CN1993342A - Intermediates for the preparation of halichondrin b - Google Patents

Abstract

The present invention provides macrocyclic compounds of formula (F-4), methods for their synthesis, and intermediates thereof. Such compounds and compositions thereof are useful for treating or preventing proliferative diseases.

Description

Intermediates for the preparation of halichondrin B

Cross References to Related Applications

This application claims priority to U.S. Provisional Patent Applications 60/576,642, filed June 3, 2004, 60/626,769, filed November 10, 2004, and 60/663,300, filed March 18, 2005, by The entire contents are incorporated herein by reference.

Technical field

The present invention relates to compounds used as intermediates in the synthesis of pharmaceutically active macrolide compounds.

Background of the Invention

The present invention relates to pharmaceutically active macrocyclic lactone, its synthesis and the intermediate used in its synthesis. Halichondrin B, a potent anticancer agent, was originally isolated from the marine sponge Halichondria okadai and subsequently found in Axinella sp. Found in the sponge genus (Lissonderndryx sp.). The total synthesis of halichondrin B was published in 1992 (Aicher, TD et al., J. Am. Chem. Soc. 114:3162-3164). Halichondrin B has been shown to inhibit tubulin polymerization, microtubule assembly, β ^s -tubulin cross-linking, GTP and vinblastine binding to tubulin and tubulin-dependent GTP hydrolysis in vitro. and have shown anticancer properties in vivo. Therefore, there is a need for the development of synthetic methods for the preparation of halichondrin B analogs useful as anticancer agents.

Invention Overview

As described herein, the present invention provides methods for preparing halichondrin B analogs having pharmaceutically useful activities such as anticancer activity or antimitotic (blocking mitosis) activity. These compounds include compounds of formula B-1939:

These compounds are useful in the treatment of cancer and other proliferative diseases including, but not limited to, melanoma, fibrosarcoma, leukemia, colon cancer, ovarian cancer, breast cancer, osteosarcoma, prostate cancer, lung cancer, and ras-transformed fibroblasts. The present invention also provides synthetic intermediates for preparing the halichondrin B analogs.

A detailed description of certain embodiments of the present invention

The methods and intermediates of the present invention are useful in the preparation of various halichondrin B analogs as described in, for example, US Patent 6,365,759 and US Patent 6,469,182 (incorporated herein by reference). As shown in Scheme I below, halichondrin B analogs were generally prepared by the assembly of three fragments F-1, F-2 and F-3.

Option I

1. Fragment F-1

According to one embodiment, the present invention provides compound F-1:

Wherein: PG ¹ and PG ² are each independently hydrogen or a suitable hydroxyl protecting group;

R ¹ is R or OR;

R ² is CHO or -CH=CH ₂ ; and

Each R is independently hydrogen, C _1-4 haloaliphatic, benzyl, or C _1-4 aliphatic, with the proviso that when R ¹ is OMe, PG ¹ and PG ² do not form an acetonide group.

In certain embodiments, R ¹ is OR. In other embodiments, R ¹ is OR, wherein R is hydrogen, methyl or benzyl.

In certain embodiments, PG ¹ and PG ² are hydrogen. In other embodiments, one of PG ¹ and PG ² is hydrogen.

Suitable hydroxyl protecting groups are well known in the art and include "Protecting Groups in Organic Synthesis" (TW Greene and PGM Wuts, 3rd Edition, John Wiley & Sons, 1999, which is incorporated herein in its entirety as those groups detailed in reference). In certain embodiments, PG ¹ and PG ² are each independently selected from esters, ethers, silyl ethers, alkyl ethers, aryl alkyl ethers, and alkoxyalkyl ethers, together with the oxygen atoms bound thereto. Examples of such esters include formates, acetates, carbonates and sulfonates. Specific examples include formate, benzoylformate, chloroacetate, trifluoroacetate, methoxyacetate, triphenylmethoxyacetate, p-chlorophenoxyacetate ester, 3-phenylpropionate, 4-oxopentanoate, 4,4-(ethylenedithio)valerate, pivalate (trimethylacetyl), crotonate, 4-Methoxycrotonate, benzoate, p-benzylbenzoate, 2,4,6-trimethylbenzoate; or carbonates such as methyl, 9-fluorenylmethyl , ethyl, 2,2,2-trichloroethyl, 2-(trimethylsilyl)ethyl, 2-(benzenesulfonyl)ethyl, vinyl, allyl and p-nitrobenzyl Carbonate. Examples of such silyl ethers include trimethylsilyl ether, triethylsilyl ether, tert-butyldimethylsilyl ether, tert-butyldiphenylsilyl ether, triisopropyl Alkyl silyl ethers and other trialkyl silyl ethers. Alkyl ethers include methyl ether, benzyl ether, p-methoxybenzyl ether, 3,4-dimethoxybenzyl ether, trityl ether, t-butyl ether, allyl ether, and allyl ether Oxycarbonyl ethers or derivatives. Alkoxyalkyl ethers include acetals such as methoxymethyl ether, methylthiomethyl ether, (2-methoxyethoxy)methyl ether, benzyloxymethyl ether, β-(tri Methylsilyl)ethoxymethyl ether and tetrahydropyranyl ether. Examples of aryl alkyl ethers include benzyl ether, p-methoxybenzyl ether (MPM), 3,4-dimethoxybenzyl ether, O-nitrobenzyl ether, p-nitrobenzyl ether, p-halogenated benzyl ether, 2,6-dichlorobenzyl ether, p-cyanobenzyl ether, 2- and 4-picolyl ether.

In certain embodiments, one or both of the PG ¹ and PG ² moieties of F-1 are silyl ethers or aryl alkyl ethers. In other embodiments, one or both of the PG ¹ and PG ² moieties of F-1 is t-butyldimethylsilyl or benzoyl. In another embodiment, the PG ¹ and PG ² moieties of F-1 are both tert-butyldimethylsilyl.

According to another embodiment, PG ¹ and PG ² together with the oxygen atoms bound to them form a diol protecting group such as a cyclic acetal or ketal, such groups include methylene, ethylene, benzylidene , isopropylidene, cyclohexylene and cyclopentylene; silylene derivatives such as di-tert-butylsilylene and 1,1,3,3-tetraisopropyldisiloxane Derivatives; cyclic carbonates and cyclic boronic esters. Methods for the addition and removal of such hydroxyl protecting groups and others are well known in the art and can be obtained from examples such as "Protecting Groups (protecting Groups)" (PJ Kocienski, Thieme, 1994) and "Protecting Groups in Organic Synthesis". (protective Groups in organicsynthesis)" (TW Greene and PGM Wuts, 3rd ed., John Wiley & Sons, 1999). According to another embodiment, PG ¹ and PG ² together form an acetonide group.

According to one embodiment, R ² is CHO.

According to another embodiment, R ² is -CH═CH ₂ .

In certain embodiments, the present invention provides compounds of formula F-1, having the stereochemistry shown in compound F-1':

Each of these variables is as defined above and described in categories and subcategories above and here.

In certain embodiments, the following compounds F-1a and F-1b are provided:

Here, "TBS" means tert-butyldimethylsilyl.

The detailed synthesis of F-1a and F-1b is illustrated in the Examples below.

2. Fragment F-2

According to another embodiment, the present invention provides compound F-2:

独立为单键或双键，条件是两个 基不同时为双键；Of which: each

independently as a single or double bond, provided that two The base is not a double bond at the same time;

LG ¹ is a suitable leaving group;

X is halogen or -OSO ₂ (R ^y );

R ^y is a C _1-6 aliphatic group or a saturated, partially unsaturated or fully unsaturated 5-7 membered ring, wherein R ^y is optionally replaced by up to 3 members selected from halogen, R, NO ₂ , CN, OR, SR or N(R) _group substitution;

each R is independently hydrogen, C _1-6 haloaliphatic, or C _1-4 aliphatic; and

PG ³ is a suitable hydroxyl protecting group.

A suitable leaving group as used herein is a chemical group that is readily substituted by the chemical group to be introduced. Suitable leaving groups are well known in the art, see for example "Advanced Organic Chemistry" (Jerry March, 4th edition, pp. 351-357, John Wiley and Sons, NY (1992)). Such Leaving groups include, but are not limited to, halogen, alkoxy, sulfonyloxy, optionally substituted alkylsulfonyloxy, optionally substituted alkenylsulfonyloxy, optionally substituted arylsulfonyloxy and diazo groups. Examples of suitable leaving groups include chlorine, iodine, bromine, fluorine, methanesulfonyl, tosyl (mesyl), triflate (triflate), nitrobenzenesulfonyl (nosyl ) and bromobenzenesulfonyl (brosyl). In certain embodiments, the LG ¹ moiety of F-2 is sulfonyloxy, optionally substituted alkylsulfonyloxy, optionally substituted alkenylsulfonyl Acyloxy or optionally substituted arylsulfonyloxy. In another embodiment, the ^LG1 moiety of F-2 is an optionally substituted alkylsulfonyloxy group. In other embodiments, F-2's The LG ¹ moiety is methylsulfonyl or tosyl.

In certain embodiments, the X moiety of F-2 is halogen. In other embodiments, the moiety X of F-2 is sulfonyloxy, optionally substituted alkylsulfonyl, optionally substituted alkenylsulfonyl, or optionally substituted arylsulfonyl. In other embodiments, the moiety X of F-2 is triflate.

In certain embodiments, the ^PG3 moiety of F-2, together with the oxygen atom bound thereto, is a silyl ether. In other embodiments, the ^PG3 moiety of F-2, together with the oxygen atom bound thereto, is an ester group. According to one aspect of the invention, the ^PG3 moiety of F-2 together with the oxygen atom bound thereto is tert-butyldimethylsilyl. According to another aspect of the invention, the ^PG3 moiety of F-2 together with the oxygen atom bound thereto is pivaloyl or benzoyl.

In certain embodiments, the present invention provides compounds of formula F-2, having the stereochemistry shown in compound F-2':

Each of these variables is as defined above and described in categories and subcategories above and here.

In certain embodiments, compound F-2a or F-2b is provided:

Wherein "MsO" refers to methanesulfonate, "TfO" refers to trifluorosulfonate, "Opv" refers to pivaloate, "OBz" refers to benzoate and "TsO" refers to toluenesulfonate.

In another embodiment, the present invention provides a compound of formula F-2b, wherein said compound is crystalline. According to another embodiment, there is provided a compound of formula F-2b, wherein said compound is crystallized from an alkane solvent. In certain embodiments, there is provided crystalline F-2b, wherein said compound is crystallized from pentane or heptane. In other embodiments, there is provided crystalline F-2b, wherein said compound crystallizes at about 0°C.

Compounds of formula F-2 are generally prepared from intermediates F-2d and F-2e as shown in Scheme A below.

Option A

Therefore, another aspect of the present invention provides compounds of formula F-2d:

Wherein: R' is -CH=CH ₂ or -C(O)H;

AlK is a linear or branched C _1-4 aliphatic group; and

PG ⁵ is a suitable hydroxyl protecting group.

Suitable hydroxy protecting groups ^PG5 are as described and defined above for the ^PG3 moiety of compound F-2. In certain embodiments, PG ⁵ together with the oxygen atom bound thereto is a silyl ether. In another embodiment, ^PG5 is tert-butyldimethylsilyl.

According to one embodiment, the AlK moiety of compound F-2d is methyl.

In certain embodiments, compounds of formula F-2d' are provided:

Another aspect of the invention provides compounds of formula F-2e:

Where: R" is OH, OPG ³ or LG ⁴ ;

LG ⁴ is a suitable leaving group; and

Each PG ³ is independently a suitable hydroxyl protecting group with the proviso that when PG ³ is tert-butyldiphenylsilyl, R" is not OMs.

One of ordinary skill in the art will recognize that the R" moiety of compound F-2e can be transformed from OH to a protected hydroxyl group, OPG ³ or directly to LG ^4. Such transformations are known to those skilled in the art and include, inter alia, those described herein Those. In certain embodiments R" is OH or ^LG4 . The leaving group LG ⁴ of formula F-2e is as described and defined above for the LG ¹ moiety of compound F-2. In certain embodiments, LG ⁴ is tosyl or methylsulfonyl.

The PG ³ moiety of compound F-2e is as defined and described above for the PG ³ moiety of compound F-2. In certain embodiments, PG ³ together with the oxygen atom bound thereto constitutes a silyl ether. In another embodiment, ^PG3 is t-butyldiphenylsilyl.

Yet another aspect of the present invention provides compounds of formula F-2f:

Wherein AlK, PG ³ and PG ⁵ are the same as the general definition and the classification and subclass descriptions above and here. Compounds of formula F-2 are prepared from compounds of formula F-2f by methods described herein and those known in the art.

The detailed synthesis of F-2a is illustrated in the Examples below.

Alternatively, compounds of formula F-2 are prepared from D-quinic acid as shown in Scheme II below. The detailed preparation of compounds of formula F-2 is illustrated in the Examples below.

Scheme II

An alternative method for the preparation of compounds of formula F-2 from D-quinic acid provides an alternative route from intermediate 12 to intermediate 17 (shown in Scheme III below).

Scheme III

Scheme III above shows an alternative method for preparing intermediate 17 from intermediate 12 via Eschenmoser-Tanabe fragmentation, wherein each Rx is independently ^OPGx or CN, where ^PGx is a suitable hydroxy protecting group as described herein. Intermediate 17 is then used to prepare compounds of formula F-2 according to Scheme II above.

Yet another method for the preparation of compounds of formula F-2 from D-quinic acid provides yet another route from intermediate 9 to intermediate 17 (shown in Scheme IV below).

Plan IV

wherein ^PGy is a suitable carboxy protecting group as described herein, and each ^PGx is independently a suitable hydroxy protecting group as described herein.

Scheme V represents yet another preparation of intermediates useful in the preparation of compounds of formula F-2 from D-quinic acid.

Plan V

In Scheme V above, Intermediate 7 (from Scheme II), wherein R is the methyl ester, was used to prepare crystalline intermediate ER-817664. As described in Scheme V above, ^PG5 and ^PG6 are each independently suitable hydroxyl protecting groups. In certain embodiments, PG ⁵ and PG ⁶ together form a cyclic diol protecting group. In other embodiments, PG ⁵ and PG ⁶ together form a cyclohexylene protecting group. As described in Scheme V above, LG ⁵ is a suitable leaving group. Such suitable leaving groups are well known in the art and include those described herein. In certain embodiments, LG ⁵ is methylsulfonyl or tosyl.

According to another embodiment, the present invention provides compounds of formula A:

in: Indicates a single or double bond;

n is 1, 2 or 3;

PG ⁵ and PG ⁶ are each independently a suitable hydroxyl protecting group;

W is CH-A or C(O);

A is oxo or C _1-6 aliphatic, wherein A is optionally substituted by one or more ^Q groups;

Each Q ¹ is independently selected from cyano, halogen, azido, oxo, OR, SR, SO ₂ R, OSO ₂ R, N(R) ₂ , NR(CO)R, NR(CO)(CO) R, NR(CO)N(R) ₂ , NR(CO)OR, (CO)OR, O(CO)R, (CO)N(R) ₂ , O(CO)N(R) _{2 ,} or OPG ¹ , where PG ¹ is a suitable hydroxyl protecting group, wherein:

^Two Q on A are optionally joined together to form a 3-8 membered saturated, partially unsaturated or aromatic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen or sulfur; and

Each R is independently selected from hydrogen or optionally substituted from C _1-6 aliphatic group; 5-10 membered saturated carbocycle, partially unsaturated carbocycle or aromatic carbocycle; or has 0-4 independently selected from 4-10 membered saturated, partially unsaturated or cyclic or aromatic ring groups of nitrogen, oxygen or sulfur heteroatoms, wherein:

Two R groups on the same nitrogen atom are optionally combined with the nitrogen atom to form a 3-8 membered saturated ring with 1-4 heteroatoms independently selected from nitrogen, oxygen or sulfur, partially inert Saturated or aromatic rings.

In certain embodiments, the present invention provides a compound of formula A, having the stereochemistry shown in formula A':

Each of these variables is as defined above and described in categories and subcategories above and here.

In certain embodiments, the present invention provides compounds of formula A', wherein A is C(O) and said compounds have the formula A'-1:

Each of these variables is as defined above and described in categories and subcategories above and here.

As generally defined above, the A group of formulas A and A' is a C _1-6 aliphatic group, wherein A is optionally substituted by Q ¹ . In certain embodiments, the group A of formulas A and A' is a C _2-5 aliphatic group, wherein A is substituted with one or more Q ¹ groups.

As generally defined above, each ^Q1 group of formulas A and A' is independently selected from cyano, halo, azido, oxo, OR, SR, _SO2R , _OSO2R , N(R) _2. NR(CO)R, NR(CO)(CO)R, NR(CO)N(R) _2. NR(CO)OR, (CO)OR, O(CO)R, (CO)N(R) ) ₂ , O(CO)N(R) ₂ or OPG ¹ , wherein PG ¹ is a suitable hydroxyl protecting group. In certain embodiments, each Q ¹ group of formulas A and A' is independently selected from cyano, halo, azido, oxo, N(R) ₂ , OR, SR, SO ₂ R or OSO ₂ R. In another embodiment, each Q ¹ group of formulas A and A' is independently selected from cyano, halogen, azido, oxo, OR, SR, SO ₂ R, OSO ₂ R, N(R) ₂ , NR(CO)OR, NR(CO)R, and O(CO)N(R) ₂ . In other embodiments, exemplary Q ¹ groups include NH(CO)(CO)-(heterocyclyl or heteroaryl), OSO ₂ -(aryl or substituted aryl), O(CO)NH-(aryl or substituted aryl), aminoalkyl, hydroxyalkyl, NH(CO)(CO)-(aryl or substituted aryl), NH(CO)-(alkyl)(heteroaryl or heterocyclyl) , O(substituted or unsubstituted alkyl)(substituted or unsubstituted aryl), and NH(CO)(alkyl)(aryl or substituted aryl).

In some embodiments, the A group of formulas A and A' has one of the following characteristics:

(1) A has at least one substituent selected from hydroxyl, amino, azido, halogen and oxo;

(2) A is a C _1-6 alkyl group with at least one substituent selected from hydroxyl, amino and azido;

(3) A has at least two substituents independently selected from hydroxyl, amino and azido;

(4) A has at least two substituents independently selected from hydroxyl and amino;

(5) A has at least one hydroxy substituent and at least one amino substituent;

(6) A has at least two hydroxyl substituents;

(7) A is a substituted _C2-4 aliphatic group;

(8) A is a substituted C _aliphatic group;

(9) A has an (S)-hydroxyl or (R)-hydroxyl group at the alpha position relative to the carbon atom attaching A to the G-containing ring; and

(10) A is a C _1-6 saturated aliphatic group having at least one substituent selected from hydroxyl and cyano.

The term "(S)-hydroxyl" refers to the configuration of the carbon atom having the hydroxyl group as (S). Embodiments of the invention also include compounds wherein A is at least substituted on each carbon atom at the (1) alpha and gamma, (2) beta and gamma, or (3) alpha and beta positions relative to the carbon atom to which A is attached once. Each alpha, beta and gamma carbon atom independently has the (R) or (S) configuration. In certain embodiments, the present invention provides such compounds, wherein A is substituted at least once on each of the carbon atoms alpha and beta relative to the carbon atom to which A is attached.

Exemplary A groups of formulas A and A' include 2,3-dihydroxypropyl, 2-hydroxyethyl, 3-hydroxy-4-perfluorobutyl, 2,4,5-trihydroxypentyl, 3- Amino-2-hydroxypropyl, 1,2-dihydroxyethyl, 2,3-dihydroxy-4-perfluorobutyl, 3-cyano-2-hydroxypropyl, 2-amino-1-hydroxyethyl Base, 3-azido-2-hydroxypropyl, 3,3-difluoro-2,4-dihydroxybutyl, 2,4-dihydroxybutyl, 2-hydroxy-2-p-fluorophenyl- Ethyl, -CH ₂ (CO) (substituted or unsubstituted aryl), -CH ₂ (CO) (alkyl or substituted alkyl, such as haloalkyl or hydroxyalkyl) and 3,3-difluoro- 2-Hydroxypent-4-enyl.

In certain embodiments, the A group of one of A and A' is 3-amino-2-hydroxypropyl.

According to one aspect, the present invention provides a compound of one of formulas A and A', wherein Q ¹ is OPG ¹ , wherein PG ¹ is a suitable hydroxyl protecting group. Suitable hydroxyl protecting groups are well known in the art and include those detailed in "Protecting Groups in Organic Synthesis" (TW Greene and PGM Wuts, 3rd Ed., John Wiley & Sons, 1999, which is incorporated herein by reference in its entirety). those groups. In certain embodiments, the PG ^moiety of one of formulas A and A', together with the oxygen atom bound thereto, is selected from esters, ethers, silyl ethers, alkyl ethers, aryl alkyl ethers, and alkoxyalkanes base ether. Examples of such esters include formates, acetates, carbonates and sulfonates. Specific examples include formate, benzoylformate, chloroacetate, trifluoroacetate, methoxyacetate, triphenylmethoxyacetate, p-chlorophenoxyacetate ester, 3-phenylpropionate, 4-oxopentanoate, 4,4-(ethylenedithio)valerate, pivalate (trimethylacetyl), crotonate, 4-Methoxycrotonate, benzoate, p-benzylbenzoate, 2,4,6-trimethylbenzoate, or carbonates such as methyl, 9-fluorenylmethyl , ethyl, 2,2,2-trichloroethyl, 2-(trimethylsilyl)ethyl, 2-(benzenesulfonyl)ethyl, vinyl, allyl and p-nitrobenzyl Carbonate. Examples of such silyl ethers include trimethylsilyl ether, triethylsilyl ether, tert-butyldimethylsilyl ether, tert-butyldiphenylsilyl ether, triisopropyl Alkyl silyl ethers and other trialkyl silyl ethers. Alkyl ethers include methyl ether, benzyl ether, p-methoxybenzyl ether, 3,4-dimethoxybenzyl ether, trityl ether, t-butyl ether, allyl ether, and allyl ether Oxycarbonyl ethers or derivatives. Alkoxyalkyl ethers include acetals such as methoxymethyl ether, methylthiomethyl ether, (2-methoxyethoxy)methyl ether, benzyloxymethyl ether, β-(tri Methylsilyl)ethoxymethyl ether and tetrahydropyranyl ether. Examples of aryl alkyl ethers include benzyl ether, p-methoxybenzyl ether (MPM), 3,4-dimethoxybenzyl ether, O-nitrobenzyl ether, p-nitrobenzyl ether, p-halogenated benzyl ether, 2,6-dichlorobenzyl ether, p-cyanobenzyl ether, 2- and 4-picolyl ether.

In certain embodiments, the ^PG1 moiety of one of formulas A and A', together with the oxygen atom bound thereto, is a silyl ether or an arylalkyl ether. In other embodiments, the ^PG1 moiety of one of formulas A and A' is t-butyldimethylsilyl or benzoyl. In another embodiment, the ^PG1 moiety of one of Formulas A and A' is tert-butyldimethylsilyl ("TBS").

As generally defined above, two Q ^'s on A are optionally joined together to form a 3-8 membered saturated ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, partially not Saturated or aromatic rings. In certain embodiments, two Q ^'s on A together form an epoxidized ring.

In certain embodiments, the ^PG5 and ^PG6 groups of formulas A and A' are independently selected from those suitable protecting groups described above for the ^PG1 groups of formulas A and A'. In other embodiments, the ^PG5 and ^PG6 groups of formulas A and A' are combined to form a cyclic diol protecting group. Such diol protecting groups are well known in the art and include those described by Greene , and include cyclohexylene and benzylidene diol protecting groups.

In certain embodiments, the present invention provides a method of preparing a compound of formula F-2 according to the following schemes V-a, V-b and V-c.

Scheme V-a

In other embodiments, the present invention provides crystalline ER-817664.

Scheme V-b

Scheme V-c

Using ER-817664 as a crystallization intermediate, Scheme VI-a shows a general procedure for using this compound to prepare intermediates useful in the preparation of compounds of formula F-2.

Scheme VI-a

In another method for the preparation of intermediates useful in the preparation of compounds of formula F-2, as shown in Scheme VI-b below, the triol intermediate described in Scheme VI-a above is used.

Scheme VI-b

In Scheme VI-b shown above, the triol intermediate is treated with periodate to generate the aldehyde. This compound is homologated with methyl-Wittig reagent and the resulting olefin is reduced to an ester compound. Treatment of the remaining free hydroxyl with N-iodosuccinimide yields the iodo intermediate and reduction of the ester with sodium borohydride yields the above hydroxyl compound. One of ordinary skill in the art will recognize that the resulting iodo compound corresponds to compound 21 described above in Scheme II, wherein compound 21 has a protecting group at the hydroxyl position. Finally, the above-mentioned lactone is obtained through zinc treatment. One of ordinary skill in the art will recognize that the resulting lactone compound corresponds to compound 22 described above in Scheme II, wherein compound 22 has a protecting group at the hydroxyl position.

In another preparation of intermediates for the preparation of compounds of formula F-2 from D-quinic acid as shown in Scheme VII below, a further route starting from intermediate 2 described in Scheme II is provided route.

Scheme VII

In Scheme VII above, intermediate 2 (from Scheme II) was used to prepare a stereoselective form of ER-812829. Preferably, other protecting groups can be used to protect the diol ER-812829. Such groups are known to those of ordinary skill in the art and include cyclohexylene and benzylidene diol protecting groups. First, in step (a), the hydroxyl group of ER-811510 is treated with 2-bromochloroacetic acid to generate ER-812771. The brominated intermediate was treated with triphenylphosphine to generate the Wittig reagent in situ in a manner substantially similar to that described by Murphy et al. (Tettrahedron Letters, 40, (1991) 3455-3456). The Wittig reagent then forms the lactone ER-812772. In step (b), the double bond is subjected to stereoselective hydrogenation to obtain ER-812829.

As shown in Scheme VII-a below, the present invention also provides a process for the preparation of intermediates for the preparation of compounds of formula F-2 from D-quinic acid, starting from the intermediates shown in Scheme VII above ER-812829.

Scheme VII-a

3. Fragment F-3

According to another embodiment, the present invention provides compound F-3:

Wherein, each PG ⁴ is independently selected from a suitable hydroxyl protecting group;

R ³ is CHO or C(O)OR ⁴ ;

^R is a suitable carboxy protecting group; and

LG ² is a suitable leaving group.

Suitable carboxylate protecting groups are well known in the art and are described in detail in "Protecting Groups in Organic Synthesis" (TW Greene and PGM Wuts, 3rd edition, John Wiley & Sons, 1999). In certain embodiments, the R ^group of F-3 is an optionally substituted C _1-6 aliphatic group or an optionally substituted aryl group. Examples of suitable ^R groups include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, benzyl and phenyl, each of which is optionally substituted.

As noted above, suitable leaving groups are well known in the art, see, for example, "Advanced Organic Chemistry" (Jerry March, 4th Edition, pp. 351-357, John Wiley and Sons, NY (1992 )). Such leaving groups include, but are not limited to, halogen, alkoxy, sulfonyloxy, optionally substituted alkylsulfonyloxy, optionally substituted alkenylsulfonyloxy, optionally substituted Arylsulfonyloxy, silyl and diazonium moieties. Examples of suitable leaving groups include chlorine, iodine, bromine, fluorine, mesyl, tosyl, trifluoromethane Triflate, nosyl, and brosyl. In certain embodiments, the LG ² moiety of F-3 is iodine.

According to another embodiment, a suitable leaving group can be generated in situ in the reaction medium. For example, LG ² in a compound of formula F-3 can be generated in situ from a precursor of the compound of formula F-3, wherein the precursor contains a group readily substituted in situ by LG ² . In a specific example of such substitution, the precursor of the compound of formula F-3 contains a group (eg trimethylsilyl) substituted in situ with LG ² such as iodine. The source of iodine can be such as N-iodosuccinimide and the like. Such in situ generation of suitable leaving groups is well known in the art, see for example Id.

As noted above, suitable hydroxyl protecting groups are well known in the art, including in "Protecting Groups in Organic Synthesis" (TW Greene and PGM Wuts, 3rd ed., John Wiley & Sons, 1999, which is incorporated herein by reference in its entirety) those groups detailed in . In certain embodiments, each PG ⁴ together with the oxygen atom bound thereto is independently selected from esters, ethers, silyl ethers, alkyl ethers, arylalkyl ethers and alkoxyalkyl ethers. Examples of such esters include formates, acetates, carbonates and sulfonates. Specific examples include formate, benzoylformate, chloroacetate, trifluoroacetate, methoxyacetate, triphenylmethoxyacetate, p-chlorophenoxyacetate , 3-phenylpropionate, 4-oxopentanoate, 4,4-(ethylenedithio)pentanoate, pivalate (trimethylacetyl), crotonate, 4 - Methoxycrotonate, benzoate, p-benzylbenzoate, 2,4,6-trimethylbenzoate, or carbonates such as methyl, 9-fluorenylmethyl, Ethyl, 2,2,2-trichloroethyl, 2-(trimethylsilyl)ethyl, 2-(phenylsulfonyl)ethyl, vinyl, allyl and p-nitrobenzyl carbonate ester. Such silyl ethers include trimethylsilyl ether, triethylsilyl ether, tert-butyldimethylsilyl ether, tert-butyldiphenylsilyl ether, triisopropylmethylsilyl ether, Silyl ethers and other trialkylsilyl ethers. Alkyl ethers include methyl ether, benzyl ether, p-methoxybenzyl ether, 3,4-dimethoxybenzyl ether, trityl ether, t-butyl ether, allyl ether, and allyl ether Oxycarbonyl ethers or derivatives. Alkoxyalkyl ethers include acetals such as methoxymethyl ether, methylthiomethyl ether, (2-methoxyethoxy)methyl ether, benzyloxymethyl ether, β-(tri Methylsilyl)ethoxymethyl ether and tetrahydropyranyl ether. Examples of aryl alkyl ethers include benzyl ether, p-methoxybenzyl ether (MPM), 3,4-dimethoxybenzyl ether, O-nitrobenzyl ether, p-nitrobenzyl ether, p-halogenated benzyl ether, 2,6-dichlorobenzyl ether, p-cyanobenzyl ether, 2- and 4-picolyl ether.

In certain embodiments, one, two or three of the ^PG4 moieties of F-3, together with the oxygen atom to which they are bound, are silyl ethers or arylalkyl ethers. In other embodiments, one, two or three of the PG ⁴ moieties of F-3 are t-butyldimethylsilyl or benzyl. In other embodiments, all three PG ⁴ moieties in F-3 are tert-butyldimethylsilyl.

According to another embodiment, a compound of formula F-3 is provided, wherein said compound has the stereochemistry shown in formula F-3':

Each of these variables is as defined above and described in categories and subcategories above and here.

In certain embodiments, compound F-3a is provided:

Where "TBS" refers to tert-butyldimethylsilyl.

The detailed synthesis of F-3a is illustrated in the Examples below.

4. Assemble F-1, F-2 and F-3 to prepare compound I

In general, fragments F-1 and F-2 were coupled as shown in Scheme VIII below. Scheme VIII

Scheme VIII above shows a general procedure for the preparation of intermediate F-5a from fragments F-1 and F-2. First, fragments F-1 and F-2 were coupled to obtain intermediate F-4 by a method substantially similar to that described by Kishi et al. (Org Lett, 4:25, p 4431 (2002)). The coupling is carried out in the presence of chiral oxazole (ER-807363) or in the absence of ER-807363. However, the coupling reaction of F-1 and F-2 in the presence of ER-807363 was more selective. Treatment of F-4 with potassium hexamethyldisilazide forms the intramolecular williams onether of F-4, which then provides tetrahydropyran F-5 as a mixture of stereoisomers. The stereoisomers are then separated to afford F-5a. The details of these three steps are described in the Examples below.

According to another embodiment, the present invention provides compound F-4:

wherein PG ¹ , PG ² , PG ³ , LG ¹ and R ¹ are as defined in general and as subclasses above and here.

In certain embodiments, the present invention provides a compound of formula F-4, wherein said compound has the stereochemistry shown in formula F-4':

wherein PG ¹ , PG ² , PG ³ , LG ¹ and R ¹ are as defined in general and as subclasses above and here.

The present invention also provides compound F-4a:

Where "MsO" refers to methanesulfonate, "TBS" refers to tert-butyldimethylsilyl, and "OPv" refers to pivalate.

The detailed synthesis of F-4a is described in the Examples below.

According to yet another embodiment, the present invention provides compound F-5.

wherein each of PG ¹ , PG ² , PG ³ , LG ¹ and R ¹ is as defined generally and as defined above and here for subclasses.

In certain embodiments, the present invention provides compounds of formula F-5 having the stereochemistry of formula F-5' or F-5a.

wherein each of PG ¹ , PG ² , PG ³ , LG ¹ and R ¹ is as defined generally and as defined above and here for subclasses.

As shown in Scheme IX below, the ^PG3 group of intermediate F-5a is removed, and the resulting hydroxyl compound F-6 is then coupled with compound F-3a (wherein ^R3 is CHO) to generate F-7.

Plan IX

Scheme IX above shows a general procedure for the preparation of intermediate F-9 from F-3a and F-6. The sulfone intermediate F-6 is treated first with n-butyllithium and then with aldehyde F-3a. The resulting diol intermediate F-7 is then oxidized with Dess-Martin reagent to give the keto-aldehyde intermediate F-8, which is then treated with _SmI to give intermediate F-9. Details of these steps are described in the Examples below.

Program X

Scheme X above describes a general method for the preparation of halichondrin B analogs of the invention from F-9a (LG ² is iodine). First, under conditions substantially similar to those described in Scheme V above, an intramolecular coupling is performed to generate the hydroxyl compound F-10. In another approach, the intramolecular coupling is performed in the presence of the chiral oxazole ligands described herein. The chiral oxazole ligand is added to increase the reaction yield and increase the reaction efficiency. The details of this reaction are described in the Examples below. Compound F-10 is then oxidized to F-11. The hydroxy protecting group of F-11 is removed by an appropriate method to obtain F-12. One of ordinary skill in the art will recognize that the appropriate method for deprotecting compound F-11 depends on the actual protecting group used, including those described by Greene . For example, when each hydroxy protecting group of F-11 is a TBS group, this removal can be achieved by optionally buffered tetrabutylammonium fluoride treatment. Details of these steps are described in the Examples below.

Intermediate F-12 was used in the preparation of various halichondrin B analogs such as those described in US Patent 6,365,759 and US Patent 6,469,182, which are hereby incorporated by reference in their entirety.

Example

Taking the preparation of halichondrin B analogue B-1939 as an example, the following examples describe the synthesis of halichondrin B analogues using the methods and compounds of the present invention.

Those of ordinary skill in the art will recognize that the various halichondrin B analogs prepared from the compounds of the invention by the methods of the invention include, but are not limited to, those described in U.S. Pat. Halichondrin B analogue. Accordingly, it should be understood that the synthetic procedures described below in the form of examples do not limit the scope of the invention as defined by the appended claims.

Example 1

Preparation of F-1a:

In an appropriately sized vessel, D-glucurono-6,3-lactone (1 wt, 1 eq) was mixed with acetonitrile (3 vol) and acetone (9 vol). Concentrated sulfuric acid catalyst was added, and the system was kept under reflux for 3 hours. Check the dissolution of D-glucurono-6,3-lactone in the system. The reaction solution was cooled to 25°C and stirred for 15 hours. Solid sodium bicarbonate (0.5 wt) was added and the reaction was stirred for an additional 3 hours. The solids were removed by filtration and the organics were partially concentrated and azeotroped with additional acetonitrile (2 wt). ER-806045 was used in the next reaction without isolation.

Crude ER-806045 (1 wt, 1 eq) was dissolved in acetonitrile (6.5 vol) at -20°C. Pyridine (1.5 vol, 4.0 eq) was added and _SO2Cl2 (0.38 vol, 1.02 eq) was added slowly keeping the internal temperature below ₅ °C. The reaction was quenched by adding back into cold water (28 vol) while flushing with acetonitrile (0.5 vol) while keeping the internal temperature below 10°C. ER-806410 (0.87 wt, 79% theory) was isolated as a white solid by filtration through heptane (2 vol) and drying.

Charge ER-806410 (1 wt, 1 eq) and THF (10 vol) in an appropriately sized vessel and cool to 10°C. Wet palladium on carbon (5%, 0.5 wt) was added and the heterogeneous solution was stirred for 10 minutes. The reaction solution was buffered with pyridine (0.44 wt, 1.3 eq) and placed under a hydrogen atmosphere for 3 hours. The reaction was filtered and the solids were rinsed with water (2 vol) and ethyl acetate (10 vol). The resulting solution was acidified with 1N hydrochloric acid (2.1 vol), mixed well, and the resulting layers were separated. The organic layer was washed sequentially with aqueous sodium bicarbonate (5 vol) and water (5 vol). The organics were concentrated under reduced pressure and the resulting product was recrystallized from IPA (3.4 vol) and further harvested by addition of heptane (3.4 vol) at 15°C to isolate ER-806047 as a white solid (67% yield).

Charge ER-806047 (1 wt, 1 eq) and toluene (8 vol) in an appropriately sized vessel and cool to -40°C. A 17 wt% solution of DIBAL in toluene (4.6 wt, 1.1 eq) was added maintaining the internal temperature below -35°C. After the reaction solution was measured, acetone (0.15 wt, 0.5 eq) was added to quench the excess reagent while keeping the temperature below 10°C. Below 10°C, the reaction was diluted with ethyl acetate (7 vol) and 15% aqueous citric acid (8 wt) and stirred at 20°C until a clear solution was obtained. The layers were separated and the aqueous layer was back extracted twice with ethyl acetate (2 x 10 vol). The combined organics were washed successively with aqueous sodium bicarbonate (5 v) and brine (5 v) and dried over magnesium sulfate (0.2 wt). After filtration, the organic layer was partially concentrated under reduced pressure and formed an azeotrope with toluene (4 vol). The product was stored as a THF solution for the next reaction.

A solution of _TMSCH2MgCl in ether (20 wt%, 2.04 wts, 3.0 equiv) was added to an appropriately sized vessel and cooled below 5°C. A solution of ER-806048 (1 wt, 1 eq) in THF (7 vol) was added to the reaction vessel keeping the internal temperature below 15°C. The temperature of the reaction solution was raised to 35° C. and maintained for 1.5 hours. The reaction was cooled, diluted below 20°C with toluene (7 vol), and quenched with acetic acid (3 vol). The reaction solution was further diluted with 10% aqueous ammonium chloride solution (6 wt), mixed well, and the layers were separated. The organic layer was washed sequentially with aqueous sodium bicarbonate (5 vol) and brine (5 vol). Drying over magnesium sulfate (0.2 wt), filtration, and concentration of the solution under reduced pressure isolated ER-807114 as a concentrated toluene solution (90% yield).

ER-807114 (1 wt, 1 eq) and THF (20 vol) were added sequentially in an appropriately sized vessel and the solution was cooled to below 5°C. A 15 wt% solution of KHMDS in toluene (9.16 wt, 2.0 equiv) was added. The reaction was quenched with 10% aqueous ammonium chloride (5 vol). The layers were separated and the organic layer was washed sequentially with ammonium chloride (5 vol), 2N hydrochloric acid (8.5 vol), aqueous sodium bicarbonate (5 vol), and water (5 vol). The organics were transferred to a concentrator with ethyl acetate and concentrated to a viscous oil (90% yield). The material was recrystallized from toluene (4 vol) and heptane (4 vol) at 35°C, and at lower temperatures (15°C and 10°C) from heptane (2 x 4 vol ) to further harvest the product (yield 94%).

KOtBu (0.67 wt, 1.2 eq) and THF (7.7 vol) were added to an appropriately sized vessel and cooled to an internal temperature of -20°C. A solution of ER-806049 (1 wt, 1 eq) in THF (2.3 vol) was added keeping the internal temperature below -7°C. Pure BnBr was added while the maximum temperature was maintained at -7°C. The reaction solution was stirred at -20°C for 2 hours and at 10°C for 10 hours. The reaction was quenched with 10% aqueous ammonium chloride (4 vol), diluted with toluene (4 vol), and mixed well. The layers were separated and the organic layer was washed with 10% brine (4 v) and dried over magnesium sulfate (0.15 wt). After concentration under reduced pressure a solution of ER-806050 in tBuOH (2.5 vol) was isolated (95% yield).

In an appropriately sized vessel, add _K3Fe (CN)6 (3.5 wt, 3.4 eq), _K2CO3 (1.5 wt, 3.4 eq), ₍ DHQ) _2AQN (0.0134 wt, 0.005 eq) , water (18 vol), tBuOH (13 vol), and ER-806050 in tBuOH (1 wt, 1 eq in 5 vol). The heterogeneous mixture was cooled to an internal temperature of 0°C, and K ₂ OsO ₄ ·2H ₂ O (0.0029 wt, 0.22 mol %) was added. After ₃₆ hours, the reaction was quenched _{with Na2S2O3} (3.5 eq, 1.7 wt) at 0 °C and the flask was allowed to warm to room temperature overnight. After 15 hours, the mixture was transferred to a workup vessel and diluted with toluene (15 vol) and water (4 vol). Stir vigorously and separate the biphasic mixture. The organic layer was washed with brine (10 vol), concentrated and solvent exchanged to give a crude mixture of diols ER-806051 and ER-806052 in 10% toluene (92% yield).

A solution of ER-806051/52 (1 wt, 1 eq) in toluene (10.1 wt%, 9.9 wts) was further diluted with additional toluene (3 wts). N-methylmorpholine (0.94 wt, 3.0 eq) and DMAP (0.075 wt, 0.2 eq) were added to the toluene solution, and the resulting mixture was cooled to below 15°C. While keeping the internal temperature below 25°C, add benzoyl chloride. The reaction was then stirred at 75°C for 12 hours. The reaction was cooled to 15°C and quenched with 1N hydrochloric acid (5 vol) keeping the temperature below 25°C. Mix well and separate layers. The organic layer was washed sequentially with brine (3 wts), aqueous sodium bicarbonate (3 wts) and brine (3 wts). The organic layer was dried (magnesium sulfate, 0.25 wt), treated with activated carbon (0.1 wt), and filtered (Celite(R), 0.3 wt) through toluene (1 wt). The product was partially concentrated under reduced pressure forming an azeotrope with toluene (3 wt). A toluene solution of the bisbenzoate ER-806053/54 (5 vol) was isolated (95% yield).

Under an inert atmosphere, a solution of 20 wt% _TiCl4 (6.42 wt, 3.6 eq) in toluene was cooled to 15°C. While keeping the internal temperature below 30°C, Ti(OiPr) ₄ (0.64 wt, 1.2 eq) was added, and the resulting solution was stirred for 15 minutes. Allyl TMS (1.03 wt, 4.8 eq) was premixed with 22 wt% toluene solution (4.55 wt, 1 eq) of ER-806053/54 (1 wt, 1 eq) obtained in the previous step, and into freshly generated Ti(OiPr)Cl ₃ . Keep the internal temperature below 30 °C during the sample addition. The reaction solution was stirred for 2 hours at 20-30°C. The reaction was cooled to -5°C and quenched with 1N hydrochloric acid (6 vol) while keeping the internal temperature below 30°C. After thorough stirring, the layers were separated and the organic layer was washed sequentially with 1N hydrochloric acid (3 vol) and brine (2 x 3 vol). The organic layer was stirred with magnesium sulfate (0.3 wt) and activated carbon (0.15 wt), rinsed with toluene (1 vol), and filtered through a plug of Celite ^(R) (0.2 wt). The product was isolated after concentration as a 3:1 mixture of C-34 in 83% yield. Recrystallization from IPA/n-heptane afforded ER-806055 (C-34>99.5%de, yield 71%)

Add alcohol ER-806055 (1 wt, 1 eq), toluene (7 vol), DMSO (0.31 wt, 2.0 eq) and triethylamine (0.78 wt, 4.0 eq) in an appropriately sized container at room temperature ). The resulting solution was cooled to -19°C. TCAA (0.84 wt, 1.4 equiv) was added dropwise keeping the internal temperature below -10°C. The reaction was stirred for an additional 10 minutes. The reaction was diluted with IPA (0.5 vol) and quenched with 1N HCl (5 vol) keeping the internal temperature below 10°C. The layers were separated and the organic layer was washed sequentially with aqueous sodium bicarbonate (5 wt) and water (3 v). The organic layer was partially concentrated under reduced pressure (crude yield 100%) and further azeotroped with additional toluene (4 vol.). The resulting ketone (ER-806058) was dissolved in the last 4 volumes of toluene, and the water content was checked for the next reaction.

A solution of ER-107446 (1 wt, 1.5 eq) in THF (2.7 vol) was cooled to 10°C, keeping the internal temperature below 15°C, and 25.2% by weight of LHMDS in THF (5.2 )deal with. In a second vessel, a solution of crude ER-806058 in toluene (21.9 wt%, 5.4 vol) was cooled to 10°C. While maintaining the internal temperature below 20°C, transfer the contents of the first container to the solution containing the substrate. The reaction was stirred for 30 minutes and then quenched by the addition of 1M hydrochloric acid (6.5 vol) while keeping the internal temperature below 20°C. The layers were separated and the organic layer was washed four times with 1:1 methanol/water (4 x 5 vol) followed by aqueous sodium bicarbonate (5 vol) and brine solution (2 x 5 vol). The product was dried over magnesium sulfate (0.52 wt), filtered (rinsing with 0.7 vol of toluene), and concentrated under reduced pressure to a heavy oil.

ER-806059 was dissolved in 1:1 toluene/acetonitrile (5 vol) at room temperature. While maintaining the initial temperature below 40°C, filtered TMSI (1.23 wt, 4 eq) was added. The reaction solution was heated to 60 °C for 2 hours. The reaction solution was cooled to -15°C, and quenched with 25% aqueous ammonium hydroxide solution below 30°C. The reaction contents were stirred overnight and the layers were separated. Additional toluene (5 vol) and water (2 vol) were added to the organic layer. Mix well and separate layers. The organic layer was then washed successively with 10% aqueous sodium sulfite (5 vol), 1N hydrochloric acid (5 vol), 5% aqueous sodium bicarbonate (5 vol), and brine (5 vol). The organic layer was dried over magnesium sulfate (0.2 wt), filtered, and partially concentrated to obtain a 50% solution in toluene, which was used in the next reaction.

In an appropriately sized vessel, mix NaBH(OAc) ₃ (1.19 wt, 3.15 eq), _Bu4NCl (1.04 wt, 2.1 eq), DME (8.2 vol), and toluene (4 vol) at 65 °C , and stirred at room temperature. The mixture was heated to 75°C for 1 hour. A 50 wt% solution of ER-806060 (1 wt, 1 eq) in toluene was added at 75°C and rinsed with additional toluene (0.3 vol). The reaction temperature was raised to 85°C, and the reaction solution was stirred for 2-4 hours. The reaction was cooled to <10°C and quenched with water (3.2 vol) while keeping the internal temperature below 20°C. Mix well and separate layers. The organic layer was washed sequentially with aqueous sodium bicarbonate (2 x 5 vol) and water (2 x 5 vol). The organic layer was concentrated and solvent exchanged to obtain a 40% by weight solution of ER-806061 in methanol.

A solution of ER-806061 (1 wt, 1.0 eq) in methanol (40 wt%) was dissolved in additional methanol (1.6 vol). Potassium carbonate (0.24 wt, 1.0 eq) was added and the reaction temperature was raised to 50°C for 1 hour. The reaction was cooled to 15°C and quenched with 1N hydrochloric acid (3.5 vol, 2 eq) at an internal temperature below 30°C. The reaction was diluted with water (3.9 vol) and toluene (3 vol). The layers were separated and the aqueous layer was back extracted with toluene (1.5 vol). NaHCO3 (0.3 wt) and NaCl (0.6 wt) were added to the aqueous phase and back extracted with nBuOH (3 v). The three organic phases were combined and concentrated to dryness to give crude triol ER-806064 and inorganic salts. The product was dissolved in 7:1 toluene/nBuOH at 80°C, filtered hot, recrystallized by cooling and stirred overnight. ER-806064(F-1b) was isolated after filtration and methanol rinse (57% overall yield over five steps). FAB(+)-MS M/z 357(M+H). The melting point is 96.2°C.

At 25 °C, the purified triol ER-806064 (1 wt, 1 eq) was dispersed in acetone (2 v), diluted with 2,2-dimethoxypropane (1 v), washed with concentrated sulfuric acid (0.0086 wt, 0.03 eq) was treated. Stir the reaction until homogeneous. The reaction was diluted with toluene (5 vol) and quenched into 5% potassium carbonate (2 vol). Mix well and separate layers. The organic layer was washed with 10% brine, dried over sodium sulfate (0.5 wt). The solution was filtered (toluene rinse) and concentrated under reduced pressure to afford ER-806126 as a yellow oil. This material was used in the next step.

Solid NaOtBu (0.34 wt, 1.4 eq) was dissolved in THF (2.7 vol) and DMF (0.3 vol) and cooled to below 10°C. A THF (2.5 vol) solution of ER-806126 (1 wt, 1 eq) was added to the NaOtBu solution under a THF (0.5 vol) rinse while keeping the internal temperature below 15°C. After stirring for 30 min, methyl iodide (0.204 vol, 1.3 eq) was added keeping the temperature below 15°C (exotherm). The reaction was allowed to warm to 25°C, quenched with water (5 vol) and diluted with toluene (7 vol). Mix well and separate layers. The organic layer was washed twice with brine (2 x 5 vol), dried over sodium sulfate (0.5 wt), filtered and concentrated under reduced pressure.

ER-806068 (1 wt, 1 eq) was dissolved in 1 vol methanol. Water (1.5 vol) and 2N hydrochloric acid (1.25 vol, 1 eq) were added and the reaction was stirred at 25°C. The reaction was quenched back into 2M NaOH (1.34 vol) at 10 °C. The reaction was diluted with isopropyl acetate (5 vol). Mix well and separate layers. The aqueous layer was back extracted with 5 volumes of isopropyl acetate and the combined organic layers were dried over magnesium sulfate (0.5 wt), filtered and concentrated under reduced pressure to afford crude diol ER-806063.

To a solution of crude ER-806063 (1 wt, 1.0 eq) in DMF (4 v) at 25°C was added imidazole (0.62 wt, 3.4 eq) followed by addition of TBSCl ( 1.02 parts by weight, 2.53 equivalents). The reaction solution was stirred at 25°C. The reaction was diluted with MTBE (10 vol) and washed with water (4 vol). The organic layer was washed sequentially with 1M hydrochloric acid (3 vol), water (3 vol), aqueous sodium bicarbonate (3 vol), and brine (3 vol). The organic layer was dried over magnesium sulfate (0.5 wt), filtered rinsed with 1 vol of MTBE, concentrated under reduced pressure, and solvent exchanged with heptane (4 vol).

ER-806065 (1 wt, 1 eq) was dissolved in heptane, isooctane or IPA (10 vol). Cool the solution to below -60°C (±10°C). Ozone was bubbled through the solution at low temperature until the solution remained blue. The solution was purged with nitrogen for 15-30 minutes, and the temperature of the reaction solution was raised to 5° C. while continuing to purse with nitrogen. 7-15% by weight of Lindlar's catalyst (5% palladium/calcium carbonate poisoned with lead, 0.1 parts by weight) was added. The reactor head was purged several times with nitrogen, evacuated, and placed under a hydrogen atmosphere (g). The reaction was then allowed to warm to room temperature (20-25°C). The reaction was stirred for 2.5 hours. The resulting heterogeneous solution was filtered through Celite( ^R) (1.0 wt, with MTBE (2 vol) as rinse). The solution was concentrated to dryness and 1.0 parts by weight of crude ER-806067 was isolated. The crude isolate was recrystallized from heptane or isooctane to obtain ER-806067 (F-1a) as a white crystalline solid (68% yield over five steps). FAB(+)-MS M/z 601(M+H). The melting point is 64.5°C.

Example 2

Preparation of F-2a

The reactor was charged with pre-rinsed Amberlyst 15 (0.05 wt) and water (4.63 vol) and cooled to an internal temperature of 0-5°C. While keeping the internal temperature at about 5°C, 2,3-dihydrofuran (1 wt, 1 eq) was added into the reactor and stirred for 1.5 hours. Water (4.63 vol) was added to the second reactor and heated to an internal temperature of 35°C. In the same reactor was charged tin powder (2.2 wt, 1.3 eq) and diluted with 2,3-dibromopropene (3.71 wt, 1.3 eq) and 48% hydrobromic acid (0.002 vol), respectively. After observing the onset of the reaction (indicated by the temperature trace reaching 36-38° C.), 2,3-dibromopropene (9×0.37 parts by weight). After the addition was complete, the contents of the second reactor were stirred for an additional 60 minutes at an internal temperature of 35°C. The filtered contents of the first reactor were added to the second reactor at such a rate that the internal temperature did not exceed 45°C. After the addition was complete, the heat source was removed, Celite ^(R) 545 (2.0 wt.) was added to the second reactor, and the resulting mixture was stirred for 30 minutes. The heterogeneous mixture was filtered through a pad of Celite ^(R) 545 (2.0 wt) and the filter cake was washed with additional water (5 vol). All filtrates were combined into one reactor and concentrated hydrochloric acid (1.5 vol) was added until the cloudy solution became clear. Under vigorous stirring, sodium chloride (3.6 wt) was added to the reactor, and the layers were separated. Separate and set aside the organic layer. The aqueous layer was extracted with n-butanol (20 vol). The aqueous layer was drained and the reactor was charged with the first isolated organics. The organics were washed with concentrated sodium bicarbonate (24 vol) and the aqueous layer was back extracted with n-butanol (20 vol). All organics were combined and concentrated in vacuo. The concentrate was dissolved in MTBE (10 vol), filtered and the filtrate concentrated to 2 vol. With stirring, the concentrate was allowed to cool to an internal temperature of 0°C, then n-heptane (4 vol.) was added. The heterogeneous mixture was stirred for 2 hours at an internal temperature of 0°C, and the target product was isolated by filtration and dried in vacuo to obtain ER-806909 (1.34 wt, 0.45 eq) as a white powder.

The reactor was charged with imidazole (0.65 wt, 2 eq), ER-806909 (1 wt, 1 eq) and anhydrous DMF (4.04 vol). With stirring, the reactor was cooled to an internal temperature of 0°C, then tert-butyldiphenylchlorosilane (1.25 wt, 0.95 eq) was added at such a rate that the internal temperature did not exceed 15°C. Keeping the internal temperature at <15°C, the reaction solution was stirred for another 1 hour. Water (3.2 vol) and n-heptane (6.4 vol) were added to the reactor. The mixture was stirred for 5-15 minutes and the layers were separated. Separate and set aside the organic layer. The aqueous layer was extracted with n-heptane (3.2 vol). All organics were combined, washed with brine (3.2 vol), and concentrated in vacuo to constant weight to afford ER-806545 (2.13 wt, 0.95 eq) as a yellow oil. This product was used in the next step without further purification.

Enantiomers ER-806909 were separated by simulated moving bed (SMB) chromatography to give ER-808373 (0.55 wt, 0.55 eq) and ER-806721 (0.45 wt, 0.45 eq) as yellow oils. The SMB chromatography protocol used to separate enantiomers ER-806909 is as follows:

Column and medium: Chiracel OD column 20μm 30mm×

                                                                                               

Solvent system: 96:4 (v/v) heptane: tert-butanol (mobile phase)

Simulated moving bed chromatograph: Knauer SMB system CSEP C912

Isotherm (Langmuir, determined by head-on analysis)

Non-target isomer: Qi ^* = 2.8768×Ci/(1+0.02327*Ci)

Target isomer: Qi＝4.5320×Ci/(1+0.0034595*Ci)

^* Where Qi = solid phase concentration (g/L), Ci = liquid phase concentration (g/L)

Column porosity: 0.658

Temperature: 27℃

Use EuroChrom 2000 for windows, SMB guide version 1.2 (Wissenschaftliche Geratebau Dr.-Ing. Herbert Knauer GmbH, D-14163 Berlin; authors H. Kniep and A. Seidel-Morgenstern) to simulate the flow rate, etc.:

Injection (feed) concentration: 36g/L (ER-806909)

Injection liquid flow rate (No. 1 pump) 15mL/min

Eluent flow rate (No. 2 pump) 76.4mL/min

Zone IV (pump No. 3) flow rate 107mL/min

Zone II (Pump No. 4) flow rate 134.3mL/min (actual flow rate =

                            143.5mL/min)

Flow rate in Zone I 183.4mL/min

Zone III flow rate 149.3mL/min

Raffinate ^* Flow Rate 42.3mL/min ^* Raffinate = weakly bound isomer

body

Extract flow rate 49.1mL/min ^* Extract = strongly bound isomer

body

Intermittent time (tact time)

(Port switching time) 0.8864 minutes (53.18 seconds, actual intermittent time

= 54 seconds)

Using the above protocol, enantiomer ER-806909 was separated in the following manner. For an 11 hour run, pump 10 L of mobile phase (36 g/L of ER-806909) (Silogmodel Chemtech) through a 142 mm diameter, 0.45 μm pore nylon filter (Cole-Parmer # 2916-48) into the injection tank (feed tank). The eluent tank was filled with 36 L of mobile phase, which had been filtered through a 1 μm glass fiber filter (Whatman GFC) with a diameter of 45 mm in the pipeline, and an additional volume was added during operation. The internal temperature of the SMB instrument was adjusted to 27°C.

During initial startup, both the sample inlet and the eluent inlet are connected to the eluent tank. Prime and rinse the injection and eluent pumps with mobile phase solvent. Start the column switching of the SMB instrument, turn on the pumps, and gradually increase the flow rate to full speed, while maintaining the absolute flow rate difference between the pumps. Once at full speed, measure the raffinate flow rate and extract flow rate and adjust the pump flow rate to correct for deviations in pump specifications. The injection pump (No. 1 pump) was decelerated to 0mL/min, the inlet was connected to the injection tank again, the injection solution was injected into the pump, and then the flow rate was gradually returned to the full speed. The outlets of raffinate and extract were connected to the separation tank, and each sample was obtained every 2 hours. The chiral purity of the samples was monitored by analytical HPLC using the HPLC method described below. Adjust the flow rate and intermittent time of No. 2, No. 3 and No. 4 pumps to obtain the required outlet purity.

At the end of the run, reduce the flow rate of the injection pump to 0 again and connect it to the eluent tank. Return the injection pump to full speed and wash the system for 20 minutes. During the washing period, the raffinate and extract outlets were kept open for 10 minutes (10 intervals). After washing, the outlets were connected to the separation tank. The column wash was gradually concentrated and added to the sample in subsequent runs.

The extracts (ER-806721) collected at the end of each run were combined with material collected from the same starting batch and the final batch pool was reanalyzed for chiral purity using the analytical HPLC method described in Table 1 below. The same operation was performed on the collected raffinate (ER-808373).

Table 1 HPLC analysis of chiral purity of ER-806721 Column: Flow rate: Temperature (°C): Injection volume: Instrument: Mobile phase composition ABCD Chiracel OD column 10 μm 250×4.6 mm, DAICEL Chemical Industries, Ltd. (DAICEL Chemical Industries, Ltd.) catalog number 140250.8 mL/min 27 (preferred)-35 usually 10 μL, sample dissolved in solvent A, 5 mg/mL Waters Alliance W2690, With UV W2487 (also with advanced laser polarimeter) (PDR-Chiral, Inc.) 99:1 heptane:2-propanol Gradient table: (%) gradient time (min) A B C D. 0 100 0 0 0 isocratic running time 30min

Detection: UV absorbance at 254nm

Triphenylphosphine (0.7 wt, 1.2 eq), p-nitrobenzoic acid (0.45 wt, 1.2 eq), ER-808373 (1 wt, 1 eq) and anhydrous toluene (8 vol ). The reaction solution was cooled to an internal temperature of 0°C, and DEAD (1.17 wt, 1.2 eq) was slowly added at such a rate that the internal temperature did not exceed 7°C. n-Heptane (3.3 vol) was added and the mixture was cooled to an internal temperature of 10°C, then stirred for 30-40 minutes. The resulting precipitate was removed by filtration. The filter cake was washed with n-heptane (3.3 vol), TBME (0.55 vol), n-heptane (1.1 vol) and MTBE (0.55 vol), respectively. All filtrates were combined and concentrated in vacuo. The crude concentrate was dissolved in THF (8 vol), then water (0.8 vol) and lithium hydroxide dihydrate (0.18 wt, 2 eq) were added. After the mixture was stirred at room temperature, n-heptane (3.3 vol.) was added and stirred for 5 minutes. Water (2.2 vol) and n-heptane (3.3 vol) were added and the biphasic mixture was stirred for 5 minutes to allow the layers to separate. The aqueous layer was separated and back-extracted with n-heptane if necessary. The organic layers were combined and concentrated in vacuo. The crude product was purified by SiO ₂ column chromatography to afford ER-806721 (0.74-0.85 wt, 0.74-0.85 eq) as pale yellow oil.

The reactor was charged with ER-806721 (1 wt, 1 eq) and anhydrous dichloromethane (4.2 vol). The reaction solution was cooled to an internal temperature of 0-5°C, then triethylamine (0.34 parts by weight, 1.5 equivalents), p-toluenesulfonyl chloride (0.51 parts by weight, 1.2 equivalents) and 4-(dimethylamino)pyridine (0.001 parts by weight, 0.25 equivalents). The resulting mixture was stirred at room temperature for 48 hours, then water (1.8 vol) and dichloromethane (1.8 vol) were added. After stirring well, the organics were separated and concentrated. The concentrate was dissolved in MTBE (1.8 vol) and washed with brine (1.8 vol). Separate and set aside the organic layer. The aqueous layer was back extracted with MTBE (1.8 vol), then all organics were combined and concentrated in vacuo. Eluting with MTBE (7 vol), the crude oil was filtered through a _plug of SiO2 (70-230 mesh, 1 wt), and the filtrate was concentrated in vacuo. The concentrate was dissolved in IPA (5 vol) and water (0.25 vol) was added. The resulting mixture was cooled to an internal temperature of 15°C, and then ER-807204 seed crystals were added. After seeding, the mixture was cooled to an internal temperature of 0° C. and stirred for 4-5 hours. The suspension was filtered, the filter cake was washed with cold IPA (1 v), and the filter cake was dried in vacuo to constant weight to afford ER-807204 (1.05 wt, 0.78 eq) as a white powder. IR (thin film, cm-1) λ2597, 1633, 1363, 1177, 907, 729. LRMS m/z 602 (M+H).

A 21% solution of sodium ethoxide in ethanol (2.97 wts, 0.9 eq) was added to the reactor. The solution was heated to an internal temperature of 65°C, then diethyl malonate (3.24 wt, 2 eq) was added at such a rate that the internal temperature did not exceed 70°C. The mixture was stirred for 30 minutes, then ER-806906 (1 wt, 1 eq) was added over 3-5 hours. After adding the sample, the reaction solution was stirred for 60 minutes, and then cooled to an internal temperature of 50°C. Concentrated hydrochloric acid (0.84 wt, 1.05 equiv) was added at such a rate that the internal temperature did not exceed 65°C. After removal of DMF (3 vol) and ethanol by distillation, a solution of magnesium chloride hexahydrate (0.21 wt, 0.1 eq) in distilled water (0.25 vol, 1.4 eq) was added. The resulting mixture was heated to an internal temperature of 135°C while removing distillate. The mixture was heated at reflux, then cooled to room temperature, and brine (12 vol) and TBME (16 vol) were added. The organic layer was separated, washed with water (1.3 vol) and brine (1.2 vol) and concentrated in vacuo. The product was purified by distillation to give ER-805552 (0.95-1.09 wt, 0.71 equiv).

A 1.0M solution of LHMDS in toluene (6.61 parts by weight, 1.04 equivalents) was added to the reactor, and cooled to an internal temperature of -75°C. ER-805552 (1 wt, 1 eq) dissolved in anhydrous THF was added to the reactor at such a rate that the internal temperature did not exceed -70°C. After the addition was complete, the resulting mixture was stirred for 30 minutes, anhydrous THF (2.5 vol) and methyl iodide (1.27 wt, 1.25 eq) were added to a second reactor and cooled to an internal temperature of -75°C. The THF solution of ER-805552 was added to the methyl iodide solution at such a rate that the internal temperature did not exceed -65°C. After adding the sample, the reaction solution was stirred for 30 minutes at an internal temperature of -78°C. The reaction was inverse quenched into 1 N hydrochloric acid solution (10 vol) and MTBE (8 vol) with vigorous stirring. After the addition was complete, the aqueous layer was separated and discarded. The organic layer was washed with brine solution (3 vol) and concentrated in vacuo. The product was purified by distillation to give ER-806724 (0.75 wt) as an approximately 6/1 diastereomeric mixture.

N,O-dimethylhydroxylamine hydrochloride (1.05 parts by weight, 1.5 equivalents) and anhydrous CH ₂ Cl ₂ (8.1 parts by volume) were added to the reactor, and cooled to an internal temperature of 0°C. A 2M solution of trimethylaluminum in toluene (3.93 wt, 1.5 eq) was added at such a rate that the internal temperature did not exceed 5°C. The reaction solution was stirred for another 10 minutes so that the internal temperature did not exceed 5°C and ER-806724 was added at a rate. The reaction solution was diluted with CH ₂ Cl ₂ (15 vol), and then back-quenched by adding to 1.3M sodium tartrate (20 vol) at 0°C at such a rate that the internal temperature did not exceed 10°C. After the addition was complete, the layers were allowed to separate and the aqueous layer was set aside. The organics were washed with water (1 v), dried over sodium sulfate (1 wt), filtered, and concentrated in vacuo to remove traces of dichloromethane. To the concentrate were added anhydrous DMF (6.3 vol), imidazole (0.64 wt, 1.5 eq) and tert-butyldimethylsilyl chloride (0.94 wt, 0.97 eq) respectively. Water (5 vol) and MTBE (10 vol) were added and the resulting mixture was stirred to allow the layers to separate. The aqueous layer was separated and discarded. The organic layer was washed with water (5 vol) and the layers were separated. 1N Sodium hydroxide (2.5 vol) and methanol (2.5 vol) were added and the resulting mixture was stirred. The aqueous layer was separated and the organic layer was washed with brine (2.5 vol) and concentrated in vacuo to afford ER-806753 (1.94 wt, 0.91 eq) as a brown oil.

The reactor was charged with ER-806753 (1 wt, 1 eq), _CH2Cl2 (5 vol) _, and a 50% aqueous solution of NMO (0.8 wt, 1.1 eq). After the mixture was cooled to an internal temperature of 10°C, a 0.197M solution of _OsO4 in toluene (0.06 vol, 0.004 eq) was added. Sodium sulfite (0.1 wt, 0.25 eq) and water (0.85 vol) were added and the reaction was stirred for 1 hour. The mixture was diluted with brine (0.85 vol), and the organics were concentrated in vacuo to about 1/3 volume. Sodium periodate (1.3 wt, 2 eq) was added to a second reactor followed by THF (2.5 vol). The mixture was diluted with pH=7 phosphate buffer (3.0 vol) and cooled to an internal temperature of 20°C. Concentrated diol was added at such a rate that the internal temperature did not exceed 30°C. After the addition was complete, the resulting mixture was stirred at room temperature. Water (1.25 vol), MTBE (7 vol) and brine solution (1.25 vol) were added and the layers were separated. The organics were washed twice with a mixture of brine solution (1 vol) and saturated sodium bicarbonate (1 vol). Finally, a mixture of organics with brine (1 vol) and 10% (w/v) sodium thiosulfate solution (1 vol) was stirred for 1 hour, then concentrated in vacuo. The crude material was purified by SiO _column chromatography to give ER-806754 (0.93 wt, 0.93 equiv) IR (thin film, cm-1) λ 2953, 2856, 1725, 1664, 1463, 1254, 1092, 833.LRMS m/ z 332(M+H).

The reactor was charged with ER-806629 (1.53 wt, 3.1 eq) and THF (10.5 vol), and the solution was degassed by sparging nitrogen for 60 minutes. In a second inert reactor, ER-807204 (1 wt, 1.0 eq), ER-806754 (0.66 wt, 1.2 eq) and THF (2.7 vol) were charged and the solution was degassed by sparging argon for 45 minutes. gas. The reactor containing ER-806906 was charged with _CrCl2 (0.63 wt, 3.1 eq) followed by triethylamine (0.52 wt, 3.1 eq). The dark green suspension was stirred at an internal temperature of 30-35°C for 1 hour, cooled to 0-5°C, and then _NiCl2 (0.1 eq.) was added. The contents of the second reactor were added slowly to the first reactor over 0.5 hours and the reaction was allowed to warm to room temperature. The reaction solution was cooled to an internal temperature of 0°C, then ethylenediamine (1.0 wt, 10 eq) was added over 30 minutes, and the reaction solution was stirred at an internal temperature of 25°C for at least 30 minutes. Water (4 vol), TBME (10 vol) and n-heptane (1 vol) were added to the reaction, the resulting mixture was stirred for 15 min and the phases were allowed to separate (approximately 30 min). The aqueous layer was separated and back extracted with TBME (ca. 7.5 vol). The organic layers were combined, washed with water (5 vol), brine (3 vol), and concentrated in vacuo to minimum volume. IPA (10 vol) and _SiO2 (1 wt) were added to the crude mixture and the resulting mixture was stirred at an internal temperature of 25°C for up to 4 days. The suspension was filtered and the filter cake was washed with IPA (2 x 1 vol). To the filtrate was added n-heptane (6.6 vol) and the mixture was concentrated in vacuo until a suspension formed. The mixture was filtered, the filter cake was washed with n-heptane, and the mixture was concentrated in vacuo. The crude product was purified by SiO _column chromatography to give ER-807524 (0.54 wt, 0.48 eq) IR (film, cm-1) λ 2934, 1668, 1471, 1108, 833.LRMS m/z as a clear yellow oil 704 (M+Na).

The reactor was charged with ER-807524 (1 wt, 1 eq) and anhydrous THF (1.25 vol). The mixture was cooled to an internal temperature of -20°C, and 3M methylmagnesium chloride (0.59 vol, 1.2 eq) was added at such a rate that the internal temperature did not exceed 0°C. After the addition was complete, the mixture was allowed to warm to an internal temperature of 0°C over 2 hours. The reaction mixture was backquenched by adding half saturated ammonium chloride (2.62 vol) and the resulting mixture was diluted with TBME (2 vol) with vigorous mixing. The aqueous layer was discarded and the organics were washed with brine (2 v) and concentrated in vacuo. The crude product was purified by SiO ₂ column chromatography to afford ER-807525 (0.79-0.82 wt, 0.85-0.88 eq) as a yellow oil.

ER-807525 (1 wt, 1 eq), N-phenylbistrifluoromethanesulfonamide (0.59 wt, 1.1 eq) and anhydrous THF (4.1 vol) were added to the reactor, and the mixture was cooled to The temperature is -75°C. A 0.5 M KHMDS solution in toluene (2.75 wt, 1 eq) was added at such a rate that the internal temperature did not exceed -60°C, and then the reaction solution was allowed to warm to -20°C over 2 hours. The reaction was quenched with half-saturated ammonium chloride (2.4 vol) at such a rate that the internal temperature did not exceed 0°C. The mixture was allowed to warm to an internal temperature of 20°C and n-heptane (2.4 vol.) was added. The mixture was stirred, and the aqueous layer was separated and discarded. The organic layer was washed three times with saturated sodium bicarbonate (2.3 vol each), then concentrated in vacuo to give ER-807526 (1.2-1.4 wt, 1.0-1.2 eq). This material was used in the next step without further purification.

The reactor was charged with ER-807526 (1 wt, 1 eq) and anhydrous methanol (3 vol) at 20°C. 1.25M hydrochloric acid in IPA (4 vol, 5 eq) was added and the mixture was stirred for 3 hours. Solid sodium bicarbonate (0.42 wt, 5 eq) was added portionwise with stirring until the pH of the reaction mixture reached 6-7. The reaction mixture was filtered with methanol (3 x 2 vol.) washes. All filtrates were concentrated in vacuo and then purified by _SiO2 column chromatography to give ER-807527 (0.43 wt, 0.79-0.85 eq).

The diastereomeric mixture ER-807527 was separated by preparative HPLC chromatography and the desired fraction was concentrated to give ER-806730 (0.56 wt, 0.56 equiv) as a bright yellow oil. The preparative HPLC chromatographic protocol used to isolate ER-806730 was as follows:

Column and medium: Kromasil spherical silica gel column, 60A, particle size 10

             μml, in a Varian of 7.7cm×60cm

Dynamax Rampak column filled to 7.7cm

(diameter)×30cm (length)

HPLC filling station: Varian(Rainin)Dynamax Rampak

                                                                   

HPLC pump: Varian (Rainin) SD-1 titanium pump head

Main (primary) HPLC detector: Waters R403 Refractive Index Detector

Deputy (secondary) HPLC detector: Varian (Rainin) UV-1 detector (with

Prepare flow cell)

Chromatographic control and acquisition software: Varian (Rainin) Dynamax DA version 1.4.6

Chromatographic data processing software: Varian (Rainin) Dynamax R4.3 version

Mobile phase: 28.5:63.7:7.85 (volume/volume) n-heptane:methane

        tert-butyl ether: 2-propanol

Flow rate: 140mL/min

Column temperature: room temperature (25°C)

Detection: Refractive Index, Cathode (attenuated at 16X), 215nm

UV absorption value at

Mobile Phase Gradient: Isocratic

Running time: 40 minutes

Injection volume: 10ml, containing 0.8g/mL of ER-807527

Using the above protocol, the diastereomer ER-807527 was separated in the following manner. Batches of ER-807527 were first diluted to 0.1 g/ml with mobile phase and filtered through a 47 mm glass fiber filter (Whatman GFC) with a pore size of 1 μm under vacuum. The filtrate was then concentrated in vacuo using a rotary evaporator. Start flow on SD-1 HPLC pump A (prime and wash with mobile phase) and gradually increase the flow rate to 140 mL/min. The system was washed until the UV and RI detectors reached baseline stabilization. Rinse the reference flow cell of the RI detector with fresh flow phase.

The equivalent batch of ER-807527 was diluted to a concentration of 0.8 g/mL with mobile phase to obtain a chromatogram of an 8 g ER-807527 injection. A 10 mL aliquot of the solute was injected corresponding to the ER-807527 peak (starting at approximately 24 min peak and continuing to 35 min) and the eluate was collected. Subsequent injections and fraction collections were continued until the starting material was exhausted.

Fractions corresponding to ER-806730 were combined and concentrated in vacuo using a rotary evaporator. The HPLC analysis described in Table 2 was used to evaluate the diastereomeric purity and area % purity area.

Table 2 HPLC analysis of diastereoisomeric purity of ER-806730 Column: Flow rate: Temperature (°C): Injection volume: Instrument: Mobile phase composition ABCD Kromasil silica gel column 250 x 4.6 mm, 5 μm, MetaChem catalog number 0475-250 x 046 1 mL/min 2710 μL, sample in solvent A, 2.5 mg/mL Waters Alliance W2690 with UV W2487 30:67:3 n-heptane:methyl tert-butyl Base ether: 2-propanol 2-propanol Gradient table: time (min) (%) gradient A B C D. 0222632 1001009090 001010 0000 0000 isocratic isocratic linear isocratic running time 32min, of which under the initial conditions, rebalance for 18min

Detection: UV absorption value at 205nm

ER-806730 (1 wt, 1 eq) and anhydrous dichloromethane (4.8 vol) were added to the reactor and cooled to an internal temperature of 0°C. 2,4,6-Collidine (1.16 wt, 4 eq) and DMAP (0.03 wt, 0.1 eq) were added, the resulting mixture was stirred for 15 minutes, then trimethylpyridine was added at such a rate that the internal temperature did not exceed 10 °C acetyl chloride (0.3 wt, 1.05 eq). Water (3 vol.) was added and the mixture was stirred for 15 minutes. TBME (10 v) was added and the mixture was stirred for an additional 10 minutes. The organic layer was washed with 1N hydrochloric acid (10 vol.) until 2,4,6-collidine was negatively charged, then water (5 vol.), saturated sodium bicarbonate (5 vol.) and saturated brine (5 vol. volume) wash. The organic layer was concentrated in vacuo, and the concentrate was purified by _SiO2 column chromatography to afford ER-806732 (1.02 wt, 0.85 eq) as a yellow oil.

The reactor was charged with ER-806732 (1 wt, 1 eq) and anhydrous THF (2.35 vol) and cooled to an internal temperature of 0°C. Triethylamine (0.22 wt, 1.1 eq) and methanesulfonyl chloride (0.24 wt, 1.05 eq) were added successively at such a rate that the internal temperature did not exceed 10°C. The reaction was stirred at 0°C internal temperature, then n-heptane (3.4 vol) was added with vigorous stirring and the layers were allowed to separate. The organics were washed with saturated brine (3.4 v), dried over saturated sodium sulfate (2 wt), filtered, and the filter cake was washed with n-heptane until ER-805973 (F-2a) was negatively charged. The filtrate was concentrated in vacuo to afford ER-805973 (1.12 wt, 0.97 equiv). Crude ER-805973 (F-2a) was used in the next step without further purification. IR (thin film, cm-1) λ 2961, 1725, 1413, 1208, 926. LRMS m/z 579 (M+H).

Example 3

Another preparation method of ER-806730

The reactor was charged with quinic acid (1 wt), cyclohexanone (2.11 eq, 1.08 wt) and concentrated sulfuric acid (0.011 eq, 0.0056 wt). The reaction mixture was heated to 160°C and water was removed by azeotropic distillation (azeotrope started at 100°C). The reaction solution was cooled to 90°C-100°C, and sodium bicarbonate (0.0096 parts by weight) and toluene (3.6 parts by weight) were added. The reaction was allowed to cool to room temperature over 4-6 hours, and the resulting precipitate was filtered, washed with toluene (2 x 0.9 wt) and dried to give 1 (0.97 wt) as a white powder.

Compound 1 (1 eq, 1 wt) and imidazole (2.5 eq, 0.80 wt) were mixed, purged with nitrogen, and suspended in anhydrous THF (10 v). TMSCl (1.2 equiv, 0.61 wt) was added at such a rate that the temperature was kept below 30°C. The reaction was cooled to room temperature, heptane (10 vol) was added and the resulting suspension was filtered. The filter cake was washed with 1:1 heptane/THF (10 v) and the filtrate solvent exchanged with toluene by atmospheric distillation to give a solution of 2a (1.34 wt, calculated) in toluene (ca. 5 wt).

Cool the 2a solution to -78°C and add DIBAL-H (1.5M in toluene, 1.2 equiv, 2.1 wts) at such a rate that the temperature remains below -65°C. Excess DIBAL-H was quenched with methanol (0.3 eq, 0.034 wt) and the solution was allowed to warm to 0°C. The solution was transferred to a solution of 30% by weight aqueous Rochelle Salt (10 wt) and sodium bicarbonate (1 wt) at such a rate that the temperature was maintained below 25°C. The mixture was stirred vigorously, resulting in a biphasic solution. The layers were separated and the aqueous layer was extracted with MTBE (5 v). The combined organic layers were washed successively with water (2.5 wt) and saturated brine (2.5 wt). The organics were concentrated and solvent exchanged with TFH to give 2b (0.98 wt, calc.) as a solution in TFH (5 v).

The 2b solution was cooled to 5°C and acetic acid (2.9 eq, 0.51 vol) was added. Water (1.0 equiv, 0.055 vol) was added and the solution was stirred at 0°C-5°C. Additional additions of up to two aliquots of acetic acid and one aliquot of water were added as needed to facilitate silyl deprotection. Triethylamine (12 eq, 3.6 wt) and DMAP (0.05 eq, 0.02 wt) were added at such a rate that the temperature was kept below 20°C. Acetic anhydride (6 eq, 2.0 wt) was added, and the reaction solution was stirred at room temperature. The reaction was cooled to 5°C and saturated aqueous sodium bicarbonate (10 vol) was added at such a rate that the temperature was kept below 30°C. The resulting mixture was stirred for 3-4 hours and the layers were separated. The aqueous layer was extracted with MTBE (5 v). The combined organics were washed with water (5 vol.). The extract was solvent exchanged with IPA by distillation to give 2c as a solution in IPA (3 v). The solution was cooled to 5°C, and the resulting crystals were filtered. The mother liquor was concentrated and a second crop was obtained after recrystallization to give 2c (0.87 wt) as a crystalline white solid.

2c (1 wt) was dissolved in acetonitrile (6 vol), and methyl 3-trimethylsilylpent-4-enoate (3.0 eq, 1.86 wt) was added followed by TFAA (0.2 eq, 0.083 portion volume). _BF3OEt2 ( _1.0 equiv, 0.42 vol) was then added to the solution at such a rate that the temperature was kept below 25 °C. Saturated aqueous sodium bicarbonate solution (10 vol.) was added to the reaction solution, and the resulting mixture was stirred for 15 minutes. The mixture was extracted with heptane (10 v) followed by MTBE (5 v) and the combined extracts were concentrated to give 3 (0.72 wt, calculated) as an orange oil.

A solution of 3 (1 wt) in THF (9 v) was treated with sodium methoxide (25% w/w in methanol (1.5 eq, 2.2 wt)) at such a rate that the temperature was kept below 25 °C. The reaction was quenched by the addition of 1N hydrochloric acid (10 vol.). The organic layer was separated and the aqueous layer was extracted with MTBE (10 vol). The combined organics were washed with water (2.5 vol), saturated sodium bicarbonate (2.5 vol) and water (2.5 vol). The solution was concentrated to give a solution of 4 (0.88 wt, calculated) in THF (2.5 vol). This solution was used directly in the next step.

To a solution of 4 (1 wt) in tetrahydrofuran (2.5 vol) was added methanol (5 vol). 1N Hydrochloric acid (0.75 eq, 2 v) was added and the reaction was allowed to warm to 60-80 °C. The reaction solution was cooled to room temperature, and saturated aqueous bicarbonate solution was added. The mixture was extracted with DCM (3 x 2.5 v) and the combined DCM extracts were solvent exchanged with ethyl acetate to give a solution of 5 in ethyl acetate (3 v). Heptane (2 vol.) was added to induce crystallization and the resulting suspension was cooled to 0°C. The solid was collected by filtration, the filter cake was washed with cold ethyl acetate/heptane (1:1 v/v), and dried to give 5 (0.55 wt) as a white powder.

Compound 5 (1 wt) was dissolved in acetonitrile (10 vol), followed by the addition of 2-acetoxy-2-methylpropanyl bromide (4.0 eq, 2.2 wt) and water (1 equiv, 0.067 wt). The resulting mixture was stirred at room temperature, then cooled to 5-10°C. NaOMe (25% w/w in methanol, 8 eq, 6.2 wt) was added and the reaction was allowed to warm to room temperature. The reaction was quenched by the addition of saturated sodium bicarbonate (10 vol) and extracted with MTBE (2 x 10 vol). Solvent exchange with methanol by atmospheric distillation afforded 6 (0.91 wt, calculated) as a solution in methanol (10 v).

A solution of 6 (1 wt) in methanol (10 v) was heated to 55°C. Sodium borohydride (5 eq, 0.68 wt) was added in 6 portions, the reaction was cooled to 5°C and quenched with 1N hydrochloric acid (10 v). Brine (5 vol) was added and the reaction was extracted with ethyl acetate (2 x 10 vol). The combined extracts were concentrated to give 7 as a brown residue.

Compound 7 (1 wt) was dissolved in _CH2Cl2 (10 v), then DMAP (0.1 eq, 0.054 wt), triethylamine ₍ 3.0 eq, 1.85 v) and TBDPSCl ( 1.2 eq, 1.38 vol). Sodium bicarbonate (10 vol) was added and the organic layer was separated. The aqueous layer was extracted again with _{CH2Cl2 (} 10 vol). The organic extracts were combined and concentrated to afford 8 (1.8 wt, calculated) as a colorless oil.

To a solution of 8 (1 wt) in THF (10 vol) was added LDA (1.5M in cyclohexane, 4 eq, 6 vol) at room temperature. The solution was allowed to warm to 50°C and quenched with 1N hydrochloric acid (5 vol) and extracted with MTBE (10 vol). Concentration of the extract gave 9 (0.9 wt) as an oil.

A solution of 9 (1 wt) in dichloromethane (5 vol) and methanol (5 vol) was cooled to -60°C and treated with _O3 keeping the temperature below -50°C. The reaction was purged with nitrogen, _NaBH4 (0.5 eq, 0.04 wt) was added, and the mixture was allowed to warm to 0 °C. Additional NaBH ₄ (1 eq, 0.08 wt) was added in portions, and the reaction solution was warmed to room temperature. After 3 hours, the mixture was quenched with 1N hydrochloric acid (10 vol), dichloromethane (5 vol) was added and the layers were allowed to separate. The aqueous layer was re-extracted with dichloromethane (10 v) and the combined organic extracts were concentrated to give 10 (0.97 wt) as a colorless oil.

Compound 10 was dissolved in THF (10 vol) and phosphate buffer (pH=7, 5 vol) was added. _NaIO4 (2 eq, 0.854 wt) was added and the reaction was allowed to warm to room temperature. Water (5 vol) and MTBE (10 vol) were added and the resulting mixture was stirred vigorously for 10 minutes. The organic layer was separated, washed with 10% w/v aqueous sodium thiosulfate (5 vol), water (5 vol), and brine (5 vol), then dried azeotropically with THF (about 200 ppm water) to give 11 (0.93 wt, calculated) in THF (10 v). This solution was used directly in the next step.

To a solution of 11 (1 wt) in THF (10 v) was added (methoxymethylene)triphenylphosphine (1 wt) and heated to 65°C. Heptane (40 vol) was added and the resulting mixture was stirred for 30 minutes. The resulting precipitate was filtered and the filtrate was concentrated to a total volume of 10 vol. _SiO2 (5 wt) was added and the suspension was filtered through a pad of _SiO2 eluting with MTBE (20-40 v). The solvent was exchanged with methanol to give a solution of 12 (0.95 wt, calculated) in methanol (10 v) which was used directly in the next step.

To a solution of 12 (1 wt) in methanol (10 v) was added 10% w/w palladium on carbon (0.23 eq, 0.37 wt) and treated with hydrogen. The suspension was filtered while rinsing the filter cake with THF (10 v). Solvent exchange with THF gave 13 (0.95 wt, calculated) in THF (10 v) which was used directly in the next step.

A solution of 13 (1 wt) in THF (10 v) was cooled to 0-5 °C and LAH (1M in THF, 0.78 eq, 1.5 v) was added at a rate to keep the temperature below 10 °C. Water (1.7 equiv, 0.06 vol) was then added at a rate to maintain the temperature below 10°C. Sodium hydroxide 10% wt/water solution (0.16 eq, 0.06 vol) and water (4.98 eq, 0.17 vol) were added sequentially at a rate to maintain the temperature below 10°C, and the resulting mixture was allowed to warm to room temperature with vigorous stirring. The suspension was filtered and the filter cake was rinsed with THF (5 v). The filtrate was partially concentrated to give 14 (0.9 wt, calculated) in THF (10 v) which was used directly in the next step.

A solution of 14 (1 wt) in THF (10 v) was cooled to 0°C, then imidazole and TrCl (1.5 eq, 0.59 wt) were added. Saturated aqueous sodium bicarbonate (5 vol) was added and the mixture was extracted with heptane (10 vol). The extract was washed with brine (10 v) and concentrated to give 15 (1.35 wt).

Compound 15 (1 wt) was dissolved in THF (10 v) and treated with TBAF (1M, 1.2 eq, 1.6 v). The reaction mixture was concentrated to 2 vol, then heptane (5 vol) and _SiO2 (5 vol) were added. The resulting suspension was filtered, eluting with heptane (5 vol) followed by THF (10 vol). The THF eluate was collected to give a solution of 16 (0.61 wt, calculated) in THF (10 v) which was used directly in the next step.

To a solution of 16 (1 wt) in THF (10 v) was added _PPh3 (5 eq, 2.3 wt), pyridine (10 eq, 1 v) and NIS (3.0 eq, 1.1 wt). Then 20% w/w aqueous citric acid (10 eq, 14 wt) was added and the resulting mixture was stirred for 10 minutes. The reaction was diluted with heptane (10 vol) and the aqueous layer was separated. The organic layer was washed with water (5 vol), 10% w/v aqueous sodium thiosulfate (5 vol), water (5 vol), and brine (5 vol). The solvent was exchanged with ethanol and concentrated to 5 volumes. Water (10 v) was added and the resulting precipitate was collected by filtration to afford 17 (0.65 wt) as a white solid.

Compound 17 (1 wt) and potassium cyanide (6 eq, 0.54 vol) were suspended in ethanol (5 vol) and water (10 vol), and the resulting suspension was heated to 80°C. The reaction was diluted with water (5 vol) and ethyl acetate (10 vol) and mixed for 10 minutes. The aqueous layer was removed and the organic layer was washed with water (5 vol) and brine (5 vol). Solvent exchange with ethanol gave a solution of 18 (0.75 wt) in ethanol (10 v) which was used directly in the next step.

To a solution of 18 (1 wt) in ethanol (10 v) was added zinc (37 eq, 3.9 wt) and the mixture was heated to 75-80°C. The reaction was partially concentrated to 2-3 vol, cooled to room temperature and partitioned between MTBE (10 vol) and water (5 vol). The aqueous layer was removed and the organic layer was washed with saturated sodium bicarbonate (5 vol), water (5 vol) and brine (5 vol). It was then dried by azeotropic distillation in THF to about 200 ppm water to give a solution of 19 (0.81 wt) in THF (10 vol). The resulting solution was used directly in the next step.

To a solution of 19 (1 wt) in THF (10 vol) was added LDA (1.0M in THF, 1.2 eq, 2.4 vol) at -78°C. The resulting mixture was stirred for 10 min, then the enolate solution was added to a solution of methyl iodide (1.5 eq, 0.19 v) in THF (5 v) at -78 °C. The reaction was back-quenched by adding saturated sodium bicarbonate (10 vol) and extracted with MTBE (15 vol). The extract was washed with brine (5 v), concentrated and purified by chromatography to give 20 (0.86 wt).

To a suspension of dimethylhydroxylamine hydrochloride (1 wt) in dichloromethane (2.5 vol) was added AlMe3 (2M in toluene, 1.5 eq, 1.5 vol) at 0°C. A solution of 20 (1 wt) in dichloromethane (5 vol) was added at a rate to maintain the reaction temperature below 5°C. The reaction mixture was then added to aqueous sodium tartrate (1.3M, 20 vol) while maintaining the temperature below 10°C. The layers were allowed to separate and the organic layer was dried over sodium sulfate (5 wt). The resulting suspension was filtered and the filtrate was concentrated. The residue was dissolved in DMF (2 v) followed by the addition of imidazole (0.19 wt) and TBSCl (0.29 wt). The reaction was diluted with water (5 vol) and MTBE (10 vol) and stirred for 10 minutes. The aqueous layer was removed and the organic layer was washed with water (5 vol). The extract was added to a solution of aqueous sodium hydroxide (1 N, 0.78 vol) and methanol (0.7 vol). After stirring the reaction, the aqueous layer was removed and the organic layer was washed with brine (2.5 vol) and concentrated to give 21 (1.2 wt).

To a solution of 2 l (1 wt) in anhydrous THF (1.11 wt, 1.25 vol) was added methylmagnesium chloride (3.0 M, 59 wt, 1.2 eq) at such a rate as to keep the reaction temperature below 0 °C. After stirring at 0°C, the reaction was backquenched by adding saturated ammonium chloride (2.5 vol) and water (2.3 vol). The resulting mixture was diluted with MTBE (10 v) and stirred vigorously. The aqueous layer was separated and the organic layer was washed with brine (2.5 vol) and concentrated to give 22 (0.84 wt).

Compound 22 (1 wt) was dissolved in THF (4 vol) and cooled to -78°C. KHMDS (1.5M in toluene, 1.01 equiv, 2.78 wts) was added keeping the temperature below -60°C. A solution of Tf ₂ NPh (0.62 wt, 1.1 eq) in THF (1.5 vol) was added and the reaction was allowed to warm to -20°C. Sat. ammonium chloride (2.5 vol), water (2.5 vol) and n-heptane (2.5 vol) were added and the mixture was allowed to warm to room temperature. Separate the layers and remove the aqueous layer. The organic extract was washed with saturated aqueous sodium bicarbonate (3 x 2.5 vol) and brine (2.5 vol), then concentrated in vacuo to give 23 (1.1 wt).

Compound 23 was dissolved in methanol (2.5 vol) and cooled to 15°C. Hydrochloric acid (5N in IPA, 1.30 equiv, 1.18 wt) was added and the resulting solution warmed to 25°C. The reaction solution was cooled to 0°C, and sodium bicarbonate (3 eq, 0.33 wt) was added. The reaction was stirred for 15 minutes, and the resulting precipitate was removed by filtration. The filter cake was washed with ACS grade methanol (1 vol), and the combined filtrates were concentrated. The crude concentrate was purified by chromatography to give ER-806730(24) (0.5 wt).

Example 4a

Using the general scheme set forth in Scheme V above, Example 4a provides an alternative method for the preparation of compounds of formula A (intermediate for the preparation of F2). This method uses ER-812935 as an intermediate (prepared according to Example 3 (compound 4)).

ER-812935 (1 wt) was dissolved in THF (10 vol) and cooled to 0°C. LAH (1.0 M in THF, 0.70 equiv, 2.0 vol) was added keeping the temperature below 5°C. With vigorous stirring, the excess reagent was quenched with water (0.078 vol) while maintaining the temperature below 5°C. While maintaining vigorous stirring, sodium hydroxide (15% w/w in water (0.078 vol)) was added followed by water (0.18 vol). After addition of Celite( ^R ) (2 wt) the suspension was filtered and the filter cake was rinsed with THF (5 v). ER-817633 (0.92 wt, calculated based on 100% conversion) was concentrated to 5 volumes and used directly in the next step.

A previously prepared solution of ER-816961 (1 wt in 5 v THF) was diluted with THF (5 v), cooled to 5°C, and triethylamine (3 eq, 0.94 wt) was added. MsCl (1.05 eq, 0.25 vol) was added at a rate to keep the temperature below 10 °C. Water (5 wt) was added to quench the reaction. Heptane (8 vol) was added and the mixture was separated. The aqueous phase was separated and extracted with MTBE (2 v). The combined organic extracts were washed with saturated sodium bicarbonate (5 vol) and water (1.9 vol). The organic layer was concentrated, and the solvent was exchanged with ethanol to prepare a solution of ER-818937 (1.23 wt, calculated based on 100% conversion) in ethanol (1 vol), which was used directly in the next step.

A previously prepared solution of ER-818937 (1 wt in 0.8 vol ethanol) was diluted with ethanol (190 proof strength, 9 vol). Potassium cyanide (3 equiv, 0.41 wt) was added and the suspension was heated to 70-80°C. The reaction was cooled to room temperature and water (10 vol) was added followed by MTBE (10 vol). The layers were separated and the aqueous layer was extracted with MTBE (5 v). The combined organics were washed with water (2 v) and saturated brine (4 wt). The extract was concentrated and used directly in the next step.

ER-818950 was dissolved in acetic acid (5 vol), hydrochloric acid (1.0 M, 1 eq, 3 vol) was added, and the reaction was stirred at room temperature. The reaction was cooled to 0°C and sodium hydroxide (50 wt%, 30 eq, 7 wts) was added at a rate to keep the temperature below 10°C. The solution was extracted with heptane (2 x 10 vol). The aqueous phase was saturated with sodium chloride and extracted with acetonitrile (2 x 10 vol). The combined acetonitrile extracts were concentrated, solvent exchanged with ethyl acetate by atmospheric distillation to give a solution of ER-817664 in ethyl acetate (3 vol.). The salt was filtered off from the hot solution and cooled to 0°C. Filtration of the suspension afforded ER-817664 as a crystalline white solid.

Example 4b

Example 4b provides an alternative method for preparing compounds of formula F-2 using the general scheme described in Schemes Vb and Vc above. This method uses ER-817664 (prepared according to Example 4a above) as an intermediate.

ER-817664 (1 wt) was dissolved in acetonitrile (10 vol), the suspension was cooled to 0°C, and 2-acetoxy-2-methylbromopropane (4.0 eq, 2.4 vol) was added followed by water ( 1.0 eq, 0.07 vol). The resulting mixture was stirred at 0°C for 2 hours. At 0°C, sodium bicarbonate (sat. aq., 8.0 equiv, 40 vol) was added slowly. The resulting mixture was stirred at room temperature for 30 min, then extracted with MTBE (2 x 20 v). The organic layer was washed with brine (5 vol) and concentrated to give the product as a colorless oil.

The bromine starter described immediately above (1 wt) was dissolved in toluene (10 vol). DBU (1.8 eq, 0.73 vol) was added and the mixture was heated at 80°C. The mixture was cooled to room temperature, diluted with MTBE (20 vol), washed with water (5 vol) and brine (5 vol). The organic layer was then concentrated to give the product as an off-white powder.

The starting olefinic compound (1 wt) (described immediately above) was dissolved in dichloromethane (5 vol) and methanol (5 vol) and cooled to -40°C to -45°C. Treat the solution with _O3 . Excess _O3 was removed by nitrogen purge and the solution was allowed to warm to -15°C. _NaBH4 (1.0 equiv, 0.18 wt) was added and the mixture was allowed to warm to 0 °C. Potassium carbonate (1.3 equiv) was added and the suspension was stirred at room temperature. The reaction was neutralized with 1N hydrochloric acid (about 4 equiv, about 20 vol) at 0°C, and the solution was extracted with MTBE (10 vol) to remove lipophilic material. The aqueous layer was concentrated to remove dichloromethane and methanol. THF (4 vol) was added followed by _NaIO4 (2 eq, 2 wt). The reaction was extracted with MTBE (10 vol) and n-BuOH (10 vol). The combined organic extracts were concentrated and the resulting powder was triturated with ethyl acetate. After filtration, lactol was isolated as light yellow powder.

ER-818638 (1 wt) and LiCl (2 eq, 0.35 wt) were stirred in acetonitrile (8.7 vol). At 25°C, Hunigs base (1.5 eq.) was added. 1N Hydrochloric acid (5 vol) was added and the mixture was extracted with MTBE (10 vol). The organics were concentrated to give ER-818640, which was used in the next step.

The starting α-enester compound (1 wt) (described immediately above) was dissolved in methanol (10 v) and 10 wt% palladium on carbon (0.09 eq, about 0.33 wt) was added under nitrogen. The suspension was then stirred under hydrogen. The suspension was filtered through a pad of Celite ^(R ) (20 wt), rinsing the filter cake with methanol (20 vol). The filtrate was concentrated and purified by flash chromatography to give the product as a colorless oil (94.3% yield).

To a solution of the ester (1 wt) in THF (15 v) was added pyridine (10 eq), _Ph3P (7 eq) and NIS (4 eq) respectively. The reaction mixture was stirred at room temperature. Aqueous citric acid (20 wt%, 10 equiv) was added and the mixture was diluted with TBME (30 v). The aqueous layer was separated and the organic layer was washed with water (5 vol), aqueous sodium thiosulfate (10% w/v, 5 vol), water (5 vol) and brine (5 vol). The organic layer was concentrated and purified by flash chromatography to give the product as a colorless oil.

The iodine starter (1 wt) was dissolved in methanol (30 vol) and heated to 55°C. At 55°C, NaBH ₄ (47 equiv) was added in 6 portions over 80 minutes. The reaction was cooled to 0°C and quenched with 1N hydrochloric acid (30 vol). After stirring for 5 min, the mixture was diluted with brine (30 vol) and extracted with DCM (50 vol x 2). The organic layer was dried over sodium sulfate and concentrated. The crude product was used directly in the next step.

The alcoholic starter (1 wt) (described immediately above) was dissolved in ethanol (70 vol) and Zn (165 eq) was added. The suspension was refluxed at 75-80°C. The reaction mixture was cooled to room temperature and 1N hydrochloric acid (70 vol) was added. The mixture was extracted with DCM (3 x 100 vol), the organic layer was washed with brine and concentrated.

The lactone starting material (described immediately above) was dissolved in DCM (50 vol) and triethylamine (5.0 eq), DMAP (0.3 eq) and TBDPSCl (1.5 eq) were added separately at room temperature under nitrogen, The resulting solution was stirred at room temperature for 2-3 hours. After the reaction was complete, the mixture was diluted with TBME (100 vol), washed with saturated aqueous sodium bicarbonate (10 vol), water (10 vol), and brine (10 vol). The organic layer was concentrated and purified by flash chromatography to give the product as a colorless oil.

Example 4c

Using the general scheme described in Scheme VII above, Example 4c provides an alternative method for the preparation of compounds of formula F-2. This method uses ER-811510 (prepared according to Example 3, wherein acetone is used instead of cyclohexanone) as an intermediate.

ER-811510 (1 wt, 1 eq) was dissolved in dichloromethane (6.3 vol) and cooled to -5°C. Pyridine (0.41 vol, 1.1 eq) was added followed by bromoacetyl bromide (0.44 vol, 1.1 eq) keeping the temperature below 0°C. The reaction was stirred for 1 hour and allowed to warm to room temperature. Water (8 vol) was added and the layers were allowed to separate. The organic layer was washed sequentially with aqueous copper sulfate pentahydrate (1.0 M, 10 vol), water (8 vol), and brine (10 vol), dried over magnesium sulfate, filtered, and concentrated in vacuo to afford ER- 812771.

ER-812771 (1 wt, 1 eq) was dissolved in acetonitrile (6 vol), triphenylphosphine was added, and the reaction was heated to 50°C for 45 minutes. After cooling the reaction solution to -10°C, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU, 0.35 vol, 0.8 eq) was added. The reaction was stirred for 15 minutes, heated to 80°C, held for 45 minutes, then cooled to room temperature. Ammonium chloride (sat aq, 10 vol) was added and the aqueous layer was extracted with ethyl acetate (3 x 10 vol). The combined organic layers were dried over magnesium sulfate and concentrated in vacuo. The crude product was purified by chromatography to afford ER-812772 as a white solid.

ER-812772 (1 weight, 1 equivalent) was dissolved in ethyl acetate (8 volumes), and 10% palladium carbon (0.05 weight, 0.01 equivalent) was added, and the reaction solution was purged with nitrogen and stirred under a hydrogen atmosphere 2 hours. The catalyst was removed by filtration through celite, washing with ethyl acetate. The combined filtrates were concentrated in vacuo to afford ER-812829 as a white solid.

Example 5

Preparation of F-3a

Into a reaction vessel was added D-gulonolactone (1 wt, 1 eq), cyclohexanone (2-3 eq), toluene (6 vol), and p-toluenesulfonic acid (0.021 wt, 0.02 eq ). While stirring, the reaction solution was heated to reflux. After azeotropic removal of water, the reaction was complete. The reaction mixture was cooled to 85-90°C with intensified stirring. With stirring, heptane (5.2 vol) was added over 20-30 minutes. Cool to 65-70°C and stir at 65-70°C for 30 minutes. The solid product was filtered at 65-70°C while maintaining the temperature of the mother liquor >35°C. Filter again at 35-40°C, and keep the mother liquor at room temperature for 30 minutes. Filter the mother liquor again. The filter cake was washed twice with heptane (2 x 1.7 v) and dried to give ER-805715 (84% yield, 1.6 wt).

In another method for preparing ER-805715, D-gulonolactone (1 wt), cyclohexanone (1.32 wt, 2.4 eq), p-toluenesulfonic acid monohydrate (0.02 wt, 0.02 equiv) and toluene (12 v) were refluxed for 19 h with simultaneous azeotropic removal of water. The mixture was washed successively with 5% aqueous sodium bicarbonate (4 vol) and saturated aqueous sodium chloride (2 vol x 2, pH=7). The organic phase was concentrated by distillation (4.5 vol calculated toluene remaining), and after cooling to 100°C, heptane (10 vol) was added while maintaining the internal temperature >80°C. Heat the mixture to reflux for at least 1 hour, then cool to 85°C and age for 3 hours, to 80°C for 3 hours, then cool to 40°C and age for 12 hours. The product was collected by filtration and the filter cake was washed with heptane (2 vol.). The filter cake was air-dried to afford ER-805715 (78% yield, 1.48 wt).

ER-805715 (1 wt, 1 eq) was added to a reaction vessel and dissolved in anhydrous THF (3.34 vol) and anhydrous toluene (2.5 vol). Cool the mixture to -15°C - 10°C. -15°C to -10°C, DIBALH (1.5M in toluene, 2.4 vol, 1.2 eq) was added over 1 hour and the mixture was stirred for 15-30 minutes. At 10°C, the reaction solution was added to a sodium potassium tartrate solution (1 part by weight of sodium potassium tartrate was dissolved in 2.9 parts by weight of water) for inverse quenching, and the resulting mixture was warmed to room temperature and stirred for 4 hours. The mixture was filtered and the layers were separated and extracted with MTBE (2 v). The organic layers were combined and the solvent was removed in vacuo to afford ER-805814 (1.02 wt, 100% yield).

ER-805814 (1 wt) was dissolved in anhydrous THF (3.3 vol) and treated with (methoxymethyl)triphenylphosphine chloride (2.11 wt, 2.1 eq). The reaction mixture was heated to 28-32°C, then a solution of KOtBu (0.66 wt, 2 eq) in anhydrous THF (2.64 vol) was added over 100-140 min maintaining the reaction temperature at 30-35°C. After 5 hours the reaction was cooled to 20-25°C, MTBE (5.11 vol) was added and the mixture was stirred. Brine (3 wts) and water (3 wts) were added (addition started exothermic, water bath controlled at 20-25°C). The organic layer was separated and treated with a solution of maleic anhydride (0.27 wt) in MTBE/THF (1/1 vol, 1.78 vol). Sodium hydroxide solution (0.88 wt in 2.5 volumes of water) was slowly added to the reaction mixture. The organic layer was concentrated to afford crude ER-805815 (0.985 wt). The residue was triturated three times with MTBE/heptane (1/4 v, 6.6 v). The extract was filtered through _SiO2 (3 wt) eluting with MTBE/heptane (1/2 v/v, 45 v). The filtrate was concentrated to afford ER-805815 (0.88 wt, 81% yield).

In another method for preparing ER-805815, (methoxymethyl)triphenylphosphine chloride (3.12 wt., 3.1 eq.) in THF (1.78 wt. wt) suspension was added a solution of t-BuOK (0.989 wt, 3 eq) in THF (4 wt). The addition vessel was rinsed with THF (2 x 0.7 wt). A solution of ER-805814 (1 wt, 1 eq) in THF (1.42 wt) was added to the reaction mixture while maintaining the temperature at 0-10°C. The sample container was rinsed with THF (2 x 0.7 wt). The mixture was stirred overnight at 20-30°C and 3 hours at 30-35°C. The reaction liquid was cooled to below 30° C., and diluted with MTBE (3.7 parts by weight) and 10% by weight aqueous sodium chloride solution (4 parts by weight) successively. The mixture was stirred for 30 minutes and the layers were separated. Maleic anhydride (0.63 wt, 2.2 eq) was added and the mixture was stirred at room temperature for 30 minutes. Keeping the reaction solution below 15°C, water (6 wt.) and sodium hydroxide solution (48 wt%, 0.64 wt., 2.6 eq.) were added dropwise. After stirring below 15°C, the lower layer was separated. Water (6 wts) was added followed by sodium hydroxide solution (48 wt%, 0.64 wts, 2.6 equiv), keeping the mixture below 15°C during addition. After stirring below 15°C, the lower layer was separated. The organic layer was washed three times with 15% by weight aqueous sodium chloride (3 x 4 wt). The organic layer was concentrated in vacuo. The residue was diluted with MTBE (1 wt) and concentrated in vacuo. The residue was diluted with IPE (3 wt) dropwise over 30 minutes at 40-50°C. Stir the suspension for 1 hour at 40-50°C, slowly cool to 0-10°C, and stir for 1 hour. The solid was filtered and the filter cake was washed with IPE (2 wt). The filtrate and washings were concentrated in vacuo. The residue was treated with methanol (2.37 wt) and water (0.4 wt) and extracted with heptane (2.74 wt). The lower layer was extracted 9 times with heptane (2.05 parts). The combined extracts were concentrated in vacuo to give ER-805815 (1.07 wt, 98.6%).

In another workup of ER-805815, the crude organic layer obtained after brine washing and concentration was treated with MTBE (2.86 wt) and Celite (0.5 wt). After stirring for 2.5 hours, heptane (1.46 wt) was added over 2 hours and the mixture was stirred overnight. Filter the precipitate. The filter cake was washed with MTBE/heptane (1:1, 5 wt). The filtrate was concentrated in vacuo until the volume was reduced to about 3 vol. The residue was dissolved in methanol (2 wts) and water (6 wts), and the mixture was extracted with heptane/MTBE (5:1, 3 x 6 wts). The organic layer was separated and concentrated to give ER-805815, which was used in the next step.

ER-805815 (1 wt) was dissolved in acetone (2.4 vol) and water (0.4 vol). N-methylmorpholine N-oxide (0.62 wt, 2 eq) was added and the mixture was cooled to 0-5°C. _OsO4 (0.15M in water, 0.065 vol) was added and the reaction was maintained at 0-5°C. The reaction mixture was stirred at 0-5°C for 12 hours. Water (0.2 vol) was added over 1 hour at 0-2°C. The mixture was stirred at 0-5°C for 1 hour. The product was filtered and the solid was washed twice with pre-cooled (0-5°C) acetone/water (1/1, vol ratio, 2 x 0.7 vol). The product was dried to afford ER-805816 (0.526 wt, 52% yield, residual Os < 17 ppm).

In an alternative method for preparing ER-805816, a solution of ER-805815 (1 wt, 1 eq) in acetone (4 vol) was added to a four-neck flask, followed by water (0.5 wt) at room temperature. To the mixture was added anhydrous N-methylmorpholine-N-oxide (0.38 wt, 1.2 eq). At 25-35°C, potassium osmate dihydrate (0.003 wt, 0.003 eq) was added in batches while cooling with water. The mixture was kept at this temperature for 4 hours. A solution of sodium thiosulfate (0.075 wt, 0.49 eq) in water (0.5 wt) was added at room temperature, and the mixture was stirred for 0.5 hr. The mixture was cooled to 0-5°C and stirred for 2 hours. The resulting precipitate was collected and the wet filter cake was washed with methanol (0.6 wt) and water (1.5 wt) to give the crude product (1.25 wt). A sample of the crude product was dried (0.611 wt). Crude ER-805816 (1.25 wt) was added to water (3.05 wt) and stirred at about 25°C for 2 hours. The precipitate was filtered and washed with water (1.53 wt) to give a crude wet cake (1.05 wt). A crude product sample was dried and sampled, ICPOs = 37 ppm. Crude ER-805816 (1.05 wt) was added to water (2.81 wt) and stirred at about 25°C for 2 hours. The precipitate was filtered, washed with water (1.4 wt) and methanol (0.45 wt) to afford crude ER-805816 (0.736 wt). The wet filter cake was dried to give the crude product (0.56 wt, ICP(Os)=28 ppm). ER-805816 (0.56 wt) was dissolved in acetone (1.76 wt) at 45-55°C. Activated carbon (0.027 parts by weight) was added to the solution, and stirred at the same temperature for 0.5 hours. The mixture was filtered and the filter cake was washed with hot acetone (0.214 wt). The filtrate was kept at 45-50°C, and water (0.83 wt) was added over 10 minutes, maintaining the temperature at 40-50°C during the addition. The mixture was cooled to 0-5°C and stirred for 1.5 hours. The white precipitate was filtered, washed with a solution of acetone (0.17 wt) and water (0.22 wt) and dried to give ER-805816 (0.508 wt, 0.49 equiv, KF 5.0%, ICP(Os) 9.6 ppm).

ER-805816 (1 wt) was suspended in acetic acid (0.89 vol, 5.8 eq) and acetic anhydride (3.57 wt, 13 eq). Anhydrous zinc chloride (0.2 wt, 0.54 equiv) was added. The reaction mixture was stirred at 18-22°C for 24 hours. The reaction was quenched between ice (5 wt) and water (5 v). Ethyl acetate (10 vol) was added with stirring and the aqueous layer was separated. The aqueous layer was back extracted with ethyl acetate (10 vol). The combined organic layers were washed sequentially with brine (10 vol), 5% aqueous sodium ethoxide (6 vol), and brine (6 vol). The organic layer was concentrated. The crude concentrate was dissolved in 25% EtOAc/hex (4 v) and filtered through _SiO2 . The filter pad was washed with 25% EtOAc/hex (2 x 12 vol) and further washed with 25% EtOAc/hex (48 vol). The organic layer was concentrated to afford ER-805819 (1 wt, 81%).

In an alternate method of preparing ER-805819, zinc chloride (0.2 wt, 0.54 eq), acetic anhydride (2.75 wt, 10 eq) and acetic acid (1 wt, 6 eq) were combined. The mixture was cooled to 15-20°C. ER-805816 (1 wt, 1 eq) was added while maintaining an internal temperature of 15-30°C. The mixture was then stirred at 35-40°C for 6 hours. Cool the reaction mixture to below 25°C. Methanol (3.2 wt, 4 vol) was added dropwise keeping the reaction temperature below 25°C. Heptane (2.7 wt, 4 vol) was added. Water (4 wt, 4 vol) was added maintaining the reaction temperature below 25°C. The mixture was stirred for 15 minutes, then the phases were separated. The lower layer was washed twice with heptane (2.7 wt, 4 vol), and the heptane layer was discarded. The lower layer was extracted twice with toluene (6.1 wt, 8 vol). The combined toluene layers were washed twice with 17 wt% aqueous potassium bicarbonate (0.82 wt potassium bicarbonate dissolved in 3.98 wt water, 4.36 vol), twice with water (4 wt) and concentrated. Methanol (3.95 wt, 5 vol) was added at 25-30°C and the mixture was stirred for 10 minutes. Water (0.3 wt) was added at 25-30°C. The mixture was cooled to 0°C and seeded. The mixture was stirred at 0°C for 1 hour, and water (0.7 wt) was added dropwise over 1 hour. Water (4 wt) was added dropwise over 1 hour. The resulting precipitate was filtered, and the filter cake was washed twice with a solution of methanol (1.03 wt) and water (0.7 wt) at 0°C. The filter cake was dried to give ER-805819 (0.99 wt, 0.84 equiv).

ER-805819 (1 wt) was dissolved in anhydrous acetonitrile (15 vol) and treated with ER-803055 (0.93 vol, 2 eq). The reaction mixture was cooled to 0-5 °C and BF3- _OEt2 (0.54 vol, 1.95 equiv) was added over 5 min while maintaining the reaction temperature at 0-5 °C. The reaction mixture was stirred at 0-5°C for 12 hours. The reaction was quenched by adding saturated sodium bicarbonate (20 vol) with vigorous stirring. Extracted twice with ethyl acetate (2 x 8 vol). The combined organics were washed with brine (12 vol) and concentrated to give ER-805821 (1 wt, 88% yield, used directly).

In an alternate method of preparing ER-805821, ER-805819 (1 wt, 1 eq) and ER-803055 (0.93 vol, 2 eq) were dissolved in anhydrous acetonitrile (5.46 wt, 7 vol) . The reaction mixture was cooled to 0-5 °C and _BF3 - _OEt2 (0.54 vol, 1.95 equiv) was added over 5 min while maintaining the reaction temperature at 0-5 °C. The reaction mixture was stirred at 0-5°C for 20 hours, then heptane (5.47 wt, 8 vol) was added at 0-5°C. The phases were separated and the lower layer was treated with heptane (5.47 wt, 8 vol) at 0-5°C. Keeping the reaction temperature at 0-15°C, 7.4% potassium bicarbonate aqueous solution (0.64 parts by weight of potassium bicarbonate dissolved in 8 parts by weight of water) was added dropwise to quench the reaction. Toluene (8.65 wt, 10 vol) was added and the mixture was stirred for 30 minutes. The lower layer was separated and the upper organic layer was washed twice with water (10 vol) and concentrated to give ER-805821 as a crude oil (1.05 wt, 0.935 eq).

ER-805821 (1 wt) was dissolved in anhydrous THF (8.4 vol) and anhydrous methyl acetate (2 vol). With the reaction maintained at 17-23°C, Triton B(OH) (3.6 vol) was added over 2 minutes. The reaction was stirred for 1.5 hours. The reaction mixture was filtered. The filtrate was concentrated and passed through a pad of _SiO2 (5 wt, 20 vol ethyl acetate eluting). The filtrate was washed with brine (2.2 vol) and evaporated to give ER-805822 (0.54 wt, 72% yield).

In another method for preparing ER-805822, ER-805821 (1 wt, 1 eq, 11.18 g, 21.81 mmol) was dissolved in anhydrous MTBE (4.4 wt, 6 vol) and cooled to 0-5 ℃. At 0-5°C, sodium methoxide (28 wt% methanol solution, 0.564 wt, 1.5 eq) was added to the mixture over 1 hour, and stirred at the same temperature range for 3 hours. Acetic acid (0.188 wt, 1.6 eq) was added to quench the reaction, maintaining 0-5°C during addition. The mixture was stirred overnight and then treated with 5 wt% aqueous potassium bicarbonate (3 wt) and ethyl acetate (3.6 wt, 4 vol) at 0-5°C, followed by stirring for 15 min. After phase separation, the lower layer was extracted twice with ethyl acetate (3.6 wt, 4 vol). The combined organic layers were concentrated. Acetone (2 wt, 2.5 vol) and IPE (2 wt, 2.7 vol) were added to the residue and stirred overnight at 0-5°C. The mixture was filtered through Celite ^(R) (0.25 wt), washing with acetone (2 wt). The filtrate was concentrated to give a crude oil (0.55 wt). Acetone (0.2 wt, 0.25 vol) and IPE (0.54 wt, 0.75 vol) were added to the residue and stirred at 40-50°C for 1 hour. The solution was seeded with ER-805822 at room temperature and stirred overnight at room temperature. To the suspension was added IPE (1.27 wt, 1.75 vol) over 2 hours at room temperature. After stirring at room temperature for 5 hours, the precipitate was collected by filtration and the filter cake was washed with acetone/IPE (1/10) (2 vol.). The obtained filter cake was dried in a tray-type chamber at 30-40° C. overnight to obtain the target product ER-805822 (0.286 wt, 0.38 eq, 38.0% yield based on ER-805819).

In an alternative workup of crude ER-805822, the residue obtained after concentrating the final ethyl acetate solution was dissolved in IPA (2 wt) and the solution was heated to 50°C. Heptane (5 wt) was added, the mixture was cooled to 20°C, and seeded. The mixture was stirred overnight at 20°C. Heptane (10 wt) was added, the mixture was cooled to -5°C over 30 minutes and stirred at -5°C for 5 hours. The mixture was filtered and the filter cake was washed with heptane (2 wt). The filter cake was air-dried under vacuum to afford ER-805822 (60%).

ER-805822 (1 wt) was dissolved in ethyl acetate or other appropriate solvent (5 vol) and water (5 vol). Keeping the reaction temperature at 0-10°C, NaIO ₄ (0.58 wt, 1.05 eq) was added in portions over 30 min-1 h. The reaction was stirred for up to 2 hours. The reaction mixture was treated with sodium chloride (1 wt) at 0-10°C and stirred for 30 minutes. The reaction mixture was filtered and the filter cake was rinsed with ethyl acetate (2 vol.). The phases were separated and the lower layer was extracted three times with ethyl acetate (5 vol.). The combined organic layers were washed with 20% aqueous sodium chloride (5 wt). The organic layer was concentrated to give ER804697 (1 wt). The residue was dissolved in toluene (2 vol) and the solution was concentrated. The residue was dissolved in acetonitrile (7 vol) and used in the next step.

Under an inert atmosphere, _NiCl2 (0.025 wt) and _CrCl2 (2.5 wt) were added to the reaction vessel. Anhydrous dichloromethane (5 vol.) was added. Stirring was started and the mixture was cooled to 0-3°C. Keeping the temperature below 20°C, anhydrous DMSO (6.7 vol) was added with vigorous stirring for 45 minutes. ER-804697 (1 wt) was dissolved in anhydrous dichloromethane (1 vol) and added to the reaction vessel. The resulting mixture was allowed to warm to 25°C and ER-806643 (2.58 wt) was added over 20 minutes. Keep the reaction temperature below 45°C. After adding the sample, the reaction solution was stirred at 25-35° C. for 30 minutes. Methanol (5 vol.) was added and the mixture was stirred for 10 minutes. MTBE (33 vol) was added and the suspension was transferred to 1N hydrochloric acid (25 vol) and water (10 vol). The mixture was stirred for 5 minutes. The aqueous layer was back extracted with MTBE (10 vol), and the combined organic layers were sequentially washed with 0.2N hydrochloric acid (17 vol), 1% NaCl solution (2 x 17 vol, washed twice) and brine (13 vol )washing. The organic layer was concentrated and purified ( _SiO2 , 25 wt, 10 column volumes of ethyl acetate/hexane, 1/3.5 vol ratio) to afford ER-804698 (0.53 wt, 61%).

In another approach, the reaction is carried out in the presence of the chiral ligand ER-807363 in a substantially similar manner to the preparation of ER-118047 described hereinafter.

In an alternative method to prepare ER-804698, DMSO (7 vol) and acetonitrile (7 vol) were degassed and cooled to 0-10°C. The solution was treated in batches with CrCl ₂ (10 eq, 3.47 wt) and NiCl ₂ (0.1 eq, 0.037 wt) with the internal temperature not exceeding 20°C. A solution of ER-804697 (1 wt, 1 eq) in acetonitrile (7 vol) and ER-806643 (5 eq, 2.5 wt) was added dropwise at 0-10°C, while the internal temperature did not exceed 15°C. The reaction mixture was stirred overnight at 5-15°C. Methanol (5.5 wt), water (7 wt) and MTBE (5.2 wt) were added to the mixture. The reaction was stirred for 1 hour and the lower layer (Layer 1) was separated. A premixed solution of sodium chloride (1.5 wt) and water (13.5 wt) was added to the upper layer. The mixture was stirred for 1 hour and the lower layer (Layer 2) was separated. Heptane (4.8 wt), methanol (2.8 wt), and a premixed solution of sodium chloride (1.5 wt) and water (13.5 wt) were added to the upper layer. The mixture was stirred for 1 hour and the lower layer (Layer 3) was separated. The upper layer was saved after dehydration (organic 1). Layer 1, methanol (2.8 wts) and MTBE (2.8 wts) were charged to the reactor. The mixture was stirred overnight. Separate and discard the lower layer. Treat the upper layer with layer 2. The mixture was stirred for 1 hour, the lower layer was separated and discarded. The upper layer was treated with layer 3 and heptane (4.8 wt). The mixture was stirred for 1 hour, the lower layer was separated and discarded. The upper layer was saved after dehydration (organic 2). Layer 3, MTBE (0.8 wt) and heptane (2.7 wt) were added to the reactor. The mixture was stirred for 1 hour, the lower layer was separated and discarded. The upper layer was combined with organoids 1 and 2. The combined organics were filtered and concentrated under reduced pressure to give crude ER-804698, which was purified by chromatography ( _SiO2 , 25 wt, 10 column volumes of ethyl acetate/hexane, 1/3.5 by volume) to give ER-804698 (0.67 wt, 57% yield).

In another method for the preparation of ER-804698, the crude material was used directly in the next step without purification.

ER804698 (1 wt, 1 eq) was treated with acetic acid (4.2 wts) and water (4.2 wts). The mixture was heated to 90-97°C for 100 minutes. The mixture was cooled below 15°C, then washed twice with heptane (2 x 2.7 wts) below 15°C. After phase separation, a mixture of 20 wt% potassium bicarbonate aqueous solution (7.7 wt, 35 eq) and MTBE (5.95 wt) was added dropwise to the lower layer under the condition that the temperature did not exceed 15°C. After phase separation, the upper layer was washed sequentially with 5 wt % aqueous potassium bicarbonate (0.2 wt) and twice with 5 wt % aqueous sodium chloride (2 x 0.2 wt). The organic layer was concentrated under reduced pressure and MTBE (1.49 wt) was added. The mixture was heated to 55°C and stirred until dissolved. Heptane (1.00 parts by weight) was added to the solution and the solution was cooled to 40-45°C. Additional heptane (4.47 wt) was added to the solution and the solution was cooled to 5-15°C and then stirred overnight. The crystals were filtered and rinsed with heptane to afford ER-807023 (0.58 wt, 71% yield).

Under nitrogen atmosphere, ER-807023 (1 wt, 1 eq) and MTBE (7.43 wt) were added to the reactor. 2,6-Lutidine (2.15 parts by weight, 7.5 equivalents) was added to the reaction solution. At 0°C, TBSOTf (2.47 wt, 3.5 eq) was added dropwise to the mixture. The reaction mixture was stirred at 0-10°C for 30 minutes, then warmed to 23°C over 1 hour and kept at 23°C for 16 hours. Methanol (0.21 wt, 2.5 eq) and water (14.8 wt) were sequentially added dropwise to the reaction mixture while keeping the temperature below 30°C. After phase separation, the upper layer was washed with 1N aqueous hydrochloric acid solution (16.2 parts by weight), 5% aqueous sodium chloride solution (14.8 parts by weight), 5% aqueous sodium bicarbonate solution (14.8 parts by weight), 5% aqueous sodium chloride solution (14.8 parts by weight), ) and 5% aqueous sodium chloride solution (14.8 wt.). The upper organic layer was concentrated by distillation under reduced pressure to obtain crude ER-804699. Methanol (7.91 wts) was added and the mixture was heated to 50°C for 30 minutes. The mixture was cooled to 0°C over 5 hours, then stirred at 0°C overnight. The solid was filtered and the filter cake was washed with cold methanol (4 wt) and dried to give ER-804699 (1.42 wt, 74% yield).

Under a nitrogen atmosphere, a solution of ER-804699 (1 wt, 1 eq) in toluene (2.60 wts) was added to the reactor. Acetonitrile (4.72 wt) was added. TBSCl (0.011 wt, 0.05 eq) was added. The reaction mixture was allowed to warm to 30°C and NIS (1.25 wt, 4 eq) was added. The reaction mixture was stirred at 30°C for 22 hours. The reaction solution was cooled to 25°C, keeping the internal temperature below 30°C, and a mixture of aqueous sodium thiosulfate and sodium bicarbonate (10.35 wts) was added over 10 minutes. The reaction solution was stirred for 30 minutes at 25°C. Separate the aqueous layer. The upper layer was washed twice with 10% aqueous sodium chloride (2 x 9.9 wt). The organic layer was concentrated under reduced pressure to give crude ER-803895, which was purified by silica gel chromatography to give ER-803895 (0.96 wt, 89.5% yield).

Example 6

Assembly of F-1a, F-2a and F-3a and preparation of B-1939

A. Preparation of (R) or (S) N-[2-(4-isopropyl-4,5-dihydroxy-oxazol-2-yl)-6-methyl-phenyl]-methanesulfonamide

Triphosgene (1 wt, 1 eq) and anhydrous THF (2 vol) were added to a pre-dried glass-lined reactor and cooled to an internal temperature of -10°C. A second pre-dried glass-lined reactor was charged with ER-807244 (1.27 wt, 2.5 eq) and anhydrous THF (3 v) and cooled to an internal temperature of -10°C. The contents of the first reactor were transferred to the second reactor at such a rate that the internal temperature did not exceed 15°C. After adding the sample, the reaction solution was stirred at an internal temperature of 0°C for 1 hour, and then the temperature was gradually raised to 25°C. Nitrogen was sparged for 18 hours to wash off excess phosgene (the offgas was collected with 2N sodium hydroxide solution). MTBE (3 vol.) was added and the solvent was distilled off at 40-46° C. under a nitrogen purge, and more MTBE was added if necessary. After complete removal of phosgene, the mixture was cooled to an internal temperature of 5-10°C and the solution was filtered with MTBE (3 v) washes to afford ER-807245 (1.12 wt, 0.97 eq) as a white crystalline solid.

In a pre-dried inert Reactor 1, ER-807245 (1 wt, 1 eq) and anhydrous DMF (4 vol) were charged. With stirring, the mixture was heated to an internal temperature of 95°C. In Reactor 2, D or L-valinol (1.05 equiv, 0.61 wt) was dissolved in anhydrous DMF (1.3 vol) by heating to an internal temperature of 90°C. The contents of Reactor 2 were transferred to Reactor 1 having an internal temperature of 90°C. Evolution of carbon dioxide was found and the reaction solution was vented by nitrogen loss. After stirring the reaction solution at 90°C for 3 hours, it was cooled to an internal temperature of 65°C. Then, a suspension of lithium hydroxide (0.47 wt, 2 eq) in water (2 v) was added to Reactor 1, and the suspension was stirred at an internal temperature of 65°C for 1 hour. Water (5 vol) was added to the reactor and cooled to an internal temperature of about 5°C over 3 hours. The mixture was stirred at an internal temperature of about 5°C for 8 hours, and the target product was collected by filtration, washed successively with water (2 x 4 vol.) and n-heptane (2 x 3 vol.). The product was dried under vacuum at 35° C. with nitrogen flow for 24 hours or until KF ≤ 250 ppm to give ER-806628 or ER-808056 (0.80 wt, 0.60 eq) as a solid crystal.

In a pre-dried inert reactor under nitrogen, ER-806628 or ER-808056 (1 wt, 1 eq), pyridine (3 wt, 11.4 eq) and DMAP (0.03 wt, 0.05 eq) were charged. After the reaction solution was cooled to an internal temperature of -10°C, methanesulfonyl chloride (1.46 parts by weight, 3 equivalents) was added at a rate such that the internal temperature was lower than 15°C. After adding the sample, the reaction solution was stirred at an internal temperature of 0-15°C for 1 hour, and then slowly raised to 25°C over 2 hours. MTBE (2.6 vol.) followed by water (2 vol.) was added at such a rate that the internal temperature did not exceed 35°C. The biphasic mixture was titrated in portions with 6N hydrochloric acid (ca. 1.9 vol.) until the pH of the aqueous layer was ca. 3-5. If the pH is below 3, add 30% by weight aqueous sodium carbonate for back titration to obtain the desired pH. The phases were separated and the aqueous phase was separated. All organics were mixed with water (0.7 vol) and the aqueous phase was discarded. MTBE was atmospherically distilled to about 2 volumes to obtain a constant boiling point of 55°C with a KF <500 ppm. Additional MTBE was added if necessary. Cool the solution to an internal temperature of 5-10°C, and add seeds to induce crystallization if necessary. n-Heptane (0.5 vol.) was added and the mixture was stirred at 5°C for 18 hours. ER-806629 or ER-807363 was collected by filtration under n-heptane (2 x 3 vol.) washes. The filtrate was concentrated to 1/2 volume and cooled to 0°C to obtain a second crop of crystals. The filter cake was dried under nitrogen for 18 hours. A crude weight of ER-806629 was added to the pre-dried reactor and MTBE (3 vol.) was added. The resulting mixture was heated to an internal temperature of 45-50°C for 45 minutes, then slowly cooled to 5°C over 3 hours, seeded if necessary. n-Heptane (0.5 vol.) was added and the mixture was stirred at an internal temperature of 5°C for 18 hours. The solid product was collected by filtration, washed with n-heptane (2 x 3 vol), and then dried under vacuum at 35°C for 24 hours to give ER-806629 or ER-807363 (1.7 wt, 0.57 eq) as a crystalline solid.

B. Assembly of F-1a and F-2a and intramolecular ether formation:

ER-807363 (1.82 wt, 3.55 eq) was added to Reactor 1 of appropriate size for nitrogen displacement. Anhydrous THF (15 vol) was added. In Reactor 2, ER-806067 (F-1a, 1.14 wt, 1.1 eq) and ER-805973 (F-2a, 1 wt, 1 eq) were mixed and dissolved in anhydrous THF (6.3 part volume). Both reactors were sparged with nitrogen for 30-45 minutes with stirring. Under an inert atmosphere, CrCl ₂ (0.75 wt, 3.55 eq) was added to Reactor 2, and then heated to an internal temperature of 30°C. Triethylamine (0.62 wt, 3.55 eq) was added to Reactor 2 at such a rate that the internal temperature did not exceed 45°C. After adding the sample, keep the internal temperature at 30°C for 1 hour. After 1 hour, Reactor 2 was cooled to 0° C., NiCl ₂ (0.02 wt, 0.1 equiv) was added to make an inert environment, and the reaction solution was allowed to warm to room temperature after adding the contents of Reactor 1 . Reactor 2 was cooled to an internal temperature of 0°C, then ethylenediamine (1.2 vol, 10 eq) was added at such a rate that the internal temperature did not exceed 10°C. CAUTION: Found exothermic. The reaction was stirred for 1 hour, then water (8 vol) and n-heptane (20 vol) were added, the biphasic mixture was stirred for 4 minutes and the layers separated. The organic layer was separated and the aqueous layer was back extracted with MTBE (20 vol). The combined organic layers were concentrated in vacuo to a crude oil, which was then azeotroped with anhydrous THF (2 x 10.5 vol). The crude product was dissolved in anhydrous THF (4.5 vol) and stored at -20°C until used in the next step.

KF analysis was performed on the ER-808227/TFH solution obtained in the previous step. If KF<1000ppm, continue the reaction. If KF > 1000 ppm, vacuum azeotrope with anhydrous THF (4.1 vol). Repeat the azeotropic operation until the requirements are met. The satisfactory final solution contained crude ER-808227 in anhydrous THF (4.1 vol). Once satisfactory, anhydrous THF (106 vol) and the ER-808227/TFH solution from the previous step were added in an appropriately sized inert reactor. The reactor was cooled to an internal temperature of -15°C to -20°C, then a 0.5M solution of KHMDS in toluene (9.1 wts, 3.0 eq) was added at such a rate that the internal temperature did not exceed -12°C. About 4.5 equivalents of KHMDS are required to drive the complete reaction. At 0 °C internal temperature, the reaction solution was added to half-saturated ammonium chloride (40 vol.) for back quenching. Add n-heptane (80 vol) and stir for 2-5 minutes before separating the layers. The organic layer was separated, the aqueous layer was back extracted with MTBE (70 vol), and the combined organic layers were washed with saturated sodium chloride solution (70 vol). The organic layer was separated and concentrated in vacuo. To the crude concentrate was added n-heptane (60 vol). Note: ER-808373 precipitated out of solution. The resulting suspension was filtered and the solid was washed with n-heptane (20 vol). The filtrate was concentrated in vacuo to afford crude ER-806746 (ca. 4 wt) as a brown oil. NOTE: Repeat the filtration operation when additional ER-808373 precipitates out of solution. Crude ER-806746 was purified by SiO ₂ column chromatography to give ER-804027 (1.16 wt, 0.55 eq) as a clear yellowish oil. Chromatography was performed as follows: the column was first flushed with sufficient MTBE to remove water, and then the column was flushed with heptane to remove MTBE. ER-806746 was packed into a column in the form of heptane solution, and it was eluted from the column with heptane/MTBE (5:1) and heptane/MTBE (4:1) successively. fraction.

The reactor was charged with ER-804027 (1 wt, 1 eq) and anhydrous dichloromethane (7.6 vol). The reactor was cooled to an internal temperature of -78°C, then a 1M solution of DIBALH in dichloromethane (3.0 wt, 2.25 equiv) was added at such a rate that the internal temperature did not exceed -60°C. Methanol (0.1 vol) was added at such a rate that the internal temperature did not exceed -60°C. NOTE: Hydrogen is selected and diluted with nitrogen flow. After the addition was complete, the mixture was allowed to warm to room temperature and 1 N hydrochloric acid (10.6 vol) and MTBE (25 vol) were added. The mixture was stirred for 20 minutes and the layers were separated. The organic layer was separated and the aqueous layer was back extracted with MTBE (15.3 vol). The combined organic layers were washed with water (3 vol), saturated sodium bicarbonate (3 vol), and saturated sodium chloride (3 vol), respectively, and concentrated in vacuo. The crude concentrate was purified by _SiO2 column chromatography to afford ER-804028 (0.84 wt, 0.93 eq) as a white foam.

C. Incorporation of F-3a and conversion to B-1939

Under argon atmosphere, ER-803895 (F-3a) was dissolved in anhydrous toluene (14 wts) and cooled to <-75°C. DIBALH (1.5M in toluene, 0.95 wt, 1.3 equiv) was then added at such a rate that the internal reaction temperature was <-70°C. The resulting mixture was stirred for 30 min and then quenched with anhydrous methanol (0.13 wt, 3.2 eq) while maintaining the internal reaction temperature <-65°C. The reaction mixture was allowed to warm to -10°C and transferred to a workup vessel containing 1N brine (10.2 wt) with a rinse of MTBE (3.74 wt). The mixture was stirred for 30 minutes and the aqueous layer was discarded. The organic phase was washed successively with 1N hydrochloric acid (10.2 wts), water (10 wts), saturated aqueous sodium bicarbonate (10 wts) and brine (10 wts), then concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to obtain ER-803896 (0.96 wt, 93% yield). The product was stored under argon at -20°C.

At 0°C, azeotropically dried sulfone ER-804028 (1.0 wt. , 1 eq) in anhydrous THF (5 vol, 4.45 wt). The mixture was stirred at an internal temperature of 0-5°C for 10 minutes, then cooled to <-75°C. The azeotropically dried aldehyde ER-803896 (1.07 wt, 1.23 eq) was dissolved in anhydrous hexane (3.53 wt, 5.35 vol) and cooled to <-75°C. Under the condition of internal temperature ≤ -65°C, add the aldehyde solution to the ER-804028 anion solution through a cannula. The mixture was stirred at -78°C internal temperature for 45 minutes, then quenched by the addition of saturated ammonium chloride (5 vol), methyl tert-butyl ether (10 vol) and water (5 vol). The aqueous layer was discarded, and the organic layer was concentrated under reduced pressure. The crude material was purified by C-18 reverse phase chromatography to give ER-804029 (84%, 1.57 wt).

Sulfone-diol ER-804029 (1 wt, 1 eq) was dissolved in wet dichloromethane (7.4 vol, 0.04 wt% water) and placed in a 20-25°C water bath. Dess-Martin reagent (0.67 wt, 2.5 eq) was added in one portion. The reaction mixture was quenched with saturated sodium bicarbonate (10 vol) and 10 wt% aqueous sodium sulfite (10 vol), and stirred for 30 minutes. The mixture was diluted with saturated NaCl (10 vol) and extracted with MTBE (25 vol). The aqueous layer was discarded, the organic layer was concentrated, and purified by silica gel chromatography to give ER-804030 (0.9 wt, 90%). At -20°C, the material was stored under an inert atmosphere.

Under an inert atmosphere, samarium diiodide solution (2.5 equivalents) was added to a pre-dried reactor, and the solution was cooled to an inner temperature <-70°C. ER-804030 (1 wt) was dissolved in anhydrous methanol (4.1 wt) and anhydrous THF (2.3 wt), then cooled to <-70°C. Add ER-804030 to the cold samarium solution at such a rate that the internal temperature does not exceed -70°C. The reaction was quenched with potassium carbonate/Rochelle salt/water (1/10/100; w/w/v, 15 vol) and MTBE (5 vol) without internal temperature exceeding -65°C. After the addition of the work-up solution was complete, the reaction was allowed to warm to room temperature and the mixture was transferred to another vessel with work-up solution (20 vol, rinse) and MTBE (20 vol, rinse). The aqueous layer was discarded, the organic layer was evaporated, and the residue was purified by silica gel chromatography to obtain ER-118049 (0.77 wt, 85%). Store the product under an inert atmosphere at -20°C.

(S)-ligand ER-807363 (2.05 wt.) was added to the pre-dried reactor for nitrogen replacement. _CrCl2 (0.85 wt, 10 eq) was added in one portion followed by anhydrous acetonitrile (21.5 wt) and the mixture was allowed to warm to and maintain at 30°C-35°C. Triethylamine (0.7 wt, 0.96 vol, 10 eq) was added in one portion and the mixture was stirred for 1 hour. A solution of ketoaldehyde ER-118049 in anhydrous THF (2.43 wt, 2.73 vol) was added over 30 minutes after one addition of _NiCl2 (0.09 wt, 1 eq). The heat was removed and heptane (20.5 wt, 30 vol) and Celite ^(R) (1.5 wt) were added. The mixture was stirred for 5 minutes, filtered through a pad of Celite ^(R) (15 wt) and the pad of ^Celite (R) was rinsed with heptane (7.3 vol) and acetonitrile (5 vol). The filtrate was transferred to another funnel and the lower layer was removed. The combined heptane layers were washed with acetonitrile (up to 47.2 wt, up to 60 vol) as needed. The heptane layer was evaporated under reduced pressure, and the product was purified by silica gel chromatography to obtain ER-118047/048 (0.64 wt, 70%).

Allyl alcohol ER-118047/048 was dissolved in dichloromethane (containing 0.04% by weight of water, 9 volumes), the reactor was placed in a water bath (20° C.), and Dess-Martin reagent (0.48 parts by weight, 1.5 eq. ) treatment solution. The reaction mixture was treated with saturated aqueous sodium bicarbonate (9 vol) and 10% aqueous sodium sulfite (9 vol), stirred for 20 min, and transferred to another funnel via DCM (10 vol). The aqueous layer was discarded and the organic layer was evaporated to a residue. The crude material was purified by flash chromatography (pretreated with 3 column volumes of DCM/heptane (1:1 volume ratio), the material was packed with 1:1 DCM/heptane, then 10/10/1 heptane/DCM /MTBE for elution). Fractions containing product were concentrated and stored at -20°C under an inert atmosphere.

Alternatively, ER-118047/48 was oxidized to the diketone ER-118046 as follows. A flask was charged with ER-118047/48 (1 wt, 1.0 eq) and toluene (10 v) and DMSO (0.15 wt, 2.5 eq) were added at room temperature. Triethylamine (0.31 wt, 4.0 eq) was added and the solution was cooled to -15°C. Neat TCAA (0.33 wt, 1.4 eq) was added and the reaction was allowed to warm to 0°C. It was then stirred at 0°C for 10 minutes. The reaction was stirred again for 10 minutes and then quenched with IPA (0.15 vol). The reaction was stirred at 0°C for 10 minutes. 1 N Hydrochloric acid (5 vol) was added over 2 minutes, the reaction was allowed to warm to room temperature and diluted with MTBE (5 vol). Two clear layers formed and the aqueous layer was removed. The organic layer was washed with 5 volumes of 5% bicarbonate (aq), concentrated on a rotary evaporator to a dark yellow oil, and purified by silica gel chromatography (91% isolated yield).

In an appropriately sized reaction vessel (vessel A) was added imidazole hydrochloride (0.39 wt, 5 eq) followed by 1 M TBAF in THF (7.6 vol, 10 eq) at room temperature. The resulting mixture was stirred until homogeneous (15-30 minutes). In a second reaction vessel (vessel B) was charged ER-118046 (1 wt, 1 eq) and THF (33 vol). The contents of Vessel B were placed under an inert atmosphere and stirred until ER-118046 was completely dissolved. In flask B (ER-118046/THF) was added the contents of flask A (TBAF/imidazole) in one portion. After 3-4 days, the reaction solution was packed into a column and purified by silica gel chromatography.

Under nitrogen atmosphere, the dried residue of ER-118064 (F-12, where ^R is methoxy) was dissolved in anhydrous dichloromethane (28 v) and treated once with PPTS (1.0 wt, 5.2 eq) . After 30-90 minutes, the reaction mixture was loaded directly from the top onto a suitable column and purified by chromatography on silica gel. The target fraction ER-076349 was concentrated in vacuo. All pure fractions were concentrated and the resulting material was azeotroped twice in toluene (20 v) to give ER-076349 as a friable colorless solid/foam (0.44 wt, 0.79 eq (p to residual toluene after correction)).

In a clean and dry reaction vessel (flask C), ER-076349 (1 wt, 1 eq) was dissolved in anhydrous toluene (20 vol) and concentrated to dryness under reduced pressure. This material was redissolved in anhydrous toluene (20 vol) and concentrated to dryness. This material was dissolved in DCM (5 v) and the solution was placed under argon atmosphere. Collidine (0.66 wt, 4.0 eq) was added in one portion. A solution of pyridine in DCM (in flask B) (5 mol %) was added in one portion. The resulting mixture in flask C was cooled to an internal temperature of -20 to -25°C. A solution of _Ts2O in DCM (1.02 equiv) was added dropwise keeping the internal temperature below -16°C. The reaction solution was stirred at -20 to -25°C for 80 minutes, then heated to 0°C over 20 minutes, and stirred for a further 20 minutes. The reaction was quenched with water (2 vol.). The water bath was removed and the reaction was allowed to warm to room temperature (15-20 °C) with stirring (20 min). The reaction was rinsed into a larger vessel with IPA (100 vol), and aqueous ammonium hydroxide (100 vol) was added to the reaction. The reaction was stirred at room temperature for 15-36 hours, monitoring the disappearance of the in situ formed tosylate (ER-082892) and epoxide (ER-809681). Under reduced pressure, the reaction solution was concentrated to dryness or nearly dryness. The resulting material was diluted with DCM (25-40 vol) and washed with pH 10 buffer (NaHCO3/NaCO3(aq), 10 vol). The aqueous phase was back extracted with 25 volumes of DCM and the combined organic layers were concentrated to dryness. The resulting free amine was purified by silica gel chromatography (acetonitrile/water buffer as mobile phase). The combined fractions were concentrated under reduced pressure to remove acetonitrile. The resulting aqueous layer was diluted with DCM (40 v) and 30 v stock buffer (pH 10, sodium bicarbonate/sodium carbonate). The layers were mixed well and separated. The aqueous phase was back extracted with 25 volumes of DCM and the combined organic layers were concentrated to dryness. The resulting free amine was polished filtered into a 3:1 DCM/pentane solution and concentrated to dryness (0.80 wt) to afford B-1939.

While we have described a number of embodiments of the invention, it will be apparent that our basic examples can be altered to provide other embodiments utilizing the compounds and methods of the invention. It is therefore to be understood that the scope of the present invention is defined by the appended claims rather than by the specific embodiments set forth by way of example.

Claims (12)

1. a kind of compound of formula F-4:

Wherein: PG¹、PG²And PG³Respectively stand alone as hydrogen or suitable hydroxyl protection base；

R¹For R or-OR；

Each R stands alone as hydrogen, C_1-4Halogenated aliphatic base, benzyl or C_1-4Aliphatic group；And

LG¹For suitable leaving group.

2, compound according to claim 1, wherein the compound is the compound of formula F-4 ':

3, compound according to claim 2, wherein R¹For OR, wherein R is hydrogen, methyl or benzyl.

4, compound according to claim 2, wherein PG¹And PG²It is hydrogen.

5, compound according to claim 2, wherein PG¹And PG²Respectively stand alone as suitable hydroxyl protection base.

6, compound according to claim 5, wherein PG¹And PG²One or two of together with the oxygen atom of each self-bonding be silyl ether or aryl alkyl ethers.

7, compound according to claim 6, wherein PG¹And PG²It is t-butyldimethylsilyl.

8, compound according to claim 5, wherein PG¹And PG²Glycerol protection base is formed together together with the oxygen atom in conjunction with them.

9, compound according to claim 8, wherein the glycerol protection base is cyclic ketal or ketal, silylene derivative, cyclic carbonate ester or ring borate.

10, compound according to claim 2, wherein LG¹For sulfonyloxy, the alkylsulfonyloxy optionally replaced, the alkenyl sulfonyloxy optionally replaced or the aryl-sulfonyl oxygen optionally replaced.

11, compound according to claim 10, wherein LG¹For mesyl or tosyl.

12, compound according to claim 2, wherein the compound are as follows: