WO2008157407A2 - Thiadiazole, oxadiazole and triazole derivatives for treating leukemia - Google Patents

Abstract

Compounds of formula (I) hat are inhibitors of transcription factors of leukemia cell lines are provided. In addition, pharmaceutical compositions including the inhibiting compounds and methods for treating leukemia are also provided.

Description

PATENT APPLICATION

COMPOUNDS AND METHODS FOR TREATING LEUKEMIA

Cross-Reference to Related Applications

[0001] This application claims priority to Provisional Application No. 60/934,332, filed June 13, 2007, the disclosure of which is incorporated by reference its entirety.

Background [0002] Leukemia is a disease of the blood and bone marrow characterized by uncontrolled accumulation of blood cells. Over 44,000 new cases of leukemia will be diagnosed in the United States this year. There are four main types of Leukemia, acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute myelogenous leukemia (AML) and chronic myelogenous leukemia (CML). The most common types of leukemia in adults are acute myelogenous leukemia (AML), with over 13,000 estimated new cases this year, and chronic lymphocytic leukemia (CLL), with over 15,000 estimated new cases this year. Chronic myelogenous leukemia (CML) is estimated to be contracted in over 4,000 persons this year. Acute lymphocytic leukemia (ALL) estimated to be contracted in about 5,200 cases this year. Other forms of leukemia are unclassified types. The cause of leukemia is not known. Some causes include exposure to carcinogenic chemicals in the workplace and exposure to high doses of radiation.

[0003] There is a history of developing small molecule inhibitors of enzymes by targeting their active sites, however, protein-protein interaction inhibitors are relatively rare. In addition, the targeting of transcription factors in particular has been quite limited (see, Arndt, H. D. Angew Chem. Int Ed Engl 45, 4552-4560 (2006)), despite the clear advantages in specificity such targeting is likely to afford (see Melnick, A., Leukemia 19, 1109-1117, (2005) and Melnick, A. M., et al. J Clin Oncol 23, 3957- 3970, (2005) [0004] Currently, there is a need for novel, potent, and selective agents which are inhibitors of transcription factors of leukemia cells. The present invention provides small molecules that are inhibitors of transcription factors in leukemia cell lines. Thus, the compounds will be suitable for use alone or in combination for treating subjects in need of cancer chemotherapy.

Summary [0005] The present invention provides compounds that are inhibitors of transcription factors of leukemia cell lines. Accordingly the invention provides compounds having formula (I):

(I) wherein each Z\ Z², Z³, Z⁴,or Z⁵ is independently C-R¹ or N; each R¹ is independently, hydrogen, halo, (Ci-C₆)alkyl, (C₂-C₆)alkenyl, (C₂- C₆)alkynyl, (C₁-C₆)alkoxy, thio(Ci-C₆)alkyl, -N(R^a)₂, halo(Ci-C₆)alkyl, (C₃-C₈)- cycloalkyl, (C₃-C₈)cycloalkenyl, -C(=O)R⁵, -Q=O)Y¹R⁵, (C₆-Cio)aryl, (C₇-Ci₆)- arylalkyl, (C₂-C₉)heterocyclic, (C₂-C₉)heteroaryl, (C₃-Cis) heteroarylalkyl, -SO₂N(R⁸)₂ or cyano; or two R¹ groups taken together with the atoms to which they are attached can form a ring; each R⁸ is independently hydrogen, (Ci-C₆)alkyl or (Cβ-Cio) aryl; Y¹ is O, S or NR^b; each R^b is independently hydrogen or (Ci-C₆)alkyl; each R^a is independently hydrogen, (Ci-C₆)alkyl, -C(=O)R , or -C(=O)Y²R⁶;

Y² is O, S or NR^C; each R^c is independently hydrogen or (Ci-Cg)alkyl; each R⁵ and R⁶ is independently hydrogen, (d-C₆)alkyl, halo(Ci-C₆)alkyl, (C₃-C₈)cycloalkyl, (C₆-C₁₀)aryl, or (C₇-C₁₆)alkylaryl; each R² is independently hydrogen, (Ci-C₆)alkyl, heterocyclic, or heterocyclic(Ci-C₆)alkyl or absent; •'^"""* represents an optional double bond;

X is S, O, or NR³; R³ is hydrogen, or N(R⁴)₂, where each R⁴ is independently hydrogen, (Ci-C₆)alkyl or -C(=O)R⁷; R⁷ is hydrogen or (Ci- C₆)alkyl; wherein at least two of Z¹, Z², Z³, Z⁴, and Z⁵ are C-R¹; where any of alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclic, or heteroaryl groups in R¹ and R², R³ or R⁴ can be optionally substituted with 1, 2, 3, or 4 substituent groups, where the substituent groups independently are halo, oxo (=O), imino (=NR^d), (d-C₁₀)alkyl, (d-C₁₀)alkoxy, or C₆-aryl, -C(=O)R⁷,

-C(=O)Y³R⁷; -N(R^d)₂ or cyano; each R⁷ is independently hydrogen, (d-C₆)alkyl, halo(Ci-C₆)alkyl, (C₃-C₈)cycloalkyl, (C₆-Ci₀)aryl, or (C₇-Ci₆)alkylaryl; Y³ is O or S; or wherein one or more of the carbon atoms in the alkyl, alkenyl, or alkynyl groups can be independently replaced with non-peroxide oxygen, sulfur or NR^d; each R is independently hydrogen or (Ci-C₆)alkyl; provided that: when X is

N-NH₂, and Z¹, Z², Z⁴, and Z⁵ are C-H, then Z³ is not C-F; when X is NH, and Z¹, Z², Z³, and Z⁵ are C-H, then Z⁴ is not N; and when X is NH, and Z¹, Z², Zf.or Z⁵ are C-H, then Z is not N; or or a pharmaceutically acceptable salt thereof. [0006] The invention also provides compounds of formula (I) for use in medical therapy.

[0007] In another aspect, the present invention also provides: a pharmaceutical composition comprising a compound of formula (I) and a pharmaceutically acceptable excipient; a method for prevention or treatment of leukemia, comprising administering to a mammal (e.g., a human) in need of such treatment, an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof; a method for inhibiting transcription factors of leukemia cell lines comprising administering to a mammal (e.g., a human) in need of such treatment an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof;. prevent cancer progression via inhibition of angiogenesis in a tumor; or the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof to prepare a medicament for preventing angiogenesis in a tumor in a mammal (e.g., a human).

[0008] The invention also provides novel intermediates and processes disclosed herein that are useful for preparing compounds of formula (I) including the generic and specific intermediates as well as the synthetic processes described in the schemes and examples herein.

[0009] In another embodiment, the disclosed compounds can be used in combination with one or more additional drugs, e.g., ATRA (all trans retinoic acid), daunorubicin, and cytarabine.

Brief Description of the Drawings

[0010] Fig. IA and Fig. IB illustrates synthetic routes to prepare compounds of the invention.

[0011] Figs. 2 and 3 are illustrations of the FRET assay for identification of inhibitors of the Runt domain-CBFβ interaction.

[0012] Fig. 2 are fluorescence emission spectra (433 nm excitation) for a Cerulean- Runt domain (C-RD) / Venus-CBFβ (V-CBFβ) complex (black), for (C-RD) / (V- CBFβ) complex plus a 20-fold excess of CBFβ (gray), and for V-CBFβ alone (dark gray). [0013] Fig. 3 is a K_d measurement for C-RD binding with V-CBFβ using a serial dilution FRET-based assay. The serial dilution method was used for K_d determination with proteins at a 1:1 ratio over the concentration range 1 μM -1 nM. Arrow indicates the concentration used for compound screening.

[0014] Fig. 4A is a competitive displacement of V - CBFβ (150 nM) from C-RD (150 nM) by RDI-2 (KG-3-275).

[0015] Fig. 4B is an illustration of the RDI-2 (KG-3-275) inhibition of the RD/CBFβ interaction ex vivo.

[0016] Fig. 5A-5D illustrate NMR validation of RDI-2 (KG-3-275) binding to the Runt domain. Fig. 5A, ID and STD NMR spectra for RDI-2 alone (800 μM). Fig. 5B, ID and STD NMR spectra for the mixture of RDI-2 (800 μM) and Cerulean-Runt domain (30 μM). Fig. 5C, ID and STD NMR spectra for the mixture of RDI-2 (800 μM) and Venus-CBFβ (30 μM). Fig. 5D. Aliphatic portion of the ID and STD NMR spectra for the mixture of RDI-2 (800 μM) and Cerulean-Runt domain (30 μM).

[0017] Fig. 6 illustrates a dose-dependent inhibition SKNO-I cell growth measured by MTT assay after 72 hrs of incubation. [0018] Fig. 7 illustrates the inhibition of Kasumi-1 cell growth analyzed as in Fig. 6.

[0019] Fig. 8 illustrates inhibition of cell growth caused by RDI-2 (KG-3-275) on U937 (non-CBF leukemia cell line), HK-2 (kidney) and HEP-G2 (liver) cell lines. [0020] Fig. 9 illustrates apoptosis measured by Annexin V/PI positivity after 72 hrs.

[0021] Fig. 10 illustrates the Kasumi-1 cell cycle distribution (BrdU incorporation) following treatment with RDI-2 (KG-3-275) and RDI-4 (KG-1-253).

[0022] Fig. 11 illustrates histograms of CDl Ib expression in 7-AAD negative cells in the absence (DMSO) or presence of RDI-2 (KG-3-275) (50 μM), ATRA (2 μM), or combinations thereof.

[0023] Fig. 12 illustrates a summary of the results from Fig. 11 compiled from two experiments, each with three samples (n=6). Bars indicate 95% confidence intervals.

[0024] Fig. 13 illustrates examples of different cell types observed after Wright- Giemsa staining of treated Kasumi-1 cells.

