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Version Date: December 28, 2017 Abbreviated Title: Lupron &18F FLT in allo-HSCT CC Protocol \#: 11-C-0136 NCT\# NCT01338987 Version Date: December 28, 2017 Amendment: T
Title of Study: Multi-institutional Prospective Pilot Study of Lupron to Enhance Lymphocyte Immune Reconstitution following Allogeneic Bone Marrow Transplantation in Post-pubertal Children and Adults with Molecular Imaging Evaluation  
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| NIH Principal Investigator:                                                | Christopher Kanakry, M.D. A-F Experimental Transplantation & Immunology Branch Center for Cancer Research, National Cancer Institute, National Institutes of Health 10 Center Drive, Rm. 4-3142, MSC 1203 Telephone: 240-760-6171 Email: christopher.kanakry@nih.gov   |
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| Lead Associate Investigator:                                               | Ronald Gress, M.D. A-F Branch Chief, ETIB, CCR, NCI, NIH                                                                                                                                                                                                               |
| Medical Advisory Investigator:                                             | Peter Choyke, M.D. A-F Branch Chief, Molecular Imaging, NCI, CCR, NIH                                                                                                                                                                                                  |
| NIH Associate Investigators:  Daniele Avila, CRNP, ETIB, CCR, NCI, NIH A-E | Meg Kenyon, CRNP, ETIB, CCR, NCI, NIH A-E                                                                                                                                                                                                                              |
| Stephanie Cotton, RN, OCD, CCR, NCI, NIH A-B                               | Jennifer Sadler, RN, OCD, CCR, NCI, NIH A-B                                                                                                                                                                                                                            |
| Bazetta Blacklock Schuver, RN, OCD, CCR, NCI,                              | Karen Kurdziel, MD, MIP, CCR, NCI, NIHA-F                                                                                                                                                                                                                              |
| NIH A-B Deborah Citrin, MD, ROB, CCR, NCI, NIHA-F                          | Seth M. Steinberg, PhD, BDMS, CCR, NCI, NIH B, E                                                                                                                                                                                                                       |
| Kristen Cole, RN, OCD, CCR, NCI, NIH A-B                                   | Constance Yuan, MD, PhD, LP, CC, NIH E-F                                                                                                                                                                                                                               |
| Dennis Hickstein, MD, ETIB, CCR, NCI, NIHA-F                               | Bonnie Yates, RN, POB, CCR, NCI, NIH A-B                                                                                                                                                                                                                               |
| Fran Hakim, PhD, ETIB, CCR, NCI, NIH E-F                                   | Cindy Delbrook, RN, OCD, CCR, NCI, NIH A-B                                                                                                                                                                                                                             |
| James Kochenderfer, MD, ETIB, CCR, NCI, NIHA-F                             | Terry Fry, MD, POB, CCR, NCI, NIHA-F                                                                                                                                                                                                                                   |
| Nirali Shah, MD, POB, CCR, NCI, NIHA-F                                     | Maya Lodish, MD, NICHDA-F                                                                                                                                                                                                                                              |
| Steven Z. Pavletic, MD, ETIB, CCR, NCI, NIHA-F                             | Jennifer Kanakry, MD, ETIB, CCR, NCI, NIHA-F                                                                                                                                                                                                                           |  
Version Date: December 28, 2017  
| Tracey Chinn RN, ETIB, CCR, NCI, NIH A-B     | David Stroncek, MD, DTM, CC, NIH E-F        |
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| Ellen Carroll, RN, OCD, CCR, NCI, NIH A-B    | Maria Liza Lindenberg, MD, MIP, CCR, NIHA-F |
| Brenna Hansen, RN, OCD, CCR, NCI, NIH A-B    | Willy Flegel, MD, DTM, CC, NIH E-F          |
| Jennifer Mann, CRNP, ETIB, CCR, NCI, NIH A-E |                                             |  
Site Principal Investigator: Jennifer Holter, M.D., University of Oklahoma Site Principal Investigator Kirsten Williams, MD., Children's National Medical Center, Washington Referral Contact: Bazetta Blacklock Schuver, RN Study Coordinator: Jennifer Sadler, RN
240-760-6175 Email: jennifer.sadler@nih.gov Non-NIH Associate Investigators:
Jennifer Wilder, RN., Leidos Inc. A-B
For each person listed above, roles are identified with the appropriate letter A. Obtain information by intervening or interacting with living individuals for research purposes B. Obtaining identifiable private information about living individuals C. Obtaining the voluntary informed consent of individuals to be subjects D. Makes decisions about subject eligibility E. Studying, interpreting, or analyzing identifiable private information or data/specimens for research purposes F. Studying, interpreting, or analyzing de-identified data or specimens for research purposes G. Some/all research activities performed outside NIH
IND Information: IND Number: 111119 IND Sponsor: Center for Cancer Research, NCI
Commercial Agents: Lupron Depot, Cyclophosphamide, Methotrexate, Tacrolimus, Fludarabine, Busulfan Version Date: December 28, 2017 Multi-institutional data:  
| University of Oklahoma Principal Investigator:   | Jennifer Holter, M.D. Associate Professor of Hematology/Oncology and BMT   |
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| Associate Investigator:                          | George Selby, MD                                                           |
| Institution:                                     | University of Oklahoma                                                     |
| MPA or SPA Number:                               | Federal-Wide Assurance program FWA#00003191.                               |
| Complete Address:                                | 920 Stanton L. Young, WP 2040 Oklahoma City, OK 73104                      |
| Phone Number:                                    | 405-271-4022 Ext 48397 Contact Elizabeth Cambron                           |
| E-mail:                                          | jholter2@ouhsc.edu                                                         |
| Name of IRB Contact:                             | Meg Ribaudo, Director                                                      |
| Complete Address:                                | 1000 Stanton L Young Blvd, Suite 176, Oklahoma City, OK 73117              |
| Phone Number:                                    | 405-271-2045                                                               |
| E-mail of Contact:                               | meg-ribaudo@ouhsc.edu                                                      |
| Name of Pharmacy Contact: N/A                    |                                                                            |  
Name of Pharmacy Contact: N/A  
| Children's National Medical Center (CNMC) Principal Investigator: Kirsten Williams, M.D. Assistant Professor of Pediatrics/Hematology/Oncology and BMT Institution: Children's National Medical Center MPA or SPA Number: Federal-Wide Assurance program FWA# 00004487. Complete Address: 111 Michigan Ave NW Washington DC, 20010 Phone Number: 202-476-2694 E-mail: kmwillia@cnmc.org Name of IRB Contact: Fahmida Hoq, MD Complete Address: 111 Michigan Ave NW   |
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Version Date: December 28, 2017 Washington DC, 20010 Phone Number: 202-476-5000 E-mail of Contact: FHoq@childrensnational.org Name of Pharmacy Contact: N/A
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precis

Background:
x Impaired lymphocyte immune reconstitution is associated with morbidity and mortality following allogeneic hematopoietic stem cell transplantation (HSCT).  
x Data suggest that one of the limitations of immunity after HSCT is the lack of thymus recovery and proper B cell development.  
x Androgen withdrawal has been shown to enhance T and B lymphopoiesis. x Lupron is an approved, safe, gonadotropin releasing hormone (GnRH)
agonist/antagonist.  
x Noninvasive imaging modalities to study immune reconstitution would be invaluable to predict optimal or impaired immune recovery permitting early institution of therapies.  
x FLT is 3'-deoxy-3 18F-fluorothymidine, a radiolabeled thymidine analogue that illustrates dividing hematopoietic cells and may predict immune recovery after allogeneic HSCT.  
x FLT has been used safely in patients who have received intensive chemotherapy.
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objectives

x Primary: To determine if Lupron improves B lymphocyte reconstitution after HSCT.  
x Primary: To assess whether 18F FLT PET/CT could predict early engraftment/immune reconstitution in marrow and thymus after allogeneic HSCT.  
x Primary: To assess safety of Lupron after 2nd HSCT evaluated in a separate arm.
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eligibility

x Patients > 9 years old and pubertal and/or >15 year and  55 years, with aggressive leukemia (Acute Myelogenous Leukemia (AML), myelodysplastic syndromes (MDS) with high risk cytogenetics, Acute Lymphocytic Leukemia (ALL), CMML, certain CML) requiring HSCT will be enrolled at NCI.  
x $W8QLYHUVLW\RI2NODKRPD$JH!\HDUVROGDQG\HDUVIRUUHFLSLHQW x Patients > 4 and < 24 years with the above diseases will be enrolled at Children's National Medical Center.
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design

