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brain research 1580 (2014) 188–198

Available online at www.sciencedirect.com

www.elsevier.com/locate/brainres

Research Report

Intranasal oxytocin in the treatment of autism


spectrum disorders: A review of literature
and early safety and efficacy data in youth

Evdokia Anagnostoua,n, Latha Sooryab, Jessica Briana, Annie Dupuisc,


Deepali Mankada, Sharon Smilea, Suma Jacobd
a
Bloorview Research Institute, University of Toronto, 150 Kilgour Road, Toronto, ON, Canada M4G 1R8
b
Rush University Medical Center, Department of Psychiatry, 2150 West Harrison Street, Chicago, IL 606103, USA
c
The Hospital for Sick Children, Dalla Lana School of Public Health, University of Toronto, 555 University Avenue,
Toronto, Canada M5G 1X8
d
University of Minnesota, Department of Psychiatry & Pediatrics, 2101 6th Street SE, Minneapolis, MN 55455, USA

ar t ic l e in f o abs tra ct

Article history: Background: There is a paucity of treatments targeting core symptom domains in Autism
Accepted 28 January 2014 Spectrum Disorder (ASD). Several animal models and research in typically developing
Available online 5 February 2014 volunteers suggests that manipulation of the oxytocin system may have therapeutic
Keywords: potential for the treatment of social deficits. We review the literature for oxytocin and ASD
Autism and report on early dosing, safety and efficacy data of multi-dose oxytocin on aspects of
Oxytocin social cognition/function, as well as repetitive behaviors and co-occurring anxiety
Clinical trial within ASD. Methods: Fifteen children and adolescents with verbal IQs Z70 were diagnosed
Social cognition with ASD using the ADOS and the ADI-R. They participated in a modified maximum
Social function tolerated dose study of intranasal oxytocin (Syntocinon). Data were modeled using
repeated measures regression analysis controlling for week, dose, age, and sex. Results:
Among 4 doses tested, the highest dose evaluated, 0.4 IU/kg/dose, was found to be well
tolerated. No serious or severe adverse events were reported and adverse events reported/
observed were mild to moderate. Over 12 weeks of treatment, several measures of social
cognition/function, repetitive behaviors and anxiety showed sensitivity to change with
some measures suggesting maintenance of effect 3 months past discontinuation of
intranasal oxytocin. Conclusions: This pilot study suggests that daily administration of
intranasal oxytocin at 0.4 IU/kg/dose in children and adolescents with ASD is safe and has
therapeutic potential. Larger studies are warranted.
This article is part of a Special Issue entitled Oxytocin and Social Behav.
& 2014 Elsevier B.V. All rights reserved.

n
Corresponding author.
E-mail address: eanagnostou@hollandbloorview.ca (E. Anagnostou).

http://dx.doi.org/10.1016/j.brainres.2014.01.049
0006-8993 & 2014 Elsevier B.V. All rights reserved.
brain research 1580 (2014) 188–198 189

