PAN-SELECTIN INHIBITOR WITH ENHANCED PHARMACOKINETIC ACTIVITY

BACKGROUND

Technical Field The present invention relates generally to compounds, compositions and methods for treating diseases or complications associated therewith, in which a selectin plays a role such as sickle cell disease or a cancer involving a selectin. More specifically, the present invention relates to particular glycomimetics and uses thereof.

Description of the Related Art Selectins play, and are being discovered to play, a role in a variety of diseases and complications associated therewith. Inhibitors of selectins may be used to treat such diseases or complications.

Sickle cell disease is an inheritable hematological disorder based on a mutation in the β-globin gene of hemoglobin. Upon deoxygenation, this mutated hemoglobin polymerizes and causes a shape change (sickling) of the red blood cell.

This change in red blood cells leads to obstruction of blood vessels causing a wide variety of complications such as stroke, pulmonary hypertension, end-organ disease and death.

In addition to the fatal or potentially fatal complications, there are serious non-fatal complications of sickle cell disease such as pain. The severity of the pain may vary, but normally requires some form of medical attention. Hospitalization may be necessary.

In the U.S. alone, approximately 70,000-80,000 people suffer from sickle cell disease. Sickle cell disease is estimated to affect one of every 1,300 infants in the general population, and one of every 400 of African descent. Currently, there is no cure for sickle cell disease. The disease is chronic and lifelong. Life expectancy is typically shortened.

Accordingly, there is a need in the art for the treatment of sickle cell disease or the complications associated therewith. The present invention fulfills these needs and further provides other related advantages.

BRIEF SUMMARY Briefly stated, compounds, compositions and methods for treating diseases or complications associated therewith, in which a selectin plays a role, are provided. In the present invention, the compounds used for treatment are particular glycomimetics. Such compounds may be combined with a pharmaceutically acceptable carrier or diluent to form a pharmaceutical composition. The present compounds are inhibitors of selectins. Such compounds may be used to treat a variety of diseases or complications associated therewith, including sickle cell disease, a disease involving the migration of T-cells to the skin, acute myocardial infarction, or a cancer involving a selectin.

In one embodiment, the present invention provides a compound having the formula:

wherein

Bz = benzoyl;

Q is H or a physiologically acceptable salt;

L = linker group;

P = O-I; and

R ¹ is one of

ϊ1 X HV ^Ar " IηY ^Ar V YV ^Ar <w T> I ψ i ^N(CH3)2

where Ar is aryl, Q is H, a physiologically acceptable salt, C ₁ -C ₈ alkanyl, C ₁ -C ₈ alkenyl, Cj-C ₈ alkynyl, aryl, or (CH ₂ ) _m -aryl where m is 1-10, n = 1-4, and Z and Y are independently selected from Ci-C ₈ alkanyl, Ci-C ₈ alkenyl, Ci-C ₈ alkynyl, halogenated Ci-C ₈ alkanyl, and aryl substituted with Me, OMe, halide, OH, and R is CN, OH, NH ₂ , Ci-C ₈ alkanyl, Cj-C ₈ alkenyl, Ci-C ₈ alkynyl, aryl, or (CH ₂ ) _m -aryl where m is 1-10. In one embodiment, a present compound may be combined with a pharmaceutically acceptable carrier or diluent to form a composition.

In one embodiment, the present invention provides a method for the treatment of sickle cell disease or a complication associated therewith in an individual who is in need thereof, comprising administering to the individual a compound in an amount effective for treatment of sickle cell disease or a complication associated therewith, wherein the compound is set forth above.

In one embodiment, the present invention provides a method for the treatment of a disease involving the migration of T-cells to the skin or a complication associated therewith in an individual who is in need thereof, comprising administering to the individual a compound in an amount effective for treatment of a disease involving the migration of T-cells to the skin or a complication associated therewith, wherein the compound is set forth above.

In one embodiment, the present invention provides a method for the treatment of acute myocardial infarction or a complication associated therewith in an

individual who is in need thereof, comprising administering to the individual a compound in an amount effective for treatment of acute myocardial infarction or a complication associated therewith, wherein the compound is set forth above.

In one embodiment, the present invention provides a method for the treatment of a cancer involving a selectin or a complication associated therewith in an individual who is in need thereof, comprising administering to the individual a compound in an amount effective for treatment of a cancer involving a selectin or a complication associated therewith, wherein the compound is set forth above.

In other embodiments, the above compounds or compositions thereof may be used, and in the manufacture of a medicament, for any of the uses recited herein.

These and other aspects of the present invention will become apparent upon reference to the following detailed description and attached drawings. All references disclosed herein are hereby incorporated by reference in their entirety as if each was incorporated individually.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is a diagram illustrating the synthesis of a glycomimetic.

Figure 2 is a diagram illustrating the synthesis of aminonaphthalene trisulfonic acid with linker. Figure 3 is a diagram illustrating the synthesis of a compound formed by coupling the compound of Figure 1 with the compound of Figure 2.

Figure 4 is a diagram illustrating the modification of the compound of Figure 3 (beginning with compound XVIII of Figure 1 and using compound C of Figure 2), whereby the -COOH group is converted to a substituent selected from R ¹ as defined herein.

Figure 5 is a diagram illustrating the modification of a compound such as the compound of Figure 3, whereby the -COOH group is converted to a substituent selected from R ¹ as defined herein.

DETAILED DESCRIPTION

As noted above, the present invention provides compounds, compositions and methods for the treatment of diseases or complications associated therewith, in which a selectin plays a role in an individual. The compounds are inhibitors of selectins, and have a variety of uses in vitro and in vivo.

The present compounds have the formula:

In the above formula and the present disclosure generally, there are several abbreviations. "Bz" is benzoyl. "Ar" is aryl. "Et" is ethyl. "Me" is methyl. As any acids in the above formula may be in the form of a free acid or a salt, the various forms are encompassed by "Q" where denoted. "Q" is hydrogen (H) or a physiologically acceptable salt, unless defined otherwise herein. Examples of physiologically acceptable salts are Na, K, Li, Mg and Ca. Where groups may be present in multiples or not at all, the arbitrary letters "p", "n", "m" and "q" represent ranges of integers as defined herein. As used herein, the term "independently selected" refers to the selection of identical or different substituents.

In the above formula, "L" represents a linker which joins the carbon of C(=O) of the glycomimetic portion of the compound to NH of the diphenyl portion (i.e., to an aminonaphthalene trisulfonic acid). There may be no linkers present (i.e., "p" is 0) or a linker may be present (i.e., "p" is 1). Where no linker is present, the compound is with the formula:

Where p is 1 , a linker is present. A linker may include a spacer group, such as -(CH ₂ ) _q - or -0(CH ₂ ) _q - where q is generally about 1-20 (including any whole integer range therein). Other examples of spacer groups include a carbonyl or carbonyl containing group such as an amide. Embodiments of linkers include the following:

Squaric acid Thiourea

Dithiadiazoleoxide Acylation via Thiofuran

H O H ₂ H O O H

I Il Il I

— N-C- (CH ₂ ) ₂ — C-NH — -N-C- (CH ₂ )n— C-N-

N-Pentenoylation and Reductive Coupling Via Bifunctional amination NHS reagent

Other linkers, e.g., polyethylene glycols (PEG) or -C(=O)-NH-(CH ₂ ) _q -C(=O)-NH ₂ where q is as defined above, will be familiar to those in the art or in possession of the present disclosure.

In another embodiment, the linker is which produces:

In another embodiment, the linker is

which produces:

In another embodiment, the linker is

which produces :

The present compounds, having the formula set forth above with (L) _p , possess the substituent "R ¹ ". R ¹ is one of

where Ar is aryl, Q is H, a physiologically acceptable salt, C ₁ -C ₈ alkanyl, C ₁ -C ₈ alkenyl, C ₁ -C ₈ alkynyl, aryl, or (CH ₂ ) _m -aryl where m is 1-10, n = 1-4, and Z and Y are independently selected from C ₁ -C ₈ alkanyl, C ₁ -C ₈ alkenyl, C ₁ -C ₈ alkynyl, halogenated Ci-C ₈ alkanyl, and aryl substituted with Me, OMe, halide, OH, and R is CN, OH, NH ₂ ,

Ci-C ₈ alkanyl, Ci-C ₈ alkenyl, Ci-C ₈ alkynyl, aryl, or (CH ₂ ) _m -aryl where m is 1-10.

O In an embodiment, R ¹ is Ji . This embodiment applies to where

' NH ₂ the linker is (L) _p or any of the specific linkers disclosed herein.

O

In another embodiment, R ¹ is /^N^ ^CN • Thi ^s embodiment applies to

H where the linker is (L) _p or any of the specific linkers disclosed herein.

O

In another embodimen tt,, RR ¹ iiss //^^ _N N' ^0H .• TThhiiss embodiment applies to

H where the linker is (L) _p or any of the specific linkers disclosed herein.

As used herein, a "Ci-Cg alkanyl" refers to an alkane substituent with one to eight carbon atoms and may be straight chain, branched or cyclic (cycloalkanyl). Examples are methyl, ethyl, propyl, isopropyl, butyl and t-butyl. A "halogenated Ci-C ₈ alkanyl" refers to a "Ci-C ₈ alkanyl" possessing at least one halogen. Where there is more than one halogen present, the halogens present may be the same or different or both (if at least three present). A "Ci-C ₈ alkenyl" refers to an alkene substituent with one to eight carbon atoms, at least one carbon-carbon double bond, and may be straight

chain, branched or cyclic (cycloalkenyl). Examples are similar to "Cj-C ₈ alkanyl" examples except possessing at least one carbon-carbon double bond. A "Ci-Cg alkynyl" refers to an alkyne substituent with one to eight carbon atoms, at least one carbon-carbon triple bond, and may be straight chain, branched or cyclic (cycloalkynyl). Examples are similar to "C ₁ -C ₈ alkanyl" examples except possessing at least one carbon-carbon triple bond. An "alkoxy" refers to an oxygen substituent possessing a "C ₁ -C ₈ alkanyl," "Ci-C ₈ alkenyl" or "C _r C ₈ alkynyl." This is -O-alkyl; for example methoxy, ethoxy, n-propoxy, i-propoxy, n-butoxy and the like; and alkenyl or alkynyl variations thereof (except for methoxy). It further refers to the group O-alkyl- W-alkyl where W is O or N; for example -O-(CH ₂ ) _n - W-(CH ₂ ) _m where n and m are independently 1-10. An "aryl" refers to an aromatic substituent with one to fourteen carbon atoms as ring atoms in one or multiple rings which may be separated by a bond or fused. As used herein, "aryl" includes "heteroaryl." A "heteroaryl" is similar to an "aryl" except the aromatic substituent possesses at least one heteroatom (such as N, O or S) in place of a ring carbon. Where an aromatic substituent is an aryl in which all the ring atoms are carbon (/. e. , not a heteroaryl), there are typically six to fourteen ring atoms. Where an aryl is a heteroaryl, there may be less than six carbon ring atoms. Examples of aryls include phenyl, naphthyl, pyridinyl, pyrimidinyl, triazolo, furanyl, oxazolyl, thiophenyl, quinolinyl and diphenyl. The following confirms what would be understood by one of skill in the art in possession of the present disclosure. The line to which no element is attached at one end of any R ¹ substituent disclosed herein is intended to be the same line (i.e., the same bond) that joins R ¹ in any of the compound formula depicted above.

All compounds of the present invention or useful thereto (e.g., for pharmaceutical compositions or methods of treating), include physiologically acceptable salts thereof. Examples of such salts are Na, K, Li, Mg, Ca and Cl.

Compounds as described herein may be present within a pharmaceutical composition. A pharmaceutical composition comprises one or more compounds in combination with (i.e., not covalently bonded to) one or more pharmaceutically or physiologically acceptable carriers, diluents or excipients. Such compositions may

comprise buffers (e.g., neutral buffered saline or phosphate buffered saline), carbohydrates (e.g., glucose, mannose, sucrose or dextrans), mannitol, proteins, polypeptides or amino acids such as glycine, antioxidants, chelating agents such as EDTA or glutathione, adjuvants (e.g., aluminum hydroxide) and/or preservatives. Within yet other embodiments, compositions of the present invention may be formulated as a lyophilizate. Compositions of the present invention may be formulated for any appropriate manner of administration, including for example, topical, oral, nasal, intravenous, intracranial, intraperitoneal, subcutaneous, or intramuscular administration. The compositions described herein may be administered as part of a sustained release formulation (i.e., a formulation such as a capsule or sponge that effects a slow release of compound following administration). Such formulations may generally be prepared using well known technology and administered by, for example, oral, rectal or subcutaneous implantation, or by implantation at the desired target site. Carriers for use within such formulations are biocompatible, and may also be biodegradable; preferably the formulation provides a relatively constant level of compound release. The amount of compound contained within a sustained release formulation depends upon the site of implantation, the rate and expected duration of release and the nature of the condition to be treated. The above described compounds including equivalents thereof are useful in methods of the present invention to treat individuals in need thereof. As used herein, such individuals include humans, as well as non-human warm-blooded animals such as non-human mammals. A preferred individual for treatment is a human. Typically a compound will be administered to an individual as a pharmaceutical composition, i.e., in combination with a pharmaceutically acceptable carrier or diluent.

In an embodiment, an individual who is in need of treatment for sickle cell disease or a complication associated therewith is administered at least one (i.e., one or more) of the above described compounds in an amount effective for the treatment. As used herein, the term "treatment" (including variations such as "treating") includes prevention. For example, a complication associated with sickle cell disease may not

have presented itself in an individual with the disease, and a compound may be administered to prevent presentation of the complication in the individual. Sickle cell disease and complications associated therewith include, for example, anemia, red blood cells becoming stuck in blood vessels, ischemia, infarction, stroke, acute chest crisis, splenic sequestration crisis, shortened life expectancy, organ damage and periodic or chronic pain.

In an embodiment, an individual who is in need of treatment for a disease involving the migration of T-cells to the skin or a complication associated therewith is administered at least one (i.e., one or more) of the above described compounds in an amount effective for the treatment. Examples of diseases involving the migration of T-cells to the skin include atopic dermatitis, psoriasis, cutaneous T-cell lymphoma, and graft vs. host disease following bone marrow transplantation. Complications associated with diseases involving the migration of T-cells to the skin include redness of the skin, swelling of the skin, and accumulation of T-cells or T-cell lymphomas in the skin.

In an embodiment, an individual who is in need of treatment for acute myocardial infarction or a complication associated therewith is administered at least one (i.e., one or more) of the above described compounds in an amount effective for the treatment. An acute myocardial infarction includes those that are post ischemia or post reperfusion. Complications associated with acute myocardial infarction include chest pain, shortness of breath and syncope.

In an embodiment, an individual who is in need of treatment for a cancer involving a selectin or a complication associated therewith is administered at least one (i.e., one or more) of the above described compounds in an amount effective for the treatment. Complications associated with a cancer involving a selectin include shortened life expectancy, organ damage, and periodic or chronic pain.

The term "treatment," as set forth above, refers to any of a variety of positive effects from the treatment including, for example, eradicating a complication associated with the disease, relieving to some extent a complication, slowing or stopping progression of the disease, and prolonging the survival time of the recipient.

The treatment may be used in conjunction with one or more other therapies for any of the illnesses (or complications associated therewith) described above.

The above described compounds may be administered in a manner appropriate to the disease to be treated. Appropriate dosages and a suitable duration and frequency of administration may be determined by such factors as the condition of the patient, the type and severity of the patient's disease and the method of administration. In general, an appropriate dosage and treatment regimen provides the compound(s) in an amount sufficient to provide therapeutic and/or prophylactic benefit. Within particularly preferred embodiments of the invention, a compound may be administered at a dosage ranging from 0.001 to 1000 mg/kg body weight (more typically 0.01 to 1000 mg/kg), on a regimen of single or multiple daily doses. Appropriate dosages may generally be determined using experimental models and/or clinical trials. In general, the use of the minimum dosage that is sufficient to provide effective therapy is preferred. Patients may generally be monitored for therapeutic effectiveness using assays suitable for the condition being treated, which will be familiar to those of ordinary skill in the art.

The following Examples are offered by way of illustration and not by way of limitation.

EXAMPLES

EXAMPLE 1 SYNTHESIS OF GLYCOMIMETIC (FIG. 1)

Synthesis of intermediate II: (-)-Shikimic acid (20 g) in MeOH (200 ml) and sulfuric acid (2 ml, 98%) are stirred at room temperature (rt) for 50 h. The reaction mixture is neutralized with 2N aqueous NaOH in the cold. After evaporation to dryness, the residue is purified by silica gel chromatography to afford II (19.2 g).

Synthesis of intermediate III: Methyl shikimate (II, 10 g), 2,2- dimethoxypropane (10 ml) and p-TsOH (0.8 g) are dissolved in acetonitrile (125 ml) and stirred at rt for 1 h. The reaction mixture is then neutralized with triethylamine (2 ml) and evaporated to dryness. The residue is chromatographed on silica gel to yield III

(i i g).

Synthesis of intermediate IV: The shikimic acid derivative III (10 g) and PtO ₂ ZC (10%, 250 mg) in MeOH (40 ml) are hydrogenated at rt under vigorous stirring. After 16 h the reaction mixture is filtered over celite and evaporated to dryness. The residue is chromatographed on silica gel to yield IV (8.6 g).

Synthesis of intermediate V: To a solution of IV (8 g) in DCM (100 ml) at O ⁰ C are added pyridine (12 ml), acetic anhydride (7 ml) and a DMAP (25 mg). The reaction mixture is stirred at rt for 1 h, and diluted with EtOAc (250 ml). After washing with 0.5 M aqueous HCl (3 x 50 ml), saturated solution of KHCO ₃ (3 x 50 ml) and brine (3 x 50 ml), the combined organic layers are dried (Na ₂ SO ₄ ) and evaporated to dryness. The residue is purified by chromatography on silica gel to yield V (6.8 g).

Synthesis of intermediate VI: A solution of V (6.0 g) in acetic acid (30 ml, 80%) is stirred at 80°C for 1 h. Solvent is evaporated off and the residue is purified by chromatography on silica gel (DCM/MeOH 14:1) to yield VI (3.6 g). Synthesis of intermediate TVID: A solution of VI (3 g) and p-TsC\ (3.5 g) in pyridine (30 ml) is stirred at rt for 6 h. MeOH (5 ml) is added and the solvent is evaporated at reduced pressure, the residue dissolved in EtOAc (3 x 150 ml) and the organic layers are washed with 0.5 M aqueous HCl (0 ⁰ C), water (cold) and brine (cold). The combined organic layers are dried (Na ₂ SO ₄ ), filtered on Celite and evaporated to

dryness. The residue is purified by chromatography on silica gel (toluene/EtOAc 4:1) to yield VII (3.7 g).

Synthesis of compound VIII: A solution of VII (3 g) and NaN ₃ (2.5 g) in DMF (20 ml) is stirred at 8O ⁰ C. The reaction mixture is cooled to rt and diluted with EtOAc (200 ml) and water (50 ml). The organic layer is additionally washed twice with water (2 x 50 ml) and once with brine (50 ml). All aqueous layers are extracted twice with EtOAc (2 x 50 ml). The combined organic layers are dried with Na ₂ SO ₄ , filtered and the solvent is evaporated off. The residue is purified by chromatography on silica gel (petroleum ether/EtOAc 5:2) to give VIII (2.2 g). Synthesis of compound X: To a solution of ethyl 2,3,4-tri-O-benzyl-α-L- fucothiopyanoside IX (1.5 g) in DCM (3 ml), bromine (150μl) is added at 0°C under argon. After 5 min the cooling bath is removed and the reaction mixture is stirred for an additional 25 min at rt. Cyclohexene (200μl) is added and the reaction mixture is added to a solution of VIII (400 mg), (Et) ₄ NBr (750 mg) and powdered 4 A molecular sieves in DCM (10 ml) and DMF (5 ml). After 16 h, triethylamine (1.5 ml) is added and stirred for an additional 10 min, diluted with EtOAc (50 ml) and washed with sat. aqueous NaHCO ₃ , water and brine. The aqueous layers are extracted twice with EtOAc (2x 50 ml). The combined organic layers are dried (Na ₂ SO ₄ ), filtered and evaporated to dryness. The residue is purified by chromatography on silica gel (toluene/EtOAc 9:1) to yield X (700 mg).

Synthesis of compound XI: To a solution of X (1.5 g) in MeOH (20 ml) is added freshly prepared NaOMe (80 mg) and the reaction mixture is stirred in a pressure tube at 8O ⁰ C for 20 h. The reaction mixture is cooled to rt and neutralized with acetic acid. Solvent is evaporated to dryness and the residue is dissolved in ether. Freshly prepared diazomethane is added and the excess diazomethane is neutralized with acetic acid. Solvent is evaporated off to give XI (1.25 g).

Synthesis of building block XV: This synthesis is done exactly in same way as described previously (Helvetica Chemica Acta 53:2893-2907 (2000)).

Synthesis of compound XVI: A mixture of XI (1.6 g), XV(3 g) and activated powdered molecular sieves 4 A (1 g) in DCM (17 ml) is stirred at rt under argon for 2 h. Then DMTST (2 g) is added in 4 equal portions over a period of 1.5 h. After 24 h the reaction mixture is filtered over Celite and the filtrate is diluted with

DCM (100 ml). The organic layer is washed with sat. aqueous NaHCO ₃ and brine and the aqueous layers are extracted twice with DCM. The combined organic layers are dried (Na ₂ SO ₄ ), filtered and evaporated to dryness. The residue is purified by chromatography on silica gel (toluene/ EtOAc 8:1) to yield XVI (1.5 g). Synthesis of compound XVII: To a solution of XVI (500 mg) and orotic acid chloride (500 mg) in dichloromethane (10 ml) is added a solution of triphenylphosphine (500 mg in 5 ml dichloromethane) dropwise during 10 min. The reaction mixture is stirred at rt for 25 h and the solvent is evaporated off. The residue is purified (chromatography on silica gel DCM/MeOH 19:1) to give XVII (250 mg). Synthesis of compound XVIII: To a solution of XVII (200 mg) in dioxane- water (5:1, 12 ml) is added 10% Pd-C (100 mg) and the reaction mixture is stirred vigorously under hydrogen (55 psi) for 24 h. Catalyst is filtered through a bed of celite and the solvent is evaporated off. Residue is purified by silica gel chromatography to give compound XVIII (150 mg). Synthesis of XIX: To a solution of compound XVIII (145 mg) in MeOH

(5 ml) is added a solution of NaOMe in MeOH (25%, 0.025 ml) and the reaction mixture is stirred at rt for 4 h, neutralized with acetic acid and the solvent is evaporated off. Residue is dissolved in water and passed through a bed of Dowex 50wX-8 (Na- form) resin. Water wash is evaporated off to afford compound XIX (100 mg). Synthesis of EDA-XIX: XIX (80 mg) is heated at 70 ⁰ C with ethylenediamine (EDA) (1 ml) with stirring for 5 h. Solvent is evaporated off and then purified by sephadex G-25 column to give EDA-XIX (82 mg).

EXAMPLE 2 SYNTHESIS OF AMINONAPHTHALENE TRISULFONIC ACID WITH LINKER (FIG. 2)

Synthesis of intermediate A: To a solution of 3,6-dioxaoctanedioic acid

(4.34 g) in DMF (220 ml) is added triethylamine (6.2 ml) and cooled to 0° C. To this solution is added a solution of benzyl bromide (2.4 ml) in DMF (220 ml) dropwise with stirring over a 2 h period at 0° C and continued to stir at the same temperature for 4 h. After 4 h, the reaction mixture is slowly warmed to room temperature and stirred at

room temperature for overnight. Solvent is removed under reduced pressure and the residue is dissolved in IM HCl in saturated NaCl solution (500 ml). The aqueous solution is extracted with EtOAc (6x200ml). The combined organic extracts are dried (Na ₂ SO ₄ ) and concentrated to dryness. The residue is purified by flush chromatography to give compound A (2g).

Synthesis of intermediate B: To a solution of compound A (Ig) in DMF (10ml) is added diisopropylethylamine (0.45ml) and then HATU (1.2g). A solution of 8-aminonaphthalene-l,3,6-trisulfonic acid (1.5 g) in DMF (10ml) and diisopropylethylamine (0.45ml) is added to the above solution after 5 min with stirring. The reaction mixture is stirred at room temperature for 1 h. The solvent is evaporated off under reduced pressure and the residue is purified by reversephase (C- 18) column to give compound B (0.5g).

Synthesis of compound C: Compound B (0.5g) is dissolved in an aqueous solution of K ₂ CO ₃ and the pH of the solution is adjusted to 10 by adding IN solution of NaOH with stirring. The reaction mixture is stirred at room temperature for 1 h, neutralized to pH 7 with IN HCl, solvent is evaporated off and the residue is purified on a sephadex G-IO column to give compound C (0.2g).

EXAMPLE 3 SYNTHESIS OF COMPOUND D (FIG. 3)

EDA-XIX (Example 1) and compound C (Example 2) are each co- evaporated once with DMF separately. Compound C (421 mg, 0.452 mmol) is dissolved in 5 mL of anhydrous DMF at it To the compound C solution is added 400 μL (2.3 mmol) of DIPEA followed by 154 mg (0.406 mmol) of HATU. The reaction solution is stirred at rt for 5 min. EDA-XIX (208 mg, 0.226 mmol) is partially dissolved in 5 mL of anhydrous DMF. The compound C reaction solution is transferred to the EDA-XIX mixture at rt. The compound C reaction flask is rinsed with an additional 5 mL of anhydrous DMF and the solution is transferred to the EDA-XIX reaction flask. After stirring for 1 hr at rt, the reaction is concentrated in vacuo. The resulting yellow residue is dissolved in dH ₂ O and purified on a Waters SepPak Cl 8 (5g)

cartridge. The Cl 8 cartridge is eluted with dH ₂ O (3 x 8OmL), 10% MeOH in dH ₂ O (1 x 8OmL), 20% MeOH in dH ₂ O (2 x 8OmL), 30% MeOH in dH ₂ O (1 x 8OmL) and 50% MeOH in dH ₂ O (I x 8OmL). All fractions containing methanol are concentrated separately and further purified by HPLC using a Gemini Cl 8 column (Phenomenex) using 1OmM NH ₄ Ac and CH ₃ CN mixture as mobile phase to give compound D as ammonium salt. Compound D is further converted to Na-salt form by passing through a column of ion-exchange resin (IR- 120 sodium form). Yield 150 mg.

EXAMPLE 4 SYNTHESIS OF COMPOUND XVIII-D (FIG. 4)

Synthesis of intermediate XVIII-A: To a cool (5°C) solution of XVIII

(100 mg ) (Example 1) in THF (0.8 ml) is added triethylamine (0.025ml) and ethylchloro formate (0.014ml) under argon. The reaction mixture is stirred for 15 min in the cold and the ice-bath is removed. A solution (2M) of (CH ₃ ) ₂ NH in THF (0.07 ml) is added and stirred for Ih at room temperature. The reaction mixture is evaporated to dryness and purified by column chromatography (silica gel) to give compound XVIII-A (55 mg).

Synthesis of intermediate XVIII-B: Compound XVIII-A (50 mg) is stirred at room temperature with a 0.1N solution of NaOMe in MeOH (ImI) for 3h. AcOH (0.02ml) is added with stirring and the solvent is evaporated to dryness. Solid residue is purified by column chromatography (silica gel) to give intermediate XVIII-B (32mg).

Synthesis of intermediate XVIII-C: Compound XVIII-B (30mg) is dissolved in ethylenediamine (0.25 ml) and heated with stirring for 5h at 70°C. Solvent is evaporated off and the residue is purified by reverse phase (sep-pak C 18) chromatography to give compound XVIII-C (20mg).

Synthesis of compound XVIII-D: Compound XVIII-C (15 mg) is treated with compound C (lOmg) (Example 2) exactly in the same way as described for compound D (Example 3) and the crude reaction mixture is purified and converted to

Na-salt form exactly in the same way as described for compound D (Example 3) to give compound XVIII-D (lOmg).

All of the U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications and non-patent publications referred to in this specification and/or listed in the Application Data Sheet, are incorporated herein by reference, in their entirety. Aspects of the embodiments can be modified, if necessary to employ concepts of the various patents, applications and publications to provide yet further embodiments. From the foregoing it will be appreciated that, although specific embodiments of the invention have been described herein for purposes of illustration, various modifications may be made without deviating from the spirit and scope of the invention.