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HPC Transplantation
D. Joe Chaffin, MD
Cedars-Sinai Medical Center, Los Angeles, CA

A. Basics of Hematopoietic Progenitor Cell Transplant

1. What’s a “stem cell?”
a. Hematopoietic stem cells (HSC)
1) HSCs divide to make more HSC and originate all hematopoietic cells
b. Hematopoietic progenitor cells (HPC)
1) More mature cells that are multipotent or committed to either myeloid
(WBCs, RBCs, PLTs) or lymphoid line (lymphs, dendritic cells)
c. “HPC” and “HSC” often used interchangeably, and the entire process is
referred to as either “HPC” or “HSC” transplantation
1) Transplants include BOTH HSCs and HPCs (which actually works
better than pure HSCs, per some studies)
2) In this lecture: “HPC” will be used instead of “HSC” or “HSC/HPC”
d. HPCs identified by flow-cytometry detectable expression of CD34
1) CD34+ cells present in small quantities in bone marrow (0.5–1%) and
in tiny quantities (0.05–0.1%) in circulating blood
2) HPCs also express human leukocyte antigen (HLA) antigens, but do
NOT carry ABO blood group antigens.

Figure 1: HSC = Hematopoietic stem cell, MPP = Multipotent precursor, HPC = Hematopoietic progenitor
cell, CMP = Common myeloid precursor, CLP = Common lymphoid precursor

2. Basic steps of HPC transplantation

a. Treat the disease as best you can and determine eligibility for transplant
b. Collect HPCs either from the person who will undergo transplant or
someone else (related or unrelated) using one of several methods
c. Prepare the recipient by using intensive therapy (chemotherapy, total body
irradiation, or both), with either complete or incomplete destruction of the
patient’s native bone marrow (see “conditioning” regimens later)
d. Replace the bone marrow by infusing previously collected HPCs (HPCs
migrate from peripheral circulation into marrow space and generate a new
marrow)

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B. HPC Transplant Types
1. By Donor:

Figure 2: US Transplant Categories (Source: Pasquini MC, Wang Z. Current use and outcome of
hematopoietic stem cell transplantation: CIBMTR Summary Slides, 2011. Available at: http://www.cibmtr.org)

a. Autologous transplant
1) The donor and recipient are the same person
2) Most common type of transplant by far (see chart above)
3) Used most often for lower grade heme malignancies (e.g., multiple
myeloma, non-Hodgkins and Hodgkins lymphoma); see fig 3, pg 3
b. Syngeneic transplant
1) Donor and recipient are identical siblings (twins, triplets, etc.)
2) By definition, HLA-identical
c. Allogeneic transplant, matched related donors (“MRD”)
1) Donors are most commonly siblings (25% HLA-identical)
2) If no match, will require unrelated donor allogeneic donation
3) Slightly less common than unrelated donor transplants
d. Allogeneic transplant, matched unrelated donors (“MUD”)
1) Many facilitated through US registry (National Marrow Donor
Program, or “NMDP”)
2) Most commonly used in acute leukemias, but also in aplastic anemia,
hemoglobinopathies and immune deficiencies
2. By Recipient Preparation:
a. Standard myeloablative conditioning
1) Total body irradiation and/or chemotherapy (historically busulfan and
cyclophosphamide; melphalan, fludarabine are options now)
2) Traditional manner of recipient preparation; deliberately toxic and has
many potential complications (see later discussion)
b. Reduced-intensity conditioning (RIC)
1) Native marrow deliberately incompletely destroyed, leading to fewer
potential complications for recipients during conditioning
2) Allows new (transplanted) immune system to destroy tumor cells
(“Graft-vs.-Tumor effect” or “GVT” for short)
3) RIC allows older and/or higher risk patients access to transplants that
may have been too toxic with standard myeloablative conditioning
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4) Potential complications of RIC:
a) Incomplete tumor cell destruction with resultant microchimerism
b) Increased risk of acute Graft-vs.-Host Disease (GVHD)
3. By Collection Technique:
a. HPC, Apheresis (HPC-A)
1) HPC harvested from donor circulation using apheresis equipment
2) Currently the most common source for HPC transplantation
a) Exception: Allogeneic transplants for recipients < age 20 (HPC-M)
b) Easier on donors; no general anesthesia required
c) Less graft failure compared to marrow source
3) Recent large study comparing HPC-A and HPC-M as source in
unrelated allogeneic HPC transplants confirmed no survival benefit
with either source (Anasetti C et al, NEJM 367;16:1487-96)
a) HPC-A has lower risk of graft failure
b) HPC-M has lower risk of GVHD
b. HPC, Marrow (HPC-M)
1) HPC harvested by direct aspiration of bone marrow material
2) Requires general or at least regional anesthesia
3) Most donors recover quickly (few days to a week or two)
4) Allogeneic donors may donate units of autologous RBCs prior to
harvest in order to correct anemia that may result from procedure
c. HPC, Cord Blood (HPC-C)
1) Umbilical cord blood rich in HPCs; collect after delivery
2) Used far less than other types, but increasing
a) Best in children and small adults (due to lower CD34+ cell content)
b) Use is increasing in all age groups, however
3) Naïve immune cells in cord blood make these transplants:
a) Less likely than HPC-A or HPC-M to cause GVHD
b) Less likely to fail due to HLA mismatch with recipient
C. Indications for HPC Transplantation
1. In adults (over age 20); 2009 US data from CIBMTR