x This is a prospective pilot study, the primary aims of which are: 1) to assess whether Lupron enhances lymphocyte recovery after HSCT and 2) whether FLT imaging can be used to predict engraftment/immune reconstitution, and 3) whether Lupron and FLT are tolerable for second HSCT.  
x At NCI and Univ of Oklahoma, post-pubertal pediatric male patients (<18 years) will be randomized to receive a 3 month (11.25 mg) injection and adult male patients will be randomized to receive 4-month preparation of Lupron (30 mg) or placebo two weeks before the preparative regimen for first BMT. Women and all individuals undergoing 2nd BMT will receive Lupron at these doses per age and be evaluated in the treated cohort. At Children's National Medical Center, the patients will not receive Lupron outside of the context of clinical care, and will receive myeloablative HSCT as per standard of care with FLT imaging for engraftment as the only primary endpoint.  
x A target of 68 evaluable adult patients will be enrolled on this trial, which may necessitate up to 118 patients (118 donors) enrolled to reach this target at NCI and University of Oklahoma. A total of 10 pediatric patients will be enrolled at Children's National Medical Center for FLT imaging only. Sixteen of these patients will be enrolled to undergo second BMT.  
x At NCI, adults greater than 18 years old both female and adult male patients undergoing 2nd BMT will receive 4-month preparation of Lupron (30 mg) two weeks before the preparative regimen. All patients will undergo FLT imaging to evaluate whether this may predict HSCT response or failure (relapse). This will be a pilot arm of 16 patients total.  
x The planned length of this trial is 7 years with interim analyses at day 100 and day 365. x Some of the patients are anticipated to be evaluated using FLT (to include only patients needed for the immunological primary endpoint, not increasing total patient numbers). 23 adult NCI patients in total will undergo FLT PET/CT imaging on day -1, at day +5 or day +9, at 4 weeks, and at a future point to include evidence of GVHD relapse, or immune recovery. An estimated 50 patients (including subset of the 23 patients undergoing serial scanning) will be imaged approximately at 1 year for evaluation of thymus reconstitution. The total possible numbers will include no more than 118 patients to achieve the 68 evaluable adults for the immunological primary endpoint. However, all NCI patients will undergo a single 1 year FLT for evaluation of thymus reconstitution. 10 pediatric patients at CNMC will undergo FLT PET/CT imaging on Day -1, day+9, and day +28 (if possible). Initial images will be correlated with engraftment and other secondary endpoints.  
x Study endpoints to include: 1) safety of Lupron in the context of allogeneic BMT, 2)
lymphocyte reconstitution after Lupron administration, 3) the incidence of acute and chronic GVHD and infectious complications, 4) remission rates after HSCT.  
| TABLE OF CONTENTS                                                                                     |                                                                                                                                                       |
|-------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------|
| PRÉCIS .                                                                                              |                                                                                                                                                       |
| TABLE OF CONTENTS.                                                                                    |                                                                                                                                                       |
| l                                                                                                     | INTRODUCTION .                                                                                                                                        |
| Study Objectives .                                                                                    |                                                                                                                                                       |
| 1.1                                                                                                   | Primary Objectives                                                                                                                                    |
| 1.1.1                                                                                                 |                                                                                                                                                       |
| 1.1.2                                                                                                 | Secondary Objectives                                                                                                                                  |
| 1.2                                                                                                   | Background and Rationale .                                                                                                                            |
| 1.2.1                                                                                                 | Impaired Immune Reconstitution after Stem Cell Transplantation ......................... 15                                                           |
| 1.2.2                                                                                                 | Androgen Withdrawal Enhances T and B Lymphopoiesis                                                                                                    |
| 1.2.3                                                                                                 | Lupron for Androgen Suppression                                                                                                                       |
| 1.2.4                                                                                                 | Lupron Safety Profile                                                                                                                                 |
| Lupron Pharmacology                                                                                   |                                                                                                                                                       |
| 1.2.5                                                                                                 |                                                                                                                                                       |
| 1.2.6                                                                                                 | FLT imaging modality ............................................................................................................................. 19 |
| 1.2.7                                                                                                 | 18F FLT Safety Profile .                                                                                                                              |
| 1.2.8                                                                                                 | 18F FLT Pharmacology .                                                                                                                                |
| 1.2.9                                                                                                 | FLT Availability .                                                                                                                                    |
| 1.2.10                                                                                                | T Trial Plan                                                                                                                                          |
| 2                                                                                                     | ELIGIBILITY ASSESSMENT AND ENROLLMENT.                                                                                                                |
| Eligibility Criteria .                                                                                |                                                                                                                                                       |
| 2.1                                                                                                   |                                                                                                                                                       |
| 2.1.1                                                                                                 | Inclusion Criteria: Transplant Recipient                                                                                                              |
| 2.1.2                                                                                                 | Exclusion Criteria: Transplant Recipient                                                                                                              |
| 2.1.3                                                                                                 | Inclusion Criteria: Matched Related Transplant Donor                                                                                                  |
| Exclusion Criteria: Matched Related Transplant Donor.............................................. 26 |                                                                                                                                                       |
| 2.1.4                                                                                                 |                                                                                                                                                       |
| 2.1.5                                                                                                 | Inclusion Criteria- Matched Unrelated Donor ...................................................................... 27                                 |
| 2.1.6                                                                                                 | Inclusion Criteria- 18F FLT Candidate Transplant Recipient ..................................... 27                                                   |
| 2.1.7                                                                                                 | Exclusion Criteria- 18F FLT Candidate Transplant Recipient ................................... 27                                                     |
| Screening Evaluation and Criteria for HSCT .                                                          |                                                                                                                                                       |
| 2.2                                                                                                   |                                                                                                                                                       |
| 2.2.1                                                                                                 | Recipient                                                                                                                                             |
| 2.2.2                                                                                                 | Matched Related Donor                                                                                                                                 |
| Registration Procedures                                                                               |                                                                                                                                                       |
| 2.3                                                                                                   |                                                                                                                                                       |
| 2.3.1                                                                                                 | Protocol Entry Date.                                                                                                                                  |  
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| Version Date: December 28, 2017                                                                                                    |                                                                                                                      |
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| Registration .                                                                                                                     |                                                                                                                      |
| 2.3.2                                                                                                                              |                                                                                                                      |
| 2.3.3                                                                                                                              | University of Oklahoma Donors and Recipients .............................................................. 30       |
| 2.3.4                                                                                                                              | Children's National Medical Center Donors and Recipients                                                             |
| 2.4                                                                                                                                | Randomization Procedures .                                                                                           |
| 2.4.1                                                                                                                              | Female Recipients                                                                                                    |
| 2.4.2                                                                                                                              | Male Recipients                                                                                                      |
| 3                                                                                                                                  | STUDY IMPLEMENTATION .                                                                                               |
| Study Design                                                                                                                       |                                                                                                                      |
| 3.1                                                                                                                                | NCI and University of Oklahoma for 1 st BMT ..................................................................... 32 |
| 3.1.1                                                                                                                              |                                                                                                                      |
| 3.1.2                                                                                                                              | Children's National Medical Center .                                                                                 |
| 3.2                                                                                                                                | D Drug Administration                                                                                                |
| 3.2.1                                                                                                                              | LUPRON                                                                                                               |
| 3.2.2                                                                                                                              | 18 F-FLT .                                                                                                           |
| 3.3                                                                                                                                | On Study Protocol Evaluation Pre-Transplant Work-up ........................................................ 34      |
| Within 4 weeks of Transplant Date ............................................................................................. 34 |                                                                                                                      |
| 3.3.1                                                                                                                              |                                                                                                                      |
| 3.3.2                                                                                                                              | FLT on Study Evaluations .                                                                                           |
| 3.4                                                                                                                                | Biologic Studies                                                                                                     |
| Immune Reconstitution                                                                                                              |                                                                                                                      |
| 3.4.1                                                                                                                              | Immunologic Studies                                                                                                  |
| 3.4.2                                                                                                                              |                                                                                                                      |
| 3.4.3                                                                                                                              | Objectives/Specific Aims                                                                                             |
| 3.4.4                                                                                                                              | Methodology                                                                                                          |
| Collaborative research Studies                                                                                                     |                                                                                                                      |
| 3.5                                                                                                                                | Donor Collection                                                                                                     |
| 3.6                                                                                                                                |                                                                                                                      |
| 3.6.1                                                                                                                              | Matched Unrelated Donors .                                                                                           |
| MMATCHED Related Donors                                                                                                            |                                                                                                                      |
| 3.6.2                                                                                                                              |                                                                                                                      |
| 3.7                                                                                                                                | Preparative Regimen for HSCT and GVHD Prophylaxis                                                                    |
| 3.7.1                                                                                                                              | C Contral Access .                                                                                                   |
| 3.7.2                                                                                                                              | Remission Status                                                                                                     |
| 3.7.3                                                                                                                              | Conditioning Regimen .                                                                                               |
| 3.7.4                                                                                                                              | GVHD Prophylaxis                                                                                                     |
| 3.7.5                                                                                                                              | Growth Factors (GCSF)                                                                                                |
| 3.8                                                                                                                                | Transplant Procedure for 1 st and 2 nd BMT .                                                                         |  
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| Version Date: December 28, 2017 3.8.1 Day 0 42 3.8.2 Transplant Outcome Determinations 42 3.9 Post-transplant monitoring (1st and 2nd BMT) 42 3.9.1 Day 0 to day +100 42 3.9.2 Specific evaluation time points (1st and 2nd BMT) 43 3.9.3 Long term follow-up guidelines (1st and 2nd BMT) 43 3.9.4 FLT on study evaluations 44 3.10 Criteria for Removal from Protocol Therapy and Off Study Criteria 44 3.10.1 Criteria for Removal from Protocol Therapy 44 3.10.2 Off-Study Criteria 45 3.10.3 Off Protocol Therapy and Off Study Procedure at NCI 45 3.10.4 Participating Site Off-Study Notification 45 4 SUPPORTIVE CARE 46 4.1 Infection Prophylaxis 46 4.1.1 Infectious Disease Surveillance: 47 4.2 Management of Engraftment Syndrome 47 4.3 Treatment of Graft-Versus-Host Disease 47 4.4 Menses Suppression and Contraception 48 4.5 Blood Product Support 48 4.6 Nutritional Support 49 4.7 Anti-Emetic Usage 49 4.8 Intravenous Immune Globulin (IVIG) 49 4.9 Hepatic Function Support 49 4.10 Central Nervous System Prophylaxis 50 4.11 Fluid Management During First 30 Days Following Transplantation 50 4.12 Ancillary Support 50 4.13 Supportive Care For Relapse 50 4.14 Additional cell infusions for graft failure, impending graft failure, or poor graft  function 50 5 DATA COLLECTION AND EVALUATION 50 5.1 Data Collection 50 5.1.1 Adverse event recording 52   |
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| Response Criteria                                                                          |                                                                                                                          |                              |
| 5.2                                                                                        |                                                                                                                          |                              |
| 5.2.1                                                                                      | Primary Endpoints                                                                                                        |                              |
| 5.2.2                                                                                      | Secondary Endpoints                                                                                                      |                              |
| Toxicity Criteria                                                                          |                                                                                                                          |                              |
| 5.3                                                                                        |                                                                                                                          |                              |
| STATISTICAL SECTION                                                                        |                                                                                                                          |                              |
| Subject Accrual                                                                            |                                                                                                                          |                              |
| 6.1                                                                                        | 6.1.1                                                                                                                    | Statistics and Feasibility . |
| HUMAN SUBJECTS PROTECTIONS .                                                               |                                                                                                                          |                              |
| RATIONALE For Subject Selection                                                            |                                                                                                                          |                              |
| 7.1                                                                                        |                                                                                                                          |                              |
| 7.2                                                                                        | Participation of Children                                                                                                |                              |
| 7.3                                                                                        | Participation of Subjects Unable to Give Consent                                                                         |                              |
| Evaluation of Benefits and Risks/Discomforts .                                             |                                                                                                                          |                              |
| 7.4                                                                                        |                                                                                                                          |                              |
| 7.4.1                                                                                      | Recipient                                                                                                                |                              |
| 7.4.2                                                                                      | Risks in relation to benefit for Donor .                                                                                 |                              |
| Pediatric Patients                                                                         |                                                                                                                          |                              |
| 7.4.3                                                                                      |                                                                                                                          |                              |
| Consent and Assent Process and Documentation .                                             |                                                                                                                          |                              |
| 7.5                                                                                        |                                                                                                                          |                              |
| 7.5.1                                                                                      | Consent for minors when they reach the age of majority .............................................. 59                 |                              |
| 7.5.2                                                                                      | Telephone consent and re-consent procedure .......................................................................... 60 |                              |
| 7.5.3                                                                                      | Informed consent of Spanish speaking subjects ................................................................ 60        |                              |
| Short form consent process for other non-English speaking patients..................... 60 |                                                                                                                          |                              |
| 7.5.4                                                                                      |                                                                                                                          |                              |
| SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING PLAN                              |                                                                                                                          |                              |
| 61                                                                                         |                                                                                                                          |                              |
| 8.1                                                                                        | DEFINITIONS                                                                                                              |                              |
| A Adverse Event                                                                            |                                                                                                                          |                              |
| 8.1.1                                                                                      |                                                                                                                          |                              |
| 8.1.2                                                                                      | Suspected adverse reaction .                                                                                             |                              |
| 8.1.3                                                                                      | Unexpected adverse reaction                                                                                              |                              |
| 8.1.4                                                                                      | Serious                                                                                                                  |                              |
| Serious Adverse Event                                                                      |                                                                                                                          |                              |
| 8.1.5                                                                                      |                                                                                                                          |                              |
| 8.1.6                                                                                      | Disability                                                                                                               |                              |
| 8.1.7                                                                                      | Life-threatening adverse drug experience                                                                                 |                              |
| Protocol Deviation (NIH Definition)                                                        |                                                                                                                          |                              |
| 8.1.8                                                                                      |                                                                                                                          |                              |
| 8.1.9                                                                                      | Non-compliance (NIH Definition)                                                                                          |                              |  
6 7 8  
8.1.10    Unanticipated Problem.  
8.2 NCI-IRB and Clinical Director Reporting 8.2.1 NCI-IRB and NCI CD Expedited Reporting of Unanticipated Problems and Deaths  
| 62                                                                                                                                    |                                                                                                                               |
|---------------------------------------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------|
| 8.2.2                                                                                                                                 | NCI-IRB Requirements for PI Reporting at Continuing Review ............................. 63                                   |
| 8.2.3                                                                                                                                 | NCI-IRB Reporting of IND Safety Reports............................................................................ 63        |
| 8.3                                                                                                                                   | NCI Guidance for Reporting Expedited Adverse Events for Multi-Center Trials ....... 64                                        |
| 8.4                                                                                                                                   | IND Sponsor Reporting Criteria .                                                                                              |
| 8.5                                                                                                                                   | Data Recording of Unrelated Donors from NMDP .................................................................... 64          |
| DATA AND SAFETY MONITORING Plan                                                                                                       |                                                                                                                               |
| 8.6                                                                                                                                   |                                                                                                                               |
| 8.6.1                                                                                                                                 | Principal Investigator/Research Team ..................................................................................... 64 |
| 8.6.2                                                                                                                                 | Sponsor Monitoring Plan                                                                                                       |
| Safety Monitoring Committee (SMC) ................................................................................................ 65 |                                                                                                                               |
| 8.6.3                                                                                                                                 |                                                                                                                               |
| 9                                                                                                                                     | Multi-Institutional Guidelines                                                                                                |
| 9.1.1                                                                                                                                 | IRB Approvals                                                                                                                 |
| 9.1.2                                                                                                                                 | Amendments and Consents                                                                                                       |
| 9.1.3                                                                                                                                 | Data and Specimen Collection Procedures                                                                                       |
| DATA Center Audits                                                                                                                    |                                                                                                                               |
| 9.1.4                                                                                                                                 |                                                                                                                               |
| 9.1.5                                                                                                                                 | Protocol Monitoring Committees .                                                                                              |
| PHARMACEUTICAL INFORMATION                                                                                                            |                                                                                                                               |
| 10                                                                                                                                    | Lupron depot                                                                                                                  |
| 10.1                                                                                                                                  |                                                                                                                               |
| 10.1.1                                                                                                                                | C Chemical name                                                                                                               |
| 10.1.2                                                                                                                                | C Chemical structure .                                                                                                        |
| Commercial Formulation .                                                                                                              |                                                                                                                               |
| 10.1.3                                                                                                                                |                                                                                                                               |
| 10.1.4                                                                                                                                | S Storage                                                                                                                     |
| 10.1.5                                                                                                                                | Stability                                                                                                                     |
| 10.1.6                                                                                                                                | Route of administration                                                                                                       |
| Dose .                                                                                                                                |                                                                                                                               |
| 10.1.7                                                                                                                                |                                                                                                                               |
| 10.1.8                                                                                                                                | Drug Interactions                                                                                                             |
| 10.1.9                                                                                                                                | Known toxicities for patients >18                                                                                             |
| 10.1.10  Known toxicities for patients <18                                                                                            |                                                                                                                               |
| 10.2                                                                                                                                  | Cyclophosphamide                                                                                                              |
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| C Chemical name                    |                                            |
| 10.2.1                             |                                            |
| 10.2.2                             | C Chemical structure .                     |
| 10.2.3                             | Commercial Formulation .                   |
| 10.2.4                             | S Storage                                  |
| 10.2.5                             | Stability                                  |
| 10.2.6                             | Route of administration                    |
| 10.2.7                             | Dose .                                     |
| D Drug Interactions                |                                            |
| 10.2.8                             |                                            |
| 10.2.9                             | Known toxicities .                         |
| 10.3                               | Total Body Irradiation                     |
| CC                                 |                                            |
| 10.3.1                             |                                            |
| 10.3.2                             | Known toxicities .                         |
| 10.4                               | FLT                                        |
| 10.4.1                             | CC                                         |
| Commercial Formulation .           |                                            |
| 10.4.2                             | S Storage                                  |
| 10.4.3                             |                                            |
| 10.4.4                             | CC                                         |
| Method of Administration           |                                            |
| 10.4.5                             |                                            |
| 10.4.6                             | Availability .                             |
| 10.4.7                             | Known Toxicites                            |
| 10.5                               | Fludarabine (Fludara, Berlex Laboratories) |
| CC                                 |                                            |
| 10.5.1                             |                                            |
| 10.5.2                             | SUPPLY .                                   |
| 10.5.3                             | Preparation                                |
| 10.5.4                             | Storage and Stability                      |
| Administration .                   |                                            |
| 10.5.5                             |                                            |
| 10.5.6                             | TOXICITIES                                 |
| 10.6                               | Busulfan (Busulfex)                        |
| 10.6.1                             | SOURCE .                                   |
| 10.6.2                             | Storage and Stability                      |
| 10.6.3                             | Administration .                           |
| 10.6.4                             | TOXICITIES                                 |  
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version date december 28 2017

| Version Date: December 28, 2017 10.6.5 Drug Interactions 73 11 REFERENCES 74 12 APPENDICES 78 12.1 Appendix 1: Performance Scales 78 12.2 Appendix 2A: Recipient (arm 1 and 2) 80 12.3 Appendix 2B: Donor (arm 1 and 2) 82 12.4 Appendix 3a: FLT ( 1st BMT) 83 12.5 Appendix 4A: Adult (>22 years) BMT Roadmap 85 12.6 Appendix 4B: Pediatric (<22 years) BMT Roadmap 89 12.7 Appendix 5: Research Specimens 93 12.8 Appendix 6: Immunization Post-Transplant 94 12.9 Appendix 7: Management of Engraftment Syndrome 95 12.10 Appendix 8A: Grading and Management of Acute Graft-Versus-Host-Disease 97 12.11 Appendix 8B 99 12.12 Appendix 8C: Signs and Symptoms of cGVHD 102 12.13 Appendix 8D: cGVHD Score Sheet 104 12.14 Appendix 9: ETIB Policy for Sample Handling 107 12.14.1 Sample Processing 107 12.14.2 Labeling 107 12.14.3 Storage/Tracking in the Preclinical Development and Clinical Monitoring Facility  (PDCMF) 107 12.14.4 Protocol Completion/Sample Destruction 108 12.14.5 Handling for subjects at other Participating Institutions 109 12.15 Appendix 10: Data Collection elements Required by Protocol 110 12.16 APPENDIX 11: Problem Report Form 113   |
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1 introduction 1.1 study objectives

1.1.1 Primary Objectives x To determine whether Lupron leads to accelerated B cell recovery after allogeneic hematopoietic stem cell transplantation (after 1st and 2nd BMT).  
x To assess whether 18F FLT PET/CT could predict early engraftment/immune reconstitution in marrow and thymus after allogeneic HSCT (1st BMT only).  
x To assess safety of Lupron after 2nd HSCT and evaluate whether FLT can predict relapse
(evaluated after 2nd BMT)
1.1.2 Secondary Objectives x To investigate whether Lupron will accelerate thymic reconstitution after allogeneic HSCT using CT scan thymic index, flow cytometric measurements of naïve T cells, peripheral blood TREC on sorted CD4+ and CD8+ T cells, and Spectratype analysis of T
cell receptor diversity on sorted CD4+ and CD8+ T cells (1st BMT)
x To assess the feasibility of 4 month Lupron in male patients after allogeneic HSCT in post-pubertal individuals (after 1st and 2nd BMT).  
x To investigate whether Lupron will decrease the incidence of acute or chronic graftversus-host disease (GVHD) after allogeneic HSCT, comparing the Lupron treated and untreated groups using NIH developed criterion for the definition of clinical and biopsyproven acute and chronic GVHD (after 1st BMT).  
x To evaluate if Lupron will decrease the incidence of infectious complications in recipients of allogeneic HSCT, comparing the number and clinical severity of infectious complications between patients who received Lupron as compared to those that did not
(after 1st BMT).  
x To evaluate if Lupron will enhance graft-versus-tumor (GVT) effects by comparing the remission rates of patients who did vs. did not receive Lupron (after 1st BMT). To compare the outcomes of the pilot patients after 2nd HSCT in terms of relapse as compared to historical controls.  
x To evaluate if Lupron treatment results in other changes in immune reconstitution (e.g.  
dendritic cell reconstitution, monocyte reconstitution, and immunoglobulin recovery)
(after 1st BMT).  
x To demonstrate that 18F FLT imaging is safe in the peri-transplant period (after 1st BMT).
FLT endpoints include: 1) Ability of 18F FLT-PET to document preparative regimen ablation success 2) Evaluate the potential of 18F FLT/PET imaging to predict engraftment at week one 3) Assessment of GVHD using 18F FLT, 4) Assess relapse using FLT (after 1st and 2nd BMT).
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1.2 background and rationale 1.2.1

Impaired immune reconstitution contributes to morbidity and mortality after allogeneic stem cell transplant. Data suggest that this is due to the delayed and often disturbed recovery of T and B cells post-transplant. Although the innate immune system regenerates functionally and quickly post-transplant, the recovery of T and B lymphocytes is delayed for many months to years. This delay in lymphocyte cell recovery compared to other leukocytes is more pronounced in adult patients as compared to children. As shown in Figure 1 and Figure 2below, the two figures compare the percentage of normal leukocyte recovery in peripheral blood between adults and children. These show that B cell recovery in children approaches 50% by day 180 and 100% by day 270 and CD4+ numbers approach 100% at 2 years. In contrast, for adults, B cell recovery is 30% at day 180 and 80% at day 270 with CD4 recovery delayed indefinitely [1]
Figure 1  
  
  
Not only are the absolute numbers of T and B lymphocytes decreased post-transplant, the function of these populations remains compromised even after normal numbers are achieved. In part, this is due to the mechanism of lymphoid reconstitution post-transplant. In contrast to normal ontogeny where the thymus provides the vast majority of T cells to the peripheral pool, post-transplant T cell regeneration relies upon the mechanism of peripheral expansion for T cell recovery. In the thymus, T cells are produced de novo from marrow precursors. These T cells undergo T cell receptor (TCR) rearrangement, resulting in naïve T cells with diverse TCRs, capable of recognizing a wide array of infectious or tumor antigens. These developing T cells undergo positive and negative selection, permitting the deletion of autoreactive clones. In contrast, when thymic function is poor as is often the case after adult allo-HSCT, T cell reconstitution is accomplished through the rapid proliferation of mature T cell clones, leading to a skewed, oligoclonal population of T cells with the potential for alloimmunity.[1] Poor thymic function has significant clinical consequences in the setting of hematopoietic stem cell transplantation. Impaired thymopoiesis is associated with poor clearance of infections and diminished responses to vaccines (due to resulting B cell dysfunction). [2-6] Successful T cell reconstitution also confers significant protection from relapse of malignant disease as well. [7-9] Furthermore, impaired thymopoiesis is linked to the development and persistence of acute and chronic graft-versus-host disease. [6, 10-12] Thus, improving thymopoiesis and thus lymphopoiesis may partially abrogate GVHD, enhance immune surveillance, and improve HSCT outcomes. Similar to the effects of peripheral expansion on T cells, mature memory B cells demonstrate a restricted repertoire post-HSCT. [13] In contrast, B cells produced in the marrow through de novo development (transitional B cells) display a diverse repertoire. [13, 14] These transitional B cells emerge gradually (within months) after HSCT and are not influenced by viral reactivations and GVHD, suggesting that this population would be best used to identify the influence of immunotherapy on lymphocyte reconstitution after HSCT. In addition, studies have suggested that transitional B cell development is delayed and diminished in older adults, those with reduced intensity conditioning for HSCT, findings similar to the influence of these factors on thymus derived T cell development. [15] Furthermore, there are data to suggest that transitional de novo B cell development would be associated with enhanced tolerance that could translate to diminished GVHD. [16] Thus, because thymus recovery could take years in young adults, is likely influenced by infections and GVHD, while transitional B cell reconstitution is faster, less modulated by other post-transplant factors, B cells will be used for the primary endpoint of lymphocyte engraftment in this study. Thymus recovery and restoration of T cell diversity will be included as important secondary endpoints though this will likely only be evaluable in the small subset of patients without relapse, GVHD, who survive greater than 2 years after HSCT. At this time, there are not data to determine the best source of HSCs between marrow and peripheral blood stem cells. Marrow is chosen as the stem cell source for this protocol due to: 1) this is the standard stem cell source in pediatrics, 2) there are evidence that marrow products lead to less GVHD without compromising graft-versus-leukemia effects [17, 18], and 3) all of the basic science work that models the lupron and FLT effects were performed with marrow grafts. In many adult oncology institutions, marrow remains the preferred source as well (including University of Oklahoma). 1.2.2 Androgen Withdrawal Enhances T and B Lymphopoiesis Androgen inhibitors are excellent candidates for augmenting B lymphopoiesis and thymopoiesis. Many groups have demonstrated increase thymic size, weight, and thymocyte number following androgen withdrawal. [19-22] Murine and clinical data show that androgen withdrawal can reverse age-related thymic atrophy and reconstitute normal, young thymocyte subsets with subsequent increased peripheral T cell numbers. [23] Similarly, B lymphopoiesis is also increased after androgen withdrawal. [24] Data suggest that these increases in B and T cell numbers following androgen suppression are due to increases in de novo production of B and T cells in the marrow and thymus respectively. [21, 25] As shown in Figure 3below, following androgen withdrawal in male mice (light grey box), B cell total numbers are dramatically increased in the marrow as compared to control (black box) at 1, 3, and 4 weeks (see Figure 3 below, B subsets defined as B220+).  

