Clinical trial NCT04088760
Conditions: Allogeneic Hematopoietic Cell Transplantation
Sponsor: Children's Hospital Medical Center, Cincinnati
Start date: 2019-09-16; Completion date: 2021-09-03
Enrollment: 15 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
The purpose of this study is to investigate the safety and efficacy of TCRαβ+/CD19+ depleted allogeneic hematopoietic stem cell transplant (HSCT) for malignant and non-malignant disorders in children and adolescent/young adult patients using the CliniMACS® immunomagnetic selection device (Miltenyi Biotec).

Detailed description:
Acute graft versus host disease (GVHD) remains a significant cause of morbidity and mortality and is the biggest barrier to successful allogeneic hematopoietic cell transplantation (HSCT) outcomes. Improved methods of acute GVHD prevention are needed. TCRαβ+/CD19+ depletion of allogeneic hematopoietic stem cell products offers an opportunity to limit the risk of acute GVHD by removing TCRαβ+ T cells and CD19+ B cells which participate in acute GVHD initiation and perpetuation. The purpose of this study is to investigate the safety and efficacy of TCRαβ+/CD19+ depleted allogeneic hematopoietic stem cell transplant (HSCT) for malignant and non-malignant disorders in children and adolescent/young adult patients using the CliniMACS® immunomagnetic selection device (Miltenyi Biotec).

Arms: arm 1: None
Interventions: intervention 1: TCRαβ+/CD19+ depleted allogeneic hematopoietic stem cell transplant (HSCT)
Intervention descriptions: intervention 1: The majority of TCRαβ+ T cells and CD19+ B cells will be removed from the allogeneic graft utilizing the CliniMACS® immunomagnetic selection device (Miltenyi Biotec). The depletion process involves two phases; cell labeling (phase 1) and the automated immunomagnetic depletion process (phase 2). The CD34+ dose may be adjusted by the need to not exceed the TCRαβ+CD3+ dose threshold.

Protocol documents:
Prot_SAP_000.pdf: 
 
 
COVER PAGE FOR CLINICALTRIALS.GOV 
 
 
Protocol Title: Safety and Efficacy of TCRαβ+/CD19+ Depleted Allogeneic 
Hematopoietic Stem Cell Transplantation for Malignant and Non-malignant 
Disorders in Children and Adolescent/Young Adult Patients 
 
 
NCT Number: NCT04088760    
 
 
Document Version Date: 01/27/2021 


Protocol v01.27.2021
IRB# 2018-8568
Page 1 of 34
Safety and Efficacy of TCRαβ+/CD19+ Depleted Allogeneic Hematopoietic Stem 
Cell Transplantation for Malignant and Non-malignant Disorders in Children and 
Adolescent/Young Adult Patients
IND SPONSOR: 
Stella Davies, MB BS, PhD, MRCP
Director, Division of Bone Marrow Transplantation 
and Immune Deficiency
Cancer and Blood Diseases Institute
Cincinnati Children’s Hospital Medical Center
3333 Burnet Avenue
Cincinnati, OH 45229
PRINCIPAL INVESTIGATORS:
Parinda A. Mehta, MD
Rebecca Marsh, MD
Division of Bone Marrow Transplantation and 
Immune Deficiency
Cancer and Blood Diseases Institute
Cincinnati Children’s Hospital Medical Center
3333 Burnet Avenue
Cincinnati, OH 45229
CO-INVESTIGATORS: 
Stella Davies, MB BS, PhD, MRCP
Jacob Bleesing, MD, PhD
Sharat Chandra, MD
Christopher Dandoy, MD, MSc
Michael Grimley, MD
Michael Jordan, MD
Pooja Khandelwal, MD
Mary Christa Krupski, DO, MPH
Ashish Kumar, MD, PhD
Thomas Leemhuis, PhD 
Kasiani Myers, MD
Ruby Khoury, MD
Sonata Jodele, MD
YunZu (Michele) Wang, MD
STATISTICIAN: 
Adam Lane, PhD
 


Protocol v01.27.2021
IRB# 2018-8568
Page 2 of 34
TABLE OF CONTENTS
I. 
PURPOSE OF STUDY
II. 
SIGNIFICANCE OF STUDY IN RELATION TO HUMAN HEALTH
III. 
PREVIOUS WORK DONE IN THIS AREA
IV.
RESEARCH PLAN
1a. 
How many subjects will be studied?
1b. 
How will subjects be selected for this study?
2a. 
Inclusion Criteria
2b. 
Exclusion Criteria
2c. 
Patients Requiring Second Allogeneic HCT or Additional
Aliquots of Stored Hematopoietic Stem Cell Product
3.
Randomization
4. 
Procedures
V. 
EVALUATION CRITERIA
1. 
Primary Endpoint
2. 
Secondary Endpoints
VI. 
OFF STUDY CRITERIA/CRITERIA FOR WITHDRAWING SUBJECTS
VII. 
STATISTICAL CONSIDERATIONS
1. 
Sample size and Study Duration
2. 
Statistical considerations for Data Analysis
3. 
Stopping Rules
VIII. REGULATORY, ETHICAL AND STUDY OVERSIGHT CONSIDERATIONS
1. 
Data and Safety Monitoring Plan
2. 
Blood Specimens
3. 
Other Previously Approved Research Studies in Which the 
Projected Patient Population May Also Be Involved
4. 
Data Collection and Monitoring
5. 
Facilities Utilized in the Study
IX. 
POTENTIAL BENEFITS
X. 
POTENTIAL RISKS, DISCOMFORTS, INCONVENIENCES AND PRECAUTIONS
1. 
Potential risks, discomforts, inconveniences specific to the 
study agent administration and research evaluations


Protocol v01.27.2021
IRB# 2018-8568
Page 3 of 34
2. 
Carcinogenesis, Mutagenesis, and Impairment of Fertility
3. 
Pregnancy/Lactation
4. 
Pediatric Use
5. 
Adverse Reactions
6. 
Precautions that will be taken to monitor and avoid the above 
mentioned risks, discomforts, and inconveniences.
7. 
Adverse Events
8. 
Adverse Event Reporting
9. 
Risk Assessment Recommendation
XI. 
CONFIDENTIALITY
XII. 
PERIOD OF TIME ESTIMATED TO COMPLETE PROJECT AS DESCRIBED
XIII. 
FUNDING
XIV. PAYMENT FOR STUDIES
XV. 
METHOD TO BE USED IN PROCURING CONSENT OF SUBJECTS
XVI. REFERENCES
XVII. APPENDIX 1
XVIII. APPENDIX 2


Protocol v01.27.2021
IRB# 2018-8568
Page 4 of 34
ABSTRACT  
The purpose of this study is to investigate the safety and efficacy of TCRαβ+/CD19+ 
depleted allogeneic hematopoietic stem cell transplant (HSCT) for malignant and 
non-malignant disorders in children and adolescent/young adult patients. This 
protocol will be utilized to provide patients who lack an HLA-genotypically matched 
related donor (MRD) with an option to undergo HSCT.  We hypothesize that 
TCRαβ+/CD19+ depleted hematopoietic stem cell products will be safe, effective, 
and limit acute grades II-IV graft versus host disease (GVHD).  TCRαβ+/CD19+ 
depleted hematopoietic stem cells will be infused with or without a preparative 
regimen clinically prescribed based on our center’s standard-of-care.  This therapy 
serves as an alternative to no HSCT or HSCT with a high risk of acute GVHD, which 
is known to be associated with significant morbidity and mortality.  TCRαβ+/CD19+ 
depletion will be performed on donor hematopoietic stem cell products using the 
CliniMACS® immunomagnetic selection device (Miltenyi Biotec).  Previous reports 
have shown this approach to be safe with results equivalent to matched unrelated 
donor (URD) transplant outcomes. 


