Clinical trial NCT02582775
Conditions: Epidermolysis Bullosa
Sponsor: Masonic Cancer Center, University of Minnesota
Start date: 2016-03-01; Completion date: 2023-07-26
Enrollment: 17 participants; Allocation: NON_RANDOMIZED; Intervention model: PARALLEL

Brief summary:
This is a single-institution, phase II study to determine the event-free survival at 1 year post allogeneic transplant and serial mesenchymal stem cell (MSC) infusions from a related donor (HLA identical, mismatched or haploidentical) or matched unrelated donor for the biochemical correction of severe epidermolysis bullosa (EB).

Detailed description:

Arms: arm 1: Recessive dominant epidermolysis bullosa (RDEB) patients treated per study regimen on a post-transplant cyclophosphamide allogeneic hematopoietic cell transplant (HCT) platform conditioned with reduced intensity chemotherapy and low dose total body irradiation (300 cGy), followed post-HCT by serial infusions of donor-derived mesenchymal stromal cells (MSC). arm 2: Junctional epidermolysis bullosa (JEB) patient treated per study regimen on a post-transplant cyclophosphamide allogeneic hematopoietic cell transplant (HCT) platform conditioned with reduced intensity chemotherapy and low dose total body irradiation (300 cGy), followed post-HCT by serial infusions of donor-derived mesenchymal stromal cells (MSC). arm 3: Recessive dominant epidermolysis bullosa (RDEB) patients treated per study regimen on a post-transplant cyclophosphamide allogeneic hematopoietic cell transplant (HCT) platform conditioned with reduced intensity chemotherapy and low dose total body irradiation (400 cGy), followed post-HCT by serial infusions of donor-derived mesenchymal stromal cells (MSC). arm 4: Epidermolysis bullosa patients treated per study regimen with chemotherapy and stem cell transplant without mesenchymal stem cell infusions. arm 5: Epidermolysis bullosa patients treated regardless of original transplant arm with re-transplant using 300 cGy of TBI. arm 6: HLA-matched epidermolysis bullosa patients treated with hematopoietic cell transplant alone using 200 cGY BID of TBI (400 cGy total). arm 7: HLA-mismatched epidermolysis bullosa patients treated with hematopoietic cell transplant alone using 200 cGy BID of TBI (400 cGy total) + addition of low dose busulfan for recipients of HLA-mismatched bone marrow arm 8: HLA-mismatched epidermolysis bullosa patients treated with hematopoietic cell transplant plus serial MSC infusions using 200 cGy BID of TBI (400 cGy total) + addition of low dose busulfan for recipients of HLA-mismatched bone marrow
Interventions: intervention 1: Thymoglobulin intervention 2: Cyclophosphamide intervention 3: Fludarabine intervention 4: Total Body Irradiation intervention 5: Bone marrow infusion intervention 6: Tacrolimus intervention 7: Mycophenolate Mofetil intervention 8: Donor mesenchymal stem cell infusions intervention 9: Busulfan
Intervention descriptions: intervention 1: 0.5 mg/kg IV over 6 hours on day -9 and 2 mg/kg IV over 4 hours on day -8 and day -7 with premeds and solumedrol through day -2 intervention 2: 14.5 mg/kg IV over 1 hour day -6 and -5 50 mg/kg IV over 2 hours with mesna 40 mg/kg IV day +2 and +3 intervention 3: 30 mg/m2 IV over 60 minutes days -6 through day -2 intervention 4: See arm description for dosing. intervention 5: Bone marrow infusion on Day 0 intervention 6: Day +5 through day +100 with goals of 5-10 ug/L (not used for HLA-identical related donors). When used in non-MSD recipients, tapered over 6-8 weeks starting at day +100. intervention 7: 15 mg/kg IV q8h (based upon actual body weight) with the maximum total daily dose not to exceed 3 grams. Day +5 through day 35 intervention 8: Day 60, 100 and 180 (collected during donor BM harvest for graft) intervention 9: busulfan IV over 3 hours on days -3 and -2 for HLA-mismatched BM recipients only (Arms F and G)

Protocol documents:
Prot_SAP_000.pdf: 
University of Minnesota 
Department of Pediatrics 
Blood & Marrow Transplant Program 
 
 
Biochemical Correction of Severe Epidermolysis Bullosa 
 by Allogeneic Cell Transplantation and Serial Donor Mesenchymal 
Cell Infusions 
 
CPRC 2015LS076 
MT2015-20 
IND 14166 
 
Principal Investigator/IND Sponsor 
Jakub Tolar, MD, PhD 
 
 
Co-Investigators: 
Bruce R Blazar, MD* 
Kathryn Dusenbery, MD* 
Christen Ebens, MD, MPH 
John A McGrath, MD* 
Kristen Hook, MD* 
David H McKenna, Jr., MD* 
Paul J Orchard, MD  
Mark J Osborn, PhD* 
John E Wagner, MD 
Angela Smith, MD 
Troy Lund, MD 
Kristina M. Nelson, PharmD* 
Cathryn Jennissen, PharmD, BCOP* 
*will not consent study participants 
 
Biostatistician: 
Todd E. DeFor, MS 
 
 
Version Date: 
February 5, 2019 
 


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CPRC #2015LS076 
 
Revision History 
Revision 
Number 
Version 
Date 
Summary of Changes 
Consent 
change? 
 
07/09/2015 
original to CPRC 
n/a 
 
10/07/2015 
In response to CPRC’s initial review: 
• Corrected post-transplant CTX dose and schedule to make 
consistent throughout document 
• Added language to section 5 permitting the use of a known 
carrier as the donor in certain situations 
• Updated section 4.1 that sexually active patients must be on 
contraception 
Additional edits to section 10 including delete day 28 and day 60 
skin biopsies and associated procedures, move several 
procedures from the research x chart to standard of care as SOC 
for this disease; clarifications in the donor chart – corrected 
chimerism sample language and clarified skin sample at point of 
BM biopsy will only be obtained on donors collected at the UMN, 
delete section 8.1 as mobilized apheresis is not used in this study 
n/a 
1 
01/05/2016 
Throughout protocol document – clarify collection of additional 
BM for MSC production at the time of collection for transplant is 
optional for the donor. Create two treatment arms, based on 
donor’s consent status – no/refuses additional BM collection = 
Arm A transplant only, yes/agrees to additional BM collection = 
Arm B transplant + MSC infusions 
Section 7.5 – MSC Administration - remove language regarding 
collection of BM cells for MSCs and their manufacturing and 
storage and move to section 8 BM Harvest and MSC Production 
Section 8 – separate into subsections of bone marrow harvest for 
transplant, risks of BM donation, additional marrow harvest for 
MSC’s in consenting donors and manufacture and storage of 
MSCs 
Section 10.2 – Research Related Procedures – move procedures 
associated with MSC infusions to new section and label this 
section as Arms A and B; delete reference to QOL as determined 
by play score/performance status in section 10.1 
Section 10.3  insert new section titled Research Related 
Procedures (Arm B) to indicate MSC only related procedures and 
assessments 
Section 11.2.2 – clarify targeted toxicity assessment to be done 
on all patients at the day 60, 100, and 180 time points  
Title page, page 3 and section 12.3 – change IND sponsor from 
John Wagner to Jakub Tolar 
yes 


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CPRC #2015LS076 
 
Revision 
Number 
Version 
Date 
Summary of Changes 
Consent 
change? 
Other minor clarifications and edits through-out 
2 
04/04/2016 
Update design throughout protocol including synopsis and 
section 3 Study Design – creates two treatment arms to 
accommodate donors who refuse to consent to the collection of 
addition bone marrow for the manufacturing of MSC cells for the 
serial infusions:  Arm A – HCT only, Arm B – HCT plus serial 
post-transplant MSC infusions  
Section 1.3, synopsis - Add correlative (research related, non-
clinical) objectives section and move two of the secondary 
objectives to this section;  
Sections 14.1 and 14.2 – update endpoints to match objectives, 
update and expand statistical analysis 
Section 5.5 – delete lactating as a donor exclusion criteria – add 
rationale with associated citations 
Clarifications and Edits: 
Schema page – redesign presentation using symbols to improve 
understanding of treatment plan 
Section 5.6 – clarify donor match selection and priority 
Section 7.1.1 and Synopsis/Schema – clarify rabbit ATG 
(hymoglobulin®) to be used, replace Solumedrol with the generic 
name methylprednisolone as the pre-med 
Section 7.1.4 – update TBI to current language 
Section 7.4.1 – add a target dose range for tacrolimus 
Section 7.5 – clarify MSC will be thawed at “bedside”, delete 
sentence regarding study stopping rule 
Section 8.1 - update (decrease) targeted cell dose during BM 
harvest. Add unrelated donor targeted cell dose 
Section 8.3 – change marrow target for MSC production from a 
range (25-35 ml) to 30 ml 
Section 10.1 – clarify SOC clinical evaluations by adding 
footnotes, add carnitine testing and other minor edits 
Section 10.2 – add description of iscoreEB   
Other minor edits and clarifications 
yes 
3 
07/06/2016 
Add option for re-transplant in event of graft failure (lack of 
engraftment, autologous recovery, loss of graft) either on this 
protocol (using Arm C) or on the UMN BMT Graft Failure protocol 
(MT2013-06) 
Though-out document – increase the window (from ±7 to ±14 
days) for each MSC infusion 
Yes, 
plus 
new 
re-
transplant 
consent 


MT2015-20:  Stem Cell Transplant and Serial MSC Infusions for EB 
 
 
 
 
 
 
 
 
 
 
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Revision 
Number 
Version 
Date 
Summary of Changes 
Consent 
change? 
Synopsis, Section 1.2, Section 14.1.2 - correct wording for 2nd 
objective/endpoint from double chimerism to CD3+ and CD15+ 
and specify all time points to be done per x-chart in section 10.1 
Synopsis, Section 4.1 and Appendix I – simplify disease related 
eligibility, simplify renal eligibility requirement 
Section 7.2 – edit for clarification infusion of BM cells 
Section 7.4.1 – expand tacrolimus administration details 
Section 10 – add baseline pregnancy testing for patients and 
donors 
Section 10.1 – move iscorEB and associated footnote from 
research x chart to SOC 
Section 10.4 – move baseline chimerism to screen 
Section 13 and 14.1.3 – indicate that skin samples may be 
collected at additional time points as clinically indicated 
Section 14.2 – clarify that Arms A and B will be combined for 
assessment of the primary, secondary and transplant related 
endpoints 
Section 14.4 – create separate stopping rules based on diagnosis 
(JEB vs RDEB) – clarify graft failure as aplastic and define TRM 
as transplant related mortality 
Section 15 – add 3 additional references 
Through-out document change iscorEB nomenclature (don’t 
capitalize the “i”) – not tracked 
Other administration edits and clarifications through-out 
5 
08/23/2016 
--Synopsis, sections 1.2 and 14.1.2: Expand secondary objective 
of comparing baseline iscoreEB to the year 2 iscoreEB, replace 
CD3+ and CD15+ with myeloid and lymphoid, respectively  
--Synopsis, sections 3 and 14:  increase annual enrollment to up 
to 25 patients 
--Schema, sections 2.3, 3, 7, 7.4.1 and 10.1:  omit tacrolimus for 
patients with an HLA identical donor 
--Schema, sections 3, 7.5, 7.8, 10.3:  clarify that the time points 
for MSC infusions are the targeted time points but may be altered 
at the discretion of the treating physician without being 
considered a protocol deviations 
--Schema, sections 3, 7.8:  clarify for a re-transplant (arm c), the 
original donor is preferred but not required 
--Section 4.1 and appendix I:  replace pulmonary function 
requirement for oxygen saturation to adequate in the opinion of 
the enrolling investigator  
yes 


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Revision 
Number 
Version 
Date 
Summary of Changes 
Consent 
change? 
--Sections 5 and 10.4 and appendix II:  clarify that unrelated 
donors will be assessed, consents and collected per local donor 
center; simplify donor criteria 
--Section 7.1.3 Remove IV hydration from pre-transplant Cytoxan 
dosing plan 
--Sections 7.5, 9.3,10.3 and 10.4 headers – include “ and if 
applicable, Arm C” 
--Section 7.7: simplify duration of study participation language 
Add Drs Angela Smith, Troy Lund and Weston Miller as co-
investigator 
5.1 
10/24/2016 
Administrative amendment (affects Section 7, Treatment Plan, 
only) 
• Allow infusion of MSCs through peripheral intravenous line (PIV) 
when a central line is not available.  
• Revise O2 monitoring guidelines to allow monitoring at regular 
intervals and if clinically indicated during the 2 hours post-MSC 
administration  
No 
6 
11/02/2016 
--Formally incorporates changes from administrative amendment 
5.1 
--Synopsis, Schema and Study Design added Arms D and E to 
capture data on subjects treated at the higher dose of TBI 
--Schema & Section 7.1.4 Increased dose of TBI to 400 cGy for 
first transplant 
--Section 3 added rationale for administering remaining MSC 
cells in the case of a second transplant and increasing the dose 
for TBI 
--Section 10.1 clarified daily assessments only until engraftment 
--Section 14.2, 14,3 and 14,4 clarified statistical analysis and 
safety monitoring plan for after TBI dose increased from 300 to 
400 cGy 
 
No 
7 
11/23/2016 
--Schema, Sections 2.3, 3, 7, and 14 clarified that TBI 
administration will be separated into two fractions of 200 cGy each 
(on a single day); other minor edits 
Yes 
8 
02/09/2017 
Section 4.1, 5, 10.1, 10.4, eligibility checklist – PRA results added 
to donor screening  
Section 4.2, eligibility checklist - beta 3 laminin JEB mutants added 
to exclusions 
Section 5 – marrow may be supplemented with cryopreserved cord 
blood 
No 
9 
09/14/2017 
 
 
Synopsis, Schema, section 2.2, 3, 7, 7.5, 8.3, 10.1, 10.3 -- 
increased number of MSC infusions 
Schema, section 1.3, 3, 7, 7.6.1, 95, 10.1, 10.4, 14.1.3, Appendix II 
– added CelluTome skin grafting to supportive care 
Section 10.1 & 13 – clarified number of skin samples taken 
Yes 


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Revision 
Number 
Version 
Date 
Summary of Changes 
Consent 
change? 
9A 
10/31/2017 
Synopsis; section 2.3 -  Clarification requested by CPRC review of 
Amendment 9 (stipulation referred to change made in Amendment 
8). 
No 
9B 
10/31/2017 
Correction of typographical errors in schema (arm names) 
No 
NA 
12/11/2018 
Consent revision as requested by IRB 
Yes 
10 
2/05/2019 
Updated investigator team 
Closure of Arms A and B to enrollment due to excess non-
neutropenic graft failure and replacement by subsequent arms 
Addition of Arms F and G for HLA-mismatched stem cell recipients 
– the treatment regimens on these arms have increased intensity 
(addition of busulfan) to prevent graft failure 
Addition of Appendix VI regarding busulfan dose selection, AUC 
monitoring and algorithm for dose modification 
Addition of clinical monitoring of CelluTome skin grafting 
Yes 
 
 
 
 
 
 
 
 
 
PI/IND Sponsor Contact Information: 
Jakub Tolar, M.D., Ph.D.  
Professor  
Hematology/Oncology and Transplantation 
Department of Pediatrics 
MMC 366 Mayo 
420 Delaware St SE 
Minneapolis, MN  55455 
612 626-2912 (phone) 
612 624-3913 (fax) 
tolar001@umn.edu (email) 
 


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TABLE OF CONTENTS 
SCHEMA ..................................................................................................................................12 
1 
Study Objectives .............................................................................................................14 
1.1 
Primary Objective 
14 
1.2 
Secondary Objectives 
14 
1.3 
Correlative Objectives 
14 
1.4 
Transplant Related Objectives 
14 
2 
Background .....................................................................................................................15 
2.1 
Dystrophic Epidermolysis Bullosa (DEB) 
15 
2.2 
Allogeneic Hematopoietic Cell Transplantation in EB 
16 
2.3 
Rationale for Treatment Plan 
26 
3 
Study Design ...................................................................................................................31 
4 
Patient Selection .............................................................................................................33 
4.1 
Inclusion Criteria 
34 
4.2 
Exclusion Criteria 
34 
5 
Donor Selection ...............................................................................................................34 
6 
Patient and Donor Registration ......................................................................................37 
6.1 
Registration with the Masonic Cancer Center Clinical Trials Office 
37 
6.2 
Patients Who Are Enrolled and Do Not Receive Study Treatment 
37 
6.3 
Re-Transplant for Graft Failure  
37 
7 
Treatment Plan ................................................................................................................38 
7.1 
Preparative Therapy (day – 9 through day -1) 
40 
7.2 
Hematopoietic Cell (Bone Marrow) Infusion (day 0) 
41 
7.3 
Post-Transplant Cyclophosphamide 
42 
7.4 
GVHD Prophylaxis 
42 
7.5 
Serial MSC Infusions (months 2, 4, 6; and optionally at 8, and 10) – Arms E, G and if 
Applicable, Arm C 
43 
7.6 
Concurrent Therapy and Supportive Care 
43 


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7.7 
Duration of Treatment 
45 
7.8 
Re-Transplant For Graft Failure 
45 
7.9 
Duration of Study Participation 
46 
8 
Bone Marrow Harvest and MSC Production ..................................................................46 
8.1 
Bone Marrow Harvest for Transplant 
46 
8.2 
Risks of Donating Bone Marrow 
47 
8.3 
Additional Marrow Harvest for MSC Production with Donor’s Consent ......................47 
8.4 
MSC Manufacture and Storage 
47 
9 
Expected Risks of Study Treatment ...............................................................................47 
9.1 
Preparative Regimen 
47 
9.2 
Transplant Procedures 
49 
9.3 
MSC Infusions (Arm E, Arm G and, if applicable, Arm C) 
49 
9.4 
GVHD Prophylaxis 
50 
9.5 
Epidermal Grafting Using the CelluTome® 
50 
10 Clinical Evaluations ........................................................................................................52 
10.1 
Clinical Evaluations – All Patients Standard of Care 
52 
10.2 
Research Related Assessments and Procedures (All Arms) 
54 
10.3 
Research Related Assessments and Procedures (Arms where MSCs are given) .....54 
10.4 
Donor 
54 
11 Adverse Event Monitoring, Recording and Reporting ..................................................55 
11.1 
Definitions 
55 
11.2 
Adverse Event Monitoring, Recording and Reporting 
56 
11.3 
Monitoring for Stopping Rule Events 
58 
12 Data Collection and Monitoring ......................................................................................59 
12.1 
Data Management 
59 
12.2 
Case Report Forms 
59 
12.3 
Data and Safety Monitoring Plan (DSMP) 
59 
12.4 
Record Retention 
60 


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13 Disease Outcome Measures ...........................................................................................60 
14 Statistical Considerations ..............................................................................................61 
14.1 
Statistical Endpoints 
61 
14.2 
Statistical Analysis 
62 
14.3 
Design and Sample-Size Justification 
63 
14.4 
Toxicity Monitoring and Stopping Rules 
64 
15 Conduct of the Study ......................................................................................................66 
15.1 
Good Clinical Practice 
66 
15.2 
Ethical Considerations 
66 
16 References .......................................................................................................................66 
Appendix I – Eligibility Checklist - Patient ............................................................................71 
Appendix II – Eligibility Checklist – Donor ............................................................................73 
Appendix III – Lansky Play Score and Karnofsky Performance Status ...............................74 
Appendix IV – GVHD Scoring .................................................................................................75 
Appendix V – Targeted Toxicities (For MSC Infusions Only) ...............................................76 
Appendix VI – Low Dose Busulfan Dose Selection (For Arms F and G Only) ....................77 
 
 
 
 
 
 


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SYNOPSIS 
Study 
Design: 
This is a single-institution, phase II study to determine the event-free survival at 1 year 
post allogeneic transplant with or without serial mesenchymal stem cell (MSC) infusions 
from a related donor (HLA identical, mismatched or haploidentical) or matched unrelated 
donor for the biochemical correction of severe epidermolysis bullosa (EB).  
 
