Clinical trial NCT02224872
Conditions: Severe Aplastic Anemia Bone Marrow Failure Syndromes
Keywords: bone marrow transplant thymoglobulin cyclophosphamide fludarabine tacrolimus Mycophenolic Acid Mofetil chemotherapy GVHD haploidentical
Sponsor: Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins
Start date: 2014-08-01; Completion date: 2021-12-01
Enrollment: 18 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
Our primary objective is to determine if it is feasible for SAA patients to be transplanted using non-myeloablative conditioning and post transplantation cyclophosphamide with partially HLA-mismatched donors.

Detailed description:
This research is being done to find out if bone marrow transplantation (BMT) followed by chemotherapy will help people with aplastic anemia who have failed other treatments.

You have a severe, life threatening disease (severe aplastic anemia) in your bone marrow. Your disease has come back or not responded after receiving one or more immunosuppressive treatments. High dose chemotherapy followed by bone marrow transplantation (BMT) has been used to treat blood diseases like yours but complications from Graft vs. Host disease (GVHD) and graft failure have limited the survival for those people.

A small study done at Johns Hopkins has shown that in subjects with other diseases (blood cancers) some immunosuppressive drugs given after the BMT have decreased how often subjects had complications of GVHD and engraftment failure.

People with aplastic anemia who have refractory disease (not responding to standard treatment) may join.

Arms: arm 1: Thymoglobulin on days -9 to -7 Fludarabine on days -6 to -2 Cyclophosphamide on days -6, -5, 3, 4 TBI on day -1 BMT on day 0 Mesna on days 3, 4 Tacrolimus on days 5-365 Mycophenolic acid mofetil on days 5-35
Interventions: intervention 1: Bone marrow transplant intervention 2: Thymoglobulin intervention 3: Fludarabine intervention 4: Cyclophosphamide intervention 5: TBI intervention 6: Mesna intervention 7: Tacrolimus intervention 8: Mycophenolic acid mofetil
Intervention descriptions: intervention 1: Day 0 intervention 2: 0.5 mg/kg IV on Day -9 2 mg/kg IV on Days -8, -7 intervention 3: 30 mg/M2 IV on days -6 to -2 intervention 4: 14.5 mg/kg IV on days -6, -5, 3, 4 intervention 5: 200 cGy on day -1 intervention 6: 40 mg/kg IV on days 3, 4 intervention 7: For patients 18 years or older, tacrolimus will be given per institutional standards; may be increased or later changed to a PO BID schedule. Treatment to continue until Day 365 or longer if GVHD present intervention 8: 15 mg/kg PO/IV TID beginning on day 5 through day 35

Protocol documents:
Prot_SAP_000.pdf: 
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A Phase II Trial of Non-Myeloablative Conditioning and Transplantation of Partially 
HLA-Mismatched/Haploidentical Related or Matched Unrelated Bone Marrow for 
Patients with Refractory Severe Aplastic Anemia and Other Bone Marrow Failure 
Syndromes 
 
 
Protocol Number:   J1424  
 
IRB Application#: IRB00031590 
NCT # NCT02224872 
 
Principal Investigator: 
Amy E. DeZern, MD, MHS 
(Coordinating Center) 
Office: 410-502-7208 
 
Pager:  410-283-7521 
 
Email: adezern1@jhmi.edu 
 
 
Co-Principal Investigator: Robert Brodsky, MD 
 
Office:  410- 955-2546 
 
Email:  brodsro1@jhmi.edu    
 
Co-Investigators: 
Please see Protocol Library for a list of approved Co-
Investigators 
 
Statisticians:  
 
Gary L Rosner, Sc.D.                
 
 
 
 
Marianna Zahurak, M.S 
 
Study Coordinator  
Tracey Graham 
Email: tgraham8@jhmi.edu 
 
Outside Site:  
 
Medical College of Wisconsin 
Children’s Hospital of Wisconsin 
 
 
 
 
Site PIs:   
David Margolis, MD 
 
 
 
 
Monica Thakar, MD 
 
 
 
 
 
 
 
 
 
 
 
 
 
TABLE OF CONTENTS 
 
 


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SCHEMA………………………………………………………. 3 
 
1.0 
OBJECTIVES…………………………………………………. 4 
 
2.0 
BACKGROUND AND RATIONALE……………………….. 4-6 
 
3.0 
DRUG INFORMATION……………………………………… 6-11 
 
4.0 
PATIENT SELECTION……………………………………… 11-13 
 
5.0 
TREATMENT PLAN………………………………………… 13-18 
 
6.0 
PATIENT MONITORING………………………………….. 
18-21 
 
7.0 
TOXICITIES TO BE MONITORED……………………….. 
21-23 
 
8.0 
STUDY PARAMETERS…………………………………….. 23-24 
 
9.0 
DATA MANAGEMENT…………………………………….. 
24-26 
 
10.0 
STATISTICAL CONSIDERATIONS……………………… 
26-30 
 
11.0 
RISKS AND BENEFITS…………………………………….. 30 
 
12.0 
INFORMED CONSENT…………………………………….. 
30-31 
 
13.0 
ON-STUDY DATE…………………………………………… 31 
 
14.0 
OFF-STUDY DATE…………………………………………. 
31 
 
 
REFERENCES………………………………………………. 
32-33 
 
 
APPENDICES………………………………………………. 
34-35 


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TREATMENT SCHEMA 
Days -9                        Thymoglobulin 0.5 mg/kg IV with pre-meds 
 
Days -8,-7 
Thymoglobulin 2 mg/kg IV qd with pre-meds 
 
Days -6, -5 
Fludarabine 30 mg/M2 iv qd 
 
Cyclophosphamide (CTX) 14.5 mg/kg IV qd*  
 
Start steroid taper from ATG  
 
 
Days –4 -2 
Fludarabine 30 mg/M2 iv qd, adjusted for CrCl 
 
 
Day –1 
TBI 200 cGy 
 
Day 0 
Infuse bone marrow per institutional standards  
 
Start appropriate antibiotic prophylaxis 
 
 
Days 3, 4 
CTX 50 mg/kg iv q d 
 
Mesna 40 mg/kg iv q d** 
(First dose of CTX must be administered 48-72 hr after START of infusion of marrow) 
 
 
Day 5    
Begin tacrolimus (section 5.6)** and  
                
MMF 15 mg/kg po tid with maximum daily dose 3 gm/d 
 
(24h after the last dose of CY) 
 
  
 
 
Day 30 
Assess Chimerism in peripheral blood  
 
 
Day 35  
 
 
 
Discontinue MMF 
 
 
Day 60          Assess Chimerism in peripheral blood and bone marrow (with biopsy) 
 
 
Day 180 
Evaluate disease 
 
 
 
     Assess Chimerism in peripheral blood 
 
 
Day 365           
Discontinue tacrolimus if no GVHD and full chimera*** 
 
Evaluate disease 
 
 
 
     Assess Chimerism in peripheral blood and bone marrow 
 
 
2 yrs 
Evaluate disease 
 
 
 
     Assess Chimerism in peripheral blood 
 
* Refer to Section 5.3 for complete dosing instructions. 
** Or as per institutional standards. 
*** Refer to Section 5.6 for complete dosing instructions. 
 
 


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1.0 Primary Objective 
Our primary objective is to determine if it is feasible for SAA patients to be 
transplanted using non-myeloablative conditioning and post transplantation 
cyclophosphamide with partially HLA-mismatched donors.  
 
2.0 Secondary Objectives 
2.1 To estimate overall survival at one year. 
2.2 To estimate full donor chimerism by day 60 
2.3 To estimate the cumulative incidence of non–relapse-related mortality 
following transplant. 
2.4 To estimate the incidences of primary and secondary graft failure following 
transplant. 
2.5 To estimate the cumulative incidences of grade II-IV and grade III-IV acute 
graft versus-host disease (GVHD).  
2.6 To estimate the cumulative incidence of chronic graft versus-host disease 
(GVHD).  
2.7 To estimate the cumulative incidence of ANC and platelet recovery.  
2.8 To estimate GVHD free relapse free survival (GRFS). 
2.9 To summarize major transplant related toxicities and to estimate transplant 
related mortality (TRM). 
 
3.0 BACKGROUND 
Bone marrow failure (BMF) syndromes (acquired and inherited) are a heterogeneous 
group of disorders marked by ineffective hematopoiesis76. They confer a significant risk of 
morbidity and death, due to their progressive natural history and complications of 
suboptimal therapy.2, 76 Acquired severe aplastic anemia (SAA) is a rare, life-threatening 
hematopoietic stem cell disorder that manifests with pancytopenia and a hypocellular bone 
marrow.6, 24 In most cases, bone marrow failure results from autoimmune destruction of 
hematopoietic stem cells. Without definitive treatment, mortality from SAA approaches 
70% at two years. Fungal infections are the leading cause of death; however, hemorrhage, 
evolution to clonal disease (myelodysplastic syndromes [MDS], leukemia, and paroxysmal 
nocturnal hemoglobinuria [PNH]), and transfusional iron overload are other causes of 
severe morbidity and mortality. Inherited bone marrow failure syndromes (IBMFS) from 
disorders such as short telomeres (dyskeratosis congenital [DKC]), Fanconi anemia and 
other congenital SAA also carries the risk of infection and clonal evolution to MDS.  These 
patients have long pursued BMT as a therapeutic avenue but are limited still by availability 
of matched sibling donor. 
 
SAA affects all ages, but is most common in children and young adults. Blood or marrow 
transplantation (BMT) from an HLA-matched sibling donor can cure most patients with 
SAA, but fewer than 30% of patients have a suitable HLA-matched sibling. Moreover, the 
best results with allogeneic BMT are in children; adults, especially those over age 40, do 
less well due to complications from graft-versus-host disease (GVHD).  Alternative donor 
transplants (mismatched and unrelated donors) also have the potential to cure SAA, but 
are currently reserved for second-line therapy because of their high rates of morbidity and 
mortality.  For SAA patients who lack matched sibling donors or are not good candidates 
for BMT, immunosuppressive therapy can also be highly effective.  Patients with acquired 
SAA, not responsive to initial IST, or inherited AA lacking other treatments options require 
additional therapeutic options that are not limited by age or donor status. Patient age and 
the type of allograft (HLA-matched sibling, unrelated, or mismatched donors) are the most 
important factors influencing the timing of BMT and outcomes post-transplant. To improve 


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survival in patients who either do not response to IST or do not have matched sibling 
donor, there needs to be an available transplant option for all that is feasible and safe.  An 
approach to BMT using post-transplant CY has allowed allogeneic BMT from matched, 
mismatched, unrelated or haplo-identical donors in other diseases.38, 39, 41 Transplant-
related mortality, graft-failure rates and risk of GVHD have been very low with this 
approach with non-myeloablative conditioning. Here we use alternative donors with 
CY post BMT to decrease GVHD to expand the donor pool in AA. 
 
