Clinical trial NCT01970345
Conditions: Autism Spectrum Disorder
Keywords: Autism Spectrum Disorder IGF-1
Sponsor: Icahn School of Medicine at Mount Sinai
Start date: 2014-05-01; Completion date: 2017-09-13
Enrollment: 3 participants; Allocation: RANDOMIZED; Intervention model: CROSSOVER

Brief summary:
The proposed project will pilot the use of IGF-1 as a novel treatment for core symptoms of autism. We will use a double-blind, placebo-controlled crossover trial design in five children with autism to evaluate the impact of IGF-1 treatment on autism-specific impairments in socialization, language, and repetitive behaviors. We expect to provide evidence for the safety and feasibility of IGF-1 in ameliorating social withdrawal in children with Autistic Disorder. Further, we expect to demonstrate that IGF-1 is associated with improvement on secondary outcomes of social impairment, language delay, and repetitive behavior, as well as on functional outcomes of global severity.

Detailed description:
IGF-1 is an FDA approved, commercially available compound that crosses the blood-brain barrier and has beneficial effects on synaptic development by promoting neuronal cell survival, synaptic maturation, and synaptic plasticity. IGF-1 is effective in reversing mouse and neuronal models of Rett syndrome and Phelan McDermid syndrome, both single gene causes of ASD and may therefore be effective in treating autism spectrum disorders (ASD) more broadly.

Arms: arm 1: Randomized, placebo-controlled, crossover format with 12 weeks in each treatment arm (IGF-1 and placebo), separated by a four-week wash-out phase.

Dose titration will be initiated at 0.04 mg/kg twice daily by subcutaneous injection, and increased, as tolerated, every week by 0.04 mg/kg per dose to a maximum of 0.12 mg/kg twice daily. Doses may be decreased according to tolerability by 0.04 mg/kg per dose. Medication will be administered twice daily with meals, and preprandial glucose monitoring will be performed by parents at treatment initiation, prior to each injection, and until a well tolerated dose is established. arm 2: Placebo
Interventions: intervention 1: IGF-1 intervention 2: Placebo/saline
Intervention descriptions: intervention 1: IGF-1 is an FDA-approved (IND exemption #113450), commercially available compound that crosses the blood-brain barrier and has beneficial effects on synaptic development by promoting neuronal cell survival, synaptic maturation, and synaptic plasticity. intervention 2: Placebo

Protocol documents:
Prot_SAP_000.pdf: 
A Pilot Treatment Study of Insulin-Like Growth Factor-1 (IGF-1) in 
Autism Spectrum Disorder
PI: Alexander Kolevzon, MD
NCT01970345
Document Date: 10-1-2021