Similarly, the thymus of mice treated with lupron was increased in size and thymocytes were increased in number as compared to controls. Notably, because lupron initially leads to androgen release, followed by suppression, the thymus first involutes then expands with subsequent increase in thymocyte production. As shown in Figure 4 below, thymus weight (left) and thymocyte number (right) are initially decreased at 1 week after lupron (grey), then similar at 2 weeks, then increased after 4 weeks as compared to controls (black).  

Additionally, recent data suggest that T and B cell reconstitution is accelerated after stem cell transplant in murine allogeneic transplant models after androgen withdrawal without decreasing graft-versus-tumor effect or worsening GVHD. [19] Our murine data demonstrate that the mechanism underlying the accelerated thymus recovery after androgen withdrawal involves: 1. Increase in thymus epithelial cell (TEC) proliferation, 2. Increased production of CCL25 by these TEC, 3. Increased early thymic progenitor entry of marrow precursors by CCL25 interactions, 4. Accelerated development of thymocytes due to CCL25 signaling, and 5. Overall enhanced thymopoiesis with increased export of naïve selected diverse T cells [25]. Similarly, B cell increases after androgen withdrawal have also been attributed to marrow stromal influences, resulting in accelerated de novo B cell development (rather than expansion of peripheral clones). [21] Given this mechanism and these murine and clinical data, androgen withdrawal has the potential to enhance de novo lymphopoiesis in young adults following HSCT with possible benefits of: improved infection and tumor clearance and diminished GVHD. Because of the timing of the androgen release and thymic involution and the possibility that these effects occur in marrow stroma as well, male randomized subjects will receive the lupron injection approximately 1-2 weeks prior to HSCT to minimize the potential additive damage of thymus involution and radiotherapy.
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1.2.3 lupron for androgen suppression

Lupron is an FDA-approved gonadotropin-releasing hormone (GnRH) analog that has been used clinically for androgen suppression in prostate cancer patients for males [23, 26] and for menses suppression during HSCT for females [27]. In males with prostate cancer, there are data to suggest enhanced thymic function following this agent. [23] Notably, most of these subjects were older individuals in whom free testosterone is substantially lower than younger subjects (0.2 +/- 0.081 vs. 0.43 +/- 0.098). [28] Thus, while patients may have diminished circulating testosterone following chemotherapy, the levels were not lower than those of older men, and the subjects are likely to still exhibit immunologic changes after androgen withdrawal. [29] Because Lupron is an analog of GnRH, it leads to an initial burst of sex steroid release followed by suppression. A study of administration and efficacy of Lupron in females for the suppression of menses revealed that the greatest efficacy was achieved when Lupron was given greater than 2 weeks prior to thrombocytopenia. Similarly, our murine data has demonstrated that optimal administration for Lupron with regards to marrow transplantation is approximately 1-2 weeks pre-transplant. This is because the initial stimulation of GnRH leads to thymus involution and 12 weeks following this will be a period of relative stabilization for the thymus, and possibly marrow stroma. Thus, thymic epithelial cells will be at basal proliferation and less harmed by radiation damage. Taken together, Lupron (4 month preparation) will be administered at approved doses 1-2 weeks prior to the preparative regimen.
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1.2.4 lupron safety profile

Lupron has a very good safety profile with only 3 reported adverse events (testicular atrophy) in greater than 20% of 94 patients studied in two clinical trials for prostate cancer (see section 10for details). There are data that HSCT alone leads to testicular atrophy in male patients, thus this particular AE is not likely to be unique in lupron-treated subjects after HSCT.
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1.2.5 lupron pharmacology

Lupron (leuprolide acetate) is a synthetic nonapeptide analog of GnRH that binds more strongly than the endogenous hormone. The 4 month preparation is 30 mg is dispensed in a prefilled dual-chamber syringe containing sterile lyophilized microspheres which, upon mixing with diluent become a suspension that can be given as an intramuscular injection. Within several days, a marked increase in serum sex steroid (testosterone) occurs followed by equilibration to baseline levels at 1 week and suppression by 2-4 weeks that is maintained until week 12 (data obtained from the prescription brochure). Although no specific drug interaction studies have been performed, drug interactions are unlikely to occur because the drug is only 46% plasma protein bound and degraded by peptidase not utilizing P-450 cytochromes. Furthermore, the drug has been used in HSCT for women for menses suppression without noted drug interactions. The effects are reversible as normal gonadal function is restored within 1-3 months upon discontinuation.
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1.2.6 flt imaging modality

Nuclear medicine imaging has been invaluable to assess tumor disease burden and response to therapy. PET/CT uses 18F-fluorodeoxyglucose (FDG) to identify cells that rapidly uptake glucose including tumor cells. This modality has revealed: tumor metastases, responses to tumor after treatment cycle, tumor recurrence, and has been useful to accurately predict true complete response, correlating PET negativity with longterm survival [30-33]. Despite these data demonstrating the utility of this test for evaluation in lymphomas and cancer, FDG-PET is limited by lack of specificity. Because FDG-PET relies upon glucose incorporation into cells, the signal will be present not only in tumor cells, but also normal tissues such as brain and heart that require active glucose uptake and in inflammatory lesions (due to uptake by macrophages and granulation tissues).  
  
  
FLT Thymidine
A novel agent, FLT is 3'-deoxy-3 18F-fluorothymidine, a radiolabeled thymidine analogue that illustrates dividing hematopoietic cells. FLT is derived from the drug azidovudine (AZT) that uses thymidine kinase 1 for uptake into cells, part of the salvage pathway for DNA synthesis. This enzyme is tightly controlled in normal cells and increases during cell division, permitting increased FLT uptake in dividing cells. Furthermore, malignant cells have lost their regulation and thus have higher thymidine kinase 1 available, leading to even greater uptake of FLT than normal cells that divide at higher rates [34]. Thus, this imaging agent has the potential to distinguish rapidly dividing cells from static cells and both of these from malignant cells based on degree of uptake of FLT. FLT has been used to successfully stage and evaluate chemotherapy response in non-hodgkins lymphoma [35-38], breast cancer [39-41], and sarcoma [42]. In many of these studies, high background activity was noted within the marrow compartment, indicative of FLT uptake by hematopoietic cells. FLT was then used to evaluate acute myelogenous leukemia in a pilot study, revealing higher SUV in patients with disease compared to controls.[43, 44] Similarly, patients with aplastic anemia or myelofibrosis associated with lack of hematopoiesis had decreased uptake in the marrow compartment [44]. FLT has been used to document response following treatment of AML [43] and has been shown  
 to be accurate and reproducible following chemotherapy in patients treated for testicular carcinoma [45]. It can further document changes in marrow following directed radiotherapy dosing, which may allow for more accurate assessment of response to therapy in marrow-borne malignancies [45]. These data suggest that FLT could be used to assess hematopoiesis in the marrow compartment, potentially illustrating empty vs. normal hematopoiesis vs. leukemia recurrence with greater sensitivity than single site marrow aspirates yield.  
Dr. Holter, our collaborator, has modeled FLT (3'-deoxy-3' 18F-fluorothymidine) in rat bone marrow transplantation and shown that by day 4, engraftment kinetics could be predicted and this time point preceded peripheral blood and full histologic evidence of marrow recovery. [46] As stated above, not only is immune reconstitution a major barriers to successful HSCT, but also our modalities for evaluating this are often invasive (bone marrow aspirate) or less informative (peripheral blood evaluation). In cord blood transplantation, this is a clinically significant problem as non-engraftment and late engraftment of the donor cells is common and there are currently no early predictive tests for whether a patient will engraft late or require a second conditioning regimen and new donor products. Furthermore, even in matched adult donor transplantation where graft failure rates are low, there are often periods post-transplant where the peripheral blood cellular counts decline and the differential includes graft loss, medication toxicities, infections, and relapse. Thus, a noninvasive imaging modality that could model engraftment kinetics would be invaluable to clinicians, potentially predicting graft failure early and narrowing the differential in the setting of cytopenias. These data prompted this clinical evaluation of FLT to determine if FLT could be used in conjunction with PET imaging at scheduled time points during HSCT for evaluation of engraftment. These data could: 1) predict the rate of marrow engraftment and the length of neutropenia, 2) demonstrate evidence of minimal residual disease at the time of transplant (the uptake should be null following myeloablative transplant preparative regimen), 3) predict thymus recovery. In this current  
  
protocol, adults and children with aggressive leukemias and MDS will have both the greatest opportunity for: 1) evaluation of residual disease that should be positive by FLT at the time of transplant, 2) a differential in uptake over time as the myeloablative negative scan increases in uptake corresponding with increasing hematopoiesis, and 3) the greatest chance for thymus recovery in adults to evaluate by FLT.
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1.2.7 18f flt safety profile

18F FLT is currently in use in clinical trials and has currently been shown to be safe at the doses proposed in this study. Although 18F FLT enters cells via an enzyme in the salvage pathway, it is not significantly incorporated into DNA due to the lack of a 3'-hydroxyl group, which is necessary for DNA propagation. Thus, although the agent remains within cells long enough for imaging, it will not persist long term nor should it be mutagenic. At the doses used in this study, several studies have been published and no adverse events reported in patients with leukemia and MDS [43, 44]. The planned dose is 0.07 mCi/kg with maximum dose of 3 mCi (maximum of 6.1 ug). At higher doses exceeding a 1000 fold increase in 18F FLT from our planned dose, 18F FLT
has been associated with grade 3 hematologic toxicity (6/10 at 0.125 mg/kg q12 hours which equals area under the curve of 417 ng-h/mL and 5/15 patients at 200 ng-h/ml), hepatic failure (2/10 at dose of 200ng-h/mL and 10mg/day). At a dose 1000 fold higher than our planned dose, mild peripheral neuropathy occurred (2/10 at dose of 50 ng-h/mL) [47]. No AEs attributable to 18F FLT administered at tracer levels have been reported in the published literature. While no related AEs are expected, patients will be monitored very closely with regards to hepatic, neurologic, and hematologic parameters following radiotracer injection.  
Patients on this study who receive 18 F FLT/CT will be exposed to a maximum whole body dose (ED) of 6.2 rem whole body dose (ED), 1.2 rem from each 18 F FLT/CT scan and 0.73 from each FDG PET/CT. For comparison, the average person in the United States receives a radiation exposure of 0.3 rem per year from natural background sources, such as from the sun, outer space, and from radioactive materials that are found naturally in the earth's air and soil. The dose that a patient will receive from this research study is about the same amount that individual would normally receive in 20.5 years from these natural sources. A routine clinical 18F FDG PET/CT is approximately 1.4 rem; for the purposes of this study, we have reduced the injected dose.
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1.2.8 18f flt pharmacology

18F FLT has a half life of 108 minutes [48]. The administered dose will be 0.07 mCi/kg with a maximum dose of 3 mCi per scan. The drug is excreted in the liver and kidney. Transport of FLT across cell membranes occurs by active transport and passive diffusion [49]. The drug solution is stored at room temperature in a gray butyl septum sealed, sterile, pyrogen-free glass vial and has an expiration time of 8 hours. The injectable dose of 18F FLT for most studies will EHP&LNJRIIOXRULQH-18, not to exceed 3 mCi with a specific activity greater than 200 Ci/mmol at the time of injection. In the dose of FLT, only a small fraction of the 18F FLT
molecules are radioactive. The amount of injected drug is < 6.1 ug (<25 nmol per dose) of 18F
FLT. 18F FLT is administered to subjects by intravenous injection of < 10mL over 30 seconds.  
There is no evidence that nonradioactive and radioactive FLT molecules display different biochemical behavior.
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1.2.9 flt availability

The Investigator-sponsored IND will be held by Karen Kurdziel, MD. 18F FLT will be obtained from commercial sources, either from Cardinal Health, Beltsville, MD, or PETNET Solutions Inc., Philadelphia, PA according to the DMFs filed with the FDA respectively for both institutions.
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1.2.10 trial plan

This study addresses 2 issues in transplantation: 1) the problem of delayed lymphoid engraftment after allogeneic transplantation in adults, and 2) the issue of evaluating engraftment of hematopoietic elements after allogeneic transplantation. Patients undergoing 1st BMT with aggressive leukemia and MDS who are $JH\HDUVROGDQGRU\HDUVROGDQGSXEHUWDODQG \HDUV will be enrolled and evaluated for remission status. If the patient is not in remission upon study entry, the patient will progress to a chemotherapy regimen with the goal of obtaining complete remission. Once the patient meets criteria for complete remission, female patients and half of the male patients (randomized) will receive Lupron prior to initiation of a standard preparative regimen with standard GVHD prophylaxis. Immunologic assessment of transitional B cells and IgM levels will be used for the primary endpoint of the issue of delayed lymphoid engraftment. This randomization and Lupron arm of the trial will be open at University of Oklahoma as well as NCI.  
A subset of these patients will undergo imaging analysis with 18F FLT as part of their evaluations. 18F FLT enrolled patients will be imaged at day -1, day 5 or 9, day 28 and a later time point to be determined by clinical events such as evidence of thymus recovery or GVHD. In addition, all newly enrolled NCI patients will be imaged at 1 year for evaluation of thymus reconstitution with the exception of those in the serial scanning subset that have been imaged at a 'later time' as discussed above. Ten FLT patients enrolled at Children's National Medical Center will be imaged at day -1, day +9, and if possible, day +28. Patients will not be imaged at University of Oklahoma. The primary endpoint for this element of the protocol will be toxicity with a secondary endpoint of imaging analysis at day 5 and day 28 and correlation or prediction of engraftment as compared to a null baseline scan at day -1. Important secondary endpoints will also include other elements of engraftment (neutrophils, T cells, and dendritic cells), relapse/remission status, presence and severity of acute and chronic GVHD, severe infectious complications, and whether the later 18F
FLT nuclear imaging scan may assist in these clinical diagnoses. Patients with aggressive leukemia and MDS who have relapsed after HSCT and are greater than or equal to 18 years and 55 will be enrolled and evaluated for remission status. Patients will undergo 2nd HSCT, after receiving Lupron and will all be evaluated with 18F FLT imaging to explore the relapse rates and ability of FLT to predict relapse after 2nd HSCT. The primary endpoints are: To assess safety of Lupron after 2nd HSCT and evaluate whether 18F FLT can predict relapse. Patients will undergo 18F FLT imaging on day -1, and FLT and FDG imaging on day 28 and day 60 to predict relapse. Patients may also be imaged before and after therapy for relapse (to not exceed 5 scans total per patient). Note patients enrolled on this protocol who relapse may receive another HSCT product on this protocol but will not be eligible to undergo FLT imaging again.
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2 eligibility assessment and enrollment

2.1.1 Inclusion Criteria: Transplant Recipient  
2.1.1.1 At NIH : Age 15 years old and/or  9 years old and pubertal and 55 years for recipient. Pubertal is defined by: prior menses at any time (females), documentation of clinical Tanner stage greater than 2 at some point pre-chemotherapy or at the current visit. (At this point, sex steroids have been produced for a few years which have driven initial pubertal development). Tanner 2 is defined as: breast buds for females with coarse pubic hair, and coarse pubic hair and testes > 2.5cm for males.
2.1.1.2 At Children's National Medical Center **only**: age > 4 years old and < 24.  
2.1.1.3 At University of Oklahoma: Age > 17 years old and 55 years for recipient. 2.1.1.4 A diagnosis of a hematologic malignancy for which stem cell transplant is standard of care:
2.1.1.4.1 Acute Lymphocytic Leukemia x Adult: (>22 years) > CR2 OR CR1 with e a high risk feature:
o Matched sibling donor for recipient treated on adult leukemia regimen [50, 51]  
o t(9:22) or bcr-abl+; t(4:11), t(1:19), t(8:14), 11q23 (MLL rearrangements)
complex cytogenetics (5 or more chromosomal abnormalities), hypodiploidy (<44 chromosomes). Note that patients with ALL blast crisis who emerge from CML are also eligible.
o Primary induction failure, defined as failure to achieve CR with primary induction chemotherapy o High WBC (>30,000 for B-cell ALL and >100,000 for T-cell ALL) at diagnosis o Persistence of minimal residual disease despite induction chemotherapy x Pediatric (< 22 years): > CR2 OR CR1 with a high risk feature:
o Matched sibling donor for recipient treated on adult leukemia regimen [50, 51]  
o Primary induction failure (M3 (>25% with greater 200 cells counted) marrow at day 29), M2 (5-25% blasts with greater than 200 cells counted) bone marrow or MRD > 1% at day 29 who then fail at day 43 with either an M2 or M3 BM or MRD > 1%
o Persistent leukemia and t(9;22) (MRD >1% day 29 or MRD > 0.01% endconsolidation)
o 11q23 (MLL) rearrangements detected by cytogenetic or PCR at initial diagnosis who are slow early responders (M2/M3 at day 14 or MRD> 0.01% at day 29)  
o Extreme hypodiploidy (< 44 chromosomes or DNA index of <0.81) detected by cytogenetic/ploidy analysis
2.1.1.4.2 Acute Myelogenous Leukemia x Adult: (>22 years) > CR2 OR CR1 with one of the following high risk features:
o Adverse or intermediate-risk cytogenetics including:
Normal cytogenetics [52] complex karytoype (>2 abnormalities) [53] inv (3) or t(3;3); t(11;19)(q23;p13.1); +13; -17/17p-; -18; -20; (t(6;9);
t(6;11); -7, 7q-; -5, 5q-; trisomy 8; t(3;5); t(9:11)(p22q23) [52, 53]
monosomy karyotype (presence of an autosomal monosomy in conjuntion with at least one other autosomal monosomy or structural abnormality.  
[52]
Any other karyotype **EXCEPT** t(8;21), t(9;11), inv(16), or t(16;16), and M3 (17; 17) [53] *unless ckit mutation present and then eligible. [*52]
AML emerging from CML (blast crisis) are eligible o Primary induction failure, defined as failure to achieve CR with primary induction chemotherapy o Secondary AML, defined as AML related to antecedent MDS, MPN, or cytotoxic chemotherapy o Hyperleukocytosis (WBC > 100,000 at diagnosis)
o Mutations in the FMS-like tyrosine kinase 3 (FLT3) gene (FLT3-LM; FLT-ITDs)
[54-56]
o Bilineage or biphenotypic leukemias are high risk features and eligible.  
x Pediatric (< 22 years): > CR2 OR CR1 with a high risk feature including:
o Primary induction failure (> 5% blasts in marrow after induction) o Persistent leukemia (>15% after first course of chemotherapy)
o Complex karyoptype, monosomy 7, or -5/-5q, FLT3 ITD-AR (>0.4) **EXCEPT** if also inv(16)/t(16;16), t(8,21)
o Normal cytogenetics or abnormal cytogenetics **EXCEPT** if also inv(16)/t(16;16), t(8,21)
are eligible for SIBLING transplant only o Bilineage or biphenotypic leukemias are high risk features and eligible.  
2.1.1.4.3 Myelodysplastic Syndrome RAEB 1 or 2; cytogenetics showing complex karyotype (3 or more abnormalities), monosomy 7/del(7q), or inv(3)/t(3q)/del(3q); or transfusion dependent.  
2.1.1.4.4 Chronic Myelomonocytic Leukemia 2.1.1.4.5 Chronic Myelogenous Leukemia who have failed 2G- tyrosine kinase inhibitors (TKI) 2.1.1.4.6 Standard pediatric indications for myeloablative transplantation for patients undergoing HSCT at Children's National Medical Center per institutional guidelines
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2.1.1.5 disease status

