Efficiency Evaluation of Allogenic Umbilical Cord Blood (UCB) Transfusion in Patients With Autism
Phase: PHASE1
Conditions: Autism Spectrum Disorder, Autism
Sponsor: State-Financed Health Facility "Samara Regional Medical Center Dinasty"

          
«APPROVED» 
The State-Financed Health Facility 
“Samara Regional Medical Center 
Dinasty” 
«AGREED AND APPROVED» 
State budgetary health care institution 
"Samara Regional Children's Clinical 
Hospital named after N.N. Ivanova " 
CEO 
O.V. Tyumina  _____________  
12.11.2018                     
CEO 
O.O.Galahova ________________ 
12.11.2018                 
IRB/IEC APPROVED 
Issue No. 5 dated November 09, 2018 
 
 
 
Efficiency Evaluation of Allogenic Umbilical Cord Blood (UCB) Transfusion 
in Patients With Autism 
 
IND 12112018  
NCT 03786744 
Date of the document 12 of November 2018 
Principal investigator 
O.V. Tyumina , MD 
The State-Financed Health Facility “Samara Regional Medical Center Dinasty”  
159 Tashkentskaya str., Samara, Russian Federation 443095. 
Co-Principal investigator 
O.O.Galahova,  MD 
State budgetary health care institution "Samara Regional Children's Clinical 
Hospital named after N.N. Ivanova " 
165 A Karl Marx st., Samara, Russian Federation 443079 
 

URGENCY OF THE RESEARCH AND LITERATURE REVIEW 
Autism Spectrum Disorders (ASDs) are heterogeneous neurodevelopmental diseases.  The 
most common disorder of the autism spectrum is autism characterized by dysfunctions in 
sympathetic social interaction and communication and the mannerisms. Recent reports indicate a 
sharp increase in the number of children with autism (57% increase compared to the rate of the 
year 2002), while the prevalence rate in the United States is approaching 1%. The exact aetiology 
of autism remains unclear. Therefore, determining effective treatments for autism is particularly 
difficult. 
Although the pathophysiology of autism remains poorly understood, the accumulated data 
suggest that the aetiology may potentially include immune dysregulation (see Onore et al., 2012). 
Extensive research points to immune system abnormalities, including active brain 
neuroinflammation, pro-inflammatory cytokines enhanced profiles, immune cells dysfunction, and 
the autoimmunity directly associated with an increase in behavioural disorders. Studies have 
shown an ongoing neuroinflammatory process with pronounced activation of microglia and 
astroglia in the cerebral cortex, alba and cerebellum of patients with autism. A unique profile of 
pro-inflammatory cytokines in autistic patients was recorded in the cerebrospinal fluid, including 
a marked increase in macrophage chemoattractant protein-1 and in peripheral plasma, such as 
significantly elevated levels of interleukin (IL) -1β, IL -8 and IL-12p40. Additional studies have 
shown altered function in subsets of immune cells (Ashwood P, Corbett BA, Kantor A, Schulman 
H, Van de Water J, Amaral DG. In search of cellular immunophenotypes in the blood of children 
with autism. , Enstrom AM, Lit L, Onore CE, Gregg JP, Hansen RL, Pessah IN, Hertz-Picciotto I, 
Van de Water JA, Sharp FR, Ashwood P. Altered gene expression and function of peripheral blood 
natural killer cells in children with autism. Brain Behav Immun. 2009.) That leads to an 
inappropriate or ineffective immune response in autism. Different "housekeeping" antibodies enter 
into reaction with proteins found in the central nervous system of the children with autism 
associated with an autoimmune process. Taken together, these data suggest that immune 
dysfunction is not only a symptom or associated disease but also an indicant of the underlying 
pathophysiological process, so targeting this pathology and changing neuroimmune reactions can 
be effective from a therapeutic point of view. However, only a few clinical research studies of 
anti-inflammatory drugs are aimed at correcting immune dysregulation or continuing 
neuroinflammation in autism. Due to its known ability to alter immune response, cord blood 
mononuclear cells can be a new therapeutic effect in clinical practice to normalize the immune 
response characteristic of some children with autism. 
There are behavioural, educational, medical, health care and optional methods of autism 
care. However, there is no definitive standard treatment for children with autism. Stem cell therapy 