Figure 3: Overall US Transplants 2009 (Source: Pasquini MC, Wang Z. Current use and outcome of
hematopoietic stem cell transplantation: CIBMTR Summary Slides, 2011. Available at: http://www.cibmtr.org)

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a. Malignant/premalignant diseases (in order of decreasing frequency)
1) Multiple myeloma (most common reason for autologous HPC
transplant)
2) Non-Hodgkins lymphoma
3) Acute myeloid leukemia (most common reason for allogeneic HPC
transplant)
4) Hodgkins disease
5) Acute lymphoid leukemia
6) Myelodysplastic/myeloproliferative disorders
7) Other malignancies (solid tumors, etc)
8) Chronic myeloid leukemia
b. Non-malignant diseases
1) Aplastic anemia
2) Systemic amyloidosis
c. Note that adults are less commonly transplanted for non-hematologic
malignancies and non-malignant disorders
2. In children (under age 20); 2009 US data from CIBMTR

Figure 4: HPC Transplants in Patients < 20 (Source: Pasquini MC, Wang Z. Current use and outcome of
hematopoietic stem cell transplantation: CIBMTR Summary Slides, 2011. Available at: http://www.cibmtr.org)

a. Childhood solid tumors (Neuroblastoma, Wilm’s, Ewing sarcoma, etc.);

vast majority of these transplants are autologous
b. Hematologic malignant/premalignant disorders (vast majority allogeneic)
1) Acute lymphoid leukemia
2) Acute myeloid leukemia
3) Myelodysplastic/myeloproliferative disorders
4) Hodgkins disease
5) Non-Hodgkins lymphoma
c. Non-malignant diseases (all allogeneic)
1) Hematologic (aplastic anemia, sickle cell disease)
2) Inherited metabolic disorders (mucopolysaccaridoses, lysosomal
disorders, adrenoleukodystrophy)
3) Immune deficiencies (e.g., severe combined immunodeficiency,
Wiscott-Aldrich)

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D. Preparation and Collection of HPC Products
1. Donor choice and preparation
a. Allogeneic donation
1) General considerations
a) HLA compatibility most important (ABO incompatibility in ~50%)
b) Perfect match = HLA-A, HLA-B, HLA-C (Cw), HLA-DRB1 alleles
(“8 of 8 match”); tested by high resolution (DNA) methods
c) Best chance of finding compatible donor is in siblings (25%)
d) NMDP used if no siblings or family matches