If patients are found to not be in remission at screening, then the patient may be returned to their primary hematologist/oncologist or may receive chemotherapy as per standard of care for the malignant disease. Patients for whom this would be their first allogeneic transplant must be in remission (< 5% malignant blasts in marrow and peripheral blood and no evidence of extramedullary disease) for transplant. Patients enrolled on this protocol for their second transplant do not need to have attained remission prior to transplant. 2.1.1.6 Performance status: Karnofsky or Lansky performance status > 60% AND life expectance of greater than 3 months.  
2.1.1.7 Ability to give informed consent. For recipients and donors < 18 years of age, their legal guardian must give informed consent. Pediatric patients will be included in an age appropriate discussion in accordance with institutional guidelines.  
2.1.1.8 Hepatic function: Patients must have evidence of adequate liver function prior to enrollment defined by total bilirubin < 2.5 mg/dL (unless documented Gilbert's syndrome) AND transaminases < 5 x the upper limit of normal for age appropriate indices.  
2.1.1.9 Renal function: Patients must have evidence of adequate renal function to proceed with stem cell transplant, creatinine clearance > 60 ml/min/1.73 m2. GFR may also demonstrate adequate renal function.  
2.1.1.10 Left ventricular ejection fraction > 50% OR shortening fraction of >27% demonstrated on 2D echocardiogram or MUGA.  
2.1.1.11 Pulmonary function of DLC0 adj/VA and FEV1 > 60% of normal indices for age and height unless the patient has a likely acute reversible etiology of decline and then DLCO adj/VA > 30% of normal. Pediatric patients unable to complete PFTs may be enrolled as per enrolling institution SOP for recipient guidelines.  
2.1.1.12 Patients with prior autologous stem cell transplants will be included. Patients with prior allogeneic stem cell transplants will be eligible for 2nd BMT if not previously transplanted with FLT on 11-c-0136.  
2.1.1.13 Prior experimental systemic therapies must have been completed greater than 2 weeks prior to study entry.  
2.1.2 Exclusion Criteria: Transplant Recipient 2.1.2.1 History of psychiatric disorder which may compromise compliance with transplant protocol, or which does not allow for appropriate informed consent.  
2.1.2.2 Active infections not responding to therapy. All efforts should be made to clear the infection prior to enrollment.  
2.1.2.3 Clinically significant systemic illness with manifestations of significant organ dysfunction which in the judgment PI or AI would render the patient unlikely to tolerate the protocol therapy or complete the study.  
2.1.2.4 Presence of active malignancy from an organ system other than hematopoietic. 2.1.2.5 HIV infection. 2.1.2.6 Chronic active hepatitis B infection. Patients may be hepatitis B core antibody positive but must be surface antigen negative and without active evidence of disease.  
2.1.2.7 Pregnant or lactating females will be excluded from this trial due to unknown risks to the developing fetus. Patients of child-bearing potential must use an effective form of contraception while on study.  
2.1.2.8 Sexually active individuals capable of becoming pregnant who are unable or unwilling to use effective form(s) of contraception during time enrolled on study and for 1 year post-transplant (see section 4.4for details).  
2.1.2.9 History of prior Lupron intolerance. Note: patients ARE eligible if prior or current lupron exposure.
2.1.3 Inclusion Criteria: Matched Related Transplant Donor  
2.1.3.1 Age 2 and 60 years old and able to give consent or assent. For donors < 18 years old, the legal guardian must be able to provide informed consent and an evaluation by a LSW or psychiatric personnel will be needed to determine willingness to participate. Pediatric patients will be included in an age appropriate discussion in accordance with institutional guidelines.
2.1.3.2 HLA-matched related donor, excluding identical twins. Donors must be matched at least 7 loci out of 8 at the allele or antigen level excluding antigen DRB1 mismatch.  
2.1.3.3 Donor selection will be in accordance with NIH/CC Department of Transfusion Medicine criteria and must be able to medically endure stem cell collection or as per local institutional guidelines.  
2.1.3.4 Donors must be HIV negative, HTLV negative, HBsAg negative.  
2.1.3.5 Donors must be physically able to and willing to tolerate marrow harvest collection preferably, or in the absence of this option, able and willing to donate via peripheral blood pheresis.
2.1.4 Exclusion Criteria: Matched Related Transplant Donor 2.1.4.1 History of medical illness that in the estimation of the PI or DTM physician precludes donation of marrow.  
2.1.4.2 Anemia (Hb < 10 gm/dl) or thrombocytopenia (< 100,000/ ul). 2.1.4.3 Pregnant females (due to risk to fetus). 2.1.4.4 Current psychiatric diagnosis that would compromise compliance with transplant protocol or precludes appropriate informed consent.  
2.1.4.5 Presence of any blood transmissible infectious disease that cannot be cleared prior to stem cell collection and poses an unacceptable risk for the recipient (excludes CMV).  
2.1.4.6 Active malignancy will exclude the donor. Any malignancy less than five years postremission will exclude the donor. Non-hematologic malignancies greater than 5 years ago will not exclude the donor. Any history of hematologic malignancy will be considered on a case by case basis.  
2.1.4.7 Any medical contraindication to anesthesia or marrow donation will exclude the donor. 2.1.4.8 Donors receiving experimental therapy or investigational agents. 2.1.4.9 Active autoimmune disease that in the opinion of the PI or AI would compromise the success of the transplant.
2.1.5 Inclusion Criteria- Matched Unrelated Donor  
2.1.5.1 Unrelated donor matched at HLA-A, B, C, and DR loci by high resolution typing (at 8/8 or 7/8 antigen/allele match) are acceptable donors.  
2.1.5.2 The evaluation of donors shall be in accordance with existing National Marrow Donor Program (NMDP) Standard Policies and Procedures at all institutions.
2.1.6 Inclusion Criteria- 18F FLT Candidate Transplant Recipient 2.1.6.1 Meets criteria Sections **2.1.1** and **2.1.2**
2.1.6.2 Age > 18 years old at NCI, and age > 4 years and < 24 years at Children's National Medical Center 2.1.6.3 Donor who is willing to undergo bone marrow or stem cell harvest.  
2.1.7 Exclusion Criteria- 18F FLT Candidate Transplant Recipient 2.1.7.1 History of prior fluorothymidine allergy or intolerance.
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2.2 screening evaluation and criteria

The following clinical, laboratory, radiological assessments must be performed on the recipient within 45 days of initiating HSCT preparative regimen with the exception of Section **2.2.2.6**
which may be performed within the preceding 3 months. These may be performed at outside institutions. 2.2.1.1 Evaluations x Complete history and physical examination.  
x CT scan of head, sinus, chest, abdomen, and pelvis.  
x An echocardiogram (2D or MUGA) an electrocardiogram. x Pulmonary function tests (PFTs) must be performed for eligibility and FEV1, FEV1/FEV
ratio, DLC02 recorded for adult patients; Oxygenation > 94% on RA may be used for pediatric patients unable to undergo PFTs at Children's National Medical Center.
x Laboratory studies:
o CBC with differential, ABO blood type, PT/PTT, fibrinogen, thrombin time o Comprehensive chemistry o Lipid profile with triglycerides, TSH, free T4 o Urine pregnancy test for women of childbearing potential o 24 hour creatinine clearance or GFR o Bone marrow biopsy and aspiration with studies appropriate to staging of malignant disease, a LP or other biopsy may be required depending upon the diagnosis (to include LP for all AML and ALL)
o Infectious disease work-up at NCI only: antibody for Hepatitis A, B, C, HIV,
T.Cruzi, HTLV-I/II, CMV, adenovirus, varicella, EBV, HSV, Toxoplasmosis, and syphilis. Work-up at other institutions may follow their policy.  
x Immunology studies: Lymphocyte phenotype panel (T, B, NK), IgG, IgA, IgM for NCI
patients only.  
x Radiation Oncology Consult (within 28 days of initiating conditioning chemotherapy for HSCT at NCI). At Children's National Medical Center and University of Oklahoma, this is per institutional practice.  
x Nutritional assessment (initial consult) and dental consult and social work consultation must be performed within 3 months of initiating preparative regimen and typing for HLA-A, -B, -C, and DR with PCR test of DNA mini-satellite regions for future determination of chimerism or as per local institutional practice. Note that when possible, the HLA and chimerism assays require 2 separate tubes of blood at NCI.
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2.2.1.2 criteria to proceed to

x Disease status: Remission as defined by < 5% blasts in marrow and no evidence of extramedullary disease for patients undergoing their first allogeneic transplant. Patients undergoing their second transplant do not have to have obtained a complete remission.  
x Performance status: Karnofsky or Lansky performance status > 60% AND life expectance of greater than 3 months.  
x Hepatic function: Patients must have evidence of adequate liver function prior to enrollment defined by total bilirubin < 2.5 mg/dL AND transaminases < 5 x the upper limit of normal for age appropriate indices.  
x Renal function: Patients must have evidence of adequate renal function to proceed with stem cell transplant, creatinine clearance > 60 ml/min/1.73 m2.  
x Left ventricular ejection fraction > 50% demonstrated on 2D echocardiogram or MUGA. x Pulmonary function of DLC0/Adj/VA and FEV1 > 60% of normal indices for height and weight for adults and >95% oxygenation on RA for pediatric patients.  
x Patients who have received prior chemotherapy must have had resolution of significant toxicities to proceed to transplant.  
x Negative pregnancy test (females)  
x No evidence of an active infection (clinical assessment and review of CT) that is not currently being treated or responding to therapy.  
x Either a HLA-matched related or unrelated donor, excluding identical twins. Donors must be matched at least 5 loci out of 6 by antigen matching or at least 8 of 10 using allele matching who met criteria for entry onto this protocol.  
2.2.2 Matched Related Donor
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(see section 12.3)

The following clinical, laboratory, radiological assessments must be performed on the donor within 4 weeks of initiating HSCT preparative regimen with the exception of Section **2.2.2.6** at NCI. Note, where these differ from institutional practice at University of Oklahoma or Children's National Medical Center, the SOPs at those institutions may be followed. 2.2.2.1 Complete history and physical examination. 2.2.2.2 Laboratory studies:
o CBC with differential, ABO blood type, PT/PTT o Comprehensive chemistry  
o Urinalysis
o Urine pregnancy test for women of childbearing potential o Screening for Hemoglobinapathy at NCI only if CBC suggests anemia due to hemoglobinapathy o Infectious disease work-up: antibody for Hepatitis A, B, C, HIV, T.Cruzi, HTLVI/II, CMV, adenovirus, EBV, HSV, varicella, Toxoplasmosis, and syphilis (RPR)
at NIH or as per institutional practice at Children's National Medical Center or University of Oklahoma.  
2.2.2.3 Immunology studies: Lymphocyte phenotype panel (TBNK) at NCI only.  
2.2.2.4 Electrocardiogram 2.2.2.5 Chest radiograph
2.2.2.6 Typing for HLA-A, -B, -C, and DR with PCR test of DNA mini-satellite regions for future determination of chimerism (may be done at anytime prior to HSCT).  
2.2.2.7 Donor Health History Screen through Dowling Apheresis Center for those undergoing peripheral blood stem cell collection at Dowling Apheresis Center at NCI, or as per institutional guidelines at: Univ of Oklahoma, or Children's National Medical Center.  
2.2.2.8 DTM venous assessment and health history screen through Dowling Apheresis Center for those undergoing PBSC collection at NCI or as per institutional practice.  
2.2.2.9 For donors undergoing bone marrow harvest, pre-anesthesia evaluation at NCI or as per local institutional practice.
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2.3 registration procedures 2.3.1 protocol

Protocol "entry date" is considered to be the day that the informed consent form has been signed by the recipient.
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2.3.2 registration

Authorized staff must register an eligible candidate with NCI Central Registration Office (CRO) within 24 hours of signing consent. A registration Eligibility Checklist from the web site (http://intranet.cancer.gov/ccr/welcome.htm) must be completed and sent via encrypted email to: NCI Central Registration Office ncicentralregistration-l@mail.nih.gov. After confirmation of eligibility at Central Registration Office, CRO staff will call pharmacy to advise them of the acceptance of the patient on the protocol prior to the release of any investigational agents. Verification of Registration will be forwarded electronically via e-mail to the research team. A recorder is available during non-working hours.  
2.3.2.1 Donors at NIH will be registered on this study prior to stem cell collection. Donors at Children's National Medical Center and University of Oklahoma need to be identified and eligible but may follow local institutional practice.
2.3.2.2 Unrelated donors from NMDP Centers will be consented on an approved NMDP
consent prior to eligibility evaluation. NIH UID numbers will be generated on unrelated donors while maintaining the anonymity required by the NMDP. The recipient may not sign consent before the matched unrelated donor has been identified and cleared by NMDP and the patient has been determined to be eligible At other institutions, this will be done as per institutional practice.  
2.3.3 University of Oklahoma Donors and Recipients 2.3.3.1 These patients will sign consent after meeting eligibility criteria at the University of Oklahoma (Site PI: Dr. Holter), may receive therapy at the National Cancer Institute or the University of Oklahoma.
2.3.3.2 Registration will be a two part process as patients are screened on this protocol. A
protocol registration form will be supplied by the CCR study coordinator and updates will be provided as needed. Subject eligibility and demographic information is required for registration. To initially register a subject, after the participant has signed consent, complete the top portion of the form and send to CCR study coordinator. Once eligibility is confirmed, after completion of screening studies, complete the remainder of the form which is the eligibility checklist, indicating that the patient is being registered for treatment and send to CCR study coordinator. In addition, source documents supporting the eligibility criteria must be sent to the CCR study coordinator. The CCR study coordinator will notify you either by e-mail or fax that the protocol registration form has been received which will include the unique patient/subject ID number. Questions about eligibility should be directed to the CCR study coordinator or PI. Questions related to registration should be directed to the CCR study coordinator. Subjects that do not meet screening criteria should be removed from the study following the procedure in section**3.10.4**.  
2.3.4 Children's National Medical Center Donors and Recipients 2.3.4.1 Patients will sign consent after meeting eligibility criteria at Children's National Medical Center (Site PI: Dr. Williams), and will receive therapy at Children's National Medical Center.  
2.3.4.2 Registration will be a two part process as patients are screened on this protocol. A
protocol registration form will be supplied by the CCR study coordinator and updates will be provided as needed. Subject eligibility and demographic information is required for registration. To initially register a subject, after the participant has signed consent, complete the top portion of the form and send to CCR study coordinator. Once eligibility is confirmed, after completion of screening studies, complete the remainder of the form which is the eligibility checklist, indicating that the patient is being registered for treatment and send to CCR study coordinator. In addition, source documents supporting the eligibility criteria must be sent to the CCR study coordinator. The CCR study coordinator will notify you either by e-mail or fax that the protocol registration form has been received which will include the unique patient/subject ID number. Questions about eligibility should be directed to the CCR study coordinator or PI. Questions related to registration should be directed to the CCR study coordinator. Subjects that do not meet screening criteria should be removed from the study following the procedure in section **3.10.4**.
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2.4 randomization procedures 2.4.1 female

2.4.1.1 All female recipients will receive Lupron 2.4.2 Male Recipients 2.4.2.1 The CRO will provide the randomized treatment assignment to the PI based on random assignments determined by the study statistician for first BMT. For 2nd BMT, all patients will receive Lupron.
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3 study implementation 3.1 study

3.1.1 NCI and University of Oklahoma for 1st BMT
(Note: FLT/FDG PET scans will not be performed at the University of Oklahoma)  
  
