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`Application Data Sheet 37 CFR 1.76
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`METHODS OF THERAPEUTIC MONITORING OF NITROGEN SCAVENGING DRUGS
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`Scharschmidt
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`METHODS OF THERAPEUTIC MONITORING OF NITROGEN SCAVENGING
`
`DRUGS
`
`RELATED APPLICATIONS
`
`[0001] The present application is a divisional of U.S. Patent Application No. 13/417,137, filed
`
`March 9, 2012 and now pending, which claims the benefit of U.S. Provisional Application No.
`
`61/564,668, filed November 29, 2011, and U.S. Provisional Application No. 61/542,100, filed
`
`September 30, 2011, the disclosures of which are incorporated by reference herein in their
`
`entirety, including drawings.
`
`BACKGROUND
`
`[0002] Nitrogen retention disorders associated with elevated ammonia levels include urea cycle
`
`disorders (UCDs) and hepatic encephalopathy (HE).
`
`[0003] UCDs include several inherited deficiencies of enzymes or transporters necessary for
`
`the synthesis of urea from ammonia, including enzymes involved in the urea cycle. The urea
`
`cycle is depicted in Figure 1, which also illustrates how certain ammonia-scavenging drugs act to
`
`assist in elimination of excessive ammonia. With reference to Figure l,N—acetyl glutamine
`
`synthetase (NAGS)—derived N-acetylglutamate binds to carbamyl phosphate synthetase (CPS),
`
`which activates CPS and results in the conversion of ammonia and bicarbonate to carbamyl
`
`phosphate. In turn, carbamyl phosphate reacts with ornithine to produce citrulline in a reaction
`
`mediated by ornithine transcarbamylase (OTC). A second molecule of waste nitrogen is
`
`incorporated into the urea cycle in the next reaction, mediated by arginosuccinate synthetase
`
`(ASS), in which citrulline is condensed with aspartic acid to form argininosuccinic acid.
`
`Argininosuccinic acid is cleaved by argininosuccinic lyase (ASL) to produce arginine and
`
`fumarate. In the final reaction of the urea cycle, arginase (ARG) cleaves arginine to produce
`
`ornithine and urea. Of the two atoms of nitrogen incorporated into urea, one originates from free
`
`ammonia (NHf) and the other from aspartate. UCD individuals born with no meaningful
`
`residual urea synthetic capacity typically present in the first few days of life (neonatal
`
`presentation). Individuals with residual function typically present later in childhood or even in
`
`adulthood, and symptoms may be precipitated by increased dietary protein or physiological
`
`stress (e.g., intercurrent illness).
`
`[0004] Hepatic encephalopathy (HE) refers to a spectrum of neurologic signs and symptoms
`
`believed to result from hyperammonemia, which frequently occur in subjects with cirrhosis or
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`certain other types of liver disease. Subjects with HE typically show altered mental status
`
`ranging from subtle changes to coma, features similar to subjects with UCDs.
`
`[0005]
`
`Subjects with nitrogen retention disorders whose ammonia levels and/or symptoms are
`
`not adequately controlled by dietary restriction of protein and/or dietary supplements are
`
`generally treated with nitrogen scavenging agents such as sodium phenylbutyrate (NaPBA,
`
`approved in the United States as BUPHENYL® and in Europe as AMMONAPS®) or sodium
`
`benzoate. These are often referred to as alternate pathway drugs because they provide the body
`
`with an alternate pathway to urea for excretion of waste nitrogen (Brusilow 1980; Brusilow
`
`l99l). NaPBA is a phenylacetic acid (PAA) prodrug. Another nitrogen scavenging drug
`
`currently in development for the treatment of nitrogen retention disorders is glyceryl tri-[4-
`
`phenylbutyrate](HPN—100), which is described in U.S. Patent No. 5,968,979. HPN—l00, which is
`
`commonly referred to as GT4P or glycerol PBA, is a prodrug of PBA and a pre—prodrug of PAA.
`
`[0006] HPN—l00 and NaPBA share the same general mechanism of action: PBA is converted
`
`to PAA via beta oxidation, and PAA is conjugated enzymatically with glutamine to form
`
`phenylacetylglutamine (PAGN), which is excreted in the urine. The structures of PBA, PAA,
`
`and PAGN are set forth below.
`
`,phe:nyib:La'iyra'E22
`
`Phresnyélaeeiée azztld
`
`EPh£~“:r:y§ace§3rl2g§u’§§a:mine
`
`[0007] The clinical benefit of NaPBA and HPN-100 with regard to nitrogen retention disorders
`
`derives from the ability of PAGN to effectively replace urea as a vehicle for waste nitrogen
`
`excretion and/or to reduce the need for urea synthesis (Brusilow l99l; Brusilow 1993). Because
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`each glutamine contains two molecules of nitrogen, the body rids itself of two waste nitrogen
`
`atoms for every molecule of PAGN excreted in the urine. Therefore, two equivalents of nitrogen
`
`are removed for each mole of PAA converted to PAGN. PAGN represents the predominant
`
`terminal metabolite, and one that is stoichiometrically related to waste nitrogen removal, a
`
`measure of efficacy in the case of nitrogen retention states. The difference between HPN-100
`
`and NaPBA with respect to metabolism is that HPN-100 is a triglyceride and requires digestion,
`
`presumably by pancreatic lipases, to release PBA (McGuire 2010).