neuronal firing (Ludwig and Leng, 2006). The mechanisms


1. Introduction by which central and peripheral release of oxytocin is
coordinated remain poorly explained.
Autism Spectrum Disorder (ASD) refers to a group of neuro- Central release of oxytocin and its closely related peptide,
developmental disorders characterized by impairments in Arginine Vasopressin (AVP), are involved in aspects of social
social communication, and repetitive behaviors and cognition and function including social recognition, social
restricted interests (http://www.DSM5.org). The rates of ASD memory, affiliative behaviors, mother–infant and male–
are on the rise with recent CDC numbers estimating the female pair-bond formation, separation distress, and other
prevalence of the disorder at 1 in 88 children (CDC, 2008). aspects of social attachment, as well as the regulation of
The social communication construct as it pertains to the stress response (Meyer-Lindenberg et al., 2011). Of note,
ASD diagnosis has been debated extensively, even within oxytocin and AVP differ only by two amino acids, share
versions of the DSM classification criteria, but there is agree- evolutionary history, have overlapping functions, influence
ment that it includes difficulties in social-emotional recipro- each other's pathways or receptors throughout development
city, including difficulties in forming or maintaining peer (Hirasawa et al., 2003; Landgraf and Neumann, 2004;
relationships, deficits in nonverbal behaviors and in play Ragnauth et al., 2004) and as such they should be considered
skills. Restricted interests and repetitive behaviors are con- together in biobehavioral contexts.
ceptualized to include repetitive motor behaviors such as arm In animal models, oxytocin has been shown to play
flapping, rocking, bouncing, and spinning behaviors, as well critical roles in social processing, recognition, and bonding,
as higher-level compulsive-like behaviors including rigid and also to influence stereotyped behaviors such as exag-
routines, ritualistic behaviors, and restrictive interests. gerated grooming (Carter, 1998; Insel et al., 1999; Winslow
Despite the high burden of this disorder, there is no et al., 2003). In mammals, the Oxytocin Receptor (OXTR) is
medication to date approved anywhere in the world for the expressed at higher levels in early development (Shapiro
treatment of social deficits or repetitive behaviors associated Shapiro and Insel, 1989; Tribollet et al., 1989). Oxytocin
with ASD. A paucity of molecular targets available for the knockout-mice have been shown to maintain olfaction and
development of novel therapeutics related to the pathophy- cognitive performance, but suffer deficits in social recogni-
siology of ASD has been postulated to contribute to this gap. tion that were recovered by intraventricular oxytocin (OXT),
Specifically, drugs that were developed for other disorders although not by AVP administration (Ferguson et al., 2000).
with potentially overlapping phenotypes were tested in ASD, OXTR knockout mice emit fewer ultrasonic vocalizations
assuming that phenotypic spectra would be associated with compared to the wild type, in response to social isolation,
neurobiologic spectra. The approach has been useful for the experience deficits in social discrimination, and demon-
treatment of associated symptoms of ASD (e.g. atypical strate more aggressive behavior (Takayanagi et al., 2005).
antipsychotics and irritability associated with ASD, Similarly, AVPR1A knock-out mice have been reported to
McCracken et al., 2002), but has not produced a single agent exhibit social memory deficits (Bielsky and Young, 2004),
shown to be effective for core symptoms. With the explosion and the expression of the receptor gene in the lateral
of findings related to the biology of ASD and the biology of septum enhances social recognition (Bielsky et al., 2005).
underlying core symptom domains associated with ASD, The pattern of AVPR1A receptor expression in the brain
there is an opportunity for translational research to facilitate appears to be determined by variation in the length of a
the development of novel therapeutics. microsatellite in the promoter region of the gene (Hammock
Oxytocin is a nine-amino-acid peptide (nonapeptide), and Young, 2005).
which is synthesized primarily in the paraventricular and
supraoptic nucleus of the hypothalamus, and released into
the bloodstream by axon terminals in the posterior pituitary.
Peripheral release of oxytocin facilitates uterine contractions 1.1. Human studies of neuropeptide hormones
during labor and milk letdown. Other peripheral targets of
oxytocin include the kidneys and the pancreas. In addition to An explosion of studies has examined the effect of adminis-
its peripheral role as a hormone, oxytocin is also widely tering a single dose on OXT and social cognition in humans.
distributed throughout the Central Nervous System (CNS) and This work has been reviewed elsewhere extensively (e.g.
functions as a neuromodulator. For example, oxytocin is Macdonald and Feifel, 2013; Guastella and MacLeod, 2012;
released within the bed nucleus of the stria terminalis, the McCall and Singer, 2012; Kumsta and Heinrichs, 2013). Single
spinal cord, the anterior commissural nucleus, and the dose studies in healthy volunteers have reported on
medial amygdala. Oxytocin fibers are evident in a variety of increased trust (Kosfeld et al., 2005), empathic accuracy
brain regions thought to be involved in social perception and (Domes et al., 2006, Guastella et al., 2009), time spent looking
cognition, as well as emotion regulation, including the amyg- at eyes (Guastella et al., 2008), and face identity recognition
dala and hippocampus and the ventral tegmental area of the memory (Savaskan et al., 2008; Rimelle et al., 2009). Attenua-
midbrain (Gordon et al., 2011). Oxytocin receptors are also tion of amydgala activity has been documented with single
widely distributed in the CNS, although their distribution is dose of OXT vs. placebo (Kirsch et al., 2005; Zink and Meyer-
highly species specific (e.g. Donaldson and Young, 2008). Lindenberg, 2012; Domes et al., 2007). Such imaging studies
Oxytocin tract studies, at least in voles, would suggest that suggest that manipulation of the oxytocin system may
oxytocin release is not limited to the synaptic cleft, that produce circuitry modification that is relevant to social
dendritic release occurs and that it is independent of deficits.
190 brain research 1580 (2014) 188–198