Immunosuppressive therapy for SAA 
Antithymocyte globulin and cyclosporine (ATG/CSA) immunosuppressive therapy (IST) is 
generally front-line therapy for SAA patients who lack matched sibling donors or are not 
good candidates for BMT. The hematopoietic response rate after ATG/CSA is 60-70% 
and the probability of survival at 5 years ranges from 60% to 85%.17-20  However, up to 
40% of patients eventually relapse and an additional 10 to 40% develop a secondary 
clonal disease.17;18;21;22   
 
High-dose cyclophosphamide (High-CY) High-CY is also highly immunosuppressive and 
has also been used to successfully treat SAA.5  A pilot study of 10 patients treated with 
high-dose cyclophosphamide demonstrated that this approach may have the potential to 
cure SAA.7 The incidence of clonal evolution after HiCY is likely less than with ATG/CsA 
but the response rates remain the same. Immunosuppressive therapy, either ATG/CSA or 
high-CY is used as initial therapy for older patients or younger patients who lack an HLA-
matched sibling donor but only 60-80% of these patients respond and have late 
complications with clonal evolution and relapse.10 
 
Bone marrow transplantation for SAA 
Allogeneic BMT from an HLA-matched sibling donor is the treatment of choice at most 
centers for young patients with SAA. A major advantage of BMT over standard IST is a 
marked reduction in the risk of relapse and the outgrowth of late clonal disorders such as 
MDS/AML and PNH21. The overall transplant related mortality (TRM) attributable to HLA-
identical sibling BMT is 20-30%, with approximately half of these deaths occurring in the 
first 100 days.  The incidence of severe, grade III-IV acute GVHD is approximately 13-
17% after HLA-identical sibling BMT.   
 
Historically, unrelated donors and mismatched or haplo- transplants have almost twice the 
transplant-related mortality and risk of GVHD as matched sibling donor transplants in 
SAA2. Despite the fact that only 30% of people have matched donors and the average 
person has 4.5 haplo-identical donors, these statistics have limited the use of these 
alternative donors. The best results with unrelated and mismatched transplants are seen 
in patients under 21 with disease duration of less than one year. The International Bone 
Marrow Transplant Registry reported on the results of 318 alternative donor transplants in 
patients with SAA between 1988 and 199818. 
 
Regimen for BMT in SAA 
The ideal BMT regimen is one that results in sustained engraftment, minimal toxicity from 
the regimen, lack of acute or chronic GVHD and allows the majority of patients (old and 
young) to proceed efficiently to this potentially curative option. 
 
Conditioning: Various conditioning regimens have been employed over the years for 
transplants to patients with SAA with failure to engraft and GVHD as the main obstacles 
to success.  The current approach used here is one that has been successfully used as 


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above in sickle cell disease.4 Historically, TBI- based conditioning regimens reduced the 
risk of graft rejection but increased GVHD and other late effects.19 Standard ATG-based 
conditioning regimens are employed to aid in engraftment but have also been associated 
with up to a 30% incidence of cGVHD.8, 14 Fludarabine has been used in conditioning for 
patients with both acquired and constitutional aplastic anemia with good results.11, 13, 17, 23  
 
Choice of Stem Cells: This trial will utilize bone marrow as the stem cell source. The EBMT 
reviewed outcomes in nearly 700 patients with SAA receiving transplants from HLA-
matched siblings. In patients younger than 20 years of age, rates of chronic GVHD 
(relative risk 2.82; p = 0.002) and overall mortality (relative risk 2.04; p = 0.024) were 
higher after transplantation of peripheral blood progenitor cell grafts than after 
transplantation of bone marrow. In younger patients, the 5-year survival was 85% after 
marrow transplants but only 73% after peripheral blood progenitor cell grafts. These data 
suggest that bone marrow grafts are preferable in this age group3, 20.  Furthermore, reports 
suggest that G-SCSF mobilization may a reasonable approach to decrease GVHD in 
SAA.9, 22 GCSF stimulation of the donor will not be mandated with this protocol but we will 
adhere to the instutitutional standards for donor preparation at the time of transplant. 
  
GVHD prophylaxis: An approach to BMT at Hopkins using post-transplant 
cyclophosphamide has allowed for transplantation of allogeneic BMT from matched, 
mismatched, unrelated or haplo-identical donors in both malignant and non-malignant 
diseases.4, 15, 16  The administration of a properly timed, high dose of CY after BMT inhibits 
both graft rejection and GVHD19-22.  It is customary for IST for GVHD to continue post-
transplant through one year in SAA.1  
 
In conclusion, the major challenge in treating SAA (inherited and acquired) is the 
management of patients who are refractory to IST, have relapsed after IST, or who have 
acquired a secondary clonal disorder (MDS/PNH) after IST.  BMT is the only curative 
option for these patients, but many are ineligible because they lack a suitable donor or are 
too high risk for BMT due to the risk GVHD. Here we seek to increase options for these 
patients by developing novel therapeutic strategies to treat refractory SAA with expansion 
of the donor pool through minimization of the post-transplant complication of GVHD. 
Present above are promising results in the non-myeloablative haplo-identical setting for 
hematologic malignancies and sickle cell disease4, 15, 16 with a low incidence of engraftment 
failure, severe acute GVHD, extensive chronic GVHD, and NRM utilizing post-
transplantation CY.  This trial will employ non-myeloablative conditioning regimen along 
with post-transplantation CY on days +3 and +4 for patients with SAA. We anticipate HiCY 
will likely ameliorate the GVHD while the combination of ATG, fludarabine and TBI will 
allow for engraftment.    Furthermore, this approach allows us to greatly expand the donor 
pool since any patient shares exactly one HLA haplotype with each biological parent or 
child and half of siblings, an eligible haplo-identical donor can be identified rapidly in the 
majority of patients.  This is of great benefit to aplastic patients as time to treatment will 
be shorter. The purpose of the current trial is to improve response rates and cure for in 
SAA above the standard 70% from immunosuppressive therapy for acquired disease. 
 
 
3.0   DRUG INFORMATION 
 
3.1 
Fludarabine 
 
Fludarabine phosphate is commercially available. 


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Fludarabine phosphate is purine antimetabolite that, after administration, 
undergoes rapid conversion in plasma to the nucleoside 2-fluoro ara-A (F-araA). 
F-araA subsequently enters cells where it is phosphorylated to F-araATP and the 
monophosphate F-araAMP.  Once activated, F-araATP inhibits DNA polymerase 
and ribonucleotide reductase. The monophosphate F-araAMP, once incorporated 
into DNA, is an effective DNA chain terminator. 
 
Fludarabine monophosphate, 50 mg/vial, is reconstituted with 2 ml of sterile water, 
resulting in a 25mg/ml solution. The desired dose is further diluted to 
concentrations of 0.04-1 mg/ml in normal saline or 5% dextrose (50-100ml) for 
injection and will be administered by IV infusion over 30 minutes or longer. 
 
Following IV administration, the drug is metabolized to 2-F-araA and widely 
distributed in tissues.  2-F-araA is excreted primarily in urine and has a terminal 
elimination half-life of 7 to 12 hours. 
 
Clinical toxicities of fludarabine monophosphate include: myelosuppression, 
primarily lymphopenia and granulocytopenia, alopecia, rash, dermatitis, nausea, 
vomiting, anorexia, stomatitis, diarrhea, somnolence, fatigue, peripheral 
neuropathy, mental status changes, cortical blindness, hepatocellular toxicity with 
elevation in serum transaminases, and interstitial pneumonitis. These effects are 
reversible when the drug is discontinued. 
 
Fludarabine will be administered by IV infusion over 30 minutes in a dose of 30 
mg/m2/day on days -6 to -2. 
 
Fludara® will be dispensed by the Oncology Pharmacy and is produced by Berlex 
Pharmaceuticals. 
 
3.2 
Cyclophosphamide (Cytoxan®) 
 
 
Cyclophosphamide is commercially available. 
 
Cyclophosphamide is an alkylating agent which prevents cell division primarily by 
cross-linking DNA strands.  Cyclophosphamide is cell cycle non-specific. 
 
Cyclophosphamide for injection is available in 2000 mg vials which are 
reconstituted with 100 ml sterile water for injection.  The concentration of the 
reconstituted product is 20 mg/ml.  The calculated dose will be diluted further in 
250-500 ml of Dextrose 5% in water.  Each dose will be infused over 1-2 hr 
(depending on the total volume). 
 
Clinical toxicities of cyclophosphamide include alopecia, nausea and vomiting, 
headache and dizziness, hemorrhagic cystitis, cardiotoxicity, immunosuppression, 
myelosuppression, pulmonary fibrosis, increased hepatic enzymes and syndrome 
of inappropriate anti-diuretic hormone (SIADH). 
 
Cyclophosphamide will be dispensed by the Oncology Pharmacy and is produced 
by Mead Johnson Pharmaceuticals. 
 


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3.3 
Mesna (sodium-2-mercapto ethane sulphonate) 
 
Mesna is a prophylactic agent used to prevent hemorrhagic cystitis induced by the 
oxasophosphorines (cyclophosphamide and ifosphamide).  It has no intrinsic 
cytotoxicity and no antagonistic effects on chemotherapy.  Mesna binds with 
acrolein, the urotoxic metabolite produced by the oxasophosphorines, to produce 
a non-toxic thioether and slows the rate of acrolein formation by combining with 4-
hydroxy metabolites of oxasophosphorines. 
 
Mesna is available in 200 mg, 400 mg and 1000 mg vials containing a 100 mg/ml 
solution.  Each dose of mesna will be diluted further in 50 ml of normal saline to be 
infused over 15 minutes (or as per institutional standards). Mesna dose will be 
based on the cyclophosphamide dose being given.  The total daily dose of mesna 
is equal to 80% of the total daily dose of cyclophosphamide. 
 
At the doses used for uroprotection mesna is virtually non-toxic.  However, adverse 
effects which may be attributable to mesna include nausea and vomiting, diarrhea, 
abdominal pain, altered taste, rash, urticaria, headache, joint or limb pain, 
hypotension and fatigue. 
 
Mesna will be dispensed by the Oncology Pharmacy and is produced by Mead 
Johnson Pharmaceuticals. 
 
3.4 
Tacrolimus (FK 506) 
 
Tacrolimus, also known as FK-506, is a macrolide immunosuppressant. It inhibits 
lymphocytes by forming a complex with FKBP-12, calcium, and calmodulin, 
leading to the decrease in the phosphatase activity of calcineurin. This drug is 
used with corticosteroids for prophylaxis of organ rejection in patients receiving 
allogeneic liver transplants. Its use is also currently being investigated in kidney, 
bone marrow, cardiac, pancreas, pancreatic islet cell and small bowel 
transplantation. This drug is well-absorbed orally. It is metabolized in the liver by 
unknown mechanisms, but demethylation and hydroxylation have been proposed 
based on in vitro studies. The metabolized products are excreted in the urine. 
Nephrotoxic drugs, antifungals, calcium channel blockers, cimetidine, danazol, 
erythromycin, methylprednisone and metoclopramide increase the bioavailabilty 
of FK-506. In contrast, phenobarbital, phenytoin, rifamycins and carbamazepine 
decrease FK-506 levels. Adverse reactions include tremor, headache, diarrhea, 
hypertension, nausea, and renal dysfunction. 
 
 
 
3.5 
Mycophenolic Acid Mofetil (Cellcept®) 
 
Mycophenolate Mofetil is an ester prodrug of the active immunosuppressant 
mycophenolic acid (MPA). This active metabolite is a noncompetitive, reversible 
inhibitor of inosine monophosphate dehydrogenase (IMPDH). There are no 
pharmacokinetic interactions with ganciclovir, cotrimoxazole, oral contraceptives 
and cyclosporine.  
 


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Side effect profiles include diarrhea, leukopenia, sepsis, allergic reactions, and 
vomiting. There is also an increase in certain types of infection mainly from the 
herpes virus family (CMV, HSV & VZV) and Candida.  
 
3.6  
Rabbit antithymocyte globulin (ATG) 
 
Thymoglobulin® [Anti-thymocyte Globulin (Rabbit)] is a purified, pasteurized, 
gamma immune globulin, obtained by immunization of rabbits with human 
thymocytes. This immunosuppressive product contains cytotoxic antibodies 
directed against antigens expressed on human T-lymphocytes. This drug is 
commonly used to treat graft rejection in kidney transplantation. It is also 
commonly used in bone marrow transplantation as part of the conditioning regimen 
to avoid graft failure and to prevent graft-versus-host disease. 
 
Thymoglobulin is a sterile, freeze-dried product for intravenous administration after 
reconstitution with Sterile Water for Injection, USP (SWFI). Each 10 mL vial 
contains 25 mg anti-thymocyte globulin (rabbit) as well as 50 mg glycine, 50 mg 
mannitol, and 10 mg sodium chloride. After reconstitution with 5 mL SWFI, each 
vial of reconstituted product contains approximately 5 mg/mL of Thymoglobulin, of 
which >90% is rabbit gamma immune globulin (IgG). The reconstituted solution 
has a pH of 7.0 ± 0.4. Human red blood cells are used in the manufacturing process 
to deplete cross-reactive antibodies to non-T-cell antigens. The manufacturing 
process is validated to remove or inactivate potential exogenous viruses. All 
human red blood cells are from US registered or FDA licensed blood banks. A viral 
inactivation step (pasteurization, i.e., heat treatment of active ingredient at 60°C/10 
hr) is performed for each lot. Each Thymoglobulin lot is released following potency 
testing (lymphocytotoxicity and E-rosette inhibition assays), and cross-reactive 
antibody testing (hemagglutination, platelet agglutination, anti-human serum 
protein antibody, antiglomerular 
 
Adverse side effects include immunodeficiency, infusion related toxicities such as 
hypertension, chills, rigors, tachycardia, capillary leak syndrome, hyperglycemia, 
cytopenias, transient hepatitis, anaphylaxis, serum sickness, myalgias, sensory 
changes including hearing loss, headaches, renal toxicity, dyspnea and bronchial 
spasm, fevers. The drug is potentially teratogenic and is unknown if it can be 
passed to children in breastfeeding. 
 
Thymoglobulin will be dispensed by the Oncology Pharmacy and is produced by 
Genzyme. ATG–rabbit must be infused through a 0.22 micro filter with 
premedications: acetaminophen 650 mg orally and diphenhydramine 25mg orally 
as well as a steroid taper (see Section 6.3). The dose to be used is 0.5 mg/kg on 
day -9 and 2 mg/kg/day on days -8 and -7. Note: Keep anaphylaxis kit at bedside 
during ATG administration.  
 
 
4.0 
PATIENT SELECTION 
 
4.1 Criteria for recipient eligibility 
 
 
4.1.1  Patients with relapsed or refractory SAA or very SAA defined: 
 


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 
Bone marrow (< 25% cellular) 
 
Peripheral cytopenias (at least 2 of 3) 
o ANC < 500 per ml 
o Platelets < 20,000 per ml 
o Absolute retic < 60,000 or corrected retic < 
1% 
 
Very severe: as above, but ANC < 200  
 
Disease may be designated as acquired or inherited 
if previous counts known (these other bone marrow 
failure disorders that are characterized by aplastic 
anemia may go by additional names such as 
dyskeratosis congenita or PNH) 
 
Failed at least one course of immunosuppressive 
therapy (if presumed acquired disease). Patients 
with inherited disease will be characterized as 
refractory and do not require immunosuppressive 
first.  
 
 
4.1.2   Age 0- upper age limit as determined by current institutional standards 
 
 
4.1.3 Good performance status (ECOG 0 or 1; Karnofsky and Lansky 70-100) 
 
4.1.4  Patients and donors must be able to sign consent forms (or if a minor 
the parent will sign). Donors should be willing to donate. 
 
 
4.1.5 Patients must be geographically accessible and willing to  
           participate in all stages of treatment. 
 
 
 
       4.1.6  Adequate end-organ function as measured by: 
a. Left ventricular ejection fraction  35%, or shortening fraction > 25% 
(For pediatric patients, a normal ejection fraction is required) 
b. Bilirubin ≤ 3.0 mg/dL (unless due to Gilbert’s syndrome or hemolysis), 
and ALT and AST < 5 x ULN 
c. FEV1 and FVC > 40% of predicted; or in pediatric patients, if unable to 
perform pulmonary function tests due to young age, oxygen saturation 
>92% on room air   
 
 
 
 
 
4.2   Criteria for recipient ineligibility 
4.2.1 Patients will not be excluded on the basis of sex, racial or 
ethnic background. 
4.2.2 Prior transfusions from selected donor (as this could have 
cause recipient alloimmunization against the donor) 


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4.2.3  Women of childbearing potential who currently are 
pregnant (HCG+) or who are not practicing adequate 
contraception. 
4.2.4 Patients who have any debilitating medical or psychiatric 
illness that would preclude their giving informed consent or 
their receiving optimal treatment and follow up. 
4.2.5 Uncontrolled viral, bacterial, or fungal infections (HIV 
infection permitted if viral load undetectable) 
 
4.3 Criteria for donor eligibility 
 
  
 
4.3.1 Donors must meet the selection criteria as defined by the 
Foundation for the Accreditation of Hematopoietic Cell 
Therapy (FAHCT) and will be screened per the American 
Association of Blood Banks (AABB).  (AABB guidelines and 
the recipients will be informed of any deviations.) 
4.3.2 Weight ≥ 20kg and patient or guardian able to provide 
consent 
4.3.3 HLA mismatched or haploidentical related donors (including 
1st degree relatives and half siblings).  
4.3.3.1 The donor and recipient must be identical at 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. 
4.3.4 Matched unrelated donors 
4.3.4.1 Unrelated volunteer donor matched for HLA-A, -B, -
C and -DRB1 defined by high resolution molecular 
typing.  
4.3.4.1.1 Mismatched unrelated volunteer donors 
may be considered if no other suitable 
donor is available. 
4.3.5 When more than one donor is available, the donor with the 
lowest number of HLA allele mismatches will be chosen, 
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, in consultation with 
the immunogenetics laboratory.  In cases where there is 
more than one donor with the least degree of mismatch, 
donors will be selected based on the most favorable 
combination of (i) HLA compatibility in cross-match testing 
and (ii) ABO compatibility. We will prioritize the lowest 
number of mismatches in the HVG direction (to potentially 
minimize graft rejection risk) 
 
 
 
 
Donor selection criteria, in decreasing order of priority: 
 
1. Donor must be medically, socially, and psychologically fit to donate. 


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2. HLA-identical sibling. 
 
3. For a partially HLA-mismatched transplant, the patient must lack antibodies 
against donor HLA molecules.  Specifically, complement dependent 
cytotoxicity and flow cytometric crossmatch assays must be negative, and the 
mean fluorescence intensity (MFI) of any anti-donor HLA antibody by solid phase 
immunoassay should be <3000.  Consult with Immunogenetics for the clinical 
significance of any anti-donor antibody.  Desensitization to remove anti-donor 
antibody should only be performed for patients who have no other donor options 
 
If there is more than one donor with the least amount of HVG allele mismatches, the 
following prioritization will be used: (Will always minimize HVG mismatch as highest 
priority) 
 
 
4. ABO compatibility (in order of priority). 
 
a. Compatible or minor ABO incompatibility 
b. Major ABO incompatibility 
 
5. CMV status 
 
The CMV status of the pair donor-recipient is frequently employed to select a potential 
donor. This is a controversial issue and the data available is somewhat limited. The 
following guidelines are recommended: 
 
a. For a CMV seronegative recipient, use a CMV seronegative donor 
b. For a CMV seropositive recipient, use a CMV seropositive donor 
 
In CMV- patients with CMV+ stem-cell donors, primary CMV infection/reactivation 
develops in about 30%. Data from the European Registry shows the following: 
Seropositive patients receiving grafts from CMV+ HLA-identical sibling donors had the 
same survival as patients grafted from CMV- donors. However, MUD recipients receiving 
grafts from CMV+ donors had an improved 5-year survival, an improved event-free 
survival, and a reduced transplant-related mortality. There was no influence on the 
relapse incidence. The effects of donor CMV status remained in multivariate analyses. 
The effect of donor status was different among different disease categories. In patients 
with chronic myelogenous leukemia, T-cell depletion abrogated the beneficial effect of 
donor status, suggesting that the effect is mediated through transfer of donor immunity. 
These data suggest that donor CMV status influences outcome of unrelated SCT. 
 
6. Other donor characteristics 
 
Donor parity and sex mismatch, have also been associated with an increased risk of 
aGVHD and decreased survival in some but not all studies(4-8). Donor age and weight 
should be also taken into consideration. 
 