A single marrow harvest is performed to collect the stem cells for the transplant 
procedure and, with the donor’s consent, the collection of an additional 40-50 ml sample 
for MSC production for post-transplant infusion on month (± 14 days) 2, 4, and 6; and 
additionally offered at 8 and 10 months. If the donor refuses to consent for the extra 
marrow collection, the patient will receive only the transplant. The patient will be enrolled, 
based HLA-matching and on the donor’s consent to one of the following arms: 
Arm A:  stem cell transplant alone using 300 cGy TBI (closed to enrollment) 
Arm B:  stem cell transplant plus serial MSC infusions using 300 cGy TBI (closed to 
enrollment) 
Arm D:  stem cell transplant alone using 200 cGy BID TBI for 8/8 HLA-matched bone 
marrow recipients 
Arm E:  stem cell transplant plus serial MSC infusions using 200 cGy AND TBI for 8/8 
HLA-matched bone marrow recipients 
Arm F: stem cell transplant along using 200 cGy BID TBI plus addition of low dose 
Busulfan for HLA-mismatched bone marrow recipients 
Arm G: stem cell transplant plus serial MSC infusions 200 cGy BID TBI plus addition of 
low dose Busulfan for HLA-mismatched bone marrow recipients 
 
In event of graft failure (failure to engraft, autologous recovery or loss of graft), patients 
may be eligible for re-transplant on this study (Arm C) or through the University of 
Minnesota BMT protocol MT2013-06 (if <1 year from first BMT) or on this study (Arms D-
G) (if >/=1 year from first BMT).  
Primary 
Objective: 
To estimate the event-free survival rate by 1 year post-transplant with an event defined as 
death or a 50% increase in a patient’s iscoreEB from baseline 
Secondary 
Objectives: 
• To document the percentage change of a patient’s iscoreEB at 1 and 2 years as 
compared with baseline 
• To determine the incidence of transplant-related mortality (TRM) at day 180  
• To describe health quality of life at 1 and 2 years as compared to pretreatment results 
• To determine proportion of lymphoid and myeloid chimerism at various time-points 
(days 28, 60, 100, 180, and year 1 and 2) among surviving patients 
Key 
Eligibility: 
Diagnosis of severe form of EB with documented collagen, laminin, integrin, keratin or 
plakin deficiency 
0-25 years of age 
An available healthy HLA matched or partially HLA matched related or unrelated donor 
for HSC graft and MSC  
Adequate renal, hepatic, cardiac and pulmonary function 


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Key 
Exclusion: 
Evidence of HIV infection or known HIV positive serology; current active serious infection; 
diagnosis of squamous cell carcinoma; beta-3 laminin JEB, donor with EB; pregnancy  
Enrollment 
Plan: 
Enroll up to 25 patients per year for a total of 69 patients 
An optimal interim analysis for futility without suspension of accrual due to the long 
primary endpoint of event-free survival at 1 year post HCT will be performed after 36 
patients are enrolled. Given the change in conditioning (300 cGy TBI to 200 cGy bid TBI, 
then inclusion of low dose busulfan), enrollment will start over for the purpose of 
analyses; the currently enrolled 15 patients plus the planned new cohort of 69 patients for 
a total of 84. 


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SCHEMA 
 
Treatment arms based on HLA-match and donor’s level of consent 
• 
ARM D or ARM F:  stem cell transplant (if donor refused collection of additional BM cells for 
MSC production) Arm D: 200 cGy BID TBI effective November 2016  Arm F: Addition of busulfan 
for HLA-mismatched BM effective January 2019 
• 
ARM E or ARM G:  stem cell transplant plus serial MSC infusions (if donor consented to 
additional BM cell collection for MSC production) Arm E: 200 cGy BID TBI effective November 
2016   Arm G: Addition of busulfan for HLA-mismatched BM effective January 2019 
 
 
HLA-matched 
HLA-mismatched 
Donor consents for MSC production 
Arm E 
Arm G 
Donor refuses MSC production 
Arm D 
Arm F 
 
 
                                                                                                                                    ARM E/G only 
                                                                         HCT                                                   MSC Infusions  
                
 
 
 
 
 
 
 
 
 
every 2 months through day +300 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
             -9      -8     -7       -6     -5     -4       -3     -2      -1       0     +1      +2    +3     +4    +5       +28            60   100   180    365      720    
 
 
 
 
 
 
 
 
            tacrolimus through day 100 (omit if HLA identical donor) 
 
 
 
 
 
 
 
 
            MMF through day 35 or 7 days after engraftment 
 
 
 
 
 
 
 
 
                            CelluTome Wound healing start d+100 
Preparative Regimen 
rabbit ATG (Thymoglobulin®) 0.5 mg/kg IV over 6 hours on day -9 with pre-meds (acetaminophen, 
diphenhydramine and methylprednisolone) and   
rabbit ATG (Thymoglobulin®) 2 mg/kg IV over 4 hours on day -8 and day -7 with pre-meds (acetaminophen, 
diphenhydramine and methylprednisolone). Continue methylprednisolone taper through day -2. 
cyclophosphamide 14.5 mg/kg IV over 1 hour on days -6 and -5 with mesna 2.9 mg/kg IV 5x/daily  
fludarabine 30 mg/m2 IV over 60 minutes on days -6 through day -2   See section 7.1.2 for dose adjustments 
based on renal function, age or weight.  
busulfan IV over 3 hours on days -3 and -2 for HLA-mismatched BM recipients only (Arms F and G). See 
section 7.1.3 and Appendix VI for dose based on weight. 
TBI:  total body irradiation 200 cGy BID (400 cGy total) day -1 (reduce to 300 cGy if re-transplanted on Arm C) 
 
Hematopoietic Cell Transplant  
HCT:  hematopoietic cell infusion 
 
Post HCT chemotherapy 
cyclophosphamide 50 mg/kg IV over 2 hours with mesna 10 mg/kg IV 5x/day on day +3 and +4 
 
GVHD Prophylaxis 
tacrolimus day +5 through day +100, then taper (omit if HLA identical donor – 8/8 match) 
mycophenolic acid mofetil (MMF) day +5 through day +35 or 7 days after engraftment 
 
Epidermal Grafting Using the CelluTome® Epidermal Harvesting System 
Offered up to 3 sessions starting day +100 then additional sessions at 12 week intervals if clinically indicated 
 
ARM E, and G only^  
Serial MSC Infusions  


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MSC: donor mesenchymal stem cell infusions offered post-transplant every 2 months (±14 days) through 10 months 
post BMT; however these time points may be altered on an individual patient basis at the discretion of the treating 
investigator. 
 
Re-Transplant Option for Graft Failure (not part of the above schema) 
In the event of graft failure, time from first transplant should be considered when determining the intensity of 
conditioning required.  
If <1 year since the initial BMT, two options are available: 
1) 
Re-transplant the patient within this protocol (ARM C) 
2) 
Re-transplant by enrolling patient on the University of Minnesota BMT protocol MT2013-06: Treatment of 
Graft Failure after HSCT (PI – T. Lund) while continuing follow-up per this protocol  
If >/= 1 year since the initial BMT: Re-transplant on this protocol as a new subject on ARM D-G, whichever is 
appropriate. 
Ideally the same donor would be used for both transplants; however it is recognized this is not always an option. 


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1 
Study Objectives 
1.1 Primary Objective 
To estimate the event-free survival rate by 1 year after transplant with an event 
defined as death or a 50% increase in a patient’s iscoreEB from baseline.  
1.2 Secondary Objectives 
• To document the percentage change of a patient’s iscoreEB at 1 and 2 years 
as compared to baseline 
• To determine the incidence of transplant related mortality (TRM) at day 180  
• To describe health quality of life at 1 and 2 years as compared to pretreatment 
results based on Lansky Play Score (Karnofsky if > 16 years) 
• To determine proportion of lymphoid and myeloid chimerism at various time-
points (days 28, 60, 100, 180, and year 1 and 2) among surviving patients 
1.3 Correlative Objectives 
• To describe the pattern of biochemical improvement as measured by an 
increase in protein expression (collagen, laminin, integrin, keratin or plakin) 
and related structural and physical changes at days 100 and 180, then at 1 
and 2 years 
• To describe the pattern and durability of donor MSC engraftment in the skin 
(ARM B, E, and G) 
• To describe wound healing for the patients who receive CelluTome as 
supportive care. 
1.4 Transplant Related Objectives 
• Estimate the probability of survival at 1 year  
• Determine the incidence of grade II-IV and grade III-IV acute graft versus host 
disease (GvHD) at day 100 
• Determine the incidence of chronic graft versus host disease at 1 year 
• Time to neutrophil engraftment (defined as the first of three consecutive days 
after HCT that the patient’s absolute neutrophil counts is ≥ 0.5x109 per liter) 
• Determine the incidence of sustained neutrophil engraftment (neutrophil 
engraftment without autologous recovery) 
• Time to platelet engraftment (defined as the first time after HCT that the 
patient can sustain platelet count ≥ 20x109 per liter for three consecutive 
days) 
• Determine incidence of bacterial, viral and fungal infections at 1 year 


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2 
Background 
2.1 
Dystrophic Epidermolysis Bullosa (DEB) 
Dystrophic EB (DEB) is a group of heritable mechanobullous skin diseases 
characterized by skin fragility, blister formation, and scarring. The most severe 
forms of DEB are characterized by mutilating scarring, blisters over large 
proportions of the body surface and later on, mitten deformities, joint contractures, 
esophageal strictures, corneal erosions, chronic cutaneous infections and 
aggressive squamous cell carcinoma (SCC)1-3. Children and adults with recessive 
dystrophic epidermolysis bullosa (RDEB) are often faced with a life of pain, often 
dying of SCC. SCC can appear as early as 13 years of age with 50% having SCC 
if they survive to age 40 years. The prognosis of a DEB patient with SCC is very 
poor with nearly all dying of metastatic disease. Patients with the severe DEB have 
profound physical disabilities. Daily activities (e.g., toileting, feeding, bathing, 
walking) are major challenges. Quality of life also tends to decline with age. 
Caretakers are consumed by the day to day care of the child with DEB. 
 
Type VII collagen (C7) is synthesized by both human keratinocytes and fibroblasts. 
It is secreted within the basement membrane zone (BMZ) lying between the 
epidermis and dermis of skin. C7 is the major component of anchoring fibrils (AF) 
which are necessary for normal epidermal-dermal adherence. Genetic defects in 
the collagen type VII gene, COL7A1, result in RDEB. Ultrastructurally, abnormal 
morphology and low numbers or absence of AFs results in cleavage and 
detachment of the sublamina densa from the underlying dermis and extreme skin 
fragility (Figure 1). DEB can be inherited as an autosomal dominant or autosomal 
recessive disease with blisters and scarring often present at birth or shortly 
thereafter. The blisters in the neonate often cover the whole body, including oral 
and esophageal mucosa, and it continues throughout life.  


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Several new therapies targeting wounds in RDEB have been developed, including 
local cell therapy with skin fibroblasts4, 5 and mesenchymal stem/stromal cells,6 
gene therapy with gamma-retroviral7 or lentiviral vectors,8-10 and C7 protein 
replacement therapy.11-13 These innovations emanated from decades of preclinical 
molecular and cellular studies by highly motivated teams around the globe, 3, 5, 14-
20. Despite the ‘impossible-to-forget’ appearance of a child with RDEB, the 
pathology of generalized severe RDEB is best understood as a global one, with 
multiple external and internal organ systems affected (cutaneous and 
gastrointestinal) and with whole body responses (such as compromised nutrition, 
anemia, pruritus, chronic inflammation, local and systemic infections) amplifying 
the injury of the underlying C7 deficiency. Therefore, as an alternative to local 
gene, cell and protein therapies, we sought systemic and possibly permanent 
cross-correction of C7 in the extracellular matrix using durable transplantation of 
self-renewing blood and marrow stem cells. 
2.2 
Allogeneic Hematopoietic Cell Transplantation in EB 
To investigate a possibility of systemic stem cell therapy for generalized severe 
RDEB, we transplanted congenic wild-type bone marrow cells (among a variety of 


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other stem cell types) into neonatal mice with a targeted disruption of the Col7a1 
gene21 that results in widespread blistering and early death due to the absence of 
C7 expression. We hypothesized that there was a stem cell population in bone 
marrow that homed to injured skin near the DEJ and produced cellular progeny 
which secreted wild-type C7 protein in the area where it was needed. While various 
non-hematopoietic and hematopoietic cell populations from bone marrow failed to 
successfully correct the disease, it turned out that an infusion of highly purified 
bone marrow progenitors (CD150+ CD48- cells),22  migrated to injured skin and 
secreted C7. AFs were partially restored, and blisters on paws healed23 within the 
14 day window prior to death of untreated affected pups. Thus, donor cells capable 
both of secreting C7 and homing to the injured mucocutaneous membranes led to 
partial correction of the disease phenotype. With this functional correction of C7 in 
a murine model of human RDEB, data from Chino et al.24 also demonstrating 
similar effects with  prenatal CD90-depleted bone marrow, and absence of any 
other curative approach for  severe RDEB, we sought to examine the safety and 
efficacy of allogeneic blood and marrow  transplantation as a treatment for  children 
with the severest forms of RDEB. 
 
Hematopoietic cell transplantation. Our first clinical trial25 of systemic cellular 
therapy (hematopoietic cell transplantation, HCT) for a genodermatosis 
demonstrated that a) donor cells home to injured skin and do so in unexpectedly 
high numbers, b) expression of C7 is increased and sustained for many years after 
blood and marrow transplantation, and c) AFs gradually appear and increase in 
numbers. Clinically, nearly all patient have experienced some improvement in the 
maintenance of overall skin integrity (Figure 2). However, none of the HCT 
recipients has become 
entirely free of wounds, 
and 
several 
children 
experienced 
severe 
complications, including 
death as a result of the 
HCT-related toxicities of 
the 
high-dose 
chemotherapy used as 
myeloablative 
conditioning (MAC) or 
RDEB 
disease 
progression, 
or 
both. 
Therefore, 
with 
the 
ultimate 
aim 
of 


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improving the efficacy and safety of HCT for the systemic and durable therapy of 
generalized severe RDEB, we sequentially modified the HCT treatment protocol 
(Figure 3) with the addition of supplemental cells and reduced intensity 
conditioning (RIC). The former, addition of mesenchymal stem/stromal cells 
(MSC), has been based on identification of non-hematopoietic donor cells in skin 
of HCT recipients (Figure 4), on the capacity of MSC to secrete C7, and on their 
immune-modulatory potential to mitigate graft-versus-host disease, one of the 
common immune complications of allogeneic HCT. The latter, RIC, has been 
introduced to assess whether less than complete hematopoietic chimerism can 
mediate adequate clinical response in the setting of conditioning regimen that is 
expected to result in less morbidity and mortality after allogeneic HCT. 
 
Patients treated to-date. In total, between 2007 and 2014, 26 individuals (0.4-
20y) with life-threatening, severe generalized form of RDEB have undergone 
allogeneic HCT. Of these, 13 were treated with a MAC (busulfan, fludarabine 
[FLU], and cyclophosphamide [CY]) and 13 were treated with RIC (CY, FLU, anti-
thymocyte globulin, and low dose total body irradiation). RDEB subjects received 
HCT from related (N=15, all bone marrow grafts), or an unrelated donor (N=11; 6 


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bone marrow grafts and 5 umbilical cord blood grafts). For the recent 20 individuals 
(the first 6 have been reported in reference 25), the patient characteristics, grafts 
and outcomes are summarized in Table 1 for MAC and Table 2 for RIC. In a cohort 
of patients with another generalized severe form of EB, junctional EB (JEB), two 
children were transplanted using the MAC, and four using the RIC. Four of six 
patients died between days 42 and 146 after HCT, predominantly from progression 
of their primary genodermatosis. As severe JEB is genetically heterogeneous, it is 
relevant to note that the two surviving individuals with severe JEB have loss-of-
function mutations in LAMA3 gene, while those who died had genetic inactivation 
of LAMB3 and, in one case ITGB4 gene. As only six JEB patients have been 
treated since the study was opened there are still too few to accurately assess 
outcome/efficacy for this subgroup. Thus the review below has been confined to 
the 26 RDEB patients. 


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Table 1. Patient characteristics, grafts and outcomes for RDEB with myeloablative conditioning (MAC). 
Patient  
RII-1 
RII-2 
RII-3 
RII-4 
RII-5 
RII-6 
RII-7 
Sex 
Age at HCT 
Female 
2 years 
Female 
9 years 
Male 
13 years 
Female 
1 years 
Male 
5 years 
Male 
4 years 
Male 
10 years 
Clinical 
features1 
Bloody emesis, corneal 
abrasions, anal fissure 
Esophageal 
strictures 
Pseudo-syndactyly, 
IDA, esophageal 
strictures, wheelchair
bound 
Neonatal seizure 
Esophageal 
strictures, 
aspiration 
pneumonia 
Neonatal sepsis, 
pneumonia, colitis, 
esophageal strictures, 
constipation 
Neonatal intubation, 
esophageal strictures, 
Pseudo-syndactyly, 
corneal erosions, 
contractures, IDA 
Donor: sex,  
HLA match 
UCB, female, 5/6 
1st UCB 
transplant, female, 
5/6; 2nd UCB 
transplant, female, 
5/6 
UCB, female, 4/6 
Sister BM, 8/8 
Brother BM, 
8/8 
Unrelated BM, 
female, 8/8 
Brother BM, 8/8 
Cell dose2 
11 x 107 
5.5 x 107 
4.4 x 107 
8.17 x 107 
5.23 x 108 
1.41 x 108 
3.1 x 108 
3.5 x 108 
HCT day 
2/19/2010 
3/25/2010 and 
4/27/2010 
4/29/2010 
7/2/2010 
9/10/2010 
12/30/2010 and 
7/14/12 
3/11/2011 
Severe 
adverse 
events 
Respiratory failure, 
PTLD/EBV viremia 
Graft failure, 
HHV6 viremia 
Chronic GvHD 
(extensive), 
seizure, colitis, 
HHV6 viremia 
RF: HD, VOD, 
mucositis: 
intubated, capillary 
leak syndrome, 
heart failure 
Upper GI 
bleeding after 
esophageal 
dilatation 
Fungemia, 
bronchopulmonary 
hemorrhage; RF: HD; 
PTLD: late graft 
failure, successful 
second transplant 
RF: HD, acute GvHD 
(skin 3, grade 2), 
seizure, upper GI 
bleeding, respiratory 
failure, fungal sepsis 
Skin donor 
chimerism3 
30% 
0% 
33% 
24% 
21% 
14% 
97% 
Biochemical 
correction 
C7 increased 
C7 increased4 
C7 not increased 
C7 not increased 
C7 not 
increased 
C7 not increased 
C7 increased 
Outcome: 
before → 
after HCT 
75% → <25% BSA 
involved 
Graft failure 
No change 
No change 
75% → <5% BSA 
involved 
90% → <5% 
BSA involved 
90% → 0% BSA 
involved 
90% → <35% → 90% 
BSA involved5 
Survival 
(as of 1/31/15) 
Alive 
Alive  
Died of cGvHD on 
day 496 
Died of VOD on 
day 101 
Alive 
Alive 
Died of sepsis on day 
212 
1 All patients had: >55% skin surface covered with erosions and blisters; history of skin colonization and infections; FTT; pruritus, and pain. 2total nucleated cells/kilogram of recipient’s body weight. 
Four hours after hematopoietic cells, all patients received single intravenous dose of 2 x 106 HLA-unmatched (3rd party) mesenchymal stromal cells. 3Highest value shown. 4The C7 increase occurred 
early after HCT (day 28) and later comparisons were not possible, or indicated. 5The erosions healed between days 60 and 100, then progressively worsened.  
Legend:  BM, bone marrow; UCB, umbilical cord blood; BSA, body surface area; C7, collagen type VII; CB, cord blood; CSA, cyclosporine; EBV, Epstein-Barr virus; FTT, failure to thrive; GI, 
gastrointestinal; GvHD, graft-versus-host disease; HD, hemodialysis; HHV6, human herpes virus type 6; HLA, human leukocyte antigen; IDA, iron-deficient anemia; MA, myeloablative; PTLD, post-
transplant lymphoproliferative disease; RDEB, recessive dystrophic epidermolysis bullosa; RF, renal failure; GI, gastrointestinal; VOD, veno-occlusive disease (sinusoid obstruction syndrome). 


