U.S. patent application number 10/238738 was filed with the patent office on 2003-06-19 for hematopoietic stem cell proliferating agents.
This patent application is currently assigned to Fujisawa Pharmaceutical Co. Ltd.. Invention is credited to Kobayashi, Masakazu, Saito, Yoshimasa, Takata, Yoko, Tamura, Kouichi, Ueda, Yoshiko, Yamada, Hisashi, Yamashita, Tatsuo.
Application Number | 20030113912 10/238738 |
Document ID | / |
Family ID | 26346424 |
Filed Date | 2003-06-19 |
United States Patent
Application |
20030113912 |
Kind Code |
A1 |
Saito, Yoshimasa ; et
al. |
June 19, 2003 |
Hematopoietic stem cell proliferating agents
Abstract
This invention relates to a hematopoietic stem cell
proliferating agent comprising IGF-I, a hematopoietic stem cell
proliferating agent comprising IGF-I and at least one protein
selected from among SCF, M-CSF, and G-CSF, and a method of growing
hematopoietic stem cells which comprises culturing hematopoietic
stem cells in a medium containing IGF-I and at least one protein
selected from the group consisting of SCF and M-CSF. The
hematopoietic stem cell proliferating agent of the invention causes
hematopoietic stem cells to proliferate in the undifferentiated
state whether in vivo or in vitro and can, therefore, be used for
amelioration of the cytopenia induced by radiotherapy or
chemotherapy using anticancer drugs, prevention of infectious
diseases associated with lymphopenia, or in vitro culture for
multiplication of hematopoietic stem cells and extrasomatic culture
of recombinant stem cells in gene therapy.
Inventors: |
Saito, Yoshimasa;
(Kawanishi-shi, JP) ; Ueda, Yoshiko; (Osaka-shi,
JP) ; Tamura, Kouichi; (Kobe-shi, JP) ;
Takata, Yoko; (Osaka-shi, JP) ; Yamada, Hisashi;
(Sanda-shi, JP) ; Yamashita, Tatsuo; (Kobe-shi,
JP) ; Kobayashi, Masakazu; (Takarazuka-shi,
JP) |
Correspondence
Address: |
OBLON SPIVAK MCCLELLAND MAIER & NEUSTADT PC
FOURTH FLOOR
1755 JEFFERSON DAVIS HIGHWAY
ARLINGTON
VA
22202
US
|
Assignee: |
Fujisawa Pharmaceutical Co.
Ltd.
Osaka
JP
|
Family ID: |
26346424 |
Appl. No.: |
10/238738 |
Filed: |
September 11, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10238738 |
Sep 11, 2002 |
|
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09147689 |
May 13, 1999 |
|
|
|
09147689 |
May 13, 1999 |
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PCT/JP97/02818 |
Aug 12, 1997 |
|
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Current U.S.
Class: |
435/372 ;
514/7.9; 514/8.6; 530/303 |
Current CPC
Class: |
A61K 38/30 20130101;
A61K 38/30 20130101; A61K 2300/00 20130101 |
Class at
Publication: |
435/372 ;
530/303; 514/2 |
International
Class: |
A61K 038/00; A61K
038/28; C12N 005/08 |
Foreign Application Data
Date |
Code |
Application Number |
Aug 13, 1996 |
JP |
8/213641 |
Jan 24, 1997 |
JP |
9/11054 |
Claims
1. A hematopoietic stem cell proliferating agent comprising
IGF-I.
2. A hematopoietic stem cell proliferating agent according to claim
1 which comprises IGF-I and at least one protein selected from the
group consisting of M-CSF, SCF, and G-CSF.
3. The hematopoietic stem cell proliferating agent according to
claim 2 which comprises M-CSF and IGF-I.
4. The hematopoietic stem cell proliferating agent according to
claim 2 which comprises SCF and IGF-I.
5. The hematopoietic stem cell proliferating agent according to
claim 2 which comprises G-CSF and IGF-I.
6. A method of growing hematopoietic stem cells which comprises
culturing hematopoietic stem cells in a medium containing IGF-I and
at least one protein selected from the group consisting of M-CSF,
SCF, and G-CSF.
7. A method of growing hematopoietic stem cells which comprises
administering IGF-I to a mammal.
8. The method of growing hematopoietic stem cells according to
claim 7 which comprises administering IGF-I and at least one
protein selected from the group consisting of M-CSF, SCF, and
G-CSF.