Protocol v01.27.2021
IRB# 2018-8568
Page 5 of 34
I. PURPOSE OF STUDY 
Acute graft versus host disease (GVHD) remains a significant cause of morbidity and 
mortality and is the biggest barrier to successful allogeneic hematopoietic cell 
transplantation (HSCT) outcomes.  Improved methods of acute GVHD prevention 
are needed.  TCRαβ+/CD19+ depletion of allogeneic hematopoietic stem cell 
products offers an opportunity to limit the risk of acute GVHD by removing TCRαβ+ 
T cells and CD19+ B cells which participate in acute GVHD initiation and 
perpetuation.  The purpose of this study is to investigate the safety and efficacy of 
TCRαβ+/CD19+ depleted allogeneic hematopoietic stem cell transplant (HSCT) for 
malignant and non-malignant disorders in children and adolescent/young adult 
patients using the CliniMACS® immunomagnetic selection device (Miltenyi Biotec).
Hypothesis 
 
TCRαβ+/CD19+ depleted hematopoietic stem cell products will be safe, effective, 
and limit acute GVHD. 
 
Primary Objective 
Assess the safety of infusion of TCRαβ+/CD19+ depleted donor stem cell 
products
Secondary Objectives
Assess engraftment, acute and chronic GVHD incidence, and 1 year GVHD-
free survival
II. SIGNIFICANCE OF STUDY IN RELATION TO HUMAN HEALTH
Allogeneic hematopoietic cell transplantation (HSCT) has been used extensively to 
treat patients with malignancies, hematologic disorders, and primary immune 
deficiencies.1  Ideally, allogeneic HSCT is performed using an HLA-matched related 
or unrelated donor.  However, only approximately 25% of patients will have an HLA-
identical sibling, and the chances of finding an HLA-matched unrelated donor vary 
from 60 to 70% for Caucasian patients to less than 10% for ethnic minorities.2  The 
lack of a suitable donor means that some patients are not offered the option of 


Protocol v01.27.2021
IRB# 2018-8568
Page 6 of 34
allogeneic HSCT, or must proceed using mismatched donors which greatly 
increases the risk of GVHD and death.  
Haploidentical transplant offers an attractive alternative because almost all 
patients will have a haploidentical family member who could potentially serve as 
the stem cell donor.  This type of transplant presents the challenge of a very high 
risk of GVHD.  Mature donor T-cells are responsible for GVHD.3,4   Methods to T-cell 
deplete the graft or suppress the donor T cells have been developed to prevent 
GVHD after HLA-haploidentical HSCT.5-8  Isolation and infusion of purified CD34+ 
cells can prevent severe GVHD in haplo-HSCT patients.9,10  However, this can result 
in an increased risk of infections early after HSCT.  In order to reduce this risk, 
alloreactive TCRαβ+ T-cells and antigen-presenting CD19+ B-cells can be selectively 
removed from the graft.11  This approach avoids GVHD but allows the retention of 
TCRγδ+ T cells and NK cells in the graft to provide early innate immunity.  The 
activation of γδ T-cell activation is not regulated by MHC molecules, which makes 
them less likely to cause GVHD.  TCRγδ+ T cells are known to participate in 
immunity against cytomegalovirus (CMV) reactivation and disease following 
HSCT,12 and expansion of CMV-reactive TCRγδ+ T-cells can be detected in patients 
who have been treated with kidney or lung transplantation.13,14  TCR γδ+ T-cells and 
NK cells may additionally offer protection against other infections and malignancy 
relapse.15-18  Importantly, pediatric studies have demonstrated that relapse rates 
are similar to rates in patients undergoing sibling or unrelated donor transplant 
while GVHD-free/relapse-free survival is better.19-21
Access to TCRαβ+/CD19+ depleted allogeneic HSCT is important as it will expand 
the access to transplant tremendously: essentially 100% of patients needing HSCT 
will have a donor option.  TCRαβ+/CD19+ depleted HSCT can also be offered to 
children for whom rapidly progressive or unstable disease does not allow the time 
to identify a suitable matched URD.  Additionally, some patients may have organ 
dysfunction prior to transplant that prevents giving common GVHD prophylaxis 
such as cyclosporine or tacrolimus.  TCRαβ+/CD19+ depleted HSCT would allow for 
HSCT without any post-HSCT immune suppression in these patients.  Similarly, rare 
subgroups of pediatric disorders have underlying defects in DNA damage repair 
pathways, telomere biology, or epithelial cell propensity for abnormal 
inflammatory responses and cell death (e.g. Fanconi anemia, dyskeratosis 
congenita, XIAP Deficiency, etc).  GVHD can be associated with increased risk of 
fatal outcomes in such patients and therefore T cell depletion has been used in 


Protocol v01.27.2021
IRB# 2018-8568
Page 7 of 34
these disorders to prevent GVHD. TCRαβ+/CD19+ depleted allogeneic HSCT will 
likely provide a safer alternative to CD34+ selection in these high-risk patients even 
in the setting of fully matched URDs.
III. PREVIOUS WORK DONE IN THIS AREA
Multiple methods of ex-vivo and in-vivo T-cell depletion have been reported in the 
HLA-haploidentical HSCT setting.22,23  CD34+ stem cells can be positively selected 
and administered as a graft product, or negative selection can be performed to 
remove specific components such as T cells from the graft product.  CD34+ positive 
selection, or combined CD3+ and CD19+ negative selection, results in the loss of 
cells that are important for preventing infections and relapse.9,10,24-26  In order to 
reduce the risk of infections and relapse, several centers have more recently 
implemented a TCRαβ+ T-cell/CD19+ B-cell depletion approach.11  Selective 
TCRαβ+/CD19+ depletion aims to remove TCRαβ+ T-cells from the graft which have 
alloreactive potential and can cause GVHD, as well as antigen-presenting B cells 
which also contribute to GVHD.27-30  However, TCRγδ+ T-cells purposefully remain 
in the graft to facilitate engraftment, reduce the risk of infections, and reduce the 
risk of leukemia relapse without causing GVHD as TCRγδ+ T-cells do not have 
alloreactive capacity.18,31-34  Donor NK cells also remain in the graft and offer activity 
during the early weeks following HSCT until mature donor graft-derived NK cell 
reconstitution occurs.16,35   The large number of stem cells typically infused with 
this type of product can also allow early robust neutrophil engraftment which may 
decrease early bacterial infections.  
Promising clinical results have been recently reported in children and young adults 
treated with TCRαβ+/CD19+ depleted HSCT (Table 1).   Bertaina et al evaluated 23 
children 
with 
non-malignant 
hematological 
disorders 
who 
underwent 
haploidentical transplantation using TCRαβ+/CD19+ depletion.36 No acute GVHD 
prophylaxis was administered.  All but 4 of the patients engrafted.  Three patients 
developed grade I-II acute GVHD of the skin, and there was no occurrence of 
visceral acute or chronic GVHD.   The 2-year probability of disease-free survival was 
91.1%.  Other pediatric experiences have been published since, and are also 
summarized in Table 1.19-21,36,37  All patients received less than 2x10^5 TCRαβ+ T 
cells per kg and the median CD34+ cell doses were 9x10^6/kg or greater.  Rates of 
Grades 3-4 acute GVHD were low.  Several studies did not use additional GVHD 
prophylaxis.  Overall survival at various timepoints ranged from 50-97%.


