Champlin, R.

Gao, H.

Korbling, M.

Lee, B.

Reuben, J.M.

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Poster  P resentat ions  - Sess ion  II 
GRAFT PROCESSING 
167 
MOBILIZED PERIPHERAL BLOOD CONTAINS PRIMITIVE HEMATOPOI- 
ETIC CELLS THAT CAN BE ENUMERATED AND ISOLATED USING A 
FLUORESCENT SUBSTRATE FOR ALDEHYDE DEHYDROGENASE 
ACTIVITY 
Fallon, P.z; GelJtO,, T.~; Balber, A.s; Boul~vm'e, D/,  ffanssen, I V.1; 
Smilee, R. z;StoJwTs, R.2; Smith, C~ l. Department of l*zterdisciplinmy 
Oncologo~, lojCfi'tt Cancer Ce*lter, TaTnpa, FL; 2. Dzlke University 
Medical Center, Durham, NC; 3. StemCo Biomedical, Durham, NC. 
High dose therapy followed by autolog'ous or allogeneic trans- 
plantation with peripheral blood stem cells (PBSCs) has been used 
to treat patients with a variety of diseases. Selection of primitive 
stem cells and progenitors from PBSC collections is useful for 
reducing the transplant volume and decreasing the number of con- 
taminating tmnor cells or T-cells. We have developed a novel 
approach for enmnerating and enriching primitive mobilized 
peripheral blood cells that express high levels of the enzyme alde- 
hyde dehydrogenase (ALDH). Mobilized cells were stained with a 
fluorescent ALDH substrate, termed BODIPY-aminoacetaldehyde 
(BAAA), and then analyzed or sorted using flow cytometry. A pop- 
ulation of cells, termed SSCloALDHbr cells, was readily discrimi- 
nated and comprised amean of 3.1 +4.8% of the collected events. 
A mean of 73.4 +11.7% of the SSCloALDHbr population 
expressed CD34 and 56 + 24.5% of all the mobilized CD34+ cells 
resided within the SSCloALDHbr population. The SSCloALD- 
Hbr population was largely depleted of cells with mature pheno- 
types and enriched for cells with immature phenotypes. The BAAA 
staining procedure did not diminish the viability or clonogenic 
activity of hematopoietic progenitors and caused no toMcity to cells 
or animals in a variety of pre-clinical toxicology studies. Sorted 
SSCloALDHbr and SSCloALDHbr CD34+ cells were enriched 
for progenitors with the ability to 1) generate CFUs and LTC 
derived CFUs, 2) expand in primary and secondary LTCs and 3) 
generate multiple cell lineages. In order to test whether the num- 
ber of PBSC SSCloALDHbr cells would predict engraftment in
actual human transplants, the total number of SSCIoALDHbr cells 
infused per kg in 21 cancer patients who had undergone autoPBSC 
transplantation were compared to the times to an ANC>500 and a 
platelet count >20, 000 using a Cox proportional hazard analysis. 
The time to neutrophil and platelet engraftment were both highly 
correlated with the number of SSCloALDHbr cells/kg (p<0.015 
and 0.003 respectively) and the number of SSCloALDHbr CD34+ 
cells/kg (p<0.013 and 0.0016 respectively). In smnmary, PBSC 
SSCloALDHbr cells have the phenotypic and functional properties 
of primitive hematopoietic cells and their number correlates with 
short erm engraftment following autoPBSCT transplantation. 
168 
NORMAL AND NEOPLASTIC B CELLS ARE UNDULY SUSCEPTIBLE TO 
PHOTODYNAMIC CELL THERAPY 
Roy, D.C/; Dallaire, N J; Krosl, GJ; Autonia, B.S; Freeman, G.ff.~; Per- 
reazdt, C.S ; T 7illeneuve, L. s 1. Hematology-Immunolo©~, Hopital 3/Iaison- 
neuve-Rosemont, Monn'eal, QC, Cmzada; 2. Dana-Fro'bet Cancer Insti- 
tztte, Boston, [vIA; 3. Celmed BioScie~zces Inc., l~/lo*ztreal, QC, Canada. 
The favorable effect associated with the infusion of a graft 
purged of its neoplastic ells might be jeopardized by the con- 
comitant elimination of cell populations responsible for (1) the 
eradication of residual lymphoma cells, (2) the prevention of viral 
and fungal infections, and (3) rapid hematologic reconstitution 
post-transplantation. In previous studies, we have shown that 
TH9402, a rhodamine-derived photosensitizer, selectively elimi- 
nates activated T ceils, and spares resting T cells. Interestingly, P-
glycoprotein (Pgp) modulates TH9402-mediated cytotoxicity by 
extruding this photosensitizer from the intracellular milieu. The 
absence of Pgp expression in most NHL cells prompted us to 
evaluate photodynamic cell therapy (PDCT) with TH9402 for the 
selective limination of B-lineage lymphoma cells. To identify 
optimum treatment conditions, the uptake/retention ki etics and 
cytotoxicity profile of TH9402 were evaluated with B-lineage cell 