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Table 2A. Patient characteristics, grafts and outcomes for RDEB with reduced intensity conditioning (RIC, 200cGy TBI). 
Patient  
RIII-1 
RIII-2 
RIII-3 
RIII-4 
RIII-5 
RIII-6 
RIII-7  
Sex 
Age at HCT 
Male 
20 years 
Female 
2 years 
Female 
6 years 
Female 
7 years 
Male 
9 months 
Male 
3 years 
Female 
11 mo 
Clinical 
features1 
Severe contractures, 
wheelchair bound, 
pseudo-syndactyly, 
IDA, dental carries, 
depression 
Bloody emesis, 
corneal 
abrasions 
Esophageal 
strictures, 
corneal 
abrasions 
Esophageal 
strictures, 
corneal 
abrasions 
Hygroma, IDA, 
FTT,  corneal 
abrasions 
Esophageal 
strictures, corneal 
abrasions,  
Bloody emesis 
Donor: sex, HLA 
match 
Unrelated BM, 8/8 
Brother BM, 8/8 
Brother BM, 
8/8 
Unrelated CB, 
4/6 
Unrelated BM, 
8/8 
Unrelated BM, 8/8 
Unrelated BM, 8/8 
Cell dose2 
5.00 x 108 
6.21 x 108 
7.28 x 108 
9.1 x 108 
9.91 x 108 
9.00 x 108 
7.20 x 108 
HCT day 
6/7/2011 
9/15/2011 
2/23/2012 
4/6/2012 
7/13/2012 
6/4/2013 
7/16/2013 
Severe adverse 
events 
Colitis 
Pneumonia, 
mixed chimerism 
(DLI) 
Bacteremia 
Graft failure 
Bacteremia, 
mixed 
chimerism (DLI) 
TEN, 
hemophagocytosis, 
fungal sepsis  
Pruritus 
Skin donor 
chimerism3 
23% 
2% 
7% 
0% 
15% 
11% 
3% 
Biochemical 
correction 
C7 increased 
C7 increased 
Continuous 
expression of 
C7 
C7 not 
increased 
C7 increased 
C7 increased 
C7 not increased 
Outcome: 
before→ 
after HCT 
90% → <10% BSA 
involved 
75% → <10% 
BSA involved 
75% → <5% 
BSA involved 
75% → <50% 
BSA involved 
75% → <5% 
BSA involved 
90% → <5% → 
>90% (secondary 
to TEN) BSA 
involved 
75% → <5% BSA 
involved 
Survival 
(as of 1/31/15) 
Alive  
Alive  
Alive  
Alive  
Alive  
Died of sepsis on 
day 152 
Alive  
 
 
 


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Table 2B. Patient characteristics, grafts and outcomes for RDEB with reduced intensity conditioning (RIC, 300cGy TBI). 
Patient 
RIII-8 
RIII-9  
RIII-10 
RIII-11 
RIII-12 
RIII-13 
Sex 
Age at HCT 
Male 
3 years 
Female 
1 year 
Male 
11 months 
Female 
4 years 
Female 
4 months 
Male 
6 months 
Clinical 
features1 
Esophageal 
strictures 
Severe oral mucosa 
blistering 
Corneal 
abrasions 
Esophageal 
strictures, 
corneal 
abrasions 
Mild oral 
mucosa 
blistering 
Moderate oral 
mucosa blistering 
Donor: sex, HLA 
match 
Sister BM, 8/8 
Sister BM, 7/8 
Mother BM, 8/8 
Sister BM, 8/8 
URD BM, 8/8 
Sister BM, 8/8 
Cell dose2 
3.53 x 108 
4.5 x 108 
2.45 x 108 
3.35 x 108 
4.27 x 108 
5.51 x 108 
5.5 x 108 
HCT day 
8/20/2013 and 
9/3/2014 
5/7/2014 
6/24/2014 
9/24/2014 
10/14/2014 
12/02/2014 
Severe adverse 
events 
Graft failure 
(successful second 
transplant) 
Acute GvHD (skin 3, 
grade 2), EBV; 
bacteremia 
Mixed chimerism 
(DLI); 
bacteremia 
Mixed 
chimerism 
(DLI); EBV 
viremia; 
bacteremia 
Graft failure; 
pericardial 
effusion 
Mixed chimerism 
(DLI); 
intussusception 
Skin donor 
chimerism3 
17% 
22% 
28% 
0% 
8% 
13% 
Biochemical 
correction 
C7 not increased 
Continuous 
expression of C7  
Continuous 
expression of C7 
C7 not 
increased 
C7 not 
increased 
Continuous 
expression of C7 
Outcome: 
before→ 
after HCT 
75% → <5% BSA 
involved 
75% → <5% BSA 
involved 
75% → <10% 
BSA involved 
75% → <10% 
BSA involved 
55% → <5% 
BSA involved 
55% → <5% BSA 
involved 
Survival 
(as of 1/31/15) 
Alive  
Alive  
Alive  
Alive  
Alive  
Alive  
 
1 All patients had: >55% skin surface covered with erosions and blisters; history of pruritus and pain. 2total nucleated cells/kilogram of recipient’s body weight. Four hours after 
hematopoietic cells all patients received single intravenous dose of 2 x 106 HLA unmatched (3rd party) mesenchymal stromal cells. 3Highest value shown. 
Legend:  BM, bone marrow; BSA, body surface area; C7, collagen type VII; FTT, failure to thrive; HLA, human leukocyte antigen; IDA, iron-deficient anemia; DLI, donor lymphocyte 
infusion; EBV, Epstein-Barr viremia; TEN, toxic epidermal necrolysis; RDEB, recessive dystrophic epidermolysis bullosa. 


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Safety outcomes. The overall probability of 2-year survival in the MAC cohort 
(N=13) was 69% (95% CI, 37-87 %) versus 90% (95% CI, 47-99%) for the RIC 
(N=13) cohort (p=0.3). Of the 20 recent RDEB patients, three individuals died 
before one year after HCT (MAC: RII-4 of veno-occlusive disease, RII-7 of fungal 
sepsis; RIC: RIII-6 of fungal sepsis); one patient died after one year (MAC: RII-3 
of extensive chronic GvHD; Tables 1 and 2). Three individuals did not engraft with 
donor cells (MAC: RII-2, RIC: RIII-4, RIII-8; Tables 1 and 2). There were no 
statistically significant survival differences in comparison of related versus 
unrelated grafts, bone marrow versus umbilical cord blood grafts, and those who 
received additional MSC versus those who have not. One individual in each cohort 
(MAC: RII-7, RIC: RIII-9) developed acute GvH (grade 2, with stage 3 skin 
involvement - erythroderma); one individual developed extensive chronic GvH 
(MAC: RII-3).  
 
Efficacy outcomes. Our primary efficacy measure was the event-free survival rate 
by 1 year after HCT with an event defined as death or failure to have demonstrable 
increase in C7 or other biochemical, structural or physical measure of 
improvement. The overall probability of 2-year event-free survival in the MAC 
cohort (N=13) was 62% (95% CI, 31-82 %) versus 82% (95% CI, 44-95%) for the 
RIC (N=13) cohort (p=0.3).  
 
Donor-recipient chimerism. Hematopoietic engraftment was near-complete in 
those treated with MAC. As expected with lower doses of conditioning 
chemotherapy, engraftment was significantly lower in those treated with RIC: on 
average 26% in CD15 cells and 55% on CD3 cells 100 days after HCT. The 
maximum skin chimerism in those with successful hematopoietic engraftment  was 
on average 31% in MAC cohort, 10% in RIC with 200cGy TBI, and 15% in RIC 
with 300cGy (Tables 1 and 2). Thus skin chimerism correlated proportionally with 
hematopoietic one, and provided structural and functional links between bone 
marrow and biochemical, structural and physical measures of mucocutaneous 
changes after HCT.  
 
Biochemical measure - C7 expression. We observed increase in C7 
immunofluorescence in two RDEB individuals treated with MAC (RII-1 and RII-7; 
in addition to those reported previously), and in four individuals treated with RIC 
(RIII-1, RIII-2, RIII-5 and RIII-6). In support of a mechanistic model whereby 
engrafted bone marrow serves continuously as a source of cells capable of 
migration to injured skin and secretion of C7, in individuals available for long term 
assessment, we were able to detect this increase at 1-3 years after HCT. In several 
others the change in C7 expression after HCT was less obvious as they showed, 


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despite severe clinical phenotype, baseline C7 immunoreactivity before HCT (RIC: 
RIII-3, RIII-9, RIII-10, RIII-13).  
 
Structural measure - Anchoring fibrils (AFs). AFs are homotrimers of C7 and 
the most direct structural foundation of the C7 function at the DEJ. In some 
individuals (for example, RII-1) we saw no AFs despite increased C7 expression 
after HCT; in others (for example, RIII-1 and RIII-10) AFs have been seen after 
HCT.  
 
Physical measure - Functional skin stability. Given that our ultimate goal for 
RDEB individuals is functional skin, we aimed to assess its integrity before and 
after HCT in a quantitative way. We employed a suction blister device whereby 
constant negative pressure is applied to skin and time to blistering is proportionate 
to capacity of skin layers to adhere to each other. As AFs mediate connectivity of 
epidermis and dermis the longer the time the skin is able to withstand the negative 
pressure applied to it is a measure of C7 expression, polymerization, and its 
functional integration in skin. Typically, individuals with RDEB experience blistering 
within 10 minutes, healthy people in approximately 60 minutes, and the obligatory 
heterozygotes (such as, the parents of RDEB children) in 30-50 minutes. Critically, 
in several representative examples from both MAC and RIC trials, serial 
assessments of RDEB subjects after HCT showed gradual increase in skin 
resilience under pressure (Figure 5).  


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Clinical assessment. The individual metrics of biochemical, structural and 
physical changes in skin, including the compound measure of functional skin 
stability with suction blister device, while critical to local quantitative assessment 
of treatment intervention, do not capture entirely the complex pathology in its 
impact on health of 
individuals 
with 
RDEB. 
However, 
clinical 
photographs 
documenting skin 
responses as they 
differ 
among 
individuals 
with 
RDEB, unveil the 
impact of HCT in 
the most definitive 
way (Figure 6). 


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2.3 
Rationale for Treatment 
Plan 
The key motivation for changing 
the 
HCT 
conditions 
in 
the 
individuals with EB is to improve 
efficacy and safety. The reduced 
intensity 
chemotherapy 
(RIC) 
approach 
has 
dramatically 
improved the safety (treatment-
related morbidity and mortality). 
However, 
hematopoietic 
chimerism has been mixed and, 
expectedly, skin chimerism has 
been lower than that seen in the MAC cohort. Assuming that production of C7 (and 
other cutaneous adhesive molecules) is directly proportionate to the number of 
donor cells in skin, the higher skin chimerism is desirable, and—based on our 
comparison between MAC versus RIC groups—achievable with complete 
hematopoietic chimerism. The possibility of an acceptable safety profile together 
with near-complete hematopoietic engraftment has recently become possible with 


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improvements in related HLA-haploidentical HCT using high dose post-transplant 
cyclophosphamide26-28.  
The key benefits of this post-grafting strategy are [1] a dramatic reduction of severe 
GvHD (which has been limiting for early versions of haploidentical HCT), [2] the 
ability to limit post-HCT immunosuppression and thus allow for earlier and 
qualitatively superior immune reconstitution, and [3] the rapid availability of a 
donor29-32. For patients with EB specifically, this translates to [1] less skin injury (by 
GvH reaction) in an already damaged skin, [2] better and faster establishment of 
immune response in people with multiple open wounds, typically already colonized 
or infected with polymicrobial drug-resistant species, and [3] shortening the time 
between the decision to transplant and the HCT itself, which is particularly relevant 
in generalized severe JEB, which has a mean survival of less than 6 months of 
age.  
Therefore,  we propose to adapt the most robust and tested approach to deplete 
alloreactive cells in vivo by [1] administering high doses of cyclophosphamide in a 
narrow window after HCT (originally developed by scientists at the John Hopkins 
University School of Medicine but now adopted nationally and internationally for 
many neoplastic as well as non-malignant indications)28, 33-35. This plan uses 
combination chemotherapy (fludarabine and cyclophosphamide), immunotherapy 
(thymoglobulin), and radiotherapy (total body irradiation) prior to HCT. Post-
transplant immunosuppressive therapy consists of high-dose cyclophosphamide 
with MESNA protective therapy, followed by tacrolimus (omit for persons with HLA 
identical (8/8) related donors) and mycophenolic acid mofetil (MMF) as described 
in detail in Section 7 (Treatment Plan).  
Induction of tolerance with post-HCT cyclophosphamide is bidirectional: 
eradication of both donor and host alloreactive T cells reduces the incidence of 
both GvH and host versus graft (HvG) reactions36, 37. This is relevant for EB 
cohorts, as we have observed graft failure in several individuals (Tables 1 and 2). 
In order to advance from the state-of-art current platform, we will prioritize 
matched-sibling HLA-identical donors over T-cell-replete haploidentical donors 
and HLA-matched unrelated donors—using the same conditioning before and after 
HCT. Therefore, with or without crossing the HLA barrier, we expect to be able to 
rapidly identify a favorable donor for nearly all patients. 
As RDEB and JEB are inherited genodermatoses, the haploidentical donor pool 
might be reduced but likely only minimally (in relatives with autosomal dominant 
mutations that are present in the compound heterozygosity state in the proband 
with autosomal recessive disease). Moreover, it is relevant that carriers of 


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autosomal recessive mutations (parents, the obligatory heterozygotes) are 
typically asymptomatic, and matched-sibling donors that are heterozygotic for 
these mutations have been used with success as HCT donors (with no discernible 
clinical difference in outcomes of such heterozygous-matched sibling HLA-
identical HCT and homozygous wild-type matched sibling HLA-identical or HLA-
matched unrelated donor HCT; Tables 1 and 2).  
Moreover, systemic infusions of mesenchymal stromal/stem cells (MSCs) have 
been found safe in a large number of disorders, including RDEB6, 38-44. As they are 
not HLA-matched to the recipient, and sometimes not to the HCT donor, as 
expected these “third party MSCs” are rejected and thus their effects are limited, 
typically to several weeks. Our present approach is based on the advantage of 
most donors being related (matched sibling donors or haploidentical family 
members) to the recipient and the ability to culture MSCs from marrow at the time 
of HCT harvest. Accordingly, we will obtain, screen, and qualify these MSC 
cultures in time for systemic infusions at months 2, 4, and 6; and additionally 
offered at 8, and 10 months. The cell dose will be 1-2 million MSCs per kilogram 
of recipient weight; a dose that has repeatedly been shown to be safe in a clinical 
setting.  
The sum of experimental and clinical data supports the conclusion that, contrary 
to the prevailing professional opinion of the last several decades, protein 
replacement therapy by allogeneic blood and marrow transplantation is not limited 
to freely diffusible molecules, such as enzymes, but also large structural proteins, 
such as collagens and perhaps laminins. While HCT improve the integrity of the 
skin and mucous membranes, future strategies will be built on the improvements 
of HCT (this protocol) and on concept of combination therapy (Figure 7). 


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Rationale for Re-Transplant in the Event of Graft Failure 
Haploidentical hematopoietic cell transplantation (haplo-HCT) has become a new 
standard of care for selected malignant disorders58,59. This has been enabled by T 
cell depletion, either ex vivo (by T cell depletion of the graft) or in vivo (by post-
transplant cyclophosphamide, PTCy, administration). Remarkably, this strategy 
has been shown to be reasonably safe and highly efficacious in non-malignant, 
typically genetic, disorders treatable by HCT. For example, sickle cell disease, 
thalassemia, mucopolysaccharidosis type I, dyskeratosis congenita, or Fanconi 
anemia have been treated successfully with haplo-HCT60-68. One of the limitations 
of haplo-HCT, especially with better tolerated reduced intensity conditioning, is 
failure to engraft. Fortunately, second transplants for either aplastic graft failure or 
recovery of autologous hematopoiesis have been almost always successful69-72. 
This is further supported by haplo-HCT data from diseases that present as 
“natural” graft failure-equivalents, that is, bone marrow failures such as seen in 
dyskeratosis congenita, Fanconi anemia or idiopathic severe aplastic anemia—all 
of which have been shown to respond favorably to allo-HCT. Therefore, our 


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approach of using second transplant after graft failure in patients with generalized 
severe EB, is expected to result in similar favorable safety and efficacy profile. This 
is likely to be further enhanced by the use of mesenchymal stromal/stem cells from 
allo-HCT donor after the second allo-HCT transplant. The plan for re-transplant 
includes administration of additional MSC doses after the second transplant, 
following the same schedule as the initial transplant. With depletion of marrow 
during the second transplant, the rationale for this repeated MSC administration is 
not different from the rationale for MSC administration in the initial transplant. MSC 
dosing at month 2, 4, 6, 8, and 10 may promote engraftment, skin healing and 
diminish the risk of graft versus host disease. In addition to this rationale, 
administration of MSCs following a second transplant will ensure consistency in 
the plan of care and outcome data.  
Rationale for Increased Dose of TBI (300 cGy to 200 cGy BID) 
As of November 2016, we have had incidences of autologous recovery after 
transplant (i.e. engraftment of the patient's own cells after transplant, rather than 
the donor cells). Although not unexpected, we do strive to reduce these 
occurrences as much as possible. Dr. Tolar, the study Sponsor-Investigator, 
consulted with other experts in BMT at the University of Minnesota, as well as 
international leaders in haploidentical transplant, to identify a plan to better 
promote engraftment of the donor cells. The resounding suggestion has been to 
increase the dose of TBI to 200 cGy BID (rather than 300 cGy). Leading BMT MDs 
in haploidentical transplants at Johns Hopkins (Drs. Symons, Luznik & Fuchs) 
have implemented this change in a similar patient population (i.e. non-malignant 
disease, marrow not previously prepped with chemotherapy/radiation), with much 
improved full donor engraftment (4/5 patients had full donor engraftment with 400 
cGy TBI vs 11/34 patients with 200 cGy TBI) and no additional ill effects 
experienced by patients. (unpublished data, personal communication) 
 
Rationale for Exclusion of beta-3 laminin JEB 
Interim analysis of early subjects found no lasting benefit for the patients with beta 
3 laminin 332 mutation.  
 
Rationale for Addition of Low Dose Busulfan for HLA-mismatched stem cell 
recipients 
As of November 2018, five of 14 (36%) enrolled patients exhibited graft failure with 
autologous recovery (no aplastic graft failure), including 2 of 8 patients receiving 
the increased dose of TBI (200 cGy BID as opposed to a single fraction of 300 cGy 
following the November 2016 protocol modification. A thorough interim evaluation 
of potential factors mediated autologous recovery was completed, considering 


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donor type, presence of recipient anti-HLA antibodies [both panel reactive 
antibodies (PRA) and donor specific antibodies (DSA)], pre-conditioning and day 
0 absolute lymphocyte counts, and bone marrow CD34+ stem cell counts. Graft 
failure with autologous recovery predominated in haploidentical parent recipients 
(4 of 4, 100%; 1 of 3 in haploidentical sibling, 0 of 1 in HLA-matched related donor, 
and 0 of 5 in HLA-matched sibling recipients). Eight of 11 tested patients (3 
receiving matched sibling bone marrow not tested) had positive PRA, though only 
3 had DSA. Of those patients with DSA, the earliest two had graft failure with 
autologous recovery. The 3rd patient with DSA underwent a debulking protocol 
including immunosuppression to decrease production of antibodies and 
plasmapheresis to remove pre-existing antibodies to limit the risk of immune 
mediated graft rejection. This patient had successful donor engraftment. There 
were no statistically significant associations between engraftment and pre-BMT 
absolute lymphocyte count (p=0.92) or day 0 absolute lymphocyte count (p=0.36), 
or CD34+ stem cell dose (p>0.99). 
 
The importance of DSA and potential role in immune mediated graft failure has 
been recognized and mitigation with debulking standardized. This is reflected in 
the current Donor Selection criteria. The engraftment barrier with haploidentical 
parent donors is likely the result of inadequate patient myeloablation. We strive to 
reduce the risk of this complication by escalating the intensity of conditioning for 
recipients of HLA-mismatched grafts with inclusion of low dose busulfan. 
3 Study Design 
This is a single-institution, phase II study to determine the event-free survival at 1 year 
post allogeneic transplant with or without serial mesenchymal stem cell (MSC) 
infusions from a related donor (HLA identical, mismatched or haploidentical) or 
matched unrelated donor for the biochemical correction of severe epidermolysis 
bullosa (EB). 
A single marrow harvest is performed to collect the stem cells for the transplant 
procedure and, with the donor’s consent; an additional 40-50 ml sample is collected 
at this time for the production of MSC for post-transplant infusions. If the donor refuses 
to consent for the extra marrow collection, the patient will receive only the transplant. 
The patient will be enrolled, based on the donor’s consent to either: 
Arm A:  hematopoietic cell transplant alone using 300 cGY of TBI (closed to accrual) 
Arm B:  hematopoietic cell transplant plus serial MSC infusions using 300 cGY of TBI 
(closed to accrual) 


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Arm C:  re-transplant using 300 cGy of TBI (regardless of original transplant arm) if 
<1 year from first BMT 
Effective with the November 2016 version of the protocol: 
Arm D:  hematopoietic cell transplant alone using 200 cGY BID of TBI (400 cGy total) 
for recipients of 8/8 HLA-matched bone marrow 
Arm E:  hematopoietic cell transplant plus serial MSC infusions using 200 cGY BID 
of TBI (400 cGy total) for recipients of 8/8 HLA-matched bone marrow 
Effective with the November 2018 version of the protocol: 
Arm F: hematopoietic cell transplant alone using 200 cGy BID of TBI (400 cGy total) 
+ addition of low dose busulfan for recipients of HLA-mismatched bone marrow 
Arm G: hematopoietic cell transplant plus serial MSC infusions using 200 cGy BID of 
TBI (400 cGy total) + addition of low dose busulfan for recipients of HLA-mismatched 
bone marrow 
A preparative regimen consisting of thymoglobulin, fludarabine, cyclophosphamide, 
and total body irradiation is administered prior to a T cell replete donor marrow 
infusion. Post-transplant cyclophosphamide is given on day 3 and 4 to deplete 
alloreactive cells. 
Graft-versus-host-disease prophylaxis consisting of tacrolimus and mycophenolic acid 
mofetil (MMF) is started on post-transplant day 5 and continued through day 35 or 7 
days after engraftment for MMF and day 100 for tacrolimus (start of taper). Tacrolimus 
is eliminated if a fully matched (HLA identical) donor is used. 
Local wound therapy will be offered as clinically indicated using the FDA approved 
vacuum device (CelluTome®, Regulation number 878.4820). Patients will receive 
local wound therapy using epidermal skin grafting from the same donor (donor 
consent required) that provided the hematopoietic graft. 
For patients enrolled in Arm E and G, MSCs will be infused every 2 months (± 14 
days) through 10 months post-transplant (months 8 and 10 offered if patient is 
available); however these time points may be altered on an individual patient basis at 
the discretion of the treating investigator.  
Disease reassessments will be performed at days 28, 60, 100, and 180, then at 12 
and 24 months post-transplant. 