2) Tacrolimus 0.02 mg/kg (IBW) continuous IV infusion from Day -1 until tolerating oral
Some of these patients are anticipated to be evaluated using FLT (to include only patients needed for the immunological primary endpoint, not increasing total patient numbers). 23 adult NCI patients in total will undergo FLT PET/CT imaging on day -1, at day +5 or day +9, at 4 weeks, and at a future point to include evidence of GVHD relapse, or immune recovery. An estimated 50 patients (including subset of the 23 patients undergoing serial scanning) will be imaged approximately at 1 year for evaluation of thymus reconstitution. The total possible numbers will include no more than 118 patients to achieve the 64 evalauble adults for the immunological primary endpoint. However, as of amendment K, all NCI patients will undergo a single 1 year FLT for evaluation of thymus reconstitution.  
For 2nd BMT, at NCI, all 16 patients will undergo FLT imaging at day -1, and FLT and FDG
imaging on day 28, day 60 and at relapse and response to therapy (if this occurs). 3.1.2 Children's National Medical Center Ten pediatric patients at CNMC will undergo FLT PET/CT imaging on Day -1, day +9, and day +28 (if possible). Initial images will be correlated with engraftment and other secondary endpoints. At Children's National Medical Center, the patients will not receive Lupron outside of the context of clinical care, and will receive myeloablative HSCT as per standard of care with FLT imaging for engraftment as the only primary endpoint.
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3.2 drug administration 3.2.1 lupron

Lupron will be administered to all female patients and half of the male patients (randomized for first BMT) and to all patients for 2nd BMT, once optimally between day -13 and day -20 pre-HSCT but definitely prior to initiation of preparative regimen as a 4 month intramuscular injection for patients > 18 years and as a 3 month injection (adult preparation) for patients < 18 years. Subsequent injections may be given if clinically indicated to females. There will be no dose modifications. Females already on lupron therapy will receive the 4 month injection within 2 weeks prior to the next scheduled injection and prior to HSCT preparative regimen.
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3.2.2 18f-flt

All NCI patients will undergo PET imaging in the NCI Molecular Imaging Clinic (B3). Pediatric patients will undergo PET imaging in the Children's National Medical Center clinical imaging department. Patients at University of Oklahoma will not undergo PET imaging per protocol A low dose transmission CT scan will be performed for the purpose of attenuation correction.  
18F FLT PET/CT will be performed per the described schedule to include 4 imaging sessions in the peri-transplant at NCI and a maximum of 3 at Children's National Medical Center. 18F FLT
PET/CT alone will be used to assess the marrow at a pre-HSCT baseline image and an image day +5 or +9 (to evaluate for early signs of engraftment). At both institutions, for first BMT, day + 28 (+/- 5 days) image may be performed and at NIH only, a later time point (to be obtained when clinically indicated either with evidence of thymus recovery or GVHD), 18F FLT and FDG
imaging will be conducted within 48 hours. At NIH only, all newly accrued patients will undergo a 18F FLT scan at 1 year post-HSCT with FDG. These will be used to distinguish dividing cells (18F FLT+) from cells high in glucose consumption (FDG+). A few patients in relapse who are consented to this protocol arm will undergo imaging analysis with 18F FLT PET/CT alone for calculation of 'positive' control for leukemia relapse. For 2nd BMT, patients will undergo 18F FLT imaging on day -1, day 28, and day 60 to predict relapse. Patients may also be imaged before and after therapy for relapse (to not exceed 5 scans total per patient).  
The expected dose of [18F]FLT is 0.07 mCi/kg with a maximum of 3 mCi. Due to potential unpredictable delays, the total activity administered may be reduced at the discretion of the PI.  
18F FLT will be provided by Cardinal Health or PETnet pharmacies. IND will be held by Dr.  
Karen Kurdziel, Molecular Imaging Program, CCR, NCI.  
18F FLT and FDG uptake greater than surrounding background (T:B>1.2) will be considered positive. For hypermetabolic lesions, an estimated smoothed maximum SUV will be reported. Analysis of uptake will be performed using current practice guidelines within the Molecular ImagingProgram, Children's National Medical Center and University of Oklahoma (on deidentified scans).
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3.3 on study protocol evaluation

See Sections **12.2** and **12.3**
3.3.1 Within 4 weeks of Transplant Date  
x History and Physical Examination x Laboratory studies must include the following:
o Complete blood counts with differential. o Complete chemistry panel: sodium, potassium, chloride, CO2, blood urea nitrogen, creatinine, calcium, phosphorus, magnesium, ALT, AST, total and direct bilirubin.  
o Toxoplasma IGG, Varicella IGG, CMV IGG, HSV IGG. o Immunologic screening: Immunoglobulin levels (IgG with subclasses, IgA),
CD4/CD8 total numbers and ratio at NCI only.  
o Flow cytometry of peripheral blood for T/B/NK and residual disease at NCI and Children's National Medical Center only o Chest and sinus CT
o Pulmonary Function Tests for adult patients and oxygen saturation on room air for pediatric patients.  
o Echocardiogram or MUGA o Electrocardiogram o BM biopsy and aspirate with flow cytometry, and cytogenetics if previously positive o Lumbar puncture with flow cytometry for patients with ALL and AML o Testosterone and Estrogen levels, LH, FSH for all adult patients.
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3.3.2 flt on study evaluations

(For patients enrolled on 18F FLT arm, Section **12.4**):
x Imaging Study time points, 1st BMT:
o Day -1: 18F FLT + low dose CT
o Day +5 or +9: 18F FLT + low dose CT
o Day +28, 29: 18F FLT + low dose CT, 18F FDG(not required at Children's National Medical Center) + CT
o Day + (GVHD vs. thymus recovery): 18F FLT + low dose CT, 18F FDG + CT at NCI only x Imaging Study time points 2nd BMT:
o Day -1: 18F FLT + low dose CT
o Day +28/29: 18F FLT + FDG (not required for Children's National Medical Center site) +
low dose CT
o Day +60/61: 18F FLT + FDG (not required for Children's National Medical Center site) +
low dose CT
o Day + of relapse and initial response to immunotherapy: 18F FLT + FDG + low dose CT
at NCI only x Laboratory studies for FLT within 24 hours prior:
o Creatinine < 2, ALT and AST within 5 x ULN
o TK1 serum measurement x 18F FLT imaging protocol:
o At NCI immediately following the IV infusion of 18F FLT, a dynamic PET
emission will be performed for 45 minutes followed by a whole body static image. An additional PET emission imaging acquisition will be performed 120 minutes (+/- 15 minutes) post infusion. A corresponding low dose transmission CT will be acquired immediately before each emission scan. At Children's National Medical Center, the dynamic image will be omitted.
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3.4 biologic studies

The total aliquot of phlebotomy will not exceed 3 mL/kg per draw and 7 ml/kg in a 4 week period for research studies. Should the research requests exceed these restrictions, research studies will be done in the order of Sections **12.5,12.6** and **12.7**.
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3.4.1 immune reconstitution

Studies to elucidate difference in immune reconstitution between Lupron treated and untreated groups to include multiparameter flow cytometry analysis of relevant cell populations (B and T) and serum analysis of relevant cytokines. If available, these will be compared with donor lymphocyte characteristics (whereby 5 million total donor nucleated cells of the donor product will be reserved for biologic studies and go to the clinical core of ETIB (Bldg 10, Room 12C216, (301- 402-3627)). At Univ of Oklahoma, these may be stored within CETI lab or Holter lab respectively, or sent directly to the Pre-Clinical Core at NCI. At Children's National Medical Center, these may be stored within CETI lab or Bollar lab respectively, or sent directly to the Pre-Clinical Core at NCI.
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3.4.2 immunologic studies

Impaired lymphocyte reconstitution is believed to contribute to the morbidity and mortality of allogeneic stem cell transplantation as evidenced by poor clearance of infections, and alloimmunity (GVHD).[57] In adult HSCT recipients, T and B cell functions can be compromised indefinitely due to lack of thymus function and poor B cell education. This study will use immunologic studies to determine if Lupron may enhance thymus reconstitution and B cell development. Exploratory studies will include immunologic evaluations of HSCT complications to include: relapse, GVHD, infections, and the influence of thymus reconstitution on these complications. For these studies, prospective collection of infections to include: HHV6, adenovirus, EBV, CMV. 3.4.3 Objectives/Specific Aims 3.4.3.1 Determine the effect of Lupron on engraftment and the recovery of T and B cell subsets following allogeneic HSCT by enumeration of cellular subsets in patients with and without Lupron. Should thymus recovery be evident in subsets, secondary analysis of thymus function using T cell receptor excision circle analysis on peripheral T cells and using spectratype techniques.  
3.4.3.2 Determine whether FLT uptake demonstrates engraftment, leukemia relapse or residual disease, and/or GVHD, and/or thymus recovery. FLT images will be compared as follows: pre-transplant image will be used for assessment of MRD (to be correlated with disease recurrence and histology at day 28), post infusion image will be used to assess homing and cellular growth at day +5 or +9 image to correlate to time to engraftment, and correlate with marrow stability at histological assessment day +28, day +28 image to be quantitatively evaluated to correlate with blast % and marrow repopulation with histology, and the final image to be performed at a time of new-onset GVHD, potential relapse, or thymus recovery. Exploratory studies to include the biology of HSCT.  
3.4.3.3 For arm 2, exploratory analyses will include: whether FLT uptake can predict relapse or response to immunotherapy and whether there is an improvement in this small number of patients in relapse rates (from 80% as designated in historical controls). Immune studies will be undertaken to explore whether there is alterations in lymphocyte subsets similar to patients who remain in remission on the primary arm of the protocol.
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3.4.4 methodology

The following studies will be performed on peripheral blood (if available) samples. Blood samples (20 cc in red/green CPT heparin and 16 cc in EDTA tubes, age adjusted for weight in pediatric patients) will be drawn for evaluations. Time points are listed in section 3.8.  
Complete blood counts and TBNK will be collected on the same day. Samples should be delivered to the lab of Dr. Fran Hakim in Building 10, Room 3E-3288 (301-402-3627) at NCI and may be processed and frozen and shipped in batch from Children's National Medical Center and University of Oklahoma. 3.4.4.1 Peripheral blood: quantity (20mL). Flow cytometry will be performed on peripheral blood samples. Commercially available antibodies will be used to characterize T cell, B cell, dendritic cell and monocyte recovery after HSCT. Samples will be frozen and thawed in batches.  
3.4.4.1.1 Research blood sample aliquot size will be minimized for patients < 18 years of age and the total amount restricted to a maximum of 3 ml/kg per draw and 7 ml/kg per 6week period (maximum 450 ml). In the event that blood draws are limited due to patient size, research studies will be performed in order of priority as listed in Section 12.7. Small volume apheresis collections will not be included in this calculation since such is performed isovolumetrically with minimal red cell loss.  
3.4.4.2 Plasma (>20 mL). Plasma (EDTA and heparin) will be spun in refrigerated centrifuge and frozen immediately and subsequently utilized for cytokine studies using immunofluorescent technique.
3.4.4.3 Tissue, urine, and other fluid specimens (optional): Any pathology or biopsy specimen or fluid that is obtained for clinical indications from the recipient and has remaining (or extra specimen) following the clinically indicated test may be stored and used for immunological characterization. This includes any biopsies of bones obtained because of new onset pain, uptake of nuclear imaging scanning concerning for relapse or infection from the recipient. Patients will have the opportunity to opt outin writing to determine if extra fluids and tissues may be used for these studies.
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3.5 collaborative research studies

Under Amendment P, coded serum samples and human peripheral blood will be sent to the Children's National Medical Center to evaluate comparisons of T and B cells between patients who received lupron and those who did not in terms of relapse, GVHD, and infections Rationale: Dr. Williams is a recognized expert in multicolor flow cytometry for the analysis of human peripheral blood from post-allogeneic-transplant patients. Specific Aims: To determine the effect of Lupron on engraftment and the recovery of T and B cell subsets following allogeneic HSCT by enumeration of cellular subsets in patients with and without Lupron.
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design/procedures

The samples will be sent to laboratories at the Children's National Medical Center to perform multicolor flow cytometry as per section **3.4.4.1** as well as cytokine studies as per section **3.4.4.2**
after an MTA is executed between the NCI and the Children's National Medical Center. All excess samples will be destroyed at the conclusion of the planned studies. The serum and human peripheral blood samples will be sent to the following address:
Kirsten Williams MD Center for Cancer and Immunology Research Program for Cell Enhancement and Technologies for Immunotherapy Children's National Medical Center 5th Flr, Suite 5220 111 Michigan Ave NW Washington, DC 20010 Coded serum samples will be sent to Dr. Williams from patients participating in the 11-C-0136 clinical protocol led by primary investigator, Dr. Christopher Kanakry at NCI. Cryopreserved heparinized serum from up to 25 patients will be transferred through this collaboration. Laboratory results will be sent back to Dr.Kanakry for data compilation and analysis. Dr.  
Williams will not have access to patient identifiers or patient information.  
Only the subject's unique identifier will be used and information about the enrolling site will be removed. Authorized investigators at NCI and Dr. Williams will maintain codes linking the laboratory specimens received to clinical information; however, Dr. Williams will not have access to any other patient identifiers or patient information. Sample identifiers are not derived from actual identifiers and Dr. Williams's research team will not have access to the reidentification key.
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3.6 donor collection 3.6.1 matched

Matched unrelated donors will be collected as per NMDP policies and procedures. The concentration of CD34+ cells will be determined by flow cytometry. The product may be infused fresh or stored frozen prior to infusion. Every attempt will be made to obtain a marrow product for first BMT. Should PBSC be the only alternative, these patients will be analyzed separately and replaced. The target dose is > 350 million total nucleated cells per kilogram. Of this product, 5 million total donor nucleated cells will be reserved for biologic studies and go to the clinical core of ETIB (Bldg 10, Room 12C-216, 301 402 3627) at NCI. Cell selection will not be performed on the HSC grafts. If samples are obtained at Children's National Medical Center and University of Oklahoma, these may be processed, frozen, and shipped in batch, or fresh to Dr. Hakim (above). MUD policies at Univ of Oklahoma and Children's National Medical Center will be followed as per SOPs of institutional practice. Please refer to Section 3.4 for research study details. For 2nd BMT, PBSC will be preferred over marrow.
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3.6.2 matched related donors

Matched related donors will undergo marrow harvest with general anesthesia. Subjects will undergo anesthesia consultation, and meet criteria for eligibility/enrollment. CD34+ fraction will be determined. Subjects will also undergo the Donor Health History Screen to determine donor eligibility using standard DTM criteria in the Dowling Apheresis Clinic by skilled staff in the Blood Services Section for adult patients and age-appropriate questioning when indicated for pediatric subjects at NCI, the Oklahoma Blood institute, and will be evaluated and exam performed by a member of the Oklahoma Transplant team for both PBSC collection and marrow donation, and by BMT staff at Children's National Medical Center as per institutional protocol. Subjects will undergo follow-up history and physical examination within 1 week of donation, at NCI or as per local institutional guidelines. Donor recipient weight discrepancy with smaller donors placing the recipient at risk for non-engraftment will necessitate PBSC collection. Calculations will be performed by weight; for obese patients, an average between ideal and actual body weight. PBSC collection will be done as per standard of care and local institutional practice; at NCI, with donors receiving 10 ug/kg of GCSF for 5 days, with first collection through large bore venous catheter on the 5th day and potential second collection on day 6 (for insufficient sample) with a 6th dose of GCSF. Should PBSC be the only alternative, these patients will be analyzed separately and may be replaced. The target dose is > 350 million total nucleated cells per kilogram; however > 100 million total nucleated cells per kilogram will be sufficient for marrow collections and between 3-4 million CD34/kg for PBSC products. Of this product, 5 million total donor nucleated cells will be reserved for biologic studies and go to the clinical core of ETIB (Bldg 10, Room 12C-216, 301 402 3627) at NCI, and may be processed and stored and shipped frozen from outside institutions. Cell selection will not be performed on the HSC grafts. Note that TTV testing sample will be drawn as per institutional practice (e.g. on the day of marrow collection at NCI). Harvested cells will be processed in DTM at NCI, within the Stem Cell laboratory at Children's National Medical Center and in the Oklahoma Stem cell processing center at Univ. of Oklahoma. For all frozen samples and all fresh samples with ABO mismatch, the product will be depleted of red cells until less than 0.5 cc/kg or 50 cc total remain (whichever is smallest) when possible. For fresh harvest products, plasma pheresis may also be required if indicated by ABO mismatch. DTM at NCI, the Stem Cell laboratory at Children's National Medical Center, and the Oklahoma Stem Cell processing unit at Univ. of OK will notify the Adjunct PI and/or NIH PI and /or site PI and research RN of any problems in regard to cell number or red cell volume prior to freezing product. Note: for 2nd BMT, PBSC products will be preferred.
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3.7.1 central access