lines, NHL patient cells harboring or not a bcl-2/IgH rearrange- 
ment, and normal B lymphocytes. In clonogenic assays, more than 
4 logs of indolent bcl-2/IgH rearranged (RL and DHL-16) and 
aggressive (Namalwa nd Rail) NHL cell lines and patient (n=27) 
NHL cells were eradicated following PDCT using TH9402. 
Mthough normal B lymphocytes retained TH9402 in anmunts 
similar to T cells, as measured by flow-cytometry, PDCT elimi- 
nated 4 logs of B ceils but only 1 log of T lymphocytes, which 
were spared for imnmne reactivity. Importantly, PDCT condi- 
tions of maximum intensity that preserved more than 50% of 
CFU-GM, BFU-E and CFU-Mix colonies, and greater than 75% 
of LTC-IC progenitors for hematopoietic reconstitution, demon- 
strated lower levels of retention of TH9402 in CD34+ cells. The 
discrepancy between TH9402 retention levels and cytotoxicity 
indicates that the exquisite sensitivity of B lineage ceils to TH9402 
phototherapy must rely on a retention-independent mechanism of 
target cell elimination. The specificity" of TH0402 for B lympho- 
cytes indicates that this strategy should be evaluated for ex vivo 
purging of autologous tem cell grafts from patients with NHL 
and also for the treatment of autoimmune B cell disorders. 
169 
MOBILIZATION OF TYPE I AND TYPE 2 DENDRITIC CELLS BY GM- 
CSF ALONE OR IN COMBINATION WITH G-CSF 
Gao, H.; Korb/iug, M.; Lee, B.; Champ~in, R.; Reuben, ff.M. 
M.D.A1~de'rsmz Crower Center, Houston, TX. 
Dendritic cells (DC) are fire only antigen-presenting cells capa- 
ble of priming naive T cells with antigens. Generally, DC does 
not express lineage-specific markers (lineage negative), but express 
HLA-DR. There are two subsets of DC: one of lymphoid lineage 
that induces Thl-like responses and expresses CD11c (DC1) and 
another of myeloid lineage that induces Th2-1ike responses and 
expresses CD123 (DC2). Previous studies reported that G-CSF 
preferentially mobilized DC2, and not DC1, in nornral subjects. 
To determine the effect of GM-CSF alone or in combination with 
G-CSF on the mobilization of DC1 and DC2, we studied DC 
markers in 7 subjects received 10 mcg/kg/day of GM-CSF for 4 
days and another 5 subjects received 10 meg/ko'/day of GM-CSF 
and 10 mcg/kg/day of G-CSF for 4 days. Peripheral blood was 
obtained from all subjects before treatment and 24 hours after the 
final dose of the mobilization regimen. The numbers of DC1 and 
DC2 were determined by staining blood with lineage cocktail 
antibodies, anti-HLA-DR, anti-CD11c, and anti-CD123, and 
analyzed by 4 color flow cytometry. Compared with the number 
of total DC prior to treatment with G CSF plus GM-CSF, the 
post-treatment number of DC was significantly hip'her (32 +/- 6 
per mcl versus 143 +/- 35 per mel; p = 0.020). The increase in 
total DC post-mobilization with G-CSF and GM-CSF was due to 
a significant increase in DC2 (8 +/- 3 per mcl versus 37 +/- 19 per 
mcl; p = 0.046), and not in the number of DC1. By comparison, 
mobilization of DC, DC1, and DC2, with GM CSF alone was 
only marginal. The preferential expansion of DC2 by the combi- 
nation of G-CSF and GNI-CSF as with G-CSF alone may have 
important consequences on sensitization of the host and on the 
engraftumnt. 
170 
EFFECTIVENESS OF METRONIDAZOLE ON CYCLOSPORINE-INDUCED 
GINGIVIAL HVPERPLASIA IN BONE MARROW AND KIDNEY TRANS- 
PLANT PATIENTS:A PROSPECTIVE, DOUBLE-BLINDED CONTROLLED 
CLINICAL TRIAL 
Heshmat, ,4.; Najafi, I.; Mog,Oadmn, K.; h'awmi, M.; Ym~esiyan, 3'1.; 
Zohrevamt, P. Shariati Hospital, Teh~wz, Teh'raJz, bwz. 
Background:  Cyclosporine-induced gingival hyperplasia 
(CIGH) regression has been reported with some antibiotic thera- 
pies, especially Metronidazole and Azithromicine. Object: To 
evaluate the efficacy of metronidazole on CIGH regression, 
Method:  we performed a prospective double-blinded study on 22 
patients with CIGH, following kidney or bone marrow transplan- 
tation. None of the patients received calcimn-channel blockers or 
anticonvulsants. Other causes of gingival enlargement were ruled 


169Mobilization of type 1 and type 2 dendritic cells by GM-CSF alone or in combination with G-CSF 

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169Mobilization of type 1 and type 2 dendritic cells by GM-CSF alone or in combination with G-CSF

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