Figure 5: Allogeneic transplant process diagram

2) Donor screening
a) HPC-A, HPC-M donors complete a donor questionnaire (cellular
therapy donor history questionnaire available from AABB)
• Mothers are questioned for HPC-C collections
b) Infectious disease screening is performed including:
• Hepatitis B (anti-HBc, HBsAg)
• Hepatitis C (anti-HCV, HCV NAT)
• HIV (anti-HIV-1,2, HIV NAT)
• HTLV-I/II (anti-HTLV-I/II)
• Cytomegalovirus (CMV)
• Also syphilis, West Nile Virus, Chagas’ Disease
c) Units from donors with reactive tests may sometimes be used (with
consent) if they are the best donor for a recipient in urgent need
3) Mobilization of CD34+ cells into peripheral blood (HPC-A collections)
a) Cytokines cause HPC to migrate from marrow into circulation
• G-CSF (granulocyte-colony stimulating factor)
– Given daily (10 µg/Kg/day); peak effect on day 5
– Complications: bone pain, enlarged spleen (with rare rupture),
thrombocytopenia, LFT increases, headache, nausea
• AMD3100 (plerixafor)
– May be used with G-CSF to aid in difficult mobilizations
b) Count peripheral CD34+ cells by flow cytometry; may use counts to
trigger start of HPC-A collection ( if >5-20 CD34+ cells/µL)
b. Autologous donation
1) Donor screening
a) Donors should be in clinical remission

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b) Infectious disease testing done for baseline
c) Full H&P, heart/lung function tests done for baseline
2) Mobilization/treatment
a) Mobilization is done in conjunction with treatment because
treatment effect makes HPCs migrate into circulation (side benefit)
b) Myeloablative regimens are used (GVT effect is not applicable)
c) Ablative chemotherapy (+/– total body irradiation) can both treat the
disease and cause dramatic increases in circulating PBPCs.
• Chemo also helps to “purify” the subsequent HPC product from
malignant cells (“in-vivo purging”)
d) G-CSF used in concert with chemotherapy to increase effect

Figure 6: Autologous HPC transplant process

2. Collection process
a. HPC-Apheresis (HPC-A)
1) Standard apheresis equipment, with specific HPC-A software
2) 2-10 x 106 CD34+ cells/Kg of recipient is targeted (5 x 106 is typical,
but larger doses are used in certain diseases, like myeloma)
a) More CD34+ cells correspond with lower risk of graft failure
b) More T-lymphs in HPC-A (10X more than HPC-M) likely is the
cause of the increased risk of chronic GVHD with HPC-A vs. –M
3) Collection days:
a) Allogeneic: Most centers use large-volume leukapheresis to collect
entire dose in one day (as tolerated by donor)
• Usually start on day 5 of G-CSF administration
• Typically requires in the range of 3-4 blood volumes processed
• Donors prone to citrate effect, and calcium is often administered
b) Autologous: More commonly smaller volumes per procedure
processed over multiple days
b. HPC-Marrow (HPC-M)
1) Multiple aspirations of the posterior iliac crest on one or both sides (50-
100 aspirations over an hour or so)
2) 1-2 liters typically withdrawn (typically 10-15 mL/Kg donor weight)
3) Quantity targeted varies depending on type
a) 2-4 x 108 nucleated cells/Kg
b) A range of 2-6 x 106 CD34+ cells/Kg weight of recipient

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4) By comparison to HPC-A, HPC-M products have many more RBCs
and much more plasma, but 10X fewer T-lymphocytes
c. HPC-Cord Blood (HPC-C)
1) Product collected with no risk to donor, after cord is cut
2) Most commonly, umbilical vessel cannulated in a sterile manner by
dedicated collection personnel after delivery of placenta
3) Collection is generally by gravity drainage of placental blood into
collection bag containing CPD anticoagulant
4) Approximately 100-150 mL of product collected (less will be infused)
E. Processing of HPC Products Before Infusion
1. Post-collection processing of allogeneic units for ABO issues (see figure 7
below for incompatibility types)
a. Major incompatibility
1) Donor with incompatible red cell antigen, such as group A donor and
group O recipient, so RBCs in HPC are hemolyzed during infusion
a) Group O recipients receiving non-O HPCs
b) Group AB HPCs going to non-AB recipients
2) Process the HPCs to decrease the number of accompanying RBCs
(usually an issue in HPC-M but not HPC-A products)
3) Options to decrease RBCs to in the range of 10-20 mL residual
(generally considered a safe level of incompatible RBCs):
a) Centrifugation and buffy coat harvest/enrichment
b) Manual RBC sedimentation with 6% hydroxyethyl starch
c) Density-gradient separation using Ficoll-Hypaque solution
d) Automatic apheresis equipment processing (HPC-A products)

Figure 7; HPC ABO Compatibility Types

b. Minor incompatibility
1) Donor with incompatible red cell antibody, such as group O donor and
group A recipient, so recipient RBCs are hemolyzed during infusion
a) Group AB recipients receiving non-AB HPCs
b) Group O HPCs going to non-O recipients
2) Process the HPCs to decrease the amount of accompanying plasma
(HPC-M much more often than HPC-A)
3) Not generally difficult; centrifugation with plasma removal
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c. Bidirectional (major and minor) incompatibility
1) Donor antigens and antibodies incompatible with recipient; so both
donor and recipient RBCs may be hemolyzed during infusion
a) Group A donor/group B recipient
b) Group B donor/group A recipient
2) Process to remove both RBCs and plasma from HPC product (HPC-M)
3) Typically done with RBC reduction methods (plasma reduction occurs
as part of process)
2. Other blood group issues
a. The presence of significant RBC antibodies (e.g., anti-D, –K, –Fya) in
either donor or recipient may lead to need for interventions as above
b. Rh incompatibility is not a transplant contraindication, though antibodies
may be induced when a D-neg donor engrafts in a D-pos recipient
3. Common post-collection processing steps
a. Filtering
1) HPC-M products filtered to remove bone fragments and debris
2) Caution: HPC products should not be filtered for leukoreduction at
infusion (some are ok with standard [170 micron] filter)
b. Freezing
1) 10% Dimethyl sulfoxide (DMSO) used as cryoprotectant
2) In autologous HPC transplants, units generally frozen in liquid or vapor
nitrogen to wait for marrow conditioning
3) HPC-C are pretty much always frozen and stored for later use
4) Allogeneic HPC donations (HPC-M or HPC-A) are less commonly
frozen; collection usually timed to recipient readiness
c. Washing
1) Frozen units may be washed to remove DMSO (optional)
2) Rare IgA deficient patients need washed units to prevent anaphylaxis
d. Centrifugation
1) Used to remove ABO-incompatible plasma from HPC units
e. Advanced methods
1) Purging
a) Reduce T-cells to reduce risk of graft-vs.-host disease
(unfortunately, this has been shown to INCREASE the relapse rate)
b) Purge malignant cells from an autologous collection
2) Enhancing
a) Isolate and purify the CD34+ cells
b) Can be done using magnetized antibodies either selecting CD34+
cells or selecting non-CD34+ cells for removal
f. HPC-A units
1) Processed to remove plasma and RBCs; around 20 mL volume
2) Frozen with 10% DMSO for final volume of 25 mL or so
4. Units that may result from post-collection processing:
a. HPC, (RBC reduced)
b. HPC, (Plasma reduced)
c. HPC, (CD34 enriched)
d. HPC, (Buffy coat enriched)
e. HPC, (Mononuclear cell enriched)
f. Cryopreserved HPC

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F. Infusion of HPC Products
1. Non-frozen products (usually allogeneic transplants)
a. Product infused similarly to blood transfusion
b. HPC-C is usually infused most rapidly due to lowest volumes, while
HPC-M may take many hours to infuse
c. Reactions to infusion (fever, chills, hives, etc.) should be documented
d. Leukocyte reduction and irradiation of HPC products is not only
forbidden but stupid!
1) Some use 170 micron “standard” blood filters to remove large clots at
time of infusion, but not all agree with this practice
2. Previously frozen products (autologous or HPC-C transplants)
a. Product may be washed or just infused directly following thaw
1) HPC-C units are nearly always washed before infusion
2) Autologous HPC units may or may not be washed
b. DMSO toxicity is the most common complication of HPC infusion
1) Flushing/rash
2) Nausea/vomiting
3) Chest tightness/wheezing/cough
4) Hypertension
5) Garlic odor on breath
c. If washing is not available, may need to divide infusion to limit DMSO
exposure to less than 1mL DMSO /Kg recipient weight/day
G. Potential Complications During and After Transplant
1. Mucositis (over 90% of patients)
2. Bacterial infections early, viral/fungal infections later
3. Graft rejection (Host-vs.-graft effect)
a. Factors that increase risk for rejection:
1) Increasing HLA disparity
2) T-cell purging of the infusion product
b. Factors that decrease risk for rejection:
1) Increased dose of HPCs in infusion
2) Dose-intense treatment regimens
4. Graft-vs.-Host Disease (GVHD)
a. Acute GVHD
1) First 100 days, presents with skin, GI tract, and liver involvement
2) Risk more with HLA differences and increased T-cells in transplant
b. Chronic (cGVHD):
1) Usually after day 50, have prominent skin and autoimmune symptoms
(like Sjögren’s and systemic sclerosis) in addition to above
2) HPC-A recipients: less aGVHD, greater cGVHD vs. HPC-M recipients
3) Chronic GVHD seen in 40-50% of long-term survivors
c. Graft-vs.-tumor effect (GVT) or “graft-vs.-leukemia” effect (GVL)
1) Newly formed lymphocytes may act to destroy residual tumor cells,
leading to better survival and decreased recurrence
2) As such, this is a beneficial form of GVHD, and is used intentionally in
reduced-intensity conditioning allogeneic HPC transplants
5. Problems inherent to ABO-mismatched HPC transplants
a. Immediate hemolytic transfusion reaction

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1) Potential consequence of major incompatibility (donor RBCs
hemolyzed) or minor incompatibility (recipient RBCs hemolyzed)
2) Process units for prevention as above
b. Delayed hemolytic transfusion reaction
1) In minor ABO incompatibility, engrafting donor B-lymphocytes form
ABO antibodies vs. circulating recipient cells
2) Potential substantial hemolysis 1-3 weeks post-transplant
c. Pure red cell aplasia
1) Consequence of the combination of major ABO incompatibility and
survival of recipient plasma cells making ABO antibodies
2) Reticulocytopenia and absent RBC precursors in marrow at 60 days
after transplant
6. Hepatic veno-occlusive disease (VOD, aka sinusoidal obstructive syndrome)
a. Hepatomegaly, fluid retention secondary to sinusoidal injury
7. Engraftment syndrome (idiopathic pulmonary syndrome)
a. Pulmonary complication similar to TRALI clinically
b. Fever, hypoxia, may progress to diffuse alveolar hemorrhage
8. Hemorrhagic cystitis
9. Risk of secondary cancers (acute leukemias, various skin and solid tumors)
H. Transfusion Issues in HPC Recipients
1. Three phases
a. Pre-transplant (phase I)
1) From decision to transplant until treatment begins
b. Peri-transplant (phase II)
1) Begins at time of recipient treatment (chemo/TBI)
2) Ends when full engraftment has occurred
3) In ABO-incompatible transplants, ends at full ABO type conversion
(DAT neg, no detectable recipient RBCs or ABO antibodies)
c. Post-transplant (phase III)
1) After engraftment
2) After full blood type conversion in ABO-incompatible
2. Pre-transplant (Phase I)
a. Main concerns:
1) HLA alloimmunization
a) HLA antibodies formed via pregnancy or transfusion; more
antibodies make compatible HPC donors harder to find
b) If transfusion needed before transplantation, leukocyte-reduced
blood products prevent HLA immunization well (but not perfectly)
c) Also, avoid family member transfusions to prevent exposure to
family HLA antigens (siblings are often best donors)
2) Transfusion-associated graft-vs.-host disease (TA-GVHD)
a) Patients will be profoundly immunosuppressed and greatly at risk
b) Transfused T-lymphocytes may stay in recipient circulation for
years after transfusion (“microchimerism”)
c) As a result, all HPC transplant candidates should receive irradiated
blood products as soon as the decision to transplant is made
3) Cytomegalovirus status
a) Both recipients and potential donors are CMV tested