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In the event of graft failure (e.g. lack of engraftment, autologous recovery, loss of 
graft), time from the first transplant should be considered when determining the 
intensity of conditioning required.  
If the 2nd transplant will occur <1 year since the first transplant, two options are 
available:  
1) Re-transplant the patient within this protocol (Arm C)   
2) Re-transplant the patient by enrolling on the University of Minnesota BMT protocol 
MT2013-06: Treatment of Graft Failure after HSCT (PI – T. Lund) while continuing 
follow-up per this protocol 
If the 2nd transplant will occur >/=1 year following the first transplant: The 2nd BMT 
could be completed on this protocol as a new subject on ARM D-G, whichever is 
appropriate based on HLA-match and donor consent for MSC production. 
Ideally the same donor will be used for the re-transplant; however it is recognized that 
this will not always be possible.  
At the discretion of the treating physician and with the donor’s permission, extra 
marrow for MSCs may be collected at the time of the bone marrow harvest. If the 
patient has donor MSCs available (either from the original transplant or the re-
transplant), they may be infused in the same manner and schedule as detailed after 
the original transplant using day 0 of the second transplant. Targeted toxicities will be 
assessed at time of all MSC infusions.  
All other tests/procedures and follow-up milestone visits would continue to be based 
on the initial transplant day for statistical analysis purposes. For those that are re-
transplanted on arm C, standard of care and research testing will restart at second 
transplant day 0 in addition to the remaining follow-up milestones from initial transplant 
day. The window for these assessments may be expanded so these time points 
coincide if in the best interest of the patient.  
4 Patient Selection 
External Advisory Panel:  Hematopoietic cell transplantation (HCT) for EB has 
spanned dermatology and transplantation biology in a novel fashion, and 
unquestionable RDEB expertise has been critical for success of the study. Therefore, 
we will continue to work with our External Advisory Panel (EAP): Professor John A. 
McGrath (St. John’s Institute of Dermatology, King’s College, London, United 
Kingdom), Professor Alain Hovnanian (Institut Federatif de Recherche Necker Enfants 
Malades, Paris, France) and Professor Katsuto Tamai (Department of Dermatology, 


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Osaka University, Osaka, Japan). The EAP reviewed data prior to and following HCT, 
and make recommendations on whether the EB phenotype of each individual 
considered for HCT is severe enough to make the potential risks of HCT acceptable. 
4.1 
Inclusion Criteria 
• 0 through 25 years of age 
• Diagnosis of a severe form of EB with documented collagen, laminin, integrin, 
keratin or plakin deficiency (by immunofluorescence staining with protein 
specific antibodies or Western blotting or by mutation analysis)  
• Adequate organ function within 4 weeks of study registration defined as: 
- 
Renal: glomerular filtration rate within normal range for age 
- 
Hepatic: bilirubin, AST/ALT, ALP < 5 x upper limit of normal 
- 
Pulmonary: adequate pulmonary function in the opinion of the enrolling 
investigator 
- 
Cardiac: left ventricular ejection fraction ≥ 45%, normal EKG or approved 
by Cardiology for transplant 
• Sexually active participants must agree to use adequate birth control for the 
during the study period (from before the start of the preparative chemotherapy 
through 1 year post-transplant) 
• 
Available donor per section 5: targeted MFI < 1,000 (MFI exceeding 1000 must 
be approved by the PI and treatment team.) 
• Voluntary written consent – adult or parent (with information sheet for minors, 
if applicable) prior to any research related procedures or treatment 
4.2 
Exclusion Criteria 
• 
beta 3 laminin JEB mutants 
• 
Active untreated systemic infection at time of transplantation (including active 
infection with Aspergillus or other mold within 30 days)  
• 
History of HIV infection 
• 
Evidence of squamous cell carcinoma 
• 
Pregnant or breast feeding. Females of child-bearing potential must have a 
negative pregnancy test prior to study registration as the agents administered 
in this study are Pregnancy Category C and D. 
5 Donor Selection 
Unrelated donors will be screened and consented per usual by the Donor Center using 
the Donor Match Selection Criteria found below.  
 


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Donors assessed at the University of Minnesota must meet the following criteria in 
addition to the Donor Match Selection Criteria.  
5.1 
Be medically, socially, and psychologically fit to donate in the opinion of the 
evaluating medical personnel 
5.2 
Does not have epidermolysis bullosa* 
5.3 
Adequate hematologic, hepatic and renal function to undergo the donor 
procedure in the opinion of the evaluating medical personnel 
5.4 
Not pregnant with confirmation of non-pregnancy status within 7 days of 
donation. Lactation is not a contra-indication to donation. Transmission of 
anesthetic medications via breast-milk is variable, dependent upon fat 
solubility of the medication, doses provided to the lactating donor, and infant 
enteral absorption. Transmission of anesthetic medications via breast milk is 
negligible and general anesthesia not a contraindication to continued breast 
feeding / pumping.51-54 
5.5 
Voluntary written consent - adult or parent (with assent for minors, if 
applicable) 
*At the discretion of the primary BMT physician, donors with known positive carrier 
status for EB may be considered if in the opinion of the primary BMT physician, 
the donor is the best available. 
 
Donor Match Selection Criteria: 
HLA-matched sibling (or 1 allele HLA-mismatched related):  HLA-A, B, C, DRB1 
genotypic identical donor, or HLA-A, B, C, DRB1 phenotypic identical donor, or 7/8 
HLA-matched donor at HLA-A, B, C, DRB1 
  
HLA mismatched or haploidentical related donors (including 1st degree relatives 
and half siblings): The donor and recipient must be identical at least one allele of 
each of the following genetic loci: HLA-A, HLA-B, HLA-Cw, HLA-DRB1, and HLA-
DQB1. A minimum match of 5/10 is therefore required, and will be considered 
sufficient evidence that the donor and recipient share one HLA haplotype. 
 
Matched unrelated donors: Unrelated volunteer donor matched for HLA-A, -B, -C 
and -DRB1 defined by high resolution molecular typing.  
 
Donor Source In Decreasing Order Of Priority 
1. Related Donor (Marrow)1, 2 


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a. HLA-A, B, C, DRB1 genotypic identical (sibling) donor 
b. HLA-A, B, C, DRB1 phenotypic identical donor 
c. 7/8 HLA-matched donor at HLA-A, B, C, DRB1 
d. Haploidentical matching at least one allele at HLA-A, HLA-B, HLA-Cw, HLA-
DRB1 & HLA-DQB1 
2. Unrelated Donor (marrow)1 
a. HLA-A, B, C, DRB1 phenotypic identical donor 
b. 7/8 HLA-matched donor at HLA-A, B, C, DRB1 
1Targeted MFI <1,000 for any anti-donor HLA antibody; MFI > 1,000 may be 
considered by PI if no alternative. The PI can approve use of a certain donor even 
if DSA+ and debulk or not based on his/her preference or institutional guidelines.  
2May be supplemented with cryopreserved UCB if available 
  
If there is more than one donor with the same degree of HLA match, the 
following prioritization will be used unless there is HLA cross-match 
incompatibility or a medical reason to select otherwise, in which case donor 
selection is the responsibility of the PI. We will prioritize the lowest number of 
mismatches in the HVG direction (to potentially minimize graft rejection risk). 
3. ABO compatibility (in order of priority) 
a. Compatible or minor ABO incompatibility 
b. Major ABO incompatibility 
4. CMV status 
a. For a CMV seronegative recipient, use a CMV seronegative donor 
b. For a CMV seropositive recipient, use a CMV seropositive donor 
5. Other donor characteristics 
a. The younger ( if over 18) and lighter weight donor should be preferred. 
b. If all else is equal, male donors may be preferred over nulliparous female 
donors who may be preferred over multiparous female donors. 
c. Other factors such as donor age and health history will be integrated into 
the donor selection process per standard practice and may be prioritized 
over HLA, ABO and CMV status. Children donors may be used if 
appropriate. 


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6 Patient and Donor Registration 
Registration will occur after the patient and/or parent/guardian has signed the 
appropriate consent form and eligibility is confirmed, but before any study related 
procedures are performed. To be eligible for registration to this study, the patient must 
meet each of the criteria listed on the eligibility checklist based on the eligibility 
assessment documented in the patient’s medical record. A copy of the eligibility 
checklist is in appendices I and II and under attachments within the study in OnCore 
the Cancer Center’s clinical database. 
Donors will be registered in OnCore per Masonic Cancer Center guidelines. 
6.1 Registration with the Masonic Cancer Center Clinical Trials Office  
Upon completion of the screening evaluation, eligibility confirmation and 
obtaining written consent, the study coordinator or designee will register the 
patient in OnCore.  
The patient will be assigned to a statistical arm based on transplant only or 
transplant plus serial MSC infusions and TBI dose.  
6.2 Patients Who Are Enrolled and Do Not Receive Study Treatment 
If a patient is registered to the study and is later found not able to begin study 
treatment, the patient will be removed from study and treated at the physician’s 
discretion. The study staff will update OnCore of the patient’s non-treatment 
status. Study data will be collected until the time the patient is taken off study. 
The reason for removal from study will be clearly indicated in OnCore. The 
patient will be replaced to complete enrollment.  
If a patient receives any study therapy and then is discontinued, regardless of 
the reason, the patient must be followed per section 7.8. 
6.3 Re-Transplant for Graft Failure  
Patients being re-transplanted due to graft failure within in this study reassigned 
to Arm C (if <1 year from 1st transplant) or Arm D-G as appropriate (if >/=1 year 
from 1st transplant) to release a new calendar and set of case report forms 
(CRFs). For patients re-transplanted on MT2013-06, no reassignment on this 
study is necessary.  
 
 


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7 Treatment Plan 
In order to provide optimal patient care and to account for individual medical 
conditions, investigator discretion may be used in the prescribing of all supportive care 
drug therapy (i.e. acetaminophen, diphenhydramine, G-CSF, antimicrobials, etc.).  
Enrollment is open to both pediatrics and adults; however, the vast majority or all 
patients will qualify for care through the University of Minnesota Masonic Children’s 
Hospital and the treatment plan is written as such. If an adult (>25 years of age) is 
enrolled, he/she would receive all of their care at the appropriate facilities within the 
University of Minnesota Medical Center, Fairview and the adult outpatient clinic(s).  
 
 


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Day 
Drug Or Procedure 
Refer To 
Section: 
-9 
Rabbit ATG (Thymoglobulin®) 0.5 mg/kg IV over 6 hours with pre-meds  
Continue methylprednisolone per taper through day -2 
7.1.1 
-8 
Rabbit ATG (Thymoglobulin®)  2 mg/kg IV over 4 hours with pre-meds 
7.1.1 
-7 
Rabbit ATG (Thymoglobulin®)  2 mg/kg IV over 4 hours with pre-meds 
7.1.1 
-6 
fludarabine 30 mg/m2 IV over 60 minutes* 
cyclophosphamide 14.5 mg/kg IV over 1 hour 
mesna 2.9 mg/kg IV 5x/daily 
7.1.2  
7.1.3 
-5 
fludarabine 30 mg/m2 IV over 60 minutes* 
cyclophosphamide 14.5 mg/kg IV over 1 hour 
mesna 2.9 mg/kg IV 5x/daily 
7.1.2 
7.1.3 
-4 
fludarabine 30 mg/m2 IV over 60 minutes* 
7.1.2 
-3 
fludarabine 30 mg/m2 IV over 60 minutes* 
busulfan IV over 3 hours** 
7.1.2 
-2 
fludarabine 30 mg/m2 IV over 60 minutes* 
busulfan IV over 3 hours** 
7.1.2 
-1 
TBI  200 cGy BID (400 cGy total)  or 300 cGy if re-transplanted on Arm C 
7.1.4 
0 
Hematopoietic Cell Infusion (Transplant) 
7.2 
1 
rest 
- 
2 
rest 
- 
3 
cyclophosphamide 50 mg/kg IV over 2 hours 
mesna 10 mg/kg IV 5x/daily 
7.3 
4 
cyclophosphamide 50 mg/kg IV over 2 hours 
mesna 10 mg/kg IV 5x/daily 
7.3 
5 
tacrolimus (continue until day 100 then taper) – omit if HLA identical 
related donor 
mycophenolic acid mofetil (continue until day 35 or 7 days after 
engraftment) 
7.4.1 
7.4.2 
Serial MSC Infusions- Arm E, Arm G, and if applicable, Arm C (± 14 days).  
60 
MSC Infusion #1 
7.5 
120 
MSC Infusion #2 
7.5 
180 
MSC Infusion #3 
7.5 
240 
MSC Infusion #4*** 
7.5 
300 
MSC Infusion #5*** 
7.5 
Wound Healing with CelluTome Epidermal Harvesting System 
At least 
100 
Offered up to three sessions at least 12 weeks apart 
7.6 
*See Section 7.1.2 for dosing in patients with renal dysfunction and/or < 10 kg body weight. 
**See Section 7.1.3 and Appendix VI for dosing. Only for patients enrolled on Arms F or G. 
***Offered if patient is available for additional infusions 
 


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7.1 Preparative Therapy (day – 9 through day -1) 
The preparative therapy will be administered as written with no plans for dose 
modifications except as described below. 
7.1.1 Rabbit ATG (Thymoglobulin®) 
Thymoglobulin (rATG) will be infused through a 0.22 micro filter. The dose 
will be 0.5 mg/kg IV on day -9 over 6 hours and 2 mg/kg IV on days -8 and 
-7 over 4 hours. 
Pre-medications:  
• Acetaminophen 15 mg/kg/dose (Max Dose = 650 mg) by mouth or 
feeding tube 30 minutes prior to rATG 
• Diphenhydramine 1mg/kg/dose (Max Dose =50 mg by mouth or 
feeding tube 30 minutes prior to rATG  
• Hypersensitivity orders per standard guidelines during rATG 
administration.  
A steroid taper will be given to prevent reactions to rATG as follows: 
• On day -9 to -7, methylprednisolone 1mg/kg IV 1 hour prior rATG. 
This dose may be repeated once, 3 hours after the first dose.  
• On day -6 and -5, methylprednisolone 0.75 mg/kg/ IV as a single 
dose;  
• On days -4 and -3, methylprednisolone 0.5 mg/kg/ IV as a single 
dose;  
• On day -2 methylprednisolone 0.25 mg/kg/ IV as a single dose.  
7.1.2 Fludarabine 
Fludarabine 30 mg/m2/dose IV every 24 hours (adjusted for renal function 
per standard guidelines). Administer over a 60 minute IV infusion on Days 
–6 through –2 (maximum cumulative dose =150 mg/m2).  
For children < 10 kg, fludarabine dosing will be 1 mg/kg IV over 1 hour. If > 
1 year of age, consider converting to mg/kg dosing after discussion between 
the protocol PI and pediatric BMT pharmacist. 
 
If the adjusted creatinine clearance < 70 ml/min, a 20% dose-reduction of 
fludarabine may be considered after discussion between the protocol PI and 
pediatric BMT pharmacist. 
 


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Fludarabine dose adjustments for the purpose of poor renal function will 
not be considered a protocol deviation, but will be noted in OnCore. 
7.1.3 Busulfan (Arms F and G only) 
Busulfan compounding, administration and monitoring should be performed per 
institutional guidelines and infused over 3 hours on days -3 and -2. Busulfan dosing 
will be determined based on the weight of the patients upon admission and per our 
institution busulfan standard of practice dose calculator and nomogram (Appendix 
VI). 
Seizure and VOD prophylaxis are prescribed according to institutional guidelines. 
Concomitant administration of azole anti-fungal agents (except fluconazole) and 
acetaminophen should be avoided within 72 hours before and after busulfan 
administration.  
7.1.4 Pre-Transplant Cyclophosphamide  
Cyclophosphamide 14.5 mg/kg/day will be administered as an IV infusion 
over 1 hour. Cyclophosphamide dosing is calculated based on actual weight 
(ABW) unless ABW is >150% above Ideal Body Weight (IBW) per institution 
guidelines.  
 
Mesna 2.9 mg/kg IV 5 times daily will be administered per institutional 
guidelines. 
7.1.5 Total Body Irradiation (TBI) 
 
For patients undergoing transplant on Arm D, E, F, or G: 
TBI will be given in two fractions on day -1 at least 6 hours apart. Each 
fraction will consist of 200 cGy given with right and left lateral beams 
prescribed to the midplane at the umbilicus at a dose rate of 10-19 
cGy/minute. The total TBI dose will be 400 cGy for patients on either of 
these arms. 
For patients undergoing second transplant on 2015-20 Arm C: 
The dose of TBI will be 300 cGy in a single fraction on day -1 with right and 
left lateral beams prescribed to the midplane at the umbilicus at a dose rate 
of 10-19 cGy/minute. 
7.2 Hematopoietic Cell (Bone Marrow) Infusion (day 0) 
Refer to section 8 for donor collection. 
 
Bone Marrow (T cell replete graft) will be harvested for infusion on day 0. 
 


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In general therefore: 
• No T cell depletion 
Infusion of fresh, unirradiated bone marrow after filtration of bone fragments 
7.3 Post-Transplant Cyclophosphamide 
Cyclophosphamide 50 mg/kg IV over 2 hours on day +3 and day +4 
Mesna and IV fluid hydration will be administered per institutional guidelines.  
Cyclophosphamide dosing is calculated based on actual weight (ABW) unless 
ABW is >150% above Ideal Body Weight (IBW) per institution guidelines.  
 
7.4 GVHD Prophylaxis 
On day 5, patients will begin prophylaxis with Tacrolimus and Mycophenolic Acid 
Mofetil (MMF). Patients will not be eligible for other GVHD prophylaxis studies. 
7.4.1 Tacrolimus (Omit if 8/8 HLA- matched donor) 
Tacrolimus will be initiated on day +5 as a continuous infusion at a starting dose 
of 0.03 mg/kg/day. The patient will be converted to oral dosing when appropriate. 
Tacrolimus will continue until day +100 with goals of 5-10 ug/L. Starting on day 
101, the dose will be tapered 10% per week to discontinue at approximately day 
+180. Therapy with tacrolimus may continue past day +100 if GVHD is present 
or if patient is still a mixed chimera.  
Tacrolimus may be discontinued earlier than Day 100 (with an appropriate taper 
plan) in the context of graft failure, or prohibitive toxicity. It is suggested that 
patients with suspected graft failure remain on tacrolimus until at least the ~Day 
60 chimerism assessment.  
7.4.2 Mycophenolic Acid Mofetil (MMF) 
Mycophenolic acid mofetil will be given at a dose of 15 mg/kg IV (based upon 
actual body weight) every 8 hours with the maximum total daily dose not to 
exceed 3 grams (1 g IV TID). BMT patients have unpredictable oral bioavailability 
of MMF. Consider consulting the pediatric BMT pharmacist when changing MMF 
doses or routes of administration. MMF pharmacokinetics are not required on 
this protocol, but may be obtained when clinically indicated. Consult with the 
pediatric BMT pharmacist for AUC interpretations and dose adjustments when 
necessary. 
 
If no acute GVHD, MMF prophylaxis will be discontinued after the last dose on 
day 35, or 7 days after engraftment, whichever is later. Definition of engraftment 
is 1st day of 3 consecutive days of absolute neutrophil count (ANC) >0.5 x109/L. 


