BG106216A - Hemisynthetic method and new compounds - Google Patents

Abstract

Methods are provided for preparing a compound with a fused ring structure of formula which comprises one or more reactions starting from a 21-cyano compound of formula where typically: R1 is an amidomethylene group or and acyloxymethylene group; R5 and R 8 are independently chosen from -H, -OH or -OCOCH2OH, or R5 and R8 are both keto and the ring A is a p-benzoquinon ring; R14a and R14b are both -H ozone and the other is -OH, -OCH3 or -OCH2CH3, or R14a and R14b together form a keto group, and R15 and R18 are independently chosen from H or -OH, or R5 and R8 are both keto and the ring A is a p-benzoquinone ring in modified starting materials, the 21-cyano group can be replaced by other groups introduced using nucleophilic reagents. 33 claims

Description

Technical field

The present invention relates to synthetic methods and, in particular, the present invention relates to semi-synthetic methods.

BACKGROUND OF THE INVENTION

European Patent 309,477 relates to ecteinascidins 729,743, 745, 759A, 759B and 770. The ecteinascin compounds have been described to possess antibacterial and other beneficial properties. Ecteinascidin 743 is currently undergoing clinical trials as an antitumor agent.

Ecteinascidin 743 has a complex tris (tetrahydroisoquinolinophenol) structure of the following formula (I):

It is obtained by isolation from marine organism extracts Ecteinascidin turbinals. The yield is low and alternative preparative methods are sought.

A synthetic method for the preparation of ecteinascidin compounds is described in US Patent 5,721,362. The method according to this patent is long and complex, there are 38 examples, each of which describes a step of the synthetic sequence until it reaches ecteinascidin 743.

Claim 25 of US Patent 5,721,362 relates to an intermediate phenolic compound of formula (11), which we call Intermediate 11 or lnt-11. It has the following bis (tetrahydroisoquinolinophenol) structure (II):

where MOM is a methoxymethyl substituent and TBDPS is a 2,3-b-butyldiphenylsilyl substituent.

From Intermediate 11 it is possible to synthesize another interesting antitumor agent, phthalacidin, see Proc. Natl. Acad.

• · · • · · · · · ·

• · · ·

Sci. USA, 96.3496-3501.1999. phthalasidine is a derivative of nais (tetrahydroisoquinolinophenol) of formula (III)

In ecteinascidin 743,1,4, the bridge has the structure of formula (IV):

Other known ecteinescidins include compounds with different bridged cyclic ring systems, such as in ecteinescidins

722 and 736, where the bridge has a structure of formula (V):

583 and 597, where the bridge has a structure of formula (VI):

The complete structure of these and the corresponding compounds is given in J. Am. Chem. Soc. (1996) 118.9017-9023. This message is given for comparison.

Other compounds known to be lacking a cyclic ring ring system are known. These include bis (tetrahydroisoquinolinequinone) antitumor-antimicrobial antibiotics safracin and saframycin, and natural seafood renieramycin and gestomycin isolated from cultivated germs or fungi. They all have the usual dimeric tetrahydroisoquinoline carbon frame. These compounds can be classified into four types, types I to IV, with respect to oxidation of the aromatic nucleus.

Type I, dimeric isoquinolinquinones, is a system of formula (VIII), and for the most common in this class of compounds, see the following Table I.

Table I

Type I structure of saframycin antibiotics

····· · ·· · 4 • · V ···· ··· • · · · · * • · · · ····

Substitutes

Compound R ^{, 4a} R ^{, 4b} R ²¹ _R 25a R ^25b _r2 5c saframycin A N. n CN 0 0 CH ₃ saframycin B .H n n 0 0 CH3 saframycin C N. OSN ₃ n 0 0 CH ₃ saframycin G N. . he CN 0 0 CH3 saframycin H n n CN he CH ₂ COCH ₃ CH3 saframycin S n n HE 0 0 CH3 saframycin Uz n n CN nh ₂ H CH3 saframycin Ydi n n CN nh ₂ H C ₂ H ₅ saframycin Adi n n CN 0 0 C ₂ H ₅ saframycin Yd ₂ n n CN NH: H N. saframycin Ugh n Q * CN NH: H CH3 saframycin Y ₂ t> d n Q * CN NH: H C ₂ H ₅ saframycin AN _g n n CN H ° OH ° CH3 saframycin AN ₂ Ac n n CN H OAc CH3 saframycin AN, n n CN OH ^fl H ^fl CH3 saframycin AHiAc n n CN OAc H CH3 saframycin AR3 n n N. H OH CH3

'denotes variables ^b where Q is of formula (IX):

dream ₃

Aromatic rings of type I are given for saframycin A, B and C; G and H; and S is isolated from Streptomyces lavendulae as side compounds. The cyan derivative of saframycin A, called cyanoquinonamine, is known from Japanese Kokai JP-A2 59/225189 and 60/084288. Saframycin Y & Ydi, Adi and Yd2 are obtained from Streptomyces lavendulae by direct biosynthesis in a suitably selected culture medium. Saframycin Y _2b and Y _2t> d dimers obtained by connecting the nitrogen at C-25 of one unit to the C-14 of the other, may also be obtained from Streptomyces lavendulae in suitably selected culture medium. Saframycin ARi (= AN ₂ ), a microbial reduction product of saframycin A at C-25 derived from Rhodococcus amidophilus, is also obtained by the non-stereoselective chemical reduction of saframycin A with sodium borohydride as an 11 mixture of epimers, followed by chromatographic separation AN _} is less polar). The following reducing product saframycin AR3,21-deciano-25-dihydro-saframycin A (= 25-dihydrosaframycin B) was obtained by the same microbial conversion. Another type of microbial conversion of saframycin A, using Nocardia, produces saframycin B, and subsequent reduction with Mycobacterium produces saframycin AHiAc. The 25-O-acetates of saframycin AHj and saframycin AH _{2 were} also chemically obtained for biological testing.

Type I compounds of formula (X) are also isolated from sea mushrooms, see Table II.

Table II

Type I structures of marine mushroom compounds

OSN ₃

L · -R14a R14b

R

Ί

Substitutes

r'43 _R1 4b “F ¹ R renieramycin A HE n n -C (CH3) = CH-CH3 renieramycin B. axis ₂ n ₅ n n -C (CH3) = CH-CH3 renieramycin C. he 0 0 -C (CH3) = CH-CH3 renieramycin D OS ₂ H ₅ 0 0 -C (CH3) = CH-CH3 reniramycin E N. n he -C (CH3) = CH-CH3 renieramycin F OCH3 n he -C (CH3) = CH-CH3 xestomycin OCH3 n n -CH3

Reneramycin A-D was isolated from the antimicrobial extract of a Reniera mushroom collected in Mexico, together with the biogenetically related monomeric isoquinolines rhenirone and the corresponding compounds. The structure of renieramycin A was initially determined by reverse stereochemistry at C-3, C-11 and C-13. Careful examination of the magnetic resonance data, however, for novel, corresponding renieramycin E and F compounds isolated from the same fungus collected in Palau reveals that the reenamycin ring binding is identical to that of saframycin. This result leads to the conclusion that the previously determined stereochemistry of renieramycin A-D should be the same as that of saframycin.

Xestomycin is found in a mushroom of the species Xestospongia collected from Sri Lanka waters.

Compounds of type II of formula (XI) with reduced hydroquinone ring include saframycin D and F isolated from Streptomyces lavendulae and saframycin Mx-1 and Mx-2 isolated from Myxococcus xanthus. See Table III.

^ R25c

Compound saframycin D saframycin F saframycin MX-1 saframycin MX-2

• · · · 9 99 9 9 9 9 9 9 9 9 9 99 9 9 9 _R 14a R ^{, 4b} 0 0 0 0 n OSN ₃ n OCH3

• 9 9 ··· ··· · · ·· · • 9 9 99

9 · 9 99

9 9 99

9999999 99 999

Substitutes

R ²¹ R ^25a

H0

CN0

ONN

R ^25b R ^25c 0 CH3 0 CH3 CH3 nh ₂ CH3 nh ₂

The type III skeleton is found in the antibiotics safracins A and B isolated from the ennobled Pseudomonas fluorescens. These antibiotics of formula (XII) consist of a tetrahydroisoquinolinquinone subunit and

where R ²¹ is -H in safracin A and -OH in safracin B.

Saframycin R, the only compound classified as a type IV skeleton, is also isolated from Streptomyces lavendulae. This compound of formula (XIII), composed of a side chain of a glycol ester side chain on one of the phenolic oxygen atoms, is, perhaps, a precursor of saframycin A because of its moderate toxicity.

OSN ₃

All these known compounds have a condensed system of five rings (A) to (E), as shown in the following structure of formula (XIV):

The rings A and E are phenolic in the ecteinascidins and some other compounds.

while other compounds, especially saframycin, rings A and E are quinoline. In known compounds, rings B and D are tetrahydro, while ring C is perhydro.

Object of the invention

The development of new active compounds with the condensed five-ring system of the known compounds is required, as well as alternative synthetic methods for the preparation of ecteinascidin compounds and corresponding compounds. Such synthetic methods can lead to more economical pathways for the production of known antitumor agents, as well as allow the production of new active compounds.

SUMMARY OF THE INVENTION

In one aspect, the present invention relates to the use of a known compound, safracin B, also called quinonamine, in a semi-synthetic method.

Generally, the present invention relates to a semi-synthetic method for the preparation of intermediates, derivatives and corresponding structures of ecteinascidin or other tetrahydroisoquinolinophenolic compounds. The compounds are derived from 6c (tetrahydroisoquinoline) alkaloids. Suitable starting materials for the semi-synthetic method include classes of otaframycin and safracin antibiotics obtained from different grown broths, as well as classes of reineramycin and xestomycin compounds derived from sea mushrooms.

The general formula (XV) of the starting compounds is as follows:

R ¹ is an amidomethylene group, such as -CHrNH-CO-CR ^25a R ^25b R ^25c , in which R ^25a and R ^25b form a ketocrop or one radical is -OH, -NH2 or -OCOCCH ₃ and the other is -CH 2 COCH 3, -H , -OH or -OCOCH 3, provided that when R ^25a is -OH or -NH 2, R ^25b is not-OH and R ^25c is -H, -CH 3 or -CH ₂ CH ₃ , or R ¹ is an acyloxymethyl group , such as -CH ₂ -O-CO-R, in which R is -C (CH ₃ ) = CH-CH ₃ or CH ₃ ;

R ⁵ and R ^{8 are} independently selected from -H, -OH or -OCOCH ₂ OH, or both R ⁵ and R ⁵ are ketogroup and ring A is a p-benzoquinone ring;

both R ^14a and R ^14b are -H or one -H and the other is -OH, -OCH ₃ or -OCH ₂ CH ₃ , or R ^14a and R ^14b together form a ketone group;

R ¹⁵ and R ^{18 are} independently selected from -H or -OH, or both R ⁵ and R ⁸ are ketogroup and ring A is a p-benzoquinone ring; and R ²¹ is -OH or -CN.

The more general formula of this class of compounds is presented below:

X

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Wherein each of the substituents defined as R _b R _2l R ₃ , R 4, Rs, Re, R 7, Re, R 9, ^R 10 is independently selected from the group consisting of H , OH, OCH3, CN, = 0, OCH _3;

in which X are different amide or ester groups contained in said natural products;

in which each dashed circle represents one, two or three possible double bonds.

Accordingly, according to the present invention, we provide novel semi-synthetic methods for the preparation of intermediates, including intermediate 11 and hence, for the preparation of ecteinascidin compounds, as well as phthalacidin and the corresponding compounds. Each of the semi-synthetic methods of the present invention involves a series of transformation steps to reach the desired product. Each step, by itself, is a method according to the present invention. The present invention is not limited to exemplary methods which are illustrative, and alternative methods may be presented, for example, by appropriately changing the order of the transformation steps.

In particular, the present invention encompasses the use of a cyanide starting material of general formula (XVI):

OCH3

^R18 \ CH3 R5 ^: Well. CH3 \ A H \ * h --N --CH? ' ⁵ CH ₃ O. Ί ^N R8 R ¹ CN

wherein R ¹ , R ⁵ , R ⁸ , R ^{, 4a} , R ^14b , R 1, and R ¹⁸ ca as defined above.

Other compounds of formula (XVI) with different substituents at the 21-position may also be suitable starting materials. In general, any derivative that can be obtained by nucleophilic displacement of the 21-hydroxyl group of the compounds of formula (XV) in which 12 to R ²¹ is a hydroxyl group in the cis position. Examples of suitable 21 substituents include, but are not limited to:

mercaptogroup;

an alkylthio group (the alkyl group contains from 1 to 6 carbon atoms);

arylthio group (aryl group contains from 6 to 10 carbon atoms and is unsubstituted or substituted by from 1 to 5 substituents selected from, for example, an alkyl group containing from 1 to 6 carbon atoms, alkoxy groups containing from 1 to 6 carbon atoms , halogen atoms, mercapto groups and nitro groups);

an amino group;

a mono- or dialkylamino group (each alkyl group contains from 1 to 6 carbon atoms);

a mono- or diarylamino group (each aryl group is as defined above for arylthio groups);

an α-carbonylalkyl group of formula -C (R ^a ) (R ^b ) -C (= O) R ^c where R ^a and R ^b are selected from hydrogen atoms, alkyl groups containing from 1 to 20 carbon atoms, aryl groups (as defined above for the arylthio groups) and aralkyl groups (in which the alkyl group containing from 1 to 4 carbon atoms is replaced by an aryl group defined above for the arylthio group), provided that one of R ^a and R ^b is a hydrogen atom;

R0 is selected from a hydrogen atom, an alkyl group containing from 1 to 20 carbon atoms, aryl groups (as defined above for the arylthio groups), an aralkyl group (in which an alkyl group containing from 1 to 4 carbon atoms is replaced by an aryl group defined above for the arylthio group), an alkoxy group containing from 1 to 6 carbon atoms, an amino group or a mono- or dialkylamino group, as defined above.

Therefore, in a general aspect, the present invention relates to methods in which the first step yields a 21-derivative using a nucleophilic reagent. We call these compounds 21-Nuc compounds.

The presence of the 21-cyano group is required for some of the end products, especially ecteinascidin 770 and phthalacidin, while for other end products, it acts as a protecting group that can easily be converted to another substituent, such as the 21-hydroxyl group of ecteinascidin 743 or of 21-hydroxyphthalacidin. Using the 21-cyan compound as a starting material effectively stabilizes the molecule during the subsequent synthetic steps until it is separated. Other 21-Nuc compounds show these and other advantages.

In one important aspect, the present invention relates to the use of a 21-cyan compound of general formula (XVI) in the preparation of bis- and tris-tetrahydroisoquinolinophenol) compounds. The products that can be obtained include intermediates, such as intermediate 11, and ecteinascidins and phthalacidin, as well as novel and known compounds of the corresponding structure.

Preferred starting materials include those compounds of formula (XV) or (XVI), wherein both R ¹⁴ ® and R ^14b are hydrogen. Preferred starting materials also include, compounds of formula (XV) or (XVI), wherein R ^'5 is hydrogen. Moreover, preferred starting materials include compounds of formula (XV) or (XVI), wherein ring E is a phenolic ring. Preferred starting materials also include compounds of formula (XV) or (XVI) wherein at least one, and more preferably, two or three of R ⁵ , R ⁸ , R ¹⁵ and R ¹⁸ are not hydrogen.

Examples of suitable starting materials of the present invention include saframycin A, saframycin B, saframycin C, saframycin G, saframycin H, saframycin S, saframycin Y ₃ , saframycin Ydi, saframycin Adi, saframycin Yd2, saframycin AH, saframycin · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

ANgAc, saframycin AN _b Saframycin ΑΗ, Αο, saframycin AR ₃ , renieramycin A renieramycin B, renieramycin C, renieramycin D, renieramycin E, renieramycin F, xestomycin, saframycin D, saframycin F, saframycin MX-1, saframycin Mx-1, saframycin A safracin B and saframycin R. Preferred starting materials have a cyano group at position 21 for the group R ²¹ .

In a particularly preferred aspect, the present invention includes a semi-synthetic method in which the transformation steps are applied to safracin B:

Safracin B is a ring system very similar to ecteinascidins. This compound has the same pentacyclic structure and the same substituents in the right aromatic nucleus, ring E. Also, safracin B is very close to some of the synthetic intermediates in the general synthesis of ET-743, in particular, to intermediate 11. This an intermediate can be transformed into ET-743 ecteinascidin using a well-known method. Synthetic conversion of safracin B into intermediate 11 therefore results in a semi-synthetic method for the preparation of ET-743.

Therefore, the present invention also relates to intermediate 11 obtained from safracin B and to the compounds derived from safracin B

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Intermediate 11, in particular, ecteinascidin compounds. The present invention also relates to phthalacidin derived from safracin B. The present invention also relates to the use of safracin B in the preparation of intermediate 11, phthalacidin and ecteinascidin compounds, and other intermediates according to the present invention. invention. The present invention also relates to the compounds described herein obtained from other proposed starting materials and the use of these compounds in the preparation of said compounds.

The more preferred starting materials of the present invention have a 21-cyano group. The most preferred compound of the present invention is the compound of formula 2. This compound is obtained directly from safracin B and is considered to be a key intermediate in the semi-synthetic method.

In a related aspect, cyanosafracin B is obtained by fermentation of safracin B obtained from the Pseudomonas fluorescens strain and the culture broth is treated with cyanide ion. The preferred strain of Pseudomonas fluorescens is strain A2-2, FERM BP-14, used in the method according to EP 055,299. A suitable source of cyanide ion is potassium cyanide. In the usual treatment, the broth is filtered off and an excess of cyanide ion is added. After an appropriate stirring time of 1 hour, the pH was left alkaline, 9.5, and extracted with an organic solvent produced a crude extract, which could then be purified to give cyanosafracin B.

Undoubtedly, the stereochemistry shown in the present description is based on an understanding of the true stereochemistry of natural products. Due to an error in the labeled stereochemistry, an appropriate correction of the formulas presented in this description should be made. Moreover, since the synthesis tends to be modified, the present invention is extended to stereoisomers.

The products of the present invention are generally of formula (XVila):

or of formula (XVIIb):

• ···· • ♦ · • ·· · ·· · · • · • ft • · • · • • • · · • · • · ft • · · • · • · • «· ♦ ft * ft ··· • ft • ft

where:

R ¹ is a possibly protected or derivative aminomethylene group, optionally protected or derivative hydroxymethylene group, with R ¹ being as defined for formula (XV);

R ⁴ eH;

or R ¹ and R ⁴ together form a group of formula (IV), (V), (VI) or (Vll):

R ⁷ is -OCH 3 and R ⁸ is -OH or R ⁷ and R® together form the group -O-CH 2 -O-; both R ^14a and R ^14b are -H or one is -H and the other is -OH, -OCH 3 or -OCH 2 CH 3, or R ^14a and R ^14b together form a ketone group; and R ^{15 is} H or -OH;

R ²¹ is -H, -OH or -CN;

and derivatives including their acyl derivatives, in particular where R ⁵ is acetyloxy or another acyloxy group containing up to 4 carbon atoms and including derivatives where the group -NCH 3 - at the 12th position is replaced by -NH- or -NCH 2 CH 3 - , and derivatives in which the -NH ₂ group in the compound of formula (VI) can give derivatives.

In formulas (XVIIa) or (XVIIb), R ¹ is usually aminomethylene, amidomethylene or R ¹ with R ⁴ form a group (IV) or (V). Suitable amidomethylene groups include those of formula -CH2-NH-CO-CHCH3-NH2- derived from alanine and similar groups derived from other amino acids, especially D and L, glycine, valine, leucine, isoleucine, phenylalanine, tyrosine, tryptophan , methionine, cysteine, aspartate, asparagine, glutamic acid, glutamine, lysine, proline, serine, threonine, histidine and hydroxyproline.

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

The general formula for the group R ¹ is -CH _r NH-aa, where aa represents an acylamino acid group.

The R ¹ group may be acylated at the -NH ₂ group and, for example, the N-acyl derivatives may be derived from the groups -CH ₂ -NH ₂ and -CH ₂ -NH-aa. Acyl derivatives can be their N-acyl or Nthioacyl derivatives, as well as cyclic amides. The acyl groups may illustratively be alkanoyl, haloalkanoyl, arylalkanoyl, alkenyl, heterocyclylacyl, aroyl, arylaroyl, haloaroyl, nitroaroyl or other acyl groups. The acyl groups may be of the formula -CO-R ^a , where R ^a may be different groups such as alkyl, alkoxy, alkylene, arylalkyl, arylalkylene, amino acid acyl, or heterocyclyl, each of which may be substituted by halo, cyano, nitro , carboxyalkyl, alkoxy, aryl, aryloxy, heterocyclyl, heterocyclyloxy, alkyl, amino or substituted amino. Other acylating agents include isothiocyanate such as aryl isothiocyanates, especially phenylisothiocyanate. The alkyl, alkoxy or alkylene groups of R ^a are suitably containing from 1 to 6 or 12 carbon atoms and may be linear, branched or cyclic. The aryl groups are usually phenyl, biphenyl or naphthyl. The heterocyclic groups may be aromatic or partially or fully unsaturated and may suitably contain from 4 to 8 ring atoms, preferably 5 or 6 ring atoms, one or more of the heteroatoms being selected from nitrogen, sulfur and oxygen.

Without limitation, the R ^a groups typically include alkyl, haloalkyl, alkoxyalkyl, haloalkoxyalkyl, arylalkylene, haloalkylarylalkylene, acyl, haloacyl, arylalkyl, alkenyl and amino acid. For example, R ^a -CO- can be acetyl, trifluoroacetyl, 2,2,2-trichloroethoxycarbonyl, izovalerilkarbonil, trans-3- (trifluoromethyl) cinnamoylcarbonyl, heptafluorbutirilkarbonil, dekanoilkarbonil, trans-cinnamoylcarbonyl, butirilkarbonil, 3-hlorpropionilkarbonil, cinnamoylcarbonyl, 4metilsinamoilkarbonil , hydrosinamoylcarbonyl or trans-hexenoyl19 • · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Carbonyl, or alanyl, arginyl, aspartyl, asparagyl, cystal, glutamyl, glutaminyl, glycyl, histidyl, hydroxypropyl, · * ··· ·· ··· isoleucyl, leucyl, lysyl, methmonyl, phenylalanil propyl, seryl, threonyl, tyronyl, tryptophil, tyrosyl, valyl, and other more unusual amino acid acyl groups, as well as phthalimidogroups and other cyclic amides. Other examples can be found in the list of protecting groups.

Compounds in which R ^a -CO- is derived from an amino acid and includes an amino group can form acyl derivatives. Suitable N-acyl compounds include dipeptides that can form N-acyl derivatives.

In one embodiment relating to an intermediate, Ring A is modified to a structure shown as formula (XX) or (XXI), discussed later.

In another embodiment relating to intermediates, the group R ¹ may be -CH 2 O-CO-CFu-CHrS-Prot ³ prepared from a compound of formula (XIX) in which Prot ³ and Fu have the meanings shown. In this case, R ⁷ and R ⁸ form an oxymethylenoxy group. The R ¹⁸ group is usually protected. Typically, R ²¹ is cyano.

R ^14a and R ^14b are preferably hydrogen. R ¹⁵ is preferably hydrogen. O-acyl derivatives are suitable aliphatic O-acyl derivatives containing from 1 to 4 carbon atoms and generally an O-acetyl group, especially in the 5-position.

Suitable protecting groups for phenols and hydroxy groups include ethers and esters, such as alkyl, alkoxyalkyl, aryloxyalkyl, alkoxyalkoxyalkyl, alkylsilylalkoxyalkyl, alkylthioalkyl, ariltaoalkil, azidoalkyl, cyanoalkyl, chloroalkyl, heterocyclic, arylacyl, haloarilatsil, cycloalkylalkyl, alkenyl, cycloalkyl, alkylarylalkyl, alkoxyarylalkyl, nitroarylalkyl, haloarylalkyl, alkylaminocarbonylarylalkyl, alkylsulfinylarylalkyl, alkylsilyl and other ethers, and arylacyl, arylalkylcarbonate, aliphatic carbonate, alkylsulfinylarylalkyl20 · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Carbonate, alkylcarbonate, arylhaloalkylcarbonate, arylalkenylcarbonate, arylcarbamate, alkylphosphinyl, alkylphosphinothioyl, arylphosphinothioyl, arylalkylsulfonates and other esters. It is possible that these groups are replaced by the groups R ¹ described above.

Suitable protecting groups for amines include carbamates, amides and other protecting groups such as alkyl, arylalkyl, sulfo or halo-

arylalkyl, alkilsililalkil, arylalkyl, cycloalkylalkyl, alkylarylalkyl, heterocyclylalkyl, nitroarilalkil, acylaminoalkyl, nitroarilditioarilalkil, ditsikloalkilkarboksamidoalkil, cycloalkyl, alkenyl, arylalkenyl, nitroarilalkenil, heterocyclylalkenyl, heterocyclyl, hydroxyheterocyclic, alkyldithio, alkoxy- or halo, or alkilsulfinilarilalkil, heterocyclylacyl, and other carbamates, and alkanoyl, haloalkanoyl, arylalkanoyl, alkenyl, heterocyclylacyl, aroyl, arylaroyl, haloaroyl, nitroaroyl and other amides, as well as alkyl, alkenyl, alkylcyl l ilalkoxyalkyl, alkoxyalkyl, cyanoalkyl, heterocyclyl, alkoxyarylalkyl, cycloalkyl, nitroalkyl, arylalkyl, alkoxy or hydroxyarylalkyl, and many other groups. It is possible that these groups are replaced by the groups R ¹ described above.

Examples of such protecting groups are given in the following tables.