1.2. Oxytocin/Vasopressin and ASD increased trust, and time spent looking at eyes. However,
single dose studies have limited ability to predict the ther-
A number of researchers have hypothesized that OXT may be apeutic potential of administering intranasal oxytocin over a
connected to autism given that repetitive behaviors and period of time and as such multi-dose studies are critical in
deficits in social interaction are core features of the disorder, evaluating the compound's long-term therapeutic potential
and that this neuropeptide is involved in the regulation of (Macdonald and Feifel, 2013).
social cognition and some repetitive behaviors. Abnormalities
in the neural pathway for OXT could account for many 1.2.4. Early multi-dose studies of intranasal oxytocin in ASD
features of autism including the early onset, predominance A small number of studies designed to evaluate the effect of
in males, genetic loading, and neuroanatomical abnormal- multi-dose intranasal oxytocin on core symptom domains in
ities (Insel et al., 1999; Domes et al., 2007). ASD is available. In a single case report, Kosaka et al. (2012),
reported on a 16 year old girl with ASD who received 8IU
1.2.1. Oxytocin plasma levels and ASD every day for 2 months. The authors reported that it was well
Individuals with ASD have been reported to have lower than tolerated and that improvements were noted both in social
average levels of blood OXT level in comparison to typically communication as well as irritability, based on clinician
developing controls matched for age (Modahl et al., 1998; judgment and parent report on a standardized scale.
Andari et al., 2010), although not universally, depending on Tachibana et al. (2013) reported on a case series of 8 male
gender differences and assay methodology utilized (Miller youth (ages 10–14 years) who received a total of 6 months
et al., 2013). Higher levels of oxytocin precursor peptides have exposure of oxytocin in the following manner: for the first 2
also been reported to be expressed in early ASD development months they received 8 IU per dose twice a day followed by 2
with subsequent decrease with age (Green et al., 2001). months of 16 IU per dose twice a day, and finally 2 months of
24 IU per dose. Before each step, a 1–2 week period of placebo
1.2.2. Oxytocin and vasopressin receptor genes (OXTR and was administered. Improvements were noted in social and
AVPR1A) and ASD communication scores based on direct observation on a
Accumulating studies are reporting that single nucleotide structured assessment, but not on parental reports of mala-
polymorphisms of the OXTR gene are associated with ASD daptive behaviors. In addition, Dadds et al. (2013) evaluated a
and related disorders (Wu et al., 2005; Jacob et al., 2007; 5-day intervention with 38 male youths with ASD (ages 7–16
Lerer et al., 2008; Yrigollen et al., 2008; Ebstein et al., 2012; Liu years). Boys were administered oxytocin (12 or 24 IU units
et al., 2010; Wermter et al., 2010). However, most SNPs depending on weight) or placebo during parent-child inter-
reported are outside the protein coding regions and their action training. No improvements were noted in emotion
functional significance remains unknown. There has been a recognition, social skills and other behavioral domains com-
recent report of a rare genetic variation of the OXTR gene pared to placebo, based on parent, clinician, or direct obser-
within the protein coding region associated with ASD (Ma vation measures. Although the model of combining oxytocin
et al., 2013). A heterozygous deletion of the OXTR gene has with a social learning activity is of great interest, this study is
also been reported in a patient with ASD and family history of limited by several factors including the use of an experi-
OCD (Gregory et al., 2009). In addition to variations in coding mental psychosocial intervention with unknown effects as a
sequence, there is some early data to suggest potential ‘monotherapy’, as well as limited exposure (e.g. 5 days) to
epigenetic modification related to the OXTR gene in ASD. treatments targeting core skills deficits. Our group also
Gregory et al. (2009) reported increased methylation of the recently published a pilot randomized trial of intranasal
OXTR gene promoter as compared to controls in two inde- oxytocin vs. placebo in 19 adults with ASD (Anagnostou
pendent samples, including postmortem temporal cortex et al., 2012). Adults with ASD demonstrated improvements
tissue from 8 ASD/control pairs. in empathic accuracy, lower order repetitive behaviors and
quality of life on both self-report and experimental testing.
1.2.3. Single dose OXT studies in ASD
There have been a growing number of OXT single dose 1.2.5. Rationale for intranasal administration
studies in the last decade. Initially, an intravenous adminis- Oxytocin is metabolized in the gut by chymotrypsin and as
tration of oxytocin vs. placebo over a 4 h period (Hollander such it cannot be administered orally. Despite its short half-
et al., 2003, 2007) facilitated the retention of social cognition life in the blood, the intravenous formulation has been found
in participants with ASD and produced significant reduction to produce behavioral effects (Hollander et al., 2003, 2007;
in repetitive behaviors – i.e., needing to know, repeating, Ring et al., 2006), but it is too invasive to administer. One
ordering, needing to tell/ask, self-injury, and touching. Sub- alternative is intranasal oxytocin; it is thought to be absorbed
sequent single dose studies have employed the intranasal through the highly permeable nasal mucosa and, in the case
formulation. Guastella et al. (2010) randomized 16 adoles- of the related peptide vasopressin, it has been shown to cross
cents to a cross-over placebo-controlled study of a single dose the blood brain barrier (Born et al., 2002), and produce rising
of intranasal oxytocin and reported significant improvements CNS levels for at least 4 h after intranasal administration,
in empathic accuracy as measured by the Reading-the-Mind- mitigating concerns about the peripheral half-life of this
in-the-Eyes task (RMET) with minimal adverse effects. Andari compound. It is also easy to self-administer.
et al. (2010) randomized 13 adults with ASD to a single dose of Given that oxytocin is involved in the regulation of social
intranasal oxytocin and reported stronger interactions with a communication and some repetitive behaviors, and based on
cooperative partner during a computerized ball game, emerging pilot data, the authors received funding by the
brain research 1580 (2014) 188–198 191