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b) Biggest worry about CMV-negative recipients getting CMV-
negative transplants
c) Most facilities use CMV-negative products; many substitute
leukocyte reduced products when necessary
b. Other concerns
1) ABO incompatible donors
a) Many begin giving group O RBCs to HPC transplant candidates that
will get ABO-incompatible transplants as soon as that fact is known
b) Others choose to use recipient type products until transplant
3. Peri-transplant (Phase II)
a. Continue:
1) Leukocyte-reduced blood products for HLA immunization prevention
and CMV transmission prevention
2) Irradiated cellular blood products for TA-GVHD prevention
3) CMV-safe blood products for CMV-seronegative recipients
b. Commonly a time of major blood product use
c. ABO incompatible transplants (40-50% of allogeneic transplants) have
special requirements (see table below)
Recipient Type Donor Type Mismatch Red Cell Type FFP/PLT Type
O A Major O A, AB
O B Major O B, AB
O AB Major O AB
A AB Major A, O AB
B AB Major B, O AB
A O Minor O A, AB
B O Minor O B, AB
AB O Minor O AB
AB A Minor A, O AB
AB B Minor B, O AB
A B Bidirectional O AB
B A Bidirectional O AB

1) Major ABO incompatibility:

a) Recipient ABO antibodies that may persist can hemolyze the new,
incompatible red cells formed by engrafting marrow
b) Until all original recipient RBCs are gone, patient may test in a
chimeric pattern on routine testing (reported as far out as two years)
c) Transfuse the recipient’s RBC type (or group O) and donor’s
FFP/platelet type (or group AB)
2) Minor ABO incompatibility:
a) Engrafting lymphs may start making ABO antibodies incompatible
with residual recipient RBCs (typically 1-3 weeks after transplant)
b) Delayed hemolytic transfusion reactions that may be severe and life-
threatening result
c) Transfuse the donor’s RBC type (or group O) and recipient’s
FFP/platelet type (or AB)
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3) Bidirectional (major and minor) ABO incompatibility:
a) Either of the potential problems outlined above may occur
b) Transfuse group O red cells and AB FFP/platelets
d. Testing issues in ABO incompatible transplants
1) ABO type will change over time; important for transfusion services to
know about transplanted patients to explain this issue
2) Original type of RBCs will decline over 2-3 weeks or so
3) New type of RBCs will be seen with mixed field reactions in donor
RBC grouping (“forward” grouping) tests
4) Recipient’s isoagglutinins (seen in serum or “reverse” grouping) will
also fade as donor B-lymphocytes start making new antibodies
e. Indications for various blood products
1) RBC, platelet, and plasma indications for transfusion do not differ
significantly from non-HPC patients
2) RBC “threshold” HGB 7 or 8 g/dL (higher in cardiac patients)
3) Platelet threshold commonly 10,000/µL for prophylactic transfusion,
50,000 for bleeding patients
4) FFP indications are not well-established (as is true elsewhere); FFP is
not widely used in HPC transplants, however
f. When is engraftment expected?
1) By definition, should occur within 100 days of transplant (usually
happens much faster (10-30 days for PMNs, > 15 days for PLTs)
2) Evidenced by recovery of neutrophil, platelet, and red cell production
(absolute neutrophil count > 500 cells/µL, platelets > 20K/µL)
3) Engraftment speed varies by source (HPC-A and –C faster than –M)
4. Post-transplant (Phase III)
a. Continue the use of:
1) Irradiated products for TA-GVHD prevention (probably forever)
2) Leukocyte-reduced products for HLA immunization prevention
3) CMV-safe products until patient is off of immunosuppressants
b. ABO-incompatible transplants:
1) Once full type conversion has occurred, may give RBC, plasma, and
platelet products consistent with new ABO type (donor type)
2) Simple crossmatch to detect residual recipient ABO antibodies; i.e., if
the patient’s serum does not react with red cells of the new ABO type
at all (a negative crossmatch), it is safe to begin using the new type.

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