An appropriate form of central access will be obtained prior to HSCT procedure. Tunneled triple lumen catheter is preferred in thorax at NCI. Other institutions will follow institutional standards. 3.7.2 Remission Status Patients undergoing first HSCT must be in remission prior to HSCT; this may include patients without evidence of disease in aplasia. 3.7.3 Conditioning Regimen 3.7.3.1 Adults (>22 years and on ETIB service at NCI, on adult BMT at Univ of Oklahoma): 3.7.3.1.1 TBI will be delivered to a total midplane dose of 12Gy and median lung dose of 6Gy.  
Radiation will be fractionated twice daily over 4 days (days -8, -7, -6, -5). Opposed lateral fields with lung compensation will be treated to 1.5 Gy for each fraction to a total cumulative dose of 12Gy. Mediastinal fields will be treated AP-PA concurrently with the opposed lateral fields at a dose of 75cGy twice daily to bring the mediastinal dose to 12Gy with a median lung dose of 6 Gy. Fractions will be delivered with at least a 6 hour interfraction interval. Gonadal shielding will not be used. Head and neck compensation and other technical modifications may be made at the discretion of the treating radiation oncologist. For patients with CNS positive disease at any time after diagnosis, **CNS boost may be delivered unless contraindicated by PI or AI.**
The CNS boost will be delivered as a 6 Gy boost in once daily 2 Gy fractions. The boost will be delivered prior to TBI. Patients with sites identified by nuclear imaging or other clinically accepted diagnostic tests may receive a boost to these areas as clinically indicated at any time point pre- or post transplantation.  
3.7.3.1.2 Cyclophosphamide: cyclophosphamide 60 mg/kg IV, Days -4, -3 with forced diureses with IV hydration to maintain urine output at 2-3ml/kg/hr and mesna 60mg/kg/dose IV at continuous intravenous infusion equal to the dose of cyclophosphamide. For patients with BMI > 35, the cyclophosphamide and mesna dosing will be based on a modified weight half way between the ideal and actual body weights.  
3.7.3.1.3 **Modifications:** For adult patients with MDS who are aplastic after induction therapy, a modified regimen may be used as the preparative regimen including: Fludarabine 30 mg/m2 for 3 days (day -4, -3, -2) and TBI 200cGy day -1.  
For patients who have already received TBI, or exceeded the radiation maximum tolerated dose to preclude TBI for first or second BMT, preferred second choice is busulfan (with goal steady state of 800-1000) with fludarabine or cyclophosphamide at myeloablative dosing or non-myeloablative dosing if recommended for 2nd BMT.
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3.7.3.2 pediatric (<22 years and

3.7.3.2.1 At NCI: TBI will be delivered to a total midplane dose of 12Gy and median lung dose of 6Gy. Radiation will be fractionated twice daily over 4 days (days -9, -8, -7, -6). Opposed lateral fields with lung compensation will be treated to 1.5 Gy for each fraction to a total cumulative dose of 12Gy. Mediastinal fields will be treated AP-PA concurrently with the opposed lateral fields at a dose of 75cGy twice daily to bring the mediastinal dose to 12Gy with a median lung dose of 6Gy. Fractions will be delivered with at least a 6 hour interfraction interval. Gonadal shielding will not be used. Head and neck compensation and other technical modifications may be made at the discretion of the treating radiation oncologist. For patients with CNS positive disease at any time after diagnosis without definitive prior cranial radiation, CNS boost may be delivered unless contraindicated by PI or AI. Adjustments to the boost dose in pediatric patients may be made if contraindicated at the discretion of the PI or AI. The CNS boost will be delivered as a 6 Gy boost in once daily 2 Gy fractions. The boost will be delivered prior to TBI. Patients with sites identified by nuclear imaging may receive a boost to these areas as clinically indicated at any time point pre- or post transplantation.  
3.7.3.2.2 At NCI: Cyclophosphamide: cyclophosphamide 50 mg/kg IV, Days -5, -4, -3, -2 with forced diureses with IV hydration to maintain urine output at 2-3ml/kg/hr and mesna as 24 hour continuous intravenous infusion equal to the dose of cyclophoshamide. For patients with BMI > 35, the cyclophosphamide and mesna dosing will be based on a modified weight half way between the ideal and actual body weights.  
3.6.3.2.3 For patients who have already received TBI, or exceeded the radiation maximum tolerated dose to preclude TBI for first or second BMT, preferred second choice is busulfan (with goal steady state of 800-1000) with fludarabine or cyclophosphamide at myeloablative dosing or non-myeloablative dosing if 2nd BMT.  
3.6.3.2.4At Children's National Medical Center, myeloablative transplantation for FLT imaging may be according to current standard practice including the regimens of: TBI
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3.7.4 gvhd prophylaxis

GVHD prophylaxis at NCI and Univ of Oklahoma will include: methotrexate 10 mg/m2 IV on day +1, and 5 mg/m2 IV Days + 3, 6, 11 and tacrolimus starting Day -1 at dose of 0.02 mg/kg/day continuous infusion (with target level between 5 and 10 ng/ml though up to 15 acceptable). Note, methotrexate dose should be discussed with pharmacy and adjusted for renal or liver dysfunction. Should either agent not be tolerated (for example, posterior reversible encephalopathy from tacrolimus) or other risks put patients at added risk, these may be substituted with standard alternatives (e.g. cyclosporine, sirolimus, cellcept). When the patient is tolerating orals, tacrolimus may be converted to oral administration. Taper of tacrolimus may be initiated on day +100 if there is no evidence of GVHD. Patients should be evaluated weekly after the initiation of tacrolimus taper and tacrolimus taper held or increased for evidence of GVHD (to include diarrhea, mouth sores, unexplained elevation of liver enzymes). If taper is not able to be initiated at day +100, by day +180, a new evaluation should occur to attempt taper. Patients are at high risk of relapse and every attempt should be made in this protocol to permit graft-versus-leukemia effects. For patients after 2nd BMT, taper may be initiated as early as day 45 and the plan will be to taper as rapidly as possible.
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3.7.5 growth factors (gcsf)

GCSF will not be administered unless clinically indicated such as life threatening infection and neutropenia or significantly delayed engraftment (greater than day 18) during the initial transplant phase (less than day 30). A PI or AI must be involved in this clinical decision. While there are data that GCSF accelerates neutrophil engraftment by several days without effects on GVHD and relapse in two randomized, placebo controlled studies [58, 59], there are also data from larger historical cohorts that suggest higher rates of GVHD and relapse without significant differences in neutrophil recovery times. [17, 60] In addition, GCSF has the potential to induce leukemic expansion in AML patients [60] and to alter the imaging evaluation by FLT studies. Notably, GCSF is not standard practice for pediatric patients receiving marrow grafts which are the primary source for pediatric transplantations currently. Given the lack of clear benefit from these studies and the likelihood that patients will recover neutrophils prior to life-threatening infections with matched marrow products, GCSF will not be part of the planned BMT support but rather an option for the few patients who may benefit from its administration (due to delayed engraftment or infections). GCSF may be used after engraftment in the setting of secondary neutropenia (often associated with medications administered post-transplant (e.g. antimicrobials).
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3.8 transplant procedure for 1st

On day 0, the patient will receive the donor product from the selected donor intravenously per DTM standard operating procedures or local institutional guidelines. 3.8.2 Transplant Outcome Determinations x **Neutrophil Recovery:** Designated by the first of 3 consecutive days with an ANC
above 500/mm3.  
x **Platelet Recovery:** Designated by the first of 7 days where the platelet count remains above 20,000/mm3 without transfusion support.  
x **Sustained Donor Engraftment:** Neutrophil recovery associated with complete donor chimerism at day 100. Any patient who dies before day 28 will not be evaluated for engraftment but will be evaluable by FLT.  
x **Primary Graft Failure:** Failure to achieve sustained donor engraftment x **Secondary Graft Failure:** Documented sustained donor engraftment as defined above followed by: 1) severe neutropenia (ANC < 500/mm3) and bone marrow biopsy revealing a cellularity of less than 25% or 2) absence of donor cells in the marrow or blood as demonstrated by a chimerism assay without subsequent improvement occurring either spontaneously or after growth factor treatment. Improvement is defined as ANC
> 500/mm3 consistently. Severe neutropenia with marrow cellularity > 25% is not secondary graft failure.  
x **Aplasia**: Less than 5% marrow cellularity as measured on bone marrow biopsy.
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3.9 post-transplant monitoring (1st and

At NCI, patients will be admitted during the preparative regimen and throughout the peritransplant period until engraftment has occurred and the patient is deemed stable for discharge.  
After discharge, the patient will be monitored as clinically indicated until day +100. 3.9.1 Day 0 to day +100 x History and physical evaluation x CMV, Adenovirus, EBV PCR
x CBC with differential x Acute, Hepatic and Mineral Panels, CK , Uric acid and LDH
x Lipid panel (days +**28, +60, and +100 (+/- 10 days**))
x Tacrolimus level as long as on tacrolimus x Monitoring for aGVHD (documented day +**28, +60, +100**) (see Section 12.10)  
x Monitoring for HHV6 in peripheral blood (all patients) and CSF (for those with leukemia diagnosis) at day +28, +60, +100 +/- 10 days for patients **at NCI only**
x Research blood (day +5, +7, +14, +21, +**28, +60, +100**)
o +/- Allowances for the drawing of research blood is +/-1 day for day +5 and +7, +/- 2 days for day +14, +/- 3 days for day +28, +/- 7 days for day +60, +/- 14 days for day +100, and +/- 30 days for later time points.  
3.9.2 Specific evaluation time points (1st and 2nd BMT)
Patients will be followed as clinically indicated. These evaluations should occur within 10 days of anticipated date for time points less than day 100 and within 1 month for time points after day 100.  
x History and physical evaluation on day **+180, +270, +365**  
x Bone marrow aspirate with studies appropriate to staging of malignant disease, a LP or other biopsy may be required depending upon the diagnosis (to include LP for all AML
and ALL) with chimerism on day **+28, +60, +100, +365**
x Bone marrow biopsy on day **+28, +60, +100, +365**
x Lymphocyte phenotyping (T,B,NK) on day **+28, +60, +100, +180, +270, +365 (**to be performed at NCI including those from outside institutions)
x Pulmonary function tests on day **+ 100, +180, +270, +365**  
x Chimerism of peripheral blood on day +28, +100, +180, +270, +365 for all adult patients
x Immunoglobulins (IgG, IgM, IgA) on day **+28, +100, +180, +270, +365**  
x Acute, Hepatic and Mineral Panels, CK , Uric acid and LDH and lipid on day **+28, +100,**
+180, +270, +365 x cGVHD evaluations on day **+100, +180, +270, +365** (see Sections **12.11, 12.12** and 12.13)
x Research blood on day **+180, +270, +365,** and when clinically significant events such as cGVHD, laboratory suspicion of relapse, or thymus recovery occur.  
x Chest CT **+100, +365**
3.9.3 Long term follow-up guidelines (1st and 2nd BMT)
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ref recommendations [61]

These guidelines are for all NIH patients. All patients may obtain these studies within 1 month of desired date and these may be done in other institutions than NIH with documentation. Please note that these should be performed yearly for 5 years after transplantation either as part of this protocol or in another setting for abnormal results. Pediatric follow-up for CNMC patients will be done as per Children's National Medical Center guidelines.  
x Dental evaluations day **+180, +365**
x Ferritin and iron studies on day +180, +270, +365
x Thyroid studies on day **+180, +270, +365**
x FSH, LH on day **+180, +270, +365**
x Ophthalomological evaluation on day **+180, +365**
x DEXA scan on day **+365**
x Echocardiogram on day **+365**
x **Pulmonary Function Test yearly for 5 years** x GYN examination (female patients) day **+365**
x Urinalysis/protein screening day **+180, +365**
3.9.4 FLT on study evaluations (For patients enrolled on FLT arm at NCI or Children's National Medical Center)
x Imaging Study time points 1st BMT:
o Day -1: 18F FLT + low dose CT o Day +5 or +9: 18F FLT + low dose CT
o Day +28/29: 18F FLT + FDG (not required for Children's National Medical Center site) +
low dose CT
o Day + (GVHD vs. thymus recovery): 18F FLT + FDG + low dose CT at NCI only x Imaging Study time points 2nd BMT:
o Day -1: 18F FLT + low dose CT
o Day +28/29: 18F FLT + FDG (not required for Children's National Medical Center site) +
low dose CT
o Day +60/61: 18F FLT + FDG (not required for Children's National Medical Center site) +
low dose CT
o Day + of relapse and initial response to immunotherapy: 18F FLT + FDG + low dose CT
at NCI only x Laboratory studies for FLT within 24 hours prior :
o Creatinine < 2, ALT and AST within 5 x ULN o TK1 serum measurement x 18F FLT imaging protocol:
o At NCI, Immediately following the IV infusion of 18F FLT, a dynamic PET emission imaging will be performed for 60 minutes, immediately followed by a whole body static image. An additional PET emission imaging acquisition will be performed 120 minutes (+/- 15 minutes) post infusion. A corresponding transmission CT will be acquired immediately before each emission scan. Children's National Medical Center will omit the dynamic protocol.  
3.10 CRITERIA FOR REMOVAL FROM PROTOCOL THERAPY AND OFF STUDY CRITERIA 3.10.1 Criteria for Removal from Protocol Therapy x Completion of 1year protocol therapy x Participant requests to be withdrawn from active therapy x The option to treat the patient after protocol therapy with alternative therapies.  
x Investigator discretion Please note: The AEs as outlined in the treatment protocol will not be collected from that point forward. 3.10.2 Off-Study Criteria x Failure to meet pre-transplant eligibility criteria as defined in section **3.3.1** and **3.7.2**.  
x Patients who do not have a suitable donor or patients for whom the collection product is deemed insufficient without opportunity for further collection of the donor (via PBSC). Patients may proceed on standard of care if this is deemed to be the best medical management by the PI or AI.  
x The donor or recipient refuses to continue therapy x Patients for whom a suitable donor cannot be found x Donor subjects who become decisionally impaired x Lost to follow-up x PI decision to end this study x Death x Completed post-transplant follow up period (5 years)
In all cases, the reason(s) for withdrawal will be documented. In addition, a patient may at anytime be removed from protocol at the principal investigator's discretion, if the P.I. deems the subject to be at unacceptable risk to remain on study. Reasons for this action may include (but are not limited to disease progression with declining organ function/performance status before transplantation; inadequate family/caregiver support; noncompliance.  
Note: Minors should remain on study until they reach the age of 18 and have an opportunity to be consented for the future use of their data and specimens. 3.10.3 Off Protocol Therapy and Off Study Procedure at NCI
Authorized staff must notify Central Registration Office (CRO) when a subject is taken offprotocol therapy and when a subject is taken off-study. A Participant Status Update Form from the web site (http://camp.nci.nih.gov/ccr/welcome.htm) main page must be completed and sent via encrypted email to: NCI Central Registration Office<ncicentralregistration-l@mail.nih.gov. 3.10.4 Participating Site Off-Study Notification The Participant Status Update Form will be supplied by the CCR study coordinator. Send the completed form to the CCR study coordinator.
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4 supportive care 4.1 infection

For a full description of infection prophylaxis for NCI patients please refer to the NIH BMT
Consortium Supportive Care Guidelines at http://intranet.cc.nih.gov/bmt/clinicalcare/guidelines.shtml) which are recommended. Infection prophylaxis for pediatric patients at NCI should follow the NIH BMT Consortium Supportive Care Guidelines found at http://intranet.cc.nih.gov/bmt/clinicalcare/guidelines.shtml or as per local institutional practice. Below is a table summarizing adult recommendations at NCI. Please note that the numbers correspond to the order in which the agents should be attempted (i.e. first choice for PJP is trimethoprim/sulfamethoxazole and the second choice is atovaquone). Whenever possible, these should be followed at University of Oklahoma. Children's National Medical Center pediatric patients should follow Children's National Medical Center guidelines (SOP). Also, please note that these should be followed for patients receiving chemotherapy on this protocol as well.  
| Table 1 Organism                                            | Prophylaxis Recommendations                                                                                                                                             | Timing                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     |
|-------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Pneumocystis                                                | 1) Trimethoprim/sulfamethoxazole                                                                                                                                        |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            |
| jiroveci pneumonia                                          | DS (or pediatric dose equivalent)  once a day three times weekly                                                                                                        |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            |
| 2) Atovaquone 1500 mg daily  3) Inhaled Pentamidine monthly | Start after immune  reconstitution or by day 28.  Stop > 6 months after  immunosuppression  withdrawal without evidence  of GVHD and greater than 1  year after HSCT.   |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            |
| Fungus                                                      | 1) Caspofungin  2) Voriconazole or Posaconazole  (Check levels and target dose to  Vori > 2 for prophylaxis, >4 for  treatment; posa > 1000) 3) Liposmal amphotericin B | Start caspofungin with  preparative regimen for mold  prophylaxis and continued  until neutrophil recovery and  not on prednisone equivalent  exceeding 0.8mg/kg/qod. Plan  to switch to or add azole for  evidence of invasive mold.  Note: azoles will need to  modify tacrolimus dosing.  History of presumed or proven  invasive fungal infection  necessitates continuous  antifungal coverage throughout  HSCT until off  immunosuppression unless  there is a significant  complication of therapy. |  
| Organism                        | Prophylaxis Recommendations                                                                                                                                                                      | Timing                     |
|---------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------|
| Virus                           | 1) Acyclovir 800 mg po BID or                                                                                                                                                                    | Positive HSVI/II serology, |
| 250mg/m2 IV q12.                | history of Herpes Simplex or  varicella infection, or CMV  infection. Starting on day -4.                                                                                                        |                            |
| Bacteria                        | 1) Ceftazidime 2 gm IV q 8 hrs                                                                                                                                                                   | Start at neutropenia,      |
| 2) Levofloxacin 500 mg PO or IV | discontinue after resolution of  neutropenia. Note: patients  should remain on PCN  (encapsulated organism  prophylaxis) until off  immunosuppression > 6  months and without ongoing  or cGVHD. |                            |  
This study will also monitor for viral infections to include adenovirus, EBV, CMV, and HHV6. If the patient develops chronic GVHD, he/she will receive penicillin V for prophylaxis against bacterial infections. He/she will undergo vaccination against *S. pneumoniae* and *H. influenzae*,
beginning 12 months after transplantation as per the NIH BMT Consortium Supportive Care Guidelines. Administration of live vaccines will be avoided during the first 2 years after transplantation or in patients with ongoing chronic GVHD or immunosuppression. Immunizations with inactive vaccines will be administered starting 6 months after withdrawal of immunosuppression or one year whichever comes first with the exception of influenza vaccine and prevnar. Response to vaccines is an exploratory endpoint on this trial. Vaccine response is one of the best markers of T and B cell function (requiring CD4+ T cells to convert B cells to IgG). 4.1.1 Infectious Disease Surveillance: Adenovirus PCR will be sent weekly during engraftment. For patients who have Toxoplasmosis titer positive (at any point prior to HSCT), toxoplasmosis will be monitored weekly during engraftment at NCI only. HHV6 will be monitored in blood and CSF at day +28, +60, +100 +/- 10 days (for those undergoing CSF evaluation at these time points at NCI only).
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4.2 management of engraftment syndrome