9. The method of growing hematopoietic stem cells according to
claim 8 which comprises administering IGF-I and M-CSF.
10. The method of growing hematopoietic stem cells according to
claim 8 which comprises administering IGF-I and SCF.
11. The method of growing hematopoietic stem cells according to
claim 8 which comprises administering IGF-I and G-CSF.
12. Use of IGF-I for stimulating proliferation of mammalian
hematopoietic stem cells.
13. Use of IGF-I and at least one protein selected from the group
consisting of M-CSF, SCF, and G-CSF for stimulating proliferation
of mammalian hematopoietic stem cells.
14. Use of IGF-I and M-CSF for stimulating proliferation of
mammalian hematopoietic stem cells.
15. Use of IGF-I and SCF for stimulating proliferation of mammalian
hematopoietic stem cells.
16. Use of IGF-I and G-CSF for stimulating proliferation of
mammalian hematopoietic stem cells.
Description
TECHNICAL FIELD
[0001] This invention relates to a hematopoietic stem cell
proliferating agent and a method for stimulating the proliferation.
More particularly, the invention relates to a hematopoietic stem
cell proliferating agent comprising insulin-like growth factor I
either alone or in combination with some or other
colony-stimulating factors and/or growth factors and to a method
for proliferating.
BACKGROUND ART
[0002] A variety of growth factors relating to the blood system are
under investigation, and erythropoietin (EPO) as an erythropoiesis
stimulating agent and granulocyte colony-stimulating factor (G-CSF)
as a leukopoiesis stimulating agent, among others, have been
clinically put to use to this day. Regarding the technology for
proliferating of hematopoietic stem cells, various cytokines such
as stem cell factor (SCF), macrophage colony-stimulating factor
(M-CSF), etc. were explored but none has been found to be effective
enough in causing hematopoietic stem cells to multiply sufficiently
in the undifferentiated form.
[0003] As the result of an intensive investigation, the inventors
of this invention found that hematopoietic stem cells can be
successfully caused to multiply in the undifferentiated state well
by using insulin-like growth factor I (IGF-I) in combination with
at least one protein selected from among SCF, M-CSF, and G-CSF.
This invention has been developed on the basis of the above
finding.
DISCLOSURE OF THE INVENTION
[0004] This invention relates to a hematopoietic stem cell
proliferating agent comprising IGF-I, a hematopoietic stem cell
proliferating agent comprising IGF-I and at least one protein
selected from among SCF, M-CSF, and G-CSF, and a method of
stimulating proliferation of hematopoietic stem cells which
comprises culturing hematopoietic stem cells in a culture medium
containing IGF-I and at least one protein selected from among SCF,
M-CSF, and G-CSF. Furthermore, this invention relates to a method
of proliferating of hematopoietic stem cells in a mammal which
comprises using IGF-I alone or in combination with at least one
protein selected from among SCF, M-CSF, and G-CSF.
[0005] Since the hematopoietic stem cell proliferating agent and
method of the invention are effective in causing hematopoietic stem
cells to multiply in the undifferentiated state whether in vivo or
in vitro, the invention finds application in the management of the
cytopenia induced by radiotherapy or chemotherapy using anticancer
drugs, prevention of infectious diseases with which lymphopenia is
associated, treatment of myelopathy inclusive of
osteomyelodysplasia and bone marrow suppression, therapy of marrow
diseases such as leukemia-advanced renal impairment-bone marrow
suppression, improvement in engraftment survival in bone marrow
transplantation, therapy of hypocytosis associated with inherited
diseases, in vitro culture for multiplication of hematopoietic stem
cells, and extrasomatic culture of recombinant stem cells in gene
therapy, among other uses.
[0006] The IGF-I which can be used in this invention includes the
corresponding proteins derived from human, bovine, and other
mammals by recombinant DNA technology (e.g. Kokai Tokkyo Koho
S61-1396 for IGF-I), peptide synthesis, cell culture, or other
technology and even the muteins having IGF-I activity which can be
derived from the recombinant or other IGF-I by partial modification
of its amino acid sequence by substitution, insertion, addition, or
deletion of one or more amino acid residues (e.g. WO89/05822).
