Biomarkers of CYP2D6 and CYP3A4 Variability in Pediatrics
Condition: Drug Metabolism Phenotype
Study type: Observational

Summary: 
      Cytochrome P450 2D6 (CYP2D6) is an important enzyme in the body for breaking down many
      medications that are commonly used in children of various ages. The purpose of this proposal
      is to investigate the relative roles of development and genetic variation in CYP2D6 activity
      in school-aged children and adolescents with attention deficit and hyperactivity disorder and
      health controls using the over-the-counter cough suppressant, dextromethorphan or "DM", a
      standard probe for determining CYP2D6 phenotype. Embedded in the study design are sub-studies
      to search for by-products of normal body metabolism that reflect differences in enzyme
      activity, and a pharmacokinetic study to assess the consequences of CYP2D6 genetic variation
      on the systemic exposure to medications used by this patient population. Ultimately, the goal
      of the research is to personalize the use of medications in children by selecting the
      appropriate dose of the correct medication for individual patients.
    

Detailed description: 
      This proposal represents a continuation of previous and ongoing longitudinal studies to
      characterize the relative contribution of pharmacogenetics and ontogeny to variability in
      CYP2D6 and CYP3A4 activity between birth and adolescence. Specifically, the overall goal of
      the research program is to test the hypothesis that genetic variability, rather than
      ontogeny, is the primary source of variability in CYP2D6 and CYP3A4 activity during
      adolescence. This study will extend our current series of longitudinal phenotyping studies
      and will be a cross-sectional study in children aged 7-15 years, studied on seven occasions,
      6 months apart.

      Children will be recruited from Children's Mercy Hospitals and Clinics, The Children's Mercy
      Hospital Northland Campus, and The Children's Mercy South Campus.

      Subjects participating in this study will consist of a total of 220 children and adolescents
      from 7 to 15 years of age with a gender and ethnic distribution representative of the patient
      population served by Children's Mercy Hospital (CMH; from a representative 12 month period)
      as follows: White, 59.6%; African-American, 31.9%; Hispanic, 4.9%; Asian, 0.6%;Native
      American, 0.1%; Other, 2.9%. The gender distribution is 44.4% female and 55.6% male. An equal
      number of pediatric patients who meet the DSM-IV criteria for a primary diagnosis of ADHD or
      ADD (~33%) and age- and sex-matched controls (~67%) will be recruited. The patients represent
      a population of children who can reasonably expect to be treated with a medication that is
      primarily dependent upon CYP2D6 for its elimination from the body. An example is atomoxetine,
      also known as Strattera®. Each patient will be asked to invite a friend to participate in the
      phenotyping study with them, providing a non-ADHD population as a reference. Additional
      phenotyping visits will occur every six months (totaling 7 visits). Justification: Assuming a
      10% attrition rate for the second visit, the sample size is expected to provide approximately
      25 patients and 25 controls (50 subjects total) in each initial 2-year age bin: 7.0 to 9.0 y,
      9.1 to 11.0 y, 11.1 to 13.0 y, and 13.1 to 15.0 y.

      Subjects will be enrolled via informed permission/assent and will be studied as outpatients
      utilizing resources provided by the Pediatric Clinical Research Unit located in the Division
      of Clinical Pharmacology and Medical Toxicology at Children's Mercy Hospital. For the ADHD
      group, patients being treated with atomoxetine at the time of study enrollment will be
      allowed to participate. Although in vitro studies indicate that atomoxetine inhibits CYP2D6
      activity by ~50%, there does not appear to be any effect in vivo as single dose
      pharmacokinetics of the CYP2D6 substrate, desipramine, were unaffected by four days of
      atomoxetine 60 mg b.i.d. Thus, we do not expect any substantial effect on DM
      biotransformation. Furthermore, if during the six month interval between phenotyping visits
      ADHD patients are prescribed medications that are known CYP2D6 inhibitors, they will be
      allowed to continue in the study; the dose and start date of these medications will be
      recorded and the effect of the inhibitor on CYP2D6 phenotype (urinary metabolite ratio and
      endogenous biomarker) will be determined. These data points will be excluded from the
      ontogeny and genotype-phenotype correlation analysis

      Pre-study visit. (about 1 hour long) On a screening visit, prospective subjects (and their
      parents) will be provided with a description of the proposed study. After all their questions
      have been answered, they will be given a copy of the permission/assent form to review and
      sign Subjects will then have their medical history and use of medications reviewed, including
      the use of non-prescription and herbal remedies. All subjects will be given a complete
      physical examination by a physician, including vital signs (blood pressure, heart rate,
      breathing rate, temperature, height and weight) and an assessment of pubertal development by
      Tanner stage. Breast development will be assessed by visual inspection. Pubic hair will be
      assessed by visual inspection. Testicular volume will be measured by direct comparison to
      orchidometer beads. Tanner Staging will be assessed by a gender appropriate physician. Blood
      will be drawn by needle stick for serum chemistries, liver function tests and a hematology
      panel; a portion will also be retained for DNA testing (approximately 2 teaspoons of blood
      total). Urine will be collected for a Urinalysis with Micro. Numbing cream may be applied to
      the site to minimize discomfort. A urine pregnancy test will be performed for females of
      child bearing age.

