DESCRIPTION
THIOPHENE COMPOUNDS AND THROMBOPOIETIN RECEPTOR
ACTIVATORS
5
BACKGROUND OF THE INVENTION TECHNICAL FIELD
The present invention relates to preventive, therapeutic and improving agents having affinity for and
10 agonistic action on the thrombopoietin receptor for
diseases against which activation of the thrombopoietin receptor is effective. Specifically, it relates to pharmaceutical compositions comprising compounds which increase platelets through stimulation of differentiation
is and proliferation of hematopoietic stem cells,
megakaryocytic progenitor cells and megakaryocytes or compounds for therapeutic angiogenesis or with anti-arteriosclerosis action that stimulate.differentiation and proliferation of vascular endothelial cells and
20 endothelial progenitor cells.
BACKGROUND ART
Thrombopoietin is.a cytokine consisting of 332 amino acids that increases platelet production by stimulating 25 differentiation and proliferation of hematopoietic stem cells, megakaryocytic progenitor cells and megakaryocytes mediated by its receptor and therefore is promising as a

drug for hematological disorders. Recent reports that it stimulates differentiation and proliferation of vascular endothelial cells and endothelial progenitor cells have raised expectations of therapeutic angiogenesis, anti-s arteriosclerosis and prevention of cardiovascular events (for example, non-patent document 1, non-patent document 2 and non-patent document 3).
Biologically active substances which have been known so far to regulate platelet production through the
10 thrombopoietin receptor include, in addition to
thrombopoietin itself, low molecular weight peptides having affinity for the thrombopoietin receptor (for example, patent document 1, patent document 2, patent document 3 and patent document 4).
15 As a result of search for nonpeptidic low molecular weight compounds that increase platelet production mediated by the thrombopoietin receptor, low molecular weight compounds having affinity for the thrombopoietin receptor have been reported (for example, patent document
20 5 to patent document 26).
1) Applications filed by Hokuriku Seiyaku Co., Ltd.
relating to 1,4-benzodiazepine derivatives (patent
documents 5 and 6)
2) International Laid-open Patent Applications filed by
25 Shionogi & Co., Ltd. (patent documents 7-10)
3) International Laid-open Patent Applications filed by
SmithKline Beecham Corp (patent documents 11-19)

4) Japanese Laid-open Patent Application filed by Torii Pharmaceutical Co., Ltd. (patent document 20)
5) International Laid-open Patent Application filed by-Roche Diagnostics GMBH (patent document 21)
s 6) International Laid-open Patent Applications filed by Yamanouchi Pharmaceutical Co., Ltd. (patent document 22 and 23)
7) Japanese Laid-open Patent Application filed by Japan Tabacco Inc. (patent document 24) 10 8) International Laid-open Patent Application filed by
Nissan Chemical Industries, Ltd. {patent documents 25 and 26)
Patent document 1 JP-A-10-72492 Patent document 2 WO96/40750 is Patent document 3 WO96/40189 Patent document 4 W098/25965 Patent document 5 JP-A-11-1477 Patent document 6 JP-A-11-152276 Patent document 7 WO01/07423 20 Patent document 8 WOOl/53267 Patent document 9 WO02/059099 Patent document 10 WO02/059100 Patent document 11 WO00/35446 Patent document 12 WO00/66112 25 Patent document 13 WOOl/34585 Patent document 14 WOOl/17349 Patent document 15 WO01/39773

Patent document 16 WO01/21180
Patent document 17 WO01/89457
Patent document 18 WO02/49413
Patent document 19 WO02/085343 5 Patent document 20 JP-A-2001-97948
Patent document 21 W099/11262
Patent document 22 W002/06277S
Patent document 23 WO03/062233
Patent document 24 JP-A-2003-238565 10 Patent document 25 WO04/033433
Patent document 26 WO04/108683
Non-patent document 1 Microvasc. Res., 1999: 58, p.108-113
Non-patent document 2 Circ. Res., 1999: 84, p. 785-15 796
Non-patent document 3 Blood 2001:98, p.71a-72a
DISCLOSURE OF THE INVENTION
Thrombopoietin and low molecular weight peptides
20 having affinity for the thrombopoietin receptor are likely to be easily degraded in the gastrointestinal tract and are usually difficult to orally administer. As to thrombopoietin itself, the appearance of anti-thrombopoietin antibodies have been reported.
25 Besides, though it is probably possible to orally
administer nonpeptidic low molecular weight compounds, no .practical drugs haye been put on the market.

Therefore, orally administrable low molecular weight compounds having excellent affinity for and agonistic action on the thrombopoietin receptor as preventive, therapeutic and improving agents for diseases against 5 which activation of the thrombopoietin receptor is effective have been demanded. Specifically, low molecular weight compounds which can serve as platelet increasing agents or increasing agents for other blood cells by stimulating differentiation and proliferation of
10 hematopoietic stem cells, megakaryocytic progenitor cells and megakaryocytes or low molecular weight compounds which can be used for therapeutic angiogenesis or as preventive and therapeutic agents for arteriosclerosis by stimulating endothelial cells and endothelial progenitor
15 cells have been demanded.
The present inventors conducted extensive research to find low molecular weight compounds having affinity for and agonistic action on the thrombopoietin receptor, and as a result, found that the compounds of the present
20 invention have high affinity and agonistic action which enable them to show potent platelet increasing action by stimulating differentiation and proliferation of megakaryocytic progenitor cells and megakaryocytes.
25
Namely, the present invention relates to: 1. A compound represented by the formula (I):


wherein R1 is a phenyl group (the phenyl group may be substituted with one or more C3.-6 alkyl groups, one or more Cj.3 alkyl groups (the C1-3 alkyl groups are 5 substituted with one or more halogen atoms), one or more C1-3 alkoxy groups (the C1.3 alkoxy groups may be substituted with one or more halogen atoms) or one or more halogen atoms), R3 is a hydrogen atom or a Ci_3 alkyl group (the Ci_3 alkyl
10 group may be substituted with one or more halogen atoms), R3 is a phenyl group, a pyridyl group or a thienyl' group (the phenyl group, the pyridyl group and the thienyl group are substituted with one or more substituents selected from the group consisting of hydrogen atoms,
is nitro groups, halogen atoms and C1-3 alkyl groups (the C1-3 alkyl groups may be-substituted with one or more halogen atoms) and with (C=0)Rs (wherein R5 is NR6R7 (wherein Rs is a hydrogen atom or a Cj^ alkyl group (the C1-3 alkyl group may be substituted with one or more halogen atoms),
20 and R7 is a C1-6 alkyl group (the C1-6 alkyl group may be substituted with one or more halogen atoms, one or more hydroxyl groups, one or more C1-3 alkoxy groups or one or more C2-14 aryl groups (the C2-14. aryl groups may be

substituted with one or more C1-3 alkyl groups, one or more C1-3 alkoxy groups, one or more carboxyl groups, one or more carbamoyl groups, one or more cyano groups or one or more halogen atoms, and in the case of aryl groups 5 containing one or more nitrogen atoms, may be N-oxides thereof)), a phenyl group, a thienyl group, a pyridyl group or a pyridyl-N-oxide group (the phenyl group, the thienyl group, the pyridyl group and the pyridyl-N-oxide group may be substituted with one or more halogen atoms),
10 or NR6R7 is, as a whole, "a nitrogen-containing
heterocyclyl group (the nitrogen-containing heterocyclyl group may be substituted with one or more hydrogen atoms, one or more C1-6 alkyl groups (the c1-6 alkyl groups may be substituted with one or more halogen atoms), one pr more
15 halogen atoms, one or more hydroxyl groups or one or more C1-3 alkoxy groups (the C1-3 alkoxy groups may be substituted with one or more halogen atoms))) or a C1-6 alkyl group (the C1-6 alkyl group may be substituted with one or more halogen atoms, one or more pyridyl groups,
20 one or more pyridyl-N-oxide groups, one or more furyl
groups, one or more thienyl groups or one or more phenyl groups and is substituted with one or more cyano groups))), and R4 is a hydrogen atom or a C1-3 alkyl group (the C1-3 alkyl
25 group may be substituted with one or more halogen atoms), a tautomer, prodrug or pharmaceuticaly acceptable salt of the compound or a solvate thereof.

2. The compound according to 1, wherein R2 is a methyl group, and R* is a hydrogen atom, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof. 5 3. The compound according to 2, wherein R1 is a 3,4-dimethyl-phenyl group, a 4-t-butyl-phenyl group, a 4-trifluoromethyl-phenyl group, a 3-chloro-phenyl group, a 4-chloro-phenyl group, a 4-fluoro-phenyl group, a 3,4-dichloro-phenyl group, a 4-bromo-phenyl group or a 4-10 trifluoromethoxy-phenyl group, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.
4. The compound according to 3, wherein R3 is
represented by the formula (II):
15
(wherein R6 is a methyl group or an ethyl group, and R7 is a C1-6 alkyl group, (the C1-6 alkyl group may be substituted with one or more methoxy groups)), a tautomer, prodrug or pharmaceutically acceptable salt of 20 the compound or a solvate thereof.
5. The compound according to 3, wherein R3 is
represented by the formula (II) :


(wherein R6 is a methyl group or an ethyl group, and R7 is a C1-3 alkyl group (the C1-3 alkyl group is substituted with one or more phenyl groups or one or more pyridyl groups)), a tautomer, prodrug or pharmaceutically 5 acceptable salt of the compound or a solvate thereof.
6. The compound according to 3, wherein R3 is
represented by the formula [II):

(wherein R6 is a hydrogen atom, and R7 is a C1-5 alkyl 10 group (the C1-6 alkyl group is substituted with one or more methoxy groups) or a pyridyl group), a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.
7. The compound according to 3, wherein R3 is
is represented by the formula (II):

(wherein NR6R7 is, as a whole, represented by the formula

20 " (wherein R9 is a C1-3 alkyl group)), a tautomer, prodrug .

or pharmaceutically acceptable salt of the compound or a solvate thereof.
8. The compound according to 3, wherein R3 is
represented by the formula (IV):
5
(wherein R6 is a hydrogen atom, R7 is a C1-6 alkyl group
(the C1-3 alkyl group may be substituted with one or more
hydroxyl groups), and R8 is a methyl group .or a chlorine
atom), a tautomer, prodrug or pharmaceutically acceptable
10 salt of the compound or a solvate thereof.
9. The compound according to 3, wherein R3 is
represented by the formula (V):

(wherein R10 is a hydrogen atom or a C1-3 alkyl group) , a is tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.
10. The compound according to 3, wherein R3 is
represented by the formula (II):


(wherein R6 is a hydrogen atom, and R7 is an isopropyl group, a methyl group, an ethyl group or a normal propyl group (the methyl group, the ethyl group and the normal propyl group "are unsubstituted or substituted with one or 5 more pyridyl groups, one or more pyridyl-N-oxide groups, one or more furyl groups, one or more pyrazinyl groups, one or more imidazolyl groups, one or more pyrazolyl groups or one or more isoxazolyl groups (the pyridyl groups, the pyridyl-N-oxide groups, the furyl groups, the
10 pyrazinyl groups, the imidazolyl groups, the pyrazolyl groups and the isoxazolyl groups may be substituted with one or more methyl groups, one or more methoxy groups, one or more carboxyl groups or one or more halogen atoms))), a tautomer, prodrug or pharmaceutically_
xs acceptable salt of the compound or a solvate thereof. 11. The compound according to any one.of 4 to 10, wherein R1 is a 3,4-dimethyl-phenyl group, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.
20 12. The compound according to any one of 4 to 10,
wherein R1 is a 3,4-dichloro-phenyl group, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof. 13. The compound according to any one of 4 to 10,
25 wherein R1" is a 4-chloro-phenyl group, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.

14. The compound according to any one of 4 to 10, wherein R1 is a 4-trifluoromethyl-phenyl group, a tautomer, prodrug or pharmaceutically acceptable salt of v. the compound or a solvate thereof. 5 IS. The compound according to any one of 4 to 10,
wherein R1 is a 4-bromo-phenyl group, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.
16. The compound according to any one of 4 to 10,
10 wherein R1 is a 4-trifluoromethoxy-phenyl group, a
tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof.
17. A thrombopoietin receptor activator containing the
compound according to any one of 1 to 16, a tautomer,
15 prodrug or pharmaceutically acceptable salt of the
compound or a solvate thereof, as an active ingredient.
18. A preventive, therapeutic or improving agent for
diseases against which activation of the thrombopoietin
receptor is effective, which contains the thrombopoietin
20 receptor activator according to 17, as an active ingredient.
19. A platelet increasing agent containing the
thrombopoietin receptor activator according to 17, as an
active ingredient.
25 20. Medicament containing the compound according to any one of 1 to 16, a tautomer, prodrug or pharmaceutically acceptable salt of the compound or a solvate thereof, as

an active ingredient.
EFFECTS OF THE INVENTION
The thiophene compounds of the present invention 5 have affinity for and agonistic action on the thrombopoietin receptor and show potent platelet increasing action through stimulation of differentiation and proliferation of megakaryocytic progenitor cells and megakaryocytes.
10 " The thiophene compounds of the present invention are easily absorbable from the gastrointestinal tract and highly stimulate formation of megakaryocyte colonies. The orally absorbable thiophene compounds are retained in blood at high levels and therefore useful especially as
is oral medicines.
Though patent document 26 discloses compounds having platelet increasing action, it does not disclose the thiophene compounds of the present invention specifically enough to predict the especially excellent oral
20 absorbability and the excellent megakaryocyte colony stimulating activity of the thiophene compounds of the pre sent invent ion.
Therefore, the thiophene compounds of the present invention are useful as medicines and used as preventive,
25 therapeutic and improving agents for diseases against which activation of the thrombopoietin receptor is effective, especially as platelet increasing agents.

BEST MODE FOR CARRYING OUT THE INVENTION
Now, the present invention will be described in detail. 5 In the present invention, °n" denotes normal, "i"
denotes iso, "s" denotes secondary, "t" denotes tertiary, "c" denotes cyclo, "o" denotes ortho, "m" denotes meta, "p" denotes para, "Ph" denotes phenyl, "Py" denotes pyridyl, "Me" denotes methyl, "Et" denotes ethyl, "Pr"
it) denotes propyl, and "Bu" denotes butyl.
First, the terms in the respective substituents R1 to R1D will be explained.
As a halogen atom, a fluorine atom, a chlorine atom, a bromine atom or an iodine atom may be mentionedl-
15 A C1-3 alkyl group may be linear, branched-or a C3 ! cycloalkyl group, and methyl, ethyl, n-propyl, i-propyl and c-propyl and the like may be mentioned.
A C1-6 alkyl group may be linear, branched or a C3-s cycloalkyl group, and in addition to those mentioned
20 above, n-butyl, i-butyl, s-butyl/ t-butyl, c-butyl, 1-
methyl-c-propyl, 2-methyl-c-propyl, n-pentyl, 1-methyl-n-butyl, 2-methyl-n-butyl, 3-methyl-n-butyl, 1,1-dimethyl-n-propyl, 1,2-dimethyl-n-propyl, 2,2-dimethyl-n-propyl, 1-ethyl-n-propyl, c-pentyl, 1-methyl-c-butyl, 2-methyl-c-
25 butyl, 3-methyl-c-butyl, 1,2-dimethyl-c-propyl, 2,3-
dimethyl-c-propyl, 1-ethyl-c-propyl, 2-ethyl-c-propyl, n-hexyl, 1-methyl-n-pentyl, .2-methyl-n-pentyl, 3-methyl-n-

pentyl, 4-methyl-n-pentyl, 1,1-dimethyl-n-butyl,1,2-dimethyl-n-butyl, 1,3-dimethyl-n-butyl, 2,2-dimethyl-n-butyl, 2,3-dimethyl-n-butyl, 3,3-dimethyl-n-butyl, 1-ethyl-n-butyl, 2-ethyl-n-butyl, 1,1,2-trimethyl-n-propyl, 5 1,2,2-trimethyl-n-propyl, 1-ethyl-1-methyl-n-propyl, 1-ethyl-2-methyl-n-propyl, c-hexyl, 1-methyl-c-pentyl, 2-methyl-c-pentyl, 3-methyl-c-pentyl, 1-ethyl-c-butyl, 2-ethyl-c-butyl, 3-ethyl-c-butyl, 1,2-dimethyl-c-butyl, 1,3-dimethyl-c-butyl, 2,2-dimethyl-c-butyl, 2,3-dimethyl- '
10 c-butyl, 2,4-dimethyl-c-butyl, 3,3-dimethyl-c-butyl, 1-n-propyl-c-propyl, 2-n-propyl-c-propyl, 1-i-propyl-c-propyl, 2-i-propyl-c-propyl, 1,2,2-trimethyl-c-propyl, 1,2,3-trimethyl-c-propyl, 2,2,3-trimethyl-c-propyl, 1-ethyl-2-methyl-c-propyl, 2-ethyl-1-methyl-c-propyl, 2-
is ethyl-2-methyl-c-propyl, 2-ethyl-3-methyl-c-propyl- and the like may be mentioned.
A Ci-3 alkoxy group may include a linear, branched or C3 cycloalkoxy group, and methoxy, ethoxy, n-propoxy, i-propoxy, c-propoxy and the like may be mentioned.
20 A C2-14 aryl group may be a C6-14aryl group containing no hetero atoms as ring constituting atoms or a C2-9 aromatic heterocyclic group, and a C2-9 aromatic heterocyclic group may be a 5 to 7-membered C2_6 heteromonocyclic group or a 8 to 10-membered C5.9 fused
25 heterobicyclic group containing from 1 to 3 oxygen atoms, nitrogen atoms or sulfur atoms singly or in combination. •As a C6_14 aryl group containing no hetero atoms, a

phenyl group, a 1-indenyl group, a 2-indenyl group, a 3-indenyl group, a 4-indenyl group, a 5-indenyl group, a 6-indenyl group, a 7-indenyl group, an a-naphthyl group, a p-naphthyl group, a l-tetrahydronaphthyl group, a 2-
s tetrahydronaphthyl group, a 5-tetrahydronaphthyl group, a 6-tetrahydronaphthyl group, an o-biphenylyl group, a m-biphenylyl group, a p-biphenylyl group, a 1-anthryl group, a 2-anthryl group, a 9-anthryl group, a 1-phenanthryl group, a 2-phenanthryl group, a 3-phenanthryl
10 group, a 4-phenanthryl group, a 9-phenanthryl group or the like may be mentioned.
A 5 to 7-membered C2-6 heteromonocyclic group may be a 2-thienyl group, a 3-thienyl group, a 2-furyl group, a 3-furyl group, a 2-pyranyl group, a 3-pyranyl group', a 4-
15 pyranyl group, a 1-pyrrolyl group, a 2-pyrrolyl group, a 3-pyrrolyl group, a 1-imidazolyl group, a 2-imidazolyl group, a 4-imidazolyl group, a 1-pyrazolyl group, a 3-pyrazolyl group, a 4-pyrazolyl group, a 2-thiazolyl group, a 4~thiazolyl group, a 5-thiazolyl group, a 3-
20 isothiazolyl group, a 4-isothiazolyl group, a 5-
isothiazolyl' group, a 2-oxazolyl group, a 4-oxazolyl group, a 5-oxazolyl group, a 3-isoxazolyl group, a 4-isoxazolyl group, a 5-isoxazolyl group, a 2-pyridyl group, a 3-pyridyl group, a 4-pyridyl group, a 2-
25 pyxazinyl group, a 2-pyrimidinyl group, a 4-pyrimidinyl group, a 5-pyrimidinyl group, a 3-pyridazinyl group, a 4-pyridazinyl group, a 2-1,3,4roxadiazolyl group, a 2-

1,3,4-thiadiazolyl group, a 3-1,2,4-oxadiazolyl group, a 5-1,2,4-oxadiazolyl group, a 3-1,2,4-thiadiazolyl group, a 5-1,2,4-thiadiazolyl group, a 3-1,2,5-oxadiazolyl group, a 3-1,2,5-thiadiazolyl group, a 3-4H-l,2,4-5 triazolyl group, a 3-1H-1,2,4-triazolyl group, a 5-1H-
1,2,4-triazolyl group, a 4-2H-1,2,3-triazolyl group, a 5-2H-l,2,3-triazolyl group, a 4-1H-1,2,3-triazolyl group, a' 5-1H-1,2,3-triazolyl group or the like.
A 8 to 10-membered C5-9 fused heterocyclic group may
10 be a 2-benzofuranyl group, a 3-ben2ofuranyl group, a 4-benzofuranyl group, a 5-benzofuranyl group, a 6-benzofuranyl group, a 7-benzofuranyl group, a 1-isobenzofuranyl group, a 4-isobenzofuranyl group, a 5-isobenzofuranyl group, a 2-benzothienyl group, a 3-
15 benzothienyl group, a 4-benzothienyl group, a 5-benzothienyl group, a 6-benzothienyl group, a 7-benzothienyl group, a l-isobenzothienyl gx-oup, a 4-isobenzothienyl group, a 5-isobenzothienyl group, a 2-chromenyl group, a 3-chromenyl group, a 4-chromenyl
20 group, a 5-chromenyl group, a 6-chromenyl group, a 7-chromenyl group, a 8-chromenyl group, a 1-indolizinyl group, a 2-indolizinyl group, a 3-indolizinyl group, a 5-indolizinyl group, a 6-indolizinyl group, a 7-indolizinyl group, a 8-indolizinyl group, a 1-isoindolyl group, a 2-
25 isoindolyl group, a 4-isoindolyl group, a S-isoindolyl group, a 1-indolyl group, a 2-indolyl group, a 3-indolyl group, a 4-indolyl group, a 5-indolyi group,, a 6-indolyl

group, a 7-indolyl group, a 1-indazolyl group, a 2-indazolyl group, a 3-indazolyl group, a 4-indazolyl group, a 5-indazolyl group, a 6-indazolyl group, a 7-indazolyl group, a 1-purinyl group, a 2-purinyl group, a s 3-purinyl group, a 6-purinyl group, a 7-purinyl group, a 8-purinyl group, a 2-quinolyl group, a 3-quinolyl group, a 4-quinolyl group, a 5-gTiinolyl group, a 6-guinolyl group, a 7-quinolyl group, a 8-quinolyl group, a 1-isoquinolyl group, a 3-isoquinolyl group, a 4-isoquinolyl
10 group, a 5-isoquinolyl group, a 6-isoquinolyl group, a 7-isoquinolyl group, a 8-isoquinolyl group, a 1-phthalazinyl group, a 5~phthalazinyl group, a 6-phthalazinyl group, a 1-2,7-naphthyridinyl group, a 3-2,7-naphthyridinyl group, a 4-2,7-naphthyridinyl group, a
is 1-2,6-naphthyridinyl group, a 3-2,6-naphthyridinyl group, a 4-2,6-naphthyridinyl group, a 2-1,8-naphthyridinyl group, a 3-1,8-naphthyridinyl group, a 4-1,8-naphthyridinyl group, a 2-1,7-naphthyridinyl group, a 3-1,7-naphthyridinyl group, a 4-1,7-naphthyridinyl group, a
20 5-1,7-naphthyridinyl-group, a 6-1,7-naphthyridinyl group, a 8-1,7-naphthyridinyl group, 2-1,6-naphthyridinyl group, a 3-1,6-naphthyridinyl group, a 4-1,6-naphthyridinyl group, a 5-1,6-naphthyridinyl group, a 7-1,6-naphthyridinyl group, a 8-1,6-naphthyridinyl group, a 2-
25 1,5-naphthyridinyl group, a 3-1,5-naphthyridinyl group, a 4-1,5-naphthyridinyl group, a 6-1,5-naphthyridinyl group, - - .a 7-1,5-naphthyridinyl group, a 8-1,5-naphthyridinyl

group, a 2-guinoxalinyl group, a 5-quinoxalinyl group, a 6-quinoxalinyl group, a 2-guinazolinyl group, a 4-quinazolinyl group, a 5-quinazolinyl group, a 6-quinazolinyl group, a 7-quina2olinyl group, a 8-5 guinazolinyl group, a 3-cinnolinyl group, a 4-cinnolinyl group, a 5-cinnolinyl group, a 6-cinnolinyl group, a 7-cinnolinyl group, a 8-cinnolinyl group, a 2-pterdinyl group, a 4-pterdinyl group, a 6-pterdinyl group, a 7-pterdinyl group or the like. 10 A nitrogen-containing heterocyclyl group is a C2.9
heteromonocyclic or fused heterobicyclic group which has one or more nitrogen atoms and may further contain one or more atoms optionally selected from oxygen atoms and sulfur atoms, and:
15
may be mentioned specifically.
Specific preferred examples of the substituent R1 are phenyl groups substituted with one or more of the 20 -following substituents.

Substituents: a C1_6 alkyl group, a C1-3 alkyl group (the C1-3 alkyl group is substituted with one or more halogen atoms) , a C1-3 alkoxy group (the C1-3 alkoxy group is substituted with one or more halogen atoms) and a 5 halogen atom.
Particularly preferred examples of the substituent R1 are a 3,4-dimethyl-phenyl group, a 4-t-butyl-phenyl group, a 4-trifluoromethyl-phenyl group, a 3-chloro-phenyl group, a 4-chloro-phenyl group, a 4-fluoro-phenyl 10 group, a 3,4-dichloro-phenyl group, a 4-bromo-phenyl group and a 4-trifluoromethoxy-phenyl group.
Specific preferred examples of the substituent R2 are a hydrogen atom, a methyl group, an ethyl group, an i-propyl group, a n-propyl group and a trifluoromethyl 15 group.
A particularly preferred example of the substituent R3 is a methyl group.
Specific preferred examples of the substituent R3 are a phenyl group, pyridyl groups (a 2-pyridyl group, a 3-20 pyridyl group and a .4-pyridyl group) and thienyl groups (a 2-thienyl group and a 3-thienyl group) substituted with one or more substituents selected from the following substituent set A and with one or more substituents selected from the following substituent set B. 25 Substituent set A: a hydrogen atom, a nitro group, a halogen atom, a C1-3 alkyl group and a C1-3 alkyl group substituted with one or more fluorine atoms.

Substituent set B: (C=0)Rs (wherein Rs iB NR6R7 (wherein Rs is a hydrogen atom or a Ci-3 alkyl group (the C1-3 alkyl group may be substituted with one or more halogen atoms) , and R7 is a C1-6 alkyl group (the Ci_6 5 alkyl group may be substituted with one or more halogen atoms, one or more hydroxyl groups, one or more C1-3 alkoxy groups or one or more C2-14 aryl groups (the C2-14 aryl groups may be substituted with one or more C1-3 alkyl groups, one or more Ci_3 alkoxy groups, one or more
10 carboxyl groups, one or more carbamoyl groups, one or more cyano groups or one or more halogen atoms, and in the case of aryl groups containing one or more nitrogen atoms, may be N-oxides thereof)), a phenyl group, a thienyl group, a pyridyl group or a pyridyl-N-oxide group
15 (the phenyl group, the thienyl group, the pyridyl group and the pyridyl-N-oxide group may be substituted with one or more halogen atoms), or NR6R7 is, as a whole, a nitrogen-containing heterocyclyl group (the nitrogen-containing heterocyclyl group may be substituted with one
20 or more hydrogen atoms, one or more C1-6 alkyl groups (the C1-6 alkyl groups may be substituted with one or more halogen atoms), one or more halogen atoms, one or more hydroxyl groups or one or more C1-3 alkoxy groups (the C1-3 alkoxy groups may be substituted with one or more halogen
25 atoms))) or a Ci-6 alkyl group (the C1-6 alkyl group may be substituted with one or more halogen atoms, one or more pyridyl groups, one or more pyridyl-N-oxide.groups, one

or more furyl groups, one or more thienyl groups or one or more phenyl groups and is substituted with one or more cyano groups)).
A particularly preferred example of the substituent 5 R3 is represented by the formula (II):

(wherein R6 is a methyl group or an ethyl group, and R7 is a Ci-6 alkyl group (the c1-6 alkyl group may be substituted with one or more methoxy groups)). 10 Another particularly preferred example of the substituent R3 is represented by the formula (II):
(wherein R6 is a methyl group or an ethyl group, and R7 is a C1-3 alkyl group (the C1-3 alkyl group is substituted 15 with one or more phenyl groups or one or more pyridyl groups)).
Still another particularly preferred example of the substituent R3 is represented by the formula (II) :

20 (wherein R6 is a hydrogen atom, and R7 is a Ci_6 alkyl group. (the.Cj.-6 alkyl.. group is substituted with one or

more methoxy groups) or a pyridyl group).
Still another particularly preferred example of the substituent R3 is represented by the formula (IV) :

5 (wherein R6 is a hydrogen atom, R7 is a C1-3 alkyl group (the C1-3 alkyl group may be substituted with one or more hydroxyl groups), and RB is a methyl group or a chlorine atom).
Still another particularly preferred example of the 10 substituent R3 is a group represented by the formula (II) or the formula (IV) wherein NRGR7 is, as a whole, represented by the formula (III) :

(wherein R9 is a C1-3. alkyl group) . 15 Still another particularly preferred example of the substituent R3 is represented by the formula (V) :

" ■ (wherein R10 is a hydrogen atom or a C1-3 alkyl group) . . _

Still another particularly preferred example of the substituent R3 is represented by the formula (II) :

(wherein Rs is a hydrogen atom, and R7 is an isopropyl 5 group, a methyl group, an ethyl group or a normal propyl group (the methyl group, the ethyl group and the normal propyl group are unsubstituted or substituted with one or' more pyridyl groups, one or more pyridyl-N-oxide groups, one or more furyl groups, one or more pyrazinyl groups,
10 one or more imidazolyl groups, one or more pyrazolyl groups or one or more isoxazolyl groups (the pyridyl groups, the pyridyl-N-oxide groups, the furyl groups, the pyrazinyl groups, the imidazolyl groups, the pyrazolyl groups and the isoxasolyl groups may be substituted with
15 one or more methyl groups, one or more methoxy groups, one or more carboxyl groups or one or more halogen atoms))}.
Specific preferred examples of the substituent R4 are a hydrogen atom, a methyl group, an ethyl group, an i-
20 propyl group, a n-propyl group and a trifluoromethyl group.
A particularly preferred example of the substituent R4 is a hydrogen atom.
Favorable compounds for use in the thrombopoietin
25 receptor activator, the preventive, therapeutic or

improving agent for diseases against which activation of the thrombopoietin receptor is effective and the platelet increasing agent of the present invention are as follows.
1) Compounds represented by the formula (I) wherein
5 R2 is a methyl group, and R4 is a hydrogen atom,
tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
2) The compounds according to 1), wherein R1 is a
3,4-diroethyl-phenyl group, a 4-t-butyl-phenyl group, a 4-
10 trifluoromethyl-phenyl group, a 3-chloro-phenyl group, a 4-chloro-phenyl group, a 4-fluoro-phenyl group, a 3,4-dichloro-phenyl group, a 4-bromo-phenyl group or a 4-trifluoromethoxy-phenyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or
15 solvates thereof.
3) The compounds according to 2), wherein R3 is
represented by the formula (II):

(wherein R6 is a methyl group or an ethyl group, and R7 20 is a Ci-e alkyl group (the Ci-6 alkyl group may be
substituted with one or more methoxy groups)), tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
4) The compounds according to 2), wherein R3 is
25 - represented, by the formula (II) :


(wherein Re is a methyl group or an ethyl group, and R7 is a Ci_3 alkyl group (the Ci_3 alkyl group is substituted with or.e or more phenyl groups or one or more pyridyl 5 groups)), tautomers, prodrugs or pharmaceutically
acceptable salts of the compounds or solvates thereof.
5) The compounds according to 2), wherein R3 is represented by the formula (II):

10 (wherein R6 is a hydrogen atom, and R7 is a Ci_6 alkyl group (the Ci-6 alkyl group is substituted with one or more methoxy groups) or a pyridyl group), tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
is 6) The compounds according to 2), wherein NR6R7 in the formula (II) is, as a whole, represented by the formula (III) :

(wherein R9 is a C1-3 alkyl group), tautomers, prodrugs or 20 pharmaceutically acceptable salts of the compounds or

WO 2007/010954

PCT/.TP2006/314317

27 solvates thereof.
7) The compounds according to 2), wherein R3 is
represented by the formula (IV):

5 (wherein R6 is a hydrogen atom, R7 is a Ci_3 alkyl gr (the C1-3 alkyl group may be substituted with one or hydroxyl groups) , and R8 is a methyl group or a chic atom), tautomers, prodrugs or pharmaceutically acce] salts of the compounds or solvates thereof. 10 8) The compounds according to 2), wherein R3 is represented by the formula (V):

(wherein R1D is a hydrogen atom or a Ci-3 alkyl group tautomers, prodrugs.or pharmaceutically acceptable i 15 of the compounds or solvates thereof.
9) The compounds according to 2), wherein R3 is represented by•the formula (II) :

•(wherein R6 is a hydrogen atom, and R7 is an isoprop;

group, a methyl group, an ethyl group or a normal propyl group (the methyl group, the ethyl group and the normal propyl group are unsubstituted or substituted with one or more pyridyl groups, one or more pyridyl-N-oxide groups, one or more furyl groups, one or more pyrazinyl groups, one or more imidazolyl groups, one or more pyrazolyl groups or one or more isoxazolyl groups (the pyridyl groups, the pyridyl-N-oxide groups, the furyl groups, the pyrazinyl groups, the imidazolyl groups, the pyrazolyl groups and the isoxazolyl groups may be substituted with one or more methyl groups, one or more methoxy groups, one or more carboxyl groups or one or more halogen atoms))), tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
10) The compounds according to any one of 3) to 9), wherein R1 is a 3,4-dimethyl-phenyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
11) The compounds according to any one of 3) to 9), wherein R1 is a 3,4-dichloro-phenyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
12) The compounds according to any one of 3) to 9}, wherein R1 is a 4-chloro-phenyl group, tautomers, prodrugs or pharmaceutically acceptable salt3 of the compounds or solvates thereof.
,-- 13) The compounds according to .any one of. 3) to 9),

wherein R1 is a 4-trifluoromethyl-phenyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
14} The compounds according to any one of 3) to 9), 5 wherein R1 is a 4-bromo-phenyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
15) The compounds according to any one of 3) to 9), wherein R1 is a 4-trifluoromethoxy-phenyl group, 10 tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
IS) The compounds wherein R4 is a hydrogen atom, and R1, R3 and R3 are any of the following combinations in Table 1, tautomers, prodrugs or pharmaceutically is acceptable salts of the compounds or solvates thereof. '
The symbols in Table 1 denote the following substituents.

17) The compounds wherein R4 is a hydrogen atom, and R1, R2 and R3 are any.of the following combinations in 5 Table 1, tautomers, prodrugs or pharmaceutically
acceptable salts of the compounds or solvates thereof (provided that in the case of 17), Qla, Qlb, Qlc, Qld, Qle, Qlf, Qlg, Qlh, Q3a, Q3b,"Q3c, Q3d, Q3e, Q3f, Q3g, Q3h, Q3i, Q3j, Q3k and Q31 in the table denote the 10 following substituents).


18) The compounds wherein R4 is a hydrogen atom, and R1, R2 and R3 are any of the following combinations in Table 1, tautomers, prodrugs or pharmaceutically 5 acceptable salts of the compounds or solvates thereof (provided that in the case of 18), Qla, Qlb, Qlc, Qld, Qle, Qlf, Qlg, Qlh, -Q3a, Q3b, Q3c, Q3d, Q3e, Q3f, Q3g, Q3h, Q3i, Q3j, Q3k and Q31 in the table denote the following substituents).


19) The compounds wherein R4 is a hydrogen atom, and R1, Ra and R3 are any of the following combinations-in Table 2, tautomers, prodrugs or pharmaceutically 5 acceptable salts of the compounds or solvates thereof.
The symbols in Table 2 denote the following substituents.

24) The compounds according to 16) to 23), wherein R2
is converted to a hydrogen atom, tautomers, prodrugs or
5 pharmaceutically acceptable salts "of the compounds or solvates thereof.
25) The compounds according to 16) to 23), wherein R2
is converted to a trifluoromethyl group, tautomers,
prodrugs or pharmaceutically acceptable salts of the
lo compounds or solvates thereof.
26) The compounds according to 16) to 23) , wherein R2
is converted to an ethyl group, tautomers, prodrugs or
pharmaceutically acceptable salts of the compounds or
solvates thereof.
15 27) The compounds according to 16) to 23), wherein R2

is converted to a n-propyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
28) The compounds according to IS) to 23), wherein R2
s is converted to an i-propylf tautomerB, prodrugs or
pharmaceutically acceptable salts of the compounds or solvates thereof.
29) The compounds according to 16) to 28), wherein R9
is converted to a methyl group, tautomers, prodrugs or
10 pharmaceutically acceptable salts of the compounds or solvates thereof.
3 0) The compounds according to 16) to 28), wherein R4 is converted to a trifluoromethyl group, tautomers, prodrugs or pharmaceutically acceptable salts of _the
15 compounds or solvates thereof.
31) The compounds according to 16} to 28), wherein R4 is converted to an ethyl group, tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof.
20 32) Thrombopoietin receptor activators containing
the compounds according to 1) to 31), tautomers, prodrugs or pharmaceutically acceptable salts of the compounds or solvates thereof, as an active ingredient.
33) Preventive, therapeutic and improving agents for
25 diseases against which activation of the thrombopoietin receptor is effective, which contain the thrombopoietin receptor activators according to 32), as an active

ingredient.
34) Platelet increasing agents containing the thrombopoietin receptor activators according to 32) , as an active ingredient. 5 35) Medicaments containing any of the compounds according to 1) to 31) or the compounds represented by the formula (I) , tautomers, prodrugs or pharmaceuticaly acceptable salts of the activators or solvates thereof, as an active ingredient.
10 In the present invention, the compounds of the
present invention.represented by the formula (1) may be present in the form of tautomers or geometrical isomers which undergo endocyclic or exocyclic isomerization, mixtures of tautomers or geometric isomers or mixtures of
15 thereof. When the compounds of the present invention
have an asymmetric center, whether or not resulting from an isomerization, the compounds of the present invention may be in the form of resolved optical isomers or in the form of mixtures containing them in certain ratios.
20 The compounds of the present invention represented by the formula (1) or pharmaceutically acceptable salts thereof may be in the form of arbitrary crystals or arbitrary hydrates, depending on the production conditions. The present invention covers these crystals,
25 hydrates and mixtures. They may be in the form of
solvates V7ith organic solvents such as acetone, ethanol and tetrahydrofuran, and the present invention covers any

of these forms.
The compounds of the present invention represented by the formula (1) may be converted to pharmaceutically acceptable salts or may be liberated from the resulting
salts, if necessary. The pharmaceutically acceptable
salts of the present invention may be, for example, salts with alkali metals (such as lithium, sodium and potassium), alkaline earth metals (such as magnesium and calcium), ammonium, organic bases and amino acids. They
10 may be salts with inorganic acids (such as hydrochloric acid, hydrobromic acid, phosphoric acid and sulfuric acid) and organic acids (such as acetic acid, citric acid, maleic acid, fumaric acid, tartaric acid, benzenesulfonic acid and p-toluenesulfonic acid)._
15 The compounds which serve as prodrugs are
derivatives of the present invention having chemically or metabolically degradable groups which give pharmacologically active compounds of the present invention upon solvolysis or under physiological
20 conditions in vivo. .Methods for selecting or producing
-appropriate prodrugs are disclosed, for example, in
Design of Prodrugs (Elsevier, Amsterdam 1985). In the
present invention, when the compound has a hydroxyl
group, acyloxy derivatives obtained by reacting the
25 compound with appropriate acyl halides or appropriate acid anhydrides may, for example, be mentioned as prodrugs. Acyloxys particularly preferred as prodrugs

include -OCOC2H5, -OCO(t-Bu), -OCOC15H31/ -OCO (m-C02Na-Ph) , -OCOCH2CH2C02Na, -OCOCH (NH2) CH3, -OCOCH2N (CH3) a and the like. When the compound of the present invention has an amino group, amide derivatives obtained by reacting the 5 compound having an amino group with appropriate acid halides or appropriate mixed acid anhydrides may, for example, be mentioned as prodrugs. Amides particularly preferred as prodrugs include -NHCO (CH2) 2oOCH3, -NHCOCH(NH2)CH3 and the like. When the compound of the
10 present invention has a carboxyl group, carboxylic acid esters with aliphatic alcohols or carboxylic acid esters obtained by the reaction with an alcoholic free hydroxyl group of 1,2- or 1,3-digylcerides may, for example, be mentioned as prodrugs. Particularly preferred prodrugs
15 are methyl esters and ethyl esters.
The preventive, therapeutic and improving agents for diseases against which activation of the thrombopoietin receptor is effective or platelet increasing agents which contain the thrombopoietin receptor activators of the
20 present invention as an active ingredient may usually be administered as oral medicines such as tablets, capsules, powder, granules, pills and syrup, as rectal medicines, percutaneous medicines or injections. The agents of the present invention may be administered as a single
25 therapeutic agent or as a mixture with other therapeutic agents. Though they may be administered as they are, they are usually administered in the form of medical

compositions. These pharmaceutical preparations can be obtained by adding pharmacologically and pharmaceutically acceptable additives by conventional methods. Namely, for oral medicines, ordinary excipients, lubricants, s binders, disintegrants, humectants, plasticizers and
coating agents may be used. Oral liquid preparations may be in the form of aqueous or oily suspensions, solutions, emulsions, syrups or elixirs or may be supplied as dry syrups to be mixed with water or other appropriate 10 solvents before use. Such liquid preparations may
contain ordinary additives such as suspending agents, perfumes, diluents and emulsifiera. In the case of rectal administration, they may be administered as suppositories. Suppositories may use an appropriate 15 substance such as cacao butter, laurin tallow, Macrogol, "glycerogelatin, Witepsol, sodium stearate and mixtures thereof as the base and may, if necessary, contain an emulsifier, a suspending agent, a preservative and the like. For injections, pharmaceutical ingredients such as ao distilled water for injection, physiological saline, 5% glucose solution, propylene glycol and other solvents or solubilizing agents, a pH regulator, an isotonizing agent and a stabilizer may be used to form aqueous dosage forms or dosage forms which need dissolution before use. 25 The dose of the agents of the present invention for administration to human is usually about from 0.1 to 1000 mg/human/day in the case of oral drugs or rectal

administration for adults and about from 0.05 mg to 500 mg/human/day in the case of injections for adults, though it depends on the age and conditions of the patient. The above-mentioned ranges are mere examples, and the dose 5 should be determined from the conditions of the patient. The present invention is used when the use of compounds which have thrombopoietin receptor affinity, and act as thrombopoietin receptor agonists are expected to improve pathological conditions. For example,
10 hematological disorders accompanied by abnormal platelet count may be mentioned. Specifically, it is effective for therapy or prevention of human and mammalian diseases caused by abnormal megakaryopoiesis, especially those accompanied by thrombocytopenia. Examples of such
is diseases include thrombocytopenia accompanying
chemotherapy or radiotherapy of cancer, thrombocytopenia accompanying antiviral therapy for diseases such as hepatitis C, thrombocytopenia caused by bone marrow transplantation, surgery and serious infections, or
20 gastrointestinal bleeding, but such diseases are not
restricted to those mentioned. Typical thrombocytopenias such as aplastic anemia, idiopathic thrombocytopenic purpura, myelodysplastic syndrome, hepatic disease, HIV infection and thrombopoietin deficiency are also targets
25 of the agents of the present invention. The present invention may be used as a peripheral stem cell mobilizer, a megakaryoblastic or megakaryocyte leukemia

cell differentiation inducer and a platelet increasing agent for platelet donors. In addition, potential applications include therapeutic angiogenesis based on differentiation and proliferation of vascular endothelial 5 cells and endothelial progenitor cells, prevention and therapy of arteriosclerosis, myocardial infarction, unstable angina, peripheral artery occlusive disease, but there is no restriction.
The compounds represented by the formula (I) are 10 prepared, for example, by the process represented by the formula (l) illustrated below.

The reaction of the compound (IV) with a -NH2 15 compound (VII) in a solvent, if necessary in the presence of a catalyst, under heating with stirring gives a desired compound or its precursor. The precursor may be, if necessary, hydrolyzed, deprotected, reduced or oxidized to a desired compound. The compounds of the 20 present invention usually can be purified by column chromatography, thin layer chromatography, high performance liquid chromatography (HPLC) or high performance .liquid chromatography-mass spectrometry (LC-

MS) and, if necessary, they may be obtained with high purity by recrystallization or washing with solvents. For the syntheses of the intermediates (VI), the following method disclosed in JP-A-48-026755 may, for 5 example, be referred to.

For synthesis of the -NH2 compounds (VTI) , for example, the methods disclosed in Synthetic Commun., 10 28(7), 1223-1231 (1998), J. Chem. Soc., 1225 (1948) and J. Chera. Soc, 2831 (1952) may be referred to.
The compounds represented by the formula (I) can also be obtained by the process represented by the formula (3) illustrated below.
15
The reaction of the compound (VIII) with the compound (IX) in a solvent, if necessary, in the presence of a catalyst, a dehydrating condensation agent or a -■ 20 base, under heating with stirring gives a desired

compound or its precursor. The precursor may be, if necessary, hydrolyzed, deprotected, reduced or oxidized to a desired compound. The compounds of the present invention usually can be purified by column 5 chromatography, thin layer chromatography, high performance liquid chromatography (HPLC) or high performance liquid chromatography-mass spectrometry (LC-MS) and, if necessary, they may be obtained with high purity by recrystallization or washing with solvents. 10 - The compound (VIII) can be obtained by stirring the compound (VI) with hydrazine or its derivative in a solvent, if necessary in the presence of a catalyst, under heating.
15 EXAMPLES
Now, the present invention will be described in
further detail with reference to Reference Synthetic
Examples, Synthetic Examples, Assay Examples and
Formulation Examples. However, it should be understood 20 that the present invention is by no means restricted by
these specific Examples.
The Hl-NMR analysis was carried out at 300 MHz, and
LC/MS was measured under the following conditions.
LC/MS condition 1 25 Column: Waters SunFire CIS (3.5 urn, 4.6x30 mm)
Eluent: acetonitrile/0.1% aqueous formic acid (10/90 —»
30/70)

LC/MS condition 2
Column: Waters SunFire C18 {3.5 urn, 4.6x30 mm) Eluent: acetonitrile/O.1% aqueous formic acid (10/90 -» 60/40) 5 LC/MS conditions 3
Column: Waters SunFire CIS (3.5. pm, 4.6x30 mm) Eluent: acetonitrile/0.1% aqueous formic acid (10/90 85/15)
LC/MS conditions 4 10 Column: Waters Xterra MSC18 (5 um, 4.6x50 mm)
Eluent: acetonitrile/0.1% aqueous formic acid (10/90
30/70)
LC/MS conditions 5
Column: Waters Xterra MSC18 (5 um, 4.6x50 mm) 15 Eluent: acetonitrile/0.1% aqueous formic acid (10/90 —
60/40)
LC/MS conditions 6
Column: Waters Xterra'MSC18- (5 pm, 4.6x50 mm)
Eluent: acetonitrile/0.1% aqueous formic acid (10/90 -* 20 85/15)
LC/MS conditions 7
Column: Waters Xterra MSC18 (5 μm, 4.6x50 mm)
Eluent: acetonitrile/0.1% aqueous formic acid (20/80 —>
100/0) 25 LC/MS conditions 8
Column: Waters Xterra MSC18 (3.5 pm, 2.1x20 mm)
Eluent: acetonitrile/0.2% aqueous formic acid (20/80 -*

90/10)
REFERENCE SYNTHETIC EXAMPLE ,1 Synthesis of 5-(4-isopropylpiperazine-l-carbonyl)thiophene~2-carbohydrazide s A solution of 59 mg (0.02 mmol) of methyl 5-(4-i sopropylpiperaz ine-1-carbony1}thiophene- 2 -carboxylate was dissolved in 2 mL of ethanol was heated with 100 uL
of hydrazine monohydrate at 80°C for 5 hours with reflux.
After cooling, the reaction solution was poured into a 10 liquid mixture of 5 mL of water and 5 mL of saturated
aqueous sodium chloride and extracted with 20 mL of ethyl
acetate and 20 mL of chloroform.
The extract was dried over magnesium sulfate, and
the solvent was evaporated at 40°C to give 30 mg of the 15 desired product, 5-(4-isopropylpiperazine-l-
carbonyl)thiophene-2-carbohydrazide (yield 51%).
Morphology: colorless solid
LC/MS: conditions 4 retention time 0.32 (min)
LC/MS (ESI+) m/z; 297 [M+l] + 20 LC/MS (ESI") m/z; 295 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 2
Synthesis of 5-(morpholine-4-carbonyl)thiophene-2-
c a rbohydra z i de
The procedure in Reference Synthetic Example 1 was 25 followed using methyl 5-(morpholine-4-carbonyl)thiophene-
2-carboxylate to give the desired product, 5-(morpholine-
4-carbonyl)thiophene-2-carbohydrazide (yield 51%).

Morphology: colorless solid
LC/MS: conditions 5 retention time 0.34 (min)
LC/MS (ESI+) m/z; 256 [M+l]+
LC/MS (ESI") m/z; 254 [M-l]~ 5 REFERENCE SYNTHETIC EXAMPLE 3
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid, diethylamide
The procedure in Reference Synthetic Example 1 was
followed using methyl 5-(diethylcarbamoyl)thiophene-2-10 carboxylate to give the desired product, 5-
hydrazinocarbonylthiophene-2-carboxylic acid diethylamide
(yield 89%).
Morphology: white solid
LC/MS: conditions 8 retention time 0.63 (min) 15 LC/MS (ESI+) m/z; 242 [M+l]+
REFERENCE SYNTHETIC EXAMPLE 4
Synthesis of 5-(pyrrolidine-1-carbonyl)thiophene-2-
carbohydrazide
The procedure in Reference Synthetic Example 1 was 20 followed using methyl 5-(pyrrolidine-1-
carbonyl)thiophene-2-carboxylate to give the desired
product, 5-(pyrrolidine-1-carbonyl)thiophene-2 -
carbohydrazide.
Morphology: white solid 25 LC/MS: conditions 8 retention time 0.50 (min)
LC/MS (ESI+) m/z; 240 [M+l]+
REFERENCE SYNTHETIC- EXAMPLE 5

Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid dimethylamide
The procedure in Reference Synthetic Example 1 was
followed using methyl 5-(dimethylcarbamoyl)thiophene-2-5 carboxylate to give the desired product, 5-
hydrazinocarbonylthiophene-2-carboxylic acid
dimethylamide (yield 23%).
Morphology: colorless solid
LC/MS: conditions 6 retention time 0.37 (min) 10 LC/MS (ESI+) m/z; 214 [M+l]+
LC/MS (ESI") m/z; 212 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 6
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid 2-methoxyethylamide 15 The procedure in Reference Synthetic Example 1- was
followed using methyl 5-(2-
methoxyethylcarbamoyl)thiophene-2-carboxylate to give the
desired product, 5-hydrazinocarbonylthiophene-2-
carboxylic acid 2-methoxyethylamide (yield 84%). 20 Morphology: white solid
LC/MS: conditions 1 retention time 0.34 (min)
LC/MS (ESI+) m/z; 244 [M+l]*
LC/MS (ESI") m/z; 242 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 7 25 Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid 3-pyridylamide
The procedure in Reference Synthetic Example 1 was

followed using methyl 5-(3-pyridylcarbamoyl)thiophene-2-
carboxylate to give the desired product, 5-
hydxazinocarbonylthiophene-2-carboxylic acid 3-
pyridylaraide (yield 78%) . 5 Morphology: colorless solid
LC/MS: conditions 1 retention time 0.34 (min)
LC/MS (ESI+) m/z; 263 [M+l]+
LC/MS (ESI") m/z; 261 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 8 io Synthesis of 2-chloro-4-hydrazinocarbonyl-N-(2-
hydroxyethy1)benzamide
The procedure in Reference Synthetic Example i was
followed using methyl 3-chloro-N-(2-
hydroxyethyl)terephthalic acid methyl ester to give the is desired product, 2-chloro-4-hydrazinocarbonyl~N-(2-
hydroxyethyl)benzamide (yield 66%).
Morphology: colorless solid
LC/MS: conditions 2 retention time 0.32 (min)
LC/MS (ESI+) m/z; 258, 260 [M+l]+ 20 LC/MS (ESI*) m/z; 256, 258 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 9
Synthesis of 5-(2-cyanobutyryl)thiophene-2-carbohydrazide
Methyl 5-(2-cyanobutyryl)thiophene-2-carboxylate To butyronitrile (957 pL,' 11 mmol) in 25 tetrahydrofuran, lithium hexamethyldisilazide (12.5 mL 1M
tetrahydrofuran solution, 12.5 mmol) was added at -7B°C, - and the resulting solution was stirred for 1 hour and

added dropwise to 5-methoxycarbonylthiophene-2-carbonyl
chloride (1.02 g, 5 mmol) in tetrahydrofuran at -78°C over 30 minutes, and the resulting reaction mixture was stirred at room temperature for 1 hour. The solvent was 5 evaporated, and the reaction solution was mixed with
ethyl acetate and washed with saturated aqueous ammonium chloride and saturated sodium chloride and purified by silica gel column chromatography (eluent hexane/ethyl acetate = 3/1) to give the desired product, methyl 5-{2-10 cyanobutyryl)thiophene-2-carboxylate (yield 41%). Morphology: yellow solid
LC/MS: conditions 3 retention time 2.45 (min) LC/MS (ESI+) m/z; 238 [M+l]+ LC/MS (ESI") m/z; 236 [M-l]" 15 5-(2-Cyanobutyryl)thiophene-2-carbohydrazide
Methyl 5-(2-cyanobutyryl)thiophene-2-carboxylate (213 mg, 0.90 mmol) in methanol was stirred with 0.1 M potassium hydroxide in methanol (9.0 mL, 0.90 mmol) at room temperature for 10 minutes and then with hydrazine 20 monohydrate (225 mg,.4.50 rag) at BO'C for 6 hours. After addition of saturated aqueous sodium chloride, the reaction solution was extracted with ethyl acetate, dried over anhydrous magnesium sulfate and concentrated to give the crude desired product. 25 Morphology: yellow solid
REFERENCE SYNTHETIC EXAMPLE 10
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic

acid methyl-2-picolylamide
The procedure in Reference Synthetic Example 1 was
followed using methyl 5-(methyl-2-picolylamido)thiophene-
2-carboxylate to give the desired product, 5-5 hydrazinocarbonylthiophene-2-carboxylic acid methyl-2-
picolylamide {yield 77%).
Morphology: white solid
LC/MS: conditions 8 retention time 0.45 (min)
LC/MS (ESI+) m/z; 291 [M+l]+ 10 LC/MS (ESI") m/z; 289 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 11
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid 3-picolylamide
Methyl 5-(3-picolylcarbamoyl)thiophene-2-carboxylate is {860 mg, 3.11 mmol) in ethanol {34 mL) was stirred with
hydrazine monohydrate (1.57 m, 31.1 mmol) at 85°C for 12
hours. The reaction solution was concentrated and
stirred with diethyl ether at 0°C for 1 hour. The
precipitated solid was recovered by filtration and washed 20 with a liquid mixture of diethyl ether and ethanol and
dried to give the desired product, 5-
hydrazinocarbonylthiophene-2-carboxylic acid 3-
picolylamide (yield 92%).
Morphology: white solid 25 LC/MS: - conditions 1 retention time 0.23 (min)
LC/MS (ESI+) m/z; 277 [M+l]4"
LC/MS {ESI*) m/z; 275 [M-l]"

REFERENCE SYNTHETIC EXAMPLE 12
Synthesis of 5-hydra2inocarbonylthiophene-2-carboxylic
acid 4-picolylamide
The procedure in Reference Synthetic Example 11 was 5 followed using methyl 5-(4-picolylcarbamoyl)thiophene-2-
carboxylate to give the desired product, 5-
hydrazinocarbonylthiophene-2-carboxylic acid 4-
picolylamide (yield 81%).
Morphology: white solid 10 LC/MS: conditions 1 retention time 0.23 (rain),
LC/MS (ESI+) m/z; 277 [M+l}+
LC/MS (ESI") m/z; 275 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 13
Synthesis of 5-hydrazinocarbonylthiophene-2-carbqxylic 15 acid (furan-2-ylmethyl)amide
The procedure in Reference Synthetic Example 11 was
followed using methyl 5-(furan-2-
ylmethylcarbamoyl)thiophene-2-carboxylate to give the
desired product", 5-hydrazinocarbonylthiophene-2-20 carboxylic acid (furan-2-ylmethyl)amide (yield 86%).
Morphology: white solid
LC/MS: conditions 3 retention time 1.50 (min)
LC/MS (ESI4) m/z; 26S [M+l]+
LC/MS (ESI") m/z; 264 [M-l]" 25 REFERENCE SYNTHETIC EXAMPLE 14
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid methylamide

The procedure in Reference Synthetic Example 11 was
followed using methyl 5-(methylcarbamoyl)thiophene-2-
carboxylate to give the desired product, 5-
hydrazinocarb'onylthiophene-2-carboxylic acid methylamide 5 (yield 83%).
Morphology: pale yellow solid
LC/MS: conditions 3 retention time 0,37 (min)
LC/MS (ESI"*") m/z; 200 [M+l}4'
LC/MS (ESI") m/z; 198 [M-l]" 10 REFERENCE SYNTHETIC EXAMPLE 15
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid isopropylamide
The procedure in Reference Synthetic Example 11 was
followed using methyl 5-(isopropylcarbamoyl)thiophene-2-15 carboxylate to give the desired product, 5-
hydrazinocarbonylthiophene-2-carboxylic acid
isopropylamide (yield 45%).
Morphology: ignite solid
LC/MS: conditions 2 retention time 1.07 (min) 20 LC/MS (ESI+) m/z; 228 [M+lK
LC/MS (ESI") m/z; 226 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 16
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid 2-picolylamide 25 The procedure in Reference Synthetic Example 11 was
followed using methyl 5-(2-picolylcarbamoyl)thiophene-2-
carboxylate to give the desired product, 5-

hydrazinocarbonylthiophene-2-carboxylic acid 2~ picolylamide (yield 81%). Morphology: white solid
IiC/MS: conditions 1 retention time 0.28 (min) S LC/MS (ESI+) m/z; 277 [M+l]+ LC/MS (ESI") m/z; 275 [M-l]" REFERENCE SYNTHETIC EXAMPLE 17
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic acid (2-pyridin-4-yl)ethylamide 10 Methyl 5-[2-(pyridin-4-yl)ethylcarbamoyl]thiophene-2-carboxylate (0.40 g, 1.4 mmol) was suspended in a liquid mixture of methanol (4.0 mL) and tetrahydrofuran
(2.0 mL) and left at 55°C until conversion to a homogeneous amber solution was confirmed. After addition
is of 80% hydrazine monohydrate (0.17 mL, 2.8 mmol), 'it was left at 55°C for 24 hours. After addition of 80% hydrazine monohydrate (0.17 mL, 2.8 mmol), it was left at 55°C for 4.5 hours and then at room temperature for 14 hours. The precipitated solid was recovered by
20 filtration and dried to give the desired product, 5-
hydrazinocarbonylthiophene-2-carboxylic acid (2-pyridin-4-yl)ethylamide (yield 82%). Morphology: white solid
^-NMR (DMSO-dfi) 6: 2.86 (t, J=7.0 Hz, 2H) , 3.50 (dt,
25 J=5.5 & 7.0 Hz, 2H), 4.52 (br s, 2H), 7.26 (d, J=6.0 Hz, 2H), 8.46 (d, J=6.0 Hz, 2H), 8.72 (t, J=5.5 Hz, 1H), 9.90 (br s, 1H).

REFERENCE SYNTHETIC EXAMPLE 18
Synthesis of 5-(hydrazinecarbonyl)-N-[(l-methyl-lH-pyrazol-5-yl)methyl] thiophene-2-carboxamide
Methyl 5-(l-methyl-lH-pyrazol-5-
5 ylmethylcarbamoyl)thiophene-2-carboxylate (0.25 g, 0.90
mmol) in methanol (2.5 mL) was stirred with hydrazine
monohydrate (0.17 mL, 3.6 mraol) at 70°C for 3.5 hours.
The precipitated solid was recovered by filtration,
washed with chloroform and dried to give the desired
10 product, 5-(hydrazinecarbonyl)-N-[{l-methyl-lH-pyrazol-5-yl)methyl]thiophene-2-carboxamide (yield 49%).
Morphology: white solid
LC/MS: conditions 2 retention time 0.40 (min)
LC/MS (ESI*) ra/z; 280 [M+l]* IS LC/MS (ESI") m/z; 278 [M-l] *
REFERENCE SYNTHETIC EXAMPLE 19
Synthesis of 5-hydrazinocarboynlthiophene-2-carboxylic
acid (5-methylisoxazol-3-ylmethyl)amide
The procedure in Reference Synthetic Example 18 was 20 followed using methyl 5-(5-methylisoxazol-3-
ylmethylcarbamoyl)thiophene-2-carbo3cylate to give the
desired product, 5-hydrazinocarboynlthiophene-2-
carboxylic acid (5-methylisoxazol-3-rylmethyl) amide (yield
47%) . as Morphology: white solid
LC/MS: conditions 2 retention time 1.00 (min)
LC/MS (ESt*) m/z; 281 [M+l]*

LC/MS (ESI") m/z; 279 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 20
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid {5-methylpyrazin-2-ylmethyl)amide s Methyl 5-(5-methylpyrazin-2-
ylmethylcarbamoyl)thiophene-2-carboxylate {304 mg, 1.04
mmol) in methanol (3 mL) was stirred with hydrazine
monohydrate at 60°C for 12 hours. After addition of
chloroform, it was stirred at room temperature for 5 10 hours. The precipitated solid was recovered by
filtration to give the desired product, 5-
hydrazinocarbony1thiophene-2-carboxylic acid (5-
methylpyrazin-2-ylmethyl)amide {yield 72%).
Morphology: white solid 15 LC/MS: conditions 3 retention time 0.69 (min)
LC/MS (ESI+).m/z; 292 [M+lT
LC/MS (ESI") m/z; 290 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 21
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic 20 acid (isoxazol-5-ylmethyl)amide
The procedure in Reference Synthetic Example 18 was
followed using methyl 5-(isoxazol-5-
ylmethylcarbamoyl)thiophene-2-carboxylate to give the
desired product, 5-hydrazinocarbonylthiophene-2-25 carboxylic acid (isoxazol-5-ylmethyl)amide (yield 46%).
Morphology: white solid
LC/MS: conditions 2 retention time 0.62 (min)

LC/MS (ESI+) m/z; 267 [M+l] +
LC/MS (ESI") m/z,- 265 [M-l]"
REFERENCE SYNTHETIC EXAMPLE 22
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic 5 acid (3-methoxyisoxazol-5-ylmethyl) amide
The procedure in Reference Synthetic Example 18 was
followed using methyl 5-(3-methoxyisoxazol-5-
ylmethylcarbamoyl)thiophene-2-carboxylate to give the
desired product, 5-hydrazinocarbonylthiophene-2-10 carboxylic acid (3-methoxyisoxazol-5-ylmethyl)amide (yield 50%) .
Morphology: colorless solid
LC/MS: conditions 2 retention time 1.25 (min)
LC/MS (ESI+) m/z; 297 [M+l]* 15 LC/MS (ESI") m/z; 295 [M-'l] "
REFERENCE SYNTHETIC EXAMPLE 23
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic
acid (1,5-dimethyl-lH-pyrazol-3-ylmethyl)amide
The procedure in Reference Synthetic Example 18 was 20 followed using methyl 5-(1,5-dimethyl-lH-pyrazol-3-
ylmethylcarbamoyl)thiophene-2-carboxylate to give the
desired product, 5-hydrazinocarbonylthiophene-2-
carboxylic acid (1,5-dimethyl-lH-pyrazol-3-ylmethyl)amide
(yield 59%). 25 Morphology: white solid
LC/MS: conditions 2 retention time 1.00 (min)
LC/MS (ESI+) m/z; 294 [M+l]+

LC/MS {ESI") m/z; 292 [M-l]" REFERENCE SYNTHETIC EXAMPLE 24
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic acid (pyrazih-2-ylTTiethyl) amide 5 The procedure in Reference Synthetic Example 18 was followed using methyl S-(pyrazin-2-
ylmethylcarbamoyl)thiophene-2-carboxylate to give the desired product, 5-hydrazinocarbonylthiophene-2-carboxylic acid (pyrazin-2-ylmethyl)amide (yield 82%). 10 Morphology: white solid
LC/MS: conditions 2 retention time 0.37 (min) LC/MS (ESI*) m/z; 278 [M+l]+ LC/MS (ESI') m/z; 276 [M-l]" REFERENCE SYNTHETIC EXAMPLE 25 15 a) Synthesis of 4-[{5-(methoxycarbonyl)thiophene-2-carboxamido}methyl]pyridine 1-oxide
5-Hydrazinocarbonylthiophene-2-carboxylic acid 4-picolylamide (0.20 g, 0.72 mmol) prepared in Reference Synthetic Example 12 in chloroform (4.0 mL) was stirred 20 with 65 wt% m-chloroperbenzoic acid (0.21 g, 0.80 mmol) at room temperature for 9 hours, then left for 18 hours and concentrated to dryness under reduced pressure. Chloroform (30 mL), saturated aqueous sodium hydrogencarbonate (3 mL) and"water (7 mL) i-jere added to 25 the residue, and the organic layer was separated. The aqueous layer was extracted with chloroform (10 mL x 2) and .hot chloroform (10 mL x 1) . The resulting organic

layer was concentrated to give the desired crude product,
4-[{5-(methoxycarbonyl)thiophene-2-
carboxamidojmethyl} pyridine 1-oxide (purity 80 wt%, yield
62%) . 5 Morphology: white solid
LC/MS: conditions 2 retention time 1.77 (min)
LC/MS (ESI+) m/z; 293 [M+l]* .
LC/MS (ESI") m/z; 291 [M-l]"
b) Synthesis of 4-[{5-hydrazniocarbonyl)thiophene-2-10 carboxamidojmethyl]pyridine 1-oxide
4-[{5-(Methoxycarbonyl)thiophene-2-
carboxamidojmethyl]pyridine 1-oxide (0.12 g, 0.34 mmol)
prepared above suspended in methanol (2.0 mL) was left at
60°C. After addition of 80% hydrazine monohydrat.e 15 (0.082 mL, 1.4 mmol), it was left at 60°C for 3.5 hours,
then at room temperature for 5.5 hours and at 0°C for
11.5 hours. The precipitated solid was recovered by
filtration and dried to give the desired product, 4-[{5-
hydrazniocarbonyl) thiophene-2-carboxamido}methyl] pyridine 20 1-oxide (yield 51%).
Morphology: pale yellow solid
LC/MS: conditions 2 retention time 0.37 (min)
LC/MS (ESI+) m/z; 293 [M+l]+
LC/MS (ESI") m/z; 291 [M-l]" 25 ^-NMR (DMSO-de) 5: 4.42 (br s, 2H) , 4.51 (br s,
0.7H), 7.32 (d, J=7.0 Hz, 2H), 7.68 (d, J=4.0Hz, 1H),
7.75(d, J=7.0 Hz, 1H), 8.17 (d, J=7.0 Hz, 2H), 9.25 (t, - -

J=6.Q Hz, Q.3H), 9.92 (br B, 0.3H). REFERENCE SYNTHETIC EXAMPLE 26
Synthesis of 5-hydrazinocarbonylthiophene-2-carboxylic acid 3-(pyridin-4-yl)propylamide 5 80% Hydrazine monohydrate (0.17 mL, 2.8 mmol) was added to methyl 5-[3-(pyxidin-4-
yl)propylcarbamoyl]thiophene-2-carboxylate (0.28 g, 0.92 mmol) in methanol (10 mL), and the reaction solution was left at 50°C for 95 hours. After addition of 80% 10 hydrazine monohydrate (0.17 mL, 2.8 mmol), it was left at
55°C for another 14 hours and concentrated to dryness by evaporating the solvent under reduced pressure. Methanol (2 mL) was added to the residue, and the resulting solution was put on a eonicator. The precipitated solid
15 was recovered by filtration and dried to give the desired product, 5-hydrazinocarbonylthiophene-2-carboxylic acid 3- (pyridin-4-yl)propylamide (yield 61%). Morphology: white solid
^H-NMR (DMSO-dfi) 5: 1.84 (tt, J=7.5 & 6.5 Hzf 2H) ,
20 2.64 (t, J=7.5 Hz, 2H), 3.25 (dt, J=5.5 & 6.5 Hz, 2H) ,
4.54 (br s, 1.6H), 7.26 (d, J=6.0 Hz, 2H), 7.65 (d, J=4.0
Hz, 1H), 7.67 (d, J-4.0 Hz, 1H), 8.45 (d, J=6.0 H2, 2H),
8.65 (br t, J=5.5 Hz, 0.9H), 9.90 (br s, 0.9H).
REFERENCE SYNTHETIC EXAMPLE 27 25 Synthesis of 5-(hydrazinecarbonyl)-N-[(l-methyl-lH-imidazol-5-y1)methyl)thiophene-2-carboxamide
The procedure in Reference Synthetic Example 18 was

followed using methyl 5-(l-raethyl-lH-imidazol-5-ylmethylcarbamoyl) thiophene-2-carboxylate to give the
desired produce, 5-(hydrazinecarbonyl)-N-[(1-methyl-1H-imidazol-5-yl)methyl]thiophene-2-carboxamide {yield 70%).
Morphology: pale yellow solid
LC/MS: conditions 3 retention time 0.30 (rain)
LC/MS (ESI*) m/z; 280 [M+l]+
LC/MS (ESI") m/z; 278 [M-l]" 10 REFERENCE SYNTHETIC EXAMPLE 28
Synthesis of 5-(3-hydroxypyrrolidine-l-
carbonyl)thiophene-2-carbohydrazide
The procedure in Reference Synthetic Example 18 was
followed using methyl 5-(3-hydroxypyrrolidine-l-15 carbonyl) thiophene-2-carboxylate to give the desired
product, 5- (3-hydroxypyrrolidine-l-carbonyl) thiophene-2-
carbohydrazide (yield 53%).
Morphology: white solid
SYNTHETIC EXAMPLE 1 20 Synthesis of 5-(4-isopropylpiperazine-l-
carbonyl) thiophene-2-carboxylic acid {l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazide
5- (4-Isopropylpiperazine-l-carbonyl) thiophene-2-25 carboxylic acid {l-[5-(3,4-dichlorophenyl)-4-
hydroxythiophen-3-yl] ethylidenejhydrazide hydrochloride 28 mg of 2- (3, 4-dichlorophenyl) -3-hydroxy-4-

methylcarbonylthiophene (synthesized in accordance with WO2004/108683) and 29.7 mg of 5-(4-isopropylpiperazine-l-carbonyl)thiophene-2-carbohydrazide prepared in Reference Synthetic Example 1 in 2 mL of isopropyl alcohol were 5 heated with 3 mg of p-tosylic acid monohydrate and 25 uL
(1 eq) of 4 M hydrogen chloride/dioxane at 105°C for 8 hours. The reaction solution was further heated with 2 mL of dimethylformamide at 105°C for 5 hours and cooled to room temperature. The precipitated solid was
10 recovered by filtration and washed with 1 mL of isopropyl alcohol and 1 mL of chloroform, and the resulting crystals were dried to give the desired product (yield 54%) . Morphology: colorless solid
15 LC/MS: conditions 5 retention time 3.80 (min) LC/MS (ESI+) m/z; 565, 567 [M+l]+ LC/MS (ESI") m/z; 563, 565 [M-l]" 5-(4-Isopropylpiperazine-l-carbonyl)thiophene-2-carboxylic acid {l- [5-{3,4-dichlorophenyl)-4-
20 hydroxythiophen-3-yl]ethylidenejhydrazide
5-(4-Isopropylpiperazine-l-carbonyl)thiophene-2-carboxylic acid {l- [5-(3,4-dichlorophenyl}-4-hydroxythiophen-3-yl]ethylidenejhydrazide hydrochloride (14 mg, 0.025 mmol) was suspended in methanol (2.7 mL),
25 and 0.1 M potassium hydroxide in methanol (0.24 mL) and methanol (5.4 mL) were added. The suspension was heated at 50°C and concentrated to dryness under reduced

pressure to give the desired product (yield 100%).
Morphology: light brown solid
SYNTHETIC EXAMPLE 2
Synthesis of "5-(morpholine-4-carbonyl)thiophene-2-5 carboxylic acid {l-[5-(3,4-dichlorophenyl)-4-
hydroxythiophen-3-yl]ethylidenejhydrazide potassium salt
5-(Morpholine-4-carbonyl)thiophene-2-carboxylic acid {l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazide 10 28 mg of 2-(3,4-dichlorophenyl)-3-hydroxy-4-
methylcarbonylthiophene and 26 mg of 5-(morpholine-4-
carbonyl)thiophene-2-carbohydrazide prepared in Reference
Synthetic Example 2 were heated in isopropyl alcohol with
3 mg of p-tosylic acid monohydrate at 105°C for 18 hours is and cooled to room temperature. The precipitated solid
was recovered by filtration and washed with 1 mL of
isopropyl alcohol, and .the resulting crystals were dried
to give the desired product (yield 86%).
Morphology: pale yellow solid 20 LC/MS: conditions' 5.retention time 4.89 (min)
LC/MS (ESI*) m/z; 524, 526 [M+l]+
LC/MS (ESI") m/z; 522, 524 [M-l]~
5-(Morpholine-4-carbonyl)thiophene-2-carboxylic acid {l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-25 yl]ethylidenejhydrazide potassium salt
5-(Morpholine-4-carbonyl)thiophene-2-carboxylic acid
{l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-

yl]ethylidene}hydrazide (20 mg, 0.038 nnnol) was suspended in methanol (2.4 raL), and 0.1 M potassium hydroxide in methanol (0.38 mL) and then methanol (5.S mL) were added.
The suspension was heated at 50°C and concentrated to s dryness under reduced pressure to give the desired product (yield 100%).
SYNTHETIC EXAMPLE 3
Synthesis of 5-[{2-{l-[5-(3,4-dichlorophenyl)-4-
10
hydroxythiophen-3-yl]ethylidenejhydrazino)carbonyl] thiophene-2-carboxylic acid diethylamide potassium salt 5-[(2-{l-[5-{3,4-Dichlorophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid diethylamide
is Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 2-(3,4-dichlorophenyl)-3-
hydroxy-4-methylcarbonylthiophene and 5-
hydrazinocarbonylthiophene-2-carboxylic acid diethylamide
prepared in Reference Synthetic Example 3 (yield 76%). 20 Morphology: pale yellow solid
LC/MS: conditions 5 retention time 5.82 (min)
LC/MS (ESI*) m/z; 510, 512 [M+l]*
LC/MS (ESI") m/z; 508, 510 [M-l]"
5-[(2-{l-[5-(3,4-Dichlorophenyl)-4-hydroxythiophen-3-25 yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid diethylamide potassium salt
Synthesis was carried out in the same manner as in

Synthetic Example 2 by using 5-[(2-{l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazino)carbonyl] thiophene-2-car'boxylic acid diethylamide (yield 100%).
5 Morphology: orange solid
SYNTHETIC EXAMPLE 4
Synthesis of 5-{pyirolidine-l-carbonyl)thiophene-2-
carboxylic acid {l-[5-(3,4-dichlorophenyl)-4-
hydroxythiophen-3-yl]ethylidene}hydrazide potassium salt 10 S- (Pyrrolidine-1-carbonyl) thiophene-2-carboxylic acid {l-[5- (3,4-dichlorophenyl) -4-hydroxythiophen-3-
yl]ethylidenejhydrazide
* Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 2-<3,4-dichlorophenyl) -3-15 hydroxy-4-methylcarbonyl)thiophene and 5- (pyrrolidine-1-
carbonyl) thiophene-2-carbohydrazide prepared in Reference
Synthetic Example 4 (yield 94%).
Morphology: . pale yellow solid
LC/MS: conditions 5 retention time 5.34 (min) 20 LC/MS (ESI4) m/z; 508, 510 [M+l]*
LC/MS (ESI") m/z; 506, 508 [M-l]'
5-(Pyrrolidine-l-carbonyl)thiophene-2-carboxylic acid {l-
[5- (3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl].ethylidene}hydrazide potassium salt 25 Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 5-(pyrrolidine-1-
carbonyl)thiophene-2-carboxylic acid {l-[5-(3,4-

dichlorophenyl) -4-hydroxythiophen-3-
yl]ethylidene}hydrazide (yield 100%).
Morphology: orange solid
SYNTHETIC EXAMPLE 5
5 Synthesis of 5- [ (2-(l- [5-(3,4-
di chloropheny1)-4 -hydroxythiophen-3 -
ylj ethylidene}hydrazino)carbonyl)thiophene-2-carboxylic acid dimethylamide potassium salt 5-[£2-{l-[5-(3,4-Dichlorophenyl)-4-hydroxythiophen-3-10 yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid dimethylamide
Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 2-(3,4-dichlorophenyl)-3-
hydroxy-4-niethylcarbonylthiophene and 5-15 hydrazinocarbonylthiophene-2-carboxylic acid
dimethylamide prepared in Reference Synthetic Example 5 (yield 65%).
Morphology: colorless solid
LC/MS: conditions 5 retention time 4.93 (min) 20 LC/MS (BSI+) m/z; 482, 484 [M+l] +
LC/MS [ESI") m/z; 480, 482 [M-1J"
5-[(2-(l-[5-(3,4- Dichlorophenyl)-4-hydroxythiophen-3-yl)ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid dimethylamide potassium salt
25 Synthesis was carried out in the same manner as in -Synthetic Example 2 by using 5-{l-[5-(3,4-dichlorophenyl) -4-hydroxythiophen-3 -

yl]ethylidene}hydrazino)carbonyl] thiophene-2-carboxylic acid dimethylamide (yield 100%) . Morphology: orange solid
SYNTHETIC EXAMPLE 6 5 Synthesis of 5-[ (2-{l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazino)carbonyl]thiophene-2-carboxylic acid 2-methoxyethylamide potassium salt 5-[(2-{l-[5-(3,4-
Dichlorophenyl)-4-hydroxythiophen-3-lo yl]ethylidene}hydra2ino)carbonyl]thiophene-2-carboxylic acid 2-methoxyethylamide
Synthesis was carried out in the same manner as in
« Synthetic Example 2 by using 2-(3,4-dichlorophenyl)-3-
hydrcocy-4-methylcarbonylthiophene and 5-15 hydrazinocarbonylthiophene-2-carboxylic acid 2-
methoxyethylaraide prepared in Reference Synthetic Example
6 {yield 80%).
Morphology: pale yellow solid
LC/MS: conditions 7 retention time 3.15 (min) 20 LC/MS (ESI*) m/z; 512, 514 [M+l]*
LC/MS (ESI') m/z; "510, 512 [M-l]"
5-[(2-{l-[5-(3,4-
Dichlorophenyl)-4-hydroxythiophen-3- %
yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic
acid 2-methoxyethylamide potassium salt
as
Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 5-[ (2-{l-{5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-

yl] ethylidenejhydrazi.no) carbonyl] thiophene-2-carboxylic acid 2-methoxyethylamide (yield 100%). Morphology: orange solid SYNTHETIC EXAMPLE 7 5 Synthesis of 5- [(2-{l-[5- (3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid 3-pyridylamide potassium salt 5-[(2-{l- (5-(3,4-Dichlorophenyl)-4-hydroxythiophen- 3 -10 yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid 3-pyridylamide
28 mg of 2-(3,4-dichlorophenyl)-3-hydroxy-4-
methylcarbonylthiophene and 26 mg of 5-
hydrazinocarbonylthiophene-2-carboxylic acid 3-
is pyridylamide were dissolved in 2 mL of dimethyl sulfoxide
and heated at 100°C for 18 hours, and the solvent was
evaporated. Recrystallization from chloroform-diethyl
ether gave the desired product (yield 94%) .
Morphology: pale yellow solid
20 LC/MS: conditions 2 retention time 4.02 (mini
LC/MS (ESI*) m/z; 531, 533 [M+l]*
LC/MS (ESI") m/z; 529, 531 [M-l]"
5-[(2-{l-[5-(3,4-
Dichlorophenyl)-4-hydroxythiophen-3-
yl] ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic
25
acid 3-pyridylamide potassium salt
Synthesis was carried out in the same manner as in-Synthetic Example 2 by using 5-({2-{l-[5-(3,4-

dichlorophenyl) -4 -hydroxythiophen-3 -
yll ethylidene}hydrazino)carbonyl]
thiophene-2-carboxylic
acid 3-pyridylaraide (yield 76%) .
Morphology: orange solid
s
SYNTHETIC EXAMPLE 8
Synthesis of 5-[ (2-{l-[5-(4-trifluoromethylphenyl)-
4 -hydroxythiophen-3-yl]ethylidene}hydrazino)
carbonyl]thiophene-2-carboxylic"
acid diethylamide 10 Synthesis was carried out in the same manner as in
Synthetic Example 2 by uBing 2-(4-trif luoromethylphenyl) -
3-hydroxy-4-methylcarbonylthiophene (synthesized in
accordance with WO2004/108683) and 5-
hydrazinocarbonylthiophene-2-carboxylic acid diethylamide is prepared in Reference Synthetic Example 3 (yield 67%) .
Morphology: pale yellow solid
LC/MJS: conditions 6 retention time 3.82 (min)
LC/MS (ESI') m/z; 508 [M-l]"
SYiJTHETIC EXAMPLE 9 20 Synthesis of 5-(4-ieopropylpiperazine-l-
carbonyl) thiophene-2-carboxylic acid {l-[5-(4~
trif luoromethylphenyl) -4-hydxoxythiophen-3-
yl]ethylidene}hydrazide potassium salt
S- (4-Isopropylpiperazine-l-carbonyl) thiophene-2-as carbqxylic acid {l- [5- (4-trifluoromethylphenyl) -4-
hydroxythiophen-3-yl]ethylidene}hydrazide
Synthesis was carried out in the same manner as in

Synthetic Example 7 by using 2-(4-trifluoromethylphenyl)-
3-hydroxy-4-methylcarbonylthiophene and 5-(4-
isopropylpiperazine-1-carbonyl)thiophene-2-carbohydrazide
prepared in Reference Synthetic Example 1 (yield 55%). 5 Morphology: yellow solid
LC/MS: conditions 3 retention time 2.49 (min)
LC/MS (ESI+) m/z; 565 [M+l]+
LC/MS (ESI") m/z; 563 [M-lJ'
5-(4-Isopropylpiperazine-l-carbonyl)thiophene-2-io carboxylic acid {l-[5-(4-trifluoromethylphenyl)-4-
hydroxythiophen-3-yl]ethylidene}hydrazide potassium salt Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 5-(4-isopropylpiperazine-l-
carbonyl)thiophene-2-carboxylic acid (l-[5-(4-15 trifluoromethylphenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazide (yield 100%).
Morphology: red solid ■SYNTHETIC EXAMPLE 10
Synthesis of 5-(pyrrolidine-1-carbonyl)thiophene-2-20 carboxylic acid {l-[5-(4-trifluoromethylphenyl)-4-
hydroxythiophen-3-yl]ethylidene}hydrazide potassium salt.
5-(Pyrrolidine-1-carbonyl)thiophene-2-carboxylic acid {l-[5-(4-trifluoromethylphenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazide 25 Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 2-(4-trifluoromethylphenyl)-
3-hydroxy-4-methylcarbonylthiophene and 5-(pyrrolidine-1-

carbonyl)thiophene-2-carbohydrazide prepared in Reference
Synthetic Example 4 (yield 82%).
Morphology: pale yellow solid
LC/MS: conditions 8 retention time 5.10 (min) 5 LC/MS (ESI+) m/z; 508 [M+l]*
LC/MS (ESI") m/z; 506 [M-l]"
5-(Pyrrolidine-l-carbonyl)thiophene-2-carboxylic acid fi¬ts- (4-trif luoromethylphenyl) -4-hydroxythiophen-3-
yljethylidene}hydrazide potassium salt 10 Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 5-(Pyrrolidine-1-
carbonyl)thiophene-2-carboxylic acid {l-[5-(4-
trif luoromethylphenyl) -4-hydroxythiophen-3 -
yl]ethylidene}hydrazide (yield 100%). 15 Morphology: red solid
SYNTHETIC EXAMPLE 11
Synthesis 2-chloro.-4- (2- (l- [5-
(3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl)ethylidene}hydrazinocarbonyl)-N-(2-
hydroxyethyl)benzamide potagsium salt 20 2-Chloro-4-(2-{l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazinocarbonyl)-N-(2-
hydroxyethyl)benzamide
To 2-(3,4-dichlorophenyl)-3-hydroxy-4-methylcarbonylthiophene (40 mg, 0.14 mraol) and 2-chloro-25 4-hydrazinocarbonyl-N-(2-hydroxyethyl)benzamide (43 mg, 0.17 mmol) prepared in Reference Synthetic Example 8 in dimethylformamide (0.7 raL), concentrated hydrochloric

acid (12 uL, 0.14 mmol) was added at room temperature,
and the resulting mixture was stirred at room temperature
for 1 day and stirred with 2-chloro-4-hydrazinocarbonyl-
N-(2-hydroxyethyl)benzamide (18 mg( 0.07 nrmol) for 1 day.
5 After addition of water, the resulting crystals were
recovered by filtration and dried. Chloroform was added,
and the resulting crystals were recovered by filtration
to give the desired product (yield 83%).
Morphology: Pale green solid
10 LC/MS: conditions 3 retention time 3.42 (min)
LC/M3 (ESI*) m/z; 526, 528 [M+l]*
LC/MS (BSI") m/z; 524, 526 [M-l]"
2-Chloro-4-(2-fi¬ts- (3,4-dichlorophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazinocarbonyl}-N-(2-hydroxyethyDbenzamide potassium salt
Synthesis was carried out in the same manner as in Synthetic Example 2 by using 2-chloro-4-(2-{l-[5-(3,4-
15 dichlorophenyl)-4-hydroxythiophen-3-
i ■
yl)'ethylidene}hydrazinocarbonyl) -N- (2-hydroxyethyl)benzamide (yield 77%).
20
Morphology: red solid SYNTHETIC EXAMPLE 12 Synthesis o£ 5-((2-{l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl] thiophene-2-carboxylic acid methyl-2-picolylamide potassium salt 25 5-[{2-{l-[5-(3,4-
Dichlorophenyl)-4 -hydroxythiophen-3 -

yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid methyl-2-picolylamide
Synthesis was carried out in the same manner as in
Synthetic Example 7 by using 2- (3,4-dichlorophenyl)-3-
5 hydroxy-4-methylcarbonylthiophene and 5-
hydrazinocarbonylthiophene-2-carboxylic acid methyl-2-
picolylamide prepared in Reference Synthetic Example 10
(yield 66%).
Morphology: pale yellow solid
lo LC/MS: conditions 3 retention time 3.57 (min)
LC/MS (ESI*) m/z; 559, 561 [M+l]*
LC/MS (ESI") m/z; 557, 559 [M-l]"
5- [(2-{l-[5- (3,4-Dichlorophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazino)carbonyl)thiophene-2-carboxylic acid methyl-2-picolylamide potassium salt
Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 5-[(2-{l-[5- (3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid methyl-2-picolylamide (yield 100%) . Morphology: red solid SYNTHETIC EXAMPLE 13 Synthesis of 5-[ (2-{i-[5-(4-trifluoromethylphenyl)-4-hydroxythiophen-3-yl]ethylidene J hydrazino)carbonylJthiophene-2 -carboxylic acid methyl-2-picolylamide potassium salt .-, 25 5-[(2-{l-[5-(4-Trifluoromethylphenyl)-4-hycroxythiophen-3-

yl]ethylidene}hydrazine)carbonylJ thiophene-2-carboxylic acid methyl-2-picolylaniide
Synthesis was carried out in the same manner as in Synthetic Example 7 by using 2-(4-trifluoromethylpheny)-5 3 -hydroxy-4-methylcarbonylthiophene and 5 -
hydrazinocarbonylthiophene-2-carboxylic acid methyl-2-picolylamide prepared in Reference Synthetic Example 10. Morphology: pale green solid
10 5-[(2-{l-[5-(4-Trifluoromethylphenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazino)carbonyl)thiophene-2-carboxylic acid methyl-2-picolylamide potassium salt
Synthesis was carried out in the same manner as in Synthetic Example 2 by using 5-[{2-[l- [5- (4-trifluoromethylphenyl)-4-hydroxythiophen-3-
15 yl)ethylidenejhydrazino)carbonyl)thiophene-2-carboxylic acid methyl-2-picolylamide {yield 100%).
Morphology: orange solid
SYNTHETIC EXAMPLE 14
Synthesis of 5-(2-cyanobutyryl)thiophene-2-carboxylic 20 acid {l- [5- (3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazide potassium salt
5-(2-Cyanobutyryl)thiophene-2-carboxylic. acid {l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazide 25 Synthesis was carried out in the same manner as in
Synthetic Example 7 by using 2-(3,4-dichlorophenyl)-3-
hydroxy-4-methylcarbonylthiophene and 5-(2-

cyanobutyryl)thiophene-2-carbohydrazide prepared in
Reference Synthetic Example 9 (yield 36%) .
Morphology: yellow solid
LC/MS: conditions 3 retention time 3.45 (min) 5 LC/MS (ESI+) m/z; 506, 508 [M+l]+
LC/MS (ESI") m/z; 504, 506 [M-l]"
5-(2-Cyanobutyryl)thiophene-2-carboxylic acid {l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazide potassium salt 10 Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 5-(2-cyanobutyryl)thiophene-
2-carboxyIic acid {l-[5-(3,4-dichlorophenyl)-4-
hydroxythiophen-3-yl]ethylidenejhydrazide (yield 75%) .
Morphology: red solid 15 SYNTHETIC EXAMPLE 15
Synthesis of 5-(2-cyanobutyryl)thiophene-2-carboxylic
acid {l-[5-(4-trifluoromethylphenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazide potassium salt
5-(2-Cyanobutyryl)thiophene-2-carboxylic acid {l-[5-(4-20 trifluoromethylphenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazide
Synthesis was carried out in the same manner as in
Synthetic Example 7 by using 2-(4-trifluoromethylphenyl)-
3-hydroxy-4-methylcarbonylthiophene and 5-(2-■ 25 cyanobutyryl)thiophene-2-carbohydrazide prepared in
Reference Synthetic Example 9 (yield 53%).
Morphology: yellow solid

LC/MS: conditions 3 retention time 3.82 (min) ■LC/MS (ESI+) m/z; 506 [M+l]+ LC/MS (ESI") m/z; 504 [M-l]"
5-(2-Cyanobutyryl)thiophene-2-carboxylic acid {l-[5-(4-5 trifluoromethylphenyl)-4-hydroxythiophen-3-yl]ethylidene}hydrazide potassium salt
Synthesis was carried out in the same manner as in Synthetic Example 2 by using 5-(2-cyanobutyryl)thiophene-2-carboxylic acid {l-[5-(4-trifluoromethylphenyl)-4-10 hydroxythiophen-3-yl]ethylidenejhydrazide (yield 91%). Morphology: red solid SYNTHETIC EXAMPLE 16
Synthesis of 5-(2-cyanobutyryl)thiophene-2-carboxylic acid {l- [5-(4-bromophenyl)-4-hydroxythiophen-3-15 yl]ethylidenejhydrazide potassium salt
5-(2-Cyanobutyryl)thiophene-2-carboxylic acid {l-[5-(4-bromophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazide
Synthesis was carried out in the same manner as in Synthetic Example 7 by using 2-(4-bromophenyl)-3-hydroxy-20 4-methylcarbonylthiophene (synthesized in .accordance with WO2004/108684) and 5-(2-cyanobutyryl)thiophene-2-carbohydrazide prepared in Reference Synthetic Example 9 (yield 51%).
Morphology: yellow solid 25 LC/MS: conditions 3 retention time 3.85 (min) LC/MS (ESI*) m/z; 516, 518 [M+l]+ . LC/MS. (ESI") m/z; 514, 516 [MT1J"

5- (2-Cyanobutyryl) thiophene-2-carboxylic acid {l-[5-(4-bromophenyl) -4-hydroxythiophen-3-yl] ethylidenejhydrazide potassium salt
Synthesis was carried out in the sane manner as in 5 Synthetic Example 2 by using 5- (2-cyanobutyryl) thiophene-2-carbcocylic acid {l-[5-(4-broraophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazide (yield 91%). Morphology: red solid SYNTHETIC EXAMPLE 17
Synthesis of 5-[(2-{l-(5- (4-brorriophenyl) -4-hydroxythiophen-3-yl] ethylidenejhydraziho)carbonyl]thiophene-2-carboxylic acid 3-picolylamide
2-(4-Bromophenyl)-3-hydroxy-4-raethylcarbonylthiophene (50.5 rag, 0.17 mmol) (synthesized
15 in accordance with WO2004/108683) and 5-
hydrazinocarbonylthiophene-2-carboxylic acid 3-picolylamide prepared in Reference Synthetic Example 11 were dissolved in dimethylsulfoxide (4.0 mL) and heated at 110°C for 24 hours. The solvent was evaporated, and
20 the residue was washed with methanol and water to give the desired product (yield 81%). Morphology: pale yellow solid LC/MS: conditions 6 retention time 4.IS (min) LC/MS (ESI*) m/z; 555, 557 [M+l]*
25 LC/MS (ESI") m/z; S53, 555*[M-l3" SYNTHETIC BXAMPLE 18 Synthesis of 5-[(2-{l-[S- (4-bromophenyl)-4-

hydroxythiophen-3-yl] ethylidene}hydrazino}carbonyl]thiophene-2-carboxylic acid 4-picolylamide
Synthesis was carried out in the same manner as in
Synthetic Example 17 by using.2- (4-bromophenyl) -3-
s hydroxyl-4-methylcarbonylthiophene and 5-
hydxazinocarbonylthiophene-2-carboxylic acid 4-
picolylamide prepared in Reference Synthetic Example 12
{yield 72%).
Morphology: pale yellow solid
10 LC/MS: conditions 6 retention time 4.03 (min)
LC/MS (ESI*) m/z; 555, 557 [M+l]*
LC/MS (ESI") m/z; 553, 555 [M-l]"
SYNTHETIC EXAMPLE 19
Synthesis of 5-[(2-{l-[5-(4-broraophenyl)-4-1S hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl] thiophene-2-carboxylic acid furan-2-ylmethylamide
2-(4-Bromophenyl) -3-hydroxy-4-methylcarbonylthiophene (50.1 mg, 0.17 mraol) and 5-hydrazinocarbonylthiophene-2-carboxylic acid (furan-2-
20 ylmethyl)amide (45.0 rag, 0.17 mmol) prepared in Reference Synthetic Example 13 in 2-propanol (4.0 mL) were heated with p-tosylic acid monohydrate (6 mg) at 100°C for 7.5 hours and cooled to room temperature. The precipitated solid was recovered by filtration, washed with 2-propanol
as (1 mL) and dried to give the desired product (yield 72%) . Morphology: pale yellow solid LC/MS: conditions 6 retention time 4.98 (min)

LC/MS (ESI*) m/z; 544, 546 [M+l]* LC/MS (ESI") m/z; 542, 544 [M-l]" SYNTHETIC EXAMPLE 20
Synthesis of 5-[(2-{1-[5-(4-bromophenyl)-4-hydroxythiophen-3-5 yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid diethylamide
Synthesis was carried out in the same manner as in
Synthetic Example 19 by using 2-(4-bromophenyl)-3-
hydroxy-4-methylcarbonylthiophene and 5-10 hydrazinocarbonylthiophene-2-carboxylic acid methylamide
prepared in Reference Synthetic Example 14 (yield 65%).
Morphology: pale yellow solid
LC/MS: conditions 6 retention time 4.67 (min)
LC/MS (ESI*) m/z; 478, 480 [M+l]* 15 LC/MS (ESI*) m/z; 476, 478 [M-l]"
SYNTHETIC EXAMPLE 21
Synthesis of 5-[(2-{l-[5-(4-bromophenyl)-4-hydroxythiophen-3-yl] ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid isopropylamide
20 Synthesis was carried out in the same manner as in Synthetic Example 19 by using 2-(4-bromophenyl)-3-hydroxy-4-methylcarbonylthiophene and 5-hydrazinocarbonylthiophene-2-carboxylic acid isopropylamide prepared in Reference Synthetic Example 15
2s (yield 70%).
Morphology: white solid
LC/MS: conditions 2 retention time 4.50 (min)

LC/MS (ESI*) m/z; 506, 508 [M+13* LC/MS (ESI") m/z; 504, 506 [M-l]"
SYNTHETIC EXAMPLE 22
Synthesis of 5-[(2-{l-[4-hydroxy-5-(4-
trifluoromethoxyphenyl)thiophen-3-
yl] ethylidenejhydrazi.no) carbonyl]thiophene-2-carboxylic
acid 2-picolylamide Synthesis was carried out in the same manner as in Synthetic Example 17 by using 3-hydroxy-2-(4-
10 trifluoromethoxyphenyl) -4-raethylcarbonylthiophene
(synthesized in accordance with W02004/108683) and 5-hydrazinocarbonylthiophene-2-carboxylic acid 2-picolylamide prepared in Reference Synthetic Example 16 (yield 76%).
15 Morphology: pale yellow solid
LC/MS: conditions 2 retention time 3.92 (min) LC/MS (ESI*) m/z; 561 [M+l] + LC/MS (ESI') m/z; 559 [M-l]"
SYNTHETIC EXAMPLE 23
20 Synthesis of 5-[ (2-{l-[5-(4-
trifluoromethylphenyl)-4-hydroxythiophen-3-
yl]ethyl idene} hydrazine*)carbonyl]thiophene-2-carboxylic
acid isopropylamide
Synthesis was carried out in the same manner as in 25 Synthetic Example 19 by using 2-(4-trif luoromethylphenyl) -3-hydroxy-4-methylcarbonylthiophene (synthesized in accordance with

WO2004/108683) and 5-hydrazinocarbonylthiophene-2-carboxylic acid isopropylamide prepared in Reference Synthetic Example 15 (yield 83%). Morphology: light gray solid s LC/MS: conditions 3 retention time 3.65 (min) LC/MS (ESI*) m/z; 496 [M+l]* LC/MS (ESI") m/z; 494 [M-l]*
SYNTHETIC EXAMPLE 24
Synthesis of 5-[(2-{l-[5-(4-chlorophenyl) -4-io hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl] thiophene- 2 -carboxylic acid 4-picolylamide
Synthesis was carried out in the same manner as in Synthetic Example 17 by using 2-^-chlorophenyD-S-hydroxy^-methylcarbonylthiophene (synthesized in
is accordance with W02004/108683) and 5-
hydrazinocarbonylthiophene-2-carboxylic acid 4-picolylamide prepared in Reference Synthetic Example 12 (yield 69%). Morphology: light brown solid
go LC/MS: conditions 3. retention time 2.55 (min) LC/MS (ESI*) m/z; 511, 513 [M+l] * LC/MS (ESI') m/z; 509, 511 [M-l]"
SYNTHETIC EXAMPLE 25
Synthesis of 5-[ (2-{l-[5-(4-chlorophenyl)-4-25 hydroxythiophen-3-yl]ethylidene)hydrazino)carbonyl]thiophene-2-carboxylic acid 2-picolylamide
Synthesis was carried out in the same manner as in

Synthetic Example 17 by using 2-(4-chlorophenyl)-3-
hydroxy-4-methylcarbonylthiophene and 5-
hydrazinocarbonylthiophene-2-carboxylic acid 2-
picolylaraide prepared in Reference Synthetic Example 16
s (yield 68%).
Morphology: yellow solid
LC/MS: conditions 3 retention time 3.10 (min)
LC/MS (ESI*) m/z; 511, 513 [M+l]*
LC/MS (ESI") m/z; 509, 511 [M-l]~ 10 SYNTHETIC EXAMPLE 26
Synthesis of 5- [ (2-{l-(5-(4-chlorophenyl)-4-
hydroxythiophen-3-yl)ethylidene}hydrazine)carbonyl] thiophene-2-
carboxylic
acid 3-picolylamide
Synthesis was carried out in the same manner as in
is Synthetic Example 17 by using 2- (4-chlorophenyl) -3-
hydroxy-4-methylcarbonylthiophene and 5-
hydrazinocarbonylthiophene-2-carboxylic acid 3-
picolylamide prepared in Reference Synthetic Example 11
(yield 50%). 20 Morphology: yellow solid
LC/MS: conditions 3 retention time 2.72 (min)
LC/MS (ESI*) m/z; 511, 513 (M+l]*
LC/MS (ESI") m/z; 509, 511 [M-H*
SYNTHETIC EXAMPLE 27 35 Synthesis of 5- [ (2-{l-[5-(4-chlorophenyl)-4-hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl]thiophene-2-carboxylic acid methylamide

Synthesis was carried out in the same manner as in Synthetic Example 19 by using 2-(4-chlorophenyl)-3-hydroxy-4-methylcarbonylthiophene and 5-hydrazinocarbonylthiophene-2-carboxylic acid methylamide 5 prepared in Reference Synthetic Example 14 (yield 82%). Morphology: pale yellow solid LC/MS: conditions 6 retention time 4.62 (min) LC/MS (ESI*) m/z; 434, 436 [M+l]* LC/MS (ESI-) m/z; 432, 434 tM-ll* 10
SYNTHETIC EXAMPLE 28
Synthesis of 5-((2-{l-(5-(3,4-dimethylphenyl)-4-hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl]thiophene-2-carboxylic acid isopropylamide
is Synthesis was carried out in the same manner as in Synthetic Example 19 by using 2-(3,4-dimethylphenyl)-3-hydroxy-4-methylcarbonylthiophene (synthesized in accordance with WO2004/108683) and 5-hydrazinocarbonylthiophene-2-carboxylic acid
20 isopropylamide prepared in Reference Synthetic Example 15 (yield 71%).
Morphology: pale yellow solid LC/MS: conditions 2 retention time 4.40 (min) LC/MS (ESI*) m/z; 456 [M+lJ*
25 LC/MS (ESI") m/z; 454 [M-l]"
SYNTHETIC EXAMPLE 29
Synthesis of 5-[ {2-{l-[5-(3,4-dichlorophenyl)-4-

hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl]thiophene-2-carboxylic acid 2-picolylamide
Synthesis was carried out in the same manner as in S Synthetic Example 17 by using 2- (3,4-dichlorphenyl-3-
hydroxy-4-methylcarbonylthiophene (synthesized in
accordance with WO2004/108683) and S-
hydrazinocarbonylthiophene-2-carboxylic acid 2-
picolylamide prepared in Reference Synthetic Example 16 10 {yield 65%)-
Morphology: yellow solid
LC/MS: conditions 3 retention time 3.30 (min)
LC/MS (ESD m/z; 545, 547 [M+lJ*
LC/MS (ESI") m/2; 543, 545 [M-l]" is
SYNTHETIC EXAMPLE 30
Synthesis of S-[ (2-{l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-ylJethylidene}hydrazino}carbonyl]thiophene-2-carboxylic acid 4-picolylamide
20 Synthesis was carried out in the same manner as in . Synthetic Example 17 by using 2- (3,4-dichlorophenyl)-3-hydroxy-4-methylcarbonylthiophene and 5-hydrazinocarbonylthiophene-2-carboxylic acid 4-picolylamide prepared in Reference Synthetic Example 12
as (yield 51%).
Morphology: pale yellow solid
LC/MS: conditions 2 retention time 3.25 (min)

LC/MS (ESI*) m/z; 545, 547 [M+l] * LC/MS (ESI") m/z; 543, 545 [M-l]"
SYNTHETIC EXAMPLE 31
Synthesis of 5- [ (2-{l- [5-(3,4-dichlorophenyl)-4-5 hydroxythiophen-3-yl)ethylidene}hydrazino)carbonyl] thiophene-2-carboxylic acid 2-(pyridin-4-yl) ethylamide
2-(3,4-Dichlorophenyl)-3-hydroxy-4-methylcarbonylthiophene (0.11 g, 0,38 tnmol) and 5-
10 hydrazinocarbonylthiophene-2-carboxylic acid (2-pyridin-4-yl)ethylamide (0.10 g, 0.34 mmol) prepared in Reference Synthetic Example 17 were suspended in N,N-dimethylformamide (2.0mL) and left at 100°C for 5 hours and then at room temperature for 20 hours. Water, (0.20
is tali) was added with stirring, and the precipitated solid
was recovered by filtration, washed w.i.th chloroform and
dried to give the desired product, 5-[(2-{l-(5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidene}hydrazino)carboriyl}thiophene-2-carboxylic 20 acid 2-(pyridin-4-yl)ethylamide (yield 66%).
Morphology: white solid
LC/MS: conditions 3 retention time 2.67 (min)
LC/MS (ESI+) m/z; 559, 561 [M+l]*
SYNTHETIC EXAMPLE 32 25 Synthesis of 5-((2-{l-[S-(3,4-dichlorophenyl)-4~ hydroxythiophen-3-yl]ethylidene}hydrazino) carbonyl]thiophene:2-carboxylic

acid [(i-methyl-lH-pyrazol-5-yDmethyl]amide
2-(3,4-Dichlorophenyl)-3-hydroxy-4-methylcarbonylthiophene (50 mg, 0.17 mmol) and 5-(hydrazinecarbonyl)-N-[(1-methyl-lH-pyrazol-5-yl)methyl]thiophene-2-carboxamide (49 5 mg, 0.17 mmol) prepared in Reference Synthetic Example 18 were dissolved in N,N-dimethylformamide (0.50 mL) and heated at 70°C for 24 hours and cooled to room temperature. After addition of water, the precipitated crystals were recovered by filtration, washed with water x-and chloroform and dried to give the desired product 10 {yield 76%) .
Morphology: pale yellow solid LC/MS: conditions 3 retention time 3.60 (min) LC/MS (ESI*) m/z; 548, 550 tM+l] + 15 LC/MS (ESI") m/z; 546, 548 (M-l]"
SYNTHETIC EXAMPLE 33
Synthesis of 5-[(2-{1-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-yl]ethylidenejhydrazino) carbonyl]thiophene-2-carboxylic acid (5-methylisoxazol-3-ylmethyl)amide
20
Synthesis was carried out in the same manner as in Synthetic Example 32 by using 2- (3,4-dichlorophenyl) -3-hydroxy-4-methylcarbonylthiophene and 5-hydrazinocarboynlthiophene-2-'carboxylic acid (5-25 methylieoxazol-3-ylmethyl) amide prepared in Reference Synthetic Example 19 (yield 67%). Morphology: yellow solid

LC/MS: conditions 3 retention time 3.77 (min)
LC/MS (ESI*) m/z; 549, 551 [M+l]*
LC/MS (ESI') m/z; 547, 549 [M-l)"
SYNTHETIC EXAMPLE 34 s Synthesis of 5-[(2-{l-[5- (3,4-dichlorophenyl)-4-
hydroxytbiophen-3-
yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic
acid (5-methylpyrazin-2-ylmethyl)araide
Synthesis was carried out in the same manner as in 10 Synthetic Example 17 by using 2-(3,4-dichlorophenyl)-3-
hydroxy- 4 -methylcarbonylthiophene and 5-
hydrazinocarbonylthiophene-2-carboxylic acid (5-
methylpyrazin-2-ylmethyl)amide prepared in Reference
Synthetic Example 20 (yield 74%). 15 Morphology: pale yellow solid
LC/MS: conditions 3 retention time 3.62 (min)
LC/MS (ESI*) m/z; 560, 562 [M+l]*
LC/MS (ESI") m/z; 558, 560 [M-l]*
SYNTHETIC EXAMPLE 35 20 Synthesis of 5-[ (2-{l-[5-(3,4-dichlorophenyl) -4-hydroxythiophen-3-
yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid {isoxazol-5-ylmethyl)amide
Synthesis was carried out in the same manner as in 25 Synthetic Example 19 by using 2-(3,4-dichlorophenyl)-3-hydroxy-4-methylcarbonylthiophene and 5-hydrazinocarbonylthiophene-2-carboxylic acid (isoxazol-5-

ylmethyl) amide prepared in Reference Synthetic Example 19 (yield 74%) .
Morphology: pale yellow solid LC/MS: conditions 3 retention time 3.69 (rain) S LC/MS (BSI*) m/z; 535, 537 [M+l] + LC/MS (ESI') m/z; 533, 535 [M-l]"
SYNTHETIC EXAMPLE 36 Synthesis of 5-[(2-{l-[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-io yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic acid (3-methoxyisoxazol-5-ylmethyl)amide
Synthesis was carried out in the same manner as in
Synthetic Example 19 by using 2-(3,4-dichlorophenyl)-3-
h.ydroxy-4-methylcarbonylthiophene and 5-15 hydrazinocarbonylthiophene-2-carboxylic acid (3-
methoxyisoxazol-5-ylmethyl) amide prepared in Reference
Synthetic Example 22 (yield 52%) .
Morphology: pale yellow solid
LC/MS: conditions 3 retention time 3.75 (min) 20 LC/MS (ESI*) m/z; 565, 567 [M+l]*
LC/MS (ESI*) m/z; 563, 565 [M-l]"
SYNTHETIC EXAMPLE 37 Synthesis of 5-[(2-{l-
[5-(3,4-dichlorophenyl)-4-hydroxythiophen-3-2S yl]ethylidenejhydrazino)carbonyl)thiophene-2-carboxylic acid (1,5-dimethyl-lH-pyrazol-3-ylmethyl)amide
Synthesis was carried out in the same manner as in

Synthetic Example 17 by using 2- (3,4-dichlorophenyl) -3-hydroxy-4-methylcarbonylthiophene and 5-hydrazinocarbonylthiophene-2-carboxylic acid (1,5-dimethyl-lH-pyrazol-3-ylmethyl) amide prepared in 5 Reference Synthetic Example 23 (yield 86%). Morphology: yellow solid
LC/MS: conditions 3 retention time 3.65 (rain) LC/MS (ESI*) m/z; 562, 564 (M+l] * LC/MS (BSI") m/z; 560, 562 [K-l] "
10
SYNTHETIC EXAMPLE 38
Synthesis of 5-[ (2-{l-[5-(3,4-dichlorophenyl) -4-hydroxythiophen-3-
ylj ethylidene}hydrazino)carbonyl)Chiophene-2-carboxylic acid (pyrazin-2-ylmethylJ amide
is Synthesis was carried out in the same manner as in
Synthetic Example 17 by using 2- (3,4-dichlorophenyl) -3-
hydroxy-4-methylcarbonylthiophene and 5-
hydrazinocarbonylthiophene-2-carboxylic acid (pyrazin-2-
ylmethyl) amide prepared in Reference Synthetic Example 24 20 (yield 88%).
Morphology: pale yellow solid
LC/MS: conditions 3 retention time 3.57 (min)
LC/MS (ESI+) m/z; 546, 548 [M+13*
LC/MS (ESI") m/z; 544, 546 [M-l]"
2S
SYNTHETIC EXAMPLE 39 Synthesis of 5- [ (2-{1- [5- (3,4 -dichlorophenyl)-4-hydroxythiophen-3-

yl]ethylidene}hydrazino)carbonyl]thiophene-2-carboxylic acid 4-picolyl-l-oxide-amide
2- (3,4-Dichlorophenyl)-3-hydroxy-4-methylcarbonylthlophene (32 mg, 0.11 mmol) and 4-[{5-s hydraztiiocarbonyl) thiophene-2-carboxamido}methyl] pyridine 1-oxide (30 rag, 0.10 mmol) prepared in Reference Synthetic Example 25 were suspended in N,N-dimethylformamide (0.60 mL) and stirred at 80°C for 90 hours. After addition of 2- (3,4-dichlorophenyl) -3-
10 hydroxy-4-methylcarbonylthiophene (26 mg, 0.091 mmol), the suspension was stirred at 80°C for 24 hours, left at room temperature for 5 hours and concentrated to dryness to give a yellow crude- paste (91 mg) . The paste was suspended in chloroform (2.0 mL), and the insolubles were
15 recovered by filtration and dried to give the desired
product, 5-[<2-{l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl]ethylidenejhydrazino)carbonyl)thiophene-2-carboxylic acid 4-
picolyl-1-oxide-amide (yield 52%).
20 Morphology: pale yellow solid
LC/MS: conditions 3 retention time 3.29 (min) LC/MS (ESI*) m/zf 561, 563 [M+lJ* LC/MS (ESI") m/z; 559, 561 IM-1]"
SYNTHETIC EXAMPLE 40 2S Synthesis of 5-I (2-{l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-
yl)ethylidene}hydrazino)carbonyl]thiophene-2 -carboxylic

acid 3-(pyxidin-4-yl)propylamide
2- (3,4-Dichlorophenyl) -3-hydroxyz¬ine thy lea rbony It hiophene (42 mg, 0.14 mmol) and 5-hydrazinocarbonylthiophene-2-carboxylic acid 3-(pyridin-5 4-yl)propylaraide (40 mg, 0.13 mmol) prepared in Reference Synthetic Example 26 were dissolved in N,N-
dimethylformamide (1.0 mL) and left at 70°C for 16 hours
and then at room temperature for 24 hours. Water (0.36
mL) was added with stirring, and the precipitated solid
10 was..recovered by filtration to give the desired product,
5'- [ (2-{l- [5- (3,4-dichlorophenyl) -4-hydroxythiophen-3-yl]ethylidene}hydrazino)carbonyl]thiophene-2-carboxylic acid 3-(pyridin-4-yl)propylamide (yield 95%).
Morphology: pale yellow solid
is LC/MS: conditions 2 retention time 3.31 (tnin)
LC/MS (ESI*) tn/z; 573, 575 [M+l]+
LC/MS (ESI") m/z; 571, 573 £M-1]"
SYNTHETIC EXAMPLE 41
Synthesis of 5-[(2-{l-(5-
(4-trifluoromethylphenyl)~4-hydroxythiophen-3-
20
yl]ethylidenejhydrazino)carbonyl]thiophene-2-carboxylic
acid [(l-methyl-lH-imidazol-5-yl)methyl]amide
Synthesis was carried out in the same manner as in .
Synthetic Example 17 by using 2-{4-
trifluoromethylphenyl)-3-hydroxy-4-
methylcarbonylthiophene and 5-(hydrazinecarbonyl)-N-[(1-methyl-lH-25
imidazol-5-yl)methyl)thiophene-2-carboxamide

prepared in Reference Synthetic Example 27 {yield 39%).
Morphology: pale yellow solid
LC/MS: conditions 2 retention time 3.02 (min)
LC/MS (ESI*) m/z; 548 [M+l]* 5 LC/MS (ESI") m/z; 546 [M-l]"
SYNTHETIC EXAMPLE 42
Synthesis of 5-(3-hydroxypyrrolidine-l-
carbonyl)thiophene-2-carboxylic acid {l-[5-(3,4-
dichlorophenyl)-4-hydroxythiophen-3-10 yl]ethylidene}hydrazide
Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 2-(3,4-dichlorophenyl)-3-
hydroxy-4-methylcarbonyl)thiophene and 5-(3-
hydroxypyrrolidine-1-carbonyl)thiophene-2-carbohydrazide is prepared in Reference Synthetic Example 28 (yield 97%).
Morphology: yellow solid
LC/MS: conditions 6 retention time 3..47 (min) *
LC/MS (ESI+) m/z; 524, 526 [M+l]*
LC/MS (ESI") m/z; 522, 524 [M-l]" 20 SYNTHETIC EXAMPLE 43.
Synthesis of 5-(3-hydroxypyrrolidine-l-
carbonyl)thiophene-2-carboxylic acid {l-[5-(4-
tr i fluoromethyIphenyl)-4-hydroxythiophen-3 -
yl]ethylidene}hydrazide 25 Synthesis was carried out in the same manner as in
Synthetic Example 2 by using 2-(4-trifluoromethyIphenyl)-
3-hydroxy-4-methylcarbonyl)thiophene and 5-(3-

hydr oxypyr rolidine -1 -carbonyl) thiophene - 2 - carbohydra z ide prepared in Reference Synthetic Example 28 (yiald 84%). y , Morphology: pale yellow solid LC/MS: conditions 6 retention time 3.34 (min) 5 LC/MS (ESI*) m/z; 524, 525 [M+l]+ LC/MS (ESI") m/z; 522, 523 [M-l] "
The structures of the compounds obtained in the Reference Synthetic Examples and the Synthetic Examples

ASSAY EXAMPLE 1
Stimulation, of Proliferation of a Thrombopoietin-5 dependent Cell Line
The reactivity of the compounds of the Synthetic Examples of the present invention with thrombopoietin (TPO) receptor was assayed using a human leukemic, cell line, UT7/EP0-mpl.
10 (1) Cells and cell culture
UT7/EPO-mpl is a stable transformed cell line obtained by introducing into human leukemic cell line UT7/EPO a vector that induces expression of human thrombopoietin receptor (c-mpl) under control of
15 cytomegalovirus' immediate-early promoter by the method of Takatoku et al. (J. Biol. Chem., 272:7259-7263 (1997)). Proliferation of this cell line is stimulated by thrombopoietin, while its mother cell line UT7/EP0 exhibits no response to thrombopoietin. These two cell
20 lines were subcultured in IMDM (GIBCO) containing 10% fetal bovine serum (Thermo Electron or BioWest) using a

C02 incubator (5% C02, 37°C) : (2) Cell proliferation assay
The subcultured cells described above were washed twice with PBS and suspended in IMDM containing 10% fetal
5 bovine serum at a cell density of S x 10* cells/mL. The cell suspension was transferred to a 96-well tissue culture plate (CORNING) in 100-ul aliquots. Then either thrombopoietin (Pepro Tech EC) or the compounds of the Synthetic Examples dissolved in dimethyl sulfoxide was
10 diluted 83-fold with IMDM containing 10% fetal bovine
serum and added to the aforementioned cell suspension in 20~ul aliquots. The cell suspension was incubated in a C02 incubator (5% C02, 37°C) for 4 days. Cell proliferation was assayed using WST-8 reagent (Kishida
15 Chemical Co., Ltd.) according to instructions by the manufacturer. A 10-ul aliquot of 5 mM WST-8 reagent solution was added to each well of the tissue culture plate, and the plate was incubated at 37°C for 4 h. The formazan pigment generated was detected by measuring the
20 absorbance at 450 nm with a 96-well microplate reader
(Nihon Molecular Devices, Spectramax 190). Proliferation of thrombopoietin-responsive UT7/EP0-mpl cells was', stimulated by the compounds of the Synthetic Examples of the pres.ent invention in a concentration-dependent
25 manner, while no effect of the compounds of the Synthetic Examples on proliferation was observed with UT7/EP0, the . mother cell line. These results indicate that the

compounds of the Synthetic Examples of the present invention act on the thrombopoietin receptor selectively as its activators.
The compounds of Synthetic Examples 1 to 43 (in the 5 free forms) were tested to determine the concentration of each compound that yields -a growth rate corresponding to 50% of the growth of human leukemic cell line UT7/EPO-mpl observed in the presence of 10 ng/ml TPO {EC50) . The compounds of Synthetic Examples 1 to 43 all had EC50 of
10 about 10 ng/mL or below. ASSAY EXAMPLE 2
Each of the compounds of the Synthetic Examples was suspended in a 99/l liquid mixture of 0.5% methylcellulose/Polyoxyethylene Sorbitan Monooleate and
15 orally administered to 7-week-old male Sprague-Dawley rats {Japan SLC, Inc.) at a dose of 10 mg/kg/10 mL through a stomach tube. Between 0.5 and 2 hours after the administration of the compounds, blood was periodically collected from the cervical vein with
20 heparin as the anticoagulant. The blood was centrifuged at 3500 min"1 for 10 minutes to obtain plasma. The plasma was added to the assay system used for assay of proliferation of a thrombopoietin-dependent cell line DT7/EP0-mpl in Assay Example i at final concentrations of
25 from 0.1 to 3 %, and the cell proliferation was assayed. The concentration of each compound in plasma was "calculated from the cell proliferation in the presence of

plasma by comparison with a standard curve of cell proliferation versus compound concentration prepared for each compound or measured by LC/MS {Agilent Technologies, Agilent 1100 series LC/MS D) . Each of the compounds of s Synthetic Examples 1 to 16, 31 and 38 (Compounds of
Synthetic Examples 31 and 38 were tested in the form of potassium salts) attained a maximum blood concentration (Cmax) of at least about 300 ng/mL 0.5 to 2 hours after the oral administration to rats. 10 ASSAY EXAMPLE 3
Megakaryocyte Colony Stimulating Activity
The action of the compounds of Synthetic Examples 1 to 43 of the present invention and Reference Synthetic Examples 29 and 30 on the proliferation, differentiation 15 and maturation of megakaryocyte cells was measured- by the megakaryocyte colony forming method using human bone marrow cells. Human bone marrow CD34* cells (Carnbrex Bio Science Walkersville) were incubated on 2-well chamber slide for 11 days in a C02 incubator (5% C02, 37°C) using 20 MegaCult^-C (StemCell Technologies) containing Q.1%
(v/v) of the compounds of Synthetic Examples dissolved in dimethyl sulfoxide. After dehydration and fixation, the cells were stained with an anti-glycoprotein Ilb/XIIa antibody in accordance with the instruction by the 25 manufacturer. The colonies consisting of at least 8 stained megakaryocyte cells in each well were counted under a microscope. The megakaryocyte colony counts in

at least 2 wells were averaged.
The results demonstrate that the compounds of the present invention have excellent megakaryocyte colony stimulating activity and increase platelets through the 5 activity.

FORMULATION EXAMPLE 1
A granule preparation containing the following
ingredients is prepared.
5 Ingredients
Compound represented by the formula (I) 10 mg
Lactose 700 mg
Corn Starch 274 mg
- HPC-L 16 mg
1000 rag A compound represented by the formula (I) and lactose
are sifted through a 60-mesh sieve. Corn starch is
sifted though a 120-mesh sieve. They are mixed in a V-
type blender. The powder mixture is kneaded with a low-
10 viscosity hydroxypropylcellulose (HPC-L) aqueous solution, granulated (extrusion granulation, die size 0.5-1 mm) and dried. The resulting dry granules are sifted through a shaking sieve (12/60 mesh) to obtain a granule preparation.
15 FORMULATION EXAMPLE 2
A powder preparation for capsulation containing the
following ingredients is prepared.
Ingredients
Compound represented by the formula (I) 10 mg
Lactose 79 mg
Corn Starch 10 mg
Magnesium Stearate 1 mg
100 mg A compound represented by the formula (I) and lactose

are sifted through a 60-mesh sieve. Corn starch is
sifted though a 120-mesh sieve. They are mixed with
magnesium stearate in a V-type blender. The 10% powder
is put in hard gelatin capsules No. 5, 100 mg each.
5 FORMULATION EXAMPLE 3
A granule preparation for capsulation containing the
following ingredients is prepared.
Ingredients
Compound represented by the formula (I) 15 mg
Lactose 90 mg
Corn Starch 42 mg
HPC-L 3 mg
150 mg A compound represented by the formula (I) and lactose
10 are sifted through a 60-mesh sieve. Corn starch is
sifted though a 120-mesh-sieve. They are mixed in a V-type blender. The powder mixture is kneaded with a low-viscosity hydroxypropylcellulose (HPC-L) aqueous solution, granulated and dried. The resulting dry granules are
15 sifted through a shaking sieve {12/60 mesh). The
granules are put in hard capsules No. 4, 150 mg each. FORMULATION EXAMPLE 4
A tablet preparation containing the following ingredients is prepared.
20 Ingredients
Compound represented by the formula (I) 10 mg
Lactose 90 mg
Microcrystalline cellulose 30 mg
Magnesium Stearate 5 mg

CMC-Na 15 mg
150 mg A compound represented by the formula (I), lactose,
microcrystalline cellulose and CMC-Na
(carboxymethylcellulose sodium salt) are sifted through a
60-mesh sieve and mixed. The powder mixture is mixed
s with magnesium stearate to give a bulk powder mixture.
The powder mixture is compressed directly into 150 mg
tablets.
FORMULATION EXAMPLE 5
An intravenous preparation is prepared as follows.
Compound represented by the formula (I) 100 mg
Saturated Fatty Acid Glyceride 1000 mL
10 Solutions having the above-mentioned composition are
usually administered to a patient- intravenously at a rate
of 1 mL per 1 minute.
INDUSTRIAL APPLICABILITY
15 The compounds of the present invention which have affinity for thrombopoietiri receptor and act as thrombopoietin receptor agonists are useful as preventive, therapeutic and improving agents for diseases against which activation of the thrombopoietin receptor is.
20 effective, especially as drugs for hematological
disorders accompanied by abnormal platelet count and as drugs for diseases treated or prevented by stimulating differentiation and proliferation of vascular endothelial cells and endothelial progenitor cells, and are useful as

medicines.
The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is 5 not, and should not be taken as an acknowledgment or admission or any ■ form of suggestion that that prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates. 10
Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of 15 integers or steps but not the exclusion of any other integer or step or group of integers or steps.

. We Claim: t5
1. A compound represented by the formula (I):
(i)
wherein R1 is a phenyl group (the phenyl group may be 5 substituted with one or more C1-6 alkyl groups, one or more C1-3 alkyl groups (the C1-3 alkyl groups are substituted with one or more halogen at.oms) , one or more . C1-3 alkoxy groups (the C1_3 alkoxy groups may be substituted with one or more halogen atoms) or one or
10 more halogen atoms),
R2 is a hydrogen atom or a C1-3 alkyl group (the C1-3 alkyl group may be substituted with one or more halogen atoms), R3 is a phenyl group, a pyridyl group or a thienyl group (the phenyl group, the pyridyl group and the thienyl
15 group" are substituted with one or more substituents selected from the group consisting of hydrogen atoms/ nitro groups, halogen atoms and C1-3 alkyl groups (the C1-3 alkyl groups may be substituted with one or more halogen atoms) and with (C=0)Rs (wherein RS is NR6R7 (wherein R6
20 is a hydrogen atom or a C1-3 alkyl group (the C2-3 alkyl
group may be substituted'with one or more.halogen atoms), and R7 is a Ci-6 alkyl group (the Ci-6 alkyl group may be substituted with one or more halogen atoms, one or more

hydroxyl groups, one or more C1-3 alkoxy groups or one or more C2-i4 aryl groups (the C2-i4 aryl groups may be substituted with one or more Ci_3 alkyl groups, one or * more C1-3 alkoxy groups, one or more carboxyl groups, one 5 or more carbamoyl groups, one or more cyano groups or one or more halogen atoms, and in the case of aryl groups containing one or more nitrogen atoms, may be N-oxides thereof)), a phenyl group, a thienyl group, a pyridyl ' group or a pyridyl-N-oxide group (the phenyl group, the
10 thienyl group, the pyridyl group and the pyridyl-N-oxide group may be substituted with one or more halogen atoms), or NR6R7 is, as a whole, a nitrogen-containing heterocyclyl group (the nitrogen-containing heterocyclyl group may be substituted with one or more hydrogen atoms,
15 one or more Ci_e alkyl groups (the C1-6 alkyl groups may be substituted with one or more halogen atoms), one or more halogen atoms, one or more hydroxyl groups or one or more C1-3 alkoxy groups (the C1-3 alkoxy groups may be substituted with one or more halogen atoms) ) ) or a Ci_6
20 alkyl group {the c1-6 alkyl group may be substituted with one or more halogen atoms, one or more pyridyl groups, one or more pyridyl-N-oxide groups, one or more furyl groups, one or more thienyl groups or one or more phenyl groups and is substituted with one or more cyano
25 groups))), and
R4 is a hydrogen atom or a Ci_3 alkyl group (the C1-3 alkyl group may be substituted with one or more halogen atoms) ,

a tautomer# or phartnaceutically. acceptable salt, of the compound or a solvate thereof.
2. The compound as claimed in - Claim 1, wherein R2 is a
methyl group, and R4 is a hydrogen atom, a tautomer,
5 . or pharmaceutically acceptable salt of the
compound or a solvate thereof.
3. The compound as claimed in Claim 2, wherein R1 is a
3,4-dimethyl-phenyl group, a 4-t-butyl-phenyl group, a 4-trifluoromethyl-phenyl group, a 3-chloro-phenyl group, a
10 4-chloro-phenyl group, a 4-fluoro-phenyl group, a 3,4-dichloro-phenyl group, a 4-bromo-phenyl group or a 4-trifluoromethoxy-phenyl group, a tautomer, or pharmaceutically acceptable salt of the compound or a solvate thereof.
15 4. The compound as claimed in claim 3, wherein R3 is represented by the formula (II) :

(wherein R6 is a methyl group or an ethyl group, and R7 is a Ci-6 alkyl group (the C1-6 alkyl group may be 20 substituted with one or more methoxy groups)), a
tautomer, " or pharmaceutically acceptable salt of the compound or a solvate thereof.
5. The compound as claimed in Claim 3, wherein R3 is represented by the formula (II):

I
L> --* /
(wherein R6 is a methyl group or an ethyl group, and R7 ° is a Ci-3 alkyl group (the Ci_3 alkyl group is substituted with one or more phenyl groups or one or more pyridyl 5 groups)), a tautomer, or pharmaceutically
acceptable salt of the compound or a solvate thereof. 6. The compound as claimed in claim 3, wherein R3 is represented by the formula (II):

10 (wherein R6 is a hydrogen atom, and R7 is a Ci-6 alkyl.
group (the Ci-6 alkyl group is substituted with one or
more methoxy groups) or a pyridyl group), a tautomer, or pharmaceutically acceptable salt of the
compound or a solvate thereof. 15 7. The compound as claimed in1 claim 3, wherein R3 is
represented by the' formula (II) :

(wherein NR6R7 is, as a whole, represented by the formula (III):


(wherein R9 is a Ci_3 alkyl group) ) , a tautomer,
or pharmaceutically acceptable salt of the compound or a
solvate thereof.
S 8. The compound as claimed in Claim 3, wherein R3 is
represented by the formula (IV) :

(wherein R6 is a hydrogen atom, R7 is a C1-3 alkyl group (the C1-3 alkyl group may be substituted with one or more 10 hydroxyl groups) , and R8 is a methyl group or a chlorine
atom), a tautomer, or pharmaceutically acceptable
salt of the compound or a solvate thereof.
9. The compound as claimed in' Claim 3, wherein R3 is
represented by the formula (V):
is
(wherein R10 is a hydrogen atom or a Ci_3 alkyl group) , a tautomer, or pharmaceutically acceptable salt of the compound or a solvate thereof.

10. The compound as claimed in Claim 3, wherein R3 is
represented by the formula (II) : *7i

(wherein R5 is a hydrogen atom, and R7 is an isopropyl 5 group, a methyl group, an ethyl group or a normal propyl group (the methyl group, the ethyl group and the normal propyl group are unsubstituted or substituted with one or more pyridyl groups, one or more pyridyl-N-oxide groups, one or more furyl groups, one or more pyrazinyl groups,
10 one or more imidazolyl groups, one or more pyrazolyl groups or one or more isoxazolyl groups (the pyridyl groups, the pyridyl-N-oxide groups, the furyl groups, the pyrazinyl groups, the imidazolyl groups, the pyrazolyl groups and the isoxazolyl groups may be substituted with
15 one or more methyl groups, one or more methoxy groups, one or more carboxyl groups or one or more halogen atoms))), a tautomer, or pharmaceutically acceptable salt of the compound or a solvate thereof.
11. • The compound as claimed in i.anY °ne of Claims 4 to 10,
20 wherein R1 is a 3,4-dimethyl-phenyl group, a tautomer,
or pharmaceutically acceptable salt of the compound or a solvate thereof.
12. The compound as claimed in, any one of Claims 4 to 10,
wherein R1 is a 3,4-dichloro-phenyl group, a tautomer,
25 or pharmaceutically acceptable salt of the

compound or a solvate thereof.
13. The compound as claimed in'- any one of Claims 4 to 10,
wherein R1 is a 4-chloro-phenyl group, a tautomer,
or pharmaceutically acceptable salt of the
s compound or a solvate thereof.
. >
14. The compound as claimed in any one-of Claims 4 to 10,
wherein R is a 4-trifluoromethyl-phenyl group, a ^
tautomer, or pharmaceutically acceptable salt of the compound or a solvate thereof. io 15. The compound as claimed in any one of Claims 4 to 10,
wherein R1 is a 4-bromo-phenyl group, a tautomer,
or pharmaceutically acceptable salt of the compound or a solvate thereof.
16. The compound as claimed in any one of Claims 4 to 10,
15 wherein R1 is a 4-trifluoromethoxy-phenyl group, a
tautomer, or pharmaceutically acceptable salt of the compound or a solvate thereof.
17. A thrombopoietin receptor activator containing the
compound as claimed in any one of Claims 1 to 16, a
20 tautomer, or pharmaceutically acceptable salt of the compound or a solvate thereof, as an active ingredient.
18.. A preventive, therapeutic or improving agent for diseases against which activation of the thrombopoietin
25 receptor is effective, which contains the thrombopoietin receptor activator as claimed in claim 17, as an active ingredient.

19. A platelet increasing agent containing the
thrombopoietin receptor activator as claimed in Claim 17,
as an active ingredient.
20. Medicament containing the compound as claimed in any
s one of Claims 1 to 16, a tautomer, or
pharmaceutically acceptable salt of the compound or a solvate thereof, as an active ingredient.

ABSTRACT

A compound represented by the.formula (I) (wherein
R1, R2, R3 and R4 are as defined in the description) , a
tautomer, prodrug or pharmaceutically acceptable salt of
5 the compound or a solvate thereof.