Protection for hydroxyl group

Ethers Coexisting methyl methoxy methyl IOM benzyloxymethyl PTO methoxyethoxymethyl MEM 2- (trimethylsilyl) ethoxymethyl SEM methylthiomethyl MTM phenylthiomethyl RTM

azidomethyl

cyanomethyl

2,2-dichloro-1,1-difluoroethyl 2-chloroethyl 2-6 romethyl tetrahydropyranyl 1-ethoxyethyl phenacyl 4-bromophenacyl cyclopropylmethyl allyl propargyl isopropyl cyclohexyl t-butyl benzyl

2,6-dimethylbenzyl 4-methoxybenzyl o-nitrobenzyl

2,6-Dichlorobenzyl 3,4-dichlorobenzyl 4- (dimethylamino) carbonylbenzyl 4-methylsulfinylbenzyl 9-anthryl methyl

4-picolyl heptafluoro-p-tolyl tetrafluoro-4-pyridyl trimethylsilyl t-butyldimethylsilyl t-butyldiphenylsilyl triisopropylsilyl

TNT

EE

MRM or PMB

Msib

TMS

TBDMS

TBDPS

TIPS

Esters arilformiat arylacetate arillevulinat arilpivaloat arilbenzoat aryl-9-fluorkarboksilat arilmetilkarbonat 1 -adamantilkarbonat t-butylcarbonate 4-metilsulfinilbenzilkarbonat 2,4-dimethylpent-H-ylcarbonyl aryl-2,2,2-trichloroethyl arilvinilkarbonat arilbenzilkarbonat arylcarbamate dimetilfosfinil dimethylphosphinothioyl difenilfosfinotioil arilmetansulfonat ariltoluensulfonat aryl-2 -formylbenzenesulfonate

• · · · · · · ·

ArOPv

BOC-OAG

Msz-Oar

Doc-Oar

Dmp-OAr

Mpt-OAr

Dpt-Oar

Amino group protection

Carbamates methyl ethyl

9-fluorenylmethyl

9- (2-sulfo) fluorenyl methyl

9- (2,7-dibromo) fluorenylmethyl

Abbreviation

Fmoc

17-tetrabenzo [a, c, d, 1] fluorenyl methyl Tbfmoc 2-chloro-3-indenylmethyl Climoc benzene [Pinden-3-ylmethyl Bimoc 2,7-di-1-butyl [9- (10,10-dioxo-10,10,10,10- tetrahydrothioxanthyl) methyl DBD-Tmoc 2,2,2-trichloroethyl Troc 2-trimethylsilylethyl Teoc 2-phenylethyl hZ 1- (1-adamantyl) -1-methylethyl Adpoc 2-chloroethyl 1,1-dimethyl-2-chloroethyl 1,1-dimethyl-2-bromoethyl 1,1-dimethyl-2,2-dibromethyl DB-t-BOC 1,1-dimethyl-2,2,2-trichloroethyl TCBOC 1-methyl-1- (4-biphenyl) ethyl Bpoc 1- (3,5-di + butylphenyl) -1 -1-methylethyl t-Burmeoc 2- (2'-and 4'-pyridyl) ethyl Pyoc 2,2-bis (4'-nitrophenyl) ethyl Bnpeoc n- (2-pivaloylamino) -1,1-dimethylethyl 2 - [(2-nitrophenyl) dithio] -1-phenylethyl NpSSPeoc 2- (n, n-Dicyclohexylcarboxamido) ethyl t-butyl BOC 1-adamantyl 1-Adoc 2-adamantyl 2-Adoc vinyl Voc allyl Aloe or Alloc 1-isopropylalyl Ipaoc synamyl Coc 4-nitrosinamyl Night 3- (3'-pyridyl) prop-2-enyl Paloc

8-quinolyl n-hydroxypiperidinyl alkyldithio benzyl p-methoxybenzyl p-nitrobenzyl p-bromobenzyl p-chlorobenzyl 2,4-dichlorobenzyl 4-methylsulfinylbenzyl

9-Antryl methyl diphenylmethyl phenothmazinyl- (10) -carbonyl p-p-toluenesulfonylaminocarbonyl p-phenylaminothiocarbonyl

Formides amide acetamide chloracetamide trifluoroacetamide phenylacetamide 3-phenylpropanamide pent-4-enamide picolinamide

3- pyridylcarboxamide benzamide p-phenylbenzamide p-phthalimide p-tetrachlorophthalimide

4- nitro-p-phthalimide

Cbz or 3 Moz

PNB

Msz

TFA

TCP

n-2,3-diphenylmaleimide n-2,5-dimethylpyrrole n-2,5-bis (triisopropylsiloxyl) pyrrole n-1,1,4,4-tetramethyldisilyazacyclopentane adduct

1,1,3,3-Tetramethyl-1,3-disilaisoindole ind Special -NH protected groups p-methylamine n-t-butylamine p-allylamine n- [2- (trimethylsilyl) ethoxy] methylamine n-3-acetoxypropylamine

Dts

BIPSOP

STABASE

BSB

SEM p-cyanomethylamine n- (1-isopropyl-4-nitro-2-oxo-3-pyrrolin-3-yl) amine p-2,4-dimethoxybenzylamine Dmb

2-Azanorbornene n-2,4-dinitrophenylamine n-benzylamine Bn n-4-methoxybenzylamine MRM p-2,4-dimethoxybenzylamine DMMP n-2-hydroxybenzylamine Nbn p- (diphenylmethyl) aminoDPM p-bis (4-methoxyphenyl) -Dibenzosuberylamine DBS n-triphenyl methylamine Tr n - [(4-methoxyphenyl) diphenylmethyl] amino MMT g n-9-phenylflurenylamine Pf n-ferrocenylmethylamine Fem

n-2-picolylamine n'-oxide n-1,1-dimethylthiomethyleneamine n-benzylidenamine n-p-methoxybenzylidenamine n-diphenylmethyleneamine n- (5,5-dimethyl-3-oxo-1-cyclohexenyl) amine n-nitroamine n- nitrosoamine diphenylphosphinamide

Dpp dimethylthiophosphinamide

Mpt

diphenyltophosphinamide dibenzylphosphoramidate

2-nitrobenzenesulfenamide

Ppt

Nps n-1- (2,2,2-Trifluoro-1,1-diphenyl) ethylsulfenamide TDE

3-nitro-2-pyridinesulfenamide Npys p-toluenesulfonamide T3 benzenesulfonamide

Safracin B includes an alanyl side chain. In one aspect, the present invention relates to the protection of a free amino group with a Boc group, which has great advantages.

Some ecteinascidines of the present invention include compounds of formula (XVIII):

wherein R ¹ and R ⁴ form a group of formula (IV), (V), (VI) or (VII):

R ²¹ represents hydrogen, a hydroxyl group or -CN, especially a hydroxyl group or CN;

and derivatives thereof, in particular 5-acyl derivatives including 5-

acetyl derivative.

Preparation of ecteinascidin 743 and related compounds

In general, conversion of the 21-cyan starting compound to an ecteinascidin product, for example, of formula (XVIII) includes:

(a) if necessary, conversion of the quinone system of ring E into a phenolic system;

(b) if necessary, conversion of the quinone system of ring A into a phenolic system;

c) conversion of the phenolic system of ring A into the methylenedioxyphenol ring;

d) forming a bridged spiro ring system of formula (IV), (VI) or (VII) through a 1-position and a 4-position in ring B; and

e) suitable transformation such as acylation.

Step a) if necessary, the conversion of the quinone ring system E to the phenolic system can be carried out by conventional reduction methods. A suitable reagent is hydrogen with a palladium-on-carbon catalyst, but other reduction systems may be used.

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

9 9 9 9 9999999 9 99 99

Step b), if necessary, conversion of the quinone system of ring A into a phenolic system is analogous to Step a) and no further details are needed.

Step c) conversion of the phenolic system of Ring A into a methylenedioxyphenol ring can be carried out in several ways, possibly with Step b). For example, the quinone ring A may be demethylated at the 7-position methoxy substituent and reduced to dihydroquinone and treated with a suitable electrophilic reagent, such as dibromomethane, bromochloromethane or similar bivalent reagents, which directly produces the methylenedioxy ring system, or with a divalent ring system, or with a divalent such as thiocarbonyldiimidazole when producing a substituted methylenedioxy ring system which can be converted to the desired ring.

Step d) usually proceeds by appropriately replacing the 1st position with a bridging reagent that can assist in the formation of the desired bridge, obtaining the exo-quinone methide in the 4th position and allowing metmd to interact with the 1 substituent to obtain a bridging structure. Preferred bridge reagents have the formula (XIX)

where Fu represents a protected functional group such as the -NHProt ⁴³ or OProt ^4a groups, Prot ³ is a protecting group and the dotted line indicates a possible double bond.

The method is first prepared at the 10th position, at the bond between rings A and B, a hydroxyl group is inserted to form a partial structure of formula (XX):

or, preferably, a partial structure of formula (XX):

wherein the group R 'is selected for the desired group of formulas (IV), (V), (VI) or (VII). For the first two of these groups, R 'represents -CHFu-CH ₂ -SProt3. The protecting groups can then be separated and suitably modified to give the desired compound.

A typical method for Step d) is shown in US Patent 5,721,362 for comparison. In particular, the passage in column 8, step (1) and Example 33 of the patent.

The modification in Step e) may involve acylation, for example with the Ra-CO- group, as well as conversion of the I2-NCH3 group to 12-NH or 12NCH2CH3. Such conversion may be carried out before or after the other steps, by appropriate methods.

To illustrate, it is possible to transform cyanosafracin B of formula 2 into ET-743, which is a shorter and more accurate method than previously used. Cyanosafracin B can be transformed into intermediate 25 and from this compound it is possible to obtain a number of cysteine derivatives, which can be further transformed into ET-743. Preferred cysteine derivatives are shown as compounds lnt-29 and lnt-37.

η * μμΜιι ·· ι mshamn

ΙΝΤ-25

lnt-29 lnt-37

The retrosynthetic analysis for the preparation of ET-743 using compound 29 is shown in Scheme I.

nh ₂

Safracin B

Scheme I • · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

According to Scheme I above, it is possible to obtain ET743 in 21 linear steps. In this method, cyanosafracin B is transformed into intermediate 25 after a series of interactions involving mainly (1) the separation of a methoxy group present in ring A, (2) the reduction of ring A and the formation of methylenedioxy group in a single vessel, (3) hydrolysis of the amide functionality found at carbon 1, (4) converting the resulting amino group into a hydroxyl group. This method also achieves the protection and deprotection of primary alcohol functionality at position 1 of ring B of compound 25 by directly using cysteine residue 29 to afford intermediate 27. Cysteine derivative 29 is protected at the amino group by β, β , β-trichloroethoxycarbonyl protecting group to obtain compatibility with existing allyl and MOM groups. Intermediate 27 is directly oxidized and cyclized. These circumstances, as well as the different strategy of deprotection in the later stages of synthesis, make the method new and more susceptible to industrial development than the method described in US 5,721,362.

The conversion of the 2-cyan compound to intermediate 25 usually involves the following steps (see Scheme II):

preparing a protected compound of formula 14 by reacting 2 with tert-butoxycarbonyl anhydride;

conversion of the 14 to a disubstituted compound of formula 15 by reaction with bromomethyl methyl ether and diisopropylethylamine in acetone etryl;

selective elimination of the methoxy group of the quinone system in 15 to obtain a compound of formula 16 by reaction with methanol sodium hydroxide solution;

transforming 16 into a methylenedioxy compound of formula 18 in the following preferred sequence: (1) the quinone group of compound 16 is reduced by 10% palladium on carbon in a hydrogen atmosphere; (2) the hydroquinone intermediate is converted to the methylenedioxy compound of formula 17 by reaction with bromochloromethane and casein carbonate in an atmosphere of hydrogen; (3) 17 is transformed into a compound of formula 18 by protecting the free hydroxyl group as an OCH2R group. The reaction is carried out with BrCH ₂ R and casein carbonate, where R can be aryl, CH = CH 2, OR 1 and so on.

eliminating the tert-butoxycarbonyl and methyloxymethyl protecting groups of 18 to afford a compound of formula 19 by reaction with a solution of hydrogen chloride in dioxane. This reaction is also carried out by mixing 18 with a solution of trifluoroacetic acid in dichloromethane;

preparing a thiourea compound of formula 20 by reacting 19 with phenylisothiocyanate;

converting a compound of formula 20 into an amine compound of formula 21 by reaction with a solution of hydrogen chloride in dioxane;

transformation of compound 21 into N-Troc derivative 22 by reaction with trichloroethyl chloroformate and pyridine;

preparing a protected hydroxy compound of formula 23 by reacting 22 with bromomethyl ether and diisopropylethylamine;

transformation of compound 23 into the N-H derivative 24 by reaction with acetic acid and zinc;

conversion of the compound of formula 24 into hydroxy compound 25 by reaction with sodium nitrite in acetic acid. Alternatively, nitric tetroxide may be used in a mixture of acetic acid and acetonitrile, followed by treatment with sodium hydroxide. Sodium nitrite may also be used in a mixture of acetic anhydride-acetic acid, followed by treatment with sodium hydroxide.

Scheme II

Conversion of intermediate 25 to ET-743 using cysteine derivative 29 is usually carried out at the following steps (see Scheme III):

transforming a compound of formula 24 into derivative 30 by protecting the primary hydroxyl functionality with (S) N-2,2,2-trichloroethoxycarbonyl-8- (9H-fluoro-9-ylmethyl) cysteine 29;

conversion of the protected compound of formula 30 into a phenolic derivative 31 by cleavage of the allyl group with tributyltin hydride and dichloropalladium bis (triphenylphosphine);

transformation of the phenolic compound of formula 31 into a compound of formula 32 by oxidation with benzene selenic anhydride at low temperature;

transforming the hydroxyl compound of formula 32 into lactone 33 in the following sequence: (1) reacting a compound of formula 32 with 2 equivalents of triflic anhydride and 5 equivalents of dimethyl sulfoxide; (2) reacts with 8 equivalents of diisopropylethylamine; (3) reacts with 4 equivalents of t-butyl alcohol; (4) reacts with 7 equivalents of 2-1-butyl-1,1,3,3-tetramethylguanidine; (5) reacts with 10 equivalents of acetic anhydride;

transforming lactone 33 into hydroxyl compound 34 by removing the MOM protecting group with TMSI;

cleavage of the N-trichloroethoxycarbonyl group of the compound of formula 34 to compound 35 by reaction with zinc / acetic acid;

transformation of the amine compound 35 into the corresponding α-ketolactone 36 by reaction with N-methylpyridinecarboxaldehyde chloride followed by DBU;

preparing ET-770 by reacting a compound of formula 36 with 3-hydroxy-4-methoxyphenylethylamine;

transformation of ET-770 into ET-743 by reacting with silver nitrate in an AcN / water mixture.

Me

from 31

AONOj

CHjCN.H, 0

Et-743

Scheme III

The method described above for the transformation of intermediate 25 into ET-743 can be efficiently modified using other cysteine derivatives, for example, compound 37, with the name 2-methoxymethoxy-3- (9H-fluorene-9-ylmethyl) tmopropene acid. This compound already has a ketone group in the form of enol ether, while in the other cysteine analogues there is an amino group that must later be transformed into a ketone group in a 55-60% average yield transamination reaction. Therefore, a significant increase in linear fusion yield is possible using compound 37, since the transamination step is missed.

The conversion of intermediate 25 to ET-743 using a cysteine derivative 37 can be carried out in the same manner and with the same reagents unlike cysteine derivative 29 in the absence of transformations (e) and (g). The reaction sequence is presented in Scheme IV:

BujSnH. (PPhjljPdClj

AcOKCHjCIj

TMSCI Hal

Lobster. CHjCU (PhSeOfeO

CH1C> 2

1) DMSO. Tf ₂ O

2) OIPEA

3) 'βυΟΗ

5) Ac, O. CHjCIj

CHiCh. chjCn o

Et-743

Compound 38 can also be prepared by reacting intermediate 12 as described in US Patent No. 4,259,259. No. 5,721,362, with intermediate 37 in improving the method described in this patent.

Preparation of Phthalasidine and Related Compounds

According to the present invention, a class of compounds include phthalacidin and have the following general formula (XX):

OSN ₃

wherein R ¹ is an amidomethylene group; R ⁵ is a short oxy-side chain; and R ² 'is a cyano group or a hydroxyl group. For phthalacidin, R ¹ is a phthalimidomethylene group; R ⁵ is an acetoxy group; and R ² 'is a cyano group. The other groups for R ¹ include mono- and di-N-substituted amidomethylenes, as well as other cyclic amidomethylenes, and the other groups for R ⁵ include further C ₁ -C ₄ acyl groups as well as C 1-6 alkyl groups.

Conversion of the 21-cyan compound to phthalacidin or the corresponding compound of formula (XX) typically involves the following steps:

(a) if necessary, conversion of the quinone ring system E into a phenolic system;

b) forming a -R ⁵ group at the 5 position of ring A;

c) forming a R ¹ group at the 1-position of ring B; and

(d) if necessary, conversion of the quinone ring system A into a phenolic system;

(e) conversion of the phenolic ring system A into a methylenedioxy phenol ring.

• ···· ·· 9 «· • · · • e · ···» ·

• · ♦ ♦ · ·· · · • 9 9 · • · • • • • e • · • • • · 9 • • ··· ···· ·· ···

These steps are closely similar to the steps shown for the preparation of ecteinascidins. Step c) typically involves the formation of the group -CH2NH2 in the first position and its acylation.

phthalacidin can be prepared using the intermediates described for the conversion of cyanosafracin B to intermediate 25. For example, intermediates 21 and 17 are suitable starting materials for the preparation of phthalacidin.

As shown below in Scheme V, the method for the synthetic preparation of phthalacidin from intermediate 21 involves the following steps:

transformation of 21 into a compound of formula 27 by reaction with phthalic anhydride in dichloromethane and carbonyl diimide sol;

conversion of 27 to phthalasidine by reaction with tributyltin hydride and dichloropalladium-bis (triphenylphosphine) or basic medium followed by reaction with acetyl chloride.

Scheme V

As shown in Scheme VI, the method for the synthetic preparation of phthalacidin from intermediate 17 includes the following steps: acetylation of the hydroxyl group of compound of formula 17 with acetalchloride and pyridine to give an acetylated intermediate of formula 42;

separation of the t-butoxycarbonyl and methoxymethyl protecting groups 42 to afford a compound of formula 43 by reaction with a solution of hydrogen chloride in dioxane. This reaction can also be carried out by mixing 42 with a solution of fluoroacetic acid in dichloromethane;

preparing a thiourea compound of formula 44 by reacting 43 with phenylisothiocyanate;

converting the compound of formula 44 into an amine compound of formula 45 by reaction with a solution of hydrogen chloride in dioxane;

transformation of 45 into phthalicidin by reaction with phthalic anhydride in dichloromethane and carbonyl diimidazole.

phthalicidin

Scheme VI

Preparation of intermediate 11 and the corresponding intermediates

The retrosynthetic assay is described with the following sequential-

Safracin B

In the present invention, a key class of intermediates include intermediate 11 and have the general formula (XXI): OCH ₃

PtO ^^^ x ^ CH3

OPrP where Prot ¹ and Prot ² are hydroxyl protecting groups, preferably different. Typically, Prot ¹ is selected from [more generalisation needed]. Typically, Prot ² is selected from [more generalisation needed]. For intermediate 11 itself, the Prot ¹ group is a methoxymethyl group and Prot ² is a t-butyldiphenylsilyl group.

Conversion of the 21-cyan compound to intermediate 11 or the corresponding intermediate of formula (XXI) typically involves the following steps:

(a) if necessary, conversion of the quinone ring system E into a phenolic system;

b) forming -OProt ¹ group at the 18th position of ring E;

«· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

c) formation of a CH2-OProt ² group in the 1st position of ring B; and

(d) if necessary, conversion of the quinone ring system A into a phenolic system;

(e) conversion of the phenolic ring system A into a methylenedioxyphenol ring.

Step (b), formation of the -OProt ¹ group at the 18th position of ring E, is a simple reaction of protecting a phenolic group and does not need to be explained. Appropriate conditions are chosen depending on the nature of the protecting group. The other steps are the same as the other reactions.

Step (c), formation of the CH2-OProt ² group in the 1st position of ring B, is usually carried out first producing the -CH ₂ NH _{2 group} in the first position and then converting the amino group into a hydroxyl group and protection. Therefore, when the starting compound has a group R ¹ which is -CH ₂ NH-CO-CR ^25a R ^2a, R ^25c , then the N-acyl group must be separated. When the parent compound has a group R ¹ which is -CH 2 -O-CO-R, then no changes in ecteinascidin are required, where the substituent R ¹ is the same. For other products, the O-acyl group must be separated. This deacylation can be accomplished by various methods. In one, the deacylation and conversion to hydroxyl functionality are carried out in one step. The hydroxyl group may then be acylated or otherwise modified to give the appropriate group R ¹ .

In U.S. Pat. No. 5,721,362 describes synthetic methods for the preparation of ET-743 by long, multi-step synthesis. One of the intermediates in this synthesis is intermediate 11. Using cyanosafracin B as a starting material, it is possible to obtain intermediate 11 in a much shorter way to obtain such an intermediate and, therefore, to improve the process for preparing ET-743.

• · · · * * * · · • • · · · · · · • • • • • • • • • • • • • • • • · · · · · · ·

Cyanosafracin B can be converted to intermediate 25 by the methods described above. From intermediate 25 it is possible to obtain intermediate 11 using the following steps, see Scheme VII:

preparing a protected hydroxyl compound of formula 26 by reacting 25 with tert-butyldiphenylsilyl chloride in the presence of a base;

final cleavage of the allyl group with tributyl tin hydride and dichloropalladium-bis (triphenylphosphine) at 26, resulting in intermediate 11.

BujSnH

Scheme VII

One aspect of the synthetic method of the present invention for transforming safracin B into intermediate 11 is modifying and extending Scheme VIII and considering the following steps:

stereospecific conversion of the compound of formula 1 (safracin B) into the compound of formula 2 by selective substitution of a hydroxyl group with a cyano group by reaction with potassium cyanide in an acidic medium;

Preparation of the tmocarbamide compound of formula 3 by reaction of the compound of formula 2 with phenylisothmocyanate ;

conversion of the thymocarbamide compound of formula 3 into acetamide of formula 5 under hydrolysis in acidic medium followed by addition of acetic anhydride. The intermediate amine of formula 4 can be isolated by fixing the hydrolysis in acidic sodium bicarbonate medium, but this intermediate is very unstable and quickly turns into a five membered cyclic imine, called compound 6;

preparing a protected compound of formula 7 by reaction with bromomethylmethyl ether and diisopropylethylamine in dichloromethane;

selective demethylation of the methoxy group of the quinone system of the compound of formula 7 in a compound of formula 8 by reaction with methanol sodium hydroxide solution;

converting the compound of formula 8 into the methylenedioxy compound of formula 9 according to the following preferred steps: (1) the quinone group of compound 8 is reduced by 10% palladium on carbon in a hydrogen atmosphere; (2) the hydroquinone intermediate is converted to the methylenedioxy compound of formula 9 by reaction with bromochloromethane and cesium carbonate in an atmosphere of hydrogen; (3) the compound of formula 9 is transformed into a compound of formula 10 by protecting the free hydroxyl group as an OCH ₂ R group. The reaction is carried out with BrCCH2L and cesium carbonate, where R may be aryl, CH = CH2, OR ', etc .;

converting the acetamide group of the compound of formula 10 into the corresponding hydroxyl group of the compound of formula 11 by reaction with nitric tetraoxide in a mixture of acetic acid and acetyl acetate, followed by treatment with sodium hydroxide;

or alternatively, sodium nitrite may be used in a mixture of acetic anhydride and acetic acid, followed by treatment with sodium hydroxide;

Wirtlir

or alternatively, the acetamide group of the compound of formula 10 may be converted to a primary amino group by reaction with hydrazine or with Boc2O, DMAP followed by dehydrazine;

such primary amine may be converted to the corresponding hydroxyl group (compound of formula 11) upon the oxidative conversion of the primary amine to the corresponding aldehyde with 4-formyl-1-methylpyridinium benzenesulfonate or other pyridinium ion, followed by treatment with DBU or other base and thereafter hydrolyzed and then reducing the aldehyde to the corresponding hydroxyl group with lithium aluminum hydride or other reducing agent;

preparing a protected compound of formula 26 by reaction with t-butyldiphenylsilyl chloride and dimethylaminopyridine in dichloromethane;

transformation of the silylated compound of formula 26 into intermediate 11 by deprotecting the OCH2R protecting group by reacting under reducing or acidic conditions.

Palladium black is usually used in an atmosphere of hydrogen or aqueous trifluoroacetic acid, or tributyl alkaline hydride and dichlorobis (griphenylphosphine palladium).

In another preferred modification, the cyan compound of formula 2 can be transformed into intermediate 11 using Extended Scheme II, comprising the steps of:

preparing a protected hydroxyl compound of formula 26 by reacting 25 with tert-butyldiphenylsilyl chloride in the presence of a base;

final cleavage of the allyl group with tributyl alkaline hydride and dichloropalladium bis (triphenylphosphine) at 26, resulting in intermediate 11.

• · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

Preparation of the active compounds

It is possible to transform cyanosafracin B into a number of intermediates and derivatives with potential antitumor therapeutic activity. These intermediates can be prepared from compounds already described or using alternative methods.

The intermediates described in the present invention include compound 47 and a number of amide derivatives obtained using compounds 45 or 43.

Scheme VIII illustrates the preparation of compound 47 using the following sequence:

preparing the thiourea compound of formula 3 by reacting the compound of formula 2 with phenylisothiocyanate;

conversion of the thiourea compound of formula 3 into acetamide of formula 5 under hydrolysis in an acidic medium followed by addition of acetic anhydride. The intermediate amine of formula 4 can be isolated by fixing the hydrolysis in acidic sodium bicarbonate medium, but this intermediate is very unstable and quickly turns into a five membered cyclic imine, called compound 6;

preparing a protected compound of formula 7 by reaction with bromomethylmethyl ether and diisopropylethylamine in dichloromethane;

selective demethylation of the methoxy group of the quinone system of the compound of formula 7 in a compound of formula 8 by reaction with methanol sodium hydroxide solution;

converting the compound of formula 8 into the methylenedioxy compound of formula 9 according to the following preferred steps: (1) the quinone group of compound 8 is reduced by 10% palladium on carbon in a hydrogen atmosphere; (2) the hydroquinone intermediate is converted to the methylenedioxy compound of formula 9 by reaction with bromochloromethane and cesium carbonate in an atmosphere of hydrogen; (3) the compound of formula 9 is transformed into a compound of formula 10 through 46 by protecting the free hydroxyl group as an allyloxy group by reaction with allyl bromide and cesium carbonate;

transforming the compound of formula 9 into acetyl derivative 46 by reaction with acetyl chloride in pyridine;

transforming the compound of formula 46 into an unprotected compound

dynoxy 47 by reaction with hydrochloric acid in dioxane.

Other useful derivatives of the amide intermediates are obtained from the already described intermediate 45 using the following scheme. The second stage is possible. The method is an important part of the present invention, especially when the group R is R ^a , as defined above. Moreover, Scheme VIII can be easily extended to show the preparation of the compounds of formula (XXIII) upon introduction of the starting material with a different group at the 5 position, or

a group that directly remains in the product, or a group that can be separated or modified to obtain the desired group.

RCOCI. ru

OMe

Scheme IX

From compound 45, a group of analogues can be obtained according to the following scheme:

acylation of the amino group of the compound of formula 45 with a large number of acyl derivatives to give the corresponding amides, the preferred acyl groups being acetyl, syamoyl chloride, p-trifluorosinamoyl chloride, isovaleryl chloride phenylisothiocyanate, or trans-aminoidamide groups, or aminylamines, or amine groups of the cyano group in the hydroxyl group by reacting with silver nitrate in an AcN / water mixture.

Other useful derivatives of amide intermediates are obtained from the already described intermediate 43 using the following scheme:

AgNO ₃

OMe

Me Oh

Scheme X

From compound 43, another group of interesting derivatives can be obtained using the following scheme:

(a) acylation of the amino group of a compound of formula 43 with a large number of acyl derivatives to afford the corresponding amides, with the preferred acyl groups being acetyl, synamoyl chloride, p-trifluorosinamoyl chloride, isovaleryl chloride or amino acids or the above acid groups or CO-dimers ;

(b) transformation of the cyano group into a hydroxyl group by reaction with silver nitrate in an AcN / water mixture.

New intermediates

In the light of the above explanations, it can be seen that the present invention relates to novel intermediates. Depending on Ring A, the intermediates are of formula (XXPa):

or with the formula (XXX):

where:

R ¹ is -CH ₂ NH ₂ or CH ₂ OH or a protected or derivative of this group and R ⁴ is -H; or

R ^1a and R ⁴ together form a group of formula (IV), (VI) or (VII):

R ⁵ is a hydroxyl group or a protected or derivative variant thereof;

both R ^14a and R ^14b are hydrogen or one is hydrogen and the other is hydrogen or a protected or derivative of this group, -OCH ₃ or -OCH ₂ CH ₃ , or both R ^14a and R ^14b together form a ketone group;

R ¹² is -NH-, -NCH ₃ - or -NCH ₂ CH ₃ -;

R ¹⁵ is a hydroxyl group or a protected or derivative thereof; and

R ¹⁸ is a hydroxyl group or a protected or derivative variant thereof.

In one aspect of the present invention, at least one of the groups R ', R ⁵ , R' ⁴ *, R ^{, 4b} , R ^{15 and 11} R ¹⁸ fla is protected or a derivative group is preferred.

In one aspect of the present invention, the R ¹ group is not a 3,5-t-6yldiphenylsilyl substituent and / or the R ¹ group is not a methoxymethyl group.

Preferably R ¹ is -CH 2 NH 2 or CH 2 OH or a protected or derivative of this group and R ⁴ is -H; or R ^1a and R ⁴ together form the group:

One preferred class of intermediates according to the present invention includes the compound we identified as compound 25 of the formula:

Therefore, the preferred class is of the general formula in which the IOM group is replaced by some other protecting group.

Other preferred intermediates according to the present invention include the compound we identified as compound 45 and 47. Other N-acyl derivatives can be readily prepared from compound 45 and are an important part of the present invention. Suitable acyl groups include the acyl groups described above. According to51 • ···· ·· · • · • · <

The 9 21-hydroxyl compound is also useful and is one of the active compounds we have obtained.

New active compounds

We have also found that some of the compounds of the present invention, which were originally obtained as intermediates, have exceptional activity in the treatment of cancer, such as leukemias, lung cancer, colon cancer, kidney cancer and melanoma.

