CD123+ dendritic cells in blood and lymphoid tissues

Details CD123+ dendritic cells in blood and lymphoid tissues CD123+ dendritic cells in blood and lymphoid tissues

1999-09-08

2001-09-25

Chan, Christina Y. (Department: 1644)

Chemistry: molecular biology and microbiology

Animal cell, per se ; composition thereof; process of...

C435S355000, C435S372000, C435S373000, C436S501000, C436S536000, C530S391100

Reexamination Certificate

active

06294381

ABSTRACT:

BACKGROUND OF INVENTION
Dendritic cells (DCs) play a key role in the immune system as initiators of primary T cell responses. DCs are difficult cells to isolate because they are sparsely distributed in the tissues, and because there is a lack of positive markers identifying the cells. There is, thus, little information concerning DCs in various organs, and, while various researchers have postulated on the origin of these cells, it is not definitively known whether these cells constitute a separate lineage of cells. Furthermore, while there are many theories regarding this, it is not known if there is a dendritic committed progenitor cell in the bone marrow. The importance of solving these questions has become evident from a clinical perspective, as their function has made them into critical targets for vaccine development against tumor cells, and, potentially, in the development of vaccines against HIV infection.
Furthermore, since some DCs have been observed to serve as reservoirs for the HIV virus, identification of subtypes of cells having differences in infectability potential and/or the ability to provide infection resistance, is becoming increasingly desirable. In addition, there is considerable interest in identifying subsets of DCs believed to be involved in the induction of donor-specific tolerance after transplantation. Thus, there exists a need for methodologies to characterize and isolate dendritic cells.
Such methodologies would also be useful in the isolation of DCs by utilization of the phenotype by use of cytometry. Flow cytometry is particularly useful in this regard, as the cells bearing a particular phenotype can be recognized and sorted by use of labeled antibodies. Thus, DCs of a known phenotype could be sorted and isolated.
SUMMARY OF INVENTION
A CD34
+
(as used herein, any cell designated as positive (
+
) for a particular marker will be a cell which expresses that marker as evidenced by the binding of an antibody directed against the recited marker; a cell designated as negative (−) does not express the marker as measured by the same methodology) mammalian, preferably human, bone marrow precursor of a distinct type of DCs that can be identified in peripheral blood, thymus, spleen, tonsil and lymph nodes solely by the positive expression of CD123 and CD4 has been characterized. This CD34
+
population of bone marrow cells contains primitive progenitor cells capable of reconstituting the entire hematopoietic system and more differentiated progenitors committed to a single lineage. Subsets of progenitor cells can be identified by specific cell surface markers. It is known that the receptor for macrophage-colony-stimulating factor (M-CSFR/CD115) is expressed on granulomonocytic progenitors, but not on primitive, erythroid, or CD19
+
B-lymphoid progenitors. A population of CD34
lo
(as used herein, the magnitude of expression of a particular antigen can be designated as lo or hi; such cells express the antigen, as measured by the binding of an antibody directed against the recited antigen, to a lesser (for lo) or greater (for hi) magnitude than at least 50%, preferably 75%, more preferably 90% of the cells which are positive for the recited antigen) cells (3±0.8% of the CD34
+
cells) was identified that stained brightly with antibodies to the interleukin 3 receptor alpha chain (IL3R alpha/CD123) and weakly with antibodies to the M-CSFR (see FIGS.
1
A and
1
B). Similar cells could be generated by short term culture of M-CSFR
+
cells, suggesting that the cells belong to the granulomonocytic lineage (see FIG.
1
C). Unlike most granulomonocytic progenitors in fetal bone marrow, the CD123
hi
cells had low levels of CD64 and CD13, but expressed high levels of CD36, a molecule found on monocytic cells. Further characterization showed that the cells had several phenotypic characteristics of DC precursors isolated from peripheral blood (CD3
−
, CD4
+
, CD11c
−
, CD13
lo
, CD14
−
, CD16
−
, CD19
−
, CD33
+
, CD45RA
+
, CD45RO
−
, CD56
−
, CD64
−
, HLA-DR
+
). Morphologically the cells had a smooth plasma membrane, abundant agranular cytoplasm and reniform or slightly lobulated nucleus (see FIG.
1
D).


REFERENCES:
patent: 0 563 485 A1 (1993-10-01), None
patent: WO 95/28479 A1 (1995-10-01), None
C. Caux Et Al.: “Interleukin-3 cooperates with tumor necrosis factor alpha for the development of human dendritic_Langerhans cells from cord blood CD34+ hematopoietic progenitor cells.” Blood, vol. 87, No. 6, pp. 2376-2385, New York, NY, USA (1996).
B. Herbst Et Al.: “In virto differentiation of CD34+ hematopoietic progenitor cells toward distinct dendritic cell subsets of the birbeck granule and MIIC-positive Langerhans cell and the interdigitating dendritic cell type.” Blood vol. 88, No. 7, pp. 2541-2548 New York, NY, USA (1996).
W. Egner Et Al.: “The phenotype of freshly isolated and cultured human bone marrow allostimulatory cells: possible heterogeneity in bone marrow dendritic cell populations.” Immunology, vol. 85, No. 4, pp. 611-620, Oxford, GB, (1995).
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Galy, A. et al. Immunity 3: 459-473 (1995).
Santiago-Schwartz, F. et al. Blood 82 (10): 3019-3028 (1993).
O'Doherty, U. et al., “Human Blood Contains Two Subsets of Dendritic Cells, One Immunologically Mature and the Other Immature,”Immunology, 82:487-493 (1994).

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