[0025] Fig. 14 summarizes results of a 200 cell differential count based on morphology (see Fig. 13) after Wright-Giemsa staining of Kasumi-1 cells treated with DMSO control, RDI-2 (KG-3-275) alone, vitamin D alone, ATRA alone, RDI-2 + vitamin D, and RDI-2 + ATRA. [0026] Fig 15 illustrates histograms of CDl Ib expression in 7-AAD negative cells in the absence (DMSO) or presence of RDI-2 (KG-3-275) (50 μM), VitD (100 nM), or combinations thereof.

Detailed Description

[0027] In describing and claiming the invention, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any materials and methods similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred materials and methods are described herein. Each of the following terms has meaning associated with it in this section. Exemplary and preferred values listed below for radicals, substituents, and ranges are for illustrations only; they do not exclude other defined values or other values within defined ranges for the radicals and substituents.

[0028] For purposes of the description of this invention, the articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0029] As used herein, an "analog" of a chemical compound is a compound that, by way of example, resembles another in structure but is not necessarily an isomer (e.g., 5- fluorouracil is an analog of thymine). [0030] A "control" cell, tissue, sample, or subject is a cell, tissue, sample, or subject of the same type as a test cell, tissue, sample, or subject. The control may, for example, be examined at precisely or nearly the same time the test cell, tissue, sample, or subject is examined. The control may also, for example, be examined at a time distant from the time at which the test cell, tissue, sample, or subject is examined, and the results of the examination of the control may be recorded so that the recorded results may be compared with results obtained by examination of a test cell, tissue, sample, or subject. The control may also be obtained from another source or similar source other than the test group or a test subject, where the test sample is obtained from a subject suspected of having a disease or disorder for which the test is being performed.

[0031] A "test" cell, tissue, sample, or subject is one being examined.

[0032] A "pathoindicative" cell, tissue, or sample is one which, when present, is an indication that the animal in which the cell, tissue, or sample is located (or from which the tissue was obtained) is afflicted with a disease or disorder. By way of example, the presence of one or more breast cells in a lung tissue of an animal is an indication that the animal is afflicted with metastatic breast cancer.

[0033] A tissue "normally comprises" a cell if one or more of the cell are present in the tissue in an animal not afflicted with a disease or disorder.

[0034] As used herein, a "derivative" of a compound refers to a chemical compound that may be produced from another compound of similar structure in one or more steps, as in replacement of hydrogen by an alkyl, acyl, or amino group. [0035] The use of the word "detect" and its grammatical variants is meant to refer to measurement of the species without quantification, whereas use of the word "determine" or "measure" with their grammatical variants are meant to refer to measurement of the species with quantification. The terms "detect" and "identify" are used interchangeably herein.

[0036] As used herein, an "effective amount" means an amount sufficient to produce a selected effect. A "therapeutically effective amount" of a compound is that amount of compound which is sufficient to provide a beneficial effect to the subject to which the compound is administered. [0037] The term "in combination therapy" includes administration of at least one disclosed compound and at least one additional active drug or agent in the same composition or in a separate composition for administration simultaneously or sequentially. Combination therapy also includes administration of at least one disclosed compound within 24 hours of the administration of at least one additional drug or agent for treating leukemia. Preferably, the disclosed compound and the additional drug are administered within at least 12 hours of each other. More preferably, the disclosed compound and the additional drug are administered within at least 6 hours of each other. Non-limiting examples of additional drugs that can be used with the disclosed compounds include ATRA (all trans retinoic acid), daunorubicin, cytarabine, and the like.

[0038] As used herein, a "detectable marker" or a "reporter molecule" is an atom or a molecule that permits the specific detection of a compound comprising the marker in the presence of similar compounds without a marker. Detectable markers or reporter molecules include, but are not limited to, radioactive isotopes, antigenic determinants, enzymes, nucleic acids available for hybridization, chromophores, fluorophores, chemiluminescent molecules, electrochemically detectable molecules, and molecules that provide for altered fluorescence-polarization or altered light-scattering.

[0039] As used herein, the term "purified" and like terms relate to an enrichment of a molecule or compound relative to other components normally associated with the molecule or compound in a native environment. The term "purified" does not necessarily indicate that complete purity of the particular molecule has been achieved during the process. A "highly purified" compound as used herein refers to a compound that is greater than 90% pure.

[0040] As used herein, the term "pharmaceutically acceptable carrier" includes any of the standard pharmaceutical carriers known in the art, such as a phosphate buffered saline solution, hydroxypropyl beta-cyclodextrins (HO-propyl beta cyclodextrins), water, emulsions such as an oil/water or water/oil emulsion, and various types of wetting agents. The term also encompasses any of the agents approved by a regulatory agency of the U.S. Federal government or listed in the US Pharmacopeia for use in animals, including humans. [0041] As used herein, the term "pharmaceutically- acceptable salt" refers to salts which retain the biological effectiveness and properties of the compounds of the present invention and which are not biologically or otherwise undesirable. In many cases, the compounds of the present invention are capable of forming acid and/or base salts by virtue of the presence of amino and/or carboxyl groups or groups similar thereto. [0042] A "sample," as used herein, refers preferably to a biological sample from a subject, including, but not limited to, normal tissue samples, diseased tissue samples, biopsies, blood, saliva, feces, semen, tears, and urine. A sample can also be any other source of material obtained from a subject, which contains cells, tissues, or fluid of interest. A sample can also be obtained from cell or tissue culture. [0043] The term "standard," as used herein, refers to something used for comparison. For example, it can be a known standard agent or compound which is administered or added to a control sample and used for comparing results when measuring said compound in a test sample. Standard can also refer to an "internal standard", such as an agent or compound which is added at known amounts to a sample and is useful in determining such things as purification or recovery rates when a sample is processed or subjected to purification or extraction procedures before a marker of interest is measured.

[0044] A "subject" of analysis, diagnosis, or treatment is an animal. Such animals include mammals, preferably a human. [0045] A "therapeutic" treatment is a treatment administered to a subject who exhibits signs of pathology for the purpose of diminishing or eliminating those signs. [0046] A "therapeutically effective amount" of a compound is that amount of compound which is sufficient to provide a beneficial effect to the subject to which the compound is administered.

[0047] As used herein, the term "treating" includes prophylaxis of the specific disorder or condition, or alleviation of the symptoms associated with a specific disorder or condition and/or preventing or eliminating said symptoms. A "prophylactic" treatment is a treatment administered to a subject who does not exhibit signs of a disease or exhibits only early signs of the disease for the purpose of decreasing the risk of developing pathology associated with the disease. [0048] As used herein, an "instructional material" includes a publication, a recording, a diagram, or any other medium of expression which can be used to communicate the usefulness of the composition of the invention for its designated use. The instructional material of the kit of the invention may, for example, be affixed to a container which contains the composition or be shipped together with a container which contains the composition. Alternatively, the instructional material may be shipped separately from the container with the intention that the instructional material and the composition be used cooperatively by the recipient.

[0049] The method of the invention includes a kit comprising an inhibitor identified in the invention and an instructional material which describes administering the inhibitor or a composition comprising the inhibitor to a cell or an animal. This should be construed to include other embodiments of kits that are known to those skilled in the art, such as a kit comprising a (preferably sterile) solvent suitable for dissolving or suspending the composition of the invention prior to administering the compound to a cell or an animal. Preferably the animal is a human. [0050] In cases where compounds are sufficiently basic or acidic to form acid or base salts, use of the compounds as salts may be appropriate. Examples of acceptable salts are organic acid addition salts formed with acids which form a physiological acceptable anion, for example, tosylate, methanesulfonate, acetate, citrate, malonate, tartarate, succinate, benzoate, ascorbate, α-ketoglutarate, and α-glycerophosphate. Suitable inorganic salts may also be formed, including hydrochloride, sulfate, nitrate, bicarbonate, and carbonate salts. [0051] Pharmaceutically acceptable acid addition salts may be prepared from inorganic and organic acids. Salts derived from inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Salts derived from organic acids include acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluene-sulfonic acid, salicylic acid, and the like. [0052] Pharmaceutically-acceptable base addition salts can be prepared from inorganic and organic bases. Salts derived from inorganic bases, include by way of example only, sodium, potassium, lithium, ammonium, calcium and magnesium salts. Salts derived from organic bases include, but are not limited to, salts of primary, secondary and tertiary amines, such as alkyl amines, dialkyl amines, trialkyl amines, substituted alkyl amines, di(substituted alkyl) amines, tri(substituted alkyl) amines, alkenyl amines, dialkenyl amines, trialkenyl amines, substituted alkenyl amines, di(substituted alkenyl) amines, tri(substituted alkenyl) amines, cycloalkyl amines, di(cycloalkyl) amines, tri(cycloalkyl) amines, substituted cycloalkyl amines, disubstituted cycloalkyl amine, trisubstituted cycloalkyl amines, cycloalkenyl amines, di(cycloalkenyl) amines, tri(cycloalkenyl) amines, substituted cycloalkenyl amines, disubstituted cycloalkenyl amine, trisubstituted cycloalkenyl aminέs, aryl amines, diaryl amines, triaryl amines, heteroaryl amines, diheteroaryl amines, triheteroaryl amines, heterocyclic amines, diheterocyclic amines, triheterocyclic amines, mixed di- and tri-amines where at least two of the substituents on the amine are different and are selected from the group consisting of alkyl, substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, heteroaryl, heterocyclic, and the like. Also included are amines where the two or three substituents, together with the amino nitrogen, form a heterocyclic or heteroaryl group. Examples of suitable amines include, by way of example only, isopropylamine, trimethyl amine, diethyl amine, tri(iso-propyl) amine, tri(n-propyl) amine, ethanolamine, 2-dimethylaminoethanol, tromethamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, N- alkylglucamines, theobromine, purines, piperazine, piperidine, morpholine, N- ethylpiperidine, and the like. It should also be understood that other carboxylic acid derivatives would be useful in the practice of this invention, for example, carboxylic acid amides, including carboxamides, lower alkyl carboxamides, dialkyl carboxamides, and the like.

