Note: Claims are shown in the official language in which they were submitted.
97
WHAT IS CLAIMED IS:
1. A method of ex-vivo expanding a population of hematopoietic stem
cells, while at the same time, substantially inhibiting differentiation of the
hematopoietic stem cells ex-vivo, the method comprising providing
hematopoietic
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells, with ex-vivo
culture
conditions for ex-vivo cell proliferation and, at the same time, for reducing
an
expression and/or activity of CD38, thereby expanding a population of said
hematopoietic stem cells, while at the same time, substantially inhibiting
differentiation of said hematopoietic stem cells ex-vivo.
2. A method of hematopoietic cells transplantation or implantation
comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a donor;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing an
expression
and/or activity of CD38, thereby expanding a population of said hematopoietic
stem
cells, while at the same time, substantially inhibiting differentiation of
said
hematopoietic stem cells ex-vivo; and
(c) transplanting or implanting said hematopoietic stem cells to a recipient.
3. The method of claim 2, wherein said donor and said recipient are a
single individual.
4. A method of genetically modifying hematopoietic stem cells with an
exogene comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells;
98
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing an
expression
and/or activity of CD38, thereby expanding a population of said hematopoietic
stem
cells, while at the same time, substantially inhibiting differentiation of
said
hematopoietic stem cells ex-vivo; and
(c) genetically modifying said hematopoietic stem cells with the exogene.
5. The method of claim 4, wherein genetically modifying is effected by a
vector which comprises the exogene.
6. The method of claim 5, wherein the vector is a viral vector or a nucleic
acid vector.
7. A method of adoptive immunotherapy comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a recipient;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing an
expression
and/or activity of CD38, thereby expanding a population of said hematopoietic
stem
cells, while at the same time, substantially inhibiting differentiation of
said
hematopoietic stem cells; and
(c) transplanting said hematopoietic stem cells to the recipient.
8. A transplantable hematopoietic cell preparation comprising an
expanded population of hematopoietic stem cells propagated ex-vivo from
hematopoietic mononuclear cells which comprise, prior to expansion, a major
fraction
of hematopoietic committed cells and a minor fraction of hematopoietic stem
and
progenitor cells, in the presence of an effective amount of an agent, said
agent
reducing an expression and/or activity of CD38, while at the same time,
substantially
inhibiting differentiation of said hematopoietic stem cells, and a
pharmaceutically
acceptable carrier.
99
9. The method of any of claims 1, 2, 4, and 7, wherein said hematopoietic
mononuclear cells are derived from a source selected from the group consisting
of
bone marrow, peripheral blood and neonatal umbilical cord blood.
10. The method of any of claims 1, 2, 4 and 7, wherein providing said
hematopoietic mononuclear cells with said conditions for ex-vivo cell
proliferation
comprises providing said hematopoietic mononuclear cells with nutrients and
with
cytokines.
11. The method of claim 10, wherein said cytokines are early acting
cytokines.
12. The method of claim 11, wherein said early acting cytokines are
selected from the group consisting of stem cell factor, FLT3 ligand,
interleukin-1,
interleukin-2, interleukin-3, interleukin-6, interleukin-10, interleukin-12,
tumor
necrosis factor-a and thrombopoietin.
13. The method of claim 10, wherein said cytokines are late acting
cytokines.
14. The method of claim 13, wherein said late acting cytokines are selected
from the group consisting of granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
15. The method of any of claims 1, 2, 4 and 7, wherein providing said
hematopoietic mononuclear cells with ex-vivo culture conditions for reducing
said
expression and/or said activity of CD38 is by providing said hematopoietic
mononuclear cells with an agent that downregulates CD38 expression.
16. The transplantable hematopoietic cell preparation of claim 8, wherein
said agent is an agent that downregulates CD38 expression.
100
17. The method of any of claims 15 and 16, wherein said agent that
downregulates CD38 expression is selected from the group consisting of a
retinoic
acid receptor antagonist, a retinoid X receptor antagonist and a Vitamin D
receptor
antagonist.
18. The method of any of claims 15 and 16, wherein said agent that
downregulates CD38 expression is an antagonist for reducing a capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoid
and/or
Vitamin D.
19. The method of any of claims 15 and 16, wherein said agent that
downregulates CD38 expression is a polynucleotide.
20. The method of claim 19, wherein said polynucleotide encodes an anti
CD38, an anti retinoic acid receptor, an anti retinoid X receptor or an anti
Vitamin D
receptor intracellular antibody.
21. The method of claim 19, wherein said polynucleotide encodes an anti
CD38, an anti retinoic acid receptor, an anti retinoid X receptor or an anti
Vitamin D
receptor antibody.
22. The method of claim 19, wherein said polynucleotide is a small
interfering polynucleotide molecule directed to cause intracellular CD38,
retinoic
acid receptor, retinoid X receptor or Vitamin D receptor mRNA degradation.
23. The method of claim 22, wherein said small interfering
polynucleotide molecule is selected from the group consisting of an RNAi
molecule,
an anti-sense molecule, a rybozyme molecule and a DNAzyme molecule.
24. The method of any of claims 15 and 16, wherein said agent that
downregulates CD38 expression is an agent that downregulates PI 3-kinase
expression.
101
25. The method of claim 24, wherein said agent that downregulates PI 3-
kinase expression is a polynucleotide.
26. The method of claim 24, wherein agent that downregulates PI 3-
kinase expression is an intracellular antibody.
27. The method of claim 25, wherein said polynucleotide is a small
interfering polynucleotide molecule directed to cause intracellular PI 3-
kinase
mRNA or gene degradation.
28. The method of claim 27, wherein said small interfering
polynucleotide molecule is selected from the group consisting of an RNAi
molecule,
an anti-sense molecule, a rybozyme molecule and a DNAzyme molecule.
29. The method of any of claims 15 and 16, wherein said agent that
downregulates CD38 expression is an agent that inhibits PI 3-kinase activity.
30. The method of claim 30, wherein said agent that inhibits PI 3-kinase
activity is selected from the group consisting of wortmannin and LY294002
31. The method of any of claims 1, 2, 4 and 7, wherein providing said
hematopoietic mononuclear cells with ex-vivo culture conditions for reducing
said
expression and/or said activity of CD38 is by providing said hematopoietic
mononuclear cells with an agent that inhibits CD38 activity.
32. The transplantable hematopoietic cell preparation of claim 8, wherein
said agent is an agent that inhibits CD38 activity.
33. The method of any of claims 31 and 32, wherein said agent that
inhibits CD38 activity is nicotinamide, a nicotinamide analog, a nicotinamide
or a
nicotinamide analog derivative or a nicotinamide or a nicotinamide analog
metabolite.
102
34. The method of claim 33, wherein said nicotinamide analog is selected
from the group consisting of benzamide, nicotinethioamide, nicotinic acid and
.alpha.-
amino-3-indolepropionic acid.
35. The method of any of claims 1, 2, 4 and 7, wherein providing said
hematopoietic mononuclear cells with ex-vivo culture conditions for reducing
said
expression and/or said activity of CD38 is by providing said hematopoietic
mononuclear cells with an agent that inhibits PI 3-kinase activity.
36. The transplantable hematopoietic cell preparation of claims 8,
wherein said agent is an agent that inhibits PI 3-kinase activity.
37. The method of any of claims 35 and 36, wherein said agent that
inhibits PI 3-kinase activity is selected from the group consisting of
wortmannin and
LY294002.
38. A method of ex-vivo expanding a population of hematopoietic stem
cells, while at the same time, substantially inhibiting differentiation of the
hematopoietic stem cells ex-vivo, the method comprising providing
hematopoietic
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells, with ex-vivo
culture
conditions for ex-vivo cell proliferation and, at the same time, for reducing
a capacity
of said hematopoietic mononuclear cells in responding to retinoic acid,
retinoids
and/or Vitamin D, thereby expanding a population of said hematopoietic stem
cells
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells ex-vivo.
39. A method of hematopoietic cells transplantation or implantation
comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a donor;
103
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing a
capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoids
and/or
Vitamin D, thereby expanding a population of said hematopoietic stem cells,
while at
the same time, substantially inhibiting differentiation of said hematopoietic
stem cells
ex-vivo; and
(c) transplanting or implanting said hematopoietic stem cells to a recipient.
40. The method of claim 39, wherein said donor and said recipient are a
single individual.
41. A method of genetically modifying hematopoietic stem cells with an
exogene comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing a
capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoids
and/or
Vitamin D, thereby expanding a population of said hematopoietic stem cells,
while at
the same time, substantially inhibiting differentiation of said hematopoietic
stem cells
ex-vivo; and
(c) genetically modifying said hematopoietic stem cells with the exogene.
42. The method of claim 41, wherein genetically modifying is effected by
a vector which comprises the exogene.
43. The method of claim 42, wherein the vector is a viral vector or a
nucleic acid vector.
44. A method of adoptive immunotherapy comprising:
104
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a recipient;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing a
capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoids
and/or
Vitamin D, thereby expanding a population of said hematopoietic stem cells,
while at
the same time, substantially inhibiting differentiation of said hematopoietic
stem cells;
and
(c) transplanting said hematopoietic stem cells to the recipient.
45. A transplantable hematopoietic cell preparation comprising an
expanded population of hematopoietic stem cells propagated ex-vivo from
hematopoietic mononuclear cells which comprise, prior to expansion, a major
fraction
of hematopoietic committed cells and a minor fraction of hematopoietic stem
and
progenitor cells in the presence of an effective amount of an agent, said
agent
reducing a capacity of said hematopoietic mononuclear cells in responding to
retinoic
acid, retinoids and/or Vitamin D, while at the same time, substantially
inhibiting
differentiation of said hematopoietic stem cells, and a pharmaceutically
acceptable
carrier.
46. The method and/or the transplantable hematopoietic cell preparation of
any of claims 38, 39, 41, 44 and 45, wherein said hematopoietic mononuclear
cells are
derived from a source selected from the group consisting of bone marrow,
peripheral
blood and neonatal umbilical cord blood.
47. The method of any of claims 38, 39, and 44, wherein providing said
hematopoietic mononuclear cells with said conditions for ex-vivo cell
proliferation
comprises providing said hematopoietic mononuclear cells with nutrients and
with
cytokines.
48. The method of claim 47, wherein said cytokines are early acting
cytokines.
105
49. The method of claim 48, wherein said early acting cytokines are
selected from the group consisting of stem cell factor, FLT3 ligand,
interleukin-l,
interleukin-2, interleukin-3, interleukin-6, interleukin-10, interleukin-12,
tumor
necrosis factor-a and thrombopoietin.
50. The method of claim 47, wherein said cytokines are late acting
cytokines.
51. The method of claim 50, wherein said late acting cytokines are
selected from the group consisting of granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
52. The method and/or the transplantable hematopoietic cell preparation of
any of claims 38, 39, 41, 44 and 45, wherein reducing said capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoids
and/or
Vitamin D is reversible.