Engraftment syndrome is characterized by noninfectious fever, rash, and vascular leak causing non-cardiogenic pulmonary edema, weight gain, and renal. Diagnostic criteria and a treatment schema for engraftment syndrome are included in Section 12.9.
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4.3 treatment of graft-versus-host disease

In patients in whom GVHD is suspected, standard clinical criteria and biopsy findings (when clinically indicated) will be used to establish the diagnosis. Acute GVHD will be assessed according to 1994 Consensus Conference on Acute GVHD Grading criteria [62]. Chronic GVHD will be assessed according to 2005 Chronic GVHD Consensus Project [62]. See Section 12.12 for details concerning the assessment of acute and chronic GVHD.  
Patients with clinical Stage 1 or 2 (Grade I) GVHD of the skin without any other organ involvement will be treated with a topical corticosteroid cream.  
In general, patients with t Grade II acute GVHD will be treated with high-dose, systemic corticosteroids. Patients who fail to respond to corticosteroids within 3 days will be considered for second-line immunosuppressive therapy. Given that the patients in this protocol are young adults with the potential for rapid response to immunosuppressive agents, every effort will be made for rapid taper of high dose steroids and rescue with other, less immunosuppressive agents. As standard treatment of steroid-refractory acute GVHD and of chronic GVHD is often unsuccessful, treatment of either can include standard systemic or local therapies. This can specifically include autologous serum eye drops when recommended by ophthalmology for ocular GVHD per NIH Consensus Guidelines.
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4.4 menses suppression and contraception

For a full description of menses suppression and contraception guidelines for NCI please refer to the NIH BMT Consortium Supportive Care Guidelines at: http://intranet.cc.nih.gov/bmt/clinicalcare/guidelines.shtml Pre-menopausal women who have not undergone hysterectomy will receive Lupron within 2 weeks of preparative regimen. Adult women will receive the 4 month (30 mg) preparation and females < 18 years old will receive the 3 month (11.25 mg) preparation. Subsequent therapy will continue until platelet recovery after transplantation (> 50,000/μl without transfusion) and may include repeat Lupron dosing or Lo-Ovral (300μg norgestrol and 30 μg ethinyl estradiol) 1 tablet daily without placebo tablets. If Lo-Ovral is started when the patient is bleeding and bleeding persists for more than 2-3 days, increase the dose to 1 tablet twice daily. If bleeding persists for more than an additional 2-3 days (total 5 to 6 days after starting hormones), consult the NIH Gynecology Consult Service. However, Lo-Ovral will be stopped as soon as possible and patients who receive it may be analyzed separately from the lupron-treated female cohort. Female transplant recipients will be advised to use an effective form of contraception for at least 1 year after transplantation, and to have their male partners use condoms.  
Male transplant recipients will be advised to use contraception, preferably condoms, for 1 year after transplantation.
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4.5 blood product support

Patient's blood counts will be monitored daily during the transplant hospitalization. Patients will receive packed red blood cells and platelets as needed to maintain **Hb > 8.0** gm/dl at NCI and CNMC and Hb> 7.0 gm/dl at Univ. of OK as per institutional practice, and **plts > 10,000/mm3**
(or higher, if clinically indicated) unless the **patient is not group 0 blood type and the donor** is, then Hb> 9.4 until after day 15 at NCI and per institutional practice. All blood products, with the exception of the stem cell product and Donor Cell Infusion (DCI), will be irradiated and leukoreduced as per DTM protocol at NIH, and per the Blood Bank at Children's National Medical Center or the University of Oklahoma. The patient, who is seronegative for CMV and whose donor is seronegative, should receive CMV-negative blood products whenever possible. Notably, all future transplant recipients should receive transfusion only of irradiated blood and cellular blood products. Transfusion of irradiated blood and cellular blood products will continue to at least one year after transplantation. Patients receiving immunosuppressive medication will continue to have all blood and cellular blood products irradiated until discontinuation of immunosuppressive treatment. Patients sensitized to HLA or platelet specific antigens should receive HLA matched apheresis platelet collections.
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4.6 nutritional support

If mucositis or GVHD prevent adequate PO intake, nasogastric tube feeds or parenteral hyperalimentation will be instituted and discontinued with input from the Pharmacy and Nutrition Departments at NCI. Oral intake will resume when clinically appropriate under the supervision of the dietary service of the Clinical Center. At Children's National Medical Center and University of Oklahoma, this should be done as per institutional practice.
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4.7 anti-emetic usage

Anti-emetic usage will follow recommendations from the NIH CC Pharmacy or as per institutional practice.
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4.8 intravenous immune globulin (ivig)

IVIG administration will follow Guidelines for Infection Management in pediatric standard practices due to the difficulties of avoidance of viral exposures in pediatric patients. For an IgG values less than 400 mg/dl, 500 mg/kg IVIG will be administered intravenously as per institutional guidelines. Administration of IVIG may also occur if clinically indicated (e.g. setting of severe viral infection and vascular leak).
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4.9 hepatic function support

All patients will receive ursodeoxycholic acid (Ursodiol) for the prevention of hepatic complications after allogeneic stem cell transplantation. Ursodiol will start by day -4 prior to the transplant and will continue until day +100 post-transplant (or longer if clinically indicated). Patients weighing less than 90 kg will receive 300mg orally twice daily (600mg total dose each day). Those patients weighing more than 90 kg will receive 300 mg orally each morning and 600 mg orally each evening (900 mg total dose each day). CNMC pediatric patients will be dosed as per Children's National Medical Center guidelines. The primary hepatic complication after transplantation is sinusoidal obstructive syndrome (also known as veno-occlusive disease or VOD). This has been defined using Seattle criteria as: unexplained weight gain, hepatic tenderness, and jaundice; these criteria have been refined by Baltimore to include: total bilirubin > 2, and 2 of the following: hepatomegaly, weight gain 5%, and ascites. Because no drugs are approved to treat VOD, and access to defibrotide, the only drug that has demonstrated promise, is limited, every effort will be made to optimize management to minimize the severity of VOD. Throughout the first 30 days of transplantation, management should include equal ins and outs without weight gain. Salt loads should not occur in the absence of clear benefit to the patient (e.g. sepsis). Close monitoring for this condition and immediate fluid restriction, use of oncotics and lasix (see Section **4.11**), and salt removal from infusions should be instituted.
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4.10 central nervous system prophylaxis

Patients with history of CNS positive disease will receive CNS boost.
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4.11 fluid management during first

Patients should be well-hydrated prior to stem cell infusion (within 12 hours prior). Following stem cell infusion, weights and abdominal girths should be followed closely. Patients should be kept EVEN in fluid status with every attempt to minimize salt and fluid loading. High salt and fluid loads have been implicated in the physiology that leads to severe engraftment syndrome and VOD. Lasix may be used to maintain this. In addition, an oncotic (e.g. blood, IVIG, platelets, albumin) may be used followed by lasix to preserve renal function while optimizing fluid management.
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4.12 ancillary support

Other supportive care to include: physical therapy, occupational therapy, psychosocial therapy as deemed clinically appropriate.
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4.13 supportive care for relapse

Patients who experience relapse on this protocol may receive therapy for relapse to include intrathecal or systemic chemotherapy, donor lymphocyte infusions or other treatments.
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4.14 additional cell infusions for

Patients who experience graft failure (primary or secondary), impending graft failure (falling counts and/or donor chimerism after initial engraftment in the absence of relapse), or poor graft function posttransplant (hypoplastic bone marrow, complete or near-complete donor chimerism, absence of relapse or cytopenias clinically thought to be caused by medication or infection, severe cytopenias in at least two lines [Hb < 10 g/dL, platelet count < 30 x 109/L, neutrophil count < 1.0 x 109/L] lasting at least 2 weeks beyond day +14 and with transfusion requirement) will be eligible to receive a CD34+-selected cell boost. The same donor for the most recent BMT must be used. The donor will have a GCSFmobilized peripheral blood stem cell product collected and CD34+-selected using the Miltenyi CliniMACS® system prior to infusion into the patient. A fresh product will be preferred although a cryopreserved product will be acceptable.
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5 data collection and evaluation

Data will be prospectively and entered into an NCI, CCR C3D Database from each institution. After obtaining Informed Consent, a file will be created in the database with standardized forms. The database will maintain complete records on each patient including any pertinent supplementary information obtained from outside laboratories, outside hospitals, radiology reports, laboratory reports, or other patient records. Patient records will be summarized at key time points in protocol visit notes at: pre-transplant, day 28, 60, 100, 6 month, 9 month, 12 months, and yearly post-transplant. These protocol notes will serve as the primary source material from which data will be collected and research analyses will be performed. These will be reviewed by the Adjunct PI or site PI and research nurse/coordinator for accuracy and include interpretation of the events prior to this note. This is done to minimize the day to day confusion that may occur (e.g. patient has diarrhea, thought initially to be GVHD but later determined to be CMV). The following will be used for source data and wherever possible will use the interpretations from the protocol visit notes as detailed below:
x Eligibility Checklist: To be completed at study entry and forwarded to the protocol research nurse.  
x Patient history and physical examinations while on study (protocol visit notes) x Patient flow sheets (protocol visit notes)
x Patient pathology, radiology, chimerism data, laboratory evaluations. (interpreted in protocol visit notes)
x Relevant data will also be sent to the Center for International Bone Marrow Transplant Registry (CIBMTR). This will include outcome data and patient characteristics, but do not include adverse events.  
x Protocol Deviations/Unanticipated problems (UPs): Any protocol deviations or UPs should be directly reported to the PI, PC, or AI. These will be submitted to each site's respective IRB. Outside institutions are required to send all protocol deviations and UPs to the Adjunct PI to be submitted to NCI IRB.  
x Confidentiality Protection: Prior to analysis, all identifiers will be removed and a random number ascribed to protect patient confidentiality.
The PI will be responsible for overseeing entry of data into an in-house password protected electronic system and ensuring data accuracy, consistency and timeliness. The principal investigator, associate investigators/research nurses and/or a contracted data manager will assist with the data management efforts. All data obtained during the conduct of the protocol will be kept in secure network drives or in approved alternative sites that comply with NIH security standards. Primary and final analyzed data will have identifiers so that research data can be attributed to an individual human subject participant. All adverse events, including clinically significant abnormal findings on laboratory evaluations, regardless of severity, will be followed until return to baseline or stabilization of event. Patients will be followed for adverse events for at least 30 days after removal from study treatment or until off-study, whichever comes first. An abnormal laboratory value will be considered an AE if the laboratory abnormality is characterized by any of the following:  
x Results in discontinuation from the study x Is associated with clinical signs or symptoms x Requires treatment or any other therapeutic intervention x Is associated with death or another serious adverse event, including hospitalization. x Is judged by the Investigator to be of significant clinical impact
x If any abnormal laboratory result is considered clinically significant, the investigator will provide details about the action taken with respect to the test drug and about the patient's outcome.  
End of study procedures: Data will be stored according to HHS, FDA regulations, and NIH Intramural Records Retention Schedule as applicable.  
Loss or destruction of data: Should we become aware that a major breach in our plan to protect subject confidentiality and trial data has occurred, the IRB will be notified. 5.1.1 Adverse event recording  
x Grade 1 adverse events will not be recorded x All serious adverse events regardless of the attribution will be recorded x If adverse events are not serious and possibly, probably, or definitely attributable to either Lupron or FLT imaging then they will be recorded.
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5.2 response criteria 5.2.1 primary

5.2.1.1 To determine whether Lupron leads to accelerated B cell recovery after allogeneic hematopoietic stem cell transplantation.  
5.2.1.2 To determine if FLT imaging can be used safely to evaluate for engraftment of transplanted cells.  
5.2.1.3 B cell recovery will be assessed on day 180 for comparison between two groups: those who have received Lupron and those who have not.
5.2.1.4 Patients will be evaluated for complete clinical response to primary malignancy at day
+28, day +100, and day +365; those with and without Lupron will be compared. Complete response will include M1 marrow and no evidence of disease by flow cytometry, cytogenetics or FISH testing in all sites evaluated (lumbar puncture, bone marrow). Minimal residual disease will be used to denote < 5 % blasts in marrow but detection of disease by one of these measures.  
5.2.2 Secondary Endpoints 5.2.2.1 Thymus studies with CT scan thymic index, flow cytometry measurements of naïve T
cells.  
5.2.2.2 Assess feasibility and tolerability of Lupron in male HSCT recipients. Assess recovery of testicular and ovarian function after HSCT.  
5.2.2.3 Rates and details of acute and chronic GVHD in Lupron vs. no-Lupron HSCT
recipients.  
5.2.2.4 Rates and details of infectious episodes in Lupron vs. no-Lupron HSCT recipients. 5.2.2.5 Recovery of other populations to include: neutrophils, red blood cells, platelets, and other cell subsets (e.g. monocytes) will be assessed and compared between groups.  
5.2.2.6 Transplant related mortality, incidence of graft rejection, disease-free and overall survival will also be collected.  
5.2.2.7 The FLT images will be used in exploratory fashion to determine if the day +5 or day
+9 image can be used to predict engraftment of donor cells by comparing the uptake of FLT at day -1 with day +5 or day +9 and day +28 (post-engraftment) and correlating this will leukocyte recovery for first BMT. Difference between patients who receive bone marrow or peripheral blood stem cell products will also be explored. For 2nd BMT, exploratory analyses will compare day -1 FLT image with relapse rates, and day +28 and +60 image abnormalities suggestive of relapse with relapse rates.
5.2.2.8 Evaluate thymic reconstitution by FLT (1st BMT).  
5.2.2.9 Evaluate whether FLT vs. FDG could distinguish GVHD vs. infection and relapse vs.  
healthy donor engraftment (1st and 2nd BMT).  
5.2.2.10 Evaluate the safety of FLT imaging in the peritransplant period (1st and 2nd BMT).
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5.3 toxicity criteria

Adverse events on this study will be reported using the NIH mechanism of NCI Common Terminology Criteria for Adverse Events. This study will utilize the CTCAE version 4.0 for toxicity and adverse event reporting. A copy of the CTCAE version 4.0 can be downloaded from the CTEP home page (http://ctep.info.nih.gov) under reporting guidelines. All appropriate treatment areas should have access to a copy of the CTCAE version 4.0. An adverse event will be defined as any novel, concerning medical condition or worsening of a previously existing medical condition. Please refer to section 8 for details on adverse event collection.
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6 statistical section 6.1 subject

Non-pregnant female and male subjects from all racial and ethnic groups are eligible for this trial if they meet entry criteria. Although every effort will be made to incorporate individuals representative of the national population with the eligible malignant diseases, issues of limited accrual may not permit this study to explore the impact of race or gender. Should the analysis suggest that there is a difference observed across race or gender, accrual may be expanded or additional studies will be designed to address these questions. 6.1.1 Statistics and Feasibility The primary objectives of this study are:  
1) To determine if there is a difference in the percentage of B cells which recover at 12 months between males who are randomized to receive Lupron or not, and to estimate the fraction of females, all of whom will receive Lupron, who have B cells which recover at 12 months.  
2) To evaluate engraftment kinetics to estimate the relationship between level of uptake of FLT at a day +5 scan and the day at which neutrophils recover to >500, statistical analyses were performed for this secondary endpoint.
6.1.1.1 Statistics on FLT
This subset of the total male and female patients receiving marrow transplants will undergo pretransplant conditioning and FLT imaging agent. The patients will have their level of FLT uptake evaluated at a day +5 scan, and will be followed until they recover their neutrophils to >500. A Spearman correlation coefficient will be used to estimate the degree of association between these two parameters. With 13 evaluable patients, who have both a day +5 scan level of uptake and a known number of days to recovery of neutrophils, a correlation coefficient will be estimated in a pilot fashion; a 0.10 one-sided Fisher's test of whether the correlation coefficient has a value of 0.85 rather than a null value of 0.50 can be performed with 80% power if 13 patients are included in the evaluation. Fifteen patients will be studied in this cohort to obtain 13 fully evaluable images given that 2 may have early events that preclude day 28 evaluation. Patients enrolled on this arm of the protocol who do not complete a day 28 evaluation may be substituted to achieve 13 evaluable patients unless the patients are not evaluable due to toxicities attributed to FLT. Interim safety analysis of AEs attributed to FLT imaging will be performed on the first 7 patients. Should greater than 1 of these 7 patients not engraft and require a back-up product, the FLT arm of this protocol will be halted. Notably, patients who experience early relapse may be analyzed separately for some statistical evaluations as leukemia cells upregulate TK1 and may alter images obtained around the time of relapse. An additional eight evaluable patients will permit us to evaluate the day +9 scan rather than a day +5 scan and correlate this with engraftment and explore differences between peripheral blood stem cell source and bone marrow stem cell source. These demonstrate different kinetics of marrow recovery (with engraftment of neutrophils typically 14 days earlier with peripheral blood stem cells). Finally, not only will these kinetics help us to understand the biology of stem cell transplantation, they will also serve as a critical piece of information for a future study with cord blood as the stem cell source (which has even greater delay in neutrophil recovery, typically engrafting 2 weeks later than a marrow product). Because the interim analysis confirmed safety in adults, up to 10 pediatric patients will now be enrolled at Children's National Medical Center. These patients will be imaged on day -1, day +9, and if possible, day +28. Analyses will mirror the adult statistics and also permit exploratory comparisons between pediatric and adult patients. The same safety stopping rules will apply: should greater than 1 of these 7 patients not engraft and require a back-up product, the FLT arm of this protocol will be halted.  
In addition, because there is a suggestion that FLT may predict relapse (showing relapse in 4/4 imaged patients to date, and prediting relapse in ¾), FLT will be incorporated into the evaluation of all patients in arm 2 (who are undergoing 2nd HSCT for relapse after 1st HSCT). This will be exploratory analyses to use FLT to try to predict response of therapy (in the day -1 image) and relapse in subsequent images in this population at high risk for relapse (80% historically). For patients who do relapse after this 2nd HSCT, a scan may be performed pre- and after therapies to treat this relapse to explore the effect of immunotherapy on FLT uptake.
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6.1.1.2 statistics on lupron