Department of Defense to conduct a series of studies of last dose (week 12) and was lost to follow-up for week 16
intranasal oxytocin in children and adolescents with ASD. assessments, due to study burden and non-efficacy. Two
First, a modified Maximum-Tolerated-Dose (MTD) study was participants dropped out before first dose was administered
agreed upon in collaboration with Health Canada to identify a and were replaced in the MTD escalation scheme.
maximum dose for multi-dose studies in children up to a There was no effect of dose on measures of efficacy.
maximum of 24 IU/per dose adjusted for weight. The study However, with only 3 children per group, the study was not
also aimed at evaluating safety of multi-dose dosing in this designed and not powered to examine efficacy with each
age group and identifying measures sensitive to change to be dose range. Several measures were insensitive to change or
used in a follow-up randomized controlled trial in this not affected by oxytocin in our sample. These tasks included
population. We report here the results of the MTD study. the DANVA, the Benton Facial Recognition Test, some subt-
ests of the LFI battery, the Real and Apparent Test and the
Second order Irony and Empathy tasks. For some, our
participants experienced ceiling effects (e.g. DANVA) and for
2. Results
some the participants experienced floor effects (e.g. 2nd order
irony and empathy tasks). Several other measures of social
Fifteen children and adolescents (11 male; 4 females; mean
function, social cognition, repetitive behaviors and anxiety
age: 13.8 (2.4) years), with a diagnosis of high-functioning
did show improvements across 12 weeks of treatment
autism or Asperger's Disorder (ADOS:SocialþCommunication:10.33
(Table 2). These include the ABC-SW scale, SRS, functional
(3.21)); (ADIsocial:19.87 (4.76)); (ADIcommunication: 15.80 (3.67));
communication and social skills subtests of the BASC, gen-
(ADIrepetitive 5.80 (1.97)) were recruited into the 16 week study
eral and separation anxiety subscales of the CASI, both
with twice daily dosing. Mean full scale IQ was 101.47 (22.60).
repetitive behavior measures (CYBOCS and RBS-R), as well
Of the 4 doses tested for 12 weeks duration (0.2, 0.26, 0.33
as measures of social recognition, empathic accuracy and
and 0.4 IU/kg/dose), the 0.4 IU/kg/dose was the maximum
theory of mind. Lastly, some of the measures of social
tolerated dose, as no serious adverse events (SAEs) or severe
cognition and function showed carry over effects 3 months
adverse events were noted at any dose level. Adverse events
after discontinuation of drug.
were mild to moderate, and either expected or typically
Ten of 15 participants were noted to be global responders
associated with the disorder (Table 1). The WRAML revealed
based on Clinical global Impression CGI-I-Global at week 12
no adverse events related to memory (Verbal Recall p ¼0.16,
and 6/14 participants remained responders 3 months after
Picture Memory Recognition p¼ 0.92, Verbal recognition
the last dose was administered. In terms of the CGI-I-Social,
p¼ 0.6). There were no clinically significant alterations in
7/15 were classified as responders at week 12 and 6/14
CBC, electrolytes, liver/renal function and osmolality, or
remained responders at week 24.
electrocardiograms. There were no discontinuations due to
adverse events (Fig. 1). One adolescent dropped out after the

Table 1 – Safety (# events severity, if present in more than


10% of participants). 3. Discussion
Neuropsychiatric disorders
The study is contributing to accumulating literature to
Emotional lability (3 mild, 4 moderate)
Irritability (6 mild)
suggest potential efficacy and safety of multi-dose intranasal
Headache (9 mild, 2 moderate) oxytocin in children and youth with ASD. Although this is not
Migraine (3 mild) a conventional dose finding protocol, doses up to 0.4 IU/kg/
dose, given twice a day over 12 weeks produced no severe or
Gastrointestinal disorders
Abdominal discomfort (3 mild)
serious adverse events, and no metabolic or EKG abnormal-
Nausea (1 mild, 1 moderate) ities. Two thirds of the sample was classified as global
responders, and almost half the sample was classified as
Infections and infestations
responders in social function by the CGI at 12 weeks. Of
Upper respiratory Infections (5 mild, 2 moderate)
particular interest, a large percent of week 12 responders
Metabolism and nutrition disorders maintained improvements 3 months after the end of the
Decreased appetite (5 mild) study, especially in the social domain.
Skin and subcutaneous tissue disorders A large number of measures were examined with respect
Rash (3 mild) to their sensitivity to change in this pilot study. The reader
Itchy nose (2 mild) should note that this study was not a randomized controlled
General disorders trial, the proper method to confidently answer questions of
Fatigue (4 mild, 1 moderate) efficacy, and as such the results on outcome measures should
be interpreted with caution. There is a lack of data on
Respiratory, thoracic, and mediastinal disorders
instruments that are sensitive to change with treatment,
Asthma attack (2 mild)
especially within the domain of social function. Our study
Tooth disorders was designed to determine instruments appropriate for
Tooth sensitivity/pain (1 mild, 1 moderate)
follow-up in a larger randomized controlled trial. Within this
Serious adverse events 0 context, improvements were noted in the social domain on
both parent/caregiver report as well as direct assessments,
192 brain research 1580 (2014) 188–198

CONSORT Flow Diagram

Enrollment
Assessed for eligibility (phone

screened (n=120)
Excluded (n=103)

Not meeting inclusion criteria

(n=62)
Consented (n=17)
Declined to participate (n=41)
Discontinued prior to

medication n=2

Received IN-OXT (n=15)

Received allocated intervention (n=15)

Lost to follow-up (n=1)

Discontinued intervention (n=0)

Analysis

Analysed (n=15)

Excluded from analysis (n= 0)

Fig. 1 – Consort flow diagram.

Table 2 – Estimates of 0–12 and 0–24 week change for the 14 outcomes of 35 with a statistically significant (unadjusted)
effect.