53. The method and/or the transplantable hematopoietic cell preparation of
any of claims 38, 39, 41, 44 and 45, wherein reducing said capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoids
and/or
Vitamin D is by ex-vivo culturing said hematopoietic mononuclear cells in a
presence
of an effective amount of at least one retinoic acid receptor antagonist, at
least one
retinoid X receptor antagonist and/or at least one Vitamin D receptor
antagonist.
54. The method of claim 53, wherein reducing said capacity of said
hematopoietic mononuclear cells in responding to retinoic acid, retinoids
and/or
Vitamin D is by ex-vivo culturing said hematopoietic mononuclear cells in a
presence
of an effective amount of at least one retinoic acid receptor antagonist, at
least one
retinoid X receptor antagonist and/or at least one Vitamin D receptor
antagonist, for a
time period of 0.1-50 % of an entire ex-vivo culturing period of said
hematopoietic
mononuclear cells.
106
55. The transplantable hematopoietic cell preparation of claim 45, wherein
said agent is selected from the group consisting of retinoic acid receptor
antagonist,
retinoid X receptor antagonist and/or Vitamin D receptor antagonist.
56. The method and/or the transplantable hematopoietic cell preparation of
any of claims 53, 54 and 55, wherein said retinoic acid receptor antagonist is
selected
from the group consisting of:
AGN 194310; AGN 193109; 3-(4-Methoxy-phenylsulfanyl)-3-methyl-butyric acid;
6-Methoxy-2,2-dimethvl-thiochroman-4-one,2,2-Dimethyl-4-oxo-thiochroman-6-
yltrifluoromethane-sulfonate; Ethyl 4-((2,2 dimethyl-4-oxo-thiochroman-6-
yl)ethynyl)-benzoate; Ethyl 4-((2,2-dimethy 1-4-triflouromethanensulfonyloxy -
(2H)-
thiochromen-6-yl)ethynyl)-benzoate(41); Thiochromen-6-yl]-ethynyl]-
benzoate(yl);
(p-[(E)-2-[3'4'-Dihydro-4,4'-dimethyl-7'-(heptyloxy)-2'H-1-benzothiopyran-
6'yl]
propenyl] benzoic acid 1'1'-dioxide; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
butoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
propoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
pentoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
hexoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
heptoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
octoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; (2E,4E,6E)-7-[3-t-butyl-5-(1-phenyl-vinyl)-
phenyl]-
3-methyl-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-{[4,5-3
H2]-n-pentoxy}phenyl)-3-methyl]-octa-2,4,6-trienoic acid; (2E,4E)-
(1RS,2RS)-
5-[2-(3,5-di-tert.butyl-2-ethoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-
dienoic acid
ethyl ester; (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert.butyl-2-ethoxy-phenyl)-
cyclopropyl]-
3-methyl-penta-2,4-dienoic acid; (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert.butyl-2-
butoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-dienoic(2E,4E,6Z)-7-[3,5-di-
acid;
tert.butyl-2-ethoxyphenyl]3-methyl-2,4,6-octatrienoic(2E,4E,6Z)-7-[3,5-di-
acid;
tert.butyl-2-butyloxyphenyl]-3-methyl-2,4,6-octatrienoic4-(5,6,7,8-tetrahydro-
acid;
5,5,8,8-tetramethyl-2-naphthalene-carboxamido)(2E,4E)-3-methyl-5-
benzoic acid;
[(1S,2S)-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-
cyclopropyl]-
penta-2,4-dienoic acid; p-[(E)-2-[3',4'-Dihydro-4',4'-dimethyl-7'-(heptyloxy)-
2'H-1-
benzothiopyran-6'-yl]propenyl]benzoic acid; 1',1'-dioxide, 4-(7,7,10,10-
Tetramethyl-
1-pyridin-3-ylmethyl-4,5,7,8,9,10-hexahydro-1H-naphto[2,3-g]indol-3-yl)-
benzoic
107
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-methoxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-ethoxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-hexyloxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-octyloxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; and (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert-butyl-2-butoxy-phenyl)-
cyclopropyl]-
3-methyl-penta-2,4-dienoic acid, (2E,4E,6Z)-7-(3-n-propoxy-5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-2,4,6-trienoic acid, 4-(5H-
2,3(2,5
dimethyl-2,5-hexano)-5-n-propyldibenzo[b,e][1,4]diazepin-11-yl)benzoic acid, 4-
(SH-2,3-(2,5-dimethyl-2,5-hexano)-5methyl-8-nitrodibenzo[b,e][1,4]diazepin-11-
yl)benzoic acid, 4-{[4-(4-Ethylphenyl)2,2-dimethyl-(2H)-thiochromen-6-
yl]ethynyl}benzoic acid, 4-[4-2methyl-1,2-dicarba-closo-dodecaboran-1-yl-
phenylcarbamoyl]benzoic acid, 4-[4,5,7,8,9,10-hexahydro-7,7,10,10-tetramethyl-
1-(3-
pyridylmethyl)-anthra[1,2-b]pyrrol-3-yl]benzoic acid, (3-pyridylmethyl)-]5-
thiaanthra[2,1-b]pyrrol-3-yl)benzoic acid, and (3-pyridylmethyl)-anthra[2m1-
d]pyrazol-3-yl]benzoic acid.
57. The method and/or the transplantable hematopoietic cell preparation of
any of claims 53, 54 and 55, wherein said retinoid X receptor antagonist is
selected
from the group consisting of:
LGN100572, LGN100574, 1-(3-hydroxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-2-yl)ethanone, 1-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-2-yl)ethanone, 3-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-2-yl)but-2-enenitrile, 3-(3-propoxy-5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene-2-yl)but-2-enal, (2E,4E,6E)-7-3[-propoxy-
5,6,7,8-
tetrahydro 5,5,8,8-tetramethyl-2-naphthalene-2-yl]-3-methylocta-2,4,6-trienoic
acid,
4-[3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl] benzoic acid,
4-[1-
(3,5, 5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl] benzoic acid, 4-
[1(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)cyclopropyl] benzoic
acid, 4-
[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl] benzenete
trazole,
2-[1-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthyl) ethenyl]pyridine-5-
carboxylic acid, 2-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)ethyl]pyridine-5-carboxylic acid, ethyl-2-[1-(3,5,5,8, 8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethenyl]pyridine-5-carboxylate, 5-[1-3,5,5,8,8-
pentamethyl-
108
5,6,7,8-tetrahydro-2-naphthyl)ethenyl]pyridine-2-carboxylic acid, 2-[1-
(3,5,5,8,8-
pentamethyl-5,6,7,8-tetrahydro-2-naphthyl) cyclopropyl]pyridine-5-carboxylic
acid,
methyl 2-[ 1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)cyclopropyl]pyridine-5-carboxylate, 4-[1-(3,5, 5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethenyl]-N-(4-hydroxyphenyl) benzamide, 2-[1-(3,5,5,8,8-
Pentamethyl-5,6,7,8-tetrahydro-2-naphthyl) ethenyl] pyridine-5-carboxylic
acid, 2-[1-
(3,5,5,8,8-Pentamethyl-5, 6,7,8-tetrahydro-2-naphthyl)cyclopropyl]pyridine-5-
carboxylic acid, 4-[(3,5, 5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)carbonyl]benzoic acid butyloxime, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-naphthyl) carbonyl]benzoic acid propyloxime, 4-[(3,5,5,8,8-
pentamethyl-5,6,7,8-terrahydro-2-naphthyl)carbonyl]benzoic acid cyanoimine, 4-
[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl]benzoic acid
allyloxime, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)carbonyl]benzoic
acid 4-(3-methylbut-2-enoic acid)oxime, and 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-naphthyl)carbonyl]benzoic acid 1-aminoethyloxime, (2E,4E,6Z)-7-(3-
n-
propoxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-
2,4,6-
trienoic acid, 4-(5H-2,3(2,5 dimethyl-2,5-hexano)-5-n-
propyldibenzo[b,e][1,4]diazepin-11-yl)benzoic acid, and 4-(5H-2,3-(2,5-
dimethyl-
2,5-hexano)-5methyl-8-nitrodibenzo[b,e][1,4]diazepin-11-yl)benzoic acid.
58. The method and/or the transplantable hematopoietic cell preparation of
any of claims 53, 54 and 55, wherein said Vitamin D receptor antagonist is
selected
from the group consisting of: 1 alpha, 25-(OH)-D3-26,23 lactone; 1alpha, 25-
dihydroxyvitamin D (3); the 25-carboxylic ester ZK159222; (23S)- 25-dehydro-1
alpha-OH-D (3); (23R)-25-dehydro-1 alpha-OH-D (3); 1 beta, 25 (OH)Z D3; 1
beta,
25(OH)2-3-epi-D3; (23S) 25-dehydro-1 alpha(OH) D3-26,23-lactone; (23R) 25-
dehydro-1 alpha(OH)D3-26,23-lactone and Butyl-(5Z,7E,22E-(1S,7E,22E-
(1 S,3R,24R)-1,3,24-trihydroxy-26,27-cyclo-9,10-secocholesta-5,7,10(19),22-
tetraene-
25-carboxylate).
59. A method of ex-vivo expanding a population of hematopoietic stem
cells, while at the same time, substantially inhibiting differentiation of the
hematopoietic stem cells ex-vivo, the method comprising providing
hematopoietic
109
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells, with ex-vivo
culture
conditions for ex-vivo cell proliferation and, at the same time, for reducing
a capacity
of said hematopoietic mononuclear cells in responding to signaling pathways
involving the retinoic acid receptor, the retinoid X receptor and/or the
Vitamin D
receptor, thereby expanding a population of said hematopoietic stem cells
while at the
same time, substantially inhibiting differentiation of said hematopoietic stem
cells ex-
vivo.
60. A method of hematopoietic cells transplantation or implantation
comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a donor;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing a
capacity of said
hematopoietic mononuclear cells in responding to signaling pathways involving
the
retinoic acid receptor, the retinoid X receptor and/or the Vitamin D receptor,
thereby
expanding a population of said hematopoietic stem cells, while at the same
time,
substantially inhibiting differentiation of said hematopoietic stem cells ex-
vivo; and
(c) transplanting or implanting said hematopoietic stem cells to a recipient.
61. The method of claim 60, wherein said donor and said recipient are a
single individual.
62. A method of genetically modifying hematopoietic stem cells with an
exogene comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing a
capacity of said
hematopoietic mononuclear cells in responding to signaling pathways involving
the
110
retinoic acid receptor, the retinoid X receptor and/or the Vitamin D receptor,
thereby
expanding a population of said hematopoietic stem cells, while at the same
time,
substantially inhibiting differentiation of said hematopoietic stem cells ex-
vivo; and
(c) genetically modifying said hematopoietic stem cells with the exogene.