Protocol v01.27.2021
IRB# 2018-8568
Page 8 of 34
    Table 1.  PREVIOUS WORK DONE IN THE AREA
Study
Bertania et al36
Balashov37
Lang19
Maschan20
Locatelli21
Year
2014
2015
2015
2016
2017
PMID
24869942
26187864
26039210
26808573
28588018
No of pts
23
37
41
33
80
Adult /pediatric
Pediatric
Pediatric
Pediatric
Pediatric
Pediatric
Disease 
Indication
Non-malignant 
disorders
Primary Immune 
Deficiencies
ALL (20) 
AML(9) 
MDS/JMML 
(3) 
Solid 
tumors (4)
Nonmalignant 
(5)
AML
ALL, AML
Donor
Haplo
MUD (n=27)
Haplo (n=10)
⩾2 HLA loci-
mismatched 
parents
MUD
MMRD
Haplo
Haplo
GvHD 
prophylaxis
None
Tac/MTX (n=34) 
Tac/MMF (N=2)
CSA/MTX (n=1)
MMF until day 
+30
None
None
CD34+ Dose
15.8x10^6/kg 
(10.2-40.0)
11.7x10^6/kg
(5.9-21.3)
14.9x10^6/kg
9x10^6/kg 
(3.8–21)
13.9x10^6/kg
(6-40.44)
Alpha/beta Dose
4x10^4/kg 
(1-9.5)
10.6x10^3/kg
(0.8-368)
16.9x10^3/kg
20x10^3/kg 
(2.9–126)
0.047x10^6/kg
(0.002-0.099)
Gamma/delta 
Dose
9.4x10^6/kg
(1.6-95.4)
11.0x10^6/kg
9x10^6/kg
(1–30)
8.1x10^6/kg
(0.86-56.7)
Neutrophil 
Engraftment
Day+13 (10-20)
Day +16 (11-28)
Day +10 (7-21)
Day +16
Day +13 (9-19)
GVHD
Grade 1–2: 3 
pts
No chronic
Grade 2: 7 pts 
Grade 4: 1 pt 
Chronic: 1 pt
Grade 2: 4 pts
Grade 3-4: 6 
pts
Chronic: 9 pts
Grade 
2-3:14 
pts
Chronic: 10 pts
Grade 1-2: 24 pts
Chronic: 4 pts
Graft Failure
2 Primary 
2 Secondary
2 Primary
8 Secondary
5
2
Malignancy 
Relapse
17 pts
10 pts
19 pts
Infection
Viral 
reactivations: 9
CMV: 16pts, 5 
with disease
Not Reported
CMV 
Reactivation:
52%
EBV 
Reactivation: 
50%
Not reported
 Survival
2yr OS 91%
2yr DFS 91%
2yr OS 97%
2yr EFS 68%
If in CR1-3: 
100%
If 
in 
active 
disease:  0%
2yr OS 68% 
2yr EFS 60%
OS 72% 5 years


Protocol v01.27.2021
IRB# 2018-8568
Page 9 of 34
RATIONALE
Based on these results we propose the use of αβ T-cell/CD19+ B-cell depleted stem 
cells in mismatched related and unrelated donor HSCT for malignant and 
nonmalignant disorders in pediatric and adolescent/young adult patients.  
Mismatched related donors provide an allogeneic HSCT option for essentially all 
patients in need of HSCT.  This approach reduces the risk of acute GVHD and 
reduces the need for GVHD prophylaxis, either of which is poorly tolerated by 
certain patient groups.  Additionally, transplantation can be performed without 
delay which can be critical in patients with high risk malignant disease or very 
severe non-malignant disorders requiring urgent treatment.    
IV. RESEARCH PLAN 
1a. How many subjects will be studied?
We will enroll up to 80 patients.
1b. How will subjects be selected for this study?
Patients attending the CCHMC bone marrow transplant (BMT) clinic who meet the 
eligibility criteria will be offered the opportunity to participate in this research 
study.  The study is open to all participants regardless of gender or ethnicity.
2a. Inclusion Criteria  
Any patient requiring an allogeneic HSCT who lacks an HLA-genotypically matched 
related donor.  Genotypically matched related donors are allowed when there is a 
clinical desire to avoid the use of GVHD prophylaxis medications. 
2b. Exclusion Criteria
Prior allogeneic transplant with active acute or chronic GVHD, or life-threatening 
infection. Patients with a prior history of allogenic transplant without active GVHD 
or life-threatening infection can be considered.
2c. Patients Requiring Second Allogeneic HCT or Additional Aliquots of Stored 
Hematopoietic Stem Cell Product
Patients may receive additional TCRa/b CD19 depleted product(s) if clinically 
indicated.


Protocol v01.27.2021
IRB# 2018-8568
Page 10 of 34
3. Randomization 
Not applicable. 
4. Procedures  
Pre-Infusion Patient Evaluation:
Patients enrolled on this protocol will undergo evaluation for disease status and 
organ function per institutional HSCT guidelines prior to starting conditioning 
regimen. 
Donor Selection:
Transplant donors will be selected based on institutional guidelines/SOPs. 
Treatment Plan:  
 
 
 
 
Informed Consent:
Informed consent will be obtained for each study participant (stem cell transplant 
recipient) and/or parent or legal guardian as appropriate. The study will be 
explained to the study subject in lay terms, in their native language. The informed 
consent process will require written consent by the patient (if ≥18 yrs of age) or 
parent/legal guardian. 
The investigator will be available to answer any questions that the patient or 
caregiver may have regarding procedures, risks and alternatives. The signed 
consent form will be maintained in the study subject’s chart with a copy being 
placed in the patient’s medical record.


Protocol v01.27.2021
IRB# 2018-8568
Page 11 of 34
Preparative therapy: 
Chemotherapy +/- radiation therapy +/- serotherapy will be administered to the 
recipient prior to infusion of donor stem cells based on patient’s underlying 
diagnosis and organ function according to institutional standard of care or per 
study requirements if participating in another study. This will include 
myeloablative, non-myeloablative, reduced intensity, or minimal intensity 
conditioning regimens on or off study. On a rare occasion no preparative 
conditioning will be administered at the discretion of patient’s primary BMT 
attending/team.
Donor Consent and Mobilization:
The stem cell donor will consent to collection of peripheral blood stem cells 
following standard clinical institutional guidelines/SOPs as the mobilization and 
collection of peripheral blood stem cells does not constitute research.  Prior to 
initiation of the stem cell collection process, donors will be thoroughly evaluated 
and determined to be an acceptable donor candidate and able to tolerate all of the 
procedures involved. This will be done on a clinical basis per current standard-of-
care. G-CSF mobilization of the donor will also be performed per institutional 
guidelines and SOPs.  
Collection Target Cell Dose:
We will aim to collect 10 x 10^6 or greater CD34+ stem cells per kg recipient body 
weight.  
Stem Cell Product Processing – TCRαβ+/CD19+ Depletion with CliniMACS 
(Miltenyi Biotech):
The TCRαβ+ T cells and the CD19+ B cells will be removed from the allogeneic graft 
utilizing the CliniMACS® immunomagnetic selection device (Miltenyi Biotec).  The 
detailed procedures are provided in the CliniMACS® User Manual that accompanies 
the CliniMACS® Instrument.  The depletion process involves two phases; cell 
labeling (phase 1) and the automated immunomagnetic depletion process (phase 
2).  A brief summary of the procedure is provided below.
Phase 1 – Labeling:  
 
 
  
 


Protocol v01.27.2021
IRB# 2018-8568
Page 12 of 34
 
 
  
Phase 2 – Cell Depletion:  
 
 
 
 
 
 
 
 
 
 
Post depletion - Stem Cell Dosing:
 
 
 
 
 
  
  
 
 
  
 
 
 


Protocol v01.27.2021
IRB# 2018-8568
Page 13 of 34
Stem Cell Processing: 
   
    
  
 
 
Product Release Parameters:
 
 
 
 
 
 
 
Possible Interventions Based on Viable TCR alpha/beta T Cell Dose and B Cell 
Dose:
 
 
 
 
  
Safety of Product:
   
     


Protocol v01.27.2021
IRB# 2018-8568
Page 14 of 34
Product testing:
 
 
 
 
 
 
 
Culture of the product:
 
 
 
Stem Cell Infusion: 
 
. 
 
Immune Suppression after stem cell infusion: 
 
 
 
 


Protocol v01.27.2021
IRB# 2018-8568
Page 15 of 34
 
 
Supportive therapy: 
•
 
  
Required Observations:
 
 
 
 
Day of Graft Infusion:
 
 
Post-HSCT: 
 
 
 
 
 
 
 
 
 
 
 
 


Protocol v01.27.2021
IRB# 2018-8568
Page 16 of 34
   
 
 
 
 
  
V. EVALUATION CRITERIA
Primary Endpoint:
The primary endpoint of the study is safety of infusion of TCRαβ+/CD19+ depleted 
donor grafts. 
We will determine the frequency and characteristics of potential infusion-related
toxicity.  Experience at our center using CD34+ selected stem cells has shown
infusion related toxicity occurs at an incidence of <5%.  We previously observed 
zero infusion reactions in 22 patients who received a CD34+ selected boost product 
as part of a CCHMC study (IRB# 2010-2344), and 2 infusion reactions among 55 
infusions which were performed as part of clinical care including 1 occurrence of 
hypertension, and 1 occurrence of urticaria that resolved with anti-histamine 
administration.  While there is little expectation of infusion-related toxicity, the 
incidence of infusion-related reactions will be recorded and evaluated. 
Secondary Endpoints:
The secondary endpoints of the study are to assess engraftment and acute GVHD 
incidence.
Engraftment:
Engraftment and graft function are monitored on a clinical basis.  Neutrophil 
recovery will be defined as the first of 3 consecutive days that the absolute 
neutrophil count is 500 cells/mcL.  Platelet recovery will be defined as the first of 
3 consecutive days that the platelet count is greater than 20 without any platelet 
cell transfusions in the previous 7 days.  Initial donor-derived hematopoietic cell 
recovery will be defined as having 95% or greater donor-derived cells.  Initial 
mixed donor and recipient-derived engraftment will be defined as having less 


Protocol v01.27.2021
IRB# 2018-8568
Page 17 of 34
than 95% donor-derived cells but more than 5% donor-derived cells on a first and 
second confirmatory engraftment study (if the second study shows ≥95% donor 
cells, the patient will be classified as having donor-derived hematopoietic cell 
recovery).  Recipient-derived hematopoietic cell recovery will be defined as 
having 5% or less donor-derived cells.  Any patient who fails to achieve neutrophil 
recovery within 42 days of allogeneic HSCT, or has 5% or less donor-derived cells 
will be considered as having primary graft failure.  We will report the cumulative 
incidence of achieving initial donor-derived hematopoietic cell recovery. 
Complete blood counts and whole blood donor chimerism studies are frequently 
performed on a clinical basis following allogeneic HSCT.  Stable hematopoietic cell 
recovery at 100 Days (+/- 2 weeks) will be defined as having hemoglobin, 
neutrophil, and platelet counts of at least 80% of the normal range in the absence 
of transfusions or clinical diagnoses which may affect the counts such as 
hemolytic anemia, auto-immune thrombocytopenia, iron deficiency, infection or 
medication induced bone marrow suppression, or other clinically relevant 
diagnosis.  Patients with these or other medical diagnoses may still be considered 
to have count recovery.  The percentage of donor cells will be recorded at Day 
+100 (preferably +/- 2 weeks) and Day +100 Donor-derived hematopoietic cell 
recovery will be defined as having 95% or greater donor-derived cells.  Day +100 
mixed donor and recipient-derived hematopoietic cell recovery will be defined as 
having less than 95% donor-derived cells but more than 5% donor-derived cells.  
Recipient-derived Day +100 hematopoietic cell recovery will be defined as having 
5% or less donor-derived cells.  These patients will be considered as having 
secondary graft failure.  We will report rates of successful donor-derived 
hematopoietic cell recovery at Day +100.  Exceptions may be made for patients 
with Severe Combined Immune Deficiency who may reconstitute with donor-
derived T cells but have autologous recovery of myeloid and other cells.  For such 
cases we will examine results of sorted T cell chimerism studies.  We will also 
record the incidence of any additional cell product administrations or second 
allogeneic HSCT.  Any patient who receives a second allogeneic HSCT will be 
considered as having graft failure.  
We will report the Day +100 event-free survival of patients where an event is 
defined as death, primary graft failure, autologous recovery (<5% donor-derived 
cells in whole blood or in the cellular compartment of interest, i.e. T cells in patients 
with SCID), secondary graft failure, receipt of an additional hematopoietic cell 


Protocol v01.27.2021
IRB# 2018-8568
Page 18 of 34
product other than for control of infection (Donor lymphocyte infusion or Virus-
Specific T cell infusion), and second allogeneic HSCT.  
Acute GVHD:
Clinical assessments for acute GVHD are frequently performed using Modified 
Glucksberg Criteria as in Appendix 1.  We will collect clinical information regarding 
incidence and severity of acute GVHD.  We will report the Day +100 cumulative 
incidences of acute GVHD grades II-IV and III-IV.  
Chronic GVHD:
Clinical assessments for chronic GVHD are also frequently performed using NIH 
Consensus criteria.  We will report the 1 year cumulative incidences of chronic 
GVHD.  
GVHD-Free Survival:
Based on the presence or not of acute or chronic GVHD at 1 year, we will report 
GVHD-free survival at 1 year.
VI. OFF STUDY CRITERIA/CRITERIA FOR WITHDRAWING SUBJECTS
Subjects (or parents or legal guardians) have the right to withdraw from the study 
at any time and for any reason without prejudice to his or her future medical care 
by the physician or the institution.  
Any subject who withdraws consent to participate in the study will be removed 
from further treatment and/or study observation immediately upon the date of 
request.  The subject will be followed for safety purposes for at least 1 week after 
infusion of the stem cell product.  
Criteria for Removal from Protocol Therapy:  
•
Patient withdrawal from protocol therapy 
Off Study Criteria:
a) Death
b) Receipt of a second stem cell product
c) Lost to follow-up


Protocol v01.27.2021
IRB# 2018-8568
Page 19 of 34
VII. 
STATISTICAL CONSIDERATIONS
1. Sample Size and Study Duration:
We chose a sample size of 80 patients based on estimates of patient need for access 
to T cell depleted grafts over 3-5 years at our Institution.  All patients will be 
followed for primary and secondary endpoints for at least 100 Days following 
allogeneic HSCT, and 1 year data will be collected regarding survival and presence 
or absence of acute or chronic GVHD.  
2. Statistical Considerations for Data Analysis:
Data will be presented with descriptive statistics.  Cumulative incidence curves will 
be created to analyze engraftment and acute GVHD.  Death will be treated as a 
competing event.  Probability of overall survival and event-free survival will be 
estimated with Kaplan-Meier curves.  
3. Stopping Rules:
A predictive probability design (Lee, Lin. Clinical Trials. 2008) will be used to 
control for safety with respect to TRM. The unacceptable rate of TRM is 30% or 
more. This will be tested against a one-sided alternative. The trial will initialize by 
enrolling 10 patients. If 3 or more out of 10 have a TRM the trial will stop. 
Otherwise the trial will continue according to the stopping rules plan (Appendix 2) 
up to a maximum of 73 patients. If the number of patients with a TRM within 100 
days following allogeneic HCT is ever greater than or equal to the number listed in 
the corresponding patients enrolled row the trial will stop and the drug will be 
declared not safe. This design yields a type I error rate 0.05 and power greater 
than 80% when the true rate of TRM is 15%. Note the stopping rule must be made 
based on the chronological order of the patients enrolled. 
We plan to meet with medical monitor approximately every quarter to review 
these safety data and stopping rules. The medical monitor will review the TRM 
data in the context of HLA mismatch; and as usual, he/she will have the final 
authority regarding continuation of the study in the event of excessive TRM.


Protocol v01.27.2021
IRB# 2018-8568
Page 20 of 34
VIII.
REGULATORY, ETHICAL, AND STUDY OVERSIGHT CONSIDERATIONS
The protocol and informed consent document for this study must be approved in 
writing by the Institutional Review Board (IRB) prior to any patient being registered 
on the study.
Changes to the protocol must also be approved by the IRB. Records of the 
Institutional Review Board review and approval of all documents pertaining to this 
study must be kept on file by the investigator and are subject to FDA inspection at 
any time during the study. Periodic status reports will be submitted to the 
Institutional Review Board at least yearly, as well as notification of completion of 
the study and a final report within 6-12 months of study completion or termination.
Informed consent will be obtained prior to the treatment of participants.
1. Data Safety Monitoring Plan:
The study will be monitored closely by a medical monitor.  This will be a qualified 
physician not associated with this particular protocol.  The monitor will have 
expertise in basic hematology and conduction of clinical trials, and will have prior 
regulatory experience for similar studies.  The monitor will work closely with the PI 
to monitor the participants on study.  He/she will meet with the PI and review study 
data/progress approximately every 3 months; more often at the discretion of the 
PI and/or medical monitor. 
The medical monitor will review the TRM data in the context of HLA mismatch; and 
as usual, he/she will have the final authority regarding continuation of the study in 
the event of excessive TRM. The medical monitor is required to review all 
unanticipated problems involving risk to subjects or others, unanticipated serious 
adverse events and all subject deaths associated with the protocol and provide an 
unbiased written report of the review.  
At a minimum, the medical monitor should comment on the relationship of the 
event or problem, to participation in the study.  Based on the review of these 
events, the monitor should make a recommendation regarding study continuation. 
Although the expected rate of serious toxicities will be low, any unanticipated 
attributable serious adverse event will be discussed with the medical monitor. If 
there are any attributable Serious Adverse Events, the study will stop accrual 


Protocol v01.27.2021
IRB# 2018-8568
Page 21 of 34
pending review by the medical monitor. The study will resume patient accrual only 
if written approval is given by the medical monitor.  
All decisions regarding study continuation, modification, or termination will be 
reported immediately or annually, as appropriate, to the IRB and other appropriate 
agencies. Reports for events determined by either the investigator or medical 
monitor to be possible or definitely related to study participation and reports of 
events resulting in death should be forwarded in compliance with current IRB policy 
and applicable federal regulations.
All reports from Medical monitor will be forwarded to Cincinnati Children's Hospital 
Medical Center IRB and other regulatory agencies as appropriate.  
2. Blood Specimens:
There are no research blood specimens for this study.
3. Other Previously Approved Research Studies in Which the Projected Patient  
Population May Also Be Involved:
Patients may be additionally involved in other studies surrounding the care of 
patients undergoing allogeneic HCT.  
4. Data Collection and Monitoring:
Data for the study will be entered into a secure, password protected database by 
the study staff. Monitoring and auditing procedures will be followed to ensure that 
the study is conducted, documented, and reported in accordance with the IRB 
approved protocol, all applicable federal regulations and guidelines, and applicable 
regulatory requirements of Cincinnati Children’s Hospital Medical Center.
Verification of eligibility will be performed and appropriate documentation of 
informed consent will be documented for all subjects enrolled into the study.  The 
timeliness of Serious Adverse Event reporting will be monitored to ensure 
regulatory compliance.  All case report forms (CRF) for the first subject enrolled into 
the study will be monitored for completeness and quality by comparing data in the 
case report forms to data in the source documents.  Thereafter, a minimum of 10% 
of enrolled subjects’ CRFs will be monitored for completeness and quality by 
comparing data in the case report forms to data in the source documents. 
 


Protocol v01.27.2021
IRB# 2018-8568
Page 22 of 34
5. Facilities Utilized in the Study:
Study participants will be cared for by the Division of Bone Marrow Transplantation 
and Immune Deficiency physicians on the Bone Marrow Transplant inpatient and 
outpatients units at Cincinnati Children’s Hospital Medical Center.
IX.
POTENTIAL BENEFITS
Patients may benefit from this study from gaining access to allogeneic HCT with a 
reduced risk of acute GVHD.  
X.
POTENTIAL RISKS, DISCOMFORTS, INCONVENIENCES AND PRECAUTIONS
1. Potential risks, discomforts, inconveniences specific to the study agent 
administration and research evaluations include:
There is a risk of infusion reactions that we estimate to be less than 5% based on 
previous experience using CD34+ selected stem cell grafts.  Every effort will be 
made to minimize any discomfort associated with TCRαβ+/CD19+ depleted stem 
cell infusion.
There is also risk of loss of confidentiality.  We will minimize this risk by keeping all 
study data stored on password protected computers.  Any paper records will be 
kept in secure office locations. Access to the files will be limited to the Principal 
Investigator and designees.
2.  Carcinogenesis, Mutagenesis, and Impairment of Fertility:
Selection of the hematopoietic stem cell product should not affect these 
complications.  Any conditioning chemotherapy prescribed on a clinical basis (not 
part of the study) may affect risk of these complications and is explained during the 
clinical informed consent procedure for clinical treatment with allogeneic HCT.
3.  Pregnancy/Lactation:
Selection of the hematopoietic stem cell product should not affect risk to 
pregnancy/lactation. However due to the high risk nature of allogeneic HCT, 
patients should not be pregnant or lactating.  
4.  Pediatric Use:
This study will enroll pediatric and adolescent and young adult patients. 