Electrophysiological markers in PMS and iASD
PI: Kolevzon
1
Electrophysiological Markers for Interventions in Phelan-McDermid Syndrome and Idiopathic 
Autism
Protocol Identifying Number:
Electrophysiological markers in PMS and iASD
Principal Investigator:  
Alexander Kolevzon, MD
Professor of Psychiatry and Pediatrics
Clinical Director, Seaver Autism Center
Icahn School of Medicine at Mount Sinai
New York, NY 10029
Tel: 212-659-9134
Email: alexander.kolevzon@mssm.edu
IND Sponsor:
Alexander Kolevzon, MD
Draft of Version Number: 
v 2.0
Date: 
August 17, 2021
Confidentiality Statement:
The trial will be carried out in accordance with Good Clinical Practice (GCP) as required by the 
following: United States (US) Code of Federal Regulations (CFR) applicable to clinical studies (45 
CFR Part 46, 21 CFR Part 50, 21 CFR Part 56, 21 CFR Part 312, and/or 21 CFR Part 812), ICH 
E6
All key personnel (all individuals responsible for the design and conduct of this trial) have 
completed Human Subjects Protection Training.
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Electrophysiological markers in PMS and iASD
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Introductory Statement and Investigational Plan
The genomic revolution in autism spectrum disorder (ASD) has positioned the field to study 
personalized approaches to treatment. Recent work suggests that up to 2% of individuals with ASD 
and intellectual disability (ID) have deletions or point mutations in the SHANK3 gene, resulting in 
Phelan-McDermid syndrome (PMS), and approximately 65% of people with PMS meet criteria for 
ASD. SHANK3 codes for a master scaffolding protein that forms a key framework in the 
postsynaptic density of glutamatergic (excitatory) synapses and plays a critical role in synaptic 
function. SHANK3 and glutamate pathways are implicated in multiple forms of ASD and this 
convergence implies shared biochemical pathways with potentially overlapping therapeutic targets. 
Thus, targeting SHANK3 deficiency in PMS as a specific genetic cause of ASD allows us to inform 
treatment in broader idiopathic ASD (iASD). Our ongoing studies provide in-depth phenotyping and 
natural history of PMS, pointing toward a specific clinical and electrophysiological (EEG) profile. 
However, the potential of EEG biomarkers as a measure of treatment response remains to be 
determined. 
In PMS, our preliminary data using visual evoked potentials (VEPs) to examine cortical 
excitatory postsynaptic potentials demonstrate promising links to disease mechanisms. 
Biomarkers and novel clinical measures for evaluating response to intervention have been piloted 
successfully at our site in cohorts of patients with PMS and iASD. We have identified a unique VEP 
profile of excitatory deficits (markedly reduced P60-N75 transient VEP amplitude) in PMS that is also 
present in a subset of children with iASD in order to stratify individuals with iASD and select those 
we predict will show response to growth hormone. Importantly, our pilot work showed clinical 
benefit of both insulin-like growth factor-1 (IGF-1) (Kolevzon et al. 2014) and growth hormone 
(which triggers release of IGF-1) in PMS, and significant improvement in gamma band activity on 
our primary VEP measure post-treatment with IGF-1. VEP biomarkers may therefore be harnessed 
as an indicator of brain engagement during intervention.
The overall objective of this proposal is to use clinical and electrophysiological features of PMS 
to characterize subtypes of iASD and inform treatment development. We will enroll children age 2-
12 years, specifically recruiting intellectually disabled and minimally verbal children given the 
prominence of this profile in PMS and the critical need to address this group in broader ASD 
research. Our short-term goal is to show that select electrophysiological markers in PMS are 
relevant to iASD and predictive of treatment response. Our long-term goal is to optimize treatment 
selection in iASD by establishing biological signature(s) derived from PMS that are: a) useful for 
predicting treatment responders, and b) responsive to intervention
Aim 1: Validate the ASD phenotype within PMS to select clinical targets for intervention. 
This Aim has been completed. Measures were selected based on: a) our preliminary data showing 
a unique profile of sensory reactivity in PMS, and b) their focus on core symptoms of ASD. We 
validated our phenotypic profile of PMS, showed that it overlaps with a subset of patients with 
iASD, and established clinical targets and appropriate outcome measures for both groups.   
Aim 2: Establish electrophysiological markers of PMS and iASD using VEPs. This aim 
has also been completed. We established the stability of VEP amplitudes across two time points, 
in order to validate this tool as a reliable stratification biomarker capturing excitatory activity. We 
also showed that individual variability detected by electrophysiological measures was associated 
with clinical symptoms of PMS and iASD measured in Aim 1, including sensory reactivity. Our data 
suggest a unique VEP profile of excitatory deficits in PMS that overlaps with a subset of children 
with iASD. Neural responses show strong test-retest reliability, associations with clinical metrics in 
PMS and iASD, and in Aim 3 we will now test whether they are useful in stratifying patients with 
iASD with excitatory deficits.
Aim 3. Evaluate the feasibility of electrophysiological markers as a measure of 
treatment response. Using a double-blind, placebo-controlled crossover design, we will evaluate 
the use of electrophysiological markers to detect neural response to growth hormone treatment in 
30 children (15 PMS; 15 iASD), all of whom show characteristic VEP waveforms reflecting deficits 
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Electrophysiological markers in PMS and iASD
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in glutamatergic activity pre-treatment. Based on our preliminary data, approximately 50% of 
patients with iASD show reduced P60-N75 responses comparable to PMS; this subset will be 
targeted for the trial. We expect gamma band VEP activity will be sensitive to growth hormone and 
will track with clinical response on sensory measures. We will also explore the relation between 
electrophysiological response, growth hormone dose, and IGF-1 level, by examining VEP activity 
by time point. 
We have completed Aims 1 and 2 of this study and this revised protocol will focus on Aim 3. 
The expected outcome of this Aim 3 is to establish the feasibility of electrophysiological 
biomarkers for use in clinical trials in PMS and iASD and define a subset of patients with iASD 
likely to show clinical response to treatment.
RESEARCH STRATEGY
Significance: Overall Scientific Premise
Development of effective treatments for autism spectrum disorder (ASD) is hampered by 
several barriers: 1) ASD is a heterogeneous disorder in which etiology and neuropathology is 
unknown in most cases, 2) existing parent-report and clinician-administered assessments often 
lack the reliability, objectivity, sensitivity, and precision necessary to be confidently translated to 
clinical trials as outcome measures, and 3) many individuals with ASD who are most severely 
affected are often excluded from research. Gene discovery approaches, followed by functional 
analysis of model systems, have elucidated the neurobiology of several genetic subtypes of ASD 
and intellectual disability (ID) and led to important opportunities for developing novel, disease-
modifying therapeutics. Phelan-McDermid syndrome (PMS) is a common monogenic form of ASD, 
accounting for 0.5-2% of ASD cases, and resulting from haploinsufficiency of SHANK3 due to 
22q13.3 deletions or mutations in the gene (Durand et al., 2007; Moessner et al., 2007; Gauthier 
et al., 2009; Marshall et al., 2008; Leblond et al., 2014; Cooper et al., 2011; Bonaglia et al., 2006; 
2011). SHANK3 codes for a key scaffolding protein in the postsynaptic density of glutamatergic 
synapses and plays a critical role in synaptic function (Boeckers et al, 2006). Indeed, recent 
evidence suggests that the SHANK3 and glutamate signaling pathway is common to multiple forms 
of ASD and that many different genetic causes of ASD and ID, including tuberous sclerosis and 
Fragile X syndrome (FXS), converge on common pathways, including SHANK3 (Darnell et al., 
2011; Sakai et al., 2011). Thus, studies in PMS offer a unique opportunity to constrain ASD 
heterogeneity using a subset of individuals in which the neuropathology is better understood and 
can more readily be targeted for treatment. Importantly, the ratio of affected males to females is 
approximately equal in PMS and therefore provides critical opportunities for comparison between 
sexes.  
Discovering the function of the SHANK3 protein has led us to translational work to develop 
novel therapeutics for ASD. Studying Shank3-deficient mice, we have identified specific deficits in 
synaptic function and plasticity in glutamate signaling (Bozdagi et al, 2010). Insulin-Like Growth 
Factor-1 (IGF-1) reversed electrophysiological and behavioral deficits in the mice (Bozdagi et al., 
2013) and, critically, improved social withdrawal and restricted behaviors in children with PMS 
(Kolevzon et al., 2014). IGF-1 has also shown benefit in human, mouse, and neuronal models of 
Rett syndrome (Khwaja et al., 2014; Tropea et al, 2009; Marchetto et al., 2010), providing evidence 
of the potential for this treatment to have significant benefit not only in PMS, but also in other forms 
of ASD with heterogeneous genetic etiologies. However, IGF-1 is difficult and costly to obtain and 
associated with significant risks of hypoglycemia. Since IGF-1 can be increased intrinsically by 
growth hormone administration, we will use recombinant human growth hormone (rhGH) instead 
and our pilot data demonstrate significant benefit with rhGH in 
children with PMS across a wide range of clinical symptoms.  
Because PMS has constrained genetic heterogeneity and the 
SHANK3 and glutamate signaling pathway underlies other forms of 
ASD, this project represents an excellent opportunity to develop 
biomarkers and test a novel therapy. Our preliminary data 
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Electrophysiological markers in PMS and iASD
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demonstrate promising electrophysiological results with links to glutamatergic dysregulation in 
PMS using visual evoked potentials (VEPs). These data show that VEPs are objective, stable, 
linked to ASD symptoms, and sensitive to treatment effects. This study will be significant in further 
probing VEP and other electrophysiological measures with links to glutamatergic functioning, in 
children with PMS and idiopathic ASD (iASD). 
Scientific Premise for Aim 3. Our preliminary work and results from data collected as part of 
Aims 1 and 2 suggests some elements of the ASD phenotype in PMS that are unique. Although 
approximately 80% of both children with PMS and iASD show sensory reactivity abnormalities, 
even in lower functioning samples, children with PMS have significantly less overall sensory 
