. Finally, these conclusions pertaining to the absence of
`a statistical relationship between plasma PAA levels and neurological AEs apply at the
`population levels and may not apply to individual UCD or HE patients [38]. Since the
`symptoms reportedly associated with elevated PAA levels are non-specific and similar to
`those associated with elevated ammonia, it is possible that PAA may occasionally cause
`reversible AEs that go clinically unrecognized or are attributed to something else.
`
`Collectively, the present findings indicate that the PAA:PAGN ratio is a useful dosing
`biomarker suitable for use with random blood draws and they suggest further that dose
`reduction may be warranted in patients receiving PAA prodrugs with an elevated plasma
`PAA:PAGN ratio who exhibit neurological adverse events not explained by elevated
`ammonia or intercurrent illness.
`
`Supplementary Material
`
`Refer to Web version on PubMed Central for supplementary material.
`
`Acknowledgments
`
`The authors gratefully acknowledge and thank the efforts of the Study Coordinators and nursing staff who made
`these trials possible, including D. Bartholomew (Nationwide Children's Hospital), S. Cederbaum, D. Wong
`(University of California), J. Vockley (Children's Hospital of Pittsburgh), S. Bart, M. Al-Ibraham (SNBL), M.S.
`Korson (Tufts Medical Center), D. Kronn (Westchester Medical Center), R. Zori (University of Florida), J.L.
`Merritt (Seattle Children's Hospital), N. Schrager (Mount Sinai School of Medicine), A. Donovan, J. Crawford,
`Pediatric TRU Staff, K. Defouw, J. Balliet (The Medical College of Wisconsin), M. Keuth, N. O’Donnell (Long
`Beach Memorial Hospital), M. Hussain, E. Bailey, M. Ambreen (The Hospital for Sick Children, University of
`Toronto, ON, Canada), C. Bailey, A. Lang (The University of Utah), J. Perry, V. de Leon, A. Niemi, K. Cusmano
`(Stanford University),T. Carlson, J. Parker, S. Elsbecker (University of Minnesota), K. Simpson (Children's
`National Medical Center), K. Regis (Nationwide Children's Hospital), A. Behrend, T. Marrone, J. Martin (Oregon
`Health Sciences University), N. Dorrani (University of California, Los Angeles), M.B. Frohnapfel, S. Bergant, J.
`Haky, C. Tasi, C. Heggie (Case Western Reserve University), S. Mortenson (Maine Medical Center), S. Deward
`(Children's Hospital of Pittsburgh), S. Burr (Children's Hospital Colorado ), K. Bart, C. Duggan (SNBL), K.
`Murray, C. Dedomenico (Tufts Medical Center), C. Gross (University of Florida), L. Brody (Seattle Children's
`Hospital), M. Mullins, S. Carter, A. Tran, J. Stuff, TCH General Clinical Research Center nursing staff (Baylor), B.
`McGuire (University of Alabama), D. Wolf (New York Medical College), C. O’Brien (University of Miami), R.
`O'shea (Cleveland Clinic), I. Zupanets (National University of Pharmacy of MH of Ukraine), Kathy Lisam
`(Hyperion), as well as the Clinical and Translational Science Awards/General Clinical Research Center Grants
`(Baylor College of Medicine, M01RR00188; Case Western Reserve University, UL1RR024989; Clinical and
`Translational Science Institute at Children's National Medical Center NIH/NCRR, UL1RR31988; Medical College
`of Wisconsin, UL1RR31973; Mount Sinai School of Medicine, UL1RR29887; Oregon Health & Science
`University, UL1RR24140; Stanford University, UL1RR25744; Tufts University, UL1RR25752; University of
`California, Los Angeles, UL1RR33176; University of Colorado, UL1RR25780; University of Florida,
`UL1RR29890; University of Minnesota, UL1RR33183; University of Pittsburgh, UL1RR24153, UL1TR000005;
`University of Utah, UL1RR25764; University of Washington, UL1RR25014), the Urea Cycle Disorders
`Consortium (NIH Grant U54RR019453) and grants from the O’Malley Foundation and Kettering Fund which
`provided support. SCS. Nagamani is an awardee of the National Urea Cycle Disorders Foundation Research
`Fellowship. The authors also thank G. Enns (Stanford) for his comments on the manuscript.
`
`Mol Genet Metab. Author manuscript; available in PMC 2014 December 01.
`
`NIH-PA Author Manuscript
`
`NIH-PA Author Manuscript
`
`NIH-PA Author Manuscript
`
`Par Pharmaceutical, Inc. Ex. 1039
`Par v. Hyperion, IPR2015-01117
`Page 9 of 21
`
`

`
`Mokhtarani et al.
`
`List of Abbreviations
`
`Page 10
`
`GEE
`
`GPB
`
`HE
`
`NaPBA
`
`PAA
`
`generalized estimating equations
`
`glycerol phenylbutyrate (generic name for glyceryl tri (4-
`phenylbutyrate), also referred to as HPN-100 or RAVICTI®)
`
`hepatic encephalopathy
`
`sodium phenylbutyrate (BUPHENYL®)
`
`phenylacetic acid
`
`PAA:PAGN ratio
`
`ratio of the concentrations in μg/mL of PAA to PAGN in plasma
`
`PAGN
`
`PBA
`
`SE
`
`SO
`
`UCD
`
`REFERENCES
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`phenylbutyric acid
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