has shown great promise in modern treatment and rehabilitation of patients with autism. Preclinical 
research studies have reported that transplantation of human cord blood mononuclear cells 
(CBMNC) in animals in models of cerebral ischemia promotes functional recovery by improving 
local blood perfusion in damaged areas through angiogenesis (Park DH, Borlongan CV, Willing 
AE, Eve DJ, Cruz LE, Sanberg CD, Chung YG, Sanberg PR, Cell Transplant, 2009) 
Some studies have confirmed that cerebral hypoperfusion is associated with many of leading 
autism symptoms (Wilcox J, Tsuang MT, Ledger E, Algeo J, Schnurr T. Brain perfusion in autism 
varies with age. Neuropsychobiology. 2002;). Generalized brain hypoperfusion, achieved in the 
frontal and prefrontal areas, was observed in children with autism and is associated with cognitive 
and neuropsychological defects. Besides, a decrease in cerebral perfusion, especially in the 
temporal-parietal areas, is associated with a cognitive deficit, such as speech deficits, cognitive 
disorder or object representation or anomalous perception and responses to sensory stimuli. 
Inadequate perfusion leading to hypoxia of brain tissue had not only caused apoptosis and necrosis 
of neurons but also led to abnormal brain tissue metabolism and accumulation of pathological 
doses of neurotransmitters. As a result of the systemic administration of stem cells CDbrain 
tissue34 + cord blood to overcome ischemia in vitro and in animals, therapeutic angiogenesis has 
been proven experimentally. Also, it was proved that the endothelial progenitor cell contained in 
the population of CD34 + cells had the ability to initiate angiogenesis in ischemic tissues. 
Circulating CD34 + progenitors in the PC with endothelium development potential has turned into 
new endothelial cells to either restore the damaged endothelium wall or germinate a new vascular 
structure. Moreover, human CD34 + cells and hematopoietic progenitors could secrete multiple 
angiogenic factors, such as vascular endothelium growth factor (VEGF), HGF, and insulin-like 
growth factor-1. 
The preclinical research study data on the mononuclear core blood cells ability to stimulate 
neoangiogenesis in ischemic areas allow introducing the method of injecting these cells in cases 
of brain hypoperfusion or hypoxia to limit and reduce the area of ischemic tissue degeneration. 
To assess the therapeutic efficacy of using core blood cells, during the research studies the 
CARS, CGI and ABC scales were used. CARS provides descriptive information about 
pathological behaviour and classifies severity in children with autism, while the CGI scale, as a 
global measure, indicates a noticeable overall treatment effect. However, further research will need 
to extend the rating scales to standardize the diagnosis and treatment of autism in detail. 
In 2011, a Chinese research group from the Shenzhen Beike Bio-Technology Co. conducted a 
scientific clinical research study of the use of intravenous mononuclear cord blood cells in patients 
diagnosed with autism. In total, 37 patients aged 3 to 12 years were tested. There were 3 clinical 
groups: Group 1 consisted of 14 patients receiving the injections of mononuclear cord blood cells 

at a dosage of 2 million cells per 1 kg of patient weight (1 intravenous injection, 3 intrathecal with 
an interval of 5-7 weeks). Group 2 consisting of 9 patients receiving the injections of mononuclear 
cord blood cells at a dosage of 2 million cells per 1 kg of patient weight and mesenchymal stem 
cells at a dosage of 1 million cells per 1 kg of patient weight (2 intrathecal and intramuscular 
injections and 2 intrathecal injections solely). Patients of Groups 1 and 2 received standard 
rehabilitation therapy. Group 3 consisted of 14 patients receiving standard rehabilitation therapy. 
The evaluation was conducted 24 weeks after the course of injections. During the treatment, all 
patients had normal and stable vitals. At the periods of injections or period after them. No allergic 
or immunological reactions were observed in any patient. There were no deviations from the 
normal ranges or significant changes in the laboratory indicators of the liver and kidneys 
functioning, in comparison with the initial level (ALAT, ASAT, bilirubin, urea, and creatinine). 
Evaluation of the psychological status was made on the CARS scale. The total CARS score was 
significantly reduced in the Group 2 (a decline of 37.9%); in Group 1, the indicator decreased by 
20.0%; in Group 3, i.e. the control group, the indicator decreased by 13.7%. The initial CARS 
score indicator in each group was at the level of 45. In Groups 1 and 2, there were improvements 
in behaviour defined for the “Attitude to people”, “Body use”, “Visual response”, “Taste, smell, 
the reaction on touch and use” and "General Impressions" sub-scales. Those changes were not 
observed in the control group. The research study proved the safety and effectiveness of the use of 
cord blood mononuclear cells in the complex treatment of patients. (Transplantation of human 
cord blood mononuclear cells and umbilical cord-derived mesenchymal stem cells in autism 
Yong-Tao Lv, Yun Zhang, Min Liu,1 Jia-na-ti Qiuwaxi,2 Paul Ashwood,3 Sungho Charles Cho,4 
Ying Huan,1 Ru-Cun Ge,1 Xing-Wang Chen,1 Zhao-Jing Wang,2 Byung-Jo Kim,5 and Xiang 
Hucorresponding). 
 