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7.5 Serial MSC Infusions (months 2, 4, 6; and optionally at 8, and 10) – Arms 
E, G and if Applicable, Arm C  
Refer to section 8 regarding MSC production and storage. 
Month 2, 4, 6, 8, and 10 post-transplant are the “targeted” infusion time points; 
however, these time points may be adjusted to best fit individual patient situations 
as deemed most appropriate by the treating investigator. Adjustments to the 
MSC schedule may be made without being considered a protocol deviation. 
Doses at 8 and 10 months are optional and would be based on patient availability 
to return to UMMC. 
MSC will be administered at a dose of 1-2 x 106/kg at the months (± 14 days) 2, 
4 and 6: and offered at 8 and 10 months post-transplant in those patients whose 
donor consented to the additional marrow collection per section 8. The cells will 
be thawed at the “bedside”. 
Premedication (with acetaminophen, diphenhydramine, anti-emetic) and/or 
additional hydration are not required. The product is infused via IV drip directly 
into the central line as tolerated by the patient without a needle or pump. MSCs 
can also be given via a peripheral IV (PIV) when a central line is not available. 
While adverse events are rare based on substantial past experiences with MSCs, 
infusional toxicity will be evaluated by continuous monitoring of the subject’s vital 
signs and O2 saturation by pulse oximetry during the MSC administration. For 
two hours after MSC administration, O2 saturation will be monitored at regular 
intervals and when clinically indicated. Infusional toxicity will be indicated by a 
decrease in oxygen saturation during the infusion period. After the product bag 
empties, the bag and IV tubing will be flushed with sterile normal saline. Saline 
may be either injected directly into the bag or a 50 cc saline IV bag may be 
attached to the product bag. Post infusion assessments will be done per section 
11.2. 
7.6 Concurrent Therapy and Supportive Care 
Supportive care will be provided per institutional guidelines. Guidelines may be 
updated based on current data/drugs without requiring a protocol amendment or 
being considered a protocol deviation.  
Due to the risk of squamous cell carcinoma associated with voriconazole, it is 
preferred that itraconazole be used for antifungal prophylaxis in this patient 
population. Additionally, empiric vancomycin therapy should only be used as 
appropriate according to the circumstances outlined in our institutional infection 
prophylaxis and therapy guidelines.  


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7.6.1 Epidermal Grafting Using the CelluTome® Epidermal Harvesting 
System 
For patients with persistent wounds, local wound therapy will be offered using 
epidermal skin grafting from the same donor that provided the hematopoietic 
graft with donor consent/assent. We will use FDA approved vacuum device 
(CelluTome®, Regulation number 878.4820) that enables scar-free harvesting 
of epidermis with negligible bleeding and minimal pain (commonly described as 
a sensation of warmth and mild pressure) and its transfer on a non-adhering 
silicone mesh dressing (i.e. Adaptic Touch™) to the recipient’s wound. 
EB Wounds  
We will identify regions indicated for grafting.  Such wounds would be open 
erosions and apparently free of infection (as documented by physical 
exam—local redness, swelling, and pain—as microbiome is inevitable and 
physiologic on any muco-cutaneous surface).   
Healthy Skin Grafts   
We will identify optimal harvesting sites on the body of the donor. Although 
the epidermal collection is essentially painless (donor will feel warmth and 
mild pressure at the site) and leaves no scar, we will not choose intensely 
exposed locations, such as head, neck, palms and soles.  As skin is 
developmentally, structurally and functionally different at different parts of 
the body we will attempt collection of epidermis from a general area on the 
donor’s body that corresponds to the site of the wound being treated.  Of 
course, we are prepared to adapt to physical specifics and psychological 
choices of the donor, and we will not collect twice from exactly the same 
site. 
Epidermal Transfer 
The grafting will be done in real time.  This FDA approved (Regulation 
number 878.4820) operating procedure will be followed in all details to 
collect the donor epidermis from a maximum of 3 sites at one sitting and 
transferred to open wound areas.   Grafts will be secured with bandages 
and ideally protected for at least 2 weeks, and preferably up to 3-4 weeks 
if possible. Wounds will be observed for any signs of infection and 
photographed weekly as able. 
Sessions will be offered to patients at least 100 days post-transplant, with 
up to three total sessions occurring at least 12 weeks apart. 
Donor Eligibility 
• 
The HCT donor for the EB patient  


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• 
Age > 2 years (based on prior safety testing of the device) 
• 
Healthy on physical examination in the opinion of the evaluating 
provider 
• 
Known negativity for Hepatitis B and C, HIV, and HTLV1/2 
• 
Voluntary written consent (donor or parent/guardian for minors 
with assent)  
7.7 
Duration of Treatment 
All patients will undergo a hematopoietic stem cell transplant on day 0 with 
standard of care post-transplant follow-up. In addition, if the donor agreed to the 
collection of additional cells for MSC production, the patient will receive MSC 
infusions post-transplant unless any of the following occurs:  
• consent is withdrawn or patient is noncompliant  
• patient experiences unacceptable toxicity 
• administering the MSCs would not be in the best interest of the patient  
7.8 Re-Transplant For Graft Failure  
Patients with graft failure (e.g. lack of engraftment, autologous recovery, loss of 
graft) may be re-transplanted with consideration of time from 1st BMT informing 
intensity of conditioning.  
If the 2nd transplant will occur <1 year since the 1st transplant, two options are 
available: 
1) Re-transplant on this study (Arm C)   
2) Re-transplant on University of Minnesota Blood and Marrow Transplantation 
(BMT) protocol MT2013-06C while maintaining post-transplant follow-up 
visits based on the original transplant date 
 
If the 2nd transplant will occur >/=1 year following the first transplant: The 2nd BMT 
could be completed on this protocol as a new subject on ARM D-G, whichever is 
appropriate based on HLA-match and donor consent for MSC production. 
Re-transplant is done out of medical need. All aspects of the procedure will be 
within the standard of care for re-transplant at this institution and charged to the 
patient and/or 3rd party payer in the usual manner,   
At the discretion of the treating physician and with the donor’s permission, extra 
marrow for MSCs may be collected at the time of the bone marrow harvest. If the 
patient has donor MSCs available (either from the original transplant or the re-
transplant), they may be infused in the same manner and schedule as detailed 


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after the original transplant using day 0 of the second transplant. Targeted 
toxicities will be assessed at time of all MSC infusions.  
All tests/procedures and follow-up milestone visits in association with the 1st 
transplant would continue.  
A formal statistical analysis in association with the re-transplant will not be used 
as these patients are already being analyzed within the context of the initial 
transplant. A descriptive analysis may also be completed for those transplanted 
on Arm C or retransplanted on Arms D-G, in regards to the standard of care and 
research testing that will restart at second transplant day 0; the window for these 
assessments may be expanded so these time points coincide if in the best 
interest of the patient. 
7.9 Duration of Study Participation 
Evaluable patients will be followed for response, toxicity, and survival for 24 
months from the transplant unless consent is withdrawn.  
After 2 years, routine post-transplant follow-up will continue. Survival status and 
other routine clinical information may be abstracted from the medical record or 
other data sources for up to 5 years from study registration. 
8 
Bone Marrow Harvest and MSC Production 
Prior to the bone marrow collection it will be established if the donor has consented to 
the collection of additional BM for the production of MSCs.  
8.1 Bone Marrow Harvest for Transplant 
Unrelated donor bone marrow will be collected in the usual manner using 
established parameters determined by the National Marrow Donor Program.  
 
Related donor bone marrow (BM) will be collected according to current institutional 
guidelines. The bone marrow harvest will be performed on the transplant day (Day 
0). The BM will be infused the same day of collection. The nucleated cell target 
range will be between 2 and 10 x 108/kg of recipient ideal body weight with the 
volume not to exceed 20 mL/kg of donor's weight once the minimal target of 2 x 
108/kg has been reached. Given that there is always uncertainty about the 
feasibility of harvesting certain patients, the harvesting team will have the ability to 
stop the collection if the minimum target of cells has not been reached and it is 
believed that it will be unsafe or technically unfeasible to reach the mentioned 
target if harvest was to continue after a reasonable effort has been made. In this 


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case, the study PI will be notified immediately and efforts should be made to assure 
at least a cell count of 2 x 108/kg of recipient ideal body weight. For unrelated 
donors, a target nucleated cell dose of 5 x 108/kg will be requested. 
8.2 Risks of Donating Bone Marrow 
The most serious risk associated with donating bone marrow involves the use and 
effects of anesthesia during surgery. Other risks associated with bone marrow 
donation include soreness at the site(s) of collection, pain with walking and 
generalized weakness after the procedure. 
8.3 Additional Marrow Harvest for MSC Production with Donor’s Consent 
At the time of BM harvest, an additional 40-50 mL BM will be taken from the donor 
for MSC culture in donors who have agreed to the extra cell collection (embedded 
in the donor consent as a yes (agree)/no (refuse) checkbox). If the donor has 
refused to the additional bone marrow collection, the transplant will proceed as 
plan, but the MSC infusions will not be done. 
8.4 
MSC Manufacture and Storage 
BM adherent cells are cultured under standardized conditions. After expansion 
culture, cells are removed for testing while the remainder is cryopreserved in 
aliquots for future use. MSCs will be banked prior to use and will not be released 
to a patient until the results of all quality control assays (sterility, epitope analysis, 
and differentiation potential) satisfy lot-release criteria (as defined in the IND). 
Details on the manufacturing procedure, including lot-release testing and 
specifications, can be found in the Chemistry, Manufacturing and Controls (CMC) 
section of the Investigational New Drug (IND) application. Only MSCs meeting lot-
release criteria will be available for clinical use. 
9 
Expected Risks of Study Treatment 
9.1 
Preparative Regimen 
Anti-Thymocyte Globulin (rATG)  
Common  
occurs in more than 20% of patients 
Less Common 
occurs in 5 to 20% of 
patients 
Rare 
occurs in fewer than 5% of 
patients 
• fever 
• chills 
• low white blood cell count with increased risk of 
infection  
• low platelet count with increased risk of bleeding  
• pain 
• headache 
• abdominal pain 
• diarrhea 
• feeling poorly (malaise)  
• dizziness  
 
• severe allergic reaction 
(anaphylaxis) 
 


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Anti-Thymocyte Globulin (rATG)  
Common  
occurs in more than 20% of patients 
Less Common 
occurs in 5 to 20% of 
patients 
Rare 
occurs in fewer than 5% of 
patients 
• high blood pressure (hypertension) 
• nausea 
• swelling of hands and/or feet (peripheral edema) 
• shortness of breath (dyspnea) 
• loss or lack of strength (asthenia) 
• high levels of potassium in the blood 
(hyperkalemia) 
• rapid heartbeat (tachycardia) 
 
Fludarabine 
Common  
occurs in more than 20% of patients 
Less Common 
occurs in 5 to 20% of patients 
Rare 
occurs in fewer than 5% of patients 
• low white blood cell count with 
increased risk of infection  
• low platelet count with 
increased risk of bleeding  
• low red blood cell count 
(anemia) with tiredness and 
weakness 
• tiredness (fatigue)  
• nausea  
• vomiting  
• fever and chills 
• infection 
 
• pneumonia  
• diarrhea  
• loss of appetite 
• weakness 
• pain 
 
• numbness and tingling in hands and/or feet 
related to irritation of nerves  
• changes in vision  
• agitation  
• confusion 
• clumsiness 
• seizures 
• coma  
• cough  
• trouble breathing  
• intestinal bleeding 
• weakness  
• death due to effects on the brain, infection, 
bleeding, severe anemia, skin blistering, or 
other causes 
 
Cyclophosphamide 
Common  
occurs in more than 20% of patients 
Less Common 
occurs in 5 to 20% of patients 
Rare 
occurs in fewer than 5% of patients 
• nausea/vomiting 
• mucositis 
• sterility 
• severe suppression of 
• blood counts 
• diarrhea 
• fluid weight gain/edema 
• alopecia 
• hemorrhagic cystitis 
 
• cardiomyopathy 
• skin rash 
• SIADH (Syndrome of Inappropriate 
Anti-diuretic Hormone) 
 
 
Busulfan (ARMS F, G only) 
Common 
Less Common 
Rare 
• Nausea/vomiting 
• Mucositis 
• Rash 
• Severe suppression of blood 
counts 
• Diarrhea 
• Veno-occlusive disease 
• Alveolar hemorrhage 
• Pulmonary fibrosis 
• Interstitial pneumonitis 
• Seizures (low frequency with anti-seizure 
prophylaxis) 
• Hepatic fibrosis/liver failure 


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Busulfan (ARMS F, G only) 
Common 
Less Common 
Rare 
• Fluid weight gain/edema 
• Alopecia 
• Hyperpigmentation 
 
Total Body Irradiation 
Common  
occurs in more than 20% of patients 
Less Common 
occurs in 5 to 20% of patients 
Rare 
occurs in fewer than 5% of patients 
• nausea and vomiting 
• diarrhea 
• cataracts 
• sterility 
• endocrinopathies 
• growth failure 
• intestinal cramps 
• mucositis 
• parotitis 
• interstitial pneumonitis 
• generalized mild erythema 
• veno-occlusive disease 
• dysphagia 
• vertebral deformities 
• nephropathy 
• risk of 2nd malignancy years later 
(when given along with chemotherapy) 
9.2 
Transplant Procedures 
 
With The Cell Infusion 
• nausea and vomiting  
• possible allergic reaction (including itching, hives, flushing [red face], shortness 
of breath, wheezing, chest tightness, skin rash, fever, chills, stiff muscles, or 
trouble breathing) 
 
General Transplant Related Risks  
• slow recovery of blood counts   
• graft failure 
• Graft-Versus-Host Disease (GVHD)  
• other complications including: 
o damage to the vital organs  
o serious infections  
o relapse of disease or a new blood cancer  
o risk to the unborn 
9.3 
MSC Infusions (Arm E, Arm G and, if applicable, Arm C) 
• 
allergic reaction to MSC (rare) 
• 
shortness of breath (rare) 
• 
ectopic tissue formation (theoretical) 
• 
malignancy (one reported case) 
• 
opportunistic infection (rare) 


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9.4 
GVHD Prophylaxis 
Mycophenolate mofetil (MMF) 
Common 
Less Common 
Rare, but may be serious 
• miscarriage 
• birth defects 
• diarrhea 
• damage to unborn baby 
• limited effectiveness of birth 
control 
• stomach pain 
• upset stomach 
• vomiting 
• headache 
• tremors 
• low white blood cell count with 
increased risk of infection 
• increased blood cholesterols 
• swelling of the hands, feet, 
ankles or lower legs 
• anemia 
• rash 
• difficulty falling asleep or 
staying asleep 
• dizziness 
• uncontrollable hand shakes 
• difficulty breathing 
• unusual bruising 
• fast heartbeat 
• excessive tiredness 
• weakness 
• blood in stool 
• bloody vomit 
• change in vision 
• secondary cancers, such as 
lymphoproliferative disease or 
lymphoma 
• Progressive Multifocal 
Leukoencephalopathy 
 
Tacrolimus   
Common 
Less Common 
Rare, but may be serious 
 kidney problems 
 loss of magnesium, calcium, 
potassium 
 high blood pressure 
 tremors 
 increases in cholesterol and 
triglyceride 
 
 nausea 
 vomiting 
 liver problems 
 changes in how clearly one 
can think 
 insomnia 
 unwanted hair growth 
 confusion 
 
seizures 
 
changes in vision 
 
dizziness 
 
red blood cell destruction 
It is very important that grapefruit or drinks with grapefruit juice are not consumed while 
taking Tacrolimus. Grapefruit has an ingredient called bergamottin, which can affect some of 
the treatment drugs used in this study. Common soft drinks that have bergamottin are Fresca, 
Squirt, and Sunny Delight.  
9.5 
Epidermal Grafting Using the CelluTome® 
9.5.1 Donor  
We do not expect any significant side-effects as this procedure has been 
accepted as safe in the field and used for treatment of number of skin conditions, 
such as vitiligo and diabetic leg ulcers, with an excellent safety profile.  Of note,  
the harvested tissue (epidermis) does not cross the basement membrane, so the 
procedure is essentially painless (donor will feel warmth and mild pressure at the 
site) and there is not typically residual visible scarring unless there is a 
malfunction of the device or other unanticipated event.We will still encourage 
examination the sites of epidermal harvest via weekly photographs until healed 
and at 12 weeks (as able) for any abnormalities detectable by physical 
examination. 


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9.5.2 Recipient 
Even though the graft and recipient are uniquely and fully HLA matched, there 
can be local (and much less likely systemic) effects of grafting.  Most prominent 
among these is non-engraftment (sloughing off the graft) because of subclinical 
infection, ongoing fibrosis and—related to that—inadequate vascular supply at 
the recipient site.  This can be accompanied by local inflammation with systemic 
febrile reaction, though none would be expected to be serious adverse events.  
 
 


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10 Clinical Evaluations 
Days and testing/assessment frequency may be altered as clinically appropriate (e.g. 
in the event of graft failure some procedures may be waived).  
10.1 Clinical Evaluations – All Patients Standard of Care 
 
Screen 
Pre-
BMT 
Day 0 to 30 
Day 31 to 100 
 
Follow-up 
 
Days  
-3, -2 
Daily11  
Weekly 
Day +28  
(± 3 days) 
Weekly 
Day +60 
(± 14 days) 
Day +100  
(± 14 days) 
 
Day +180  
(± 14 days) 
1 and 2 
years 
(± 30 days)& 
 
Patient’s General Assessments 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
Medical history 
X 
 
X 
 
 
X 
 
 
X 
X 
Physical exam 
X 
 
X 
 
 
X 
 
 
X 
X 
iscorEBa 
X 
 
 
 
 
 
 
X 
X 
X 
Play Score/Performance 
status1 
X 
 
 
 
X 
 
 
X 
X 
X 
Height/Weight12 
X 
 
 
 
 
 
 
X 
X 
X 
GVHD evaluation2 
 
 
 
X 
 
X 
 
 
X 
X 
Adverse event notation 
 
 
X 
 
 
X 
 
 
X 
X 
 
Patient’s Laboratory Assessments3 
CBC, diff4 
X 
 
X 
 
 
X 
 
 
X 
X 
Platelet 
X 
 
X 
 
 
X 
 
 
X 
X 
PT/PTT, CH50, C3 & C4 
X 
 
 
 
 
 
 
X 
 
 
Serum chemistries5 
X 
 
 
X 
 
X 
 
 
X 
X 
Iron studies6 
X 
 
 
 
 
 
 
X 
X 
X 
Zinc, Selenium, Vitamin 
D3, Carnitine 
X 
 
 
 
 
 
 
 
X 
1 year 
only 
Viral serology7 
X 
 
 
 
 
 
 
X 
X 
X 
Pregnancy test for females 
of childbearing potential 
X 
 
 
 
 
 
 
 
 
 
Blood DNA to Molecular 
Diagnostics 
Lab 
for 
Chimerism Assessment 
X 
 
 
 
X 
 
X 
X 
X 
X 
Skin DNA to Molecular 
Diagnostics Lab for 
Chimerism Assessment 
 
 
 
 
 
 
 
X 
X 
X 
Immune Panel and Ig 
levels8 
X 
 
 
 
X 
 
X 
X 
X 
X 
PRA: initial (for all donors 
except fully matched) 
X 
 
 
 
 
 
 
 
 
 
PRA: Confirmatory w/in 21 
days of treatment (for all 
donors except fully 
matched) 
X 
 
 
 
 
 
 
 
 
 
Busulfan TDM9 
 
X 
 
 
 
 
 
 
 
 
 
Patient’s Procedures 
EKG 
X 
 
 
 
 
 
 
 
 
 
ECHO  
X 
 
 
 
 
 
 
 
 
 
Chest Xray or CT 
X 
 
 
 
 
 
 
 
 
 
Oxygen saturation 
X 
 
 
 
 
 
 
 
 
 
Renal Function (eGFR) 
X 
 
 
 
 
 
 
 
 
 
Ophthalmology consult* 
 
 
 
 
 
 
 
X 
X 
X 
GI endoscopy and biopsy* 
X 
 
 
 
X 
 
X 
X 
X 
X 
Skin biopsy specimens10 
X 
 
 
 
 
 
 
X 
X 
X 


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Screen 
Pre-
BMT 
Day 0 to 30 
Day 31 to 100 
 
Follow-up 
 
Days  
-3, -2 
Daily11  
Weekly 
Day +28  
(± 3 days) 
Weekly 
Day +60 
(± 14 days) 
Day +100  
(± 14 days) 
 
Day +180  
(± 14 days) 
1 and 2 
years 
(± 30 days)& 
Skin fragility testing 
X 
 
 
 
 
 
 
X 
X 
X 
Photographs of skin 
X 
 
 
 
 
 
 
X 
X 
X 
Wound Healing Using the 
CelluTome® Device12, 13 
 
 
 
 
 
 
 
X 
X13  
X  
day 24014 
Photographs of treated 
wounds15 
 
 
 
 
 
 
 
X16 
*Only if clinically indicated  
^Arm B and E only (if donor agreed to the collection of additional bone marrow for MSC production – refer to 
sections 7.5 and 10.3) 
& After 2 years, patient will continue with standard of care post-transplant follow-up from which information 
may be abstracted from the medical record or other sources for the purpose of this study 
a iscorEB surveys are a validated, standard of care tool used to assess disease status in all patients with 
Epidermolysis Bullosa. Patients &/or patient representatives may be asked to complete the patient portion of 
the survey corresponding with additional standard of care assessments as determined by the treating provider. 
All completed iscorEB surveys are intended to be used to further understand and describe changes in disease 
severity of epidermolysis bullosa throughout the trajectory of treatment. 
1 Assessment tool in Appendix III. The Lansky scale will be used for patients less than or equal to 16 years of 
age and the Karnofsky scale will be used for patients greater than 16 years of age. 
2 Assessment tool in Appendix IV. 
3 A sample that is not obtained or obtained outside the protocol window will not be considered a deviation from 
protocol if this is related to daily blood volume limitations. 
4 Complete blood count with leukocyte differential will be measured. 
5Basic metabolic panel to be completed daily through engraftment and comprehensive metabolic panel will 
be measured weekly from day 0 – 30. Comprehensive metabolic panel will be measured at all additional 
specified time points. 
6 Ferritin, TIBC, and serum iron will be measured. 
7 Hepatitis B surface antigen (HBsAg), Hepatitis B core antibody (HBcAb), Hepatitis C virus (HCV), CMV & 
EBV will be measured. CMV and EBV may be assessed at additional intervals per institutional guidelines. 
8 IgA, IgE, IgG, IgM & IgG subclasses will be measured. 
9 Busulfan therapeutic drug monitoring (TDM) per Appendix VI (Arms F and G only)  
10 5 samples: DNA for chimerism assessment; IF or Western to assess protein expression (i.e. collagen, 
laminin, integrin, keratin or plakin); EM to assess structural changes 
11 Daily until engraftment 
12As able 
13 The donor must provide consent/assent 
14 Up to 3 sessions at least 12 weeks apart 
15 For patients who elect wound healing with CelluTome 
16 At minimum, photographs will be taken at baseline grafting and ideally also at wk 6 (+/- 1 wk), *wk 12 (+/- 
1 wk), and *1 year (+/1 3 months).[*Ideally at UMN site unless patient is absolutely unable to return, in which 
case local or remote assessment should be completed with results sent to the PI and study staff] . 
Photographs are also requested weekly between day of grafting and 12-week evaluations; photographs not 
received at these target dates or outside of these target dates that are outside of the study team’s control 
(i.e. patient unable to return to study site and photographs not sent via email despite request from study 
team), will not be considered reportable deviations.   
 


