Drs. Calarge and DiSilvestro and Ms. Farmer report no competing interests. Dr.

Arnold has received research funding from Celgene, Curemark, Lilly, Neuropharm, Novartis, Noven, Shire, Sigma Tau, and Targacept, the National Institute of Health, and Autism Speaks; has consulted to Abbott, Neuropharm, Novartis, Noven, Organon, Shire, and Sigma Tau; and has been speaker for Abbott, Shire, McNeil, Novartis and Targacept.

Iron Stores, ADHD Severity, and Stimulant Optimal Dose

Chadi Calarge, M.D.1; Cristan Farmer, M.A.2, Robert DiSilvestro, Ph.D.2, L. Eugene Arnold, M.D.2;
1

The University of Iowa and 2 Ohio State University

Table 3: Anthropometric and Dietary Data In the Total Sample and across Medication-Nave and Formerly-Medicated Participants Total Sample Med-Nave Past-Med p Weight z score 0.2 (1.0) 0.4 (1.0) 0.0 (1.1) >.1 Height z score 0.0 (1.0) 0.1 (0.9) -0.1 (1.0) >.3 BMI z score 0.3 (1.1) 0.5 (1.0) 0.0 (1.1) >.1 Dietary Intake: Total Calories, kcal/d Protein, g/d Fat, g/d Carbohydrates, g/d Iron, g/d 1721 (609) 61.9 (21.5) 63.3 (22.2) 231.4 (89.3) 12.2 (4.4) 1746 (607) 62.8 (21.8) 63.2 (22.5) 236.9 (87.6) 12.7 (4.5) 1690 (625) 60.8 (21.6) 63.5 (22.4) 224.3 (93.2) 11.5 (4.3) >.7 >.7 >.9 >.6 >.3

Introduction
Background: Iron deficiency in rats results in reduced density in the basal ganglia of the dopamine transporter as well as the D1 and D2 dopamine receptors (1). Moreover, the response to cocaine, a potent inhibitor of the dopamine transporter, is attenuated in iron-deficient rats (1). In addition, prospective studies have shown that infants with iron deficiency anemia exhibit impairment in socio-emotional and academic functioning (2). Finally, studies in patients with ADHD have reported an inverse association between serum ferritin concentration and the severity of inattention, hyperactivity/impulsivity, and sleep disturbances (3,4). Objective: Thus, we here attempt a replication of serum ferritin association with ADHD symptom severity and explore a possible effect of ferritin concentration on response to stimulant treatment (5).

The average rating per item ( sd) on the SNAP-IV and the mean scale scores on the CPRS-R at baseline and at the end of each treatment phase are reported in Table 1.
Table1: ADHD Symptom Severity Ratings Baseline N=52 SNAP-IV Inattention Hyperactivity/Impulsivity Total CPRS-R ADHD Scale Cognitive Scale Hyperactivity Scale ODD Scale 2.3 0.5 1.9 0.7 2.1 0.5 26.6 5.6 25.5 5.7 14.0 7.1 12.2 6.8 Week 8 N=51 2.0 0.6 1.7 0.8 1.8 0.6 24.2 6.7 23.1 7.0 12.3 6.8 11.3 6.3 Week 10 N=49 1.6 0.6 1.4 0.6 1.5 0.5 16.9 7.0 16.5 8.2 8.9 6.1 8.9 6.3 Week 13 N=49 1.2 0.6 0.9 0.6 1.0 0.5 12.9 6.7 12.6 6.5 5.7 4.9 6.7 5.2

The mean dose of amphetamine by week 13 was 13.4 mg (sd=6.4) with the mean weight-adjusted dose being 0.37 mg/kg (sd=0.13). After partialing out the effect of age and sex, both baseline as well as week 8 serum ferritin concentrations were inversely associated with the weight-adjusted dose of amphetamine at week 13 (Partial Spearmans r= -0.45, p<0.007 (Figure 3) and r= -0.48, p=0.003, respectively). These associations were not different between psychotropic-nave and previously-medicated participants.
Weight-adjusted daily dose of amphetamine (mg/kg/d)

Methods
Participants: This analysis is based on data collected in a multiphase double-blind placebo-controlled pilot trial investigating the efficacy of zinc supplementation in treating ADHD and optimizing response to stimulant treatment (5).

Serum ferritin concentration at baseline and week 8 were strongly correlated (Pearsons r=0.65, p<.0001) and changed little during the course of the study (p>.3) (Table 2).
Table 2: Serum Ferritin Concentration in the Total Sample and across Medication-Nave and Formerly-Medicated Participants Total Sample Med-Nave Past_Med Serum Ferritin, mean (sd) Week 0, ng/ml 18.4 (11.4) 18.7 (12.1) 18.0 (10.6) Week 8, ng/ml 17.1 (11.1) 19.2 (12.6) 14.0 (7.8) Low Baseline Ferritin, n (%) < 30 ng/ml Cutoff < 7 ng/ml Cutoff 45 (87) 12 (23) 26 (87) 7 (23) 19 (86) 5 (23)

Study Design
WK4 WK8

p > .8 > .09 1.0 1.0

0.75

Partial Spearmans r= -0.45, p<0.007

0.6 0.45 0.3 0.15 0 0 10 20 30 40 50 Serum Ferritin Concentration (ng/ml)

Zin

c
WK1 WK10 WK13

Plac eb

o
Monotherapy

Standard Amphetamine Dose

Zinc vs. PBO randomized throughout

Amphetamine Dose Optimization

Due to missing data, the samples consisted of 27 med-nave and 19 past-med participants at baseline and of 26 and 18, respectively, at week 8.