63. The method of claim 62, wherein genetically modifying is effected by
a vector which comprises the exogene.
64. The method of claim 63, wherein the vector is a viral vector or a
nucleic acid vector.
65. A method of adoptive immunotherapy comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a recipient;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing a
capacity of said
hematopoietic mononuclear cells in responding to signaling pathways involving
the
retinoic acid receptor, the retinoid X receptor and/or the Vitamin D receptor,
thereby
expanding a population of said hematopoietic stem cells, while at the same
time,
substantially inhibiting differentiation of said hematopoietic stem cells; and
(c) transplanting said hematopoietic stem cells to the recipient.
66. A transplantable hematopoietic cell preparation comprising an
expanded population of hematopoietic stem cells propagated ex-vivo from
hematopoietic mononuclear cells which comprise, prior to expansion, a major
fraction
of hematopoietic committed cells and a minor fraction of hematopoietic stem
and
progenitor cells in the presence of an effective amount of an agent, said
agent
reducing a capacity of said hematopoietic mononuclear cells in responding to
signaling pathways involving the retinoic acid receptor, the retinoid X
receptor and/or
the Vitamin D receptor, while at the same time, substantially inhibiting
differentiation
of said hematopoietic stem cells, and a pharmaceutically acceptable carrier.
111
67. The method and/or the transplantable hematopoietic cell preparation of
any of claims 59, 60, 62, 65 and 66, wherein said hematopoietic mononuclear
cells are
derived from a source selected from the group consisting of bone marrow,
peripheral
blood and neonatal umbilical cord blood.
68. The method of any of claims 59, 60, and 65, wherein providing said
hematopoietic mononuclear cells with said conditions for ex-vivo cell
proliferation
comprises providing said hematopoietic mononuclear cells with nutrients and
with
cytokines.
69. The method of claim 68, wherein said cytokines are early acting
cytokines.
70. The method of claim 69, wherein said early acting cytokines are
selected from the group consisting of stem cell factor, FLT3 ligand,
interleukin-1,
interleukin-2, interleukin-3, interleukin-6, interleukin-10, interleukin-12,
tumor
necrosis factor-a and thrombopoietin.
71. The method of claim 68, wherein said cytokines are late acting
cytokines.
72. The method of claim 71, wherein said late acting cytokines are
selected from the group consisting of granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
73. The method and/or the transplantable hematopoietic cell preparation of
any of claims 59, 60, 62, 65 and 66, wherein reducing said capacity of said
hematopoietic mononuclear cells in responding to signaling pathways involving
the
retinoic acid receptor, the retinoid X receptor and/or the Vitamin D receptor
is
reversible.
112
74. The method and/or the transplantable hematopoietic cell preparation of
any of claims 59, 60, 62, 65 and 66, wherein reducing said capacity of said
hematopoietic mononuclear cells in responding to signaling pathways involving
the
retinoic acid receptor, the retinoid X receptor and/or the Vitamin D receptor
is by ex-
vivo culturing . said hematopoietic mononuclear cells fraction in a presence
of an
effective amount of at least one retinoic acid receptor antagonist, at least
one retinoid
X receptor antagonist and/or at least one Vitamin D receptor antagonist.
75. The method of claim 74, wherein reducing said capacity of said
hematopoietic mononuclear cells in responding to signaling pathways involving
the
retinoic acid receptor, the retinoid X receptor and/or the Vitamin D receptor
is by ex-
vivo culturing said hematopoietic mononuclear cells in a presence of an
effective
amount of at least one retinoic acid receptor antagonist, at least one
retinoid X
receptor antagonist and/or at least one Vitamin D receptor antagonist, for a
time
period of 0.1-50 % of an entire ex-vivo culturing period of said hematopoietic
mononuclear cells.
76. The transplantable hematopoietic cell preparation of claim 66, wherein
said agent is selected from the group consisting of retinoic acid receptor
antagonist,
retinoid X receptor antagonist and/or Vitamin D receptor antagonist.
77. The method and/or the transplantable hematopoietic cell preparation of
any of claims 74, 75 and 76, wherein said retinoic acid receptor antagonist is
selected
from the group consisting of:
AGN 194310; AGN 193109; 3-(4-Methoxy-phenylsulfanyl)-3-methyl-butyric acid;
6-Methoxy-2,2-dimethvl-thiochroman-4-one,2,2-Dimethyl-4-oxo-thiochroman-6-
yltrifluoromethane-sulfonate; Ethyl 4-((2,2 dimethyl-4-oxo-thiochroman-6-
yl)ethynyl)-benzoate; Ethyl 4-((2,2-dimethy 1-4-triflouromethanensulfonyloxy -
(2H)-
thiochromen-6-yl)ethynyl)-benzoate(41); Thiochromen-6-yl]-ethynyl]-
benzoate(yl);
(p-[(E)-2-[3'4'-Dihydro-4,4'-dimethyl-7'-(heptyloxy)-2'H-1-benzothiopyran-
6'yl]
propenyl] benzoic acid 1'1'-dioxide; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
butoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
propoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
pentoxyphenyl)-
113
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
hexoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
heptoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
octoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; (2E,4E,6E)-7-[3-t-butyl-5-(1-phenyl-vinyl)-
phenyl]-
3-methyl-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-{[4,5-3
H2]-n-pentoxy}phenyl)-3-methyl]-octa-2,4,6-trienoic acid; (2E,4E)-
(1RS,2RS)-
5-[2-(3,5-di-tert.butyl-2-ethoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-
dienoic acid
ethyl ester; (2E,4E)-(1RS,2RS)-S-[2-(3,5-di-tert.butyl-2-ethoxy-phenyl)-
cyclopropyl]-
3-methyl-penta-2,4-dienoic acid; (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert.butyl-2-
butoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-dienoic acid; (2E,4E,6Z)-7-[3,5-
di-
tert.butyl-2-ethoxyphenyl]3-methyl-2,4,6-octatrienoic acid; (2E,4E,6Z)-7-[3,5-
di-
tert.butyl-2-butyloxyphenyl]-3-methyl-2,4,6-octatrienoic acid; 4-(5,6,7,8-
tetrahydro-
5,5,8,8-tetramethyl-2-naphthalene-carboxamido) benzoic acid; (2E,4E)-3-methyl-
5-
[( 1 S,2S)-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-
cyclopropyl]-
penta-2,4-dienoic acid; p-[(E)-2-[3',4'-Dihydro-4',4'-dimethyl-7'-(heptyloxy)-
2'H-1-
benzothiopyran-6'-yl]propenyl]benzoic acid; 1',1'-dioxide, 4-(7,7,10,10-
Tetramethyl-
1-pyridin-3-ylmethyl-4,5,7,8,9,10-hexahydro-1 H-naphto[2,3-g]indol-3-yl)-
benzoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-methoxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-ethoxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-hexyloxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-octyloxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; and (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert-butyl-2-butoxy-phenyl)-
cyclopropyl]-
3-methyl-penta-2,4-dienoic acid, (2E,4E,6Z)-7-(3-n-propoxy-5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-2,4,6-trienoic acid, 4-(5H-
2,3(2,5
dimethyl-2,5-hexano)-5-n-propyldibenzo[b,e][1,4]diazepin-11-yl)benzoic acid, 4-
(5H-2,3-(2,5-dimethyl-2,5-hexano)-5methyl-8-nitrodibenzo[b,e][ 1,4]diazepin-11-
yl)benzoic acid, 4-{[4-(4-Ethylphenyl)2,2-dimethyl-(2H)-thiochromen-6-
yl]ethynyl}benzoic acid, 4-[4-2methyl-1,2-dicarba-closo-dodecaboran-1-yl-
phenylcarbamoyl]benzoic acid, 4-[4,5,7,8,9,10-hexahydro-7,7,10,10-tetramethyl-
1-(3-
pyridylmethyl)-anthra[1,2-b]pyrrol-3-yl]benzoic acid, (3-pyridylmethyl)-]5-
thiaanthra[2,1-b]pyrrol-3-yl)benzoic acid, and (3-pyridylmethyl)-anthra[2m1-
d]pyrazol-3-yl]benzoic acid.
114
78. The method and/or the transplantable hematopoietic cell preparation of
any of claims 74, 75 and 76, wherein said retinoid X receptor antagonist is
selected
from the group consisting o~
LGN100572, LGN100574, 1-(3-hydroxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-2-yl)ethanone, 1-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-2-yl)ethanone, 3-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-2-yl)but-2-enenitrile, 3-(3-propoxy-5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene-2-yl)but-2-enal, (2E,4E,6E)-7-3[-propoxy-
5,6,7,8-
tetrahydro 5,5,8,8-tetramethyl-2-naphthalene-2-yl]-3-methylocta-2,4,6-trienoic
acid,
4-[3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl] benzoic acid,
4-[1-
(3,5, 5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl] benzoic acid, 4-
[1(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)cyclopropyl] benzoic
acid, 4-
[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl] benzenete
trazole,
2-(1-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthyl) ethenyl]pyridine-5-
carboxylic acid, 2-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)ethyl]pyridine-5-carboxylic acid, ethyl-2-[1-(3,5,5,8, 8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethenyl]pyridine-5-carboxylate, 5-[1-3,5,5,8,8-
pentamethyl-
5,6,7,8-tetrahydro-2-naphthyl)ethenyl]pyridine-2-carboxylic acid, 2-[1-
(3,5,5,8,8-
pentamethyl-5,6,7,8-tetrahydro-2-naphthyl) cyclopropyl]pyridine-5-carboxylic
acid,
methyl 2-[ 1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)cyclopropyl]pyridine-5-carboxylate, 4-[1-(3,5, 5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethenyl]-N-(4-hydroxyphenyl) benzamide, 2-[1-(3,5,5,8,8-
Pentamethyl-5,6,7,8-tetrahydro-2-naphthyl) ethenyl] pyridine-5-carboxylic
acid, 2-[1-
(3,5,5,8,8-Pentamethyl-5, 6,7,8-tetrahydro-2-naphthyl)cyclopropyl]pyridine-S-
carboxylic acid, 4-[(3,5, 5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)carbonyl]benzoic acid butyloxime, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-naphthyl) carbonyl]benzoic acid propyloxime, 4-[(3,5,5,8,8-
pentamethyl-5,6,7,8-terrahydro-2-naphthyl)carbonyl]benzoic acid cyanoimine, 4-
[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl]benzoic acid
allyloxime, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)carbonyl]benzoic
acid 4-(3-methylbut-2-enoic acid)oxime, and 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-naphthyl)carbonyl]benzoic acid 1-aminoethyloxime, (2E,4E,6Z)-7-(3-
n-
propoxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-
2,4,6-
115
trienoic acid, 4-(5H-2,3(2,5 dimethyl-2,5-hexano)-5-n-
propyldibenzo[b,e][1,4]diazepin-11-yl)benzoic acid, and 4-(5H-2,3-(2,5-
dimethyl-
2,5-hexano)-Smethyl-8-nitrodibenzo[b,e][1,4]diazepin-11-yl)benzoic acid.