sensitivity but more weak/low-energy symptoms as compared to 
children with iASD (Mieses et al., 2016). Sensory symptoms are now 
included in core DSM-5 diagnostic criteria for ASD; they are 
especially important to this project based on relation to VEP neural 
responses in both PMS and iASD populations, and may represent a 
phenotypic profile in PMS that informs which subset of individuals 
with iASD are most likely to benefit from intervention with IGF-1. Our 
preliminary data also show a unique pattern of repetitive behaviors and sensory symptoms that 
overlaps with that seen in iASD but has some key defining features. Assessment of sensory 
aspects of the ASD phenotype has been particularly hampered by the lack of objective, 
observational measurements of functioning in this domain.
To address this gap, we developed the Sensory Assessment for Neurodevelopmental 
Disorders (Siper et al., 2017), which is a clinician-administered assessment and corresponding 
caregiver interview, not reliant on verbal or cognitive capacity and therefore appropriate for 
severely affected populations. The SAND captures hyperreactivity, hyporeactivity, and seeking 
behaviors across visual, tactile, and auditory domains. In an initial sample of 45 children with iASD 
and 33 TD controls, the SAND showed high internal consistency (Cronbach’s α=.90), strong inter-
rater reliability (ICC’s=.87-.91), strong test-test reliability (ICC’s=.82-.97), and good convergent 
validity with the Short Sensory Profile (SSP; Dunn, 1999) (r=-.82, p<.0001). Across sensory 
modalities, increased hyperreactivity, hyporeactivity, and seeking behaviors all characterized 
children with iASD compared to TD controls (ps<.0001), highlighting the prominence of widespread 
sensory reactivity abnormalities in ASD. In comparison to the broader iASD population, children 
with PMS (n=24) are unique in their display of more visual, auditory, and tactile hyporeactivity as 
compared to both iASD (p=.001) and TD (p=.001), as well as less sensory hyper-reactivity and 
seeking behavior compared to iASD (p=.04). Thus, sensory hyporeactivity may be a key feature of 
the ASD profile in PMS and is an important treatment target in PMS. Sensory hyporeactivity, and 
in particular, high pain tolerance in PMS can complicate injuries in PMS and in the absence of 
functional communication, lead to unnecessary infection. Furthermore, results suggest that the 
SAND can identify sensory reactivity subtypes that are a hallmark of PMS and also characterize a 
subset of iASD (Fig.1). Our results suggest that approximately 30% of children with iASD display 
SAND hyporeactivity scores within one standard deviation of the PMS mean (Fig.1; purple iASD 
circles above red dotted line), reflecting “PMS-like” sensory reactivity. It is also important to note 
that both SAND and SSP scores are significantly correlated with our electrophysiological marker 
(Aim 2).
Further, our electrophysiological measures of interest are sensory-related, based on the 
premise that sensory abnormalities are common in ASD (APA, 2013) and significantly impact daily 
functioning. Furthermore, our preliminary data demonstrate that individuals with PMS show a 
relatively consistent pattern of sensory hypo-reactivity in response to visual, auditory, and tactile 
input. VEPs provide a noninvasive technique to evaluate the functional integrity of visual pathways 
in the brain from the retina to the visual cortex via the optic nerve/optic radiations. The VEP is 
recorded 
from 
occipital 
scalp 
locations, is extracted from ongoing 
Figure 2. Repeated testing with (A) TD child and (B) child with PMS using 
contrast-reversing checkerboard stimulus at two time points. Across the epoch,
test-retest correlation in amplitude was >.93 in both groups 
Figure 1: Increased hyporeactivity, hyperreactivity, and 
sensory seeking behaviors are seen in iASD vs. TD.
Children with PMS show significantly more hyporeactivity 
compared to both iASD and TD. Children with PMS also 
show 
less 
hyperreactivity 
and 
seeking 
behaviors 
compared to children with iASD. Intellectual ability did not 
significantly impact any of these findings.
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Electrophysiological markers in PMS and iASD
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EEG through signal averaging, and reflects the sum of excitatory and inhibitory postsynaptic 
potentials occurring on apical dendrites, which modulate excitatory and inhibitory signals received 
by pyramidal cells (Zemon et al., 1986). The major positive and negative peaks and troughs in VEP 
waveforms reflect different cellular events (Zemon et al., 1986; Creutzfeldt et al., 1973). This study 
focuses on a conventional transient VEP response, produced by abruptly modulating the contrast 
of a spatial pattern at low frequencies. The contrast-reversing checkerboard stimulus used in this 
study produces a positive peak at approximately 60ms (P60), reflecting activation of the primary 
visual cortex from the lateral geniculate nucleus. A negative peak at approximately 75ms (N75) 
reflects depolarization and glutamatergic postsynaptic activity spreading to the superficial layers of 
primary visual cortex, and a positive peak at approximately 100ms (P100) reflects superficial 
hyperpolarization and GABAergic activity (Zemon et al., 1980). Coherence of high-frequency 
oscillatory responses, reflected in magnitude-squared coherence (MSC), can be quantified from 
VEPs and provide a critical metric of excitatory activity (Zemon et al., 2009). VEPs have been used 
in a variety of disorders for diagnostic and prognostic purposes (e.g., epilepsy, schizophrenia; 
Schechter et al, 2005; Sheppard et al., 2013; Zemon et al., 2008), and pharmacological studies 
provide evidence for VEP electrogenesis. When GABA-blocking agents are topically applied to 
cortex, there is an enhancement in N75 and attenuation or elimination of P100. In contrast, when 
GABA is applied to cortex, N75 is attenuated and P100 enhanced (Zemon et al., 1980; Purpura et al., 
1959; Purpura et al, 1953). VEPs are used throughout the lifespan, mature early, and are stable 
across development (Garcia-Quispe et al., 2009; Moskowitz et al., 1983).  
Our preliminary findings demonstrate strong test-retest reliability of VEP recordings in TD, 
iASD, and PMS samples using repeated measures within a single recording session and across 
multiple time points (Fig. 2). We have established feasibility of our VEP battery in both individuals 
with PMS and iASD. Results indicate that while children with iASD (on average) and TD controls 
both demonstrate the characteristic waveform, VEP 
amplitudes are significantly smaller in iASD for both P60-N75 
(p=.005) and N75-P100 (p=.009; Fig.3). Moreover, results from 
frequency domain analyses indicate that children with iASD 
showed significantly weaker activity in the 30-36 Hz range 
(p<.01) and 38-48 Hz range (p=.034), corresponding to 
gamma band activity. Cognitive ability has no effect on the 
results. In PMS, we see distinct VEP waveforms with smaller 
P60-N75 amplitudes reflecting deficits in glutamatergic activity 
(Fig. 2B & 3). All PMS participants displayed reduced 
excitatory responses (absence of P60-N75), compared to both 
TD and iASD (p’s<.001). Across groups, reduced P60-N75 
amplitude 
corresponded 
to 
greater 
sensory 
hypo-
responsiveness on the SAND (n=32, r=-.529, p=.001) and 
SSP (n=32, r=.403, p=.022), suggesting VEP response 
marks variability in clinical symptoms. P60-N75 amplitude also 
correlated with SHANK3 deletion size (rho = .414; Fig.3 inlay) 
where there was significantly reduced amplitude in patients 
with larger deletions as compared to point mutations in 
SHANK3 (p=.03). Yet, all patients with PMS display the expected GABAergic positive peak at P100, 
indicating a specific, dissociable, glutamatergic VEP signature. These results are consistent with 
work in animal (Bozdagi et al., 2010) and human neuronal (Shcheglovitov et al., 2013) model 
systems demonstrating the deleterious effects of SHANK3 deficiency on glutamatergic system 
function. In an effort to examine the use of VEPs as a stratification biomarker to aid personalized 
medicine approaches among an otherwise heterogeneous group of children with iASD (n=27), our 
results suggest that approximately 30% display P60-N75 amplitudes within one standard deviation 
of the PMS mean (Fig.4; purple iASD circles below red dotted line), reflecting “PMS-like” excitatory 
Figure 3. PMS shows significantly smaller 
VEP amplitudes relative to TD and iASD.
Inset: P60-N75 amplitude is further reduced
with increased size of SHANK3 deletion.
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Electrophysiological markers in PMS and iASD
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deficits.
Work with IGF-1 in PMS at our Center began with Shank3-deficient mice showing a reduction 
in basal neurotransmission reflecting decreased glutamate (i.e., AMPA receptor-mediated) 
transmission. Long-term potentiation was impaired with no significant 
change in long-term depression (Bozdagi et al., 2010), and intraperitoneal 
injection of IGF-1 reversed the electrophysiological and motor deficits seen 
in the Shank3-deficient mice (Bozdagi et al., 2013). Subsequently, we 
began a controlled trial of IGF-1 in PMS using the design proposed herein, 
and providing evidence of improvement in core ASD symptoms (Kolevzon 
et al., 2014) using the Aberrant Behavior Checklist (ABC; Aman et al., 
1985) and the Repetitive Behavior Scale-Revised (RBS-R; Bodfish et al., 
2000). IGF-1 also reversed phenotypic and electrophysiological changes in 
human neuronal models of PMS (Shcheglovitov et al., 2013) and iASD 
(Marchetto et al., 2016), providing additional support for this intervention.
IGF-1 enters the brain from the circulation where it is released mainly 
by the liver upon growth hormone stimulation. Blood-borne IGF-1 is found 
in the CNS and promotes brain vessel growth (Lopez-Lopez et al., 2004), 
neurogenesis, and synaptogenesis (O’Kusky et al., 2000). Once IGF-1 
binds to the IGF-1 receptor, activation of the PI3K/mTOR/AKT1 and 
MAPK/ERK pathways induces its downstream effects (Costales & 
Kolevzon, 2016). A recent study in Rett syndrome provides evidence of IGF-
1 brain penetrance: cerebrospinal fluid and serum analysis revealed 
significant increases in IGF-1 levels after treatment (Khwaja et al., 2014). 
Because of the exorbitant cost, limited availability due to manufacturing challenges, and 
burdensome safety monitoring required with IGF-1, we performed a pilot study with rhGH to 
demonstrate feasibility in a) elevating IGF-1 levels reflected in peripheral blood assays, and b) 
showing clinical improvement in symptom domains. We enrolled six children (2 males, 4 females) 
between 3.2 to 11.4 years of age in an open-label study of rhGH. At baseline, all children were of 
normal weight (-0.85 to 1.15 SD), height (-1.38 to 0.75 SD) and BMI (-0.82 to 1.27SD). Bone ages 
ranged between -0.5 to +1.5 years and were all within the normal range. Baseline IGF-1 Z-scores 