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10.2 Research Related Assessments and Procedures (All Arms) 
 
Screen 
Day +28 (± 
3 days) 
Day +60 
(± 14 days) 
Day +100 
(± 14 days) 
Day +180 
(± 14 days) 
1 and 2 years 
(± 30 days) 
Monitor for Early Stopping Rules 
per section 14.4 
Through day 100 
 
 
Research related bloods for Tolar 
Lab: 
≥40kg patient weight: draw One 5 mL 
red top tube (serum) and One 5mL 
purple top tube (plasma/PBL) 
<40kg patient weight: draw One 0.2 
mL/10kg red top tube (serum) and 
One 0.2 mL/10kg purple top tube 
(plasma/PBL) 
X 
X 
X 
X 
X 
X 
10.3 Research Related Assessments and Procedures (Arms where MSCs 
are given) 
 
Day +60 
(± 14 days) 
Day +120 
(± 14 days) 
Day +180 
(± 14 days) 
Day +240 
(± 14 days)* 
Day +300 
(± 14 days)* 
MSC infusion 
X 
X 
X 
X 
X 
Targeted Toxicity at pre and 4 
hours (+/- 30 minutes) post MSC 
infusion 
X 
X 
X 
X 
X 
*Optional based on patient availability 
Day 60, 120, 180, 240 and 300 post-transplant are the “targeted” infusion time 
points; however, these time points may be adjusted to best fit individual patient 
situations as deemed most appropriate by the treating investigator. Adjustments 
to the MSC schedule may be made without being considered a protocol deviation. 
10.4 Donor 
Unrelated donors are screened, consented and collected per their usual local 
donor center procedures. 
 
Screen 
After Consent  
Donor’s General Assessments 
Informed consent 
X 
 
Medical history 
X 
 
Physical exam 
X 
 
Donor’s Laboratory Assessments 
CBC, diff  
X 
 
Platelet 
X 
 
PT/PTT 
X 
 
Viral serology  
X 
X1 
donor blood to molecular 
diagnostics for their 
baseline chimerism 
X 
 
PRA* 
X 
 


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Screen 
After Consent  
Pregnancy test if of child 
bearing potential 
X 
X1 
Donor’s Procedures 
BM harvest  
 
X 
Skin specimen from BM 
site (UMN related donors 
only)2 
 
X 
For Donors who consent to Skin Cell Harvest 
Donor infectious disease 
panel within 30 days of 
Harvest 
 
X 
Skin Cell Harvest Using 
the CelluTome® Device** 
 
X 
1- Repeat if not done in previous 7 days 
2- As able 
 
 
 
* Not required for fully-matched related donors per institutional SOP 
 
 
 
**as clinically indicated 
 
11 
Adverse Event Monitoring, Recording and Reporting 
Toxicity and adverse events will be classified according to NCI's Common 
Terminology Criteria for Adverse Events V 4.0 (CTCAE) and reported on the schedule 
below. 
A 
copy 
of 
the 
CTCAE 
events 
can 
be 
downloaded 
from 
http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm#ctc_40 
11.1 Definitions 
The following definitions are based on the Code of Federal Regulations Title 21 
Part 312.32 (21CFR312.32(a)). 
Adverse Event:  Any untoward medical occurrence associated with the use of a 
drug in humans, whether or not considered drug related.  
Suspected Adverse Reaction:  Any adverse event for which there is a 
reasonable possibility that the drug caused the adverse event.  
Treatment-Emergent Adverse Event:  Any event not present prior to the initiation 
of the treatment or any event already present that worsens in either intensity or 
frequency following exposure to the treatment. A treatment emergent AE refers to 
an event temporally related to the study treatment regardless of the causality 
assessment by the investigator. 


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Life-Threatening Adverse Event Or Life-Threatening Suspected Adverse 
Reaction: An adverse event or suspected adverse reaction is considered “life-
threatening” if, in the view of either the investigator or sponsor, its occurrence 
places the patient or subject at immediate risk of death. Note:  a life-threatening 
event does not necessarily equate to a CTCAE grade 4. 
Serious Adverse Event Or Serious Suspected Adverse Reaction:  An adverse 
event or suspected adverse reaction is considered “serious” if, in the view of either 
the investigator or sponsor, it results in any of the following outcomes:  
• Death 
• A life-threatening adverse event 
• Inpatient hospitalization or prolongation of existing hospitalization 
• A persistent or significant incapacity or substantial disruption of the ability 
to conduct normal life functions 
• A congenital anomaly/birth defect.  
• Important medical events that may not result in death, be life-threatening, 
or require hospitalization may be considered serious when, based upon 
appropriate medical judgment, they may jeopardize the patient or subject 
and may require medical or surgical intervention to prevent one of the 
outcomes listed in this definition.  
Unexpected adverse event or unexpected suspected adverse reaction: An 
adverse event or suspected adverse reaction is considered "unexpected" if it is not 
listed in the investigator brochure or is not listed at the specificity or severity that 
has been observed; or, if an investigator brochure is not required or available, is 
not consistent with the risk information described in the general investigational plan 
or elsewhere in the current application, as amended. Thus, as defined by the FDA 
adverse events that occur as part of the disease process or underlying medical 
conditions are considered unexpected; however, for the purposes of this study they 
will not be documented or reported. 
11.2 Adverse Event Monitoring, Recording and Reporting 
11.2.1 Event Monitoring  
Monitoring for adverse events will begin with the start of the preparative 
regimen and continue through 1 year post-transplant anniversary visit.  
11.2.2 Event Recording/Documentation 
Due to the intentional clearing of the marrow with chemotherapy as 
preparation for the HCT, it is expected that all patients will experience 
severe depression of their blood counts and other related toxicities as 


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detailed in section 9. Therefore, adverse event documentation in 
OnCore for the purposes of this study will focus on: 
• All serious non-hematologic, non-infectious, non-dermatologic 
adverse events through neutrophil engraftment or Day 42, 
whichever is shorter 
• All serious unexpected suspected adverse reactions after Day 42 
(or engraftment) through 1 year post-transplant upon knowledge  
• Targeted toxicities (appendix V) and unexpected suspected 
adverse reactions will be recorded for patients receiving MSCs at 
the following time points: 
o Prior to each MSC infusion 
o 4 hours (± 30 minutes) after each MSC infusion 
Transplant related outcomes will continue to be collected as routine by 
the BMT Database. 
 
 


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11.2.3 Event Reporting to the U of MN IRB, the FDA, and MCC 
Certain events, in addition to recording on the study’s eCRF’s, will require 
prompt report to the University of Minnesota and/or expedited reporting to 
the FDA according to the table below: 
 
At the time of continuing review/IND annual report, relevant events recorded 
in OnCore will be reported in summary format by those persons responsible 
for such reporting.  
11.3 Monitoring for Stopping Rule Events 
In addition to the event monitoring and reporting described in the previous section, 
the following events will be considered excess toxicity per section 14.4 and be 
reported as a stopping rule event using the Event Form found in OnCore: 
• Aplastic graft failure by day 42 post-transplant 
• Transplant related mortality (TRM) by day 100 post-transplant 
Agency 
Criteria for reporting 
Timeframe 
Form to 
Use 
Submission 
information 
U of MN IRB 
Events requiring prompt reporting 
including, but not limited to unanticipated 
death of a locally enrolled subject(s); new 
or increased risk; any adverse event  that 
require a change to the protocol or consent 
form or any protocol deviation that 
resulting in harm refer to  
http://www.research.umn.edu/irb/forms.htm
l 
within 5 business 
days of discovery 
Report 
Form 
irb@ 
umn.edu 
FDA 
 
Unexpected  fatal or unexpected life 
threatening suspected adverse reaction 
 
As soon as 
possible but no 
later than 7 
Calendar-Days 
UMCC 
SAE form  
Submit as an 
amendment 
to IND 
1) Serious and unexpected suspected 
adverse reaction or 
2)  increased occurrence of serious 
suspected adverse reactions over that 
listed in the protocol or investigator 
brochure or 
3) findings from other sources (other 
studies, animal or in vitro testing) 
As soon as 
possible but no 
later than 15 
Calendar-Days 
Note:  Events due to the disease under treatment or an underlying medical condition 
will not require expedited reporting to the FDA for the purposes of this study 
SAE 
Coordinator 
Study stopping rule events per section 
11.3:  
Aplastic graft failure by day 42 
Transplant related mortality by day 100 
At time of 
reporting  
Event 
Form 
SAE 
Coordinator 
mcc-
saes@umn.
edu 


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Stopping rule events only apply to the initial transplant procedure.  
A copy of the completed Event Form must be sent to the SAE Coordinator (mcc-
saes@umn.edu) at the time it is submitted to the study statistician.  
As these events are outcomes of the treatment, not toxicity as found in CTCAE, 
they should only be recorded/reported as an adverse event, if they meet the criteria 
found in section 11.2.    
12 
Data Collection and Monitoring 
12.1 Data Management 
This study will report clinical data using The Online Enterprise Research 
Management Environment (OnCoreTM), a web based Oracle® database utilizing 
study specific electronic case report forms. Key study personnel will be trained on 
the use of OnCore and will comply with protocol specific instructions embedded 
within the OnCore forms. Patient demographics, patient specific study treatment 
calendars, adverse events, reporting of deaths, and other information required for 
IND annual reporting will be placed in OnCore and other research databases 
maintained by MCC IT.  
12.2 Case Report Forms 
Participant data will be collected using protocol specific electronic case report 
forms (e-CRFs) developed within OnCore based on its library of standardized 
forms. The e-CRF will be approved by the study’s Principal Investigator and the 
Biostatistician prior to release for use. The Study Coordinator or designee will be 
responsible for completing e-CRFs based on the patient specific calendar, and 
updating the patient record until patient death or end of required study 
participation. 
12.3 Data and Safety Monitoring Plan (DSMP) 
The study’s Data and Safety Monitoring Plan will be in compliance with the 
University of Minnesota Masonic Cancer Center's Data & Safety Monitoring Plan 
(DSMP), which can be accessed at http://z.umn.edu/dmsp 
 
For the purposes of data and safety monitoring, this study is classified as high risk 
(investigator initiated protocol under an IND). Therefore the following requirements 
will be fulfilled: 
 The Masonic Cancer Center Data and Safety Monitoring Council (DSMC) will 
review the trial’s progress quarterly. 


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 The PI will comply with at least twice yearly monitoring of the clinical protocol 
by the Masonic Cancer Center monitoring services.  
 The PI will oversee the submission of all reportable adverse events per the 
definition of reportable in section 11.2.3 to the Masonic Cancer Center’s SAE 
Coordinator, the University of Minnesota IRB, and the FDA.  
 
In addition, at the time of the continuing review with the University of Minnesota 
IRB, a copy of the report with any attachments will be submitted to the Cancer 
Protocol Review Committee (CPRC). 
 
IND Annual Reports 
In accordance with regulation 21 CFR § 312.33, the IND sponsor (Dr. Tolar) will 
submit a progress report annually. The report will be submitted within 60 days of 
the anniversary date that the IND went into effect.  
12.4 Record Retention 
The investigator will retain study records including source data, copies of case 
report form, consent forms, HIPAA authorizations, and all study correspondence 
in a secured facility for at least 6 years after the study file is closed with the IRB 
and FDA.  
In addition, the University of Minnesota Clinical Trials Office (CTO) will keep a 
master log of all patients participating in the study with sufficient information to 
allow retrieval of the medical records for that patient.  
Please contact the U of MN CTO before destroying any study related records. 
13 
Disease Outcome Measures 
Five skin samples for tissue analysis will be obtained before the HCT and at day 
100, 180, and 1 year  then annually after HCT (additional skin samples may be 
taken as clinically indicated), from areas approximately 1 mm from a wound border. 
Histological changes will be assessed with light microscopy and the expression of 
C7 with a panel of anti-C7 antibodies;25 ultrastructural changes assessed with 
transmission electron microscopy and presence of C7-containing anchoring fibrils 
with immuno-electron microscopy;45 and donor engraftment in skin assessed by 
competitive polymerase-chain-reaction analysis of polymorphic variable-number 
tandem-repeat regions.  
For functional assessment of skin stability, the time needed to create blisters using 
suction with a vacuum device (NP2 model, Electronic Diversities, Finksburg, USA) 


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will be measured. To test for anti-C7 antibodies in blood samples, a quantitative 
enzyme-linked immunosorbent assay will be used with the assessment of serum 
reactivity with the dermal side of the salt-split human skin46.  
For clinical evaluations, we will used instruments validated for scoring treatment 
outcomes in individuals with epidermolysis bullosa: disease severity tool 
(iscorEB)47.  
Photographs. A specialized camera or photograph with standard measuring 
system will be used at these time-points (day of harvest, 6 weeks, 12 weeks, and 
1 year after grafting) to allow for quantitative assessment of wound size over time 
in correspondence with CelluTome treatments. Weekly photographs will also be 
requested as able until the week 12 visit to assess healing. 
14 
Statistical Considerations 
14.1 Statistical Endpoints 
14.1.1 Primary Endpoint 
Event-free survival by 1 year post-transplant. An event will be defined as 
death or a 50% increase in a patient’s iscoreEB from baseline. 
14.1.2 Secondary Endpoints 
• Percentage change in iscoreEB at 1 year and 2 years as compared to 
baseline calculated using each iscoreEB total score 
• Probability of transplant-related mortality (TRM) at 180 days  
• Change in health quality of life pre-transplant as compared to at 1 and 2 
years post-transplant as measured by the Lansky or Karnofsky score (10-
100)  
• Proportion of lymphoid and myeloid chimerism at various time-points (days 
28, 60, 100, 180, and year 1 and 2) among surviving patients 
14.1.3 Correlative Endpoints 
• Levels of biochemical improvement as measured by protein expression 
(collagen, laminin, integrin, keratin or plakin) and related structural and 
physical changes as derived from biopsy samples at days 100 and180, then 
at 1 and 2 years using a binary result provided by Dr. Tolar’s lab 
• Chimerism results from skin biopsy samples at days 100 and 180, then at 1 
and 2 years (and additional time points if collected as clinically indicated) as 
performed by the Molecular Laboratory  


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• Wound healing will be evaluated at milestone clinic visits (day 180, 1 and 2 
years post-transplant and as clinically indicated) by relative change of 
wound area. It is measured as number of square cm of open wound area at 
the each time point/open wound area before skin grafting. It is also included 
in the total body surface are as noted in the iscorEB. 
• For patients who receive CelluTome, wound healing over time will be 
described by photographs taken prior to graft, at 6 weeks, 12 weeks, and 1 
year after treatments. 
14.1.4 Transplant Related Endpoints 
The following endpoints are abstracted from the BMT database: 
• Survival by 1 year 
• Grade II-IV and grade III-IV acute graft versus host disease (GvHD) at 
Day 100 
• Chronic graft versus host disease at 1 year 
• Time to neutrophil engraftment (defined as the first of three consecutive 
days after HCT that the patient’s absolute neutrophil counts is ≥ 0.5x109 
per liter) 
• The incidence of sustained neutrophil engraftment (neutrophil 
engraftment without autologous recovery) 
• Time to platelet engraftment (defined as the first time after HCT that the 
patient can sustain platelet count ≥ 20x109 per liter for three consecutive 
days) 
• Incidence of bacterial, viral and fungal infections at 1 year  
14.2 Statistical Analysis 
Arms A and B will be closed after enrollment of seven patients. Analyses of these 
patients will remain descriptive. Arms D and E will remain open but we expect to 
enroll approximately 8 additional patients due to the eligibility restriction to HLA 
matched siblings. The total enrollment on these two arms should be approximately 
16 so analyses will also primarily be descriptive. Due to the change/justification in 
conditioning (addition of Busulfan) for future patients receiving an HLA-
mismatched or haploidentical transplant as described in section 2.3, arms F and 
G will now be combined for assessment of the primary endpoint, secondary 
endpoints and the transplant related endpoints since we do not expect the addition 
of MSCs to alter these endpoints based on our large past experience of more than 
1,000 patients42,44,55-57. Arms F and G may be separated, however, when 
assessing levels of biochemical improvement since MSCs may assume a role in 
collagen and laminin expression. The planned 69 patients in Arms F and G will be 
analyzed as follows: Kaplan-Meier curves will be used to estimate event-free 


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survival and survival. Cumulative incidence will be used to estimate the probability 
of TRM, treating time to events as a competing risk. Ninety-five percent confidence 
intervals will be estimated from respective standard errors. The proportion of donor 
chimerism or donor cells (including MSCs) at various time-points will be estimated 
among patients who have survived to the evaluated time-point along with 
descriptive plots. Medians, ranges and inter-quartile ranges will be given for actual 
chimerism values and percentage change and absolute values of iscoreEB 
between baseline and 1 year along with descriptive box-plots. iscoreEB 
assessments will be evaluated among patients surviving to 1 year post transplant. 
The Lansky or Karnofsky score will be measured by the proportion of patients with 
a score < 90 at each time-point. Specific focus will be on survivors at 1 and 2 years 
post-transplant. Simple proportions from binary measures will be used to describe 
the patterns of biochemical improvement over time from 5 different protein 
expressions. Wound healing will be assessed at 6 months, 1 year and 2 years 
descriptively by use of box-plots and estimating the average or median percent 
change in open wound area in relation to pre-skin grafting. In addition to primary 
and secondary endpoints, the transplant-related endpoints of acute and chronic 
GvHD, primary and sustained neutrophil and platelet engraftment and infections 
will be estimated by cumulative incidence treating non-event death as a competing 
risk. Analyses and descriptive plots will be performed with SAS 9.3 (SAS Institute, 
Cary, NC) and/or R 3.0.2.  
14.3 Design and Sample-Size Justification 
This study is designed as a two-stage phase II trial to estimate event-free survival 
(EFS). Since the goal is to estimate the EFS at a long-term time-point (1 year post-
transplant), our design is a generalization of the Simon design. Rather than 
suspension of the trial for evaluation after stage 1, this design uses an optimal 
interim analysis for futility without suspension of accrual as proposed by Huang, 
Talukder and Thomas48. The goal of this design minimizes the expected sample 
size under the assumed parameters.  
Our trial assumes that 40% 1 year EFS is unacceptably low and 60% 1 year EFS 
is a rate worthy of further study. Both the null and alternative hypotheses assume 
that the EFS follow an exponential distribution. Given an overall one-sided type I 
error of 5%, 69 total patients with an interim analysis for futility after accrual of 36 
patients will provide 90% statistical power. These 69 patients do not include the 
currently enrolled 7 patients on Arms A and B nor the expected 16 patients on 
arms D and E as enrollment is starting over for purposes of analyses. Therefore 
total enrollment will be approximately 92 patients. 