Accordingly, the present invention relates to a method of treating mammals, in particular, to a human suffering from cancer, comprising administering to a diseased individual a therapeutically effective amount of a compound of the present invention, or a pharmaceutical composition, that contains it.

The present invention also relates to pharmaceutical compositions containing as active ingredient a compound or compounds of the present invention, and methods for their preparation.

Examples of pharmaceutical compositions include solids in the form of tablets, pills, capsules, granules, etc., or a liquid (solutions, suspensions or emulsions) of a suitable composition for oral, topical or parenteral administration, which may contain pure a compound or a mixture thereof with a carrier or other pharmacologically active compounds. These formulations need to be sterile when administered parenterally.

The administration of the compounds or compositions of the present invention can be accomplished by any suitable method such as intravenous infusion, oral compositions, intraperitoneal and intravenous administration. The infusion time is preferably up to 24 hours, more preferably 2 to 12 hours, and most preferably 2

• ···· • · · · · • · • · • · · · • · • · • • · · • · · • • · · · • · · • • · · • · · · • ·

up to 6 hours. Short infusion times that allow treatment to be carried out without a hospital stay are particularly desirable. However, if necessary, the infusion may last from 12 to 24 hours or longer. The infusion can be given at appropriate intervals of 2 to 4 weeks. pharmaceutical compositions containing compounds of the present invention can be delivered by liposomal or nanospheric encapsulation, in restraining compositions or by other standard means.

The dose of the compounds will vary depending on the individual composition, the route of administration and the individual situs, host, and tumor to be treated. Other factors such as age, body weight, gender, diet, administration time, rate of discharge, patient's condition, drug combination, response sensitivity, and severity of the disease are also considered. Administration may be carried out continuously or periodically with the maximum tolerated dose.

The compounds and compositions of the present invention may be used in other medicines for combination therapy. The other drugs may be part of the same composition or administered as a separate composition at the same time or at another time. The nature of the other medicine is not limited, with suitable ones including:

(a) antimitotic drugs, especially those intended for cytoskeletal elements, including microtubule modulators, such as taxane drugs (such as taxol, paclitaxel, taxotere, dosetaxel), podophyllotoxins or vinca alkaloids (vincristine, vinblastine);

b) antimetabolite drugs such as 5-fluorouracil, cytarabine, gemcitabine, purine analogues (such as pentostatin, methotrexate);

c) alkylating agents such as nitrogen mustards (such as cyclophosphamide or ifosfamide);

• · · · · · · · · · · · · · · · · · · · · · · · · · · · · 9 · 9 9 9 99

9 9 9 9 9 99 • 99 99 999 9999 99999

(d) drugs intended for deoxyribonucleic acid, such as anthracyclines, adriamycin, doxorubicin, pharmorubicin or epirubicin;

e) drugs intended for topoisomerases, such as etoposide;

f) hormones and hormone agonists and antagonists such as estrogens, antiestrogens (tamoxifen and related compounds) and androgens, flutamide, leuprorelin, goserelin, cyprotron or octreotmd;

(g) medicines intended for signal transduction in tumor cells, including antibody derivatives such as herceptin;

h) alkylating agents such as platinum drugs (cis-platinum, carboplatin, oxaliplatin, paraplatin) or nitrosocarbamides;

i) drugs that potentially act on tumor metastases, such as matrix metalloproteinase inhibitors;

(j) gene therapy agents;

(k) antibody therapists;

(l) other bioactive compounds of marine origin, in particular didemnin, such as aaplidine;

m) steroid analogues, in particular dexamethasone;

n) anti-inflammatory agents, in particular dexamethasone; and

o) vomiting medicines, in particular dexamethasone.

The present invention also relates to the use of the compounds of the present invention in the method of treatment and to the use of the compounds in the preparation of a pharmaceutical composition for the treatment of cancer.

Cytotoxic activity

Cell cultures. The cells were kept in logarithmic growth phase in Eagle's Minimum Essential Medium, with Earle's Balanced Salts, with 2.0 mM L-glutamine, with basic amino acids, without sodium bicarbonate

• ···· • ·· ·· • · ·· · · • · • · • · • · • · • · · • · • · · • · · • · • · • · · · ··· ···· • · • ·

(EMEM / neaa); supplemented with 10% fetal calf serum (FCS), ΙΟ ' ² Μ sodium bicarbonate and 0.1 g / l penicillin-G + streptomycin sulfate.

A simple screening procedure was performed to determine and compare the antitumor activity of these compounds using the adapted form of the method described by Bergeron et al (1984). Tumor cells used were P-388 (suspension culture of DBA / 2 mouse lymphoma neoplasm), A-549 (human lung carcinoma monolayer), HT-29 (human colon carcinoma monolayer) and MEL-28 (monolayer). human melanoma culture).

P-388 cells were seeded in 16 mm wells in an amount of 1x10 * well cells in 1 ml aliquots of MEM 5FCS containing the drug at the indicated concentration. Non-drug cultures are planted in a separate set to monitor control growth and to determine whether cells remain in the exponential phase of growth. All determinations are made twice. After three days of incubation at 37 ° C, under an atmosphere of 10% carbon dioxide in 98% moisture, the approximate IC50 was determined to compare growth in drug wells with that in control wells.

A-544, HT-29, and MEL-28 cells were seeded in 16 mm wells in an amount of 1x10 * well cells in 1 ml aliquots of MEM 5FCS containing the drug at the indicated concentration. Non-drug cultures are planted in a separate set to monitor control growth and to determine whether cells remain in the exponential phase of growth. All determinations are made twice. After three days of incubation at 37 ° C, under a atmosphere of 10% carbon dioxide in 98% moisture, the wells were stained with 0.1% Crystal Violet. The approximate IC50 was determined to compare growth in drug wells with that in control wells.

1. Raymond J. Bergeron, Paul F. Cavanaugh, Jr., Steven J. Kline, Robert G. Hughes, Jr., Gary T. Elliot, Carl W. Porter. Antineoplastic and antiherpetc

• ···· • ·· • · • · ·· · · • · • · • · • · • · • · · • · • · · • · · • · • · is · ·· ··· ···· ·· • ·

activity of spermidine catecholamide iron chelators. Biochem. Bioph. Really. Comm. 1984, 121 (31848-854.

2. Alan C. Schroeder, Robert G. Hughes, Jr., Alexander Bloch. Effects of Acyclic Pyrimidine Nucleoside Analoges. J. Med. Chem. 1981, 24, 1078-1083.

It can be seen from the data on cytotoxic activity and other perceptions that the active compounds of the present invention include a preferred class of compounds of general formula (XXIII):

where:

R ¹ is as defined for formula (XVIIb) and is preferably a medium-sized aminomethyl group;

R ⁵ is as defined for formula (XVIIb) and is preferably a small size hydroxyl group;

R ¹² is as defined above and is preferably -NCH- and R ²¹ is hydroxyl or cyano.

R ¹ is a suitable hydrophobic group and not free amino, hydroxyl or other hydrophilic functionality. Typically, R ¹ is the group -CHrNH ₂ -CO-R ^a in which R ^a is as defined, but preferably has a linear chain of less than 20 atoms, more preferably less than 15, or 10 atoms, with 1,4-phenyl being counted as a chain with a length of 4 atoms, and this applies to other cyclic groups (for example, 1,2-cyclohexyl is counted as a chain with a length of 2 atoms), and the linear chain from - less than 10,15 or 20 atoms can be substituted. In particular, the data show that a balance is achieved between the non-existent

MKMWAY

IC 50 (pM) P-388 A-549 HT-29 MEL-28 CV-1 DU-145 ohm · ^ m · |) || mH-m · m «o '^ pV' ^N 's ^z ^ ΟΙ & N nor ₂ 0.009 * 0.018 0.018 0.018 0.023 OM * ko ^ jl ^ m · m · a || χ · Ι-Μ · ⁰ AM M 14 0.15 > 0.15 0.15 > 0.15 1 OMe ° ^ ° ^ y ^and · Me. Αχ>. Lg ^ ϊι itl ^z ^ ^N j ' ^M * “ΎΪ - ⁰ AM 15 1.44 1.44 1.44 1.44 1 ohm · Ο ^ ΟγΛγ “· m · ,. JL> ^ 7 τ ** ^ 0 k bi 0. ι ⁰ A 16 > 1.5 > 1.5 > 1.5 > 1.5 1? "· Γ η 1.4 1.4 1.4 1.4 \ ί? "· Οχ ^ Οη ^ γ ** ° ^ A.m- • In the 18th 0.01 0.01 0.01 0.01 oh · M ·. Jl ^ z- ^> r ^ r Υ ']! H '* W 'V' 19 0.08 0.16 0.01 0.16

I

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ······ ·· ···

ι “* Οη ^ Ύ“ · WyP ^V ° l ^ CN _e A> »« c »M4> ⁰ τ 20 0.01 0.01 0.01 0.01 1 “· ^ -0 L Sn a * hh, 21 0.019 0.019 0.019 i 0.019 4.? '· 1 HQ ^ Js ^ Me Me. xL xs ^ M- | -Me HH ₀ Λ ₀ .ον: ο, 22 0.014 0.014 0.014 0.014 0.014 0.014 OMe H ^ ^o ^ ^o uf ' ^w Me V η | ^N 2l '' · V-0 X ^ CN ₀ L ₀ .ss 23 0.13 0.13 0.13 0.13 0.13 0.13 % ^0Me Me xL ^hh . N- | - Me ^ -0 I CN «<> l 0.18 1.8 1.8 1.8 1.8 1.8 h, 1? "· Me N- | -Me ° ^Vo M 25 0.2 0.2 0.2 0.2 0.2 H4 « j. 9 “* OX''ή * o \ s Τ 'J UK .WZ Z <Y XXkiJ> “* ° Co b, 35 0.008 0.008 0.008 0.008 o 1 ° “· HO ^ Jy ^ Me * ° \ i ΤΎ Me ^ ll / y ^ TjjPk * jJJr “* ^ -0 CN 36 0.01 0.01 0.01 0.01 OMe -Me Oh ^ -0 J Sn UG w 28 0.001 0.001 0.001 0.001 0.001 0.001 1? "· 0 ___ O ^ JL ^ Me oh Τ J Me« xL x- <. ^v f ^Z q ^{z <X} J ^ ^N 4 “ ¹ * · '42 0.13 0.13 0.13 0.13 0.13

OM · OAc I J 43 0.008 0.016 0.008 0.008 0.016 Ohm · HO ^ A ^ Μ »ο ** 2l J V-Q h Ϊν MH ° \ 44 0.001 0.001 0.001 0.001 0.001 0Μ »HO ^ J ^ IM OAC Tj and ^ ί ^ ΊΓ ** ^“ ^{|, and} Vo £ n as 0.01 0.01 0.01 0.01 0.01 OM · ΗΟ ^ ^, Μ · 0 1 GN * M * 3 0.015 0.015 0.015 0.015 0.018 CM · 2.171 2.171 2.171 2.171 2.171 Ohm · MO ^^ L, · »· * ^ * 'Y ^ t ***'! ** ^ **! m · IMO If ^N, ^ < ⁰ <A 5 0.005 0.005 0.005 0.005 1 ® * 0 ^ 0 ^ A. M · 'Fez ItoO ^ fY> T » <A 7 0.22 0.22 0.22 0.22 0.22 *? "· O ^ O ^ Js ^ M · Mu .Jk. Υττ ^ ίΗΜί · χ, ΧΧ, Ν ^ Χ ⁰ oA 8 > 9 > 18.1 > 18.1 > 18.1 > 18.1

't ° 4z% ZV “· He 1 J m ·. ± · »-» · * · ^Vo _O A 9 > 1.77 > 1.77 > 1.77 > 1.77 > 1.77 \ i _v <4A—. W ~ ΜΗ <A 10 > 1.65 > 1.65 > 1.65 > 1.65 > 1.65 <m + y "· * sA <vci. <ηχ «· <46 0.016 0.016 0.016 0.016 0.016 Ohm · MO ^^ ush О * с Γ Т -yV'sX / “· <A 47 0.001 0.001 0.001 0.001 0.001 oh · HO ^ Jy ^ m · ols' J M- JL ^ <- 4. Υττ n-1-w oVTs ^ ^ -o to £ n OAL 48 0.0008 0.001 0.0008 0.0008 0.001 OAe G himself, j M · ^ ^ -s. ΓιΓΤ * * οΑ \ Υ ^ \ -0 X CN MM ° ^L ! ° * 49 0.007 0.007 0.007 0.007 0.007 ohm · ojv J “• ^ ΑγΖ ^ ΧΧ. oVy * ^ \ —0 ”^ О ₅ 0 0.0001 0.0001 0.0001 0.0001 0.0001 Osh oas X j ΊγΎίγ * 7 · * о ^ к ^ и · '' About CN IM yLy “· 51 0.0001 0.0001 0.0001 0.0001 0.0001 Osh MO ^ J ^ W OAC J2 J M ·. J <^ «h. ιΓΊι * ** ** · M / V-0 X CN NM o + y- 0.001 0.001 0.001 0.001 0.001

oh · HO ^ Js ^ M · OAC j AV / —0 \ CN μ »o 53 0.0001 0.0001 0.0001 0.0001 0.0001 ' • hours ° “· hsb ^ - ^ -0 \ CN №4 oA ^ nh w, I 0 54 0.001 0.001 0.001 0.001 0.001 OM · oi 2Σ J *** oW / \ -0 \ CN NM ο <γ * γ ^ μ · o 55 0.01 0.01 0.01 0.01 0.01 OM · HO ^ k ^ M · ° M j M · ^ L · ^ * 4. Uyy * "!" "· '-Ό L in • in 56 0.18 0.9 0.18 0.8 0.9 Ohm · hO ^^ L ^ m · cm j2 J M · ^ A. .χ-κ. ΊΓ u ^ i ⁴ ! d * ⁴ * · o L £ n Ml ^ X ^ NMCSM ** 57 0.14 0.14 0.14 0.14 0.14 -X ^ '\ -ok δΝ nor ⁰ ' “58 0.001 0.001 0.001 0.001 0.001 oh · HO ^ Js ^ -M · OAC J ^^ ίΤΤ ^ Η-ϊΜ · οΛγΛγΝγ ^ IM 0 and 60 0.001 0.001 0.0005 0.001 0.0005 OM · - ΊίΓ ΛΥΖ '-o 1 CN A * 61 0.001 0.001 0.001 0.001 0.001 OM · HO ^ J ^ M · OAC TG Jt oS \ ^H · ^ 4-0 0N NN _u ^ 1 ^ 62 0.001 0.00 i 0.0005 0.0005 0.001

Yeeeeee

· · · · · · · · · · · · · · · · · · · · · · 9 9 9 ··

UDF ί 7μΤ * 7 * o'VS'V ** '—Ο * CN NK -with 63 0.0001 • · · · · 0.0001 0.0001 • · · · · 0.0001 0.0001 OAC J j) ΪΙ4Ι7 * '-O * CN. * »Η a ^a w ° 64 0.001 0.001 0.001 0.001 0.001 OAC jZ j Π ' '-o (ch 0 ^ 65 0.0001 0.0005 0.0001 0.0001 0.0005 OAC JZ T JL Pi? " V-0 (0H ιΛ inuAAC, 0 66 0.0001 0.0001 0.0001 0.0001 0.0001 OAC JZ T v 0 V on o ^< S * ^NH Y ° ^! '' · 0 67 0.0001 0.0001 0.0001 0.0001 0.0001

the R'-CO group and having a large group.

In one aspect of the present invention, it is preferred that in R 'there are no cyclic groups, especially aromatic groups. In the same sense, the present invention does not refer to the preparation of the compounds described in Proc. Natl. Acad. Sci. USA, 96,3496-3501,1999 given for comparison. Our preferred groups for R 'exclude the corresponding substituents CH ₂ R ₂ shown in Table 1 of the above message, especially groups A, B, C and D for R2.

R ⁵ is preferably an acetyl group.

In the particularly preferred compounds of the present invention, the R ¹ group is acylated on the -NH ₂ group, for example, N-acyl derivatives can be obtained from the groups -CH ₂ NH ₂ and -CH ₂ NH-aa.

The acyl derivatives may be N-acyl or N-thioacyl

• · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ··· ········ Derivatives. The acyl groups may be of formula -C0-R ³ in which R ^a is as defined above and is selected to satisfy the criteria shown. Suitable acyl groups include alanyl, arginyl, aspartyl, asparagyl, cystyl, glutamyl, glutaminyl, glycyl, histidyl, hydroxyprolyl, isoleucyl, leucyl, lysyl, methionyl, phenylalanyl, prolyl, seryl, threyl, threyl, threyl, threyl, threyl, threyl, threyl, threyl, threyl, threyl, threyl, threyl other acyl groups of amino acids. Such acyl groups of amino acids are preferably formed on the amino group to give hydrophobicity.

In one aspect of the present invention, the R ¹ group is a hydroxymethylene group derivative. It may also be a derivative of an aminomethylene group.

An important method of the present invention in which the active compounds are prepared is the following:

where R ⁵ for the final product is as defined for compound (XXXII) and may be different in the starting material and altered during the process,

R ¹⁸ is a hydroxyl group in the final product but may be a protected hydroxyl group in the starting material and altered during the process, R ¹² in the finished product may be the same as in the starting material or altered in the process,

R ¹² in the final product is as defined above, and if it is a hydroxyl group, is obtained from a cyano group during the process,

R ^a is as defined above and can then be acylated as a step of the process to obtain the end product with an acylated R ^a group.

It is preferred that in the starting material R ⁵ Aa is acetyl or another small acyl group and does not change during the course of the reaction. It is preferred that in the starting material R ¹⁸ is a hydroxyl group and does not change during the course of the reaction. Preferably, in the starting material R ^{, 2} fla is -NCHy and does not change during the course of the reaction. In the final product, R ²¹ is as defined above and, if a hydroxyl group, is obtained from a cyano group during the process. Preferably, in the end product R ^a is as defined for the compound of formula (XXIII).

Another important method of the present invention includes

interaction:

Another important method of the present invention involves the interaction of:

Another important method of the present invention involves an interaction wherein the R ¹ group is aminomethylene and converted to a hydroxymethylene group.

Another important method of the present invention involves reacting wherein the compound of group R ¹ which is hydroxymethylene is reacted with a reagent of formula (XIX):

ft ···· · ·· «« · ·· · · · · * ···· • ft · · · ·· • ·· ············· · · · · · ··· ········ where Fu is a protected functional group, Prot ³ is a protecting group and the dotted line indicates a possible double bond.

Another important method of the present invention involves reacting to obtain a 21-cyan compound of formula (XVI), characterized in that it involves reacting a compound of formula (XV):

in which the groups R ¹ , R ⁵ , R ⁸ , R ¹ **, R ¹⁴ **, R ¹⁵ h R ¹⁸ are as defined above and R ²¹ is a hydroxyl group, with a cyanide ion source, to give the desired 21-cyan compound.

In addition, methods employing other nucleophil-containing compounds to obtain similar compounds of formula (XVI) in which position 21 is protected by another nucleophilic group, the 21-NuC group, are also contemplated. For example, a 21-Nu compound of formula (XVI) with an alkylamino substituent at the 21-position can be prepared by reacting a compound of formula (XV) in which R ²¹ is a hydroxyl group, with a suitable alkylamine. The 21-Nuc compound of formula (XVI) with an alkylthio substituent at the 21-position can also be prepared by reacting the compound of formula (XV), in which R ²¹ is a hydroxyl group, with a suitable alkanethiol. Alternatively, the 21-Nuc compound of formula (XVI) with the α-carbonylalkyl substituent at the 21-position can be prepared by reacting the compound of formula (XV) in which R ²¹ is a hydroxyl group, with a suitable carbonyl compound, usually in the presence of a base. Other synthetic methods are known for other 21-Nuc compounds.

Another important reaction of the present invention includes the use of the present invention. Treatment of the 21-cyan compound according to the present invention to obtain a 21-hydroxyl compound. These compounds have interesting in vivo properties.

Examples of carrying out the invention

Me

(Boss ^ Oh

EtOH, 7h. 23 ^e C

14

To a solution of 2 (21.53 g, 39.17 ml) in ethanol (200 ml) was added tert-butoxycarbonyl anhydride (7.7 g, 35.25 ml) and the mixture was stirred for 7 hours at 23 ° C. The reaction mixture was then concentrated in vacuo and the residue purified by flash chromatography (silica gel, hexane: ethyl acetate 6: 4) to give 14 (20.6 g, 81%) as a yellow solid.

Rf: 0.52 (ethyl acetate chloroform 5: 2).

1 H NMR (300 MHz, CDCl 3): δ 6.49 (s, 1H), 6.32 (bs, 1H), 5.26 (bs, 1H), 4.60 (be, 1H), 4.14 (d, J = 2.4 Ηζ, ΙΗ) , 4.05 (d, J = 2.4 Ηζ, ΙΗ), 3.94 (s, 3H), 3.81 (d, J = 4.8 Hz, 1H), 3.7 (s, 3H), 3.34 (brd, J = 7.2 Hz, 1H) , 3.18-3.00 (m, 5H), 2.44 (d, J = 18.3 Hz, 1H), 2.29 (S, 3H), 2.24 (S, 3H), 1.82 (S, 3H), 1.80-1.65 (m, 1H) ), 1.48 (S, 9H), 0.86 (d, J = 5.7 HZ.3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 185.5, 180.8, 172.7, 155.9,154.5, 147.3, 143.3, 141.5,

135.3, 130.4, 129.2, 127.5, 120.2, 117.4, 116.9, 80.2, 60.7, 60.3, 58.5, 55.9, 55.8,

54.9, 54.4, 50.0, 41.6, 40.3, 28.0, 25.3, 24.0, 18.1, 15.5, 8.5.

ESI-MS m / z: Calcd. For C34H43N5O8: 649.7. Found (M + H) ⁺ : 650.3.

15

With stirring to a solution of 14 (20.6 g, 31.75 ml) in CHsCN (159 ml), diisopropylethylamine (82.96 ml, 476.2 ml), methoxymethylene bromide (25.9 ml, 317.5 ml) and dimethylaminopyridine (155 mg, 1.27) were added at 0 ° C. ml). The mixture was stirred at 23 ° C for 24 hours. The reaction was fixed at 0 ° C with aqueous 0.1N hydrochloric acid (750 ml) (pH = 5) and extracted with methylene chloride (2x400 ml). The organic phase was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography (silica gel, gradient of hexane: ethyl acetate 4: 1 doxane: ethyl acetate 3: 2) to give 15 (17.6 g, 83%) as a yellow solid.

Rf: 0.38 (hexane: ethyl acetate 3: 7).

Ή NMR (300 MHz, CDCl3): δ 6.73 (s, 1H), 5.35 (bs, 1H), 5.13 (s, 2H), 4.40 (bs, 1H), 4.25 (d, J = 2.7 Hz, 1H). 4.03 (d, J = 2.7 Ηζ, ΙΗ), 3.97 (s, 3H), 3.84 (bs, 1H), 3.82-3.65 (m, 1H), 3.69 (s, 3H), 3.56 (s, 3H), 3.39 -3.37 (m, 1H), 3.20-3.00 (m, 5H), 2.46 (d, J = 18 Hz, 1H), 2.33 (s, 3H), 2.23 (s, 3H), 1.85 (s, 3H). 1.73-1.63 (m, 1H), 1.29 (s, 9H), 0.93 (d, J = 5.1 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 185.4, 180.9, 172.4, 155.9,154.5, 149.0, 148.4, 141.6,

135.1, 131.0,129.9, 127.6,124.4, 123.7,117.3, 99.1, 79.3, 60.7, 59.7, 58.4, 57.5,

56.2, 55.9, 55.0, 54.2, 50.0, 41.5, 39.9, 28.0, 25.2, 24.0, 18.1, 15.6, 8.5.

ESI-MS m / z: Calculated for C31H26N3O: 693.8. Found (M + H) *: 694.3.

Example 3

To a flask containing 15 (8 g, 1.7 ml) in methanol (1.61), at 0 ° C, was added 1M aqueous sodium hydroxide solution (3.21). The reaction mixture was stirred for 2 hours at this temperature and then fixed with 6M hydrochloric acid to pH 5. The mixture was extracted with ethyl acetate (3x1 I) and the combined organic layers were dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography (silica gel, chloroform gradient: ethyl acetate 2: 1) to give 16 (5.3 g, 68%).

Rf: 0.48 (CHaCNiBOfla 7: 3, RP-C18).

¹ H NMR (300 MHz, CDCl 3): δ 6.73 (s, 1H), 5.43 (bs, 1H), 5.16 (s, 2H), 4.54 (bS, 1H), 4.26 (d, J = 1.8 Ηζ, ΙΗ) , 4.04 (d, J = 2.7 Ηζ, ΙΗ), 3.84 (bs, 1H), 3.80-3.64 (m, 1H), 3.58 (S, 3H), 3.41-3.39 (m, 1H), 3.22-3.06 (m , 5H), 2.49 (d, J = 18.6Hz, 1H), 2.35 (s, 3H), 2.30-2.25 (m, 1H), 2.24 (s, 3H), 1.87 (s, 3H), 1.45-1.33 ( m, 1H), 1.19 (s, 9H), 1.00 (br d, J = 6.6 HZ, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 184.9, 180.9,172.6,154.7, 151.3,149.1,148.6,144.7,

132.9,131.3,129.8, 124.5, 123.7,117.3, 116.8, 99.1, 79.4, 59.8, 58.6, 57.7, 56.2,

55.6, 54.9, 54.5, 50.1, 41.6, 40.1, 28.0, 25.3, 24.4, 18.1, 15.7, 8.0.

ESI-MS m / z: Calcd. For C35H45N5O9: 679.7. Found (M + H) ⁺ : 680.3.

Example 4

To a degassed solution of compound 16 (1.8 g, 2.64 ml) in dimethylformamide (221 ml) was added 10% palladium on carbon (360 mg) and stirred under an atmosphere of hydrogen (at atmospheric pressure) for 10 minutes.

17 woman in 45 minutes. The reaction mixture was filtered through celite under argon into a flask containing anhydrous cesium carbonate (2.58 g, 7.92 ml). Bromochloromethane (3.40 ml, 52.8 ml) was then added and the vessel was closed and stirred at 100 ° C for 2 hours. The reaction mixture was cooled, filtered through a pad of celite and washed with methylene chloride. The organic layer was concentrated and dried over sodium sulfate to give 17 as a brown oil, which was used in the next step without further purification.

Rf: 0.36 (hexane: ethyl acetate 1: 5, silica gel).

1 H NMR (300 MHz, CDCl 3): δ 6.68 (s, 1H), 6.05 (bs, 1H), 5.90 (s, 2H), 5.79 (s, 1H), 5.40 (bs, 1H), 5.31-5.24 ( m, 2H), 4.67 (d, J = 8.1 Ηζ, ΙΗ), 4.19 (d, J = 2.7 Ηζ, ΙΗ), 4.07 (bs, 1H), 4.01 (bs, 1H), 3.70 (s, 3H). 3.67 (s, 3H), 3.64-2.96 (m, 5H), 2.65 (d, J = 18.3 Ηζ, ΙΗ), 2.33 (S, 3H), 2.21 (s, 3H), 2.04 (s, 3H), 2.02 -1.95 (m, 1H), 1.28 (s, 9H), 0.87 (d, J = 6.3 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 172.1, 162.6,154.9,149.1, 145.7,135.9,130.8, 130.7,

125.1, 123.1, 117.8, 100.8, 99.8, 76.6, 59.8, 59.2, 57.7, 57.0, 56.7, 55.8, 55.2, 49.5,

41.6, 40.1, 36.5, 31.9, 31.6, 29.7, 28.2, 26.3, 25.0, 22.6, 18.2, 15.8, 14.1, 8.8. ESI-MS m / z: Calcd. For C36H47N5O9: 693.34. Found (M + H) ⁺ : 694.3.

Example 5

To a flask containing a solution of 17 (1.83 g, 2.65 ml) in dimethylformamide (13 ml), cesium carbonate (2.6 g, 7.97 ml) and allyl bromide (1.15 ml, 13.28 ml) were added at 0 ° C. Filter the resulting reaction mixture ·

AllylBr, Cs ₂ CO ₃

DMF, 23 ° C

The mixture was washed with a pad of celite and washed with methylene chloride. The organic layer

dried over sodium sulfate and concentrated. The residue was purified by flash chromatography (silica gel, chloroform: ethyl acetate 1: 4) to give 18 (1.08 g, 56%) as a white solid.

