Cdc7 kinase inhibitors and uses thereof
11045446 · 2021-06-29
Assignee
Inventors
- Mark G. Frattini (Ridgewood, NJ, US)
- Hakim Djaballah (Scarsdale, NY, US)
- Thomas J. Kelly (New York, NY, US)
Cpc classification
A61P29/00
HUMAN NECESSITIES
A61N5/10
HUMAN NECESSITIES
A61P9/10
HUMAN NECESSITIES
C09B61/00
CHEMISTRY; METALLURGY
A61P43/00
HUMAN NECESSITIES
A61P7/02
HUMAN NECESSITIES
A61P35/00
HUMAN NECESSITIES
C09B13/02
CHEMISTRY; METALLURGY
A61K31/122
HUMAN NECESSITIES
A61K9/0019
HUMAN NECESSITIES
A61P21/00
HUMAN NECESSITIES
C12Q2600/106
CHEMISTRY; METALLURGY
A61K31/352
HUMAN NECESSITIES
International classification
C09B13/02
CHEMISTRY; METALLURGY
A61K9/00
HUMAN NECESSITIES
A61K31/122
HUMAN NECESSITIES
C09B61/00
CHEMISTRY; METALLURGY
A61K31/352
HUMAN NECESSITIES
Abstract
The invention provides compounds, methods, pharmaceutical compositions, and kits for the treatment of proliferative disorders such as cancer. In one aspect, the methods comprise compounds that inhibit the activity of protein kinases, such as cell division cycle (Cdc) kinase. In another aspect, the methods comprise compounds that inhibit Cdc7 and/or Dbf4 activity. In another aspect, the methods comprise compounds that exhibit anti-proliferative properties useful in treating diseases such as cancer. Compounds useful for any of the methods include compounds of the Formula (A) or (B): ##STR00001##
or pharmaceutically acceptable salts thereof. Exemplary compounds of Formula (A) or (B) include granaticin A, granaticin B, dihydrogranaticin A, dihydrogranaticin B, medermycin, and actinorhodin.
Claims
1. A compound of Formula (A) or (B): ##STR00049## or a pharmaceutically acceptable salt thereof, wherein: each instance of R.sup.1 and R.sup.4 is independently selected from the group consisting of hydrogen, carbonyl, silyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; each instance of R.sup.2 and R.sup.3 is independently selected from the group consisting of hydrogen, halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; or R.sup.2 and R.sup.3 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group; R.sup.5 is hydrogen and R.sup.6 is selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; or R.sup.5 and R.sup.6 are joined to form a direct bond; R.sup.7 is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; R.sup.12 is carbonyl, silyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; and each instance of R.sup.13 and R.sup.14 is independently selected from the group consisting of hydrogen, halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; or R.sup.13 and R.sup.14 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group; with the proviso that the compound cannot be: ##STR00050##
2. A pharmaceutical composition for treating a proliferative disorder comprising a pharmaceutically acceptable excipient and a compound of the Formula (A) or (B): ##STR00051## or a pharmaceutically acceptable salt thereof, wherein: each instance of R.sup.1 and R.sup.4 is independently selected from the group consisting of hydrogen, carbonyl, silyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; each instance of R.sup.2 and R.sup.3 is independently selected from the group consisting of hydrogen, halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; or R.sup.2 and R.sup.3 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group; R.sup.5 is hydrogen and R.sup.6 is selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; or R.sup.5 and R.sup.6 are joined to form a direct bond; R.sup.7 is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; R.sup.12 is carbonyl, silyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; and each instance of R.sup.13 and R.sup.14 is independently selected from the group consisting of hydrogen, halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; or R.sup.13 and R.sup.14 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group; with the proviso that the compound cannot be: ##STR00052##
Description
BRIEF DESCRIPTION OF THE DRAWINGS
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(5)
(6)
(7)
(8)
(9)
(10)
(11)
(12)
(13)
(14)
(15)
(16)
(17)
(18)
(19)
(20)
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(22)
DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS OF THE INVENTION
(23) The current invention derives from the discovery that the benzoisochromane quinones (BIQs), such as the granaticins, inhibit protein kinase activity. The granaticins, in particular, were found to inhibit Cdc kinase activity based upon hits identified from high-thoroughput screening (HTS) of over 300,000 compounds for their ability to inhibit a heterodimer of a kinase (Cdc7) and an activator (Dbf4) that phosphorylates serine and threonine residues.
(24) The present invention provides methods for the inhibition of protein kinase activity, such as Cdc kinase activity, and particularly, Cdc7 kinase activity. The inhibitory compounds useful in the methods include, but are not limited to, granaticin A, granaticin B, dihydrogranaticin A, dihydrogranaticin B, medermycin, actinorhodin, benzoisochromanequinones, as shown herein. The methods of the present invention are useful in the treatment of kinase-related diseases or disorders. Specifically, the methods are useful in the treatment of proliferative disorders, such as cancer, benign tumors, inflammatory disorders, and complications thereof. The present invention also provides pharmaceutical compositions and kits for the treatment of various diseases.
(25) Compounds Useful in the Invention
(26) The present invention utilizes compounds of the Formula (A):
(27) ##STR00008##
or pharmaceutically acceptable salts thereof,
wherein:
(28) each instance of R.sup.1 and R.sup.4 is independently selected from the group consisting of hydrogen, carbonyl, silyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl;
(29) each instance of R.sup.2 and R.sup.3 is independently selected from the group consisting of hydrogen, halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; or R.sup.2 and R.sup.3 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group;
(30) R.sup.5 is hydrogen and R.sup.6 is selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; or R.sup.5 and R.sup.6 are joined to form a direct bond; and
(31) R.sup.7 is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl.
(32) In certain embodiments, R.sup.1 is hydrogen. In certain embodiments, R.sup.1 is carbonyl, silyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl.
(33) In certain embodiments, R.sup.4 is hydrogen. In certain embodiments, R.sup.4 is optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl.
(34) In certain embodiments, both R.sup.1 and R.sup.4 are hydrogen, e.g., to provide a compound of the Formula (A-1):
(35) ##STR00009##
or a pharmaceutically acceptable salt thereof,
wherein R.sup.3, R.sup.4, R.sup.5, R.sup.6, and R.sup.7 are as defined herein.
(36) In certain embodiments, R.sup.2 is hydrogen. In certain embodiments, R.sup.2 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.2 is optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.2 is optionally substituted heterocyclyl (e.g., optionally substituted tetrahydropyranyl).
(37) In certain embodiments, R.sup.2 is an optionally substituted tetrahydropyranyl group of the formula (i):
(38) ##STR00010##
wherein:
(39) each instance of R.sup.8 is independently selected from optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, —N.sub.2, carbonyl, silyl, sulfonyl, and sulfinyl; and
(40) m is 0 or an integer of between 1 and 5, inclusive.
(41) In certain embodiments, each instance of R.sup.8 is independently selected from optionally substituted alkyl, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, carbonyl, and silyl. In certain embodiments, each instance of R.sup.7 is independently selected from optionally substituted alkyl, —OH, substituted hydroxyl, —NH.sub.2, and substituted amino. In certain embodiments, each instance of R.sup.8 is independently selected from optionally substituted alkyl, —OH, and substituted amino (e.g., disubstituted amino). In certain embodiments, each instance of R.sup.8 is independently selected from —CH.sub.3, —OH, and —N(CH.sub.3).sub.2.
(42) In certain embodiments, m is an integer of between 1 and 4, inclusive. In certain embodiments, m is an integer of between 1 and 3, inclusive. In certain embodiments, m is an integer of between 1 and 2, inclusive. In certain embodiments, m is 3.
(43) In certain embodiments, each instance of R.sup.8 is independently selected from —CH.sub.3, —OH, and —N(CH.sub.3).sub.2, and m is an integer of between 1 and 3, inclusive. For example, in this instance, in certain embodiments, R.sup.2 is a substituted tetrahydropyranyl group of the formula (ii):
(44) ##STR00011##
(45) In certain embodiments, the substituted tetrahydropyranyl group of the formula (ii) is of the formula (iii):
(46) ##STR00012##
(47) Alternatively, in certain embodiments, R.sup.2 is an optionally substituted aryl or optionally substituted heteroaryl. In certain embodiments, R.sup.2 is an optionally substituted aryl (e.g., a benzoisochromanequinone).
(48) In certain embodiments R.sup.2 is substituted benzoisochromanequinone of the formula (iv):
(49) ##STR00013##
wherein R.sup.1, R.sup.3, R.sup.4, R.sup.5, R.sup.6, and R.sup.7 are as defined herein.
(50) In certain embodiments, the substituted benzoisochromanequinone of formula (iv) is of the formula (v):
(51) ##STR00014##
(52) Alternatively, in certain embodiments, R.sup.2 and R.sup.3 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group. In certain embodiments, R.sup.2 and R.sup.3 are joined to form an optionally substituted carbocyclyl or optionally substituted heterocyclyl group. In certain embodiments, R.sup.2 and R.sup.3 are joined to form an optionally substituted heterocyclyl group (e.g., an optionally substituted 2-oxabicyclo[2.2.2]octenyl group).
(53) In certain embodiments, R.sup.2 and R.sup.3 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (vi):
(54) ##STR00015##
wherein each instance of R.sup.9 and R.sup.10 is independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, carbonyl, silyl, sulfonyl, and sulfinyl.
(55) In certain embodiments, R.sup.9 is hydrogen. In certain embodiments, R.sup.9 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.9 is optionally substituted carbocyclyl or optionally substituted heterocyclyl. In certain embodiments, R.sup.9 is optionally substituted heterocyclyl (e.g., optionally substituted tetrahydropyranyl).
(56) In certain embodiments, R.sup.9 is optionally substituted tetrahydropyranyl group of the formula (vii):
(57) ##STR00016##
wherein:
(58) each instance of R.sup.11 is independently selected from optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, —N.sub.2, carbonyl, silyl, sulfonyl, and sulfinyl; and
(59) p is 0 or an integer of between 1 and 5, inclusive.
(60) In this instance, in certain embodiments, the R.sup.2 and R.sup.3 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (viii):
(61) ##STR00017##
(62) In certain embodiments, each instance of R.sup.11 is independently selected from optionally substituted alkyl, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, carbonyl, and silyl. In certain embodiments, each instance of R.sup.10 is independently selected from optionally substituted alkyl, —OH, and substituted hydroxyl. In certain embodiments, each instance of R.sup.11 is independently selected from optionally substituted alkyl and —OH. In certain embodiments, each instance of R.sup.11 is independently selected from —CH.sub.3 and —OH.
(63) In certain embodiments, p is an integer of between 1 and 4, inclusive. In certain embodiments, p is an integer of between 1 and 3, inclusive. In certain embodiments, p is an integer of between 1 and 2, inclusive. In certain embodiments, p is 2.
(64) In certain embodiments, each instance of R.sup.11 is independently selected from —CH.sub.3 and —OH, and p is an integer of between 1 and 2, inclusive. For example, in this instance, in certain embodiments, R.sup.11 is a substituted tetrahydropyranyl group of the formula (ix):
(65) ##STR00018##
(66) In certain embodiments, the substituted tetrahydropyranyl group of the formula (ix) is of the formula (x):
(67) ##STR00019##
(68) In this instance, in certain embodiments, the R.sup.2 and R.sup.3 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xi):
(69) ##STR00020##
(70) In certain embodiments, R.sup.10 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, carbonyl, silyl, sulfonyl, or sulfinyl.
(71) However, in certain embodiments, R.sup.10 is hydrogen. In this instance, in certain embodiments, the R.sup.2 and R.sup.3 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xii):
(72) ##STR00021##
(73) In certain embodiments, both R.sup.9 and R.sup.10 are hydrogen. In this instance, in certain embodiments, the R.sup.2 and R.sup.3 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xiii):
(74) ##STR00022##
(75) In certain embodiments, R.sup.5 is hydrogen and R.sup.6 is selected from the group consisting of selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl.
(76) In certain embodiments, both R.sup.5 and R.sup.6 are hydrogen. However, in certain embodiments, R.sup.5 is hydrogen and R.sup.6 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl.
(77) Alternatively, in certain embodiments, R.sup.5 and R.sup.6 are joined to form a direct bond, e.g., to provide a dihydrofuran-2-one of the formula (xiv):
(78) ##STR00023##
(79) In certain embodiments, the dihydrofuran-2-one of the formula (xiv) is of the formula (xv):
(80) ##STR00024##
(81) In certain embodiments, the dihydrofuran-2-one of the formula (xiv) is of the formula (xvi):
(82) ##STR00025##
(83) In certain embodiments, R.sup.7 is hydrogen. In certain embodiments, R.sup.7 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl. In certain embodiments, R.sup.7 is optionally substituted alkyl (e.g., —CH.sub.3, —CH.sub.2CH.sub.3, —CH.sub.2CH.sub.2CH.sub.3, —CH(CH.sub.3).sub.2, and the like). In certain embodiments, R.sup.7 is —CH.sub.3.
(84) In certain embodiments, wherein R.sup.5 and R.sup.6 are joined to form a direct bond, i.e., to provide a dihydrofuran-2-one of the formula (xiv), the compound of Formula (A) is of the Formula (A-2):
(85) ##STR00026##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3. In certain embodiments, R.sup.1 and R.sup.4 are hydrogen.
(86) In certain embodiments, wherein R.sup.5 and R.sup.6 are joined to form a dihydrofuran-2-one of the formula (xv), the compound of Formula (A-2) is of the Formula (A-3):
(87) ##STR00027##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3. In certain embodiments, R.sup.1 and R.sup.4 are hydrogen.
(88) In certain embodiments, wherein R.sup.5 and R.sup.6 are joined to form a dihydrofuran-2-one of the formula (xvi), the compound of Formula (A-2) is of the Formula (A-4):
(89) ##STR00028##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3. In certain embodiments, R.sup.1 and R.sup.4 are hydrogen.
(90) In certain embodiments, wherein R.sup.1 and R.sup.4 are hydrogen, the compound of Formula (A-3) is of the Formula (A-5):
(91) ##STR00029##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(92) In certain embodiments, wherein R.sup.1 and R.sup.4 are hydrogen, the compound of Formula (A-4) is of the Formula (A-6):
(93) ##STR00030##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(94) In certain embodiments, wherein R.sup.2 and R.sup.3 are joined to form a substituted oxabicyclo[2.2.2]octenyl group of the formula (vi), the compound of Formula (A-5) is of the Formula (A-7):
(95) ##STR00031##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(96) Exemplary compounds of the Formula (A) include, but are not limited to:
(97) ##STR00032## ##STR00033##
or pharmaceutically acceptable salts thereof.
(98) The present invention also utilizes compounds of the Formula (B):
(99) ##STR00034##
or pharmaceutically acceptable salts thereof,
(100) wherein:
(101) R.sup.12 is hydrogen, carbonyl, silyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl;
(102) each instance of R.sup.13 and R.sup.14 is independently selected from the group consisting of hydrogen, halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl; or R.sup.13 and R.sup.14 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group;
(103) R.sup.5 is hydrogen and R.sup.6 is selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl; or R.sup.5 and R.sup.6 are joined to form a direct bond; and
(104) R.sup.7 is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl.
(105) In certain embodiments, R.sup.12 is hydrogen. In certain embodiments, R.sup.12 is carbonyl, silyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl.
(106) In certain embodiments, R.sup.13 is hydrogen. In certain embodiments, R.sup.13 is halogen, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfinyl, sulfonyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl. In certain embodiments, R.sup.13 is —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino. In certain embodiments, R.sup.13 is —OH or substituted hydroxyl. In certain embodiments, R.sup.13 is —OH.
(107) In certain embodiments, R.sup.14 is hydrogen. In certain embodiments, R.sup.14 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.14 is optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.14 is optionally substituted heterocyclyl (e.g., optionally substituted tetrahydropyranyl).
(108) In certain embodiments, R.sup.14 is an optionally substituted tetrahydropyranyl group of the formula (xvii):
(109) ##STR00035##
wherein:
(110) each instance of R.sup.15 is independently selected from optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, —CN, —NO.sub.2, carbonyl, silyl, sulfonyl, and sulfinyl; and
(111) q is 0 or an integer of between 1 and 5, inclusive.
(112) In certain embodiments, each instance of R.sup.15 is independently selected from optionally substituted alkyl, —OH, substituted hydroxyl, —SH, substituted thiol, —NH.sub.2, substituted amino, carbonyl, and silyl. In certain embodiments, each instance of R.sup.15 is independently selected from optionally substituted alkyl, —OH, substituted hydroxyl, —NH.sub.2, and substituted amino. In certain embodiments, each instance of R.sup.15 is independently selected from optionally substituted alkyl, —OH, and substituted hydroxyl. In certain embodiments, each instance of R.sup.15 is independently selected from —CH.sub.3 and —OH.
(113) In certain embodiments, q is an integer of between 1 and 4, inclusive. In certain embodiments, q is an integer of between 1 and 3, inclusive. In certain embodiments, q is an integer of between 1 and 2, inclusive. In certain embodiments, q is 3.
(114) In certain embodiments, each instance of R.sup.15 is independently selected from —CH.sub.3, —OH, and m is an integer of between 1 and 3, inclusive. For example, in this instance, in certain embodiments, R.sup.15 is a substituted tetrahydropyranyl group of the formula (xviii):
(115) ##STR00036##
(116) Alternatively, in certain embodiments, R.sup.13 and R.sup.14 are joined to form an optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl group. In certain embodiments, R.sup.13 and R.sup.14 are joined to form an optionally substituted carbocyclyl or optionally substituted heterocyclyl group. In certain embodiments, R.sup.13 and R.sup.14 are joined to form an optionally substituted heterocyclyl group (e.g., an optionally substituted 2-oxabicyclo[2.2.2]octenyl group).
(117) In certain embodiments, R.sup.13 and R.sup.14 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xix):
(118) ##STR00037##
wherein each instance of R.sup.16 and R.sup.17 is independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, carbonyl, silyl, sulfonyl, and sulfinyl.
(119) In certain embodiments, R.sup.16 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.16 is optionally substituted carbocyclyl or optionally substituted heterocyclyl. However, in certain embodiments, R.sup.16 is hydrogen, e.g., and R.sup.13 and R.sup.14 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xx):
(120) ##STR00038##
(121) In certain embodiments, R.sup.17 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, or optionally substituted heteroaryl. In certain embodiments, R.sup.17 is optionally substituted carbocyclyl or optionally substituted heterocyclyl. However, in certain embodiments, R.sup.17 is hydrogen, e.g., and R.sup.13 and R.sup.14 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xxi):
(122) ##STR00039##
(123) In certain embodiments, both R.sup.16 and R.sup.17 are hydrogen. In this instance, in certain embodiments, the R.sup.13 and R.sup.14 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xxii):
(124) ##STR00040##
(125) In certain embodiments, R.sup.5 is hydrogen and R.sup.6 is selected from the group consisting of selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl.
(126) In certain embodiments, both R.sup.5 and R.sup.6 are hydrogen. However, in certain embodiments, R.sup.5 is hydrogen and R.sup.6 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl.
(127) Alternatively, in certain embodiments, R.sup.5 and R.sup.6 are joined to form a direct bond, e.g., to provide a dihydrofuran-2-one of the formula (xiv):
(128) ##STR00041##
(129) In certain embodiments, the dihydrofuran-2-one of the formula (xiv) is of the formula (xv):
(130) ##STR00042##
(131) In certain embodiments, the dihydrofuran-2-one of the formula (xiv) is of the formula (xvi):
(132) ##STR00043##
(133) In certain embodiments, R.sup.7 is hydrogen. In certain embodiments, R.sup.7 is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl. In certain embodiments, R.sup.7 is optionally substituted alkyl (e.g., —CH.sub.3, —CH.sub.2CH.sub.3, —CH.sub.2CH.sub.2CH.sub.3, —CH(CH.sub.3).sub.2, and the like). In certain embodiments, R.sup.7 is —CH.sub.3.
(134) In certain embodiments, wherein R.sup.12 is hydrogen, the compound of Formula (B) is of the Formula (B-1):
(135) ##STR00044##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(136) In certain embodiments, wherein R.sup.5 and R.sup.6 are joined to form a direct bond, the compound of Formula (B-1) is of the Formula (B-2):
(137) ##STR00045##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(138) In certain embodiments, wherein R.sup.13 and R.sup.14 are joined to form a substituted 2-oxabicyclo[2.2.2]octenyl group of the formula (xix), the compound of Formula (B-1) is of the Formula (B-3):
(139) ##STR00046##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(140) In certain embodiments, wherein R.sup.13 is —OH and R.sup.14 is an optionally substituted tetrahydropyranyl group of the formula (xvii), the compound of Formula (B-1) is of the Formula (B-4):
(141) ##STR00047##
or a pharmaceutically acceptable salt thereof. In certain embodiments, R.sup.7 is —CH.sub.3.
(142) Exemplary compounds of the Formula (B) include, but are not limited to:
(143) ##STR00048##
or pharmaceutically acceptable salts thereof.
(144) In certain embodiments, the compounds of Formula (A) and (B) have an IC.sub.50 of less than approximately 100 μM, e.g., less than approximately 10 μM, e.g., less than approximately 1 μM, e.g., less than approximately 0.1 μM, or e.g., less than approximately 0.01 μM.