The study enrolled healthy 6 to 14 year-old unmedicated children with a primary diagnosis of ADHD (combined or inattentive subtype) and without other serious psychiatric or medical conditions requiring treatment. The diagnosis of ADHD was based on the Childrens Interview for Psychiatric Symptoms; parent version (P-ChIPS) and clinical DSM-IV-based assessment. The parents completed the Swanson, Nolan, and Pelham, Version IV (SNAP-IV), and the Conners Parent Rating Scale-Revised (CPRS-R), long version. At baseline and week 8, serum ferritin was measured using the Ferritin EIA kit from Alpco Diagnostics (Salem, NH, USA) . The study was approved by the local IRB and written consents and assents were obtained. Statistical Analysis: The sample was divided based on prior treatment with psychotropics. Differences across the two groups were compared using a Student t-test for continuous variables and Fishers Exact test for categorical ones. Partial (adjusting for age and sex) Pearsons correlations were computed. Multiple linear regression analysis was used to investigate whether baseline ferritin was associated with ADHD symptom severity after adjusting for age, sex, and prior psychotropic treatment status, and whether it predicted stimulant dose at week 13, the end of dose optimization.

Because there were no significant differences between the two intervention groups and because zinc supplementation had a negligible effect on ADHD symptoms and no effect on ferritin concentration, all participants were pooled for the following analyses. At baseline, after partialing out the effect of age and sex, serum ferritin concentration was inversely correlated with the SNAP-rated inattention, hyperactivity/impulsivity, and total ADHD symptom scores (Partial Spearmans r= -0.31, p=0.04; r= -0.42, p<0.006; and r= -0.43, p<0.004, respectively ). However, this effect was different depending on the status of prior treatment with psychotropics (Figures 1 & 2). This finding is not accounted for by a difference in nutritional status (Table 3).
3

Figure 3: Association between baseline ferritin and weight-adjusted dose of dextro-amphetamine needed to optimize clinical response.

Conclusions
1. Our findings are consistent with a growing body of evidence linking iron stores to ADHD symptom severity. 2. This association, however, seems more prominent in patients who have already received ADHD treatment, though dietary and anthropometric factors do not fully account for the difference. 3. Iron status might predict sensitivity to psychostimulants. 4. Future studies should explore whether iron supplementation improves response to pharmacotherapy in ADHD.
References:
1. 2. 3. 4. Erikson KM, Jones BC, Hess EJ, Zhang Q and Beard JL. Iron deficiency decreases dopamine D1 and D2 receptors in rat brain. Pharmacol Biochem Behav 69:409-418, 2001. Lozoff B, Jimenez E, Hagen J, Mollen E and Wolf AW. Poorer behavioral and developmental outcome more than 10 years after treatment for iron deficiency in infancy. Pediatrics 105:E51, 2000. Konofal E, Lecendreux M, Arnulf I and Mouren MC. Iron deficiency in children with attentiondeficit/hyperactivity disorder. Arch Pediatr Adolesc Med 158:1113-1115, 2004. Cortese S, Konofal E, Bernardina BD, Mouren MC and Lecendreux M. Sleep disturbances and serum ferritin levels in children with attention-deficit/hyperactivity disorder. Eur Child Adolesc Psychiatry 18:393-399, 2009. Arnold, L.E., DiSilvestro RA, Bozzolo D, Bozzolo H, Crowl L, Fernandez S, Ramadan Y, Thompson S, Mo X, Abdel-Rasoul M, Joseph E. Zinc for Attention-Deficit/Hyperactivity Disorder: Placebo-Controlled Double-Blind Pilot Trial Alone and Combined with Amphetamine. Journal of Child and Adolescent Psychopharmacology, In Press, 2010.

SNAP-Based Hyperactivity/Impulsivity Score

Partial Spearmans r= -0.02, p>0.9 Med-Naive

SNAP-Based Inattention Score

2.6

Partial Spearmans r= -0.38, p<0.07

2.6

Med-Naive
2.2

2.2

1.8

1.8

Past-Med
1.4

1.4

1 0

Partial Spearmans r= -0.73, p<0.001

5 10 15 20 25 30 35 40 45 50 Baseline Serum Ferritin Concentration (ng/ml)

Past-Med Partial Spearmans r= -0.44, p<0.08

0 10 20 30 40 50

1 Baseline Serum Ferritin Concentration (ng/ml)

Results
Fifty two children (83% males, n=43) were randomized: 24 to placebo and 28 to zinc glycinate. Their mean age was 10.0 years (sd=2.6). Most (73%, n=38) had ADHD, combined type.

5.

Figures 1 & 2: Association of baseline ferritin with inattention and hyperactivity/ impulsivity across medication-nave and formerly-medicated participants. All participants were unmedicated for at least one week prior to randomization.

Acknowledgements: This work was funded by National Institute of Mental Health (5R34MH071683 and K23MH085005) and the National Center for Research Resources (UL1RR025755). Contact: chadi-calarge@uiowa.edu