Protocol v01.27.2021
IRB# 2018-8568
Page 23 of 34
5.  Adverse reactions:
There is a risk of infusion reactions that we estimate to be less than 5% based on 
previous experience using CD34+ selected stem cell grafts.  To minimize this risk, 
pre-medication with anti-histamines and/or steroids will be used before infusion 
as per standard clinical guidelines. Every effort will be made to minimize any 
discomfort associated with TCRαβ+/CD19+ depleted stem cell grafts.
6.  Precautions that will be taken to monitor and avoid the above mentioned risks, 
discomforts, and inconveniences:
Patients will be clinically monitored during the infusion of TCRαβ+/CD19+ depleted 
stem cell grafts and any complications will be clinically treated.  
All study data will be stored on password protected computers.  Any paper records 
will be kept in secure office locations.
7.  Adverse Events:
Unanticipated severe adverse event/s must be reported to the principal 
investigator who is responsible for the reporting to the Cincinnati Children's 
Hospital Medical Center IRB, FDA, and the medical monitor as applicable. 
All Adverse Event reporting is to comply with the current CCHMC IRB policy and 
applicable federal regulations. If any serious adverse events occur, current 
guidelines will be followed for expedited reporting to the Sponsor, IRB, FDA and/or 
the medical monitor.  
The investigator should notify the IRB of serious adverse events occurring at the 
site, in accordance with local procedures. 
Definitions:
Adverse Event
An adverse event is any new, undesirable medical occurrence or change of an 
existing condition in a subject that occurs during treatment (which may include a 
specified post treatment period), whether or not considered to be product related.  
Abnormal laboratory findings considered by the investigator to be clinically 
significant, e.g. those that are unusual or unusually severe for the population being 
studied, should be recorded as adverse events. 


Protocol v01.27.2021
IRB# 2018-8568
Page 24 of 34
Serious Adverse Event
A serious adverse event is defined by regulatory/clinical criteria.  It is one that 
suggests a significant hazard or side effect, regardless of the investigator’s or 
sponsor’s opinion on the relationship to study material.  This includes, but may not 
be limited to, any event that (at any dose):
•   is fatal;
•
is life threatening (places the subject at immediate risk of death);
•
requires intensive care;  
•
is persistent and results in significant disability/incapacity; 
•
Hospitalization
•
Disability or Permanent Damage
•
Congential Anomaly/Birth Defect
•
Require Intervention to Prevent Permanent Impairment or Damage 
(this one may only apply to investigational devices)
•
Other Serious (Important Medical Events)
•
For an event that may not be immediately life threatening or result 
in death or hospitalization, but may jeopardize the patient or 
require intervention to prevent one of the outcomes listed above, 
or result in urgent investigation, may be considered serious.
Important medical events that may not be immediately life threatening or result in 
death or hospitalization, but may jeopardize the patient or require intervention to 
prevent one of the outcomes listed above, or result in urgent investigation, may be 
considered serious.
8.  Adverse Event Reporting:
Safety and tolerability will be assessed according to the Common Terminology 
Criteria for Adverse Events v5.0 (CTCAE published November 27, 2017). The 
principal investigator(s) or designee will review adverse events closely and assess 
the event’s relationship to study procedures to determine whether the event is 
unrelated or, unlikely, possibly, probably or definitely related to the study 
procedures, especially in context of the underlying disease and baseline lab values 
for the patient.
Allogeneic HSCT is known to be a high-risk procedure with many complications and 
a gross mortality risk of approximately 70%.  Only adverse events of grade 4 or 


Protocol v01.27.2021
IRB# 2018-8568
Page 25 of 34
higher will be recorded on case report forms. Because the primary aim of this study 
is to assess safety of the infusion of TCRαβ+/CD19+ depleted HSCT, and infusion 
reactions or other adverse events are expected to occur within 48 hours of the 
initial product infusion, we will only monitor and record AEs for the first week of 
the study.  Any “attributable” events noted after this first week will be recorded 
and reported according to applicable institutional and federal regulations. Patients 
will be followed through Day +100 for endpoints including engraftment, acute 
GVHD, and event-free survival.
Expected events are those that have been previously identified as resulting from 
administration of the protocol therapy, those that can be attributed to the 
underlying condition, and/or those associated with transplant. An adverse event is 
considered unexpected, for reporting purposes only, when either the type of event 
or the severity of the event is not listed in the protocol and/or investigational 
brochure. 
Please note that for the purpose of this study we will exclude the report of 
hematologic toxicities, as these subjects, by virtue of their underlying disease 
and/or condition and the transplant regimen, will develop hematologic 
abnormalities that otherwise would qualify as toxicity.  Hematologic toxicities 
which are attributable to the underlying hematological disease and/or HCT 
preparative regimen will not be considered adverse events. In the event that any 
of these known complications of transplant or hematologic toxicities are thought 
to be at least possibly attributable to the study, the event will be recorded and 
subsequently reported to the IRB and FDA according to current reporting 
requirements.
Additionally for this study, events as previously identified as resulting from 
administration of a stem cell transplant will also be considered expected.  The 
transplant regimen is well known to commonly affect systems such as the 
hematologic, immunologic, and gastrointestinal systems.  The following are known 
complications of transplant: feeling tired, anemia, disseminated intravascular 
coagulation, febrile neutropenia, hemolysis, thrombotic thrombocytopenia 
purpura, adrenal insufficiency, colitis, constipation, diarrhea, enterocolitis, 
gastrointestinal pain, malabsorption, mucositis, nausea, pancreatitis, vomiting, 
dyspepsia, hepatitis, jaundice, hepatic veno-occlusive disease, thrombotic 
microangiopathy, fever, pain (including but not limited to headache, muscle 


Protocol v01.27.2021
IRB# 2018-8568
Page 26 of 34
cramps, abdominal pain, throat pain etc), cholecystitis, allergic reaction, serum 
sickness, entercolitis, infections, viral reactivations, upper respiratory infection, 
urinary 
tract 
infection, 
leukocytopenia, 
lymphocytopenia, 
neutropenia, 
thrombocytopenia, bleeding complications such as epistaxis, hematemesis, 
petechiae, subconjunctival hemorrhage, weight gain/loss, fluid retention, acidosis, 
alkalosis, anorexia, dehydration, glucose intolerance, electrolyte disturbances, iron 
overload, generalized muscle weakness, reversible posterior leukoencephalopathy 
syndrome, bladder spasm, cystitis, noninfective, hematuria, urinary frequency, 
urinary urgency, irregular menstruation, pruritis, hypertension, depression, 
anxiety, purpura, and petechiae.  It is notable that adverse events following 
allogeneic hematopoietic cell transplantation can be severe and life-threatening.  
Acute graft versus host disease, infections, mixed chimerism, cytopenias, and graft 
failure are all known complications of allogeneic HCT, and can result in death.  
Organ failure my also occur due to toxicities of chemotherapy, infections, 
thrombotic microangiopathy, or other reasons.  For the purposes of this study, the 
listed events that are </= grade 3 will not be considered adverse events.  All listed 
events that are grade 4 or 5 will be recorded on the AE case report forms (CRF) and 
reported according to current IRB guidelines.  In the event that any of these known 
complications are thought to be at least possibly attributable to the study 
procedure, and not to the allogeneic hematopoietic cell transplantation and 
expected complications, the event will be recorded and subsequently reported to 
the IRB and FDA according to current reporting requirements.  Acute GVHD, mixed 
chimerism, and graft failure are not considered AEs during the study (they are 
captured as endpoints). 
 
Medical conditions/diseases present before the infusion of TCRαβ+/CD19+ 
depleted HSCT are only considered adverse events if they worsen after the infusion 
of TCRαβ+/CD19+ depleted HSCT. Abnormal laboratory values or test results 
constitute adverse events only if they induce clinical signs or symptoms, are 
considered clinically significant, or require therapy.
Adverse Events will be reported according to current CCHMC IRB policy and 
applicable federal regulations. If any serious adverse events occur, current 
guidelines will be followed for expedited reporting to the IRB, medical monitor, and 
sponsor. 