(145) The compounds described herein may be useful in methods, pharmaceutical compositions, and kits, for example, in the treatment and prevention of proliferative disorders, such as cancer, benign tumors, inflammatory disorders, and complications thereof.
(146) Protein Kinases, Cdc7, and Dbf4
(147) Compounds useful in the present invention are protein kinase inhibitors. The protein kinases constitute a group of enzymes that catalyse the phosphorylation of hydroxy groups of specific protein residues such as tyrosine, serine or threonine residues. Said phosphorylations can extensively modify the function of proteins; thus, the protein kinases play an important role in the regulation of a great variety of cellular processes, notably including metabolism, cellular proliferation, cell differentiation or cell survival. Among the various cellular functions in which the activity of a protein kinase is involved, certain processes represent attractive targets for treating certain diseases. As an example, we may notably mention angiogenesis and control of the cell cycle, in which the protein kinases can play an essential role. These processes are essential for the growth of solid tumours as well as for the development of other diseases.
(148) The protein kinases participate in signalling events that control the activation, growth and differentiation of cells in response either to extracellular mediators, or to changes in the environment. In general, these kinases belong to two groups: those which phosphorylate serine and/or threonine residues preferentially, and those which phosphorylate tyrosine residues preferentially (S. K. Hanks and T. Hunter, FASEB. J., 1995, 9, pages 576-596). The serine/threonine kinases, for example, are isoforms of protein kinases C (A. C. Newton, J. Biol. Chem., 1995, 270, pages 28495-28498) and a group of cyclin-dependent kinases, such as Cdc2 (Cdk1) (J. Pines, Trends in Biochemical Sciences, 1995, 18, pages 195-197). The tyrosine kinases include growth factor receptors, such as the epidermal growth factor (EGF) receptor (S. Iwashita and M. Kobayashi, Cellular Signalling, 1992, 4, pages 123-132), and cytosolic kinases such as p56tck, p59fYn, ZAP-70 and the csk kinases (C. Chan et al., Ann. Rev. Immunol., 1994, 12, pages 555-592).
(149) Abnormally high levels of protein kinase activity have been associated with many diseases, resulting in abnormal cellular functions. This can occur either directly or indirectly, from a malfunction in the control mechanisms of kinase activity, connected for example with a mutation, overexpression or inappropriate activation of the enzyme, or with overproduction or underproduction of cytokines or of growth factors, which are also involved in signal transduction upstream or downstream of the kinases. In all these cases, selective inhibition of the action of the kinases offers hope of a beneficial effect. Among these protein kinases, we may mention quite particularly protein kinase Cdc7. Cdc7 is a serine/threonine kinase that has been characterized at the molecular level as a factor that is essential for initiating DNA replication.
(150) The catalytic activity of Cdc7, which is conserved throughout the eukaryotes, is dependent on its Dbf4 regulatory subunit. Although the expression of Cdc7 (at the level of messenger and protein RNA) is constant throughout the cell cycle, the level of expression of Dbf4 is in contrast dependent on the cell cycle, which induces an increase in Cdc7 kinase activity during the G1-S transition. For this reason, Cdc7 is given the designation DDK (Dbf-4-dependent kinase).
(151) The principal activity of the Cdc7/Dbf4 complex occurs on initiation of DNA replication during the S phase. It phosphorylates MCM2 which thus activates the MCM (MiniChromosome Maintenance) complex, which is an essential component of DNA-helicase activity.
(152) Cdc7 also plays an important role in mutagenesis, mainly induced by action at the level of the DNA-damage pathways and checkpoints, in particular at the ATR-dependent checkpoint, which prevents the initiation of DNA replication in response to damage of the single-stranded type caused by chemical agents such as etoposide.
(153) Cdc7 and Dbf4 are overexpressed in human tumour cell lines and in many tumour samples (lung, breast, thyroid, colon-rectum, oesophagus, uterus, testicle, liver) in comparison with the corresponding normal tissues (Hess, G. F., Drong, R. F., et al. 1998).
(154) Experiments in suppressing Cdc7 expression using RNA interference (RNAi) technology show that inhibition of Cdc7 expression induces arrest of the cell cycle and prevents cellular proliferation of the human tumour cell lines HeLa and HCT116, but has a limited effect on normal cells (normal human skin fibroblasts). This is reflected in a prolonged stoppage in G1 that induces apoptosis in cells lacking p53 (>50% of tumours) but is reversible in normal cells [A. Montagnoli et al., CANCER RESEARCH 64, 7110-7116, Oct. 1, 2004].
(155) The inhibitors of Cdc7 kinase activity can constitute a novel category of targeted cytotoxic therapy as well as of inhibitors of DNA replication. Such inhibitors would inhibit replication before the replication forks are established, thus blocking replication without damaging the DNA. The present application thus relates in particular to novel inhibitors of protein kinase Cdc7 that can be used notably for the treatment of abnormal cellular proliferation and more particularly in oncology.
(156) Methods of Treatment
(157) The methods involve uses of a compound of Formula (A) or (B), or pharmaceutically acceptable salt or pharmaceutical composition thereof.
(158) In one aspect, the present invention provides a compound of Formula (A) or (B), or a pharmaceutically acceptable salt thereof, for use in the treatment or prevention of a proliferative disorder.
(159) In another aspect, the present invention provides use of a compound of Formula (A) or (B), or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for treating or preventing a proliferative disorder.
(160) In yet another aspect, the present invention provides a method for treating or preventing a proliferative disorder in a subject in need thereof, wherein the subject is administered a therapeutically effective amount of a compound of the Formula (A) or (B), or a pharmaceutically acceptable salt thereof.
(161) In certain embodiments, the subject is human.
(162) In certain embodiments, the compound induces apoptosis. In certain embodiments, compound induces S-phase arrest.
(163) In certain embodiments, the compound is an inhibitor of a protein kinase; and wherein the inhibition of a protein kinase is useful for the treatment or prevention of the proliferative disorder. In certain embodiments, the protein kinase is Cdc7 kinase or the Dbf4 regulatory subunit of Cdc7 kinase, and wherein the inhibition of Cdc7 kinase or the Dbf4 regulatory subunit of Cdc7 kinase is useful for the treatment or prevention of the proliferative disorder. In some embodiments, the protein kinase inhibitor is a Cdc7 kinase inhibitor or the Dbf4 regulatory subunit of Cdc7 kinase.
(164) In certain embodiments, the proliferative disorder is selected from the group consisting of cancer, myeloproliferative disorders (e.g., polycythemia vera, essential thrombocytosis, meylofibrosis, and chronic myelogenous leukemia (CML)), benign prostate hyperplasia, familial adenomatosis, polyposis, neuro-fibromatosis, psoriasis, vascular smooth cell proliferation associated with atherosclerosis, fibrotic disorders, pulmonary fibrosis, arthritis, rheumatoid arthritis, glomerulonephritis, and post-surgical stenosis, restenosis, disorders of proliferation of blood vessels, disorders of proliferation of mesangial cells, metabolic disorders, allergies, asthmas, thromboses, diseases of the nervous system, retinopathy, diabetes, and muscular degeneration. In certain embodiments, the proliferative disorder is cancer.
(165) In certain embodiments, the cancer is selected from the group consisting of cancer of the bone, brain, connective tissue, endocrine glands, adrenal cortex, endometrium, germ cells, head and neck, larynx and hypopharynx, mesothelioma, muscle, rectum, renal, small intestine, soft tissue, testis, ureter, vagina, and vulva; bladder cancer; breast cancer; colon cancer; kidney cancer; liver cancer; lung cancer; esophagus cancer; gallbladder cancer; ovarian cancer; pancreatic cancer; stomach cancer; cervical cancer; thyroid cancer; prostate cancer; papillary thyroid carcinoma; genitourinary malignancies; retinoblastoma; Wilms tumor; myelodysplastic syndrome; plasma cell neoplasia; paraneoplastic syndromes; renal cell carcinoma; Ewing's sarcoma; desmoplastic small round cell tumors; mesothelioma; skin cancer, wherein said skin cancer is squamous cell carcinoma; hematologic cancers [e.g., hematopoietic cancers of lymphoid lineage, wherein said cancers are leukemia, acute lymphocytic leukemia (ALL), acute lymphoblastic leukemia, lymphoma, B-cell lymphoma, T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma (e.g., mantle cell lymphoma (MCL), hairy cell lymphoma, and Burkitt's lymphoma; chronic lymphocytic leukemia (CLL); hematopoietic cancers of myeloid lineage, wherein said cancers are multiple myeloma, chronic myeloid leukemia (CML) and acute myeloid leukemia (AML) (e.g., acute megakaryoblastic leukemia (AMKL); myelodysplastic syndrome and promyelocytic leukemia]; tumors of mesenchymal origin, wherein the tumors are fibrosarcoma and rhabdomyosarcoma; tumors of the central and peripheral nervous system, wherein said tumors are astrocytoma, neuroblastoma, glioma (e.g., glioblastoma) and schwannomas; and other tumors, wherein said tumors are melanoma, cutaneous melanoma, seminoma, teratocarcinoma, osteosarcoma, xeroderma pegmentosum, keratoxanthoma, thyroid follicular cancer, Kaposi's sarcoma, cancers of unknown primary site; solid tumors, hematologic cancers, and AIDS-related malignancies.
(166) In certain embodiments, the cancer is a multi-drug resistant (MDR) cancer.
(167) In certain embodiments, the cancer is relapsed and/or refractory cancer. Relapsed cancer refers to a cancer which has returned after a patient has enjoyed a remission. Refractory cancer refers to a cancer which does not respond to other therapies or therapeutic agents. In certain embodiments, the cancer is resistant to (i.e., does not respond to) therapies or chemotherapeutic agents. In certain embodiments, the hematologic cancer is resistant to therapies or chemotherapeutic agents.
(168) In certain embodiments, the cancer is leukemia, lymphoma, melanoma, cancer of the breast, stomach, ovaries, colon, rectum, lung, brain, larynx, lymphatic system, thyroid, oesophagus, liver, uterus, testis, bladder, prostate, bones or pancreas. In certain embodiments, the cancer is leukemia, cancer of the breast, the colon or the lung.
(169) In certain embodiments, the cancer is a hematologic cancer. In certain embodiments, the compound inhibits the growth of hematologic cancers. In certain embodiments, the compound inhibits the growth of hematologic cancers with IC50s in the nanomolar range. In certain embodiments, the hematologic cancer is a hematopoietic cancer of lymphoid lineage. In certain embodiments, the hematologic cancer is a relapsed and/or refractory hematopoietic cancer of lymphoid lineage. In certain embodiments, the cancer is refractory to multiple cycles of cancer therapy (e.g., including allogenic bone marrow transplantation). In certain embodiments, the hematologic cancer is relapsed and/or refractory ALL, CLL, AML, or CML.
(170) In certain embodiments, the cancer is ovarian cancer. In certain embodiments, the ovarian cancer is ovarian carcinoma.
(171) In certain embodiments, the cancer is lung cancer. In certain embodiments, the lung cancer is non-small cell lung cancer. In certain embodiments, the lung cancer is small cell lung cancer. In certain embodiments, the lung cancer is lung adenocarcinoma.
(172) In certain embodiments, the cancer is prostate cancer.
(173) In certain embodiments, the cancer is renal cell carcinoma.
(174) In certain embodiments, the cancer is cervical cancer. In certain embodiments, the cervical cancer is cervical adenocarcinoma. In certain embodiments, the cervical cancer is positive for human papillomavirus (HPV) infection.
(175) In certain embodiments, the cancer is glioblastoma.
(176) In certain embodiments, the cancer is retinoblastoma.
(177) In certain embodiments, the cancer is rhabdomyosarcoma.
(178) In certain embodiments, the cancer is a desmoplastic small round cell tumor.
(179) In certain embodiments, the cancer is breast cancer. In certain embodiments, the breast cancer is breast ductal carcinoma. In certain embodiments, the breast cancer is breast adenocarcinoma. In certain embodiments, the breast cancer is metastatic breast adenocarcinoma. In certain embodiments, the breast cancer is HER2 negative. In certain embodiments, the breast cancer is HER2 positive. In certain embodiments, the breast cancer is NEU receptor negative.
(180) In certain embodiments, the cancer is mesothelioma.
(181) In certain embodiments, the cancer is melanoma.
(182) In certain embodiments, the cancer is thyroid carcinoma.
(183) In certain embodiments, the cancer is Ewing's sarcoma.
(184) In certain embodiments, the cancer is a solid tumor. In certain embodiments, the compound inhibits the growth of solid tumors. In certain embodiments, the compound inhibits the growth of solid tumors with IC50s in the nanomolar range.
(185) In certain embodiments, the cancer comprises a genetic mutation.
(186) In certain embodiments, the cancer comprises a RAS mutation. In certain embodiments, the cancer comprises wild-type RAS.
(187) In certain embodiments, cancer comprises an EGFR mutation. In certain embodiments, the EGFR mutation is an L858R EGFR mutation. In certain embodiments, the EGFR mutation is an DelE746 EGFR mutation. In certain embodiments, the EGFR mutation is an DelE746-A750 EGFR mutation. In certain embodiments, the EGFR mutation is an DelE746-E749 EGFR mutation. In certain embodiments, the EGFR mutation is an T790M EGFR mutation. In certain embodiments, the EGFR mutation is an T790ML858R EGFR mutation. In certain embodiments, the cancer comprises wild-type EGFR.
(188) In certain embodiments, the cancer comprises a KRAS mutation. In certain embodiments, the cancer comprises a G13C KRAS mutation. In certain embodiments, the cancer comprises a G12C KRAS mutation. In certain embodiments, the cancer comprises a G12C KRAS mutation. In certain embodiments, the cancer comprises a Q61H KRAS mutation. In certain embodiments, the cancer comprises wild-type KRAS.
(189) In certain embodiments, the cancer comprises a p53 mutation. In certain embodiments, the p53 mutation is a R273H p53 mutation. In certain embodiments, the p53 mutation is a G262V p53 mutation. In certain embodiments, the p53 mutation is a G16L p53 mutation. In certain embodiments, the p53 mutation is a C176F p53 mutation. In certain embodiments, the p53 mutation is a M246I p53 mutation. In certain embodiments, the cancer comprises wild-type p53.
(190) In certain embodiments, the cancer comprises a BRAF mutation. In certain embodiments, the BRAF mutation is a BRAF V600E mutation.
(191) In certain embodiments, the cancer comprises a EVI1 mutation.
(192) In certain embodiments, the cancer comprises a Flt-3 mutation.
(193) In certain embodiments, the cancer comprises a WT-1 mutation.
(194) In certain embodiments, the cancer comprises a cyclin D mutation.
(195) In certain embodiments, the cancer comprises a PTEN mutation.
(196) In certain embodiments, the cancer comprises a ABL kinase mutation.
(197) In certain embodiments, the mutation comprises a chromosomal abnormality. In certain embodiments, the chromosomal abnormality is a chromosome deletion or inversion. In certaine embodiments, the cancer comprises a chromosome 17p deletion. In certain embodiments, the cancer comprises an inversion of chromosome 16. In certain embodiments, the cancer comprises a trisomy of chromosome 8. In certain embodiments, the cancer comprises a monosomy of chromosome 7. In certain embodiments, the cancer comprises a chromosome 11q23 abnormality. In certain embodiments, the cancer comprises a Philadelphia chromosome positive abnormality.
(198) In certain embodiments, the cancer comprises a fusion transcript. In certain embodiments, the fusion transcript is a reciprocal ASPL-TFE3 fusion transcript.
(199) Assays
(200) In some embodiments, the present invention provides a method of determining dose response using a cytotoxicity assay. In some embodiments, the present invention includes a method of performing dose response studies comprising the steps of providing a test compound, contacting the test compound with a cell, and incubating the cell with the compound under suitable conditions to determine the cytotoxicity of the compound. The antiproliferative activity of the test compound can then be assessed using a method known to those of ordinary skill in the art. This process can then be repeated using different concentrations of a test compound in order to calculate the IC.sub.50. In certain embodiments, the test compound is an compound. In certain embodiments, the cells are retinoblastoma cells.
(201) In certain embodiments, the cells are incubated with a test compound (e.g., an compound) for approximately 1 minute to approximately 1 week. In certain embodiments, the cells are incubated with a test compound for approximately 1 hour to approximately 1 week. In certain embodiments, the cells are incubated with a test compound for approximately 12 hours to approximately 1 week. In certain embodiments, the cells are incubated with a test compound for approximately 24 hours to approximately 1 week. In certain embodiments, the cells are incubated with a test compound for approximately 36 hours to approximately 1 week. In certain embodiments, the cells are incubated with a test compound for approximately 48 hours to approximately 1 week. In certain embodiments, the cells are incubated with a test compound for approximately 48 hours to approximately 120 hours. In certain embodiments, the cells are incubated with a test compound for approximately 48 hours to approximately 96 hours. In certain embodiments, the cells are incubated with a test compound for approximately 62 hours to approximately 82 hours. In certain embodiments, the cells are incubated with a test compound for approximately 72 hours. In certain embodiments, the cells are incubated with a test compound for approximately 1, 2, 3, 4, 5, 6, or 7 days.
(202) In certain embodiments, after a specified amount of time (e.g., 72 hours) an indicator of cell viability (e.g., Alamar Blue) is added, and the mixture is incubated for an additional period of time. In some embodiments, this additional period of time ranges from approximately 1 hour to approximately 48 hours. In some embodiments, this additional period of time ranges from approximately 12 hour to approximately 36 hours. In some embodiments, this additional period of time is approximately 24 hours.
(203) The inhibition of cell proliferation may be measured using methods or technology known in the art. In some embodiments, inhibition of cell proliferation is measured using a substance which produces a detectable signal that is proportional to the amount of inhibition of cell proliferation. In some embodiments, inhibition of cell proliferation is quantified using one of any indicators known to those of ordinary skill in the art that produces a quantifiable signal, the intensity of which is detectable and proportional to the amount of inhibition. In some embodiments, inhibition of cell proliferation is quantified using an indicator which fluoresces. Exemplary indicators include Tyramide-Alexa Fluor 488, Alamar Blue, etc.
(204) In some embodiments, the efficacy of a compound is measured by measuring tumor size over a period of time before, during, and/or after treatment with said compound. In some embodiments, tumor size is measured once a week. In some embodiments, tumor size is measured twice a week. In some embodiments, tumor size is measured daily. In some embodiments, tumor size is measured once a day. In some embodiments, tumor size is measured twice a day. In some embodiments, tumor size is measured once every other day. In some embodiments, tumor size is measured once every three days. In certain embodiments, tumor size is measured at intervals as required by any one of the methods known to those of skill in the art. In some embodiments, tumor size is measured externally twice a week with a caliper. In certain embodiments, tumor size is measured once a week using an imaging technique (e.g., MRI, X-ray, CT). In some embodiments, the imaging technique is bioluminescent imaging. In certain embodiments, bioluminescent imaging comprises anesthetization of the host animal, injection of a bioluminescent compound, and subsequent measurement of photonic emission. In some embodiments, imaging of the tumor is achieved using any of the methods known in the medical arts.
(205) As detailed herein, in assays to determine the ability of a compound (e.g., an compound) to inhibit cancer cell growth certain compounds may exhibit IC.sub.50 values ≤100 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤50 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤40 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤30 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤20 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤10 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤7.5 μM. In certain embodiments, the compound exhibits IC.sub.50 values ≤5 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤2.5 μM. In certain embodiments, the compound exhibits IC.sub.50 values ≤1 μM. In certain embodiments, the compound exhibits IC.sub.50 values ≤0.75 μM. In certain embodiments, the compound exhibits IC.sub.50 values ≤0.5 μM. In certain embodiments, the compound exhibits IC.sub.50 values ≤0.25 μM. In certain embodiments, the compound exhibits IC.sub.50 values ≤0.1 μM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤75 nM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤50 nM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤25 nM. In certain other embodiments, the compound exhibits IC.sub.50 values ≤10 nM. In other embodiments, the compound exhibits IC.sub.50 values ≤7.5 nM. In other embodiments, the compound exhibits IC.sub.50 values ≤5 nM.
(206) Pharmaceutical Compositions
(207) Certain aspects of the invention include use of a pharmaceutical composition in any of the above methods, comprising a compound of Formula (A) or (B) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.
(208) Pharmaceutically acceptable excipients include any and all solvents, diluents or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like, as suited to the particular dosage form desired. General considerations in the formulation and/or manufacture of pharmaceutical compositions agents can be found, for example, in Remington's Pharmaceutical Sciences, Sixteenth Edition, E. W. Martin (Mack Publishing Co., Easton, Pa., 1980), and Remington: The Science and Practice of Pharmacy, 21.sup.st Edition (Lippincott Williams & Wilkins, 2005).
(209) Pharmaceutical compositions described herein can be prepared by any method known in the art of pharmacology. In general, such preparatory methods include the steps of bringing the active ingredient into association with a carrier and/or one or more other accessory ingredients, and then, if necessary and/or desirable, shaping and/or packaging the product into a desired single- or multi-dose unit.
(210) Pharmaceutical compositions can be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses. As used herein, a “unit dose” is discrete amount of the pharmaceutical composition comprising a predetermined amount of the active ingredient. The amount of the active ingredient is generally equal to the dosage of the active ingredient which would be administered to a subject and/or a convenient fraction of such a dosage such as, for example, one-half or one-third of such a dosage.
(211) Relative amounts of the active ingredient, the pharmaceutically acceptable carrier, and/or any additional ingredients in a pharmaceutical composition of the invention will vary, depending upon the identity, size, and/or condition of the subject treated and further depending upon the route by which the composition is to be administered. By way of example, the composition may comprise between 0.1% and 100% (w/w) active ingredient.
(212) Pharmaceutically acceptable excipients used in the manufacture of provided pharmaceutical compositions include inert diluents, dispersing and/or granulating agents, surface active agents and/or emulsifiers, disintegrating agents, binding agents, preservatives, buffering agents, lubricating agents, and/or oils. Excipients such as cocoa butter and suppository waxes, coloring agents, coating agents, sweetening, flavoring, and perfuming agents may also be present in the composition.
(213) Exemplary diluents include calcium carbonate, sodium carbonate, calcium phosphate, dicalcium phosphate, calcium sulfate, calcium hydrogen phosphate, sodium phosphate lactose, sucrose, cellulose, microcrystalline cellulose, kaolin, mannitol, sorbitol, inositol, sodium chloride, dry starch, cornstarch, powdered sugar, etc., and combinations thereof.
(214) Exemplary granulating and/or dispersing agents include potato starch, corn starch, tapioca starch, sodium starch glycolate, clays, alginic acid, guar gum, citrus pulp, agar, bentonite, cellulose and wood products, natural sponge, cation-exchange resins, calcium carbonate, silicates, sodium carbonate, cross-linked poly(vinyl-pyrrolidone) (crospovidone), sodium carboxymethyl starch (sodium starch glycolate), carboxymethyl cellulose, cross-linked sodium carboxymethyl cellulose (croscarmellose), methylcellulose, pregelatinized starch (starch 1500), microcrystalline starch, water insoluble starch, calcium carboxymethyl cellulose, magnesium aluminum silicate (Veegum), sodium lauryl sulfate, quaternary ammonium compounds, etc., and combinations thereof.
(215) Exemplary surface active agents and/or emulsifiers include natural emulsifiers (e.g. acacia, agar, alginic acid, sodium alginate, tragacanth, chondrux, cholesterol, xanthan, pectin, gelatin, egg yolk, casein, wool fat, cholesterol, wax, and lecithin), colloidal clays (e.g. bentonite [aluminum silicate] and Veegum [magnesium aluminum silicate]), long chain amino acid derivatives, high molecular weight alcohols (e.g. stearyl alcohol, cetyl alcohol, oleyl alcohol, triacetin monostearate, ethylene glycol distearate, glyceryl monostearate, and propylene glycol monostearate, polyvinyl alcohol), carbomers (e.g. carboxy polymethylene, polyacrylic acid, acrylic acid polymer, and carboxyvinyl polymer), carrageenan, cellulosic derivatives (e.g. carboxymethylcellulose sodium, powdered cellulose, hydroxymethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, methylcellulose), sorbitan fatty acid esters (e.g. polyoxyethylene sorbitan monolaurate [Tween 20], polyoxyethylene sorbitan [Tween 60], polyoxyethylene sorbitan monooleate [Tween 80], sorbitan monopalmitate [Span 40], sorbitan monostearate [Span 60], sorbitan tristearate [Span 65], glyceryl monooleate, sorbitan monooleate [Span 80]), polyoxyethylene esters (e.g. polyoxyethylene monostearate [Myrj 45], polyoxyethylene hydrogenated castor oil, polyethoxylated castor oil, polyoxymethylene stearate, and Solutol), sucrose fatty acid esters, polyethylene glycol fatty acid esters (e.g. Cremophor), polyoxyethylene ethers, (e.g. polyoxyethylene lauryl ether [Brij 30]), poly(vinyl-pyrrolidone), diethylene glycol monolaurate, triethanolamine oleate, sodium oleate, potassium oleate, ethyl oleate, oleic acid, ethyl laurate, sodium lauryl sulfate, Pluronic F 68, Poloxamer 188, cetrimonium bromide, cetylpyridinium chloride, benzalkonium chloride, docusate sodium, etc. and/or combinations thereof.
(216) Exemplary binding agents include starch (e.g. cornstarch and starch paste), gelatin, sugars (e.g. sucrose, glucose, dextrose, dextrin, molasses, lactose, lactitol, mannitol, etc.), natural and synthetic gums (e.g. acacia, sodium alginate, extract of Irish moss, panwar gum, ghatti gum, mucilage of isapol husks, carboxymethylcellulose, methylcellulose, ethylcellulose, hydroxyethylcellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, microcrystalline cellulose, cellulose acetate, poly(vinyl-pyrrolidone), magnesium aluminum silicate (Veegum), and larch arabogalactan), alginates, polyethylene oxide, polyethylene glycol, inorganic calcium salts, silicic acid, polymethacrylates, waxes, water, alcohol, etc., and/or combinations thereof.
(217) Exemplary preservatives include antioxidants, chelating agents, antimicrobial preservatives, antifungal preservatives, alcohol preservatives, acidic preservatives, and other preservatives.
(218) Exemplary antioxidants include alpha tocopherol, ascorbic acid, acorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene, monothioglycerol, potassium metabisulfite, propionic acid, propyl gallate, sodium ascorbate, sodium bisulfite, sodium metabisulfite, and sodium sulfite.
(219) Exemplary chelating agents include ethylenediaminetetraacetic acid (EDTA) and salts and hydrates thereof (e.g., sodium edetate, disodium edetate, trisodium edetate, calcium disodium edetate, dipotassium edetate, and the like), citric acid and salts and hydrates thereof (e.g., citric acid monohydrate), fumaric acid and salts and hydrates thereof, malic acid and salts and hydrates thereof, phosphoric acid and salts and hydrates thereof, and tartaric acid and salts and hydrates thereof. Exemplary antimicrobial preservatives include benzalkonium chloride, benzethonium chloride, benzyl alcohol, bronopol, cetrimide, cetylpyridinium chloride, chlorhexidine, chlorobutanol, chlorocresol, chloroxylenol, cresol, ethyl alcohol, glycerin, hexetidine, imidurea, phenol, phenoxyethanol, phenylethyl alcohol, phenylmercuric nitrate, propylene glycol, and thimerosal.
(220) Exemplary antifungal preservatives include butyl paraben, methyl paraben, ethyl paraben, propyl paraben, benzoic acid, hydroxybenzoic acid, potassium benzoate, potassium sorbate, sodium benzoate, sodium propionate, and sorbic acid.
(221) Exemplary alcohol preservatives include ethanol, polyethylene glycol, phenol, phenolic compounds, bisphenol, chlorobutanol, hydroxybenzoate, and phenylethyl alcohol.
(222) Exemplary acidic preservatives include vitamin A, vitamin C, vitamin E, beta-carotene, citric acid, acetic acid, dehydroacetic acid, ascorbic acid, sorbic acid, and phytic acid.
(223) Other preservatives include tocopherol, tocopherol acetate, deteroxime mesylate, cetrimide, butylated hydroxyanisol (BHA), butylated hydroxytoluened (BHT), ethylenediamine, sodium lauryl sulfate (SLS), sodium lauryl ether sulfate (SLES), sodium bisulfite, sodium metabisulfite, potassium sulfite, potassium metabisulfite, Glydant Plus, Phenonip, methylparaben, Germall 115, Germaben II, Neolone, Kathon, and Euxyl. In certain embodiments, the preservative is an anti-oxidant. In other embodiments, the preservative is a chelating agent.
(224) Exemplary buffering agents include citrate buffer solutions, acetate buffer solutions, phosphate buffer solutions, ammonium chloride, calcium carbonate, calcium chloride, calcium citrate, calcium glubionate, calcium gluceptate, calcium gluconate, D-gluconic acid, calcium glycerophosphate, calcium lactate, propanoic acid, calcium levulinate, pentanoic acid, dibasic calcium phosphate, phosphoric acid, tribasic calcium phosphate, calcium hydroxide phosphate, potassium acetate, potassium chloride, potassium gluconate, potassium mixtures, dibasic potassium phosphate, monobasic potassium phosphate, potassium phosphate mixtures, sodium acetate, sodium bicarbonate, sodium chloride, sodium citrate, sodium lactate, dibasic sodium phosphate, monobasic sodium phosphate, sodium phosphate mixtures, tromethamine, magnesium hydroxide, aluminum hydroxide, alginic acid, pyrogen-free water, isotonic saline, Ringer's solution, ethyl alcohol, etc., and combinations thereof.
(225) Exemplary lubricating agents include magnesium stearate, calcium stearate, stearic acid, silica, talc, malt, glyceryl behanate, hydrogenated vegetable oils, polyethylene glycol, sodium benzoate, sodium acetate, sodium chloride, leucine, magnesium lauryl sulfate, sodium lauryl sulfate, etc., and combinations thereof.
(226) Exemplary oils include almond, apricot kernel, avocado, babassu, bergamot, black current seed, borage, cade, camomile, canola, caraway, carnauba, castor, cinnamon, cocoa butter, coconut, cod liver, coffee, corn, cotton seed, emu, eucalyptus, evening primrose, fish, flaxseed, geraniol, gourd, grape seed, hazel nut, hyssop, isopropyl myristate, jojoba, kukui nut, lavandin, lavender, lemon, litsea cubeba, macademia nut, mallow, mango seed, meadowfoam seed, mink, nutmeg, olive, orange, orange roughy, palm, palm kernel, peach kernel, peanut, poppy seed, pumpkin seed, rapeseed, rice bran, rosemary, safflower, sandalwood, sasquana, savoury, sea buckthorn, sesame, shea butter, silicone, soybean, sunflower, tea tree, thistle, tsubaki, vetiver, walnut, and wheat germ oils. Exemplary oils include, but are not limited to, butyl stearate, caprylic triglyceride, capric triglyceride, cyclomethicone, diethyl sebacate, dimethicone 360, isopropyl myristate, mineral oil, octyldodecanol, oleyl alcohol, silicone oil, and combinations thereof.
(227) Liquid dosage forms for oral and parenteral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredients, the liquid dosage forms may comprise inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (e.g., cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents. In certain embodiments for parenteral administration, the conjugates of the invention are mixed with solubilizing agents such as Cremophor, alcohols, oils, modified oils, glycols, polysorbates, cyclodextrins, polymers, and combinations thereof.
(228) Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions can be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation can be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that can be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.
(229) The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.
(230) In order to prolong the effect of a drug, it is often desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This can be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is accomplished by dissolving or suspending the drug in an oil vehicle.
(231) Compositions for rectal or vaginal administration are typically suppositories which can be prepared by mixing the conjugates of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active ingredient.
(232) Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active ingredient is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may comprise buffering agents.
(233) Solid compositions of a similar type can be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally comprise opacifying agents and can be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions which can be used include polymeric substances and waxes. Solid compositions of a similar type can be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like.
(234) The active ingredients can be in micro-encapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active ingredient can be admixed with at least one inert diluent such as sucrose, lactose or starch. Such dosage forms may comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may comprise buffering agents. They may optionally comprise opacifying agents and can be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions which can be used include polymeric substances and waxes.
(235) Dosage forms for topical and/or transdermal administration of a compound of this invention may include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants and/or patches. Generally, the active ingredient is admixed under sterile conditions with a pharmaceutically acceptable carrier and/or any needed preservatives and/or buffers as can be required. Additionally, the present invention contemplates the use of transdermal patches, which often have the added advantage of providing controlled delivery of an active ingredient to the body. Such dosage forms can be prepared, for example, by dissolving and/or dispensing the active ingredient in the proper medium. Alternatively or additionally, the rate can be controlled by either providing a rate controlling membrane and/or by dispersing the active ingredient in a polymer matrix and/or gel.
(236) Suitable devices for use in delivering intradermal pharmaceutical compositions described herein include short needle devices such as those described in U.S. Pat. Nos. 4,886,499; 5,190,521; 5,328,483; 5,527,288; 4,270,537; 5,015,235; 5,141,496; and 5,417,662. Intradermal compositions can be administered by devices which limit the effective penetration length of a needle into the skin, such as those described in PCT publication WO 99/34850 and functional equivalents thereof. Jet injection devices which deliver liquid vaccines to the dermis via a liquid jet injector and/or via a needle which pierces the stratum corneum and produces a jet which reaches the dermis are suitable. Jet injection devices are described, for example, in U.S. Pat. Nos. 5,480,381; 5,599,302; 5,334,144; 5,993,412; 5,649,912; 5,569,189; 5,704,911; 5,383,851; 5,893,397; 5,466,220; 5,339,163; 5,312,335; 5,503,627; 5,064,413; 5,520,639; 4,596,556; 4,790,824; 4,941,880; 4,940,460; and PCT publications WO 97/37705 and WO 97/13537. Ballistic powder/particle delivery devices which use compressed gas to accelerate vaccine in powder form through the outer layers of the skin to the dermis are suitable. Alternatively or additionally, conventional syringes can be used in the classical mantoux method of intradermal administration.
(237) Formulations suitable for topical administration include, but are not limited to, liquid and/or semi liquid preparations such as liniments, lotions, oil in water and/or water in oil emulsions such as creams, ointments and/or pastes, and/or solutions and/or suspensions. Topically-administrable formulations may, for example, comprise from about 1% to about 10% (w/w) active ingredient, although the concentration of the active ingredient can be as high as the solubility limit of the active ingredient in the solvent. Formulations for topical administration may further comprise one or more of the additional ingredients described herein.
(238) A pharmaceutical composition of the invention can be prepared, packaged, and/or sold in a formulation suitable for pulmonary administration via the buccal cavity. Such a formulation may comprise dry particles which comprise the active ingredient and which have a diameter in the range from about 0.5 to about 7 nanometers or from about 1 to about 6 nanometers. Such compositions are conveniently in the form of dry powders for administration using a device comprising a dry powder reservoir to which a stream of propellant can be directed to disperse the powder and/or using a self propelling solvent/powder dispensing container such as a device comprising the active ingredient dissolved and/or suspended in a low-boiling propellant in a sealed container. Such powders comprise particles wherein at least 98% of the particles by weight have a diameter greater than 0.5 nanometers and at least 95% of the particles by number have a diameter less than 7 nanometers. Alternatively, at least 95% of the particles by weight have a diameter greater than 1 nanometer and at least 90% of the particles by number have a diameter less than 6 nanometers. Dry powder compositions may include a solid fine powder diluent such as sugar and are conveniently provided in a unit dose form.
(239) Low boiling propellants generally include liquid propellants having a boiling point of below 65° F. at atmospheric pressure. Generally the propellant may constitute 50 to 99.9% (w/w) of the composition, and the active ingredient may constitute 0.1 to 20% (w/w) of the composition. The propellant may further comprise additional ingredients such as a liquid non-ionic and/or solid anionic surfactant and/or a solid diluent (which may have a particle size of the same order as particles comprising the active ingredient).
(240) Pharmaceutical compositions of the invention formulated for pulmonary delivery may provide the active ingredient in the form of droplets of a solution and/or suspension. Such formulations can be prepared, packaged, and/or sold as aqueous and/or dilute alcoholic solutions and/or suspensions, optionally sterile, comprising the active ingredient, and may conveniently be administered using any nebulization and/or atomization device. Such formulations may further comprise one or more additional ingredients including, but not limited to, a flavoring agent such as saccharin sodium, a volatile oil, a buffering agent, a surface active agent, and/or a preservative such as methylhydroxybenzoate. The droplets provided by this route of administration may have an average diameter in the range from about 0.1 to about 200 nanometers.
(241) The formulations described herein as being useful for pulmonary delivery are useful for intranasal delivery of a pharmaceutical composition of the invention. Another formulation suitable for intranasal administration is a coarse powder comprising the active ingredient and having an average particle from about 0.2 to 500 micrometers. Such a formulation is administered. by rapid inhalation through the nasal passage from a container of the powder held close to the nares.
(242) Formulations suitable for nasal administration may, for example, comprise from about as little as 0.1% (w/w) and as much as 100% (w/w) of the active ingredient, and may comprise one or more of the additional ingredients described herein. A pharmaceutical composition of the invention can be prepared, packaged, and/or sold in a formulation suitable for buccal administration. Such formulations may, for example, be in the form of tablets and/or lozenges made using conventional methods, and may contain, for example, 0.1 to 20% (w/w) active ingredient, the balance comprising an orally dissolvable and/or degradable composition and, optionally, one or more of the additional ingredients described herein. Alternately, formulations suitable for buccal administration may comprise a powder and/or an aerosolized and/or atomized solution and/or suspension comprising the active ingredient. Such powdered, aerosolized, and/or aerosolized formulations, when dispersed, may have an average particle and/or droplet size in the range from about 0.1 to about 200 nanometers, and may further comprise one or more of the additional ingredients described herein.
(243) A pharmaceutical composition of the invention can be prepared, packaged, and/or sold in a formulation suitable for ophthalmic administration. Such formulations may, for example, be in the form of eye drops including, for example, a 0.1/1.0% (w/w) solution and/or suspension of the active ingredient in an aqueous or oily liquid carrier. Such drops may further comprise buffering agents, salts, and/or one or more other of the additional ingredients described herein. Other opthalmically-administrable formulations which are useful include those which comprise the active ingredient in microcrystalline form and/or in a liposomal preparation. Ear drops and/or eye drops are contemplated as being within the scope of this invention.
(244) Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and/or perform such modification with ordinary experimentation. General considerations in the formulation and/or manufacture of pharmaceutical compositions can be found, for example, in Remington: The Science and Practice of Pharmacy 21.sup.st ed., Lippincott Williams & Wilkins, 2005.
(245) Provided compounds can be administered using any amount and any route of administration effective for treatment. The exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the infection, the particular composition, its mode of administration, its mode of activity, and the like.
(246) Compounds provided herein are typically formulated in dosage unit form for ease of administration and uniformity of dosage. It will be understood, however, that the total daily usage of the compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular subject or organism will depend upon a variety of factors including the disease, disorder, or condition being treated and the severity of the disorder; the activity of the specific active ingredient employed; the specific composition employed; the age, body weight, general health, sex and diet of the subject; the time of administration, route of administration, and rate of excretion of the specific active ingredient employed; the duration of the treatment; drugs used in combination or coincidental with the specific active ingredient employed; and like factors well known in the medical arts.
(247) The compounds and compositions provided herein can be administered by any route, including oral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (as by powders, ointments, creams, and/or drops), mucosal, nasal, bucal, enteral, sublingual; by intratracheal instillation, bronchial instillation, and/or inhalation; and/or as an oral spray, nasal spray, and/or aerosol. Specifically contemplated routes are systemic intravenous injection, regional administration via blood and/or lymph supply, and/or direct administration to an affected site, in a bolus or continuous infusion regimen. In general the most appropriate route of administration will depend upon a variety of factors including the nature of the agent (e.g., its stability in the environment of the gastrointestinal tract), the condition of the subject (e.g., whether the subject is able to tolerate oral administration), etc.
(248) The exact amount of a compound required to achieve a therapeutically effective amount will vary from subject to subject, depending, for example, on species, age, and general condition of a subject, severity of the side effects or disorder, identity of the particular compound(s), mode of administration, and the like. The desired dosage can be delivered three times a day, two times a day, once a day, every other day, every third day, every week, every two weeks, every three weeks, or every four weeks. In certain embodiments, the desired dosage can be delivered using multiple administrations (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, or more administrations).
(249) In certain embodiments, a therapeutically effective amount of a compound for administration one or more times a day to a 70 kg adult human may comprise about 0.0001 mg to about 3000 mg, about 0.0001 mg to about 2000 mg, about 0.0001 mg to about 1000 mg, about 0.001 mg to about 1000 mg, about 0.01 mg to about 1000 mg, about 0.1 mg to about 1000 mg, about 1 mg to about 1000 mg, about 1 mg to about 100 mg, about 10 mg to about 1000 mg, or about 100 mg to about 1000 mg, about 0.001 mg to about 100 mg, from about 0.01 mg to about 50 mg, from about 0.1 mg to about 40 mg, from about 0.5 mg to about 30 mg, from about 0.01 mg to about 10 mg, from about 0.1 mg to about 10 mg, and from about 1 mg to about 25 mg, per kilogram, of a compound. It will be appreciated that dose ranges as described herein provide guidance for the administration of provided pharmaceutical compositions to an adult. The amount to be administered to, for example, a child or an adolescent can be determined by a medical practitioner or person skilled in the art and can be lower or the same as that administered to an adult. The desired dosage may be delivered three times a day, two times a day, once a day, every other day, every third day, every week, every two weeks, every three weeks, or every four weeks. In certain embodiments, the desired dosage may be delivered using multiple administrations (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, or more administrations).
(250) The method comprises administration of a compound in a therapeutically effective dose to a host subject. In some embodiments, the subject is an animal. In some embodiments, a therapeutically effective dose comprises an amount ranging from approximately 0.1 mg/kg to approximately 50.0 mg/kg. In some embodiments, a therapeutically effective dose comprises an amount ranging from approximately 0.5 mg/kg to approximately 50.0 mg/kg. In some embodiments, a therapeutically effective dose comprises an amount ranging from approximately 0.5 mg/kg to approximately 40.0 mg/kg. In some embodiments, a therapeutically effective dose comprises an amount ranging from approximately 0.5 mg/kg to approximately 30.0 mg/kg. In some embodiments, a therapeutically effective dose comprises an amount ranging from approximately 1.0 mg/kg to approximately 25.0 mg/kg. In some embodiments, a therapeutically effective dose comprises an amount ranging from approximately 1.5 mg/kg to approximately 15.0 mg/kg. In some embodiments, treatment is administered locally. In some embodiments, treatment is administered by continuous infusion over a certain period of time. In certain embodiments, administration is via intraarterial infusion. In certain embodiments, administration is via intraarterial infusion via an artery feeding the tumor being treated.
(251) Some embodiments of the invention further comprise the administration of at least one other therapy or therapeutic agents. The compound or composition can be administered concurrently with, prior to, or subsequent to, the other therapy or therapeutic agents. In general, each therapy or agent will be administered at a dose and/or on a time schedule determined for that agent. In will further be appreciated that the additional therapeutically active agent utilized in this combination can be administered together in a single composition or administered separately in different compositions. The particular combination to employ in a regimen will take into account compatibility of the compound with the therapy or therapeutic agent and/or the desired therapeutic effect to be achieved. In general, it is expected that additional therapy or therapeutic agent utilized in combination be utilized at levels that do not exceed the levels at which they are utilized individually. In some embodiments, the levels utilized in combination will be lower than those utilized individually.
(252) The compounds or compositions can be administered in combination with a therapy or therapeutic agent that improves their bioavailability, reduce and/or modify their metabolism, inhibit their excretion, and/or modify their distribution within the body. It will also be appreciated that the therapy or therapeutic agent employed may achieve a desired effect for the same disease or disorder, and/or it may achieve different effects (e.g., control of adverse side-effects).
(253) Cancer therapies include, but are not limited to, surgery and surgical treatments, radiation therapy, and therapeutic agents (e.g., biotherapeutic agents and chemotherapeutic agents). In certain embodiments, the method comprises administration of radiation.
(254) Exemplary biotherapeutic agents include, but are not limited to, interferons, cytokines (e.g., tumor necrosis factor, interferon α, interferon γ), vaccines, hematopoietic growth factors, monoclonal serotherapy, immunostimulants and/or immunodulatory agents (e.g., IL-1, 2, 4, 6, or 12), immune cell growth factors (e.g., GM-CSF) and antibodies (e.g. HERCEPTIN (trastuzumab), T-DM1, AVASTIN (bevacizumab), ERBITUX (cetuximab), VECTIBIX (panitumumab), RITUXAN (rituximab), BEXXAR (tositumomab)).
(255) Exemplary chemotherapeutic agents include, but are not limited to, anti-estrogens (e.g. tamoxifen, raloxifene, and megestrol), LHRH agonists (e.g. goscrclin and leuprolide), anti-androgens (e.g. flutamide and bicalutamide), photodynamic therapies (e.g. vertoporfin (BPD-MA), phthalocyanine, photosensitizer Pc4, and demethoxy-hypocrellin A (2BA-2-DMHA)), nitrogen mustards (e.g. cyclophosphamide, ifosfamide, trofosfamide, chlorambucil, estramustine, and melphalan), nitrosoureas (e.g. carmustine (BCNU) and lomustine (CCNU)), alkylsulphonates (e.g. busulfan and treosulfan), triazenes (e.g. dacarbazine, temozolomide), platinum containing compounds (e.g. cisplatin, carboplatin, oxaliplatin), vinca alkaloids (e.g. vincristine, vinblastine, vindesine, and vinorelbine), taxoids (e.g. paclitaxel or a paclitaxel equivalent such as nanoparticle albumin-bound paclitaxel (ABRAXANE), docosahexaenoic acid bound-paclitaxel (DHA-paclitaxel, Taxoprexin), polyglutamate bound-paclitaxel (PG-paclitaxel, paclitaxel poliglumex, CT-2103, XYOTAX), the tumor-activated prodrug (TAP) ANG1005 (Angiopep-2 bound to three molecules of paclitaxel), paclitaxel-EC-1 (paclitaxel bound to the erbB2-recognizing peptide EC-1), and glucose-conjugated paclitaxel, e.g., 2′-paclitaxel methyl 2-glucopyranosyl succinate; docetaxel, taxol), epipodophyllins (e.g. etoposide, etoposide phosphate, teniposide, topotecan, 9-aminocamptothecin, camptoirinotecan, irinotecan, crisnatol, mytomycin C), anti-metabolites, DHFR inhibitors (e.g. methotrexate, dichloromethotrexate, trimetrexate, edatrexate), IMP dehydrogenase inhibitors (e.g. mycophenolic acid, tiazofurin, ribavirin, and EICAR), ribonuclotide reductase inhibitors (e.g. hydroxyurea and deferoxamine), uracil analogs (e.g. 5-fluorouracil (5-FU), floxuridine, doxifluridine, ratitrexed, tegafur-uracil, capecitabine), cytosine analogs (e.g. cytarabine (ara C), cytosine arabinoside, and fludarabine), purine analogs (e.g. mercaptopurine and Thioguanine), Vitamin D3 analogs (e.g. EB 1089, CB 1093, and KH 1060), isoprenylation inhibitors (e.g. lovastatin), dopaminergic neurotoxins (e.g. 1-methyl-4-phenylpyridinium ion), cell cycle inhibitors (e.g. staurosporine), actinomycin (e.g. actinomycin D, dactinomycin), bleomycin (e.g. bleomycin A2, bleomycin B2, peplomycin), anthracycline (e.g. daunorubicin, doxorubicin, pegylated liposomal doxorubicin, idarubicin, epirubicin, pirarubicin, zorubicin, mitoxantrone), MDR inhibitors (e.g. verapamil), Ca.sup.2+ ATPase inhibitors (e.g. thapsigargin), imatinib, thalidomide, lenalidomide, tyrosine kinase inhibitors (e.g., axitinib (AG013736), bosutinib (SKI-606), cediranib (RECENTIN™, AZD2171), dasatinib (SPRYCEL®, BMS-354825), erlotinib (TARCEVA®), gefitinib (IRESSA®), imatinib (Gleevec®, CGP57148B, STI-571), lapatinib (TYKERB®, TYVERB®), lestaurtinib (CEP-701), neratinib (HKI-272), nilotinib (TASIGNA®), semaxanib (semaxinib, SU5416), sunitinib (SUTENT®, SU11248), toceranib (PALLADIA®), vandetanib (ZACTIMA®, ZD6474), vatalanib (PTK787, PTK/ZK), trastuzumab (HERCEPTIN@), bevacizumab (AVASTIN®), rituximab (RITUXAN®), cetuximab (ERBITUX®), panitumumab (VECTIBIX®), ranibizumab (Lucentis®), nilotinib (TASIGNA®), sorafenib (NEXAVAR®), everolimus (AFINITOR®), alemtuzumab (CAMPATH®), gemtuzumab ozogamicin (MYLOTARG®), temsirolimus (TORISEL®), ENMD-2076, PCI-32765, AC220, dovitinib lactate (TKI258, CHIR-258), BIBW 2992 (TOVOK™), SGX523, PF-04217903, PF-02341066, PF-299804, BMS-777607, ABT-869, MP470, BIBF 1120 (VARGATEF®), AP24534, JNJ-26483327, MGCD265, DCC-2036, BMS-690154, CEP-11981, tivozanib (AV-951), OSI-930, MM-121, XL-184, XL-647, and/or XL228), proteasome inhibitors (e.g., bortezomib (VELCADE)), mTOR inhibitors (e.g., rapamycin, temsirolimus (CCI-779), everolimus (RAD-001), ridaforolimus, AP23573 (Ariad), AZD8055 (AstraZeneca), BEZ235 (Novartis), BGT226 (Norvartis), XL765 (Sanofi Aventis), PF-4691502 (Pfizer), GDC0980 (Genetech), SF1126 (Semafoe) and OSI-027 (OSI)), oblimersen, gemcitabine, carminomycin, leucovorin, pemetrexed, cyclophosphamide, dacarbazine, procarbizine, prednisolone, dexamethasone, campathecin, plicamycin, asparaginase, aminopterin, methopterin, porfiromycin, melphalan, leurosidine, leurosine, chlorambucil, trabectedin, procarbazine, discodermolide, carminomycin, aminopterin, and hexamethyl melamine.
(256) Kits
(257) In still another aspect, the present invention also provides kits (e.g., pharmaceutical packs) for treating a proliferative disorder comprising a compound of the Formula (A) or (B) and instructions for administration to a subject for treating a proliferative disorder. Kits may comprise a provided composition and a container (e.g., a vial, ampoule, bottle, syringe, and/or dispenser package, or other suitable container). In some embodiments, provided kits may optionally further include a second container comprising a suitable aqueous carrier for dilution or suspension of the provided composition for preparation of administration to a subject. In some embodiments, contents of provided formulation container and solvent container combine to form at least one unit dosage form.
(258) Optionally, a single container may comprise one or more compartments for containing a provided composition, and/or appropriate aqueous carrier for suspension or dilution. In some embodiments, a single container can be appropriate for modification such that the container may receive a physical modification so as to allow combination of compartments and/or components of individual compartments. For example, a foil or plastic bag may comprise two or more compartments separated by a perforated seal which can be broken so as to allow combination of contents of two individual compartments once the signal to break the seal is generated. A pharmaceutical pack or kit may thus comprise such multi-compartment containers including a provided composition and appropriate solvent and/or appropriate aqueous carrier for suspension.
(259) Optionally, instructions for use are additionally provided in such kits of the invention. Such instructions may provide, generally, for example, instructions for dosage and administration. In other embodiments, instructions may further provide additional detail relating to specialized instructions for particular containers and/or systems for administration. Still further, instructions may provide specialized instructions for use in conjunction and/or in combination with additional therapy.
EXAMPLES
(260) In order that the invention described herein may be more fully understood, the following examples are set forth. It should be understood that these examples are for illustrative purposes only and are not to be construed as limiting this invention in any manner.
Example 1. Inhibition of Primary Patient Samples
(261) Table 1 highlights a series of primary patient leukemia samples-both acute and chronic and treatment naïve and refractory and the effect of granaticin A, granaticin B, derivative 1, and derivative 2 to inhibit cellular proliferation. The IC.sub.50 data reveals that granaticin B is approximately one log more active than granaticin A against all the samples tested and inhibits cellular proliferation in the low nanomolar range. The two starred samples represent serial samples taken from the same patients after they received 2-3 cycles of additional high dose salvage chemotherapy and indicate that although refractory to salvage chemotherapy, the Cdc7 pathway remains a potentially efficacious target as the sensitivity to these compounds remains unchanged. Importantly, each patient sample was resistant to the prior therapy received using this assay.
(262) TABLE-US-00001 TABLE 1 Inhibition of patient leukemia samples IC.sub.50 (microM) Leukemia sample Granaticin A Deriv. 1 Deriv. 2 Granaticin B AML, de novo 0.72 0.62 2.01 0.18 AML, primary 0.75 0.62 1.55 0.11 refractory 1 AML, primary 0.63 0.47 1.44 0.10 refractory 1* AML, primary 0.77 0.84 1.45 0.14 refractory AML, relapsed 1.20 0.70 2.23 0.10 AML, relapsed 0.21 0.23 0.35 0.05 AML, relapsed s/p 1.88 1.86 8.10 0.62 HSCT AMMoL 0.99 0.79 5.88 0.49 ALL, Ph+ 0.56 0.47 2.57 0.19 Biphenotypic acute 0.30 0.69 0.90 0.04 leukemia 1 Biphenotypic acute 0.54 0.76 1.02 0.10 leukemia 1* CML, untreated 0.26 0.33 1.24 0.08 CLL, untreated, 0.46 0.36 1.21 0.31 chromosome 17p del. PhALL3.1 0.14 0.12 0.47 0.03 *Indicates a second sample from the same patient following high-dose chemotherapy
(263) Table 2 highlights cell lines and primary patient samples tested for granaticin A and granaticin B to inhibit cellular proliferation (summarized from
(264) TABLE-US-00002 TABLE 2 IC.sub.50 (microM) Cell Line Description Granaticin A Granaticin B ALL3 Hematopoietic-Human 0.14 0.03 Acute Lymphoblastic leukemia; Philadelphia chromosome positive HL60 Hematopoietic-Human 0.14 0.09 Acute Promyelocytic leukemia HL60 MX1 Hematopoietic-Human 0.17 0.08 Acute Promyelocytic leukemia; MDR variant of HL60 selected with mitoxantrone-clone 1 HL60 MX2 Hematopoietic-Human 0.11 0.07 Acute Promyelocytic leukemia; MDR variant of HL60 selected with mitoxantrone-clone 2 HL60 RV Hematopoietic-Human 0.09 0.07 Acute Promyelocytic leukemia: MDR variant of HL60 selected with vincristine JEKO Hematopoietic-Human 0.04 0.02 Mantle Cell Lymphoma JURKAT E61 Hematopoietic-Human 0.05 0.02 Acute Lymphoblastic T-cell leukemia K562 Hematopoietic-Human 0.12 0.08 Chronic Myelogenous leukemia transformed to acute erythroleukemia KASUMI4 Hematopoietic-Human 0.30 0.18 Chronic Myelogenous leukemia transformed to acute myeloid leukemia- overexpression of EVI1 MEG01 Hematopoietic-Human 0.17 0.08 Chronic Myelogenous leukemia transformed to acute Megakaryoblastic leukemia MOLT3 Hematopoietic-Human Acute Lympholastic 0.03 0.01 T-cell Leukemia NCEB1 Hematopoietic-Human 0.33 0.26 Mantle Cell Lymphoma 132 Primary-Patient sample 0.13 0.01 of Chronic Lymphocytic Leukemia ALBU Primary-Patient sample 0.43 0.03 of essential thrombocythemia transformed to refractory acute myelogenous leukemia DOGO Primary-Patient sample 0.15 0.07 of Chronic Lymphocytic Leukemia with trisomy of chromosome 12 and unmutated Ig heavy chain locus GLHU Primary-Patient sample 0.26 0.08 of Chronic Myelogenous leukemia JABR Primary-Patient sample 0.21 0.05 of relpased/refractory acute myelogenous leukemia with normal cytogenetics and Flt-3 ITD JAKL Primary-Patient sample 0.05 0.01 of Chronic Lymphocytic Leukemia with trisomy of chromosome 12 and unmutated Ig heavy chain locus JAMC Primary-Patient sample 0.06 0.04 of acute myelogenous leukemia with inversion of chromosome 16 JAQU Primary-Patient sample 0.05 0.01 of relapsed/refractory pre-B cell acute lymphoblastic leukemia JOBL Primary-Patient sample 0.56 0.19 of relapsed/refractory acute myelogenous leukemia JOHO Primary-Patient sample 0.88 0.17 of primary refractory acute myelogenous leukemia with inv3 and monosomy 7 (refractory to allogeneic BMT) JOQU Primary-Patient sample 0.77 0.14 of relapsed/refractory acute myelogenous leukemia with trisomy of chromosome #8 and Flt-3 ITD JUCO Primary-Patient sample 0.08 0.01 of pre-B cell acute lymphoblastic leukemia with t(4;11) involving the MLL gene MAWI Primary-Patient sample 0.72 0.18 of relpased/refractory acute myelogenous leukemia with normal cytogenetics MAWI B Primary-Patient sample 0.42 0.16 of secondary chronic myelogenous leukemia MIHA Primary-Patient sample 0.30 0.04 of relapsed/refractory acute biphenotypic leukemia (patient was refractory to allogeneic bone marrow transplant) PAPR Primary-Patient sample 0.99 0.49 of relapsed acute myelomonocytic leukemia RADO Primary-Patient sample 0.06 0.03 of primary refractory acute myelogenous leukemia with multiple chromosomal abnormalities (−5,−7) RORI Primary-Patient sample 1.20 0.10 of relapsed/refractory acute myelogenous leukemia with trisomy of chromosome #8 SOPA Primary-Patient sample 0.10 0.09 of secondary acute myelogenous leukemia with chromsome 11q23 abnormality STGL Primary-Patient sample 1.88 0.62 of refractory acute myelogenous leukemia with multiple chromosomal abnormalities(refractory to allogeneic BMT) A2780 Solid-Human ovarian 0.12 0.02 carcinoma A549 Solid-Human lung 0.28 0.09 adenocarcinoma; G12S KRAS mutation BE(2)C Solid-human 0.07 0.04 neuroblastoma; p53 mutant CWR22 Solid-human prostate 0.09 0.04 carcinoma FUUR1 Solid-human renal cell 0.48 0.34 carcinoma with the reciprocal ASPL-TFE3 fusion transcript H11-18 Solid-Human lung 0.42 0.06 adenocarcinoma; L858R EGFR mutation H1650 Solid- Human lung 0.29 0.06 adenocarcinoma; DelE746- A750 EGFR mutation; wild-type RAS; PTEN null H1734 Solid-Human lung 0.24 0.04 adenocarcinoma; G13C KRAS mutation H1975 Solid-Human lung 0.12 0.03 adenocarcinoma; T790M/L858R EGFR mutation; R273H p53 mutation; PTEN null; wild-type KRAS H2030 Solid-human lung 0.49 0.22 adenocarcinoma; wild-type EGFR; G12C KRAS mutation; G262V P53 mutation; PTEN null H2122 Solid-human lung 1.19 0.21 adenocarcinoma; G12C KRAS mutation; Q16L and C176F P53 mutations H23 Solid-Human lung 0.12 0.05 adenocarcinoma; G12c KRAS mutation; M2461 P53 mutation H2444 Solid-human lung 0.31 0.03 adenocarcinoma; G12V KRAS mutation H3255 Solid-Human lung 0.21 0.07 adenocarcinoma; L858R EGFR mutation; wild-type KRAS; wild-type P53 H358 Solid-Human lung 0.21 0.05 adenocarcinoma; G13C KRAS mutation H460 Solid-human lung 0.39 0.08 adenocarcinoma; Q61H KRAS mutation H820 Solid-human lung 0.09 0.05 adenocarcinoma; Del E746-E749 and T790M EGFR mutations HCC4011 Solid-Human lung 0.14 0.05 adenocarcinoma; L858R EGFR mutation; wild-type KRAS HCC827 Solid-human lung 0.28 0.12 adenocarcinoma; DelE746- A750 EGFR mutation HELA N10 Solid-human cervical 1.27 0.97 adenocarcinoma (positive for human papillomavirus infection) HPLD1 Solid-Immortalized 0.18 0.04 human bronchiolar epithelial cell with SV-40 Large T antigen HTB15 Solid-Human 0.65 0.46 Glioblastoma (U-118MG), classified as Grade IV JNDSRCT1 Solid-Human 0.07 0.05 desmoplastic small round cell tumor cell line MCF-10A Solid-human normal 0.26 0.24 mammary epithelium MCF-7 Solid-human invasive 0.17 0.13 breast ductal carcinoma, estrogen and progesterone receptor positive MDA-MB-231 Solid-human metastatic 0.47 0.29 breast adenocarcinoma; estrogen, progesterone, and HER2/NEU receptor negative MESO47 Solid-human 0.61 0.38 mesothelioma; WT-1 overexpression OVCAR3 Solid-human ovarian 0.21 0.11 adenocarcinoma PC9 Solid-Human lung 0.30 0.07 adenocarcinoma; DelE746- A750 EGFR mutation RB355 Solid-Human 0.09 0.04 retinoblastoma RH30 Solid-Human 0.13 0.08 rhabdomyosarcoma; P53 mutation SKMEL28 Solid-Human melanoma 0.54 0.34 SKOV3 Solid-Human 0.12 0.04 adenocarcinoma from the ovary derived from ascites; hypodiploid SW1736 Solid-Human anaplastic 0.16 0.02 thyroid carcinoma with BRAF V600E mutation TC71 Solid-Human Ewing's 0.15 0.09 sarcoma Y79 Solid-Human 0.06 0.05 retinoblastoma
Example 3. Stability Studies Using Granaticin A
(265) Standard liver microsome stability tests were performed with mouse and human microsomes with granaticin A and revealed that this drug is cleared quickly by the liver. The stability of granaticin A was determined in mouse and human liver microsomes. In mouse liver microsomes, recovery at 30 minutes was 3.68%. In mouse liver microsomes, recovery at 60 minutes was 3.84%. In human liver microsomes, recovery at 30 minutes was 23.88%. In human liver microsomes, recovery at 60 minutes was 12.01%.
Example 4. Preliminary Screening of Granaticin B
(266) Standard solubility studies were conducted with granaticin B. In aqueous solution, 4.0 mg/ml of granaticin B is soluble. In alcohol, >188 mg/ml of granaticin B is soluble. In methanol, >252 mg/ml of granaticin B is soluble.
(267) Different formulations of granaticin B were also tested and PK data measured (see Table 3).
(268) TABLE-US-00003 TABLE 3 Preliminary Mouse PK Screening Granaticin B, Mouse PK AUC.sub.inf Half-life C.sub.max CL screening 5 mg/kg (hr − μ/mL) (t.sub.2/1.sup.β) (μg/mL) (mL/hr/kg) A: 1.25% of Ethanol 3532.5 6.9 2834.7 1415.4 & Tween-20 B: 0.6% GDO-12 477981.5 3445.5 2537.2 10.5 C: 5% GDP-12 2689.9 6.6 2449.9 1858.8 D: DMSO 2724.5 2.1 2210.9 1835.2
OTHER EMBODIMENTS
(269) All patents, patent applications, and literature references cited herein are incorporated herein by reference.
(270) The foregoing has been a description of certain non-limiting embodiments of the invention. Those of ordinary skill in the art will appreciate that various changes and modifications to this description may be made without departing from the spirit or scope of the present invention, as defined in the following claims.