79. The method and/or the transplantable hematopoietic cell preparation of
any of claims 74, 75 and 76, wherein said Vitamin D receptor antagonist is
selected
from the group consisting of: 1 alpha, 25-(OH)-D3-26,23 lactone; lalpha, 25-
dihydroxyvitamin D (3); the 25-carboxylic ester ZK159222; (23S)- 25-dehydro-1
alpha-OH-D (3); (23R)-25-dehydro-1 alpha-OH-D (3); 1 beta, 25 (OH)2 D3; 1
beta,
25(OH)2-3-epi-D3; (23S) 25-dehydro-1 alpha(OH) D3-26,23-lactone; (23R) 25-
dehydro-1 alpha(OH)D3-26,23-lactone and Butyl-(5Z,7E,22E-(1S,7E,22E-
(1S,3R,24R)-1,3,24-trihydroxy-26,27-cyclo-9,10-secocholesta-5,7,10(19),22-
tetraene-
25-carboxylate).
80. A method of ex-vivo expanding a population of hematopoietic stem
cells, while at the same time, substantially inhibiting differentiation of the
hematopoietic stem cells ex-vivo, the method comprising providing
hematopoietic
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells, with ex-vivo
culture
conditions for ex-vivo cell proliferation and with nicotinamide, a
nicotinamide analog,
a nicotinamide or a nicotinamide analog derivative or a nicotinamide or a
nicotinamide analog metabolite thereby expanding a population of said
hematopoietic
stem cells while at the same time, substantially inhibiting differentiation of
said
hematopoietic stem cells ex-vivo.
81. A method of hematopoietic cells transplantation or implantation
comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a donor;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and with nicotinamide, a nicotinamide
analog, a
nicotinamide or a nicotinamide analog derivative or a nicotinamide or a
nicotinamide
116
analog metabolite, thereby expanding a population of said hematopoietic stem
cells,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells ex-vivo; and
(c) transplanting or implanting said hematopoietic stem cells to a recipient.
82. The method of claim 81, wherein said donor and said recipient are a
single individual.
83. A method of genetically modifying hematopoietic stem cells with an
exogene comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and with nicotinamide, a nicotinamide
analog, a
nicotinamide or a nicotinamide analog derivative or a nicotinamide or a
nicotinamide
analog metabolite, thereby expanding a population of said hematopoietic stem
cells,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells ex-vivo; and
(c) genetically modifying said hematopoietic stem cells with the exogene.
84. The method of claim 83, wherein genetically modifying is effected by
a vector which comprises the exogene.
85. The method of claim 84, wherein the vector is a viral vector or a
nucleic acid vector.
86. A method of adoptive immunotherapy comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a recipient;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and with nicotinamide, a nicotinamide
analog, a
117
nicotinamide or a nicotinamide analog derivative or a nicotinamide or a
nicotinamide
analog metabolite, thereby expanding a population of said hematopoietic stem
cells,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells; and
(c) transplanting said hematopoietic stem cells to the recipient.
87. A transplantable hematopoietic cell preparation comprising an
expanded population of hematopoietic stem cells propagated ex-vivo from
hematopoietic mononuclear cells which comprise, prior to expansion, a major
fraction
of hematopoietic committed cells and a minor fraction of hematopoietic stem
and
progenitor cells in the presence of an effective amount of an agent selected
from the
group consisting of nicotinamide, a nicotinamide analog, a nicotinamide or a
nicotinamide analog derivative or a nicotinamide or a nicotinamide analog
metabolite,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells, and a pharmaceutically acceptable carrier.
88. The method and/or the transplantable hematopoietic cell preparation of
any of claims 80, 81, 83, 86 and 87, wherein said hematopoietic mononuclear
cells are
derived from a source selected from the group consisting of bone marrow,
peripheral
blood and neonatal umbilical cord blood.
89. The method of any of claims 80, 81, and 86, wherein providing said
hematopoietic mononuclear cells with said conditions for ex-vivo cell
proliferation
comprises providing said hematopoietic mononuclear cells with nutrients and
with
cytokines.
90. The method of claim 89, wherein said cytokines are early acting
cytokines.
91. The method of claim 90, wherein said early acting cytokines are
selected from the group consisting of stem cell factor, FLT3 ligand,
interleukin-l,
interleukin-2, interleukin-3, interleukin-6, interleukin-10, interleukin-12,
tumor
necrosis factor-a and thrombopoietin.
118
92. The method of claim 89, wherein said cytokines are late acting
cytokines.
93. The method of claim 92, wherein said late acting cytokines are
selected from the group consisting of granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
94. The method and/or the transplantable hematopoietic cell preparation of
any of claims 80, 81, 83, 86 and 87, wherein said nicotinamide analog is
selected
from the group consisting of benzamide, nicotinethioamide, nicotinic acid and
oc-
amino-3-indolepropionic acid.
95. A method of ex-vivo expanding a population of hematopoietic stem
cells, while at the same time, substantially inhibiting differentiation of the
hematopoietic stem cells ex-vivo, the method comprising providing
hematopoietic
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells, with ex-vivo
culture
conditions for ex-vivo cell proliferation and, at the same time, for reducing
an
expression and/or activity of PI 3-kinase, thereby expanding a population of
said
hematopoietic stem cells while at the same time, substantially inhibiting
differentiation of said hematopoietic stem cells ex-vivo.
96. A method of hematopoietic cells transplantation or implantation
comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a donor;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing an
expression
and/or activity of PI 3-kinase, thereby expanding a population of said
hematopoietic
stem cells, while at the same time, substantially inhibiting differentiation
of said
hematopoietic stem cells ex-vivo; and
119
(c) transplanting or implanting said hematopoietic stem cells to a recipient.
97. The method of claim 96, wherein said donor and said recipient are a
single individual.
98. A method of genetically modifying hematopoietic stem cells with an
exogene comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing an
expression
and/or activity of PI 3-kinase, thereby expanding a population of said
hematopoietic
stem cells, while at the same time, substantially inhibiting differentiation
of said
hematopoietic stem cells ex-vivo; and
(c) genetically modifying said hematopoietic stem cells with the exogene.
99. The method of claim 98, wherein genetically modifying is effected by
a vector which comprises the exogene.
100. The method of claim 99, wherein the vector is a viral vector or a
nucleic acid vector.
101. A method of adoptive immunotherapy comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a recipient;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, for reducing an
expression
and/or activity of PI 3-kinase, thereby expanding a population of said
hematopoietic
stem cells, while at the same time, substantially inhibiting differentiation
of said
hematopoietic stem cells; and
(c) transplanting said hematopoietic stem cells to the recipient.
120
102. A transplantable hematopoietic cell preparation comprising an
expanded population of hematopoietic stem cells propagated ex-vivo from
hematopoietic mononuclear cells which comprise, prior to expansion, a major
fraction
of hematopoietic committed cells and a minor fraction of hematopoietic stem
and
progenitor cells in the presence of an effective amount of an agent, said
agent
reducing an expression and/or activity of PI 3-kinase, while at the same time,
substantially inhibiting differentiation of said hematopoietic stem cells, and
a
pharmaceutically acceptable carrier.
103. The method of any of claims 95, 96, 98, and 101, wherein said
hematopoietic mononuclear cells are derived from a source selected from the
group
consisting of bone marrow, peripheral blood and neonatal umbilical cord blood.
104. The method of any of claims 95, 96, 98 and 101, wherein providing
said hematopoietic mononuclear cells with said conditions for ex-vivo cell
proliferation comprises providing said hematopoietic mononuclear cells with
nutrients
and with cytokines.
105. The method of claim 104, wherein said cytokines are early acting
cytokines.
106. The method of claim 105, wherein said early acting cytokines are
selected from the group consisting of stem cell factor, FLT3 ligand,
interleukin-1,
interleukin-2, interleukin-3, interleukin-6, interleukin-10, interleukin-12,
tumor
necrosis factor-a and thrombopoietin.
107. The method of claim 104, wherein said cytokines are late acting
cytokines.
108. The method of claim 107, wherein said late acting cytokines are
selected from the group consisting of granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
121
109. The method of any of claims 95, 96, 98 and 101, wherein providing
said hematopoietic mononuclear cells with ex-vivo culture conditions for
reducing
said expression and/or said activity of PI 3-kinase is by providing said
hematopoietic
mononuclear cells with an agent that downregulates PI 3-kinase expression.
110. The transplantable hematopoietic cell preparation of claim 102,
wherein said agent is an agent that downregulates PI 3-kinase expression.
111. The method and/or transplantable hematopoietic cell preparation of
any claim 109 and 110, wherein said agent that downregulates PI 3-kinase
expression is a polynucleotide.
112. The method and/or transplantable hematopoietic cell preparation of
any claim 109 and 110, wherein said agent that downregulates PI 3-kinase
expression is an intracellular antibody.
113. The method of claim 112, wherein said polynucleotide is a small
interfering polynucleotide molecule directed to cause intracellular PI 3-
kinase
mRNA or gene degradation.
114. The method of claim 113, wherein said small interfering
polynucleotide molecule is selected from the group consisting of an RNAi
molecule,
an anti-sense molecule, a rybozyme molecule and a DNAzyme molecule.
115. The method and/or transplantable hematopoietic cell preparation of
any claim 109 and 110, wherein providing said hematopoietic mononuclear cells
with ex-vivo culture conditions for reducing said expression and/or said
activity of PI
3-kinase is by providing said hematopoietic mononuclear cells with an agent
that
inhibits PI 3-kinase activity.
116. The transplantable hematopoietic cell preparation of claim 102,
wherein said agent is an agent that inhibits PI 3-kinase activity.
122
117. The method and/or the transplantable hematopoietic cell preparation
of any of claims 115 and 116, wherein said agent that inhibits PI 3-kinase
activity is
selected from the group consisting of wortmannin and LY294002
118. A method of ex-vivo expanding a population of hematopoietic stem
cells, while at the same time, substantially inhibiting differentiation of the
hematopoietic stem cells ex-vivo, the method comprising providing
hematopoietic
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells, with ex-vivo
culture
conditions for ex-vivo cell proliferation and, at the same time, with at least
one copper
chelator or chelate, thereby expanding a population of said hematopoietic stem
cells
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells ex-vivo.
119. A method of hematopoietic cells transplantation or implantation
comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a donor;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, with at least one
copper
chelator or chelate, thereby expanding a population of said hematopoietic stem
cells,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells ex-vivo; and
(c) transplanting or implanting said hematopoietic stem cells to a recipient.
120. The method of claim 119, wherein said donor and said recipient are a
single individual.
121. A method of genetically modifying hematopoietic stem cells with an
exogene comprising:
123
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, with at least one
copper
chelator or chelate, thereby expanding a population of said hematopoietic stem
cells,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells ex-vivo; and
(c) genetically modifying said hematopoietic stem cells with the exogene.