\ 
 
СLINICAL PURPOSES 
Estimation of the safety and efficiency of the method of transfusion of umbilical blood hemopoetic 
cells to patients with diagnosed autism spectrum disorder without depending on the degrees of 
compatibility of donor and recipient.  
 
TASKS  
1. Develop an algorithm for using the method of transfusion of hematopoietic umbilical cord blood 
cells in the treatment of patients with autistic type disorder.  
2. To formulate criteria for the selection of patients for this technique.  

3. To analyze the efficacy and safety of transfusion of hematopoietic cord blood cells in patients 
with autistic type disorder using rating scales.  
4. Introduce the method of transfusion of cord blood hematopoietic cells to patients with autistic 
type disorder in the complex therapy of patients with autistic type disorder. 
 
 
OBJECT OF STUDY  
The protocol for testing the method will be carried out on 10 patients with autism spectrum 
disorder by transfusion of hematopoietic cord blood cells. As a control group, 10 patients with a 
similar pathology will be examined, comparable in age, gender and degree of autistic disorder 
according to the “paracopy” type against the background of standard therapy.  
Type of study: interventional, prospective, non-randomized with a control group. 
Patient selection criteria (indications for this type of treatment): 
 
• age of the patient is from 3 to 15 years.  
• Diagnosis: autistic disorder.  
• Systemic speech underdevelopment  
• The presence of attention deficit hyperactivity disorder as a comorbid state  
• Cognitive impairment 
Patient exclusion criteria (contraindications for this type of treatment): 
 
• The patient's under 3 years, after 15 years; 
 • The presence of organic pathology of the brain according to CT, MRI  
• The presence of the following diseases in the anamnesis: heart failure in the stage of 
decompensation, a history of stroke less than 1 year ago, anemia and other blood diseases;  
• Decompensation of chronic and endocrinological diseases;  
• Acute respiratory viral and bacterial infections, period less than 1 month after the acute phase. 
 • HIV infection, hepatitis B and C.  
• Cancer, chemotherapy and history of cancer; 
• Tuberculosis.  
• Severe form of intellectual disability as a concomitant disease (diagnosis can be ignored, 
according to the decision of the Medical Committee of the Center); 
• Cerebral palsy.  
• Increased convulsive readiness, epilepsy. 

Indications and contraindications to this type of treatment for each patient are determined at a 
consultation of doctors with the participation of a paediatrician or a psychologist (psychiatrist), 
and transfusiologist. 
 
Research methods 
Evaluation scales CARS, ATEC (to assess the dynamics). Neuropsychological examination, 
examination of a speech therapist-pathologist. Assessment of neurological status. Determination 
of the immune status by flow cytometry (CD3; CD4; CD8; CD19; CD16-56). Determination of 
the level of cytokines in peripheral blood: interleukin 1 beta, interleukin 6, tumor necrosis factor 
alpha, interleukin 8, gamma interferon. Determination of the HLA genotype of the subject 
parents and siblings. 
 
The results will be statistically processed using parametric and non-parametric statistics methods 
using STATA version 9.0, Statistica for Windows version 6.0 and MS Office Excel 2007. Changes 
from the start of therapy and after 1, 2, 6, 12 months will be compared using the Wilcoxon criterion, 
comparing data between the main group and the control group using non-parametric method, using 
the Mann-Whitney test (data are presented in Me - median format with an indication of 25% (q1) 
and 75% (q3) quartile. 
 
Treatment protocol 
 
Selection of patients and assessment of their psychological status, transfusion of umbilical cord 
blood HSC in the conditions of day hospital is carried out by specialists of the State budgetary 
institution of public health "Samara Regional Children's Clinical Hospital N.N. Ivanova 
Laboratory examination, selection of biomaterial will be carried out by the State Budgetary 
Institution "MC Dynasty". 
Stage 1 - selection and examination of the patient, determination of indications and 
contraindications to this method of treatment is carried out by a child neurologist, pediatrician 
(pediatrician), medical psychologist. Assessment of autism disorder is carried out according to the 
assessment scales of CARS, ATEC. 
            A prerequisite for inclusion: 
- diagnosed with autism spectrum disorder by a psychiatrist, 
- Dispensary supervision by a child psychiatrist at the place of residence.   

Mandatory general clinical trials (clinical and diagnostic laboratory):  
a) General clinical blood tests (erythrocytes, hemoglobin, platelets, clotting time, leukocytes + 
formula, COE) - shelf-life 14 days. 
b) general clinical urine test - shelf life 14 days. 
c) feces per helminth eggs - shelf life 14 days. 
d) blood type and rhesus factor. 
A patient who meets the selection criteria and has no contraindications for this type of treatment 
enters into a protocol and signs an informed consent (or his/her legal representative). 
 