Rf: 0.36 (chloroform-ethyl acetate 1: 3).

Ή NMR (300 MHz, CDCl3): δ 6.70 (s, 1H), 6.27-6.02 (m, 1H), 5.94 (s, 1H), 5.83 (s, 1H), 5.37 (dd, Ji = 1.01 Hz, J ₂ = 16.8 Ηζ, ΙΗ), 5.40 (bs, 1H), 5.25 (dd, J, = 1.0 Hz, J ₂ =

10.5 Hz, 1H), 5.10 (s, 2H), 4.91 (bs, 1H), 4.25-4.22 (m, 1H), 4.21 (d, J = 2.4 Ηζ, ΙΗ), 4.14-4.10 (m, 1H). 4.08 (d, J = 2.4 Ηζ, ΙΗ), 4.00 (bS, 1H), 3.70 (s, 3H), 3.59 (s, 3H), 3.56-3.35 (m, 2H), 3.233.20 (m, 2H) , 3.05-2.96 (cM, H = 8.1 Hz, J ₂ = 18 Hz, 1H), 2.63 (d, J = 18 Ηζ, ΙΗ), 2.30 (s, 3H), 2.21 (s, 3H), 2.09 (s , 3H), 1.91-1.80 (m, 1H), 1.24 (s, 9H), 0.94 (d, J = 6.6 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 172.0, 154.8, 148.8, 148.6,148.4, 144.4,138.8, 133.7,

130.9, 130.3, 125.1, 124.0, 120.9, 117.8, 112.8, 112.6, 101.1, 99.2, 73.9, 59.7, 59.3,

57.7, 56.9, 56.8, 56.2, 55.2, 40.1, 34.6, 31.5, 28.1, 26.4, 25.1, 22.6, 18.5, 15.7, 14.0,

9.2.

ESI-MS m / z: Calcd. For C39H51N5O9: 733.4. Found (M + H) ⁺ : 734.4.

Example 6

To a solution of 18 (0.1 g, 0.137 ml) in dioxane (2 ml) was added 4.2M hydrochloric acid / dioxane (1.46 ml) and the mixture was stirred for 1.2 hours at 23 ° C. The reaction mixture was fixed at 0 ° C with sat.

day solution of sodium bicarbonate (60 ml) and extracted with ethyl acetate (2x70 ml). The organic layer was dried over sodium sulfate, concentrated in vacuo to give 19 (267 mg, 95%) as a white solid, which was used in the next step without further purification. Rf: 0.17 (ethyl acetate: methanol 10: 1, silica gel).

¹ H NMR (300 MHz, CDCl 3): δ 6.49 (s, 1 Η), 6.12-6.00 (m, 1H), 5.94 (s, 1 Η), 5.86 (s, 1H), 5.34 (dd.J ^ IO ΗζΛ = 17.4 Ηζ, ΙΗ), 5.25 (dd, 0, = 1.0 Hz, = = 10.2 Ηζ, ΙΗ), 4.18-3.76 (m, 5H), 3.74 (s, 3H), 3.71-3.59 (m, 1H) , 3.36-3.20 (m, 4H), 3.01-2.90 (m, 1H), 2.60 (d, J = 18.0 Hz, 1H), 2.29 (s, 3H), 2.24 (s, 3H), 2.11 (s, 3H) ), 1.97-1.86 (m, 1H), 0.93 (d, J = 8.7 Hz, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 175.5,148.4, 146.7,144.4, 142.4,138.9, 133.7,131.3,

128.3, 120.8, 117.9,117.4, 113.8, 112.4, 101.1, 74.2, 60.5, 59.1, 56.5, 56.1, 56.3, 56.0, 55.0, 50.5, 41.6,39.5, 29.5, 26.4, 24.9, 21.1, 15.7,9.33.

ESI-MS m / z: Calcd. For (WW 2 SO 4: 589. Found (M + H) *: 590.

Example 7

Me phenylisothiocyanate

1.23 ° C

Me

f ♦ ··· • ·· • · • · ·· · · • · • · • · • · • e • · · • · • · · is · · • · • · · ♦ · ♦ ··· ··· ♦ • ·

To a solution of 19 (250 mg, 0.42 ml) in methylene chloride (1.5 ml) was added phenylisothiocyanate (0.3 ml, 2.51 ml) and the mixture was stirred at 23 ° C for 1 hour. The reaction mixture was concentrated in vacuo and the residue purified by flash chromatography (silica gel, gradient hexane to 5: 1 hexane: ethyl acetate) to give 20 (270 mg, 87%) as a white solid.

Rf: 0.56 (chloroform: ethyl acetate 1: 4).

1 H NMR (300 MHz, CDCl 3): δ 8.00 (bs, 1H), 7.45-6.97 (m, 4H), 6.10 (s, 1H), 6.086.00 (m, 1H), 5.92 (s, 1H). 5.89 (s, 1H), 5.82 (s, 1H), 5.40 (dd, 4 = 1.5 Hz, 4 = 17.1 Hz, 1H), 5.38 (bs, 1H), 5.23 (dd, 4 = 1.5 Hz, J ₂ = 10.5 Hz, 1H), 4.42-4.36 (m, 1H), 4.19-4.03 (m, 5H), 3.71 (S, 3H), 3.68-3.17 (m, 4H), 2.90 (dd, Ji = 7.8 Hz, J ₂ = 18.3 Hz, 1H), 2.57 (d, J = 18.3 Ηζ, ΙΗ), 2.25 (s, 3H), 2.12 (s, 3H), 2.10 (s, 3H), 1.90 (<* 1.4-12.3) Hz, J ₂ =

16.5 Hz, 1H), (m, 1H), 0.81 (d, J = 6.9 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 178.4, 171.6,148.6, 146.8,144.3,142.7,138.7, 136.2, 133.6, 130.7,129.8, 126.6, 124.2, 124.1,120.9, 117.7, 117.4, 116.7, 112.6, 112.5 , 101.0, 74.0, 60.6, 59.0, 57.0, 56.2, 56.1,55.0, 53.3,41.4,39.7, 26.3, 24.8, 18.3,15.5,

9.2.

ESI-MS m / z: Calculated for C ^ H44N ₆ O6S: 724.8. Found (M + H) ⁺ . 725.3.

Me

To a solution of 20 (270 mg, 0.37 ml) in dioxane (1 ml) was added 4.2M hydrochloric acid / dioxane (3.5 ml) and the mixture was stirred for minutes at 23 ° C. Ethyl acetate (20 ml) and water (20 ml) were then added.

• ·

mi) and the organic layer is decanted. The aqueous phase was basified at 0 ° C with saturated aqueous sodium bicarbonate solution (60 ml) (pH 8) and then extracted with methylene chloride (2x50 ml). The combined organic extracts were dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography (silica gel, ethyl acetate methanol 5: 1) to give 21 (158 mg, 82%) as a residue.

lo solid.

Rf: 0.3 (ethyl acetate: methanol 1: 1).

1 H NMR (300 MHz, CDCl 3): 56.45 (s, 1 Η), 6.12-6.03 (m, 1 Η), 5.91 (s, 1H), 5.85 (s, 1H), 5.38 (dd, Ji = 1.2 Hz) , 42 = 17.1 Ηζ, ΙΗ), 5.24 (dd, Ji = 1.2 Hz, J ₂ = 10.5 Ηζ, ΙΗ), 4.23-4.09 (m, 4H), 3.98 (d, J = 2.1 Ηζ, ΙΗ), 3.90 ( bs, 1H), 3.72 (s, 3H), 3.36-3.02 (m, 5H), 2.72-2.71 (m, 2H), 2.48 (d, J = 18.0 Ηζ, ΙΗ), 2.33 (s, 3H), 2.22 (s, 3H), 2.11 (s, 3H), 1.85 (60.3, = 11.7 Hz, J ₂ = 15.6 Ηζ, ΙΗ).

¹³ C NMR (75 MHz, CDCl 3): δ 148.4, 146.7,144.4, 142.8, 138.8, 133.8, 130.5, 128.8,

121.5, 120.8, 118.0, 117.5, 116.9, 113.6, 112.2, 101.1, 74.3, 60.7, 59.9, 58.8, 56.6,

56.5, 55.3, 44.2, 41.8, 29.7, 26.5, 25.7, 15.7, 9.4.

ESI-MS m / z: Calculated for C ₂₉ H ₃ 4N ₄ O ₅ : 518.3. Found (M + H) ⁴ : 519.2.

Example 9

TrocCI, py, CH ₂ CI ₂ -10 ° C. 1h.

Pyridine (0.104 ml, 1.28 ml) was added to a solution of 21 (0.64 g, 1.22 ml) in methylene chloride (6.13 ml) at -10 ° C. The mixture was stirred at this temperature for 1 hour, then fixed by the addition of

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ············ cation of 0.1 N hydrochloric acid (10 ml) and extraction with methylene chloride (2 x 10 ml). The organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography (silica gel, hexane: ethyl acetate 1: 2) to give 22 (0.84 g, 98%) as a white porous solid.

«

Rf: 0.57 (ethyl acetate. Methanol 5: 1).

Ή NMR (300 MHz, CDCl 3): δ 6.50 (s, 1 Η), 6.10-6.00 (m, 1H), 6.94 (d, J = 1.5 Ηζ, ΙΗ),

5.87 (d, J = 1.5 Ηζ, ΙΗ), 5.73 (bs, 1H), 5.37 (dq, Ji = 1.5 Hz, J ₂ = 17.1 Ηζ, ΙΗ), 5.26 (dq, Ji = 1.8 Hz, J2 = 10.2 Ηζ , ΙΗ), 4.60 (d, J = 12 Ηζ, ΙΗ), 4.22-4.10 (m, 4H), 4.19 (d, J = 12 Ηζ, ΙΗ), 4.02 (m, 2H), 3.75 (s, 3H) , 3.37-3.18 (m, 5H), 3.04 (dd, J, = 8.1 Hz, J ₂ = 18 Hz, 1H), 2.63 (d, J = 18 Ηζ, ΙΗ), 2.31 (s, 3H), 2.26 ( s, 3H), 2.11 (s, 3H), 1.85 (dd, J, = 12.3 Hz, J ₂ = 15.9 Ηζ, ΙΗ).

¹³ C NMR (75 MHz, CDCl 3): δ 154.3, 148.5,146.7,144.5, 142.8,139.0, 138.8,130.7,

128.7, 121.3,120.8, 117.8, 117.7,116.8,112.7, 101.2, 77.2, 74.3, 60.7,59.9, 57.0,

56.4, 55.3,43.3, 41.7,31.6, 26.4, 25.3, 22.6,15.9,14.1, 9.4.

ESI-MS m / z: Calcd for C 30 H 31 N 3 O 3: 694.17. Found (M + H) ⁺ : 695.2.

Example 10

To a solution of 22 (0.32 g, 0.46 ml) in CH3CN (2.33 ml) at 0 ° C was added diisopropylethylamine (1.62 ml, 9.34 ml), bromomethylmethyl ether (0.57 ml, 7.0 ml) and dimethylaminopyridine (6 mg, 0.046 ml). . The mixture was stirred at 30 ° C for 10 hours. The reaction mixture was then diluted with dichloromethane (30 ml) and poured into an aqueous solution of brine. · · · · · · · · · · · · · · · · · · · · · · · · Acid (10 ml) (pH = 5). The organic layer was dried over sodium sulfate and the solvent was removed under reduced pressure to give a residue which was purified by flash chromatography (silica gel, hexane: ethyl acetate 2: 1) to give 23 (0.304 g, 88%) as a white porous solid.

Rf: 0.62 (hexane: ethyl acetate 1: 3).

Ή NMR (300 MHz, CDCI3): δ 6.73 (s, 1 Η), 6.10 (m, 1H), 5.94 (d, J = 1.5 Ηζ, ΙΗ), 5.88 (d, J = 1.5 Ηζ, ΙΗ), 5.39 (dq, Ji = 1.5 Hz, 4 = 17.1 Hz, 1H), 5.26 (dq, J, = 1.8 Hz, J ₂ =

10.2 Ηζ, ΙΗ), 5.12 (s, 2H), 4.61 (d, J = 12 Hz, 1H), 4.55 (t, J = 6.6 Ηζ, ΙΗ), 4.25 (d, J = 12 Ηζ, ΙΗ), 4.22 -4.11 (m, 4H), 4.03 (m, 2H), 3.72 (8.3H), 3.58 (s, 3H), 3.38-3.21 (m, 5H), 3.05 (dd, Ji = 8.1 Hz, J ₂ = 18 Ηζ, ΙΗ), 2.65 (d, J = 18 Ηζ, ΙΗ), 2.32 (s, 3H), 2.23 (s, 3H), 2.12 (s, 3H), 1.79 (dd, J, = 12.3 Hz, J ₂ = 15.9 Ηζ, ΙΗ).

¹³ C NMR (75 MHz, CDCl3): δ 154.3, 148.6,148.4, 144.5,139.0,133.6,130.6, 130.1, 125.07, 124.7, 124.0, 121.1,117.7, 112.6,101.2, 99.2, 77.2, 74.4, 74.1, 59.8 , 59.8,

57.7, 57.0, 56.8, 56.68, 55.3, 43.2, 41.5, 26.4, 25.2, 15.9,9.3.

ESI-MS m / z: Calcd. For C34H39Cl3N4O8: 738.20. Found (M + H) *: 739.0.

Example 11

To a suspension of 23 (0.304 g, 0.41 ml) in 90% aqueous acetic acid (4 ml) was added powdered zinc (0.2 g, 6.17 ml) and the reaction mixture was stirred for 7 hours at 23 ° C. The mixture was filtered through a pad of celite, which was washed with dichloromethane. The organic layer was washed with aqueous saturated sodium bicarbonate solution (15 ml) (pH = 9) and washed with brine. · · · · · · · · · · · · · · · · · · · · · · · · · ··· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · The solvent was removed under reduced pressure to give 24 (0.191 g, 83%) as a white solid.

Rf: 0.3 (ethyl acetate: methanol 5: 1).

¹ H NMR (300 MHz, CDCl 3): δ 6.68 (s, 1 Η), 6.09 (m, 1H), 5.90 (d, J = 1.5 Ηζ, ΙΗ), 5.83 (d, J = 1.5 Ηζ, ΙΗ). 5.39 (dq, Ji = 1.5 Hz, Ja = 17.1 Ηζ, ΙΗ), 5.25 (dq, J, = 1.5 Hz, 02 =

10.2 Ηζ, ΙΗ), 5.10 (S, 2H), 4.22-4.09 (m, 3H), 3.98 (d, J = 2.4 Hz, 1H), 3.89 (m, 1H), 3.69 (s, 3H), 3.57 ( s, 3H), 3.37-3.17 (m, 3H), 3.07 (dd, J, = 8.1 Hz, J ₂ = 18 Ηζ, ΙΗ), 2.71 (m, 2H), 2.48 (d, J = 18 Ηζ, ΙΗ ), 2.33 (s, 3H), 2.19 (s, 3H), 2.17 (s, 3H), 1.80 (dd, J, =

12.3 Hz, 02 = 15.9 Ηζ, ΙΗ).

¹³ C NMR (75 MHz, CDCl 3): δ 148.5,148.2,144.3, 138.7,133.7,130.7,129.9,125.0,

123.9, 121.3,117.9, 117.5,113.6, 112.0,101.0, 99.2, 74.0, 59.8, 59.7, 58.8, 57.6, 57.0, 56.2, 55.2, 44.2, 41.5, 31.5, 26.4, 25.6, 22.5, 16.7, 14.0, 9.2.

ESI-MS m / z: Calculated for Ο ^ Η ^ Ν ^: 562.66. Found (M + H) ⁺ : 563.1.

Example 12

To a solution of 24 (20 mg, 0.035 ml) in water (0.7 ml) and tetrahydrofuran (0.7 ml) at 0 ° C was added sodium nitrite (1.62 mg, 0.17 ml) and 90% aqueous acetic acid (0.06 ml) and the mixture. was stirred at 0 ° C for 3 hours. The reaction mixture was then diluted with dichloromethane (5 ml) and the organic layer was washed with water (1 ml), dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography (silica gel, hexane: ethyl acetate 2: 1) to give 25 (9.8 mg, 50%) as a white solid.

«· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

Rf: 0.34 (hexane: ethyl acetate 1: 1).

¹ H NMR (300 MHz, CDCl 3): δ 6.71 (s, 1H), 6.11 (m, 1H), 5.92 (d, J = 1.5 Ηζ, ΙΗ), 5.87 (d, J = 1.5 Ηζ, ΙΗ), 5.42 (dq, Ji = 1.5 NgD = 17.1 Ηζ, ΙΗ), 5.28 (dq, J, = 1.8 Hz, J ₂ =

10.2 Hz, 1H), 5.12 (s, 2H), 4.2 & 4.09 (m, 3H), 4.05 (d, J = 2.4 Ηζ, ΙΗ), 3.97 (t, J = 3.0 Ηζ, ΙΗ), 3.70 (s , 3H), 3.67-3.32 (m, 4H), 3.58 (s, 3H), 3.24 (dd, Ji = 2.7 Hz, J ₂ = 15.9 *

Ηζ, ΙΗ), 3.12 (dd, 4 = 8.1 Hz, J ₂ = 18.0 Ηζ, ΙΗ), 2.51 (d, J = 18 Hz, 1H), 2.36 (s, 3H).

2.21 (s, 3H), 2.12 (s, 3H), 1.83 (dd, 4 = 12.3 Hz, J ₂ = 15.9 Ηζ, ΙΗ).

^{, 3} C NMR (75 MHz, CDCl 3): δ 148.7, 148.4,138.9,133.7, 131.1,129.4,125.1,123.9,

120.7, 117.6, 117.5, 113.2, 112.3, 101.1, 99.2, 74.0, 63.2, 59.8, 59.7, 57.9, 57.7, 57.0,

56.5, 55.2, 41.6, 29.6, 26.1, 25.6, 22.6, 15.7,9.2.

ESI-MS m / z: Calcd. For C31H37N3O7: 563.64. Found (M + H) *: 564.1.

Example 13

2,2,2-TricNoroethyl chloroformate

NaH, THF, reflux

The starting material (2.0 g, 5.90 ml) was added to a suspension of sodium hydride (354 mg, 8.86 ml) in tetrahydrofuran (40 ml) at 23 ° C, after which the suspension was treated with allyl chloroformate (1.135 ml, 8.25 ml) at 23 And then heated at reflux for 3 hours. The suspension was cooled, filtered, the solid was washed with ethyl acetate (100 ml) and the filtrate was concentrated. The crude oil was stirred with hexane (100 ml) and kept at 4 ° C overnight. The solvent was then decanted and the light yellow slurry treated with dichloromethane (20 ml) and precipitated with hexane (100 ml). After 10 minutes the solvent was decanted again. The operation is repeated until a white solid appears. The white solid was filtered off and dried to give 29 (1.8 g, 65%) as a white solid.

• · 4 · · · · · · <. · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ···

Ή NMR (300 MHz, CDCl3): δ 7.74 (d, J = 7.5 Hz, 2H), 7.62 (d, J = 6.9 Hz, 2H), 7.33 (t, J = 7.5 HZ.2H), 7.30 (t. J = 6.3 Hz, 2H), 5.71 (d, J = 7.8 Ηζ, ΙΗ), 4.73 (d, J = 7.8 Hz, 2H), 4.59 (m, 1H), 4.11 (t, J = 6.0 Ηζ, ΙΗ) , 3.17 (dd, J, - & 0 Hz, 4 = 2.7 Hz, 2H), 3.20 (dd, 4 = 5.4 Hz, 4 = 2.1 Hz, 2H).

¹³ C NMR (75 MHz, CDCl 3): δ 173.6, 152.7, 144.0, 139.7, 137.8, 126.0, 125.6, 123.4,

118.3, 73.4, 52.4,45.5, 35.8, 33.7.

ESI-MS m / z: Calculated for C ₂₀ H ₁₈ Cl ₃ NO ₄ S: 474.8. Found (M + Na) *: 497.8.

Example 14

A mixture of compound 25 (585 mg, 1.03 ml) and compound 29 (1.47 mg, 3.11 ml) was azeotroped with anhydrous toluene (3x10 ml). To a solution of 25 and 29 in anhydrous dichloromethane (40 ml) was added DM AP (633 mg, 5.18 ml) and EDC hydrochloride (994 mg, 5.18 ml) at 23 ° C. The reaction mixture was stirred at 23 ° C for 3 hours. The mixture was separated using saturated aqueous sodium bicarbonate solution (50 ml) and the layers were separated. The aqueous layer was washed with dichloromethane (50 ml). The combined organic layers were dried over sodium sulfate, filtered and concentrated. The crude product was purified by flash chromatography (silica gel, hexane: ethyl acetate 3: 1) to give 30 (1.00 g, 95%) as a pale yellow creamy solid.

¹ H NMR (300 MHz, CDCl 3): δ 7.72 (m, 2H), 7.52 (m, 2H), 7.38 (m, 2H), 7.28 (m, 2H), 6.65 (s, 1H), 6.03 (m. 1H), 5.95 (d, J = 1.5 Hz, 1H), 5.79 (d, J = 1.5 Ηζ, ΙΗ), 5.39 (m, 1H), 5.29 (dq, J = 10.3 Hz, J = 1.5 Hz, 1H) , 5.10 (s, 2H), 4.73 (d, J = 11.9 Hz, 1H), 4.66 (d, J = 11.9 Ηζ, ΙΗ), 4.53 (m, 1H), 4.36-3.96 (m, 9H), 3.89 ( t, J = 6.4 Ηζ, ΙΗ), 3.71 (s,

3H), 3.55 (s, 3H), 3.33 (m, 1H), 3.20 (m, 2H), 2.94 (m, 3H), 2.59 (m, 1H), 2.29 (s, 3H).

2.23 (s, 3H), 2.02 (s, 3H), 1.83 (dd, J, = 16.0 Hz, J ₂ = 11.9 Ηζ, ΙΗ).

^{, 3} C NMR (75 MHz, CDCl 3): δ 109.7,154.0,148.8,148.4,145.7,144.5,140.9,139.0,

133.7, 130.9,130.6, 127.6,127.0, 124.8,124.6, 124.1,120.8, 119.9,118.2,117.7,

117.3.112.7, 112.1,101.3,99.2, 74.7, 73.9, 64.4, 59.8, 57.7, 57.0,56.8, 55.4,53.3,

46.7,41.4, 36.5,34.7, 31.5, 26.4, 24.9, 22.6,15.7, 14.0,9.1.

ESI-MS m / z: Calcd. For CeIH ^ CyttOmS: 1020.4. Found (M + H) *:

1021.2.

Example 15

31

To a solution of 30 (845 mg, 0.82 ml), acetic acid (500 mg, 8.28 ml) and (PPI3) _g PICCl ₂ (29 mg, 0.04 ml) in anhydrous dichloromethane (20 ml) was added at 23 ° C. drops of BuaSnH (650 mg, 2.23 ml). The reaction mixture was stirred at this temperature for 15 minutes, whereby bubbles were removed. The crude product was fixed with water (50 ml) and extracted with dichloromethane (3x50 ml). The organic layers were dried over sodium sulfate, filtered and concentrated. The crude product was purified by flash chromatography (ethyl acetate / hexane in a gradient of 1: 5 to 1: 3) to give compound 31 (730 mg, 90%) as

pale yellow creamy solid.

Ή NMR (300 MHz, CDCl 3): δ 7.72 (m, 2H), 7.56 (m, 2H), 7.37 (m, 2H), 7.30 (m, 2H).

6.65 (s, 1H), 5.89 (S, 1H), 5.77 (s, 1H), 5.74 (s, 1H), 5.36 (d, J = 5.9 Ηζ, ΙΗ), 5.32 (d, J =

5.9 Ηζ, ΙΗ), 5.20 (d, J = 9.0 Ηζ, ΙΗ), 4.75 (d, J = 12.0 Ηζ, ΙΗ), 4.73 (m, 1H), 4.48 (d, J =

11.9 Hz, 1H), 4.08 (m, 4H), 3.89 (m, 1H), 3.86 (t, J = 6.2 Ηζ, ΙΗ), 3.70 (s, 3H), 3.69 (s.

3H), 3.38 (m, 1H), 3.25 (m, 1H), 3.02-2.89 (m, 4H), 2.67 (s, 1H), 2.61 (s, 1H), 2.51 (dd,

Ji = 14.3 Hz, J ₂ = 4.5 Ηζ, ΙΗ), 2.29 (s, 3H), 2.23 (s, 3H), 1.95 (s, 3H), 1.83 (m, 1H). ¹³ C NMR (75 MHz, CDCl 3): δ 168.2, 152.5,148.1, 146.2,144.4, 144.3,143.3, 139.6,

134.6, 129.7,129.6, 126.2, 125.6, 123.4,123.3, 121.6,118.5, 116.3,110.7, 110.2,

40.1,35.6, 33.3, 24.8, 23.3, 14.5, 7.3.

ESI-MS m / z: Calcd. For C 22 H 10 Cl 2 O 5 S: 980.3. Found (M + H) ⁺ : 981.2.

Example 16

To a solution of 31 (310 mg, 0.32 ml) in anhydrous dichloromethane (15 ml), at -10 ° C was added, via cannula, a solution of benzene selenic anhydride 70% (165 mg, 0.32 ml) in anhydrous dichloromethane (7 ml), such as the temperature is maintained at -10 ° C. The reaction mixture was stirred at -10 ° C for 5 minutes. At this temperature, saturated sodium bicarbonate solution (30 ml) was added. The aqueous solution was washed with more dichloromethane80

tan (40 ml) · The organic layers were dried over sodium sulfate, filtered and concentrated. The crude product was purified by flash chromatography (ethyl acetate / hexane in a gradient of 1: 5 to 1: 1) to give compound 32 (287 mg, 91%, HPLC: 91.3%) as a pale yellow creamy solid and as a a mixture of two isomers (65:35),

which are used in the next step.

1 H NMR (300 MHz, CDCl 3): δ (mixture of isomers) 7.76 (m, 4H), 7.65 (m, 4H), 7.39 (m, 4H), 7.29 (m, 4H), 6.62 (s, 1H) ), 6.55 (s, 1H), 5.79-5.63 (t, 6H), 5.09 (s, 1H), 5.02 (d, J = 6.0 Ηζ, ΙΗ), 4.99 (d, J = 6.0 Ηζ, ΙΗ), 4.80 -4.63 (m, 6H), 4.60 (t, 1H), 4.50 (t, 1H), 4.38 (dd, J = 12.8 Hz, J = 7.5 Ηζ, ΙΗ), 4.27 (dd, J = 12.8 Hz, J = 7.5 Ηζ, ΙΗ), 4.16-

3.90 (m, 10H), 3.84 (s, 3H), 3.62 (s, 3H), 3.50 (s, 3H), 3.49 (s, 3H), 3.33-2.83 (m, 14H), 2.45-2.18 (m. 2H), 2.21 (s, 6H), 2.17 (s, 6H), 1.77 (s, 6H), 1.67 (m, 2H).

¹³ C NMR (75 MHz, CDCl 3): δ (mixture of isomers) 168.6,168.4,158.6,154.8,

152.8, 152.5,147.3, 147.2,146.8, 144.1,144.0, 140.8,139.7, 137.1,129.8, 129.3,

128.4, 128.7,126.5, 125.5,123.7, 123.6,123.5, 123.4,122.2, 121.3,118.3,115.8,

115.5, 110.2, 106.9, 103.5, 103.2, 100.1,99.6, 97.9, 97.7, 93.8, 73.4, 70.9, 69.2, 64.9,

62.5, 59.3, 58.9, 58.4, 56.7, 56.3, 56.2, 55.4, 55.2, 55.1, 54.9, 54.7, 54.3, 54.1, 53.8,

52.8, 45.5, 40.5, 40.0, 39.8, 35.8, 35.5, 33.9, 33.7, 30.1, 28.4, 24.2, 24.1, 21.2, 14.5,

14.4, 12.7, 6.0, 5.7.

ESI-MS m / z: Calcd. For C48H49Cl3N4OHS: 996.3. Found (M + H) ⁺ : 997.2.

Example 17

• ♦ ··· · ·· · * f ·· · ···· ···· • · · ♦ · · · • · · · · · · · · · · · · · · · · · · · · · · · · · · ·

The reaction vessel was washed twice, purged with argon in vacuo and kept in an argon atmosphere for the reaction. To a solution of dimethyl sulfoxide (39.1 ml, 0.55 ml, 5 equivalents) in anhydrous dichloromethane (4.5 ml) at -78 ° C was added dropwise triflic anhydride (37.3 ml, 0.22 ml, 2 equivalents). The reaction mixture was stirred at -78 ° C for 20 minutes, then a solution of 32 (110 mg, 0.11 ml, HPLC: 91.3%) anhydrous dichloromethane (1 ml for the main addition and 0.5 ml for washing). The temperature was maintained at -78 ° C during the addition and the color changed from yellow to brown. The reaction mixture was stirred at -40 ° C for 35 minutes. During this period the solution changes from yellow to dark green. IPrzNEt (153 ml, 0.88 ml, 8 equivalents) was then added dropwise and the reaction mixture was kept at 0 ° C for 45 minutes, turning the color of the solution to brown. Then t-butanol (41.6 ml, 0.44 ml, 4 equivalents) and r -butyl N3.3-tetramethylguanidine (132.8 ml, 0.77 ml, 7 equivalents) were added dropwise and the reaction was stirred at 23 ° C for 23 h. 40 minutes. Acetic anhydride (104.3 ml, 1.10 ml, 10 equivalents) was then added dropwise and the reaction mixture was maintained at 23 ° C for another 1 hour. The reaction mixture was then diluted with dichloromethane (20 ml) and washed with aqueous saturated ammonium chloride solution (50 ml), sodium bicarbonate (50 ml) and sodium chloride (50 ml). The combined organic layers were dried over sodium sulfate, filtered and concentrated. The residue was purified by flash chromatography (ethyl acetate / hexane gradient from 1: 3 to 1: 2) to give compound 33 (54 mg, 58%) as a pale yellow solid.

Ή NMR (300 MHz, CDCl3): δ 6.85 (s, 1H), 6.09 (s, 1H), 5.99 (s, 1H), 5.20 (d, J = 5.8)

Ηζ, ΙΗ), 5.14 (d, J = 5.3 Ηζ, ΙΗ), 5.03 (m, 1H), 4.82 (d, J = 12.2 Ηζ, ΙΗ), 4.63 (d, J =

12.0 Ηζ, ΙΗ), 4.52 (m, 1H), 4.35-4.17 (m, 4H), 3.76 (s, 3H), 3.56 (s, 3H), 3.45 (m, 2H).

2.91 (m, 2H), 2.32 (s, 3H), 2.28 (s, 3H), 2.21 (s, 3H), 2.12 (m, 2H), 2.03 (s, 3H).

· · · · · · · · · · · · · · · · · · · · 9 9 9 9 99

9 9 9 9 9 9 99

9 9 9 9 9 99

999 99 999 9999 99999 ¹³ C NMR (75 MHz, CDCl 3): δ 168.5, 167.2, 152.7,148.1,147.1,144.5,139.6,139.1,

130.5, 129.0,123.7, 123.5,123.3,118.8, 116.5, 112.1, 100.6,97.8, 73.3, 60.5, 59.4,

59.2, 58.3, 57.6, 57.4, 56.1,53.3, 53.1,40.6, 40.0,31.0, 22.2,18.9,14.4, 8.1.

ESI-MS m / z: Calcd. For C36H39Cl3N4O11S: 842.1. Found (M + H) ⁺ : 843.1.

Example 18

To a solution of 33 (12 mg, 0.014 ml) in anhydrous dichloromethane (1.2 ml) and in pure high-performance liquid chromatography acetonitrile (1.2 ml), sodium iodide (21 mg, 0.14 ml) was added and freshly distilled (at 23 ° C). atmospheric calcium hydride) trimethylsilyl chloride (15.4 mg, 0.14 ml). The reaction mixture turned orange. After 15 minutes, the solution was diluted with dichloromethane (10 ml) and washed with fresh aqueous saturated sodium bisulfite solution (3x10 ml). The organic layer was dried over sodium sulfate, filtered and concentrated. Compound 34 (13 mg, quantitative) was obtained as a pale yellow solid, which was used without further purification.

1 H NMR (300 MHz, CDCl 3): δ 6.85 (s, 1H), 6.09 (s, 1H), 5.99 (s, 1H), 5.27 (d, J = 5.8 Hz, 1H), 5.14 (d, J = 5.3 Ηζ, ΙΗ), 5.03 (d, J = 11.9 Ηζ, ΙΗ), 4.82 (d, J = 12.2 Hz, 1H),

4.63 (d, J = 13.0 Ηζ, ΙΗ), 4.52 (m, 1H), 4.34 (m, 1H), 4.27 (bs, 1H), 4.18 (m, 2H), 3.76 (s, 3H), 3.56 (s , 3H), 3.44 (m, 1H), 3.42 (m, 1H), 2.91 (m, 2H), 2.32 (s, 3H), 2.28 (s, 3H), 2.21 (S, 3H), 2.03 (S. 3H).

ESI-MS m / z: Calcd. For CmIW ^ OiqS: 798.1. Found (M + H) ⁺ : 799.1.

• · · · · · ·

Example 19

35

To a solution of 34 (13 mg, 0.016 ml) in a mixture of acetic acid and water (90: 10.1 ml) was added zinc powder (5.3 mg, 0.081 ml) at 23 ° C. The reaction mixture was heated at 70 ° C for 6 hours. It was then cooled to 23 ° C, diluted with dichloromethane (20 ml) and washed with aqueous saturated sodium bicarbonate solution (15 ml) and aqueous triethylamine solution (15 ml). The organic layer was dried over sodium sulfate, filtered and concentrated. The residue was purified by flash chromatography on SiHca-NH ₂ (eluent: ethyl acetate / hexane gradient 0: 100 to 50:50) to give compound 35 (6.8 mg, 77% in two steps) as a pale yellow solid.

1 H NMR (300 MHz, CDCl 3): δ 6.51 (s, 1H), 6.03 (dd, J = 1.3 Hz, J = 26.5 Hz, 2H), 5.75 (bs, 1H), 5.02 (d, J = 11.6 Ηζ , ΙΗ), 4.52 (m, 1H), 4.25 (m, 2H), 4.18 (d, J = 2.5 Ηζ, ΙΗ), 4.12 (dd, J = 1.9 Hz, J = 11.5 Hz, 1H), 3.77 (s , 3H), 3.40 (m, 2H), 3.26 (t, J = 6.4 Hz, 1H), • 2.88 (m, 2H), 2.30-2.10 (m, 2H), 2.30 (s, 3H), 2.28 (s , 3H), 2.18 (s, 3H), 2.02 (s, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 174.1, 168.4,147.8, 145.4,142.9, 140.8,140.1, 131.7,

130.2, 129.1, 128.3, 120.4, 111.3, 117.9, 113.8, 111.7, 101.7, 61.2, 59.8, 59.2, 58.9,

54.4, 53.8, 54.4, 41.3, 41.5, 34.1, 23.6, 20.3, 15.5, 9.4.

ESI-MS m / z: Calcd. For C31H34N4O8S: 622.7. Found (M + H) ⁺ : 623.2.

Example 20

To a solution of 36 (49 mg, 0.08 ml) and 2- [3-hydroxy-4-methoxyphenyl] ethylamine (46.2 mg, 0.27 ml) in ethanol (2.5 ml) was added silica gel (105 mg). ft · · «· ·· · · ft · · · ft ft ft • * · ft ft ft ft • ft ft ft ft ft ft ft ft ft • ft · ft ft ft ·· ··· ·· ··· ft · · · Ft · · · ·

at 23 ° C. The reaction mixture was stirred at 23 ° C for 14 hours. It was diluted with hexane and poured into a chromatography column (ethyl acetate / hexane 1/3 to 1/1) to give Et-770 (55 mg, 90%) as a pale yellow solid.

Ή NMR (300 MHz, CDCl3): δ 6.60 (s, 1H), 6.47 (s, 1H), 6.45 (s, 1H), 6.05 (s, 1H).

5.98 (s, 1H), 5.02 (d, J = 11.4 Ηζ, ΙΗ), 4.57 (bs, 1H), 4.32 (bs, 1H), 4.28 (d, J = 5.3 Hz, 1H), 4.18 (d, J = 2.5 Ηζ, ΙΗ), 4.12 (dd, J = 2.1 Hz, J = 11.5 Ηζ, ΙΗ), 3.78 (s, 3H), 3.62 (s, 3H), 3.50 (d, J = 5.0 Ηζ, ΙΗ). 3.42 (m, 1H), 3.10 (ddd, J = 4.0 Hz, J = 10.0 Hz, J = 11.0 Ηζ, ΙΗ), 2.94 (m, 2H), 2.79 (m, 1H), 2.61 (m, 1H). 2.47 (m, 1H), 2.35 (m, 1H), 2.32 (s, 3H), 2.27 (s, 3H), 2.20 (s, 3H), 2.09 (m, 1H), 2.04 (s, 3H).

ESI-MS m / z: Calcd. For C40H42N4O10S. 770.7. Found (M + H) ⁺ : 771.2.

© Example 22

To a solution of 21 (22 mg, 0.042 ml) in dichloromethane (0.8 ml) was added phthalic anhydride (6.44 mg, 0.042 ml) and the reaction mixture was stirred for 2 hours at 23 ° C. Carbonyldiimidazole (1 mg, 0.006 ml) was then added and the mixture stirred at 23 ° C for 7 hours. Carbonyldiimidazole (5.86 mg, 0.035 ml) was then added and the reaction stirred for a further 17 hours at 23 ° C. The solution was diluted with dichloromethane (15 ml) and washed with 0.1N hydrochloric acid (15 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash chromatography (silica, hexane: ethyl acetate 2: 1) to give compound 27 (26.4 mg, 96%) as a white solid.

Rf: 0.58 (ethyl acetate).

¹ H NMR (300 MHz, CDCl 3): δ 7.73-7.64 (m, 4H), 6.40 (s, 1H), 6.12-6.01 (m, 1H), 5.63 (s, 1H), 5.58 (d, J = 1.5) Hz, 1H), 5.37 (dd, 4 = 1.8 Hz, 4 = 17.4 Ηζ, ΙΗ), 5.23 (dd, Ji =

1.8 Hz, 4 = 10.5 Ηζ, ΙΗ), 5.12 (d, J = 1.5 Ηζ, ΙΗ), 4.22-4.15 (m, 3H), 4.08 (d, J = 1.8 Hz, 1H), 3.68 (S, 3H) , 3.59-3.55 (m, 2H), 3.35 (d, J = 8.1 Ηζ, ΙΗ), 3.27-3.16 (m, 2H), 3.05 (dd, 4 = 8.1 Hz, 4 = 18.3 Ηζ, ΙΗ), 2.64 ( d, J = 18.0 Ηζ, ΙΗ), 2.30 (s, 3H), 2.24 (s, 3H), 2.09 (s, 3H), 1.80 (dd, 4 = 11.4 Hz, 4 = 15 Ηζ, ΙΗ).

¹³ C NMR (75 MHz, CDCl 3): δ 167.7, 148.9,146.4,144.2, 142.6,139.5, 134.0,133.5, 132.0, 131.0, 128.3, 123.0, 121.3, 120.9, 118.1, 117.5, 116.8, 113.6,112.4, 100.8 ,

74.5, 60.6, 60.5, 57.7, 56.6, 55.6, 55.5, 42.3, 41.7, 26.6, 25.5, 15.9,9.46.

ESI-MS m / z: Calcd. For C37H35N4O7: 648.79. Found (M + H) ⁺ : 649.3.

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

To a solution of 27 (26 mg, 0.041 ml) in dichloromethane (11 ml) at 23 ° C was added acetic acid (11 ml) and (PPhs ^ dCh (2.36 mg) and BusSnH (28 ml, 0.10 ml). stirred at this temperature for 2 hours and poured into a pressure column (silica, gradient hexane to hexane: ethyl acetate 2: 1) to give 28 (24.7 mg, 99%) as a white solid.

Rf: 0.33 (hexane: ethyl acetate 2: 1).

1 H NMR (300 MHz, CDCl 3): δ 7.75-7.70 (m, 2H), 7.69-7.65 (m, 2H), 6.39 (s, 1H), 5.82 (bs, 1H), 5.50 (d, J = 1.5) Ηζ, ΙΗ), 5.0 (d, J = 1.5 Ηζ, ΙΗ), 4.45 (bs, 1H), 4.23-4.19 (m, 2H), 4.10-4.09 (m, 1H), 3.73 (s, 3H), 3.60 -3.48 (m, 2H), 3.36-3.33 (m, 1H), 3.263.20 (m, 1H), 3.14-3.08 (m, 1H), 3.98 (d, J = 14.4 Ηζ, ΙΗ), 2.61 (d , J = 18.8 Ηζ, ΙΗ), 2.30 (s, 3H), 2.23 (s, 3H), 2.06 (s, 3H), 1.85 (dd, J, = 12 Hz, J ₂ = 15.3 Ηζ, ΙΗ) .

¹³ C NMR (75 MHz, CDCl 3): δ 167.8, 146.4,145.1, 143.9, 142.7,137.1, 133.5,131.9,

130.8, 128.4,122.9,120.8, 118.0,116.8,114.0, 113.4,106.4, 100.4, 60.6,60.5,57.8,

56.6, 55.6, 55.5, 42.6, 41.5, 25.6, 25.5, 15.8, 8.9.

ESI-MS m / z: Calcd. For C34H32N4O7: 608.6. Found (M + H) ⁴ : 609.2.

Example 24

phthalicidin

Acetyl chloride (41.58 ml, 0.58 ml) and pyridine (47.3 ml, 0.58 ml) were added to a solution of 28 (357 mg, 0.058 ml) in dichloromethane (3 ml) at 0 ° C. Reaction87 ····· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · The mixture was stirred at this temperature for 1 hour and then the solution was diluted with dichloromethane (15 ml) and washed with 0.1 N hydrochloric acid (15 ml). . The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue is purified by flash chromatography «

(RP-18, CH 3 Cl 2 H 2 O 60:40) to give phthalacidin (354 mg, 94%) as a white solid.

Rf: 0.37 (CH3 Cl2Hz 7: 3, RP-18).

¹ H NMR (300 MHz, CDCl 3): δ 7.72-7.68 (m, 2H), 7.67-7.63 (m, 2H), 6.38 (s, 1H), 5.69 (d, J = 1.2 Ηζ, ΙΗ), 5.64 ( d, J = 1.2 Ηζ, ΙΗ), 5.30 (bs, 1H), 4.25-4.21 (m, 2H), 4.02 (d, J = 2.1 Ηζ, ΙΗ), 3.64-3.62 (m, 5H), 3.33 (d , J = 8.4 Hz, 1H), 3.21-3.16 (m, 1H), 3.02 (dd, 4 = 8.1 Hz, J ₂ = 18 Ηζ, ΙΗ), 2.76 (dd, 4 = 18 Hz, 4 = 15.6 Ηζ. ΙΗ), 2.63 (d, J = 17.7 Hz, 1H), 2.29 (s, 3H), 2.28 (s, 3H), 2.21 (s, 3H), 2.0 (s, 3H), 1.73 (dd, 4 = 12.0) Hz, 4 = 15.3 Hz, 1H).

¹³ C NMR (75 MHZ, CDCl 3): δ 168.5, 167.6,146.2, 144.2,142.5,141.0, 140.5,133.4,

131.8, 130.7, 128.2, 120.9, 120.8, 117.9, 116.4, 113.6, 101.1, 60.4, 60.0, 57.0, 56.3,

55.6, 55.4, 41.6, 41.5, 26.5, 25.2, 20.2, 15.7, 9.4.

ESI-MS m / z: Calcd. For ΟχΗχΝ ^: 650. Found (M + H) ⁺ : 651.2.

Example 25

To a solution of 17 (300 mg, 0.432 ml) in dichloromethane (2 ml) at 0 [deg.] C. was used. Acetyl chloride (30.7 ml, 0.432 ml) and pyridine (34.9 ml, respectively) were added. 0.432 ml). The reaction mixture was stirred at this temperature for 2 hours and then the solution was diluted with dichloromethane (15 ml) and washed with 0.1N hydrochloric acid (15 ml). The organic layer was dried over sodium sulfate, filtered and the solvent removed under reduced pressure to afford 42 (318 mg, 100%) as a white solid, which was used in subsequent reactions without further purification.

Rf: 0.5 (ethyl acetate: methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.66 (s, 1 Η), 5.93 (d, J = 1.2 Ηζ, ΙΗ), 5.83 (d, J = 1.2 Ηζ, ΙΗ), 5.42 (t, J = 6.6 .07, ΙΗ), 5.07 (d, J = 5.7 Ηζ, ΙΗ), 4.98 (d, J = 5.7 Ηζ, ΙΗ), 4.16 (d, J = 1.8 Ηζ, ΙΗ), 4.11 (d, J = 2.7 Ηζ. ΙΗ), 3.98 (bs, 1H), 3.73-3.61 (m, 2H), 3.64 (s, 3H), 3.52-3.48 (m, 1H), 3.50 (s, 3H), 3.33 (d, J = 9.6 Ηζ , ΙΗ), 3.17-3.14 (m, 1H), 2.97-

2.87 (m, 1H), 2.75-2.70 (d, J = 16.8 Hz, 1H), 2.26 (s, 6H), 2.16 (s, 3H), 1.96 (s, 3H), 1.70 (dd, ^ = 11.7 Hz) , J2 = 15.6 Ηζ, ΙΗ), 1.33 (s, 6H), 0.59 (d, J = 6.0 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 172.0, 168.3,162.3, 148.2,144.4, 140.4, 140.2,130.9,

130.5, 125.3,123.4,120.8, 117.6,112.7,111.7, 101.4,99.1, 79.2, 59.5, 58.8, 57.5,

57.4, 56.4, 55.5, 55.0,41.3, 39.0, 28.2, 26.4, 24.6, 19.9, 18.4,15.4,9.1.

ESI-MS m / z: Calcd. For C38H49N5O10: 735.82. Found (M + H) ⁺ : 736.3.

Example 26

• e ··· • 99 9 ' 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9

To a solution of 42 (318 mg, 0.432 ml) in dichloromethane (2.16 ml) was added trifluoroacetic acid (1.33 ml, 17.30 ml) and the reaction mixture was stirred at 23 ° C for 3.5 hours. The reaction mixture was fixed at 0 ° C with saturated aqueous sodium bicarbonate solution (60 ml) and extracted with dichloromethane (2x70 ml). The combined organic layers were dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate: methanol 20: 1) to give 43 (154 mg, 60%) as a white solid.

Rf: 0.22 (EtOAc 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.47 (s, 1H), 6.22 (bs, 1H), 5.95 (d, J = 1.2 Ηζ, ΙΗ),

5.88 (d, J = 1.2 Ηζ, ΙΗ), 4.084.06 (m, 2H), 4.01 (be, 1H), 3.69 (s, 3H), 3.49 (d, J = 3.6 Ηζ, ΙΗ), 3.33 (d , J = 8.1 Ηζ, ΙΗ), 3.26-3.22 (m, 1H), 2.95 (dd, J, = 8.1 Hz, 4 = 18 Hz, 1H), 2.80-2.76 (m, 2H), 2.58 (d, J = 18 Ηζ, ΙΗ), 2.29 (s, 3H), 2.27 (s, 3H), 2.21 (s, 3H), 1.96 (s, 3H), 1.77 (dd, 4 = 12.3 Hz, 4 = 15.6 Ηζ, ΙΗ ), 0.90 (d, J = 6.9 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3). 5 174.8, 169.0,146.8,144.4,142.8,140.5, 140.2,131.1,

128.8, 120.8, 120.5, 117.1, 112.9,111.6, 101.5, 60.3, 59.0, 56.5, 56.3, 55.6, 55.1,

50.2, 41.6, 39.5, 26.8, 26.3, 24.9, 20.2, 15.4, 9.2.

ESI-MS m / z: Calcd. For C31H37N5O7: 591.65. Found (M + H) ⁺ : 592.3.

Example 27

Me Me

• f · »· • 9 9 9 9 9 9 99 9 9 9 9 99 9 9 9 9 9 9 9 9 9 9 · 9 9 9 9 9 9 • · 9 9 99 99 999 9999 99 ··

To a solution of 43 (154 mg, 0.26 ml) in dichloromethane (1.3 ml) was added phenylisothiocyanate (186 ml, 1.56 ml) and the reaction mixture was stirred at 23 ° C for 2 hours. The reaction mixture was concentrated in vacuo and the residue purified by flash column chromatography (silica, gradient hexane to hexane, ethyl acetate 1: 1) to give 44 (120 mg, 63%) as a white solid. Rf: 0.41 (ethyl acetammethanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 8.17 (s, 1 Η), 7.49-7.44 (m, 3H), 7.31-7.24 (m, 3H), 7.05 (d, J = 6.9 Ηζ, ΙΗ), 5.98 (d, J = 1.2 Ηζ, ΙΗ), 5.87 (d, J = 1.2 Ηζ, ΙΗ), 5.52 (bs, 1H), 4.54 (t, J = 6.6 Ηζ, ΙΗ), 4.15 (d, J = 2.1 Ηζ , ΙΗ), 4.03 (d, J = 2.7 Hz, 2H), 3.80 (bs, 1H), 3.66 (s, 3H), 3.40 (bs, 1H), 3.32 (d, J = 7.8 Ηζ, ΙΗ), 3.16 (d, J = 11.7 Ηζ, ΙΗ), 2.82-2.61 (m, 3H), 2.29 (s, 3H), 2.20 (s, 3H), 2.01 (s, 3H), 1.99 (s, 3H), 1.80 ( ddJ ^ I ^ O Hz, J ₂ =

15.9 Ηζ, ΙΗ), 0.62 (d, J = 6.0 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 178.5, 171.9,168.7, 146.7,144.5,142.6,140.6,140.3,

136.3, 131.0, 129.9, 128.9, 126.7, 124.4, 120.9, 120.6,117.7,116.6,112.7,111.9,

101.4, 60.4, 58.7, 57.5, 56.1, 55.7, 55.1, 53.3, 41.4, 38.8, 26.3, 24.4, 20.2, 18.1,15.3,

9.2.

ESI-MS m / z: Calcd. For CuhUsNeOrS: 726.3. Found (M + H) ⁺ : 727.3.

Example 28

NHCSNnrn

To a solution of 44 (120 mg, 0.165 ml) in dioxane (0.9 ml) was added

5.3N hydrochloric acid / dioxane (1.8 ml) and the reaction mixture was stirred · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · At 23 ° C for 2.5 hours. Dichloromethane (10 ml) and water (5 ml) were then added to the reaction mixture and the organic layer was decanted. The aqueous phase was basified with saturated aqueous sodium bicarbonate solution (20 ml) (pH 8) at 0 ° C and then extracted with dichloromethane (2x15 ml). The combined organic extracts were dried over sodium sulfate and dried.

were concentrated in vacuo to give 45 (75 mg, 87%) as a white solid, which was used in subsequent reactions without further purification.

Rf: 0.23 (EtOAc 5: 1).

Ή NMR (300 MHz, CDCl 3): δ 6.43 (s, 1H), 5.94 (d, J = 1.2 Ηζ, ΙΗ), 5.87 (d, J = 1.2 Ηζ, ΙΗ), 4.10 (d, J = 2.1 Ηζ. ΙΗ), 3.98 (d, J = 2.4 Hz, 2H), 3.91 (bs, 1H), 3.89 (s, 3H), 3.34-3.25 (m, 2H), 3.05 (dd, 4 = 1.8 Hz, 4 = 8.1 Hz, 1H), 2.80-2.73 (m, 3H), 2.46 (d, J = 18 Ηζ, ΙΗ), 2.30 (s, 3H), 2.28 (s, 3H), 2.20 (s, 3H), 1.98 (s , 3H), 1.79 (dd, J, = 12.6 Hz, 4 = 16.2 Ηζ, ΙΗ).

^{, 3} C NMR (75 MHz, CDCl 3): δ 168.7, 146.7, 144.4,142.9, 140.4,130.4, 128.9,121.1,

120.8, 117.8,116.8,113.6, 111.5,101.4, 67.6,60.5, 59.8, 58.4, 56.6, 55.8, 55.3,43.6,

41.8,31.3,25.6,20.2, 15.6,9.2.

ESI-MS m / z: Calcd. For CaNagI6Cl2: 520.58. Found (M + H) ⁺ : 521.3.

Example 29

Pht Anh „CH ₂ CI ₂ , CDI

17h. 23 ° C

phthalicidin

To a solution of 45 (10 mg, 0.02 ml) in dichloromethane (0.4 ml) was added

phthalic anhydride (2.84 mg, 0.02 ml) and the reaction mixture was stirred at

23 ° C for 2 hours. Carbonyldiimidazole (0.5 mg, 0.003 ml) was then added and the reaction mixture was stirred at 23 ° C for 7 hours. Carbonyldiimidazole (2.61 mg, 0.016 ml) was then added and the reaction stirred at 23 ° C for a further 17 hours. The solution was diluted with dichloromethane (10 ml) and washed with 0.1N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate,

filtered and the solvent removed under reduced pressure. The residue was purified by flash chromatography (RP-18, CH ₃ CN: H ₂ O 60:40) to give phthalacidin (11.7 mg, 93%) as a white solid.

Rf: 0.37 (CHsChLHaO 7: 3, RP-18).

Ή NMR (300 MHz, CDCl 3): δ 7.72-7.68 (m, 2H), 7.67-7.63 (m, 2H), 6.38 (s, 1H), 5.69 (d, J = 1.2 Ηζ, ΙΗ), 5.64 (d , J = 1.2 Ηζ, ΙΗ), 5.30 (bs, 1H), 4.25-4.21 (m, 2H), 4.02 (d, J = 2.1 Hz, 1H), 3.64-3.62 (m, 5H), 3.33 (d. J = 8.4 Ηζ, ΙΗ), 3.21-3.16 (m, 1H), 3.02 (dd, Jt = 8.1 Hz, Jz = 18 Hz, 1H), 2.76 (dd, Ji = 1.8 Hz, J ₂ = 15.6 Ηζ, ΙΗ ), 2.63 (d, J = 17.7 Hz, 1H), 2.29 (S, 3H), 2.28 (s, 3H), 2.21 (s, 3H), 2.0 (s, 3H), 1.73 (dd, Ji = 12.0 Hz) , J ₂ = 15.3 Hz, 1H).

¹³ C NMR (75 MHz, CDCl 3): δ 168.5,167.6,146.2,144.2,142.5,141.0, 140.5,133.4,

131.8, 130.7,128.2, 120.9, 120.8, 117.9, 116.4, 113.6, 101.1, 60.4,60.0, 57.0, 56.3,

55.6, 55.4,41.6,41.5, 26.5, 25.2, 20.2, 15.7,9.4.

ESI-MS m / z: Calcd for C14H26N3O6: 650. Found (M + H) ⁺ : 651.2.

Example 30

• · · · · · · · 4 · · · · · · · · · · · · · · · · · · · • · · · · · · · · · · · · · · · · · · · · · · · · · ·

To a solution of 25 (18 mg, 0.032 ml) in dimethylformamide (0.05 ml) at 0 ° C was added cat dimethylaminopyridine (0.5 mg, 0.004 ml), imidazole (5 mg, 0.08 ml) and t-butyldiphenylsilyl chloride (12.5 ml, 0.048 ml) and the reaction mixture was stirred at 23 ° C for 6 hours. Water (10 ml) at 0 ° C was added and the aqueous phase was extracted with hexane: ethyl acetate 1:10 (2x10 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash chromatography (silica, hexane ethyl acetate 3: 1) to give 26 (27 mg, 88%) as a white solid.

Rf: 0.29 (hexane: ethyl acetate 3: 1).

1 H NMR (300 MHz, CDCl 3): δ 7.61-7.58 (m, 2H), 7.42-7.28 (m, 8H), 6.71 (s, 1H), 6.19-6.02 (m, 1H), 5.78 (d, J = 1.2 Ηζ, ΙΗ), 5.64 (d, J = 1.2 Ηζ, ΙΗ), 5.40 (dd, J <= 1.2 Hz, J ₂ = 17.1 Ηζ, ΙΗ), 5.27 (dd, J, = 1.2 Hz, J ₂ = 10.2 Ηζ, ΙΗ), 5.13 (s, 2H), 4.45 (d, J =

2.4 Ηζ, ΙΗ), 4.24 (d, J = 2.1 Ηζ, ΙΗ), 4.17-4.06 (m, 3H), 3.75 (s, 3H), 3.64 (dd, J <= 2.4 Hz, J ₂ = 9.9 Ηζ. ΙΗ), 3.59 (s, 3H), 3.42-3.21 (m, 4H), 3.10 (dd, Ji = 8.1 Hz, J ₂ = 17.7 Hz, 1H), 2.70 (d, J = 17.7 Ηζ, ΙΗ), 2.33 (s, 3H), 2.26 (s, 3H), 2.11 (S, 3H), 2.08-1.89 (m, 1H), 0.87 (s, 9H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 148.5, 148.3, 148.1, 144.0, 139.0,135.6, 135.4,133.8,

133.1, 132.6, 130.5,130.3, 129.6,129.4, 127.5, 127.4, 125.1,124.3,121.6, 118.5,

117.5, 112.9, 111.7,100.8, 99.2, 74.0, 67.7, 61.5, 59.6, 59.0, 57.7, 57.1, 55.4,41D

29.6, 26.6, 25.5, 18.8, 15.8, 9.2.

ESI-MS m / z: Calculated for ¢ 47 ^ 5 ^ 0781: 801.3. Found (M + H) ⁺ : 802.3.

Example 31

* · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ···

To a solution of 26 (7 mg, 0.0087 ml) in dichloromethane (0.15 ml) at 23 ° C was added acetic acid (2.5 ml, 0.044 ml), (PPIb ^ PdCh (0.5 mg, 6.96 × 10 * ⁴ ml) and BusSnH. (3.5 ml, 0.013 ml) The reaction mixture was stirred at this temperature for 1 hour. The solution was diluted with a mixture of hexane: ethyl acetate 5: 1 (0.5 ml) and poured into a pressure column (silica, gradient hexane ethyl acetate from 5: 1 to 1: 1) to give ET-11 (5 mg, 75%) as a white solid.

Rf: 0.36 (hexane: ethyl acetate 1: 5, silica).

1 H NMR (300 MHz, CDCl 3): δ 7.56 (m, 2H), 7.41-7.25 (m, 8H), 6.67 (s, 1H), 5.72 (d, J = 1.0 Ηζ, ΙΗ), 5.58 (d. J = 1.0 Ηζ, ΙΗ), 5.51 (s, 1H), 5.38 (d, J = 5.75 Ηζ, ΙΗ), 5.16 (d, J = 5.7 Ηζ, ΙΗ), 4.57 (d, J = 2.9 Ηζ, ΙΗ) , 4.21 (m, 1H), 4.09 (m, 1H), 3.72 (s, 3H), 3.71 (s, 3H), 3.68 (dd, 4 = 2.1 Hz, 4 = 10.4 Hz, 1H), 3.38-3.26 ( m, 3H), 3.11 (dd, 4 = 2.5 Hz, 4 = 15.7 Ηζ, ΙΗ), 3.01 (dd, 4 = 8.9 Hz, 4 = 17.9 Ηζ, ΙΗ), 2.70 (d, J = 17.9 Ηζ, ΙΗ) , 2.31 (s, 3H), 2.25 (s, 3H), 2.06 (s, 3H), 1.89 (dd, 4 = 12.1 Hz, 4 = 15.7 Ηζ, ΙΗ), 0.9 (s, 9H). ¹³ C NMR (75 MHz, CDCl 3): δ 149.0, 147.4,145.3, 144.3,136.3,135.7, 135.4, 133.2,

130.9, 130.5, 129.6, 129.5, 127.5, 125.0, 118.6, 112.5, 112.1, 105.7,100.5, 99.8, 68.5,

61.5, 59.7, 58.8, 57.7, 56.9, 56.5, 55.4, 41.7, 26.6, 26.2, 25.5, 18.9, 15.8, 14.2, 8.7. ESI-MS m / z: Calcd. For CuHsiNaCbSi: 761. Found (M + H) ⁺ : 762.

Example 32

A solution of 2 (3.0 g, 5.46 ml) and phenylisothiocyanate (3.92 ml, 32.76 ml) in dichloromethane (27 ml) was stirred at 23 ° C for 1.5 hours. The reaction mixture was partitioned into dichloromethane (10 ml) and water (5 ml). Organ95

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · The layer was dried over sodium sulfate, filtered and concentrated. The residue was purified by flash chromatography (silica, gradient hexane to 2: 3 hexane: ethyl acetate) to give 3 (3.29 g, 88%) as a yellow solid.

Rf: 0.27 (CAN: water 3: 2, RP-C18).

Ή NMR (300 MHz, CDCl3): δ 7.77 (bs, 3H), 7.41-7.11 (m, 5H), 6.65 (d, 1H), 6.29 (s, 1H), 5.6-5.5 (m, 1H), 4.19 -4.14 (m, 2H), 4.08 (d, 1H), 3.92 (s, 3H), 3.87-3.65 (m, 6H), 3.77 (s, 3H), 3.37-2.98 (m, 8H), 2.50 (d , 1H), 2.31 (s, 3H), 2.20 (s, 3H), 1.96 (d, 1H), 1.87 (s, 3H), 1.81-1.75 (m, 1H), 0.96 (d, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 185.7, 180.9,178.9,172.0, 155.7,147.1,143.2,142.4, 136.0, 135.1,130.5, 129.9, 129.3,128.5, 126.9,124.4, 120.2, 117.4, 116.3. 77.1, 60.9,

58.6, 56.2, 55.8, 55.0, 54.6, 53.5, 41.7, 40.3, 25.1, 24.5, 18.4, 15.8, 8.7.

ESI-MS m / z: Calcd. For C 23 H 27 F 3 O 5 S: 684.8. Found (M + H) ⁴ : 685.2.

Example 33

PhHNSCHN

HjN

A solution of 3 (0.143 g, 0.208 ml) in 6.5M hydrochloric acid / dioxane (150 ml) was stirred at 23 ° C for 6 hours. Toluene (3 ml) was then added to the reaction mixture and the organic layer was decanted. The residue was partitioned between saturated aqueous sodium bicarbonate solution (3 ml) and chloroform (3x3 ml). The organic layers were dried and concentrated to give the title compound as a mixture of 4 and 6 (4: 6 90:10), which slowly cyclized to 6 on standing.

Rf: 0.4 (ethyl acetate: methanol 5: 1, silica).

1 H NMR (300 MHz, CDCl 3): δ 6.45 (s, 1H), 4.16 (m, 1H), 4.02 (d, 1H), 3.96 (s, 3H),

3.79 (m, 2H), 3.75 (S, 3H), 3.35 (m, 1H), 3.20-3.00 (m, 3H), 2.87 (d, 1H), 2.75 (d, 1H).

2.43 (d, 1H), 2.34 (s, 3H), 2.30 (s, 3H), 1.93 (s, 3H), 1.72-1.5 (m, 3H).

ESI-MS m / z: Calcd. For C26H30N4O5: 478.5. Found (M + H) ⁺ : 479.2.

Example 34

A solution of 3 (0.143 g, 0.208 ml) in 6.5M hydrochloric acid / dioxane (150 ml) was stirred at 23 ° C for 1 hour. Evaporation of the solvent gave a residue which was purified by flash chromatography (ethyl acetate / methanol / triethylamine 100: 25: 0.1) to give 6 (80 mg, 83%) as a yellow solid.

Rf: 0.26 (ACI: water 3: 2, RP-C18).

1 H NMR (500 MHz, CDCl 3): δ 6.46 (s, 1H), 5.09 (bs, 1H), 4.67 (dd, Ji = 18.3 Hz, J ₂ =

7.8 Ηζ, ΙΗ), 4.24 (d, 1H), 4.16 (s, 3H), 3.93 (d, J = 2.7 Ηζ, ΙΗ), 3.8 (m, 2H), 3.77 (s, 3H), 3.45 (m. 2H), 3.08 (dd, 4 = 17.9 Hz, 4 = 3.6 Hz, 1H), 2.78 (m, 1H), 2.55 (d, 1H),

2.3 (m, 1H), 2.3 (s, 3H), 2.28 (s, 3H), 1.90 (s, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 186.2, 162.1, 154.9, 146.9, 145.3, 143.0, 130.1, 129.4,

128.1, 125.0, 121.4, 116.4, 116.2, 66.6, 60.7, 60.7, 60.1, 59.6, 58.8, 55.6, 54.9, 41.9, 25.3, 24.7, 15.7, 8.9.

ESI-MS m / z: Calcd. Found (M + H) ⁺ : 461.1.

Example 35

To a solution of 3 (2.38 g, 3.47 ml) in dioxane (5 ml) was added 5.3M hydrochloric acid in dioxane (34 ml) and the reaction mixture was stirred at 23 ° C for 45 minutes. Acetic anhydride (51 ml, 539.5 ml) was then added and the reaction stirred for 4 hours. The reaction mixture was cooled to 0 ° C and partitioned between aqueous saturated sodium carbonate solution (300 ml) and ethyl acetate (300 ml) at this temperature. The organic phase was dried over sodium sulfate, filtered and concentrated. The residue was purified by flash chromatography (silica, gradient dichloromethane to dichloromethane. Ethyl acetate 1: 2) to give 5 (1.75 g, 97%) as a yellow solid.

Rf: 0.53 (ACI: water 3: 2, RP-C18).

1 H NMR (300 MHz, CDCl 3): δ 6.51 (S, 1H), 5.98 (bs, 1H), 4.84 (dd, 1H), 4.17 (d, 1H), 4.00 (d, 1H), 3.99 (s. 3H), 3.85 (bs, 1H), 3.81 (m, 1H), 3.74 (s, 3H), 3.70 (d, 1H), 3.23 (m, 1H), 3.11 (dd, 1H), 3.09 (m, 1H) ), 2.93 (m, 2H), 2.44 (d, 1H), 2.67 (s, 3H), 2.25 (s, 3H), 1.70 (S, 3H), 1.60-1.50 (m, 2H), 1.29 (s. 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 185.9, 180.8, 169.9,160.2, 156.2,147.0, 143.1, 140.4,

136.1, 130.6, 129.6, 127.9, 120.4, 117.2, 61.0, 60.7, 58.6, 56.1, 55.7, 55.1, 54.3, 41.8,

41.1, 25.7, 23.9,22.2, 15.7, 8.7.

ESI-MS m / z: Calcd. For C28H32N4O6: 520.6l Found (M + H) ⁺ : 521.1.

Example 36

To a solution of 5 (1.75 g, 3.36 ml) in dichloromethane (17 ml) at 0 ° C was added diisopropylethylamine (11.71 ml, 67.23 ml), dimethylaminopyridine (20 mg, 0.17 ml) and bromomethyl ether (4.11 ml, 50.42 ml). After 6 hours at 23 ° C, the reaction mixture was partitioned between aqueous saturated sodium bicarbonate solution (25 ml) and dichloromethane (50 ml). The organic layer was dried over sodium sulfate. The solvent was removed under reduced pressure. The crude product was purified by flash chromatography (RP-18, CHaCN / water 1: 1) to give 7 (1.32 g, 70%) as a yellow solid.

Rf: 0.34 (AC N: water 2: 3, RP-C18).

Ή NMR (300 MHz, CDCl ₃ ): δ 6.74 (s, 1H), 5.14 (s, 2H), 4.82 (m, 1H), 4.22 (d, 1H), 4.00 (s, 3H), 4.0 (m. 1H), 3.83 (m, 2H), 3.7 (s, 3H), 3.58 (s, 3H), 3.4 (m, 1H), 3.2-2.95 (m, 6H), 2.43 (d, 1H), 2.37 (S , 3H), 2.22 (S, 3H), 1.890 (S, 3H), 1.5-1.4 (m, 2H), 1.31 (s, * 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 185.9, 180.7,169.6, 156.2, 148.9,148.5, 140.3,136.2, 131.3, 130.1, 127.7,124.6,123.7, 117.3, 99.5,99.2, 60.9, 59.7, 58.8, 57.7 , 56.4, 55.7, 55.0, 54.2, 51.0, 41.6, 41.0, 40.5, 25.5, 23.9, 22.3,19.3, 15.6, 14.6, 8.6. ESI-MS m / z: Calcd. For C30H36N4O7: 520.6 Found (M + H) ⁺ : 565.3.

Example 37

To a solution of 7 (0.37 g, 0.65 ml) in methanol (74 ml) at 0 ° C was added a 1M solution of sodium hydroxide (130 ml). The reaction mixture was dissolved.

A, stirred for 15 minutes and then fixed at 0 ° C with 6M hydrochloric acid to pH 5. The reaction mixture was extracted with ethyl acetate (3 x 50 ml) and the combined organic layers dried over sodium sulfate and concentrated under vacuum. The residue was purified by flash chromatography (RP-18, CHsCN / water 1: 1) to give 8 (232 mg, 65%) as a yellow oil.

Rf: 0.5 (ACOD 3: 2, RP-C18).

1 H NMR (300 MHz, CDCl 3): δ 6.75 (s, 1 Η), 5.15 (s, 2H), 4.86 (m, 1H), 4.26 (d, 1 Η), 4.01 (d, 1H), 3.88- 3.81 (m, 2H), 3.70 (s, 3H), 3.58 (S, 3H), 3.39 (m, 1H), 3.27-3.21 (m, 1H), 3.18-3.08 (m, 2H), 3.03-2.97 ( m, 1H), 2.47 (d, 1H), 2.37 (s, 3H), 2.22 (s, 3H), 1.90 (S, 3H), 1.57-1.46 (m, 2H), 1.33 (s, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 185.3, 180.6, 175.9,170.1, 151.5,148.9,148.6, 143.3,

133.7, 131.5, 129.9, 124.7, 123.5, 117.1, 117.0, 99.2, 59.8, 58.7, 57.8, 56.3, 55.3, ® 54.9, 54.3, 41.5, 40.7, 29.6, 25.5, 24.4, 22.2, 20.7, 15.7, 8.0.

ESI-MS m / z: Calcd. For C29H34N4O7: 550.6. Found (M + H) ⁺ : 551.2.

CICH ₂ Br / Cs2CO ₃

DMF / 90 ° C

100 · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

To a degassed solution of compound 8 (240 mg, 0.435 ml) in dimethylformamide (30 ml) was added 10% palladium on carbon (48 mg) and the reaction mixture was stirred under a hydrogen atmosphere (atmospheric pressure) for 1 hour. The reaction mixture was filtered through a pad of selite under argon in a Schlenk tube as a colorless solution containing anhydrous cesium carbonate (240 mg, 0.739 ml). Bromochloromethane (0.566 ml, 8.71 ml) was then added. The tube was sealed and stirred at 90 ° C for 3 hours. The reaction mixture was cooled and filtered through Celite and washed with dichloromethane. The organic layer was concentrated and dried over sodium sulfate to give 9 as a brown oil, which was used in the next step without further purification.

Rf: 0.36 (hexane: ethyl acetate 1: 5, silica).

Ή NMR (300 MHz, CDCl3): 56.71 (s, 3H), 5.89 (d, 1H), 5.81 (d, 1H), 5.63 (bs, 1H), 5.33 (d, 1H), 5.17 (d, 1H) , 4.97 (m, 1H), 4.20 (d, 1H), 4.09 (m, 1H), 3.99 (m, 1H), 3.68 (m, 1H), 3.65 (s, 6H), 3.59-3.47 (m, 4H) ), 3.37-3.27 (m, 2H), 3.14-2.97 (m, 2H), 2.62 (d, 1H), 2.32 (S, 3H), 2.20 (s, 3H), 2.08 (s, 3H), 1.72 ( m, 1H), 1.36 (S, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 169.8, 149.1,147.4,145.3, 136.2,130.9, 130.8,125.0,

122.9,117.7, 112.6, 111.8, 106.4, 100.8, 99.8, 59.8, 58.9, 57.7, 56.6, 56.4, 55.5, 55.2,

41.6, 40.1, 29.6, 25.9, 25.0, 22.6, 15.6, 8.8.

ESI-MS m / z: Calcd. For CaoHgeN 6 Si: 564.6. Found (M + H) ⁺ : 563.3.

Example 39

AllylBr / Cs2CO ₃

DMR23 * C

A)

101

····· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

To a flask containing 9 (245 mg, 0.435 ml) in dimethylformamide (4 ml) at 0 ° C was added cesium carbonate (425 mg, 1.30 ml) and allyl bromide (376 ml, 4.35 ml) and the reaction mixture was stirred at 23 ° C for 1 hour. The reaction mixture was filtered through a filter pad and partitioned between dichloromethane (25 ml) and water (10 ml). The organic phase was dried over sodium sulfate and concentrated under reduced pressure to give a residue which was purified by flash chromatography (silica, chloroform: ethyl acetate 1: 2) to give 10 (113 mg, 43%) as a yellow butter.

Rf: 0.36 (hexane: ethyl acetate 1: 5).

1 H NMR (300 MHz, CDCl 3): δ 6.74 (S, 1H), 6.3-6.0 (m, 1H), 5.94 (d, 1H), 5.87 (d, 1H), 5.43-5.36 (m, 2H). 5.22 (s, 2H), 5.00 (m, 1H), 4.22 (m, 1H), 4.17-4.01 (m, 1H), 3.98 (m, 2H), 3.71-3.67 (m, 1H), 3.69 (s. 6H), 3.62-3.51 (m, 3H), 3.58 (s, 3H), 3.39-3.37 (m, 1H), 3.31-3.26 (m, 3H), 3.09 (dd, 1H), 2.56 (d, 1H) , 2.36 (s, 3H), 2.21 (s, 3H), 2.11 (s, 3H), 2.24-2.10 (m, 1H), 1.82-1.73 (m, 1H), 1.24 (bs, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 169.4, 148.8, 148.3, 139.1,133.7,130.9,130.3,125.2,

120.2, 117.7, 113.1, 112.6, 101.3,99.3, 74.1, 59.7, 59.3, 57.8, 57.0, 56.1, 56.1, 55.2,

41.6, 41.0, 40.9, 29.7, 26.3, 22.5, 15.6, 9.3.

ESI-MS m / z: Calculated for C30H10IDO: 604.7. Found (M + H) *: 605.3.

Example 40

oA

To a solution of 9 (22 mg, 0.039 ml) in dichloromethane (0.2 ml) at 0 ° C

102 were added acetyl chloride (2.79 ml, 0.039 ml) and pyridine (3.2 ml, 0.039 ml). The reaction mixture was stirred for 1 hour and then dissolved

The mouth was diluted with dichloromethane (10 ml) and washed with 0.1N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and filtered.

the solvent was removed under reduced pressure to give 46 (22 mg, 93%) as a white solid.

Rf: 0.4 (hexane: ethyl acetate 1: 5).

Ή NMR (300 MHz, CDCl3): δ 6.74 (s, 1H), 5.97 (d, J = 0.9 Hz, 1H), 5.91 (d, J = 0.9 Ηζ, ΙΗ), 5.12 (d, J = 5.7 Hz. 2H), 5.04 (d, J = 5.7 Hz, 1H), 4.90 (t, J = 6 Ηζ, ΙΗ), 4.17 (d, J = 2.7 Ηζ, ΙΗ), 4.05 (d, J = 2.7 Ηζ, ΙΗ) , 4.01 (bs, 1H), 3.71 (s, 3H), 3.57 (s, 3H), 3.503.44 (m, 2H), 3.38-3.36 (m, 1H), 3.30-3.26 (m, 1H), 3.00 (dd, J, = 7.8 Hz, 4 = 18.0 Hz, 1H), 2.79 (d, J = 12.9 Ηζ, ΙΗ), 2.60 (d, J = 18.0 Ηζ, ΙΗ), 2.35 (s, 3H), 2.32 ( s, 3H), 2.21 (s, 3H), 2.00 (s, 3H), 1.68 (dd, 4 = 11.7 Hz, 4 = 15.6 Ηζ, ΙΗ).

ESI-MS m / z: Calcd for C23H36N3O3: 606.67. Found (M + H) ⁺ : 607.3.

Example 41

To a solution of 46 (8 mg, 0.013 ml) in dioxane (0.1 ml) was added 5.3N hydrochloric acid / dioxane (0.5 ml) and the reaction mixture was stirred at 23 ° C for 1 hour. The solution was then diluted with dichloromethane (5 ml) and washed with 0.1N hydrochloric acid (3 ml). The organic layer

103

• · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · The mixture was dried over sodium sulfate, filtered and the solvent removed under reduced pressure to give 47 (5 mg, 70%) as a white solid.

Rf: 0.4 (hexane: ethyl acetate 1: 5).

¹ H NMR (300 MHz, CDCl 3): δ 6.51 (s, 1H), 5.97 (d, J = 1.2 Ηζ, ΙΗ), 5.91 (d, J = 1.2 Ηζ, ΙΗ), 4.97 (bs, 1H), 4.11 (bs, 1H), 4.04-4.02 (m, 2H), 3.75 (s, 3H), 3.65 (d, J = 2.1 Hz, 2H), 3.56-3.30 (m, 2H), 3.04 (dd, 4 = 7.5) Hz, Ja = 18.0 Hz, 1H), 2.80 (d, J = 14.4 Hz, 1H), 2.59 (d, J = 18.3 Ηζ, ΙΗ), 2.33 (s, 3H), 2.24 (s, 3H), 2.00 ( s, 3H), 1.76 (dd, 4 = 12.0 Hz, J _z = 15.9 Ηζ, ΙΗ), 1.33 (s, 3H), 1.25 (s, 3H).

ESI-MS m / z: Calcd. For C30H34N4O7: 562.61. Found (M + H) ⁴ : 563.3.

Example 42

To a solution of 45 (10 mg, 0.0192 ml) in dichloromethane (0.3 ml) at 0 ° C was added isovaleryl chloride (2.34 ml, 0.0192 ml) and pyridine (1.55 ml, 0.0192 ml). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane: ethyl acetate 1: 2) to give 48 (11 mg, 95%) as a white solid.

Rf: 0.12 (hexane: ethyl acetate 1: 2).

104 ¹ H NMR (300 MHz, CDCl 3): δ 6.50 (s, 1H), 5.98 (d, J = 1.5 Hz, 1H), 5.91 (d, J = 1.5 Ηζ, ΙΗ), 5.75 (s, 1H). 5.02 (t, J = 5.4 Ηζ, ΙΗ), 4.10 (d, J = 1.5 Ηζ, ΙΗ), 4.06 (d, J = 2.7 Ηζ, ΙΗ), 4.02 (d, J = 2.7 Ηζ, ΙΗ), 3.77 ( s, 3H), 3.76 ^ .71 (m, 1H), 3.86-3.28 (m, 3H), 3.04 (dd, Ji = 8.1 Ηζ, 4 = 18.3 Ηζ, ΙΗ), 2.78 (d, J = 15.9 Ηζ. ΙΗ), 2.55 (d, J = 18 Ηζ, ΙΗ), 2.32 (S, 6H), 2.26 (S, 3H), 1.98 (s, 3H), 1.84-1.68 (m, 2H), 1.36 (d, J = 7.2 Hz, 2H), 0.69 (d, J = 6.6 Hz, 3H), 0.62 (d, J = 6.6 Hz, 3H).

ESI-MS m / z: Calculated for SNF: 604.69. Found (M + H) ⁺ : 605.3.

Example 43

Me decanoyl chloride, py, CH ₂ Cl ₂ , 0 ° C. 1h

OMe

I

To a solution of 45 (10 mg, 0.0192 ml) in dichloromethane (0.3 ml) at 0 ° C was added isovaleryl chloride (3.98 mi, 0.0192 ml) and pyridine (1.55 ml, 0.0192 ml). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane: ethyl acetate 1: 2) to give 49 (12.4 mg, 96%) as a white solid.

Rf: 0.7 (ethylacetammethanol 10: 1).

¹ H NMR (300 MHz, CDCl 3): δ 6.50 (s, 1H), 5.98 (d, J = 1.5 Ηζ, ΙΗ), 5.91 (d, J = 1.5 Ηζ, ΙΗ), 5.73 (s, 1H), 5.08 (t, J = 5.4 Ηζ, ΙΗ), 4.10 (d, J = 1.5 Ηζ, ΙΗ), 4.05 (m, 1H), 4.01

105

• ft ··· • ft ft ft · • ft ♦ ft ft ft • ft ft ft • ft • · ft · • · ft • ft • · · ft ft ft ft ft • ft ft ft ft «· · · · · • ft • ft

(m, 1H), 3.76 (s, 3H), 3.65-3.61 (m, 1H), 3.40-3.27 (m, 3H), 3.03 (dd, Ji = 8.1 Hz, J ₂ =

18.6 Ηζ, ΙΗ), 2.78 (d, J = 13.2 Ηζ, ΙΗ), 2.57 (d, J = 18.8 Ηζ, ΙΗ), 2.32 (s, 6H), 2.31 (s, 3H), 2.25 (s. 3H), 1.99 (s, 3H), 1.79 (dd, Ji »12.0 Hz, J ₂ = 16.5 Ηζ, ΙΗ), 1.73-1.42 (m, 4H), 1.33-1.18 (m, 10 OH), 1.03 (m. 2H), 0.87 (t, J = 6.6 Hz, 3H).

ESI-MS m / z: Calcd. For C38H50N4O7: 674.83. Found (M + H) ⁺ : 675.5.

Example 44

50

To a solution of 45 (14.5 mg, 0.0278 ml) in dichloromethane (0.3 ml) at 0 ° C was added trans-3-trifluoromethylsinamoyl chloride (4.76 ml, 0.0278 ml) and pyridine (2.25 ml, 0.0278 ml). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane: ethyl acetate 1: 1) to give 50 (18.7 mg, 94%) as a white solid.

Rf: 0.64 (ethyl acetate: methanol 5: 1).

1 H NMR (300 MHz, CH 3 OD): δ 7.74-7.55 (m, 4H), 7.23 (d, J = 16.0 Ηζ, ΙΗ), 6.34 (s, 1H), 6.12 (d, J = 16.0 Ηζ, ΙΗ) , 6.07 (d, J = 0.9 Ηζ, ΙΗ), 5.96 (d, J = 0.9 Ηζ, ΙΗ), 4.39 (d, J = 2.4 Ηζ, ΙΗ), 4.07-4.05 (m, 1H), 3.81 (bs. 1H), 3.46-3.51 (m, 3H), 3.42 (s, 3H), 3.09

106 »· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·· ··········· (brd, J = 12.0 Ηζ, ΙΗ), 2.94-2.85 (m, 2H), 2.74 (d, J = 18.3 Ηζ, ΙΗ), 2.38 (s. 3H), 2.23 (s, 3H), 2.02 (S, 3H), 1.80 (s, 3H), 1.84-1.75 (m, 1H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 168.7,165.3,146.5,144.7, 142.6,140.6, 138.0,135.9, 131.0, 130.9, 129.1, 128.6, 125.8, 125.7,124.5, 124.4, 122.7, 121.2,117.8, 116.5, 113.0, 112.0, 101.7, 60.4, 59.1, 56.5, 56.4, 55.6, 55.3, 41.8, 40.3, 26.6, 25.1, 20.3, "

15.4, 9.3.

ESI-MS m / z: Calcd. For CssHsqN4: 674.83. Found (M + H) ⁺ : 675.5.

Example 45

Isovaleryl chloride. Py, CH ₂ Ci ₂ , 0 ° C, 1h

To a solution of 43 (33 mg, 0.0557 ml) in dichloromethane (0.4 ml) at 0 ° C was added isovaleryl chloride (6.79 ml, 0.0557 ml) and pyridine (4.5 ml, 0.0557 ml). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane. Ethyl acetate 1: 2) to give 51 (34 mg, 91%) as a white solid.

Rf: 0.09 (hexane: ethyl acetate 1: 2).

1 H NMR (300 MHz, CDCl 3): δ 6.46 (s, 1H), 6.10 (bs, 1H), 5.99 (d, J = 0.9 Hz, 1H),

5.90 (d, J = 0.9 Hz, 1H), 5.30 (t, J = 6.0 Ηζ, ΙΗ), 4.10-4.05 (m, 3H), 3.81 (bs, 3H), 3.74

107

(s, 3H), 3.54 (bs, 1H), 3.38-3.36 (m, 1H), 3.29-3.21 (m, 1H), 3.00 (dd, J, = 8.0 Hz, J ₂ = 18.0 Ηζ, ΙΗ). 2.25 (s, 3H), 2.20 (s, 3H), 2.00 (s, 3H), 1.95-1.90 (m, 3H), 0.87 (d, J, =

6.6 Hz, 6H), 0.76 (d, J = 6.0 Hz, 3H).

ESI-MS m / z: Calculated for C14H23N3Oa: 675.77. Found (M + H) ⁺ : 676.3.

Example 46

To a solution of 43 (33 mg, 0.0557 ml) in dichloromethane (0.4 ml) at 0 ° C was added trans-3-trifluoromethylsinamoyl chloride (9.52 ml, 0.0557 ml) and pyridine (4.5 ml, 0.0557 ml). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane: ethyl acetate 1: 2) to give 52 (40 mg, 92%) as a white solid.

Rf: 0.21 (hexane: ethyl acetate 1: 2).

Ή NMR (300 MHz, CDCl3): δ 7.74-7.47 (m, 4H), 6.49 (s, 1H), 6.40 (d, J = 15.6 Hz, 1H), 6.00 (d, J = 1.5 Ηζ, ΙΗ). 5.90 (d, J = 1.5 Ηζ, ΙΗ), 5.47 (t, J = 6.0 Ηζ, ΙΗ), 4.12-4.09 (m, 3H), 3.93 (bs, 1H), 3.71 (s, 3H), 3.59-3.58 (m, 1H), 3.38 (d, J = 7.8 Ηζ, ΙΗ), 3.29 (d, J = 12.0 Hz, 1H), 3.00 (dd, Ji = 8.1 Hz, ^ = 18.3 Hz, 1H), 2.79-2.78 (m, 1H), 2.65 (d, J =

108

18.3 Ηζ, ΙΗ), 2.29 (S, 6H), 2.28 (s, 3H), 2.22 (s, 3H), 1.84-1.80 (m, 1H), 0.85-0.84 (m, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 171.9, 168.8,164.4,146.9, 144.6,143.0, 140.5,140.5,

139.3, 135.7, 131.1, 131.0, 129.4,129.1, 126.0,124.1, 124.0,122.4, 121.1, 120.7,

120.6, 117.7, 116.9, 112.8, 112.0, 101.6, 60.6, 59.3, 57.1, 56.3, 55.9, 55.2, 49.0, 41.7,

39.9, 26.5, 25.1, 20.2, 18.4, 15.7,9.3.

ESI-MS m / z: Calculated for C 22 H 27 N 5 O 3: 789.8. Found (M + H) ⁺ : 790.3.

Example 47

To a solution of 43 (10 mg, 0.0169 ml) in dichloromethane (0.2 ml) at 23 ° C was added trifluoroacetic anhydride (2.38 μΙ, 0.0169 ml). The reaction mixture was stirred for 5 hours and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane: ethyl acetate 3: 2) to give 55 (10.7 mg, 93%) as a white solid.

Rf: 0.57 (ethyl acetate. Methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.45 (s, 1H), 6.00 (d, J = 1.2 Ηζ, ΙΗ), 5.90 (d, J = 1.2 Ηζ, ΙΗ), 5.87 (bs, 1H), 5.32 (bs.lH), 4.12 (d, J = 2.1 Ηζ, ΙΗ), 4.08 (d, J = 1.8 Ηζ, ΙΗ), 3.78-3.56 (m, 3H), 3.72 (s, 3H), 3.40 (d. J = 8.1 Ηζ, ΙΗ), 3.25 (d, J = 9.3 Hz, 1H), 3.00

I

109

(dd, Ji = 8.4 Hz, 02 = 18.0 Hz, 1H), 2.77 (dd, Ji = 2.1 Hz, J ₂ = 15.9 Hz, 1H), 2.68 (d, J =

18.6 Ηζ, ΙΗ), 2.30 (s, 3H), 2.28 (s, 3H), 2.22 (S, 3H), 2.00 (s, 3H), 1.75 (dd, Ji = 11.4

Hz, J ₂ = 15.9 Hz, 1H), 0.69 (d, J = 6.3 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 170.1,168.6,156.0, 147.0, 144.6,143.0,140.6,140.4,

131.0, 129.4, 120.9,120.7, 117.6, 116.8, 112.4, 112.1, 101.6, 60.5, 59.0, 57.1, 56.3,

55.6, 55.2, 48.7, 41.6, 39.4, 26.5, 24.9, 20.2, 17.8, 15.4, 9.2.

ESI-MS m / z: Calculated for 687.63. Found (M + H) ⁺ : 688.66.

Example 48

To a solution of 19 (11 mg, 0.0169 ml) in dichloromethane (0.2 ml) at 23 ° C was added trifluoroacetic anhydride (2.38 ml, 0.0169 ml). The reaction mixture was stirred for 5 hours and then the solution was diluted with dichloromethane (5 ml) and washed with 0.1 N hydrochloric acid (3 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, hexane: ethyl acetate 3: 2) to give 54 (10.7 mg, 93%) as a white solid.

Rf: 0.6 (ethyl acetate-methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): 57.33 (d, J = 6.3 Ηζ, ΙΗ), 6.04 (m, 1H), 5.95 (d, J = 1.5 Ηζ, ΙΗ), 5.84 (d, J = 1.5 Ηζ. ΙΗ), 5.32 (m, 2H), 5.21 (m, 1H), 4.11 (m, 4H), 3.73 (s, 3H),

110 ····· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·

3.64 (m, 2H), 3.51 (m, 1H), 3.37 (d, J = 7.8 Ηζ, ΙΗ), 3.22 (m, 2H), 3.03 (dd, 4 = 8.1 Hz, 4 = 18.3 Ηζ, ΙΗ). 2.60 (d, J = 18.3 Ηζ, ΙΗ), 2.29 (s, 3H), 2.24 (s, 3H), 2.08 (S, 3H), 1.86 (dd, 4 = 12 Hz, 4 = 16.2 Ηζ, ΙΗ). 0.82 (d, J = 7.2 Hz, 3H).

&Lt; 1 &gt; C NMR (75 MHz, CDCl3): δ 170.0,156.0, 148.4, 147.1, 144.3,143.0, 138.7,133.8,

130.5, 129.4, 120.6, 120.4, 111.6, 117.5, 117.0, 113.5, 112.5, 112.4, 101.1, 74.1, 66.8,

60.4, 59.3, 56.9, 56.6, 56.3, 55.4,48.7, 41.6,40.1, 26.2, 25.0,17.6, 15.4, 9.1. ESI-MS m / z: Calcd. For C 25 H 16 FaN 5 Cl 2: 685.69. Found (M + H) *: 686.3.

Example 49

55

To a solution of 54 (100 mg, 0.415 ml) in dichloromethane (4 ml) at 23 ° C was added acetic acid (40 ml), (PPl 3) rPcCl 2 (8.4 mg, 0.012 ml) and BibSnH (157 ml, 0.56 ml). . The reaction mixture was stirred for 2 hours at this temperature and then poured into a pressure column (silica, gradient hexane to hexane: ethyl acetate 2: 1) to give 55 (90 mg, 96%) as a white solid. .

Rf: 0.6 (hexane: ethyl acetate 1: 2).

Ή NMR (300 MHz, CDCl 3): δ 7.55 (d, J = 7.2 Ηζ, ΙΗ), 6.45 (s, 1H), 5.90 (d, J = 1.2)

82, ΙΗ), 5.82 (d, J = 1.2 Ηζ, ΙΗ), 5.37 (t, J = 6.0 Ηζ, ΙΗ), 4.15 (d, J = 2.1 Ηζ, ΙΗ), 4.04 (d, J = 1.8 Ηζ. ΙΗ), 3.70 (s, 3H), 3.663.53 (m, 2H), 3.37-3.21 (m, 2H), 3.19-3.15 (d, J =

11.7 Ηζ, ΙΗ), 3.08-3.00 (m, 2H), 2.56 (d, J = 18.3 Ηζ, ΙΗ), 2.30 (s, 3H), 2.24 (s, 3H).

2.04 (s, 3H), 1.91 (dd, Ji = 12.0 Hz, Jz = 15.6 Ηζ, ΙΗ), 0.84 (d, J = 6.9 Hz, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 170.1,15 & 3, 147.3, 144.9, 144.4, 143.3,136.7, 130.7,

111

129.3, 120.6, 111.6, 117.4, 114.4, 112.1, 107.7, 101.0, 85.8, 60.5, 59.3, 56.5, 56.4,

56.2, 55.2, 48.9, 41.6, 40.9, 25.7, 25.3, 18.0, 15.6, 8.7.

ESI-MS m / z: Calcd. For C32H35F3N5O7: 645.63. Found (M + H) ⁺ : 646.2.

Example 50

TFA, CH ₂ Cl ₂

4h, 23 ° C

OMe

To a solution of 17 (200 mg, 0.288 ml) in dichloromethane (1.44 ml) was added trifluoroacetic acid (888 ml, 11.53 ml) and the reaction mixture was stirred for 4 hours at 23 ° C. The reaction mixture was fixed at 0 ° C with saturated aqueous sodium bicarbonate solution (60 ml) and extracted with ethyl acetate (2x70 ml). The combined organic layers were dried over sodium sulfate and concentrated in vacuo to give 56 (147 mg, 93%) as a white solid, which was used in subsequent reactions without further purification.

Rf: 0.19 (ethyl acetate: methanol 5: 1).

1 H NMR (300 MHz, CH 3 OD): δ 6.48 (S, 1H), 5.88 (d, J = 0.9 Ηζ, ΙΗ), 5.81 (d, J = 0.9 Ηζ, ΙΗ), 4.35 (d, J = 2.4 Ηζ , ΙΗ), 4.15 (d, J = 1.8 Ηζ, ΙΗ), 3.99-3.98 (m, 1H), 3.70 (s, 3H), 3.52-2.96 (m, 7H), 2.68 (d, J = 18.3 Hz. 1H), 2.24 (s, 3H), 2.23 (s, 3H), 2.06 (s, 3H), 1.85 (dd, J, »11.7 Hz, J ₂ = 15.6 Ηζ, ΙΗ), 0.91 (d, J = 6.6 Hz, 3H).

^{, 3} C NMR (75 MHz, CH 3 OD): δ 173.2, 149.1,145.6, 144.9, 138.0,132.2, 130.6,

112

121.4, 120.6,119.6, 117.4,114.3,109.2,102.5, 82.3, 60.4, 58.4, 58.3, 57.8, 56.6,

50.1, 42.3, 41.6, 27.8, 26.2, 19.5, 15.5, 9.8.

ESI-MS m / z: Calcd. For C29H35N5O6: 549.62. Found (M + H) *: 550.3.

Example 51

57

To a solution of 56 (10 mg, 0.018 ml) in dichloromethane (0.4 ml) was added phenylisothiocyanate (13 ml, 0.109 ml) and the reaction mixture was stirred for 1.5 hours at 23 ° C. The reaction mixture was concentrated in vacuo and the residue purified by flash column chromatography (silica, gradienthexane to hexane: ethyl acetate 1: 1) to give 57 (8 mg, 65%) as a white solid.

Rf: 0.57 (ethyl acetatemethanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 7.88 (be, 1H), 7.41-7.36 (m, 2H), 7.27-7.22 (m, 1H), • 7.02-7.00 (d, J = 7.8 Hz, 2H) , 6.71 (d, J = 7.2 Hz, 1H), 6.31 (s, 1H), 6.17 (bs, 1H), 5.93 (d, J = 1.2 Ηζ, ΙΗ), 5.83 (d, J = 1.2 Ηζ, ΙΗ) , 5.55 (bs, 1H), 5.20-5.17 (m, 1H), 4.16 (d, J = 1.8 Hz, 1H), 4.05 (bs, 1H), 4.02 (d, J = 2.4 Ηζ, ΙΗ), 3.79 ( s, 3H), 3.75-3.71 (m, 1H), 3.35 (d, J = 7.8 Ηζ, ΙΗ), 3.28-3.19 (m, 2H), 3.12-2.97 (m, 2H), 2.50 (d, J = 18.3 Hz, 1H), 2.32 (s, 3H), 2.21 (s, 3H), 2.15 (dd, Ji = 11.4 Hz, J ₂ = 15.9 Hz, 1H), 1.95 (s, 3H), 0.88 (d, J = 6.9 Hz, 3H).

^{, 3} C NMR (75 MHz, CDCl 3): δ 178.5, 171.7, 147.2, 145.0, 144.3, 143.3, 137.0,135.7,

130.6, 130.4, 129.6, 127.5, 124.3, 120.6, 111.7, 117.2, 115.3, 112.1, 108.3, 100.9,

60.9, 59.5, 56.7, 56.5, 56.2, 55.2, 54.1, 41.7,41.1, 26.3, 25.4, 18.5, 15.8,9.0.

ESI-MS m / z: Calculated for Ο ^ Η ^ οΝεΟββ: 684.81. Found (M + H) *: 685.3.

113

Example 52

Me

Me

Acetyl chloride (4.67 ml, 0.065 ml) and pyridine (5.3 ml, 0.065 ml) were added to a solution of 57 (45 mg, 0.065 ml) in dichloromethane (0.5 ml) at 0 ° C. The reaction mixture was stirred for 3 hours and then the solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (RP-18, CH ₃ CN: water 40:60) to give 58 (14 mg, 28%) as a white solid. Rf: 0.34 (CHgCN: water 7:15).

1 H NMR (300 MHz, CDCl 3): δ 11.90 (d, J = 6.6 Ηζ, ΙΗ), 7.45-7.40 (m, 3H), 7.18-7.15 (m, 2H), 6.58 (s, 1H), 6.00 (d, J = 1.2 Ηζ, ΙΗ), 5.89 (d, J = 1.2 Hz, 1H), 5.70 (s, 1H), 5.37 (t, J = 4.8 Hz, 1H), 4.48 (m, 1H), 4.23 (bs, 3H), 4.07 (bs, 2H), 3.85-3.75 (m, 1H), 3.70 (s, 3H), 3.46-3.41 (m, 2H), 3.24-3.20 (m, 1H), 3.00-2.95 (m, 1H), 2.87-2.75 (m, 1H), 2.31 (s, 3H), 2.28 (s, 3H), 2.24 (s, 3H), 2.00 (s, 3H), 1.85 (dd, Ji = 11.4 Hz, J ₂ = 15.6 Ηζ, ΙΗ), 1.66 (s, 3H), 0.82 (¢ 4 = 6.0 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 182.6, 174.3, 171.0, 146.6, 144.6, 142.7, 142.3, 140.7,

140.2, 131.3,129.8, 129.3,128.9, 128.8,121.5, 120.4,117.3, 116.6,112.8, 112.0,

111.3, 101.5, 60.5, 59.0, 57.6, 56.2, 55.9, 55.3, 55.1, 41.6,39.4, 27.8, 26.5, 24.8,

114

20.2, 17.1, 15.5, 9.3.

ESI-MS m / z: Calcd. For C ₄ oH 44 N ₆ O ₈ S: 768.88. Found (M + H) *: 769.2.

Example 53

Me

Me

To a solution of 57 (130 mg, 0.189 ml) in dioxane (1 ml) was added 5.3 N hydrochloric acid / dioxane (1.87 ml) and the reaction mixture was stirred at 23 ° C for 4 hours. Dichloromethane (15 ml) and water (10 ml) were then added to the reaction mixture and the organic layer was decanted. The aqueous phase was basified with saturated aqueous sodium bicarbonate solution (60 ml) (pH 8) at 0 ° C and then extracted with ethyl acetate (2x50 ml). The combined organic extracts were dried over sodium sulfate and concentrated in vacuo to give 59 (63 mg, 70%) as a white solid.

Rf: 0.15 (EtOAc 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.67 (s, 1H), 5.99 (d, J = 0.9 Ηζ, ΙΗ), 5.91 (d, J = 1.2 Ηζ, ΙΗ), 5.10 (bs, 1H), 4.32 (d, J = 7.2 Ηζ, ΙΗ), 4.25 (dd, 4 = 3.6 Hz, 4 = 9.3 Ηζ, ΙΗ), 3.7 (s, 3H), 3.71 (m, 2H), 3.50 (dd, 4 = 2.4 Hz) , 4 = 15.9 Hz, 1H), 3.42-3.37 (m, 2H), 3.16 (dd, 4 = 3.6 Hz, 4 = 12.9 Hz, 1H), 2.57 (dd, 4 = 9-3 Hz, 4 = 12.9 Hz , 1H), 2.27 (s, 3H), 2.11 (s, 3H), 1.91 (dd, 4 = 12.0 Hz, 4 = 15.9 Hz, 1H).

ESI-MS m / z: Calcd. For C26H30N4O5: 478.5. Found (M + H) *: 479.3.

115

Example 54

Me

Trans-CgHgClO, Py CH ₂ CI ₂ .1h, 0 ^e C

To a solution of 43 (20 mg, 0.0338 mmol) in dichloromethane (0.3 ml) at 0 ° C was added sinamoyl chloride (5.63 ml, 0.0338 mmol) and pyridine (2.73 ml, 0.0338 mmol). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, ethyl acetate methanol 20: 1) to give 60 (22 mg, 90%) as a white solid.

Rf: 0.56 (ethyl acetate. Methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 7.51 (s, 1H), 7.50-7.47 (m, 2H), 7.36-7.35 (m, 2H), 6.43 (s, 1H), 6.36 (brd, J = 15.9) Hz, 2H), 6.01 (d, J = 1.5 Ηζ, ΙΗ), 5.90 (bid, J = 1.5 Hz, 2H), 5.42 (t, J = 6.0 Hz, 1H), 4.12-4.07 (m, 3H). 3.96-3.95 (m, 1H), 3.73 (bs, 3H), 3.58 (bs, 2H), 3.39 (d, J = 8.7 Ηζ, ΙΗ), 3.25 (d, J = 11.7 Hz, 1H), 3.00 (dd , 4 = 7.5 Hz, J ₂ = 17.7 Hz, 1H), 2.78 (d, J = 15.9 Hz, 1H), 2.67 (d, J = 16.5 Hz, 1H), 2.29 (S, 6H), 2.23 (S. 3H), 1.99 (s, 3H), 1.82 (dd, 4 = 11.4 Hz, J ₂ = 15.6 Hz, 1H), 0.83 (d, 4 = 6.0 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 172.0, 165.0, 146.9,144.6, 143.1, 141.0, 140.5, 134.8, 131.0, 129.7, 129.1, 128.8, 127.8, 125.5, 123.8, 123.0, 121.1, 120.5, 117.7, 116.9 ,

112.8, 112.0, 101.9, 60.6, 59.2, 57.1, 56.4, 55.9, 55.3, 48.8, 41.7, 40.0, 26.5, 25.1,

20.3, 18.5, 15.7,9.3.

ESI-MS m / z: Calcd. For C40H43N5O8: 721.8. Found (M + H) ⁺ : 722.3.

· · · · · · · · · · · · · · · · · · · · · · · 9 ·

116

Example 55

61

To a solution of 45 (19 mg, 0.0364 mmol) in dichloromethane (0.3 ml) at 0 ° C was added heptafluorobutyryl chloride (5.44 ml, 0.0364 mmol) and pyridine (2.95 ml, 0.0364 mmol). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, ethyl acetate: methanol 20: 1) to give 61 (11.7 mg, 45%) as a white solid.

Rf: 0.76 (ethyl acetate: methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.46 (s, 1H), 6.12 (bs, 1H), 5.98 (d, J = 1.2 Hz, 1H), 5.93 (d, J = 1.2 Hz, 1H), 5.72 (bs, 1H), 4.13-4.11 (m, 2H), 4.0 (d, J = 2.4 Ηζ, ΙΗ), 3.989 3.96 (m, 1H), 3.73 (s, 3H), 3.39 (d, J = 7.5 Ηζ , ΙΗ), 3.39-3.28 (m, 2H), 3.09 (dd, J, = 8.1

Hz, J ₂ = 18.0 Ηζ, ΙΗ), 2.80 (d, J = 16.2 Ηζ, ΙΗ), 2.46 (d, J = 18.3 Ηζ, ΙΗ), 2.32 (s, 6H), 2.21 (s, 3H), 1.99 (s, 3H), 1.80 (dd, J, = 12.0 Hz, J ₂ = 16.2 Ηζ, ΙΗ).

ESI-MS m / z: Calcd. For C32H31F7N4O7: 716.6. Found (M + H) ⁺ : 717.2.

Example 56

To a solution of 43 (24 mg, 0.04 mmol) in dichloromethane (0.3 ml) at 0 ° C was added butyryl chloride (4.15 ml, 0.04 mmol) and pyridine (3.28 ml, 0.04 mmol). The reaction mixture was stirred for 1 hour and after

117

this solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, ethyl acetate methanol 20: 1) to give 62 (24 mg, 90%) as a white solid.

Rf: 0.35 (EtOAc 5: 1).

Ή NMR (300 MHz, CDCl3): δ 6.47 (s, 1H), 6.10 (d, J = 6.5 Ηζ, ΙΗ), 6.0 (d, J = 1.5 Hz, 1H), 5.91 (d, J = 1.5 Ηζ. ΙΗ), 5.86 (bs, 1H), 5.31 (d, J = 6.9 Ηζ, ΙΗ), 4.11-4.06 (m, 3H), 3.85-3.81 (m, 1H), 3.75 (s, 3H), 3.59-3.53 (m, 2H), 3.38 (d, J = 7.5 Ηζ, ΙΗ), 3.27 (m, 2H), 3.0 (dd, 4 = 7.8 Hz, 4 = 17.4 Hz, 1H), 2.79 (d, J = 15.3 Ηζ , ΙΗ), 2.63 (d, J = 17.7 Ηζ, ΙΗ), 2.31 (s, 3H), 2.0 (s, 3H), 1.80 (dd, Ji = 12.0 Hz, 4 = 15.9 Ηζ, ΙΗ), 1.58 (q , J =

7.2 Hz, 2H), 0.89 (t, J = 7.2 Hz, 3H), 0.76 (d, J = 6.6 Hz, 3H).

ESI-MS m / z: Calcd. For C35H43N5O8: 661.64. Found (M + H) ⁺ : 662.3.

Example 57

С ₉ Н ₇ СЮ, Ру.

CH ₂ CI ₂ .0 ·, 1h.

118

• ···· · 9 9 «C ·

9 99 9 9 9 999

9 9 9 9 99

9 9 9 9 9 9 99

9 9 9 9 9 99

999 99 999 9999 99999

To a solution of 43 (19 mg, 0.0364 mmol) in dichloromethane (0.3 ml) at 0 ° C was added sinamoyl chloride (6.06 ml, 0.0364 mmol) and pyridine (2.95 ml, 0.0364 mmol). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica, ethyl acetate methanol 20: 1) to give 63 (20.1 mg, 85%) as a white solid.

Rf: 0.65 (ethyl acetate-methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 7.39-7.29 (m, 5H), 6.42 (s, 1H), 6.01 (d, J = 1.5 Ηζ, ΙΗ), 5.92 (d, J = 1.5 Ηζ, ΙΗ) , 5.73 (bs, 1H), 5.24 (t, J = 6.8 Hz, 1H), 4.12-4.08 (m, 3H), 3.66-

3.64 (m, 2H), 3.58 (bs, 3H), 3.36 (d, J = 8.7 Ηζ, ΙΗ), 3.29 (d, J = 12.0 Ηζ, ΙΗ), 2.98 (dd, 4 = 8.1 Hz, 4 = 18.0 33, ΙΗ), 2.33 (s, 6H), 2.29 (s, 3H), 2.01 (s, 3H), 1.84 (dd, 4 = 12.0 Hz, 4 = 15.9 Ηζ, ΙΗ).

ESI-MS m / z: Calcd. For C37H38N4O7: 650.72. Found (M + H) ⁺ : 651.2.

Example 58

OMe

NH

Me

To a solution of 43 (20 mg, 0.0338 mmol) in dichloromethane (0.3 ml) at 0 ° C was added 3-chloropropionyl chloride (3.22 ml, 0.0338 mmol) and pyridine (2.73 ml, 0.0338 mmol). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium

119

sulfate, filtered, and the solvent removed under reduced pressure.

The residue was purified by flash chromatography

not (silica, ethyl acetate methanol 20: 1) to give 64 (20.5 mg, 89%) as a white solid.

Rf: 0.32 (ethyl acetate: hexane 5: 1).

Ή NMR (300 MHz, CDCl3): δ 6.48 (s, 3H), 6.28 (m, 1H), 5.99 (d, J = 1.2 Ηζ, ΙΗ), 5.91 (d, J = 1.2 Ηζ, ΙΗ), 5.86 ( bs, 1H), 5.31 (m, 1H), 4.08-4.07 (m, 3H), 3.75 (s, 3H), 3.723.53 (m, 5H), 3.39 (d, J = 8.1 Ηζ, ΙΗ), 3.24 (d, J = 12.0 Hz, 1H), 3.00 (dd, Ji = 8.1 Hz, J ₂ = 18.0 Hz, 1H), 2.79 (d, J = 13.5 Ηζ, ΙΗ), 2.50 (t, J = 6.3 Hz. 2H), 2.32 (s, 3H), 2.28 (s, 3H), 2.25 (s, 3H), 2.0 (s, 3H), 1.79 (dd, Ji = 12.3 Hz, J ₂ = 14.8 Hz, 1H), 0.81 (d, J = 6.3

Hz, 3H).

Example 59

butyryl chloride

Py, CH ₂ CI ₂ . 1h.O ° C

To a solution of 43 (19 mg, 0.0364 mmol) in dichloromethane (0.3 ml) at 0 ° C was added butyryl chloride (3.78 ml, 0.0364 mmol) and pyridine (2.95 ml, 0.0364 mmol). The reaction mixture was stirred for 1 hour and then the solution was diluted with dichloromethane (10 ml) and washed with 0.1 N hydrochloric acid (5 ml). The organic layer was dried over sodium sulfate, filtered and the solvent was removed under reduced pressure. The residue was purified by flash column chromatography (silica 120

· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · 9 e. Dioxide, ethyl acetate: methanol 20: 1) to give 64 (19 mg, 87%) as a white solid.

Rf: 0.32 (EtOAc 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.50 (s, 1H), 5.98 (d, J = 1.5 Ηζ, ΙΗ), 5.91 (d, J = 1.5 Ηζ, ΙΗ), 5.75 (s, 1H), 5.01 (t, J = 6.4 Hz, 1H), 4.10-4.09 (m, 1H), 4.06 (d, J = 2.1 Hz, 1H), 4.03-4.02 (m, 1H), 3.76 (s, 3H), 3.67- 3.60 (m, 1H), 3.42-3.35 (m, 2H), 3.29 (d, J = 12.0 Ηζ, ΙΗ), 3.02 (dd, Ji = 7.8 Hz, J ₂ = 17.7 Ηζ, ΙΗ), 2.79 (d. J = 14.1 Ηζ, ΙΗ), 2.56 (d, J =

18.3 HZ, 1H), 2.32 (s, 3H), 2.31 (s, 3H), 2.25 (s, 3H), 1.78 (dd, Ji = 12.0 Hz, J ₂ = 15.9 Hz, 1H), 1.63 (s, 3H) ), 1.53-1.46 (m, 2H), 1.28-1.16 (m, 2H), 0.68 (t, J = 7.2 Hz, 3H). ESI-MS m / z: Calcd for C 30 H 30 N 3 O: 590.67. Found (M + H) ⁺ : 591.2.

Example 60

To a solution of 50 (31.7 mg, 0.044 mmol) in CHaCN / water (1.5 m | /0.5 ml) was added silver nitrate (225 mg, 1.32 mmol) and the reaction mixture was stirred at 23 ° C for 17 hours. Then, brine (10 ml) and aqueous saturated sodium bicarbonate solution (10 ml) were added to the reaction mixture at 0 ° C and the reaction mixture was stirred for 15 minutes, filtered through a pad of celite and washed with dichloromethane ( 20 ml). The solution was decanted and the organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate: methanol 5: 1) to give 66 (16 mg, 51%) as a white solid. ft ft ··· • · · · ft ft ft • · ft · ft ···· ··· ·· ··· ··· ·· ··· ···· ft · ··· to substance.

Rf: 0.26 (ethyl acetate: methanol 5: 1).

Ή NMR (300 MHz, CDCl 3): δ 7.66-7.42 (m, 4H), 7.20 (bs, 1H), 6.44 (s, 1H), 5.97 (b, J = 1.2 Hz, 1H), 5.90 (d, J = 1.2 Ηζ, ΙΗ), 5.76 (bs, 1H), 5.28 (bs, 1H), 4.54 (bs, 1H), 4.43 (bs, 1H), 4.00 (bs, 1H), 3.68-3.57 (m, 4H) , 3.47 (d, J = 3.3 Ηζ, ΙΗ), 3.40 (d, J =

11.7 Hz, 1H), 3.17 (d, J = 6.9 Hz, 1H), 2.92 (dd, Ji = 8.1 Hz, J2 = 17.7 Ηζ, ΙΗ), 2.74 (d, J = 17.1 Hz, 1H), 2.48 (d , J = 1.8 Hz, 1H), 2.32 (s, 6H), 2.28 (s, 3H), 1.99 (s, 3H), 1.76 (dd, J = 12.0 Hz, J ₂ = 16.2 Hz, 1H).

ESI-MS m / z: Calcd. For C37H38F3N3O8: 709. Found (M ⁺ -17): 692.3.

Example 61

Me Oh

To a solution of 53 (57 mg, 0.0828 mmol) in CHsCN / water (1.5 ml / 0.5 ml) was added silver nitrate (650 mg, 3.81 mmol) and the reaction mixture was stirred at 23 ° C for 24 hours. Then, brine (10 ml) and aqueous saturated sodium bicarbonate solution (10 ml) were added to the reaction mixture at 0 ° C and the reaction mixture was stirred for 15 minutes, filtered through a pad of celite and washed with dichloromethane ( 20 ml). The solution was decanted and the organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate: methanol 5: 1) to give 67 (28 mg, 50%) as a white solid.

122

Rf: 0.28 (ethyl acetate-methanol 10: 1).

¹ H NMR (300 MHz, CDCl 3): δ 6.47 (s, 1H), 5.97 (s, 1H), 5.35 (bs, 1H), 4.51 (bs, 1H), 4.41 (bs, 1H), 4.12-4.05 ( m, 1H), 4.00 (d, J = 2.7 Hz, 1H), 3.77 (s, 3H), 3.64 (bs, 1H), 3.46 (d, J = 3.3 Hz, 1H), 3.34 (d, J = 11.4 Ηζ, ΙΗ), 3.18 (d, J = 7.5 Hz, 1H), 2.95 (dd, Ji =

8.4 Hz, 4 = 18.3 Hz, 1H), 2.70 (d, J = 15.6 Hz, 1H), 2.48 (d, J = 17.7 Ηζ, ΙΗ), 2.28 (s, 3H), 2.27 (s, 3H), 2.26 (s, 3H), 1.98 (s, 3H), 1.68 (dd, 4 = 12.0 Hz, 4 = 15.6 Ηζ, ΙΗ), 0.86 (d, J = 6.3 Hz, 3H).

ESI-MS m / z: Calcd. For C32H37F3N4O9: 678.66. Found (M ⁺ -17): 661.2.

Example 62

To a solution of 48 (32 mg, 0.0529 mmol) in CH 3 Cl 2 / water (1.5 ml / 0.5 ml) was added silver nitrate (270 mg, 1.58 mmol) and the reaction mixture was stirred at 23 ° C for 24 hours. Then, brine (10 ml) and aqueous saturated sodium bicarbonate solution (10 ml) were added to the reaction mixture at 0 ° C and the reaction mixture was stirred for 15 minutes, filtered through a pad of celite and washed with dichloromethane ( 20 ml). The solution was decanted and the organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate: methanol 5: 1) to give 68 (18 mg, 56%) as a white solid.

123

Rf: 0.40 (ethyl acetate: methanol 5: 1).

¹ H NMR (300 MHz, CDCl 3): δ 6.50 (S, 1H), 5.95 (d, J = 1.2 Ηζ, ΙΗ), 5.88 (d, J = 1.2)

3, ΙΗ), 5.23 (d, J = 6.9 Ηζ, ΙΗ), 4.45 (d, J = 3.3 Ηζ, ΙΗ), 4.38 (s, 1H), 4.01 (d, J = 2.4

78, ΙΗ), 3.78 (m, 3H), 3.41-3.37 (m, 1H), 3.17-3.15 (m, 1H), 2.96 (dd, Ji = 7.8 Hz, J ₂ = 18.0 Ηζ, ΙΗ), 2.70 ( d, J = 15.3 Hz, 1H), 2.40 (d, J = 18.0 Hz, 1H), 2.30 (s, 6H), 2.27 (s,

3H), 1.76-1.65 (m, 1H), 1.35-1.25 (m, 2H), 0.89-0.82 (m, 1H), 0.69 (d, J = 6.6 Hz, 3H).

0.58 (d, J = 6.6 Hz, 3H).

To a solution of 51 (27 mg, 0.04 mmol) in CHaCN / water (1.5 ml / 0.5 ml) was added silver nitrate (204 mg, 1.19 mmol) and the reaction mixture was stirred at 23 ° C for 24 hours. Then, brine (10 ml) and aqueous saturated sodium bicarbonate solution (10 ml) were added to the reaction mixture at 0 ° C and the reaction mixture was stirred for 15 minutes, filtered through a pad of celite and washed with dichloromethane ( 20 ml). The solution was decanted and the organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate: methanol 5: 1) to give 69 (10 mg, 38%) as a white solid.

Rf: 0.38 (ethyl acetate: methanol 5: 1).

1 H NMR (300 MHz, CDCl 3): δ 6.48 (s, 1H), 6.16 (bs, 1H), 5.98 (d, J = 1.5 Hz, 1H),

124

5.89 (d, J = 1.5 HZ.1H), 5.33 (t, J = 6.0 Hz, 1H), 4.50 (m, 1H), 4.40 (m, 1H), 4.11-4.09 (m, 1H), 4.00 (d , J = 2.6 Hz, 1H), 3.78 (s, 3H), 3.41-3.32 (m, 3H), 3.18 (d, J = 8.4 Hz,

1H), 2.94 (dd, Ji = 8.4 Hz, J ₂ = 18.3 Ηζ, ΙΗ), 2.70 (d, J = 14.4 Hz, 1H), 2.45 (d, J = 18.3

Hz, 1H), 2.31 (S, 3H), 2.28 (s, 3H), 2.27 (s, 3H), 2.04 (s, 3H), 2.01-1.86 (m, 3H), 1.73 (m, 1H), 0.87 (d, J = 6.3 Hz, 6H).

Example 64

To a solution of 63 (15 mg, 0.023 mmol) in CH 2 DN / water (1.5 ml / 0.5 ml) was added silver nitrate (118 mg, 0.691 mmol) and the reaction mixture was stirred at 23 ° C for 24 hours. Then, brine (10 ml) and aqueous saturated sodium bicarbonate solution (10 ml) were added to the reaction mixture at 0 ° C and the reaction mixture was stirred for 15 minutes, filtered through a pad of celite and washed with dichloromethane ( 20 ml). The solution was decanted and the organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate methanol 5: 1) to give 70 (20.1 mg, 85%) as a white solid.

Rf: 0.43 (ethyl acetate-methanol 5: 1).

125 • · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · H NMR (300 MHz, CDCl3): δ 7.38-7.28 (m, 5H), 6.48 (s, 1H), 5.98 (d, J = 1.5 Ηζ). ΙΗ),

5.91 (d, J = 1.5 Ηζ, ΙΗ), 5.75 (bs, 1H), 5.38 (brd, 1H), 5.30 (bs, 1H), 4.53 (m, 1H), 4.42 (m, 1H), 4.02 (d , J = 2.7 Hz, 1H), 3.78-3.65 (m, 5H), 3.46-3.40 (m, 2H), 3.17 (d, J = 7.8 Ηζ, ΙΗ), 2.94 (dd, 4 = 7.8 Hz, 4 = 17.7 Hz, 1H), 2.73 (d, J = 16.8 Hz, 1H), 2.45 (d, J =

18.0 Hz, 1H), 2.31 (s, 3H), 2.28 (s, 3H), 1.97 (s, 3H), 1.77 (dd, 4 = 12.0 Hz, 4 = 15.3

Ηζ, ΙΗ).

Example 65

AgNO3, CHaCN / H10

OMe

To a solution of 65 (25 mg, 0.042 mmol) in CH 3 Cl 2 / water (1.5 m | /0.5 ml) was added silver nitrate (215.56 mg, 1.269 mmol) and the reaction mixture was stirred at 23 ° C for 24 hours. Then, brine (10 ml) and aqueous saturated sodium bicarbonate solution (10 ml) were added to the reaction mixture at 0 ° C, and the reaction mixture was stirred for 15 minutes, filtered through a filter pad and washed with dichloromethane (20 ml). The solution was decanted and the organic layer was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash column chromatography (silica, ethyl acetate methanol 5: 1) to give 71 (16 mg, 65%) as a white solid.

Rf: 0.05 (ethyl acetate-methanol 5: 2).

¹ H NMR (300 MHz, CDCl 3): δ 6.50 (s, 1H), 5.95 (d, J = 1.5 Ηζ, ΙΗ), 5.78 (s, 1H), 5.19 (bs, 1H), 4.45 (d, J = 3.3 Ηζ, ΙΗ), 4.37 (bs, 1H), 4.11 (brd, J = 4.8 Hz, 1H), 4.01 (d, J =

126

• ···· • ·· «· F ·· · ·· · · • · ·· • · • ♦ fff • · · • · fff · • · · • · • · f • · · · ♦ ··· ···· ·· ··

2.1 Ηζ, ΙΗ), 3.76 (s, 1 Η), 3.71-3.69 (m, 1H), 3.49-3.35 (m, 1H), 3.24 (d, J = 13.5 Hz.

1H), 3.15 (d, J = 9.3 Hz, 1H), 2.95 (dd, 4 = 8.1 Hz, J _z = 17.7 Hz, 1H), 2.70 (d, J = 15.6

Hz, 1H), 2.40 (d, J = 18.0 Hz, 1H), 2.31 (s, 3H), 2.29 (s, 3H), 2.26 (s, 3H), 1.96 (s, 3H).

1.75-1.66 (m, 3H), 1.52-1.17 (m, 2H), 0.66 (t, J = 7.2 Hz, 3H).

fermentation methods

Example A

YMP3 embryo medium containing 1% glucose, 0.25% bovine extract, 0.5% bactopeptone, 0.25% sodium chloride, 0.8% calcium carbonate was inoculated with 0.1% frozen vegetative mass of microorganisms, strain A2-2 of Pseudensonas and Pseudensonas flu, and is incubated in a rotary shaker (250 rpm) at 27 ° C. After incubation for 30 hours, the culture was added with stirring to a fermentation vessel containing medium consisting of 2% dextrose, 4% mannitol, 2% dried beer yeast (Vitalevor® Biolux, Belgium), 1% ammonium sulfate, 0.04% acidic potassium phosphate, 0.8% potassium chloride, 0.001% ferric chloride, 0.1% L-Tyr, 0.8% calcium carbonate, 0.05% polypropylene glycol PPG-2000, 0.2% silicone (ASSAF-100, RHODIA UK). Sterilization was carried out at 122 ° C for 30 minutes. The inoculated volume is 2% (v / v). The temperature is 27 ° C (0 to 16 hours) and 24 ° C 16 to the end of the process (41 hours). Dilution oxygen pressure is above 25%. The pH was adjusted to 6.0 using dilute sulfuric acid from the 28th hour until the end of the process. The overpressure is 0.5 bar. From the 16th hour to the end of the process (running for 2 days), 1% mannitol or sorbitol was added and 2% for the three-day fermentation process.

After 41 or 64 hours, the fermentation broth should be extracted to restore safracin B or the clarified broth treated with potassium cyanide to restore safracin B-cyano.

127

• · · · 9 99 9 9 • e 99 9 9 9 9 9 9 • 9 • 9 9 9 9 9 9 • 9 9 9 9 9 9 9 • 9 9 9 99 99 999 9999 99 9 9

Example B

Preparation of safracin B-cyano from crude extract

Solids are separated by clarification or filtration of the fermentation broth at pH 6. The clarified broth was adjusted to pH 9.5 using a dilute sodium hydroxide solution and extracted twice with 2: 1 (v / v) ethyl acetate, methylene chloride or butyl acetate. The extraction was carried out in a stirring vessel for 20 minutes while maintaining the temperature of the mixture from 8 to 10 ° C. The two phases are separated by liquid-liquid centrifugation. The organic phase was dried over anhydrous sodium sulfate or by freezing and then filtered to remove ice. The organic phase (ethyl acetate layer) was evaporated until a crude oil extract was obtained.

Example B

Preparation of safracin B-cyano from clear broth

Solids are separated by clarification or filtration of the fermentation broth at pH 6. The clarified broth was adjusted to pH 3.9 using concentrated acetic acid. To the clarified broth was added 0.5 grams per liter of potassium cyanide and incubated at 20 ° C for 1 hour with stirring. The temperature was then lowered to 15 ° C and the pH adjusted to 9.5 using a dilute sodium hydroxide solution and extracted with 2: 1.5 (v / v) ethyl acetate. The extraction was carried out in a stirring vessel for 20 minutes while maintaining the temperature of the mixture from 8 to 10 ° C. The two phases are separated by liquid-liquid centrifugation. The organic phase was dried over anhydrous sodium sulfate. The organic phase (ethyl acetate layer) was evaporated until a crude oil extract was obtained. This extract was purified by flash chromatography (silica, g128).

9 ···· ♦ ·· • · ·· · ·· · · • • ·· • · • · • • • · · • · • · • • · · • · • • ··· ·· ··· ···· ·· ··

diethyl 20: 1 to 10: 1 to 5: 1 ethyl acetate / methanol) to quantitatively obtain compound 2 as a light yellow solid.

Rf: 0.55 (ethyl acetate: methanol 5: 1): tR = 19.9 min [HPLC, Delta Pack C4, 5 μπι, 300 A, 150 x 3 mm, λ = 215 nm, flow = 0.7 ml / min, temperature = 50 ° C, gradient: aqueous CH3CN NaOAc (10 mM) 85% -70% (20 ')].

1 H NMR (300 MHz, CDCl 3): δ 6.54 (dd, 4 = 4.4 Hz, J ₂ = 8.4 Hz, 1H), 6.44 (s, 1H), 4.12 (d, J = 2.4 Hz, 1H), 4.04 ( d, J = 2.4 Hz, 1H), 4.00 (s, 3H), 3.87 (bs, 3H), 3.65 (ddd, 4 =

1.5 Hz, 4 = 8.7 Hz, 4 = 9.9 Hz, 1H), 3.35 (brd, J = 8.4 Hz, 1H), 3.15-2.96 (m, 4H), 2.92 (q, J = 7.2 Hz, 1H), 2.47 (d, J = 18.3 Hz, 1H), 2.29 (s, 3H), 2.18 (s, 3H), 1.83 (s, 3H),

1.64 (ddd, 4 = 2.7 Hz, 4 = 11.1 Hz, 4 = 14.1 Ηζ, ΙΗ), 0.79 (d, J = 7.2 Hz, 3H).

¹³ C NMR (75 MHz, CDCl 3): δ 186.0 (q), 175.9 (q), 156.2 (q), 146.8 (q), 142.8 (q),

140.7 (q), 136.6 (q), 130.5 (q), 128.8 (q), 127.0 (q), 120.5 (s), 117.4 (q), 116.5 (q),

60.8 (t), 60.4 (s), 58.7 (t), 56.2 (s), 55.7 (s), 54.8 (s), 54.8 (s), 54.4 (s), 50.0 (s), 41.6 (t). 39.8 (d), 25.2 (d), 24.4 (d), 21.2 (t), 15.5 (t), 8.4 (t).

ESI-MS m / z: Calcd. For C29H35N5O6: 549.6. Found (M + Na) ⁺ : 572.3.

Example D

Medium (501) consisting of 2% dextrose, 4% mannitol, 2% dry brewer's yeast (Vitalevor® Biolux, Belgium), 1% ammonium sulfate, 0.04% acid potassium phosphate, 0.8% potassium chloride, 0.001% ferric chloride 6 -hydrate, 0.1% L-tyrosine, 0.8% calcium carbonate, 0.05% polypropylene glycol PPG2000 and 0.2% silicone (ASSAF-100, RHODIA UK), poured into a 75 I fermentation vessel and, after sterilization, inoculated with culture (2%) of A2-2 strain (FERM BP-14) and the aerated stirring cultivation was carried out at from 27 ° C to 24 ° C for 64 hours (aeration at 75 liters per minute and stirring at from 350 to 500 rpm). The pH was controlled by automatic feeding with dilute sulfuric acid from the 27th hour until the end of the process. From the 16th hour until the end of the process, 2% mannitol was added. The culture medium thus obtained (45 I)

129 »· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · After separation of the cells by centrifugation, the pH was adjusted to pH 9.5 using a dilute sodium hydroxide solution and extracted twice with 25 liters of ethyl acetate. The mixture was placed in a stirring vessel at 8 ° C for 20 minutes. The two phases are separated by liquid-liquid centrifugation. The organic phase was frozen at -20 ° C and filtered to remove ice and evaporated to give 40 g of a dark crude oil extract. After introduction of the cyanide group and purification, 3 g of safracin B-cyano are obtained.

Example E

Medium (501) consisting of 2% dextrose, 4% mannitol, 2% dry brewer's yeast (Vitalevor® Biolux, Belgium), 1% ammonium sulfate, 0.02% acid potassium phosphate, 0.2% potassium chloride, 0.001% ferric chloride 6 -hydrate, 0.1% L-tyrosine, 0.8% calcium carbonate, 0.05% polypropylene glycol PPG2000 and 0.2% silicone (ASSAF-100, RHODIA UK), poured into a 75 I fermentation vessel and, after sterilization, inoculated with a culture (2%) of A2-2 strain (FERM BP-14) and the aerated stirring cultivation was carried out at from 27 ° C to 24 ° C for 41 hours (aeration at 75 liters per minute and stirring at from 350 to 500 rpm). The pH was controlled by automatic feeding with dilute sulfuric acid from the 28th hour until the end of the process. From the 16th hour until the end of the process, 1% mannitol was added. The culture medium thus obtained (45 I), after separation of the cells by centrifugation, was adjusted to pH 3.9 with 200 ml of concentrated acetic acid. 25 g of potassium cyanide 97% were added and, after stirring for 1 hour at 20 ° C, the pH was adjusted to 9.5 with the help of 1500 ml of 10% sodium hydroxide solution. It is then extracted with 35 liters of ethyl acetate. The mixture was placed in a stirring vessel at 8 ° C for 20 minutes. The two phases are separated by liquid-liquid centrifugation. The organic phase is dried over

130

anhydrous sodium sulfate and evaporated to give 60 g of a dark oil crude extract.

Chromatography afforded 4.9 g of safracin B-cyano.

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Use of a 21-Nuc compound of structure of formula (XIV), wherein at least one of rings A or E is quinoline, in syn-

Claims (33)

thesis as a starting compound 2. Method for the preparation of a compound of the fused ring structure of formula (XIV): characterized by Cxi ') that it involves one or more interactions starting from a 21-cyano compound of formula (XVI): CH ·., -X A CHO AXI R ¹⁸ .. CH. H _ \ R ¹⁵ k - CIS. N \ I R ¹ CN where: R ¹ is an amidomethylene group or an acyloxymethylene group; 133 R ^s and R ^{8 are} independently selected from -H, -OH or -OCOCH ₂ OH, or both R ⁵ and R ⁸ are ketogroup and ring A is a p-benzoquinone ring; both R ^14a and R ^14b are -H or one is -H and the other is -OH, -OCH ₃ or -OCH ₂ CH ₃ , or R ^14a and R ^14b together form a ketone group; and R ^{15 and} R ^{18 are} independently selected from -H or -OH, or both R ⁵ and R ⁸ are ketogroup and ring A is a p-benzoquinone ring. A method according to claim 2, wherein R ¹ is -CH 2 -NH-CO-CR ^25a R ^2a> R ^25c , in which R ^25a and R ^25b form a ketocrop or one radical is -OH, -NH 2 or -OCOCCH3 and the other is -CH2COCH3, -H, -OH or -OCOCCH3, provided that when R ^25a is -OH or -NH2, R ^25b is not-OH and R ^25c is -H, -CH 3 or -CH2CH3, or R ¹ is -CH2-O-CO-R, wherein R is -C (CH ₃₎ = CH-CH ₃ or CH _3. A method according to claim 2, characterized in that the cyan compound of formula (XVI) is cyanosafracin B. A method according to any of the preceding claims, characterized in that the compound of the fused ring structure of formula (XIV) is a compound of formula (XVIla): or of formula (XVIIb): 134 • · · · · ft ·· ft ft · • ft · ···· ft · ft ft • ft ft ft ft ft ft ft ·· ft ft ···· ··· ·· ··· ··· · · · · · Ft · · · · · · · where: R ¹ is possibly a protected or derivative aminomethylene group, optionally a protected or derivative hydroxymethylene group; R * is -H; or R ¹ and R ⁴ together form a group of formula (IV), (V), (VI) or (VII): R ⁷ is -OCH 3 and R ⁸ is -OH or R ⁷ and R® together form the group -O-CH 2 -O-; both R ^{, 4a} and R ^14b are -H or one is -H and the other is -OH, -OCH ₃ or -OCH ₂ CH ₃ , or R ^14a and R ^14b together form a ketone group; and R ^{15 is} H or -OH; R ^{21 is} H, -OH or -CN; and derivatives including their acyl derivatives and including derivatives wherein the group -NCH ₃ - in the 12th position is replaced by -NH- or -NCH2CH3-, and derivatives in which the -NH ₂ group in the compound of formula (VI) can give derivatives. A process according to claim 6, wherein R ⁵ is alkanoyloxy having from 1 to 5 carbon atoms. 7. The method of claim 6, wherein R ⁵ is acetyloxy. A process according to claim 5, 6 or 7, characterized in that R ^14a and R ^14b are hydrogen. A method according to any one of claims 5 to 8, characterized 135 is that R ^{14 15 16 17 *} is hydrogen. A method according to any one of claims 5 to 9, characterized in that R ²¹ is hydroxyl or cyano. A method according to any one of claims 5 to 10, characterized in that the compound is of formula (XVIIb). A method according to claim 11, wherein R ^{7 and} R ⁸ together form the group -O-CH ₂ -O-. A method according to any one of claims 4 to 11, characterized in that R ¹ and R ⁴ together form the group of formula (IV), (V), (VI) or (VII): A method according to any one of claims 4 to 11, characterized in that R 'is a possibly protected or derivative aminomethylene group, possibly protected or derivative hydroxymethylene group and R ⁴ is hydrogen. The method of claim 14, wherein R 'is the group -CH ₂ -NH ₂ or -CH ₂ -NH-aa, where aa is an acylamino acid group. The method of claim 15, wherein the compound is in the form of an N-acyl derivative of the group -CH ₂ -NH ₂ or -CH ₂ -NH-aa. The method of claim 16, wherein R 'is an N-acyl derivative, the acyl group being of formula -CO-R ^a , 136 • · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Wherein R ^a is alkyl, alkoxy, alkylene, arylalkyl, arylalkylene, amino acid acyl, or heterocyclyl, each of which may be substituted by halo, cyano, nitro, carboxyalkyl, alkoxy , aryl, aryloxy, heterocyclyl, heterocyclyloxy, alkyl, amino or substituted amino; or R ^a is aa. A method according to claim 15, 16 or 17, characterized in that one or more aa groups are present and are alanyl, arginyl, aspartyl, asparagyl, cystyl, glutamyl, glutaminyl, glycyl, histidyl, hydroxypropyl, isoleucyl, leucyl, lysyl, methionyl, phenylalanil, prolyl, seryl, threonyl, tyronyl, tryptophil, tyrosyl, valyl, or other amino acid acyl group. A method according to any one of claims 5 to 18, characterized in that one or more substituent groups are protected by a protecting group. A method according to any one of the preceding claims, characterized in that the product has the formula (XXPa): or formula (XXXN): '' Liirt + iMriM 137 ····· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Where R ¹ is -CH ₂ NH ₂ or CH ₂ OH, or a protected or derivative variant of this group and R ⁴ is -H; or R ^1a and R ⁴ together form a group of formula (IV), (VI) or (VII): R ⁵ is a hydroxyl group or a protected or derivative variant thereof; both R ^14a and R ^14b are hydrogen or one is hydrogen and the other is hydrogen or a protected or derivative of this group, -OCH ₃ or -OCH ₂ CH ₃ , or both R ^14a and R ^14b together form a ketone group; R ¹² is -NH-, -NCH ₃ - or -NCH ₂ CH ₃ -; R ¹⁵ is a hydroxyl group or a protected or derivative thereof; and R ¹⁸ is a hydroxyl group or a protected or derivative variant thereof. A method according to any of the preceding claims, wherein R ¹ is as defined for formula (XVIIb) and is preferably a medium-sized aminomethylene group; R ⁵ is as defined for formula (XVIIb) and is preferably derived from: 138 small hydroxyl group; R ¹² is as defined above and is preferably -NCH ₃ - and R ²¹ is hydroxyl or cyano. A method according to claim 21, wherein R ¹ is a hydrophobic group. The method of claim 22, wherein R ¹ is the group -CH ₂ -NH ₂ -CO-R ^a , in which R ^a has a linear chain of less than 20 atoms, preferably less than of 15 or 10 atoms. A method according to claim 20, 21 or 22, wherein R ⁵ is an acetyl group. A method according to any of the preceding claims, characterized in that it comprises the step: wherein R ⁵ for the final product is as defined for compound (XXXII) and may be different in the starting material and changed in the course of the process, R ¹⁸ is a hydroxyl group in the final product but may be a protected hydroxyl group in the starting material and altered during the process, R ¹² in the finished product may be the same as in the starting material or altered in the process, R ²¹ in the final product is as defined above, and if it is a hydroxyl group, is obtained from a cyano group during the process, R ^a is as defined above and can then be acylated as a step of the process to obtain the end product with an acylated R ^a group. A method according to any one of the preceding claims, characterized in that aa is alanyl. A method according to claim 26, characterized in that the starting compound contains an alanyl group which is protected by a Boc group. A method according to any one of the preceding claims, characterized in that it involves the interaction of: A method according to any one of the preceding claims, characterized in that it involves the interaction of: A method according to any of the preceding claims, characterized in that it involves an interaction wherein the group R ¹ is aminomethylene and converted to a hydroxymethylene group. A method according to any of the preceding claims, characterized in that the compound of group R ¹ , which is a hydroxymethylene group, is reacted with a reagent of formula (XIX) ° n ^ ° • ··· « • ·· • · • · ·· · · • · • · • · • · • · • · · is · ♦ · · • · · • · • · • · · · · · · · · · • · ·· where Fu is a protected functional group, Prot ³ is a protecting group and the dotted line indicates a possible double bond. 32. A process for preparing a 21-cyan compound of formula (XVI) according to claim 1, characterized in that it involves reacting a compound of formula (XV): in which the groups R ¹ , R ^s , R ⁸ , R ^14a , R ^14b , R ¹⁵ h R ¹⁸ are as defined above and R ²¹ is a hydroxyl group, with a cyanide ion source, to give the desired 21-cyan compound . 33. 21-Cyan compound of formula (XVI): wherein R ¹ is an amidomethylene group or an acyloxymethylene group; R ⁵ and R ^{8 are} independently selected from -H, -OH or -OCOCH ₂ OH, or both R ⁵ and R ⁸ are ketogroup and ring A is a p-benzoquinone ring; both R ^14a and R ^14b are -H or one is -H and the other is -OH, -OCH ₃ or -OCH2CH3 or R ^{, 4a} and R ^{, 4b} together form a ketone group; and R ^{15 and} R ^{18 are} independently selected from -H or -OH, or both R ⁵ and R ⁸ are ketogroup and ring A is a p-benzoquinone ring, with the exception of safracin B.