`
`[0008]
`
`In contrast to NaPBA or HPN-100, sodium benzoate acts when benzoic acid is
`
`combined enzymatically with glycine to form hippuric acid. For each molecule of hippuric acid
`
`excreted in the urine, the body rids itself of one waste nitrogen atom.
`
`[0009] Methods of determining an effective dosage of PAA prodrugs such as NaPBA or HPN-
`
`l00 for a subject in need of treatment for a nitrogen retention disorder are described in
`
`W009/l 134460 and W010/025303. Daily ammonia levels, however, may vary greatly in a
`
`subject. This can lead to overestimation by the physician of the average daily ammonia levels,
`
`which may result in overtreatment. Thus, there is a need in the art for improved methods for
`
`PAA prodrug dose determination and adjustment based on ammonia levels in subjects with
`
`nitrogen retention disorders such as UCDs or HE.
`
`SUMMARY
`
`[0010] Provided herein in certain embodiments are methods for determining whether to
`
`increase a dosage of a nitrogen scavenging drug in a subject with a nitrogen retention disorder by
`
`measuring a fasting blood ammonia level and comparing the fasting blood ammonia level to the
`
`upper limit of normal (ULN) for blood ammonia, where a fasting blood ammonia level that is
`
`greater than halfthe ULN for blood ammonia indicates that the dosage needs to be increased. In
`
`certain embodiments, the nitrogen retention disorder is a UCD or HE. In certain embodiments,
`
`the nitrogen scavenging drug is HPN-100, PBA, NaPBA, sodium benzoate, or any combination
`
`thereof (i.e., any combination of two or more of HPN-100, PBA, NaPBA). In certain
`
`embodiments, the ULN is around 35 umol/L or 59 ug/mL. In certain embodiments, the methods
`
`include an additional step of administering an increased dosage of the nitrogen scavenging drug
`
`if the need exists, and in certain of these embodiments administration of the nitrogen scavenging
`
`drug produces a normal average daily ammonia level in the subject. In certain embodiments
`
`wherein a determination is made to administer an increased dosage of nitrogen scavenging drug
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`and wherein the nitrogen scavenging drug is a PAA prodrug, the methods include an additional
`
`step of measuring urinary PAGN excretion and determining an effective dosage of the PAA
`
`prodrug based on a mean conversion of PAA prodrug to urinary PAGN of 60-75%.
`
`[0011] Provided herein in certain embodiments are methods for determining whether to
`
`administer a nitrogen scavenging drug to a subject with a nitrogen retention disorder by
`
`measuring a fasting blood ammonia level and comparing the fasting blood ammonia level to the
`
`ULN for blood ammonia, where a fasting blood ammonia level that is greater than half the ULN
`
`for blood ammonia indicates that the nitrogen scavenging drug needs to be administered. In
`
`certain embodiments, the nitrogen retention disorder is a UCD or HE. In certain embodiments,
`
`the nitrogen scavenging drug is HPN-100, PBA, NaPBA, sodium benzoate, or any combination
`
`thereof (i.e., any combination of two or more of HPN—100, PBA, NaPBA). In certain
`
`embodiments, the ULN is around 35 umol/L or 59 ug/mL. In certain embodiments, the methods
`
`include an additional step of administering a nitrogen scavenging drug if the need exists, and in
`
`certain of these embodiments administration of the nitrogen scavenging drug produces a normal
`
`average daily ammonia level in the subject. In certain embodiments wherein a determination is
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`made to administer a nitrogen scavenging drug and wherein the nitrogen scavenging drug is a
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`PAA prodrug, the methods further include a step of determining an effective initial dosage of the
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`PAA prodrug by determining a target urinary PAGN output based on a target nitrogen output and
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`calculating an effective initial dosage that results in the target urinary PAGN output based on a
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`mean conversion of PAA prodrug to urinary PAGN of 60-75%. In certain embodiments, the
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`methods include a step of administering the calculated effective initial dosage.
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`[0012] Provided herein in certain embodiments are methods for treating a nitrogen retention
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`disorder in a subject who has previously been administered a nitrogen scavenging drug by
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`measuring a fasting blood ammonia level, comparing the fasting blood ammonia level to the
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`ULN for blood ammonia, and administering an increased dosage of the nitrogen scavenging drug
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`if the fasting ammonia level is greater than half the ULN for blood ammonia. In certain
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`embodiments, administration of an increased dosage of the nitrogen scavenging drug produces a
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`normal average daily ammonia level in the subject. In certain embodiments, the nitrogen
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`retention disorder is a UCD or HE. In certain embodiments, the nitrogen scavenging drug is
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`HPN—lOO, PBA, NaPBA, sodium benzoate, or any combination thereof (ie, any combination of
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`two or more of HPN—100, PBA, NaPBA). In certain embodiments, the ULN is around 35
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`umoL’L or 59 ug/mL. In certain embodiments wherein the nitrogen scavenging drug is a PAA
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`prodrug, the methods include an additional step of measuring urinary PAGN excretion and
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`determining an effective dosage of the PAA prodrug based on a mean conversion of PAA
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`prodrug to urinary PAGN of 60-75%. In certain embodiments, the methods include a step of
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`administering the calculated effective dosage.
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`BRIEF DESCRIPTION OF DRAWINGS
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`[0013] Figure 1: The urea cycle and how certain nitrogen-scavenging drugs may assist in
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`elimination of excessive ammonia.
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`[0014] Figure 2: Relationship between fasting ammonia and average ammonia UCD patients.
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`[0015] Figure 3: Venous blood ammonia values over 24 hours in (A) adult and (B) pediatric
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`UCD patients.
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`DETAILED DESCRIPTION
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`[0016] The following description of the invention is merely intended to illustrate various
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`embodiments of the invention. As such, the specific modifications discussed are not to be
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`construed as limitations on the scope ofthe invention. It will be apparent to one skilled in the art
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`that various equivalents, changes, and modifications may be made without departing from the
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`scope of the invention, and it is understood that such equivalent embodiments are to be included
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`herein.
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`[0017]
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`In subjects with a nitrogen retention disorder, the desired effect of treatment with a
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`nitrogen scavenging drug is control of blood ammonia level. Control of blood ammonia level
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`generally refers to ammonia values within the normal range and avoidance of hyperammonemic
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`crises, which are often defined in the art as transient ammonia values exceeding l00 umol/L or
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`178 ug/mL accompanied by clinical signs and symptoms ofhyperammonemia. Dosing of
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`nitrogen scavenging drugs is usually based upon clinical assessment and measurement of
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`ammonia. However, assessment of treatment effect and interpretation of ammonia levels is
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`confounded by the fact that individual ammonia values vary several—fold over the course of a day
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`and are impacted by timing of the blood draw in relation to the last meal and dose of drug (see,
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`eg, Lee 2010; Lichter—Konecl:i 2011; Diaz 2011).
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`[0018] A random ammonia value obtained during an outpatient visit may fail to provide a
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`reliable measure of a subject's status and the drug effect. For example, basing treatment on a
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`blood sample taken after eating a meal might overestimate average daily ammonia level and
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`result in overtreatment. Conversely, basing treatment on a blood sample taken after drug
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`administration might underestimate average daily ammonia level and result in undertreatment. A
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`fasting ammonia level at or near the ULN might be taken as an indication of satisfactory control
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`without appreciating the fact that the ammonia burden during the day (average and/or highest
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`possible value) might be significantly higher. Thus, a fasting level at or near the ULN may
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`actually reflect undertreatment in a subject already a receiving nitrogen scavenging drug or the
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`need for treatment in a subject not currently prescribed a nitrogen scavenging drug. A more
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`accurate view of daily ammonia level could be obtained by multiple blood draws in a controlled
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`setting over an extended period of time. Although this is currently done in clinical trials, it is
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`clinically impractical.
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`[0019] As set forth below, the relationship between fasting ammonia levels and daily ammonia
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`exposure was evaluated in subjects with nitrogen retention disorders. It was found that fasting
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`ammonia correlates strongly with daily ammonia exposure, assessed as a 24 hour area under the
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`curve for ammonia, daily average, or maximal daily concentration, and that a target fasting value
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`which does not exceed half ofthe ULN is a clinically useful and practical predictor of ammonia
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`values over 24 hours. As such, provided herein are clinically practical methods of evaluating
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`ammonia exposure in subjects with nitrogen retention disorders based on fasting ammonia levels,
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`as well as methods of using the resultant information to adjust the dosage of a nitrogen
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`scavenging drug, determine whether to administer a nitrogen scavenging drug, treat a nitrogen
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`retention disorder, and predict daily ammonia burden. The use of fasting ammonia levels to
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`predict ammonia exposure provides a significant advantage over previously developed methods
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`by reducing the number of required blood draws and eliminating the confusion associated with
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`conflicting ammonia levels over the course ofthe day.
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`[0020] As further disclosed herein, the relationship between ammonia control and
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`neurocognitive outcome was evaluated in UCD patients. Previous research has demonstrated
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`that UCD patients often exhibit lower IQ overall and deficient executive function manifested by
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`difficulty in goal setting, planning, monitoring progress and purposeful problem solving. As set
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`forth herein, it was found that ammonia control with GPB resulted in a significant improvement
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`in executive functions in pediatric patients. Based on these results, methods are provided herein
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`for improving executive function in a pediatric subject with a UCD by administering one or more
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`nitrogen scavenging drugs.
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`[0021] As further disclosed herein, the relationship between elevated PAA levels and
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`neurological adverse events (AEs) was analyzed. Many of the over 30 reports of administration
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`of NaPBA and/or sodium PAA to humans describe AEs, particularly when administered
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`intravenously. IV administration of PAA to cancer patients was shown previously to result in
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`AEs that included fatigue, dizziness, dysgeusia, headache, somnolence, lightheadedness, pedal
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`edema, nausea, vomiting, and rash (Thibault 1994; Thibault 1995). These AEs correlated with
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`PAA leve