Preventing Stem Cell Transplant Complications With a Blood Separator Machine

Peripheral blood stem cell transplant research carried out by the NHLBI BMT Unit focus on
transplant techniques designed to decrease graft versus host disease (GVHD), increase the
graft-versus-leukemia (GVL) effect and reduce the risk of post-transplant graft rejection.

Through incremental transplant clinical trials we have shown that by controlling the stem
cell (CD34+ cell) and T lymphocyte (CD3+ cell) dose, severe GVHD can be reduced whilst
beneficial GVL effects can be preserved. We found that T cell depleted transplants using the
Nexell/Baxter Isolex 300i system and subsequently, the Miltenyi CliniMACS[registered] CD34+
system to obtain high CD34+ doses depleted of lymphocytes were safe to administer and
associated with less severe acute GVHD and promising response rates and overall survival. Our
previous trials have helped us to create the transplant environment (significant
lymphodepletion and minimal post transplant immunosuppression) that make for an ideal
platform for adoptive cellular immunotherapy. Adoptive cell transfer is the passive transfer
of immune cells, into a new recipient host with the goal of transferring the immunologic
functionality and characteristics into the new host.

This protocol is designed to evaluate the safety and efficacy of the Miltenyi
CliniMACS[registered] CD 34 selection system in HLA-matched sibling allogeneic peripheral
blood stem cell transplant. The manipulation of the graft is the primary research
intervention, subject to IDE# 15632, and all other aspects of clinical management on this
protocol are standard care. The target CD34+ dose range will be >3 x 10(6)/kg and the target
CD3+ dose range will be 5 x 10(4)/kg to 1 x 10(6)/kg. Once we demonstrate adequacy of this
platform for engraftment and absence of significant GVHD in ten consecutive recipients, we
will seek IRB permission to proceed with planned adoptive cellular therapies.

The protocol will accrue up to 96 transplant recipients aged 10-80 with a hematological
malignancy and their HLA-matched sibling donors, in whom allogeneic stem cell transplantation
from an HLA-matched sibling would be routinely indicated. Diagnostic categories will include
acute and chronic leukemia, myelodysplastic syndromes, lymphomas, multiple myeloma and
myeloproliferative syndromes.

Subjects will receive a myeloablative conditioning regimen of cyclophosphamide (120 mg/kg
total), fludarabine (125 mg/m(2) total) and total body irradiation (1200 cGy with lung
shielding to 600 cGy), followed by an infusion of a stem cell product selected for CD34+
progenitors using the Miltenyi CliniMACS[registered] system. Older subjects will receive a
lower dose of irradiation (800 or 600 cGy based on age) to reduce the regimen intensity.

The overall objective is to assess the feasibility of using this system as a platform for
cellular immunotherapy initiatives. The primary study endpoint will be overall survival at
day +200. Stopping criteria for safety will monitor non-relapse mortality at day +200 and
late disease free survival at 2 years. Secondary endpoints will be standard transplant
outcome variables such as non-hematologic toxicity, incidence and severity of acute and
chronic GVHD and relapse of disease.

- INCLUSION CRITERIA RECIPIENT

5.1.1 Ages 10-80 years inclusive

5.1.2 Any one of the following hematologic conditions, confirmed by pathology, meeting a
standard indication for allogeneic stem cell transplant:

5.1.2.1 Chronic myelogenous leukemia (CML): Subjects under the age of 21 in chronic phase
OR Subjects ages 10-80 in chronic phase who have failed or are intolerant to treatment with
second generation tyrosine inhibitors OR Subjects ages 10-80 in accelerated phase or blast
transformation. OR

5.1.2.2 Acute lymphoblastic leukemia (ALL): any of these categories: Adult ALL including
standard risk; Pediatric ALL in first remission with high-risk features (presenting
leukocyte count >100,000/cu mm, karyotypes t(9; 22), t4, t19, t11, biphenotypic leukemia).
All second or subsequent remissions, primary induction failure, partially responding or
untreated relapse. OR

5.1.2.3 Acute myelogenous leukemia (AML): AML in first remission - except AML with good
risk karyotypes: AML M3 (t15; 17), AML M4Eo (inv 16), c-kit unmutated AML t (8; 21). All
AML in second or subsequent remission, primary induction failure and resistant relapse. OR

5.1.2.4 Myelodysplastic syndromes(MDS): any of these categories - refractory anemia with
transfusion dependence, refractory anemia with ANC<500/microL, refractory anemia with
excess of blasts, transformation to acute leukemia, chronic myelomonocytic leukemia,
atypical MDS/myeloproliferative syndromes. OR

5.1.2.5 Myeloproliferative disorders including atypical (Ph-negative) chronic myeloid and
neutrophilic leukemias, progressing myelofibrosis, and polycythemia vera, essential
thrombocythemia either in transformation to acute leukemia or with progressive transfusion
requirements or pancytopenia. OR

5.1.2.6 Chronic lymphocytic leukemia refractory to fludarabine treatment and with bulky
progressive disease or with thrombocytopenia (less than or equal to 100,000 / microl) or
anemia (less than or equal to 10g/dl) not due to recent chemotherapy. OR

5.1.2.7 Non-Hodgkin s lymphoma including Mantle cell lymphoma relapsing or refractory to
standard of care treatments. OR

5.1.2.8 Multiple myeloma, Waldenstroms macroglobulinemia, unresponsive or relapsed
following standard of care treatments. OR

5.1.2.9 Hodgkin's Lymphoma relapsing following an autologous transplant. OR

5.1.2.10 Other rare hematologic malignancies for which hematopoietic stem cell
transplantation has been performed and offers a durable remission or as the only option for
potential for cure.

- Chemotherapy-resistant multisystem Langerhans cell histiocytosis (MSLCH) especially
involving organs like the bone marrow, liver, spleen, and lungs

- Aggressive systemic mastocytosis, and mast cell leukemia (MCL) in first CR (CR1)

- Hypereosinophilic syndrome who have failed imatinib therapy or FIP1L1-PDGFRa-negative
patients who develop end-organ dysfunction

- Adult T-cell leukemia/lymphoma at first diagnosis

- Refractory or disseminated nasal-type extranodal NK/T-lymphoma or aggressive Natural
killer cell leukemia/lymphoma

- Mycosis fungoides and S(SqrRoot)(Copyright)zary syndrome after failure of two or three
initial therapies

- Primary or relapsed refractory Angioimmunoblastic T-cell lymphoma at first diagnosis

- Hepatosplenic T-cell lymphoma (gamma/delta T-cell lymphoma) at first diagnosis

- T-cell prolymphocytic leukemia at first diagnosis

- Subcutaneous panniculitic T-cell lymphoma at first diagnosis

- Hematodermic neoplasm (blastic natural killer cell lymphoma or Blastic plasmacytoid
dendritic cell neoplasm) at first diagnosis

5.1.3 HLA identical (6/6) related donor.

5.1.4 For adults: ability to comprehend the investigational nature of the study and provide
informed consent. For minors: written informed consent from one parent or guardian.
Informed oral assent from minors: the process will be explained to the minor on a level of
complexity appropriate for their age and ability to comprehend.

EXCLUSION CRITERIA RECIPIENT (any of the following)

5.2.1 Major anticipated illness or organ failure incompatible with survival from transplant

5.2.2 Severe psychiatric illness or mental deficiency sufficiently severe as to make
compliance with the transplant treatment unlikely and making informed consent impossible.

5.2.3 Positive pregnancy test for women of childbearing age

5.2.4 DLCO adjusted for Hb and ventilation< 50% predicted

5.2.5 Left ventricular ejection fraction < 40% (evaluated by ECHO) or < 30% (evaluated by
MUGA)

5.2.6 AST/SGOT > 10 times ULN

5.2.7 Total bilirubin > 5 times ULN

5.2.8 Estimated GFR < 15 mL/min

5.2.9 Recipients who have active infections with HIV or active hepatitis C (HCV)

INCLUSION CRITERIA DONOR

5.3.1 Related donor, HLA identical (6/6) with recipient

5.3.2 Weight greater than or equal to 18 kg

5.3.3 Age greater than or equal to 2 or less than or equal to 80 years old

5.3.4 For adults: ability to comprehend the investigational nature of the study and provide
informed consent. For minors: written informed consent from one parent or guardian and
informed assent: The process will be explained to the minor on a level of complexity
appropriate for their age and ability to comprehend.

EXCLUSION CRITERIA DONOR (any of the following)

5.4.1 Pregnant or breast-feeding. Lactating donors are permitted provided breast milk is
discarded during the days filgrastim (G-CSF) is given

5.4.2 Unfit to receive G-CSF and undergo apheresis (abnormal blood counts, history of
stroke, uncontrolled hypertension)

5.4.3 Sickling hemoglobinopathy including HbSS, HbSC

5.4.4 Donors who are positive for HIV, active hepatitis B (HBV), hepatitis C (HCV) or human
Tcell lymphotropic virus (HTLV-I/II)

5.4.5 Severe psychiatric illness or mental deficiency sufficiently severe as to make
compliance with the donation process unlikely, and making informed consent impossible.