Protocol v01.27.2021
IRB# 2018-8568
Page 27 of 34
•
If an event meets all of the below criteria, an IND Safety Report will be 
generated and submitted to the FDA by the sponsor no later than 15 calendar 
days after the sponsor receives the safety information and determines that 
the information qualifies for reporting:Suspected Adverse reaction
•
Serious
•
Unexpected
9.  Risk assessment recommendation:
Minimal risk but with direct benefit to participants
Minimal risk but without direct benefit to participants
More than minimal risk, but potential direct benefit to participants
More than minimal risk but without direct benefit to participant
XI.
CONFIDENTIALITY
All records and documents pertaining to the study will be maintained by the 
Investigator and designee(s), and will be available for inspection by the Cincinnati 
Children's Hospital Institutional Review Board, the sponsor and the Medical 
monitor. All documents will be kept in a locked cabinet or on password protected 
computers in the office of the Clinical Management Research Support Core 
(CMRSC). Access to the files will be limited to the Principal Investigator and 
designees. All information provided to the investigator dealing with the study and 
information obtained during the course of the study will be regarded as 
confidential.
XII.
PERIOD OF TIME ESTIMATED TO COMPLETE PROJECT AS DESCRIBED
The period of time to complete enrollment for this study is estimated to be 
approximately 5 years.  Total time for completion of follow up is estimated to be 6 
years.  
XIII.
FUNDING 
The institution to which funding will be made: CCHMC


Protocol v01.27.2021
IRB# 2018-8568
Page 28 of 34
XIV.
PAYMENT FOR STUDIES
1.
Third party payers
All costs associated with the routine clinical care of patients will be the 
responsibility of the patient or their insurance carrier.  
2.
Reimbursement to participants
The patients will receive no financial compensation for participation in this research 
protocol. All study visits will be scheduled at the same time clinical visits are 
planned.  
The FDA authorized cost recovery for this study. The FDA determined that the cost 
of Alpha Beta CD19 Depleted Stem Cell graft using Miltenyi Biotech CliniMACS 
Immunomagnetic Selection Device may be billed to patients and/or their insurance 
company. 
XV.
METHOD TO BE USED IN PROCURING CONSENT OF SUBJECTS
All prospective patients will have the study explained by a member of the research 
team.  All the potential hazards and possible adverse reactions will be explained to 
patient/parent.
Prior to the initiation of the study, acknowledgement of the receipt of this 
information and the subject's freely tendered offer to participate will be obtained 
in writing from each subject in the study.  Assent for subjects 7-10 years old will not 
be obtained due to the complex nature of the therapy and the limited capability of 
the minor to reasonably consider all the implications associated with this treatment 
that may be of direct benefit to the subject. 
This protocol, informed consent, assent form, and any amendments to the protocol 
will be reviewed and approved by the CHMCC IRB prior to initiation.  The study will 
not be initiated without the approval of the IRB.
At reaching the age of majority (typically 18 years of age), an attempt will be made 
to reconsent the research participant at their next visit to the CCHMC BMT clinic. 
If it will be more than 12 months from the time of their 18th birthday to their next 
planned visit, contact via phone and via mail will be made for their consideration 


Protocol v01.27.2021
IRB# 2018-8568
Page 29 of 34
to continue participation. The call and letter will explain that an attempt will be 
made to reconsent the research participant at their next visit. In addition, study 
staff contact information will be provided in case they have questions or wish to 
inform us of their desire to withdraw from participation prior to their next planned 
visit.


Protocol v01.27.2021
IRB# 2018-8568
Page 30 of 34
XVI.
REFERENCES
1.
Copelan EA. Hematopoietic stem-cell transplantation. N Engl J Med. 2006;354(17):1813-
1826.
2.
Rocha V, Locatelli F. Searching for alternative hematopoietic stem cell donors for pediatric 
patients. Bone Marrow Transplant. 2008;41(2):207-214.
3.
Ferrara JL, Levine JE, Reddy P, Holler E. Graft-versus-host disease. Lancet. 
2009;373(9674):1550-1561.
4.
Petersdorf EW. Which factors influence the development of GVHD in HLA-matched or 
mismatched transplants? Best Pract Res Clin Haematol. 2017;30(4):333-335.
5.
Pulsipher MA. Haplo is the new black. Blood. 2014;124(5):675-676.
6.
Bertaina A, Pitisci A, Sinibaldi M, Algeri M. T Cell-Depleted and T Cell-Replete HLA-
Haploidentical Stem Cell Transplantation for Non-malignant Disorders. Curr Hematol Malig Rep. 
2017;12(1):68-78.
7.
Reisner Y, Hagin D, Martelli MF. Haploidentical hematopoietic transplantation: current 
status and future perspectives. Blood. 2011;118(23):6006-6017.
8.
Martelli MF, Aversa F. Haploidentical transplants using ex vivo T-cell depletion. Semin 
Hematol. 2016;53(4):252-256.
9.
Aversa F, Tabilio A, Terenzi A, et al. Successful engraftment of T-cell-depleted 
haploidentical "three-loci" incompatible transplants in leukemia patients by addition of 
recombinant human granulocyte colony-stimulating factor-mobilized peripheral blood progenitor 
cells to bone marrow inoculum. Blood. 1994;84(11):3948-3955.
10.
Aversa F, Tabilio A, Velardi A, et al. Treatment of high-risk acute leukemia with T-cell-
depleted stem cells from related donors with one fully mismatched HLA haplotype. N Engl J Med. 
1998;339(17):1186-1193.
11.
Chaleff S, Otto M, Barfield RC, et al. A large-scale method for the selective depletion of 
alphabeta T lymphocytes from PBSC for allogeneic transplantation. Cytotherapy. 2007;9(8):746-
754.
12.
Knight A, Madrigal AJ, Grace S, et al. The role of Vdelta2-negative gammadelta T cells 
during cytomegalovirus reactivation in recipients of allogeneic stem cell transplantation. Blood. 
2010;116(12):2164-2172.
13.
Halary F, Pitard V, Dlubek D, et al. Shared reactivity of V{delta}2(neg) {gamma}{delta} T 
cells against cytomegalovirus-infected cells and tumor intestinal epithelial cells. J Exp Med. 
2005;201(10):1567-1578.
14.
Pitard V, Roumanes D, Lafarge X, et al. Long-term expansion of effector/memory Vdelta2-
gammadelta T cells is a specific blood signature of CMV infection. Blood. 2008;112(4):1317-1324.
15.
Moretta L, Pietra G, Vacca P, et al. Human NK cells: From surface receptors to clinical 
applications. Immunol Lett. 2016;178:15-19.
16.
Locatelli F, Moretta F, Brescia L, Merli P. Natural killer cells in the treatment of high-risk 
acute leukaemia. Semin Immunol. 2014;26(2):173-179.
17.
Airoldi I, Bertaina A, Prigione I, et al. gammadelta T-cell reconstitution after HLA-
haploidentical hematopoietic transplantation depleted of TCR-alphabeta+/CD19+ lymphocytes. 
Blood. 2015;125(15):2349-2358.
18.
Lamb LS, Jr., Henslee-Downey PJ, Parrish RS, et al. Increased frequency of TCR gamma 
delta + T cells in disease-free survivors following T cell-depleted, partially mismatched, related 
donor bone marrow transplantation for leukemia. J Hematother. 1996;5(5):503-509.
19.
Lang P, Feuchtinger T, Teltschik HM, et al. Improved immune recovery after 
transplantation of TCRalphabeta/CD19-depleted allografts from haploidentical donors in pediatric 
patients. Bone Marrow Transplant. 2015;50 Suppl 2:S6-10.


Protocol v01.27.2021
IRB# 2018-8568
Page 31 of 34
20.
Maschan M, Shelikhova L, Ilushina M, et al. TCR-alpha/beta and CD19 depletion and 
treosulfan-based conditioning regimen in unrelated and haploidentical transplantation in children 
with acute myeloid leukemia. Bone Marrow Transplant. 2016;51(5):668-674.
21.
Locatelli F, Merli P, Pagliara D, et al. Outcome of children with acute leukemia given HLA-
haploidentical HSCT after alphabeta T-cell and B-cell depletion. Blood. 2017;130(5):677-685.
22.
Mancusi A, Ruggeri L, Velardi A. Haploidentical hematopoietic transplantation for the cure 
of leukemia: from its biology to clinical translation. Blood. 2016;128(23):2616-2623.
23.
Liu JH, Kanakry CG, Luznik L. Have haploidentical transplants replaced umbilical cord 
transplants for acute leukemias? Curr Opin Hematol. 2018;25(2):103-111.
24.
Lang P, Teltschik HM, Feuchtinger T, et al. Transplantation of CD3/CD19 depleted 
allografts from haploidentical family donors in paediatric leukaemia. Br J Haematol. 
2014;165(5):688-698.
25.
Huenecke S, Bremm M, Cappel C, et al. Optimization of individualized graft composition: 
CD3/CD19 depletion combined with CD34 selection for haploidentical transplantation. 
Transfusion. 2016;56(9):2336-2345.
26.
Bethge WA, Faul C, Bornhauser M, et al. Haploidentical allogeneic hematopoietic cell 
transplantation in adults using CD3/CD19 depletion and reduced intensity conditioning: an 
update. Blood Cells Mol Dis. 2008;40(1):13-19.
27.
Locatelli F, Bauquet A, Palumbo G, Moretta F, Bertaina A. Negative depletion of 
alpha/beta+ T cells and of CD19+ B lymphocytes: a novel frontier to optimize the effect of innate 
immunity in HLA-mismatched hematopoietic stem cell transplantation. Immunol Lett. 2013;155(1-
2):21-23.
28.
Locatelli F, Pende D, Mingari MC, et al. Cellular and molecular basis of haploidentical 
hematopoietic stem cell transplantation in the successful treatment of high-risk leukemias: role of 
alloreactive NK cells. Front Immunol. 2013;4:15.
29.
Korngold R, Sprent J. T cell subsets and graft-versus-host disease. Transplantation. 
1987;44(3):335-339.
30.
Shlomchik WD. Graft-versus-host disease. Nat Rev Immunol. 2007;7(5):340-352.
31.
Handgretinger R, Schilbach K. The potential role of gammadelta T cells after allogeneic 
HCT for leukemia. Blood. 2018;131(10):1063-1072.
32.
Dolstra H, Fredrix H, van der Meer A, de Witte T, Figdor C, van de Wiel-van Kemenade 
E. TCR gamma delta cytotoxic T lymphocytes expressing the killer cell-inhibitory receptor p58.2 
(CD158b) selectively lyse acute myeloid leukemia cells. Bone Marrow Transplant. 
2001;27(10):1087-1093.
33.
Godder KT, Henslee-Downey PJ, Mehta J, et al. Long term disease-free survival in acute 
leukemia patients recovering with increased gammadelta T cells after partially mismatched 
related donor bone marrow transplantation. Bone Marrow Transplant. 2007;39(12):751-757.
34.
Bertaina A, Zorzoli A, Petretto A, et al. Zoledronic acid boosts gammadelta T-cell activity 
in children receiving alphabeta(+) T and CD19(+) cell-depleted grafts from an HLA-haplo-identical 
donor. Oncoimmunology. 2017;6(2):e1216291.
35.
Pende D, Marcenaro S, Falco M, et al. Anti-leukemia activity of alloreactive NK cells in 
KIR ligand-mismatched haploidentical HSCT for pediatric patients: evaluation of the functional 
role of activating KIR and redefinition of inhibitory KIR specificity. Blood. 2009;113(13):3119-3129.
36.
Bertaina A, Merli P, Rutella S, et al. HLA-haploidentical stem cell transplantation after 
removal of alphabeta+ T and B cells in children with nonmalignant disorders. Blood. 
2014;124(5):822-826.
37.
Balashov D, Shcherbina A, Maschan M, et al. Single-Center Experience of Unrelated and 
Haploidentical Stem Cell Transplantation with TCRalphabeta and CD19 Depletion in Children with 
Primary Immunodeficiency Syndromes. Biol Blood Marrow Transplant. 2015;21(11):1955-1962.


Protocol v01.27.2021
IRB# 2018-8568
Page 32 of 34
APPENDIX 1.  ACUTE GVHD GRADING SCALE – MODIFIED GLUCKSBERG INDEX
Grading of acute GVHD will be based on the following 2 tables and recorded as I, II, III, or IV as 
defined in table 2.
  
Table 1. Staging of Organ Systems in aGVHD
Stage
Skin
Liver
(Bilirubin)
Intestinal tract
(Diarrhea) 
0
No rash
<35 μmol/l or
<2.0 mg/dl
None or
<500 ml/day or 
280ml/m2/day
1
Maculopapular rash, 
<25% of body surface
35-52 μmol/l or
2.0-3.0 mg/dl 
500-1000 mL/day or 
280-555ml/m2/day
2
Maculopapular rash, 
25-50% of body 
surface
53-103 μmol/l or
3.1-6.0 mg/dl
1000-1500 ml/day or 
556-833 ml/m2/day
3
Generalized 
erythroderma
104-256 μmol/l or
6.1-15.0 mg/dl
>1500 ml/day or
 >833ml/m2/day
4
Generalized 
erythroderma with 
bullae formation and 
desquamation
>256 μmol/l or
>15.0 mg/dl
Severe abdominal pain with 
or without ileus
•
Use burns chart or rule of nines:
o
Arm (all over) 9% Front 18% Head 9% Palm 1% Leg (all over) 18% Back 18% 
Genitals 1%
•
Range given as total bilirubin.  Downgrade one stage if an additional cause of elevated 
bilirubin has been documented.
•
If urine contamination suspected do not count
•
General total volume of diarrhea applies for adults.  Downgrade one stage if an 
additional cause of diarrhea has been documented.
•
Histological evidence of aGVHD in the stomach or duodenum.
Table 2. Glucksberg grading of aGVHD 
Skin
Liver
Intestinal Tract
Grade 0
Stage 0
Stage 0
Stage 0
Grade I
Stage 1-2
None
None
Grade II
Stage 3
Stage 1
Stage 1
Grade III
Stage 2-3
Stage 2-4
Grade IV
Stage 4
Stage 4
-


Protocol v01.27.2021
IRB# 2018-8568
Page 33 of 34
APPENDIX 2: STOPPING RULES PLAN
Patients 
Enrolled
Stop if # with TRM 
within 100 days 
following HCT is 
greater than or equal 
to
10
3
11
3
12
3
13
4
14
4
15
4
16
5
17
5
18
5
19
5
20
6
21
6
22
6
23
6
24
7
25
7
26
7
27
7
28
7
29
8
30
8
31
8
32
8
33
9
34
9
35
9
36
9
37
9
38
10
39
10
40
10
41
10
42
11
43
11
44
11
45
11
46
11


Protocol v01.27.2021
IRB# 2018-8568
Page 34 of 34
47
12
48
12
49
12
50
12
51
12
52
12
53
13
54
13
55
13
56
13
57
13
58
14
59
14
60
14
61
14
62
14
63
14
64
14
65
15
66
15
67
15
68
15
69
15
70
15
71
15
72
15
73
15