122. The method of claim 121, wherein genetically modifying is effected by
a vector which comprises the exogene.
123. The method of claim 122, wherein the vector is a viral vector or a
nucleic acid vector.
124. A method of adoptive immunotherapy comprising:
(a) obtaining hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells from a recipient;
(b) providing said hematopoietic mononuclear cells with ex-vivo culture
conditions for cell proliferation and, at the same time, with at least one
copper
chelator or chelate, thereby expanding a population of said hematopoietic stem
cells,
while at the same time, substantially inhibiting differentiation of said
hematopoietic
stem cells; and
(c) transplanting said hematopoietic stem cells to the recipient.
125. A transplantable hematopoietic cell preparation comprising an
expanded population of hematopoietic stem cells propagated ex-vivo from
hematopoietic mononuclear cells which comprise, prior to expansion, a major
fraction
of hematopoietic committed cells and a minor fraction of hematopoietic stem
and
progenitor cells in the presence of an effective amount of at least one copper
chelate
124
or chelator, while at the same time, substantially inhibiting differentiation
of said
hematopoietic stem cells, and a pharmaceutically acceptable carrier.
126. The method and/or the transplantable hematopoietic cell preparation of
any of claims 118, 119, 121, 124 and 125, wherein said hematopoietic
mononuclear
cells are derived from a source selected from the group consisting of bone
marrow,
peripheral blood and neonatal umbilical cord blood.
127. The method of any of claims 118, 119, and 124, wherein providing
said hematopoietic mononuclear cells with said conditions for ex-vivo cell
proliferation comprises providing said hematopoietic mononuclear cells with
nutrients
and with cytokines.
128. The method of claim 127, wherein said cytokines are early acting
cytokines.
129. The method of claim 128, wherein said early acting cytokines are
selected from the group consisting of stem cell factor, FLT3 ligand,
interleukin-1,
interleukin-2, interleukin-3, interleukin-6, interleukin-10, interleukin-12,
tumor
necrosis factor-a and thrombopoietin.
130. The method of claim 127, wherein said cytokines are late acting
cytokines.
131. The method of claim 130, wherein said late acting cytokines are
selected from the group consisting of granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
132. The method of any of claims 118, 119, 121 and 124, wherein providing
said hematopoietic mononuclear cells with at least one copper chelator or
chelate is
by providing said hematopoietic mononuclear cells at least one copper
chelator.
125
133. The method of claim 132, further comprising providing said
hematopoietic mononuclear cells fraction zinc.
134. The transplantable hematopoietic cell preparation of claim 125,
wherein said expanded population of said hematopoietic stem cells fraction is
propagated ex-vivo from hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells in the presence of an effective amount of at least one
copper
chelator.
135. The transplantable hematopoietic cell preparation of claim 134,
wherein said expanded population of said hematopoietic stem cells fraction is
propagated ex-vivo from hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells in the presence of an effective amount of zinc.
136. An assay of determining whether a transition metal chelate or chelator
causes substantial inhibition or induction of differentiation of hematopoietic
stem
cells, the assay comprising:
culturing hematopoietic mononuclear cells which comprise a major fraction of
hematopoietic committed cells and a minor fraction of hematopoietic stem and
progenitor cells, in the presence of the transition metal chelate or chelator
and
monitoring differentiation of said hematopoietic stem cells, wherein if
differentiation
is increased as is compared to non-treated hematopoietic mononuclear cells,
said
transition metal chelate induces differentiation, whereas if differentiation
is decreased
as is compared to non-treated hematopoietic mononuclear cells, or if
differentiation is
absent altogether, said transition metal chelate inhibits differentiation.
137. The method, the assay and/or the transplantable hematopoietic cell
preparation of any of claims 118, 119, 121, 124, 125, 136, and 138-141,
wherein said
at least one copper chelate or chelator comprises a polyamine chelator.
126
138. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 137, wherein said polyamine chelator is capable of
forming an
organometallic complex with a transition metal other than copper.
139. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 138, wherein said transition metal is selected from the
group
consisting of zinc, cobalt, nickel, iron, palladium, platinum, rhodium and
ruthenium.
140. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 137, wherein said polyamine chelator is a linear
polyamine.
141. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 140, wherein said linear polyamine has a general formula
I:
HX-Am-(Y1B1)1.....(YnBn)n-ZH
Formula I
wherein:
m is an integer from 1 to 10;
n is an integer from 0 to 20;
X and Z are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
Y1 and Yn are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
A is an alkylene chain having between 1 and 10 substituted and/or non-
substituted carbon atoms; and
B, and Bn are each independently an alkylene chain having between 1 and 20
substituted and/or non-substituted carbon atoms,
provided that at least one of said X, Z, Y1 and Yn is a -NH group and/or at
least one of said carbon atoms in said alkylene chains is substituted by an
amine
group.
127
142. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 141, wherein said A is an alkylene chain having a general
formula II:
Image
wherein:
g is an integer that equals 0 or 3-10;
each of R1, R2 and Rg is independently selected from the group consisting of
hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrile, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfuric acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
128
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
143. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 142, wherein each of B1 and Bn is independently an
alkylene
chain having a general formula III:
Image
wherein:
p is an integer that equals 0 or g+1;
q is an integer from g+2 to g+20; and
each of Rp, Rp+1 and Rq is independently selected from the group consisting
of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
129
thiophosphate, phosphite, pyrophosphite, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
144. The method, the assay, the pharmaceutical composition, the kit, the
expanded population and/or the assay of claim 141, wherein said linear
polyamine is
tetraethylenepentamine.
145. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 142, wherein at least one of said C1, C2 and Cg is a
chiral carbon
atom.
146. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 143, wherein at least one of said Cp, Cp+1 and Cq is a
chiral
carbon atom.
147. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 137, wherein said polyamine chelator is a cyclic
polyamine.
148. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 147, wherein said cyclic polyamine is cyclam.
149. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 147, wherein said cyclic polyamine has a general formula
IV:
Image
130
wherein:
m is an integer from 1 to 10;
n is an integer from 0 to 20;
X and Z are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
Y1 and Yn are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
A is an alkylene chain having between 1 and 10 substituted and/or non-
substituted carbon atoms;
B1 and Bn are each independently an alkylene chain having between 1 and 20
substituted and/or non-substituted carbon atoms; and
D is a bridging group having a general formula V:
U-W-V
Formula V
whereas:
U and V are each independently selected from the group consisting of
substituted hydrocarbon chain and non-substituted hydrocarbon chain; and
W is selected from the group consisting of amide, ether, ester, disulfide,
thioether, thioester, imine and alkene,
provided that at least one of said X, Z, Y1 and Yn is a -NH group and/or at
least one of said carbon atoms in said alkylene chains is substituted by an
amine
group.
150. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 149, wherein said A is an alkylene chain having a general
formula II:
Image
131
wherein:
g is an integer that equals 0 or 3-10; and
each of R1, R2 and Rg is independently selected from the group consisting of
hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrile, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
151. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 150, wherein each of B1 and Bn is independently an
alkylene
chain having a general formula III:
Image
wherein:
p is an integer that equals 0 or g+1;
q is an integer from g+2 to g+20; and
each of Rp, Rp+1 and Rq is independently selected from the group consisting
of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfuric acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
133
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
152. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 150, wherein at least one of said C1, C2 and Cg is a
chiral carbon
atom.
153. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 151, wherein at least one of said Cp, Cp+1 and Cq is a
chiral
carbon atom.
154. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 147, wherein said cyclic polyamine has a general formula
selected from the group consisting of:
Image
Image
wherein:
m is an integer from 1 to 10;
n is an integer from 0 to 20;
X and Z are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
Y1 and Yn are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
A is an alkylene chain having between 1 and 10 substituted and/or non-
substituted carbon atoms;
B1 and Bn are each independently an alkylene chain having between 1 and 20
substituted and/or non-substituted carbon atoms; and
D is a bridging group having a general formula V:
U-W-V
Formula V
whereas:
U and V are each independently selected from the group consisting of
substituted hydrocarbon chain and non-substituted hydrocarbon chain; and
W is selected from the group consisting of amide, ether, ester, disulfide,
thioether, thioester, imine and alkene,
and further wherein should said D is attached at one end to A (Formulas VI,
VII and X), said U or said V are being attached to one carbon atom in said
alkylene
chain and should said D is attached at one end to B1 or Bn (Formulas VIII, IX
and X),
said U or said V are being attached to one carbon atom in said alkylene chain,
provided that at least one of said X, Z, Y1 and Yn is a -NH group and/or at
least one of said carbon atoms in said alkylene chains is substituted by an
amine
group.
135
155. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 154, wherein said A is an alkylene chain having a general
formula II:
Image
wherein:
g is an integer that equals 0 or 3-10; and
each of R1, R2 and Rg is independently selected from the group consisting of
hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrile, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
136
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
156. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 155, wherein each of B1 and Bn is independently an
alkylene
chain having a general formula III:
Image
wherein:
p is an integer that equals 0 or g+1;
q is an integer from g+2 to g+20; and
each of Rp, Rp+1 and Rq is independently selected from the group consisting
of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfuric acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
137
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
157. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 155, wherein at least one of said C1, C2 and Cg is a
chiral carbon
atom.
158. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 156, wherein at least one of said Cp, Cp+1 and Cq is a
chiral
carbon atom.
159. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 137, wherein said polyamine chelator includes at least
one linear
polyamine and at least one cyclic polyamine.
160. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 159, wherein said polyamine chelator has a general
formula XI:
{E~f ~QlyGl~g~~ti ~~E2~~ ~Q2yCT2~j~}k_......._f~En~l'fQn_(Gn)ol~t
Formula XI
wherein:
n is an integer greater than 1;
each of f, g, h, i, j, k, l, o and t is independently an integer from 0 to 10;
each of E,, EZ and En is independently a linear polyamine;
each of G,, GZ and Gn is independently a cyclic polyamine; and
each of Q,, QZ and Qn is independently a linker linking between two of said
polyamines,
138
provided that at least one of said Q1, Q2 and Qn is an amine group and/or at
least one of said linear polyamine and said cyclic polyamine is having at
least one free
amine group.
161. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 160, wherein each of said Q1, Q2 and Qn is independently
selected from the group consisting alkylene, alkenylene, alkynylene, arylene,
cycloalkylene, hetroarylene, amine, azo, amide, sulfonyl, sulfinyl,
sulfonamide,
phosphonyl, phosphinyl, phosphonium, ketoester, carbonyl, thiocarbonyl, ester,
ether,
thioether, carbamate, thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl,
siloxy and silaza.
162. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 159, wherein each of said E1, E2 and En is independently
a linear
polyamine having a general formula I:
HX-Am-(Y1B1)1....(YnBn)n-ZH
Formula I
wherein:
m is an integer from 1 to 10;
n is an integer from 0 to 20;
X and Z are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
Y1 and Yn are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
A is an alkylene chain having between 1 and 10 substituted and/or non-
substituted carbon atoms; and
B1 and Bn are each independently an alkylene chain having between 1 and 20
substituted and/or non-substituted carbon atoms,
provided that at least one of said X, Z, Y1 and Yn is a -NH group and/or at
least one of said carbon atoms in said alkylene chains is substituted by an
amine
group.
139
163. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 162, wherein said A is an alkylene chain having a general
formula II:
Image
wherein:
g is an integer that equals 0 or 3-10;
each of R1, R2 and Rg is independently selected from the group consisting of
hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
140
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
164. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 163, wherein each of B1 and Bn is independently an
alkylene
chain having a general formula III:
Image
wherein:
p is an integer that equals 0 or g+1;
q is an integer from g+2 to g+20; and
each of Rp, Rp+1 and Rq is independently selected from the group consisting
of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrile, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
141
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphite, pyrophosphite, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
165. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 163, wherein at least one of said C1, C2 and Cg is a
chiral carbon
atom.
166. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 164, wherein at least one of said Cp, Cp+1 and Cq is a
chiral
carbon atom.
167. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 160, wherein each of said G1, G2 and Gn is independently
a
cyclic polyamine having a general formula IV:
Image
wherein:
m is an integer from 1 to 10;
n is an integer from 0 to 20;
X and Z are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
Y1 and Yn are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
142
A is an alkylene chain having between 1 and 10 substituted and/or non-
substituted carbon atoms;
B1 and Bn are each independently an alkylene chain having between 1 and 20
substituted and/or non-substituted carbon atoms; and
D is a bridging group having a general formula V:
U-W-V
Formula V
whereas:
U and V are each independently selected from the group consisting of
substituted hydrocarbon chain and non-substituted hydrocarbon chain; and
W is selected from the group consisting of amide, ether, ester, disulfide,
thioether, thioester, imine and alkene,
provided that at least one of said X, Z, Y1 and Yn is a -NH group and/or at
least one of said carbon atoms in said alkylene chains is substituted by an
amine
group.
168. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 167, wherein said A is an alkylene chain having a general
formula II:
Image
wherein:
g is an integer that equals 0 or 3-10; and
each of R1, R2 and Rg is independently selected from the group consisting of
hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
143
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
169. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 168, wherein each of B1 and Bn is independently an
alkylene
chain having a general formula III:
Image
wherein:
p is an integer that equals 0 or g+1;
q is an integer from g+2 to g+20; and
144
each of Rp, Rp+1 and Rq is independently selected from the group consisting
of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphoric acid, aryl
phosphoric acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
170. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 168, wherein at least one of said C1, C2 and Cg is a
chiral carbon
atom.
145
171. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 169, wherein at least one of said Cp, Cp+1 and Cq is a
chiral
carbon atom.
172. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 160, wherein said cyclic polyamine has a general formula
selected from the group consisting of:
Image
wherein:
m is an integer from 1 to 10;
n is an integer from 0 to 20;
X and Z are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
146
Y, and Yn are each independently selected from the group consisting of an
oxygen atom, a sulfur atom and a -NH group;
A is an alkylene chain having between 1 and 10 substituted and/or non-
substituted carbon atoms;
B1 and Bn are each independently an alkylene chain having between 1 and 20
substituted and/or non-substituted carbon atoms; and
D is a bridging group having a general formula V:
U-W-V
Formula V
whereas:
U and V are each independently selected from the group consisting of
substituted hydrocarbon chain and non-substituted hydrocarbon chain; and
W is selected from the group consisting of amide, ether, ester, disulfide,
thioether, thioester, imine and alkene,
and further wherein should said D is attached at one end to A (Formulas VI,
VII and X), said U or said V are being attached to one carbon atom in said
alkylene
chain and should said D is attached at one end to B1 or Bn (Formulas VIII, IX
and X),
said U or said V are being attached to one carbon atom in said alkylene chain,
provided that at least one of said X, Z, Y1 and Yn is a -NH group and/or at
least one of said carbon atoms in said alkylene chains is substituted by an
amine
group.
173. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 172, wherein said A is an alkylene chain having a general
formula II:
Image
wherein:
g is an integer that equals 0 or 3-10; and
147
each of R1, R2 and Rg is independently selected from the group consisting of
hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide, phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrile, alkyl isonitrile, aryl isonitrile, nitrate, nitrile, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, carboxylic acid, alkyl carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphonic acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
174. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 173, wherein each of B, and Bn is independently an
alkylene
chain having a general formula III:
148
Image
wherein:
p is an integer that equals 0 or g+1;
q is an integer from g+2 to g+20; and
each of Rp, Rp+1 and Rq is independently selected from the group consisting
of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroalicyclic,
heteroaryl, halo,
amino, alkylamino, arylamino, cycloalkylamino, heteroalicyclic amino,
heteroarylamino, hydroxy, alkoxy, aryloxy, azo, C-amido, N-amido, ammonium,
thiohydroxy, thioalkoxy, thioaryloxy, sulfonyl, sulfinyl, N-sulfonamide, S-
sulfonamide phosphonyl, phosphinyl, phosphonium, carbonyl, thiocarbonyl, C-
carboxy, O-carboxy, C-thiocarboxy, O-thiocarboxy, N-carbamate, O-carbamate, N-
thiocarbamate, O-thiocarbamate, urea, thiourea, borate, borane, boroaza,
silyl, siloxy,
silaza, aquo, alcohol, peroxo, amine oxide, hydrazine, alkyl hydrazine, aryl
hydrazine,
nitric oxide, cyanate, thiocyanate, isocyanate, isothiocyanate, cyano,
alkylnitrile, aryl
nitrite, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkyl
sulfonic acid, aryl
sulfonic acid, alkyl sulfoxide, aryl sulfoxide, alkyl aryl sulfoxide, alkyl
sulfenic acid,
aryl sulfenic acid, alkyl sulfinic acid, aryl sulfinic acid, alkyl thiol
carboxylic acid,
aryl thiol carboxylic acid, alkyl thiol thiocarboxylic acid, aryl thiol
thiocarboxylic
acid, alkyl carboxylic acid, carboxylic acid, aryl carboxylic acid, sulfate,
sulfite,
bisulfate, thiosulfate, thiosulfite, alkyl phosphine, aryl phosphine, alkyl
phosphine
oxide, aryl phosphine oxide, alkyl aryl phosphine oxide, alkyl phosphine
sulfide, aryl
phosphine sulfide, alkyl aryl phosphine sulfide, alkyl phosphonic acid, aryl
phosphoric acid, alkyl phosphinic acid, aryl phosphinic acid, phosphate,
thiophosphate, phosphate, pyrophosphate, triphosphate, hydrogen phosphate,
dihydrogen phosphate, guanidino, S-dithiocarbamate, N-dithiocarbamate,
bicarbonate,
carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate,
bromite,
hypobromite, tetrahalomanganate, tetrafluoroborate, hexafluoroantimonate,
hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkyl
borate,
149
tartarate, salicylate, succinate, citrate, ascorbate, saccharirate, amino
acid, hydroxamic
acid and thiotosylate.
175. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 173, wherein at least one of said C1, C2 and Cg is a
chiral carbon
atom.
176. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 174, wherein at least one of said Cp, Cp+1 and Cq is a
chiral
carbon atom.
177. The method, the assay and/or the transplantable hematopoietic cell
preparation of claim 137, wherein said polyamine chelator is selected from the
group
consisting of ethylendiamine, diethylenetriamine, triethylenetetramine,
triethylenediamine, tetraethylenepentamine, aminoethylethanolamine,
aminoethylpiperazine, pentaethylenehexamine, captopril, penicilamine, N,N'-
bis(3-
aminopropyl)-1,3-propanediamine, N,N'-Bis-(2-animoethyl)-1,3-propanediamine,
1,7-dioxa-4,10-diazacyclododecane, 1,4,8,11-tetraaza cyclotetradecane-5,7-
dione,
1,4,7-triazacyclononane, 1-oxa-4,7,10-triazacyclododecane, 1,4,8,12-
tetraazacyclopentadecane, and 1,4,7,10-tetraazacyclododecane.
178. An assay of determining whether a retinoic acid receptor antagonist is
an effective hematopoietic stem cell expansion agent, the assay comprising
culturing
hematopoietic mononuclear cells which comprise a major fraction of
hematopoietic
committed cells and a minor fraction of hematopoietic stem and progenitor
cells in the
presence of the retinoic acid receptor antagonist and monitoring expansion of
said
hematopoietic stem cells, wherein if increased expansion and decreased
differentiation of said hematopoietic stem cells occurs, as compared to non-
treated
hematopoietic mononuclear cells, the retinoic acid receptor antagonist is an
effective
hematopoietic stem cell expansion agent.