Variable 0–12 week (95%CL) p Value 0–24 week (95%CL) p Value

Social function
ABC_SW 3.00 (0.05;5.96) 0.05 2.91 ( 1.98;7.80) 0.2
Srs_total_tscore 9.8 (3.8;15.8) 0.002 8.2 ( 0.4;16.8) 0.06
Basc_social skills tscore 2.84 ( 5.31;  0.38) 0.03 2.15 ( 5.42; 1.13) 0.2
Basc_functional communication tscore 4.10 ( 6.23;  1.97) 0.0006 4.19(  7.29; 1.10) 0.01

Social cognition
RMET_hard items 0.14 ( 0.82;0.53) 0.7 1.15 ( 1.99; 0.31) 0.009
LFI_match_maker_ID 10.9 ( 19.3;  2.6) 0.01 7.8 ( 18.0;2.5) 0.1
LFI same different faces 9.21(  15.26–3.15) 0.005 7.4 ( 15.5;0.6) 0.07
LFI same different houses 10.73 ( 18.5;  2.91) 0.009 10.2(  20.3; 0.2) 0.05
Irony_Empathy_first order 1.64 ( 3.02;  0.27) 0.02 2.31 ( 4.08; 0.54) 0.01
Strange Stories Task 1.36 ( 2.72; 0.008) 0.05 0.28 ( 2.00;1.44) 0.7

Anxiety
ASI – general anxiety tscore 11.0 (4.0;18.0) 0.004 6.5 ( 2.5;15.6) 0.4
ASI – separation anxiety tscore 7.5 (0.2;14.8) 0.04 5.7 ( 3.3;14.7) 0.2

Repetitive behaviors
RBS-R total 17.3 (6.3; 28.3) 0.003 6.0 ( 11.8;23.9) 0.5
CYBOCS 2.93 (0.95;4.91) 0.004 2.52 (0.11;4.93) 0.04

suggesting some consistency across measures using different most likely a measure of global ASD severity, as well as the
measurement approaches. social skills and functional communication domains of the
Positive signals in the social domain were found on the BASC. Of interest, gains noted by the SRS and the functional
ABC-SW, a measure of social withdrawal, the SRS total score, communication domain of the BASC were maintained 3
brain research 1580 (2014) 188–198 193

months post end of treatment, potentially suggesting a social


learning effect. A similar pattern was noted in aspects of 4. Experimental procedures
social perception/cognition. Improvements in face recogni-
tion, and theory of mind were noted at 12 weeks. Some This was a modified Maximum Tolerated Dose (MTD), open
persisted at 24 weeks and empathic accuracy improvements label trial of intranasal oxytocin in children and adolescents
were most evident at 24 weeks, further supporting the with ASD.
hypothesis already in the literature that oxytocin may facil-
itate social learning. Of note, some non-specific improve- 4.1. Participants
ments were noted in the “same-different” tasks of the LFI
that were not specific to social stimuli and therefore cannot Children and adolescents, ages 10–17 inclusive, with a diag-
be interpreted as social recognition gains. Most likely such nosis of an ASD, and verbal IQ equal to or above 70, were
effects represent a nonspecific effect on attention and gen- enrolled in a 16 week study (12 weeks of medication exposure
eral, categorical learning. Anxiolytic effects were pronounced plus four weeks of follow-up off medication). Participants
during treatment, but disappeared with discontinuation of were recruited through the Holland Bloorview Kids Rehabili-
treatment. Lastly improvements were noted in repetitive tation Hospital clinical database as well as presentations at
behavior over the 12 weeks with some maintenance of effect local conferences and media. Diagnosis was established using
3 months later. a diagnostic interview to established DSM-IV criteria for an
ASD, utilizing the Autism Diagnostic Observation Schedule
(ADOS) (Lord et al., 2000) and the Autism Diagnostic
3.1. Limitations Interview-Revised (ADI-R) (Lord et al., 1994) that were con-
ducted by research reliable administrators. All eligibility
This is a small modified MTD study. As such, the sample assessments were completed prior to randomization into
size is too small to view both safety and efficacy data the study.
without caution. In addition, as the MTD design was For inclusion in this study, participants had to have base-
modified in collaboration with regulatory agencies to not line severity score (Clinical Global Impression – Severity
exceed the commonly used 24IU dose, it is not clear that CGI-S) of Z4 (moderately ill), be on stable pharmacological
doses higher than that would not confer benefit within and or non-pharmacologic educational, behavioral, and/or
acceptable safety parameters. Further, in the absence of dietary interventions (4 3 months prior to screening), and
placebo control, adverse events associated with oxytocin have normal physical examination and laboratory test
may be inflated. Lastly, given the sample size, interactions results. We excluded patients born prior to 35 weeks gesta-
with concomitant medications (Table 3) could not be tional age, patients with any primary psychiatric diagnosis
examined. other than autism at screening, those with current severe
neurological disease, including, but not limited to, epilepsy/
seizure disorder (except simple febrile seizures), severe
3.2. Conclusions
movement disorder, tuberous sclerosis, fragile X, and any
other known genetic syndromes, or known abnormal MRI/
Accumulating data suggest that oxytocin manipulation may
structural lesion of the brain, pregnant female patients,
have therapeutic potential in ASD. Early single dose studies
sexually active female patients on hormonal birth control
as well as case series and early multidose data support this
and sexually active females who do not use two types of non-
hypothesis. Our data extends this data set, adds to the safety
hormonal birth control, patients with a medical condition
profile of the multidose intranasal oxytocin administration in
that might interfere with the conduct of the study, confound
children and youth, establishes maximum tolerated dosage
interpretation of the study results, or endanger their own
within a range established by regulatory agencies and sug-
well-being, those who are sensitive to Syntocinon or any
gests several measures as potential candidates to be used in
components of its formulation, those with HIV, HBV, HCV,
follow-up randomized controlled trials in this population,
hemophilia, abnormal blood pressure, drug abuse (as per
already funded.
DSM criteria), immunity disorder or severe depression and
those unable to tolerate venipuncture procedures for blood
sampling. All patients or their legal guardians signed the
Table 3 – Concomitant medications. institutionally approved informed consent/assent according
Medication (indication) Frequency (out of 15) to the Helsinki agreement and tri-council policy.

Stimulantsa (ADHD like symptoms) 6 4.2. Dose selection and MTD design
Melatonin (insomnia) 4
Clonidine (insomnia) 1
Atomoxetine (ADHD like symptoms) 1
A modified dose finding method was used to determine
Sertraline (anxiety) 1 safety among four dose levels. In adults, the dose with most
Risperidone (impulse control) 2 evidence is 24 IU/dose. This is the equivalent of 0.4 IU/kg for a
60 kg person. Therefore, the maximum dose to be tested in
a
Methylphenidate (Concerta:1); Lisdexamfetamine (Vyvance):1;
youth approved by Health Canada was 0.4 IU/kg/dose for a
Methylphenidate (Ritalin):1; Methylphenidate (Biphentin):2;
maximum of 24 IU/dose. This is different from a traditional
Dextroamphetamine (Adderal):1.
MTD design where the upward titration does not typically
194 brain research 1580 (2014) 188–198

stop unless the participants experience serious adverse administration and determine safety of first dose. Partici-
events. Half the dose (0.2 IU/kg/dose) was the minimum dose pants were advised to administer one spray every 30 s until
approved and 2 intermediate doses were also evaluated: 0.26 they have reached the total number of sprays directed. They
and 0.33 IU/kg/dose. were told to sit upright, hold the bottle up right, insert the
Dose-finding escalations were done in groups of three nozzle into one nostril and inhale gently through the nose
patients. The following dose escalation rules were applied: while pushing down on the nozzle to activate the pump.

1. Three patients were studied at the first dose level (0.2 IU/ 4.5. Medications
kg bid).
2. If none of these patients experienced Dose Limiting Oxytocin (Syntocinon, NOVARTIS, Switzerland) was adminis-
Toxicity (DLT), then the dose was escalated to the next tered in the form of IN-OXT.
higher level in the three subsequent patients.
3. If one of three patients had experienced DLT at the current 4.6. Safety assessments
dose, then up to three more patients were to be accrued at
the same level. (a) If none of these three additional Participants were seen every two weeks during the 12-week
patients experienced DLT, then the dose were to be study and then again at 16 weeks, 4 weeks after last exposure
escalated in subsequent patients. (b) If one or more of to drug, for Clinical Global Improvement (CGI-I) ratings, vital
these additional patients experienced DLT, then patient signs and adverse event monitoring. Adverse events were
entry at that dose level would be stopped, the MTD elicited using the SMURF (Greenhill et al., 2004) at every visit
exceeded and dose escalation would be stopped. Up to and the Wide Range Assessment of Memory and Learning
three more patients would be treated at the next lower (WRAML) (Sheslow and Adams, 2005) was done at baseline
dose. If zero out of three patients experience DLT at a dose and end visits (weeks 12 and 24) to document any potential
of 0.4 IU/kg, an additional three patients were to be treated adverse events related to memory, based on early animal
at that dose. Using this escalation scheme, the probability model concerns (Engelmann et al., 1996). Safety blood work
of escalating to the next level if the true proportion of and EKG were done every 4 weeks.
adverse effects was 10%, 20%, 30%, 40%, or 50% was 91%,
71%, 49%, 31% and 17% respectively. Each participant only 4.7. Efficacy assessments
received one of the preset doses.
As noted above, the aim of this study was to establish
maximum tolerated dose and safety for IN-OXT in this age
group in ASD. In addition, we piloted several outcome
Twice a day dosing (morning and afternoon) was chosen
measures related to social cognition/function, repetitive
given that CNS levels of vasopressin seem to rise for approxi-
behaviors and co-occurring anxiety to select those with
mately 4 h post intranasal administration (Born et al., 2002),
documented sensitivity to change. These outcomes will be
and in order to capture the most hours in which children and
used in the follow-up randomized controlled trial in a larger
youth are most likely to be involved in social interactions.
sample with ASD and this study will also be funded by the
Department of Defense.
4.3. Definition of toxicity for MTD design
4.7.1. Social cognition measures
Based on existing data, it was difficult to find a definition of All social cognition measures were administered at baseline
toxicity for this compound. Although no standard toxicity and then week 12 and week 24.
studies have been published for IN-OXT in children, the Side Diagnostic Analysis of Nonverbal Accuracy (DANVA-2)
Effect (SE) profile of IN-OXT in adults after decades of post (Baum and Nowicki, 1989). The DANVA-2, a measure of
marketing use has been very benign. Standard toxicity pro- emotion recognition across multiple modalities with estab-
tocols are typically based on Serious Adverse Events (SAEs) to lished reliability and validity for age 3–100, was administered
determine the Maximum Tolerated Dose (MTD). In our case, to participants at baseline, week 12 and week 24.
we decided to determine toxicity as the presence of SAEs or Let's Face It! Skills Battery (or Victoria/Yale Face Proces-
severe adverse effects that are deemed by the investigators to sing Battery; VYFPB; Wolf et al., 2008) is a battery of compu-
be likely or probably related to drug. terized face processing tasks. The full battery consists of 11
separate computer-administered tests. It assesses 2 broad
4.4. Dosing schedule domains involving (1) the perception of facial identity and (2)
the perception of facial expression, with good split-half
We selected morning and afternoon dosing to try to influence reliabilities (40.75).
the greatest number of hours when youth are in settings with Benton Face Recognition Task (BFRT) (Benton et al., 1994)
increased potential for social interaction (school, afterschool). is a standardized test frequently used to assess perceptual
Moreover, this regimen was associated with improvements in skill in developmentally delayed populations, with good
our adult pilot study (Anagnostou et al., 2012). Medication reliability. It is appropriate for participants aged 6–74 and
was administered by the parents before school and right after provides a standardized and objective procedure for asses-
school. All patients received their child's first dose by the sing the capacity to identify and discriminate photographs of
study physician to educate parents and themselves on proper unfamiliar human faces.
brain research 1580 (2014) 188–198 195