Stage 2 - umbilical cord blood sampling. It is performed by a transfusion doctor of the State 
Budgetary Institution "MC Dynasty". 
The selection criteria for the sample will be the following parameters: 
1. Blood group compatibility by system AB0 and Rh 
2.  Infectious safety (negative HIV test results, hepatitis B and C, syphilis, herpes simplex virus, 
cytomegalovirus, toxoplasmosis)  
3. No bacterial or fungal contamination. 
4. Number of hematopoietic cells (mononuclears) in the range of 20-50 million cells per kg of 
patient weight. 
 
Stage 3 - intravenous injection of hematopoietic cells in umbilical cord blood. 
Preliminary examination of the patient and relatives: 
a) Determination of immune status by flow cytometry (SD3; SD4; SD8; SD19; SD16-56) - patient. 
b) Determination of cytokine levels in peripheral blood: interleukin 1 beta, interleukin 6, tumor 
necrosis factor alpha, interleukin 8, gamma-interferon - patient. 
c) Determination of HLA genotype by loci A, B, DRB1 - patient and parent, sibling. 
The sample consists of a suspension of hematopoietic cells diluted in 10 ml 0.9% NaCl + 10 ml 
6% hydroxyethyl starch in a disposable syringe labeled by the manufacturer (label). 
The patient is injected in drops with the use of infusion. The rate of administration is 60 - 70 ml 
per hour. Immediately at the beginning of injection, a biological compatibility test should be 
performed. 
Three-fold injection is planned at an interval of 1 month. 
 
 Possible area of application 
    Specialized hospitals. 
Data to be obtained during implementation 

- correct selection of criteria for selecting patients for treatment of patients with autism type 
disorder; 
- developed algorithm for using the method of transfusion of hematopoietic cells of umbilical cord 
blood in complex therapy of patients with autism type disorders; 
- information on the efficacy and safety of treatment of patients with autism-type disorder by 
transfusion of hematopoietic cells of umbilical cord blood; 
- appropriateness and effectiveness of this method of treatment. To evaluate its impact on treatment 
tactics and results; 
- Association of HLA genes with the development of autism spectrum disorders; 
- relationship between immune status and cytokine profile and severity of the disease. 
Therapy evaluation criteria 
Evaluation of the effect of treatment is carried out 1, 2, 3 and 6 months after treatment. 
Evaluation of the effect is carried out using the scales used in the introduction to the protocol, 
evaluation of the neurological and psychological status of the patient. 
Evaluation of immune status indicators is carried out 6 months after the last injection. 
Cytokine profile is assessed 1, 2, 3 months after the first injection. 
  Expected results: 
It is assumed that transfusions of hematopoietic cord blood cells in patients with autism spectrum 
disorders will not cause adverse reactions and complications (safe) and will lead to improved social 
interaction and communication of patients, increased social adaptation, reduced stereotyping, 
increased self-service skills, increased cognitive functions. 
 
 
STATISTICAL CONSIDERATIONS 
This study will enroll autistic children age 3 to 15 years of age. 
Participants  will be carried out on 10 patients with autism spectrum disorder by transfusion of 
hematopoietic cord blood cells. As a control group, 10 patients with a similar pathology will be 
examined, comparable in age, gender and degree of autistic disorder according to the “paracopy” 
type against the background of standard therapy. 
This is interventional, prospective, non-randomized with a control group study with three 
intravenous allogeneic, unrelated CB infusion in children ages 3-15 years with ASD/ 
The primary outcomes will be assessed 1, 2,3,6 and 12 months after the initial 
infusion in the sequence. Duration of study participation will be 12 months from the time 
of baseline infusion. 
Accrual 

It is estimated that up to 2-3 research participants will be enrolled each week and that 
approximately 4 weeks of accrual will be necessary to enroll 10 participants. 
Study Duration 
Research participants will be followed for safety for 12 months after the third study infusion. 
Demographics and Baseline Characteristics 
Demographics and baseline characteristics will be summarized for all research 
participants. Characteristics to be examined include age, sex, and baseline behavioral status 
(ATEK). 
The results will be statistically processed using parametric and non-parametric statistics methods 
using STATA version 9.0, Statistica for Windows version 6.0 and MS Office Excel 2007. Changes 
from the start of therapy and after 1, 2, 6, 12 months will be compared using the Wilcoxon criterion, 
comparing data between the main group and the control group using non-parametric method, using 
the Mann-Whitney test (data are presented in Me - median format with an indication of 25% (q1) 
and 75% (q3) quartile.