 
Suggestions (in no order of priority): 


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a. Younger (18 years of age or older) and lighter donors should be 
preferred. 
If all else is equal, male donors may be preferred over nulliparous female donors who 
may be preferred over multiparous female donors. 
 
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. 
 
5.0        TREATMENT PLAN 
 
5.1  
Indwelling central venous catheter 
 
A central venous catheter may be placed for administration of IV 
medications and transfusion of blood products as per BMT standard. 
 
5.2  
Pre-treatment Evaluation 
 
All patients will require documentation of a detailed history and physical 
examination and standard evaluation of cardiac, pulmonary, liver and renal 
function.  
 
Baseline disease evaluation will be performed by collecting the following 
laboratory tests:  complete blood count, reticulocyte count, LDH, PNH 
panel, bone marrow biopsy (please see Table 1) 
 
5.3  
Preparative regimen 
 
 
Thymoglobulin will be infused through a 0.22 micro filter with 
premedications: acetaminophen 650 mg orally and diphenhydramine 25mg 
orally. Keep anaphylaxis kit at bedside during ATG administration. The 
dose will be 0.5 mg/kg IV on day -9 over 6 hours and 2mg/kg IV on days -
8 and -7 over 4 hours. A steroid taper will be  
given 
to 
prevent 
reactions to ATG as follows (or per institutional standards): Solumedrol 
1mg/kg IV 1 hour prior ATG on days -9 to -7. This dose may be repeated 
once 3 hours after the first dose. On day -6 and -5, solumedrol 0.75 mg/kg/ 
IV as a single dose; on days -4 and -3, solumedrol 0.5 mg/kg/ IV as a single 
dose; on day -2 solumedrol 0.25 mg/kg/ IV as a single dose.  
 
Fludarabine 30 mg/m2/day (adjusted for renal function) is administered over a 30-
60 minute IV infusion on Days –6 through –2 (maximum cumulative dose, 150 
mg/m2). 
 
The body surface area (BSA) for fludarabine dosing is based on actual body 
weight.   
 
For patients > 18 years old, CrCl will be estimated by the Cockcroft Formula, 
based on weight:  
 
 
CrCl = (140 – age) x weight (kg)   x   0.85 for females 
 
 
                       PCr x 72 
 


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When calculating CrCl:  if Actual Body Weight is less than Ideal Body Weight, 
Actual Body Weight will be used; if Actual Body Weight is between 100-120% of 
Ideal Body Weight, Ideal Body Weight will be used; and if Actual Body Weight is 
> 120% of Ideal Body Weight, 25% Adjusted Body Weight 
 
For decreased creatinine clearance (CrCl), fludarabine dosage is reduced as 
follows: 
CrCl ≥ 70 ml/min – fludarabine 30 mg/m2 
CrCl 40-70 ml/min -  fludarabine 24 mg/m2 
CrCl 20-40 ml/min – fludarabine 20 mg/m2 
CrCl < 20 ml/min – fludarabine 15 mg/m2 
 
For patients <18 years old, CrCl will be estimated by the Schwartz equation: 
Schwartz equation: CrCl (mL/min/1.73m2)= [length (cm) x k] / serum creatinine 
k = 0.45 for infants 1 to 52 weeks old 
k = 0.55 for children 1 to 13 years old 
k = 0.55 for adolescent females 13-18 years old 
k = 0.7 for adolescent males 13-18 years old
A measured CrCl or a glomerular filtration rate may be substituted to determine 
CrCl. 
CrCl  may change during the days fludarabine is given.  The dose should be 
adjusted accordingly; however a fludarabine dose may be given based on the 
preceding day’s estimated CrCl .  
 
Cyclophosphamide will be administered as an IV infusion over 1 hour for the 
pretransplant CY and over 2 hours for post-transplant CY on D-6 and D-5. The 
dose of pre-transplantation cyclophosphamide is 14.5 mg/kg/day.  Dose is 
calculated based on the ideal body weight or actual body weight whichever is less. 
(Refer to Appendix 2.)  Body weight and height are measured directly.  An 
approximate weight for height would be calculated from a standard table or 
equations that reflect ideal “values”. 
 
 
IBW is calculated by the following equation: 
 
𝐼𝐵𝑊= √ [ℎ𝑡 (𝑐𝑚)𝑥 𝑤𝑡 (𝑘𝑔)]
3600
 
 
 
 
Note:  Mesna will be utilized for the Day 3 and Day 4 post BMT cyclophosphamide 
doses, not for the pre-BMT cyclophosphamide doses. 
 
Total body irradiation: 200 cGy AP/PA with 4MV or 6MV photons at 8-12 cGy/min 
at the point of prescription (average separation of measurements at mediastinum, 
abdomen, hips) will be administered in a single fraction on day -1. 


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Hydration and antiemetic administration will follow standard operating procedures. 
 
5.4  
Bone marrow transplantation and graft information 
 
Bone Marrow (T cell replete graft) will be harvested and infused on day 0.  
The BM will be infused the same day of collection. The nucleated cell target 
range will be between 8 to 16 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 8 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 surgery if the minimum 
target of 8 x 108 cells has not been reached and it is believed that will be 
unsafe or technically unfeasible to reach the mentioned target if the surgery 
was to continue after a reasonable effort has been made. In this case, the 
study PI will be notified immediately and efforts should be made to assure 
at least a cell count of 2-4 x 108/kg of recipient ideal body weight. 
 
Major incompatible ABO graft will have red blood cell depleted by buffy coat 
preparation. Minor ABO incompatible graft will have plasma removed.  
Guidelines for the infusion of bone marrow have been established and are 
outlined in the ABO compatible/minor mismatched allo BMT or the ABO 
incompatible allo BMT standing orders.   
 
5.5 
Post-transplantation cyclophosphamide 
 
Cyclophosphamide [50mg/kg] will be given on D+3 post-transplant (within 
48-72 hr of start of marrow infusion as per institutional standards) and on 
D+4 post-transplant. Cyclophosphamide will be given as an iv infusion over 
1- 2 hr (depending on volume).  
 
Mesna will be given in divided doses iv 30 min pre- and at 3, 6, and 8 hr 
post-cyclophosphamide or administered per institutional standards. Mesna 
dose will be based on the cyclophosphamide dose being given.  The total 
daily dose of mesna is equal to 80% of the total daily dose of 
cyclophosphamide.  
 
It is crucial that no immunosuppressive agents are given from the 
time of the transplant until at least 24 hours after the completion of 
the post-transplant Cy.  This includes steroids as anti-emetics. 
 
5.6 
GVHD prophylaxis 
 
On day 5, patients will begin prophylaxis with Tacrolimus (PO or IV as per 
institutional standards for starting this prophylaxis) and Mycophenolic Acid 
Mofetil (MMF).   
 
Tacrolimus begins on Day 5, at least 24 hours after completion of post-
transplantation Cy.   
 


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The tacrolimus starting dose will be given per institutional standards for 
adult or pediatric patients.  The starting dose of tacrolimus may be 
increased with PI or co-PI permission should institutional practice 
guidelines change..  Dose is adjusted to maintain a serum trough level of 
10 – 15 ng/Ml.  If there is tacrolimus toxicity at these troughs, lower 
troughs will be permitted to allow for tolerance of the drug after 
consultation with the PI.  If tacrolimus is completely impossible for the 
patient to tolerate, alternative immunosuppression may be chosen after 
consultation with the PI or Co-PI.   
 
 
***Tacrolimus is discontinued after the last dose on Day 365 without 
taper, or may be continued if GVHD is present or if patient is still a mixed 
chimera.  At PI or co-PI discretion, cyclosporine (target concentration 
200-400 ng/Ml) may be substituted for tacrolimus if the patient is 
significantly intolerant of tacrolimus. 
 
***Tacrolimus may be discontinued earlier than Day 365 in the context of 
relapse, 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.  Decisions regarding early discontinuation of 
immunosuppression will be made on a case-by-case basis in consultation 
with the PI or co-PI.  
 
Mycophenolic acid mofetil will be given at a dose of 15 mg/kg po/ IV TID 
(based upon actual body weight) with the maximum total daily dose not to 
exceed 3 grams (1 g po TID). 
 
MMF prophylaxis will be discontinued after the last dose on D35 and 
Tacrolimus prophylaxis will be discontinued after the last dose around day 
365. 
 
5.7 
Infection prophylaxis and therapy 
 
During pre-transplant evaluation patients will be screened for respiratory 
16yncytial virus, influenza A, B and parainfluenza viruses if symptomatic or 
during the flu season.  Assays of these viruses must be negative for 
symptomatic patients to be admitted for transplant.  Strong consideration 
should be given to institution of ribavirin therapy if positive for adenovirus 
or nalidixic acid if positive for BK virus. 
 
Oral hygiene will be maintained according to institutional standards. 
 
Prophylactic anti-microbial therapy will be given per the BMT unit 
standards. 
 
 
Empiric therapy with broad-spectrum antibiotics will be instituted for the first 
neutropenic fever (specific agents as per current practice). 
 


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Antifungal prophylaxis will be administered according to institutional 
preference. It is important to follow tacrolimus levels on patients receiving 
azoles as the combination of both drugs can raise the levels of the 
immunosuppressant to toxic levels. If the patient on tacrolimus is started 
on azoles, a dose reduction of tacrolimus is required and levels should be 
obtained to be sure the levels are not in the toxic range.  
 
Pneumocystis jiroveci pneumonia (PCP) prophylaxis will be administered 
according to institutional preference starting around Day 21 (or later if WBC 
not recovering) and should continue for at least the first year following BMT 
and while on immunosuppression.  If the patient cannot tolerate po then a 
comparable dose of Bactrim will be given iv.  Patients intolerant of Bactrim 
will receive dapsone, atovaquone, or pentamidine as PCP prophylaxis. 
 
 
Viral prophylaxis for HSV/ VZV will be administered according to 
institutional preference and should continue for at least the first year 
following BMT and while on immunosuppression. 
 
CMV viremia (by PCR) or antigenemia (by ELISA) should be documented 
weekly or every other week beginning once the WBC>1000 and until 
discharge. Monitoring of CMV viremia or antigenemia is recommended to 
continue on a weekly or every other week basis until day 100, then bi-
weekly until day 180. Patients who are viremic or antigenemic will be 
treated according to institutional preference.  
 
 
5.8  
Growth factor support 
 
GCSF (filgrastim) begins on Day 5 at a dose of 5 mcg/kg/day (actual body 
weight) IV or subcutaneously (rounding to the nearest vial dose is allowed), until 
the absolute neutrophil count (ANC) is ≥ 1,000/mm3 over the course of three 
days.  Additional GCSF may be administered as warranted.  Pegfilgrastim 
(Neulasta®) and GM-CSF are not permitted. 
 
5.9 
Transfusion support 
 
 
Packed red cell transfusions and platelet transfusions will be given per 
current 
institutional 
recommendations. 
If 
the 
patient 
becomes 
alloimmunized, the blood bank service will select the best blood products 
to transfuse and will coordinate with the medical floor the transfusion 
needs. 
 
 
5.11 
Anti-ovulatory treatment  
 
Menstruating females should be started on an anti-ovulatory agent prior to 
the initiation of the preparative regimen.  The treatment administered will 
be at the discretion of the treating physician. 
 
 


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5.0 PATIENT MONITORING 
 
The following parameters will be obtained according to this schedule: (for details 
of these evaluations, see text sections 6.1-6.3) 
 
TABLE 1: 
 
Initial 
Allowable 
time 
frame 
from date 
of 
consent** 
Day  
< 60 
Day 
30  7 
Day 
60  7 
Day 
180 
 
30 
 
Day 
365 
and 
Day 
730 
 
60 
 
History and Physical 
X 
within 30 
days 
 
X 
X 
X 
X 
Performance status 
(ECOG or Karnofsky) 
X 
within 30 
days 
 
 
X 
X 
X 
Disease staging 
(including bone 
marrow biopsy) 
Unless clinically 
indicated by CBC or 
physician discretion
X 
within 30 
days 
 
 
 
 
 
CBC & Diff. 
X 
within 
7 
days
*weekly
X 
X 
X 
X 
Comprehensive 
Metabolic Panel
X 
within 
7 
days
  
weekly
X 
X 
X 
X 
reticulocyte count, 
LDH, PNH panel, 
X 
 
 
 
 
 
 
CXR 
(May be Chest CT if 
clinically indicated) 
X 
within 30 
days 
 
 
 
 
 
Sinus CT 
X 
within 30 
days 
 
 
 
 
 
Pregnancy test 
(women, childbearing 
age)
X 
within 30 
days 
 
 
 
 
 
Peripheral blood 
chimerism, both total 
leukocyte (unsorted) 
and T-cell sorted) 
X 
 
 
X 
X 
X 
X 
PT, PTT 
X 
within 30 
days 
 
 
 
 
 
EKG 
X 
within 60 
days 
 
 
 
 
 
ECHO (or MUGA) 
X 
within 60 
days 
 
 
 
 
 
Hep B Surface Ag, 
HBC Ab, HCV Ab, 
X 
within 30 
days 
 
 
 
 
 


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HSV IgG, CMV IgG, 
RPR, HIV, VZV IgG (if 
possible)
Toxicity assessment
X
X
X
X
X
HLA 
typing/lymphocytotoxic 
screen
X 
Must 
be 
done 
at 
JHH
 
 
 
 
 
PFTs (Spirometry and 
DLCO)
X 
within 30 
days
 
 
 
 
 
GVHD questionnaire
X
X
 
* Once ANC >100, this will be obtained daily until ANC >500 for three days or two 
consecutive measurements over a three day period, then weekly. 
**Baseline laboratory tests and radiology studies time frame will follow BMT standards.  
*** In order to minimize the need for research-only in-person visits, telemedicine visits may be 
substituted for in-person clinical trial visits or portions of clinical trial visits where determined to 
be appropriate and where determined by the investigator not to increase the participant’s risks. 
Prior to initiating telemedicine for study visits the study team will explain to the participant, what 
a telemedicine visit entails and confirm that the study participant is in agreement and able to 
proceed with this method. Telemedicine acknowledgement will be obtained in accordance with 
the Guidance for Use of Telemedicine in Research. In the event telemedicine is not deemed 
feasible, the study visit will proceed as an in-person visit. Telemedicine visits will be conducted 
using HIPAA compliant method approved by the Health System and within licensing restrictions. 
 
 
6.1  
Pre-transplant Evaluation 
These represent the basic baseline studies required on all patients prior to 
starting their preparative regimen. Additional investigations may be 
clinically indicated in certain individuals.  
 
6.1.1. Complete medical history which should include particular attention 
to the following details: 
a) previous medical conditions 
b) previous transfusions and transfusion reactions 
c) previous serious infections 
d) allergies 
e) current medications 
f) assessment of performance status 
g) focused history for evidence of constitutional aplastic anemia 
 
 
6.1.2. Thorough general medical evaluation which should include: 
a) a careful physical examination 
b) evaluation for placement of a central venous access device, if the patient 
does not already have such a catheter. 
 
6.1.3. Baseline investigations including: 


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a) Hematologic 
i.   CBC with platelets, differential, reticulocyte count, LDH, PNH 
Panel 
ii.  PT, PTT 
iii. ABO and Rh typing 
iv: bone marrow aspirate and biopsy 
 
b) Chemistries 
i.  Comprehensive chemistry panel 
ii. Routine and microscopic urinalysis with C&S 
 
c) Cardiac 
i.  EKG 
ii. Echocardiogram or MUGA scan with Left Ventricular Ejection 
Fraction (LVEF) + Right Ventricular Systolic Pressure and 
evaluation for pulmonary hypertension 
 
d) Pulmonary 
i.   Chest X-ray (CT scan may be performed as clinically indicated 
but some form of chest imaging is required in advance of BMT) 
ii.  Sinus CT scan 
iii. Pulmonary function tests including at least spirometry with FEV1 
and FVC and DLCO. (pediatric patients under the age of 8 are 
excluded from this test) 
 
e) Immunologic / Infections 
i.   HBsAg, anti-HBC, anti-HCV 
ii.  RPR 
iii. HIV antibody 
iv. Serology for CMV and HSV (plus VZV – blood samples 
permitting) 
v. HLA typing/lymphocytotoxic antibody screen 
 
 
f) Chimerism studies will be drawn as a baseline for subsequent 
engraftment studies including myeloid (unsorted) and CD3 chimerism. 
 
 
 
6.2  
Post-transplant Evaluation 
 
6.2.1. Day 0 through Day 60 (+ 7 days) evaluation. These represent the 
minimum required. More frequent determinations and additional investigations 
may be indicated by the clinical condition of the patient. 
1. CBC daily with a WBC differential once the total WBC is greater than 
100 until ANC > 500 for three days or two consecutive measurements over 
a three day period; then CBC weekly with differential. 
2. Comprehensive metabolic panel once a week. 
3. Patients will have evaluations for infectious complications as clinically 
indicated. Surveillance cultures according to JHOC BMT program 
standards are recommended. 


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4. Evaluations by history and physical examination for GVHD will be 
performed as per BMT unit standards.  (see also section 6.2).  For study 
purposes, weekly GVHD summaries will be taken from these standard 
examinations from day 14 through day 60. 
 
6.2.2 Evaluations on day ~ 30 (+7 days) 
1. History and physical examination. 
2. donor chimerism on peripheral blood (including myeloid (unsorted) and 
CD3 chimerism). 
3. Disease evaluation. 
4. CBC and differential, comprehensive panel. 
5. GVHD questionnaire. 
 
 
6.2.3 Evaluations on day ~60 (+7 days) 
1. History and physical examination. 
2. Disease evaluation. 
3. Studies for donor cell chimerism on peripheral blood. 
4. CBC and white blood cell differential, reticulocyte count, 
comprehensive panel. 
5. GVHD questionnaire. 
 
 
6.2.4  Evaluations for suspected GVHD: 
 
1. Comprehensive panel 
 
2. Biopsies if needed.  
 
 
In the event that the patient is unable to return to Johns Hopkins for these visits, every 
attempt will be made to obtain data from the patient and referring physician. After 
completion of the trial at Day 60  7, patient will be followed in accordance with the JHOC 
BMT Policy and Procedure Manual.  
 
 
7.0 
POST-BMT EVALUATION 
 
Patients will be followed during (i) the initial post-BMT period (ii) IPOP care and 
(iii) after discharge to the referring physician as per standard practice.  
 
7.1      Chemotherapy toxicities  
 
The agents being used in the study are FDA approved.  These agents are used 
extensively in the Bone Marrow Transplant setting and have well defined toxicity 
profiles.  In addition, there are many expected toxicities related to a bone marrow 
transplant.  For these reasons, toxicities will be captured and recorded/graded if 
the adverse event interferes with the subject’s daily function and are considered 
clinically significant.  We will capture and grade all these events structured around 
the categories of the Common Terminology Criteria for Adverse Events (CTCAE) 
Version 4.0 for the first 60 days post BMT.  
 
Since this trial is an IPOP, inpatient and out-patient trial, the definition of an 
adverse event ‘interfering with daily function’ and ‘clinically significant’ will be 


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events that require hospitalization (outside of institutional standards for IPOP 
needs or pediatric re-hospitalization standards for this ambulatory care based on 
the outpatient clinic resources and availability).  Due to the nature of marrow 
transplantation, 
readmissions 
to 
the 
hospital 
are 
expected; 
therefore 
hospitalization (either inpatient or IPOP) during the first 60 days post-transplant 
will not be considered a SAE.  For example, if a patient has a neutropenic fever 
that requires hospitalization, then ‘neutropenic fever’ will be captured and graded 
as an adverse event.  An example of a non-captured event is if a patient has 
hypotension that is corrected by fluid administration in the outpatient setting.  This 
will not be captured as an adverse event unless the patient requires a hospital 
admission for further treatment of the hypotension. 
 
Once the patient becomes hospitalized, the above definition of ‘requiring 
hospitalization’ cannot be used to capture adverse events.  For these already 
hospitalized patients, events will only be recorded once the event is greater than 
a grade 3 or 4 as stated below. 
 
List of categories that will be recorded only if the event becomes a grade 3 
or grade 4 or meets the criteria of a SAE 
 
Cardiac disorders  
Congenital, familial and genetic disorders  
Ear and labyrinth disorders  
Endocrine disorders  
Eye disorders  
Gastrointestinal disorders  
General disorders and administration site conditions  
Hepatobiliary disorders  
Investigations  
Metabolism and nutrition disorders  
Musculoskeletal and connective tissue disorders  
Nervous system disorders  
Pregnancy, puerperium and perinatal conditions  
Psychiatric disorders  
Renal and urinary disorders  
Reproductive system and breast disorders  
Respiratory, thoracic and mediastinal disorders  
Skin and subcutaneous tissue disorders  
Vascular disorders 
 
 
The Blood/Bone Marrow category is captured as endpoints to the study.  Thus for 
this category, we will not record data according to the NCI Common Toxicity 
Criteria. 
 
7.2 
GVHD 
 
A major toxicity of allogeneic BMT from an unrelated or mismatched donor is 
GVHD.  Acute graft-versus-host disease (GVHD) shall be graded clinically 
according to the criteria developed by the consensus conference on acute GVHD14 
(Appendix 1).  All suspected cases of acute GVHD must be confirmed histologically 


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by biopsy of an affected organ (skin, liver, or gastrointestinal tract).  For purposes 
of reporting, a pathologist at SKCCC will be ultimately responsible for determining 
whether a patient does or does not have histologic evidence of GVHD. Diarrhea 
and/or hyperbilirubinemia in a patient with histologically documented skin GVHD 
may be assumed to be a manifestation of visceral GVHD and will be graded as 
such.  All patients with histologically documented, clinical grade >2 acute GVHD 
should receive initial treatment with corticosteroids (or a corticosteroid containing 
regimen if a protocol is available) according to institutional preference.  If skin 
GVHD resolves with treatment but suspected visceral GVHD does not, biopsy of 
the affected organ (liver or gastrointestinal tract) should be obtained to rule out 
other causes of hyperbilirubinemia and/or diarrhea.  Steroid refractory acute GVHD 
will be treated according to institutional preferences. 
 
Acute GVHD:  Acute GVHD is graded by standard clinical criteria (Appendix 1). 
All suspected cases of acute GVHD must be confirmed histologically by biopsy of 
an affected organ (skin, liver, or gastrointestinal tract).  Date of symptom onset, 
date of biopsy confirmation of GVHD, maximum clinical grade, and dates and 
types of treatment will be recorded.  Dates of symptom onset of grade II or higher 
GVHD and grade III-IV GVHD will be recorded. 
 
Chronic GVHD:  Chronic GVHD is graded by NIH consensus criteria12. Date of 
onset, date of biopsy proof (if any), dates and types of treatment, and extent will 
be recorded. The cumulative incidence of chronic GVHD (overall, and according 
to extent) will be determined through competing risk analysis.   
 
The occurrence and severity of acute and chronic GVHD after Day 60 will be 
captured at the patients six month and annual evaluations. 
 
 
 
7.3 
Non relapse mortality (NRM) 
 
Causes of NRM, i.e., death in the absence of relapse, will be documented as 
important indicators of procedure-associated toxicity, particularly as these causes 
relate directly or indirectly to GVHD.  Analysis will stratify mortality with respect to 
the peri-transplant period (<100 d post-BMT) or later times post-BMT. 
 
7.4 
Disease Evaluation 
 
Disease evaluations will be performed at ~Day 30, ~Day 60, ~Day 180, ~Day 365, 
year 2 and then yearly thereafter per Johns Hopkins standards. (Dates are 
approximated based on patient availability)  Disease evaluations will be conducted 
as per table 1. 
 
8.0    STUDY PARAMETERS 
 
8.1 
Transplant-related mortality 
 
Transplant-related mortality, which is defined as death in the absence of relapse 
or progression, will be characterized at 100 days, 6 months and at one year after 
BMT. 
 


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8.2 
Hematologic toxicity 
 
A secondary endpoint of this Phase II trial is time to recovery of circulating neutrophils 
and platelets (following chemotherapy).  Neutrophil recovery is defined as the first day 
of three consecutive laboratory values on different days, after the conditioning 
regimen-induced nadir of blood counts, that the absolute neutrophil count is > 500/µL.  
Platelet recovery is defined as the first day of three consecutive laboratory values on 
different days, after the conditioning regimen-induced nadir of blood counts, that the 
platelet count is  20,000 L without platelet transfusion support in the seven days 
prior. 
 
 
8.3 
Donor Chimerism 
 
Definitions:  
 
Donor chimerism: Mixed donor chimerism is defined as > 5%, but < 95%, donor.  
Full donor chimerism is defined as > 95% donor. 
 
 
Graft failure: < 5% donor chimerism in blood and/or bone marrow on ~Day 30 or 
after and on all subsequent measurements.   
 
Primary graft failure: < 5% donor chimerism in blood and/or 
bone marrow by ~ Day 56  
 
Secondary graft failure: achievement of > 5% donor 
chimerism, followed by sustained <5% donor chimerism in 
blood and/or bone marrow. 
 
< 5% donor T cell chimerism, but with > 5 % donor chimerism in total 
leukocytes, is not considered graft failure. 
 
 
Donor chimerism will be measured in the peripheral blood around day 30 and day 
60.  Patients with >5% donor chimerism around day 60 will be considered as 
having engrafted. Chimerism determinations will be made on peripheral blood by 
a number of different methods depending on the specific patient.  Methods may 
include (i) the usual standard of restriction fragment length polymorphism (RFLP) 
if the donor and recipient RFLPs are informative, (ii) fluorescence in-situ 
hybridization (FISH) for Y-chromosome markers on PBMC if the donor is male, (iii) 
cytogenetic analysis, (iv) flow cytometric analysis of HLA-A, B or DR on 
lymphocytes in the peripheral blood if haploidentical and suitable reagents exist or 
(v) PCR analysis of variable nucleotide tandem repeats (VNTR) in PBMC if 
informative. Mixed donor chimerism is defined as > 5%, but < 95%, donor.  Full 
donor chimerism is defined as > 95% donor. 
 
 
8.4 
Disease Status 
 
Disease status will be evaluated through the following laboratory tests:  complete 
blood count, reticulocyte count.  These disease evaluations will be performed at 
~Day 30, ~Day 60, ~Day 180, ~Day 365, and then yearly per Johns Hopkins 


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standards. Relapse will be defined as the loss of donor chimerism given a graft 
loss is analogous to relapse in AA. 
 
 
8.5 
GVHD 
 
Patients will be followed for development of acute and chronic GVHD using 
standard criteria.  Chronic GVHD is assessed according to standard criteria. 12 
Treatment of GVHD is outlined in Section 6.2. 
 
To allow for flexibility in patient scheduling, all time points may be approximated.  
 
 
9.0 
DATA MANAGEMENT AND COLLECTION GUIDELINES 
 
Data will be maintained on uniform case report forms and appropriate Graft 
Engineering Laboratory spreadsheets provided by the SKCCC at Johns Hopkins.  
The research team at each site will make assessments of GVHD. GVHD 
assessment will be evaluated and scored by the GVHD team, the Research Nurse, 
the attending BMT physician and PI.  Hematopoietic engraftment will be assessed 
by the BMT attending and the PI at each site.  The PI at each site will be 
responsible for evaluation of chimerism data and weekly overall toxicities.  
 
For outside sites, please notify Donna Dorr when a patient has been enrolled to 
the study (within 3 days of consent).  The CRFs are to be completed as required 
by protocol and faxed or transmitted by the Internet to JH SKCCC within 30 days 
of Day 0, and then in a timely fashion for all additional study-related visits, to: 
 
 
Donna Dorr, RN, MSN 
Ph 410-502-2547 
Fax 410-955-0185 
Email:  ddorr1@jhmi.edu 
 
  
9.1 
Data and Safety Monitoring  
 
At the Sidney Kimmel Comprehensive Cancer Center (SKCCC) at Johns Hopkins, 
the Associate Director for Clinical Research, Clinical Research Review Committee 
(CRC), SKCCC Safety Monitoring Committee (SMC), CRO Quality Assurance 
Group, and the PI share monitoring responsibilities.    
 
9.1.1  
Internal Data Monitoring  
 
The PI at each site will review data to assure the validity of data, as well as, 
the safety of the subjects. The PI will also monitor the progress of the trial.  The 
PI will review safety reports and clinical trial efficacy endpoints and to confirm 
that the safety outcomes favor continuation of the study. 
 
The PI at each site will be responsible for maintaining the clinical protocol, 
reporting adverse events, assuring that consent is obtained and documented, 
reporting of unexpected outcomes, and reporting the status of the trial in the 
continuing renewal report submitted to the IRB and to the trial monitoring 


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Page 26 of 37 
review group. Content of the continuing renewal report at a minimum should 
include year-to-date and full trial data on: accrual and eligibility, protocol 
compliance, treatment administration, toxicity and ADR reports, response, 
survival, regulatory compliance, compliance with prearranged statistical goals.  
The report should be submitted in a timely manner according to the schedule 
defined by Johns Hopkins Medicine Institutional Review Board.  
 
Adverse Event reporting – Serious Adverse Events that will be reported 
should include: any death within the first 100 days post BMT, any graft failures 
associated with failure of neutrophil recovery to >500/mm³ by day ~60 after 
transplantation, and any unexpected events as deemed significant by the PI.  
The PI will be responsible for reporting events to their local CRO/IRB, as well 
as to the primary study site and other investigators. 
 
An external Data Safety and Monitoring Board (DSMB), comprised of three 
independent external experts, will convene as requested by the PI to review 
serious toxicities and adverse events for the purpose of determining whether 
the trial should be modified or stopped. Triggers for referral to the DSMB are 
described in the Stopping Rules Criteria of section 10.0.  Each site where the 
trial is being conducted will have its own local DSMB. 
Monthly conference calls between the coordinating center and outside site, and 
in-person meetings every 6 months, will be conducted to review enrollment, 
data, and any safety issues, and to discuss study progress and the need for 
any protocol amendments. 
 
 
9.1.2  
External Data Monitoring and Auditing  
 
This is a Level I study under the SKCCC Data and Safety Monitoring Plan.  The 
SKCCC Clinical Research Office, Quality Assurance Group will perform 
periodic study audits.  All trial monitoring and reporting will also be reviewed 
annually by the SKCCC Safety Monitoring Committee. 
 
9.1.3  
Safety Monitoring  
 
The SKCCC Safety Monitoring Committee (SMC) performs an annual review 
of this Level I study. 
 
The SMC is charged with ensuring the safety of participants and the validity 
and integrity of the data and the appropriate closure of studies for which 
significant benefits or risks have been uncovered.  The Committee is 
responsible for continuous, ongoing review of the conduct of the trial, including 
adherence to study design, documentation of appropriate monitoring, and 
proper reporting of protocol problems and events.  Inherent in this process is 
the goal of enhancing the quality of the research by providing the investigator 
with constructive criticism.  The SMC membership includes physicians and 
other representatives from various Center Programs, biostatistics, data 
management, nursing, and quality assurance.   
 