varied between -2.4 to 2.3 SD. Growth hormone doses were initiated at 0.14 to 0.16 mg/kg/week 
for two weeks and then increased to 0.27 to 0.31 mg/kg/week once tolerated. Further titrations 
were based on IGF-1 Z-scores with a target of 2 SD. During the course of the trial, IGF-1 Z-scores 
increased in all six participants, reaching a maximum of 6 SD, and demonstrating feasibility of this 
approach. Further, using the Wilcoxon signed rank test, rhGH was associated with statistically 
significant improvement in social withdrawal using the Aberrant Behavior Checklist – Social 
Withdrawal subscale (ABC-SW; p = 0.028), hyperactivity using the ABC - Hyperactivity subscale 
(p = 0.027), sensory reactivity using the Sensory Profile (SP; p = 0.042), and global improvement 
using the Clinical Global Impressions Improvement scale (CGI-I; p = 0.024). 
We will use VEPs as our primary electrophysiological outcome. VEPs have already been used 
as outcome measures in clinical trials: gabapentin (Conte et al., 2009) 
and an infant formula (O’Connor et al., 2001) received FDA approval 
based in part on positive findings from VEP studies. In patients with 
epilepsy, augmentation with both gabapentin and vagus nerve 
stimulation were associated with VEP changes that reversed on 
discontinuation of treatment (Conte et al., 2009). These results suggest 
that VEP is sensitive to change with 
treatment and a viable outcome measure 
for clinical trials. 
Our preliminary results suggest that 
VEP amplitude is a robust marker of 
Figure 4. A subset of 50% of iASD patients fall 
within 1 SD of the PMS P60-N75 mean and are 
defined as “PMS-like” for Aim 3.
Figure 5. Improved low gamma 
power with IGF-1 in PMS (bars show
group averages; lines are individual
patients (n=6; p=.048). 
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Electrophysiological markers in PMS and iASD
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glutamatergic dysfunction in PMS and that approximately 50% of children with iASD display deficits 
in the glutamatergic P60-N75 VEP response (Fig.4) akin to those seen in PMS. Given that VEP 
amplitude is associated with sensory symptoms and that IGF-1 improves clinical symptoms of PMS 
(Kolevzon et al., 2014), we hypothesize that P60-N75 VEP can be used as a stratification 
biomarker to predict which individuals with iASD have alterations in neural pathways 
overlapping with PMS and are therefore likely to respond to rhGH, a treatment that 
stimulates the release of IGF-1, and consequently, glutamatergic pathways. Data from 
ongoing clinical trials at our center provide preliminary support for the use of transient VEPs as a 
biomarker of target engagement and treatment response. We piloted VEPs in a subset of patients 
in the context of our IGF-1 clinical trial in PMS. Results from frequency-domain analyses using 
MSC statistics indicated that IGF-1 consistently increased low gamma power (Fig.5) as measured 
by change in the band which includes frequencies from 30-36 Hz. There was a significant increase 
in low gamma MSC following IGF-1 relative to baseline (n=6; p=.048); when data were inspected 
individually for consistency of directional trends, MSC increased in 5 of 6 patients (Fig. 5). Increase 
in high frequency responses from baseline reflects stronger, more consistent excitatory 
contributions of the VEP, confirming glutamatergic pathway enhancement as a result of treatment, 
and suggesting that MSC in the low gamma range represents an ideal biomarker for target 
engagement and treatment response for rhGH in a 12-week trial.  
Critically, we also saw clinical improvement in sensory hyporeactivity on the SAND with IGF-1 
in the same six patients (p=.037; Fig.6), which itself is significantly correlated with VEP response. 
Given that a greater number of visual hyporeactivity symptoms is significantly correlated with 
smaller P60-N75 (r=-.561, p=.024), in combination with our results 
showing treatment response on both measures, VEPs and the SAND 
may represent robust electrophysiological and behavioral measures for 
evaluating effects of rhGH. 
Significance of the Expected Research Contribution. 
Through completion of Aims 1 and 2, we have: 1) validated the ASD phenotype within a sample 
of individuals with PMS, 2) established electrophysiological biomarkers of PMS, and 3) used 
biomarkers associated with PMS to identify an overlapping subset in iASD. In Aim 3, we will 
evaluate the feasibility of our biomarkers in a clinical trial with rhGH. Results will demonstrate 
evidence of our biomarkers’ utility for: 1) stratifying subsets within iASD, 2) detecting target 
engagement with rhGH via neural responses, and 3) predicting treatment responders based on 
electrophysiological profiles. We have demonstrated feasibility of our biomarkers in individuals with 
ID who are minimally verbal, and this study will specifically focus on this underserved population 
in ASD research. Upon successful completion of the proposed research, we expect to clarify an 
underlying neural mechanism in PMS common to subsets of iASD, validate methods to predict 
treatment response for future clinical trials, and demonstrate efficacy of rhGH for individuals with 
PMS and iASD with glutamatergic dysfunction. 
Innovation
To date, the development of treatments in ASD has mainly relied on strategies only loosely 
related to what is known about the neurobiology, using etiologically heterogeneous samples, and 
delivering intervention broadly and with mixed success. More recently, genetic discovery and 
model systems have elucidated the neurobiology of several subtypes of ASD, including PMS, and 
led to opportunities for developing biomarkers and novel therapeutics. This proposal is innovative, 
in our opinion, on several technical and conceptual levels. First, this study will leverage existing 
work in animal models of PMS and include electrophysiological tests. Second, we have the largest 
known cohort of individuals with PMS and have identified both a unique clinical phenotype and a 
striking VEP deficit that characterizes this population. Thus, we are uniquely situated to 
systematically assess the reliability of both clinical and electrophysiological measures in PMS. 
Third, the phenotype and electrophysiological profile identified in PMS is critically relevant to ASD. 
We will use the knowledge gained in an ASD-related syndrome to inform stratification of a subset 
Figure 6. Children with PMS show significant 
improvement in sensory symptoms following 12 
weeks of treatment with IGF-1. Greatest improvement
was noted in the hyporeactivity domains (n=6; 
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Electrophysiological markers in PMS and iASD
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of iASD with constrained heterogeneity based on a biological marker. Fourth, given our strong 
preliminary data in PMS and iASD and existing infrastructure and pipeline for conducting clinical 
trials, we will test the feasibility and robustness of electrophysiological markers in the context of 
treatment with rhGH, a step which has yet to be taken successfully in the ASD literature. Fifth, we 
expect to predict a subset of iASD with high potential to respond to treatment with rhGH based on 
biological profiles related to PMS, making progress toward a more personalized medicine approach 
for ASD. 
Approach 
Aim 3: Evaluate the feasibility of electrophysiological markers as a measure of treatment 
response.
Introduction. The objective of this aim is to administer rhGH to participants with PMS and 
iASD to evaluate the feasibility of select electrophysiological biomarkers to: 1) detect change in 
neural response (engagement) and 2) predict treatment responders on clinical assessments 
(efficacy). We will also explore the relationship between rhGH dose, IGF-1 level, and 
electrophysiological response. Participants will be selected based on electrophysiological profiles 
with reduced P60-N75 VEP amplitudes, reflecting impaired glutamatergic function. Approach: We 
will employ a randomized, placebo-controlled, double-blind, crossover design with a four-week 
wash-out period between phases. Rationale: Successful completion of this aim will establish 
feasibility of select biomarkers for detecting treatment response and advance knowledge about 
developing targeted treatments for PMS and related subsets of iASD with impaired glutamatergic 
activity. We expect, based on preliminary data, that the knowledge gained will provide evidence of 
targeted brain engagement with rhGH, a mechanistic pathway, and evidence of clinical efficacy. 
Research Design. The proposed intervention will recruit 15 children with PMS and 15 children 
with iASD (2-12 years old). Treatment will follow a randomized, placebo-controlled, crossover 
format with 12 weeks in each treatment phase (rhGH and placebo). This duration was selected 
based on evidence with IGF-1 in PMS suggesting our clinical outcomes will be sensitive to change 
within this time frame (Kolevzon at al., 2014). The primary electrophysiological outcomes will be 
VEP gamma band activity as our preliminary data with PMS in IGF1 suggests it is sensitive to drug 
effects (Fig. 5). The primary clinical outcome will be the SAND because: a) our preliminary studies 
suggest improvement on this measure following IGF-1 (Fig. 6), b) it accurately reflects a sensory 
phenotype characteristic of PMS and is relevant to a substantial subset of iASD, c) it is associated 
with electrophysiological measures, and d) it has been validated in ASD and ID. 
Inclusion criteria: All participants with PMS will have pathogenic SHANK3 deletions or 
sequence variants confirmed in clinical genetics laboratories approved by the Clinical Laboratory 
Improvement Amendments (CLIA). Genotyping will also be used to confirm the absence of known 
pathogenic copy number variants in the iASD group. Participants with iASD will meet gold standard 
diagnostic criteria for ASD, confirmed by the Autism Diagnostic Observation Schedule – 2nd Edition 
(ADOS-2; Lord et al., 2012) and the Autism Diagnostic Interview-Revised (ADI-R; Lord et al., 1994). 
All iASD subjects will have a reduced neural response defined by P60-N75 amplitudes within one 
standard deviation of the PMS mean. Subjects will be on stable medication and psychosocial 
treatment regimens for at least three months prior to enrollment. 
Exclusion criteria: 1) closed epiphyses; 2) active or suspected neoplasia; 3) intracranial 
hypertension; 4) hepatic insufficiency; 5) renal insufficiency; 6) cardiomegaly/valvulopathy; 7) 
allergy to growth hormone or any component of the formulation; 8) comorbid conditions which 
prevent participation; 8) visual problems which preclude use of VEP. 
Drug: Growth hormone is secreted by the somatotrophs of the anterior pituitary gland in a 
pulsatile manner. It binds to its receptor, signaling a cascade of intracellular events which directly 
stimulated the synthesis and release of IGF-1 (Ashpole et al; 2014, Grimberg et al; 2016). Growth 
hormone is FDA approved for use in children with growth hormone deficiency (including idiopathic 
growth hormone deficiency), Turner syndrome, Noonan syndrome, Prader-Willi syndrome, short 
stature homeobox-containing gene (SHOX) deficiency, chronic renal insufficiency, idiopathic short 
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Electrophysiological markers in PMS and iASD
PI: Kolevzon
9
stature 
and 
children 
small 
for 
gestational 
age 
(https://www.fda.gov/Drugs/DrugSafety/ucm237839.htm). 
Drug Administration: Growth hormone is commercially available for subcutaneous injection as 
a 10 mg vial to be reconstituted with in-package syringe of 1 mL of bacteriostatic water (preserved 
with 0.33% metacresol) and a 25G reconstitution needle. We have previously received an IND 
exemption from the FDA to conduct trials in PMS (#142585) and will submit this revised protocol 
for an exemption in ASD. Growth hormone will be administered subcutaneously once daily. 
Approved doses range from 0.16 - 0.25 mg/kg/week for growth hormone deficient patients 
(Grimberg et al.; 2016) and up to 0.47 mg/kg/week for patients born small for gestational age. In 
our proposed trial, we expect a majority of our patients to be growth hormone sufficient and 
therefore similar to the population of children with idiopathic short stature. We will use a starting a 
dose of 0.15 mg/kg/week divided daily for 2 weeks to ensure safety and tolerance. The dose will 
then be increased to 0.3 mg/kg/week for 10 weeks, an average growth hormone dose in children 
with idiopathic short stature. Dose will be titrated based on IGF-1 levels monitored every four weeks 
up to a maximum dose of 0.45 mg/kg/week based on the package insert. Drug dose is calculated 
on a mg/kg/week basis. Using Center for Disease Control average Weight-for-age statistics in 
typically developing children ages 2 to 12 years-old (www.cdc.gov/growthcharts/cdc_charts.htm), 
the overall average weight across a representative sample of males and females in this age range 
is 24.65kg. Therefore, average growth hormone dose for weeks 1-2 (0.15 mg * 24.65 kg per week) 
is 3.7 mg per week, and for weeks 3-10 (0.3 mg * 24.65 kg per week), is 7.4 mg per week. We 
anticipate each participant to require about 81.4 mg of growth hormone for 12 weeks, a total of 
2442 mg for 30 participants, or 250 vials (10 mg per vial).
Outcome measures: The primary electrophysiological outcome will be VEP gamma band 
activity as measured by MSC, which includes frequencies between 30-36 Hz. The secondary 
electrophysiological outcome will be VEP P60-N75 amplitude. VEP stimuli, data preprocessing, and 
variable extraction. The primary stimulus condition will be a contrast-reversing checkerboard 
(100% contrast) of one-minute duration to elicit a transient VEP. VEPs will be analyzed in both time 
and frequency domains. Waveforms will be standardized into z-scores, segmented to evoking 
stimuli, filtered 1-100Hz with a 60Hz notch, and averaged across epochs. For time-domain 
analyses, VEP amplitudes over occipital cortex (Oz) will be measured peak-to-trough (P60-N75, N75-
P100) and latency (P60, N75, P100) will be measured by time-to-peak.  Frequency-domain analyses 
will be conducted using MSC statistics (Zemon et al., 2009, Siper et al. 2016) to quantitatively 
assess the integrity of overall responses in different frequency bands. MSC reflects response 
reliability by providing an estimate of signal power relative to the combined power of signal+noise. 
As such, MSC measures consistency from one trial to the next at a given frequency. A pure signal 
produces a value of 1 and no signal produces a value of approximately 0.1 (bias level for pure 
noise); thus, higher MSC values reflect stronger and more consistent oscillatory activity in a given 
frequency band. Six distinct frequency mechanisms for the tVEP waveform include: Band 1, 6-10 
Hz, Band 2, 12-28 Hz, Band 3, 30-36 Hz, Band 4, 38-48 Hz, Band 5, 50-64 Hz (60 Hz electrical 
noise omitted), Band 6, 66-84 Hz (Siper et al., 2016; Zemon et al, 2009, Zemon & Gordon, in prep.). 
These MSC frequency bands meet criteria for reliable components regardless of sample size as 
demonstrated by Monte Carlo computer simulations (Guadagnoli & Velicer, 1988; Stevens, 1996). 
Based on preliminary data, we expect both P60-N75 amplitude and MSC values in bands 
encompassing gamma wave activity (Bands 3 and 4) to be decreased in PMS relative to iASD.
Primary clinical outcome: SAND hyporeactivity subscale (Siper et al., 2017). Secondary and 
exploratory outcomes targeting ASD symptom domains: ABC-SW (Aman et al., 1985) and other 
measures employed in Aim 1 (Table 1) based on results from Aim 1. Clinical outcomes will be 
measured by an independent evaluator who is blind to side effects to prevent the risk of bias, and, 
within 
subjects, 
the 
same 
evaluator 
will 
complete 
assessments 
across 
all 
visits. 
Electrophysiological markers and clinical measures will be collected at baseline, weeks 4, 8, and 
12 of each phase. Safety and tolerability will be measured after two weeks in each phase and then 
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Electrophysiological markers in PMS and iASD
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10
every four weeks thereafter during monitoring visits and phone calls using an adapted semi-
structured interview, the Safety and Monitoring Uniform Report Form (SMURF); extensive clinical 
and 
laboratory 
assessments will occur every four weeks during visits (see Protection of Human Subjects). 
Language testing will include standardized measures reliant on caregiver report and objective 
standardized assessments. Specifically, caregivers will report on a child’s expressive and receptive 
verbal and non-verbal communication repertoire on the MacArthur-Bates Communication 
Development Inventory (MCDI; Fenton et al., 2007), which is a valid assessment of young 
children’s language (Charman et al., 2003; Luyster et al., 2007). Although not normed in children 
above 30 months of age, preliminary data collected on this measure suggest that raw scores on 
the MCDI are useful in capturing the range of language functioning in our minimally verbal and 
intellectually disabled sample. The MSEL and Vineland will provide standardized assessments of 
receptive and expressive language, by direct assessment and parent interview, respectively. 
The repetitive behavior domain will be evaluated using the RBS-R (Bodfish et al., 2000) and 
the ABC-Motor Stereotypies subscale. Sensory sensitivity will be measured using the SP (Dunn, 
1999) and the SAND (Siper et al., 2017). The SP is a widely used caregiver questionnaire with 125 
items investigating daily life sensory experiences. The SAND includes both a clinician-administered 
observation, involving standardized presentation of 15 sensory stimuli, with 5 sensory toys per 
modality (visual, auditory and tactile), and a corresponding caregiver interview. Behavioral 
responses are rated by a trained examiner on an algorithm measuring specific, discrete sensory 
hyperreactivity, hyporeactivity, and seeking behaviors across domains. The corresponding 
caregiver interview consists of 36 items and indicates whether a given sensory behavior is present 
(1) or absent (0). For any domain with behaviors coded “present,” caregivers rate the severity of
symptoms within that domain. Total SAND scores are based on the combination of the observation
and interview, including scores by sensory modality and by DSM-5 symptom domain
(hyperreactivity, hyporeactivity, seeking). Domain scores range from 0-30 and higher scores
represent a higher level of sensory symptoms.
Behavioral assessments will be performed using the Aberrant Behavior Checklist (ABC; Aman 
et al., 1985) as a clinical outcome measure for Aim 3. The ABC is a parent checklist that has been 
well validated in children with ASD and ID and is frequently used as an outcome measure in ASD 
clinical trials. As part of studies in Aim 1, we compared 25 PMS patients with ID 57 participants 
with iASD and ID and found no significant differences between groups on any of the ABC Lethargy, 
Stereotypies, or Hyperactivity subscales. These results highlight commonalities across the PMS 
and broader iASD behavioral phenotype and support the ABC as a relevant measure. 
Data Analysis. Following the procedures of a 2x2 cross-over design, each subject is 
randomized to one of two treatment sequences. First we will use graphical and numerical statistics 
to characterize treatment-, period- and cross-over effects (Jones & Kenward, 1989). Under the 
assumption of no cross-over effects and data following an approximate normal distribution, we will 
fit generalized linear mixed models (GLMMs) to the data examining the effects of group (PMS, 
iASD), condition (placebo, drug), time (baseline, week-12). Interactions on clinical and VEP 
measures will be checked for differential treatment effects, including subject level random 
Table 1: Clinical Outcome Measures
Domain
Instrument 
Measurement modality
Social
Aberrant Behavior Checklist - Social Withdrawal Subscale
Caregiver report
Language 
/Communication
MacArthur-Bates Communication Development Inventory
Vineland Adaptive Behavior subscales
Mullen Scales for Early Learning subscales
Caregiver report
Caregiver report
Direct assessment
Repetitive Behavior
Repetitive Behavior Scales-Revised 
Aberrant Behavior Checklist - Motor Stereotypies Subscale
Sensory Profile
Sensory Assessment for Neurodevelopmental Disorders
Caregiver report
Caregiver report
Caregiver report
Direct assessment
Adaptive Behavior
Vineland Adaptive Behavior Scales-3
Caregiver report
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Electrophysiological markers in PMS and iASD
PI: Kolevzon
11
intercepts, to control for within subject correlations. Next, we will add baseline VEP gamma band 
activity, along with its interactions, to the models for clinical outcomes. In particular, a VEP by group 
by time interaction would suggest that the electrophysiological marker is predictive of differential 
treatment response. We will repeat these analyses with secondary electrophysiological markers 
and any derived composites. To examine whether electrophysiological markers track changes in 
clinical profiles, we will fit an additional GLMM with VEP values included as a time-varying rather 
than baseline covariate. If treatment effects become less significant it will suggest that rhGH is 
acting on clinical outcomes through the biomarker. Finally, we will directly correlate week-12 
change scores for EEG and clinical measures. In case of major cross-over effects, we will base 
our primary analysis on phase 1 data only as a parallel group study. All tests will use a two-sided 
α of .05. 
Power Analysis: Power is calculated for drug group differences in baseline to week 12 change 
scores in VEP gamma power since we are primarily interested in an overall treatment effect rather 
than a differential effect by group. These correspond to paired t-tests, taking advantage of the 
crossover design. With α at .05, we have 80% power to detect moderate to large effects (d=.77 
within each group, d=.53 across groups), a reasonable expectation based on the potentially 
disease modifying effects of IGF-1 and our pilot study, which found a large effect (d=1.05) of IGF-