      The DNA testing, referred to "genotyping" will be conducted by a combination of technologies
      designed to detect variations from single nucleotide level (including polymorphisms,
      insertions and deletions), to larger gene deletions, rearrangements, hybrid structures and
      copy number variations using established methods in the Developmental Pharmacology and
      Experimental Therapeutics Laboratory. As next-generation sequencing technology becomes
      established, existing technologies will be replaced by next-generation sequencing for its
      ability to assess the entire gene region rather than at individual locations.

      Study Day 1 (about 5 hours long) Within approximately one week of the pre-study visit,
      subjects will be admitted to the study unit. The child must refrain from eating 2 hours prior
      to the scheduled visit. Prior to dosing, the subjects will completely empty their bladders
      and provide a blank urine sample. Each subject will then be given a single 0.5 mg/kg dose of
      DM as Robitussin Cough (7.5 mg/5 ml; alcohol-free, fruit punch flavor). All urine produced
      over the 4-hour study period will be collected. Urine will be retained for analysis of DM and
      its three metabolites, and for metabolomic analyses. One hour after dosing, the subjects will
      be provided with a small meal that they will select from the standard hospital menu. A
      pregnancy test will be performed for females of child bearing age. Subjects will be asked to
      provide breaths (inspiratory and expiratory) for ten minutes with a handheld indirect
      calorimeter (ie. MedGen/Body Gem: also known as "GEM") with a clip on their nose. The
      measurements obtained for resting metabolic rates will occur prior to dosing and 4 hours post
      dose.

      Subjects will be asked how they are feeling during the study and before going home.

      Study Day 2-7 (about 5 hours long) The remaining visits will occur every 6 months following
      Study Day 1; the study sessions will be delayed for 1-2 weeks if the subjects are suffering
      from flu-like symptoms. The child must refrain from eating 2 hours prior to the scheduled
      visit. For females, a pregnancy test will be conducted using the pre-dose urine sample prior
      to DM dosing.

      Subjects will then have their medical history and use of medications reviewed, including the
      use of non-prescription and herbal remedies. Subjects will be given a complete physical
      examination by a physician, including vital signs (blood pressure, heart rate, breathing
      rate, temperature, height and weight) and an assessment of pubertal development by Tanner
      stage. Breast development will be assessed by visual inspection. Pubic hair will be assessed
      by visual inspection. Testicular volume will be measured by direct comparison to orchidometer
      beads. Tanner Staging will be assessed by a gender appropriate physician.

      Subjects will be admitted to the study unit. Prior to dosing, the subjects will completely
      empty their bladders and provide a blank urine sample. Each subject will then be given a
      single 0.5 mg/kg dose of DM as Robitussin Cough (7.5 mg/5 ml; alcohol-free, fruit punch
      flavor). All urine produced over the 4-hour study period will be collected. Urine will be
      retained for analysis of DM and its three metabolites, and for metabolomic analyses. One hour
      after dosing, the subjects will be provided with a small meal that they will select from the
      standard hospital menu. A pregnancy test will be performed for females of child bearing age.
      Subjects will be asked to provide breaths (inspiratory and expiratory) for ten minutes with a
      handheld indirect calorimeter (ie. MedGen/Body Gem: also known as "GEM" with a clip on their
      nose. The measurements obtained for resting metabolic rates will occur prior to dosing.
      Subjects will be asked how they are feeling during the study and before going home.
    

Observational model: Case-Control
Time perspective: Prospective
Sampling method: Non-Probability Sample
Study population: 
        Cases: Pediatric patients who have a primary diagnosis of ADHD, combined type, hyperactive
        impulsive, or inattentive type (ADD). (110 subjects)

        Controls: Age and gender matched subjects who do not meet any of the exclusion criteria
        (110 subjects)
      
Biospecimen description: 
      Blood or saliva for DNA testing. Blood samples will be obtained for safety labs throughout
      the study.
    
Keywords: CYP2D6; CYP3A4; cytochrome P450; children; pediatrics; dextromethorphan; phenotype; genotype