 
10.0 
STATISTICAL CONSIDERATIONS 
Overall Study Design 


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This is a single-arm study of non-myeloablative conditioning and post transplantation 
cyclophosphamide with partially HLA-mismatched donors in patients with SAA.  Without 
BMT, the one year survival for refractory SAA is very poor.  Seventy percent of 
Caucasians and 10% of African Americans and patients of mixed racial backgrounds will 
have a partially HLA-mismatched donor.  In children and young adults with this type of 
transplant the one year survival is at best 70% and in other series as low as 40%.  In this 
trial, we hope to improve survival, but even if survival was unchanged we would 
markedly advance the field by expanding the donor pool for these patients.    
 
The primary goal of this study is to determine whether this type of transplantation for 
SAA is feasible and safe.  There will be continuous monitoring for feasibility and for 
safety.  Feasibility will have been met with these benchmarks: the patient has the 
transplant, is assessed for the safety endpoint, and survives one year.  The safety 
monitoring plan is included to monitor graft failure (day 60), grade 2-4 acute graft versus 
host disease (day100), 6 month mortality (day 180), and chronic graft versus host 
disease (day 180). 
 
10.1 Primary Objective 
Our primary objective is to determine if it is feasible for SAA patients to be transplanted 
using non-myeloablative conditioning and post transplantation cyclophosphamide with 
partially HLA-mismatched donors.  
 
10.2 Secondary Objectives 
10.3.1 To estimate overall survival at one year. 
10.3.2 To estimate full donor chimerism by day 60 
10.3.3 To estimate the cumulative incidence of non–relapse-related mortality following 
transplant. 
10.3.4 To estimate the incidences of primary and secondary graft failure following 
transplant. 
10.3.5 To estimate the cumulative incidences of grade II-IV and grade III-IV acute graft 
versus-host disease (GVHD).  
10.3.6 To estimate the cumulative incidence of chronic graft versus-host disease 
(GVHD).  
10.3.7 To estimate the cumulative incidence of ANC and platelet recovery.  
10.3.8 To estimate GVHD free relapse free survival (GRFS). 
10.3.9 To summarize major transplant related toxicities and to estimate transplant 
related mortality (TRM). 
 
10.3 Early stopping guideline for feasibility 
Feasibility of non-myeloablative, partially HLA-mismatched transplantation in SAA will be 
based on three benchmarks: the patient has the transplant, is assessed for the safety 
endpoint, and survives one year.  We will use a probability-based decision rule for the 
study to decide if the probability of successfully proceeding through the study is 
convincingly less than 0.70.  We expect, a priori, the feasibility to be high and that 80% 
of patients will meet the feasibility benchmarks.  The monitoring rule will therefore use an 
a priori optimistic Beta(8,2) prior distribution.  This distribution corresponds to an 
assumption that 8 out of 10 patients will proceed successfully through the study as 
planned and 90% certainty that feasibility is between .57 and .96.  This stopping rule will 
hold enrollment if, given the data, there is at least 90% probability that fewer than 70% of 
patients are meeting the feasibility criterion. 
 


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Page 28 of 37 
The table below gives the numbers of patients out of the number on study that would 
cause the study to be reviewed for feasibility.  For example, If only two of the first nine 
patients are able to meet feasibility criterion and complete the study as expected, the 
study would be paused. 
 
Stop 
if 
number 
of 
patients 
successful  
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
in N patients
5
7
9
10
12
13
15
16
18
20
 
Operating characteristics for feasibility:  The operating characteristics of this 
feasibility rule have been calculated based on 5000 simulations.  If the posterior certainty 
that feasibility is 70% or less, based on Bayes rule and the assumption of a Beta(8.2) 
prior, is 90% or higher (≥ 9:1 odds against the patients proceeding through the study as 
planned), further study will be reconsidered.  For data simulated with known probabilities 
of feasibility (θ), the table shows the percent of time that the feasibility rule will determine 
that the underlying proportion of patients who continue successfully through is below 
70%. 
 
True feasibility 
(θ)
0.30 
0.40 
0.50 
0.60 
0.70 
0.75 
0.80 
0.85 
% studies 
stopped
96.5
%
82.8
%
51.0
%
20.3
%
3.5%
1.2%
0.2%
0.1%
 
10.4 Sample size and accrual 
We estimate two to three years of accrual to enroll 20 patients.   At the end of the study, 
according to the stopping rule, at least 10 patients will have met the feasibility 
benchmarks for the study to complete without a pause.  We would probably not 
recommend this treatment if, given the data, there is a 90% probability that fewer than 
70% of patients could continue successfully through the study.  If 11 out of 20 patients 
fail to complete the study successfully, there would be less than 7% probability that the 
underlying feasibility is 70% or higher. 
 
10.5 Stopping Rules for safety 
Mortality, graft failure, and acute and chronic graft versus host disease will be 
monitored after every patient.  We will use probability-based decision rules with 
assumptions outlined in the table below.   Independence was assumed between the 
four types of toxicities to establish the stopping boundaries.  Historical probabilities of 
these events were obtained from Bacigalupo1 and Eapen2.  The prior probabilities for 
this study are modeled by beta distributions which have means that correspond to 
the historical estimates.  The spread of the prior distributions is described in column 
5 as intervals over which we are 90% certain that the estimates of the historical 
means are within.   The last two columns give thresholds for our stopping rule and 
the certainty with which we must be that the threshold is exceeded before we pause 
for a review of safety.  For all of the endpoints, if there are 3:1 odds that the safety 
threshold has been exceeded, the study will be paused for a review.      
 


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Page 29 of 37 
Endpo
int 
Tim
e 
(da
y) 
Ref
.* 
Prior 
Spread 
of prior 
(90% 
certainty
)
Thresh
old 
Certainty 
required 
for 
stopping 
Graft 
fail
60 
17
%
Beta(2,
10)
3.3 - 
36.4%
25% 
75% 
aGVH
D
100 
31
%
Beta(3,
6.5)
10.4 -
57.4%
35% 
75% 
Mortal
ity
180 
20
%
Beta(2,
8)
4.1 – 
42.9%
25% 
75% 
cGVH
D
180 
50
%
Beta(5,
5)
25.1 – 
74.9%
50% 
75% 
 
* Historical estimates taken from references.  
 
 
Stopping rule for graft failure: 
Stop if GF
4
5
6
7
8
and N 
patients
4-6 
7-9 
10-13 
14-17 
18-20 
 
Stopping rule for acute graft versus host disease: 
Stop if 
aGVHD
3 
4 
5 
6 
7 
8 
9 
10 
and N 
patients
3-
4
5-
6
7-
9
10-
11
12-
14
15-
16
17-
19
20 
 
Stopping rule for death: 
Stop if 
death
3 
4 
5 
6 
7 
8 
and N 
patients
3-4 
5-8 
9-11 
12-15 
16-19 
20 
 
Stopping rule for chronic graft versus host disease: 
Stop if 
cGVHD
3 
4 
5 
6 
7 
8 
and N 
patients
3 
4-5 
6-7 
8-9 
10 
11-12 
 
Stop if 
cGVHD
9 
10 
11 
12 
and N 
patients
13-14 
15-16 
17-18 
19-20 
 


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Operating characteristics of safety stopping rule: 
We have assumed independence for establishing our safety monitoring stopping rules.  
Operating characteristics for each event, assuming the others do not censor or otherwise 
mask the event, are given below.  In practice, these endpoints are not independent and 
the study stopping criterion could be anti-conservative.  While this is a limitation, the lack 
of preliminary data precluded realistic simulations based on cumulative incidence or 
multinomial probabilities.   
 
True event 
prob. (θ)
0.10
.15 
.20 
.25 
.30 
.35 
.40 
.50 
.55 
.60 
Graft failure 
stopped (%)
0.4 
1.7 
6.5 
15.7
31.7
48.2
64.7
90.0
96.1
98.4
aGVHD 
stopped  (%)
0.4 
2.0 
5.2 
9.9 
19.8
32.9
48.9
77.1
88.0
94.1
Death 
Stopped (%)
0.9 
4.3 
12.3
26.7
43.1
59.3
75.5
93.4
97.2
99.2
cGVHD 
Stopped (%)
0.1 
0.6 
1.6 
3.4 
6.6 
12.5
19.5
44.5
58.8
74.1
 
10.6 Analysis of Primary Endpoint 
10.6.1
The following table shows the 90% credible intervals for the underlying 
probability of feasibility, based on different numbers of patients 
successfully meeting the feasibility benchmarks, using a Beta(8,2) prior.  
The analysis plan is two-sided (5% in each tail), allowing for the full 
range of possible outcomes, while sample size is based on a one-sided 
consideration (10% in the upper tail).  As an example of the final 
inference, if 17 out of 20 patients meet the feasibility criterion, we will be 
fairly confident that feasibility is 70% or higher.
 
 
 
 
 
Outcome and 90% Credible 
Interval
Outcome and 90% Credible 
Interval
 10 out of 20        (45.1, 74.1)
16 out of 20        (67.1, 90.6)  
 11 out of 20        (48.6, 77.1)
17 out of 20        (71.2, 93.0)
 12 out of 20        (52.1, 80.0)
18 out of 20        (75.4, 95.1)
 13 out of 20        (55.7, 82.8)
19 out of 20        (79.8, 97.1)
 14 out of 20        (59.4, 85.5)
20 out of 20        (87.2, 100.0)
 15 out of 20        (63.2, 88.1)
 
10.7 Analysis of Secondary Endpoints 
10.7.1
Standard life table methods will be used to report OS.  We will report 
the six-month, one, and two year OS.  
10.7.2
We will estimate full donor chimerism by day 60 with an exact 90% 
binomial confidence interval.


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10.7.3
NRM: To estimate the cumulative incidence of non–relapse-related 
mortality following transplant, a cumulative incidence curve will be 
produced. Incidence of NRM will be estimated at 60 days, 100 days, 
six months, and one year along with 90% confidence intervals.  
Relapse or death will be considered as competing events.
10.7.4
Graft failure: To estimate the day 60 and overall incidence of primary 
and secondary graft failure following transplant.  Exact binomial 90% 
confidence intervals will be reported.
10.7.5
Acute GVHD: To estimate the cumulative incidence of grade II-IV and 
grade III-IV acute GVHD from day of transplant.  The first day of acute 
GVHD onset for a given grade will be used to estimate the cumulative 
incidence curves.  Overall cumulative incidence will be estimated along 
with a 90% confidence interval as well as at 100 days post-transplant.  
Graft failure, disease progression or death prior to occurrence of acute 
GVHD will be considered as competing events.
10.7.6
Chronic GVHD: To estimate the cumulative incidence and severity of 
chronic GVHD from day of transplant, the first day of clinical onset of 
chronic GVHD will be used to estimate a cumulative incidence curve. 
Incidences of chronic GVHD at one and two years post-transplant will 
be estimated along with 90% confidence intervals. Death, disease 
progression, or graft failure prior to occurrence of chronic GVHD will be 
considered as competing events.
10.7.7
The cumulative incidence of ANC and platelet recovery will be 
reported.  Death before count recovery will be considered a competing 
event.  Platelet recovery to both 20K and 50K will be reported. 
10.7.8
GRFS is defined as the interval from Day 0 to date of first grade 3-4 
aGVHD, or chronic GVHD, or relapse, death from any cause, or last 
patient evaluation. Patients without grade 3-4 aGVHD, chronic GVHD, 
or who have not progressed or died will be censored at the last date 
they were assessed.  Standard life table methods will be used to report 
GRFS.  We will report the six-month, one, and two year GRFS.  
10.7.9
Transplant related toxicities will be summarized descriptively with 
proportions and exact binomial confidence intervals.  TRM: Estimate 
the probability of death due to causes unrelated to the underlying 
disease.  Standard life table methods will be used to report TRM. 
 