1 in PMS on a different primary outcome (ABC-SW). 
Potential pitfalls and alternative strategies. We expect loss of some electrophysiological 
data due to the severely affected nature of the population; however, this study is well powered and 
our proposed sample size can accommodate attrition and still detect meaningful differences across 
a range of responses. We have been successful in recording VEP in more than 70% of patients 
with PMS seen in our center. We will implement a sensory desensitization and behavioral training 
protocol and have extensive experience collecting these data from children with ASD, PMS, and 
severe ID. Moreover, for the VEP, short-duration stimulus conditions are being utilized. In addition, 
crossover trial designs carry risks of contamination effects, so we have provided a four-week 
washout period to minimize this risk. We have not observed significant carryover effects in our 
preliminary data. We believe this design is important because all patients will be afforded the 
opportunity to receive active medication. Second, the burden of frequent monitoring visits and 
subcutaneous injections risks increase the risk of subject withdrawal. Given the rigid control of the 
study procedures, relatively simple measurement techniques, and based on our experience to 
date, we do not foresee any major loss of data due to drop-outs or missing values. Still, in case of 
drop-outs we will use the intent to treat principle so all randomized patients will be included in the 
data analysis. Third, because of concerns about recruitment feasibility, we have established an 
excellent working relationship with the national PMS Foundation. Fourth, because we are only 
enrolling patients with iASD with PMS-like VEP amplitudes, we will not be able to test the specificity 
of target engagement and successful treatment response. Finally, if VEPs cannot be established 
as a biomarker for stratification or efficacy, we still have a wealth of other electrophysiological and 
clinical outcomes that will provide critical information. 
Expected outcomes. We expect to provide evidence for the feasibility of using VEPs to detect 
neural change related to glutamatergic activity in response to treatment with rhGH. Specifically, we 
expect rhGH will be associated with increased low gamma band activity as compared to placebo 
in both PMS and the PMS-like iASD subset. We further anticipate enhanced gamma band activity 
to be associated with changes on clinical outcome measures, particularly those that reflect sensory 
reactivity given our preliminary data. Finally, we expect to demonstrate that rhGH treatment will 
result in changes on primary and secondary outcomes of sensory reactivity, social impairment, 
repetitive behavior, and functional outcomes of adaptive behavior. If positive, in addition to 
demonstrating efficacy of rhGH for PMS, results from this clinical trial will demonstrate the feasibility 
of using electrophysiological markers to stratify patients with iASD and identify early biomarkers of 
potentially disease-modifying effects on core symptoms of ASD.
Timeline
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Electrophysiological markers in PMS and iASD
PI: Kolevzon
12
It is anticipated that 30 months will be required to complete Aim 3. Months 0-4 will be dedicated 
to study startup, regulatory, and Review Board approval. Months 5-20 will be required to enroll 30 
participants (15 PMS; 15 iASD) at a rate of 1 participant per group per month. Months 21-27 will 
be required to complete data collection and months 28-30 will be reserved for data analysis. This 
timeline is considered feasible based on our experience and assessment of current interest among 
families on our waiting list and registered with the PMS Foundation. 
Protection of Human Subjects
Risks to the Subjects 
Side effects of recombinant human growth hormone (rhGH) include: headaches, muscle pain, 
extremity stiffness, and swelling of arms and legs due to retaining water in body. Less common 
side effects include bruising, lumps under the skin, gynecomastia (enlarged breast tissue in 
males), progression of previously present curvature of the spine, resistance to insulin hormone 
and low thyroid hormone levels, allergic reaction (such as hives, itching, breathing difficulty, 
swelling of face/lips), slipped capital femoral epiphysis (a fracture through the growing part of the 
bone at the hip), and benign intracranial hypertension (increased pressure in the brain with 
headaches and blurring of sight). Cancer has been found to be associated with long-term 
treatment with rhGH only in children with a previous history of cancers or with certain syndromes 
or genetic conditions which are predisposed to cancers. Because rhGH is administered 
subcutaneously, there is risk of pain and irritation at the injection site. There is also risk of 
inflammation and increased growth of fat cells in the skin at the injection site. It is recommended 
to alternate injection sites (upper arm, thigh, buttock or abdomen) with each injection to reduce 
these risks. 
In terms of somatic effects, rhGH is expected to have significant positive effects on growth. 
However, growth improvement is not the goal of therapy in this study. Moreover, the rate of growth 
in subjects who have an exuberant response is unwanted. Anthropometric measurements, 
including height, weight, body mass index (BMI) and head circumference will be collected 
throughout the study. Growth will be defined as the height velocity and change in height SD score 
during treatment.   
Another risk to participating in this project is the blood drawing procedures. The risks of a blood 
draw include pain, bruising, and the slight possibility of infection at the needle insertion site. 
Some children may also feel dizziness following the blood draw. Blood sampling may be 
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Electrophysiological markers in PMS and iASD
PI: Kolevzon
13
particularly uncomfortable for children with developmental disabilities and will be kept to a 
minimum to reduce risks. The procedures for minimizing risk include assisting parents in keeping 
their child still, the use of a special lounge chair to help the child relax during the procedure, the 
use of numbing cream if needed to reduce pain, and the use of professionals in the Mount Sinai 
Clinical Research Center who are trained and certified in blood drawing procedures in children 
with developmental disabilities.
The risks in participating in the electrophysiology aims of this study are minimal. The physical 
contacts with the instruments are limited to the standard EEG electrodes placed on the scalp. 
Electrodes are disinfected between participants. It is possible that as a result of participating in 
electrophysiological testing, participants may experience fatigue or anxiety. We will attempt to 
minimize potential discomfort from testing/evaluation by providing frequent breaks during the visits. 
If the participant finds any discomfort during the EEG intolerable, he or she will be withdrawn from 
the study. If the participant feels uncomfortable, the participant or his/her parents can tell the 
investigators and ask to stop the EEG at any time. We will implement a sensory desensitization 
and behavioral training protocol in which participants will have the opportunity to practice mock 
EEG procedures over the course of visits. Participants will also receive a social story describing 
the components of their visit as needed. Visual schedules and positive reinforcement systems (e.g., 
token economies) will be used throughout testing.
A Data and Safety Monitoring Board (DSMB) has been convened and has primary responsibility 
for developing the oversight necessary to promptly identify and act upon any adverse events. The 
DSMB is made up of three physicians who will meet routinely every six months and immediately 
should a severe adverse event occur (please see attached DSMB Charter). 
Adequacy of Protection Against Risks 
Recruitment and Informed Consent: 
The investigator will describe the protocol to potential subjects’ parents/guardians in person, 
although general information and assessment for eligibility can be carried out by phone if 
necessary. The Informed Consent may be read to the subjects’ parent/guardians, but, in any event, 
the investigator or designee shall give the subjects’ parents/guardians ample opportunity to inquire 
about details of the study and ask any questions before dating and signing the Informed Consent 
Form. The Informed Consent will be created with a level of language fully comprehensible to the 
prospective subjects’ parents/guardians. Informed consent will be documented by the use of a 
written consent form approved by the IRB and signed and dated by the subjects’ parents/guardians 
and by the person who conducted the informed consent discussion. The signature confirms the 
consent is based on information that has been understood. Each subject’s signed informed consent 
form will be kept on file by the investigator for possible inspection by regulatory authorities. The 
parents/guardians will receive a copy of the signed and dated written informed consent form and 
any other written information provided to the subjects’ parents/guardians, and will receive copies 
of any signed and dated consent form updates. Any amendments to the written information will be 
provided to parents/guardians. 
Assent:
Our preliminary phenotyping studies and clinical trials that include characterizing cognitive and 
adaptive functioning in Phelan-McDermid syndrome (PMS) suggest that these children have 
severe to profound intellectual disability (ID). As such, these patients do not have the capacity to 
provide assent. We will specifically be recruiting patients with idiopathic autism spectrum disorder 
(iASD) who are also intellectually disabled and anticipate that they will not have capacity to provide 
assent. 
Ethics and Regulatory Considerations: 
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Electrophysiological markers in PMS and iASD
PI: Kolevzon
14
The study will be conducted according to Good Clinical Practice (GCP), the 1996 Declaration of 
Helsinki, and local rules and regulations of the United States. 
Confidentiality of Source Documents and Study Data: 
A subject identification code will be used in lieu of the subject’s name on all study data compiled 
and delivered to the secure database. All source documents and study data will be kept 
confidential, in accordance with all requirements of the laws. 
Adverse Events 
Definition of an Adverse Event
An adverse event (AE) will be defined as any untoward medical occurrence in a study subject, 
temporally associated with the use of the experimental medication, whether or not considered 
related to the medication. An adverse event can therefore be any unfavorable and unintended sign 
(including an abnormal laboratory finding), symptom, or disease (new or exacerbated) temporally 
associated with the use of the experimental medication. A serious adverse event (SAE) will be 
defined as an AE that meets any of the following criteria:

results in death;

is life threatening;

requires inpatient hospitalization;

results in a persistent or significant disability/incapacity;

any other adverse event that, based upon appropriate medical judgment, may jeopardize
the subject’s health and may require medical or surgical intervention to prevent one of the
outcomes listed above.
Adverse Events Reporting 
The Principal Investigator (PI) will be responsible for the detection and documentation of events 
meeting the criteria and definition of an adverse event or serious adverse event, as provided in this 
protocol. During the study, when there is a safety evaluation, the investigator or team member will 
be responsible for reporting adverse events and serious adverse events. Each subject’s 
parents/guardians will be instructed to contact the investigator immediately should the subject 
manifest any signs or symptoms they perceive as serious. 
Clinical laboratory parameters and other abnormal assessments qualifying as adverse events and 
serious adverse events 
Abnormal laboratory findings or other abnormal assessments that are judged by the investigator to 
be clinically significant will be recorded as AEs or SAEs if they meet the definition of an AE. 
Clinically significant abnormal laboratory findings or other abnormal assessments that are detected 
during the study or are present at baseline and significantly worsen following the start of the study 
will be reported as AEs or SAEs. The investigator will exercise his or her medical and scientific 
judgment in deciding whether an abnormal laboratory finding or other abnormal assessment is 
clinically significant. 
Medically attended visits 
For each AE the subject experiences, the subject’s parents/guardians will be asked if they received 
medical attention defined as hospitalization, an emergency room visit or a visit to or form medical 
personnel (medical doctor/doctor of osteopathy or nurse practitioner) for any reason. This 
information will be recorded in the CRF. 
Lack of Efficacy 
Lack of efficacy per se will not be reported as an AE. The signs and symptoms or clinical sequelae 
resulting from lack of efficacy will be reported if they fulfill the AE or SAE definition. 
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Electrophysiological markers in PMS and iASD
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15
Time Period, Frequency, and Method of Detecting AEs 
All AEs occurring from the initiation of therapy until completion will be recorded on the Adverse 
Event form in the subject’s CRF, irrespective of severity or whether or not they are considered 
medication-related. Onset of chronic illness (e.g. autoimmune disorders, asthma, type 1 diabetes 
and allergies) and conditions prompting emergency room (ER) visits or physician office visits that 
are not related to well-child care, injury, or common acute illnesses (e.g., upper respiratory tract 
infection, otitis media, pharyngitis, and gastroenteritis) will be reported during the entire study 
period. The investigator will inquire about the occurrence of AEs at every visit/contact during the 
study as appropriate. All AEs either observed by the investigator or a clinical collaborator or 
reported by the subject’s parent/guardian spontaneously or in response to a direct question will be 
evaluated by the investigator. AEs not previously documented in the study will be recorded in the 
Adverse Event form within the subject’s CRF. The nature of each event, date and time (where 
appropriate) of onset, outcome, intensity and relationship to drug administration should be 
established. Details of any corrective treatment should be recorded on the appropriate page of the 
CRF. 
When an AE/SAE occurs, it will be the responsibility of the investigator to review all documentation 
(e.g., hospital progress notes, laboratory, and diagnostic reports) relative to the event. The 
investigator will then record all relevant information regarding an AE/SAE on the CRF or SAE 
Report Form as applicable. The investigator will attempt to establish a diagnosis of the event based 
on signs, symptoms, and/or other clinical information. 
Follow-up of Adverse Events and Serious Adverse Events and Assessment of Outcome:
After the initial AE/SAE report, the investigator will actively follow each subject and provide further 
information on the subject’s condition. All AEs and SAEs documented at a previous visit/contact 
and designated as not recovered/not resolved or recovering/resolving will be reviewed at 
subsequent visits/contacts. Investigators will follow-up subjects with SAEs or subjects withdrawn 
from the study as a result of an AE, until the event has resolved, subsided, stabilized, disappeared, 
the event is otherwise explained, or the subject is lost to follow-up. In the case of other non-serious 
AEs, until they complete the study or they are lost to follow-up. Clinically significant laboratory 
abnormalities will be followed until they have returned to normal, or a satisfactory explanation has 
been provided. The investigator may perform or arrange for the conduct of supplemental 
measurements and/or evaluations to elucidate as fully as possible the nature and/or causality of 
the AE or SAE. 
Regulatory Reporting Requirements for Serious Adverse Events:
All safety data will be reported to Safety Monitoring Board and IRB every six months, or, in the 
case of any major safety concern or question, immediately.  If any study stopping condition occurs, 
this will be reported immediately, and the study will be halted, pending review by these agencies. 
The investigator has a legal responsibility to promptly notify, as appropriate, both the local 
regulatory authority and other regulatory agencies about the safety of a product under clinical 
investigation. Prompt notification of SAEs by the investigator to the Study Contact for Reporting 
SAEs is essential so that legal obligations and ethical responsibilities towards safety of other 
subjects are met. The investigator, or responsible person according to local requirements, will 
comply with the applicable local regulatory requirements related to the reporting of SAEs to 
regulatory authorities and the IRB. The protocol will be filed under Investigational New Drug (IND) 
exemption status with the US FDA. 
Safety Measures
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Electrophysiological markers in PMS and iASD
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16
Participants will undergo comprehensive medical evaluation by the study physicians. Medical 
history, family history, physical and neurological examination (including anthropometric 
measurements and fundoscopic exams), routine hematology, thyroid function, and blood chemistry 
(including liver profile), bone X-ray for bone age, and electrocardiography will be performed to 
determine eligibility for participation. Patients will be monitored for safety and tolerability at weeks 
2, 4, 8, and 12 in both treatment phases – see Table 1 and 2.   There are a number of tertiary 
outcomes designed to monitor the safety of rhGH (see Tables 1 and 2) and will be reported to the 
DSMB (see Stopping Conditions). 
Our preliminary phenotyping studies characterizing cognitive and adaptive functioning in Phelan-
McDermid syndrome suggest that these children and adults have ID. This sample was significantly 
delayed in receptive language, expressive language, and overall functional communication skills. 
We will specifically be recruiting children with iASD who also have ID. As such, it is impossible to 
directly query these children using verbal methods with respect to safety and tolerability. Instead, 
we rely on parent report using open-ended methods and a modified SMURF, in addition to physical 
and neurological exams, and laboratory measures.
Monitoring for AEs will be conducted during scheduled and unscheduled visits per clinical and 
laboratory assessments. If a subject develops significant neurological signs and symptoms (e.g.  
cerebrovascular event), they will be seen immediately for a comprehensive evaluation, appropriate 
treatment, and removal from active participation in the study. We will also apply criteria for 
attribution of AEs by means of the following descriptors and codes.