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When 36 patients have been enrolled at the new TBI dose of 200 cGy bid with the 
inclusion of Busulfan, accrual will stop if the 1-year Kaplan-Meier estimate of EFS 
is less than 41.6%. The probability of stopping under the null hypothesis is 56%. 
The probability of stopping if the alternative hypothesis is true is 5%. If the trial 
reaches the end of stage II, significant clinical activity would require at least a 1 
year Kaplan-Meier estimate of EFS of 52%. The R package ‘OptInterim’ with 
functions ‘OptimDes’ and ‘SimDes’ were used to create design parameters49. 
We do not expect the February 2019 amendment to alter the outcome of EFS as 
any differences will be in degree not in kind. Stated differently, differences will not 
be qualitative. Therefore, enrollment will continue as planned using only patients 
in arms F and G in the interim decision to continue enrollment as well as in the 
final estimate of EFS at the conclusion of the trial. 
As many as 25 patients may be enrolled per year allowing study enrollment to be 
completed in approximately 3 years. 
14.4 Toxicity Monitoring and Stopping Rules 
Monitoring guidelines are developed to monitor excess toxicity using a continuous 
monitoring strategy based on an adaptation of Pocock stopping boundaries50. In 
the event that a stopping rule is triggered, enrollment will be halted. The study will 
be reviewed by the full study committee and if appropriate, by the IRB, prior to re-
initiating enrollment. Although we do not expect different toxicity rates for the 
diagnoses of JEB and RDEB, monitoring guidelines will be applied separately 
based on EB diagnosis (JEB versus RDEB). We expect that 1/3 (n=23) of the 
patients will be transplanted for JEB and 2/3 (n=46) for RDEB. Continuous 
monitoring will start over with the use of the new conditioning regimen (adjustment 
from 300 cGy TBI to 200 cGy bid TBI plus the addition of Busulfan. Neither 
monitoring boundary was triggered in the 1st fifteen patients. Arms D and E will be 
monitored separately but with both diagnoses combined due the small expected 
enrollment (n=16). 
14.4.1 Aplastic Graft Failure 
14.4.1.1 JEB 
Stopping rules were developed for excessive graft failure by day 42 post-
transplant. The goal is to construct a boundary based on graft failure such 
that the probability of early stopping is at most 10% if the true graft failure 
rate is equal to 10% and our sample size is 23. Given these parameters, 
the upper stopping boundary for graft failure is 2 failures out of 3 patients, 
3 out of 8, 4 out of 14, 5 out of 21 or 6 anytime. Graft failure will be defined 


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as failure to reach an ANC ≥ 500 neutrophils/uL for three consecutive days 
prior to day 42. If the actual rate of graft failure is 20% and 30% respectively, 
the probability of early stopping is 49% and 84% respectively. 
14.4.1.2 RDEB 
Stopping rules were developed for excessive graft failure by day 42 post-
transplant. The goal is to construct a boundary based on graft failure such 
that the probability of early stopping is at most 10% if the true graft failure 
rate is equal to 10% and our sample size is 46. Given these parameters, 
the upper stopping boundary for graft failure is 2 failures out of 3 patients, 
3 out of 7, 4 out of 12, 5 out of 18, 6 out of 24, 7 out of 31, 8 out of 38 or 9 
anytime. Graft failure will be defined as failure to reach an ANC ≥ 500 
neutrophils/uL for three consecutive prior to day 42. If the actual rate of graft 
failure is 20% and 30% respectively, the probability of early stopping is 66% 
and 97% respectively. 
14.4.1.2  Arms D & E 
Stopping rules were developed for excessive graft failure by day 42 post-
transplant. The goal is to construct a boundary based on graft failure such 
that the probability of early stopping is at most 10% if the true graft failure 
rate is equal to 10% and our sample size is 16. Given these parameters, 
the upper stopping boundary for graft failure is 2 failures out of 4 patients, 
3 out of 9, 4 out of 14, or 5 anytime. Graft failure will be defined as failure to 
reach an ANC ≥ 500 neutrophils/uL for three consecutive days prior to day 
42. If the actual rate of graft failure is 20% and 30% respectively, the 
probability of early stopping is 40% and 73% respectively. 
14.4.2 Transplant Related Mortality by 100 Days 
14.4.2.1 JEB 
Stopping rules were developed for excessive TRM by day 100 post-
transplant. The goal is to construct a boundary based on mortality such that 
the probability of early stopping is at most 10% if the true TRM is equal to 
10% and our sample size is 23. Given these parameters, the upper stopping 
boundary for TRM is 2 events out of 3 patients, 3 out of 8, 4 out of 14, 5 out 
of 21 or 6 anytime. The probability of early stopping of TRM is same as that 
for graft failure. 
14.4.2.2 RDEB 
Stopping rules were developed for excessive TRM by day 100 post-
transplant. The goal is to construct a boundary based on TRM such that the 


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probability of early stopping is at most 10% if the true TRM rate is equal to 
10% and our sample size is 46. Given these parameters, the upper stopping 
boundary for TRM is 2 events out of 3 patients, 3 out of 7, 4 out of 12, 5 out 
of 18, 6 out of 24, 7 out of 31, 8 out of 38 or 9 anytime. The probability of 
early stopping of TRM is same as that for graft failure. 
14.4.2.3 Arms D & E 
Stopping rules were developed for excessive TRM by day 100 post-
transplant. The goal is to construct a boundary based on mortality such that 
the probability of early stopping is at most 10% if the true TRM is equal to 
10% and our sample size is 16. Given these parameters, the upper stopping 
boundary for TRM is 2 events out of 4 patients, 3 out of 9, 4 out of 14, or 5 
anytime. The probability of early stopping of TRM is same as that for graft 
failure. 
Due to additional stopping rules and re-initiation of the study, it is 
understood that the overall power may be slightly reduced for this study. 
15 Conduct of the Study 
15.1 Good Clinical Practice 
The study will be conducted in accordance the appropriate regulatory 
requirement(s). Essential clinical documents will be maintained to demonstrate the 
validity of the study and the integrity of the data collected. Master files should be 
established at the beginning of the study, maintained for the duration of the study 
and retained according to the appropriate regulations.  
15.2 Ethical Considerations 
The study will be conducted in accordance with ethical principles founded in the 
Declaration of Helsinki. The IRB will review all appropriate study documentation in 
order to safeguard the rights, safety and well-being of the patients. The study will 
only be conducted at sites where IRB approval has been obtained. The protocol, 
consent, written information given to the patients, safety updates, progress reports, 
and any revisions to these documents will be provided to the IRB by the 
Investigator.  
16 References 
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17. Petrova A, Ilic D, McGrath JA. Stem cell therapies for recessive dystrophic epidermolysis 
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21. Heinonen S, Mannikko M, Klement JF, Whitaker-Menezes D, Murphy GF, Uitto J. Targeted 
inactivation of the type VII collagen gene (Col7a1) in mice results in severe blistering 
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22. Kiel MJ, Yilmaz OH, Morrison SJ. CD150- cells are transiently reconstituting multipotent 
progenitors with little or no stem cell activity. Blood 2008; 111(8): 4413-4; author reply 4-5. 
23. Tolar J, Ishida-Yamamoto A, Riddle M, et al. Amelioration of epidermolysis bullosa by 
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24. Chino T, Tamai K, Yamazaki T, et al. Bone marrow cell transfer into fetal circulation can 
ameliorate genetic skin diseases by providing fibroblasts to the skin and inducing immune 
tolerance. Am J Pathol 2008; 173(3): 803-14. 
25. Wagner JE, Ishida-Yamamoto A, McGrath JA, et al. Bone marrow transplantation for 
recessive dystrophic epidermolysis bullosa. N Engl J Med 2010; 363(7): 629-39. 
26. Luznik L, Bolanos-Meade J, Zahurak M, et al. High-dose cyclophosphamide as single-
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27. Luznik L, Fuchs EJ. High-dose, post-transplantation cyclophosphamide to promote graft-
host tolerance after allogeneic hematopoietic stem cell transplantation. Immunol Res 2010; 
47(1-3): 65-77. 
28. Luznik L, O'Donnell PV, Symons HJ, et al. HLA-haploidentical bone marrow transplantation 
for hematologic malignancies using nonmyeloablative conditioning and high-dose, 
posttransplantation cyclophosphamide. Biol Blood Marrow Transplant 2008; 14(6): 641-50. 
29. Luznik L, O'Donnell PV, Fuchs EJ. Post-transplantation cyclophosphamide for tolerance 
induction in HLA-haploidentical bone marrow transplantation. Semin Oncol 2012; 39(6): 
683-93. 
30. McCurdy SR, Kanakry JA, Showel MM, et al. Risk-stratified outcomes of nonmyeloablative 
HLA-haploidentical BMT with high-dose posttransplantation cyclophosphamide. Blood 
2015; 125(19): 3024-31. 
31. Munchel A, Kesserwan C, Symons HJ, et al. Nonmyeloablative, HLA-haploidentical bone 
marrow transplantation with high dose, post-transplantation cyclophosphamide. Pediatr 
Rep 2011; 3 Suppl 2: e15. 
32. Solomon SR, Sizemore CA, Sanacore M, et al. Total Body Irradiation-Based Myeloablative 
Haploidentical Stem Cell Transplantation Is a Safe and Effective Alternative to Unrelated 
Donor Transplantation in Patients Without Matched Sibling Donors. Biol Blood Marrow 
Transplant 2015. 
33. Castagna L, Bramanti S, Furst S, et al. Nonmyeloablative conditioning, unmanipulated 
haploidentical SCT and post-infusion CY for advanced lymphomas. Bone Marrow 
Transplant 2014; 49(12): 1475-80. 
34. Jaiswal SR, Chatterjee S, Mukherjee S, Ray K, Chakrabarti S. Pre-transplant sirolimus 
might improve the outcome of haploidentical peripheral blood stem cell transplantation with 
post-transplant cyclophosphamide for patients with severe aplastic anemia. Bone Marrow 
Transplant 2015; 50(6): 873-5. 
35. Kanda J, Ichinohe T, Fuji S, et al. Impact of HLA mismatch direction on the outcome of 
unrelated bone marrow transplantation: a retrospective analysis from the Japan Society for 
Hematopoietic Cell Transplantation. Biol Blood Marrow Transplant 2015; 21(2): 305-11. 
36. Kanakry CG, Ganguly S, Zahurak M, et al. Aldehyde dehydrogenase expression drives 
human regulatory T cell resistance to posttransplantation cyclophosphamide. Sci Transl 
Med 2013; 5(211): 211ra157. 


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37. Kanakry CG, Tsai HL, Bolanos-Meade J, et al. Single-agent GVHD prophylaxis with 
posttransplantation cyclophosphamide after myeloablative, HLA-matched BMT for AML, 
ALL, and MDS. Blood 2014; 124(25): 3817-27. 
38. El-Darouti M, Fawzy MM, Amin IM, Abdel Hay RM, Hegazy RA, Gabr H. Allogeneic bone 
marrow derived mesenchymal stem cells for the treatment of dystrophic epidermolysis 
bullosa. Hum Gene Ther 2013; 24: A54. 
39. Fibbe WE, Dazzi F, LeBlanc K. MSCs: science and trials. Nat Med 2013; 19(7): 812-3. 
40. Chen L, Tredget EE, Wu PY, Wu Y. Paracrine factors of mesenchymal stem cells recruit 
macrophages and endothelial lineage cells and enhance wound healing. PLoS One 2008; 
3(4): e1886. 
41. Prockop DJ. Repair of tissues by adult stem/progenitor cells (MSCs): controversies, myths, 
and changing paradigms. Mol Ther 2009; 17(6): 939-46. 
42. Tolar J, Le Blanc K, Keating A, Blazar BR. Concise review: hitting the right spot with 
mesenchymal stromal cells. Stem Cells 2010; 28(8): 1446-55. 
43. Walter MN, Wright KT, Fuller HR, MacNeil S, Johnson WE. Mesenchymal stem cell-
conditioned medium accelerates skin wound healing: an in vitro study of fibroblast and 
keratinocyte scratch assays. Exp Cell Res 2010; 316(7): 1271-81. 
44. Petrof G, Lwin SM, et al. Potential of Systemic Allogeneic Mesenchymal Stromal Cell 
Therapy for Children with Recessive Dystrophic Epidermolysis Bullosa. Journal of . 
Investigational Dermatology;  2015. 
45. Tidman MJ, Eady RA. Evaluation of anchoring fibrils and other components of the dermal-
epidermal junction in dystrophic epidermolysis bullosa by a quantitative ultrastructural 
technique. The Journal of investigative dermatology 1985; 84(5): 374-7. 
46. Chen M, Marinkovich MP, Veis A, et al. Interactions of the amino-terminal noncollagenous 
(NC1) domain of type VII collagen with extracellular matrix components. A potential role in 
epidermal-dermal adherence in human skin. The Journal of biological chemistry 1997; 
272(23): 14516-22. 
47. Pope E, iscorEB (iphone app) 2014. 
48. Huang B, Talukder E, Thomas N. Optimal two-stage phase II designs with long-term 
endpoints. Statistics in Biopharmaceutical Research 2010: 2(1): 51-61. 
49. Huang B, Thomas N. Optimal Two and Three Stage Designs for Single-Arm and Two-Arm 
Randomized Controlled Trials with a Long-Term Binary Endpoint, 2012 
50.  Ivanova A, Qaqish BF, Schell MJ. Continuous toxicity monitoring in phase II trials in 
oncology. Biometrics 2005;61(2):540-5. 
51. Howie and McMullen, 2006, Journal of Perinatal Education 15(3): 50-7.  
52 American Academy of Pediatrics: The transfer of drugs and other chemicals into human milk. 
Pediatrics Vol 108 No. 3 September 2001; 776–789. 
53 Gotsch, G.  Maternal medications and breastfeeding New Beginnings March–April 2000; 
55–56. http://www.llli.org/NB/NBMarApr00p55.html 
54. Schneider P, Reinhold P. Anesthesia in breast feeding. Which restrictions are justified? 
Anasthesiol Intensivmed Notfallmed Schmerzther 2000 Jun; 35(6):356–374 
55. LeBlanc K, Ringden O. Immunomodulation by mesenchymal stem cells and clinical 
experience. J Intern Med 2007; 262(5):509-25. 
56. Tolar J, Villeneuve P, Keating A. Mesenchymal stromal cells for graft-versus-host disease. 
Hum Gene Ther 2011; 22(3):257-62. 
57. Prockop DJ, Oh Jy. Medical therapies with adult stem/progenitor cells (MSCs): a backward 
journey from dramatic results in vivo to the cellular and molecular explanations. J Cell 
Biochem 2012; 113(5):1460-9. 
58. Teltschik HM, Heinzelmann F, Gruhn B, et al. Treatment of graft failure with TNI-based 
reconditioning and haploidentical stem cells in paediatric patients. Br J Haematol. 2016. 


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59. Tang BL, Zhu XY, Zheng CC, et al. Successful early unmanipulated haploidentical 
transplantation with reduced-intensity conditioning for primary graft failure after cord blood 
transplantation 
in 
hematologic 
malignancy 
patients. 
Bone 
Marrow 
Transplant. 
2015;50(2):248-252. 
60. Arnold SD, Bhatia M, Horan J, Krishnamurti L. Haematopoietic stem cell transplantation for 
sickle cell disease - current practice and new approaches. Br J Haematol. 2016. 
61. Bolanos-Meade J, Fuchs EJ, Luznik L, et al. HLA-haploidentical bone marrow transplantation 
with posttransplant cyclophosphamide expands the donor pool for patients with sickle cell 
disease. Blood. 2012;120(22):4285-4291. 
62. Gaziev J, Lucarelli G. Allogeneic cellular gene therapy for hemoglobinopathies. Hematol 
Oncol Clin North Am. 2010;24(6):1145-1163. 
63. Gaziev J, Lucarelli G. Hematopoietic stem cell transplantation for thalassemia. Curr Stem 
Cell Res Ther. 2011;6(2):162-169. 
64. Kapelushnik J, Mandel H, Varadi G, Nagler A. Fludarabine-based protocol for haploidentical 
peripheral blood stem cell transplantation in Hurler syndrome. J Pediatr Hematol Oncol. 
2000;22(5):433-436. 
65. Algeri M, Comoli P, Strocchio L, et al. Successful T-cell-depleted haploidentical 
hematopoietic stem cell transplantation in a child with dyskeratosis congenita after a 
fludarabine-based conditioning regimen. J Pediatr Hematol Oncol. 2015;37(4):322-326. 
66. Zecca M, Strocchio L, Pagliara D, et al. HLA-haploidentical T cell-depleted allogeneic 
hematopoietic stem cell transplantation in children with Fanconi anemia. Biol Blood Marrow 
Transplant. 2014;20(4):571-576. 
67. Trujillo JP, Surralles J. Savior siblings and Fanconi anemia: analysis of success rates from 
the family's perspective. Genet Med. 2015;17(11):935-938. 
68. Yabe H, Inoue H, Matsumoto M, et al. Unmanipulated HLA-haploidentical bone marrow 
transplantation for the treatment of fatal, nonmalignant diseases in children and adolescents. 
Int J Hematol. 2004;80(1):78-82. 
69. Zhu H, Luo RM, Luan Z, et al. Unmanipulated haploidentical haematopoietic stem cell 
transplantation for children with severe aplastic anaemia. Br J Haematol. 2016. 
70. Sarmiento M, Ramirez PA. Unmanipulated haploidentical hematopoietic cell transplantation 
with post-transplant cyclophosphamide in a patient with paroxysmal nocturnal 
hemoglobinuria and secondary aplastic anemia. Bone Marrow Transplant. 2016;51(2):316-
318. 
71. Im HJ, Koh KN, Seo JJ. Haploidentical hematopoietic stem cell transplantation in children 
and adolescents with acquired severe aplastic anemia. Korean J Pediatr. 2015;58(6):199-
205. 
72. Esteves I, Bonfim C, Pasquini R, et al. Haploidentical BMT and post-transplant Cy for severe 
aplastic anemia: a multicenter retrospective study. Bone Marrow Transplant. 2015;50(5):685-
689. 


MT2015-20:  Stem Cell Transplant and Serial MSC Infusions for EB 
 
 
 
 
 
 
 
 
 
 
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Appendix I – Eligibility Checklist - Patient 
 
Biochemical Correction of Severe Epidermolysis Bullosa by Allogeneic Cell Transplantation 
and Serial Donor Mesenchymal Cell Infusions (MT2015-20) 
Eligibility Checklist – page 1 of 2 
Patient initials   
     
 
 
Patient ID
 
 
 
 
1st 2 initials of first name + 1st 2 initials of last name 
        Seq # (i.e. 01, 02, 03, etc.) 
 
 
INCLUSION CRITERIA  
A "NO" response to any of the following disqualifies the patient from study entry. 
 
 
 
 
 
Yes      No 
1. 0 through 25 years of age 
 
 
2. 
Diagnosis of severe form of EB with documented collagen, laminin, integrin, keratin or plakin 
deficiency (by immunofluorescence staining with protein specific antibodies or Western blotting 
and by mutation analysis)  
 
 
3. 
Adequate organ function within 4 weeks of study registration defined as: 
test 
requirement 
glomerular 
filtration rate 
within normal range for age 
total bilirubin 
< 5 x UNL 
AST  
< 5 X UNL 
ALT 
< 5 X UNL 
ALP 
< 5 X UNL 
oxygen saturation  adequate pulmonary function in the opinion of the enrolling investigator 
left ventricular 
ejection fraction 
≥ 45% 
normal EKG or approved by Cardiology 
 
 
 
4. Available donor PRA results (Not applicable for fully matched donors): 
 Target MFI level <1,000 or approved by PI  
 
 
N/A 
5. 
Sexually active participants must agree to use adequate birth control for the during the study 
period (from before the start of the preparative chemotherapy through 1 year post-transplant) 
check if not applicable 
 
 
 
6. Voluntary written consent (patient or parent/guardian for minors) prior to any research related 
procedures or treatment 
 
 


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Biochemical Correction of Severe Epidermolysis Bullosa by Allogeneic Cell Transplantation 
and Serial Donor Mesenchymal Cell Infusions (MT2015-20) 
 
Eligibility Checklist – page 2 of 2 
Patient initials   
     
 
 
Patient ID
 
 
 
 
EXCLUSION CRITERIA 
A "YES" response to any of the following disqualifies the patient from study entry. 
 
 
Yes      No 
1. Beta 3 laminin JEB mutants 
 
 
2. 
 Active untreated systemic infection at time of transplantation (including active infection with 
Aspergillus or other mold within 30 days) 
 
 
3. 
 History of HIV infection 
 
 
4. 
 Evidence of squamous cell carcinoma 
 
 
5. 
 
Pregnant or breast feeding. Females of child-bearing potential must have a negative pregnancy 
test prior to study registration as the agents administered in this study are Pregnancy Category 
C and D 
 
 
 
  
 
Having obtained consent and reviewed each of the inclusion/exclusion criteria, I verify that this patient is 
eligible. 
 
 
 
 
 
 
 
 
 
 
 
 
Signature of enrolling investigator 
 
 
Date 
 
 
 


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Appendix II – Eligibility Checklist – Donor 
 
 
Biochemical Correction of Severe Epidermolysis Bullosa by Allogeneic Cell Transplantation 
and Serial Donor Mesenchymal Cell Infusions (MT2015-20) 
Eligibility Checklist – page 1 of 1 
Donor initials   
     
 
 
Patient ID
-D  
 
 
1st 2 initials of first name + 1st 2 initials of last name 
         
 
 
INCLUSION CRITERIA  
A "NO" response to any of the following disqualifies the patient from study entry. 
 
Additional Eligibility to donate skin graft 
A "NO" response to any of the following disqualifies the donor from the CelluTome Procedure. 
 
 
Did donor consent to collection of additional BM cells for MSC production? 
  no – enroll patient in Arm D or F:  transplant only 
  yes – enroll patient in Arm E of G:  transplant plus serial MSC infusions 
 
 
Having obtained consent and reviewed each of the inclusion/exclusion criteria, I verify that this donor is 
eligible. 
 
 
 
 
 
 
 
 
 
 
 
 
 
Signature of person verifying eligibility 
 
 
Date 
 
 
Yes      No 
1. 
 
Be medically, socially, and psychologically fit to donate  in the opinion of the evaluating medical 
personnel 
 
 
2. 
 
Does not have epidermolysis bullosa (or does have but will be used as the donor per section 5) 
 
 
3. 
 
Adequate hematologic, hepatic and renal function to undergo the donor procedure in the opinion of the 
evaluating medical personnel 
 
 
4. 
 
Not pregnant with confirmation of non-pregnancy status within 7 days of donation 
 
 
5. 
 
Voluntary written consent - adult or parent (with assent for minors, if applicable)  
 
 
6. 
 
PRA results (Not applicable for fully matched donors): targeted MFI level within protocol bounds or 
approved by PI (The PI can approve use of a certain donor even if DSA+ and debulk or not based on 
his/her preference or institutional guidelines.) 
 
 
N/A 
 
 
Yes      No 
1. 
 Age > 2 years (based on prior safety testing of the device) 
 
 
2. 
 
Known negativity for Hepatitis B and C, HIV, and HTLV1/2 
 
 
3. 
 
Voluntary written consent (donor or parent/guardian for minors with assent) 
 
 


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Appendix III – Lansky Play Score and Karnofsky Performance Status 
 
For patients < 16 years of age: 
Lansky 
Score 
Play Score 
100 
Fully active, normal 
90 
Minor restrictions in physically strenuous activity 
80 
Active, but tires more quickly 
70 
Both greater restriction of and less time spent in play activity 
60 
Up and around, but minimal active play; keeps busy with quieter activities 
50 
Gets dressed but lies around much of the day, no active play but able to 
participate in all quiet play and activities 
40 
Mostly in bed; participates in quiet activities 
30 
In bed; needs assistance even for quiet play 
20 
Often sleeping; play entirely limited to very passive activities 
10 
No play; does not get out of bed 
0 
Unresponsive 
 
For patients 16 years of age and older: 
Karnofsky Performance Scale 
Percent 
Description 
100 
Normal, no complaints, no evidence of disease. 
90 
Able to carry on normal activity; minor signs or symptoms of disease. 
80 
Normal activity with effort; some signs or symptoms of disease. 
70 
Cares for self, unable to carry on normal activity or to do active work. 
60 
Requires occasional assistance, but is able to care for most of his/her needs. 
50 
Requires considerable assistance and frequent medical care. 
40 
Disabled, requires special care and assistance. 
30 
Severely disabled, hospitalization indicated. Death not imminent. 
20 
Very sick, hospitalization indicated. Death not imminent. 
10 
Moribund, fatal processes progressing rapidly. 
0 
Dead. 
 
 


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Appendix IV – GVHD Scoring 
Acute GVHD 
Organ involvement will be staged using the criteria outlined in the table below. Biopsy of each organ site at 
diagnosis or major change in disease activity will be performed unless clinical circumstances make it 
impossible. 
Consensus Clinical Stage and Grade of Acute GVHD (Przepiorka et al, 1995) 
Stage 
Skin 
Liver 
Lower 
Gastrointestinal 
Tract 
Upper 
Gastrointestinal 
Tract 
1 
Maculopapular rash 
<25% of body 
surface 
Bilirubin 2.0 – 3.0 
mg/dl 
Diarrhea 500 – 1000 
mL/day or 280 – 555 
mL/m2 
No protracted 
nausea and 
vomiting 
2 
Maculopapular rash 
25-50% body 
surface 
Bilirubin 3.1 – 6.0 
mg/dl 
Diarrhea 1000 – 1500 
mL/day or 556 – 833 
mL/m2 
Persistent nausea, 
vomiting or anorexia 
3 
Generalized 
erythroderma 
Bilirubin 6.1 – 15.0 
mg/dl 
Diarrhea >1500 
mL/day or    >833 
mL/m2 
 
4 
Generalized 
erythroderma with 
bullous formation 
and desquamation 
Bilirubin > 15 mg/dl 
Severe abdominal 
pain, with or without 
ileus, or stool with 
frank blood or melena 
 
 
Grading for Treatment Criteria: 
Mild GVHD  
 
Skin stage I-II only (Equivalent to Seattle Grade I). 
Moderate GVHD 
Skin stage I-III and/or liver I-IV and/or Gastrointestinal tract (GI) I-III and/or Upper GI 
(UGI). (Equivalent to Seattle Grade II, III). 
Severe GVHD Any stage IV along with severe clinical illness. 
Late Acute and Chronic GVHD 
Late acute and chronic GVHD will be assessed using the National Institutes of Health (NIH) Consensus 
Criteria. 
 
 
 
 


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Appendix V – Targeted Toxicities (For MSC Infusions Only) 
MT2015-20 
 
 
  
 
 
 
 
 
Refer to section 11.2.2 for time points  
 Patient Initials:  _______ Date of Assessment:  _________     Assessment Time point:  _________      
Toxicity 
Grade 0 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Infusion related 
reaction 
None 
Mild transient reaction; 
infusion interruption not 
indicated; intervention 
not indicated 
Therapy or infusion interruption 
indicated but responds promptly to 
symptomatic treatment (e.g., 
antihistamines, NSAIDS, narcotics, IV 
fluids); prophylactic medications 
indicated for ≤ 24 hrs 
Prolonged (e.g., not rapidly 
responsive to symptomatic 
medication and/or brief interruption 
of infusion); recurrence of symptoms
following initial improvement; 
hospitalization indicated for clinical 
sequelae 
Life-threatening 
consequences; urgent 
intervention indicated 
Sinus Bradycardia 
None 
Asymptomatic, 
intervention not indicated 
Symptomatic, medical intervention 
indicated 
Severe, medically significant, 
medical intervention indicated 
Life-threatening 
consequences; urgent 
intervention indicated 
Sinus Tachycardia 
 
None 
Asymptomatic, 
intervention not indicated 
Symptomatic; non-urgent medical 
intervention indicated 
Urgent medical intervention 
indicated 
-------------- 
Bleeding/ 
Hemorrhage 
 
Mild; intervention not 
indicated 
Moderate symptoms; medical 
intervention or minor cauterization 
indicated 
Transfusion, radiologic, 
endoscopic, or elective operative 
intervention indicated 
Life-threatening 
consequences; urgent 
intervention indicated 
Hypertension 
None 
Pre-hypertension 
(systolic BP 120 - 139 
mm Hg or diastolic BP 
80 - 89 mm Hg) 
Stage 1 hypertension (systolic BP 
140 - 159 mm Hg or diastolic BP 90 - 
99 mm Hg); medical intervention 
indicated; recurrent or persistent  ≥ 
24 hrs); symptomatic increase by >20 
mm Hg (diastolic) or to >140/90 mm 
Hg if previously WNL; monotherapy 
indicated 
Stage 2 hypertension (systolic BP 
≥ 160 mm Hg or diastolic BP ≥ 100 
mm Hg); medical intervention 
indicated; more than one drug or 
more intensive therapy than 
previously used indicated.  
Life-
threateningconsequences 
(e.g.,malignant 
hypertension,transient or 
permanentneurologic 
deficit,hypertensive 
crisis); urgent intervention 
indicated.  
Hypotension 
None 
Asymptomatic, 
intervention 
not indicated 
Non-urgent medical 
intervention indicated 
Medical intervention or 
hospitalization indicated 
Life-threatening and 
urgent intervention 
indicated 
Fever  
None 
38.0 - 39.0◦ C (100.4 -
102.2◦ F) 
> 39.0 - 40.0◦C (102.3 - 104.0◦F) 
> 40.0 ◦ C (>104.0◦ F) for ≤ 24 hrs 
> 40.0 ◦ C (>104.0◦ F) for 
> 24 hrs 
Chills 
None 
Mild sensation of cold; 
shivering; chattering of 
teeth 
Moderate tremor of the entire body; 
narcotics indicated 
Severe or prolonged, not 
responsive to narcotics 
-------------- 
Dyspnea 
None or 
 no change 
Shortness of breath with 
moderate exertion 
Shortness of breath with minimal 
exertion; limiting instrumental ADL 
Shortness of breath at rest; 
limiting self care ADL 
Life-threatening 
consequences; urgent 
intervention indicated 
Hypoxia 
None 
……………… 
Decreased O2 saturation with 
exercise (e.g., pulse oximeter < 88%) 
intermittent supplemental oxygen 
Decreased oxygen saturation at 
rest (e.g., pulse oximeter < 88% or 
PaO2 ≤ 55 mm Hg) 
Life-threatening airway 
compromise; urgent 
intervention indicated  
Rash 
None 
Covering < 10% body 
surface area (BSA) 
Covering 10-30% body surface area 
(BSA) 
>30% body surface area (BSA) 
Generalized exfoliative, 
ulcerative, or bullous 
dermatitis 
 
Person Completing Form:  
 
 
 
 
 
 
 
ADL = activities of daily living 
 
 


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Appendix VI – Low Dose Busulfan Dose Selection (For Arms F and G Only) 
 
Busulfan Dose Selection, AUC Monitoring and Algorithm for Dose 
Modification Using Once Daily IV Dosing 
Goal AUC 7560 [31 (mg · h)/L] 
This protocol will use once daily (every 24 hour) intravenous busulfan dosing.  Two (2) total doses of busulfan 
will be given over 2 days in the preparative regimen.  AUC determination after doses #1 and #2 (AUC1 and 
AUC2). For all patients, each busulfan dose will be administered over 3 hours by central venous line per 
University of Minnesota BMT Program standard guidelines.  
 
THE TOTAL regimen busulfan AUC (AUCcum) targeted will be = 6950-8170 (µM·Min)/L [28.5-33.5 (mg · h)/L] 
with ideal target AUCcum of 7560 (µM·Min)/L [31 (mg · h)/L] in order to best optimize engraftment while 
minimizing toxicity. If at any time upon interim analysis and review of patients transplanted on this protocol with 
the principal investigators and pharmacy team, the subsequent busulfan levels following dose #2 (AUC2) are 
deemed reasonable and consistent, the decision may be made to only do kinetics with the dose#1 for all 
remaining patients treated on this protocol. In that event, kinetics for a dose#2  (i.e. AUC monitoring) may be 
performed whenever a dose change is required but will not be mandatory. These protocol changes (if enacted) 
will not require a treatment deviation or protocol amendment. 
 
Steady State Concentration (Css) equivalent to 6950-8170 (µM·Min)/L = 600-700 ng/ml 
 
DETERMINATION OF FIRST BUSULFAN DOSE (DOSE #1)  
The initial empiric busulfan dosing (Dose #1) for ALL patients will be determined as follows:  
a. For patients weighing <12 kg: initial dose will be based on the formula according to the population PK 
model developed by Long-Boyle.  
b. For patients weighing ≥12kg and <66 kg: initial dose will be determined by the nomogram adapted from 
Bartelink.  
c. For patients weighing ≥66kg: initial busulfan dose = 3 mg/kg IV Once Daily 
 
1. For patients <12 kg: the first busulfan dose will be based on the population PK formula 
developed by Long-Boyle which calculates the first dose as a function of weight and age:  
• 
Use the Dosing Calculator (Long-Boyle Busulfan 1st Dose) on the BMT website to 
determine the precise dose for the patient;  
• 
Age: enter the age in years (to the hundredths place) by dividing the patient’s age in days by 365 
• 
Weight: enter the weight in kg (to the tenths place)  
• 
The Calculator is constructed using the following function:  
o Dose (mg) = AUC(target) x CLi  
o AUC(target) = 15.6 mg·hr·L-1 (equivalent to 3800 μM·min∙L-1)  
o CLi is a function of weight and age  
 
CLi = 2.3 L/h x (Matmag + (1- Matmag) x [1-e(-age x Kmat)]) x (weight/8 kg)0.75 
 
Maturation magnitude (Matmag) = 0.46; Maturation rate constant (kmat) = 1.4 


MT2015-20:  Stem Cell Transplant and Serial MSC Infusions for EB 
 
 
 
 
 
 
 
 
 
 
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• 
See Table A-1 below for EXAMPLE first Busulfan doses for hypothetical children under 12 kg; this 
table can be used to ensure your patient's calculator output does not carry significant error. 
However, the patient's first dose should be determined directly from the calculator.  
 
 
Table A-1: Sample First Busulfan Doses for Various Ages and Weights Using the Long-Boyle Method 
for Determination of First Busulfan Dose for patients < 12 kg & for ideal target AUCcum of 7560 
(µM·Min)/L [31 (mg · h)/L]. Use this table to check against error from the calculator. 
 
 
Sample Age 
 
Sample Weight (kg) 
 
1st Busulfan Dose Per  
Long-Boyle Method (mg) 
1 month 
(0.08 years) 
 
4 
11 
4.4 
11.9 
5 
13 
3 months 
(0.25 years) 
 
5.8 
17.4 
6.4 
18.7 
6.8 
19.6 
6 months 
(0.50 years) 
 
7.2 
24.1 
7.8 
25.5 
8.4 
27 
9 months 
(0.75 years) 
 
8.2 
29.4 
9 
31.6 
9.6 
33.1 
12 months 
(1 years) 
 
9.6 
35.4 
10.2 
37.1 
11 
39.2 
15 months 
(1.25 years) 
 
10.2 
38.8 
11 
41 
11.8 
43.3 
 
2. For patients >/= 12 kg to <66 kg: the first busulfan dose will be according to weight-based dosing 
nomogram developed by Bartelink, which determines the first dose as a function of weight only.  
• 
Determine the first dose directly from the nomogram chart (no calculator is used)  


MT2015-20:  Stem Cell Transplant and Serial MSC Infusions for EB 
 
 
 
 
 
 
 
 
 
 
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CPRC #2015LS076 
 
• 
Round the patient's weight to the nearest kg. For weights in kg ending in "x.5", round UP to 
the nearest kg. For instance, for a weight of 43.5 kg, use 44 kg to determine the first 
busulfan dose from the nomogram.  
• 
See Table A-2 for nomogram 
 
Table A-2: Nomogram for the Determination of First Busulfan Dose for Patients ≥ 12 kg and < 66kg for ideal 
target AUCcum of 7560 (µM·Min)/L [31 (mg · h)/L].  (Adapted from Bartelink).  
Round the patient's weight to the nearest kg in order to determine the first busulfan dose from this nomogram 
 
Weight 
(kg) 
Dose 
(mg) 
Weight 
(kg) 
Dose 
(mg) 
Weight 
(kg) 
Dose 
(mg) 
Weight 
(kg) 
Dose 
(mg) 
Weight 
(kg) 
Dose 
(mg) 
Weight 
(kg) 
Dose 
(mg) 
 
 
20 
64.7 
30 
86.3 
40 
 103.5 
50 
 117.8 
60 
130.5 
 
 
21 
67.1 
31 
88.2 
41 
105 
51 
119.1 
61 
131.6 
12 
41.8 
22 
69.5 
32 
90.2 
42 
106.5 
52 
120.5 
62 
132.6 
13 
45.1 
23 
71.9 
33 
92.1 
43 
108 
53 
 121.8 
63 
133.7 
14 
48.2 
24 
74.3 
34 
94.1 
44 
109.5 
54 
123.2 
64 
134.7 
15 
51.2 
25 
76.5 
35 
96 
45 
111 
55 
 124.5 
65 
135.8 
16 
54.1 
26 
78.5 
36 
97.5 
46 
112.4 
56 
125.7 
 
 
17 
56.8 
27 
80.4 
37 
99 
47 
113.7 
57 
126.9 
 
 
18 
59.5 
28 
82.4 
38 
100.5 
48 
115.1 
58 
128.1 
 
 
19 
62.2 
29 
84.3 
39 
102 
49 
116.4 
59 
129.3 
 
 
 
 
3.  For patients weighing ≥ 66kg: first busulfan dose = 3 mg/kg IV. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


MT2015-20:  Stem Cell Transplant and Serial MSC Infusions for EB 
 
 
 
 
 
 
 
 
 
 
February 5, 2019 
Page 80 of 81 
CPRC #2015LS076 
 
BUSULFAN THERAPEUTIC DRUG MONITORING (TDM)  
 
Area under the curve (AUC) analyses will be calculated in-house per University of Minnesota BMT standard 
guidelines for all patients. The AUC will be calculated for each dose using serum busulfan concentrations (in 
ng/ml) performed by the University of Minnesota Medical Center Drug Analysis Lab and obtained at the 
following time-points:  
• 
Immediately prior to busulfan infusion (*ONLY to be drawn if obtaining levels with doses 2) 
• 
Immediately after the end of the busulfan infusion  
• 
15 minutes after the end of the busulfan infusion  
• 
1 hour after the end of the busulfan infusion  
• 
3 hours after the end of the busulfan infusion  
• 
5 hours after the end of the busulfan infusion  
• 
7 hours after the end of the busulfan infusion  
 
* “trough” or pre-dose serum busulfan level will only be obtained when performing AUC on 2nd or 
higher doses, as serum busulfan level prior to dose #1 is assumed to be zero.  
 
After determination of the patient's clearance and AUC, subsequent doses will be calculated linearly to achieve 
a goal cumulative AUC exposure.  
 
For patients receiving 2 total busulfan doses the goal AUCcum = 6950-8170 (µM·Min)/L [28.5-33.5 (mg · 
h)/L] with ideal target AUCcum of 7560 (µM·Min)/L [31 (mg · h)/L] 
 
 
GUIDELINES FOR CHANGING SUBSEQUENT BUSULFAN DOSES BASED ON A PATIENT'S TDM 
RESULTS  
 
Subsequent busulfan dose changes will be made according to the following guidelines:  
 
1. The goal cumulative AUC range (AUCcum for the entire 2-dose course) will be 6950-8170 
(µM·Min)/L [28.5-33.5 (mg · h)/L] with ideal target AUCcum of 7560 (µM·Min)/L [31 (mg · h)/L] 
2. Cumulative AUC is defined as the sum of each individual dose AUC:  
AUCcum = AUC1 + AUC2  
3. Changes will be made to subsequent doses only if the projected AUCcum falls outside of the goal 
range.   
a. After the first busulfan dose, projected AUCcum = AUC1 x 2 
b. If dose adjustment is necessary, new doses will be calculated based on an ideal target 
AUCcum of exactly 7560 (µM·Min)/L [31 (mg · h)/L] 
4. Total busulfan exposure will be recorded from busulfan AUC analyses following both doses:  
AUCcum = AUC1 + AUC2  
 
 
REFERENCES 
1) Bartelink IH. Body weight-dependent pharmacokinetics of busulfan in paediatric haematopoietic stem cell 
transplantation patients: towards individualized dosing. Clin Pharmacokinet. 2012;51:331-45. 
2) Bartelink IH, van Kesteren C, Boelens JJ, Egberts TC, Bierings MB, Cuvelier GD, Wynn RF, Slatter MA, 
Chiesa R, Danhof M, Knibbe CA. Predictive performance of a busulfan pharmacokinetic model in children 
and young adults. Ther Drug Monit. 2012;34:574-83. 


MT2015-20:  Stem Cell Transplant and Serial MSC Infusions for EB 
 
 
 
 
 
 
 
 
 
 
February 5, 2019 
Page 81 of 81 
CPRC #2015LS076 
 
3) Savic RM, Cowan MJ, Dvorak CC, Pai SY, Pereira L, Bartelink IH, Boelens JJ, Bredius RGM, Wynn RF, 
Cuvelier GDE, Shaw PJ, Slatter MA, Long-Boyle J.  Effect of weight and maturation on busulfan clearance 
in infants and small children undergoing hematopoietic cell transplantation.  Biol Blood Marrow Transplant. 
2013;19:1608-1614. 
 

ICF_001.pdf: