Sources of stem cells:

• Bone marrow
• Peripheral blood stem cells
• Umbilical cord blood

Options of donor

• Matched sibling donor: Likelihood of sibling match is approximately 25%
• Matched unrelated donor: Likelihoodof match:
  • Whites- 90%
  • Asian/ Hispanics-70% 
  • Africans-60% 
  • This is because in evolution humans migrated from Africa-first to Asia-then some of them to United States.
• Haplo matched family donor

Points to note prior to collection of stem cells from donor:

• Safety of donor is of at most importance
• Informed consent must be fully and deeply explained
• Detailed evaluation to be done to look for all possible medical contraindications.
• No risk must be taken however great is the need of the patient.
• Age is not a contraindication. Donation can be done at any age (But  cell yield decreases with age)

Donor Evaluation and preparation for stem cell donation

• Education of donor, including education regarding procedures, risks, alternatives, and possible future collections 
• Medical history, including special attention to history of autoimmune disorders, arthritis, cardiac and vascular diseases, and history of cancer
• History of high risk behaviors, such as recent tattoos, body piercing, sexual practices and travel
• Physical examination, including vein assessment (PBSC donors) and oral examination for mouth opening (as marrow donors need intubation, inhalational anesthesia and procedure is done  in prone position)
• Laboratory studies, including verification HLA typing, ABO typing, CBC, chemistry panel, infectious disease panel, urinalysis, ECG,CXR
• Consent for collection procedures and the release of protected health information to the stem cell recipient

Consider following while choosing a donor

• As far as possible full HLA match. Single mismatch, but matched at HLA DQ-B1 is acceptable.
• Age- Improved transplant results with younger donor.
• Parity of female donor- If donor is multiparous female there is higher risk of GVHD
• Size of donor-Larger (Male) donors provide larger harvest
• CMV Sero status of both recipient and donor
• There is no effect of gender or ABO blood group.

Collection of stem cells:

Bone marrow 

• Collected by large bore needles placed in post iliac crest.
• Technique is same as diagnostic bone marrow aspiration.
• 50 to 100 aspirations simultaneously on both sides under regional anesthesia/ general anesthesia are done
• Both heparin and ACD-A can be used for anticoagulation.
• Lowest cell dose is not established.
• Generally>2x10⁸nucleated marrow cells/kg of recipient is adequate. (20ml/kg of donor body weight is safe)
• It is a very safe procedure
• Side effects include anesthesia complication, hemorrhage and infection at puncture site
• Do not aspirate a volume more than 5 ml in each aspiration to avoid dilution of bone marrow with blood.
• After collection, bone marrow is filtered and is then infused intravenously directly to the patient 
• Usual volume harvested – 10-15 ml of marrow per kg of recipient body weight
• Correction for blood loss
  • < 10 ml / kg of donor- volume replacement with salt solutions. 
  • 10 ml – 20ml / kg- colloid solutions like hydroxy ethyl starch
  • >20 ml/ kg- Replacement with blood, preferably autologous, that is collected prior to harvest

Peripheral Blood Stem cells (PBSC)

• Stem cells mobilized into peripheral blood using GCSF. Dose = 10 microg/ kg/day. Rarely GM-CSF, IL-3, Stem cell factor thrombopoietin are used.
• Collected by apheresis -2 types
  • Continuous- Cobe spectra, Fenwal CS3000
  • Discontinuous- Haemotonics
• Timing of apheresis-On 5^th day of G-CSF administration (Before and after that CD34+count is less). Desirable CD34+cells count in peripheral bloodismore than 5-10/microL.
• Procedure is safe (1 in 10000 have splenic rupture)
• Other side effects- skeletal pain and flare up of auto immune disease
• No risk of leukemia
• Preferred target dose - >2x10⁶- CD34 cells /kg of recipient
• Circadian rhythm in hypothalamus affects stem cell release from BM (By altering expression of CXCL-12 m marrow microenvironment ) Peak time for release of stem cells is evening (So start procedure in late afternoon)
• Regimen of chemotherapy followed by G-CSF results in higher number of CD34+cells (Choice of chemotherapy must be appropriate for the disease) Stem cells in recovering marrow is high. Cyclophosphamide and ifosmide are commonly used in these protocols.
• Venous access 
  • Single access is enough for discontinuous PBSC collection
  • For continuous flow apheresis devices, 2 lumen accessis needed.
  • Stable blood flow capacity with generally > 20µl/min is sufficient
  • Hickman central line is avoided due to risk of fracture
  • Temporary percutaneous dialysis catheter is preferred (Have adequate wall thickness to prevent collapse during aspiration of blood as well as a tip design that decreases local circulation of blood)
  • Catheters of 10F or larger calibers are appropriate (Pediatric patients –who have slower flow rate-may use catheters of 5 to 7 F)
• Citrate toxicity can be prevented by
  • Limiting quantity of citrate infused by decreasing the blood flow rate or by decreasing blood-citrate ratio
  • IV calcium gluconate (but beware of cardiac dysfunction)
  • Use heparin as anticoagulant 
• When blood in the extracorporeal circuit is expected to exceed 15% of donor’s blood volume, prime the machine with ABO compatible, irradiated PCV. Blood remaining in the circuit after completion of procedure can be rinsed back.
• Sedation may be needed for pediatric patient.

