Abstract 3022

Flow cytometric enumeration of CD34+ stem cells is a critical step in evaluation of graft adequacy prior to harvest for hematopoietic progenitor stem cell transplantation (HPSCT). The International Society of Hematotherapy and Graft Engineering (ISHAGE) established a standardized protocol in 1996 that is widely used for this purpose. Use of the sequential gating strategy routinely identifies a separate population of CD34+ events, with low side scatter and immunophenotype characteristic of hematogones (B-lymphocyte precursors) that are not excluded from the total CD34% according to the guideline. Whether including hematogones, presumably committed to lymphoid development, impacts engraftment is unknown. We investigated the effect of hematogones on engraftment.

Apheresis samples for CD34+ stem cell enumeration were analyzed for the presence (≥0.01%) of the additional CD34+ population by flow cytometry prospectively for 12 months. Patients who underwent HPSCT in that time frame were categorized according to whether their infused product(s) contained hematogones (negative= <0.01%, positive= ≥0.01%, or mixed (combination of collection products, some positive and some negative for hematogones)). Stepwise Cox proportional hazards analysis with a variable entry criterion of P≤0.10 and a variable retention criterion of P≤0.05 was used to identify multivariable prognostic factors for each outcome. Outcomes analyzed were neutrophil engraftment (days to absolute neutrophil count (ANC) >500 K/μL) and platelet (PLT) engraftment (days to platelet count >20 K/μL).

In a 12 month review of 274 apheresis samples, 61% harbored hematogones, comprising a median of 7.1% of total CD34+ cells (range 1.3 – 78.0). Patients (n=147, Table 1) received stem cell infusions that were negative, mixed, or positive for hematogones. The positive group had hematogones ranging from 0.22 – 6.31% of total CD34+ cells; hematogone % in the mixed group could not be quantitated. On multivariable analysis, patients in the mixed and positive groups had a marginally greater likelihood of neutrophil engraftment than those with negative hematogones (Table 2). Hematogones had no impact on platelet engraftment.

Hematogones are frequently included in the ISHAGE gating protocol. Reporting CD34% values that are inclusive of hematogones does not have an adverse effect on engraftment. Preliminary analysis suggests that receiving graft products that contain hematogones may predict for better neutrophil engraftment. Whether this is a direct or indirect effect of hematogones in the sample is unknown.

Table 1.

Descriptive Data by Hematogone Group (n=60, n=24, n=63)

Negative # (range) %Mixed # (range) %Positive # (range) %
Median age at transplant, years 58 (27–76) 54 (38–69) 54 (15–76) 
Male 40 66.7 16 66.7 34 54.0 
Diagnosis type 48 80.0 23 95.8 39 61.9 
Lymphoid    
    Transplant type 47 78.3 23 95.8 27 42.9 
    Autologous (Auto) 4 6.7 – 31 49.2 
    Myeloablative allogeneic (MA) 5 8.3 1 4.2 2 3.2 
    Reduced intensity allogeneic (RIC) 4 6.7 – 3 4.8 
Mini allogeneic (Mini)    
    Cell source 55 91.7 24 100.0 36 57.1 
    Peripheral stem cells (PSC) 1 1.7 – 24 38.1 
    Bone marrow (BM) 4 6.7 – 3 4.8 
    Umbilical cord (UC)    
    Median CD34+ dose, × 106/kg 5.06 (0.01–27.60) 4.38 (1.81–8.59) 4.00 (0.02–11.91) 
    Total nucleated cell dose (TNC), × 108/kg 9.00 (0.27–46.88) 18.20 (8.55–37.13) 4.54 (0.27–24.93) 
Negative # (range) %Mixed # (range) %Positive # (range) %
Median age at transplant, years 58 (27–76) 54 (38–69) 54 (15–76) 
Male 40 66.7 16 66.7 34 54.0 
Diagnosis type 48 80.0 23 95.8 39 61.9 
Lymphoid    
    Transplant type 47 78.3 23 95.8 27 42.9 
    Autologous (Auto) 4 6.7 – 31 49.2 
    Myeloablative allogeneic (MA) 5 8.3 1 4.2 2 3.2 
    Reduced intensity allogeneic (RIC) 4 6.7 – 3 4.8 
Mini allogeneic (Mini)    
    Cell source 55 91.7 24 100.0 36 57.1 
    Peripheral stem cells (PSC) 1 1.7 – 24 38.1 
    Bone marrow (BM) 4 6.7 – 3 4.8 
    Umbilical cord (UC)    
    Median CD34+ dose, × 106/kg 5.06 (0.01–27.60) 4.38 (1.81–8.59) 4.00 (0.02–11.91) 
    Total nucleated cell dose (TNC), × 108/kg 9.00 (0.27–46.88) 18.20 (8.55–37.13) 4.54 (0.27–24.93) 
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Table 2.

Predictors for Neutrophil Engraftment

Univariable HR 95% CI P-valueMultivariable HR 95% CI P-value
Hematogones   
    Mixed/Negative 1.8 1.1–2.9 0.022 1.9 1.2–3.2 0.012 
    Positive/Negative 0.6 0.4–0.9 0.012 1.5 0.9–2.2 0.09 
Age   
    Per 10 year increase 1.2 1.0–1.4 0.024  
Diagnosis type   
    Lymphoid/others 2.1 1.4–3.2 <0.001  
Transplant type   
    MA/Auto 0.1 0.1–0.2 <0.001 0.1 0.1–0.2 <0.001 
    Mini+RIC/Auto 0.3 0.1–0.5 <0.001 0.3 0.1–0.5 <0.001 
Cell source   
    BM/PSC 0.2 0.1–0.3 <0.001  
    UC/PSC 0.1 0.1–0.3 <0.001  
CD34+ dose, × 106/kg   
    Per 1 × 106/kg increase 1.98 1.61–2.43 <0.001 1.77 1.38–2.26 <0.001 
TNC, × 108/kg   
    Per 1 × 108/kg increase 1.33 1.21–1.47 <0.001  
Univariable HR 95% CI P-valueMultivariable HR 95% CI P-value
Hematogones   
    Mixed/Negative 1.8 1.1–2.9 0.022 1.9 1.2–3.2 0.012 
    Positive/Negative 0.6 0.4–0.9 0.012 1.5 0.9–2.2 0.09 
Age   
    Per 10 year increase 1.2 1.0–1.4 0.024  
Diagnosis type   
    Lymphoid/others 2.1 1.4–3.2 <0.001  
Transplant type   
    MA/Auto 0.1 0.1–0.2 <0.001 0.1 0.1–0.2 <0.001 
    Mini+RIC/Auto 0.3 0.1–0.5 <0.001 0.3 0.1–0.5 <0.001 
Cell source   
    BM/PSC 0.2 0.1–0.3 <0.001  
    UC/PSC 0.1 0.1–0.3 <0.001  
CD34+ dose, × 106/kg   
    Per 1 × 106/kg increase 1.98 1.61–2.43 <0.001 1.77 1.38–2.26 <0.001 
TNC, × 108/kg   
    Per 1 × 108/kg increase 1.33 1.21–1.47 <0.001  
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Disclosures:

No relevant conflicts of interest to declare.

Topics:
apheresis, engraftment, hematopoietic stem cell transplantation, tissue transplants, transplantation, adverse effects, flow cytometry, immunophenotyping, prognostic factors, absolute neutrophil count

Author notes

*

Asterisk with author names denotes non-ASH members.

© 2012 by The American Society of Hematology
2012
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