[0007] The SCF, M-CSF, or G-CSF which can be used in this invention
includes the corresponding proteins derived from human, bovine, or
other mammals by recombinant DNA technology, peptide synthesis,
cell culture, or other technology and even those muteins having
SCF, M-CSF, or G-CSF activity which can be derived from the
recombinant or other SCF, M-CSF or G-CSF by partial modification of
its amino acid sequence by substitution, insertion, addition, or
deletion of one or more amino acid residues. Here, the sugar chain
may be present or absent.
[0008] The hematopoietic stem cell proliferating agent of this
invention which comprises IGF-I and either SCF, M-CSF, or G-CSF can
be administered virtually without limitations on the dosage form or
forms, sequence of administration, or route of administration, all
that is necessary being to insure that those factors will be
concurrently available in the recipient's body. For example, they
can be administered as a mixture in a single dosage form or in
independent dosage forms, whether concurrently or one after
another, and whether by the same route or by different routes.
[0009] The hematopoietic stem cell proliferating agent of the
invention is generally provided in an oral dosage form or in a
nonoral dosage form, e.g. an injection, drip infusion, transdermal
therapeutic system, transnasal therapeutic system, external
preparation, suppository, etc., each containing IGF-I alone or
IGF-I plus at least one protein selected from among SCF, M-CSF and
G-CSF together with a carrier (e.g. distilled water for injection,
physiological saline, glucose injection, etc.), a stabilizer (e.g.
albumin, sodium citrate, arginine, dextran, etc.), a pH control
agent (e.g. sodium monohydrogen phosphate, sodium dihydrogen
phosphate, etc.) and other additives. Such dosage forms or systems
may further contain one or more growth factors such as SCF, M-CSF,
G-CSF, EPO, and IL-3.
[0010] The dosage of this hematopoietic stem cell proliferating
agent depends on the patient's body weight, sex, and clinical
condition but the dose level for an adult human is generally about
1.about.1000 .mu.g/kg in terms of IGF-I and preferably about
5.about.500 .mu.g/kg on the same basis. When SCF, M-CSF, or G-CSF
is used concomitantly, SCF, M-CSF or G-CSF can be formulated in the
same amount as IGF-I or in an amount ranging from 0.01 to 100 times
the amount of IGF-I.
[0011] This hematopoietic stem cell proliferating agent can be
administered orally or otherwise, e.g. by intravenous injection,
intravenous drip, subcutaneous injection, coronary intraarterial
administration, transdermal administration, transnasal
administration, or rectal administration.
[0012] IGF-I and any of SCF, M-CSF and G-CSF can be formulated in
one and the same dosage form but may be administered independently
one after the other or concurrently, either by the same route or
different routes. When they are administered in sequence, it does
not matter which is administered first.
[0013] When the hematopoietic stem cell proliferating agent of the
invention is used for ameliorating the cytopenia induced by
radiotherapy or chemotherapy using anticancer drugs, the
hematopoietic stem cell proliferating agent of the invention can be
used alone or concurrently with EPO, M-CSF, SCF, IL-3, G-CSF,
and/or the like. When it is used for the prevention of infectious
diseases associated with lymphopenia, GM-CSF or the like can be
used concomitantly. Furthermore, in the treatment of myelopathies
such as osteomyelodysplasia and bone marrow suppression or marrow
diseases such as leukemia-advanced kidney impairment-bone marrow
suppression, for improvement in engraftment survival in bone marrow
transplantation, or in the treatment of hypocytosis associated with
inherited diseases, suitable growth factors such as EPO, G-CSF,
GM-CSF, etc. can be used as concomitant medications.
[0014] In the in vitro culture for multiplication of hematopoietic
stem cells or extrasomatic culture of recombinant stem cells in
gene therapy, among other applications, the hematopoietic stem cell
proliferating agent of the invention can be used in combination
with other suitable growth factors. In vitro culture of marrow
cells can be essentially carried out in accordance with the method
described in Shin Seikagaku Jikken Koza [New Biochemical Experiment
Series] 18 Saibo Baiyo Gijutsu [Cell Culture Technology] (ed. by
Japanese Biochemical Society, Tokyo Kagaku Dojin, 1989). For
example, using a CO.sub.2 incubator, marrow cells can be cultured
in RD medium [RPMI1640:DMEM=1:1 (v/v)] supplemented with insulin,
transferrin, 2-mercaptoethanol, ethanolamine, selenious acid,
HEPES, etc. in the presence of IGF-I (1.about.1000 .mu.g/ml) and at
least one protein selected from among SCF (1.about.1000 .mu.g/ml)
M-CSF (1.about.1000 .mu.g/ml), and G-CSF (1.about.1000
.mu.g/ml).
BEST MODE FOR CARRYING OUT THE INVENTION
[0015] This invention is now described in further detail by way of
the following examples.
EXAMPLE 1
Procurement of Hematopoietic Stem Cells
[0016] Using the femora of male C57BL mice (10 individuals), marrow
cells were recovered in .alpha.-MEM (5 ml, Nikken Biomedical
Research Institute). This cell suspension was centrifuged (1,200
rpm, 10 min.), the supernatant was aspirated off, and the cells
were resuspended in 10% FCS-.alpha.-MEM (5 ml). By this procedure,
approximately 3.times.10.sup.7 marrow cells per mouse were
harvested.
EXAMPLE 2
FACS
[0017] From the marrow cells thus procured, Sca-1.sup.+, Lin.sup.-,
and c-kit.sup.+cells were collected using Fac-Scan flow cytometer
(Becton-Dickinson). The labeled antibodies used are shown in Table
1. It is known from the literature that the cell fraction thus
obtained contains murine hematopoietic stem cells [Okada, S. et al.
(1992), Blood 80, 3044-3050].
1TABLE 1 Labeled antibodies used in FACS Marker Labeled antibody
Specificity Lin Biotinylated anti-CD3 .epsilon. (Clone T cell
500A2) Biotinylated anti-CD45R (B220) B cell Biotinylated
anti-mouse Erythrocyte erythrocytes (TER119) Biotinylated
anti-CD11b (Mac-1) Monocyte/ macrophage Biotinylated anti-myelocyte
Granulocyte differentiation antigen (Gr1) Sca-1 PE-labeled
anti-mouse SCA-1 (E13- Stem cell 161.7) c-kit PITC-labeled
anti-mouse CD117 (3CI) SCF receptor
EXAMPLE 3
Action of IGF-I
[0018] A U-bottomed 96-well microtiter plate (Nunc, Denmark) was
seeded with hematopoietic stem cells at a density of 50 cells/10%
FCS-.alpha.-MEM (100 .mu.g 1)/well.
[0019] To each well was added IGF-I (Mecasermin, recombinant,
Fujisawa Pharmaceutical) (100 ng/ml) SCF (recombinant, Genzyme)
(1.5, 3.0, 6.0, 12.5, 25 ng/ml)+IGF-I (100 ng/ml), or M-CSF
(recombinant, R&D System) (0.1, 0.3, 1.0, 3.0, 10, 30, 100
ng/ml)+IGF-I (100 ng/ml) and the plate was incubated in a CO.sub.2
incubator at 37.degree. for 10 days.
[0020] After 6 days of culture, the cells in each well were
counted.
[0021] After 10 days of culture, the medium was aspirated off and
the cytoplasmic acid phosphatase activity was assayed by the method
described in the literature [Ueda et al. (1994), Neurosci. Lett.,
165, 203-207].
EXAMPLE 4
Results
[0022] The results are summarized in Table 2. Table 2 Effect of the
combination of IGF-I with either
2 SCF or M-CSF IGF-I (ng/ml) Factor added (ng/ml) .DELTA. 450 means
.+-. SD 100 SCF 25.0 1.387 .+-. 0.207 100 SCF 12.5 0.654 .+-. 0.273
100 SCF 6.0 0.056 .+-. 0.015 100 SCF 3.0 0.035 .+-. 0.010 0 SCF
25.0 0.176 .+-. 0.074 100 M-CSF 100.0 0.906 .+-. 0.645 100 M-CSF
30.0 0.778 .+-. 0.649 100 M-CSF 10.0 0.564 .+-. 0.402 100 M-CSF 3.0
0.064 .+-. 0.050 0 M-CSF 100.0 0.052 .+-. 0.045 100 None ND ND:
below detection limit
[0023] It will be apparent from Table 2 that IGF-I as used in
combination with SCF or M-CSF increased the cytoplasmic acid
phosphatase activity of the marrow cell fraction obtained in
Example 2. It is known that cytoplasmic acid phosphatase activity
increases in proportion to an increase in the cell population.
Therefore, it is clear that IGF-I as used in combination with SCF
or M-CSF exerted a hematopoietic stem cell proliferation
stimulating action. It is known from the literatrue that SCF and
M-CSF each independently does not show hematopoietic stem cell
proliferation stimulating activity [Okada, S. et al. (1992), Blood
80, 3044-3050] and the inventors verified the finding.
EXAMPLE 5
FACS
[0024] Using the same procedure as used in Example 2, Lin.sup.-,
Sca-1.sup.+, c-kit.sup.+, and CD34.sup.- cells were harvested. For
selection of CD34-cells, biotinylated anti-mouse CD34 (RAM34)
(Fermigen, San Diego, Calif.) was newly used. Those cells accounted
for about 0.04% of the total marrow cell population. It is known
from the literature that cells of this group are hematopoietic stem
cells (Ohsawa & Nakauchi 1995, Japanese Molecular Biochemical
Society S4B-3; Ohsawa et al. 1995, Synopsis of Blood Stem Cell
Symposium).
EXAMPLE 6
Effect of IGF-I
[0025] Using ACDU, the cells obtained in Example 5 were transferred
to 96-well plates, one cell per well, and about 50 wells were used
as one group.
[0026] To each well was added SCF (25 ng/ml)+IGF-I (100 ng/ml) ,
M-CSF (100 ng/ml)+IGF-I (100 ng/ml), or SCF (25 ng/ml)+IL-3 (10
ng/ml), and the plates were incubated in a CO.sub.2 incubator at
37.degree. C. for 10 days.
[0027] After 6 days of culture, the cells in each well were
counted.
[0028] After 10 days of culture, the medium was aspirated off from
each well and the cytoplasmic acid phosphatase activity was
assayed.
EXAMPLE 7
Results
[0029] The results are summarized in Table 3. Table 3 Effect of the
combination of IGF-I with either
3 SCF or M-CSF on the single cell Mean number of Surviving wells/
surviving all wells cells/ (survival rate) well IGF-I (100 ng) +
25/47 (53%) 10 SCF (25 ng/ml) IGF-I (100 ng) + 3/47 (6.0%) 2 M-CSF
(100 ng/ml) Medium 0/23 (0.0%) 0 IL-3 (10 mg) + 33/48 (69%) NT SCF
(25 ng/ml) NT: not counted
[0030] It will be apparent from Table 3 that IGF-I as used in
combination with SCF or M-CSF is capable of causing proliferation
of singular hematopoietic stem cells.
EXAMPLE 8
Immunostaining
[0031] Sca-1.sup.+, Lin.sup.-, and c-kit.sup.+ cells were collected
by the same procedure as in Examples 1 and 2. The harvested cells
were seeded on a 100-well chamber slide (Lab-Tek, Nunc). In a
CO.sub.2 incubator at 37.degree. C., the cells were cultured in 10%
FCS-.alpha.-MEM (100 .mu.l) containing growth factors [IGF-I (100
ng/ml) and SCF (25 ng/ml)] for 8 days.
[0032] The cultured cells were fixed with acetone (-20.degree. C.)
on the slide. After addition of 1% BSA-phosphate buffer for
inhibition of nonspecific binding, the cells were treated with the
antibodies shown in Table 4.
[0033] The cells were not stained by any of the antibodies used,
indicating that they had no tendency toward differentiation to any
specific kinds of offspring cells. The augmentation of cell
population by IGF-I plus SCF was not the proliferation due to
differentiation of stem cells but the proliferation of
undifferentiated stem cells.
4TABLE 4 Labeled antibodies used in FACS Marker Labeled antibody
Specificity Lin Biotinylated anti-CD3 .epsilon. (Clone T cell
500A2) Biotinylated anti-CD45R (B220) B cell Biotinylated
anti-mouse Erythrocyte erythrocytes (TER119) Biotinylated
anti-CD11b (Mac-1) Monocyte/ macrophage Biotinylated anti-myelocyte
Granulocyte differentiation antigen (Gr1) Sca-1 PE-labeled
anti-mouse SCA-1 (E13- Stem cell 161.7)
EXAMPLE 9
The Action of Human G-CSF Plus Human IGF-I
[0034] Hematopoietic stem cells were transferred to a U-bottomed
96-well microtiter plate (Nunc, Denmark), 50 cells/10%
FCS-.alpha.-MEM (100 .mu.l)/well.
[0035] To each well was added human IGF-I (Mecasermin, recombinant,
Fujisawa Pharmaceutical) (100 ng/ml) or human G-CSF (recombinant,
R&D System) (5, 50, or 500 mg/ml)+IGF-I (100 ng/ml), and the
plate was incubated in a CO.sub.2 incubator at 37.degree. C.
[0036] After 7 days of culture, the medium was aspirated off from
each well and the cytoplasmic acid phophatase activity was assayed
by the method described in the literature (Ueda et al. (1994),
Neurosci. Lett., 165, 203-207).
EXAMPLE 10
Results
[0037] The results are summarized in Table 5.
5TABLE 5 G-CSF (ng/ml) IGF-I (ng/ml) .DELTA. 450 mean .+-. SE 500
100 0.164 .+-. 0.033 50 100 0.132 .+-. 0.086 5.0 100 0.126 .+-.
0.071 0.50 100 0.038 .+-. 0.020 0 100 0.001 .+-. 0.004 500 0 0.054
.+-. 0.007 50 0 0.015 .+-. 0.019 5.0 0 0.010 .+-. 0.014 0.50 0
0.008 .+-. 0.003 0 0 0.000 .+-. 0.002
[0038] It will be apparent from Table 5 that IGF-I as used in
combination with G-CSF enhanced the cytoplasmic acid phosphatase
activity of the marrow cell fraction obtained in Example 2. It is
known that cytoplasmic acid phosphatase activity is positively
correlated with the number of cells. Therefore, it is evident that
IGF-I as used in combination with G-CSF exhibited a hematopoietic
stem cell proliferation stimulating action. No effect was found
with G-CSF alone.
EXAMPLE 11
The Action of Human SCF Plus Human IGF-I
[0039] Hematopoietic stem cells were transferred to a U-bottomed
96-well microtiter plate (Nunc, Denmark), 50 cells/10%
FCS-.alpha.-MEM (100 .mu.l)/well.
[0040] To each well was added human SCF (recombinant, R&D
System) (5, 50, or 500 mg/ml)+IGF-I (100 ng/ml) and the plate was
incubated in a CO.sub.2 incubator at 37.degree. C.
[0041] After 7 days of culture, the cells in each well were
counted.
EXAMPLE 12
Results
[0042] The results are summarized in Table 6.
6 TABLE 6 Mean .+-. SE Mean .+-. SE SCF (ng/ml) (IGF-I 100 ng/ml)
(IGF-I 0 ng/ml) 800 200 .+-. 0.0 75.7 .+-. 26.4 400 175 .+-. 21.5
36.0 .+-. 14.3 200 58.0 .+-. 16.7 14.8 .+-. 8.2 100 31.5 .+-. 6.2
9.8 .+-. 1.6 50 16.0 .+-. 5.5 6.5 .+-. 5.0 25 6.3 .+-. 2.9 3.5 .+-.
0.5 12.5 3.8 .+-. 1.5 2.0 .+-. 0.7 6.25 3.3 .+-. 0.8 3.0 .+-. 3.7
3.0 3.5 .+-. 1.1 2.3 .+-. 1.1 0 2.8 .+-. 2.2 1.0 .+-. 1.0
[0043] It will be apparent from Table 6 that IGF-I as used in
combination with SCF stimulated proliferation of hematopoietic stem
cells to a remarkable extent. This combination is, therefore,
useful for the maintenance and multiplication of hematopoietic stem
cells.
EXAMPLE 13
Pharmaceutical Preparations
[0044] The components indicated in Table 7 were dissolved in water
and vials filled with 5 ml portions of the respective solutions
were lyophilized to provide hematopoietic stem cell proliferating
agents.
7TABLE 7 Examples of the hematopoietic stem cell proliferation
stimulating composition Human IGF-I SCF M-CSF G-CSF albumin (mg)
(mg) (mg) (mg) (mg) Composition 1 10 0 0 0 0 Composition 2 10 0 0 0
50 Composition 3 10 20 0 0 50 Composition 4 10 0 10 0 50
Composition 5 10 0 0 10 50 Composition 6 5 10 10 0 0 Composition 7
5 0 5 5 100 Composition 8 5 10 0 5 100 Composition 9 5 5 5 5 50
INDUSTRIAL APPLICABILITY
[0045] Thus, the hematopoietic stem cell proliferating agent and
method of the invention are of great utility value, for
hematopoietic stem cells can be caused to proliferate in the
undifferentiated state whether in vivo or in vitro.
* * * * *