unrelated to treatment
1

unlikely related to treatment
2

possibly related to treatment
3

probably related to treatment
4

definitely related to treatment
5
Stopping Conditions for Individual Subjects
The following criteria will be used to identify possible adverse treatment events, which will indicate 
the need to halt active participation of the subject in the rhGH study

Withdrawal of Consent

PI or any regulatory authority (Safety Monitoring Board, IRB) believe withdrawal is
necessary for the subject’s health, well-being, or best interests.

PI or any regulatory authority believe that withdrawal is necessary for the subject’s health,
well-being, or best interests

Laboratory: any abnormality on any test with adverse event (AE, Common Terminology
Criteria for Adverse Events, National Cancer Institute, scales) ≥ 3 or greater at any time in
the study

Any AE of any sort (clinical, laboratory) ≥ 4 will result in halting for the individual and also
for the study as a whole.
To avoid bias, all analyses will include all subjects, including those withdrawn from the study, 
regardless of adherence to study protocol.  Especially in an exploratory trial of a novel, unproven 
treatment, the safety of subjects is our primary overriding concern.  If there is doubt concerning a 
subject’s safety, the default mode will be withdrawal from treatment or active study participation, 
followed by close observation (safety follow up visits) and recommendation of standard treatment.
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Electrophysiological markers in PMS and iASD
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Return of a subject to active study participation will not be permitted, except if a transient clinical 
or laboratory abnormality unrelated to study treatment has occurred and subsequent permission of 
the Safety Monitoring Board to return the patient to active study has been provided.
If a subject is withdrawn from active study during screening or observation phases of the study, 
they will be returned to the referring physician and standard care will be recommended; in addition 
we will request the subject’s participation in an end-of-study visit.
If a subject is withdrawn from active study during rhGH treatment, they will also be returned to the 
referring physician and standard care will be recommended; however, in this case, we will in 
addition request the subject’s participation in clinical and laboratory safety assessments and follow 
up per protocol schedule.  
Stopping Conditions for Study as a Whole
The study will be halted* if two patients experience stopping conditions.  (Exceptions for this 
criterion: Patients who withdraw voluntarily for reasons not directly related to or intrinsic to the 
study, e.g. incidental considerations such as concerns about travel time to study visits, unexpected 
pregnancy in the family, etc.  Clearly, patients who experience adverse effects during rhGH 
treatment would count towards this criterion for whole study stopping).
In addition, the study will be halted if one patient experiences a serious related adverse effect 
(grade ≥ 4 AE).
All safety data will be reported to the DSMB every six months and to the IRB annually, or, in the 
case of any major safety concern or question, immediately.  If any study stopping condition occurs, 
this will be reported immediately and the study halted, pending review by the DSMB and IRB, and 
until the decision by regulatory authorities to resume, suspend or close the study has been made.
*Operationally, “halting” will ordinarily mean that no further screening of new subjects and no
treatment initiation will occur until the safety issue has been investigated and resolved (i.e., a final
decision has been made to resume, suspend or close the study has been made).  Enrolled study
participants who have no new symptoms or adverse effects will ordinarily be allowed to continue
study observation or treatment without interruption while the safety issue is being investigated,
unless it is the contemporaneous judgment of the PI or subsequent judgment of the Safety
Monitoring Board or IRB that it is unsafe to do so, in which case all observation or treatment
interventions will be suspended forthwith.
Effective Date: 10/1/2021
End Date:9/27/2022


Tables
Table 1. Safety and Screening Measures
Measures* 
Baseline
Week 4
Week 8
Week 12
Physical and neurological exam 
X
X
Fundoscopy
X
X
Electrocardiography
X
X
Bone X-ray for bone age
X
X
Pregnancy Test
X
X
Height
X
X
Weight
X
X
Head circumference
X
X
Side effect monitoring**
X
X
X
X
*Safety measures will be performed at these time points in both phases of the crossover trial.
**In addition to the above, side effect monitoring will occur at week 2 by phone, or at any point in person as 
needed,  in both phases. 
Table 2. Laboratory Safety Measures
Measures
Baseline*
Week 4*
Week 8*
Week 12*
Electrolytes (Na, K, Cl, CO2) 
X
X
Complete blood count 
X
X
Liver function panel 
X
X
Thyroid function (TSH/T3/T4)
X
X
Renal function (BUN/CR)
X
X
Hemoglobin A1c
X
X
Albumin
X
X
Glucose (fasting)
X
X
Insulin (fasting)
X
X
Lipid profile
X
X
IGF-1 / IGF-BP3
X
X
X
X
*Laboratory safety measures will be performed at these time points in both phases of the crossover trial
Table 3. Dose Titration Schedule
Week
Dose 
Week 1 and 2
0.15 mg/kg/week divided into daily doses
Week 3 to 12
0.30 mg/kg/week divided into daily doses
Effective Date: 10/1/2021
End Date:9/27/2022


Electrophysiological markers in PMS and iASD
PI: Kolevzon
19
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Effective Date: 10/1/2021
End Date:9/27/2022