For 1st BMT the larger cohort of patients (of which some of these patients may undergo FLT
imaging as well) will be eligible male subjects who will be randomized in a 1:1 ratio to receive Lupron vs. no Lupron prior to receiving their pre-transplant preparative regimen. The objective will be to determine if there is a difference in the percentage of these subjects who recover their B cells by 12 months according to whether they receive Lupron or not. With 17 evaluable patients per arm (34 total evaluable patients), there is 80% power to detect a difference equal to one standard deviation (one SD effect size) between the two arms using a 0.05 two-sided twosample t-test (that is, that the difference in the means of the two groups differs by one SD,
assumed to be the same in both groups). In practice, if the data from the two groups are not normally distributed (p<0.05 by a Shapiro-Wilks test in either group), then a Wilcoxon rank sum test will be used. Since only patients who have not relapsed by 12 months would be eligible for this definitive evaluation, up to 68 patients may need to be enrolled in order to obtain this definitive evaluation.  
For 1st BMT, a second cohort of patients (which may include FLT exposed patients) will be eligible female subjects who will each receive Lupron prior to receiving their pre-transplant preparative regimen. The objective in this cohort will be to estimate the fraction of patients who recover their B cells by 12 months. If there are 30 eligible female patients who have this value determined at 12 months, then a two-sided 95% confidence interval could have width of approximately +/- 4 if the standard deviation was equal to 10. Since the female patients may also relapse prior to 12 months and not provide a definitive value for evaluation, up to 50 females will be enrolled into this cohort to try to ensure that at least 30 are evaluable.  
For 1st BMT, for the female and male cohorts, all enrolled patients will be evaluated at the latest time point possible for a variety of immune reconstitution parameters. Thus, even if they are unable to ultimately contribute information to the definitive evaluation at 12 months, the values that are obtained at any time point will be used to estimate the recovery at various time points prior to 12 months. In order to complete accrual to these cohorts, up to 68+50=118 patients may need to be enrolled. If 2-3 patients per month are able to be accrued onto this trial, accrual should be completed within 3 to 5 years. Patients who do not undergo allogeneic transplantation may be substituted. Toxicity of FLT and Lupron in this population will be evaluated in all exposed patients.  
For 1st BMT, exploratory analyses will evaluate other leukocyte subsets of immune reconstitution with and without Lupron (including transitional B cells, dendritic cells, recent thymus emigrants, naïve and memory T cells), disease control by treatments pre- and peri-transplant, whether FLT can identify relapse disease, or GVHD. The effect of cell dose and age on lymphoid engraftment will also be explored. In addition, evidence of functional immune recovery as per vaccine response will also be explored. For patients to receive benefit from vaccines after myeloablative HSCT, the immune system must mount a durable response. If the patient receives a vaccine without evidence of a rise in antibody titers, it is possible or even likely that the patient does not receive benefit from this vaccine. In pediatric HSCT recipients in many institutions, it is standard practice to test for a vaccine response and repeat vaccinations when one is achieved to maximize the potential for long-lasting immunity to this pathogen. In adult HSCT recipients, this is not standard of care because both there is less likelihood that older adults can achieve this after myeloablative transplantation and high likelihood that residual plasma cells (of host origin) will respond after non-myeloablative transplantation which may not be meaningful for long-lasting immunity. Because the immune modulation of lupron may improve the immune system of these young adults similar to those often achieved in pediatric patients, the optimal approach for long lasting immunity is to attempt to determine a functional benefit to the vaccines, to give the vaccine to the patient when they can respond and develop durable benefit (prophylaxis from infections).  
Interim analyses will be conducted on FLT treated patients after 7 patients have completed day 28 imaging and on the lupron treated vs. untreated male subjects after 15 have been enrolled to each arm to evaluate for safety (lupron and FLT) and efficacy (lupron). Using the CIBMTR (international registry data), the expected survival is <30% for acute myelogenous leukemia and 30% for acute lymphoid leukemia at 2 years. Because 10% of these patients die of nonrelapse causes, this means that the expected relapse rate is 60% for similar patients, suggesting that our relapse rate of less than half this is considerably better. The relapse rate for patients after 2nd BMT is even higher, so these patients will need to be analyzed separately from patients who have only undergone 1 BMT. These patients will be added via amendment L, which will have the following characteristics: males and females who have previously failed a transplant will be enrolled onto the trial and receive Lupron. The patients will be followed for progression and for survival. With 16 patients, there would be 82% power to detect an improvement from 60% who relapse by one year to 30% who relapse by one year (using data for 2nd BMT where the relapse rates exceed 60%), using an exact binomial test with a 0.10 one-sided significance level. In practice, the fraction of the 16 evaluable patients who survive without relapse to one year will be estimated; this fraction will be reported along with appropriate confidence intervals, such as a one-sided 90% interval to show if the fraction is below 60%, as well as a two-sided 95% confidence interval to help describe the observed fraction of patients who are relapse free at one year. In addition, a Kaplan-Meier curve of relapse free survival will be constructed to estimate the relapse free probabilities at various time points. To ensure that patients are not continued to enroll onto this arm in the event that results are not adequate, the following early stopping rule will apply: if among the first 8 patients enrolled, 0 of 8 are able to survive to one year, then no further patients will be enrolled onto this trial as soon as this can be determined, as this will indicate worse than expected results: the upper one sided 90% Confidence interval on 0/8 is 0.25, which would indicate that this result would not be consistent with tolerable results.
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7 human subjects protections 7.1

All patients with eligible malignant diseases who may benefit from allogeneic stem cell transplantation are eligible for this trial. All ethnic groups and both sexes may be enrolled. Ethnic enrollment may vary as ethnic groups afflicted by these diseases may vary. Pregnant females will be excluded from this trial. It is unlikely that our patient numbers will permit statistics of individual ethnic or age groups, however this will be attempted should numbers permit.
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7.2 participation of children

In addition, this trial is open to pediatric as well as adult patients; however, at Univ of Oklahoma only adult patients will be enrolled and at Children's National Medical Center only pediatric patients will be enrolled. Patients > 15 years old and/or > 9 years and after Tanner stage 2 are eligible at NCI, because these; patients who are pre-pubertal are unlikely to receive benefit (and may be harmed) by exposure to Lupron. At Children's National Medical Center pediatric patients > 4 years of age will be eligible. Donors less than 2 years of age will not be included on this protocol. Physicians, nurses, and multidisciplinary support teams of the POB, NCI, and CC at NIH, and Oklahoma Transplant Team, Children's Hospital, Stephenson Cancer Center, and HCA at Univ of Oklahoma, and the BMT team, Blood Bank at Children's National Medical Center will provide patient care. The staff of the POB has expertise in the management of children with complex oncologic disorders and complications of therapy. Full pediatric support and subspecialty services are available at the NIH CC. Note: arm 2 is for adults only at NIH.
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7.3 participation of subjects unable

.Re-consent may be necessary and there is a possibility, though unlikely, that subjects could become decisionally impaired. For this reason and because there is a prospect of direct benefit from research participation (section 7.4), all recipients VXEMHFWV  DJH  ZLOO EH RIIHUHG WKH opportunity to fill in their wishes for research and care, and assign a substitute decision maker on the "NIH Advance Directive for Health Care and Medical Research Participation" form so that another person can make decisions about their medical care in the event that they become incapacitated or cognitively impaired during the course of the study. Note: The PI or AI will contact the NIH Ability to Consent Assessment Team for evaluation. For those subjects that become incapacitated and do not have pre-determined substitute decision maker, the procedures described in MAS Policy 87-4 for appointing a surrogate decision maker for adult subjects who are (a) decisionally impaired, and (b) who do not have a legal guardian or durable power of attorney, will be followed. Donor subjects who become incapacitated or cognitively impaired during the course of the study will come off study since there are no direct benefit to the donor subjects.
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7.4 evaluation of benefits and

Patients are likely to obtain direct benefit from the HSCT as the eligible diseases for this protocol are aggressive and likely to lead to death without HSCT. The transplantation preparative regimen and GVHD prophylaxis are standard of care for patients of these ages with these diseases. These regimens carry an acceptable risk of direct toxicities including: organ toxicity (lung, heart, kidney, endocrinological, neurological, and liver), immunosuppression with increased risk of infections, potential for graft versus host disease. The patients will be monitored for toxicities using the current best practice guidelines and will receive prophylaxis for those complications that we can prevent (e.g. infectious, mesna for cyclophosphamide to minimize kidney toxicity). See section 10 for details. The greatest risk to mortality remains relapse in this population. Thus, a myeloablative regimen has been chosen that confers the greatest potential for disease control. Lupron may not impact on disease control, or it may improve disease control by decreasing androgen levels (leukemias often have androgen receptor and may respond to this substrate) and through enhanced post-transplant lymphocyte reconstitution (with potential to improve GVL). 7.4.1.1 FLT risks to exposed recipient The greatest risks of clinical FLT scans to patients enrolled on imaging studies to date have included bruising at the injection site; no significant adverse events were reported in the literature. Please see section **1.2.7** for details and toxicities at much higher doses of FLT than patients on this protocol will receive, none of which are expected. The one theoretical risk to patients undergoing HSCT is that the newly dividing hematopoietic stem cells could be affected by FLT delaying engraftment or contributing to failure to engraft. However, the actual radiation dose to which the bone marrow is exposed is equivalent to the dose received in an abdominal CT with contrast, a test routinely performed for clinical indications without adverse effects on engraftment. Stopping rules are in place for nonengraftment.
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7.4.2 risks in relation to

Healthy HLA-matched donors will be co-enrolled onto this study. The collection aspect of this protocol is not investigational. There is potential benefit for donors, as they may derive psychological benefit from participating in a clinical trial designed to improve the health of the recipient of the prior transplant donation. Other potential benefits include the diagnosis of previously unknown illnesses (such as viral hepatitis) at the time of donor screening.
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7.4.3 pediatric patients

Pediatric patients are eligible to participate in this trial if they meet criteria including a disease for which BMT is the recommended therapy. The diseases addressed in this protocol are the current diseases recommended for transplantation by the national cooperative groups (Children's Oncology Group). The preparative regimen is one of the standard recommended regimens. Pediatric patients at Children's National Medical Center are now eligible for FLT, because this testing did not increase the risk to recipients in the adult study (14 adult patients have undergone FLT without attributable toxicities at NCI). Female patients who have begun puberty will receive the standard of care for menses prophylaxis, lupron at the dose of 11.25 mg for patients < 18 years. Thus, the only additive risk to pediatric patients beyond standard of care is the exposure of male patients to lupron. Given that female children have tolerated this agent well, the risks of lupron in male subjects are expected to be mild and reversible (the drug will clear in 3 months and not redosed in this study in male subjects). The potential benefits include: 1) improved immune reconstitution, 2) potential anti-tumor effects, and 3) possible benefit for preservation of fertility.
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7.5 consent and assent process

Informed consent will be obtained from all patients, and/or the patient's parents or legal guardian (if he/she is < 18 years of age) and matched related donors prior to entry.. The procedures and treatments involved in this protocol, with their attendant risks and discomforts, potential benefits, and potential alternative therapies will be carefully explained to the recipient and/or parent/guardian. Where deemed appropriate by the clinician and the child's parent(s) or guardian, the child will also be included in all discussions about the trial and age-appropriate language will be used to describe the procedures and tests involved in this study, along with the risks, discomforts and benefits of participation. Verbal assent will be obtained as appropriate for children ages (12 years and older) and the parent or guardian will sign the designated line on the informed consent attesting to the fact that the child has given assent. Children under the age of 12 years will not be required to provide assent as they typically do not have the cognitive ability to fully understand the nature of research. The consent/assent process will be documented in the child's medical record, including the assessment of the child's ability to provide verbal assent. All children will be contacted after they have reached the age of 18 to determine whether they wish to continue on the trial and informed consent will be obtained from them at that time. Note that NMDP consents will be used for unrelated donor patients. The investigators are requesting a waiver from the IRB to allow only one parent to sign the informed consent to enter a child on the protocol. Because many patients must travel to the NIH from long distances at substantial expense, requiring both parents to be present for the consent process could be a financial hardship for many families. When guardianship status of the child is uncertain, a social worker will be asked to investigate and, if necessary, seek documentation of custody status. In situations where there is joint custody of a child, both parents must sign consent. If only one parent can be present at NIH, the other parent's consent can be obtained by telephone via the procedure described in section 7.5.1 Similarly, the procedures and treatments involved in this protocol, with their attendant risks and discomforts, will be carefully explained to the matched unrelated donors at the respective donor center as required by NMDP. The original signed informed consent goes to Medical Records; a copy will be kept for the research record. The Central Registration Office (CRO), Data Management Section will also retain a copy of the informed consent document. A copy of the signed informed consent document will also be given to the recipient (and matched related donor, when applicable). b) The Central Registration Office (CRO) will ascertain the date of IRB approval before registering the first patient. The Principal Investigator and/or an authorized designee will consent the patient and matched related donor (when applicable) and will be available to answer all patient questions. If any new information becomes available relating to risks, adverse events, or toxicities, while patients are participating in this protocol, this information will be provided orally and/or in writing to all enrolled and prospective patient participants. Documentation will be provided to the IRB and if necessary the informed consent amended to reflect relevant information. The possible risks and benefits of this trial will be explained in layman's terms for patients and guardians. For patients who elect to enroll, consent will be obtained. A signed consent must be obtained by patient or legal guardian prior to study entry. The PI, an Associate investigator, or an appropriate designee will answer all questions regarding this medication and the purpose of the trial prior to obtaining consent
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7.5.1 consent for minors when

When a pediatric subject reaches age 18, continued participation (including ongoing interactions with the subject or continued analysis of identifiable data) will require consenting of the now adult with the standard protocol consent document to ensure legally effective informed consent has been obtained. We request waiver of informed consent for those individuals who have completed their participation in the research study and subjects who have been lost to follow-up Requirements for Waiver of Consent consistent with 45 CFR 46.116 (d):
1) The research involves no more than minimal risk to the subjects. a. Analysis of samples and data from this study involves no additional risks to subjects.  
3) The research could not practicably be carried out without the waiver or alteration. a.  
Considering the length of time between the minor's last contact with the research team and their age of majority, it will likely be very difficult to locate them again. A significant reduction in the number of samples analyzed is likely to impact the quality of the research.  
4) Whenever appropriate, the subjects will be provided with additional pertinent information after participation. a. We only plan to request a waiver of reconsent for those subjects who have been lost to follow-up or prior to amendment T, have been taken off study prior to reaching the age of majority.
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7.5.2 telephone consent and re-consent

The following procedure may be used in cases of re-consent or for intial consent in cases where the consent of both parents of a minor participant is required as discussed in section 7.5. the informed consent document will be sent to the subject. An explanation of the study will be provided over the telephone after the subject has had the opportunity to read the consent form. The subject will sign and date the informed consent. A witness to the subject's signaturewill sign and date the consent. The original informed consent document will be sent back to the consenting investigator who will sign and date the consent form with the date the consent was obtained via telephone. A fully executed copy will be returned via mail for the subject's records. The informed consent process will be documented on a progress note by the consenting investigator. 7.5.3 Informed consent of Spanish speaking subjects We anticipate the enrollment of Spanish speaking research participants into our study. The IRB approved full consent document will be translated into that language in accordance with the Clinical MAS Policy M77-2. 7.5.4 Short form consent process for other non-English speaking patients If there is an unexpected enrollment of a research participant for whom there is no translated extant IRB approved consent document, the principal investigator and/or those authorized to obtain informed consent will use the Short Form Oral Consent Process as described in MAS Policy M772, OHSRP SOP 12, 45 CFR 46.117 (b) (2). The summary that will be used is the English version of the extant IRB approved consent document. Signed copies of both the English version of the consent and the translated short form will be given to the subject or their legally authorized representative and the signed original will be filed in the medical record. Unless the PI is fluent in the prospective subject's language, an interpreter will be present to facilitate the conversation. Preferably someone who is independent of the subject (i.e., not a family member) will assist in presenting information and obtaining consent. Whenever possible, interpreters will be provided copies of the relevant consent documents well before the consent conversation with the subject (24 to 48 hours if possible). We request prospective IRB approval of the use of the short form process and will notify the IRB at the time of continuing review of the frequency of the use of the Short Form.
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8 safety reporting requirements/data and

Any untoward medical occurrence in a human subject, including any abnormal sign (for example, abnormal physical exam or laboratory finding), symptom, or disease, temporally associated with the subject's participation in research, whether or not considered related to the subject's participation in the research.  
8.1.2 This study incorporates a standard myeloablative bone marrow transplantation platform that has been in use for ~50 years and has many expected adverse events. As the purpose of this study is to assess the integration of lupron and FLT imaging with this transplantation platform, only 1) serious adverse events and 2) adverse events not attributable to the standard BMT platform and either possibly, probably, or definitely attributable to either lupron or FLT imaging will be reportedSuspected adverse reaction
Suspected adverse reaction means any adverse event for which there is a reasonable possibility that the drug caused the adverse event. For the purposes of IND safety reporting, 'reasonable possibility' means there is evidence to suggest a causal relationship between the drug and the adverse event. A suspected adverse reaction implies a lesser degree of certainty about causality than adverse reaction, which means any adverse event caused by a drug.