The Revised Eyes Test (Baron-Cohen et al., 2001) assesses Behavior Assessment System for Children (BASC) (Reynolds,
the ability to decipher mental states of others. Participants 1992). We used the parent report scale with a focus on
are presented with photographs of the eye-region of male functional communication and social skills subscales.
and female actors accompanied by four descriptive words
(e.g., “serious,” “ashamed,” “alarmed,” or “bewildered”) and 4.7.3. Repetitive behaviors
must select the word that best describes what the actors are Both measures were administered at every visit
thinking or feeling. In addition to restricting stimulus pre-
sentation to the eye region, this test differs from others in  Child Yale-Brown Obsessive-Compulsive Scale (C-YBOCS)
that it assesses more complex emotional states (e.g., shame, (Goodman et al., 1989, 65, modified by Scahill et al., 1997)
bewilderment). The Yale-Brown Obsessive-Compulsive Scale is a
Strange Stories Task (Happé, 1994) assesses the ability to clinician-rated questionnaire measuring the time spent,
interpret nonliteral statements from stories read to the distress, interference, resistance, and control in relation
participant. Questions probe whether the child understands to obsessions and compulsions based on a 5-point scale.
that (a) a nonliteral statement has been made and (b) the The Compulsion subscale has been shown to be a reliable
intent behind the statement (i.e., was the speaker was lying, and valid scale in ASD, and in measuring change in
being sarcastic, or joking?). treatment studies of autism, and we used the “adapted
Real and Apparent Emotion task: (Dennis et al., 2000) for PDD” version.
evaluates children's understanding of real and deceptive  Repetitive Behavior Scale-Revised (RBS-R) (Bodfish et al.,
emotion in short narratives. The task, typically mastered 2000). The RBS-R is a parent-report measure developed to
between the 6–8 years of age, represents an advanced capture the breadth of repetitive behaviors that are specific
assessment of theory of mind in that it requires judgments to autism. It consists of 43-items that tap six repetitive
about socially expressed emotion in the context of belief. behavior subtypes: Stereotyped, Self-injurious, Compulsive,
Irony and Empathy Task (Dennis et al., 2001) measures Ritualistic, Sameness, and Restricted Interests.
children's understanding of first and second order intention-
ality associated with literal truth, ironic criticism, and decep-
tive praise. Six everyday situations depict a task (e.g. tidying a
room, baking a cake), with two participants, a speaker who 4.7.4. Anxiety measures
makes a comment about the task and hearer who did the Child and Adolescent Symptom Inventory (CASI) (Gadow and
task. The task has been well published in children with Sprafkin, 2009): The CASI is a clinician-reviewed, caregiver
traumatic brain injury. report form based on the DSM-IV used to assess comorbid
psychiatric symptoms. The anxiety subscales were used.
4.7.2. Social function measures Participants used a medication diary to mark down every
Aberrant Behavior Checklist (ABC) (Aman et al., 1985) The time they took the medication. The diary was reviewed at
ABC is a symptom checklist for assessing problem behaviors every visit and the study clinician and the participant
(ages 6–54 years). The checklist consists of 58 items broken problem solved together in the case of missed doses to
down into 5 subscales; only the Social Withdrawal subscale improve compliance.
was used in this study. It was administered at baseline and
weeks 12 and 24. 4.8. Statistical approach
Social Responsiveness Scale (SRS) (Constantino, 2002) is a
caregiver/educator rating scale of social behaviors specific to Data were analyzed using SAS 9.3 (ref 2002–2010 by SAS
ASD. It is a sensitive and reliable assessment that is designed Institute Inc., Cary, NC, USA). We plotted all outcomes across
to aide diagnosis and measure treatment response with 4–18 time to visually assess any trends across dose. Data were
years of age. The SRS measures social awareness, informa- modeled using repeated measures regression analysis con-
tion processing, and social motivation and yields a quantita- trolling for week, dose, age, and sex. The changes from
tive score that has been useful in endophenotyping studies of baseline to week 12 and baseline to week 24 and their 95%
ASD and was administered at every visit. confidence limits were estimated from the model for each of
CGI-I-Social: The CGI-I is a well validated measure the 35 outcomes variables examined. Eight of the outcomes
employing a 7-point scale of clinical global impression of were also available as raw scores– in all cases, the results
improvement (very much improved, much improved, mini- were similar using either score and only t-scores are pre-
mally improved, no change, minimally worse, much worse, sented in the results.
very much worse) that the clinician fills out after considering
all the available information on the subject including the
parent history, the examination in clinic, reports from the Competing interests/Financial disclosures
school and other sources (Guy, 1976). In this case, the focus of
the interview was on aspects of social function: approach, Dr. Evdokia Anagnostou has received consultation fees from
turn taking, peer relationships and was administered at Seaside Therapeutics and Novartis and an unrestricted grant
every visit. from Sanofi – Aventis Canada.
Behavioral Assessment System for Children-2 is used to Dr. Latha Soorya, Dr. Jessica Brian, Dr. Annie Dupuis, Dr.
evaluate the behavior and perceptions of children and young Deepali Mankad, and Dr. Sharon Smile have nothing to
adults (ages 2–25 years). The BASC-2 is a revision of the disclose. Dr. Suma Jacob is a consultant to CogCubed games
196 brain research 1580 (2014) 188–198