PBSC Vs BMSC

• Accelerated engraftment occurs with PBSC (Hence less PRBC/platelet transfusions and early ANC recover)
• Higher GVH reaction with PBSC- due to greater number of T cells. GVL with PBSCT is same as bone marrow harvest, hence the relapse rate.Despite this, number of PBSC harvest is increasing, which is due to logistic reasons. Centers give reason of donor safely. But adverse effects with PBSC harvest are higher than bone marrow harvest and also mortality of donor
• Factors determining which method to use.
  • Availability of OT
  • Availability of personnel to perform bone marrow aspiration
  • Contraindications to general anaesthesia
  • Choice of donor
  • Bone marrow collection is better in children because of possible side effects of G-CSF or problems with venous access
• Although PBSC containing 1-2x10⁶ CD34 cells/kg of recipient will achieve hematopoietic stem cell reconstitutions, for most patients, cell doses of 2.5-5x10⁶ CD34+/kg of recipient is desirable.

Poor mobilization

• Risk factors
  • Older age
  • Marrow disease
  • Prior radiotherapy
  • Prior chemotherapy
  • Previous use of drugs such as Lenalidomide
• Strategies used in poor mobalizers
  • High dose (15 mg / kg-BD) GCSF after 2-4 weeks  if initial collection
  • Plerixafor (CXCR4 antagonist –AMD3100)– 240 mcg /kg given 10 hrs. before apheresis 
  • Cyclophosphamide based mobilization
    • Do not use if creatinine clearance is <40ml/min
    • Inj. Mesna- 4000mg/m2- in 500ml d5% over 12 hrs
    • Inj. Cyclophosphamide- 2000mg/m2- in 500ml D5% over 2 hrs
    • Prehydrate- 1000ml- over 6 hrs
    • Posthydrate- 1000ml- over 6 hrs
    • Start Inj. G-CSF- 300mcg- SC- BD- From day 5- continue till adequate harvest is done.
    • Start monitoring CD34 count daily from day 11. Do harvest once CD34 count is >50/microlit. Ensure platelet count of >40,000/cmm on the day of harvest.
• High volume apheresis if  CD34 count in PB is 10-20/ml- > 2-3 times  patient’s blood volume is processed 

Umbilical cord blood

• Blood is collected from umbilical vessels in placenta at the time of delivery of baby.
• Cells are immunologically relatively naïve.(Hence better results even with HLA incompatibility )
• Early cord clamping, collect UCB from placental vein by usual venesection method. Perfusion of placenta with salt solutions may increase the cell number collected. Product volume is reduced by removing plasma and RBCs to decreased storage space
• Most banks prefer to bank only the largest units of more than 70ml, in order to obtain at least 3x10⁷  nucleated cells / kg recipient body weight
• Histocompatible cord blood can be searched and obtained from registries.
• If the criterion for the minimal number of cells for single UCB transplantation is not achieved .A double cord blood transplantation should be considered. (Both together TNC should be >3x10⁷/kg)
• Advantages
  • Cells are immediately available.
  • Absence of risk to mother / donors.
  • Decreased risk of transmission of diseases such as CMV.
  • Incidence of GVHD is low
  • Strong protection against relapse
• Disadvantages
  • Small number of cells per unit, hence can be used only in pediatric age groups.
  • Prolonged engraftment time.
  • High chances of graft failure (Back up graft must be kept ready)

Exclusion criteria for collection of cord blood

• Multiple gestation.
• Premature delivery-Baby needs that blood
• Active chorioamnionitis or sepsis.
• Mother being recipient of an organ transplant
• Mother with history of cancer.
• Mother with high risk behaviors or previously diagnosed with HIV, hepatitis or syphilis
• Mother with active veneral disease such as vaginal herpes simplex and delivering vaginally.

Matched Unrelated Donor

• If there are several potential donors, better to recruit a donor who is,
  • Male
  • Young
  • ABO matched
  • CMV negative if recipient is negative or CMV positive if recipient is positive.
• MUD search can be done through www.bmdw.org.
• Preliminary search is free but needs registration

Combining haploidentical stem cells with umbilical cord blood.

• UCB transplant is associated with slow hematologic recovers and delayed immune reconstitution.
• Co-transplanting with CD34+cell selected haploidentical PBSC causes rapid neutrophil recovery usually at medium of 10 days. Later once cord immune cells engraft, they remove haploidentical cells.

Recent advances:

Source of stem cells and transplant outcomes in patients over 50 years of age with myelodysplastic syndrome

This retrospective study aimed to determine the most suitable donor type for allogeneic hematopoietic cell transplantation (HCT) in elderly patients (over 50 years old) with myelodysplastic syndrome (MDS). Among 1787 patients in Japan, four donor types were compared: HLA-matched sibling donors (MSDs), 8/8 allele-matched unrelated donors (MUDs), 7/8 allele-MUDs, and unrelated cord blood (UCB) donors. While MSD transplants were not found to be superior, 8/8MUD transplants exhibited a lower risk of relapse, and UCB transplants had higher non-relapse mortality. However, donor type did not significantly impact overall survival, disease-free survival, or graft-versus-host disease (GVHD)-free, relapse-free survival. 

https://doi.org/10.1038/s41409-023-01997-3

Impact of donor age and relationship on outcomes of haploidentical HSCT

This retrospective analysis assessed outcomes in 299 patients who underwent peripheral blood haploidentical hematopoietic cell transplantation (haplo-HCT) with post-transplant cyclophosphamide (PTCy) for acute leukemias between July 2009 and May 2021. Donor characteristics, such as age, sex, relationship, ABO status, CMV status, and HLA match grade, were examined in multivariate analyses for associations with various outcomes. Higher donor age was linked to increased non-relapse mortality (NRM) and reduced relapse risk. No significant differences were observed in cytokine release syndrome (CRS), acute or chronic graft-versus-host disease (GVHD), or overall survival (OS) based on donor age. 

https://doi.org/10.1038/s41409-023-01984-8

Clinical significance of total nucleated cell count in bone marrow of patients with acute lymphoblastic leukemia who underwent allogeneic hematopoietic stem cell transplantation

In a multicenter retrospective study focusing on patients with acute lymphoblastic leukemia who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) while in remission, the recipients were stratified into high- and low-bone marrow nucleated cell count (NCC) groups based on a cut-off of 10 × 10^4/µL. The high-NCC group exhibited a 3-year overall survival (OS) of 51.2% compared to 84.5% in the low-NCC group (p < 0.001). The high-NCC group also had a significantly higher non-relapse mortality (NRM) of 27.5% compared to 6.5% in the low-NCC group (p < 0.001). There was no significant difference in relapse rates between the groups. 

https://doi.org/10.1007/s12185-023-03688-7

Etoposide combined with cytarabine and pegfilgrastim for poorly mobilizing patients with multiple myeloma and lymphoma

This prospective study was conducted to evaluate the efficacy and safety of the EAP (etoposide, cytarabine, and pegfilgrastim) regimen in poorly mobilizing patients with multiple myeloma (MM) or lymphoma. The trial included 58 patients, with 91.4% achieving adequate mobilization (≥2.0 × 10^6 CD34+ cells/kg) and 70.7% achieving optimal mobilization. The median number of cumulative collected CD34+ cells was 9.2 × 10^6/kg, with a median apheresis count of 1.2 per patient. While 8.6% required plerixa for rescue, successful mobilization was achieved. Infections occurred in 20.7% of patients, and 43.1% required platelet transfusions, with a median engraftment duration of 11 days. 

https://doi.org/10.1111/bjh.19367