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Viability of non cryopreserved
CD34+Stem Cells
On behalf of the « Western Algerian Group of Bone Marrow Transplant »
WAG-BMT
Haematology and Cell Therapy Depatment
Hemobiology Department
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Introduction
 Autologous stem cell transplantation (ASCT) after a
high dose conditioning is an established treatment
modality with definitive indications for many
hematological disorders.
 However, this line of treatment requires many
expensive resources, such as freezing of the harvest
product in order to maintain cell viability until stemcell reinfusion and the use of growth factors for the
management of neutropenia.
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Aim of the study
 To study the viability of CD34+ stem
cells stored at +4°C
 To demonstrate the feasibility and
the safety of the ASCT without any
cryopreservation neither the use of
growth factors.
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Study design
 STEP 1: Short Conditioning regiments
ASCT in MM (MEL200) in 75 patients
ASCT in HL (CBV) in 17 patients
 STEP 2: Long conditioning regiments
ASCT in HL (BEAM) in 02 patients
ASCT in HL and NHL (EAM) in 15 patients
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STEP 1
Material and methods
75 patients with Multiple Myeloma
Figure 1. Schema of procedure.
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STEP 1
Material and methods
17 patients with Hodgkin Lymphoma
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STEP 1
Material and methods
Wannesson L et al. Annals of Oncology 18: 623–632, 2007
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STEP 1
Material and methods
 A 2ml sample was taken from 10 collection bags and
then was sent to the flow cytometry laboratory where
it was stored in the refrigerator at +4°C.
 A CD34+ stem cells count were performed using a
“Lyse no wash double platform CD34+ flow cytometry
assay”
 The viability of CD34+ cells was assessed with 7'AAD
immediately after the end of apherisis (H0) then at
(H24), (H48) and finally at (H72).
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H0
H24
Viability (median) 98%
97.6%
Viability (range) 97.4 – 100 92.4 – 99.4
STEP 1
Results
H48
96%
91.6-98
H72
95%
91-96
STEP 1
Results
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y = -1,06x + 99,3
R2 = 0,9571
p=0.05
CD34+ stem cells viability kenetic curve
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STEP 1
Results
Brahimi M et al. Revue Algerienne D’hematologie 2011 ; 5 :46-48.
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STEP 1
Results
n=200
Time of storage of the Harvest bags at +4°C
STEP 1
Results
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CBV (HODGKIN) Mel200 (MYELOMA)
(n = 17)
(n=75)
CD34+ cells/Kg
Duration of neutropenia
Duration of thrombocytopenia,
No. of platelet transfusions,
No. of red blood cell transfusions,
Duration of intravenous antibiotics,
Duration of hospitalization,
Transplant Related Mortality
3,66 (2,3-13,22)
13(9-24)
12(9-37)
3,66 (1,5 - 9,7)
10(06-17)
13(9-24)
1(0-2)
2(0-8)
9(0-15)
21(15-37)
01
1(0-3)
2(0-9)
4(0-13)
17(12-30)
00
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Mel200 protocol
Bekadja MA et al. Hematol Oncol Stem Cell Ther 2012; 5(1): 49-53
Bekadja MA et al. EBMT 38th meetting: Geneva, Switzerland 1-4 april 2012.page
140 (Abstract 905)
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CBV protocol
Bekadja MA et al. Revue Algerienne D’hematologie 2011 ; 5 :50-53.
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STEP 2
«BEAM» VS «-EAM»
BEAM (n=2)
-EAM (n=15)
BCNU (mg/m2)
300 (D-2)
---
VP16 (mg/m2)
200 (D-5 to D-2)
200 (D-5 to D-2)
ARAC (mg/m2)
400 (D-5 to D-2)
2000 (D-5 to D-2)
140 (D-1)
140 (D-1)
MELPHALAN (mg/m2)
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STEP2
Material and methods
15 Patients with Hodgkin Lymphoma
Bekadja MA et al. EBMT 38th meetting: Geneva, Switzerland 1-4 april 2012.page 162
(Abstract 1140)
Talhi S et al. Hématologie, Vol. 1, Supplement 1, Mars 2012: pp 213 (Abstract)
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STEP2
RESULTS
n=30
Time of storage of the Harvest bags
STEP2
RESULTS
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CD34+ cells/Kg
Duration of neutropenia
Duration of thrombocytopenia,
No. of platelet transfusions,
No. of red blood cell transfusions
Transplant Related Mortality
BEAM (HODGKIN)
(n = 2)
-EAM (HODGKIN)
(n=15)
6,98 (4,42-9,53)
12(11-13)
11(11-12)
3,68 (2,71 - 6,15)
13(10-32)
13(9-24)
2(1-4)
2(2-3)
00
3(2-4)
4(2-9)
00
STEP2
RESULTS
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Viability ( % )
Per Bag
MNC ( % )
Per Bag
CD34 (% )
Per Bag
DAY : 0 (n=10)
DAY : 6 (n=10)
93,77 ( 70-100 )
82,18 ( 54,24 – 94 )
51,24 ( 25,85 – 71,21 )
57,05 ( 39,71 – 75 )
0,73 ( 0,16 – 1,94 )
0,34 ( 0,19 – 0,54 )
Brahimi M et al. Unpublished data.
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Conclusion
 We conclude that high-dose
chemotherapy with non-frozen
peripheral stem cells is safe in terms
of haematopoietic reconstitution even
without using growth factors.
Department of Hemobiology
EHU 1st November, Oran, Algeria
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Department of Hematology and Stem Cell Therapy
EHU 1st November Oran, Algeria
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23
Chief of Biology Pôle in EHU 1st November
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Professor Mohamed Amine BEKADJA
Bekadja MA et al. Revue Algérienne d’Hématologie 2011 ; 5 :50-53.
Bekadja MA et al. Hematol Oncol Stem Cell Ther 2012; 5(1): 49-53
Bekadja MA et al. EBMT 38th meetting: Geneva, Switzerland 1-4 april 2012.,
(Abstract 905)
Bekadja MA et al. EBMT 38th meetting: Geneva, Switzerland 1-4 april 2012,
(Abstract 1140)