and has as nothing to disclose with regards to pharmaceu- Benton, A.L., Sivan, A.B., KdeS, Hamsher, Varney, N.R., Spreen, O.,
tical companies. 1994. Contributions to Neuropsychological Assessment, 2nd
edition Oxford University Press, New York.
Bodfish, J.W., Symons, F.J., Parker, D.E., Lewis, M.H., 2000. Varieties
of repetitive behavior in autism: comparisons to mental
Funding retardation. J. Autism Dev. Disord. 30, 237–243.
Born, J., Lange, T., Kern, W., McGregor, G.P., Bickel, U., Fehm, H.L.,
This study was funded by a Grant from the Department of 2002. Sniffing neuropeptides: a transnasal approach to the
human brain. Nat. Neurosci. 5, 514–516.
Defense (DOD) (AR093387) to Dr. Anagnostou.
Carter, C.S., 1998. Neuroendocrine perspectives on social
attachment and love. Psychoneuroendocrino 23, 779–818.
CDC-Autism and Developmental Disabilities Monitoring Network
Surveillance Year; 2008 Principal Investigators; Centers for
Authors' contributions
Disease Control and Prevention 30;61(3):1–19. PubMed PMID:
22456193.
EA was the principal investigator for the study. She designed Constantino, J.N., 2002. The Social Responsiveness Scale. Western
and ran the study, and prepared the manuscript. LS con- Psychological Services, Los Angeles.
tributed to designing the study and manuscript preparation. Dadds, M.R., Macdonald, E., Cauchi, A., Williams, K., Levy, F.,
JB contributed to the design of the study, supervised psycho- Brennan, J., 2013. Nasal oxytocin for social deficits in
logical assessments, and contributed to manuscript prepara- childhood autism: a randomized controlled trial. J. Autism
tion. SJ contributed to designing the study, preparing the Dev. Disord..
Dennis, M., Lockyer, A., Lazenby, A., 2000. How high-functioning
manuscript and is a site-PI on the subsequent RCT treatment
children with autism understand real and deceptive emotion.
trial. AD contributed to study design, data analysis and
Autism 4, 370–381.
manuscript preparation. DM and SS were critical to running Dennis, M., Purvis, K., Barnes, M.A., Wilkinson, M., Winner, E.,
the study and contributed to manuscript preparation. All 2001. Understanding of literal truth, ironic criticism, and
authors read and approved the final manuscript. deceptive praise following childhood head injury. Brain Lang.
78, 1–16.
Domes, G., Heinrichs, M., Gläscher, J., Büchel, C., Braus, D.F.,
Herpertz, S.C., 2007. Oxytocin attenuates amygdala responses
Acknowledgments to emotional faces regardless of valence. Biol. Psychiatry 62,
1187–1190.
The authors would like to thank Ms. Nadia Tanel, Ellen Domes, G., Heinrichs, M., Michel, A., Berger, C., Herpertz, S.C.,
Drumm, Lisa Genore and Dina Zaghloul as well as Mr. Brian 2006. Oxytocin improves “Mind-Reading” in humans. Biol.
Psychiatry 61, 731–733.
Crowe for their assistance in running the study and in the
Donaldson, Z.R., Young, L.J., 2008. Oxytocin, vasopressin, and the
preparation of this manuscript.
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