179. The assay of claim 178, wherein said retinoic acid receptor antagonist
is selected from the group consisting of: AGN 194310; AGN 193109; 3-(4-Methoxy-
150
phenylsulfanyl)-3-methyl-butyric acid; 6-Methoxy-2,2-dimethyl-thiochroman-4-
one,2,2-Dimethyl-4-oxo-thiochroman-6-yltrifluoromethane-sulfonate; Ethyl 4-
((2,2
dimethyl-4-oxo-thiochroman-6-yl)ethynyl)-benzoate; Ethyl 4-((2,2-dimethy 1-4-
triflouromethanensulfonyloxy -(2H)- thiochromen-6-yl)ethynyl)-benzoate(41);
Thiochromen-6-yl]-ethynyl]-benzoate(yl); (p-[(E)-2-[3'4'-Dihydro-4,4'-dimethyl-
7'-
(heptyloxy)-2'H-1-benzothiopyran-6'yl] propenyl] benzoic acid 1'1'-dioxide;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-butoxyphenyl)-3-methyl]-octa-2,4,6-trienoic
acid;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-propoxyphenyl)-3-methyl]-octa-2,4,6-trienoic
acid;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-pentoxyphenyl)-3-methyl]-octa-2,4,6-trienoic
acid;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-hexoxyphenyl)-3-methyl]-octa-2,4,6-trienoic
acid;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-heptoxyphenyl)-3-methyl]-octa-2,4,6-trienoic
acid;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-octoxyphenyl)-3-methyl]-octa-2,4,6-trienoic
acid;
(2E,4E,6E)-7-[3-t-butyl-5-(1-phenyl-vinyl)-phenyl]-3-methyl-octa-2,4,6-
trienoic acid;
2E,4E,6E-[7-(3,5-Di-t-butyl-4-{[4,5-3 H2]-n-pentoxy}phenyl)-3-
methyl]-
octa-2,4,6-trienoic acid; (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert.butyl-2-ethoxy-
phenyl)-
cyclopropyl]-3-methyl-penta-2,4-dienoic acid ethyl ester; (2E,4E)-(1RS,2RS)-5-
[2-
(3,5-di-tert.butyl-2-ethoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-dienoic
acid;
(2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert.butyl-2-butoxy-phenyl)-cyclopropyl]-3-
methyl-
penta-2,4-dienoic acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-ethoxyphenyl]3-
methyl-
2,4,6-octatrienoic acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-butyloxyphenyl]-3-
methyl-
2,4,6-octatrienoic acid; 4-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-
naphthalene-
carboxamido) benzoic acid; (2E,4E)-3-methyl-5-[(1S,2S)-2-(5,5,8,8-tetramethyl-
;
5,6,7,8-tetrahydro-naphthalen-2-yl)-cyclopropyl]-penta-2,4-dienoic acid; p-
[(E)-2-
[3',4'-Dihydro-4',4'-dimethyl-7'-(heptyloxy)-2'H-1-benzothiopyran-6'-
yl]propenyl]benzoic acid; 1',1'-dioxide, 4-(7,7,10,10-Tetramethyl-1-pyridin-3-
ylmethyl-4,5,7,8,9,10-hexahydro-1H-naphto[2,3-g]indol-3-yl)-benzoic acid;
(2E,4E,6Z)-7-[3,5-di-tert.butyl-2-methoxyphenyl]-3-methyl-2,4,6-octatrienoic
acid;
(2E,4E,6Z)-7-[3,5-di-tert.butyl-2-ethoxyphenyl]-3-methyl-2,4,6-octatrienoic
acid;
(2E,4E,6Z)-7-[3,5-di-tert.butyl-2-hexyloxyphenyl]-3-methyl-2,4,6-octatrienoic
acid;
(2E,4E,6Z)-7-[3,5-di-tert.butyl-2-octyloxyphenyl]-3-methyl-2,4,6-octatrienoic
acid;
and (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert-butyl-2-butoxy-phenyl)-cyclopropyl]-3-
methyl-penta-2,4-dienoic acid, (2E,4E,6Z)-7-(3-n-propoxy-5,6,7,8-tetrahydro-
5,5,8,8-
tetramethylnaphthalene-2-yl)-3-methylocta-2,4,6-trienoic acid, 4-(5H-2,3(2,5
151
dimethyl-2,5-hexano)-5-n-propyldibenzo[b,e][1,4]diazepin-11-yl)benzoic acid, 4-
(5H-2,3-(2,5-dimethyl-2,5-hexano)-5methyl-8-nitrodibenzo[b,e] [1,4]diazepin-11-
yl)benzoic acid, 4-{[4-(4-Ethylphenyl)2,2-dimethyl-(2H)-thiochromen-6-
yl]ethynyl}benzoic acid, 4-[4-2methyl-1,2-dicarba-closo-dodecaboran-1-yl-
phenylcarbamoyl]benzoic acid, 4-[4,5,7,8,9,10-hexahydro-7,7,10,10-tetramethyl-
1-(3-
pyridylmethyl)-anthra[1,2-b]pyrrol-3-yl]benzoic acid, (3-pyridylmethyl)-]5-
thiaanthra[2,1-b]pyrrol-3-yl)benzoic acid, and (3-pyridylmethyl)-anthra[2ml-
d]pyrazol-3-yl]benzoic acid.
180. An assay of determining whether a retinoid X receptor antagonist is an
effective hematopoietic stem cell expansion agent, the assay comprising
culturing
hematopoietic mononuclear cells which comprise a major fraction of
hematopoietic
committed cells and a minor fraction of hematopoietic stem and progenitor
cells in the
presence of the retinoid X receptor antagonist and monitoring expansion of
said
hematopoietic stem cells, wherein if increased expansion and decreased
differentiation of said hematopoietic stem cells occurs, as compared to non-
treated
hematopoietic mononuclear cells, the retinoid X receptor antagonist is an
effective
hematopoietic stem cell expansion agent.
181. The assay of claim 180, wherein said retinoid X receptor antagonist is
selected from the group consisting of LGN100572, LGN100574, 1-(3-hydroxy-
5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)ethanone, 1-(3-propoxy-
5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)ethanone, 3-(3-propoxy-
5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)but-2-enenitrile, 3-(3-
propoxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)but-2-enal,
(2E,4E,6E)-7-3[-propoxy-5,6,7,8-tetrahydro 5,5,8,8-tetramethyl-2-naphthalene-2-
yl]-
3-methylocta-2,4,6-trienoic acid, 4-[3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-
2-
naphthyl)carbonyl] benzoic acid, 4-[1-(3,5, 5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-
naphthyl)ethenyl] benzoic acid, 4-[1(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-
2-
naphthyl)cyclopropyl] benzoic acid, 4-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-
naphthyl)ethenyl] benzenete trazole, 2-[1-(5,5,8,8-tetramethyl-5,6,7,8-
tetrahydro-2-
naphthyl) ethenyl]pyridine-5-carboxylic acid, 2-[1-(3,5,5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethyl]pyridine-5-carboxylic acid, ethyl-2-[1-(3,5,5,8, 8-
152
pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl]pyridine-5-carboxylate, 5-[1-
3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl]pyridine-2-
carboxylic
acid, 2-[1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)
cyclopropyl]pyridine-5-carboxylic acid, methyl 2-[1-(3,5,5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)cyclopropyl]pyridine-5-carboxylate, 4-[1-(3,5, 5,8,8-
pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl]-N-(4-hydroxyphenyl)
benzamide,
2-[1-(3,5,5,8,8-Pentamethyl-5,6,7,8-tetrahydro-2-naphthyl) ethenyl] pyridine-5-
carboxylic acid, 2-[1-(3,5,5,8,8-Pentamethyl-5, 6,7,8-tetrahydro-2-
naphthyl)cyclopropyl]pyridine-5-carboxylic acid, 4-[(3,5, 5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)carbonyl]benzoic acid butyloxime, 4-[(3,5,5,8,8-
pentamethyl-
5,6,7,8-tetrahydro-2-naphthyl) carbonyl]benzoic acid propyloxime, 4-
[(3,5,5,8,8-
pentamethyl-5,6,7,8-terrahydro-2-naphthyl)carbonyl]benzoic acid cyanoimine, 4-
[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl]benzoic acid
allyloxime, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)carbonyl]benzoic
acid 4-(3-methylbut-2-enoic acid)oxime, and 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-naphthyl)carbonyl]benzoic acid 1-aminoethyloxime, (2E,4E,6Z)-7-(3-
n-
propoxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-
2,4,6-
trienoic acid, 4-(5H-2,3(2,5 dimethyl-2,5-hexano)-5-n-
propyldibenzo[b,e][1,4]diazepin-11-yl)benzoic acid, and 4-(5H-2,3-(2,5-
dimethyl-
2,5-hexano)-5methyl-8-nitrodibenzo[b,e][1,4]diazepin-11-yl)benzoic acid.
182. An assay of determining whether a vitamin D receptor antagonist is an
effective hematopoietic stem cell expansion agent, the assay comprising
culturing
hematopoietic mononuclear cells which comprise a major fraction of
hematopoietic
committed cells and a minor fraction of hematopoietic stem and progenitor
cells in the
presence of the vitamin D receptor antagonist and monitoring expansion of said
hematopoietic stem cells, wherein if increased expansion and decreased
differentiation of said hematopoietic stem cells occurs, as compared to non-
treated
hematopoietic mononuclear cells, the vitamin D receptor antagonist is an
effective
hematopoietic stem cell expansion agent.
183. The assay of claim 182, wherein said Vitamin D receptor antagonist is
selected from the group consisting of: 1 alpha, 25-(OH)-D3-26,23 lactone; 1
alpha, 25-
153
dihydroxyvitamin D (3); the 25-carboxylic ester ZK159222; (23S)- 25-dehydro-1
alpha-OH-D (3); (23R)-25-dehydro-1 alpha-OH-D (3); 1 beta, 25 (OH)2 D3; 1
beta,
25(OH)2-3-epi-D3; (23S) 25-dehydro-1 alpha(OH) D3-26,23-lactone; (23R) 25-
dehydro-1 alpha(OH)D3-26,23-lactone and Butyl-(5Z,7E,22E-(1S,7E,22E-
(1S,3R,24R)-1,3,24-trihydroxy-26,27-cyclo-9,10-secocholesta-5,7,10(19),22-
tetraene-
25-carboxylate).
184. An assay of determining whether an agent that inhibits PI 3-kinase
activity is an effective hematopoietic stem cell expansion agent, the assay
comprising
culturing hematopoietic mononuclear cells which comprise a major fraction of
hematopoietic committed cells and a minor fraction of hematopoietic stem and
progenitor cells in the presence of the agent that inhibits PI 3-kinase
activity and
monitoring expansion of said hematopoietic stem cells, wherein if increased
expansion and decreased differentiation of said hematopoietic stem cells
occurs, as
compared to non-treated hematopoietic mononuclear cells, the agent that
inhibits PI 3-
kinase activity is an effective hematopoietic stem cell expansion agent.
185. An assay of determining whether a nicotinamide analog, a
nicotinamide or a nicotinamide analog derivative or a nicotinamide or a
nicotinamide
analog metabolite is an effective hematopoietic stem cell expansion agent, the
assay
comprising culturing hematopoietic mononuclear cells which comprise a major
fraction of hematopoietic committed cells and a minor fraction of
hematopoietic stem
and progenitor cells in the presence of a nicotinamide analog, a nicotinamide
or a
nicotinamide analog derivative or a nicotinamide or a nicotinamide analog
metabolite
and monitoring expansion of said hematopoietic stem cells, wherein if
increased
expansion and decreased differentiation of said hematopoietic stem cells
occurs, as
compared to non-treated hematopoietic mononuclear cells, the a nicotinamide
analog,
a nicotinamide or a nicotinamide analog derivative or a nicotinamide or a
nicotinamide analog metabolite is an effective hematopoietic stem cell
expansion
agent.
154
186. The assay of any of claims 178, 180, 182, 184 and 185, wherein
culturing said hematopoietic mononuclear cells is performed in a presence of
an
effective amount of a cytokine.
187. The assay of claim 186, wherein said cytokine is an early acting
cytokines.
188. The assay of claim 187, wherein said early acting cytokine is selected
from the group comprising stem cell factor, FLT3 ligand, interleukin-1,
interleukin-2,
interleukin-3, interleukin-6, interleukin-10, interleukin-12, tumor necrosis
factor-a
and thrombopoietin.
189. The assay of claim 186, wherein said cytokine is a late acting
cytokines.
190. The assay of claim 189, wherein said late acting cytokines are selected
from the group comprising granulocyte colony stimulating factor,
granulocyte/macrophage colony stimulating factor, erythropoietin, FGF, EGF,
NGF,
VEGF, LIF, Hepatocyte growth factor and macrophage colony stimulating factor.
191. The assay of any of claims 178, 180, 182, 184 and 185, wherein said
hematopoietic mononuclear cells are derived from a source selected from the
group
consisting of bone marrow, peripheral blood and neonatal umbilical cord blood.
192. The assay of any of claims 178, 180, 182, 184 and 185, wherein said
monitoring decreased differentiation is by determining hematopoietic cell
surface
expression of CD34.
193. The assay of any of claims 178, 180, 182, 184 and 185, wherein said
monitoring decreased differentiation is by determining an absence, or
significantly
diminished hematopoietic cell surface expression of CD38, CD3, CD61, CD19,
CD33, CD14, CD15 or CD4.
155
194. A hematopoietic stem cells collection/culturing bag supplemented with
an effective amount of a retinoic acid receptor antagonist, a retinoid X
receptor
antagonist and/or a Vitamin D receptor antagonist, which substantially
inhibits cell
differentiation of a hematopoietic stem cells fraction of hematopoietic
mononuclear
cells which comprise a major fraction of hematopoietic committed cells and a
minor
fraction of hematopoietic stem and progenitor cells.
195. The hematopoietic stem cells collection/culturing bag of claim 194,
wherein said retinoic acid receptor antagonist is selected from the group
consisting of
AGN 194310; AGN 193109; 3-(4-Methoxy-phenylsulfanyl)-3-methyl-butyric acid;
6-Methoxy-2,2-dimethvl-thiochroman-4-one,2,2-Dimethyl-4-oxo-thiochroman-6-
yltrifluoromethane-sulfonate; Ethyl 4-((2,2 dimethyl-4-oxo-thiochroman-6-
yl)ethynyl)-benzoate; Ethyl 4-((2,2-dimethy 1-4-triflouromethanensulfonyloxy -
(2H)-
thiochromen-6-yl)ethynyl)-benzoate(41); Thiochromen-6-yl]-ethynyl]-
benzoate(yl);
(p-[(E)-2-[3'4'-Dihydro-4,4'-dimethyl-7'-(heptyloxy)-2'H-1-benzothiopyran-
6'yl]
propenyl] benzoic acid 1'1'-dioxide; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
butoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
propoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
pentoxyphenyl)-
3-methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
hexoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
heptoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-n-
octoxyphenyl)-3-
methyl]-octa-2,4,6-trienoic acid; (2E,4E,6E)-7-[3-t-butyl-5-(1-phenyl-vinyl)-
phenyl]-
3-methyl-octa-2,4,6-trienoic acid; 2E,4E,6E-[7-(3,5-Di-t-butyl-4-{[4,5-3
H2]-n-pentoxy}phenyl)-3-methyl]-octa-2,4,6-trienoic acid; (2E,4E)-
(1RS,2RS)-
S-[2-(3,S-di-tert.butyl-2-ethoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-
dienoic acid
ethyl ester; (2E,4E)-(1RS,2RS)-S-[2-(3,5-di-tert.butyl-2-ethoxy-phenyl)-
cyclopropyl]-
3-methyl-penta-2,4-dienoic acid; (2E,4E)-(1RS,2RS)-S-[2-(3,5-di-tert.butyl-2-
butoxy-phenyl)-cyclopropyl]-3-methyl-penta-2,4-dienoic acid; (2E,4E,6Z)-7-[3,5-
di-
tert.butyl-2-ethoxyphenyl]3-methyl-2,4,6-octatrienoic acid; (2E,4E,6Z)-7-[3,5-
di-
tert.butyl-2-butyloxyphenyl]-3-methyl-2,4,6-octatrienoic acid; 4-(5,6,7,8-
tetrahydro-
5,5,8,8-tetramethyl-2-naphthalene-carboxamido) benzoic acid; (2E,4E)-3-methyl-
5-
[(1S,2S)-2-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-naphthalen-2-yl)-
cyclopropyl]-
penta-2,4-dienoic acid; p-[(E)-2-[3',4'-Dihydro-4',4'-dimethyl-7'-(heptyloxy)-
2'H-1-
156
benzothiopyran-6'-yl]propenyl]benzoic acid; 1',1'-dioxide, 4-(7,7,10,10-
Tetramethyl-
1-pyridin-3-ylmethyl-4,5,7,8,9,10-hexahydro-1H-naphto[2,3-g]indol-3-yl)-
benzoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-methoxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-ethoxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-hexyloxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; (2E,4E,6Z)-7-[3,5-di-tert.butyl-2-octyloxyphenyl]-3-methyl-2,4,6-
octatrienoic
acid; and (2E,4E)-(1RS,2RS)-5-[2-(3,5-di-tert-butyl-2-butoxy-phenyl)-
cyclopropyl]-
3-methyl-penta-2,4-dienoic acid. (2E,4E,6Z)-7-(3-n-propoxy-5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-2,4,6-trienoic acid, 4-(5H-
2,3(2,5
dimethyl-2,5-hexano)-5-n-propyldibenzo[b,e] [1,4]diazepin-11-yl)benzoic acid, 4-
(5H-2,3-(2,5-dimethyl-2,5-hexano)-5methyl-8-nitrodibenzo[b,e][1,4]diazepin-11-
yl)benzoic acid, 4-{[4-(4-Ethylphenyl)2,2-dimethyl-(2H)-thiochromen-6-
yl]ethynyl}benzoic acid, 4-[4-2methyl-1,2-dicarba-closo-dodecaboran-1-yl-
phenylcarbamoyl]benzoic acid, 4-[4,5,7,8,9,10-hexahydro-7,7,10,10-tetramethyl-
1-(3-
pyridylmethyl)-anthra[1,2-b]pyrrol-3-yl]benzoic acid, (3-pyridylmethyl)-]5-
thiaanthra[2,1-b]pyrrol-3-yl)benzoic acid, and (3-pyridylmethyl)-anthra[2m1-
d]pyrazol-3-yl]benzoic acid.
196. The hematopoietic stem cell collection bag of claim 194, wherein said
retinoid X receptor antagonist is selected from the group consisting of:
LGN100572,
LGN100574, 1-(3-hydroxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-
yl)ethanone, 1-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-
yl)ethanone, 3-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-
yl)but-2-enenitrile, 3-(3-propoxy-5,6,7,8-tetrahydro-5,5,8,8-
tetramethylnaphthalene-
2-yl)but-2-enal, (2E,4E,6E)-7-3[-propoxy-5,6,7,8-tetrahydro 5,5,8,8-
tetramethyl-2-
naphthalene-2-yl]-3-methylocta-2,4,6-trienoic acid, 4-[3,5,5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)carbonyl] benzoic acid, 4-[1-(3,5, 5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethenyl] benzoic acid, 4-[1(3,5,5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)cyclopropyl] benzoic acid, 4-[1-(3,5,5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl)ethenyl] benzenete trazole, 2-[1-(5,5,8,8-tetramethyl-
5,6,7,8-
tetrahydro-2-naphthyl) ethenyl]pyridine-5-carboxylic acid, 2-[1-(3,5,5,8,8-
pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethyl]pyridine-5-carboxylic acid,
ethyl-2-
[1-(3,5,5,8, 8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)ethenyl]pyridine-5-
157
carboxylate, 5-[1-3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)ethenyl)pyridine-2-carboxylic acid, 2-[1-(3,5,5,8,8-pentamethyl-
5,6,7,8-
tetrahydro-2-naphthyl) cyclopropyl]pyridine-5-carboxylic acid, methyl 2-[1-
(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)cyclopropyl]pyridine-5-
carboxylate, 4-[1-(3,5, 5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)ethenyl]-N-
(4-hydroxyphenyl) benzamide, 2-[1-(3,5,5,8,8-Pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl) ethenyl] pyridine-5-carboxylic acid, 2-[1-(3,5,5,8,8-Pentamethyl-5,
6,7,8-
tetrahydro-2-naphthyl)cyclopropyl]pyridine-5-carboxylic acid, 4-[(3,5, 5,8,8-
pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)carbonyl]benzoic acid butyloxime, 4-
[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl) carbonyl]benzoic acid
propyloxime, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-terrahydro-2-
naphthyl)carbonyl]benzoic acid cyanoimine, 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-
tetrahydro-2-naphthyl)carbonyl]benzoic acid allyloxime, 4-[(3,5,5,8,8-
pentamethyl-
5,6,7,8-tetrahydro-2-naphthyl)carbonyl]benzoic acid 4-(3-methylbut-2-enoic
acid)oxime, and 4-[(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-
naphthyl)carbonyl)benzoic acid 1-aminoethyloxime, (2E,4E,6Z)-7-(3-n-propoxy-
5,6,7,8-tetrahydro-5,5,8,8-tetramethylnaphthalene-2-yl)-3-methylocta-2,4,6-
trienoic
acid, 4-(5H-2,3(2,5 dimethyl-2,5-hexano)-5-n-propyldibenzo[b,e][1,4]diazepin-
11-
yl)benzoic acid, and 4-(5H-2,3-(2,5-dimethyl-2,5-hexano)-5methyl-8-
nitrodibenzo[b,e][1,4]diazepin-11-yl)benzoic acid.
197. The hematopoietic stem cell collection bag of claim 194, wherein said
Vitamin D receptor antagonist is selected from the group consisting of 1
alpha, 25-
(OH)-D3-26,23 lactone; 1 alpha, 25-dihydroxyvitamin D (3); the 25-carboxylic
ester
ZK159222; (235)- 25-dehydro-1 alpha-OH-D (3); (23R)-25-dehydro-1 alpha-OH-D
(3); 1 beta, 25 (OH)2 D3; 1 beta, 25(OH)2-3-epi-D3; (235) 25-dehydro-1
alpha(OH)
D3-26,23-lactone; (23R) 25-dehydro-1 alpha(OH)D3-26,23-lactone and Butyl-
(5Z,7E,22E-(1S,7E,22E-(1S,3R,24R)-1,3,24-trihydroxy-26,27-cyclo-9,10-
secocholesta-5,7,10(19),22-tetraene-25-carboxylate).
198. A hematopoietic stem cells collection/culturing bag supplemented with
an effective amount of nicotinamide, a nicotinamide analog, a nicotinamide or
a
nicotinamide analog derivative or a nicotinamide or a nicotinamide analog
metabolite,
158
which substantially inhibits cell differentiation of a hematopoietic stem
cells fraction
of hematopoietic mononuclear cells which comprise a major fraction of
hematopoietic
committed cells and a minor fraction of hematopoietic stem and progenitor
cells.
199. A hematopoietic stem cells collection/culturing bag supplemented with
an effective amount of an agent that inhibits PI 3-kinase activity, which
substantially
inhibits cell differentiation of a hematopoietic stem cells fraction of
hematopoietic
mononuclear cells which comprise a major fraction of hematopoietic committed
cells
and a minor fraction of hematopoietic stem and progenitor cells.
200. An ex-vivo expanded population of hematopoietic stem cells, obtained
by the method of any of claims 1, 38, 59, 80, 95 and 118.