[0053] Acceptable salts may be obtained using standard procedures well known in the art, for example by reacting a sufficiently basic compound such as an amine with a suitable acid affording a physiologically acceptable anion. Alkali metal (for example, sodium, potassium or lithium) or alkaline earth metal (for example calcium) salts of organic (e.g., carboxylic) acids can also be made.

[0054] Processes for preparing compounds of formula (I) or for preparing intermediates useful for preparing compounds of formula (I) are provided as further embodiments of the invention. Intermediates useful for preparing compounds of formula (I) are also provided as further embodiments of the invention.

[0055] As used herein, the term "halogen" or "halo" includes bromo, chloro, fluoro, and iodo.

[0056] The term "haloalkyl" as used herein refers to an alkyl radical bearing at least one halogen substituent, for example, chloromethyl, fluoroethyl, trifluoromethyl, pentafluoroethyl, and the like.

[0057] The term "alkyl or (Ci-C₆)alkyl," as used herein, represents a branched or linear alkyl group having from one to six carbon atoms. Typically Ci-C₆ alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, butyl, iso-butyl, sec- butyl, tert-butyl, pentyl, hexyl, and the like.

[0058] The term "alkenyl or (C₂-C6)alkenyl," as used herein, represents an olefinically unsaturated branched or linear group having from two to six carbon atoms and at least one double bond. Examples of such groups include, but are not limited to, 1-propenyl, 2-propenyl, 1, 3-butadienyl, 1-butenyl, hexenyl, pentenyl, and the like. [0059] The term "alkynyl or (C₂-C₆)alkynyl," refers to an unsaturated branched or linear group having from two to six carbon atoms and at least one triple bond. Examples of such groups include, but are not limited to, 1-propynyl, 2-propynyl, 1- butynyl, 2-butynyl, 1-pentynyl, and the like.

[0060] The term "C₃-Cs cycloalkyl" represents cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like.

[0061] The term "C₃-Cs cycloalkenyl" represents cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl, and the like. [0062] As used herein, the term "optionally substituted" refers to zero, 1, 2, 3, or 4 substituents, wherein the substituents are each independently selected. Each of the independently selected substituents may be the same or different than other substituents. [0063] As used herein the term "aryl" refers to a mono or bicyclic C₆-Ci_O carbocyclic ring system having one or two aromatic rings including, but not limited to, phenyl, benzyl, naphthyl, tetrahydronaphthyl, indanyl, indenyl, and the like.

[0064] As used herein "optionally substituted aryl" includes aryl compounds having zero, 1, 2, 3, or 4 substituents, and a substituted aryl includes aryl compounds having 1, 2, 3, or 4 substituents, wherein the substituents include groups such as, for example, alkyl, halo or amino substituents.

[0065] The term "(C₇-C i₆)arylalkyl" refers to any aryl group which is attached to the parent moiety via the alkyl group, e.g., aryl(Ci-Cg)alkyl. Thus, the term (C₅- C₆)aryl(C₅-C₈)alkyl refers to a five or six membered aromatic ring that is attached to the parent moiety via the Cs-Cs alkyl group. Non-limiting examples of arylalkyl include benzyl or phenethyl (Ph-CH₂-CH₂-) and the like.

[0066] The term "heterocyclic group" refers to an optionally substituted mono- or bicyclic carbocyclic ring system containing from one to three heteroatoms wherein the heteroatoms are selected from the group consisting of oxygen, sulfur, and nitrogen. [0067] As used herein the term "heteroaryl" refers to an optionally substituted mono- or bicyclic carbocyclic ring system having one or two aromatic rings containing from one to three heteroatoms and includes, but is not limited to, furyl, thienyl, pyridyl and the like.

[0068] The term "heteroarylalkyl" refers to any aryl group which is attached to the parent moiety via the alkyl group, e.g., heteroaryl (C i-Cg)alkyl. Thus, the term (C₄- C₅)heteroaryl(C₅-C₈)alkyl refers to a five or six membered aromatic ring that is attached to the parent moiety via the C₅-Cs alkyl group. Non-limiting examples of heteroalkyl include pyridylmetnyl (Pyr-CH₂-) or pyridylethyl (Pyr-CH₂-CH₂-) and the like. [0069] The term "bicyclic" represents either an unsaturated or saturated stable 7- to 12-membered bridged or fused bicyclic carbon ring. The bicyclic ring may be attached at any carbon atom which affords a stable structure. The term includes, but is not limited to, naphthyl, dicyclohexyl, dicyclohexenyl, and the like.

[0070] The compounds of the present invention can contain one or more asymmetric centers in the molecule. In accordance with the present invention any structure that does not designate the stereochemistry is to be understood as embracing all the various optical isomers, as well as racemic mixtures thereof.

[0071] It will be appreciated by those skilled in the art that compounds of the invention having a chiral center may exist in and be isolated in optically active and racemic forms. Some compounds may exhibit polymorphism. It is to be understood that the present invention encompasses any racemic, optically-active, polymorphic, or stereoisomeric form, or mixtures thereof, of a compound of the invention, which possess the useful properties described herein, it being well known in the art how to prepare optically active forms (for example, by resolution of the racemic form by recrystallization techniques, by synthesis from optically-active starting materials, by chiral synthesis, or by chromatographic separation using a chiral stationary phase) and how to determine SlP agonist activity using the standard tests described herein, or using other similar tests which are well known in the art.

[0072] The compounds of the present invention may exist in tautomeric forms and the invention includes both mixtures and separate individual tautomers. For example, the following structure:

]sr NH is understood to represent a mixture of the structures:

^ NH HN ^ N

' ' as well as ' ' .

[0073] The terms 16:0, 18:0, 18:1, 20:4 or 22:6 hydrocarbon refers to a branched or straight alkyl or alkenyl group, wherein the first integer represents the total number of carbons in the group and the second integer represent the number of double bonds in the group.

[0074] In one embodiment, of the compounds of formula (I) have the structure of Formula (II):

(H)

wherein Z¹, Z², Z³, Z⁴, Z⁵, X and R² are as defined above or a pharmaceutically acceptable salt thereof. [0075] In a preferred embodiment, in compounds having formula (II) each R is independently hydrogen, halo, (Ci-C₆)alkyl, (d-C₆)alkoxy, thio(Ci-C₆)alkyl, -N(R^a)₂, halo(Ci-C₆)alkyl, (C₃-C₈)cycloalkyl, (C₃-C₈)cycloalkenyl, -SO₂N(R⁸)₂, or (C₆-Cio)aryl; or two R groups taken together with the atoms to which they are attached can form a ring. R² is hydrogen, (Ci-C₆)alkyl, heterocyclic, or heterocyclic(Ci-C₆)alkyl. [0076] In another embodiment, the compounds of formula (I) have the structure of Formula (III):

(III)

wherein Z¹, Z², Z³, Z⁴, Z⁵, R² and R³ are as defined above or a pharmaceutically acceptable salt thereof.

[0077] In another embodiment, the compounds of formula (I) have the structure of Formula (IV):

(IV)

wherein Z , Z , Z , Z , Z and R² are as defined above or a pharmaceutically acceptable salt thereof. [0078] In another embodiment, the compounds of formula (I) have the structure of Formula (V):

(V)

wherein Z¹, Z², Z³, Z⁴, Z⁵ and R² are as defined above or a pharmaceutically acceptable salt thereof.

[0079] The present invention is also includes pharmaceutical compositions comprising the compounds of the present invention. More particularly, such compounds can be formulated as pharmaceutical compositions using standard pharmaceutically acceptable carriers, fillers, solubilizing agents and stabilizers known to those skilled in the art. For example, a pharmaceutical composition comprising a compound of the invention, or analog, derivative, or modification thereof, as described herein, is used to administer the appropriate compound to a subject.

[0080] The compounds of the invention are useful for treating a disease or disorder including administering to a subject in need thereof of a therapeutically acceptable amount of a compound of formula (I), or a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I), and a pharmaceutically- acceptable carrier.

[0081] Exemplary values for R¹ are hydrogen, methyl, ethyl, propyl, methoxy, ethoxy, propoxy, fluoro, chloro, bromo, trifluoromethyl, pentafluoroethyl, -SO₂N(R )₂ or -N(R^a)₂; R⁸ is hydrogen, methyl, ethyl, or phenyl; and R^a is hydrogen, methyl, ethyl, propyl, -C(=O)CH₃, -C(=O)CH₂CH₃, -C(=O)OCH₃, C(_^O)OCH₂CH₃. -C(=O)NHCH₃, or -C(=O)NHCH₂CH₃. In addition, two R¹ groups are taken together with the atoms to which they are attached to form a ring. [0082] Additional exemplary values for R¹ are hydrogen, methyl, ethyl, methoxy, ethoxy, fluoro, chloro, trifluoromethyl, -SO₂N(R⁸)₂ or -N(R^a)₂.

[0083] Additional exemplary values for R¹ are hydrogen, methyl, ethyl, methoxy, ethoxy, fluoro, trifluoromethyl, -SO₂N(R⁸)₂ or -N(R^a)₂. [0084] Additional exemplary values for R¹ are hydrogen, methyl, fluoro, trifluoromethyl, -N(CH₃)₂. or SO₂NH₂.

[0085] In addition, two R¹ groups are taken together with the atoms to which they are attached to form a ring, e.g., naphthyl.

[0086] Exemplary values for R^a are hydrogen, methyl, or ethyl. [0087] Additional Exemplary values for R^a are hydrogen or methyl. [0088] Exemplary values for R are hydrogen, methyl, ethyl or propyl. [0089] Exemplary values for X are NR³, O or S. [0090] Exemplary values for R³ are hydrogen, or N(R⁴)₂. [0091] An additional exemplary value for R is NH₂. [0092] Exemplary values for R⁴ are hydrogen or (Ci-C₆)alkyl. [0093] Additional exemplary values for R⁴ are hydrogen or methyl. [0094] Exemplary values for R⁵ are hydrogen or (Ci-C₆)alkyl. [0095] Additional exemplary values for R⁵ are hydrogen or methyl. [ [00009966]1 Exemplary values for R are hydrogen or (Ci-C₆)alkyl. [0097] Additional exemplary values for R are hydrogen or methyl. [0098] Exemplary values for R⁷ are hydrogen or (C]-C₆)alkyl. [0099] Additional exemplary values for R are hydrogen or methyl. [00100] Exemplary values for R⁸ are hydrogen, methyl, ethyl or phenyl. [00101] Additional exemplary values for R are hydrogen or methyl. [00102] Exemplary values for R^b are hydrogen, methyl, or ethyl. [00103] Additional Exemplary values for R are hydrogen or methyl. [00104] Exemplary values for R^c are hydrogen, methyl, or ethyl. [00105] Additional Exemplary values for R^c are hydrogen or methyl. [00106] Exemplary values for R^d are hydrogen, methyl, or ethyl.

[00107] Additional Exemplary values for R^d are hydrogen or methyl. [00108] Exemplary values for halo are fluorine or chlorine.

[00109] A synthetic route to prepare exemplary compounds disclosed herein is provided in the scheme in Figs. IA and IB. Additional compounds of formula (I), formula (II), formula (III), formula (IV) or formula (V) can be prepared by a person skilled in the art using known modifications to procedures from the schemes and detailed descriptions in the specific examples herein.

[00110] Exemplary compounds of formula (I) are illustrated in Table 1 , below.

Table 1

ND - Not determined NA - No activity NS - No saturation

[00111] Pharmaceutical compositions comprising one of more compounds of the invention can be administered to a subject in need thereof by any number of routes and means including, but not limited to, topical, oral, buccal, intravenous, intramuscular, intra arterial, intramedullary, intrathecal, intraventricular, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, vaginal, ophthalmic, pulmonary, or rectal means. The oral route is typically employed for most conditions requiring the compounds of the invention. Preference is given to intravenous injection or infusion for the acute treatments. For maintenance regimens, the oral or parenteral, e.g. intramuscular or subcutaneous, route is preferred.

[00112] In one embodiment, the pharmaceutical compositions useful for practicing the invention may be administered to deliver a dose of between 1 ng/kg/day and 200 mg/kg/day. In another embodiment, the pharmaceutical compositions useful for practicing the invention may be administered to deliver a dose of between 1 ng/kg/day and 200 mg/kg/day. [00113] Pharmaceutically acceptable carriers which are useful include, but are not limited to, glycerol, water, saline, ethanol, and other pharmaceutically acceptable salt solutions such as phosphates and salts of organic acids. Examples of these and other pharmaceutically acceptable carriers are described in Remington's Pharmaceutical Sciences (1991, Mack Publication Co., New Jersey).

[00114] The pharmaceutical compositions may be prepared, packaged, or sold in the form of a sterile injectable aqueous or oily suspension or solution. This suspension or solution may be formulated according to the known art, and may comprise, in addition to the active ingredient, additional ingredients such as the dispersing agents, wetting agents, or suspending agents described herein. Such sterile injectable formulations may be prepared using a non toxic parenterally acceptable diluent or solvent, such as water or 1,3 butane diol, for example. Other acceptable diluents and solvents include, but are not limited to, Ringer's solution, isotonic sodium chloride solution, and fixed oils such as synthetic mono- or di-glycerides. [00115] Compounds which are identified using any of the methods described herein may be formulated and administered to a subject for treatment of any of the diseases and disorders described herein. However, the use of compounds of the invention should not be construed to include only the diseases and disorder described herein. Preferably, the subject is a human. [00116] The formulations of the pharmaceutical compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology. In general, such preparatory methods include the step of bringing the active ingredient into association with a carrier or one or more other accessory ingredients, and then, if necessary or desirable, shaping or packaging the product into a desired single- or multi- dose unit.

[00117] Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions which are suitable for ethical 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 perform such modification with merely ordinary, if any, experimentation. Subjects to which administration of the pharmaceutical compositions of the invention is contemplated include, but are not limited to, humans and other primates, and mammals, including commercially relevant mammals such as cattle, pigs, horses, sheep, cats, and dogs.

[00118] A pharmaceutical composition of the invention may be prepared, packaged, or sold in bulk, as a single unit dose, or as a plurality of single unit doses. As used herein, a "unit dose" is a 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 or a convenient fraction of such a dosage such as, for example, one-half or one-third of such a dosage.

[00119] The relative amounts of the active ingredient, the pharmaceutically acceptable carrier, and any additional ingredients in a pharmaceutical composition of the invention will vary, depending upon the identity, size, and 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 99.9% (w/w) active ingredient.

[00120] In addition to the active ingredient, a pharmaceutical composition of the invention may further comprise one or more additional pharmaceutically active agents. Particularly contemplated additional agents include anti-emetics and scavengers such as cyanide and cyanate scavengers.

[00121] Controlled- or sustained-release formulations of a pharmaceutical composition of the invention may be made using conventional technology. In some cases, the dosage forms to be used can be provided as slow or controlled-release of one or more active ingredients therein using, for example, hydropropylmethyl cellulose, other polymer matrices, gels, permeable membranes, osmotic systems, multilayer coatings, microparticles, liposomes, or microspheres or a combination thereof to provide the desired release profile in varying proportions. Suitable controlled-release formulations known to those of ordinary skill in the art, including those described herein, can be readily selected for use with the pharmaceutical compositions of the invention. Thus, single unit dosage forms suitable for oral administration, such as tablets, capsules, gelcaps, and caplets that are adapted for controlled-release are encompassed by the present invention.

[00122] Controlled-release formulations can be designed to initially release an amount of drug that promptly produces the desired therapeutic effect, and gradually and continually release of other amounts of drug to maintain this level of therapeutic effect over an extended period of time. In order to maintain this constant level of drug in the body, the drug must be released from the dosage form at a rate that will replace the amount of drug being metabolized and excreted from the body. In addition, controlled- release of an active ingredient can be stimulated by various inducers, for example pH, temperature, enzymes, water, or other physiological conditions or compounds.

[00123] Powdered and granular formulations of a pharmaceutical preparation of the invention may be prepared using known methods. Such formulations may be administered directly to a subject, used, for example, to form tablets, to fill capsules, or to prepare an aqueous or oily suspension or solution by addition of an aqueous or oily vehicle thereto. Each of these formulations may further comprise one or more of dispersing or wetting agent, a suspending agent, and a preservative. Additional excipients, such as fillers and sweetening, flavoring, or coloring agents, may also be included in these formulations. As used herein, an "oily" liquid is one which comprises a carbon-containing liquid molecule and which exhibits a less polar character than water.

[00124] A formulation of a pharmaceutical composition of the invention suitable for oral administration may be prepared, packaged, or sold in the form of a discrete solid dose unit including, but not limited to, a tablet, a hard or soft capsule, a cachet, a troche, or a lozenge, each containing a predetermined amount of the active ingredient. Other formulations suitable for oral administration include, but are not limited to, a powdered or granular formulation, an aqueous or oily suspension, an aqueous or oily solution, a paste, a gel, a toothpaste, a mouthwash, a coating, an oral rinse, or an emulsion. The terms oral rinse and mouthwash are used interchangeably herein.

[00125] A tablet comprising the active ingredient may, for example, be made by compressing or molding the active ingredient, optionally with one or more additional ingredients. Compressed tablets may be prepared by compressing, in a suitable device, the active ingredient in a free flowing form such as a powder or granular preparation, optionally mixed with one or more of a binder, a lubricant, an excipient, a surface- active agent, and a dispersing agent. Molded tablets may be made by molding, in a suitable device, a mixture of the active ingredient, a pharmaceutically acceptable carrier, and at least sufficient liquid to moisten the mixture. Pharmaceutically acceptable excipients used in the manufacture of tablets include, but are not limited to, inert diluents, granulating and disintegrating agents, binding agents, and lubricating agents. Known dispersing agents include, but are not limited to, potato starch and sodium starch glycollate. Known surface-active agents include, but are not limited to, sodium lauryl sulphate. Known diluents include, but are not limited to, calcium carbonate, sodium carbonate, lactose, microcrystalline cellulose, calcium phosphate, calcium hydrogen phosphate, and sodium phosphate. Known granulating and disintegrating agents include, but are not limited to, corn starch and alginic acid. Known binding agents include, but are not limited to, gelatin, acacia, pre-gelatinized maize starch, polyvinylpyrrolidone, and hydroxypropyl methylcellulose. Known lubricating agents include, but are not limited to, magnesium stearate, stearic acid, silica, and talc.

[00126] Tablets may be non-coated or they may be coated using known methods to achieve delayed disintegration in the gastrointestinal tract of a subject, thereby providing sustained release and absorption of the active ingredient. For example, a material such as glyceryl monostearate or glyceryl distearate may be used to coat tablets. Further by way of example, tablets may be coated using methods described in U.S. Patent Nos. 4,256,108; 4,160,452; and 4,265,874 to form osmotically-controlled release tablets. Tablets may further include a sweetening agent, a flavoring agent, a coloring agent, a preservative, or some combination of these in order to provide for pharmaceutically elegant and palatable preparation.

[00127] Hard capsules comprising the active ingredient may be made using a physiologically degradable composition, such as gelatin. Such hard capsules comprise the active ingredient, and may further comprise additional ingredients including, for example, an inert solid diluent such as calcium carbonate, calcium phosphate, or kaolin. [00128] Soft gelatin capsules comprising the active ingredient may be made using a physiologically degradable composition, such as gelatin. Such soft capsules comprise the active ingredient, which may be mixed with water or an oil medium such as peanut oil, liquid paraffin, or olive oil. [00129] Liquid formulations of a pharmaceutical composition of the invention which are suitable for oral administration may be prepared, packaged, and sold either in liquid form or in the form of a dry product intended for reconstitution with water or another suitable vehicle prior to use. [00130] Injectable formulations may be prepared, packaged, or sold in unit dosage form, such as in ampules or in multi dose containers containing a preservative. Formulations for parenteral administration include, but are not limited to, suspensions, solutions, emulsions in oily or aqueous vehicles, pastes, and implantable sustained- release or biodegradable formulations. Such formulations may further comprise one or more additional ingredients including, but not limited to, suspending, stabilizing, or dispersing agents. In one embodiment of a formulation for parenteral administration, the active ingredient is provided in dry {e.g., powder or granular) form for reconstitution with a suitable vehicle {e.g., sterile pyrogen free water) prior to parenteral administration of the reconstituted composition. [00131] A pharmaceutical composition of the invention may be prepared, packaged, or sold in a formulation suitable for buccal administration. Such formulations may, for example, be in the form of tablets or lozenges made using conventional methods, and may, for example, 0.1 to 20% (w/w) active ingredient, the balance comprising an orally dissolvable or degradable composition and, optionally, one or more of the additional ingredients described herein. Alternately, formulations suitable for buccal administration may comprise a powder or an aerosolized or atomized solution or suspension comprising the active ingredient. Such powdered, aerosolized, or aerosolized formulations, when dispersed, preferably have an average particle 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.

[00132] As used herein, "additional ingredients" include, but are not limited to, one or more of the following: excipients; surface active agents; dispersing agents; inert diluents; granulating and disintegrating agents; binding agents; lubricating agents; sweetening agents; flavoring agents; coloring agents; preservatives; physiologically degradable compositions such as gelatin; aqueous vehicles and solvents; oily vehicles and solvents; suspending agents; dispersing or wetting agents; emulsifying agents, demulcents; buffers; salts; thickening agents; fillers; emulsifying agents; antioxidants; antibiotics; antifungal agents; stabilizing agents; and pharmaceutically acceptable polymeric or hydrophobic materials. See Genaro, ed., 1985, Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, which is incorporated herein by reference.

[00133] The compound can be administered to a subject as frequently as several times daily, or it may be administered less frequently, such as once a day, once a week, once every two weeks, once a month, or even less frequently, such as once every several months or even once a year or less. The frequency of the dose will be readily apparent to the skilled artisan and will depend upon any number of factors, such as, but not limited to, the type and severity of the disease being treated, the type, and age of the subject, etc.

[00134] The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the pharmaceutical compositions of the invention. In accordance with one embodiment, a kit is provided for treating a subject in need of prevention or treatment of leukemia. Preferably, the subject is a human. In one embodiment, the kit comprises one or more of active compounds and may also include one or more known anti-cancer agents. These pharmaceuticals can be packaged in a variety of containers, e.g., vials, tubes, microtiter well plates, bottles, and the like. Other reagents can be included in separate containers and provided with the kit; e.g., positive control samples, negative control samples, buffers, cell culture media, etc. Preferably, the kits will also include instructions for use.

[00135] Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are described herein. [00136] One skilled in the art will readily appreciate that the present invention is well adapted to carry out the objects and obtain the ends and advantages mentioned, as well as those inherent therein. The present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof.

[00137] Chemicals for syntheses can be purchased at commercial suppliers such as TCI America, Portland, OR, 97203 USA, Sigma- Aldrich, St. Louis, MO, 63178 USA, Alfa Aesar, Ward Hill, MA, 01835 USA, Thermo Fisher Scientific, Geel, 2440 Belgium,Wako Chemicals USA, Inc., Richmond, VA, 23237 USA, Matrix Scientific, Columbia, SC, 29224-5067 USA, and Oakwood Products, Inc., West Columbia, SC, 29172 USA, and unless noted were used without further purification.

Examples

[00138] The invention is now described with reference to the following examples. These examples are provided for the purpose of illustration only and the invention should in no way be construed as being limited to these examples, but rather should be construed to encompass any and all variations which become evident as a result of the teachings provided herein.

Example 1: Synthesis of 4-Amino-5-(pyridin-4-yl)-2,4-dihvdro-ri,2,41triazole-3-thione (RDI-4 (KG-1-253). 1).

[00139] To a solution of 2.0 g (14.6 mmol) of isonicotinic hydrazide in 20 mL of ethanol was added 1.2 g (21.9 mmol) of potassium hydroxide followed by 1.3 mL (21.9 mmol) of carbon disulfide. The solution was allowed to stir at room temperature for 30 minutes. The resulting precipitate was filtered and washed several times with ethanol. The precipitate was then taken up in 20 mL of water and 1.4 mL (29.2 mmol) of hydrazine monohydrate was added. The reaction mixture was heated at reflux for 4 hours. After cooling, the solution was diluted with 10 mL of cold water and acidified with concentrated hydrogen chloride solution. The immediately formed precipitate was filtered and recrystallized from ethanol to provide 1.1 g (41%) of the desired triazole. M.P.: 218-220 ⁰C. ¹H NMR (300 MHz, DMSOd₆) δ 14.17 (broad s, IH), 8.75 (d, J = 9.0 Hz, 2H), 8.01 (d, J = 6.3 Hz, 2H), 5.85 (s, 2H); ¹³C NMR (300 MHz, DMSOd₆) δ 167.7, 150.1, 147.4, 133.0, 121.6; ESI m/z (rel intensity): 193.8 (100).

Example 2: Synthesis of RDI-3 (KG-4-039).

[00140] 6-Methyl-nicotinic acid hvdrazide (2). To a solution of 1.0 g (6.6 mmol) of methyl 6-methylnicotinate in 20 mL of methanol was added 0.8 mL (16.5 mmol) of hydrazine monohydrate. The reaction was allowed to stir at room temperature for 2 hours. The solvent was removed under reduced pressure and the remaining solid was recrystallized with ethanol to provide 1.0 g (98%) of the hydrazide, 2. ¹H NMR (300 MHz, DMSOd₆): δ 8.82 (s, IH), 8.02 (d, J = 8.1 Hz, IH), 7.30 (d, J = 8.1 Hz, IH), 3.40 (broad s, 2H), 2.48 (s, 3H); ¹³C NMR (300 MHz, DMSOd₆): δ 167.3, 157.9, 151.2, 136.5, 127.1, 121.2, 22.8. [00141] 4-Amino-5-(6-methyl-pyridin-3-vn-2,4-dihvdro-ri,2,41triazole-3-thione (3). To a solution of 1.0 g (6.6 mmol) of hydrazide 2 in 10 mL of ethanol was added 0.6 g (9.9 mmol) of potassium hydroxide followed by 0.6 mL (9.9 mmol) of carbon disulfide. The solution was allowed to stir at room temperature for 30 minutes. The resulting precipitate was filtered and washed several times with ethanol. The precipitate was then taken up in 10 mL of water and 0.7 mL (13.2 mmol) of hydrazine monohydrate was added. The reaction mixture was refluxed for 4 hours. Once cool, the solution was diluted with 10 mL of cold water and acidified with concentrated hydrogen chloride solution. The immediately formed precipitate was filtered and recrystallized from ethanol to provide 0.3 g (20%) of the desired triazole. M.P.: 200-203 ⁰C. ¹H NMR

(300 MHz, DMSO-d₆) δ 14.00 (s, IH), 9.01 (s, IH), 8.25 (d, J = 8.1 Hz, IH), 7.44 (d, / = 6.0 Hz, IH), 5.78 (s, 2H), 2.53 (s, 3H); ¹³C NMR (300 MHz, DMSOd₆) δ 166.2, 156.4, 145.0, 143.2, 139.7, 127.3, 124.9, 23.2; ESI m/z (rel intensity): 207.8 (100).

Example 3: Synthesis of RDI-2 (KG-3-275) [00142] Methyl-6-methyl-pyridine-2-carboxylate (4). To a solution of 0.9 g (6.6 mmol) of 6-methylpicolinic acid in 7 mL of methanol and 3 mL of dichloromethane was added 6.6 mL (13.2 mmol) of a 2.0 M solution of trimethylsilyldiazomethane in ether. The reaction was allowed to stir at room temperature for 30 minutes. The solvent was then removed under reduced pressure and the remaining residue was taken up in ethyl acetate. The solution was washed with saturated sodium bicarbonate (2 x 10 mL), dried over magnesium sulfate and filtered. The solvent was removed under reduced pressure and the residue was purified by flash chromatography (1:5 ethyl acetate-hexanes) to provide 0.9 g (91%) of the methyl ester. ¹H NMR (300 MHz, CDCl₃): δ 7.95 (d, J = 7.7 Hz, IH), 7.77 (t, J = 7.7 Hz, IH), 7.38 (d, J = 7.7 Hz, IH), 4.11 (s, 3H), 2.65 (s, 3H); ¹³C NMR (300 MHz, CDCl₃): δ 165.3, 158.4, 147.0, 136.8, 126.4, 122.0, 52.2, 24.1.

[00143] 6-Methyl-pyridme-2-carboxylic hydrazide (5). To a solution of 0.9 g (6.0 mmol) of ester 4 in 15 mL of methanol was added 0.7 mL (14.9 mmol) of hydrazine monohydrate. The reaction was allowed to stir at room temperature for 30 minutes. The solvent was removed under reduced pressure and the remaining solid was recrystallized with ethanol to provide 0.8 g (94%) of the desired hydrazide. ¹H NMR (300 MHz, DMSOd₆): δ 9.67 (broad s, IH), 7.76 (m, 2H), 7.38 (d, J = 7.5 Hz, IH), 4.50 (broad s, 2H), 2.49 (s, 3H); ¹³C NMR (300 MHz, DMSOd₆): δ 163.5, 157.9, 149.8, 138.5, 126.5, 119.5, 24.5.

[00144] 4-Amino-5-(6-methyl-pyridin-2-yl)-2,4-dihydro-ri,2.41triazole-3-thione (6). To a solution of 2.O g (13.2 mmol) of hydrazide, 5, in 20 mL of ethanol was added 1.1 g (19.9 mmol) of potassium hydroxide followed by 1.2 mL (19.9 mmol) of carbon disulfide. The solution was allowed to stir at room temperature for 30 minutes. The resulting precipitate was filtered and washed several times with ethanol. The precipitate was then taken up in 20 mL of water and 1.3 mL (26.5 mmol) of hydrazine monohydrate was added. The reaction mixture was refluxed for 4 hours. Once cool, the solution was diluted with 20 mL of cold water and acidified with concentrated hydrogen chloride solution. The immediately formed precipitate was filtered and recrystallized from ethanol to provide 1.0 g (35%) of the desired triazole. mp: 210-212 ⁰C. ¹H NMR (300 MHz, DMSO-d₆) δ 14.00 (s, IH), 7.88 (m, 2H), 7.41 (d, / = 7.6 Hz, IH), 6.31 (s, 2H), 2.55 (s, 3H); ¹³C NMR (300 MHz, DMSO-d₆) δ 165.2, 158.7, 147.4, 145.2, 138.5, 125.5, 121.2, 24.7; ESI m/z (rel intensity): 207.7 (100).

[00145] Additional compounds wherein X is an amine or hydrazide group can be prepared starting with a suitable amine or hydrazide following the procedures outlined above.

General procedure for synthesis of oxadiazole derivatives [00146] To a solution of benzohydrazide (lOmmol) in absolute ethanol (2O mL) was added a solution of KOH (0.84g, 15mmol ) in water (2 mL) followed by carbon disulfide (0.9 mL, 15 mmol) and the resulting solution was heated under reflux for 4 - 8 hours till, release of hydrogen sulphide gas ceases. The reaction mixture was added to cold water and neutralized with concentrated hydrogen chloride. The solid thus separated was filtered, washed with water and dried. The dried solid was taken into ethyl acetate (10 mL) and stirred for 2 hours at room temperature and filtered the solid and dried.

Example 4: 5-(2-Ethoxyphenyl)-l, 3, 4-oxadiazole-2(3H)-thione; [00147] Following the general procedure outlined above the title compound was prepared. Yield (1.3g, 60%); M.P. 196-198°C; ¹H NMR (300 MHz, DMSO): δ 7.73- 7.69 (dd,lH, J = 1.8, 7.8 Hz), 7.6-7.5 (dd,lH, 7 = 9 Hz), 7.24-7.21 (d, IH, J = 8.4 Hz), 7.11-7.06 (dd, IH, J = 8.4 Hz), 4.19-4.12(q, 2H, J = 6.9 Hz ), 1.36-1.31(t, 3H, / = 6.9 Hz ) ; ¹³C NMR (300 MHz, CDCl₃): δ 177.8, 160.0, 157.5, 134.4, 130.3, 121.4, 114.3, 112.0, 64.9, 15.3; HRMS (EI) m/z 222.0461[M]⁺.

Example 5: 5-(Naphthalen-2-yl)-l,3,4-oxadiazole-2(3H)-thione. [00148] Following the general procedure outlined above the title compound was prepared. Yield (0.9g, 40%); M.P. 204-206⁰C; ¹H NMR (300 MHz, DMSO): δ 8.52 (s, IH), 8.17-8.15 (d,lH, J = 7.8 Hz), 8.12-8.09 (d, IH, J = 8.7 Hz), 8.03-8.01 (d, IH, J = 7.8 Hz), 7.94-7.91 (d, IH, J = 8.7 Hz), 7.70-7.62 (m, 2H); ¹³C NMR (300 MHz, CDCl₃): δ 178.2, 161.1, 134.9, 133.0, 129.9, 129.6, 129.0, 128.5, 128.0, 127.4, 122.7, 120.4; ESI m/z: 229.0[M+H]⁺.

Example 6: 5-(Naphthalen-2-yl)-l,3,4-oxadiazole-2(3H)-thione. [00149] Following the general procedure outlined above the title compound was prepared. Yield (0.9g, 40%); M.P. 204-206⁰C; ¹H NMR (300 MHz, DMSO): δ 8.52 (s, IH), 8.17-8.15 (d,lH, J = 7.8 Hz), 8.12-8.09 (d, IH, / = 8.7 Hz), 8.03-8.01 (d, IH, / = 7.8 Hz), 7.94-7.91 (d, IH, J = 8.7 Hz), 7.70-7.62 (m, 2H); ¹³C NMR (300 MHz,

CDCl₃): δ 178.2, 161.1, 134.9, 133.0, 129.9, 129.6, 129.0, 128.5, 128.0, 127.4, 122.7, 120.4; ESI m/z: 229.0[M+H]⁺.

General procedure for synthesis of thiadiazole derivatives

[00150] Potassium 2-benzoylhydrazinecarbodithioate (2 mmol) was added portion wise to concentrated sulfuric acid (1 mL) at O⁰C over a period of 30 minutes under argon and stirred at same temperature for 3 - 5hours and then reaction mixture was warmed up to room temperature. The reaction mixture was added to ice cold water and neutralized with sodium bicarbonate. The solid thus separated was filtered, washed with water, dried and purified by Flash column chromatography.

Example 7: 4-(Fluorophenyl)-l, 3, 4-thiadiazole-2(3H)-thione.

Following the general procedure outlined above the title compound was prepared. Yield (0.21g, 50%); M.P. 144-145⁰C; ¹H NMR (300 MHz, DMSO): δ 8.07-8.03 (dd,lH, y = 7.8 Hz), 7.43-7.38 (dd, IH, / = 7.2 Hz); ¹³C NMR (300 MHz, CDCl₃): δ 170.8, 165.4, 163.1, 130.9, 126.2, 117.3; ESI m/z: 212.5[M+H]⁺. Example 8: 5-(6-Methylpyridin-2-yl)-l ,3.4-thiadiazole-2(3H)-thione. Following the general procedure outlined above the title compound was prepared. Yield (0.15g, 35%); M.P. 208-210⁰C; ¹H NMR (300 MHz, DMSO): δ 7.89-7.82 (m, 2H), 7.45-7.42 (d,lH, J = 7.8 Hz), 2.51 (s, 3H) ¹³C NMR (300 MHz, CDCl₃): δ 170.5, 158.9, 148.5, 145.3, 136.0, 126.2, 113.5, 23.5; ESI m/z: 210.5[M+H]⁺.

Example 9: Evaluation of compound efficacy using a fluorescence resonance energy transfer (FRET) assay.

[00151] In order to assess efficacy, test compounds were evaluated using a FRET- based assay disclosed in Gorczynski, M. J., et al, (2007), Chem Biol 14, 1186-1197, and Matheny, C. J., et al, (2007), EMBO. J. 26, 1163-1175 (Fig. 2). A Cerulean-Runt domain and Venus-CBFβ complex at a 150 nM concentration (3*K_d, 1:1 ratio) was used for compound screening (Fig. 3). Test compounds were screened at a series of concentrations in a competition experiment using the fluorescence emission ratio (emission intensity at 525 nm / emission intensity at 474 nm, FRET ratio) as a read-out (Fig. 2). The dynamic range for the FRET assay was determined by adding a 20-fold excess of untagged CBFβ, and the associated change in the FRET ratio (~ 0.5) was defined as 100 % inhibition (Fig. 2).

Example 10: Development of inhibitors with increased potency.

[00152] The library of analogs synthesized above was used to identify several compounds with IC₅0 values in the low μM range, including RDI-2 (KG-3-275: IC50 = 2.0 ± 0.2 μM) and RDI-3 (KG-4-039: IC₅₀ = 4.6 ± 0.5 μM) (Fig. 4A). RDI-2 was the most potent compound within the series and therefore was used in the majority of the experiments described below. In addition, one compound, RDI-4 (KG-1-253), that shares the same molecular scaffold and virtually all the same functional groups but was a very weak inhibitor (IC₅0 > 500 μM) was used as a control. All of these compounds meet Lipinski's criteria (Lipinski, C. A. (2000), J Pharmacol Toxicol Methods 44, 235- 249) for orally bioavailable drugs. Active compounds were tested in the presence of detergent (0.1% TRITON X-100) to exclude the possibility of promiscuous inhibition by aggregate (micelle) formation (Feng, B. Y., et al, (2005), Nat Chem Biol 1, 146- 148). An ELISA assay to confirm the activity of these compounds by an independent method was performed. IC₅₀ values measured by ELISA correlated very well with FRET data (RDI-2: IC₅₀ (ELISA) = 1.0 ± 0.3 μM; RDI-3: IC₅₀ (ELISA) = 1.7 ± 0.4 μM; RDI-4: IC₅₀ (ELISA) > 300 μM).

Example 11 : Inhibition in HEK293 cells analyzed by FACS/FRET.

[00153] A FACS/FRET assay with Cerulean-Runt domain and Venus-CBFβ (Gorczynski et al., 2007) was used to test the cell permeability of the RDI compounds as well as their ability to disrupt the Runt domain-CBFβ interaction in mammalian cells. RDI-2 inhibited the Runt domain-CBFβ interaction in HEK293 cells in a dose dependent manner (Fig. 4B) whereas RDI-4 showed no inhibition, consistent with our in vitro results. There was no evidence of toxicity to the cells following two days of compound treatment based on forward/side scatter profiles.

Example 12: Verification of ligand binding to the Runt domain by NMR spectroscopy.

[00154] Nuclear Magnetic Resonance (NMR) spectroscopy was employed to establish that these compounds do indeed bind to the Runt domain. The saturation transfer difference (STD) method disclosed in Mayer, M., et al. (1999), Angewandte Chemie-International Edition 38, 1784-1788, and Mayer, M., et al. (2001), J. Am. Chem. Soc. 123, 6108-6117, was used to detect binding to proteins as well as to localize the site of binding. To enhance the sensitivity of the experiment, the size of the proteins was increased by using Cerulean-RD and Venus-CBFβ. A saturation transfer effect was observed only for the RDI-2 plus Cerulean-Runt domain mixture (Figs. 5A, 5B, 5C, and 5D). All protons from RDI-2, including the aromatic protons and methyl group, were observed in the STD spectrum (Fig. 5B, 5D), confirming that RDI-2 binds to the Runt domain. In contrast, the signal for DMSO (-2.5 ppm) is absent in the STD spectrum (Fig. 5D), consistent with its lack of interaction with Cerulean-RD and serving as a valid negative control. Importantly, the absence of signals on the RDI-2 plus Venus-CBFβ STD spectrum (Fig. 5C) confirms that RDI-2 does not interact with either CBFβ or Venus (nor by analogy to Cerulean). This was confirmed by collecting high quality ¹⁵N-¹H HSQC spectra for CBFβ in the presence of RDI-2. These results confirm selective binding of RDI-2 to the Runt domain. Example 13: RDI-2 decreases proliferation of the t(8;21) cell lines Kasumi-1 and SKNO-I.

[00155] The impact of the RDI compounds on growth of the t(8;21) containing cell lines Kasumi-1 and SKNO-I was assessed using an MTT (3-(4,5-Dimethylthiazol-2- yl)-2,5-diphenyltetrazolium bromide) assay (Fig. 6, Fig 7). Test compounds, RDI-2 and RDI-3 inhibited SKNO-I growth in a dose-dependent manner following 72 hours of treatment, whereas the inactive RDI-4 displayed no inhibition. Because RDI-4 is chemically very similar to the active compounds, its lack of activity supports the notion that the observed effects are due to specific targeting of AMLl-ETO (and possibly wildtype RUNX proteins) rather than off -target effects. RDI-2 also inhibited the growth of Kasumi-1 cells whereas RDI-4 only did so very modestly at the highest concentration (100 μM) (Fig. 7). The effects observed for RDI-2 with SKNO-I cells are more pronounced than those seen with Kasumi-1 cells. The concentration necessary for RDI-2 to achieve effects in cells is ~ 10-fold higher than the FRET IC₅₀ values, similar to what has been observed for the kinase assay versus cellular effects of Imatinib (Druker, B. J., et al, (1996), Nat Med 2, 561-566.) Since CBFβ is reported to protect Runxl from proteasome degradation (Huang, G., et al, (2001), EMBO J 20, 723-733), Kasumi-1 cells were treated with RDI-2 in combination with the proteasome inhibitor Z-Leu-Leu-Leu-al (MG132) (Palombella, V. J., et al, (1994), Cell 78, 773- 785.) but no attenuation of activity was observed, suggesting that RDI-2's effects are not caused by increased proteasome degradation of RUNXL

[00156] The activity of RDI-2 on a hematopoietic cell line lacking AMLl-ETO (U937), a kidney carcinoma cell line (HK-2), and a liver carcinoma cell line (HepG2) was determined to further assess the specificity. RDI-2 had no effect on the growth of HK-2 and HepG2 cells, and only a modest effect on U937 cells at the highest concentration (100 μM) (Fig. 8), indicating that it is not generally cytotoxic and has a significant degree of selectivity.

Example 14: RDI-2 increases apoptosis and alters cell cycle.

[00157] To determine the cause of growth inhibition the effects of RDI-2 on apoptosis and proliferation was examined. Treatment of Kasumi-1 and SKNO-I cells with RDI-2 caused a dose-dependent increase in Annexin V/PI positive cells (Fig. 9), similar to that caused by a ribozyme knockdown of AMLl-ETO (Matsushita, H., et al, (1999), J Cancer 79, 1325-1331) although not consistent with siRNA results (Martinez, N., et al, (2004), BMC Cancer 4, 44). A consistent decrease of cells in the S and G0/G1 phases of the cell cycle, a concomitant increase of cells in G2/M was observed, and a higher percentage of cells in the subG0/Gl compartment (Fig. 10), consistent with the Annexin V/PI results. No significant changes in any of these parameters were observed with the control RDI-4 compound.

Example 15: Microarray analysis identifies gene expression changes consistent with reduced AMLl-ETO activity.

[00158] As a further test of the RDI compounds' specificity a microarray analysis of Kasumi-1 cells was carried out following treatment with 50 μM RDI-2. A 6 hour treatment time was employed to focus on the likely direct effects of the compound on gene expression rather than more downstream events, as was also done in the compound signature studies used for the development of the Connectivity Map by Golub and co-workers (Lamb, J., et al, (2006), Science 313, 1929-1935). 1978 annotated genes whose expression was altered were identified. The table identifies compounds whose expression was shown to be altered both by introduction of AMLl- ETO into Lin- BM (Liu, Y., et al, (2007), Cancer Cell 11, 483-497) as well as by treatment with RDI-2 in Kasumi-1 cells and whose changes in gene expression are in opposite directions in these two data sets. [00159] Several genes dysregulated by AMLl-ETO and whose expression was affected by RDI-2 encode proteins that regulate proliferation. Notably, the expression of cyclin-dependent kinase inhibitor 1C (CDKNlC) decreased upon introduction of AMLl-ETO in Lin^" BM, but increased when Kasumi-1 cells were exposed to RDI-2 (Table 1). CDKNlC upregulation may contribute to the observed inhibitory effect of RDI-2 on proliferation. RAB33B, a member of the RAS oncogene family which is frequently mutated in human leukemia and provides a proliferative advantage, is upregulated with AMLl-ETO and down-regulated with RDI-2. Changes in the expression of genes that are regulated during myeloid cell differentiation were also observed. The expression of CD34, which encodes a well-established marker for immature cells that is highly expressed on the blastic fraction of cells from t(8;21) patients (Kita, K., et al, (1992), Blood 80, AlQ-AIl) increased in Lin^" BM upon introduction of AMLl-ETO, but decreased upon treatment with RDI-2 (Table 2). Conversely the expression of C/EBPα, a transcription factor essential for granulocyte differentiation and mutated in leukemia (Pabst, T., et al, (2001), Nat Genet 27, 263- 270), is decreased by AMLl-ETO and increased by RDI-2. The same trends in CD34 and C/EBPα expression were also documented upon siRNA knockdown of AMLl- ETO in Kasumi-1 cells (Dunne, J., et al, (2006), Oncogene 25, 6067-6078, and Martinez et al, 2004). Cyclin D3 (CCND3) is down with AMLl-ETO and up with RDI-2 (Table 2). Recent studies have shown that Cyclin D3 is essential for T-cell and B-cell differentiation (Cooper, A. B., et al, (2006), Nat Immunol 7, 489-497, and Sicinska, E., et al, (2003), Cancer Cell 4, 451-461). Importantly, the cyclin D3 gene has been clearly established as a direct target of RUNXl (Bernardin-Fried, F., et al,

(2004), J Biol Chem 279, 15678-15687), providing a clear example of upregulation of a direct RUNX target with RDI-2 treatment. The expression of RXRA, which heterodimerizes with the retinoic acid (RAR) and Vitamin D receptors (VDR) to regulate gene expression, was also upregulated by RDI-2. RXRA is an important regulator of myelopoiesis (Collins, S. J. (2002), Leukemia 16, 1896-1905), and was previously identified as a key component of an expression signature for differentiation based on a comparison of primary AML cells to normal donor neutrophils (Stegmaier, K., et al, (2004), Nat Genet 36, 257-263.). Thus RDI-2 treatment caused multiple changes in gene expression associated with increased myeloid differentiation.

Table 2. RDI-2 si nature following acute 6 hour) treatment of Kasumi-1 cells.

Kasumi-1 cells.

[00160] AMLl-ETO inhibits both vitamin D (vitD) and all-trans retinoic acid (ATRA)-induced differentiation of leukemic cell lines (Heidenreich, O., et al., (2003), Blood 101, 3157-3163 and Fazi, F., et al, (2007), Blood 109, 4432-4440). The inhibitory effect on vitD induced differentiation could be reversed in Kasumi-1 cells by reducing the AMLl-ETO concentration 2.5-fold with siRNA (Heidenreich et al., 2003). Similarly, the inhibitory effect on ATRA induced differentiation could be ameliorated in SKNO-I cells by reducing the AMLl-ETO concentration 2-fold with siRNA (Fazi et al., 2007). If RDI-2 was specifically targeting AMLl-ETO it was believed that it should restore vitD and ATRA sensitivity to Kasumi-1 or SKNO-I cells. ATRA only modestly promoted the differentiation of Kasumi-1 cells (measured by increased cell surface expression of CDl Ib), as reported previously (Fazi et al., 2007) (Fig. 11, Fig. 12). RDI-2 also modestly but significantly increased the percentage of CDl Ib⁺ cells. However, the addition of RDI-2 to either ATRA (Figs. 11, 12) or 9-cis retinoic acid (data not shown) caused a synergistic increase in the percentage of CDl Ib⁺ cells. In addition changes in cell morphology consistent with augmented differentiation

(decreased cell size, denser chromatin with folded and/or clefted nuclei, less prominent nucleoli, lower nuclear to cytoplasmic ratio, and granules in some cells) were observed upon treatment with RDI-2 plus ATRA (Fig. 13, Fig. 14). RDI-2 also synergized with 1 ,25-dihydroxy-vitamin D3 to increase the percentage Of CDl Ib⁺ cells (Fig. 12, Fig. 15) and induced similar morphological changes indicative of differentiation (decreased cell size, denser chromatin with folded and/or clefted nuclei, less prominent nucleoli, lower nuclear to cytoplasmic ratio, and increased granulation) (Figs. 13, Fig. 14). Thus, treatment with RDI-2 restored the ability of Kasumi-1 cells to respond to retinoid and vitamin D signaling. Our microarray analysis shows changes in the levels of NRIPl, NCOAl, and RXRA (Table 1), all of which are consistent with this increased sensitivity to ATRA and vitamin D. Since AMLl-ETO suppresses retinoid and vitamin D signaling, these data support the hypothesis that RDI-2 specifically inhibits AMLl- ETO activity.

[00161] Other methods which were used but not described herein are well known and within the competence of one of ordinary skill in the art of clinical, chemical, cellular, histochemical, biochemical, molecular biology, microbiology and recombinant DNA techniques.

[00162] The abbreviations used herein have their conventional meaning within the chemical and biological arts. AU publications, patents, and patent documents cited in the specification are incorporated by reference herein, as though individually incorporated by reference. In the case of any inconsistencies, the present disclosure, including any definitions therein will prevail. The invention has been described with reference to various specific and preferred embodiments and techniques. However, it should be understood that many variations and modifications may be made while remaining within the spirit and scope of the invention.

Claims

We Claim:

1. A compound of formula (I):

(D wherein each of Z¹, Z², Z³, Z⁴, or Z⁵ is independently C-R¹ or N; each R¹ is independently, hydrogen, halo, (Ci-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl, (Ci-C₆)alkoxy, thio(Ci-C₆)alkyl, -N(R^a)₂, halo(C₁-C₆)alkyl, (C₃-C₈)cycloalkyl, (C₃-C₈)cycloalkenyl, -C(=O)R⁵, -Q=O)Y¹R⁵, (C₆- Cio)aryl, (C₇-Ci₆)arylalkyl, (C₂-C₉)heterocyclic, (C₂-C₉)heteroaryl, (C₃- Ci₅)heteroarylalkyl, -SO₂N(R⁸)₂ or cyano; or two R¹ groups taken together with the atoms to which they are attached can form a ring; each R⁸ is independently hydrogen, (Ci-C₆)alkyl or (C₆-CiO) aryl;

Y¹ is O, S or NR^b; each R^b is independently hydrogen or (C_rC₆)alkyl; each R^a is independently hydrogen, (C_rC₆)alkyl, -C(=O)R⁶, or -C(=O)Y²R⁶;

Y² is O, S or NR^C; each R^c is independently hydrogen or (C]-C₆)alkyl; each R⁵ and R⁶ is independently hydrogen, (Ci-C₆)alkyl, halo(Ci-C₆)alkyl, (C₃-C₈)cycloalkyl, (C₆-C₁₀)aryl, or (C₇-C i₆)alkylaryl; each R² is independently hydrogen, (Ci-Ce)alkyl, heterocyclic, or heterocyclic(Ci-C₆)alkyl or absent; '^~~s represents an optional double bond;

X is S, O, or NR³; R³ is hydrogen, or N(R⁴)₂, where each R⁴ is independently hydrogen, (Ci-C₆)alkyl or -C(=O)R⁷; R⁷ is hydrogen or (Q- C₆)alkyl; wherein at least two of Z¹, Z², Z³, Z⁴, and Z⁵ are C-R¹; where any of alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclic, or heteroaryl groups in R¹ and R², R³ or R⁴ can be optionally substituted with 1, 2, 3, or 4 substituent groups, where the substituent groups independently are halo, oxo (=O), imino (=NR ), (Ci-Cio)alkyl, (Ci-Cio)alkoxy, or C₆-aryl, -C(=O)R⁷, -C(=O)Y³R⁷; -N(R^d)₂ or cyano; each R⁷ is independently hydrogen, (Ci-C₆)alkyl, halo(C_rC₆)alkyl, (C₃-C₈)cycloalkyl, (C₆-C₁₀)aryl, or (C₇-Ci₆)alkylaryl; Y³ is O or S; or wherein one or more of the carbon atoms in the alkyl, alkenyl, or alkynyl groups can be independently replaced with non-peroxide oxygen, sulfur or NR^d; each R^d is independently hydrogen or (Ci-C₆)alkyl; provided that: when X is N-NH₂; and Z¹, Z², Z⁴, and Z⁵ are C-H; then Z³ is not C-F; when X is NH; and Z¹, Z², Z³, and Z⁵ are C-H, then Z⁴ is not N; and when X is NH; and Z¹, Z², Z⁴^r Z⁵ are C-H, then Z³ is not N; or a pharmaceutically acceptable salt thereof.

2. The compound of claim 1 , of the formula:

wherein each Z¹ is independently C-R¹ or N; each R¹ is independently hydrogen, halo, (Ci-Ce)alkyl, (Ci-C₆)alkoxy, thio(Ci-C₆)alkyl, -N(R^a)₂, halo(C_rC₆)alkyl, (C₃-C₈)cycloalkyl, (C₃- C₈)cycloalkenyl, -SO₂N(R⁸)₂, or (C₆-C₁₀)aryl; or two R¹ groups taken together with the atoms to which they are attached can form a ring; and

R² is hydrogen, (Ci-C₆)alkyl, heterocyclic, or heterocyclic (Ci -Ce)alkyl.

3. The compound of claims 1 or 2, wherein each R¹ is independently hydrogen, methyl, ethyl, propyl, methoxy, ethoxy, propoxy, fluoro, chloro, bromo, trifluoromethyl, pentafluoroethyl, -N(R^a)₂ or -SO₂N(R⁸)₂; and R^a is hydrogen, methyl, ethyl, propyl, -C(=O)CH₃, -C(_^O)CH₂CH₃, -C(=O)OCH₃, C(=O)OCH₂CH₃. -C(=O)NHCH₃, or -Q=O)NHCH₂CH₃; or two R¹ groups are taken together with the atoms to which they are attached to form a ring.

4. The compound of claims 1 or 2, wherein each R¹ is independently hydrogen, methyl, ethyl, methoxy, ethoxy, fluoro, chloro, trifluoromethyl, -N(R^a)₂ -SO₂N(R⁸)₂; R^a is hydrogen, methyl, or ethyl; and R⁸ is hydrogen, methyl, ethyl, or phenyl; or two R¹ groups are taken together with the atoms to which they are attached to form a ring.

5. The compound of any of claims 1-4, wherein X is NR³; R³ is hydrogen, or N(R⁴)₂, where each R is independently hydrogen or (Ci-Cio)alkyl;.

6. The compound of claim 5, wherein each R¹ is independently hydrogen, methyl, fluoro, trifluoromethyl, -N(CHs)₂ or -SO₂NH₂; or two R¹ groups are taken together with the atoms to which they are attached to form a ring; and R is N(R⁴)₂, where each R⁴ is independently hydrogen or methyl.

7. The compound of claim 6, wherein R is NH₂.

8. The compound of claim 7, of the formula:

9. The compound of claim 8, of the formula:

10. The compound of any of claims 1-4, wherein X is S.

11. The compound of claim 10, wherein each R¹ is independently hydrogen, methyl, ethyl, methoxy, ethoxy, fluoro, trifluoromethyl or -N(CHs)₂ or - SO₂NH₂; or two R¹ groups are taken together with the atoms to which they are attached to form a ring.

12. The compound of claim 11 , of the formula:

13. The compound of claim 12, of the formula:

14. The compound of any of claims 1-4, wherein X is O.

15. The compound of claim 14, wherein R is hydrogen, methyl, ethyl, methoxy, ethoxy, fluoro, trifluoromethyl or -N(CHs)₂ or -SO₂NH₂; or two R¹ groups are taken together with the atoms to which they are attached to form a ring.

16. The compound of claim 15, of the formula:

17. The compound of claim 16, of the formula:

18. A composition comprising a compound of any of claims 1-17 and a pharmaceutically acceptable carrier.

19. The composition of claim 18, further comprising at least one additional antileukemic drug or agent.

20. The composition of claim 19, wherein the anti-leukemic drug or agent is all trans retinoic acid, daunorubicin, or cytarabine.

21. A method for prevention or treatment of leukemia, comprising administering to a mammal in need of such treatment, an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof.

22. A method for inhibiting transcription factors of leukemia cell lines comprising administering to a mammal (e.g., a human) in need of such treatment an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof.

23. The method of claim 21 or 22, where the leukemia is lymphocytic leukemia, chronic lymphocytic leukemia, acute myelogenous leukemia or chronic myelogenous leukemia.

24. The method of claim 21 - 23, further comprising administering at least one additional drug in a combination therapy.

25. The method of any of claims 21 - 24, where the leukemia is acute myelogenous leukemia.

26. A compound of any of claims 1-17, for use in medical therapy.

27. Use of a compound of any of claims 1-17, to prepare a medicament for use in the prevention or treatment of a pathological condition or symptom in a mammal, wherein the pathological condition or symptom is a leukemia.

28. The use of claim 26, wherein the leukemia is lymphocytic leukemia, chronic lymphocytic leukemia, acute myelogenous leukemia or chronic myelogenous leukemia.

29. The use of claim 26 or 27, wherein the medicament comprises a liquid carrier.

30. The use of claim 26 or 27, wherein the medicament comprises a solid carrier.

31. A compound of any of claims 1-17, for use in the prevention or treatment of a pathological condition or symptom in a mammal, wherein the pathological condition or symptom is as defined in claim 26 or 27.