 
 
 
References: 
[1] Bacigalupo, A., Socie’, G., Lanino, E., Prete, A., Locatelli, F., Locasciulli, A., Cesaro, 
S.,  Shimoni, A., Marsh, J., Brune, M., Van Lint, M.,  Oneto, R., and Passweg, J.  
Fludarabine, cyclophosphamide, antithymocyte globulin, with or without low dose total 
body irradiation, for alternative donor transplants, in acquired severe aplastic anemia: a 
retrospective study from the EBMT-SAA working party.  Haematologica, 95(6): 976–982, 
2010. 
 


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[2] Eapen, M., Le Rademacher, J., Antin, J.H., Champlin, R.E., Carreras, J., Fay, 
J., Passweg, J.R., Tolar, J., Horowitz, M.M., Marsh, J.C., Deeg, H.J.   Effect of stem cell 
source on outcomes after unrelated donor transplantation in severe aplastic anemia.  
Blood, 118(9):2618-21, 2011.  
  
 
 
 
 
11.0 
RISKS AND BENEFITS 
 
11.1 
Risks and toxicity 
 
The major toxicity of using bone marrow from HLA-mismatched donors is GVHD.  
Using non-myeloablative conditioning regimens and stem cell products from 
peripheral blood, we would expect the incidence of GVHD in this study to be in the 
40-50% range using unmanipulated bone marrow as the source of stem cells.   
 
Another significant risk is failure-to-engraft due to rejection by host lymphocytes.  
However, because of the non-myeloablative nature of the conditioning regimen we 
would expect patients to have full autologous, hematologic recovery.   
 
Infection is a major cause of morbidity and mortality in the peri-transplant period 
(<100d post-BMT).  However, given current supportive care and the intensive 
infection prophylaxis of this protocol, we expect the risk to be acceptable.  
Prolonged neutropenia may increase this risk in the case of graft rejection, 
however. 
 
Other risks that may be associated with fludarabine chemotherapy include 
prolonged immunosuppression of T-lymphocytes increasing the incidence of PCP 
and viral infections.  The extent of this risk is unclear at present.  Patients will 
receive appropriate PCP prophylaxis and will be monitored carefully for evidence 
of infection by viruses such as CMV, BK and adenovirus.  Major risks associated 
with cyclophosphamide chemotherapy include hemorrhagic cystitis and 
congestive heart failure.  
 
Relapse of the underlying disease also may occur.   
 
11.2 
Benefits 
 
The potential benefits of this trial are prolongation of overall survival. 
 
12.0 
INFORMED CONSENT 
 
Patients eligible for marrow grafting are completely evaluated and then presented 
and approved for transplant at the Bone Marrow Transplant group conference.  
The group's recommendations are discussed with the patient.  If the patient is 
approved for BMT, the marrow processing procedure itself, the risks of the 
preparative regimen, risks of BMT complications including infection and GVHD and 
alternate forms of therapy are presented as objectively as possible.  For pediatric 


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Page 33 of 37 
patients (<18 yr of age) assent is obtained from the patient and informed consent 
is obtained from all parents.  Informed consent is obtained  from 
the 
recipient 
using the forms approved by the IRB.          
 
13.0 
ON-STUDY DATE:  
 
 
Date of consent signing.  
 
 
14.0     OFF-STUDY DATE: 
 
Upon completion of “Day 60” evaluations, patients have completed their treatment 
except for tacrolimus, which continues until day 365. Patient follow-up beyond day 
60 will consist of collecting information regarding ongoing engraftment, disease 
status, late effects of this protocol, acute and chronic graft-vs-host disease, 
immune reconstitution, additional therapies, and survival. Patients will go off study 
early in the event of:  
 
1. Death  
2. Patient decision (or decision by a parent or guardian on behalf of a minor) 
There will be long term follow up, regardless of disease status, to all patients for a 
minimum of two years. 
 
An eligibility form must be completed for every subject and must be kept in the research 
chart. 
 


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Appendix 1.  Consensus conference clinical grading of acute GVHD 
 
Clinical Staging 
Stage 
Skin 
Liver: 
Total 
Bilirubin
Intestinal Tract: Diarrhea 
0 
No rash 
<2.0 mg/dL 
<500 ml/day 
1
<25% of skin surface
2.0-3.0
500-1000 ml/day
2
25-50%
3.1-6.0
1001-1500 ml/day
3 
Erythroderma 
6.1-15.0 
>1500 ml/day 
4 
Erythroderma with bullae 
and desquamation
>15.0 
Severe abdominal pain with 
or without ileus
 
Clinical Grading 
Grade
Skin*
Liver
GI
I 
1-2 
0 
0 
II
3
1
1
III 
- 
2-3 
2-4 
IV
4
4
-
*Each column identifies minimum stage for organ grade 
 
 


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Appendix 2.  Ideal Body Weight and Adjusted Ideal Body Weight Calculations 
 
 
Ideal Body Weight Formula 
 
Males: 50 kg + (2.3 x the number of inches > 5 feet) 
Females: 45 kg + (2.3 x the number of inches > 5 feet) 
 
 
Adjusted Ideal Body Weight Formula 
 
[(actual weight – ideal weight) x 25%] + ideal weight 
 
Note:  If actual weight < ideal, use actual weight. 
 
If actual weight > ideal, use corrected ideal. 
 
 
 
1 
A. Bacigalupo, R. Brand, R. Oneto, B. Bruno, G. Socie, J. Passweg, A. Locasciulli, 
M. T. Van Lint, A. Tichelli, S. McCann, J. Marsh, P. Ljungman, J. Hows, P. Marin, 
and H. Schrezenmeier, 'Treatment of Acquired Severe Aplastic Anemia: Bone 
Marrow Transplantation Compared with Immunosuppressive Therapy--the 
European Group for Blood and Marrow Transplantation Experience', Semin 
Hematol, 37 (2000), 69-80. 
2 
A. Bacigalupo, R. Oneto, B. Bruno, G. Socie, J. Passweg, A. Locasciulli, M. T. Van 
Lint, A. Tichelli, S. McCann, J. Marsh, P. Ljungman, J. Hows, P. Marin, and H. 
Schrezenmeier, 'Current Results of Bone Marrow Transplantation in Patients with 
Acquired Severe Aplastic Anemia. Report of the European Group for Blood and 
Marrow Transplantation. On Behalf of the Working Party on Severe Aplastic 
Anemia of the European Group for Blood and Marrow Transplantation', Acta 
Haematol., 103 (2000), 19-25. 
3 
A. Bacigalupo, G. Socie, H. Schrezenmeier, A. Tichelli, A. Locasciulli, M. Fuehrer, 
A. M. Risitano, C. Dufour, J. R. Passweg, R. Oneto, M. Aljurf, C. Flynn, V. Mialou, 
R. M. Hamladji, J. C. Marsh, Blood Aplastic Anemia Working Party of the European 
Group for, and Transplantation Marrow, 'Bone Marrow Versus Peripheral Blood as 
the Stem Cell Source for Sibling Transplants in Acquired Aplastic Anemia: Survival 
Advantage for Bone Marrow in All Age Groups', Haematologica, 97 (2012), 1142-
8. 
4 
J. Bolanos-Meade, E. J. Fuchs, L. Luznik, S. M. Lanzkron, C. J. Gamper, R. J. 
Jones, and R. A. Brodsky, 'Hla-Haploidentical Bone Marrow Transplantation with 
Posttransplant Cyclophosphamide Expands the Donor Pool for Patients with 
Sickle Cell Disease', Blood, 120 (2012), 4285-91. 
5 
R. A. Brodsky, A. R. Chen, D. Dorr, E. J. Fuchs, C. A. Huff, L. Luznik, B. D. Smith, 
W. H. Matsui, S. N. Goodman, R. F. Ambinder, and R. J. Jones, 'High-Dose 
Cyclophosphamide for Severe Aplastic Anemia: Long-Term Follow-Up', Blood, 
115 (2010), 2136-41. 
6 
R. A. Brodsky, and R. J. Jones, 'Aplastic Anaemia', The Lancet, 365 (2005), 1647-
56. 
7 
R. A. Brodsky, L. L. Sensenbrenner, and R. J. Jones, 'Complete Remission in 
Severe Aplastic Anemia after High-Dose Cyclophosphamide without Bone Marrow 
Transplantation', Blood, 87 (1996), 491-94. 


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8 
R. E. Champlin, W. S. Perez, J. R. Passweg, J. P. Klein, B. M. Camitta, E. 
Gluckman, C. N. Bredeson, M. Eapen, and M. M. Horowitz, 'Bone Marrow 
Transplantation for Severe Aplastic Anemia: A Randomized Controlled Study of 
Conditioning Regimens', Blood, 109 (2007), 4582-85. 
9 
R. Chu, R. Brazauskas, F. Kan, A. Bashey, C. Bredeson, B. Camitta, K. Y. Chiang, 
H. Frangoul, R. P. Gale, A. Gee, B. George, F. D. Goldman, T. G. Gross, V. Gupta, 
G. A. Hale, L. Isola, A. U. Ispizua, H. Lazarus, J. Marsh, J. Russell, M. Sabloff, E. 
K. Waller, and M. Eapen, 'Comparison of Outcomes after Transplantation of G-
Csf-Stimulated Bone Marrow Grafts Versus Bone Marrow or Peripheral Blood 
Grafts from Hla-Matched Sibling Donors for Patients with Severe Aplastic Anemia', 
Biol Blood Marrow Transplant, 17 (2011), 1018-24. 
10 
A. E. Dezern, and R. A. Brodsky, 'Clinical Management of Aplastic Anemia', Expert 
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